NUT 551

FOOD SAFETY AND HYGIENE

FOOD MICROBIOLOGY-2

F. C. MILLS-ROBERTSON (PhD)
0208970091
 MICROBIAL GROWTH RESPONSE IN FOOD ENVIRONMENT
• Microbes grow or multiply in numbers when exposed to a favourable
environment such as food

• Growth is associated with food spoilage, food-borne diseases and food bio-
processing

• In general, under optimum conditions of growth, bacteria have the shortest

generation time, followed by yeasts and moulds

• Also, among bacterial species and strains, generation time under optimum
conditions varies greatly; some species, such as Vibrio parahaemolyticus,
under optimum conditions can have a generation time as low as 10 to 12 min
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D
• During their growth cycles, microorganisms undergo reproduction many
times, causing the numbers in the population to increase dramatically

• In fungi, unicellular algae, and protozoa, reproduction involves a duplication

of the nucleus through the asexual process of mitosis and a splitting of the cell
in cytokinesis

• Bacteria reproduce by the asexual process of binary fission

• One of the remarkable attributes of bacteria is the relatively short generation

time, the time required for a microbial population to double in numbers

• Escherichia coli, for example, has a generation time of about 20 minutes when
dividing under optimal conditions
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

• The growth of a bacterial population can be expressed in various phases of a
growth curve

• The logarithms of the actual numbers in the population are plotted in the growth
curve along the side axis, and the time is plotted at the base

• Four phases of growth are recognized in the growth curve

✓Lag phase
✓Log phase
✓Stationary phase
✓Decline/death phase
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D
❑The Growth Curve
▪ Lag Phase
• When bacteria are inoculated into a new medium, reproduction does not start
immediately

• The population remains at the same number as the bacteria become accustomed to
their new environment

• This period can last from 1 hour to several days

• The period depends on kind of bacteria, the age of culture and the available
nutrients in the new medium

• If the organisms are young and actively growing, the lag phase is very short,
because the cells do not have to equilibrate
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

▪ Lag Phase
• If the new medium is different from the old one, the lag is long, because the cells
need to get acclimatized to the new environment

• For spore formers, the lag phase is long because the spores have to germinate
first

• Organisms from old cultures may be so depleted of enzymes or metabolic

intermediates that the full growth cannot be obtained until optimal
concentrations of these substances are restored by synthesis
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

▪ Log Phase
• The log phase or logarithmic phase is the phase of exponential increase where
bacterial growth occurs at its optimal level and the population doubles rapidly

• The typical characteristics of active cells can be observed at this stage where the
cells divide at a steady/constant rate; and the generation time is constant

• This continues until nutrient supply becomes limiting

• A plot of the logarithm of the numbers of cells against time is a straight line
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

▪ Log Phase
• The generation time varies with species of organism, available nutrients and
temperature of incubation

• pH and O2 availability for aerobes can also influence the generation time

• The shape of the slope of log phase is affected by growth conditions such as
temperature, pH, osmotic pressure

• Bacterial cells can be maintained in the log phase by continuously adding fresh
medium of the same constitution
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

▪ Log Phase
• At any point in time during the log phase some bacteria are just beginning to
divide, others are half-finished and still others have finished

• Certain techniques have been used by researchers to make all the cells divide
simultaneously, known as the synchronous growth

• This synchrony lasts for only a few generation since the daughter cells soon go out
of phase

• Some of the techniques involve chilling for a period followed by bringing the
culture back to their growth temperature
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

▪ Log Phase
• During chilling, the organisms metabolize slowly but do not divide

• When the temperature is raised the cells undergo synchronized division

• In ordinary liquid cultures the log phase lasts for a relatively short time

• When the organisms are numerous their activities begin to have an effect on the
medium, that is, nutrients become exhausted or toxic metabolites accumulate

• As the medium becomes less favourable for growth the rate decreases, leading to
the stationary phase
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D
❑The Growth Curve
▪ Stationary Phase
• The reproduction of bacterial cells is offset by their death, and the population
reaches a plateau

• The reasons for bacterial death include the accumulation of waste, the lack of
nutrients, and the unfavorable environmental conditions that may have developed

• At this stage, endospores develop spore-formers

• If the conditions are not altered, the population will enter its decline, or death
phase (Figure 3)
 MICROBIAL GROWTH RESPONSE IN FOOD CONT’D

❑The Growth Curve

▪ Decline/Death Phase
• When the death rate exceeds the growth rate the actual number of live bacteria
decreases

• Thus, the bacteria die off rapidly, the curve turns downward, and the last cell in the
population soon dies

• Important factors contributing to the death of cells are the depletion of nutrients
and accumulation of inhibitory substances such as acids thereby decreasing the pH

• The number of viable cells also decreases geometrically and they die at different
rates and this may take hours, weeks or months to reach the end of this phase
Figure 3. A growth curve of a bacterial population showing the four major phases of the curve
 NATURE OF MICROBIAL GROWTH IN FOOD

• Normally, food harbours a mixed population of microorganisms that can include

different species and strains of bacteria, yeasts and moulds

• Some species can be present in relatively higher numbers than others

• The growth characteristics of a mixed population differ in several respects from

that of a pure culture (a single strain of a species)

• Depending on the environment, which includes both the food environment

(intrinsic) and the environment in which the food is stored (extrinsic), some of the
species or strains can be in optimum or near-optimum growth condition
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D
• Because of rapid rate of growth during storage, such strains will outnumber the
others and become predominant

• This can occur even if they are initially present in low numbers and this is often the
case in foods kept for a long time under a specific condition, such as at refrigerated
temperature

• If the product is enumerated initially, it may show that the majority of microbial
populations is able to grow at 35 0C and only a few grow at 4 0C (refrigerated
temperature)

• If the product is enumerated after a few weeks of refrigerated storage, usually

populations that grow at 4 0C outnumber those that grow at 35 0C
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

• Another situation can arise if a food contains, among the mixed population, two
species initially present in equal numbers and both growing optimally under the
specific intrinsic and extrinsic environments of the food, but one having a shorter
generation time than the other

• After a storage period, the one with shorter generation time becomes predominant

• Many foods most often get spoiled by bacteria than by yeasts and moulds because,
in general, bacteria have shorter generation time

• Thus, in a mixed population, the intrinsic and extrinsic environments dictate which
one, two, or a few in the initial mixed population will become predominant and
produce specific changes in a food
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑INTRINSIC FACTORS
• Intrinsic factors that affect the rate of proliferation relate more to the characteristics
of the substrate (foodstuff) that support the growth of microorganisms

• Intrinsic factors of a food include

✓Nutrients
✓Growth factors and inhibitors (or anti-microbials)
✓Water activity
✓pH
✓Oxidation–reduction potential
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Intrinsic Factors
▪ Nutrients
• Microbial growth is accomplished through the synthesis of cellular components and
energy

• Most microbes need external sources of nitrogen, energy (carbohydrates, proteins,

or lipids), minerals and vitamins to support their growth

• Nitrogen is usually obtained from amino acids and non protein nitrogen sources

• Moulds are the most effective in the utilization of proteins, complex carbohydrates
and lipids because they contain enzymes capable of hydrolyzing these molecules
into less complex components
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D
❑Intrinsic Factors
▪ Nutrients
• All microorganisms need minerals but requirement for vitamins vary

• Moulds and some bacteria can synthesize enough B vitamins for their needs,
whereas other microorganisms require a ready-made supply

• In general, meat is rich in protein, lipids, minerals, and vitamins but poor in
carbohydrates

• Foods from plant sources are rich in carbohydrates but can be poor sources of
proteins, minerals, and some vitamins

• Some foods such as milk and many prepared foods have all five nutrient groups in
sufficient amounts for microbial growth
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Intrinsic Factors
✓Growth factors and inhibitors (or anti-microbials)
• Foods can also have some factors that either stimulate growth or adversely affect
growth of microorganisms

• The exact nature of growth factors is not known, but they are naturally present in
some foods

• An example is the growth factors in tomatoes that stimulate growth of some

Lactobacillus species

• These factors can be added to raw materials during food bio-processing or to media
to isolate some fastidious bacteria from foods
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Intrinsic Factors
✓Growth factors and inhibitors (or anti-microbials)
• Foods also contain many chemicals, either naturally or added that adversely affect
microbial growth

• Some of the natural inhibitors are lysozyme in egg, agglutinin in milk, and eugenol
in cloves

• The inhibitors, depending on their mode of action, can prevent or reduce growth or
may inhibit the growth of microorganisms
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Intrinsic Factors
▪ Water Activity (Aw)
• Water activity (Aw) is a measure of the availability of water for biological functions
and relates to water present in a food in free form

• In a food system, total water or moisture is present in free and bound forms

• Bound water is the fraction used to hydrate hydrophilic molecules and to dissolve
solutes, and is not available for biological functions

• Thus, it does not contribute to water activity (Aw)

 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Intrinsic Factors
▪ Water Activity (Aw)
• The free water in a food is necessary for microbial growth

• A reduction of water availability will reduce microbial proliferation

• It is necessary to transport nutrients and remove waste materials, carry out

enzymatic reactions, synthesize cellular materials, and take part in other
biochemical reactions, such as hydrolysis of a polymer to monomers (proteins to
amino acids)

• Each microbial species (or group) has an optimum, maximum, and minimum Aw
level for growth
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Intrinsic Factors
▪ Water Activity (Aw)
• The approximate optimal water activity for the growth of many microorganism is
0.99 and most bacteria require Aw higher than 0.91 for growth

• Most natural food products have Aw of approximately 0.99

• Most spoilage bacteria do not grow at Aw below 0.91, but moulds and yeasts can
grow at Aw of 0.80 or lower

• Moulds and yeasts can grow on partially dehydrated surfaces (including food),
whereas bacterial growth is retarded
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D
❑Intrinsic Factors
▪ Oxidation Reduction Potential
• Oxidation reduction potential is an indication of the oxidizing and reducing power
of the substrate

• Aerobes grow more rapidly under a high oxidation reduction potential while a low
oxidation reduction potential favours the growth of anaerobes

• Facultative microorganisms are capable of growth under either condition

• Some microbes can alter the oxidation reduction potential of food to the extent
that the activity of other microorganism is restricted

• For example, anaerobes can decrease the oxidation reduction potential to such a
low level that the growth of aerobes can be inhibited
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑EXTRINSIC FACTORS
• Extrinsic factors important in microbial growth in a food include the environmental
conditions in which the food is stored

• Extrinsic factors of a food environmental include

✓Temperature
✓Relative humidity
✓Gaseous environment
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Extrinsic Factors
▪ Temperature
• Microbial growth is accomplished through enzymatic reactions

• It is well known that within a certain range, with every 10 0C rise in temperature,
the catalytic rate of an enzyme doubles

• Similarly, the enzymatic reaction rate is reduced to half by decreasing the

temperature by 10 0C

• Thus, because temperature influences enzyme reactions, it has an important role in

microbial growth in food
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D
❑Extrinsic Factors
▪ Temperature
• Foods are exposed to different temperatures from the time of production until
consumption

• Depending on processing conditions, a food can be exposed to high heat, from 65 0C

(roasting of meat) to more than 100 0C (in ultra high temperature processing)

• For long-term storage, a food can be kept at 5 0C (refrigeration) to–20 0C or below

(freezing)

• Some relatively stable foods are also kept between 10 and 35 0C (cold to ambient
temperature)
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Extrinsic Factors
▪ Temperature
• Some ready-to-eat foods are kept at warm temperature (50 0C to 60 0C) for several
hours (such as, in the supermarket deli)

• Different temperatures are also used to stimulate desirable microbial growth in

food fermentation

• When the foods are exposed to temperatures beyond the maximum and minimum
temperatures of growth, microbial cells die rapidly at higher temperatures and
relatively slowly at lower temperatures

• Microbial growth and viability are important considerations in reducing food

spoilage and enhancing safety against pathogens, as well as in food bio-processing
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Extrinsic Factors
▪ Gaseous Environment
• Some microorganisms have an absolute requirement for oxygen, others grow in the
total absence of oxygen and others grow either with or without available oxygen

• Microorganisms that require free oxygen are called aerobic microorganisms

(example Pseudomonas species)

• Those that thrive in the absence of oxygen are called anaerobic microorganisms
(such as, Clostridium species)

• Microorganisms that grow with or without the presence of free oxygen are called
facultative microorganisms (example Lactobacillus species)
 NATURE OF MICROBIAL GROWTH IN FOOD CONT’D

❑Extrinsic Factors
▪ Relative Humidity
• All microorganisms have high requirement for water to support their growth and
activity

• A high relative humidity can cause moisture condensation on food, equipments,

walls and ceilings which is conducive to microbial growth and spoilage

• Bacteria require a higher humidity than do yeast and moulds

• Optimal relative humidity for bacteria is to be 92% or higher; whereas yeast prefer it
to be 90% or higher

• Moulds thrive more if the relative humidity is 85% to 90%