4.4 Microbiology

Classifying Bacteria - shape

According to shapeCoccus - plural Cocci

Bacillus - plural Bacilli

Spirillum – plural Spirilli

Classifying Bacteria – Gram Stain

Classifying Bacteria – Gram Positive or Gram Negative

Cell wall gives a bacterium shape and prevents them from bursting osmotically.

Gram Positive Bacteria• Retain the Gram Stain• Have a thick layer of peptidoglycan• Are affected by antibiotics such as Penicillin and

the enzyme lysozyme (in tears) which interfere with the cross linking of peptidoglycan

Gram Negative Bacteria• Do not retain the Gram Stain, only pick up the

counter stain• Have a thinner but more complex 2 layered cell

wall• Very little peptidoglycan but outer layer is another

membrane.

Classifying Bacteria – Gram Stain

Culturing Bacteria - Growth

Lag Phase – Cells adapting to new environment – synthesising new enzymes etc.

Log Phase / Exponential Growth – No limiting factors

Stationary Phase – Limiting factors affecting growth death rate equals birth rate

Death / Decline Phase – All nutrients used, toxin accumulating. Oxygen concentration decreasing.

Culturing Bacteria – Conditions Needed for Growth• Nutrients – Carbon / Nitrogen Sources as

well vitamins and minerals• Water - Solvent for biochemical

reactions, cell expansion• Temperature & pH - Particular to specific species• Oxygen- Obligate aerobes (need O2 all

the time)Obligate anaerobes find oxygen toxic

Facultative anaerobes – grow better in the presence of oxygen

Aseptic Technique• Compete with cultured microbe, may produce toxins and/or

contaminate the product / Protect from pathogenic species

Measuring Bacterial Growth• Turbidity – indirect method (total or viable?)

Measuring Bacterial Growth• Haemocytometer (total or viable?)

May need to dilute beforehand and then factor in the dilution factor.

Total count.

Measuring Bacterial Growth

Plating and Counting Colonies

Fermentation

Culturing of microorganisms both aerobically and anaerobically in fermentation tanks.Metabolism – reactions of living organismsMetabolites – products from the reactionsTwo types of metabolite:• Primary metabolite – produced by an

organism as part of its normal growth• Secondary metabolite – substances produced

by an organism that are not part of its normal growth (usually begins after the log phase)

Industrial Scale Fermenters

Batch Versus ContinuousNutrients added

Product removed

Growth rates

Product quality

Repercussions if contaminated

Penicillin Production• Secondary metabolite• Batch culture• Penicillium chrysogenum or Penicillium

notatum• Fungus – Aerobic / 24oC / pH6.5

• Inhibits peptidoglycan cross links – Cell wall weakened – osmotic bursting