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    Microbial Analysis of Water

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    Stephen Moore
    The document discusses microbial analysis of water samples to detect indicator organisms like coliform bacteria. It outlines the ideal characteristics of an indicator organism and describes several common tests used to detect coliforms, including the presumptive, confirmed and completed tests, membrane filter technique, presence-absence test, and Colilert defined substrate test. Coliforms are facultatively anaerobic, gram-negative rods that ferment lactose with gas within 48 hours at 35°C, and fecal coliforms grow at 44.5°C to distinguish them from other sources.

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    Microbial Analysis of Water

    Uploaded by

    Stephen Moore
    226 views9 pages
    The document discusses microbial analysis of water samples to detect indicator organisms like coliform bacteria. It outlines the ideal characteristics of an indicator organism and describes several common tests used to detect coliforms, including the presumptive, confirmed and completed tests, membrane filter technique, presence-absence test, and Colilert defined substrate test. Coliforms are facultatively anaerobic, gram-negative rods that ferment lactose with gas within 48 hours at 35°C, and fecal coliforms grow at 44.5°C to distinguish them from other sources.

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    The document discusses microbial analysis of water samples to detect indicator organisms like coliform bacteria. It outlines the ideal characteristics of an indicator organism and describes several common tests used to detect coliforms, including the presumptive, confirmed and completed tests, membrane filter technique, presence-absence test, and Colilert defined substrate test. Coliforms are facultatively anaerobic, gram-negative rods that ferment lactose with gas within 48 hours at 35°C, and fecal coliforms grow at 44.5°C to distinguish them from other sources.

    Copyright:

    © All Rights Reserved
    Download as PPTX, PDF, TXT or read online from Scribd
    Download as pptx, pdf, or txt
    226 views9 pages

    Microbial Analysis of Water

    Uploaded by

    Stephen Moore
    The document discusses microbial analysis of water samples to detect indicator organisms like coliform bacteria. It outlines the ideal characteristics of an indicator organism and describes several common tests used to detect coliforms, including the presumptive, confirmed and completed tests, membrane filter technique, presence-absence test, and Colilert defined substrate test. Coliforms are facultatively anaerobic, gram-negative rods that ferment lactose with gas within 48 hours at 35°C, and fecal coliforms grow at 44.5°C to distinguish them from other sources.

    Copyright:

    © All Rights Reserved
    Download as PPTX, PDF, TXT or read online from Scribd
    Download as pptx, pdf, or txt
    of 9

    Microbial Analysis of water

    Indicator organism:
    The indicator bacterium should be suitable for the analysis of all types of
    water: tap, river, ground, impounded, recreational, estuary, sea, and waste.
    It should be present whenever enteric pathogens are present.
    It should survive longer than the hardiest enteric pathogen.
    It should not reproduce in the contaminated water, as this would produce an
    inflated value.
    It should be harmless to humans.
    Its level in contaminated water should have some direct
    relationship to the degree of fecal pollution.
    The assay procedure for the indicator should have great specificity; in other
    words, other bacteria should not give positive results. In addition, the
    procedure should have high sensitivity and detect low levels of the indicator.
    The testing method should be easy to perform.

    Coliforms are defined as facultatively aerobic, gram-negative, nonspore-forming, rod-shaped bacteria that ferment lactose with gas
    formation within 48 hours at 35C.
    The coliforms include a wide range of bacteria whose primary
    source may not be the intestinal tract. To address this, tests have
    been developed that assay for the presence of fecal coliforms.
    These are coliforms derived from the intestine of warm blooded
    animals, which grow at the more restrictive temperature of 44.5C.
    Enterobacter; Escherichia coli, a common intestinal organism and
    occasional pathogen; and Klebsiella pneumoniae

    The original test for coliforms involved the


    presumptive, confirmed, and completed tests
    The presumptive step is carried out by means of tubes
    inoculated with three different sample volumes to give an
    estimate of the most probable number (MPN) of coliforms
    in the water.
    The complete process, including the confirmed and
    completed tests, requires at least 4 days of incubations
    and transfers.

    The confirmation test:


    The confirmed test is done by selective and differential media such
    as eosinmethylene blue (EMB) or Endo agar.
    The medium will be streaked from a positive lactose broth tube
    obtained from the presumptive test.
    Eosinmethylene blue contains the dye methylene blue, which
    inhibits the growth of gram-positive organisms.
    In the presence of an acid environment, EMB forms a complex that
    precipitates out onto the coliform colonies, producing dark centers
    and a green metallic sheen. The reaction is characteristic for
    Escherichia coli,

    The membrane filter technique:


    The water sample is passed through a membrane filter.
    The filter with its trapped bacteria is transferred to the
    surface of a solid medium or to an absorptive pad
    containing the desired liquid medium.
    A resuscitation step often is needed with chlorinated
    samples, where the microorganisms are especially
    stressed. Membrane filters have been widely used with
    water that does not contain high levels of background
    organisms, sediment, or heavy metals.

    The presence-absence (P-A) test:


    This is a modification of the most probable number
    procedure in which a larger water sample (100 milliliters) is
    incubated in a single culture bottle with a triple-strength
    broth containing lactose broth, lauryl tryptose broth, and
    bromocresol purple indicator.
    The P-A test is based on the assumption that no coliforms
    should be present in 100 milliliters of drinking water. A
    positive test results in the production of acid (a yellow
    color), which requires confirmation.

    Colilert defined substrate test:


    A water sample of 100 milliliters is added to
    a specialized medium containing onitrophenyl-13-ogalactopyranoside (ONPG)
    and 4-methylumbelliferyl-13-oglucuronide
    (MUG) as the only nutrients.
    If coliforms are present, the medium will
    turn yellow within 24 hours at 35C due to
    the hydrolysis of ONPG, which releases onitrophenol (figure 43.3).
    To check for E. coli, the medium is observed
    under long-wavelength ultraviolet (UV) light
    for fluorescence.
    When E. coli is present, MUG is modified to
    yield a fluorescent product.

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