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Microbiology of Water and Waste Water Management

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Water Microbiology NOTES (SRTMUN Nanded/BAMU A'Bad)
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       MICROBIOLOGY OF WATER AND WASTE WATER MANAGEMENTMicrobial Communities in Natural Water The three biological organisms present in wastewater are bacteria, viruses, and parasites. Sewageconsists of vast quantities of bacteria, most of which are harmless to man.  However, pathogenic (disease-causing) organisms such as typhoid, dysentery, and other intestinal disorders may be present inwastewater. Bacteria are the most widely distributed life forms. Pathogenic bacteria range in length fromapproximately 0.4 to 14 mm (a mm or “micrometer” equals one one -thousandth of a millimeter) and 0.2 to1.2 mm in width. Key bacterial pathogens responsible for waterborne disease include Legionella,Salmonella typhi, Shigella, and Vibrio cholerae. Viruses are inactive when outside of a living host cell.Viruses linked to waterborne disease have protein coats that provide protection from environmentalhazards and range in size from 0.02 to 0.09 mm. Unlike bacteria and protozoa, they contain only one typeof nucleic acid (RNA or DNA). Key pathogens include hepatitis A. Protozoa , common in bodies of water,are much larger than bacteria and viruses. To survive harsh environmental conditions, some species can secrete a protective covering and form a resting stage called a “cyst.” Encystment can protect protozoafrom drinking water disinfection efforts and facilitate the spread of disease. Key protozoa being studied asagents of waterborne disease include Giardia and Cryptosporidium . Fecal pollution of water Fecal pollution of water from a health point of view is the contamination of water with disease-causing organisms (pathogens) that may inhabit the gastrointestinal tract of mammals, but with particularattention to human fecal sources as the most relevant source of human illnesses globally. Ingestion of water contaminated with feces is responsible for a variety of diseases important to humans via what isknown as the fecal-oral route of transmission. Food, air, soil, and all types of surfaces can also beimportant in the transmission of fecal pathogens, and thereby implicated in disease outbreaks. Most fecalmicroorganisms, however, are not pathogenic. Indeed, some are considered beneficial to the host as theycan out compete pathogens for space and nutrients, complement the bioc hemical potential of the host’s gastrointestinal tract, and help in the development of the host immune system. Nonetheless, animal fecescan also carry a number of important frank and opportunistic pathogens, capable of inflicting debilitatingillnesses and, in some cases, death. Indicators of Feacal Pollution Traditionally, indicator micro-organisms have been used to suggest the presence of pathogens.Today, however, we understand a many reasons for indicator presence and pathogen absence, or viceversa. In short, there is no direct correlation between numbers of any indicator and enteric pathogens. Toeliminate the vagueness in the term ‘microbial indicator’, the following three groups are now recognized.       1. Process indicator: A group of organisms that demonstrates the efficacy of a process. 2. Faecal indicator: A group of organisms that indicates the presence of faecal contamination.Ex: E. coli . 3. Index and model organisms: A group/or species indicative of human pathogen. Feacal Coliforms as index of water Pollution The use of bacteria as indicators of the sanitary quality of water probably dates back to 1880 when VonFritsch described Klebsiella pneumoniae and K. rhinoscleromatis as micro-organisms characteristicallyfound in human faeces. In 1891, the Franklands came up with the concept that organisms characteristic of  sewage must be identified to provide  evidence of potentially dangerous pollution. By 1893, the ‘Wurtzmethod’ of enumerating B. coli by direct plating of water samples on litmus lactose agar w as being usedby sanitary bacteriologists, using the concept of acid from lactose as a diagnostic feature. This wasfollowed by gas production, with the introduction of the Durham tube (Durham 1893). The concept of  ‘coliform’ bacteria, those bacteria resem bling B. coli, was in use in Britain in 1901. The colony count forbacteria in water, however, was not formally introduced until 1934. Therefore, the sanitary significance of finding various coliforms along with streptococci and C. perfringens was recognised by bacteriologists bythe start of the twentieth century. It was not until 1905, however, that Mac Conkey (1905) described hisnow famous Mac Conkey’s broth , which was diagnostic for lactose-fermenting bacteria tolerant of bilesalts. Nonetheless, coli-forms were still considered to be a heterogeneous group of organisms, many of which were not of faecal srcin.It is almost impossible to isolate from water the organisms responsible for water-borne diseases.Few organisms are present and they do not multiply in water. The only safe method to prevent waterbornedisease is to condemn fecally polluted water as being unfit for human use, as it may contain harmfulorganisms. Fecal pollution can be determined by examination of water for colon bacilli ( E.coli ). E.coli isabundant in feces and not found outside intestinal tract in nature. The E.coli in water indicates the presenceof pathogenic microorganisms in water, which may be responsible for a number of water-borne diseases.Hence, E.coli is known as indicator organism. Water also contains bacteria that resemble E.coli but may ormay not be of fecal srcin. These bacteria also ferment lactose with formation of gas like E.coli . The otherindicator organisms are Streptococcus faecalis Streptococcus faecium, Streptococcus bovis , Streptococcusequinus etc., and Clostridium perfringenes. Significance of Feacal Coliforms The group of coliform bacteria as an indicator of other pathogenic micro-organisms, specificallyorganisms of faecal srcin, has had much emphasis in all countries. This is due primarily to the fact thatthe coliform bacteria groups meets many of the criteria for a suitable indicator organism, and are thus asensitive indicator of faecal pollution:       ã They are abundant in faeces ã They are generally found only in polluted waters, ã They are easily detected by simple laboratory tests, ã Can be detected in low concentrations in water ã The number of indicator bacteria seems to be correlated with the extent of contamination.It is important to remember, however, that not all coliforms srcinate from human faeces as theycan srcinate from other mammalian species or from other environmental sources (e.g., bird droppings).When coliforms are discharged to the aquatic environment they will tend to die at a rate which depends,amongst other things, on the temperature and turbidity of the water and the depth to which solar radiationpenetrates. Therefore, it is not safe to conclude that the lack of coliforms in a water means that it has notbeen subject to faecal pollution. It is necessary to be familiar with a number of terms are as follows: Total coliforms : The Total coliform group comprises several distinct types (genera) of bacteria. Thesebacteria have been isolated from the faeces of humans and other warm-blooded animals, as well ascontaminated and non-contaminated soils. This group of bacteria is widely used as a measure of healthhazard from faecal contamination. The total coliform group comprises the aerobic and facultative, gramnegative, nonspore-forming, rod shaped bacteria that ferment lactose with gas formation within 48 hours at35 °C. Faecal coliforms : The Faecal coliform group of bacteria are indicative of faeces of humans and otherwarm blooded animals. The specific bacterium Escherichia coli is part of this group. The test for faecalcoliform is at an elevated temperature, 44.5°C, where growth of other non-faecal bacteria is suppressed.However, some non-faecal bacteria may be also be identified in the faecal coliform test, though a smallpercentage (<5%). Faecal streptococci : This group of bacteria includes several species or varieties of streptococci and thenormal habitat of these bacteria is the intestines of humans and animals. Examples include Streptococcifaecalis which represents bacteria of humans and Streptococci bovis and Streptococci equinus whichrepresent bacteria that are indicators of cattle and horses. Thermotolerant coliforms : This is a more precise definition of coliforms which are determined by the testfor faecal coliforms. In practise not all such coliforms are faecal in srcin although most (> 95%) are. Escherichia coli ( E. coli ) This bacterium is a particular member of the faecal coliform group of bacteria;this organism in water indicates the presence of faecal contamination. E. coli reside in human intestinaltracts. They are excreted in large numbers in faeces, averaging about 50 million per gram. Untreateddomestic wastewater generally contains 5 to 10 million coliforms per 100 ml.Pathogenic bacteria and viruses causing enteric diseases in humans srcinate from faecaldischarges of diseased persons. Consequently, water containing coliform bacteria is identified as       potentially dangerous. Coliform bacteria are, therefore, considered as an indicator of bacteriologicalquality of water for the following reasons: ã Coliform bacteria far outnumbers the pathogenic micro-organisms, ã They do not multiply in natural waters, ã The die-off rate of pathogenic bacteria is greater than the death rate of coliforms ã Test for coliform bacteria is relatively simple and can be performed in water quality laboratoriesThe bacterium E.coli is exclusively of faecal srcin. Some coliform bacteria are normal inhabitantsof soil and water. In testing for conforms, therefore, tests may be run in conjunction to verify their faecalsrcin. However, unconfirmed testing, indeed, would provide a factor of safety. The degree to whichindicator organisms represent the presence of individual pathogens (such as Salmonella ) has been thesubject of continuing investigation. There does seem to be a genera correlation between the concentrationof Faecal coliform bacteria and the occurrence of Salmonella. When faecal coliform numbers are about1000 per 100 ml, Salmonella occurrence is about 95 % Relationships between total coliform and individualpathogens is not so quantitative. Thus the test of total coliform is not so effective for an indicator. The totalcoliform test is complicated by the presence of non-faecal bacteria. As a general rule, faecal coliformlevels are about 20% of total coliform concentrations, although a wide spread exists. BACTERIOLOGICAL EXAMINATION OF WATER The test for coliform bacteria is usually conducted using a liquid culture. Enumeration employing solidculture media is not commonly done in Ind ia. The liquid culture ‘multiple tube technique’ consists of  mainly 2 stages (third test is optional). These are 1. Presumptive test 2. Confirmed test 3.Completed test(Optional) 1. Presumptive test The first step in water examinations is known as the presumptive test. The presumptive test is based on gasproduction during fermentation of lauryl tryptose broth which contains beef extract, peptone and lactosewithin 48 hour of incubation at 35°C. The confirmed test is used to accept or reject the presence of coliforms in a positive presumptive test. A small inoculum from a positive lactose broth is transferred to atube containing brilliant green lactose bile broth. The green dye and bile salts in this broth inhibit non-coliform growth. The presence of coliform is confirmed by growth and gas production within 48 hour at35°C. The Most Probable Number (MPN) of coliform can then be calculated from the number of confirmed tubes. False Positive Presumptive Tests: A positive, presumptive test does not necessarily mean that members of the colon group are present. In most cases it is true, but there are exceptions. False, positive, presumptivetests are caused by (1) the presence of other organisms capable of fermenting lactose with the production

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