Cow Bacteroidetes ID™

The phylum Bacteroidetes is composed of three large groups of bacteria with the best-known category being Bacteroidaceae. This family of gram-negative bacteria is found primarily in the intestinal tracts and mucous membranes of warm-blooded animals and is sometimes considered pathogenic.

Comprising Bacteroidaceae are the genus Bacteroides and Prevotella. The latter genus was originally classified within the former (i.e. Bacteroides), but since the 1990's it has been classified in a separate genus because of new chemical and biochemical findings. Bacteroides and Prevotella are gram-negative, anaerobic, rod-shaped bacteria that inhabitant of the oral, respiratory, intestinal, and urogenital cavities of humans, animals, and insects. They are sometimes pathogenic.

Fecal Bacteroidetes are considered for several reasons an interesting alternative to more traditional indicator organisms such as E. coli and Enterococci. Since they are strict anaerobes, they are indicative of recent fecal contamination when found in water systems. This is a particularly strong reference point when trying to determine recent outbreaks in fecal pollution. They are also more abundant in feces of warm-blooded animals than E. coli and Enterococci. Furthermore, these latter two organisms are facultative anaerobes and as such they can be problematic for monitoring purposes since it has been shown that they are able to proliferate in soil, sand and sediments.

The Cow Bacteroidetes ID™ service is designed around the principle that fecal Bacteroidetes are found in large quantities in feces of warm-blooded animals. Furthermore, certain categories of Bacteroidetes have been shown to be predominately detected in cattle. Within these Bacteroidetes, certain strains of the Bacteroides and Prevotella genus have been found in cattle. As such, these bacterial strains can be used as indicators of cattle fecal contamination.

One of the advantages of the Cow Bacteroidetes ID™ service is that the entire water is sampled and filtered for fecal Bacteroidetes. As such, this method avoids the randomness effect of culturing and selecting bacterial isolates off a petri dish. This is a particular advantage for highly contaminated water systems with potential multiple sources of fecal contamination.

Accuracy of the results is possible because the method uses PCR DNA technology. PCR allows quantities of DNA to be amplified into large number of small copies of DNA sequences. This is accomplished with small pieces of DNA called primers that are complementary and specific to the genomes to be detected.

Through a heating process called thermal cycling, the double stranded DNA is denatured and inserted with complementary primers to create exact copies of the DNA fragment desired. This process is repeated rapidly many times ensuring an exponential progression in the number of copied DNA. If the primers are successful in finding a site on the DNA fragment that is specific to the genome to be studied, then billions of copies of the DNA fragment will be available for detection by gel electrophoresis.

The gel electrophoresis apparatus uses an electrical field to distinguish different DNA fragments according to their molecular weights. Lighter DNA fragments will move farther along the gel than their heavier counterparts. At the end of the procedure different bands of accumulated DNA fragments will aggregate at different parts of the gel. It is this accumulation of DNA fragments that creates a band on the gel. Researchers use these bands to distinguish certain genomes such as the cattle gene biomarker from the Bacteroides and Prevotella genus.

These banding patterns confirm or negate the presence of the fecal Bacteroidetes cattle gene biomarker. As such, the banding patterns can be a good indicator of cattle fecal contamination. Nonetheless, in order to strengthen the validity of the results, the Cow Bacteroidetes ID™ service should be combined with other DNA analytical services such as the Cow Fecal Virus ID™ services since the fecal Bacteroidetes cattle gene biomarker has been detected occasionally in other ruminants.