Development of discriminatory multiple-locus variable number tandem repeat analysis for Bartonella henselae

Bartonella henselae is a zoonotic bacterium that infects cats and humans. Several attempts have been made to develop typing techniques for epidemiological purposes; however, most of the techniques developed do not appear to be sufficiently discriminatory or easy to use. In order to develop multilocus variable number tandem repeat (VNTR) analysis (MLVA) for B. henselae, 30 VNTR candidates were selected from the genome sequence of the reference strain Houston 1 (H1). The VNTR candidates were initially tested by PCR on six B. henselae isolates from different geographical areas. Five VNTRs were selected from those that showed two or more alleles. These five B henselae VNTRs (BHVs) were tested on 42 feline B. henselae isolates and strains from France (23 isolates), Denmark (17 isolates), the Philippines (one isolate) and the USA (F1 strain), on one human isolate from Germany, and on the H1 reference strain. These BHVs were sufficiently discriminatory to obtain 31 different profiles (corresponding to two different groups) among the 44 isolates and strains of B. henselae tested. Thirty-five profiles were obtained using these BHVs and two variant alleles. The combination of the five markers led to a diversity index of 0.98. The stability of the five BHVs was demonstrated on the feline F1 strain, with no change in stability observed after 2, 21 and 41 passages. This is believed to be the first study conducted on B. henselae typing using MLVA, and it demonstrates the high quality of this technique for discriminating between B. henselae isolates.