Maximum Containment Laboratory, ICMR-National Institute of Virology, Pune, Maharashtra, India.
Maximum Containment Laboratory, ICMR-National Institute of Virology, Pune, Maharashtra; ICMR-National Institute of Virology Kerala Unit, Alappuzha, Kerala, India.
Indian J Med Res. 2020;151(2 & 3):226-235. doi: 10.4103/ijmr.IJMR_795_20.
BACKGROUND & OBJECTIVES: Bats are considered to be the natural reservoir for many viruses, of which some are potential human pathogens. In India, an association of Pteropus medius bats with the Nipah virus was reported in the past. It is suspected that the recently emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) also has its association with bats. To assess the presence of CoVs in bats, we performed identification and characterization of bat CoV (BtCoV) in P. medius and Rousettus species from representative States in India, collected during 2018 and 2019.
Representative rectal swab (RS) and throat swab specimens of Pteropus and Rousettus spp. bats were screened for CoVs using a pan-CoV reverse transcription-polymerase chain reaction (RT-PCR) targeting the RNA-dependent RNA polymerase (RdRp) gene. A single-step RT-PCR was performed on the RNA extracted from the bat specimens. Next-generation sequencing (NGS) was performed on a few representative bat specimens that were tested positive. Phylogenetic analysis was carried out on the partial sequences of RdRp gene sequences retrieved from both the bat species and complete viral genomes recovered from Rousettus spp.
Bat samples from the seven States were screened, and the RS specimens of eight Rousettus spp. and 21 Pteropus spp. were found positive for CoV RdRp gene. Among these, by Sanger sequencing, partial RdRp sequences could be retrieved from three Rousettus and eight Pteropus bat specimens. Phylogenetic analysis of the partial RdRp region demonstrated distinct subclustering of the BtCoV sequences retrieved from these Rousettus and Pteropus spp. bats. NGS led to the recovery of four sequences covering approximately 94.3 per cent of the whole genome of the BtCoVs from Rousettus bats. Three BtCoV sequences had 93.69 per cent identity to CoV BtRt-BetaCoV/GX2018. The fourth BtCoV sequence was 96.8 per cent identical to BtCoV HKU9-1.
INTERPRETATION & CONCLUSIONS: This study was a step towards understanding the CoV circulation in Indian bats. Detection of potentially pathogenic CoVs in Indian bats stresses the need for enhanced screening for novel viruses in them. One Health approach with collaborative activities by the animal health and human health sectors in these surveillance activities shall be of use to public health. This would help in the development of diagnostic assays for novel viruses with outbreak potential and be useful in disease interventions. Proactive surveillance remains crucial for identifying the emerging novel viruses with epidemic potential and measures for risk mitigation.
蝙蝠被认为是许多病毒的天然宿主,其中一些可能是人类病原体。在印度,过去曾有报道称,中澳大拉西亚狐蝠与尼帕病毒有关。据怀疑,最近出现的严重急性呼吸系统综合征冠状病毒 2(SARS-CoV-2)也与蝙蝠有关。为了评估蝙蝠中冠状病毒的存在,我们对 2018 年和 2019 年在印度代表性邦采集的中澳大拉西亚狐蝠和果蝠属蝙蝠进行了蝙蝠冠状病毒(BtCoV)的鉴定和特征描述。
使用针对 RNA 依赖性 RNA 聚合酶(RdRp)基因的泛冠状病毒逆转录-聚合酶链反应(RT-PCR)对代表性的中澳大拉西亚狐蝠和果蝠属蝙蝠的直肠拭子(RS)和咽拭子标本进行 CoV 筛查。从蝙蝠标本中提取的 RNA 进行一步 RT-PCR。对检测呈阳性的少数代表性蝙蝠标本进行下一代测序(NGS)。从这两个蝙蝠种的 RdRp 基因序列部分序列和从果蝠属种回收的完整病毒基因组中进行系统发育分析。
对来自七个邦的蝙蝠样本进行了筛查,发现 8 只果蝠属和 21 只中澳大拉西亚狐蝠属的 RS 样本 CoV RdRp 基因呈阳性。其中,通过 Sanger 测序,可以从 3 只果蝠和 8 只中澳大拉西亚狐蝠属蝙蝠标本中获得部分 RdRp 序列。对从这些果蝠和中澳大拉西亚狐蝠属蝙蝠中分离出的 BtCoV 序列的部分 RdRp 区进行系统发育分析,表明 BtCoV 序列明显分为亚群。NGS 导致从 Rousettus 蝙蝠中回收了大约 94.3%的整个 BtCoV 基因组的四个序列。三种 BtCoV 序列与 CoV BtRt-BetaCoV/GX2018 的同源性为 93.69%。第四个 BtCoV 序列与 BtCoV HKU9-1 的同源性为 96.8%。
本研究是了解印度蝙蝠中冠状病毒循环的一步。在印度蝙蝠中检测到潜在的致病性冠状病毒,强调需要加强对它们的新型病毒进行筛查。动物健康和人类健康部门在这些监测活动中的合作,采用“同一健康”方法,将有助于公共卫生。这将有助于开发具有潜在爆发性的新型病毒的诊断检测方法,并有助于疾病干预。主动监测对于识别具有流行潜力的新兴新型病毒和缓解风险的措施仍然至关重要。