Analysis ofBorrelia burgdorferivlsEGene Expression and Recombination in the Tick Vector

Abstract

Expression and recombination of the antigenic variationvlsEgene of the Lyme disease spirocheteBorrelia burgdorferiwere analyzed in the tick vector. To assessvlsEexpression,Ixodes scapularisnymphs infected with theB. burgdorferistrain B31 were fed on mice for 48 or 96 h or to repletion and then crushed and acetone fixed either immediately thereafter (ticks collected at the two earlier time points) or 4 days after repletion. Unfed nymphs also were examined. At all of the time points investigated, spirochetes were able to bind a rabbit antibody raised against the conserved invariable region 6 of VlsE, as assessed by indirect immunofluorescence, but not preimmune serum from the same rabbit. This same antibody also bound to B31 spirochetes cultivated in vitro. Intensity of fluorescence appeared highest in cultured spirochetes, followed by spirochetes present in unfed ticks. Only a dim fluorescent signal was observed on spirochetes at the 48 and 96 h time points and at day 4 postrepletion. Expression ofvlsEin vitro was affected by a rise in pH from 7.0 to 8.0 at 34°C. Hence,vlsEexpression appears to be sensitive to environmental cues of the type found in theB. burgdorferinatural history. To assessvlsErecombination, nymphs were capillary fed theB. burgdorferiB31 clonal isolate 5A3. Ticks thus infected were either left to rest for 4 weeks (Group I) or fed to repletion on a mouse (Group II). The contents of each tick from both groups were cultured and 10B. burgdorfericlones from the spirochetal isolate of each tick were obtained. ThevlsEcassettes from several of these clones were amplified by PCR and sequenced. Regardless of whether the isolate was derived from Group I or Group II ticks, no changes were observed in thevlsEsequence. In contrast,vlsEcassettes amplified fromB. burgdorfericlones derived from a mouse that was infected with B31-5A3 capillary-fed nymphs showed considerable recombination. It follows thatvlsErecombination does not occur in the tick vector.