Koizumi Shunjiro, Nishi Tatsuya, Morioka Kazuki, Fukai Katsuhiko
Saitama Prefectural Chuo Livestock Hygiene Service Center, Saitama, Japan.
Kodaira Research Station, National Institute of Animal Health, National Agriculture and Food Research Organization, Tokyo, Japan.
Vet Med Sci. 2025 Sep;11(5):e70491. doi: 10.1002/vms3.70491.
Bovine rhinitis A virus (BRAV) is a virus of the genus Aphthovirus, family Picornaviridae. BRAV has recently attracted interest as a causative agent in bovine respiratory disease complex (BRDC). However, only a few PCR systems, designed based on the limited genome information, are available to detect BRAV. Therefore, we aimed to construct a new reverse transcription-PCR (RT-PCR) system for the broad detection of BRAV and compared its sensitivity and specificity with two existing real-time RT-PCR systems. Initially, we designed a new primer pair targeting the highly conserved 3D region. The detection limit of RT-PCR using the primers against strain H-1 was 10 TCID/mL, and the system was able to detect gene amounts equivalent to those detectable by existing systems. Moreover, the new system could detect synthetic DNA referring to the emerging BRAV3 strain NSWL8. The new RT-PCR system did not show cross-reactivity against non-BRAV BRDC-related viruses or taxonomically related viruses. To assess the validity of the new RT-PCR system for analysing clinical samples, nasal swabs, conjunctival swabs, and lung homogenates were collected from dairy and beef cattle in the field from 2019 to 2023 in Japan. Among the 125 clinical samples, the new RT-PCR detected BRAV from 15 samples, a higher number than two existing real-time RT-PCR systems, which identified only two and six positives. In conclusion, this RT-PCR system was demonstrated to detect currently circulating BRAVs in a highly sensitive and broad manner.
牛A型鼻炎病毒（BRAV）是小RNA病毒科口疮病毒属的一种病毒。最近，BRAV作为牛呼吸道疾病综合征（BRDC）的病原体引起了人们的关注。然而，基于有限的基因组信息设计的PCR系统只有少数几种可用于检测BRAV。因此，我们旨在构建一种新的逆转录PCR（RT-PCR）系统，用于广泛检测BRAV，并将其敏感性和特异性与现有的两种实时RT-PCR系统进行比较。最初，我们针对高度保守的3D区域设计了一对新引物。使用针对H-1株的引物进行RT-PCR的检测限为10 TCID/mL，该系统能够检测到与现有系统可检测到的基因量相当的基因量。此外，新系统能够检测参考新出现的BRAV3株NSWL8的合成DNA。新的RT-PCR系统对非BRAV的BRDC相关病毒或分类学相关病毒未表现出交叉反应性。为了评估新的RT-PCR系统分析临床样本的有效性，2019年至2023年期间从日本的奶牛和肉牛身上采集了鼻拭子、结膜拭子和肺匀浆。在125份临床样本中，新的RT-PCR从15份样本中检测到了BRAV，检测到的阳性样本数量高于现有的两种实时RT-PCR系统，后者仅鉴定出2份和6份阳性样本。总之，该RT-PCR系统被证明能够以高度敏感和广泛的方式检测目前正在传播的BRAV。
