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6095224 
Journal Article 
Use of discriminatory probes for strain typing of formalin-fixed, rabies virus-infected tissues by in situ hybridization 
Nadin-Davis, SA; Sheen, M; Wandeler, AI 
2003 
Yes 
Journal of Clinical Microbiology
ISSN: 0095-1137
EISSN: 1098-660X 
41 
9 
4343-4352 
English 
12958267 
An in situ hybridization (ISH) method has been developed to overcome difficulties encountered in the viral typing of formalin-fixed rabies virus-infected brain tissue. Rabies viruses representative of all strains normally encountered in diagnostic submissions throughout Canada, including 3 strains of terrestrial hosts (arctic fox, western skunk, mid-Atlantic raccoon), 10 strains circulating in several bat reservoirs (BBCAN1 to BBCAN7, LACAN, SHCAN, and MYCAN), and the Evelyn-Rokitniki-Abelseth (ERA) strain, used as an oral vaccine for fox rabies control in Ontario, were targeted. Partial phosphoprotein gene fragments generated from reverse transcription (RT)-PCR products of specimens of each viral type were molecularly cloned and used to produce negative-sense digoxigenin-labeled RNA transcripts. Conditions permitting the use of these transcripts as strain-specific probes were optimized by blotting analyses with RT-PCR amplicons generated with representative rabies viruses and by ISH applied to mouse brains inoculated with these strains. The successful application of this methodology to two rabies virus-positive specimens that were also identified by traditional methods and the retrospective typing of two archival rabies virus-positive equine specimens is described. This technique provides a typing regimen for rabies virus isolates submitted in a form that is normally recalcitrant to alternate typing strategies.