The Complex Microbiota of Raw Milk

REVIEW ARTICLE
The complex microbiota of raw milk

Lisa Quigley1,2, Orla O’Sullivan1, Catherine Stanton1,3, Tom P. Beresford1, R. Paul Ross1,3,
Gerald F. Fitzgerald2,3 & Paul D. Cotter1,3
1
Teagasc Moorepark Food Research Centre, Fermoy, Cork, Ireland; 2Microbiology Department, University College Cork, Cork, Ireland; and
3
Alimentary Pharmabiotic Centre, Cork, Ireland
Downloaded from https://academic.oup.com/femsre/article-abstract/37/5/664/541439 by guest on 08 August 2019

Correspondence: Paul D. Cotter, Teagasc Abstract
Food Research Centre, Moorepark, Fermoy,
Cork, Ireland. Tel.: +353 25 42 694; Here, we review what is known about the microorganisms present in raw milk,
fax: +353 25 42 340; including milk from cows, sheep, goats and humans. Milk, due to its high
e-mail: paul.cotter@teagasc.ie nutritional content, can support a rich microbiota. These microorganisms enter
milk from a variety of sources and, once in milk, can play a number of roles,
Received 20 November 2012; revised 14 June such as facilitating dairy fermentations (e.g. Lactococcus, Lactobacillus, Strepto-
2013; accepted 18 June 2013. Final version
coccus, Propionibacterium and fungal populations), causing spoilage (e.g.
published online 24 July 2013.
Pseudomonas, Clostridium, Bacillus and other spore-forming or thermoduric
DOI: 10.1111/1574-6976.12030 microorganisms), promoting health (e.g. lactobacilli and bifidobacteria) or
causing disease (e.g. Listeria, Salmonella, Escherichia coli, Campylobacter and
Editor: Dieter Haas mycotoxin-producing fungi). There is also concern that the presence of antibi-
otic residues in milk leads to the development of resistance, particularly among
Keywords pathogenic bacteria. Here, we comprehensively review these topics, while com-
dairy; spoilage; safety; health; probiotic; paring the approaches, both culture-dependent and culture-independent, which
milkborne pathogens.
can be taken to investigate the microbial composition of milk.
MICROBIOLOGY REVIEWS
frequently include Pseudomonas and Acinetobacter spp.

Introduction
Other strains of non-LAB genera are also encountered in
Milk is a highly nutritious food that can be obtained milk, as well as various yeasts and moulds (Quigley et al.,
from a variety of animal sources such as cows, goats, 2011). Human milk on the other hand is typically domi-
sheep and buffalo, as well as humans, for human con- nated by Streptococcus, Staphylococcus, Lactobacillus and
sumption. However, the high nutrient content of these Bifidobacterium spp. (Martın et al., 2007).
milks, which includes proteins, fats, carbohydrates, vita- The specific composition of the milk microbiota
mins, minerals and essential amino acids (Supporting directly impacts on the subsequent development of dairy
information, Table S1), all at a near neutral pH and at a products (Fig. 1). Microorganisms can bring about the
high water activity, provides an ideal environment for the fermentation of milk through the production of lactate
growth of many microorganisms. Some of these nutrients and have a variety of different impacts on the sensory,
are directly available to all microorganisms, while others texture, flavour and organoleptic properties of resultant
are provided following the metabolism of major compo- products (Wouters et al., 2002). Microorganisms can also
nents by specific populations to release components and negatively impact on milk quality and shelf life; for exam-
metabolites that are used by others (Frank, 1997). It is ple, psychrotolerant bacteria can proliferate during refrig-
generally accepted that the lactic acid bacteria (LAB), a eration and, through the production of extracellular
group of bacteria that ferment lactose to lactate, are a lipases and proteases, result in spoilage (Desmasures &
dominant population in bovine, goat, sheep and buffalo Gueguen, 1997; Hantsis-Zacharov & Halpern, 2007). The
milk, prior to pasteurisation. The most common LAB microbial composition of milk can also have health-
genera in milk include Lactococcus, Lactobacillus, Leuco- related implications in that the consumption of raw milk
nostoc, Streptococcus and Enterococcus. Psychrotrophic contaminated with pathogens can lead to, in some cases,
populations, which particularly establish themselves dur- severe illness (Oliver et al., 2009). In contrast, it is
ing cold storage, are also a major component and claimed that other raw milk microorganisms can
ª 2013 Federation of European Microbiological Societies FEMS Microbiol Rev 37 (2013) 664–698
Published by John Wiley & Sons Ltd. All rights reserved
Raw milk microbiota 665
Sources Role/ Significance

Food Technology
Lactococcus
Lactobacillus
Streptococcus
Leuconostoc
Enterococcus
Propionibacterium
Health Promotion
Lactococcus
Lactobacillus
Streptococcus
Leuconostoc
Enterococcus
Some yeasts

Spoilage
Pseudomonas
Acinetobacter
Chryseobacterium
Clostridium
Phage
Milk-associated
microorganisms Illness
Listeria
Staphylococcus
Escherichia coli
Campylobacter
Mycobacterium
Fungi - Aflatoxins
Microorganisms can enter milk from contact with the Once in the milk these microorganisms can play an important role
animal including teat, hides, faeces; also from the in dairy product manufacture; they may contribute to promoting
housing, bedding, feed, air and water. Contact with human health or enhancing food safety. On the other hand these
farm equipment and milking equipment as well as microorganisms can lead to spoilage of milk and dairy products or
insufficient farm or personnel hygiene may influence they may contribute to disease and illness in humans.
the microbial content of milk.
Fig. 1. The potential sources of the microorganisms that are present in raw milk and the role/significance that some of these have when present
in milk.
contribute to health by aiding digestion or by reducing traditionally employed and involve the growth of microor-
the frequency of allergies, including asthma and atopic ganisms in microbiological media (either general or selec-
diseases, in individuals who consume raw milk during the tive) supplemented with morphological, biochemical or
early years of life (Debarry et al., 2007; Braun-Fahrl€ander physiological characterisation (Quigley et al., 2011). These
& Von Mutius, 2011). This review will highlight the vari- testing methods are still the standard in industrial settings
ous microbial populations found in raw milk and the and typically involve tests to determine total bacteria
methods employed for their detection. It also addresses counts, reflecting general milk quality, or to detect specific
their sources, their subsequent significance with respect to pathogens or other microorganisms, which indicate
industrial applications and the contribution of specific whether contamination has occurred. Populations fre-
populations to food quality and health. quently tested for include thermoduric populations (resist-
ing pasteurisation), sulphate-reducing clostridia, Listeria
monocytogenes, Salmonella, coagulase-positive staphylo-
Methods employed to determine the
cocci, Escherichia coli, Enterobacteriaceae, coliforms and
microbial composition of milk
Bacillus cereus among others. These standard methods are
Many microbial communities are complex; that is, they legislated and accredited by National and International
are comprised of many different taxonomical groups of Accreditation Boards (e.g. http://eur-lex.europa.eu/LexUri
microorganisms. Raw milk is an example of an environ- Serv/LexUriServ.do?uri=OJ:L:2005:338:0001:0026:EN:PDF;
ment that contains a diverse and complex microbial popu- http://www.inab.ie/). These tests generally rely heavily on
lation (Quigley et al., 2011; Vacheyrou et al., 2011). Most the use of microbiological broths or agars that selectively
of our knowledge with respect to the identity of the support the growth of the target microbial population and
microorganisms that are present in raw milk, and resul- often include further confirmatory biochemical analysis.
tant dairy products, has been gained through the growth These approaches are usually low-tech and inexpensive
or ‘culturing’, and subsequent analysis, of these microor- but are relatively labour intensive and time-consuming,
ganisms. The ultimate identification of these cultured and in some cases, insufficient discriminatory power can
microorganisms involves phenotypic and/or genotypic be a problem. More recently, considerable efforts have
methods. Phenotypic methods are those which have been been made to develop more rapid, high-throughput tests
FEMS Microbiol Rev 37 (2013) 664–698 ª 2013 Federation of European Microbiological Societies
666 L. Quigley et al.
that rely on DNA-based, genotypic analysis. Such technol- isms from a variety of sources, including the teat apex,
ogies, which usually rely, at least to some extent, on the milking equipment, air, water, feed, grass, soil and other
application of polymerase chain reaction (PCR) technol- environments (Fig. 1; Coorevits et al., 2008; Lejeune &
ogy, can be used to confirm the results generated through Rajala-Schultz, 2009; Vacheyrou et al., 2011).
traditional tests, but their ability to serve as an alternative The bovine teat surface can contain a high diversity of
to culture-based analysis is increasingly being appreciated. bacteria (Braem et al., 2012; Monsallier et al., 2012;
One of the key benefits of replacing the culturing step Verdier-Metz et al., 2012). In one particularly detailed
relates to the fact that many microorganisms are averse to study, culture-dependent methods revealed that the bacte-
isolation using common culturing methods, thus poten- ria present could be classified at the phylum level as
tially leading to a significant underestimation of the micro- Firmicutes (76%), Actinobacteria (4.9%), Proteobacteria

bial communities. A number of factors must be considered (17.8%) and Bacteroides (1.3%). When this approach was
when applying these culture-independent methods. The supplemented by a clone library sequencing–based
selection of a protocol that efficiently extracts nucleic acids approach, some additional phyla, that is, Planctomycetes,
from as many of the microorganisms present as possible is Verrucomicrobia, Cyanobacteria, Chloroflexi and unclassi-
critical. One must also consider the use of strategies, such fied Bacteria, were detected at low levels (Verdier-Metz
as the use of DNA-binding agents or an alternative focus et al., 2012). Notably, a large percentage of the reads from
on RNA, to limit the risk of false positives resulting from this and other studies (Fricker et al., 2011) corresponded
the amplification of DNA from dead cells (Quigley et al., to as yet unidentified bacteria. Of those which could be
2011). Finally, a decision has to be made regarding the identified, many corresponded to technologically impor-
genes to be targeted. The oligonucleotides or probes can tant bacteria such as Lactobacillus, Leuconostoc and Entero-
be selected to detect target-specific genes or to provide an coccus spp. Bacteria that can be involved in flavour, aroma
overview of the microbiota within a particular niche and colour development in cheese such as coagulase-nega-
through the non-target-specific amplification of highly tive staphylococci as well as Arthrobacter, Brevibacterium
conserved genes, such as the 16S or 23S rRNA genes. In and Corynebacterium spp. were also detected. However,
the latter case, amplified products can then be analysed by some of the microorganisms detected on the teat surface,
techniques such as denaturing gradient gel electrophoresis for example, Solobacterium, Clavibacter and Arcanobacteri-
(DGGE), temporal temperature gradient gel electrophore- um spp., have not been identified in milk (Verdier-Metz
sis (TTGE) or single-stranded conformation polymor- et al., 2012), presumably reflecting a lack of competitive-
phism (SSCP; Quigley et al., 2011) to highlight similarities ness in milk environments should transfer occur. It was
or differences in the populations. These approaches may also noted that the composition of the microbial commu-
be used in conjunction with Sanger (first generation) DNA nity on the teat surface varied qualitatively and quantita-
sequencing, to help specifically identify the populations tively from one farm to another (Verdier-Metz et al.,
present. More recently, there has been a rapid evolution in 2012). This can be attributed to many different factors; for
next-generation DNA sequencing technologies that pro- example, microorganisms associated with bedding material
duce millions of sequence reads in a single run, thus allow- can contaminate the surface of teat and thus potentially
ing a much more in-depth and accurate estimation of enter milk (Vacheyrou et al., 2011). Similarly, milking
microbial diversity. The ever-increasing number and length machines can contain a reservoir of microorganisms, and
of the sequence reads provided by these technologies cou- thus, unsurprisingly, differences between machines and
pled with the availability of databases and bioinformatic related practices can influence the microbial population of
tools have been hugely beneficial with respect to the taxo- the milk collected (Michel et al., 2006). With respect to
nomic assignment of the microorganisms present (Loman more general environmental factors, it has been observed
et al., 2012). Although, to date, high-throughput sequenc- that the microorganisms present in cows’ milk depend on
ing approaches have not been extensively applied to assess whether animals are fed indoors or outdoors, with an
the microbiota of dairy-based environments, there have increase in Staphylococcus spp. during outdoor feeding
been a number of recent publications, which suggest that (Hagi et al., 2010), on the location of the animals (Bonizzi
this situation will change dramatically in the coming years et al., 2009) and on the lactation stage (Callon et al., 2007).
(Masoud et al., 2011, 2012; Alegria et al., 2012; Quigley An intense study was carried out to relate the microorgan-
et al., 2012). isms detected in milk to where they can be found on the
farm (Vacheyrou et al., 2011). These results highlighted
141 bacterial species, representing 54 genera, from
Sources of milk microorganisms
throughout the farm. There were 25 genera detected in
Milk in healthy udder cells is thought to be sterile (Tolle, these milk samples, and many of these, including Aerococ-
1980) but thereafter becomes colonised by microorgan- cus, Streptococcus, Propionibacterium, Acinetobacter, Bacillus,
Ochrobactrum, Pseudomonas, Psychrobacter, Staphylococcus, the United States with 90 million tonnes (http://www.
Sphingomonas, Enterobacter, Pantoea, Brachybacterium, dairyco.org.uk/market-information/supply-production/
Corynebacterium, Kocuria, Microbacterium and Pseudoclav- milk-production/world-milk-production/). This milk is
ibacter, were also detected in different areas throughout the employed in many ways, including direct consumption
farm including teat surfaces, milking parlours, hay, air and and the manufacture of dairy products and milk pow-
dust. Also present in milk, but not detected in the farm ders. Raw cows’ milk has the potential to contain a
environment, were technologically relevant bacteria such as diverse bacterial population as highlighted previously
Lactococcus, Lactobacillus and Enterococcus as well as Leu- (Quigley et al., 2011). Typically, cows’ milk contains a
cobacter, Deinococcus and Paracoccus. Similarly, a large significant LAB population that includes Lactococcus
number of other taxa were detected in the farm environ- (8.2 9 101–1.4 9 104 CFU mL 1), Streptococcus (1.41 9
101–1.5 9 104 CFU mL 1), Lactobacillus (1.0 9 102–

ment, but not in milk (Vacheyrou et al., 2011). Finally, it is
notable that the implementation of strict hygiene standards 3.2 9 104 CFU mL 1), Leuconostoc (9.8 9 101–2.5 9 103
brings about a reduction in the microbial load of milk, CFU mL 1) and Enterococcus spp. (2.57 9 101–1.58 9
including a reduction in populations of technological 103 CFU mL 1; Fig. 2). A number of other microorgan-
importance, which can, in turn, impact negatively on isms can be present in significant proportions. These
cheese manufactured using traditional or artisanal include psychrotrophs, such as Pseudomonas, Acinetobacter
approaches (Monsallier et al., 2012). Indeed, Mallet et al. and Aeromonas spp., which flourish during cold storage
(2012) recently reported a one-magnitude reduction in the (Raats et al., 2011). However, while the bacterial composi-
levels of technologically relevant lactococci present in raw tion of cows’ milk has been extensively studied for quite
milk relative to what had been detected 15 years before in some time, new developments with respect to DNA
raw milk collected from the same area (Desmasures & sequencing technologies have highlighted that the diversity
Gueguen, 1997). These populations seem to be particularly of these bacteria is greater than that originally appreciated
sensitive to the evolution of farm practices, as other popu- (Table 1). Indeed, a recent study applied high-throughput
lations, such as Pseudomonas, Lactobacillus and yeast popu- DNA sequencing to examine the bacterial population of
lations, did not differ across the two studies. While it is raw cows’ milk that was to be used for cheese production
important to ensure that the quality of milk is maintained (Masoud et al., 2012); 256 bacterial species were detected,
at high levels, producers of traditionally manufactured raw of which Streptococcus thermophilus and Lactococcus lactis
milk cheese should be aware that certain farming practices dominated in the milk, representing 43.7% and 19% of
may negatively impact on distinctive flavours and aromas reads, respectively. A number of other microorganisms that
as a consequence of limiting the numbers of specific micro- had previously been associated with raw milk, including
organisms and may need to compensate through the intro- Acinetobacter, Aeromonas, Brevibacterium, Corynebacterium,
duction of starters and adjunct strains. Lactobacillus, Pseudoalteromonas, Pseudomonas and Staphy-
lococcus, which represented between 1.3% and 3.7% of the
total reads, were also detected. A large subpopulation of
The microbial composition of different
taxa, which each corresponded to <1% of the total reads,
milk types
was also highlighted (Masoud et al., 2012). We have also
Although the largest proportion of commercially pro- recently compared the bacterial population present in
duced milk worldwide comes from cows, there are a cows’ milk pre- and postpasteurisation using high-
number of other animal sources of milk that is used for throughput sequencing to reveal the presence of a previ-
human consumption. These include quite common ously unrecognised and diverse bacterial population in
sources such as goats, sheep, buffalo and others utilised unpasteurised cows’ milk. While the milk, sourced from a
in more specific regions such as camel milk in African variety of commercial producers throughout Ireland, was
and Arab countries and yak milk in Asian countries. This dominated by Lactococcus, Pseudomonas and Leuconostoc,
section will review recent findings on the microbial con- we also detected a number of anaerobic taxa, including
tent of these various milks. We will also discuss an issue Bacteroides, Faecalibacterium, Prevotella and Catenibacteri-
that has been receiving ever more attention in recent um, which are more typically associated with the gut mic-
years; that is, the microbial composition of the human robiota and may be entering the milk through faecal
milk that is consumed by infants only (Box 1). contamination. Our analyses indicate that the bacterial
population of pasteurised milk is more diverse than previ-
ously appreciated, but that the nonthermoduric bacteria
Cows’ milk
that are present within these populations are likely to be in
Cows’ milk is produced on a massive scale. In 2012, the EU a damaged, nonculturable form (Quigley et al., 2013).
produced c. 139 million tonnes of cows’ milk followed by Thus, high-throughput sequencing approaches can provide
100% Other
Weisella
Streptococcus
% of genera detected
80%
Staphylococcus
Pseudomonas
60% Pseudoalteromonas
Propionibacterium
40% Leuconostoc
Lactococcus
20% Lactobacillus
Escherichia

Enterococcus
0% Brevibacterium
CD NGS CD NGS CD NGS CD NGS
Bifidobacterium
Cow Sheep* Buffalo Human Acinetobacter
Fig. 2. Relative abundance [This graph provides merely an overview derived from a number of separate studies (Heikkila & Saris, 2003; Delbes
et al., 2007; Devirgiliis et al., 2008; Maniruzzaman et al., 2010; Hunt et al., 2011; Masoud et al., 2011; Alegria et al., 2012; Cabrera-Rubio
et al., 2012; Ercolini et al., 2012; Quigley et al., 2013) using different methods and from various locations. To more accurately assess the outputs
generated by these respective approaches one should assess identical samples using both approaches.] of the most common bacteria in raw milk
samples as detected by culture-dependent (CD) or next-generation DNA sequencing (NGS)-based technologies. CD results are represented as a
percentage of total isolates. NGS results are represented as a percentage of total reads. The minor bacterial populations and species detected can
be viewed in the tables corresponding to each milk type found throughout the manuscript and, in particular, Tables 1–4. *Sheep milk NGS
information was extrapolated from data relating to naturally ripened cheeses manufactured using raw sheep milk.
a detailed insight into the bacterial composition of milk, media, SSCP analysis as well as restriction fragment
and it is likely that these technologies will be used increas- length polymorphism (RFLP) typing of isolates to exam-
ingly in future to investigate the factors that influence the ine the microbial diversity of 118 goats’ milk samples
composition of cows’ milk. taken from one herd throughout one lactation year to
reveal the presence of a diverse bacterial population in the
milk (Table 2). In addition to microorganisms commonly
Goats’ milk
encountered in milk, such as those listed above, some spe-
Goats’ milk production represents about 2.1% of global cies were identified that are not typically associated with
milk production (Tsakalidou & Odos, 2012). It is an goats’ milk or that had previously only been associated
important commodity that has gained increased interest with cheeses, including a number of corynebacteria and
as an alternative to cows’ milk, due to evidence that it is brachybacteria. Another unexpected finding was the pres-
less likely to induce allergies (Park, 1994). Goats’ milk ence of several halophilic species not previously associated
also differs from cows’ and sheep’s milk by virtue of hav- with milk, including Jeotgalicoccus psychrophilus, Salinicoc-
ing greater levels of iron bioavailability (Boyazoglu & cus sp., Dietzia maris, Exiguobacterium, Ornithinicoccus
Morand-Fehr, 2001) as well as containing smaller fat sp. and Hahella chejuensis. The significance of the presence
globules, having a higher content of fatty acids and form- of these microorganisms with respect to health, safety
ing a softer curd during subsequent fermentations, in or product development is not known. Through this
turn leading to greater digestibility (Klinger & Rosenthal, approach, it was also revealed that milks collected during
1997). Goats’ milk is most frequently used for cheese winter were dominated by the presence of Lactococcus and
making, usually at farm level or in small dairies. Goats’ Pseudomonas, those from summer by Pantoea agglomerans
milk cheeses are particularly common in Mediterranean and Klebsiella and those from autumn by Chryseobacteri-
countries and south-east Europe (Pirisi et al., 2007). um indologenes, Acinetobacter baumannii, Staphylococcus,
Goats’ milk is also typically dominated by LAB, including Corynebacteria and yeasts. While these variations can be
species of Lactococcus (3.7 9 106 CFU mL 1), Lactobacil- attributed to differences in feed, the authors suggested that
lus (1.34 9 105 CFU mL 1), Leuconostoc (3.27 9 103 other factors, such as weather conditions and the health of
CFU mL 1) and Enterococcus (2.95 9 102 CFU mL 1), the animal, were also important (Callon et al., 2007).
as well as Enterobacteriaceae, Micrococcaceae, moulds (fila- There has not been an in-depth assessment of the
mentous fungi) and yeasts (Alonso-Calleja et al., 2002; microbiology of goats’ milk since this study, perhaps next-
Tamagnini et al., 2006; Nikolic et al., 2008). Callon et al. generation sequencing technologies could have the poten-
(2007) relied on the use of selective microbiological tial to be very revealing.
Table 1. Bacterial populations detected in raw cows’ milk using culture-dependent, culture-independent and next-generation DNA sequencing
methods
Culture-dependent* Culture-independent† Next-generation sequencing‡
Acinetobacter species/johnsonii/junii/ Acinetobacter species/johnsonii/ Acinetobacter species

haemolyticus/lwoffii baumannii/junii
Aerococcus species/viridans Aerococcus Aerococcus species
Bacillus species/cereus Bacillus thuringiensis Bacillus subtilis
Brevibacterium helvolum/linens Brevibacterium species/samyangensis Brevibacterium linens
Chryseobacterium species Chryseobacterium species/joostei/ Chryseobacterium piscium
bifermentans/freundii
Corynebacterium ammoniagenes/freneyi/ Corynebacterium freneyi/casei/variabile/ Corynebacterium casei

glutamicum/ macginleyi
variabilis/casei
Enterococcus species/faecalis/gallinarum/ Enterococcus aquimarinus Enterococcus faecalis
saccharominimus
Escherichia coli Escherichia coli Escherichia coli
Lactobacillus casei/curvatus/mindensis/animalis/ Lactobacillus plantarum/pentosus/ Lactobacillus casei/helveticus/
coryneformis/curvatus/delbrueckii/johnsonii/ delbrueckii/acidophilus/fermentum plantarum/sakei/rhamnosus
paracasei/paraplantarum/plantarum/
rhamnosus/amylovorus
Lactococcus lactis/garvieae Lactococcus lactis/garvieae Lactococcus lactis
Leuconostoc mesenteroides Leuconostoc carnosum/ Leuconostoc
pseudomesenteroides
Staphylococcus species/capitis/cohnii/ Staphylococcus aureus/epidermidis/ Staphylococcus saprophyticus/
saprophuticus/equorum/xylosus/aureus/ fleuretii/sciuri succinus
haemolyticus/hominis/epidermis
Streptococcus species/uberis/parauberis Streptococcus species/uberis/dysgalactiae/ Streptococcus thermophilus
parauberis/thermophilus
Pseudomonas species/alcalophila/stutzeri/ Pseudomonas species/fragi/psychrophila/ Pseudomonas species/gessardii
synxantha/fluorescens/putida brenneri/synxantha/putida/pertucinogena
Microbacterium liquefaciens/oxydans/ Microbacterium species/xinjiangensis Macrococcus equipercicus
lacticum
Rhodococcus erythropolis Rothia Rothia mucilaginosa
Serratia liquefaciens/odorifera Stenotrophomonas species/koreensis Stenotrophomonas maltophilia
Enterobacter species/gergoviae Empedobacter brevis Enterobacter cloacae
Klebsiella ozaenae/oxytoca Klebsiella oxytoca Kurthia gibsonii
Kocuria species/carniphila/kristinae/ Kocuria species/pneumoniae Leptotrichia hofstadii
rhizophila
Frigoribacterium species Facklamia Facklamia tabacinasalis
Paracoccus species Nocardioides dubius Paracoccus carotinifaciens
Micrococcus species Ornithinicoccus species Marinomonas
Ochrobactrum anthropi/tritici Pandoraea species/norimbergensis Meiothermus species
Pantoea species/agglomerans Phyllobacterium myrsinacearum Methylobacterium extorquens
Propionibacterium freudenreichii/jensenii Propionibacterium Pediococcus pentosaceus
Providencia stuartii Proteobacteria Prevotella
Psychrobacter species/maritimus Ralstonia species/pickettii Psychrobacter
Pseudoclavibacter species/helvolus Sphingomonas melonis Pseudoalteromonas agarivorans
Rahnella aquatilis Thauera species Ruminococcus flavefaciens
Renibacterium salmoninarum Trichococcus Weissella hellenica
Sphingomonas species Yania halotolerans Actinomyces radicidentis
Achromobacter delicatulus Acidobacteria Alistipes finegoldii
Aeromonas hydrophila Adhaeribacter aquaticus Aeromonas species/
hydrophila/popoffii
Arthrobacter species/arilaitensis/ Arthrobacter species/arilaitensis/ Anaerococcus octavius
psychrolactophilus psychrolactophilus
Brachybacterium species/nesterenkovii Bacteroidetes Bacteroides
Deinococcus species Bosea thiooxidans Bifidobacterium species/
pseudolongum
Table 1. Continued
Culture-dependent* Culture-independent† Next-generation sequencing‡
Dermacoccus species Bradyrhizobium species Carnobacterium species

Hafnia alvei Caryophanon latum Empedobacter brevis
Clavibacter michiganensis Delftia Catenibacterium
Comamonas testosteroni Clostridium perfringens/lituseburense Caulobacter crescentus
Enhydrobacter aerosaccus Janibacter anophelis Faecalibacterium
Halomonas species Janthinobacterium lividum Jeotgalicoccus psychrophilus
Leucobacter species Paenibacillus apiarius Unassigned
Unassigned

The bacterial names emphasised in red highlight the most prevalent bacterial populations detected.
The bacterial names emphasised in blue highlight the less prevalent, but frequently isolated bacterial populations detected.
The bacterial names in black highlight the occasional bacteria that are detected.
Where there are bacterial names in bold, these are detected by two of the three methods.
Where the bacterial name is underlined, these were detected by only one of the methods.
All other bacteria were detectable by all of the methods.
References used for data in this table are Verdier-Metz et al. (2009), Raats et al. (2011), Vacheyrou et al. (2011), Mallet et al. (2012), Masoud
et al. (2012) and Quigley et al. (2013).
*Culture-dependent methods are based on isolation of bacteria using agar-based methods followed by identification using phenotypic or geno-
typic methods.
†
Culture-independent methods are based on direct extraction of bacterial DNA from the milk followed by identification using various techniques
including DGGE/SSCP/clone libraries etc.
‡
Next-generation sequencing methods are based on direct extraction of bacterial DNA from the milk sample followed by identification using
pyrosequencing.
(Alegria et al., 2012; Table 3). As this is a naturally fermented

Sheep milk
raw milk cheese, it is likely that these cheese-associated
Sheep milk is rarely consumed but still constitutes bacteria were also present in the corresponding raw milk.
c. 1.3% of global milk production as it is often employed A culture-based approach established that lactococci
throughout Europe in the development of cheese (Tsaka- (Lactococcus lactis ssp. lactis and ssp. cremoris) dominated
lidou & Odos, 2012). Sheep milk is dominated by LAB, (c. 109 CFU g 1), with lactobacilli (Lactobacillus casei,
with mesophilic bacteria representing 102–106 CFU mL 1, Lactobacillus plantarum, Lactobacillus parabuchneri and
while psychrotrophic populations correspond to 102– Lactobacillus brevis) also being common (107–108 CFU g 1).
104 CFU mL 1 (Fotou et al., 2011). Studies assessing the Leuconostoc (Leuconostoc citreum, Leuconostoc lactis and
impact of storing sheep milk at refrigeration temperature Leuconostoc mesenteroides) were detected at levels of 105–
highlighted increases in psychrophiles, but also in meso- 108 CFU g 1, fungal populations were present between
philes. Unsurprisingly, the thermoduric population did 105 and 106 CFU g 1 and Enterobacteriaceae, including
not increase. These general trends are also affected by Enterobacter kobei, were at 103–106 CFU g 1, but were
temperature and the length of storage (de Garnica et al., reduced during processing. A parallel DGGE investigation
2011). Other bacteria that have been detected on occasion confirmed the dominance of Lactococcus lactis but also
can include microorganisms of concern from a milk highlighted the presence of a significant population of
safety perspective including E. coli, Salmonella, Staphylo- Lactococcus garvieae, which had not been detected by cul-
coccus aureus, Bacillus and Clostridium perfringens (Fotou turing. This approach also revealed a number of minor
et al., 2011). The location can affect both the nutritional populations including Tetragenococcus halophilus, Strepto-
composition and microbial composition of sheep milk. A coccus salivarius, S. thermophilus and Streptococcus vestibu-
correlation has been noted between milks with a higher laris. A high-throughput sequencing–based approach
fat content and greater counts of LAB, coliforms and revealed the presence of 40 different genera in the cheese.
moulds. In populations of streptococci and S. aureus, This included 9 dominant genera, including 6 from the
there was an increase and a decrease in counts, respec- order Lactobacillales (which include the lactococci, lacto-
tively, in regions where the milk was more acidic and bacilli and related genera), which constituted 97% of
nutrient levels were lower (Yabrir et al., 2013). Some assigned sequences. The other dominant genera were the
insight into the microbiology of sheep milk was also pro- Bifidobacteriaceae, Enhydrobacter and unclassified Bacilli
vided by a recent study of the raw sheep milk cheese, (Fig. 2). The benefits of employing this technology were
Oscypek, which is manufactured without a starter culture again highlighted when previously overlooked populations
Table 2. Bacterial populations detected in raw goats milk using of Kocuria, Sanguibacter, Flavobacteria, Chryseobacterium,
culture-dependent and culture-independent methods Exiguobacterium, Staphylococcus and Chromohalobacter
Culture-dependent* Culture-independent† were detected. Notably, a considerable proportion,
Acinetobacter baumannii Acinetobacter baumannii c. 20%, of sequence reads could not be assigned, and so
Arthrobacter species Arthrobacter species the identity of these bacteria, and the importance of the
Bacillus thuringiensis/cereus Bacillus thuringiensis/cereus other subpopulations, will require further attention (Ale-
Brachybacterium Brachybacterium
paraconglomeratum paraconglomeratum
gria et al., 2012; Table 3).
Brevibacterium stationis Brevibacterium stationis
Chryseobacterium indologenes Chryseobacterium indologenes
Citrobacter freundii Citrobacter freundii Buffalo milk
Corynebacterium variable Corynebacterium variable

Delftia acidovorans Delftia acidovorans Buffalo milk is consumed in various countries around
Enterococcus faecalis/ Enterococcus faecalis/ the world, with India and Pakistan being the highest
saccharominimus saccharominimus consumers. It is not as common in Europe, but it does have
Exiguobacterium Exiguobacterium
Jeotgalicoccus psychrophiles Jeotgalicoccus psychrophiles
an important market in some Mediterranean countries
Kocuria rhizophila/Kristinae Kocuria rhizophila/Kristinae where it is utilised in making traditional mozzarella cheese.
carniphila carniphila The microbial content of raw buffalo milk has been
Lactobacillus casei Lactobacillus casei
assessed, through culturing, and found to contain a large
Lactococcus lactis/garvieae Lactococcus lactis/garvieae
Leuconostoc mesenteroides Leuconostoc mesenteroides population of LAB, including lactococci and lactobacilli, as
Microbacterium oxydans Microbacterium oxydans well as coliforms, E. coli, S. aureus and bacterial endosp-
Micrococcus species/caseolyticus Micrococcus species/caseolyticus ores, highlighting that while technologically relevant bacte-
Pantoea agglomerans Pantoea agglomerans
Pseudomonas species/putida/ Pseudomonas putida/aeruginosa
ria are present, microorganisms of concern with respect to
aeruginosa/fulgida quality and safety can also be found (Ercolini et al., 2004;
Salinicoccus species Salinicoccus species Han et al., 2007). Culture-independent methods, that is,
Staphylococcus epidermidis/ Staphylococcus epidermidis/
DGGE, have revealed that raw buffalo milk contains a rich
simulans/caprae/equorum caprae/simulans/equorum
Stenotrophomonas maltophilia Stenotrophomonas maltophilia diversity of bacteria that changes during subsequent fer-
Streptococcus mitis Streptococcus mitis mentation to manufacture traditional mozzarella (Ercolini
Dietzia maris et al., 2001). More recently, high-throughput sequencing
Enterobacter species/absuria
Hahella chejuensis has been applied to identify the bacterial populations pres-
Klebsiella milletis/oxytoca ent in buffalo milk and throughout the manufacture of
Ornithinicoccus species mozzarella cheese (Table 3; Ercolini et al., 2012). The
Rothia species
dominant microorganisms in the milk were Lactococcus
Next-generation sequencing methods are not applicable in contrast to spp. (30%), Acinetobacter spp. (21%), Pseudomonas spp.
Table 1, as at the time of writing this article, next-generation DNA (20%), Streptococcus macedonicus (10%) and Lactococcus
sequencing technology had not been applied to monitor the microbial lactis (10%; Fig. 2). A number of other microorganisms
content of raw goat milk or any source which may represent the
were detected in low abundance including Brochothrix,
microbial content of raw goat milk.
The bacterial names emphasised in red highlight the most prevalent
Carnobacterium, Chryseobacterium, Clostridium, Corynebac-
bacterial populations detected. terium, Enterobacteriaceae, Gammaproteobacteria and
The bacterial names emphasised in blue highlight the less prevalent, Haloanella. There was also a large percentage of unassigned
but frequently isolated bacterial populations detected. reads (c. 20%) corresponding to the raw milk. This
The bacterial names in black highlight the occasional bacteria that are percentage was much greater than that associated with the
detected. corresponding cheese (Ercolini et al., 2012).
Where there are bacterial names in bold, these are detected by two
of the three methods.
Where the bacterial name is underlined, these were detected by only Milk from other animal sources
one of the methods.
All other bacteria were detectable by all of the methods. Other milks that are consumed by humans around the
References used for data in this table are Alonso-Calleja et al. (2002), world include those produced by camels, yaks, donkeys
Callon et al. (2007), Goetsch et al. (2011). and, to a lesser extent and in only some countries, mares.
*Culture-dependent methods are based on isolation of bacteria using Camel milk is commonly consumed in African and Arab
agar-based methods followed by identification using phenotypic or
countries where it is a valuable food resource for pastoral
genotypic methods.
†
Culture-independent methods are based on direct extraction of
people. The microbial population of camel milk, like that
bacterial DNA from the milk followed by identification using various of other milks, can play a role in subsequent fermentations,
techniques including DGGE/SSCP/clone libraries etc. health promotion and milk spoilage. The milk is typically
dominated by mesophiles, including lactobacilli (such as
Table 3. Bacterial populations detected in raw milk of sheep or buffalo using culture-dependent, culture-independent and next-generation DNA
sequencing methods
Milk source Culture-dependent* Culture-independent† Next-generation sequencing‡
Sheep Milk§ Lactococcus lactis Lactococcus lactis/garvieae/raffinolactis Lactococcus

Lactobacillus casei/plantarum/parabuchneri/ Lactobacillus plantarum/paraplantarum/ Lactobacillus
brevis helveticus/crispatus
Leuconostoc citreum/lactis/mesenteroides/ Leuconostoc citreum/mesenteroides/ Leuconostoc
pseudomesenteroides pseudomesenteroides/lactis
Streptococcus thermophilus/agalactiae Streptococcus vestibularis/salivarius/uberis- Streptococcus
parauberis/thermophilus
Enterococcus faecalis/durans/italicus/kobei Enterococcus Enterococcus

Bacillus simplex Tetragenococcus halophilus Tetragenococcus
Pediococcus Actinobacteria
Corynebacterium species Bifidobacterium
Staphylococcus species Chromohalobacter
Salmonella species Chryseobacterium
Escherichia coli Enhydrobacter
Enterobacter kobei Flavobacteria
Kocuria
Sanguibacter
Staphylococcus
Unassigned
Buffalo Milk Lactobacillus species/plantarum/paracasei/ Lactobacillus species/delbrueckii Lactobacillus species/kefiranofaciens
fermentum/delbrueckii
Lactococcus species Lactococcus lactis Lactococcus species/lactis
Staphylococcus species Leuconostoc lactis Pseudomonas species/fragi
Streptococcus Streptococcus thermophilus Streptococcus thermophilus/macedonicus
Bacillus species Enterococcus species/faecalis Acinetobacter species/johnsonii
Escherichia coli Carnobacteria species
Unassigned Clostridium species/hiranonis
Corynebacteria species
Enterobacteria species
Unassigned
Reference used for data on sheep milk is Alegria et al. (2012). For buffalo milk, the references are Ercolini et al. (2001), Devirgiliis et al. (2008),
Maniruzzaman et al. (2010), and Ercolini et al. (2012).
typic methods.
†
‡
pyrosequencing.
§
Sheep milk information is from naturally ripened cheeses manufactured using raw sheep milk, indicating that the microbial population is
potentially from the raw sheep milk microbial communities.
Lactobacillus helveticus, L. casei ssp. casei and L. planta- rally fermented camel milk, Streptococcus infantarius ssp.
rum), lactococci (such as Lactococcus lactis ssp. lactis), infantarius dominates, with significant populations of Lac-
streptococci (such as S. salivarius) and leuconostoc (all at tococcus lactis ssp. lactis, S. thermophilus and L. helveticus
102–107 CFU mL 1; Khedid et al., 2009). As with other also detected (Jans et al., 2012). Another animal that is typ-
milks, camel milk can also contain human pathogens, ically associated with difficult climates is the yak. c. 92% of
including Salmonella, Listeria and E. coli, the prevalence of the world’s yaks are located in China, and yak milk can be
which can vary with location (Abeer et al., 2012). In natu- an important commodity in some regions of China. Again
Box 1. Human milk
Human milk has the potential to protect a newborn against logical changes in the mother during the labour process may
infectious disease through the provision of active components influence the composition of the bacterial community. Finally, as a
including immunoglobulins, immunocompetent cells, fatty acids, consequence of nursing periods, the bacterial population of breast
oligosaccharides and glycoproteins (Saaverda, 2002; Newburg, milk eventually became dominated by microorganisms from the oral
2005). Furthermore, breast milk is also a source of microorganisms cavity and skin (Cabrera-Rubio et al., 2012). Following on from
and of growth factors that contribute to their growth in the gut. these recent investigations, it will be important to continue to
Traditionally, it has been thought that these microorganisms are unravel the roles of these individual microbial components with
transferred from the mother’s skin (West et al., 1979). However, respect to mammary gland health, colonisation of the infant gut

recent studies have indicated that bacteria present in the maternal a n d , s u b s e q u e n t l y, m a t e r n a l a n d i n f a n t h e a l t h .
gut can reach the mammary gland by vertical transfer (Albesharat Importantly, it is already known that human milk bacteria can play
et al., 2011). This topic requires further investigation. Regardless of important roles in the infant gut. They can contribute to a reduction
the origin of these microorganisms, human milk constitutes one in the incidence and severity of infections by different mechanisms
of the primary sources of the bacteria that colonise the gut of through competitive exclusion, the production of antimicrobial
breastfed infants. Indeed, an infant consuming c. 800 mL day 1 of compounds or improving intestinal barrier function (Olivares et al.,
human milk would be predicted to ingest between 105 and 107 2006). Many human milk lactobacilli and bifidobacteria can
bacteria daily (Heikkila & Saris, 2003). This may explain why some contribute to infant digestion by aiding in the breakdown of
studies have shown that the bacterial composition of the gut complex foods such as proteins and sugars; some lactobacilli
microbiota of breastfed infants closely resembles that found in the increase the production of functional metabolites such as butyrate,
breast milk of their mothers (Albesharat et al., 2011). As for other which is utilised as an energy source and can improve intestinal
milks, a variety of different approaches have been taken to function (Asakuma et al., 2011; Zivkovic et al., 2011; Gil-Campos
investigate the microbiology of human milk. Traditional culture- et al., 2012), while various bifidobacteria have positive effects on
based methods have indicated that staphylococci, LAB and propi- health, including prevention of infection by pathogenic bacteria
onibacteria dominate in human milk with a significant Bifidobac- (e.g. protection against diarrhoea), immunostimulatory and anti-
terium population also present (Table 4; Martin et al., 2009). carcinogenic capabilities, lowering of serum cholesterol and allevi-
Similarly, several studies have demonstrated the transfer of ation of lactose maldigestion (Fernandez et al., 2012).
Staphylococcus, Lactobacillus, Bifidobacterium as well as Enterococ- While breast milk can introduce a number of potentially health-
cus spp. from mother to infant through breastfeeding (Martin et al., promoting bacteria, it may also contain pathogens (Jones, 2001).
2004, 2009; Albesharat et al., 2011). The application of culture- Mastitis can be a common disease, with incidence rates of up to 33%
independent molecular techniques, and particularly those based on in lactating mothers (Foxman et al., 2002; the issue of bovine mastitis
analysis of 16S rRNA gene, has facilitated even more detailed is addressed later in the review). Mastitis results in the inflammation
analyses. These approaches have provided quite similar findings, of the mammary lobules, usually due to the presence of staphylo-
that is, a dominance of staphylococci and streptococci and the cocci, streptococci or corynebacteria. Traditionally, S. aureus has
presence of LAB, propionibacteria and bifidobacteria. However, DNA been considered as the main causative agent; however, Staphylo-
from other bacterial groups, including Weissella, Clostridium and coccus epidermidis is emerging as a leading cause of subacute and
Serratia, was also detected (Table 4; Martin et al., 2004, 2009). acute mastitis in both women and veterinary medicine (Delgado
More recently, through the application of high-throughput sequenc- et al., 2009).
ing, an even more diverse population has been uncovered (Table 4; The possible presence of pathogenic or spoilage microorganisms in
Hunt et al., 2011; Cabrera-Rubio et al., 2012). Some taxa that were human breast milk also needs to be considered in the context that
consistently found across all samples included species of Strepto- mothers may have to store milk for a variety of different reasons.
coccus, Staphylococcus, Serratia, Pseudomonas, Corynebacterium, Human milk may be stored at neonatal units and used as a life-
Ralstonia, Propionibacterium, Sphingomonas and Bradyrhizobiaceae saving therapy to high-risk infants (Silvestre et al., 2006). The
(Fig. 2; Hunt et al., 2011; Cabrera-Rubio et al., 2012). The microbial influence of refrigeration on the bacterial content of human milk,
composition of breast milk changed from prelabour to postbirth. over a 4-day period, has recently been assessed; refrigeration
Where birth was by nonelective caesarean section, the bacterial prevented the growth of total aerobic bacteria, LAB and Entero-
profile was more comparable with milk from mothers who had a bacteriaceae, while reducing the levels of coagulase-positive
natural labour than that of milk from mothers who underwent staphylococci (Giribaldi et al., 2013).
elective caesarean section. These results suggest that the physio-
the LAB dominate, with yeasts and coliforms also being Lactobacillus fermentum, Lactobacillus kefiranofaciens,
common (Zhang et al., 2008a). Other studies have L. plantarum ssp. plantarum, L. brevis, Lactobacillus buchneri,
identified L. casei, Lactobacillus delbrueckii ssp. bulgaricus, Leuconostoc lactis, L. mesenteroides, Lactococcus lactis ssp.
cremoris, S. thermophilus, Enterococcus faecalis, Enterococcus microbial content of donkey milk has been found to be
durans and Weissella cibaria as being among the most similar to that of other milks, consisting of LAB, coliforms
common species in yak milk (Watanabe et al., 2008; Yu and fungi, with mesophiles being detected at levels up to
et al., 2011). Donkey milk is less commonly consumed 104 CFU mL 1 and psychrotrophs at 102 CFU mL 1
and, for the same reason, is less extensively studied. The (Zhang et al., 2008b; Sarno et al., 2012).
Table 4. Bacterial populations detected in raw human milk using culture-dependent, culture-independent and next-generation DNA sequencing
methods
Next-generation
Human milk Culture-dependent* Culture-independent† sequencing‡

Hunt et al. (2011), Bifidobacterium species/adolescentis/ Bifidobacterium species/longum/ Bifidobacterium
Cabrera-Rubio et al. bifidum/breve/longum adolescentis/animalis/bifidum/breve/
(2012) and Fernandez catenulatum
et al. (2012) Corynebacterium species Corynebacterium species Corynebacterium
Enterococcus species faecium/faecalis/ Enterococcus faecalis/faecium Enterococcus
durans/hirae/mundtii
Lactobacillus species/acidophilus/fermentum/ Lactobacillus species/fermentum/gasseri/ Lactobacillus
plantarum/gasseri/crispatus/rhamnosus/ rhamnosus
salivarius/reuteri/casei/gastricus/vaginalis/
animalis/brevis/helveticus/oris
Lactococcus species/lactis Lactococcus species/lactis Lactococcus
Leuconostoc species/mesenteroides Leuconostoc species/citreum/fallax Leuconostoc
Streptococcus species/mitis/salivarius/oris/ Streptococcus species/mitis/parasanguis/ Streptococcus
parasanguis/lactarius/australis/gallolyticus/ salivarius
vestibularis
Staphylococcus species/epidermidis/aureus/ Staphylococcus species/epidermidis/hominis Staphylococcus
capitis/hominis
Rothia species/mucilaginosa Clostridium species Rothia
Pediococcus species/pentosaceus Weissella species/cibaria/confusa Weissella
Peptostreptococcus species Propionibacterium species/acnes Propionibacterium
Kocuria species/rhizophila Acinetobacter Acinetobacter
Pseudomonas Pseudomonas
Sphingomonas
Ralstonia
Serratia
Stenotrophomonas
Veillonella
Actinomyces
Bradyrhizobium
Carnobacterium
Citrobacter
Gemella
Granulicatella
Lysinibacillus
Prevotella
Unassigned
typic methods.
†
‡
pyrosequencing.
present, including a number of genes involved in fermen-

Technologically relevant bacteria of raw tation as well as a novel gene, poxL, encoding pyruvate
milk oxidase, which may play a role in switching between fer-
As described above, raw milk can contain a diverse bacte- mentation modes. Forty-three insertion elements were
rial population. Many such bacteria can contribute subse- identified, the distribution of which suggests that recent
quently to natural fermentations. In some situations, recombination between two closely related genomes may
specific strains have been so successful in this regard that have occurred (Bolotin et al., 2001). Sequencing of Lacto-
they have been isolated from milk and consciously added coccus lactis ssp. cremoris MG1363 revealed some similari-
as starters or adjuncts designed to confer desirable traits ties to strain IL1403, including proteolytic systems and
on fermented products. This can be particularly impor- genes associated with the utilisation of lactose. The

tant in situations where regulations require the use of absence of virulence genes from these genomes is consis-
pasteurised milk, and thus, the re-introduction of dairy tent with their ‘generally regarded as safe’ (GRAS) status
microorganisms can compensate for the removal of com- (Wegmann et al., 2007). Genome sequencing of Lactococ-
mensal populations and the associated adverse effect on cus lactis ssp. cremoris strain A76 revealed that it has
the flavour of resultant products. This section will review 99.2% identity to the industrially important strain crem-
what is known about the most technologically important oris SK11 and identified two contiguous regions associ-
genera of raw milk bacteria. ated with the ssp. lactis lineage. The first contains genes
involved in cell wall biosynthesis. The second region cor-
responds to a prophage. The presence of these regions
Lactococcus
suggests that they were introduced through a recombina-
Lactococcus consists of seven species, two subspecies and tion event and highlights the potential importance of
one biovar (www.bacterio.cict.fr; as of November 2012). such events among strains of industrial importance (Bolo-
Of these, Lactococcus lactis ssp. lactis and Lactococcus lactis tin et al., 2012). The Lactococcus lactis ssp. cremoris SK11
ssp. cremoris can dominate in raw milk, cheese and other strain contains a number of unique plasmids, pSK11A,
(unheated) dairy products. pSK11B, pSK11L and pSK11P, which encode important
In dairy foods, Lactococcus lactis, and Lactococcus lactis traits related to dairy adaptation and utilisation, including
ssp. lactis and Lactococcus lactis ssp. cremoris in particular, lactose utilisation, a complex proteolytic system and an
are primarily known for their role as starter cultures for oligopeptide permease system (Siezen et al., 2005). Lacto-
the cheese industry. Lactococcus lactis ssp. lactis biovar coccus lactis ssp. lactis biovar diacetylactis strain DPC 3901
diacetylactis is also recognised for its production of contains four unique plasmids: pVF18, pVF21, pVF22
flavour-developing compounds (Hugenholtz & Starrenburg, and pVF50. While sequence analysis of these plasmids has
1992). These microorganisms are distinguished from one revealed that this bacterium most likely originated from a
another on the basis of arginine and citrate utilisation, plant origin, there are some features that highlight its
growth temperature and salt tolerance (Kahala et al., adaption to milk, for example plasmid pVF59, which
2008). While these microorganisms are naturally present contains genes of relevance for growth on milk. These
in raw milk and artisanally produced cheeses (Gaya et al., include genes encoding a lactose phosphotransferase
2001), they are frequently added to pasteurised milk to operon, a protein predicted to function in D-lactate util-
facilitate the commercial manufacture of cheeses (Smit isation, a system for uptake of oligopeptides generated
et al., 2005). Their primary role during cheese production from casein degradation as well as oligopeptidases, which
is acidification through the production of L-lactate. How- allow this strain to utilise casein as a nitrogen source
ever, they also contribute to proteolysis, the conversion of (Fallico et al., 2011).
amino acids into flavour compounds (alcohols, ketones, A number of other species of Lactococcus are naturally
aldehydes), citrate utilisation and/or fat metabolism present in raw milk. Although Lactococcus raffinolactis is
(Smit et al., 2005). A comparison of 20 Lactococcus lactis typically not used by the dairy industry because of a lack
strains, 10 of the ssp. lactis phenotype and 10 of the ssp. of caseinolytic activity (Holler & Steele, 1995), a recent
cremoris phenotype, confirmed two major subspecies lin- study observed synergism between L. raffinolactis and Lac-
eages that were distinguished on the basis of the presence tococcus lactis strains, whereby L. raffinolactis improved
or absence of 4571 gene orthologs (Bayjanov et al., 2009; acid production thanks to its ability to utilise metabolic
Fernandez et al., 2011). Thus, it is estimated that these products generated by Lactococcus lactis (Kimoto-Nira
phenotypically similar subspecies diverged c. 17 million et al., 2012), presumably thanks to the presence of a
years ago (Bolotin et al., 2004). Sequencing of the gen- complete set of genes for lactate fermentation and genes
ome of Lactococcus lactis ssp. lactis IL1403 revealed that responsible for an oligopeptide ABC transporter (Meslier
all known genes required for energy metabolism were et al., 2012b;). Even though L. garvieae is a recognised
fish pathogen (Vendrell et al., 2006), it has been detected manufacture of dairy products. The release of free fatty
in raw milk, some natural mixed starter cultures and acids following lipolysis introduces important flavour
artisanal cheeses (Fortina et al., 2007; Foschino et al., compounds (Hickey et al., 2007). Genome sequencing of
2008). Genomic studies have shown that L. garvieae L. helveticus DPC4571, a Swiss cheese isolate, revealed
isolates from dairy and fish sources form two distinct that the presence of a high percentage of pseudogenes
clusters and that only the dairy isolates possess the ability that are associated with loss-of-function events and
to utilise lactose (Fortina et al., 2007, 2009). It is hypoth- presumably reflect adaptation to the dairy niche (Callanan
esised that this key phenotype has been gained by dairy et al., 2008). The growth of L. helveticus in milk is
isolates through lateral gene transfer (Ferrario et al., dependent on a complex system of proteolytic enzymes,
2012). However, strains from both clusters lack proteo- which collectively enable strains to access essential amino

lytic activity (Fortina et al., 2007). Finally, Lactococcus acids (Christensen et al., 1999). Genome sequencing of
piscium has been detected in raw milk and raw milk DPC4571 revealed the presence of a number of peptidase
cheese (Carraro et al., 2011). This is typically regarded to genes (pepE, pepQ, pepT and pepD) that are likely involved
be a salmon-associated species (Williams et al., 1990), in the proteolytic activity of this strain. PepQ is a proline-
which is psychrotrophic and has been investigated with a specific peptidase, which may be significant given that
view to its use as a biopreservative, albeit not in a dairy milk casein has a relatively high number of proline resi-
context (Fall et al., 2010). Little is known regarding its dues. Also, PepE, an endopeptidase, may have a role in
contribution to dairy products and how it has adapted to reducing bitter defects during cheese ripening. DPC4571
grow in dairy environments. lacks genes for colonisation and interaction with the
mucosal surface that are present in related gastrointestinal
probiotic species. These traits include surface proteins, cell
Lactobacillus
wall anchoring proteins and mucus-binding proteins
The genus Lactobacillus is very diverse and, according to (Callanan et al., 2008). An isolate from naturally
the most recent estimations, consists of 174 different spe- fermented milk, L. helveticus H10, has a larger genome
cies and 27 subspecies (www.bacterio.cict.fr). Lactobacilli than DPC4571 (Zhao et al., 2011). Most of the functional
can be found in rich, carbohydrate-containing niches, genes of this strain are conserved with DPC4571, but there
including those associated with plants, animals, silage and are also 300 unique genes and 130 genes that are absent
raw milk (Bernardeau et al., 2008). An ever greater compared with DPC4571. Lactobacillus delbrueckii can be
understanding of Lactobacillus biology has led to the use divided into three major subspecies, that is, ssp.
of strains of Lactobacillus for an increasing range of delbrueckii, which is plant derived, ssp. bulgaricus and ssp.
industrial dairy applications. In particular, their proteo- lactis (Giraffa et al., 2008). Representatives of the latter
lytic activity and ability to produce aroma compounds two subspecies have been regularly detected in raw milk
and exopolysaccharides can contribute to the quality and samples, as well as being dominant populations in many
nutritional value of dairy products (Leroy & De Vuyst, traditionally manufactured cheeses and protected designa-
2004). Lactobacilli that are of particular importance tion of origin (PDO) cheeses (Torriani et al., 1999;
within the dairy industry are L. helveticus, L. delbrueckii Randazzo et al., 2002; Morandi et al., 2011). Both exhibit
ssp. bulgaricus and L. delbrueckii ssp. lactis (the latter two strong proteolytic activity (Giraffa et al., 2004; Kholif
species will be referred to as L. bulgaricus and Lactobacillus et al., 2011; Agyei & Danquah, 2012). Lactobacillus bulgar-
lactis hereafter). icus is, as a consequence of its worldwide use in yoghurt
Lactobacillus helveticus was first described by Orla- production, one the most important dairy-associated
Jensen in 1919 as an isolate from an Emmental cheese lactobacilli. It acts synergistically with S. thermophilus,
(Naser et al., 2006), but it has since been evident that allowing rapid growth and acidification with desired orga-
representatives of this species are commonly isolated from noleptic properties (Herve-Jimenez et al., 2009). A num-
raw milk and raw milk based products (Quigley et al., ber of factors are known to play a role in this
2011). Lactobacillus helveticus has a number of traits that ‘protocooperation’ process including degradation of milk
are desirable with respect to cheese production. These proteins by L. bulgaricus; also, the production of formate
include rapid autolysis of the strains, which results in the and CO2 by S. thermophilus can stimulate the growth of
release of intracellular enzymes and a reduction in bitter- L. bulgaricus. Genomics has revealed important factors,
ness and increased flavour notes in cheese (Broadbent which may play a role in this beneficial interaction. While
et al., 2011). Lactobacillus helveticus is also characterised L. bulgaricus encodes a full set of genes for biosynthesis of
by its ability to grow at relatively high temperatures folate, it is thought that S. thermophilus is required to
(55 °C; Kiernan et al., 2000; Hannon et al., 2003) and is produce p-aminobenzoate, which feeds this pathway. Also,
the most proteolytic of the LAB frequently used in the in silico analysis predicted the existence of a limited
number of cell wall–bound and extracellular proteins, second most important industrial dairy starter after Lacto-
which could contribute to direct contact between L. bul- coccus lactis. Its importance in dairy products is due to its
garicus and S. thermophilus (van de Guchte et al., 2006). ability to rapidly convert lactose to lactate; bringing about
As with other milk-associated microorganisms, genomic a rapid decrease in pH; and the production of important
studies suggest that the genome of L. bulgaricus is in an metabolites including low levels of formate, acetoin,
active state of gene elimination and size reduction but diacetyl, acetaldehyde and acetate (Ott et al., 2000). Many
does contain genes encoding complete transport systems S. thermophilus strains produce exopolysaccharide that
for lactose as well as mannose, glucose, fructose and glyc- contributes to the desirable viscous texture and rheologi-
erol (van de Guchte et al., 2006). As expected, strain- cal properties of fermented milk products, particularly
specific features are evident. For example, the industrially yoghurt. In cheese, S. thermophilus is used alone or in

important bacterium L. bulgaricus strain 2038 has a num- combination with several lactobacilli and mesophilic
ber of unique features including a gene set involved in starters, but in yoghurt, it is always used with L. bulgari-
exopolysaccharide synthesis, which may improve synaere- cus. While S. thermophilus is, in general, readily isolated
sis (separation of liquid) as well as texture, viscosity and by traditional microbiological methods, there are some
mouthfeel of the final product. This strain also contains a instances where it has been overlooked, and rapid molec-
larger genome than other strains (Hao et al., 2011). ular and culture-independent methods, such as SSCP
There are several other lactobacilli in raw milk that (Randazzo et al., 2002) and DGGE (Duthoit et al., 2005),
increase in number during the manufacture of dairy were required to detect its presence.
products and can become particularly dominant during Whole-genome sequencing of S. thermophilus has
the ripening of cheese (Henri-Dubernet et al., 2008). revealed that this bacterium exhibits 80% similarity with
These populations, which are often referred to as non- other streptococci, indicating that it shares a substantial
starter LAB (or NSLAB), have an ability to adapt to the part of its overall physiology and metabolism with its
conditions of cheese ripening, where many nutrients are pathogenic relatives. The analyses suggest that their com-
depleted, pH is reduced, and moisture content is low. mon ancestor dates back 3000–30 000 years, roughly
Here, they are able to carry out proteolysis and lipolysis corresponding to the origin of human dairy activity (i.e.
to produce many end products that contribute to flavour c. 7000 years ago). The species lacks a number of func-
and texture development of cheese (Smit et al., 2005). tional genes typically found in other streptococci, includ-
These include L. casei, Lactobacillus paracasei, L. planta- ing many genes involved in carbohydrate utilisation as
rum/paraplantarum, Lactobacillus rhamnosus and Lactoba- well as virulence-related genes, for example, genes for
cillus curvatus, L. brevis, Lactobacillus sake, Lactobacillus some surface proteins that are required for adhesion to
pentosus, Lactobacillus acidophilus, Lactobacillus reuteri, mucosal surfaces, and antibiotic modification genes. This
Lactobacillus johnsonii, Lactobacillus crispatus, L. fermen- lack provides strong evidence to support the GRAS status
tum, L. buchneri and Lactobacillus gasseri. While the levels of S. thermophilus. Genome sequencing has also revealed
of these species can be underestimated if culture-dependent the presence of more than 50 insertion sequences, which
methods are relied upon exclusively, the use of culture- have shaped the genome and, in part, are associated with
independent techniques such as DGGE, TTGE and qPCR, genes involved in the adaptation of the species to milk.
or a combination of these with culturing, has addressed this Indeed, a number of genes appear to have been acquired
issue in a number of instances. from other dairy microorganisms through horizontal gene
transfer (Bolotin et al., 2004). Streptococcus thermophilus
strain ND03, isolated from naturally fermented yak milk,
Streptococcus
contains unique genes not previously found in other
The genus Streptococcus consists of 97 species and 17 sub- strains. These are contained within a large insertion island
species (www.bacterio.cict.fr). Although many genera of and encode a transposase, a glutamate decarboxylase, an
streptococci are pathogenic, S. thermophilus carries a acetyltransferase, a polysaccharide biosynthesis protein
‘GRAS’ status (Facklam, 2002) and is frequently isolated and proteins associated with exopolysaccharide biosynthe-
from dairy environments, including raw milk, natural sis (Sun et al., 2011). A comparative genome analysis of
starter cultures and cheese curds (Duthoit et al., 2005; 47 S. thermophilus strains has revealed that the gene con-
Randazzo et al., 2006; Santarelli et al., 2008). Strains of tent can be split into core genes (58%; present in all
S. thermophilus have also been detected in the teats of strains) and noncore genes. The noncore genes can be
cows, cowsheds and dairy facilities (Vacheyrou et al., split into conserved genes (14%), variable genes (20%)
2011; Braem et al., 2012). Streptococcus thermophilus is a and acquired genes (8%). The latter genes – in addition
thermophilic LAB widely used as a starter culture in the to those referred to above with respect to ND03 –
manufacture of dairy products. It is often regarded as the encode, for example, bacteriocins, efflux/uptake pumps,
proteins involved in peptide metabolism, phage proteins peptones of milk prior to milking and consequently
as well as phage resistance genes (Rasmussen et al., 2008). results in a reduced ability of milk to coagulate during
Other streptococci that have been associated with milk fermentations (Merin et al., 2008). Similarly, S. agalactiae
and milk products include Streptococcus uberis, Streptococ- is a major causative agent associated with mastitis in
cus agalactiae, Streptococcus dysgalactiae Streptococcus bovis cows. It has been proposed that a number of genes,
and S. macedonicus. Streptococcus macedonicus has been including a lac operon, have been acquired through lat-
isolated from artisanal raw milk cheeses (Pacini et al., eral gene transfer to allow this bacterium to adapt to the
2006; De Vuyst & Tsakalidou, 2008) and has displayed bovine host. An unusually high number of insertion ele-
some desirable characteristics from a dairy technology ments have also been detected suggesting frequent geno-
perspective. These include the ability to acidify and to mic rearrangement (Richards et al., 2011). Finally,

produce peptidases and the generation of inhibitory com- S. bovis is an opportunistic human pathogen, which is
pounds while, importantly, lacking antibiotic resistance often associated with infections in immunocompromised
and haemolytic activity (Lombardi et al., 2004; De Vuyst patients or patients with cancer. It can also be detected in
& Tsakalidou, 2008). Genome sequencing of a dairy iso- other environments including fermented foods (Barile
late of S. macedonicus revealed that the bacterium is et al., 2012).
undergoing regular genome decay as indicated by the
presence of a large number of pseudogenes. The genus
Propionibacterium
also appears to lack the pil1 locus that is involved in
instances of infectious endocarditis caused by pathogenic The genus Propionibacterium comprises two distinct
relatives (Papadimitriou et al., 2012). Nonetheless, further groups from different habitats, that is, strains typically
investigations are required to definitively establish the found on human skin, referred to as the ‘acnes group’,
safety status of the species with respect to its use in dairy and strains isolated from milk and dairy products,
products. An African dairy isolate, Streptococcus infantari- referred to as ‘dairy’ or ‘classical’ propionibacteria
us ssp. infantarius strain CJ18, contains plasmids with (Meile et al., 2008). Notably, it has also been claimed
high sequence identity to Lactococcus lactis sequences. The that dairy propionibacteria possess health-promoting
presence of a gal-lac operon suggests an evolutionary characteristics (Cousin et al., 2011). The dairy group of
adaptation to the dairy niche (Jans et al., 2012). More propionibacteria comprises four species: Propionibacte-
recently, comparative genomic analysis has revealed that rium freudenreichii, Propionibacterium acidipropionici,
the gal-lac operon of CJ18 has 91% identity with that of Propionibacterium jensenii and Propionibacterium thoenii
S. thermophilus. CJ18 also contains an oligopeptide trans- (www.bacterio.cict.fr). Propionibacterium freudenreichii
port operon, which is important during growth in milk serves as a starter in Swiss-style cheeses. It was first iso-
for the uptake of peptides and amino acids, and lacks lated over a century ago from an Emmental cheese and
classical streptococcal virulence factors (Jans et al., 2013). contributes to hole or ‘eye’ formation and flavour
Many of the other streptococci regularly detected in formation in these cheeses (Langsrud & Reinbold, 1973).
bovine milk are associated with mastitis infection. Masti- The other species of dairy Propionibacterium are usually
tis-associated pathogens typically infect the teat canal of isolated from milk and different cheese types (Meile
cows and pass into the milk during milking. The presence et al., 2008). The characteristic trait of P. freudenreichii
of these microorganisms impairs milk quality and the is the fermentation of lactate into propionate, acetate
quality of subsequent products (Barbano et al., 2006). and CO2, while associated distinctive flavours arise from
Streptococcus uberis is an animal pathogen and one of the the formation of fatty acids, through lipolysis, and of
major causes of bovine mastitis worldwide (Bradley et al., branched-chain acids from the catabolism of amino
2007). Genomic analysis provides evidence of the nutri- acids. Due to its long documented use in cheese manu-
tional flexibility of S. uberis that allows it to occupy vari- facture, P. freudenreichii has a GRAS status (Cousin
ous ecological niches, including mammary glands (Ward et al., 2011). Whole-genome sequencing of P. freud-
et al., 2009). Streptococcus dysgalactiae, a contagious and enreichii CIRM-BIA1T revealed its ability to cope with
environmental pathogen, also accounts for a notable pro- different stresses including oxidative, bile salt and tem-
portion of bovine mastitis infections (Todhunter et al., perature stresses and an ability to resist phage attack, to
1995). A number of genes are involved in its ability to accumulate nutrients and to mobilise these during peri-
adhere to the mammary gland, where it can survive and ods of starvation and to synthesise most vitamins and
is protected from the immune system as well as therapeu- amino acids. Sequencing also revealed the presence of a
tics. The severity of disease differs with genotype (Beecher number of genes that are thought to encode surface
et al., 2012). It has been suggested that S. dysgalactiae proteins potentially involved in adhesion and immuno-
present in the mammary gland degrades the proteose regulatory activity. The ability of P. freudenreichii to
utilise lactose has been found to be strain dependent. ecological niches that include the gastrointestinal tracts of
CIRM-BIA1T possesses a lactose utilisation locus encod- humans and animals (Giraffa, 2002) and, depending on
ing a b-galactosidase, a galactose transporter and an the strain in question, can be considered to be starter cul-
UDP-glucose isomerise. This locus is surrounded by tures, probiotics, spoilage or pathogenic organisms
transposable elements, is highly similar to corresponding (Bhardwaj et al., 2009). Due to their psychrotrophic
regions in strains of Clostridium and Mannheimia and, nature, ability to survive adverse conditions, including
thus, is believed to have been acquired through gene high-temperature and high-salinity environments, and
transfer to facilitate the adaptation of the microorganism adaptability to different growth substrates and growth
to the dairy environment. Importantly, genes involved in conditions, enterococci can survive refrigeration. In labo-
pathogenicity of Propionibacterium acnes are absent from ratory-based experiments, strains of this bacterium have
CIRM-BIA1T (Falentin et al., 2010).

been shown to potentially survive pasteurisation and thus
may be part of the microbial populations in both raw
and pasteurised milk as well as in subsequent products
Leuconostoc
(Giraffa, 2003; Ladero et al., 2011; McAuley et al., 2012).
The genus Leuconostoc consists of 23 species and 4 subspe- Studies on raw milk cheeses indicate that enterococci are
cies (www.bacterio.cict.fr). Leuconostoc spp. are frequently a common, and frequently important component of the
associated with plant material, but some, and in particular natural cultures involved in fermentations and contribute
the species mesenteroides and pseudomesenteroides, are also to ripening, taste and flavour (Foulquie Moreno et al.,
found in milk. However, it is possible that this is due to 2006). The most common enterococcal species in milk
their introduction during the collection of milk or subse- and dairy products are E. faecalis and Enterococcus
quent storage and processing. Notably, in this regard, faecium, but others, including E. durans (Franciosi et al.,
Leuconostoc spp. have the ability to survive on surfaces, 2009), Enterococcus italicus and Enterococcus mundtii, are
tools and pasteurisers for long periods of time and to resist also encountered. Enterococci contribute to fermentations
heat treatments and refrigeration temperatures (Hemme & due to their proteolytic activity; ability to hydrolyse milk
Foucaud-Scheunemann, 2004). Leuconostoc spp. grow fat; and contribution to the development of flavour com-
poorly in milk due to a lack of sufficient proteolytic activity pounds, including acetaldehyde, acetoin and diacetyl
and thus require the addition, or generation by other (Franz et al., 1999). Recent genome sequencing projects
microorganisms, of amino acids or peptides to stimulate identified large gene sets related to dairy adaption,
growth (Vedamuthu, 1994; Hemme & Foucaud-Scheune- including genes involved in lactose and galacto-oligosac-
mann, 2004). Leuconostoc spp. have the ability to produce charide utilisation, in E. mundtii. Furthermore, a large
gas (CO2), which is responsible for eye formation in some number of putative antibiotic resistance determinants
artisanal raw milk or blue-veined cheeses (Cardamone have also been found (Magni et al., 2012). Importantly,
et al., 2011); metabolise lactose and citrate; and produce food isolates of E. faecalis lack a large number of genes,
lactate, acetate, ethanol, acetaldehyde, diacetyl, acetoin and which are present in clinical isolates. These traits (includ-
2,3-butanediol, which contribute to the organoleptic prop- ing genes for adhesion and an entire prophage) are
erties of fermented dairy products (Vedamuthu, 1994; San- believed to contribute to the development of human
chez et al., 2005). Due to these attributes, Leuconostoc spp. infection (Lepage et al., 2006).
can act as beneficial NSLAB cultures. Genome sequencing
of the dairy isolate L. pseudomesenteroides strain 4882 has
Other milk microorganisms with
further highlighted the beneficial attributes, for example,
potential technological relevance
genes involved in carbohydrate fermentation, protein and
amino acid metabolism and a key pathway in production There are some other groups of microorganisms of tech-
of aromatic compounds from citrate (Meslier et al., nological relevance, albeit not regarded as being as impor-
2012a). Notably, while phenotypic assays do not always tant as those discussed above, which are present in low
reliably differentiate between species and subspecies of Leu- quantities in raw milk. These include a number of bacte-
conostoc, molecular methods can facilitate the rapid charac- ria, including Gram-positive and Gram-negative bacteria,
terisation of Leuconostoc to species level (Duthoit et al., as well as yeast and mould populations.
2003; Sanchez et al., 2006; Martin-Platero et al., 2009).
Gram-positive subpopulations
Enterococcus
Corynebacterium spp. were detected in milk over 40 years
Enterococci are the most controversial group of food- ago (Jayne-Williams & Skerman, 1966) and have also
associated LAB. Enterococci occupy a diverse range of been found on the teat surface and throughout the farm
environment (Vacheyrou et al., 2011; Braem et al., 2012). starter cultures but can be considered to be beneficial
Coryneform bacteria are generally regarded as being NSLAB due to their aromatic and flavour contributing end
important components of the surface of smear-ripened products (Afzal et al., 2010). These include malty aromas
cheese but can also be located in cheese cores (Duthoit as well as alcohol and fruity odours in cheese, although
et al., 2003). These bacteria can contribute to cheese some strains have been linked with sweat, faecal and rot-
flavour and aroma due to their ability to produce volatile ten-fruit-associated flavours (Marilley & Casey, 2004). The
sulphur compounds giving notes of garlic, onion and occurrence of Carnobacterium in dairy products is proba-
even cabbage to the cheese. These compounds result from bly underreported due to the frequent use of acetate-con-
the production of methanethiol, sulphides, thiols and taining media, such as MRS medium, as acetate inhibits
thioesters (Bloes Breton & Bergere, 1997). Sequencing of the growth of these microorganisms (Leisner et al., 2007).

Corynebacterium casei UCMA 3821, Corynebacterium vari- Bifidobacterium represents an important genus, which
able DSM 44702 and Chryseobacterium bovis DSM 20583 is generally regarded as a health-promoting genus and is
has revealed genes involved in iron acquisition and most commonly associated with the gastrointestinal tract
uptake, an important feature of cheese surface bacteria. of humans and animals (Lamendella et al., 2008). It is
Also present are genes involved in utilisation of alterna- also frequently detected in raw milk and fermented dairy
tive carbon and sulphur sources, amino acid metabolism products (Delcenserie et al., 2005), despite the fact that
and fatty acid degradation. The genetic repertoire of these many Bifidobacterium strains have stringent nutrient
strains also highlights their ability to catabolise lactate requirements and are generally thought to grow poorly
and propionate; to utilise external caseins; and to produce outside of the gut (Lamendella et al., 2008). In dairy
acetoin, butanediol and methanethiol, which are impor- products, the presence of bifidobacteria results in
tant with respect to the flavour of smear-ripened cheeses increased levels of lactate and acetate but does not influ-
(Monnet et al., 2010; Schr€ oder et al., 2011, 2012). ence sensory or textural properties (Dinakar & Mistry,
Arthrobacter spp. are commonly isolated from raw milk 1994). Finally, the significance of coagulase-negative
(Verdier-Metz et al., 2009; Masoud et al., 2012) and are staphylococci (CNS) with respect to dairy fermentations
thought to enter from the dairy facility as well as the teat has been the subject of much debate. The species typically
surface (Vacheyrou et al., 2011). While little is known isolated from milk include Staphylococcus equorum, Staph-
with respect to the influence of these bacteria on cheese ylococcus xylosus and Staphylococcus carnosus. These bacte-
development, they are an important microorganism on ria are salt tolerant and acid tolerant. Although no CNS
the surface of smear-ripened cheese where they contribute of dairy origin have been associated with food poisoning
to colour, flavour and textural development. Arthrobacter or human pathology, a few cases of nosocomial infection
arilaitensis is perhaps the most important cheese-associ- caused by Staphylococcus caprae, Staphylococcus capitis or
ated species, and genome sequencing of strain Re117 Staphylococcus sciuri have been reported in patients with
reveals the presence of several genes that reflect its adap- depressed immune systems (Irlinger, 2008). A recent
tation to growth in dairy/cheese environments including study of isolates from milk and cheese revealed 17 CNS
salt tolerance, galactose metabolism and enzymes for species. Ten of these contained transferable antibiotic
catabolism of fatty acids, amino acids and lactate. As with resistance genes, and one-third exhibited haemolytic
other smear-associated bacteria, Arthrobacter spp. possess activity (Ruaro et al., 2012), indicating that the safety of
a gene set involved in iron transport (Monnet et al., these bacteria must continue to be assessed.
2010). Similarly, Brevibacterium, which is commonly
detected in raw milk (Desmasures & Gueguen, 1997;
Gram-negative subpopulations
Lafarge et al., 2004; Raats et al., 2011; Masoud et al.,
2012), is known for its association with characteristic Gram-negative bacteria are very common in dairy foods.
taste, aroma and colour of smear-ripened cheeses. They can reach high levels (106–107 CFU g 1) in cheeses
Brevibacterium linens is particularly important in this and usually consist of a diverse number of species.
regard (Irlinger & Mounier, 2009). Another genus detected Although Gram-negative bacteria are regularly considered
in raw milk, Carnobacterium, consists of 11 species (www. as indicators of poor hygiene and may constitute a health
bacterio.cict.fr; Cailliez-Grimal et al., 2005). The risk if pathogenic species are present, some may play roles
Carnobacterium species most frequently isolated from in dairy fermentations by contributing positively or nega-
dairy environments is Carnobacterium maltaromaticum. tively to the sensory quality of dairy products (Delbes-
Carnobacterium maltaromaticum was first isolated from Paus et al., 2011, 2012). These issues are addressed in
milk in 1974 and was originally named Lactobacillus malt- greater depth in this section.
aromaticus (Mora et al., 2003). Carnobacteria are slow The presence of high numbers of Gram-negative bacte-
acidifiers and therefore are not suitable for the use as ria in milk has been noted in situations where hygiene
standards are low and generally reflect poor udder prepa- animal, as well as the weather, feeding and season (Callon
ration, poor sanitation or deficiencies with respect to the et al., 2007; Vacheyrou et al., 2011). As with bacteria, the
hygiene of equipment. In one instance, milk sampled at extent of the fungal population in raw milk and dairy
the farm, at milk collection and at milk transportation products is often underestimated. However, the develop-
was found to be contaminated with E. coli (29.6%), Pseu- ment of culture-independent DNA-based methods, such as
domonas aeruginosa (18.5%) and Klebsiella pneumoniae those targeting the internal transcribed spacer (ITS) region
(16.7%) and, to a lesser extent, Enterobacter aerogenes, of fungi, has addressed this issue (Callon et al., 2006;
Alcaligenes faecalis, Proteus mirabilis and Citrobacter freun- Alessandria et al., 2010). This ITS region of fungi is partic-
dii. It was noted that no Gram-negative bacteria were ularly useful because of its high copy number, phylogenetic
isolated from pasteurised milk samples (Garedew et al., utility and the availability of universal primers to generate

2012). In a study investigating the presence of Gram- PCR amplicons. This and other culture-independent
negative bacteria in different cheeses produced in France, approaches, such as denaturing high-performance liquid
173 isolates were isolated. Nearly half of all isolates were chromatography, have led to the detection of fungi, for
representatives of Enterobacteriaceae. Overall, 26 different example, Torrubiella and Malassezia, which had not previ-
genera were present. The most frequent isolates included ously been detected in milk (Delavenne et al., 2011). While
Proteus, Psychrobacter, Halomonas, Serratia and Pseudomo- a relatively small number of yeast species occur in raw
nas, representing almost 54% of the total isolates. Milk milk, they are persistent and can be detected at relatively
and cheese core samples also contained Chryseobacterium, high levels, that is, 102–104 CFU mL 1 (Lagneau et al.,
Enterobacter and Stenotrophomonas, while surface samples 1996). Yeasts species that have been detected in raw milk
were dominated by Proteus, Psychrobacter, Halomonas and include Kluyveromyces marxianus, Kluyveromyces lactis,
Serratia (Coton et al., 2012). When a model cheese sys- Rhodotorula mucilaginosa, Debaryomyces hansenii, Geotri-
tem was employed to assess the consequences of the pres- chum candidum, Geotrichum catenulate, Pichia fermentans,
ence of some of these Gram-negative bacteria, it was Candida sake, Candida parapsilosis, Candida inconspicua,
established that the majority had little influence on col- Trichosporon cutaneum, Trichosporon lactis, Cryptococcus
our, odour and volatile compounds (Delbes-Paus et al., curvatus, Cryptococcus carnescens and Cryptococcus victoriae
2011). However, Hafnia alvei did contribute to the pro- (Delavenne et al., 2011). Yeasts can play a major role in
duction of volatile compounds and of volatile sulphur dairy fermentations due to a number of their physiological
compounds in particular. Furthermore, Psychrobacter celer and biochemical characteristics, including the ability to
was found to flourish within the cheese surface smear utilise lactose or galactose, for example, in D. hansenii
during ripening, contributing to the production of vola- (Van den Tempel & Jakobsen, 2000); high proteolytic or
tile compounds such as aldehydes, ketones and sulphur lipolytic activity, for example, in Yarrowia lipolytica and
compounds (Delbes-Paus et al., 2011; Irlinger et al., G. candidum (Sacristan et al., 2012); and the ability to
2012). Another Gram-negative species, Proteus vulgaris grow at low temperatures and to tolerate high salt concen-
1 M10, has been shown to produce high concentrations trations. In cheese, yeasts secrete enzymes that play a key
of flavour compounds, particularly branched-chain alco- role in texture and produce various aromas during ripen-
hols during ripening (Deetae et al., 2009). These studies ing. Kluyveromyces marxianus is of particular interest due
reveal the high biodiversity of Gram-negative bacteria to its fast growth rate, thermotolerance, the ability to
among raw milk and dairy products and suggest that they assimilate a wide range of sugars, the secretion of lytic
may play a role in dairy fermentations. However, the fact enzymes and the production of ethanol by fermentation
that one of the studies referred to above revealed that (Lane & Morrissey, 2010). Moulds are typically present at
c. 50% of the Gram-negative strains isolated were resis- lower levels than yeasts (Arora et al., 1991). Moulds have
tant to several antibiotics is a particular cause for concern the ability to enhance the flavour and aroma and modify
(Delbes-Paus et al., 2011). Given this observation, and the texture and structure of milk-derived products as a
the association of particular Gram-negative bacteria with consequence of bringing about extensive proteolysis and
milk spoilage or disease, the presence of Gram-negative lipolysis. The mould genera that are most commonly
bacteria in these products will in general continue to be detected in raw milk include Penicillium, Geotrichum,
regarded as undesirable. Aspergillus, Mucor and Fusarium (Lavoie et al., 2012).
At the species level, Fusarium merismoides, Penicillium
glabrum, Penicillium roqueforti, Aspergillus fumigatus, Engy-
Fungal populations
odontium album, as well as species of Cladosporium and
Yeasts and moulds can also be important microbial popu- Torrubiella are common (Delavenne et al., 2011).
lations within raw milk. The fungal composition of raw In recent years, genome sequencing has been particu-
milk can be influenced by the physiological state of the larly useful with respect to enhancing our understanding
of milkborne microorganisms by, for example, highlight- increases in the number of Listeria innocua, L. monocytog-
ing the phenomenon of genomic decay and adaptation to enes, L. fermentum, Staphylococcus epidermidis, P. fluores-
the milk environment, revealing potential sources of evo- cens, E. faecium, Enterococcus hirae, E. durans, Leuconostoc
lutionary origin and identifying genes that contribute to carnosum, S. dysgalactiae, H. alvei, Serratia marcescens,
flavour development in dairy products. This information K. pneumoniae, Kocuria rosea, propionic acid bacteria and
can allow industry to develop novel starter cultures, Aeromonas (Lafarge et al., 2004). Notably, some of the
determine how best to enhance the efficacy of existing latter would be typically regarded as thermophilic micro-
strains and provide re-assurances in terms of the ‘GRAS’ organisms. A similar study, but carried out over a 48-h
status of some of these technologically important micro- period, specifically highlighted increases in Pseudomonas
organisms. and Acinetobacter spp. (Raats et al., 2011). Newly identi-

fied psychrotrophs, such as Chryseobacterium (Hantsis-
Zacharov & Halpern, 2007; Hantsis-Zacharov et al.,
Impact of storage conditions and
2008a, b) and Epilithonimonas spp. (Shaked et al., 2009),
downstream treatments on the
have also been detected in raw milk. However, their
microbiology of raw milk
involvement in milk spoilage is unclear. A number of
psychrotrophic spore-forming species have also been
Cold storage
identified and will be discussed in the next section. Yeasts
It is important to understand the changes that can occur and moulds have also been associated with milk spoilage
in the microbiology of raw milk during its storage and as (Agarwal et al., 2012).
a consequence of subsequent treatments. Milk is typically
stored at refrigeration temperatures that reduce the
Pasteurisation
growth of most bacteria, with the exception of psychro-
tolerant microorganisms that can proliferate under these Pasteurisation of raw milk is carried out to reduce the
conditions and become a major cause of milk spoilage microbial load of milk and, in particular, to limit the num-
(Eddy, 1960; Morita, 1975; De Jonghe et al., 2011). This ber of spoilage microorganisms and to prevent foodborne
is primarily a consequence of the production of extracel- disease. However, this process also reduces the number of
lular enzymes, with lipases and proteases being most microorganisms that would typically contribute to desir-
important. These lipases degrade milk fat causing rancid- able sensory properties associated with raw milk cheeses. In
ity, while proteases degrade casein producing a grey these instances, starter cultures that are known to generate
colour and bitter off-flavours (De Jonghe et al., 2011). desirable flavours and aromas, as discussed above, are
Investigations into seasonal variations of microbial added to the milk postpasteurisation. The typical milk pas-
growth in raw milk have, unsurprisingly, established that teurisation treatment is a ‘high-temperature short-time’
psychrotolerant bacteria exhibit better growth and prote- (HTST) approach involving heating to 72 °C for 15 s.
ase production in winter milk rather than in summer Some countries have increased the exposure temperature
milk (Marchand et al., 2008). Pseudomonas spp., which and/or time (Martin et al., 2011). While this can help to
are commonly found in raw milk, are the most common further reduce bacterial counts (Fromm & Boor, 2004) and
cause of milk spoilage (Ercolini et al., 2009). The to eliminate microorganisms of concern including Myco-
Pseudomonas species most commonly detected in milk bacterium avium ssp. paratuberculosis (MAP; Grant et al.,
and cheeses are Pseudomonas fluorescens, Pseudomonas 2002) and L. monocytogenes (Doyle et al., 1987), there have
gessardii, Pseudomonas fragi and Pseudomonas lundensis also been some suggestions that this approach can encour-
(Mallet et al., 2012). These bacteria can become the pre- age the activation of spores, which may be dormant in milk
dominant microorganisms in raw milk stored at low tem- (Ranieri et al., 2009). The heat treatment of milk typically
peratures, constituting up to 70–90% of the microbial reduces psychrotrophic and mesophilic populations, leav-
population (Sorhaug & Stepaniak, 1997). Many other ing two main groups to consider thereafter, that is, ther-
psychrotolerant microorganisms are present in milk but moduric microorganisms and bacteria introduced through
are generally less important than Pseudomonas with postpasteurisation contamination. Following pasteurisa-
respect to milk spoilage. In one study, these were identi- tion, some microorganisms may enter into a ‘viable but
fied as being strains of Acinetobacter, Microbacterium, nonculturable’ state, meaning that they may be underesti-
Aeromonas, Enterobacter, Flavobacterium, Corynebacte- mated by traditional culture methods (Bartoszcze, 2009).
rium, Clostridium, Bacillus, Staphylococcus and some LAB The findings of a recent culture-independent study con-
(Hantsis-Zacharov & Halpern, 2007). Another study, ducted by our group are consistent with this theory, reveal-
assessing the overall impact of refrigeration (24 h) on the ing a more diverse bacterial population in pasteurised milk
microbial content of raw milk, particularly noted than expected (Quigley et al., 2013).
When one considers thermoduric bacteria, it is particu- dairy phages are able to survive the pasteurisation of milk
larly important to keep the issue of spore-forming micro- (Suarez & Reinheimer, 2002; Abedon, 2009). Due to the
organisms in mind. These bacteria may enter the milk severe economic loss that phages can cause, constant
chain from soil, silage and bedding material and, signifi- monitoring of the environment is required. Traditionally,
cantly, are resistant to pasteurisation. Spore formers such standard microbiological methods were employed. How-
as Clostridium sporogenes, Clostridium butyricum and Clos- ever, molecular methods based on PCR and quantitative
tridium tyrobutyricum have the potential to survive and PCR allow the rapid detection and classification of phages
grow at refrigeration temperature, as well as the potential from different dairy matrices (Binetti et al., 2008). While
to utilise carbohydrates, proteins and lactate from milk many dairy bacteria can be infected by phage, there are a
(Driehuis, 2013). Indeed, clostridia have been identified number of industrially important strains that have inher-

in raw milk quite frequently (Herman et al., 1995; Lopez- ent resistance to phage. Genome sequencing studies have
Enriquez et al., 2007; Cremonesi et al., 2012) and can revealed that a number of strains, such as L. helveticus
contribute to the spoilage of subsequent cheese products DPC 4571, P. freudenreichii CIRM-BIA1T and some
by causing a late blowing defect, which is particularly S. thermophilus strains to name but a few, possess features
associated with C. tyrobutyricum, leading to off-flavours to help these bacteria withstand phage attack.
and textural defects in cheese (Cocolin et al., 2004; Le
Bourhis et al., 2005). Culture-independent DGGE/TTGE
Biopreservative potential of raw milk
has identified C. tyrobutyricum, but also C. sporogenes,
microorganisms
C. butyricum and Clostridium beijerinckii as possible
causes. Other spore-forming contaminants of milk The production of an antimicrobial can be regarded as a
include B. cereus, Bacillus sporothermodurans and Geoba- beneficial probiotic trait. The diverse populations in raw
cillus stearothermophilus. Bacillus cereus is a major spoilage milk produce many antimicrobials including bacteriocins,
organism of pasteurised milk and milk products stored at antifungals, organic acids and hydrogen peroxide. As
refrigeration temperature, causing off-flavour and cur- noted above, raw milk also contains phages. These might
dling. This bacterium is also a concern for food safety as also be regarded as biopreservative agents that could be
it can produce different types of toxins and is a potential used to extend the shelf life and safety of fermented and
food-poisoning agent (Driehuis, 2013). In the EU in other foods (Stiles, 1996).
2010, 3.8% of all milk samples tested were positive for Bacteriocins are antimicrobial peptides or proteins pro-
Bacillus toxin (European Food Safety Authority, 2012). duced by bacteria and are typically active against closely
related species, but can exhibit activity across broad gen-
era. Bacteriocin producers are naturally immune to their
Bacteriophage
own bacteriocins (Cotter et al., 2005). The most recent
Lytic bacteriophages are other spoilage agents that are classification groups LAB bacteriocins into two classes;
naturally present in raw milk. These viruses infect bacte- class I are post-translationally modified bacteriocins, and
ria and, after intracellular replication, lyse their host cells class II are unmodified bacteriocins (Rea et al., 2011).
(Marc o et al., 2012). Phages can survive at low tempera- Many LAB isolated from raw milk produce putative
ture; resist a variety of different treatments; and nega- bacteriocin-like compounds and exhibit activity against
tively affect the quality, safety and value of dairy products L. monocytogenes, S. aureus, C. tyrobutyricum, C. sporogenes,
(cheese in particular). Phages that target important starter E. faecalis, E. faecium and E. durans (Alegrıa et al., 2010;
or adjunct species, such as Lactococcus lactis, S. thermo- Ortolani et al., 2010; Perin et al., 2012). Some potential
philus, L. helveticus and L. delbrueckii, have long been raw milk-derived bacteriocins have been characterised
associated with causing a delay or disruption of fermenta- in-depth. Lactococcus lactis strains produce the best
tion processes, resulting in slow acidification, undesirable characterised bacteriocin, nisin. Nisin is well known due
organoleptic properties or complete loss of batches to its use in biopreservation throughout the world because
(Emond & Moineay, 2007). Raw milk is the most promi- of its wide spectrum of activity (Delves-Brougthon,
nent source of phages within the dairy environment, with 1990). Lactococcus lactis strains isolated from raw milk
concentrations ranging from 101 to 104 phage mL 1 and raw milk products are capable of producing nisin
(Madera et al., 2004). Phages can also gain access to the with activity against L. monocytogenes as well as other
dairy environment by aerosols, personnel, equipment, pathogens including E. coli and Staphylococcus spp.
work surfaces and dairy byproducts (Verreault et al., (Bravo et al., 2009; Alegrıa et al., 2010; Ortolani et al.,
2011). Thus, attaining a phage-free environment is not a 2010; Cosentino et al., 2012; Perin et al., 2012). A huge
realistic goal. While the phage concentration is higher in variety of other bacteriocins are produced by L. garvieae
raw milk products, it has also been reported that many (Villani et al., 2001; Florez et al., 2012), S. thermophilus
(Gul et al., 2012), S. macedonicus (Georgalaki et al., 2002, throughout different countries, found that the percentage
2013), enterococci and Leuconostoc spp. (Mathieu et al., of tanks containing the different pathogens varied greatly.
1993; Casaus et al., 1997; Giraffa & Carminati, 1997; The occurrence of Salmonella ranged between 0% and
Achemchem et al., 2006; Izquierdo et al., 2009; Mirhosse- 11.8% of milk samples in the USA/Canada and between
ini et al., 2010) and are active against many spoilage and 1.4% and 4% of samples in Asia, while the percentage of
pathogenic microorganisms. In addition to contributing milk samples that was positive for the presence of Listeria
to the control of pathogens and spoilage microorganisms ranged from 0% to 7% in USA/Canada and from 0% to
in raw milk and resultant products, these bacteriocins can 1.9% in Asia. Campylobacter jejuni was detected at 2%
also be employed, through the addition of producing LAB, and 9.2% of milk samples from the USA, while the
fermentates or semi-purified preservatives, to enhance the percentage of bulk tank milk worldwide that was positive

safety of other foods (Cotter et al., 2005; Deegan et al., for shiga-toxin-producing E. coli varied from 0% to
2006). 33.5%. Mycobacterium avium ssp. paratuberculosis was
Raw milk LAB isolates also produce organic acids, present in European samples at frequencies of 1.6–19.7%
hydrogen peroxide and diacetyl (Stiles, 1996). These com- and in Asia at 8.6–23%. Brucella was present in milk in
pounds can inhibit many potential pathogens and food Africa at 0–10%, increasing with increasing farming inten-
spoilage microorganisms (Batdorj et al., 2007). Some sity (Oliver & Murinda, 2011).
strains of L. acidophilus, L. casei, L. helveticus, L. bulgari- Regardless of the frequency at which they are present,
cus, S. thermophilus and Lactococcus lactis are also able to these pathogens can impact, in some instances severely,
transform hippuric acid, which is naturally present in on health. Staphylococcus aureus can be transferred to
milk, into benzoic acid, thereby providing another natural milk through the teat canal, equipment, the environment
preservative in milk products (Garmeine et al., 2010). or human handling (Rosengren et al., 2010) and cause
Finally, common food-spoiling moulds and, to a lesser illness through the production of heat-stable enterotoxins,
extent, yeasts can be inhibited by LAB metabolites from which can withstand pasteurisation (Balaban & Rasooly,
strains of L. casei, L. reuteri, L. plantarum and L. buchneri 2000). Staphylococcal toxins were detected in 18.4% of
and strains of dairy propionibacteria (Lind et al., 2005, cheeses assessed across the EU in 2010 (European Food
2007; Voulgari et al., 2010; Delavenne et al., 2011). Safety Authority, 2012). Coxiella burnetii, the causative
agent of Q fever, can infect many animal species, and it is
thought that its association with cows, sheep and goats is
Human health associations
the main source of human infection. The infection may
be acute, presenting flu-like symptoms that are self-limit-
Pathogenic bacteria associated with raw milk
ing or chronic, leading to endocarditis and hepatitis
Milk and dairy products are important staples of a healthy (Maurin & Raoult, 1999). Coxiella burnetii is shed by the
diet. However, if pathogenic microorganisms are not animal host through birth products, vaginal mucus,
removed by pasteurisation, consumption of these products semen, faeces, urine and milk (Guatteo et al., 2006).
can represent a serious health risk. As mentioned above, While this bacterium can persist in dairy cattle popula-
these pathogens can originate from the mammary gland tions (Astobiza et al., 2012; Tilburg et al., 2012), the con-
or associated lymph nodes of cows suffering from systemic sumption of raw or insufficiently pasteurised milk is
diseases or infections (Oliver & Murinda, 2011; Hunt rarely identified as a source of Q fever (Guatteo et al.,
et al., 2012) or from equipment, raw milk tankers and 2006). Another zoonotic bacterium of health concern is
personnel (Rosengren et al., 2010; Teh et al., 2011; Giaco- Mycobacterium bovis. This bacterium causes the disease
metti et al., 2012). Ingestion of these microorganisms can bovine tuberculosis in animals, with symptoms including
lead to illnesses of varying severity. Typical symptoms can fever, weakness, emaciation, inappetence and respiratory
include fever, nausea, vomiting, diarrhoea and abdominal distress, and can lead to severe economic loss yearly
pains; in extreme cases, death can occur (Langer et al., (Thoen et al., 2006). This pathogen can also spread to
2012). Indeed, food poisoning from consumption of raw humans through the ingestion of raw milk causing zoo-
milk and such products over a period of 13 years (1993– notic tuberculosis, which is indistinguishable from human
2006) in the United States resulted in 1571 reported inci- tuberculosis (Thoen et al., 2006). Typically, this concern
dences with 202 hospitalisations and 2 deaths. The main is removed with pasteurisation, but remains a problem in
cause of illness was consumption of raw milk products instances where raw milk is still consumed daily (Coker
contaminated with Salmonella spp., Listeria spp., E. coli, et al., 2006; De la Rua-Domenech, 2006). Mycobacterium
Campylobacter spp., Brucella spp. or Shigella spp. (Langer avium ssp. paratuberculosis (MAP) is the causative agent
et al., 2012). A recent review, which examined multiple of paratuberculosis or Johne’s disease, which primarily
reports of milkborne pathogen detection in bulk tanks infects domestic animals. MAP survives and multiplies in
the intestinal tract mucosa, where it causes both a Healthy adults are typically not at risk although they may
decrease in the absorption of nutrients and chronic diar- experience flu-like or gastrointestinal symptoms (Liatsos
rhoea with consequent ‘wasting away’ of the animals. et al., 2012). There is evidence to suggest that raw milk
Animals may harbour this bacterium for a long period purchased from retailers represents a greater risk of Liste-
before symptoms arise, making them dangerous vectors ria-associated illness than milk obtained directly from
of infection. MAP can be shed into the external environ- milk tanks on farms, most likely as a consequence of the
ment in animal faeces or milk. Recently, there has been growth of the pathogen over the extended storage period
increased concern with the association of MAP and Cro- (Latorre et al., 2011). Brucella spp. primarily cause disease
hn’s disease in humans, an inflammatory bowel disease, in animals and from there are thought to enter into the
whose symptoms include abdominal pain, diarrhoea, milk supply. On consumption, these pathogens can pro-

vomiting and weight loss. High prevalence of MAP has voke brucellosis, which leads to fever, abdominal pain,
been reported in raw milk in developed countries (Argen- headaches and personality changes (Roop et al., 2004).
tina 8.3%, Czech Republic 2%, Ireland 0.3%, UK 6.9%, Like Listeria, Brucella can survive and multiply in milk
USA 0–28.6%; Slana et al., 2008). Current commercial (Falenski et al., 2011), also following contamination after
pasteurisation standards may reduce the number of viable pasteurisation (Oliver et al., 2005). Due to the severe nat-
MAP, but do not ensure destruction (Gao et al., 2002). ure of many of the illnesses caused by milkborne patho-
However, the link between Crohn’s disease and MAP still gens, it is important to test rigorously for their presence.
remains controversial and unclear (Chiodini et al., 2012). While traditional methods can be laborious and time-
Another, relatively new, pathogen of concern to the dairy consuming, newer culture-independent methods have
industry is Shiga-toxin-producing E. coli (STEC). been investigated, with quantitative PCR being particu-
Although cows are the main reservoir for STEC, other larly rapid and sensitive (Quigley et al., 2011). However,
domestic animals including goats and sheep can also har- these have yet to be implemented on a large scale by the
bour this bacterium in their gastrointestinal tracts with- dairy industry. While detection is important, the practices
out any symptoms of disease and shed them from their that prevent or limit the presence of pathogens are more
faeces. If hygiene standards are not sufficiently high, milk crucial. Thus, it is important to implement a hygiene sys-
can become contaminated during milking or processing. tem that begins at the farm level and includes a focus on
There are nine virulence genes associated with STEC cow health and hygiene, equipment cleanliness, overall
strains. Two toxin genes, stx1 and stx2, appear to be asso- farm and personnel sanitation, correct storage and subse-
ciated with bovine dairy products, while stx1c and stx2b quent processing of milk.
are more frequently associated with strains from sheep Mycotoxins, that is, low molecular weight compounds
and goat milk (Martin & Beutin, 2011). produced as secondary metabolites by filamentous fungi,
Of the milkborne pathogens, L. monocytogenes, Yersinia can lead to illness in humans with symptoms such as
enterocolitica and Brucella spp. are a particular cause for nausea, vomiting, diarrhoea and headache (Creppy,
concern as they are able to survive and multiply at refrig- 2002), and some mycotoxins have carcinogenic potential
eration temperatures and may cause severe diseases. Yer- (Murphy et al., 2006). The most important genera of
sinia enterocolitica is a major cause of acute gastroenteritis food mycotoxigenic fungi are Aspergillus, Alternaria, Fusa-
(Schiemann & Toma, 1978). The symptoms of illness can rium and Penicillium. Examples of mycotoxins of greatest
include diarrhoea, abdominal pain and fever and may public health and agro-economic significance include afla-
mimic appendicitis, occasionally leading to misdiagnosis toxin, trichothecenes, zearalenone, fumonisins and ochra-
(Ackers et al., 2000). Although pasteurisation will kill toxin. After their intake by cows, mycotoxins follow a
Y. enterocolitica, if insufficient pasteurisation or recon- typical pharmacokinetic cascade of uptake from the gas-
tamination occurs, the bacterium can multiply under trointestinal test to the blood, internal distribution,
refrigeration temperatures (Schiemann & Toma, 1978). metabolism, storage, remobilisation and excretion. The
Similarly, during dairy product manufacture, where rumen has an important function in the metabolism of
hygiene standards are poor, Y. enterocolitica can become mycotoxins with some mycotoxins being rapidly metabo-
prevalent (Harakeh et al., 2012). In 2010, Yersinia inci- lised to less toxic metabolites (e.g. ochratoxin); some are
dence in raw milk and low-heat-treated milk products transformed into equally toxic or more toxic metabolites
was low, with only two positive results reported within (e.g. zearalenone), while some are not transformed at all
the EU (European Food Safety Authority, 2012). Listeria (e.g. fumonisins). Aflatoxin B1 is transformed into afla-
monocytogenes, a common environmental isolate, causes toxin M1 in the liver of ruminants. While M1 is less
the human disease listeriosis, which targets highly suscep- mutagenic and genotoxic than B1, the cytotoxicity of M1
tible individuals, including pregnant, immunocompro- and B1 is similar. Notably, aflatoxin B1 is the only myco-
mised or elderly people, and has a high fatality rate. toxin with significant carry-over into milk. Between 1%
and 6% is excreted in milk as aflatoxin M1. Sixty countries drug-resistant strains are a serious concern (Dijkshoorn,
now have regulations with respect to the presence of afla- 2013). Other antibiotic-resistant raw milk isolates include
toxin M1 in milk, with limits of 0.05–0.5 lg kg 1; the EU Lactococcus lactis displaying resistance to tetracycline, clin-
has a legal limit of 50 ng L 1 (Driehuis, 2013). Testing is damycin and erythromycin and L. garvieae exhibiting
important given that one study has shown that a high resistance to tetracycline, streptomycin and quinupristin–
percentage (83.2%) of raw milk samples in Portugal were dalfopristin (Walther et al., 2008).
positive for aflatoxins (Martins & Martins, 2000) and that The use of antibiotics to treat animals that are in the
the levels of aflatoxin B1 may frequently exceed recom- food chain can obviously compound this issue by select-
mended limits (Nordkvist & Hoorfar, 2012). Finally, ing for the development of antibiotic resistance among
yeasts, and especially Candida species, can be opportunis- food microorganisms (in particular in cows’ milk) and by

tic pathogens, causing infections in immunocompromised exposing consumers to antibiotic residues in milk and
patients. Debaryomyces hansenii and Y. lipolytica may also other dairy foods (Doyle et al., 2013). The use of antibi-
be emerging dairy pathogens. These microorganisms otics to treat mastitis during lactation is common, as
cause rare infections in immunocompromised patients, between 2% and 55% of cows encounter a mastitis infec-
which are generally mild and either self-limiting or easily tion during this period (Kelton et al., 1998). Notably,
treated (Jacques & Casaregola, 2008). bacterial strains associated with bovine mastitis, including
many S. aureus isolates, have demonstrated resistance to
antibiotics such as penicillins, oxytetracycline, streptomy-
Antibiotic residues and antibiotic-resistant
cin and/or gentamicin (Thaker et al., 2013). These prob-
bacteria in milk
lems can be limited through the withholding of milk
Antibiotics have been employed to treat bacterial diseases from sale in situations where a cow has mastitis and is
over the past 70 years. The greatest threat to the success- being treated with antibiotics or during a compulsory
ful application of antibiotics has been the development of withdrawal period after antibiotic treatment. Safeguards,
resistance, particularly in pathogenic bacteria. Resistance such as those introduced by Codex Alimentarius 2009
can be intrinsic or acquired. Intrinsic resistance is a natu- and European Union Council Regulation 37/2010/EC,
ral characteristic of a microorganism that allows it to require the monitoring of milk and provide limits with
grow in the presence of the corresponding antibiotic. respect to the concentration of antibiotic residues that are
Acquired resistance results either from spontaneous tolerated in milk for commercial use.
mutation in the bacterial genome or from the acquisition
of genes encoding resistance through transduction, conju-
Health-promoting microorganisms
gation or transformation (Davies, 1997).
Although lactococci, enterococci and lactobacilli are Some raw milk isolates can have health-promoting abili-
intrinsically resistant to some antibiotics (Mathur & Singh, ties; for millennia, it has been suggested that fermented
2005), the strains of these that are found in foods are typi- milk can cure some disorders of the digestive system, and
cally quite sensitive to clinically important antibiotics such biblical scriptures highlight the use of milk to treat body
as ampicillin, penicillin, gentamicin and vancomycin ailments (Lourens-Hattingh & Viljoen, 2001). In 1907, the
(Franz et al., 1999; Mannu et al., 2003; Can zek Majhenic Russian scientist Elie Metchnikoff pointed out the benefits
et al., 2005; Herreros et al., 2005; Mathur & Singh, 2005). of consuming a diet of fermented milk (Rasic & Kurmann,
Furthermore, Leuconostoc strains are generally sensitive to 1983). Health-promoting bacteria isolated from these
antibiotics (Swenson et al., 1990). However, it is still beverages and other sources are commonly referred to as
important to assess the frequency with which antibiotic- ‘probiotics’, that is, ‘live bacteria which when adminis-
resistant isolates occur in milk. A recent study determined tered in adequate amounts confer a health benefit on the
that psychrotrophs, including P. fluorescens, P. aeruginosa, host’ (FAO/WHO, 2002). The selection of such bacteria
Stenotrophomonas maltophilia, Burkholderia species, as well for commercial probiotic application relies on criteria
as a number of unidentified psychrotrophs, isolated from relating to safety, technological and digestive stress sur-
milk harbour resistance to several b-lactam and non-b- vival; intestinal cell adhesion; and human origin. The lat-
lactam antibiotics. This trait appears to increase in occur- ter two criteria are controversial, and it is now recognised
rence through the cold chain transportation of raw milk that adhering to these criteria should not be mandatory,
(Munsch-Alatossava & Alatossava, 2007). Bacteria of the although may be desirable in certain instances. Notably,
genus Acinetobacter isolated from raw milk have also many raw milk isolates have desirable probiotic traits.
exhibited antibiotic resistance (Gurung et al., 2013). These These include the ability to survive bile juice, to tolerate
bacteria are widespread in nature, including soil and gastric acid conditions and to adhere to intestinal cells
water, and are opportunistic pathogens in humans; multi- (Jamaly et al., 2011). Probiotic lactobacilli typically inhibit
pathogenic organisms, reduce lactose intolerance, increase Raw milk and the raw milk microbiota have also been
the immune response and often are gastrointestinal iso- the focus of attention with respect to alleviating allergy.
lates (Kopp-Hoolihan, 2001; Maragkoudakis et al., 2006). Allergy to cows’ milk affects 2.5% of children below
However, there are a number of dairy lactobacilli isolates, 3 years of age due to the presence of caseins and b-lacto-
which also have demonstrated efficiency as probiotic globulins (Cocco et al., 2003; Gaudin et al., 2008). The
strains (Maragkoudakis et al., 2006). Other milk and dairy bacterial fermentation of milk proteins, particularly by
isolates that exhibit probiotic properties include strains of highly proteolytic Lactobacillus populations, results in a
Lactococcus lactis as well as a variety of Pediococcus, Leuco- reduction in the allergenic properties of cows’ milk (El-
nostoc, Enterococcus and Streptococcus isolates (Kim et al., Ghaish et al., 2011). Others have suggested a link between
2006; Premalatha & Dhasarathan, 2011; Espeche et al., farm living, including the consumption of raw milk and

2012; Floros et al., 2012; Forghani et al., 2012). Strains of raw milk microorganisms, and protection against the
P. freudenreichii, and to a lesser extent P. acidipropionici, development of asthma and atopy later in life (Debarry
have begun to attract attention as potential probiotics as a et al., 2007; Ege et al., 2012). If confirmed, further inves-
consequence of studies revealing an ability, either alone or tigations will be required to determine whether overall
in combination with other probiotics, to reduce pathogen microbial load or specific components of the microbial
adhesion to mucus (Collado et al., 2008), increase bifido- population are responsible.
bacteria counts in the gut, aid in restoring a healthy gut
microbiota, improve bowel movement, alleviate inflamma-
Conclusion
tory disorders and reduce allergy development in infants
(Jan et al., 2002; Cousin et al., 2012). Finally and from a The microbial community within raw milk is complex.
fungal perspective, the dairy yeast Pichia fermentans has The dominant, and subdominant, microorganisms pres-
demonstrated some probiotic potential, and it has been ent in raw milk can have a variety of influences on the
suggested that together with strains of Pichia kudriavzevii flavour, taste and texture of raw milk-derived products
and Y. lipolytica, P. fermentans could serve as probiotics (Fig. 1). A number of these microorganisms also have
that assimilate cholesterol (Chen et al., 2010). Regardless the potential to contribute to health through the
of the specific microorganism in question, dairy products production of antimicrobials or possessing other probi-
are an excellent vehicle for probiotics, regardless of their otic-associated traits. Through modern genomics-based
source, due to their buffering capacity and fat content, analysis, it has been established that many of these
which can help protect the bacteria during gastric transit. microorganisms have become adapted to milk niches
There has been quite a degree of focus on the use of from various sources, including plant and gut environ-
dairy microorganisms to control hypertension. The ren- ments, through genomic evolution and gene gain and/or
nin–angiotensin–aldosterone system is a key factor in the loss. Despite the beneficial impact of many milk-associ-
maintenance of arterial blood pressure. One of the main ated microorganisms from a flavour, technological or
components of this system is angiotensin-converting health-related perspective, it is clear that there can be
enzyme (ACE). As ACE plays an important role in the significant risks associated with the consumption of raw
regulation of arterial blood pressure, inhibition of this milk and raw milk-derived products or, more specifi-
enzyme can generate an antihypertensive effect. ACE- cally, of the pathogens that can be found therein. While
inhibitory drugs are commonly used to control arterial many of these microorganisms gain entry to the milk
blood pressure. Raw cows’ milk can be a source of anti- from equipment and/or personnel, zoonotic pathogens
hypertensive activity (Meisel, 2005). LAB that release bio- can also be introduced into milk from unhealthy ani-
active peptides with this activity include strains of mals. As a consequence of this risk, pasteurisation or
E. faecalis, Lactococcus lactis ssp. cremoris, L. helveticus, other treatments are employed to remove disease-causing
L. fermentum, L. rhamnosus, L. paracasei and L. acidophilus microorganisms. In the food industry, the negative
(Muguerza et al., 2006). The antihypertensive properties impact of removing LAB and other bacteria on subse-
of these microorganisms are being investigated and quent food fermentations has been addressed for some
exploited by industry with a view to producing health-pro- time through their re-introduction in the form of starter
moting drinks. Indeed, L. helveticus is currently used in and adjunct cultures. Similarly, once established defini-
the production of fermented drinks such as Evolus (Valio tively, it may be possible to restore the benefits associ-
Ltd., Valio, Finland) and Calpis (Calpis Food Industry Co. ated with the consumption of raw milk and specific
Ltd., Tokyo, Japan), which have properties associated with microorganisms therein, through the re-introduction of
a reduction in blood pressure through the inhibition of these microorganisms after processing. Thus, the micro-
ACE as a consequence of the production of bioactive trip- bial composition of raw milk is likely to continue to be
eptides (Slattery et al., 2010). the focus of much attention into the future.
Arora DK, Mukerji KG & Marth EH (1991) Handbook of Applied

Acknowledgements Mycology: Volume 3: Foods and Feeds. Marcel Dekker Inc,
Lisa Quigley is funded by the Teagasc Walsh Fellowship New York.
Scheme. Dr Catherine Stanton is funded by EFRAIM – Asakuma S, Hatakeyama E, Urashima T et al. (2011)
Physiology of consumption of human milk oligosaccharides
mechanisms of early protective exposures on allergy
by infant gut-associated bifidobacteria. J Biol Chem 286:
development FP7-KBBE-2007-1.
34583–34592.
Astobiza I, Ruiz-Fons F, Pi~ nero A, Barandika J, Hurtado A &
Garcıa-Perez A (2012) Estimation of Coxiella burnetii
References
prevalence in dairy cattle in intensive systems by serological
Abedon ST (2009) Phage evolution and ecology. Adv Appl and molecular analyses of bulk-tank milk samples. J Dairy

Microbiol, 67: 1–45. Sci 95: 1632–1638.
Abeer AA, Gouda AS, Dardir H & Ibrahim A (2012) Balaban N & Rasooly A (2000) Staphylococcal enterotoxins.
Prevalence of some milk borne bacterial pathogens Int J Food Microbiol 61: 1–10.
threatening camel milk consumers in Egypt. Global Vet 8: Barbano D, Ma Y & Santos M (2006) Influence of raw milk
76–82. quality on fluid milk shelf life. J Dairy Sci 89: E15.
Achemchem F, Abrini J, Martinez-Bueno M, Valdivia E & Barile S, Devirgiliis C & Perozzi G (2012) Molecular
Maqueda M (2006) Control of Listeria monocytogenes in characterization of a novel mosaic tet(S/M) gene encoding
goat’s milk and goat’s Jben by the bacteriocinogenic tetracycline resistance in foodborne strains of Streptococcus
Enterococcus faecium F58 strain. J Food Prot 69: 2370–2376. bovis. Microbiology 158: 2353–2362.
Ackers ML, Schoenfeld S, Markman J et al. (2000) An Bartoszcze M (2009) Bacteria in the state of VBNC-a threat to
outbreak of Yersinia enterocolitica O: 8 infections associated human health. Med Weter 65: 228–231.
with pasteurized milk. J Infect Dis 181: 1834–1837. Batdorj B, Trinetta V, Dalgalarrondo M et al. (2007) Isolation,
Afzal MI, Jacquet T, Delaunay S, Borges F, Milliere JB, taxonomic identification and hydrogen peroxide production
Revol-Junelles AM & Cailliez-Grimal C (2010) by Lactobacillus delbrueckii subsp. lactis T31, isolated from
Carnobacterium maltaromaticum: identification, isolation Mongolian yoghurt: inhibitory activity on food-borne
tools, ecology and technological aspects in dairy products. pathogens. J Appl Microbiol 103: 584–593.
Food Microbiol 27: 573–579. Bayjanov JR, Wels M, Starrenburg M, van Hylckama Vlieg JE,
Agarwal A, Awasthi V, Dua A, Ganguly S, Garg V & Marwaha Siezen RJ & Molenaar D (2009) PanCGH: a
SS (2012) Microbiological profile of milk: impact of genotype-calling algorithm for pangenome CGH data.
household practices. Indian J Public Health 56: 88–94. Bioinformatics 25: 309–314.
Agyei D & Danquah MK (2012) Carbohydrate utilization Beecher C, Daly M, Ross RP, Flynn J, McCarthy TV & Giblin
affects Lactobacillus delbrueckii subsp lactis 313 L (2012) Characterization of the bovine innate immune
cell-enveloped-associated proteinase production. Biotechnol response in milk somatic cells following intramammary
Bioprocess Eng 17: 787–794. infection with Streptococcus dysgalactiae subsp.
Albesharat R, Ehrmann MA, Korakli M, Yazaji S & Vogel RF dysgalactiae.J Dairy Sci 95: 5720–5729.
(2011) Phenotypic and genotypic analyses of lactic acid Bernardeau M, Vernoux JP, Henri-Dubernet S & Gueguen M
bacteria in local fermented food, breast milk and faeces of (2008) Safety assessment of dairy microorganisms: the
mothers and their babies. Syst Appl Microbiol 34: 148–155. Lactobacillus genus. Int J Food Microbiol 126: 278–285.
Alegria A, Szczesny P, Mayo B, Bardowski J & Kowalczyk M Bhardwaj A, Kapila S, Mani J & Malik RK (2009) Comparison
(2012) Biodiversity in Oscypek, a traditional Polish cheese, of susceptibility to opsonic killing by in vitro human
determined by culture-dependent and -independent immune response of Enterococcus strains isolated from dairy
approaches. Appl Environ Microbiol 78: 1890–1898. products, clinical samples and probiotic preparation. Int J
Delgado S, Roces C, L
Alegrıa A, opez B & Mayo B (2010) Food Microbiol 128: 513–515.
Bacteriocins produced by wild Lactococcus lactis strains Binetti AG, Capra ML, Alvarez MA & Reinheimer JA (2008)
isolated from traditional, starter-free cheeses made of raw PCR method for detection and identification of Lactobacillus
milk. Int J Food Microbiol 143: 61–66. casei/paracasei bacteriophages in dairy products. Int J Food
Alessandria V, Dolci P, Rantsiou K, Pattono D, Dalmasso A, Microbiol 124: 147–153.
Civera T & Cocolin L (2010) Microbiota of the Planalto de Bloes Breton S & Bergere J (1997) Volatile sulfur compounds
Bolona: an artisanal cheese produced in uncommon produced by Micrococcaceae and coryneform bacteria
environmental conditions in the Cape Verde Islands. World isolated from cheeses [methanethiol]. Lait 77: 543–559.
J Microbiol Biotechnol 26: 2211–2221. Bolotin A, Wincker P, Mauger S et al. (2001) The
Alonso-Calleja C, Carballo J, Capita R, Bernardo A & complete genome sequence of the lactic acid bacterium
Garcı́ a-L
opez MaL (2002) Changes in the microflora of Lactococcus lactis ssp. lactis IL1403. Genome Res 11: 731–753.
Valdeteja raw goat’s milk cheese throughout manufacturing Bolotin A, Quinquis B, Renault P et al. (2004) Complete
and ripening. LWT-Food Sci Technol 35: 222–232. sequence and comparative genome analysis of the dairy
bacterium Streptococcus thermophilus. Nat Biotechnol 22: Cardamone L, Quiberoni A, Mercanti D, Fornasari M,
1554–1558. Reinheimer J & Guglielmotti D (2011) Adventitious dairy
Bolotin A, Quinquis B, Ehrlich SD & Sorokin A (2012) Leuconostoc strains with interesting technological and
Complete genome sequence of Lactococcus lactis subsp. biological properties useful for adjunct starters. Dairy Sci
cremoris A76. J Bacteriol 194: 1241–1242. Technol 91: 457–470.
Bonizzi I, Buffoni JN, Feligini M & Enne G (2009) Carraro L, Maifreni M, Bartolomeoli I et al. (2011) Comparison
Investigating the relationship between raw milk bacterial of culture-dependent and -independent methods for bacterial
composition, as described by intergenic transcribed community monitoring during Montasio cheese
spacer-PCR fingerprinting, and pasture altitude. J Appl manufacturing. Res Microbiol 162: 231–239.
Microbiol 107: 1319–1329. Casaus P, Nilsen T, Cintas LM, Nes IF, Hernandez PE & Holo
Boyazoglu J & Morand-Fehr P (2001) Mediterranean dairy H (1997) Enterocin B, a new bacteriocin from Enterococcus

sheep and goat products and their quality: a critical review. faecium T136 which can act synergistically with enterocin A.
Small Rum Res 40: 1–11. Microbiology UK 143: 2287–2294.
Bradley AJ, Leach KA, Breen JE, Green LE & Green MJ Chen L-S, Ma Y, Maubois J-L, He S-H, Chen L-J & Li H-M
(2007) Survey of the incidence and aetiology of mastitis (2010) Screening for the potential probiotic yeast strains
on dairy farms in England and Wales. Vet Rec 160: from raw milk to assimilate cholesterol. Dairy Sci Technol
253–257. 90: 537–548.
Braem G, De Vliegher S, Verbist B, Heyndrickx M, Leroy F & Chiodini RJ, Chamberlin WM, Sarosiek J & McCallum RW
De Vuyst L (2012) Culture-independent exploration of the (2012) Crohn’s disease and the mycobacterioses: a quarter
teat apex microbiota of dairy cows reveals a wide bacterial century later. Causation or simple association? Crit Rev
species diversity. Vet Microbiol 157: 383–390. Microbiol 38: 52–93.
Braun-Fahrl€ander C & Von Mutius E (2011) Can farm milk Christensen JE, Dudley EG, Pederson JA & Steele JL (1999)
consumption prevent allergic diseases? Clin Exp Allergy 41: Peptidases and amino acid catabolism in lactic acid bacteria.
29–35. Anton Leeuw Int JG 76: 217–246.
Bravo D, Rodriguez E & Medina M (2009) Nisin and lacticin Cocco RR, Jarvinen KM, Sampson HA & Beyer K (2003)
481 coproduction by Lactococcus lactis strains isolated from Mutational analysis of major, sequential IgE-binding
raw ewes’ milk. J Dairy Sci 92: 4805–4811. epitopes in alpha(s1)-casein, a major cow’s milk allergen.
Broadbent J, Cai H, Larsen R et al. (2011) Genetic diversity J Allergy Clin Immunol 112: 433–437.
in proteolytic enzymes and amino acid metabolism Cocolin L, Innocente N, Biasutti M & Comi G (2004) The late
among Lactobacillus helveticus strains. J Dairy Sci 94: blowing in cheese: a new molecular approach based on PCR
4313–4328. and DGGE to study the microbial ecology of the alteration
Cabrera-Rubio R, Collado MC, Laitinen K, Salminen S, process. Int J Food Microbiol 90: 83–91.
Isolauri E & Mira A (2012) The human milk microbiome Coker R, McKee M, Atun R, Dimitrova B, Dodonova E,
changes over lactation and is shaped by maternal weight Kuznetsov S & Drobniewski F (2006) Risk factors for
and mode of delivery. Am J Clin Nutr 96: 544–551. pulmonary tuberculosis in Russia: case–control study. BMJ
Cailliez-Grimal C, Miguindou-Mabiala R, Leseine M, 332: 85–87.
Revol-Junelles AM & Milliere JB (2005) Quantitative Collado MC, Meriluoto J & Salminen S (2008) Adhesion and
polymerase chain reaction used for the rapid detection of aggregation properties of probiotic and pathogen strains.
Carnobacterium species from French soft cheeses. FEMS Eur Food Res Technol 226: 1065–1073.
Microbiol Lett 250: 163–169. Coorevits A, De Jonghe V, Vandroemme J et al. (2008)
Callanan M, Kaleta P, O’Callaghan J et al. (2008) Genome Comparative analysis of the diversity of aerobic
sequence of Lactobacillus helveticus, an organism spore-forming bacteria in raw milk from organic and
distinguished by selective gene loss and insertion sequence conventional dairy farms. Syst Appl Microbiol 31: 126–140.
element expansion. J Bacteriol 190: 727–735. Cosentino S, Fadda ME, Deplano M, Melis R, Pomata R &
Callon C, Delbes C, Duthoit F & Montel MC (2006) Pisano MB (2012) Antilisterial activity of nisin-like
Application of SSCP-PCR fingerprinting to profile the yeast bacteriocin-producing Lactococcus lactis subsp. lactis isolated
community in raw milk Salers cheeses. Syst Appl Microbiol from traditional sardinian dairy products. J Biomed
29: 172–180. Biotechnol 2012: 376428.
Callon C, Duthoit F, Delbes C, Ferrand M, Le Frileux Y, De Coton M, Delbes-Paus C, Irlinger F et al. (2012) Diversity and
Cremoux R & Montel MC (2007) Stability of microbial assessment of potential risk factors of Gram-negative isolates
communities in goat milk during a lactation year: molecular associated with French cheeses. Food Microbiol 29: 88–98.
approaches. Syst Appl Microbiol 30: 547–560. Cotter PD, Hill C & Ross RP (2005) Bacteriocins: developing
zek Majhenic A, Rogelj I & Perko B (2005) Enterococci
Can innate immunity for food. Nat Rev Microbiol 3: 777–788.
from Tolminc cheese: population structure, antibiotic Cousin FJ, Mater DDG, Foligne B & Jan G (2011) Dairy
susceptibility and incidence of virulence determinants. Int J propionibacteria as human probiotics: a review of recent
Food Microbiol 102: 239–244. evidence. Dairy Sci Technol 91: 1–26.
Cousin FJ, Jouan-Lanhouet S, Dimanche-Boitrel M-T, Corcos microbial consortium on biogenic amine content and
L & Jan G (2012) Milk fermented by Propionibacterium sensory characteristics of an uncooked pressed cheese. Food
freudenreichii induces apoptosis of HGT-1 human gastric Microbiol 30: 74–82.
cancer cells. PLoS One 7: e31892. Delcenserie V, Bechoux N, China B, Daube G & Gavini F
Cremonesi P, Vanoni L, Silvetti T, Morandi S & Brasca M (2005) A PCR method for detection of bifidobacteria in raw
(2012) Identification of Clostridium beijerinckii, milk and raw milk cheese: comparison with culture-based
Cl. butyricum, Cl. sporogenes, Cl. tyrobutyricum isolated methods. J Microbiol Meth 61: 55–67.
from silage, raw milk and hard cheese by a multiplex PCR Delgado S, Arroyo R, Jimenez E, Marin ML, del Campo R,
assay. J Dairy Res 79: 318–323. Fernandez L & Rodriguez JM (2009) Staphylococcus
Creppy EE (2002) Update of survey, regulation and toxic epidermidis strains isolated from breast milk of
effects of mycotoxins in Europe. Toxicol Lett 127: 19–28. women suffering infectious mastitis: potential virulence

Davies JE (1997) Origins, acquisition and dissemination of traits and resistance to antibiotics. BMC Microbiol
antibiotic resistance determinants. Ciba Found Symp 207: 9: 82.
15–27; discussion 27–35. Delves-Brougthon J (1990) Nisin and its uses as a food
de Garnica ML, Santos JA & Gonzalo C (2011) Short preservative. Food Technol 44: 100–117.
communication: influence of storage and preservation on Desmasures N & Gueguen M (1997) Monitoring the
microbiological quality of silo ovine milk. J Dairy Sci 94: microbiology of high quality milk by monthly sampling
1922–1927. over 2 years. J Dairy Res 64: 271–280.
De Jonghe V, Coorevits A, Van Hoorde K, Messens W, Van Devirgiliis C, Caravelli A, Coppola D, Barile S & Perozzi G
Landschoot A, De Vos P & Heyndrickx M (2011) Influence (2008) Antibiotic resistance and microbial composition
of storage conditions on the growth of Pseudomonas species along the manufacturing process of Mozzarella di Bufala
in refrigerated raw milk. Appl Environ Microbiol 77: 460– Campana. Int J Food Microbiol 128: 378–384.
470. Dijkshoorn L (2013) Acinetobacter baumannii. Molecular
De la Rua-Domenech R (2006) Human Mycobacterium bovis Typing in Bacterial Infections (de Filippis I & McKee ML,
infection in the United Kingdom: incidence, risks, control ed.), pp. 433–456. Humana Press Inc., New York.
measures and review of the zoonotic aspects of bovine Dinakar P & Mistry VV (1994) Growth and viability of
tuberculosis. Tuberculosis (Edinb) 86: 77. Bifidobacterium bifidum in cheddar cheese. J Dairy Sci 77:
De Vuyst L & Tsakalidou E (2008) Streptococcus macedonicus, 2854–2864.
a multi-functional and promising species for dairy Doyle MP, Glass K, Beery J, Garcia G, Pollard D & Schultz R
fermentations. Int Dairy J 18: 476–485. (1987) Survival of Listeria monocytogenes in milk during
Debarry J, Garn H, Hanuszkiewicz A et al. (2007) high-temperature, short-time pasteurization. Appl Environ
Acinetobacter lwoffii and Lactococcus lactis strains isolated Microbiol 53: 1433–1438.
from farm cowsheds possess strong allergy-protective Doyle MP, Loneragan GH, Scott HM & Singer RS (2013)
properties. J Allergy Clin Immunol 119: 1514–1521. Antimicrobial resistance: challenges and perspectives. Compr
Deegan LH, Cotter PD, Hill C & Ross P (2006) Bacteriocins: Rev Food Sci Food Saf 12: 234–248.
biological tools for bio-preservation and shelf-life extension. Driehuis F (2013) Silage and the safety and quality of dairy
Int Dairy J 16: 1058–1071. foods: a review. Agric Food Sci 22: 16–34.
Deetae P, Spinnler H-E, Bonnarme P & Helinck S (2009) Duthoit F, Godon JJ & Montel MC (2003) Bacterial
Growth and aroma contribution of Microbacterium foliorum, community dynamics during production of registered
Proteus vulgaris and Psychrobacter sp. during ripening in a designation of origin Salers cheese as evaluated by 16S
cheese model medium. Appl Microbiol Biotechnol 82: 169– rRNA gene single-strand conformation polymorphism
177. analysis. Appl Environ Microbiol 69: 3840–3848.
Delavenne E, Mounier J, Asmani K, Jany J-L, Barbier G & Le Duthoit F, Callon C, Tessier L & Montel MC (2005)
Blay G (2011) Fungal diversity in cow, goat and ewe milk. Relationships between sensorial characteristics and microbial
Int J Food Microbiol 151: 247–251. dynamics in “Registered Designation of Origie’ Salers
Delbes C, Ali-Mandjee L & Montel MC (2007) Monitoring cheese. Int J Food Microbiol 103: 259–270.
bacterial communities in raw milk and cheese by Eddy B (1960) The use and meaning of the term
culture-dependent and -independent 16S rRNA gene-based ‘psychrophilic’. J Appl Microbiol 23: 189–190.
analyses. Appl Environ Microbiol 73: 1882–1891. Ege MJ, Mayer M, Schwaiger K et al. (2012) Environmental
Delbes-Paus C, Irlinger F, Coton M et al. (2011) Benefits and bacteria and childhood asthma. Allergy 67: 1565–1571.
risks associated with Gram-negative bacteria within cheese El-Ghaish S, Ahmadova A, Hadji-Sfaxi I et al. (2011) Potential
microbial communities. 10th International Meeting on use of lactic acid bacteria for reduction of allergenicity and
Mountain Cheese 14–15 September 2011. Ed. University of for longer conservation of fermented foods. Trends Food Sci
Turin, Dronero (CN), Italy. Technol 22: 509–516.
Delbes-Paus C, Pochet S, Helinck S et al. (2012) Impact of Emond E & Moineay S (2007) Bacteriophages and food
Gram-negative bacteria in interaction with a complex fermentations. Bacteriophage: Genetics and Molecular Biology
(McGrath S & van Sinderen D, eds), pp. 93–124. Horizon Ferrario C, Ricci G, Borgo F, Rollando A & Fortina MG
Scientific Press/Caister Academic Press, Poole, UK. (2012) Genetic investigation within Lactococcus garvieae
Ercolini D, Moschetti G, Blaiotta G & Coppola S (2001) The revealed two genomic lineages. FEMS Microbiol Lett 332:
potential of a polyphasic PCR-DGGE approach in 153–161.
evaluating microbial diversity of natural whey cultures for Florez AB, Reimundo P, Delgado S, Fernandez E, Alegria A,
water-buffalo mozzarella cheese production: bias of Guijarro JA & Mayo B (2012) Genome sequence of
culture-dependent and culture-independent analyses. Syst Lactococcus garvieae IPLA 31405, a bacteriocin-producing,
Appl Microbiol 24: 610–617. tetracycline-resistant strain isolated from a raw-milk cheese.
Ercolini D, Mauriello G, Blaiotta G, Moschetti G & Coppola S J Bacteriol 194: 5118–5119.
(2004) PCR-DGGE fingerprints of microbial succession Floros G, Hatzikamari M, Litopoulou-Tzanetaki E &
during a manufacture of traditional water buffalo Tzanetakis N (2012) Probiotic and technological properties

mozzarella cheese. J Appl Microbiol 96: 263–270. of facultatively heterofermentative Lactobacilli from Greek
Ercolini D, Russo F, Ferrocino I & Villani F (2009) Molecular traditional cheeses. Food Biotechnol 26: 85–105.
identification of mesophilic and psychrotrophic bacteria Forghani F, Nazemi A, Oh D, Saffar S, Taghinezhad M &
from raw cow’s milk. Food Microbiol 26: 228–231. Hamidkholgh K (2012) Novel lactic Acid Bacteria in raw
Ercolini D, De Filippis F, La Storia A & Iacono M (2012) cow’s milk from highland farms. Ann Biol Res 3: 3055–3061.
“Remake” by high-throughput sequencing of the microbiota Fortina MG, Ricci G, Foschino R et al. (2007) Phenotypic
involved in the production of water buffalo mozzarella typing, technological properties and safety aspects of
cheese. Appl Environ Microbiol 78: 8142–8145. Lactococcus garvieae strains from dairy environments. J Appl
Espeche MC, Pellegrino M, Frola I, Larriestra A, Bogni C & Microbiol 103: 445–453.
Nader-Macıas MEF (2012) Lactic acid bacteria from raw Fortina MG, Ricci G & Borgo F (2009) A study of lactose
milk as potentially beneficial strains to prevent bovine metabolism in Lactococcus garvieae reveals a genetic marker
mastitis. Anaerobe 18: 103–109. for distinguishing between dairy and fish biotypes. J Food
European Food Safety Authority (EFSA) (2012) Scientific Prot 72: 1248–1254.
Report of EFSA and ECDC. The European Union Summary Foschino R, Nucera D, Volponi G, Picozzi C, Ortoffi M &
Report on Trends and Sources of Zoonoses, Zoonotic Agents Bottero MT (2008) Comparison of Lactococcus garvieae
and Food-borne Outbreaks in 2011. EFSA J 10: 2597. strains isolated in northern Italy from dairy products and
Facklam R (2002) What happened to the streptococci: fishes through molecular typing. J Appl Microbiol 105:
overview of taxonomic and nomenclature changes. Clin 652–662.
Microbiol Rev 15: 613. Fotou K, Tzora A, Voidarou C et al. (2011) Isolation of
Falenski A, Mayer-Scholl A, Filter M, Gollner C, Appel B & microbial pathogens of subclinical mastitis from raw sheep’s
Nockler K (2011) Survival of Brucella spp. in mineral water, milk of Epirus (Greece) and their role in its hygiene.
milk and yogurt. Int J Food Microbiol 145: 326–330. Anaerobe 17: 315–319.
Falentin H, Deutsch SM, Jan G et al. (2010) The complete Foulquie Moreno MR, Sarantinopoulos P, Tsakalidou E &
genome of Propionibacterium freudenreichii CIRM-BIA1T, a De Vuyst L (2006) The role and application of enterococci
hardy actinobacterium with food and probiotic applications. in food and health. Int J Food Microbiol 106: 1–24.
PLoS One 5: e11748. Foxman B, D’Arcy H, Gillespie B, Bobo JK & Schwartz K
Fall PA, Leroi F, Cardinal M, Chevalier F & Pilet MF (2010) (2002) Lactation mastitis: occurrence and medical
Inhibition of Brochothrix thermosphacta and sensory management among 946 breastfeeding women in the United
improvement of tropical peeled cooked shrimp by States. Am J Epidemiol 155: 103–114.
Lactococcus piscium CNCM I-4031. Lett Appl Microbiol 50: Franciosi E, Settanni L, Cavazza A & Poznanski E (2009)
357–361. Presence of enterococci in raw cow’s milk and “Puzzone Di
Fallico V, McAuliffe O, Fitzgerald GF & Ross RP (2011) Moena” cheese. J Food Proc Pres 33: 204–217.
Plasmids of raw milk cheese isolate Lactococcus lactis subsp. Frank JF (1997) Milk and dairy products. Food Microbiology –
lactis biovar diacetylactis DPC3901 suggest a plant-based Fundamental and Frontiers (Doyle P, Beuchat R & Montville J,
origin for the strain. Appl Environ Microbiol 77: 6451–6462. eds), pp. 169–186. ASM Press, Washington, DC.
Delgado S, Martın MC & Mayo B
Fernandez E, Alegrıa A, Franz C, Holzapfel WH & Stiles ME (1999) Enterococci at
(2011) Comparative phenotypic and molecular genetic the crossroads of food safety? Int J Food Microbiol 47:
profiling of wild Lactococcus lactis subsp. lactis strains of the 1–24.
L. lactis subsp. lactis and L. lactis subsp. cremoris genotypes, Fricker M, Skanseng B, Rudi K, Stessl B & Ehling-Schulz M
isolated from starter-free cheeses made of raw milk. Appl (2011) Shift from farm to dairy tank milk microbiota
Environ Microbiol 77: 5324–5335. revealed by a polyphasic approach is independent from
Fernandez L, Langa S, Martın V, Maldonado A, Jimenez E, geographical origin. Int J Food Microbiol 145: S24–S30.
Martın R & Rodrıguez JM (2012) The human milk Fromm HI & Boor K (2004) Characterization of
microbiota: origin and potential roles in health and disease. pasteurized fluid milk shelf-life attributes. J Food Sci 69:
Pharmacol Res 69: 1–10. M207–M214.
Gao A, Mutharia L, Chen S, Rahn K & Odumeru J (2002) Goetsch AL, Zeng SS & Gipson TA (2011) Factors affecting goat
Effect of pasteurization on survival of Mycobacterium milk production and quality. Small Rumin Res 101: 55–63.
paratuberculosis in milk. J Dairy Sci 85: 3198–3205. Grant IR, Hitchings EI, McCartney A, Ferguson F & Rowe MT
Garedew L, Berhanu A, Mengesha D & Tsegay G (2012) (2002) Effect of commercial-scale high-temperature,
Identification of gram-negative bacteria from critical control short-time pasteurization on the viability of Mycobacterium
points of raw and pasteurized cow milk consumed at paratuberculosis in naturally infected cows’ milk. Appl
Gondar town and its suburbs, Ethiopia. BMC Public Health Environ Microbiol 68: 602–607.
12: 950. Guatteo R, Beaudeau F, Berri M, Rodolakis A, Joly A &
Garmeine G, Salomskiene J, Jasutiene I, Macioniene I & Seegers H (2006) Shedding routes of Coxiella burnetii in
Miliauskiene I (2010) Production of benzoic acid by lactic dairy cows: implications for detection and control. Vet Res
acid bacteria from Lactobacillus, Lactococcus and 37: 827–833.

Streptococcus genera in milk. Milchwissenschaft 65: 295–298. Gul S, Masud T, Maqsood S, Latif A, Irshad I & ul Haque I
Gaudin JC, Rabesona H, Choiset Y et al. (2008) Assessment of (2012) Streptococcus thermophilus bacteriocin, from
the immunoglobulin E-mediated immune response to production to their application: an overview. Afr J Microbiol
milk-specific proteins in allergic patients using microarrays. Res 6: 859–866.
Clin Exp Allergy 38: 686–693. Gurung M, Nam HM, Tamang MD, Chae MH, Jang GC, Jung
Gaya P, Babın M, Medina M & Nu~ nez M (2001) Diversity SC & Lim SK (2013) Prevalence and antimicrobial
among lactococci isolated from ewes’ raw milk and cheese. susceptibility of Acinetobacter from raw bulk tank milk in
J Appl Microbiol 87: 849–855. Korea. J Dairy Sci 96: 1997–2002.
Georgalaki MD, Van Den Berghe E, Kritikos D et al. (2002) Hagi T, Kobayashi M & Nomura M (2010) Molecular-based
Macedocin, a food-grade lantibiotic produced by analysis of changes in indigenous milk microflora during
Streptococcus macedonicus ACA-DC 198. Appl Environ the grazing period. Biosci Biotechnol Biochem 74: 484–487.
Microbiol 68: 5891–5903. Han B-Z, Meng Y, Li M, Yang Y-X, Ren F-Z, Zeng Q-K &
Georgalaki M, Papadimitriou K, Anastasiou R, Pot B, Van Robert Nout MJ (2007) A survey on the microbiological
Driessche G, Devreese B & Tsakalidou E (2013) and chemical composition of buffalo milk in China. Food
Macedovicin, the second food-grade lantibiotic produced by Control 18: 742–746.
Streptococcus macedonicus ACA-DC 198. Food Microbiol 33: Hannon JA, Wilkinson MG, Delahunty CM, Wallace JM,
124–130. Morrissey PA & Beresford TP (2003) Use of autolytic starter
Giacometti F, Serraino A, Finazzi G et al. (2012) systems to accelerate the ripening of Cheddar cheese. Int
Foodborne pathogens in in-line milk filters and associated Dairy J 13: 313–323.
on-farm risk factors in dairy farms authorized to produce Hantsis-Zacharov E & Halpern M (2007) Chryseobacterium
and sell raw milk in northern Italy. J Food Prot 75: haifense sp. nov., a psychrotolerant bacterium isolated from
1263–1269. raw milk. Int J Syst Evol Microbiol 57: 2344–2348.
Gil-Campos M, Lopez MA, Rodriguez-Benitez MV et al. Hantsis-Zacharov E, Senderovich Y & Halpern M (2008a)
(2012) Lactobacillus fermentum CECT 5716 is safe and well Chryseobacterium bovis sp nov., isolated from raw cow’s
tolerated in infants of 1–6 months of age: a Randomized milk. Int J Syst Evol Microbiol 58: 1024–1028.
Controlled Trial. Pharmacol Res 65: 231–238. Hantsis-Zacharov E, Shaked T, Senderovich Y & Halpern M
Giraffa G (2002) Enterococci from foods. FEMS Microbiol Rev (2008b) Chryseobacterium oranimense sp nov., a
26: 163–171. psychrotolerant, proteolytic and lipolytic bacterium isolated
Giraffa G (2003) Functionality of enterococci in dairy from raw cow’s milk. Int J Syst Evol Microbiol 58:
products. Int J Food Microbiol 88: 215–222. 2635–2639.
Giraffa G & Carminati D (1997) Control of Listeria Hao P, Zheng H, Yu Y et al. (2011) Complete sequencing and
monocytogenes in the rind of Taleggio, a surface-smear pan-genomic analysis of Lactobacillus delbrueckii subsp.
cheese, by a bacteriocin from Enterococcus faecium 7C5. Sci bulgaricus reveal its genetic basis for industrial yogurt
Aliments 17: 383–391. production. PLoS One 6: e15964.
Giraffa G, Andrighetto C, Antonello C et al. (2004) Genotypic Harakeh S, Saleh I, Barbour E & Shaib H (2012) Highly
and phenotypic diversity of Lactobacillus delbrueckii subsp. resistant Yersinia enterocolitica isolated from dairy based
lactis strains of dairy origin. Int J Food Microbiol 91: foods in Lebanon. Int Arabic J Antimicrob Agents 2: 1–6.
129–139. Heikkila MP & Saris PEJ (2003) Inhibition of Staphylococcus
Giraffa G, De Vecchi P & Rossetti L (2008) Note: aureus by the commensal bacteria of human milk. J Appl
identification of Lactobacillus delbrueckii subspecies Microbiol 95: 471–478.
bulgaricus and subspecies lactis dairy isolates by amplified Hemme D & Foucaud-Scheunemann C (2004) Leuconostoc,
rDNA restriction analysis. J Appl Microbiol 85: 918. characteristics, use in dairy technology and prospects in
Giribaldi M, Ortoffi MF, Giuffrida MG et al. (2013) Effect of functional foods. Int Dairy J 14: 467–494.
prolonged refrigeration on the protein and microbial profile Henri-Dubernet S, Desmasures N & Gueguen M (2008)
of human milk. Int Dairy J 31: 121–126. Diversity and dynamics of lactobacilli populations during
ripening of RDO Camembert cheese. Can J Microbiol 54: colorectal carcinoma cells via short-chain fatty acids acting
218–228. on mitochondria. Cell Death Differ 9: 179.
Herman LM, De Block JH & Waes GM (1995) A direct PCR Jans C, Bugnard J, Njage PMK, Lacroix C & Meile L (2012)
detection method for Clostridium tyrobutyricum spores in up Lactic acid bacteria diversity of African raw and fermented
to 100 milliliters of raw milk. Appl Environ Microbiol 61: camel milk products reveals a highly competitive, potentially
4141–4146. health-threatening predominant microflora. LWT-Food Sci
Herreros M, Sandoval H, Gonzalez L, Castro J, Fresno J & Technol 47: 371–379.
Tornadijo M (2005) Antimicrobial activity and antibiotic Jans C, Follador R, Hochstrasser M, Lacroix C, Meile L &
resistance of lactic acid bacteria isolated from Armada Stevens MJ (2013) Comparative genome analysis of
cheese (a Spanish goats’ milk cheese). Food Microbiol 22: Streptococcus infantarius subsp. infantarius CJ18, an African
455–459. fermented camel milk isolate with adaptations to dairy

Herve-Jimenez L, Guillouard I, Guedon E et al. (2009) environment. BMC Genomics 14: 200.
Postgenomic analysis of Streptococcus thermophilus Jayne-Williams D & Skerman T (1966) Comparative studies
cocultivated in milk with Lactobacillus delbrueckii subsp. on Coryneform bacteria from milk and dairy sources. J Appl
bulgaricus: involvement of nitrogen, purine, and iron Microbiol 29: 72–92.
metabolism. Appl Environ Microbiol 75: 2062–2073. Jones CA (2001) Maternal transmission of infectious pathogens
Hickey DK, Kilcawley KN, Beresford TP & Wilkinson MG in breast milk. J Paediatr Child Health 37: 576–582.
(2007) Lipolysis in cheddar cheese made from raw, Kahala M, Maki M, Lehtovaara A et al. (2008)
thermized, and pasteurized milks. J Dairy Sci 90: 47–56. Characterization of starter lactic acid bacteria from the
Holler BJ & Steele JL (1995) Characterization of lactococci Finnish fermented milk product viili. J Appl Microbiol 105:
other than Lactococcus-lactis for possible use as starter 1929–1938.
cultures. Int Dairy J 5: 275–289. Kelton DF, Lissemore KD & Martin RE (1998)
Hugenholtz J & Starrenburg MJ (1992) Diacetyl production by Recommendations for recording and calculating the
different strains of Lactococcus lactis subsp. lactis var. incidence of selected clinical diseases of dairy cattle. J Dairy
diacetylactis and Leuconostoc spp. Appl Microbiol Biotechnol Sci 81: 2502–2509.
38: 17–22. Khedid K, Faid M, Mokhtari A, Soulaymani A & Zinedine A
Hunt KM, Foster JA, Forney LJ et al. (2011) Characterization (2009) Characterization of lactic acid bacteria isolated from
of the diversity and temporal stability of bacterial the one humped camel milk produced in Morocco.
communities in human milk. PLoS One 6: e21313. Microbiol Res 164: 81–91.
Hunt K, Drummond N, Murphy M, Butler F, Buckley J & Kholif A, Mahran G, El-Nawawy M, Ismail A, Salem M &
Jordan K (2012) A case of bovine raw milk contamination Zaky W (2011) Evaluation of proteolytic activity of some
with Listeria monocytogenes. Ir Vet J 65: 13. dairy lactobacilli. World J Dairy Food Sci 6: 21–26.
Irlinger F (2008) Safety assessment of dairy microorganisms: Kiernan RC, Beresford TP, O’Cuinn G & Jordan KN (2000)
coagulase-negative staphylococci. Int J Food Microbiol 126: Autolysis of lactobacilli during Cheddar cheese ripening.
302–310. Irish J Agric Food Res 39: 95–106.
Irlinger F & Mounier J (2009) Microbial interactions in cheese: Kim H, Shin H, Ha W, Yang H & Lee S (2006)
implications for cheese quality and safety. Curr Opin Characterization of lactic bacterial strains isolated from raw
Biotechnol 20: 142–148. milk. Asian Aust J Anim Sci 19: 131.
Irlinger F, In Yung SAY, Sarthou A-S et al. (2012) Ecological Kimoto-Nira H, Aoki R, Mizumachi K, Sasaki K, Naito H,
and aromatic impact of two Gram-negative bacteria Sawada T & Suzuki C (2012) Interaction between
(Psychrobacter celer and Hafnia alvei) inoculated as part of Lactococcus lactis and Lactococcus raffinolactis during growth
the whole microbial community of an experimental smear in milk: development of a new starter culture. J Dairy Sci
soft cheese. Int J Food Microbiol 153: 332–338. 95: 2176–2185.
Izquierdo E, Marchioni E, Aoude-Werner D, Hasselmann C & Klinger I & Rosenthal I (1997) Public health and the safety of
Ennahar S (2009) Smearing of soft cheese with Enterococcus milk and milk products from sheep and goats. Rev Sci Tech
faecium WHE 81, a multi-bacteriocin producer, against 16: 482.
Listeria monocytogenes. Food Microbiol 26: 16–20. Kopp-Hoolihan L (2001) Prophylactic and therapeutic uses of
Jacques N & Casaregola S (2008) Safety assessment of dairy probiotics: a review. J Am Dietetic Assoc 101: 229–241.
microorganisms: the hemiascomycetous yeasts. Int J Food Ladero V, Sanchez-Llana E, Fernandez M & Alvarez MA
Microbiol 126: 321–326. (2011) Survival of biogenic amine-producing dairy LAB
Jamaly N, Benjouad A, Bouksaim M et al. (2011) Probiotic strains at pasteurisation conditions. Int J Food Sci Technol
potential of Lactobacillus strains isolated from known 46: 516–521.
popular traditional Moroccan dairy products. Br Microbiol Lafarge V, Ogier JC, Girard V, Maladen V, Leveau JY, Gruss A
Res J 1: 79–94. & Delacroix-Buchet A (2004) Raw cow milk bacterial
Jan G, Belzacq A, Haouzi D, Rouault A, Metivier D, Kroemer population shifts attributable to refrigeration. Appl Environ
G & Brenner C (2002) Propionibacteria induce apoptosis of Microbiol 70: 5644–5650.
Lagneau PE, Lebtahi K & Swinne D (1996) Isolation of yeasts Lombardi A, Gatti M, Rizzotti L, Torriani S, Andrighetto C &
from bovine milk in Belgium. Mycopathologia 135: 99–102. Giraffa G (2004) Characterization of Streptococcus
Lamendella R, Santo Domingo JW, Kelty C & Oerther DB macedonicus strains isolated from artisanal Italian raw milk
(2008) Bifidobacteria in feces and environmental waters. cheeses. Int Dairy J 14: 967–976.
Appl Environ Microbiol 74: 575–584. Lopez-Enriquez L, Rodriguez-Lazaro D & Hernandez M (2007)
Lane MM & Morrissey JP (2010) Kluyveromyces marxianus: a Quantitative detection of Clostridium tyrobutyricum in milk
yeast emerging from its sister’s shadow. Fungal Biol Rev 24: by real-time PCR. Appl Environ Microbiol 73: 3747–3751.
17–26. Lourens-Hattingh A & Viljoen BC (2001) Yogurt as probiotic
Langer AJ, Ayers T, Grass J, Lynch M, Angulo FJ & Mahon BE carrier food. Int Dairy J 11: 1–17.
(2012) Nonpasteurized dairy products, disease outbreaks, Madera C, Monjardın C & Suarez JE (2004) Milk
and state laws-United States, 1993-2006. Emerg Infect Dis contamination and resistance to processing conditions

18: 385–391. determine the fate of Lactococcus lactis bacteriophages in
Langsrud T & Reinbold G (1973) Flavor development and dairies. Appl Environ Microbiol 70: 7365–7371.
microbiology of Swiss cheese – a review. III. Ripening and Magni C, Espeche C, Repizo GD et al. (2012) Draft genome
flavor production. J Milk Food Technol 36: 593–609. sequence of Enterococcus mundtii CRL1656. J Bacteriol 194:
Latorre AA, Pradhan AK, Van Kessel JAS et al. (2011) 550.
Quantitative risk assessment of listeriosis due to Mallet A, Gueguen M, Kauffmann F, Chesneau C, Sesboue A
consumption of raw milk. J Food Prot 74: 1268–1281. & Desmasures N (2012) Quantitative and qualitative
Lavoie K, Touchette M, St-Gelais D & Labrie S (2012) microbial analysis of raw milk reveals substantial diversity
Characterization of the fungal microflora in raw milk and influenced by herd management practices. Int Dairy J 27:
specialty cheeses of the province of Quebec. Dairy Sci 13–21.
Technol 92: 455–468. Maniruzzaman M, Khan M, Amin M, Paul A & Islam M (2010)
Le Bourhis AG, Saunier K, Dore J, Carlier JP, Chamba JF, Isolation and identification of bacterial flora from milk of
Popoff MR & Tholozan JL (2005) Development and apparently health buffalo–cows. Int J BioRes 1: 13–16.
validation of PCR primers to assess the diversity of Mannu L, Paba A, Daga E, Comunian R, Zanetti S, Dupre I &
Clostridium spp. in cheese by temporal temperature gradient Sechi LA (2003) Comparison of the incidence of virulence
gel electrophoresis. Appl Environ Microbiol 71: 29–38. determinants and antibiotic resistance between Enterococcus
Leisner JJ, Laursen BG, Prevost H, Drider D & Dalgaard P faecium strains of dairy, animal and clinical origin. Int J
(2007) Carnobacterium: positive and negative effects in the Food Microbiol 88: 291–304.
environment and in foods. FEMS Microbiol Rev 31: Maragkoudakis PA, Zoumpopoulou G, Miaris C,
592–613. Kalantzopoulos G, Pot B & Tsakalidou E (2006) Probiotic
Lejeune JT & Rajala-Schultz PJ (2009) Food safety: potential of Lactobacillus strains isolated from dairy
unpasteurized milk: a continued public health threat. Clin products. Int Dairy J 16: 189–199.
Infect Dis 48: 93–100. Marchand S, Heylen K, Messens W et al. (2008) Seasonal
Lepage E, Brinster S, Caron C et al. (2006) Comparative influence on heat-resistant proteolytic capacity of
genomic hybridization analysis of Enterococcus faecalis: Pseudomonas lundensis and Pseudomonas fragi, predominant
identification of genes absent from food strains. J Bacteriol milk spoilers isolated from Belgian raw milk samples.
188: 6858–6868. Environ Microbiol 11: 467–482.
Leroy F & De Vuyst L (2004) Lactic acid bacteria as functional Marco MB, Moineau S & Quiberoni A (2012) Bacteriophages
starter cultures for the food fermentation industry. Trends and dairy fermentations. Bacteriophage 2: 0–1.
Food Sci Technol 15: 67–78. Marilley L & Casey MG (2004) Flavours of cheese products:
Liatsos GD, Thanellas S, Pirounaki M, Ketikoglou I & metabolic pathways, analytical tools and identification of
Moulakakis A (2012) Listeria monocytogenes peritonitis: producing strains. Int J Food Microbiol 90: 139–159.
presentation, clinical features, treatment, and outcome. Scan Martin A & Beutin L (2011) Characteristics of Shiga
J Gastroenterol 47: 1129–1140. toxin-producing Escherichia coli from meat and milk
Lind H, Jonsson H & Schn€ urer J (2005) Antifungal effect of products of different origins and association with food
dairy propionibacteria – contribution of organic acids. Int J producing animals as main contamination sources. Int J
Food Microbiol 98: 157–165. Food Microbiol 146: 99–104.
Lind H, Sj€ ogren J, Gohil S, Kenne L, Schn€ urer J & Broberg A Martin R, Langa S, Reviriego C et al. (2004) The commensal
(2007) Antifungal compounds from cultures of dairy microflora of human milk: new perspectives for food
propionibacteria type strains. FEMS Microbiol Lett 271: bacteriotherapy and probiotics. Trends Food Sci Technol 15:
310–315. 121–127.
Loman NJ, Constantinidou C, Chan JZ et al. (2012) Martin R, Jimenez E, Heilig H, Fernandez L, Marin ML,
High-throughput bacterial genome sequencing: an Zoetendal EG & Rodriguez JM (2009) Isolation of
embarrassment of choice, a world of opportunity. Nat Rev Bifidobacteria from breast milk and assessment of the
Microbiol 10: 599–606. bifidobacterial population by PCR-denaturing gradient gel
electrophoresis and quantitative real-time PCR. Appl Meslier V, Loux V & Renault P (2012b) Genome sequence of
Environ Microbiol 75: 965–969. Lactococcus raffinolactis strain 4877, isolated from natural
Martin NH, Ranieri ML, Murphy SC, Ralyea RD, dairy starter culture. J Bacteriol 194: 6364.
Wiedmann M & Boor KJ (2011) Results from raw milk Michel V, Hauwuy A & Chamba J (2006) Raw milk microbial
microbiological tests do not predict the shelf-life composition: differences in links with microbial practices.
performance of commercially pasteurized fluid milk. Rencontres Rech Rumin 13: 309–312.
J Dairy Sci 94: 1211–1222. Mirhosseini M, Nahvi I, Emtiazi G & Tavassoli M (2010)
Martın R, Heilig HG, Zoetendal EG, Jimenez E, Fernandez L, Characterisation of anti-Listeria monocytogenes bacteriocins
Smidt H & Rodrıguez JM (2007) Cultivation-independent from Enterococcus faecium strains isolated from dairy
assessment of the bacterial diversity of breast milk among products. Int J Dairy Technol 63: 55–61.
healthy women. Res Microbiol 158: 31–37. Monnet C, Loux V, Gibrat JF et al. (2010) The Arthrobacter

Martin-Platero AM, Maqueda M, Valdivia E, Purswani J & arilaitensis Re117 genome sequence reveals its genetic
Martinez-Bueno M (2009) Polyphasic study of microbial adaptation to the surface of cheese. PLoS One 5: e15489.
communities of two Spanish farmhouse goats’ milk Monsallier F, Verdier-Metz I, Agabriel C, Martin B & Montel
cheeses from Sierra de Aracena. Food Microbiol 26: M-C (2012) Variability of microbial teat skin flora in
294–304. relation to farming practices and individual dairy cow
Martins ML & Martins HM (2000) Aflatoxin M1 in raw and characteristics. Dairy Sci Technol 92: 265–278.
ultra high temperature-treated milk commercialized in Mora D, Scarpellini M, Franzetti L, Colombo S & Galli A
Portugal. Food Addit Contam 17: 871–874. (2003) Reclassification of Lactobacillus maltaromicus (Miller
Masoud W, Takamiya M, Vogensen FK, Lillevang S, Abu et al. 1974) DSM 20342 (T) and DSM 20344 and
Al-Soud W, Sorensen SJ & Jakobsen M (2011) Carnobacterium piscicola (Collins et al. 1987) DSM 20730(T)
Characterization of bacterial populations in Danish raw and DSM 20722 as Carnobacterium maltaromaticum comb.
milk cheeses made with different starter cultures by nov. Int J Syst Evol Microbiol 53: 675–678.
denaturating gradient gel electrophoresis and Morandi S, Brasca M & Lodi R (2011) Technological,
pyrosequencing. Int Dairy J 21: 142–148. phenotypic and genotypic characterisation of wild lactic acid
Masoud W, Vogensen FK, Lillevang S, Abu Al-Soud W, bacteria involved in the production of Bitto PDO Italian
Sorensen SJ & Jakobsen M (2012) The fate of indigenous cheese. Dairy Sci Technol 91: 341–359.
microbiota, starter cultures, Escherichia coli, Listeria innocua Morita RY (1975) Psychrophilic bacteria. Bacteriol Rev 39:
and Staphylococcus aureus in Danish raw milk and cheeses 144–167.
determined by pyrosequencing and quantitative real time Muguerza B, Ramos M, Sanchez E et al. (2006)
(qRT)-PCR. Int J Food Microbiol 153: 192–202. Antihypertensive activity of milk fermented by Enterococcus
Mathieu F, Suwandhi IS, Rekhif N, Milliere JB & Lefebvre G faecalis strains isolated from raw milk. Int Dairy J 16: 61–69.
(1993) Mesenterocin 52, a bacteriocin produced by Munsch-Alatossava P & Alatossava T (2007) Antibiotic
Leuconostoc mesenteroides ssp. mesenteroides FR 52. J Appl resistance of raw-milk-associated psychrotrophic bacteria.
Bacteriol 74: 372–379. Microbiol Res 162: 115–123.
Mathur S & Singh R (2005) Antibiotic resistance in food lactic Murphy PA, Hendrich S, Landgren C & Bryant CM (2006)
acid bacteria – a review. Int J Food Microbiol 105: 281–295. Food mycotoxins: an update. J Food Sci 71: R51–R65.
Maurin M & Raoult D (1999) Q fever. Clin Microbiol Rev 12: Naser SM, Hagen KE, Vancanneyt M, Cleenwerck I, Swings J
518–553. & Tompkins TA (2006) Lactobacillus suntoryeus (Cachat and
McAuley CM, Gobius KS, Britz ML & Craven HM (2012) Priest 2005) is a later synonym of Lactobacillus helveticus
Heat resistance of thermoduric enterococci isolated from (Orla-Jensen 1919) Bergey et al. 1925 (Approved Lists
milk. Int J Food Microbiol 154: 162–168. 1980). Int J Syst Evol Microbiol 56: 355–360.
Meile L, Le Blay G & Thierry A (2008) Safety assessment of Newburg DS (2005) Innate immunity and human milk. J Nutr
dairy microorganisms: propionibacterium and 135: 1308–1312.
Bifidobacterium. Int J Food Microbiol 126: 316. Nikolic M, Terzic-Vidojevic A, Jovcic B, Begovic J, Golic N &
Meisel H (2005) Biochemical properties of peptides encrypted Topisirovic L (2008) Characterization of lactic acid bacteria
in bovine milk proteins. Curr Med Chem 12: 1905–1919. isolated from Bukuljac, a homemade goat’s milk cheese. Int
Merin U, Fleminger G, Komanovsky J, Silanikove N, Bernstein J Food Microbiol 122: 162–170.
S & Leitner G (2008) Subclinical udder infection with Nordkvist E & Hoorfar J (2012) An unforeseen source of
Streptococcus dysgalactiae impairs milk coagulation aflatoxins in milk in Sweden. Case Studies in Food Safety
properties: the emerging role of proteose peptones. Dairy Sci and Authenticity: Lessons From Real-Life Situations, pp.
Technol 88: 407–419. 110–115. Woodhead Publishing Ltd., Cambridge, UK.
Meslier V, Loux V & Renault P (2012a) Genome sequence of Olivares M, Diaz-Ropero MP, Martin R, Rodriguez M & Xaus
Leuconostoc pseudomesenteroides strain 4882, isolated from a J (2006) Antimicrobial potential of four Lactobacillus strains
dairy starter culture. J Bacteriol 194: 6637. isolated from breast milk. J Appl Microbiol 101: 72–79.
Oliver SP & Murinda SE (2011) Milk and raw milk Randazzo CL, Torriani S, Akkermans ADL, de Vos WM &
consumption as a vector for human disease. Zoonotic Vaughan EE (2002) Diversity, dynamics, and activity of
Pathogens in the Food Chain (Krause DO & Hendrick S, bacterial communities during production of an artisanal
eds), pp. 99–118. CAB International, Oxfordshire, UK. sicilian cheese as evaluated by 16S rRNA analysis. Appl
Oliver SP, Jayarao BM & Almeida RA (2005) Foodborne Environ Microbiol 68: 1882–1892.
pathogens in milk and the dairy farm environment: food Randazzo CL, Vaughan EE & Caggia C (2006) Artisanal and
safety and public health implications. Foodborne Pathog Dis experimental Pecorino Siciliano cheese: microbial dynamics
2: 115–129. during manufacture assessed by culturing and PCR-DGGE
Oliver SP, Boor KJ, Murphy SC & Murinda SE (2009) Food analyses. Int J Food Microbiol 109: 1–8.
safety hazards associated with consumption of raw milk. Ranieri ML, Huck JR, Sonnen M, Barbano DM & Boor KJ
Foodborne Pathog Dis 6: 793–806. (2009) High temperature, short time pasteurization

Ortolani M, Moraes P, Perin L, Vicßosa G, Carvalho K, Silva temperatures inversely affect bacterial numbers during
J
unior A & Nero L (2010) Molecular identification of refrigerated storage of pasteurized fluid milk. J Dairy Sci 92:
naturally occurring bacteriocinogenic and 4823–4832.
bacteriocinogenic-like lactic acid bacteria in raw milk and Rasic J & Kurmann JA (1983) Bifidobacteria and their role.
soft cheese. J Dairy Sci 93: 2880–2886. Microbiological, nutritional-physiological, medical and
Ott A, Germond JE & Chaintreau A (2000) Origin of technological aspects and bibliography. Experientia Suppl
acetaldehyde during milk fermentation using (13)C-labeled 39: 1.
precursors. J Agric Food Chem 48: 1512–1517. Rasmussen TB, Danielsen M, Valina O, Garrigues C, Johansen
Pacini F, Cariolato D, Andrighetto C & Lombardi A (2006) E & Pedersen MB (2008) Streptococcus thermophilus core
Occurrence of Streptococcus macedonicus in Italian cheeses. genome: comparative genome hybridization study of 47
FEMS Microbiol Lett 261: 69–73. strains. Appl Environ Microbiol 74: 4703–4710.
Papadimitriou K, Ferreira S, Papandreou NC, Mavrogonatou Rea M, Ross RP, Cotter P & Hill C (2011) Classification of
E, Supply P, Pot B & Tsakalidou E (2012) Complete bacteriocins from gram-positive bacteria. Prokaryotic
genome sequence of the dairy isolate Streptococcus Antimicrobial Peptides (Drider D & Rebuffat S, eds), pp. 29–
macedonicus ACA-DC 198. J Bacteriol 194: 1838–1839. 53. Springer, New York, NY.
Park Y (1994) Hypo-allergenic and therapeutic significance of Richards VP, Lang P, Bitar PD, Lefebure T, Schukken YH,
goat milk. Small Rum Res 14: 151–159. Zadoks RN & Stanhope MJ (2011) Comparative genomics
Perin LM, Moraes PM, Vicßosa GN, Silva J unior A & Nero LA and the role of lateral gene transfer in the evolution of
(2012) Identification of bacteriocinogenic Lactococcus bovine adapted Streptococcus agalactiae. Infect Genet Evol 11:
isolates from raw milk and cheese capable of producing 1263–1275.
nisin A and nisin Z. Int Dairy J 25: 46–51. Roop RM, Bellaire BH, Valderas MW & Cardelli JA (2004)
Pirisi A, Lauret A & Dubeuf J (2007) Basic and incentive Adaptation of the brucellae to their intracellular niche. Mol
payments for goat and sheep milk in relation to quality. Microbiol 52: 621–630.
Small Rum Res 68: 167–178. Rosengren A, Fabricius A, Guss B, Sylven S & Lindqvist R
Premalatha M & Dhasarathan P (2011) Probiotic action of (2010) Occurrence of foodborne pathogens and
Lactobacillus isolated from the milk sample against some characterization of Staphylococcus aureus in cheese produced
human pathogens. Res J Immunol 4: 31–37. on farm-dairies. Int J Food Microbiol 144: 263–269.
Quigley L, O’Sullivan O, Beresford TP, Ross RP, Fitzgerald GF Ruaro A, Andrighetto C, Torriani S & Lombardi A (2012)
& Cotter PD (2011) Molecular approaches to analysing the Biodiversity and characterization of indigenous
microbial composition of raw milk and raw milk cheese. Int coagulase-negative staphylococci isolated from raw milk and
J Food Microbiol 150: 81–94. cheese of North Italy. Food Microbiol 34: 106–111.
Quigley L, O’Sullivan O, Beresford TP, Ross RP, Fitzgerald GF Saaverda JM (2002) Probiotic agents: clinical applications in
& Cotter PD (2012) High-throughput sequencing for infants and children. Infant Formula: Closer to the Reference
detection of subpopulations of bacteria not previously (Raiha N & Rubaltelli FF, eds), Lippincott Williams &
associated with artisanal cheeses. Appl Environ Microbiol 78: Wilkins, Philadelphia (PA). Nestle Nutrition Workshop
5717–5723. Series. Pediatric Program Vol 47 Supplement.
Quigley L, O’Sullivan O, Beresford TP, Ross RP, Fitzgerald GF Sacristan N, Gonzalez L, Castro JM, Fresno JM & Tornadijo
& Cotter PD (2013) The microbial content of raw and ME (2012) Technological characterization of Geotrichum
pasteurised cow’s milk as determined by molecular candidum strains isolated from a traditional Spanish goats’
approaches. J Dairy Sci 96: 1–10. milk cheese. Food Microbiol 30: 260–266.
Raats D, Offek M, Minz D & Halpern M (2011) Molecular Sanchez JI, Martinez B & Rodriguez A (2005) Rational
analysis of bacterial communities in raw cow milk and the selection of Leuconostoc strains for mixed starters based on
impact of refrigeration on its structure and dynamics. Food the physiological biodiversity found in raw milk
Microbiol 28: 465–471. fermentations. Int J Food Microbiol 105: 377–387.
Sanchez JI, Rossetti L, Martinez B, Rodriguez A & Giraffa G Swenson JM, Facklam RR & Thornsberry C (1990)
(2006) Application of reverse transcriptase PCR-based Antimicrobial susceptibility of vancomycin-resistant
T-RFLP to perform semi-quantitative analysis of Leuconostoc, Pediococcus, and Lactobacillus species.
metabolically active bacteria in dairy fermentations. J Antimicrob Agents Chemother 34: 543–549.
Microbiol Meth 65: 268–277. Tamagnini L, De Sousa G, Gonzalez R & Budde C (2006)
Santarelli M, Gatti M, Lazzi C, Bernini V, Zapparoli GA & Microbiological characteristics of Crottin goat cheese made
Neviani E (2008) Whey starter for Grana Padano cheese: in different seasons. Small Rum Res 66: 175–180.
effect of technological parameters on viability and Teh KH, Flint S, Palmer J, Lindsay D, Andrewes P & Bremer P
composition of the microbial community. J Dairy Sci 91: (2011) Thermo-resistant enzyme-producing bacteria isolated
883–891. from the internal surfaces of raw milk tankers. Int Dairy J
Sarno E, Santoro AM, Di Palo R & Costanzo N (2012) 21: 742–747.

Microbiological quality of raw donkey milk from Campania Thaker H, Brahmbhatt M, Nayak J & Thaker HC (2013)
Region. Ital J Anim Sci 11: e49. Isolation and identification of Staphylococcus aureus from
Schiemann D & Toma S (1978) Isolation of Yersinia enterocolitica milk and milk products and their drug resistance patterns in
from raw milk. Appl Environ Microbiol 35: 54–58. Anand, Gujarat. Vet World 6: 10–13.
Schr€oder J, Maus I, Trost E & Tauch A (2011) Complete genome Thoen C, LoBue P & de Kantor I (2006) The importance of
sequence of Corynebacterium variabile DSM 44702 isolated Mycobacterium bovis as a zoonosis. Vet Microbiol 112:
from the surface of smear-ripened cheeses and insights into 339–345.
cheese ripening and flavor generation. BMC Genomics 12: 545. Tilburg JJHC, Roest HJIJ, Nabuurs-Franssen MH, Horrevorts
Schr€oder J, Glaub A, Schneider J, Trost E & Tauch A (2012) AM & Klaassen CHW (2012) Genotyping reveals the
Draft genome sequence of Corynebacterium bovis DSM presence of a predominant genotype of Coxiella burnetii in
20582, which causes clinical mastitis in dairy cows. consumer milk products. J Clin Microbiol 50: 2156–2158.
J Bacteriol 194: 4437. Todhunter DA, Smith KL & Hogan JS (1995) Environmental
Shaked T, Hantsis-Zacharov E & Halpern M (2009) streptococcal intramammary infections of the bovine
Epilithonimonas lactis sp. nov., isolated from raw cow milk. mammary gland. J Dairy Sci 78: 2366–2374.
Int J Syst Evol Microbiol 60: 675–679. Tolle A (1980) The microflora of the udder. Bull Int Dairy Fed
Siezen RJ, Renckens B, Van Swam I, Peters S, Van Kranenburg 120: pp. 4.
R, Kleerebezem M & De Vos WM (2005) Complete Torriani S, Zapparoli G & Dellaglio F (1999) Use of
sequences of four plasmids of Lactococcus lactis subsp. PCR-based methods for rapid differentiation of Lactobacillus
cremoris SK11 reveal extensive adaptation to the dairy delbrueckii subsp bulgaricus and L-delbrueckii subsp lactis.
environment. Appl Environ Microbiol 71: 8371–8382. Appl Environ Microbiol 65: 4351–4356.
Silvestre D, Lopez MC, March L, Plaza A & Martinez-Costa C Tsakalidou E & Odos I (2012) Microbiota of Goat’s Milk and
(2006) Bactericidal activity of human milk: stability during Goat’s Milk Cheese. Proceedings of First Asia Dairy Goat
storage. Br J Biomed Sci 63: 59–62. Conference, pp. 40–41. University Putra Malaysia and the
Slana I, Paolicchi F, Janstova B, Navratilova P & Pavlik I Food and Agricultural Organisation on the United Nations
(2008) Detection methods for Mycobacterium avium subsp (FAO), Kuala Lumpur, Malaysia.
paratuberculosis in milk and milk products: a review. Vet Vacheyrou M, Normand A-C, Guyot P, Cassagne C, Piarroux
Med (Praha) 53: 283. R & Bouton Y (2011) Cultivable microbial communities in
Slattery L, O’Callaghan J, Fitzgerald GF, Beresford T & Ross raw cow milk and potential transfers from stables of sixteen
RP (2010) Invited review: Lactobacillus helveticus – a French farms. Int J Food Microbiol 146: 253–262.
thermophilic dairy starter related to gut bacteria. J Dairy Sci van de Guchte M, Penaud S, Grimaldi C et al. (2006) The
93: 4435–4454. complete genome sequence of Lactobacillus bulgaricus reveals
Smit G, Smit BA & Engels WJM (2005) Flavour formation by extensive and ongoing reductive evolution. PNAS 103:
lactic acid bacteria and biochemical flavour profiling of 9274–9279.
cheese products. FEMS Microbiol Rev 29: 591–610. Van den Tempel T & Jakobsen M (2000) The technological
Sorhaug T & Stepaniak L (1997) Psychrotrophs and their characteristics of Debaryomyces hansenii and Yarrowia
enzymes in milk and dairy products: quality aspects. Trends lipolytica and their potential as starter cultures for
Food Sci Technol 8: 35–41. production of Danablu. Int Dairy J 10: 263–270.
Stiles ME (1996) Biopreservation by lactic acid bacteria. Anton Vedamuthu ER (1994) The dairy Leuconostoc – use in
Leeuw Int JG 70: 331–345. dairy-products. J Dairy Sci 77: 2725–2737.
Suarez VB & Reinheimer JA (2002) Effectiveness of thermal Vendrell D, Balcazar JL, Ruiz-Zarzuela I, de Blas I, Girones O
treatments and biocides in the inactivation of Argentinian & Muzquiz JL (2006) Lactococcus garvieae in fish: a review.
Lactococcus lactis phages. J Food Prot 65: 1756–1759. Comp Immunol Microbiol Infect Dis 29: 177–198.
Sun Z, Chen X, Wang J et al. (2011) Complete genome Verdier-Metz I, Michel V, Delbes C & Montel MC (2009) Do
sequence of Streptococcus thermophilus strain ND03. milking practices influence the bacterial diversity of raw
J Bacteriol 193: 793–794. milk? Food Microbiol 26: 305–310.
Verdier-Metz I, Gagne G, Bornes S, Monsallier F, Williams AM, Fryer JL & Collins MD (1990) Lactococcus
Veisseire P, Delbes-Paus C & Montel MC (2012) Cow piscium sp-nov a new Lactococcus-species from salmonid
teat skin, a potential source of diverse microbial fish. FEMS Microbiol Lett 68: 109–114.
populations for cheese production. Appl Environ Microbiol Wouters JTM, Ayad EHE, Hugenholtz J & Smit G (2002)
78: 326–333. Microbes from raw milk for fermented dairy products. Int
Verreault D, Gendron L, Rousseau GM et al. (2011) Detection Dairy J 12: 91–109.
of airborne lactococcal bacteriophages in cheese Yabrir B, Hakem A, Laoun A, Labiad M, Nazek E-G, Hamadi
manufacturing plants. Appl Environ Microbiol 77: 491–497. A & Abderrahmane M (2013) Does the aridity of Algerian
Villani F, Aponte M, Blaiotta G, Mauriello G, Pepe O & Steppe affect the Ewe’s raw milk quality? Bull Univ Agric Sci
Moschetti G (2001) Detection and characterization of Vet Med Cluj Napoca 69: 1–9.
a bacteriocin, garviecin L1-5, produced by Lactococcus Yu J, Wang W, Menghe B et al. (2011) Diversity of lactic acid

garvieae isolated from raw cow’s milk. J Appl Microbiol 90: bacteria associated with traditional fermented dairy products
430–439. in Mongolia. J Dairy Sci 94: 3229–3241.
Voulgari K, Hatzikamari M, Delepoglou A, Georgakopoulos P, Zhang H, Xu J, Wang J, Sun T, Li H & Guo M (2008a) A
Litopoulou-Tzanetaki E & Tzanetakis N (2010) Antifungal survey on chemical and microbiological composition of
activity of non-starter lactic acid bacteria isolates from dairy kurut, naturally fermented yak milk from Qinghai in China.
products. Food Control 21: 136–142. Food Control 19: 578–586.
Walther C, Rossano A, Thomann A & Perreten V (2008) Zhang X-Y, Zhao L, Jiang L, Dong M-L & Ren F-Z (2008b)
Antibiotic resistance in Lactococcus species from bovine The antimicrobial activity of donkey milk and its microflora
milk: presence of a mutated multidrug transporter mdt (A) changes during storage. Food Control 19: 1191–1195.
gene in susceptible Lactococcus garvieae strains. Vet Zhao WJ, Chen YF, Sun ZH et al. (2011) Complete genome
Microbiol 131: 348–357. sequence of Lactobacillus helveticus H10. J Bacteriol 193:
Ward PN, Holden MTG, Leigh JA et al. (2009) Evidence for 2666–2667.
niche adaptation in the genome of the bovine pathogen Zivkovic AM, German JB, Lebrilla CB & Mills DA (2011)
Streptococcus uberis. BMC Genomics 10: 54. Human milk glycobiome and its impact on the infant
Watanabe K, Fujimoto J, Sasamoto M, Dugersuren J, gastrointestinal microbiota. PNAS 108: 4653–4658.
Tumursuh T & Demberel S (2008) Diversity of lactic acid
bacteria and yeasts in Airag and Tarag, traditional
fermented milk products of Mongolia. World J Microbiol Supporting Information
Biotechnol 24: 1313–1325.
Wegmann U, O’Connell-Motherway M, Zomer A et al. (2007) Additional Supporting Information may be found in the
Complete genome sequence of the prototype lactic acid online version of this article:
bacterium Lactococcus lactis subsp. cremoris MG1363.
Table S1. Representative nutritional content of four
J Bacteriol 189: 3256–3270.
major milk types consumed throughout the world.
West PA, Hewitt JH & Murphy OM (1979) Influence of
methods of collection and storage on the bacteriology of
human milk. J Appl Bacteriol 46: 269–277.

The Complex Microbiota of Raw Milk

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The complex microbiota of raw milk

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frequently include Pseudomonas and Acinetobacter spp.

Sources Role/ Significance

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Culture-dependent* Culture-independent† Next-generation sequencing‡

Acinetobacter species/johnsonii/junii/ Acinetobacter species/johnsonii/ Acinetobacter species

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Culture-dependent* Culture-independent† Next-generation sequencing‡

Dermacoccus species Bradyrhizobium species Carnobacterium species

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(Alegria et al., 2012; Table 3). As this is a naturally fermented

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Milk source Culture-dependent* Culture-independent† Next-generation sequencing‡

Sheep Milk§ Lactococcus lactis Lactococcus lactis/garvieae/raffinolactis Lactococcus

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Box 1. Human milk

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present, including a number of genes involved in fermen-

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Arora DK, Mukerji KG & Marth EH (1991) Handbook of Applied

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