MICROBIAL GROWTH

Veterinary MedicineBrawijaya University

The Requirements for Growth

The growth of a population is an increase in the number of cells

The requirements for microbial growth are both physical and chemical

Physical Requirements

Temperature Psychrophiles (cold loving) Mesophiles (moderate-temperature loving) Thermophiles (heat-loving)

pH Most bacteria grow best at pH 6.5 - 7.5 Acidophiles tolerant to acidity

Osmotic pressure They require water for growth and made up of

80-90% water In hypertonic solution,most microbes undergo

plasmolysis In hypotonic solution wrinkle Halophiles, can tolerate high salt concentrations

Chemical Requirements

Carbon Carbon is the structural back-bone of living

matter Half the dry weight of bacterial cell is carbon Chemoheterotrophs, use an organic molecule Autotroph (chemoautotroph or photoautotroph)

derive their carbon from carbon dioxide

Nitrogen Nitrogen is needed for protein and nucleic acid

synthesis Nitrogen can be obtained from the

decomposition of proteins or from NH4+ or NO3

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A few bacteria are capable of nitrogen (N2) fixation

Chemical RequirementsOxygen On the basis of oxygen requirements, organisms

are classified as obligate aerobes facultative anaerobes aerotolerant anaerobes microaerophilic capnophilic Aerobes, facultative anaerobes, and aerotolerant

anaerobes must have the enzymes superoxide dismutase (2O2

- + 2H+ O2 + H2O2) and either catalase (2H2O2 2H2O + O2) or peroxidase (H2O2 + 2H2 2H2O)

Other chemicals required for microbial growth include

sulfur, phosphorus, trace elements, and, for some organisms, organic growth factors

Culture Media

A culture medium is any material prepared for the growth of bacteria in a laboratory

Microbes that grow and multiply in or on a culture medium known as a culture

- Agar is a common solidifying agent for a culture medium - Broth medium is a liquid medium - Semisolid medium

Culture MediaTYPE PURPOSE

Chemically defined media Growth of chemoautotrophs andphotoautotrophs, and microbiolo-gical assay

Complex media, made up ofnutrient such as extract fromyeast, meat, or digest of proteinexp: nutrient agar or nutrient broth

Most heterotrophic bacteria and fungi are routinely grown on complex media

Reducing media Cultivation of anaerobic bacteriaContain ingredients, such as sodium thioglycholate, that chemically combine with dissolved oxygen and depleted the oxygen in the culture medium

TYPE PURPOSE

Differential mediaMcConkey, Urease medium, Salmonella Shigella Agar (SSA), Blood Agar Plate (BAP)

Differentiation of colonies of desired microbes from others

Selective mediaBismuth Sulfite Agar (BSA), Eosin Methylene Blue (EMB), Thiosulphate Citrate Bile Salt Sucrose(TCBS)

Suppression of unwanted microbes; encouraging desired microbes

Enrichment mediaSelenite brothAlkali Pepton Water (APW)

Similar to selective media but designed to increase numbers of desired microbes to detectable level

Enriched mediaBAP, Chocolate Agar Plate (CAP),Brain Heart Infusion (BHI)

For fastidious bacteria

Transport mediaCary-Blair (V. cholerae)Amis (Nesseria gonorrhoe)

It was needed if the specimens was taken from far places

The Growth of Bacterial CultureBacterial Division Bacteria normally reproduce by binary fission,

in which a single cell divides into two identical cells

Fungi reproduce by budding, aerial spore formation, or fragmentation

Bacterial division occurs according to a logarithmic progression : 2 cells 4 cells 8 cells etc.

Generation Time The time required for a cell to divide or a

population to double is known as the generation time

Most bacteria have generation time 1 – 3 hours; other require more than 24 hours per generation

Pathogenic Mycobacterium require > 8 weeks

Binary Fission in Bacteria

Phases of Growth

Phases of Growth

During the lag phase, there is little or no change in the number of cells, but metabolic activity is high

During the log phase (exponential growth phase), bacteria multiply at the fastest rate possible under the conditions provided

During the stationary phase, there is equilibrium between cell division and death

During the death phase, the number of deaths exceeds the number of new cells formed

Direct Measurement of Microbial Growth A standard plate count reflects the number

of viable microbes and assumes that each bacterium grows into a single colony; plate count are reported as number of colony forming units (CFU)

In filtration, bacteria are retained on the surface of a membrane filter and then transferred to a culture medium to grow and subsequently be counted

The most probable number (MPN) method can be used for microbes that will grow in a liquid medium; it is statistically estimation

In a direct microscopic count, the microbes in a measured volume of a bacterial suspension are counted with the use of a specially designed slide Petroff-Hausser cell counter

Estimating Bacterial Numbers by Indirect Methods

Turbidity A spectrophotometer is used to determine

turbidity by measuring the amount of light that passes through a suspension of cells

McFarland standard

Metabolic Activity Acid production or oxygen consumption

Dry Weight For filamentous organisms such as fungi,

measuring dry weight is a convenient method of growth measurement

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