Visualization of Borrelia burgdorferi sensu lato by fluorescence in situ hybridization (FISH) on whole-body sections of Ixodes ricinus ticks and gerbil skin biopsies

The objective of this study was to visualize borreliae directly in whole-bo dy sections of Ixodes ricinus by fluorescence in situ hybridization (FISH), Borrelia afzelii mono-infected or Borrelia burgdorferi sensu stricto (ss)I B, afzelii double-infected nymphs were fixed, embedded in cold polymerizing resin and sectioned. The same sample processing was applied to skin biopsi es taken from a Mongolian gerbil after an infectious tick-bite, FISH was ca rried out using 16S-rRNA-directed, fluorescence-labelled oligonucleotide pr obes specific for the genus Borrelia and specific within the group of Lyme borreliosis-associated genospecies a. afzelii, a, burgdorferi ss, Borrelia garinii and Borrelia valaisiana, Sensitivity and specificity of the newly d esigned probes were evaluated using PCR, dot-blot hybridizations and FISH. Despite significant autofluorescence of certain tick tissues (which allowed good histological orientation within the sections), borreliae showing the typical spirochaetal morphotype were clearly visible in five out of six put atively infected ticks, These findings were confirmed by electron microscop y of ticks from the same infected batch as used for FISH. Attempts to produ ce ticks infected by two different Borrelia genospecies were not successful . FISH on whole-body sections of resin-embedded ticks offers the possibilit y of visualizing and identifying borreliae within tick tissues, This techni que has great potential for the investigation of the transmission of bacter ia or other micro-organisms by arthropod vectors, Furthermore, clear visual ization of single spirochaetes distributed along subcutaneous fat cell memb ranes in gerbil skin biopsies suggests that FISH might also be suitable for the detection of borreliae in clinical tissue specimens.