Mitochondrial DNA marker: A PCR approach for tracking rat (Rattus rattus and Rattus norvegicus) fecal pollution in surface water systems

Abstract

Rats act as reservoirs for a wide range of zoonotic pathogens and can negatively impact human health. In this study, we developed a novel dye base mitochondrial DNA (mtDNA) PCR-assay (RatMt) specifically targeting a 180 bp fragment of the NADH dehydrogenase subunit 2 gene for detecting fecal pollution from two species of rats (Rattus rattus and Rattus norvegicus) in environmental samples. Estimation of Escherichia coli concentrations in Rattus norvegicus fecal pellets suggested that there were approximately 2.24 × 10 4 ± 4.86 × 103 MPN/g of fecal pellet. The RatMt PCR assay was robust, had a detection limit of rat feces in water of 0.274 ± 0.14 mg/100 mL and was 100 % specific for detecting Rattus rattus and Rattus norvegicus fecal mtDNA. Fecal Indicator Bacteria (FIB) along an urbanized gradient in Pensacola-Bay was assessed by the IDEXX Colilert™ − 18 and indicated that the majority of the fifteen sampling sites in the Pensacola-Bay area had E. coli concentrations >410 MPN/100 mL. Rattus rattus and Rattus norvegicus mtDNA were detected in all the urban marine sites, three of the urban freshwater sampling areas, and three of the forested sampling sites. The RatMt PCR assay is a useful tool for rapidly detecting Rattus rattus and Rattus norvegicus fecal pollution in environmental samples.