Detection and characterization of a novel bat-borne coronavirus in Singapore using multiple molecular approaches

Lim, Xiao Fang; Lee, Chengfa Benjamin; Pascoe, Sarah Marie; How, Choon Beng; Chan, Sharon; Tan, Jun; Yang, Xing; Zhou, Peng; Z, Shi; Sessions, October M.; Wang, Lin‐Fa; Ng, Lee Ching; Anderson, Danielle E.; Yap, Grace

doi:10.1099/jgv.0.001307

Cited by 33 publications

(37 citation statements)

References 43 publications

(50 reference statements)

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“…This virus was only detected by unbiased NGS. Similar observations have been made in previous studies, where full-length genome sequencing of human herpesvirus 1, West Nile virus, and MERS-CoV was achieved only when high viral titers were present (15,18). We could improve the capture efficiency in two ways in the future: by using newly designed probes that bind better to their targets or by changing the steps that affect binding.…”

Section: Discussionsupporting

confidence: 80%

“…Most data generated from unbiased NGS can be attributed to non-CoV-specific reads. To assess whether samples can be enriched to allow sequencing of only CoV-specific reads, we utilized NGS in conjunction with viral nucleic acid capture specifically targeting most of the known CoVs using a pool of 4,303 unique baits (18). These baits were designed from 90 representative CoV genomes, and in silico analysis determined that these baits should target all known CoV species tested here ( [MHV]) were amplified in cell culture, and RNA was extracted from the supernatants.…”

Section: Resultsmentioning

confidence: 99%

“…Minimal reads were obtained for the genomes of BtTyCoV-HKU4r and BtHpCoV-HKU10r, and both samples had small amounts of viral RNA (C T value of Ͼ30). The technical procedure of the capture is itself a limitation of enrichment NGS (15,18). Key steps in the hybridization protocol, such as washes, could result in the loss of viral nucleic acid.…”

Section: Discussionmentioning

confidence: 99%

“…Virus enrichment NGS has been successfully used for various viral families, and the most common protocols rely on predesigned viral probes that share more than 60% homology with the target virus sequence (15)(16)(17). In this study, we utilized an enrichment NGS approach with predesigned probes targeting most of the CoV species (18). Our aim was to strategically perform bat CoV surveillance in which high-throughput sample processing for virus discovery would be balanced with cost effectiveness.…”

mentioning

confidence: 99%

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Discovery of Bat Coronaviruses through Surveillance and Probe Capture-Based Next-Generation Sequencing

Hao

Zhu

et al. 2020

mSphere

Self Cite

104

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Coronaviruses (CoVs) of bat origin have caused two pandemics in this century. Severe acute respiratory syndrome (SARS)-CoV and Middle East respiratory syndrome (MERS)-CoV both originated from bats, and it is highly likely that bat coronaviruses will cause future outbreaks. Active surveillance is both urgent and essential to predict and mitigate the emergence of these viruses in humans. Next-generation sequencing (NGS) is currently the preferred methodology for virus discovery to ensure unbiased sequencing of bat CoVs, considering their high genetic diversity. However, unbiased NGS is an expensive methodology and is prone to missing low-abundance CoV sequences due to the high background level of nonviral sequences present in surveillance field samples. Here, we employ a capture-based NGS approach using baits targeting most of the CoV species. Using this technology, we effectively reduced sequencing costs by increasing the sensitivity of detection. We discovered nine full genomes of bat CoVs in this study and revealed great genetic diversity for eight of them. IMPORTANCE Active surveillance is both urgent and essential to predict and mitigate the emergence of bat-origin CoV in humans and livestock. However, great genetic diversity increases the chance of homologous recombination among CoVs. Performing targeted PCR, a common practice for many surveillance studies, would not reflect this diversity. NGS, on the other hand, is an expensive methodology and is prone to missing low-abundance CoV sequences. Here, we employ a capture-based NGS approach using baits targeting all CoVs. Our work demonstrates that targeted, cost-effective, large-scale, genome-level surveillance of bat CoVs is now highly feasible. 2020. Discovery of bat coronaviruses through surveillance and probe capture-based next-generation sequencing. mSphere 5:e00807-19. https://doi .org/10.1128/mSphere.2020. Correction for Li et al., "Discovery of bat coronaviruses through surveillance and probe capture-based next-generation sequencing." mSphere 5: e00170-20. https://doi.

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Section: Discussionsupporting

confidence: 80%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Discovery of Bat Coronaviruses through Surveillance and Probe Capture-Based Next-Generation Sequencing

Hao

Zhu

et al. 2020

mSphere

Self Cite

104

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“…Coronaviruses belong to the family of Coronaviridae . Bats have been recognized as natural reservoir and vectors of a variety of coronaviruses and the viruses have crossed species barriers to infect humans and many other different kinds of animals, including avians, rodents, and chiropters 50,51 . Coronavirus may cause respiratory and neurological diseases 52 .…”

Section: Coronavirus and Its Diagnostic Approachesmentioning

confidence: 99%

Recent advances in the detection of respiratory virus infection in humans

Zhang

Wang

Deng

et al. 2020

Journal of Medical Virology

429

363

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Respiratory tract viral infection caused by viruses or bacteria isone of the most common diseases in human worldwide, while those caused by emerging viruses, such as the novel coronavirus, 2019-nCoV that caused the pneumonia outbreak in Wuhan, China most recently, have posed great threats to global public health. Identification of the causative viral pathogens of respiratory tract viral infections is important to select an appropriate treatment, save people's lives, stop the epidemics, and avoid unnecessary use of antibiotics. Conventional diagnostic tests, such as the assays for rapid detection of antiviral antibodies or viral antigens, are widely used in many clinical laboratories. With the development of modern technologies, new diagnostic strategies, including multiplex nucleic acid amplification and microarray-based assays, are emerging. This review summarizes currently available and novel emerging diagnostic methods for the detection of common respiratory viruses, such as influenza virus, human respiratory syncytial virus, coronavirus, human adenovirus, and human rhinovirus. Multiplex assays for simultaneous detection of multiple respiratory viruses are also described. It is anticipated that such data will assist researchers and clinicians to develop appropriate diagnostic strategies for timely and effective detection of respiratory virus infections. K E Y W O R D S adenovirus, coronavirus, diagnostic methods, influenza virus, respiratory syncytial virus, respiratory viral infection, rhinovirus

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Optimizing noninvasive sampling of a zoonotic bat virus

Giles

Peel

Wells

et al. 2021

Ecology and Evolution

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Outbreaks of infectious viruses resulting from spillover events from bats have brought much attention to bat‐borne zoonoses, which has motivated increased ecological and epidemiological studies on bat populations. Field sampling methods often collect pooled samples of bat excreta from plastic sheets placed under‐roosts. However, positive bias is introduced because multiple individuals may contribute to pooled samples, making studies of viral dynamics difficult. Here, we explore the general issue of bias in spatial sample pooling using Hendra virus in Australian bats as a case study. We assessed the accuracy of different under‐roost sampling designs using generalized additive models and field data from individually captured bats and pooled urine samples. We then used theoretical simulation models of bat density and under‐roost sampling to understand the mechanistic drivers of bias. The most commonly used sampling design estimated viral prevalence 3.2 times higher than individual‐level data, with positive bias 5–7 times higher than other designs due to spatial autocorrelation among sampling sheets and clustering of bats in roosts. Simulation results indicate using a stratified random design to collect 30–40 pooled urine samples from 80 to 100 sheets, each with an area of 0.75–1 m2, and would allow estimation of true prevalence with minimum sampling bias and false negatives. These results show that widely used under‐roost sampling techniques are highly sensitive to viral presence, but lack specificity, providing limited information regarding viral dynamics. Improved estimation of true prevalence can be attained with minor changes to existing designs such as reducing sheet size, increasing sheet number, and spreading sheets out within the roost area. Our findings provide insight into how spatial sample pooling is vulnerable to bias for a wide range of systems in disease ecology, where optimal sampling design is influenced by pathogen prevalence, host population density, and patterns of aggregation.

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Detection and characterization of a novel bat-borne coronavirus in Singapore using multiple molecular approaches

Cited by 33 publications

References 43 publications

Discovery of Bat Coronaviruses through Surveillance and Probe Capture-Based Next-Generation Sequencing

Discovery of Bat Coronaviruses through Surveillance and Probe Capture-Based Next-Generation Sequencing

Recent advances in the detection of respiratory virus infection in humans

Optimizing noninvasive sampling of a zoonotic bat virus

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