title: Application of mitochondrial DNA analysis for microbial source tracking purposes in shellfish harvesting waters.
abstract: We present a method for the reliable detection and source characterisation of faecal pollution in water and shellfish matrices, utilising real-time PCR analysis of mitochondrial DNA targets. In this study we designed real-time PCR (TaqMan) probes to target human, bovine, ovine and swine mtDNA. PCR amplification using species-specific TaqMan probes on faecal matter and mixed effluent slurries revealed no cross-reactions between species of interest and other vertebrate faecal matter. Performed as a single blind experiment we were able to correctly identify faecal material in 17/20 effluents (85% correct). mtDNA degrades relatively quickly in faecally-spiked water samples (approximately 2 weeks), a similar timeframe of environmental persistence to several bacterial faecal indictors, highlighting its applicability. The procedure described here is specific, rapid (<5 hours) and sensitive. These results confirm the suitability of using species-specific mtDNA as an indicator in source tracking studies in surface waters, shellfish harvesting areas and shellfish matrices.
authors.affiliation: Centre for Environment, Fisheries and Aquaculture Science, Weymouth Laboratory, Weymouth, Dorset, DT4 8UB, UK. craig.baker-austin@cefas.co.uk
authors.forename: Craig
authors.initials: C
authors.lastname: Baker-Austin
authors.forename: Rachel
authors.initials: R
authors.lastname: Rangdale
authors.forename: James
authors.initials: J
authors.lastname: Lowther
authors.forename: David N
authors.initials: DN
authors.lastname: Lees
mesh_terms: D000818:Animals; D002417:Cattle; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D017931:DNA Primers; D004272:DNA, Mitochondrial / Q000235:genetics*; D005243:Feces / Q000382:microbiology; D006801:Humans; D012756:Sheep; D012758:Shellfish / Q000382:microbiology*; D013045:Species Specificity; D013552:Swine
chemical_list: D017931:DNA Primers; D004272:DNA, Mitochondrial; D004247:DNA