BSc 2nd Year Microbiology of Water Notes Study Material

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BSc 2nd Year Microbiology of Water Notes Study Material

In Microbiology of water we shall briefly examine the main pollutants that make water unfit for drinking and the methods of its purification. Some microbiological tests of water for its safety in consumption will also be described. We shall also examine briefly the sewage treatment since sewage and water are important source of microbial transmission.

Types of Water

There are two major types of water.

1. Ground water. It originates from deep wells and subterranean springs. This is virtually free of bacteria due to filtering action of soil, deep sand and rock. However, it may become contaminated when it flows along the channels.

2. Surface water. It is found in streams, lakes, and shallow wells. The air through which the rain passes may contaminate the water. Other sources are the various types of establishments and agricultural farms etc. by the sides the water flows. Possible sources of microbial contamination of a body of water are soil and agricultural run off, farm animals, rain water, industrial waste, discharges from sewage treatment plants and storm water run off from urban areas.

In water microbiology the water is contaminated when it contains a chemical or biological poison or an infectious agent. These conditions also apply to water which is polluted except that the agent or poison is often obvious and the water carries an unpleasant taste or appearance. Potability refers to the drinkability of water. When potable, it is fit for drinking. When unpotable it is unfit due to some contaminant or pollutant.(BSc Microbiology of Water Notes Study Material)

Microbial Components of Water

The types of microorganisms differ in unpolluted, polluted and marine waters.

We shall briefly examine such differences. Typical organisms in these water environments are shown in Table.

In unpolluted water of mountain lake or stream there are usually low organic nutrients. The number of bacteria is very much limited, a few thousand per ml. Actinomycetes are typical. Other organisms shown in Table are also present. Autotrophic bacteria are common along with free-living protozoans as Euglena, Paramecium and various amoebae.

In polluted waters, there are large amounts of organic matter from sewage, feces and industrial complex. The microbes are usually heterotrophic. The digestion of organic matter by these organisms is incomplete, due to which there accumulate acids, bases, alcohols and various gases.

The major types of bacteria are coliform bacteria, the Gram-negative non-sporeforming bacilli usually found in the intestine. This group includes E. coli and species of Enterabacter. They ferment lactose to acid and gas. Non-coliform bacteria-Streptococcus, Proteus and Pseudomonas are also present.

Under some conditions, the polluting organisms multiply rapidly and consume most of the available oxygen. For instance, nutrients enter the river from sources like sewage treatment plants or urban/suburban runoff. Thus river suddenly develops a high nutrient content. Under these conditions algae may bloom rapidly. This leads to depletion of oxygen in water.

There is very little oxygen available to the protozoa, small animals, fish and plants. Due to this non-availability of oxygen, a layer of dead organisms, mud and silt accumulate at the bottom, and anaerobic species of Clostridium, Desulfovibrio etc. will flourish. They produce gases. One gas, H₂S combines with lead or iron to give a precipitate which makes the mud black and the water poisonous. Due to complete depletion of oxygen, the suspended bacteria die in their own waste products. There is hardly any life in water at this stage. The gas bubbles from the anaerobes in the mud break! the surface. Such processes lead to death of a river.

The marine water is high in salt and only halophilic organisms survive. Since temperature is very low, most of them are psychrophilic. Diatoms and protozoa as dinoflagellates are important components in food chain. In the offshore oceanic zone, photosynthetic organisms are present. These are diatoms and dinoflagellates. Most marine microbes are found along the shoreline or littoral zone where nutrients are in abundance. Some unusual types are also present on the ocean floor in the benthic zone and even at the bottom of several miles-deep trenches, the abyssal zone.(BSc Microbiology of Water Notes Study Material)

Water Pollution

Polluted waters can be classified in different ways. In microbiological context, there are generally recognised physical, chemical and biological pollution of water. Of these biological pollution is of much concern to us.

1. Physical pollution. Here the water becomes cloudy due to presence of some particulate matter such as sand or soil. Some plant material as algal-bloom may also be involved, that turns the consistency of water like a pea soup. Presence of some phosphates or nitrates may encourage the growth of blooms, leading finally to a condition known as eutrophication. This upsets the ecological balance.

2. Chemical pollution. It occurs due to introduction of inorganic and organic waste to water. For example, water passing out of a mine contains large amount of copper, iron or other ore. Another source of pollution is metal pipelines. Corrosion of metals of the pipes may yield rusty or black precipitates. Other chemical pollutants include laundry detergents, radioactive wastes and nitrates from fertilisers etc. An important group of chemicals are polychlorinated biphenyls or PCBs, a group of industrial chemicals used in the production of electric insulators and paper products. They are carcinogenic and necessary to be eliminated from water. Main sources of chemical pollutants are mining wastes, air pollutants (washed down by rain), power plants, industries, sewage and storm water runoff, wash from farm lands (pesticides, fertilisers), hospitals, road salts and exhausts from automobiles.(BSc Microbiology of Water Notes Study Material)

3. Biological pollution. It develops from the microorganisms that enter waters from such sources as human waste, food operations, meat-packing plants and medical facilities etc. Under normal conditions, the water body is able to handle biological material, because heterotrophs digest the organic matter to simple phosphates, nitrates and sulphates which fertilise the plants. Carbohydrates are converted to CO₂ and water, which are also useful to plants. The plants then release O₂ through photosynthesis. Due to rapid movement of water, aeration is more or less constant and the wastes eliminated.

However, there arises the problem when water becomes stagnant or is overloaded with waste. Water is now unable to handle the biological material, and it soon becomes polluted. We have already seen how a lake may turn into a swamp due to overabundance of organic matter. A critical factor operating under such a situation is the biological oxygen demand (BOD). It refers to the oxygen requirement by the metabolising organisms. As the number of organisms increases, the demand for O₂ increases proportionally. Depletion of O₂ in water may lead to death of fish and other aquatic animals, a flat taste of water, and eventual death of aerobes.(BSc Microbiology of Water Notes Study Material)

The BOD is therefore an important indication of the levels of biological pollution in the water. In laboratory, BOD is determined by measuring the amount of dissolved oxygen in test samples of water distributed to BOD bottles. Initial determination of dissolved oxygen in sample is made. The bottles are then stoppered and incubated at 20°C for five days, after which the amount of dissolved oxygen is again determined. The difference in the oxygen content represents the amount of oxygen used up in the test sample, which is the BOD. Results are expressed as parts per million (ppm). A BOD of several handred taken usually high.(Microbiology of Water Notes study Material)

Diseases transmitted by water

Water is infact a vehicle for the transfer of a wide range of diseases of microbial origin. The important bacterial diseases include typhoid fever, choler and bacterial dysentry that are generally transmitted when human feces from carriers or patients contaminates the water. Viral diseases transmitted by water include virus A hepatitis and polio. Many protozoa form cysts which survive for long periods in water. The causal agents of amoebiasis, giardiasis etc. are important concerns in water pollution.(BSc Microbiology of Water Notes Study Material)

Water Treatment

We have seen that water and sewage harbour a variety of organisms responsible for diseases in man. It is thus necessary to remove or rather interrupt the disease cycle in nature through treatment of water and sewage. Water purification and sanitary sewage disposal are two major methods for interrupting disease cycle. In water purification organisms are prevented from reaching the body whereas in sewage disposal, they are removed from body waste products.

The various steps in the purification of drinking water in municipal water supplies are shown in Figure. It may be seen that there are three basic steps in the purification of drinking water: sedimentation, filtration and chlorination.

1. Sedimentation. The objective is to remove bulky objects such as leaves, sand and gravel particles and the materials that have run off from the soil.

Sedimentation is done in large reservoirs or in a restricted area of a settling tank. Aluminium sulphate (alum) or iron sulphate may be added to hasten sedimentation process. This is done in a mixing chamber (C) in Fig. After mixing, the chemicals form jellylike masses of coagulated material called flocs. They fall through the water and cling to organic particles and microorganisms, dragging a major portion to the bottom sediment in a process called flocculation. Coagulated material is allowed to settle in a settling tank.

2. Filtration. This process removes the remaining microorganisms from the water. Although, different types of filter material are available, mostly a layer of sand and gravel is utilised to trap the microorganisms. A slow sand filter uses several feet deep layer of fine sand particles. Within the sand there forms a layer of microorganisms which acts as an additional filter. This layer is called a Schmutzdecke or dirty layer.(BSc Microbiology of Water Notes Study Material)

To cleanse the filter, the top layer is removed and replaced with fresh sand. A rapid sand filter contains coarser particles of gravel. The schmutzdecke does not develop; the filtration rate is higher (over 200 million gallons per acre per day). This type of filter is generally used in municipal supplies.

3. Chlorination. In this final step, chlorine gas is added to the water. This gas is an active oxidant which reacts with organic matter in the water. Gas is added until any residue is present. A residue of 0.2 ppm-1.0 ppm of water is often the standard used. Under these conditions, most remaining microbes die within 30 minutes.(BSc Microbiology of Water Notes Study Material)

Bacteriological Analysis of Water

There are methods to detect bacterial contamination of water. The chief objective is to identify coliform organisms as Escherichia coli. Their presence indicates that water contains fecal pollution and is unsafe for consumption. The standard methods used are as follows:

1. Membrane filter technique. It may be used in the field also. A special, collecting bottle is held against the water current and a 100-ml sample is taken. The water is filtered through a membrane filter and the filter pad is then transferred to a plate of bacteriological medium. Bacteria trapped in the filter will form colonies which may be counted.(Microbiology of Water Study Material)

2. Standard plate count (SPC). Water sample is diluted in sterile buffer. Measured amounts are pipetted into Petri dishes. Agar medium is added and plates incubated. Colony counts are made and multiplied by the reciprocal dilution factor to have total bacteria per ml water. This method is similar to that used for milk.(BSc Microbiology of Water Notes Study Material)

3. Most probable number (MPN). In this procedure, water in 10 ml, 1 ml and 0.1 ml amounts is inoculated into lactose broth tubes. The tubes are incubated and coliform organisms may be identified by their production of gas from the lactose. By referring to a MPN table, a statistical range of the number of coliform may be determined by observing how many broth tubes showed gas. This method does not detect total number of bacteria in the water nor it locates non-coliforms like Salmonella. However it indicates the presence and quantity of coliforms.

4. Specific tests (Coliform counts) for assessing water safety. The most frequently used indicator organism is the normally nonpathogenic coliform bacterium Escherichia coli. Positive tests for E. coli do not prove the presence of enteropathogens, such as Salmonella and Shigella, but do establish the possibility. E. coli is more numerous and easier to be grown. For fecal contamination of water, using E. coli as an indicator organism, a three-stage test procedure is followed.(BSc Microbiology of Water Notes Study Material)

(a) Presumptive test. In the first stage, lactose broth tubes are inoculated with undiluted or appropriately diluted water samples. The tubes showing gas formation are marked as positive and used to calculate the most probable number (MPN) in the sample. Gas formation in inverted test tubes (Durham tubes), indicates fecal contamination by coliforms.

(b) Confirmed test. Gas formation in lactose broth at 37°C is characteristic not only of Salmonella, Shigella and E. coli strains but also to the non-fecal coliform Enterobacter aerogenes and some Klebsiella spp. In second stage, the presence of enteric bacteria is confirmed by streaking samples from the positive lactose broth culture onto a medium, such as eosin methylene blue (EMB) agar. Fecal coliform colonies on this medium acquire a greenish metallic sheen, Enterobacter spp. form reddish, and non-lactose fermenters form colourless colonies respectively.(BSc Microbiology of Water Notes Study Material)

(c) Completed test. The confirmed test can be accomplished by using brilliant green lactose-bile broth (BGLB). If BGLB is used, it is then sub cultured onto EMB. Sub-culturing colonies showing a green metallic sheen on EMB into lactose broth incubated at 35°C should produce gas, completing a positive test for fecal coliforms.(BSc Microbiology of Water Notes Study Material)

BSc Microbiology of Water Notes Study Material

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