Effect of oxygen on the growth and distribution of microorganisms

 Microorganisms being unicellular and poikilothermic, respond to variations in environment. The growth of microorganisms affected by their surroundings. So, an understanding of environmental factors is crucial in the control of microbial growth and in the study of the ecological distribution of microorganisms. 
    Oxygen concentration has a critical role in the growth and distribution of microorganisms also. Almost all multicellular organisms are completely dependent on atmospheric O₂ for growth— that is, they are obligate aerobes. In case of microorganisms, based on their oxygen requirement or the lack of it, microorganisms fall under five groups namely, aerobes, anaerobes, facultative anaerobes, aerotolerant anaerobes and microaerophiles. 
    An organism able to grow in the presence of atmospheric O₂ is an aerobe, whereas one that can grow in its absence is an anaerobe. 

    Facultative anaerobes such as E.coli, do not require O₂ for growth but do grow better in its presence. In the presence of oxygen, they will use aerobic respiration. 

    Aerotolerant anaerobes such as Enterococcus faecalis simply ignore O₂ and grow equally well whether it is present or not. 

    In contrast, strict or obligate anaerobes (e.g., Bacteroides, Fusobacterium, Clostridium pasteurianum, Methanococcus) do not tolerate O₂ at all and die in its presence. 

    Finally, there are organisms such as Campylobacter, called microaerophiles, that are damaged by the normal atmospheric level of O₂ (20%) and require O₂ levels below the range of 2 to 10% for growth. 

Campylobacter is a capnophile- require high levels of CO₂ for optimum growth 

    A microbial group may show more than one type of relationship to O₂. All five types are found among the procaryotes and protozoa. Fungi are normally aerobic, but a number of species— particularly among the yeasts—are facultative anaerobes. Algae are almost always obligate aerobes. 

    The oxygen requirement of an organism is related to the type of metabolism that it is using to generate ATP which is the energy currency. Energy (ATP) generation is linked to the movement of electrons through the electron transport chain (ETC), where the final electron acceptor can be oxygen or a non-oxygen molecule. Oxygen serves as the terminal electron acceptor for the electron-transport chain in aerobic respiration. 

    Aerobes which use oxygen as the final electron acceptor use aerobic respiration for energy generation. Obligate anaerobes use various organic and even inorganic materials as electron acceptors - - use fermentation or anaerobic respiration pathways for energy generation - do not require oxygen. Aerotolerant and strict anaerobes cannot generate energy through respiration and must employ fermentation or anaerobic respiration pathways for this purpose. 

    Facultative anaerobes have both sets of enzymes for aerobic and anaerobic respiration and can grow in both aerobic and anaerobic environments. This gives these organisms considerable flexibility and is an ecological advantage.

Based on the effect of oxygen on growth, microorganisms occupy different regions when grown  in a culture tube, as demonstrated in the figure.

The different relationships with O₂ appear due to 

•  the inactivation of proteins 
•  the effect of toxic O₂ derivatives.

Enzymes can be inactivated when sensitive groups like sulfhydryls are oxidized. An example is the nitrogen-fixation enzyme nitrogenase, which is very oxygen sensitive.

Oxygen accepts electrons and is readily reduced because its two outer orbital electrons are unpaired. Flavoproteins, several other cell constituents, and radiation promote oxygen reduction. The result is usually some combination of the reduction products superoxide radical, hydrogen peroxide, and hydroxyl radical.

O₂ + e– → O₂ ^– (superoxide radical)

O₂ ^– + e^– + 2H⁺ → H₂O₂ (hydrogen peroxide)

H₂O₂ + e^– + H⁺ → H₂O + OH^– (hydroxyl radical)

These reactive oxygen species (ROS) are extremely toxic because they are powerful oxidizing agents and rapidly destroy cellular constituents. 

Neutrophils and macrophages use these toxic oxygen products to destroy invading pathogens.

A microorganism must be able to protect itself against such oxygen products or it will be killed. Many microorganisms possess enzymes that afford protection against toxic O₂ products.

Obligate aerobes and facultative anaerobes usually contain the enzymes superoxide dismutase (SOD) and catalase, which catalyze the destruction of superoxide radical and hydrogen peroxide, respectively. Peroxidase also can be used to destroy hydrogen peroxide.

2O₂ ^– + 2H+ O₂ → H₂O₂ (superoxide dismutase)

 2H₂O₂ → 2H₂O + O₂ (catalase)

H₂O₂ + NADH + H⁺ → 2H₂O + NAD (peroxidase)

Aerotolerant microorganisms may lack catalase but almost always have superoxide dismutase. The aerotolerant Lactobacillus plantarum uses manganous ions instead of superoxide dismutase to destroy the superoxide radical.

All strict anaerobes lack both enzymes or have them in very low concentrations and therefore cannot tolerate O₂. 

Although strict anaerobes are killed by O₂, they may be recovered from habitats that appear to be aerobic. In such cases they associate with facultative anaerobes that use up the available O₂ and thus make the growth of strict anaerobes possible. For example, the strict anaerobe Bacteroides gingivalis lives in the mouth where it grows in the anaerobic crevices around the teeth.

Different approaches must be used when growing aerobes and anaerobes since aerobes need O₂ and anaerobes are killed by O₂. When culturing aerobic microorganisms, either the culture vessel is shaken to aerate the medium or sterile air must be pumped through the culture vessel.

With anaerobes, all O₂ must be excluded using

(1) Special anaerobic media containing reducing agents such as thioglycollate or cysteine may be used. The reducing agents will eliminate any dissolved O₂ remaining within the medium so that anaerobes can grow beneath its surface.

(2) The medium is boiled during preparation to dissolve its components; boiling also drives off oxygen very effectively.

(3) Oxygen also may be eliminated from an anaerobic system by removing air with a vacuum pump and flushing out residual O₂ with nitrogen gas or CO₂. Many anaerobes require a small amount of CO₂ for best growth.

(4) One of the most popular ways of culturing small numbers of anaerobes is by use of a GasPak/Gas generator envelope. Water is added to chemicals in envelope to generate Hydrogen and carbon dioxide. Carbon dioxide promotes more rapid growth of microorganisms. The palladium catalyst catalyzes the formation of water from hydrogen and oxygen, thereby removing oxygen if at all it is present.

(5) Plastic bags or pouches can be used when only a few samples are to be incubated anaerobically. These have a catalyst and calcium carbonate to produce an anaerobic, carbon-dioxide rich atmosphere. A special solution is added to the pouch’s reagent compartment; petri dishes or other containers are placed. Anaerobic indicator strip Methylene blue becomes colorless in absence of O₂.

 Reference: Prescott's Microbiology