Microbial Analysis

Microbial Analysis



  1. Samples are observed as wet mounts to estimate numbers and types present
  2. An appropriate aliquot is fixed and stained with Lugol's Iodine
  3. Organisms are identified and enumerated using a Sedgewick-Rafter Counting Cell, or a settling chamber. A professional phycologist identifies the organisms using a large reference library, and culture collection.


  1. Samples for identification are prepared & dispensed onto a number of media for specific or general fungal isolation and/or enumeration. Agar plates are incubated at several temperatures and checked on a daily basis.
  2. Organisms are enumerated (either from agar isolations or direct) using an appropriate microscope (with/without stains; epi-fluoresence, phase, DF, Nomarski, or dissection microscopy).
  3. A professional mycologist identifies the organisms using a large reference library & culture collection.


A. Standard Methods for Coliforms (Total & Fecal) & Enterococci (Fecal Streptococcus)
  1. Multiple tube fermentation (15 or 10 tube)
  2. Membrane filter techniques
  3. Modified MacConkey for Fecal Coliforms
  4. Enzyme or antibody techniques

Methodology as per
Standard Methods for the Examination of Water & Wastewater, APHA, AWWA, WEF
Laboratory Procedures for the Examination of Seawater &_Shellfish 5th ed., 1985
FDA/AOAC Biological Analytical Manual 8th ed, 1995 + Revision, 1998
Compendium of Analytical Methods, Laboratory Procedures of Microbiological Analysis of Food, HC
U.S. Pharmacopeia & National Formulary USP 38 NF 33

All media and components are pre-tested for sterility and ability to support the appropriate growth. Test results are checked with gram stains and alternative tests for confirmation.

B. Microbial Identification for Bacteria, & Yeast
  1. Appropriate volume of sample is filtered through sterile 450nm membrane filter, or blended/diluted.
  2. The filter contents are suspended in 2mL heart infusion broth.
  3. Agar plates and liquid media are inoculated using appropriate volumes. Sample volumes are dispensed using calibrated loops (platinum APHA) & pipettes.
  4. Inoculated plates and tubes are incubated at appropriate temperatures (such as 4, 20, 28, 35, 44.5, 55C) in aerobic, C02 enriched or anaerobic atmospheres -C02, H2 or N).
  5. Colonies are enumerated and recorded.
  6. Representative colonies are removed and identified using routine microbiological tests.
  7. A professional microbiologist identifies the organisms using the tests, interactive identification programs and an extensive reference library and culture collection.
General Comments:

All filters are sterilized in individual filter holders to eliminate all cross-contamination between samples. All media are prepared in our laboratory and are pre-tested for sterility and ability to support the appropriate growth. Test results are checked with gram stains and alternative tests for confirmation.

Time Required for Microbial Identification:

Time will vary between a few days to several weeks. This is because the number and types of tests required and the organism(s)’s rate of growth will be dictated by the microbe not the laboratory.


All methods conform to established protocols derived from -
International Subcommittees on Taxonomy, IJSB
FDA/AOAC Bacteriological Analytical Manual 8th ed, 1995 + Revision A, 1998
Bergey’s Manual of Systematic Bacteriology vol 1-4
Yeasts Characteristics and Identification, Barnett, Payne & Yarrow, 1983
The Prokaryotes, Starr et.al. 1981

Micro Identification Tests for Bacteria and Yeast

Typical Tests or determinations required to identify an organism

  • hemolysis
  • colony description
  • atmospheric requirements (temp, 02)
  • gram stain + confirmation with KOH
  • tellurite reduction
  • H2S production (TSI)
  • urease
  • catalase
  • oxidase
  • motility
  • nitrate-nitrite reduction
  • citrate
  • O/F carbohydrates:
    glucose, lactose, mannitol, sucrose
  • indole
  • decarboxylase
  • litmus milk
  • coagulase
  • lysostaphin
  • bile solubility
  • gelatin
  • aesculin
  • O/F carbohydrates (additional):
    arabinose, rhamnose, fructose, galactose, mannose, maltose, trehalose, melibiose, starch cellibiose, raffinose, simmons inulin, dextrin, glycogen, glycerol, sorbitol, dulcitol
  • selective stains:
    DAPI, geimsa, acid-fast
  • antibiotic sensitivities (18)
  • digitonin (sensitivity)
  • fatty acid profile (MIDI)
  • VP methyl-red
Organisms identified as per: 

Beregy's Manual of Systematic Bacteriology vols 1-4
Bergey's Manual of Determinative Bacteriology 8th edition
International Journal of Systematic Bacteriology
International Subcommittees on Microbial Taxonomy + IJSB
The Prokaryotes Vol1
The Genera of Bacteria 2nd Ed (Skerman)
Yeasts Characteristics and Identification (Barnett, Payne,
and Yarrow)


  • all tests adhere to internationally recognized methods. Where no ‘official’ methodology exists, all analysis are conducted with verifiable scientific practices
  • all test media are pre-tested for ability to support appropriate growth
  • positive and negative controls are used routinely
  • temperatures are monitored electronically on a continuous basis
  • shellfish are prepared as per Laboratory Procedures for the Examination of Seawater & Shellfish APHA 5th ed 1985. 6. 3-5 sub-samples are used throughout the protocols.
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