Subtyping of Canadian isolates of Salmonella Enteritidis using Multiple Locus Variable Number Tandem Repeat Analysis (MLVA) alone and in combination with Pulsed-Field Gel Electrophoresis (PFGE) and phage typing

Salmonella enterica subspecies enterica serovar Enteritidis (SE) is one of the most common causes of human salmonellosis and in Canada currently accounts for over 40% of human cases. Reliable subtyping of isolates is required for outbreak detection and source attribution. However, Pulsed-Field Gel Electrophoresis (PFGE), the current standard subtyping method for Salmonella spp., is compromised by the high genetic homogeneity of SE. Multiple Locus Variable Number Tandem Repeat Analysis (MLVA) was introduced to supplement PFGE, although there is a lack of data on the ability of MLVA to subtype Canadian isolates of SE. Three subtyping methods, PFGE, MLVA and phage typing were compared for their discriminatory power when applied to three panels of Canadian SE isolates: Panel 1: 70 isolates representing the diversity of phage types (PTs) and PFGE subtypes within these PTs; Panel 2: 214 apparently unrelated SE isolates of the most common PTs; and Panel 3: 27 isolates from 10 groups of epidemiologically related strains. For Panel 2 isolates, four MLVA subtypes were shared among 74% of unrelated isolates and in Panel 3 isolates, one MLVA subtype accounted for 62% of the isolates. For all panels, combining results from PFGE, MLVA and PT gave the best discrimination, except in Panel 1, where the combination of PT and PFGE was equally as high, due to the selection criteria for this panel. However, none of these methods is sufficiently discriminatory alone for reliable outbreak detection or source attribution, and must be applied together to achieve sufficient discrimination for practical purposes. Even then, some large clusters were not differentiated adequately. More discriminatory methods are required for reliable subtyping of this genetically highly homogeneous serovar. This need will likely be met by whole genome sequence analysis given the recent promising reports and as more laboratories implement this tool for outbreak response and surveillance.