Environmental factors

Environmental conditions and bacterial growth

The prokaryotes exist in nature under enormous range of physical condition such as O₂concentration, hydrogen ion concentration (pH), temperature, salt concentration and water activity. The exclusive limit of life on the planet is set by the prokaryotes especially the archaea. An understanding of these influences aids in the control of microbial growth and the study of the ecological distribution of microorganisms. Prokaryotes are present or grow anywhere life can exist. The environments in which some prokaryotes grow would kill most other organisms. For example Bacillus infernus is able to live over 1.5 miles below the earth's surface without O₂ and 60^oC temperature. These microorganisms which can thrive and grow in such harsh conditions are often called extremophiles. In this chapter, we will see the relativeness of environmental factors and bacterial growth.

Temperature

Temperature profoundly affects microorganisms as the most important factor influencing the effect is temperature sensitivity of enzyme-catalyzed reactions. Beyond a certain point of higher temperature, slow growth takes place and damages the microorganisms by denaturing enzymes, transport carriers and other proteins. The plasma membrane also is disrupted as lipid bilayer simply melts and the damage is such an extent that it cannot be repaired. At very low temperature, membranes solidify and enzymes don't work rapidly. In summary, when organisms are above their optimum temperature, both the function and cell structure is affected at low temperature, function is affected. The cardinal temperatures vary greatly between microorganisms. For example, microorganisms have been found growing in virtually all environments where there is liquid water, regardless of its temperature. In 1966, Professor Thomas D. Brock at Indiana University made the amazing discovery in boiling hot springs of Yellowstone National Park that bacteria were not just surviving there, they were growing and flourishing. Boiling temperature could not inactivate any essential enzyme. Subsequently, prokaryotes have been detected growing around black smokers and hydrothermal vents in the deep sea at temperatures at least as high as 115°C. Microorganisms have been found growing at very low temperatures as well. In super cooled solutions of H₂O as low as -20°C, certain organisms can extract water for growth, and many forms of life flourish in the icy waters of the Antarctic, as well as household refrigerators, near 0°C. Based on the growth of bacteria at different temperatures, there are three different temperatures are available.

Minimum temperature – The temperature below which the organism will not grow at all.

maximum temperature – above which there won’t be any growth of organism

Optimum temperature – The temperature in which maximum growth occurs.

Based on the optimum temperature requirement, the organisms can be classified as :

Psychrophiles: The organisms which prefer low temperature (15°C) as optimum temperature are referred as psychrophiles. They have about 0°C and 20°C as their minimum and maximum temperature respectively. Ex. Poloromonas vaculata

Psychrotolerants: They are the mesophiles but can grow even at lower temperature of 20-40°C (as the maximum temperature not optimum) Example: Refrigerator bacteria (Food borne pathogens)

Mesophiles: The organisms which prefer 35°C (room temperature) as optimum temperature are referred as mesophiles. They have about 10°C and 48°C as their minimum and maximum temperature respectively. Ex. Bacillus, Pseudomonas, E. coli

Thermotolerants - They are the mesophiles but can grow even at higher temperatures (Ex. 45° to 55°C)

Thermophiles – The organisms which prefer high temperature (55-65°C) as optimum temperature are referred as thermophiles. They have 40°C and 70°C as their minimum and maximum temperatures respectively. Ex. Bacillus stearothermophilus

Hyper thermophiles – The organisms which prefer very high temperatures (80 – 100°C) as their optimum temperature are referred as hyper thermophiles. They have 60°C and 110°C as their minimum and maximum temperature respectively. (Ex. Thermococcus)

pH

It refers to the acidity or alkalinity of a solution. It is a measure of the hydrogen ion activity of a solution and is defined as the negative logarithm of the hydrogen ion concentration. The pH scale ranges from 1.0 to 14.0 and most microorganisms grow vary widely from pH 0 to 2.0 at the acid end to alkaline lakes and soil that may have pH values between 9.0 and 10. The pH can affect the growth of microorganisms and each species has a definite pH growth range and pH growth optimum. Acidophiles have their growth optimum between pH 0 and 5.5; neutrophiles between 5.5 and 8.0 and alkalophiles prefer pH range of 8.5 to 11.5 (Example Bacillus halodurans). Most bacteria and protozoans are neutrophiles, fungi prefer acid surroundings about pH 4 to 6; algae also seem to favour slight acidity. Cyanidium caldarium (algae) and archaeon Sulfolobus acidocaldarium are inhabitants of acidic hot springs; both grow well around pH 1 to 3 and at high temperature. Drastic changes/variations in cytoplasmic pH can harm microorganisms by disrupting the plasma membrane or inhibiting the activity of enzymes and membrane transport proteins. Prokaryotes die if the internal pH drops much below 5.0 to 5.5. External pH alterations also might alter the ionization of nutrient molecules and thus reduce their availability to the organism. The microorganism needs to maintain a neutral cytoplasmic pH and for this the plasma membrane may be relatively impermeable to protons. Neutrophiles appear to exchange potassium for protons using an antiport transport system. Extreme alkalophiles maintain their internal pH closer to neutrality by exchanging internal sodium ions for external protons. The antiport systems probably correct small variations in pH. In case of too much acidity (below 5.5 to 6.0) S. typhimurium and E.coli synthesize an array of new proteins as part of what has been called as their acidic tolerance response. If the external pH decreases to 4.5 or lower, chaperones such as acid shock proteins and heat shock proteins are synthesized. Microorganisms can change the pH of their own habitat by producing acidic or basic metabolic waste products. In order to maintain the pH, buffers are often included in the media to prevent growth inhibition. Phosphate is commonly used buffer and a good example of buffering agent. Peptides and amino acids in complex media also have a strong buffering effect.

The range of pH over which an organism grows is defined by three cardinal points: the minimum pH, below which the organism cannot grow, the maximum pH, above which the organism cannot grow, and the optimum pH, at which the organism grows best. For most bacteria there is an orderly increase in growth rate between the minimum and the optimum and a corresponding orderly decrease in growth rate between the optimum and the maximum pH, reflecting the general effect of changing [H⁺] on the rates of enzymatic reaction.

Microorganisms which grow at an optimum pH well below neutrality (7.0) are called acidophiles. Those which grow best at neutral pH are called neutrophiles and those that grow best under alkaline conditions are called alkalophiles. Obligate acidophiles, such as some Thiobacillus species, actually require a low pH for growth since their membranes dissolve and the cells lyse at neutrality. Several genera of Archaea, including Sulfolobus and Thermoplasma, are obligate acidophiles. Among eukaryotes, many fungi are acidophiles, and the champion of growth at low pH is the eukaryotic alga Cyanidium which can grow at a pH of 0.

Oxygen

Because animals require molecular oxygen (O₂), it is easy to assume that all organisms require O₂. However, this is not true; many microorganisms can, and some must, live in the total absence of oxygen. Oxygen is poorly soluble in water, and because of the constant respiratory activities of microorganisms in aquatic habitats, O₂ can quickly become exhausted. Thus, anoxic (O₂-free) microbial habitats are common in nature and include muds and other sediments, bogs, marshes, water-logged soils, intestinal tracts of animals, sewage sludge, the deep subsurface of Earth, and many other environments. In these anoxic habitats, microorganisms, particularly prokaryotes, thrive.

Oxygen is a universal component of cells and is always provided in large amounts by H₂O. However, prokaryotes display a wide range of responses to molecular oxygen O₂. Molecular oxygen is both beneficial and harmful for biologicals. It act as terminal electron acceptor in the respiratory chain – Electron transport chain reaction enable to form ATP. However, the oxygen derivatives such as hydrogen peroxide, singlet oxygen and super oxide are highly toxic and oxidizing agents.

Microorganisms vary in their need for, or tolerance of, O₂. In fact, microorganisms can be grouped according to their relationship with O₂. Aerobes can grow at full oxygen tensions (air is 21% O₂) and respire O₂ in their metabolism (Example Micrococcus). Many aerobes can even tolerate elevated concentrations of oxygen (hyperbaric oxygen). Microaerophiles, by contrast, are aerobes that can use O₂only when it is present at levels reduced from that in air (microoxic conditions) (Example Spirillum volutans). This is because of their limited capacity to respire or because they contain some O₂-sensitive molecule such as an O₂-labile enzyme. Many aerobes are facultative, meaning that under the appropriate nutrient and culture conditions they can grow under either oxic or anoxic conditions (Example: E. coli, fungi: Yeast). Some organisms cannot respire oxygen; such organisms are called anaerobes. There are two kinds of anaerobes: aerotolerant anaerobes, which can tolerate O₂ and grow in its presence even though they cannot use it (Streptococcus), and obligate anaerobes, which are inhibited or even killed by O₂. The reason obligate anaerobes are killed by O₂ is unknown, but it is likely because they are unable to detoxify some of the products of O₂ metabolism (Example: Clostridium).

So far as is known, obligate anaerobiosis is found in only three groups of microorganisms: a wide variety of Bacteria and Archaea, a few fungi, and a few protozoa. The best-known group of obligatorily anaerobic Bacteria belongs to the genus Clostridium, a group of gram-positive endospore-forming rods. Clostridia are widespread in soil, lake sediments, and the intestinal tracts of warm-blooded animals, and are often responsible for spoilage of canned foods.

Oxygen detoxification : Most of the aerobes (and aerotolerant organisms too) have three major enzymes either all or two to detoxify the oxygen derivatives. They are catalase, peroxidase and super oxide dismutase. Obligate anaerobes lack these enzymes, and therefore undergo lethal oxidations by various oxygen radicals when they are exposed to O₂.

Water activity

Water is the solvent in which the molecules of life are dissolved, and the availability of water is therefore a critical factor that affects the growth of all cells. The availability of water for a cell depends upon its presence in the atmosphere (relative humidity) or its presence in solution or a substance (water activity). The water activity (A_w) of pure H₂O is 1.0 (100% water). Water activity is affected by the presence of solutes such as salts or sugars, that are dissolved in the water. The higher the solute concentration of a substance, the lower is the water activity and vice-versa. Microorganisms live over a range of A_w from 1.0 to 0.7. The Aw of human blood is 0.99; seawater = 0.98; maple syrup = 0.90; Great Salt Lake = 0.75. Water activities in agricultural soils range between 0.9 and 1.0.

The water requirement (A_w) of few bacteria are as follows: Caulobacter - 1.00; Pseudomonas, Salmonella, E. coli - 0.91; Lactobacillus - 0.90; Bacillus -0.90; Staphylococcus -0.85; Halococcus - 0.75

(Note: The concept of lowering water activity in order to prevent bacterial growth is the basis for preservation of foods by drying (in sunlight or by evaporation) or by addition of high concentrations of salt or sugar.)

Salt concentrations

The only common solute in nature that occurs over a wide concentration range is salt [NaCl], and microorganisms are named based on their growth response to salt. Microorganisms that require some NaCl for growth are halophiles.

Mild halophiles require 1-6% salt, (Ex: Staphylococcus areus)

Moderate halophiles require 6-15% salt; (Ex. Vibrio fisheri)

Extreme halophiles require 15-30% NaCl for growth. (Ex. Halobacterium salanarum)

Bacteria that are able to grow at moderate salt concentrations, even though they grow best in the absence of NaCl, are called halotolerant. Although halophiles are "osmophiles" (and halotolerant organisms are "osmotolerant") the term osmophiles is usually reserved for organisms that are able to live in environments high in sugar.

Organisms which live in dry environments (made dry by lack of water) are called xerophiles.

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