Detection of Bovine Group A Rotavirus Using Rapid Antigen Detection Kits, RT-PCR and Next-Generation DNA Sequencing

MINAMI-FUKUDA, Fujiko; NAGAI, Makoto; TAKAI, Hikaru; MURAKAMI, Toshiaki; OZAWA, Tadashi; TSUCHIAKA, Shinobu; OKAZAKI, Sachiko; KATAYAMA, Yukie; OBA, Mami; NISHIURA, Naomi; SASSA, Yukiko; OMATSU, Tsutomu; FURUYA, Tetsuya; KOYAMA, Satoshi; SHIRAI, Junsuke; TSUNEMITSU, Hiroshi; FUJII, Yoshiki; KATAYAMA, Kazuhiko; MIZUTANI, Tetsuya

There are many assays, such as those using electron microscopy, virus isolation and gel electrophoresis of genomes, for the detection of rotavirus in specimens [12, 18, 20] . Though these methods are gold standards for diagnosis of bovine RVA, they require expensive equipment and complicated techniques. In addition, it is difficult to isolate RVA using cell lines [18] . In the clinical field, clinical veterinarians need rapid and sensitive detection methods that are routinely available in daily clinical practice. Of the seven commercial kits in this study, the Dipstick showed the highest sensitivity to the three bovine RVA strains, and these results were consistent with those obtained with equine RVA [11] . Although the sensitivity of the Dipstick was lower that than with RT-PCR, the Dipstick would be useful for field diagnosis because of its simple, easy and rapid procedure. RT-PCR has been employed as a useful method for the detection of pathogens. In Japan, the primer pair Beg9/End9, which has been applied to each human serotype virus [4] , is usually used in livestock hygiene service centers for the detection and genotyping of VP7 of bovine RVA. Genotyping VP7 using this primer pair is useful. In a limited dilution study, the primer pair Beg9/End9 amplified VP7 of KK3 up to a 10 3 dilution; on the other hand, the primer pair BRVF/BRVR amplified VP6 of KK3 up to a 10 5 dilution. The reason for this may be the variability of the VP7 reverse primer region, and this indicates that amplification of VP6 is better for detection of bovine RVA.