Rapid Detection of Heterotropic Bacteria in Water

What categorizes Heterotrophic organisms?

Heterotrophic organisms are categorized by the requirement of organic carbon sources to produce energy.  These organisms can include bacteria, yeasts and molds, and can be found throughout the environment in both natural and treated water (including tap water).  They occur normally and are unique in their ability to proliferate in the nutrient-poor environment of water systems at various temperatures. 

Why Test Water for Heterotrophs?


The concentration of Heterotrophs in treated water is an indicator of water treatment efficacy.  The presence of these microorganisms does not necessarily indicate concentrations high enough to cause disease.  However, the concentration of these microbes is important when testing for coliforms in water.  If Heterotrophic organisms are found at a concentration of approximately 500 colony forming units (cfu) per mL or higher, then the detection of coliforms may be compromised.  An increase of Heterotrophic counts may indicate the presence of coliforms in the water being tested.

Traditional methods for testing water

Heterotrophic Plate Count (HPC) is the procedure traditionally used to estimate the number of viable organisms in a sample of water.  Three different methods – (i) spread plate; (ii) pour plate; and (iii) membrane filtration – are utilized with different types of media.  Although these methods are widely accepted, there are some disadvantages.

  • In the pour plate technique, organisms are suspended in agar until the media cools.  During this time, damaged organisms are susceptible to heat shock from the higher temperature of the agar.  Also, because the colonies are surrounded by agar, the count of obligate anaerobes becomes compromised.
  • If an automated spreader is used in the spread plate method, heavy growth of colonies may make counting of colonies difficult, rendering counts non-applicable.
  • When filtering water, samples may contain suspended solids that cannot pass through the filter, interfering with accurate counts.

Due to different techniques and different types of media used, the counts achieved may vary significantly.  In addition to multiple protocols and media, temperature and incubation time vary.  Samples may be incubated anywhere from a few hours to 7 days or even a few weeks at temperatures ranging from 20°C to 40°C.  This makes processing water samples laborious with much data collection.

Rapid Detection with the BioLumix System 

BioLumix is the most advanced microbiological testing system of its kind. This automated, all-in-one microbial testing system is extremely easy to operate. The system is both simple and cost-effective, revolutionizing your current testing methodology. A novel optical system sensing color and fluorescence in ready-to-use vials provides faster results, labor savings, automation, and connectivity. The system has a large repertoire of assays that it can perform including: Total Aerobic Count, Yeast and Mold, Coliforms, Enterobacteriaceae, E. coli, Pseudomonas, Staphylococcus, and Salmonella.

Recently a new assay for the detection of Heterotrophic bacteria in water was developed for the BioLumix system.  The developed assay was validated by testing 50 samples of multiple types of water that were tested by the BioLumix method and the plate count method side-by-side.  The BioLumix vials were directly inoculated with 0.1 mL of the water sample, or 1.0 mL of a 1:100 dilution (depending on the desired specified level), and a few water samples were inoculated with heterotrophic bacteria.  These samples were monitored in the BioLumix instrument for 35 hours.  Figure 1 shows the curves obtained.

Final results were seen in the BioLumix system roughly 13 hours faster than the plate count method using Standard Methods Agar.  These particular samples were tested at specified levels <10 cfu/mL and <100 cfu/mL, but the BioLumix method can detect organisms at a level of <1 cfu/mL of water.  The BioLumix system is faster, less labor-intensive, and more sensitive than the plate count method. 

  Caron Ockerman