Microarray for Identification of the Chiropteran Host Species of Rabies Virus in Canada

Abstract

Species identification through genetic barcoding can augment traditional taxonomic methods, which rely on morphological features of the specimen. Such approaches are especially valuable when specimens are in poor condition or comprise very limited material, a situation that often applies to chiropteran (bat) specimens submitted to the Canadian Food Inspection Agency for rabies diagnosis. Coupled with phenotypic plasticity of many species and inconclusive taxonomic keys, species identification using only morphological traits can be challenging. In this study, a microarray assay with associated PCR of the mitochondrial cytochrome c oxidase subunit I (COI) gene was developed for differentiation of 14 bat species submitted to the Canadian Food Inspection Agency from 1985–2012 for rabies diagnosis. The assay was validated with a reference collection of DNA from 153 field samples, all of which had been barcoded previously. The COI gene from 152 samples which included multiple specimens of each target species were successfully amplified by PCR and accurately identified by the microarray. One sample that was severely decomposed failed to amplify with PCR primers developed in this study, but amplified weakly after switching to alternate primers and was accurately typed by the microarray. Thus, the chiropteran microarray was able to accurately differentiate between the 14 species of Canadian bats targeted. This PCR and microarray assay would allow unequivocal identification to species of most, if not all, bat specimens submitted for rabies diagnosis in Canada.

The phylogeography of Myotis bat-associated rabies viruses across Canada

“…To confirm species identification, genetic barcoding that targeted a portion of the cytochrome oxidase subunit 1 (COI) gene was performed. The DNA was extracted from tissue (lung or skin that included several hair follicles) recovered from each carcass, and employed as template for COI amplification as previously described [ 20 ] or, in some cases, a universal primer pair suitable for COI PCR of all indigenous Canadian bat species was employed [ 23 ]. For some specimens for which a carcass was no longer available, total RNA recovered from original bat brain tissue was used as template for a reverse transcriptase-PCR (RT-PCR) in which cDNA was primed with the reverse sense primer.…”

Rabies – epidemiology, pathogenesis, public health concerns and advances in diagnosis and control: a comprehensive review

“…DNA microarray has been used as a diagnostic tool for detection of nucleic acid segment of lyssaviruses including RABV (Lung et al 2013). The tailored microarray has got the capability of detecting as well as distinguishing all the seven recognised members of the genus Lyssavirus by use of 624 70mer probes, both in humans and animals.…”