New diagnostic methods for Escherichia marmotae and the first report of its identification in clinical isolates in North America

Abstract

Background: Genomic sequences of E. marmotae and E. coli differ by 10%. Discovered as an environmental "cryptic clade" of Escherichia, E. marmotae also occurs in human infections. Microbiological and MALDI-TOF-MS methods frequently misidentify E. marmotae as E.coli. Our goal was to develop methods that reliably distinguish E. marmotae from E. coli to improve therapeutic decisions and treatments.

Methods: A Taqman PCR method was developed to distinguish E. marmotae from E. coli based on genomic sequences of uidA, uidB, and a positive control targeting adk in E. marmotae and E. coli. MALDI-TOF-MS spectra were obtained for environmental and clinical isolates using a bioMérieux VITEK MALDI-TOF-MS system.

Results: UidA- and uidB species-specific PCR amplified DNA from E. marmotae with 100% specificity, and not from E. coli or other Escherichia species. The Biomérieux VITEK MALDI-TOF-MS consistently misidentified E. marmotae as E. coli, with median IVD confidence scores for both E. marmotae and E. coli of 99.9%; however, RUO scores for E. marmotae (median 0%) were significantly lower (P < 0.0001) than for E. coli (median = 87.4%). The spectral peak between m/z 7,250 to 7,280 consistently occurred between 7,260 and 7,268 in E. marmotae and only between 7,268 and 7,280 in E. coli, with no overlap (p < 0.001). Application of these spectral criteria to 176 clinical isolates revealed the first identification of a E. marmotae isolate from a human infection in North America. The isolate had originally been diagnosed as E. coli based on a 99.1% IVD confidence score. This first North American clinical isolate was confirmed as E. marmotae by Taqman-PCR and whole genome sequencing. This isolate had numerous antibiotic resistance gene markers and unlike most clinical E. coli, this E. marmotae isolate lacked motility at 37°C.

Conclusion: Clinical tests based on these methods of differentiating E. marmotae and E. coli may assist in determining the prevalence of this emerging pathogen and making therapeutic decisions.

Keywords: Escherichia marmotae; MALDI-TOF-MS; VITEK MS; bacterial identification; bioMérieux; qPCR; species-specific qPCR; spectral peaks.