Oxygen requirements of different bacteria
Almost all plants and animals are dependent upon supply of atmospheric oxygen which is unlikely in bacteria. Depending upon the oxygen need in bacteria, they are classified into 4 groups: strict or obligate aerobic that grow only in the presence of oxygen. Facultative anaerobic bacteria grow both in presence and absence of free oxygen. Some bacteria are microaerophilic which grow best in the presence of low concentration of molecular oxygen. However, not all bacteria require oxygen for growth. Strict or obligate anaerobic bacteria can grow only in the absence of oxygen.
Amount of oxygen required is different for growth and for other metabolic activities. Aerobic bacteria requires large surface growth area and exposure to contact available atmospheric oxygen, therefore under in vitro conditions, they are grown on shallow agar plates. Broth culturing of such bacteria is always accompanied by shaking and bubbling by spargers or baffles. Both of these mechanical actions are important during fermentation reactions to increase the availability and consumption of oxygen to growing aerobes. For the cultivation of anaerobic bacteria special techniques are employed which are meant to exclude atmospheric oxygen from growth medium. For this purpose, prereduced growth media contained with reducing agents like thioglycollate, formaldehyde, and sulfoxalate or cysteine hydrochloride which effectively absorb molecular oxygen are employed. Mechanical removal of oxygen by various means from an enclosed vessel containing tubes or plates of inoculated medium is another option. In one method, air is pumped out of vessel and replaced by gases like N2, helium or mixture of CO2 and N2. Burning of a candle to utilize oxygen present in the growth vessel is also one of the simplest ways to create oxygen free atmosphere. Addition of pyrogallol over the plug followed by rubber corking of test tube is used for slant cultivation of anaerobic bacteria. Anaerobic gaspak jars are routinely employed in various laboratories to cultivate anaerobes like Clostridium, Bacteroids and microaerophilic bacteria like Borrelia, Streptococci or Campylobacter.
Role of oxygen: Oxygen is elemental constituent of water and organic compounds. Obligatory aerobic bacteria are dependent on aerobic respiration for fulfillment of their energetic needs; wherein molecular oxygen functions as terminal electron acceptor or oxidising agent. Anaerobic bacteria do not obtain energy by using molecular oxygen. In metabolic terms, facultative anaerobic bacteria can use oxygen as terminal oxidising agent only when available but can also obtain energy in its absence by fermentative reactions such as that in all enterobacteria. Some of them are not sensitive to presence of oxygen and hence have exclusively fermentative energy yielding metabolism; lactic acid bacteria are representative of such fermentative metabolism. Microaerophilic bacteria grow best at oxygen concentration of 0.2atm pressure and possess enzymes that are inactivated during strong oxidising conditions, hence functional only at low oxygen pressure. Oxygen is also co-substrate for oxygenase enzymes that catalyze degradation of aromatic and alkane compounds. Oxygenative cleavage of aromatics is very useful for their dissimilation. Oxidative dissimilation of recalcitrant pollutant aromatics is hence one of the potential requirements for their efficient biodegradation. Oxygenases also mediate sterol and unsaturated fatty acid synthesis. They catalyze direct addition of one or two oxygen atoms to organic substrate compounds.
Oxygen toxicity: The role of nodulation in nitrogen fixing species of Rhizobium strains was understood only when researchers came to know about oxygen toxicity. Biological nitrogen fixation is catalyzed by nitrogenase enzyme system which is very sensitive to the presence of oxygen. It is readily inactivated by presence of molecular oxygen halting the vital process of fixation of atmospheric nitrogen. Similarly, hydrogen utilizing reactions catalyzed by enzyme hydrogenase accompanying the nitrogen fixation are also inhibited by oxygen. If oxygen is found at high concentration greater than atmosphere it can be toxic to aerobic bacteria; oxygenases of aerobes are irreversibly denatured by exposure to oxygen. Therefore, bacteria especially aerobic, possessing oxygen sensitive enzymes have developed special mechanisms to protect functional enzymes from oxygen inactivation. Some mechanisms include high respiration rate, heterocyst formation, and synthesis of exopolysaccharides or nodulation. Enzymes like superoxide dismutase (SOD), peroxidase, and catalase are also used by these bacteria as shield against toxic forms of oxygen like superoxide radical (O2.-), hydrogen peroxide (H2O2) and hydroxyl radical (OH.). Lactic acid bacteria that don't possess catalase degrade H2O2 by peroxidase to H2O. Lethal accumulation of superoxide is prevented by SOD which catalyses its conversion to O2 and H2O2. Oxidations of flavoproteins by O2 results in the formation of toxic compounds like superoxide radicals. Oxygen is also toxic to anaerobic bacteria as they do not contain SOD (very little if present) or catalase; therefore they have adapted to fermentative metabolism whereby they avoid presence of oxygen. Oxygen in its nascent or singlet state (1O2) is very toxic and powerful oxidant. Toxic singlet state is generated when triplet state of photosensitizer reacts with oxygen during photo-oxidation, a process similar to the production of superoxide radical. Its toxicity is enhanced in presence of light and photosensitive pigments or photosensitizers. Carotenoid pigments quench this form of oxygen and protect the cell from photo oxidative death. The chlorophylls are powerful photosensitizers and hence carotenoids are always present alongwith chlorophylls. Nonphotosynthetic aerobic bacteria like Micrococcus and Serratia also produce carotenoids in cell membrane to nullify the effect of photosensitizer cytochromes during their growth in high light intensified environment. Knowledge of oxygen requirements of bacteria is critical factor not only for their cultivation and classification but it also represents ecological significance.
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