The Lyme Disease Network
Conference Abstract


Title:Bb sensu lato Isolates from Missouri
Authors:Oliver JH; Kollars TM; Chandler FW
Conference:10th Annual International Scientific Conference on Lyme Disease & Other Tick-Borne Disorders, National Institutes of Health, Bethesda, MD April 28-30, 1997
Presenter:James H. Oliver, Jr., Ph.D.
Institute of Arthropodology & Parisitology
Georgia Southern University
Statesboro, GA

Abstract:
A total of 31 species of vertebrate animals were examined for presence of ticks and spirochetes at 20 study sites in southeastern Missouri. These included 22 mammals, 6 birds, and 3 reptile species. Ticks were also obtained by flagging/dragging the vegetation. Tick and/or host tissues were inoculated into BSKII medium in hopes of establishing laboratory cultures of spirochetes.

A total of 45 isolates were obtained from ticks attached to 15 eastern cottontail rabbits from 8 sites located in 5 counties. All isolates were from various developmental stages of Ixodes dentatus except one was from a Amblyomma americanum larva and one from a Haemaphysalis leporispalustris larva. The spirochetes were routinely screened by IFA monoclonal antibodies specific for Bb and included OspA (H5332, H3TS), OspB (H5TS,H6831, H614), and theBorrelia genus-specific H9724; two B. hermsii specific antibodies (9826, H4825) were also tested. The spirochete isolates were also analyzed by the PCR using several Bb specific ospA primers (788/944, 149/319, 149/459,3'/5'), fla (245/855), and Rosa's chromosomal primer (147/520). Several of the isolates were subjected to SDS-PAGE analysis and pulsed-field gel electrophoresis (PFGE).

Based on the above analyses all of the Missouri spirochete isolates are considered to be Bb sensu lato but are phenotypically and genetically different from Bb sensu stricto and in fact, appear to be rather similar to the genospecies B. andersonii. There are interesting differences among the Missouri isolates themselves. There are at least 5 immunological types and a dendrogram based on those data including 5 isolates from the Dowd farm and B. burgdorferi s.s., B. garinii, and B. hermsii suggests that MOD-5 is more closely related to B-31 B. burgdorferi s.s. than to MOD-1, MOD-2, MOD-3, and MOD-6. This relationship appears to be similar when a dendrogram is constructed based on data from pulsed-field gel electrophoresis. There are also at least 5 PCR types among the Missouri isolates thus far screened.

An ELISA for detecting antibodies against Missouri isolate MOD-1 was developed and analyses of sera from mammals collected in southeast Missouri thus far indicate high prevalences of antibodies in several species of hosts of ticks with the highest in cottontail rabbits followed by white-tailed deer, rodents, and raccoons. Western blots are being conducted to confirm positive ELISA results. Rabbits, deer, white-footed mice, cotton mice, and deer all show a prevalence of 65% or more positive for presence of IgG antibodies to the MOD-1 Bb isolate.

Five of the Missouri isolates were obtained from the Dowd farm which is the site of a human patient with a classical erythema migrans (EM) lesion and clinically diagnosed with LD. Interestingly, however, preliminary experiments involving needle injection of 2 of the isolates into mice failed to infect the mice.

Unique ID: 97LDF004


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