A Microarray Based Approach for the Identification of Common Foodborne Viruses

Ayodeji, Mobolanle; Kulka, Michael; Jackson, Scott A; Patel, Isha; Mammel, Mark; Cebula, Thomas A; Goswami, Biswendu B

We investigated whether hybridization of fluorescently labeled amplified DNA (target) to a microarray containing many oligonucleotide probes representing many different viral genomes can identify a virus without sequencing. Unlike sequencing, these arrays can interrogate thousands of bases of a viral genome in a single experiment [15] [16] [17] . We determined the feasibility of this approach by using labeled targets amplified from either the DNA (i.e. as recombinant plasmid) or RNA from several strains of HAV and CV. As shown in Figs. (4-6) , a single hybridization experiment using a multi-well array with different samples loaded in different wells of a 12-well sample pod can identify HAV and CVB by the unique profile generated with no ambiguity or crosshybridization to oligonucleotides representing an unrelated virus. Within the broad genus of hepatovirus of which HAV is the only species member, different genotypes which differ from each other by 5% to 8% of base positions (Fig. 1) can be identified (Figs. 4-6) . Within the same subgenotye Ib, strains such as wild type HM175 and the cell culture adapted variants including the cytopathic 18f strain differ by only 0.5% of base positions. We have shown that differentiation of these strains is possible by analyzing the ratio of the signal probe intensities generated by the isolates when hybridized to the probe sets present on this tiling array ( Fig. 6 and Table 5 ). A sequence based identification of the same 3.7 kb amplicon would require several sequencing reactions with multiple primers in order to identify nucleotide differences. In addition, mutations accumulating in the HM175 genome during its evolution into the cytopathic 18f strain can be identified by ratio analysis (Fig. 6 and Table 5 ). Thus, the present array design is suitable for identification of species (e.g. CV and HAV) and HAV subgenotypes since in the latter case the nucleotide differences are very few.