Going Back To The Basics: Understanding Microbes in Food

25/07/2018
Going Back to the Basics:

Understanding Microbes
in Food
Jun Barnes
Scientific Affairs & Education Manager
3M Philippines Food Safety
jbarnes@mmm.com
What makes
Microbiology
Microbes
relevant as a
What comes
into your field of
mind? Science?
25
© July
3M 2018 . All Rights Reserved. 3M Confidential. 15
What is Microbiology?
Study of organisms and agents too small to be seen clearly by the As a Basic Microbiology uses and develops tools for
unaided eye (Prescott, 2005) probing the fundamental processes of life
Biological
Size = micrometer (µm) Includes specialties such as:
Science
• Microbial physiology
• Microbial genetics
• Microbial ecology
1
25/07/2018
Microbiology is at the center of many

important aspects of human and
veterinary medicine, agriculture, and As an
industry (Brock et al, 2012) Applied
• Industrial microbiology
Biological Prokaryotic Eukaryotic
• Pharmaceutical microbiology
Science
• Medical microbiology
• Dairy microbiology
• Food microbiology
Microorganisms are diverse…
Prokaryotic Eukaryotic
Do we
test for all Molds
of them in
food?
Bacteria Yeasts
2
25/07/2018
Rhizopus
Common bread mold

Spoilage of many foods including berries, fruits vegetables, etc.
Molds Tempeh
General Characteristics
Filamentous fungi
Fuzzy/ cottony growth
Aerobic
Sometimes pigmented/colored
Molds- Importance in Food Penicillium
Causes spoilage Soft rots in fruits

Produces toxins Ripening of cheese (P. roqueforti- blue cheese)
Source of industrially important metabolites Some can produce mycotoxins
enzymes
antibiotics
Production of food ingredients
Used as animal feed
3
25/07/2018
Penicillin: 15 September 1928 Yeasts
General Characteristics
• Non-filamentous
• Unicellular
• Ovoid or spherical
• Reproduces asexually by
budding or fission
https://www.historychannel.com.au/articles/penicillin-is-discovered/
Aspergillus Yeasts- Importance in Foods
Yellow to green to black rots of fruits and meats • Involved in the manufacture of foods such
A. niger produces industrially important enzymes such as as bread, beer, wines, vinegar and surface-
β-galactosidase, invertase, lipase and peptinase) ripened cheese and as source of enzymes
Some can produce mycotoxins
• Causes spoilage of sauerkraut, fruit juices,
syrups, honey, jellies, meats, wine, beer and
other foods
Aflatoxin Saccharomyces
• Produced in foods by molds, including Aspergillus flavus and A. Food and beverage production
parasiticus Employed in many food industries with many strains that can be used
• High-level aflatoxin exposure produces an acute hepatic necrosis, in leavening of bread, manufacture of wines, production of alcohol,
resulting later in cirrhosis, or carcinoma of the liver. glycerol and invertase
• Aflatoxins are toxic and among the most carcinogenic substances
known (Hudler, G. W. (1998). Magical Mushrooms, Mischievous Molds: The Remarkable Story of
the Fungus Kingdom and Its Impact on Human Affairs. Princeton University Press)
4
25/07/2018
Bacteria Major Groups of Bacteria
General Characteristics • Gram-positive bacteria

• Gram-negative bacteria
• Mainly unicellular What cell
• Prokaryotic structure
differentiates
G+ & G-
bacteria?
Some Forms of Bacteria Gram + or Gram -?
Neisseria gonnorhoeae Streptococcus pyogenes Staphylococcus aureus Bacillus megaterium
A B
Vibrio Escherichia coli Campylobacter jejuni Helicobacter pylori
Some Bacteria of Industrial Importance Gram Positive & Gram Negative Bacteria
Gram Staining
Developed by Christian Gram in 1888
Differential staining procedure

Staphylococcus aureus Listeria monocytogenes
• Differentiates one species/group of bacteria from another
species/group of bacteria
Escherichia coli Salmonella typhi
5
25/07/2018
Gram Staining Microorganisms Grow Fast…
Primary Stain
Mordant
What are the
reagents
used in Gram
Staining?
Gram + Bacteria Binary Division
Appear violet to purple under the 1 to 2 to 4 to 8 to ?

microscope after Gram-staining
Above: Clostridium perfringens

Below: Staphylococcus aureus
Gram - Bacteria Binary Division
Appear red to pink under the

microscope after Gram-staining
Log Growth
Above: Escherichia coli

Below: Neisseria gonnorheae
25
© July
6
25/07/2018
Bacterial Growth Curve

Lag
• Adapt to nutrients
Log
• Active growth
Stationary
• Death = Growth rate
Death
• Nutrients consumed
• pH too low (why?)
Bacterial Growth and Nutrition
Bacterial nutrition and culture media √

F ood
Chemical and physical factors affecting growth √
What nourishes us may also support microorganisms…
The nature of bacterial growth
http://diverge.hunter.cuny.edu/~weigang/Images/0611_binaryfission_1.jpg
25
© July
Factors Affecting Growth
Control Conditions = Prevent Growth Nutrients

• Macronutrients: needed in larger amounts
• Needed in large quantities: CHONPS
• Carbon, hydrogen, oxygen, nitrogen, phosphorous, and sulfur. H and O are common. Sources of C, N, P, and S must also be provided.
FAT–TOM
• Macronutrients needed in smaller amounts:
• Mineral salts such as Ca+2, Fe+3, Mg+2, K+
• Micronutrients = trace elements;

• needed in very tiny amounts; e.g. Zn+2, Mo+2, Mn+2
• Growth Factors
• Required in small amounts by cells because they fulfill specific roles in biosynthesis.
• The need for a growth factor results from either a blocked or missing metabolic pathway in the cells.
• amino acids: required for the synthesis of proteins
• vitamins: needed as coenzymes and functional groups of certain enzymes
7
25/07/2018
Element % dry wgt Source

Carbon 50 organic compounds or CO2
Oxygen 20 H2O, organic compounds, CO2, and O2
Nitrogen 14 NH3, NO3, organic compounds, N2
Hydrogen 8 H2O, organic compounds, H2

Phosphorus 3 inorganic phosphates (PO4)
Sulfur 1 SO4, H2S, So, organic sulfur compounds
Potassium 1 Potassium salts

Magnesium 0.5 Magnesium salts
Calcium 0.5 Calcium salts
Iron 0.2 Iron salts
http://textbookofbacteriology.net/nutgro.html
25
© July
A cidity T ime
pH indicates the level of acidity
or alkalinity
pH
T emperature
• Lowest = 0 (very acid); highest = 14 (very basic); neutral is pH 7.
• Acidophiles/acidotolerant grow at low pH
• Alkalophiles/alkalotolerant grow at high pH
• Most bacteria prefer a neutral pH
8
25/07/2018
Temperature
O xygen
Aerobes: use oxygen in metabolism; obligate.
• Low temperature
Microaerophiles: require oxygen (also obligate), but in small
• Enzymatic reactions too slow; enzymes too stiff
amounts.
• Lipid membranes no longer fluid
Obligate (strict) anaerobes: killed or inhibited by oxygen.
• High temperature
• Enzymes denature, lose shape and stop functioning
Aerotolerant anaerobes: do not use oxygen, but not killed by it.
• Lipid membranes get too fluid, leak Facultative anaerobes: can grow with or without oxygen
• DNA denatures
Classification of Bacteria Based on Temperature
25
© July
Terms Related to Temperature

Special cases:
M oisture
Water Activity (Aw)
• Psychrotrophs: bacteria that grow at “normal” (mesophilic)
temperatures (e.g. room temperature” but can also grow in the • The availability of water in solution or a substance
refrigerator; responsible for food spoilage.
• Thermoduric: more to do with survival than growth; bacteria that can
withstand brief heat treatments.
25
© July
9
25/07/2018
Properties of the
storage
Intrinsic Extrinsic
environment that
Parameters Parameters
affect both the
foods and the
microorganisms
therein
Water Intrinsic Parameters Extrinsic Parameters

• Structural barriers • Temperature of storage
• Halophiles/halotolerant: relationship to high salt. • Hydrogen ion concentration
• Atmospheric condition of
• Marine bacteria (pH)
storage
• Osmophiles: can stand hypertonic environments whether salt, • Moisture content (Aw)
• Gasses
sugar, or other dissolved solutes
• Nutrient content
• Fungi very good at this • Relative humidity (%R.H.)
• Antimicrobial constituents
• Xerotolerant: dry. Subject to desiccation. • Presence of other organisms
• Biological Structures
• Fungi best
• Survival of bacterial endospores.
Microorganisms are everywhere…
Inherent part
of animal Intrinsic
& Extrinsic
Parameters
plant the Parameters
tissues
10
25/07/2018
Foodborne Illness
Food infection
If microbes are
everywhere, • Presence of bacteria or other microbes which infect the body after
including food consumption
and the Why still
environment Bacteria in food ► Bacteria continues to ► Illness
test for grow in intestine
microbes? • Example: Salmonella, Campylobacter, hemorrhagic E. coli, Listeria
Why still test for microbes in food & the environment? Foodborne Illness
Food intoxication
• Ingestion of toxins contained within food

• Can happen even when the microbe that produced the toxin is no
longer present or able to cause infection
Toxins in food ► Ingestion ► Illness
• Example: Staphyloccoccus aureus, Clostridium botulinum
Microbes in Food Manifests Their Presence

in One of Several Ways
Transform
food • Fermentation
properties in
a beneficial
way
• Changes in
color, texture,
Safety Spoilage Quality
Spoilage
smell, taste, etc.
• Foodborne
Foodborne infection
Illness
• Foodborne
intoxication
25
© July
3M 2018 . All Rights Reserved. 3M Confidential. 83 25
© July
11
25/07/2018
Indicator Test
A test of food quality

• Examples: APC/SPC, yeasts, molds, Enterobacteriaceae
Purpose:
Foodborne
Safety Illness
Quality • Not to determine the species of microorganism in food, but rather to
determine if there are any bacteria or fungi in the foods
Implications:
• The presence of indicators in given numbers means manufacturing
facility failed to comply with GMPs
• Depending on the indicator organism, it may suggest that the
environment is capable of harboring a pathogen
25
© July
Types of Microbiological Tests Pathogen Test
• Enumeration “How much is there?” A test of food safety

• Example: Salmonella, Listeria monocytogenes
• Detection “Is something there?”-
Presence/Absence Purpose:
• Identification “What is there?” – Provides a name • To determine if a pathogen is present.
• genus, species
Implication:
• Characterization • The presence of pathogens in given numbers (sometimes as little as
• strains (subspecies level) 1 colony forming organism in 25 grams) means the food is unsafe to
• microflora (ecology of food) consume.
Food Testing Group:
There are two areas of food testing that processors focus on: 1. Indicator Tests
1. Indicator testing 2. Pathogen Tests
2. Pathogen testing Group 3. Environmental Monitoring
Discussion
Discuss the parameters you are testing.
Methods used.
Challenges.
12
25/07/2018
Aseptic Techniques Isolated Colonies
Procedures used by microbiologists to prevent:

• Contamination of the specimen/pure cultures
• Infection of the personnel
• Contamination of the environment
Challenge
Generally, viable counts are

made on samples that contain
Indicator significant numbers of bacteria
Tests
Failure to form isolated colonies
Failure to quantify colonies
Measure of Growth Solution
Viability Count Serial Dilution

• Determination of the number of living cells (able to reproduce) • Thinning out of microbial population to achieve a population that is
present in a suspension countable
• Can be accomplished by plating • Diluents: sterile buffered peptone water
• Form isolated colonies Visible cluster sterile saline solution (0.85% NaCl)
of cells on a
sterile phosphate buffer
solid medium
derived from a sterile 0.1% peptone water
single parent
cell
13
25/07/2018
Assumptions
Dilutions to
assist Control • Each cell will form a visible colony
interpretation
• The sample is homogeneous
• There are no aggregates or clumps of cells
-3 -2
-6 -5 -4
Example of Dilution Scheme CFU/mL
11mL/11g 1mL 0.1mL CFU = colony-forming unit
CFU/mL = Average number of colonies x D.F.

Liquid/ Solid Sample volume plated
Homogenates
D.F. = dilution factor
99mL 9mL 9.9mL
= reciprocal of the dilution or 1
? ? ?
dilution
In each transfer, there is a dilution of the microbial
population from the original sample. How diluted is
your sample?
Dilution Example
Dilution (D) = volume transferred x previous dilution

total volume Dilution Bacterial Colony
Counts
11mL/11g 10-1 TNTC, TNTC
Example: D= 11 x 100
11 + 99 10-2 400, 375
D= 11 x 1 10-4 35, 40
110
Liquid/ Solid Sample
Homogenates
If the volume plated is 1.0 mL, what is the CFU/mL of
D= 1 or 10-1 the sample?
10
99mL
?
14
25/07/2018
Solution Solution
To choose the dilution to use for the computation of CFU/mL, consider the dilution CFU/mL = ∑n x lower D.F.
which gave colony counts that fall within: [(n1 x 1) + (n2 x 0.1)]
• 25-250 colonies for bacteria volume plated
• 15-150 for molds
CFU/mL = 245 + 28 + 30 X 101
In the example, since only 10-4 gave colony counts that fall within 25-250, use [ (1 x 1) + (2 x 0.1)
this dilution in computing for CFU/mL. 1 mL
CFU/mL = 2.52 x 103
Solution Aerobic Plate Count
CFU/mL = Average number of colonies x D.F. • Indicator of bacterial populations on a sample

volume plated • Limitations:
CFU/mL = 35 + 40 X 104 • Does not represent the entire bacterial population but only
those capable of growing in a particular physical environment,
2 nutrients supplied to the medium, etc.
1 mL • Does not differentiate types of bacteria
CFU/mL = 3.75 X 105 • Importance:
• Gauge sanitary quality
• Adherence to GMPs
• Indicator of safety
What if two dilutions gave counts falling within the range? Coliform Definitions
Enterobacteriaceae
Salmonella
Yersinia
Shigella
Coliform is not a taxonomic
Total Coliform
Proteus
Serratia
classification but rather a
Dilution Bacterial Colony work definition used to
Counts describe a group of:
Citrobacter Thermotolerant Coliform
10-1 260, 245 Enterobacter - Gram-negative
Hafnia
Klebsiella
10-2 28, 30 Serratia
- facultative anaerobic
GUD+
GUD - rod shaped bacteria
10-3 0, 0 Inducible
E. coli
- ferment lactose to
produce acid and gas
Anaerogenic Anaerogenic
E. coli GUD + E. coli GUD - within 48 h at 35°C. (FDA
Pathogenic
BAM and CMMEF)
E. coli
25
© July
15
25/07/2018
Coliform Definitions E. coli as a quality indicator

TYPICAL E. coli TYPICAL E. coli
β-glucuronidase positive β-glucuronidase positive (inducible)
Coliforms are bacteria which, at the 87-97% of strains 91-96% of strains
specified temperature (i.e., 30°C or

37°C, as agreed) form characteristic
colonies in crystal violet neutral red
bile lactose agar and which in the Existing methods: VRBL with MUG, 3M Petrifilm EC/CC Plate Existing methods: TBX with inducer, 3M Petrifilm SEC Plate
confirmation test cause
fermentation of lactose with the ATYPICAL E. coli ATYPICAL E. coli
Anaerogenic β-glucuronidase positive β-glucuronidase negative; ferment lactose to gas
production of gas under the test Do not produce gas from lactose
conditions specified in this
International Standard. (ISO 4832) Existing methods: VRBL with MUG, 3M Petrifilm EC/CC Plate
Existing methods: None

25
© July
© July
Coliform Definitions Why test for coliforms?

Enterobacteriaceae
In 1914, the US Public Health Service adopted the enumeration of coliforms as a
Salmonella
Yersinia - Fecal coliform is a subset of total convenient standard of sanitary significance.
Shigella Total Coliform
Proteus coliforms that grows and
Serratia
ferments lactose at elevated
The greatest application of coliform and E. coli testing is in assessing the overall
incubation temperature,
temperature hence
Citrobacter Thermotolerant Coliform quality of a food and the hygienic conditions present during food processing. For
Enterobacter also referred to as thermotolerant
Hafnia example, enumeration of these organisms in heat-pasteurized foods can be used to
Klebsiella coliforms.
Serratia
assess the adequacy of a heating process. (CMMEF)
GUD - Consists mostly of E. coli but
GUD+
Inducible some other enterics such as
E. coli Klebsiella, Enterobacter, and It is important to note that most classical methods
Anaerogenic Anaerogenic Citrobacter freundii can also for coliforms and E. coli are targeted at “typical”
E. coli GUD + E. coli GUD -
ferment lactose at these organisms. (CMMEF)
Pathogenic
E. coli temperatures.
25
© July
© July
Escherichia coli Enterobacteriaceae
• Common inhabitants of the intestinal tract of animals and man

• Presence in food/water indicates fecal contamination
Nowadays, there has been a trend to monitor
for Enterobacteriaceae because of their
increasing relevance to health problems
16
25/07/2018
Enterobacteriaceae Enterobacteriaceae
Examining an environment for the presence of Enterobacteriaceae

Potential instead of coliforms may give a better and more accurate
Enterobacteriaceae
indicator indication of the likelihood of the presence of pathogens.
organisms
Coliforms
Enterobacteriaceae are excellent sanitation indicators,
particularly in dry food processing environments where coliform
Fecal Coliforms
organisms may not survive, but where non-coliform pathogenic
organisms could survive
Enterobacteriaceae commonly colonize both wet and dry factory
E. coli environments
Concern: fecal
contamination
25
© July
Enterobacteriaceae Staphylococcus aureus
• The family Enterobacteriaceae is widely used as an indicator of Contamination of foods by Staphylococcus aureus
hygiene, sanitation and food safety because this group of usually results after process or cooking
organism includes: • Mishandling, improper refrigeration
• Intestinal pathogens
• Important food spoilage agents Foods subjected to post-process contamination with enterotoxigenic
• Has long been a convention in European food processing plants types of S. aureus represent a significant hazard because of the
as a general indicator for sanitation. absence of competitive organisms.
• It is becoming an accepted global standard.
Enterobacteriaceae Staphylococcus aureus
Enterobacteriaceae is a broader category than coliforms and includes:

Staphylococcal Intoxication
• Escherichia spp.
• Citrobacter spp.
• Enterobacter spp.
Staphyloccoccus
aureus
► Multiplication in food ► Toxin production
• Proteus spp.
• Salmonella spp. Enterotoxin
• Shigella spp. Enterobacter cloacae Proteus vulgaris • Not detectable by taste or smell
Citrobacter diversus
• Yersinia spp. • Heat resistant
• Also produced by S. intermedius and S. hyicus
Shigella Salmonella typhi Yersinia pestis
17
25/07/2018
Characteristics of Staphylococcus aureus

The Staphylococcus aureus Toxin
Facultatively anaerobic There are 11 forms of the ENTEROTOXIN (Staph Enterotoxin, SE).
SE most associated with food poisoning is type A (62.5% of
Gram-positive coccus outbreaks caused by S.aureus), with the next most common C and B
less frequently involved.
Catalase positive Heat treatment and sanitisers used in the food industry generally are
good means of inactivating S.aureus vegetative cells.
• Can produce the enzyme "catalase" and able to The SE can be inactivated by heating to 121oC for 3-8min
convert H2O2 to water and oxygen
Primarily coagulase-positive
• Can produce "coagulase", a protein product,
which is an enzyme that causes clot formation
25
© July
. 126
Sources of Staphylococcus aureus Symptoms of Staphylococcal Intoxication
Contamination from food handlers At 5000 cells → produce toxins
• Found on the skin and in the nose an throat of most people (about 40%)
Abdominal cramps
• People with colds and sinus infections
Vomiting
• Infected wounds, pimples, boils, and acne
Severe diarrhea
Exhaustion Symptoms occur within 1-8 hours
after eating contaminated food
Duration: 1-2 days
The illness is seldom fatal
. .
Sources of Staphylococcus aureus Precautions to Prevent Staph Toxin Production
In raw foods Keep S. aureus from growing

Keep food clean
• Raw milk and unpasteurized milk may contain large numbers of S. aureus as a
Keep food hot (above 60°C/140°F) or cold (below 4°C/40 °F) during serving time
result of staphylococcal mastitis
Refrigerate or freeze leftovers and foods to be served later.
• Staphylococcal contamination of hides, feathers and skins is common
. .
18
25/07/2018
Fungi Culture Media
• Eukaryotic A medium used for the growth & cultivation of microorganisms

• Heterotrophic (Nonphotosynthetic) organisms Uses:
• Characterized by a chitinous cell wall • Microbial enumeration
• Unicellular or Multicellular (Filamentous) • Isolation and maintenance of pure cultures of microorganisms
• Identification of bacteria according to their biochemical and
physiological properties
Nature of Fungi Pour Plating
• Versatile environmental requirements

• Acid/alkaline requirement for growth is quite broad, ranging from pH Cover and mix
thoroughly by
2 to above pH 9 gentle tilting
and swirling
the dish
• Temperature range (10-35°C) is also broad, with a few species
Inoculate labeled empty petri dish Pour 15 mL of melted agar
capable of growth below or above this range (45°C) into the inoculated petri
dish
Count, record, calculate Invert (except for fungi) Allow the agar to completely gel
and incubate
Why are fungi such a threat in many industries? Spread Plating
• Cause various degrees of deterioration

and decomposition of foods
Inoculate diluted
• May cause abnormal flavors and odors sample on

prepoured &
solidified agar
• Some produce toxic metabolites known as

mycotoxins. Sterilize L-rod by dipping in Cool L-rod & spread inoculated sample
alcohol & flaming
• Certain foodborne molds and yeasts may
also elicit allergic reactions or may cause
infections
Count, record, calculate Invert (except for fungi)

and incubate
19
25/07/2018
Productive, fast & easy

What are some
• Increase productivity of lab staff by an average of 80.5%
challenges with
using traditional Media preparation • Eliminate in-house agar preparation; save labour, energy and QC
agar?
• Compact size = cost savings in storage & equipment requirements
Pouring plates
• Easy to inoculate and interpret (a built-in grid facilitates counting)
Washing & sterilizing tubes • Save energy and reduce waste in the laboratory
• Reduce testing variability
Confirming results
Decontaminating loads of waste
25
© July
And 34 years later, the Petrifilm Plate portfolio

continues to grow
Rapid Yeast and Mold

Count Plate
Rapid Aerobic Count

Plate
Rapid E. coli/Coliform
25
© July
Count Plate
The Petrifilm Plate portfolio grew Forms of 3M™ Petrifilm™ Plates
Petrifilm without foam dam/barrier

- Aerobic Count - High Sensitivity Coliform Count
- Yeast & Mold Count
E.coli/Coliform
Yeast
Coliform
Aerobic
and
Rapid E.Mold
Coliform
Enterobacteriace
High-Sensitivity
Environmental
Staph
SelectExpress
coli
ae Count
Count
Count
Coliform
Listeria Plate
CountPlate
Count Plate
Plate
Plate
Count
Count
Count
Count Plate
Plate
Plate
Plate Petrifilm with foam dam/barrier
- Coliform Count - Rapid Coliform Count
- E. coli/Coliform Count - Rapid E. coli/Coliform Count
- Enterobacteriaceae Count - Rapid Yeast & Mold*
- Staph Express Count - Rapid Aerobic Count
- Environmental Listeria - Salmonella Express System
25
© July
20
25/07/2018
The Bad Bugs…

Who Salmonella L. monocytogenes E.coli O157:H7 S. aureus Campylobacter
Widespread Widespread ruminants Widespread soil, water, farm

intestines Produce animals,
including Enterotoxins domesticated
Reservoirs
cattle, goats, (SET) animals, birds,
Pathogen sheep, deer,

filth flies
rodents, sewage
Tests What
pathogens Raw meats, poultry,
eggs, milk and dairy
Raw and
pasteurized milk,
Undercooked or
raw ground
salads, cream-
filled bakery
raw poultry -
chicken, turkey,
are you
products, fish,
cheese*, ice beef, alfalfa items, deli meat, duck, goose,
What Food?
shrimp, frog legs,
yeast, coconut, cream, raw sprouts, milk and dairy game fowl - meat
sauces and salad vegetables, unpasteurized and juices, water,
fermented raw- fruit juices, dry- raw milk,
testing in
dressing, cake
mixes, cream-filled meat, raw and cured meat, undercooked
desserts and cooked poultry, lettuce, game meats such as
toppings, dried
raw meats, raw meat, and beef, pork, lamb,
the lab.?
gelatin, peanut
butter, cocoa, and and smoked fish cheese curds and occasionally
chocolate shellfish
For more information refer to the Bad Bug Book
The Bad Bugs…

Who Salmonella L. monocytogenes E.coli O157:H7 S. aureus Campylobacter
Nausea, Septicemia, Severe Nausea, Bloody diarrhea,

vomiting, meningitis, cramping and vomiting, nausea,
abdominal encephalitis, watery, bloody, abdominal periodontitis or
cramps, diarrhea, spontaneous diarrhea, cramps, dysentery
Symptoms
fever, and abortion or vomiting, fever prostration and syndrome,

headache. Onset stillbirth, influenza- diarrhea cramps, fever
12-72 hours after like symptoms and pain.
exposure. including fever., Worst cases:
Chronic nausea, vomiting, hemolytic uremic
consequences -- and diarrhea syndrome and
arthritic thrombotic
symptoms may thrombocytopeni
follow 3-4 weeks c purpura
after onset of
acute symptoms
Infective dose
As few as 15-20 Unknown but is Unknown, but Toxin dose of About 400-500
cells; depends believed to vary. It from a less than 1.0 μg. bacteria may
upon age and is safe to assume compilation of This toxin level is cause illness in
health of host, that, fewer than outbreak data, reached when S. some individuals,
10 “Pinaka” Significant and strain
differences
1,000 total
organisms may
the dose may
be as few as 10
aureus
populations
while in others,
greater numbers
Foodborne Pathogens
cause disease organisms exceed 100,000 are required
per gram
For more information refer to the Bad Bug Book

.
"Ten Least Wanted Pathogens“(CDC) Pathogen Testing Trends
Two approaches can be used as part of a comprehensive

microbiological testing program
• Use of microbiological indicator testing
• Testing for specific pathogens or spoilage organisms in the product or the
environment
Campylobacter Clostridium botulinum E. coli O157:H7 Listeria monocytogenes Norovirus
Many operations monitor the processing environment rather than
relying only on finished product testing.
• Implementation of a Pathogen Environmental Monitoring Program (PEMP)
Salmonella Staphylococcus aureus Shigella Toxoplasma gondii Vibrio vulnificus • Provides dynamic information on the state of a processing line and area
• Identify microbial niches or critical points
• Trend analysis and measurement of sanitation conditions or practices
21
25/07/2018
• Low prevalence of pathogen in

sample
What are the • Low number of pathogen in sample
challenges in • Non–homogenous distribution of
pathogen in sample
pathogen
• Possibility of matrix interference
testing?
• Low regulatory detection limit 1
CFU/25 g
• Sensitivity of the method selected
Current Challenges in Detection

Time to result
Cost
Expertise required for testing
• Interpretation of colony morphology on plating media
False positives
• Citrobacter that are phenotypically very similar to Salmonella
• Selective media will not necessarily other microorganisms with similar features
to the one we are interested in
False negatives
• Due to low bacterial numbers or due to bacteria that behave atypically
25
© July
3M 2018 . All Rights Reserved. 163
Thank you
22

Going Back To The Basics: Understanding Microbes in Food

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Going Back To The Basics: Understanding Microbes in Food

Uploaded by

Copyright:

Available Formats

25/07/2018

Going Back to the Basics:

Microbiology is at the center of many

Microorganisms are diverse…

Common bread mold

Molds- Importance in Food Penicillium

Causes spoilage Soft rots in fruits

Penicillin: 15 September 1928 Yeasts

Aspergillus Yeasts- Importance in Foods

Bacteria Major Groups of Bacteria

General Characteristics • Gram-positive bacteria

Some Forms of Bacteria Gram + or Gram -?

Neisseria gonnorhoeae Streptococcus pyogenes Staphylococcus aureus Bacillus megaterium

Differential staining procedure

Escherichia coli Salmonella typhi

Gram Staining Microorganisms Grow Fast…

Gram + Bacteria Binary Division

Appear violet to purple under the 1 to 2 to 4 to 8 to ?

Above: Clostridium perfringens

Gram - Bacteria Binary Division

Appear red to pink under the

Above: Escherichia coli

Bacterial Growth Curve

Bacterial Growth and Nutrition

Bacterial nutrition and culture media √

Factors Affecting Growth

Control Conditions = Prevent Growth Nutrients

• Micronutrients = trace elements;

Element % dry wgt Source

Oxygen 20 H2O, organic compounds, CO2, and O2

Nitrogen 14 NH3, NO3, organic compounds, N2

Hydrogen 8 H2O, organic compounds, H2

Sulfur 1 SO4, H2S, So, organic sulfur compounds

Potassium 1 Potassium salts

Factors Affecting Growth

Factors Affecting Growth

Classification of Bacteria Based on Temperature

Terms Related to Temperature

Factors Affecting Growth

Water Intrinsic Parameters Extrinsic Parameters

Microorganisms are everywhere…

microbes? • Example: Salmonella, Campylobacter, hemorrhagic E. coli, Listeria

• Ingestion of toxins contained within food

Toxins in food ► Ingestion ► Illness

• Example: Staphyloccoccus aureus, Clostridium botulinum

Microbes in Food Manifests Their Presence

A test of food quality

Types of Microbiological Tests Pathogen Test

• Enumeration “How much is there?” A test of food safety

Food Testing Group:

Aseptic Techniques Isolated Colonies

Procedures used by microbiologists to prevent:

Generally, viable counts are

Failure to quantify colonies

Measure of Growth Solution

Viability Count Serial Dilution

Example of Dilution Scheme CFU/mL

11mL/11g 1mL 0.1mL CFU = colony-forming unit

CFU/mL = Average number of colonies x D.F.

Dilution (D) = volume transferred x previous dilution

CFU/mL = 2.52 x 103

Solution Aerobic Plate Count

CFU/mL = Average number of colonies x D.F. • Indicator of bacterial populations on a sample