Development of a qPCR assay to identify and differentiate insect-associated strains of the <i>Serratia marcescens</i> complex

The <i>Serratia marcescens</i> complex contains important opportunistic pathogens of humans and vertebrate animals, as well as insects and other invertebrates. To date, the methods used for the identification of species within the genus <i>Serratia</i>, including PCR assays, have poor discriminatory power and may require further molecular typing or genomic sequence analysis to determine clinical relevance. We developed a duplex TaqMan probe-based quantitative real-time PCR (qPCR) assay targeting the <i>chiP</i> gene, which is involved in chitin degradation and transport, and the <i>ureD</i> gene, which is involved in urease production. This allowed us to simultaneously identify all members of the <i>S. marcescens</i> complex (<i>chiP</i> positive) and differentiate those most likely to act as insect pathogens (<i>chiP</i> and <i>ureD</i> positive). We applied our assay to identify potentially entomopathogenic members of the <i>S. marcescens</i> complex in the context of a conservation program for the critically endangered insect <i>Dryococelus australis</i> and found it to be a useful aid for rapid and accurate detection of infection with <i>S. marcescens</i> complex strains in insects and determination of their clinical relevance. By targeting 2 genes likely to be virulence factors, this assay may also be of use for research investigating the pathogenesis of entomopathogenic <i>Serratia</i> spp.