Khalafalla Abdelmalik Ibrahim, Ishag Hassan Zackaria Ali, Albalushi Hamdoon Ismail Abdulla, Al-Hammadi Zulaikha Mohamed Abdel-Hameed, Al Yammahi Saeed Mohamed Saeed, Shah Asma Abdi Mohamed, Al Muhairi Salama Suhail Mohammed
Biosecurity Affairs Division, Development and Innovation Sector, Abu Dhabi Agriculture and Food Safety Authority (ADAFSA), Abu Dhabi, United Arab Emirates.
Front Vet Sci. 2023 Aug 31;10:1182165. doi: 10.3389/fvets.2023.1182165. eCollection 2023.
The study of coronaviruses has grown significantly in recent years.Middle East respiratory syndrome coronavirus (MERS-CoV) replicates in various cell types, and quick development has been made of assays for its growth and quantification. However, only a few viral isolates are now available for investigation with full characterization. The current study aimed to isolate MERS-CoV from nasal swabs of dromedary camels and molecularly analyze the virus in order to detect strain-specific mutations and ascertain lineage classification.
We isolated the virus in Vero cells and adapted it for cultivation. The isolates were subjected to complete genome sequencing using next-generation sequencing followed by phylogenetic, mutation, and recombination analysis of the sequences.
A total of five viral isolates were obtained in Vero cells and adapted to cultures. Phylogenetic analysis classified all the isolates within clade B3. Four isolates clustered close to the MERS-CoV isolate camel/KFU-HKU-I/2017 (GenBank ID: MN758606.1) with nucleotide identity 99.90-99.91%. The later isolate clustered close to the MERS-CoV isolate Al-Hasa-SA2407/2016 (GenBank ID: MN654975.1) with a sequence identity of 99.86%. Furthermore, the isolates contained several amino acids substitutions in ORF1a (32), ORF1ab (25), S (2), ORF3 (4), ORF4b (4), M (3), ORF8b (1), and the N protein (1). The analysis further identified a recombination event in one of the reported sequences (OQ423284/MERS-CoV/dromedary/UAE-Al Ain/13/2016).
Data presented in this study indicated the need for continuous identification and characterization of MERS-CoV to monitor virus circulation in the region, which is necessary to develop effective control measures. The mutations described in this investigation might not accurately represent the virus's natural evolution as artificial mutations may develop during cell culture passage. The isolated MERS-CoV strains would be helpful in new live attenuated vaccine development and efficacy studies.
近年来，对冠状病毒的研究有了显著增长。中东呼吸综合征冠状病毒（MERS-CoV）可在多种细胞类型中复制，并且在其生长和定量检测方面已取得快速进展。然而，目前仅有少数病毒分离株可用于全面特征分析的研究。本研究旨在从单峰骆驼的鼻拭子中分离MERS-CoV，并对该病毒进行分子分析，以检测菌株特异性突变并确定谱系分类。
我们在Vero细胞中分离病毒并使其适应培养。使用下一代测序对分离株进行全基因组测序，随后对序列进行系统发育、突变和重组分析。
在Vero细胞中总共获得了五个病毒分离株并使其适应培养。系统发育分析将所有分离株归类于B3进化枝。四个分离株聚集在与MERS-CoV分离株骆驼/KFU-HKU-I/2017（GenBank编号：MN758606.1）接近的位置，核苷酸同一性为99.90 - 99.91%。后一个分离株聚集在与MERS-CoV分离株Al-Hasa-SA2407/2016（GenBank编号：MN654975.1）接近的位置，序列同一性为99.86%。此外，这些分离株在开放阅读框1a（32个）、开放阅读框1ab（25个）、刺突蛋白（S，2个）、开放阅读框3（4个）、开放阅读框4b（4个）、膜蛋白（M，3个）、开放阅读框8b（1个）和核衣壳蛋白（N，1个）中包含多个氨基酸替换。分析还在其中一个报告序列（OQ423284/MERS-CoV/单峰骆驼/阿联酋 - 艾因/13/2016）中鉴定出一个重组事件。
本研究提供的数据表明，需要持续鉴定和表征MERS-CoV，以监测该地区的病毒传播，这对于制定有效的控制措施是必要的。本研究中描述的突变可能无法准确代表病毒的自然进化，因为在细胞培养传代过程中可能会产生人工突变。分离出的MERS-CoV菌株将有助于新型减毒活疫苗的开发和疗效研究。
