Francisella tularensis strain typing using multiple-locus, variable-numbertandem repeat analysis

Francisella tularensis, the etiological agent of tularemia, is found throug hout the Northern hemisphere. After analyzing the F. tularensis, genomic se quence for potential variable-number tandem repeats (VNTRs), we developed a multilocus VNTR analysis (MLVA) typing system for this pathogen. Variation was detected at six VNTR loci in a set of 56 Isolates from California, Okl ahoma, Arizona, and Oregon and the F. tularensis live vaccine strain. PCR a ssays revealed diversity at these loci with total allele numbers ranging fr om 2 to 20, and Nei's diversity index values ranging from 0.36 to 0.93. Clu ster analysis identified two genetically distinct groups consistent with th e current biovar classification system of F. tularensis. These findings sug gest that these VNTR markers are useful for identifying F. tularensis isola tes at this taxonomic level. In this study, biovar B isolates were less div erse than those in biovar A, possibly reflecting the history of tularemia i n North America. Seven isolates from a recent epizootic in Maricopa County, Ariz., were identical at all VNTR marker loci. Their identity, even at a h ypervariable VNTR locus, indicates a common source of infection. This demon strates the applicability of MLVA for rapid characterization and identifica tion of outbreak isolates. Future construction of reference databases will allow faster outbreak tracking as well as providing a foundation for deciph ering global genetic relationships.