Microbiological

culture

A microbiological culture, or microbial culture, is a met hod of mult iplying microbial organisms
by let t ing t hem reproduce in predet ermined cult ure medium under cont rolled laborat ory
condit ions. Microbial cult ures are foundat ional and basic diagnost ic met hods used as a research
t ool in molecular biology.

Microbial cultures on solid and liquid media

Microbial cult ures are used t o det ermine t he t ype of organism, it s abundance in t he sample being
t est ed, or bot h. It is one of t he primary diagnost ic met hods of microbiology and used as a t ool t o
det ermine t he cause of infect ious disease by let t ing t he agent mult iply in a predet ermined
medium. For example, a t hroat cult ure is t aken by scraping t he lining of t issue in t he back of t he
t hroat and blot t ing t he sample int o a medium t o be able t o screen for harmful microorganisms,
such as Streptococcus pyogenes, t he causat ive agent of st rep t hroat .[1] Furt hermore, t he t erm
cult ure is more generally used informally t o refer t o "select ively growing" a specific kind of
microorganism in t he lab.

It is oft en essent ial t o isolat e a pure cult ure of microorganisms. A pure (or axenic) cult ure is a
populat ion of cells or mult icellular organisms growing in t he absence of ot her species or t ypes. A
pure cult ure may originat e from a single cell or single organism, in which case t he cells are
genet ic clones of one anot her. For t he purpose of gelling t he microbial cult ure, t he medium of
agarose gel (agar) is used. Agar is a gelat inous subst ance derived from seaweed. A cheap
subst it ut e for agar is guar gum, which can be used for t he isolat ion and maint enance of
t hermophiles.

Bacterial culture

A culture of Bacillus anthracis

There are several t ypes of bact erial cult ure met hods t hat are select ed based on t he agent
being cult ured and t he downst ream use.

Broth cultures
One met hod of bact erial cult ure is liquid cult ure, in which t he desired bact eria are suspended in a
liquid nut rient medium, such as Luria Brot h, in an upright flask. This allows a scient ist t o grow up
large amount s of bact eria for a variet y of downst ream applicat ions.

Liquid cult ures are ideal for preparat ion of an ant imicrobial assay in which t he experiment er
inoculat es liquid brot h wit h bact eria and let s it grow overnight (t hey may use a shaker for uniform
growt h). Then t hey would t ake aliquot s of t he sample t o t est for t he ant imicrobial act ivit y of a
specific drug or prot ein (ant imicrobial pept ides).

Liquid cultures of the cyanobacterium Synechococcus PCC 7002

As an alt ernat ive, t he microbiologist may decide t o use st at ic liquid cult ures. These cult ures are
not shaken and t hey provide t he microbes wit h an oxygen gradient .[2]

Agar plates

Microbiological cult ures can be grown in pet ri dishes of differing sizes t hat have a t hin layer of
agar-based growt h medium. Once t he growt h medium in t he pet ri dish is inoculat ed wit h t he
desired bact eria, t he plat es are incubat ed at t he opt imal t emperat ure for t he growing of t he
select ed bact eria (for example, usually at 37 degrees Celsius, or t he human body t emperat ure,
for cult ures from humans or animals, or lower for environment al cult ures). Aft er t he desired level
of growt h is achieved, agar plat es can be st ored upside down in a refrigerat or for an ext ended
period of t ime t o keep bact eria for fut ure experiment s.

There are a variet y of addit ives t hat can be added t o agar before it is poured int o a plat e and
allowed t o solidify. Some t ypes of bact eria can only grow in t he presence of cert ain addit ives.
This can also be used when creat ing engineered st rains of bact eria t hat cont ain an ant ibiot ic-
resist ance gene. When t he select ed ant ibiot ic is added t o t he agar, only bact erial cells
cont aining t he gene insert conferring resist ance will be able t o grow. This allows t he researcher
t o select only t he colonies t hat were successfully t ransformed.

Agar based dipsticks

Miniat urised version of agar plat es implement ed t o dipst ick format s, eg. Dip Slide, Digit al
Dipst ick [3] show pot ent ial t o be used at t he point -of-care for diagnosis purposes. They have
advant ages over agar plat es since t hey are cost effect ive and t heir operat ion does not require
expert ise or laborat ory environment , which enable t hem t o be used at t he point -of-care.

Stab cultures

Motile and non-motile bacteria can be differentiated along the stab lines. Motile bacteria will grow out from the stab line
while non-motile bacteria are present only along the stab line.

St ab cult ures are similar t o agar plat es, but are formed by solid agar in a t est t ube. Bact eria is
int roduced via an inoculat ion needle or a pipet t e t ip being st abbed int o t he cent er of t he agar.
Bact eria grow in t he punct ured area.[4] St ab cult ures are most commonly used for short -t erm
st orage or shipment of cult ures.

Culture collections

Microbial cult ure collect ions focus on t he acquisit ion, aut hent icat ion, product ion, preservat ion,
cat alogueing and dist ribut ion of viable cult ures of st andard reference microorganisms, cell lines
and ot her mat erials for research in microbial syst emat ics.[5][6] Cult ure collect ion are also
reposit ories of t ype st rains.

Major national culture collections.[5][6]

Collection
Name Location
Acronym

ATCC American Type Cult ure Collect ion Manassas, Virginia

Belgian Co-ordinat ed Collect ions of Decent ralized, Coordinat ion Cell in

BCCM
Micro-organisms Brussels, Belgium

Cult ure Collect ion Universit y of

CCUG Got henburg, Sweden
Got henburg

CECT Colección Española de Cult ivos Tipo Valencia, Spain

CIP Collect ion d'Inst it ut Past eur Paris, France

Deutsche Sammlung von

DSMZ Braunschweig, Germany
Mikroorganismen und Zellkulturen

Int ernat ional Collect ion of

ICMP Auckland, New Zealand
Microorganisms from Plant s

JCM Japan Collect ion of Microorganisms Tsukuba, Ibaraki, Japan

Public Healt h England, London,

NCTC Nat ional Collect ion of Type Cult ures
Unit ed Kingdom

Nat ional Collect ion of Indust rial, Food and

NCIMB Aberdeen, Scot land
Marine Bact eria

Solid plate culture of thermophilic microorganisms

For solid plat e cult ures of t hermophilic microorganisms such as Bacillus acidocaldarius, Bacillus
stearothermophilus, Thermus aquaticus and Thermus thermophilus et c. growing at t emperat ures
of 50 t o 70 degrees C, low acyl clarified gellan gum has been proven t o be t he preferred gelling
agent comparing t o agar for t he count ing or isolat ion or bot h of t he above t hermophilic
bact eria.[7]

Virus and phage culture

Virus or phage cult ures require host cells in which t he virus or phage mult iply. For bact eriophages,
cult ures are grown by infect ing bact erial cells. The phage can t hen be isolat ed from t he result ing
plaques in a lawn of bact eria on a plat e. Virus cult ures are obt ained from t heir appropriat e
eukaryot ic host cells. The st reak plat e met hod is a way t o physically separat e t he microbial
populat ion, and is done by spreading t he inoculat e back and fort h wit h an inoculat ing loop over
t he solid agar plat e. Upon incubat ion, colonies will arise and single cells will have been isolat ed
from t he biomass. Once a microorganism has been isolat ed in pure cult ure, it is necessary t o
preserve it in a viable st at e for furt her st udy and use. St ock cult ures have t o be maint ained, such
t hat t here is no loss of t heir biological, immunological and cult ural charact ers.

Eukaryotic cell culture

Isolation of pure cultures

For single-celled eukaryot es, such as yeast , t he isolat ion of pure cult ures uses t he same
t echniques as for bact erial cult ures. Pure cult ures of mult icellular organisms are oft en more
easily isolat ed by simply picking out a single individual t o init iat e a cult ure. This is a useful
t echnique for pure cult ure of fungi, mult icellular algae, and small met azoa, for example.

Developing pure cult ure t echniques is crucial t o t he observat ion of t he specimen in quest ion.
The most common met hod t o isolat e individual cells and produce a pure cult ure is t o prepare a
st reak plat e. The st reak plat e met hod is a way t o physically separat e t he microbial populat ion,
and is done by spreading t he inoculat e back and fort h wit h an inoculat ing loop over t he solid agar
plat e. Upon incubat ion, colonies will arise and single cells will have been isolat ed from t he
biomass. Once a microorganism has been isolat ed in pure cult ure, it is necessary t o preserve it in
a viable st at e for furt her st udy and use. St ock cult ures have t o be maint ained, such t hat t here is
no loss of t heir biological, immunological and cult ural charact ers.

See also

Colony-forming unit

Blood cult ure

Microbial dark mat t er

Microbial Food Cult ures

Screening cult ures

Sput um cult ure

Synchronous cult ure

Gellan gum

References

1. Healthwise, Incorporated (2010-06-28). "Throat Culture" (http://www.webmd.com/oral-health/throat-c

ulture) . WebMD. Archived (https://web.archive.org/web/20130317050528/http://www.webmd.com/or
al-health/throat-culture) from the original on 2013-03-17. Retrieved 2013-03-10.

2. Old, D.C.; Duguid, J.P. (1970). "Selective Outgrowth of Fimbriate Bacteria in Static Liquid Medium" (http
s://www.ncbi.nlm.nih.gov/pmc/articles/PMC248102) . Journal of Bacteriology. American Society for
Microbiology. 103 (2): 447–456. doi:10.1128/JB.103.2.447-456.1970 (https://doi.org/10.1128%2FJB.1
03.2.447-456.1970) . PMC 248102 (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC248102) .
PMID 4914569 (https://pubmed.ncbi.nlm.nih.gov/4914569) .

3. Iseri, Emre; Biggel, Michael; Goossens, Herman; Moons, Pieter; van der Wijngaart, Wouter (2020). "Digital
dipstick: miniaturized bacteria detection and digital quantification for the point-of-care" (https://doi.org/
10.1039/D0LC00793E) . Lab on a Chip. doi:10.1039/D0LC00793E (https://doi.org/10.1039%2FD0LC0
0793E) . ISSN 1473-0197 (https://www.worldcat.org/issn/1473-0197) .

4. "Addgene: Streaking a Plate from an Addgene Stab Culture" (https://www.addgene.org/recipient-instruct

ions/streak-plate/) . www.addgene.org. Archived (https://web.archive.org/web/20180408232824/htt
p://www.addgene.org/recipient-instructions/streak-plate/) from the original on 8 April 2018. Retrieved
21 March 2018.

5. Madigan, Michael T. (2012). Brock biology of microorganisms (13th ed.). San Francisco: Benjamin
Cummings. ISBN 9780321649638.

6. Uruburu, F. (2003). "History and services of culture collections" (http://roderic.uv.es/bitstream/10550/1

2955/1/10.Uruburu_International%20Microbiology_6_2003.pdf) (PDF). International Microbiology. 6
(2): 101–103. doi:10.1007/s10123-003-0115-2 (https://doi.org/10.1007%2Fs10123-003-0115-2) .
hdl:10550/12955 (https://hdl.handle.net/10550%2F12955) . PMID 12811589 (https://pubmed.ncbi.nl
m.nih.gov/12811589) .

7. Lin, Chi Chung and Casida, L. E. (1984) GELRITE as a Gelling Agent in Media for the Growth of
Thermophilic Microorganisms. Applied and Environmental Microbiology 47, 427-429.

External links

Wikimedia Commons has media relat ed t o Microbiological cultures.

 EFFCA (ht t p://www.effca.org) - European Food and Feed Cult ut es Associat ion. Informat ion
about product ion and uses of microbial cult ures as well as legislat ive aspect s.

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