Factors affecting the microbial growth

THE INFLUENCE OF ENVIRONMENTAL FACTORS ON GROWTH

Microbial growth is greatly affected by chemical and physical nature of their surroundings instead of variations in nutrient levels and particularly the nutrient limitation. For successful cultivation of microorganisms it is not only essential to supply proper and balanced nutrients but also it is necessary to maintain proper environmental conditions. Thus, understanding of environmental influences on the growth of microorganisms becomes mandatory. As bacteria shows divers food habits, it also exhibits diverse response to the environmental conditions. Growth and death rates of microorganisms are greatly influenced by number of environmental factors such as solutes and water acidity, temperature, oxygen requirement, pH, pressure and radiation.

Solutes and Water Acidity

Water is one of the most essential requirements for life. Thus, its availability becomes most important factor for the growth of microorganisms. The availability of water depends on two factors - the water content of the surrounding environment and the concentration of solutes (salts, sugars etc.) dissolved in the water.

In most cases, the cell cytoplasm possesses higher solute concentration in comparison to its environment. Thus, water always diffuses from a region of its higher concentration to a region of the lower concentration. This process is called osmosis which keeps the microbial cytoplasm in positive water balance. When a microbial cell is placed in hypertonic solution (or, solution of low water activity), it loses water and shrinkage of membrane takes place.

This phenomenon is called plasmolysis. Microorganisms show variability in their ability to adapt the habitats of low water activity. Microorganisms like S. aureus can survive over a wide range of water activity and are called as osmotolerant (as water activity is inversely related to osmotic pressure). However, most microorganisms grow well only near pure water activity (i.e. around 0.98-1). Thus, drying of food or addition of his concentration of salts and sugars is the most popular way of preventing spoilage of food. Seawa, microorganisms are called as halophiles since they require high concentration of salts (between 2.b 6.2 M) to grow. Halobacterium, a halophilic archaebacterium, inhabits Dead Sea (a salt lake situated between Israel and Jordan and the lowest lake in the world), the Great Salt Lake in Utah and other aquatic habitats possessing salt concentrations approaching salt water.

The quantitative availability of water can be expressed in physical term called water activity (aw). The water activity for a sample solution is the ratio of vapour pressure of the sample solution to the vapour pressure of the water at the same temperature (aw = Psoln/Pwater) The relative humidity of a test sample (at equilibrium) can be obtained, after sealing it in a closed chamber. This determines the water activity of a solution. For example, if after treating by above method relative humidity of air over the sample is 95% then, the water activity of the sample is 0.95

Temperature

All forms of life are greatly influenced by temperature. In fact, the microorganisms are very sensitive to the temperature since their temperature varies with that of environment (poikilothermic). Temperature influences the rate of chemical reactions and protein structure integrity thus affecting rates of enzymatic activity. At low temperature enzymes are not w denatured, therefore, every 10°C rise in temperature results in rise of metabolic activity and growth of microorganisms. However, enzymes have a range of thermal stability and beyond it their denaturation takes place. Thus, high temperature kills micro- CJ organism by denaturing enzymes, by inhibiting transport carrier molecules or by change in membrane integrity. Each microbe shows characteristic temperature dependence and possesses its own cardinal temperatures i. e. minimal, maximum and optimal growth temperatures.

The values for cardinal temperature vary widely among bacteria. For convenience, bacteria isolated from hot springs can survive even at temperature of 100°C and above, while those isolated from snow can survive below -10°C. On the basis of susceptibility to the thermal conditions, microorganisms are classified into three categories: thermophiles. Meshophiles and psychrophiles.

Thermophiles are microorganisms that show growth optima at 55°C. They often have growth maxima of 65°C, while few can grow even at 100°C and higher temperatures. Their growth minima is 45°C. The vast majority of thermophiles belongs to prokaryotes although a few microalgae (e.g., Cyanidium caldarium) and microfungi (e.g. Mucor pusillus) are also thermophiles. A few microorganisms are hyperthermophiles as they possess growth optima between 80°C and about 113°C. Hyper­thermophiles usually do not grow well below 55°C (e.g.. Pyrococcus abyssi, Pyrodictium occultum). Mesophiles are microorganisms that have growth minima between 15°C-20°C; optima between 20­45°C and maxima at 45°C. Most microorganisms fall within this category. Almost all human pathogens are mesophiles as they grow at a fairly constant temp. of 37°C. Psychrophiles have optimum temperature for growth at 15°C, however, few can grow even below 0°C. The maximum growth temperature of psychrophiles is around 20°C. The spoilage of refrigerated food takes place because of facultative psychrophiles. These are microorganisms that can grow at 0°C but have growth optima temperature between 20°- 30°C

Oxygen Requirements

The atmosphere of earth contains about 20% (v/v) of oxygen. Microorganisms capable of growing in the presence of atmospheric oxygen are called aerobes whereas those that grow in the absence of atmospheric oxygen are called as anaerobes. The micro-organisms that are completely dependent on atmospheric oxygen for growth are called obligate aerobes whereas those that do not require oxygen for growth but grow well in its presence are called as facultative anaerobes. Aerotolerants (e.g. Enterococcus faecalis) ignore O₂ and can grow in its presence or absence. In contrast, obligate anaerobes (e.g., Bacteroids, Clostridium pastewianum, Furobacterium) do not tolerate the presence of oxygen at all and ultimately die. Few microorganisms (e.g., Campylobacter) require oxygen at very low level (2-10%) of concentration and are called as microaerophiles. The latter are damaged by the normal atmospheric level of oxygen (20%)

Relationship of Oxygen and growth (Shake tubes method)

1. Aerobic

2. Microaerophilic

3. Facultative

4.Anaerobic

These varying relationships between microbes (especially the bacteria) and O₂ appear due to different factors such as protein-inactivation and the effect of toxic oxygen-derivatives. Bacterial enzymes can be inactivated when interact with oxygen. Nitrogen-fixing enzyme introgenese is very sensitive to oxygen and represents a good example of interaction between enzyme and oxygen. During metabolic process, flavoprotein reduces oxygen to form hydrogen peroxide (H₂O₂), superoxide radical (O₂^-) and hydroxy radical (OH^-). These compounds are highly toxic and being powerful oxidizing agent, can cause destruction of cellular macromolecules.

O₂ + e^- = O₂^- (superoxide radical)
O₂^- + e^- + 2H⁺ = H₂O₂ (hydrogen peroxide)
H₂O₂ + e^- + H+ = H₂O + OH^- (hydroxy radical)

In order to survive, therefore, bacteria must be able to protect itself from oxidising agents. All aerobes and facultative anaerobes contain two enzymes namely superoxide dismutase and catalase. These enzymes protect microbes against lethal effects of oxygen products. The oxidizing property hydrogen peroxide. The enzyme catalase decomposes hydrogen peroxide into oxygen and water. The aerotolerant bacteria like lactic acid bacteria possess enzyme peroxides instead of catalase to decompose the accumulated hydrogen peroxide.

2O₂^- 2H⁺ -----Superoxidase dismutase-----O₂ +H₂O₂
2H₂O₂ -----Catalase------ 2H₂O +O₂
H₂O₂ +NADH + H⁺ ----Peroxidase---- 2H₂O + NAD⁺

Since all obligate anaerobes lack these enzymes or have them in very low concentration and, therefore, are susceptible to oxygen.

Pressure

Normal life of microorganisms on land or on the water surface is always subjected to a pressure of 1 atmosphere. But, they are many microbes that survive in extremes of hydrostatic pressure in deep sea. Others are there that not only survive rather grow more rapidly at high pressures (e.g., Protobacterium, Colwellia, Shewanella) and are called barophilic. Some archaebacteria are thermobarophiles (e.g., Pyrococcus spp., Methanococcus jannaschii). However, A barophile has been recovered from the depth about 10,500 m in sea near Philippines and has been found unable to grow at 2°C temperature and below about 400-500 atmospheric pressure

Radiation

Some electromagnetic radiations, particularly the ionizing radiation (e.g., X-rays, gamma rays) are very harmful to microbial growth. Low levels of these radiations may cause mutations and may indirectly result in death whereas high levels may directly cause death of the microbes. Ionizing radiation, however, destroys ring-structures, breaks hydrogen bonds, oxidizes double bonds and polymerizes certain molecules. Ultraviolet (UV) radiation is lethal to all categories of microbial life due to its short wavelength and high-energy; the most lethal UV radiation has a wavelength of 260 nm. Ultraviolet radiation primarily forms thymine dimers in DNA to cause damage. Two adjacent thymines in a DNA strand join each other covalently and inhibit DNA replication and function. Microbial photosynthetic pigments (chlorophyll. bacteriochlorophyll, cytochromes and flavins), sometimes, absorb light energy, become excited or activated, and act as photosensitizers. The excited photosensitizer (P) transfers its energy to oxygen which then results in singlet oxygen (¹O₂), The latter is very reactive and powerful oxidizing agent and quickly destroys a cell. The singlet oxygen is probably the main weapon employed by phagocytes to destroy engulfed bacteria.