New Molecular Quantitative PCR Assay for Detection of Host-Specific Bifidobacteriaceae Suitable for Microbial Source Tracking

Abstract

Bifidobacterium spp. belong to the commensal intestinal microbiota of warm-blooded animals. Some strains of Bifidobacterium show host specificity and have thus been proposed as host-specific targets to determine the origin of fecal pollution. Most strains have been used in microbial-source-tracking (MST) studies based on culture-dependent methods. Although some of these approaches have proved very useful, the low prevalence of culturable Bifidobacterium strains in the environment means that molecular culture-independent procedures could provide practical applications for MST. Reported here is a set of common primers and four Bifidobacterium sp. host-associated (human, cattle, pig, and poultry) probes for quantitative-PCR (qPCR) assessment of fecal source tracking. This set was tested using 25 water samples of diverse origin: urban sewage samples, wastewater from four abattoirs (porcine, bovine, and poultry), and water from a river with a low pollution load. The selected sequences showed a high degree of host specificity. There were no cross-reactions between the qPCR assays specific for each origin and samples from different fecal origins. On the basis of the findings, it was concluded that the host-specific qPCRs are sufficiently robust to be applied in environmental MST studies. Waterborne diseases transmitted by the fecal-oral route make a significant contribution to the burden of disease worldwide (49). Consequently, fecal pollution in water is a global public health concern. This pollution has several origins: direct discharge of fecal waste or raw wastewater, secondary effluents from wastewater treatment works, combined sewer and sanitary overflows, animal husbandry activities, wastewater from abattoirs and the meat industry, and wildlife (47). Source-tracking methods allow the origin of fecal pollution in a particular body of water to be determined (19). The identification of fecal-pollution sources accompanied by appropriate water resource management policies could contribute to improving the microbial quality of water (9,22).Several chemical and microbiological methods have been proposed for tracking the origin of fecal pollution in water (10). Unfortunately, the most commonly used fecal indicators of microorganisms, such as fecal coliforms (FC), Escherichia coli, enterococci, and sulfite-reducing clostridia (SRC), are found in both human and animal feces, and thus, they are of limited use in identifying the origin of waste. Alternative microbial indicators may provide information about the presence and origin of fecal pollution. They include antibiotic-resistant enterococcus phenotypes (18), bacteriophages of Bacteroides spp. (17,23,38), F-RNA bacteriophage subgroups (21), Rhodococcus coprophilus (41), Bacteroides group (8, 39), Bifidobacterium spp. (3,6,7,11,25,31), and biochemical fingerprinting of bacterial populations (32).Bifidobacterium spp. were first proposed as microbial-sourcetracking (MST) indicators in 1983, when sorbitol fermentation was related to a possible human source (33). Bifidobacte...

Discovering new indicators of fecal pollution

“…By far, the most explored taxonomic group are the Bacteroidales [7–10], which are detailed later. Studies have also focused on Firmicutes [11, 12], Bifidobacteria [11, 13–16], and Methanobrevibacter smithii , a common archaeon in the intestine [17, 18]. Fecal anaerobes may be more indicative of the presence of pathogens because it is unlikely they will grow once released from their host into the environment, in contrast to E. coli and enterococci, which have been shown to persist and even grow in beach sand [19–22], algal mats [20, 23], and sediment [24–26].…”

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