PHYSICAL FACTORS THAT AFFECT MICROBIAL GROWTH

TEMPERATURE

Generally,an increase in temperature will increase enzyme activity. But if temperatures get too high, enzyme activity will diminish and the protein (the enzyme) will denature.

On the other hand, lowering temperature will decrease enzyme activity. At freezing temperatures enzyme activity can stop. Repeated cycles of freezing and thawing can denature proteins. In addition, freezing causes water to expand and also forms ice crystals, hence cells begin to rupture.

Every bacterial species has specific growth temperature requirements which is largely determined by the temperature requirements of its enzymes. Each organism will have:

• a MINIMUM GROWTH TEMPERATURE,
• an OPTIMUM GROWTH TEMPERATURE,
• and a MAXIMUM GROWTH TEMPERATURE.

Organisms can be classified according to their optimum growth temperature.:

• PSYCHROPHILES grow best between -5^oC and 20^oC,
• MESOPHILES grow best between 20^oC and 45^oC and
• THERMOPHILES grow best at temperatures above 45^oC.
• THERMODURIC organisms can survive high temperatures but don't grow well at such temperatures. Organisms which form endospores would be considered thermoduric.

Some organisms have exotic temperature requirements. Thermus aquaticus is a bright orange gram negative rod isolated from hot water and steam vents at Yellowstone Park. This organism grows best at temperatures between 70-75^oC (158-167^oF). Some of its unique enzymes are in demand for molecular biological and industrial applications.

OXYGEN

Microbes display a great diversity in their ability to use and to tolerate oxygen. In part this is because of the paradoxical nature of oxygen which can be both toxic and essential to life.

• OBLIGATE AEROBES rely on aerobic respiration for ATP and they therefore use oxygen as the terminal electron acceptor in the electron transport chain. Pseudomonas is an example of this group of organisms.
• MICROAEROPHILES require O₂ for growth but they are damaged by normal atmospheric levels of oxygen and they don't have efficient ways to neutralize the toxic forms of oxygen such as superoxide (O_2-) and hydrogen peroxide (H₂O₂). The Streptococci are examples of this group.
• OBLIGATE ANAEROBES will die in the presence of oxygen because they lack enzymes like superoxide dismutase and catalase. Superoxide dismutase catalyzes the following reaction:

  2O₂^- + 2H⁺ ----> H₂O₂ + O₂

  and catalase catalyzes:

  2H₂O₂ ---> 2H₂O + O₂

  Organisms like Clostridium, metabolize through fermentation and / or anaerobic respiration.
• AEROTOLERANT organisms like Lactobacillus ferment and therefore do not use oxygen, however they do tolerate it.
• FACULTATIVE ANAEROBES are the most adaptable. They are capable of both fermentation and aerobic respiration. Escherichia coli is an example of this class of organisms.

Oxygen levels can be manipulated in the laboratory in several ways:

• 1. The CO₂ jar or CANDLE JAR lowers O₂ levels below 10% (from 20%) and raises CO₂ levels from 0.03% to about 10%. In addition to lowered oxygen levels, some organisms flourish better in an enriched CO₂ environment. Such organisms are called CAPNOPHILIC.

   

• 2. The ANAEROBE JAR uses hydrogen gas and a palladium catalyst to convert oxygen into water. An indicator strip impregnated with methylene blue is used to indicate when reducing conditions (anaerobic conditions) have been achieved. When reduced, methylene blue is colorless, when oxidized this dye is blue.

   

• 3. THIOGLYCOLLATE is a chemical reducing agent often put into broth media to eliminate free oxygen.

Representative Anaerobic Pathogens:

1. Clostridium tetani - agent of tetanus, puncture wounds, produces a toxin which enters the spinal column and blocks the inhibitory spinal motor neurons. This produces generalized muscle spasms or spastic paralysis. The muscle of the jaw are often the first affected, hence the name LOCKJAW.

2. Clostridium botulinum - this soil organism is the causative agent of botulism which typically occurs after eating home canned alkaline vegetables which were not heated enough during canning. The neurotoxin blocks transmission across neuromuscular junctions and this results in flaccid paralysis.

3. Clostridium perfringes and Clostridium sporogenes - these organisms are associated with invasive infections known as GAS GANGRENE.

4. Clostridium difficile - the causative agent of pseudomembranous colitis, a side effect of antibiotic treatment which eliminates the normal flora.

The Streptococci - Microaerophiles

These organisms are all catalase negative, therefore the catalase test is useful in identification. They also have distinctive colonial morphology on blood agar which is differential for them. It is important to note if the colonies are alpha, beta, or gamma hemolytic.

1. Group A Streptococcus - Streptococcus pyogenes

This beta hemolytic organism is also bacitracin sensitive. It is the cause of strep throat, rheumatic fever, glomerulonephritis and scarlet fever.

2. Group D Streptococcus - Enterococcus - Streptococcus faecalis

This organism is a normal inhabitant of the large intestine. It is also a frequent cause of bladdder infections

3. Streptococcus pneumoniae.

This organism is a normal inhabitant of the repiratory tract. It is a frequent cause of pneumonia in people who have been compromised by other illness.