Molecular characterization ofBorrelia burgdorferi sensu lato from Slovenia revealing significant differences between tick and human isolates

Abstract

One hundred twenty-nine Slovenian isolates of Borrelia burgdorferi sensu lato derived from patients (69 strains) or Ixodes ricinus ticks (60 strains) were characterized. All of the strains were first- or second-passage isolates obtained in 1992 and 1993 from the same endemic region. The techniques used for the molecular analysis of strains included species-specific polymerase chain reaction (PCR) typing, and pulsed-field gel electrophoretic separation of undigested and MluI-digested genomic DNA. Isolates were identified to the species level by large restriction fragment pattern (LRFP) analysis and the results compared with the species-specific PCR result. Fifty-two patient isolates (75%) were typed as Borrelia afzelii (LRFP MLa1), 6 (9%) as Borrelia garinii (LRFPs MLg1-4), and 11 (16%) as Borrelia burgdorferi sensu stricto. The latter included 9 isolates (13%) with a new LRFP that is not typical of Borrelia burgdorferi sensu stricto and for which the designation MLx is suggested. In contrast, only 32 of 60 (53%) tick isolates were typed as Borrelia afzelii, while 20 strains (33%) were typed as Borrelia garinii and 8 strains (13%) as Borrelia burgdorferi sensu stricto. Three new LRFPs were found among the Borrelia garinii (MLg5 and 6) and Borrelia burgdorferi sensu stricto (MLb15) tick isolates. Large restriction fragment pattern analysis identified new groups of Borrelia burgdorferi sensu lato and revealed an apparent difference in the isolation frequency of different species from patients and ticks in the same endemic region.

Infection of Ixodes ricinus (Acari: Ixodidae) by Borrelia burgdorferi sensu lato in North Africa

“…burgdorferi sensu stricto, B. garinii, and B. afzelii. These genospecies have been isolated from patients, rodents, birds, and I. ricinus from various areas in Europe (Humair et al 1995, Olsen et al 1995, Picken et al 1996, Zhioua et al 1996. Two additional genospecies have been observed in I. ricinus and identiÞed as B. valaisiana and B. lusitaniae (formerly the genospecies VS116 and PotiB2, respectively) (Postic et al 1994Pé ter et al 1995;Rijpkema et al 1995;Le Flè che et al 1997;Wang et al 1997).…”

Comparison of Borrelia burgdorferi Sensu Lato Strains Isolated from Specimens Obtained Simultaneously from Two Different Sites of Infection in Individual Patients

“…After exhaustive block washing, DNA was restricted with 30 U of MluI restriction enzyme and restriction patterns were separated for 24 h with ramping pulse times of 1 to 40 s. Molecular size markers of 50 to 1,000 kb (Sigma) were included in each electrophoresis. The Borrelia species as well as the type within the particular species were determined according to restriction fragment length polymorphism (RFLP) as described by Belfaiza et al (2) and Picken et al (25). Fragments interpreted as being species specific were those with molecular sizes of 440, 320, and 90 kb for B. afzelii; 220 and 80 kb for B. garinii; and 145 kb for B. burgdorferi sensu stricto.…”

“…Fragments interpreted as being species specific were those with molecular sizes of 440, 320, and 90 kb for B. afzelii; 220 and 80 kb for B. garinii; and 145 kb for B. burgdorferi sensu stricto. Types within species (Mla for B. afzelii, Mlg for B. garinii, and Mlb for B. burgdorferi sensu stricto) were determined according to additional fragments of different molecular sizes (2,25).…”

“…), ticks, and reservoir hosts (9,17,25,29,42,49) obtained worldwide have been analyzed by many different phenotypic methods, such as well-established protein profile and serotyping system based on the heterogeneity of OspA and OspC proteins (33)(34)(35)(42)(43)(44)(45), and several genotypic approaches (42). Analysis of borrelial chromosomal or plasmid DNA, as performed by genotypic methods, provides more precise information on the genetic relationships among Borrelia strains (2, 5, 7, 22, 33-35, 42, 47).…”

“…It is of interest that the majority of such mixed infections were demonstrated by PCR, while as a rule only one species appeared after culturing of the sample (18,24,25,29,47,49). Theoretically this discrepancy could have been explained by false-positive PCR results and/or by an overgrowth of one species over the other in culture.…”