When the Sum of the Parts Tells You More Than the Whole: The Advantage of Using Metagenomics to Characterizespp. Infections in Norway Rats () and Their Fleas.

Abstract

Urban Norway rats () are a reservoir forspp. - a genus of zoonotic bacteria transmitted by hematophagous vectors, particularly fleas. Rats and fleas may be infected with more than onespecies; however, mixed infections may be difficult to detect using culture and/or mono-locus PCR. We set out to characterizespp. usingPCR and Sanger sequencing on blood (= 480) andflea pools (= 200) obtained from a population of urban Norways rats from Vancouver, Canada. However, when contamination of a subset of flea pools necessitated the use of a second target () and the results ofandwere discordant, a metagenomic approach was used to better characterize thespp. present in these samples and our objective transitioned to comparing data obtained via metagenomics to those from PCR/sequencing. Among thespp.-positive rats (= 95), 52 (55.3%), and 41 (43.6%) had Sanger sequences consistent withand, respectively. One rat had a mixed infection. All sequences fromspp.-positive flea pools (= 85), were consistent with, and re-analysis of 34 bloods of varyingspp. infection status (basedPCR and sequencing) usingPCR showed that the assay was capable of identifyingbut not. Metagenomics analysis of a subset of PCR-positive blood samples (= 70) and flea pools (= 24) revealed that bothandwere circulating widely in the study population with 31/70 (44.3%) rats and 5/24 (2.1%) flea pools infected with both species., however, made up a smaller relative proportion of the reads for samples with mixed infections, which may be why it was generally not detected by genus-specific PCR and Sanger sequencing. Further analysis of 16S-23S ITS sequences amplified from a subset of samples identified thestrain assubsp.type II. This demonstrates the value of a metagenomic approach for better characterizing the ecology and health risks associated with this bacterium, particularly given that the less dominant species,is associated with greater pathogenicity in people.