PMC:7151644 / 23660-31009 JSONTXT

{"project":"2_test","denotations":[{"id":"32296543-25914383-144433908","span":{"begin":188,"end":192},"obj":"25914383"},{"id":"32296543-27510566-144433909","span":{"begin":206,"end":210},"obj":"27510566"},{"id":"32296543-28934931-144433910","span":{"begin":679,"end":683},"obj":"28934931"},{"id":"32296543-22090114-144433911","span":{"begin":996,"end":1000},"obj":"22090114"},{"id":"32296543-24237667-144433912","span":{"begin":1038,"end":1042},"obj":"24237667"},{"id":"32296543-27876899-144433913","span":{"begin":1468,"end":1472},"obj":"27876899"},{"id":"32296543-31168752-144433914","span":{"begin":1487,"end":1491},"obj":"31168752"},{"id":"32296543-30896364-144433915","span":{"begin":1708,"end":1712},"obj":"30896364"},{"id":"32296543-16839713-144433916","span":{"begin":2338,"end":2342},"obj":"16839713"},{"id":"32296543-28786787-144433917","span":{"begin":3095,"end":3099},"obj":"28786787"},{"id":"32296543-20101443-144433918","span":{"begin":4145,"end":4149},"obj":"20101443"},{"id":"32296543-29618816-144433920","span":{"begin":4321,"end":4325},"obj":"29618816"},{"id":"32296543-204240-144433921","span":{"begin":4977,"end":4981},"obj":"204240"},{"id":"32296543-16318717-144433922","span":{"begin":4997,"end":5001},"obj":"16318717"},{"id":"32296543-27158509-144433923","span":{"begin":5106,"end":5110},"obj":"27158509"},{"id":"32296543-27158509-144433924","span":{"begin":5326,"end":5330},"obj":"27158509"},{"id":"32296543-28923111-144433925","span":{"begin":5345,"end":5349},"obj":"28923111"},{"id":"32296543-30939106-144433926","span":{"begin":5369,"end":5373},"obj":"30939106"},{"id":"32296543-28964878-144433927","span":{"begin":5999,"end":6003},"obj":"28964878"},{"id":"32296543-28923111-144433928","span":{"begin":6130,"end":6134},"obj":"28923111"},{"id":"32296543-28923111-144433929","span":{"begin":6349,"end":6353},"obj":"28923111"},{"id":"32296543-24753611-144433930","span":{"begin":6573,"end":6577},"obj":"24753611"},{"id":"32296543-32015508-144433931","span":{"begin":7133,"end":7137},"obj":"32015508"},{"id":"32296543-32015507-144433932","span":{"begin":7151,"end":7155},"obj":"32015507"}],"text":"4. Discussion\nPangolins are insectivorous, predominantly nocturnal, and predate almost exclusively on ants and termites, with a strong preference for particular insect species (Lin et al. 2015; Choo et al. 2016). Eight species of pangolins are present in Africa and Asia, with habitat loss and changes in their living environment seriously affecting their population status. Most importantly, pangolins have been greatly impacted by the illegal international and domestic wildlife trade for traditional medicine or meat (Zhang et al. 2015). Hence, pangolins are listed as vulnerable, endangered or critically endangered on the IUCN Red List of Threatened Species (du Toit et al. 2017). Notably, pangolins may also be threatened by epizootic pathogens, either present in their original habitats or in their new environments following translocation. Indeed, the translocation or trafficking of domestic and wild animals plays an important role in the spread of many epizootic pathogens (Lin et al. 2012; Kosmider et al. 2013; Peeler et al. 2015).\nIn this study, four pangolins—three M. javanica likely from Indonesia, Malaysia, and Thailand, and one M. pentadactyla probably of local origin—were found to be suffering disease and were sent to a local rescue station for treatment. Unfortunately, three animals died, whereas one recovered. To date, reports on pangolin disease have been rare and mainly limited to those caused by bacteria and parasites (Mohapatra et al. 2016; Jabin et al. 2019). There is no available literature on viral infections of pangolins until the recent identification of Parainfluenza Virus 5, sendai virus, and coronavirus from Sunda pangolins in China (Liu et al. 2019; Wang et al. 2019). Using a combination of meta-transcriptomic and PCR methods, we identified two novel RNA viruses—a pestivirus and a coltivirus—in two dead pangolins. Based on their clinical signs (Table 1), all these four pangolins appeared to suffer infectious disease. However, as no complete clinical data and laboratory parameters were obtained from these animals, we could not clearly define the disease they suffered.\nPestiviruses are well-known animal pathogens that cause significant economic loss, infecting both domestic (e.g. pigs, cattle, sheep, and goats) and wild (e.g. wild boars and ruminants) animals (Vilcek and Nettleton 2006). Pestivirus infections may be subclinical or cause a range of clinical signs including acute diarrhea, acute hemorrhagic syndrome, acute fatal disease, as well as a wasting disease. Herein, a novel pestivirus, designated DYPV, was identified in multiple organs in one of the sick pangolins [1-Dongyang (M. javanica)] that had clear pathological changes. Phylogenetic analysis indicates that it represents a novel member of the genus Pestivirus. Although more detailed confirmatory results are required, these data suggest that DYPV-like pestiviruses may be responsible for the hemorrhagic disease observed in the pangolins.\nTo date, it is commonly believed that vertebrates, rather than invertebrates, are the main hosts of pestiviruses (Smith et al. 2017). Recently, however, a novel pestivirus, named Fairfax Lookout virus, was identified in ticks (Ixodes trichosuri). However, given its phylogenetic position next to mammalian pestiviruses, as well as its the extremely low abundance, it was proposed that the virus was associated with the vertebrate host rather than from the tick itself (Harvey et al. 2018). In this study, DYPV was not only identified in the pangolins, but also in nymph ticks collected from the pangolins. However, DYPV was not found in the adult ticks also sampled from pangolin 1-Dongyang. Interestingly, although DYPV from ticks was closely related to that sampled from pangolins, there were ∼2 per cent nt differences across the viral genome suggesting that they are separated by multiple transmission events. Phylogenetic analysis indicated that Sunda pangolins, including the pangolin 1-Dongyang, were most likely imported into China from Indonesia, Malaysia, and the Philippines (Fig. 6). In addition, A. javanense ticks have not found in Zhejiang province (Chen et al. 2010), DYPV was not identified in locally collected ticks, and we did not observe this virus in previous large scale tick sampling studies (Li et al. 2015; Shi et al. 2016a,b, 2018). Together, these observations suggest that the virus was probably imported from abroad with the illegal trafficked pangolin.\nFigure 6. ML tree based on nt sequences of the mt-cyt b gene of four pangolins and other known pangolins. The numbers at nodes indicate bootstrap support values after 1,000 replications. Bootstrap values higher than 70 per cent were considered significant and shown in the trees. Coltiviruses are well-known tick-borne pathogens that can cause human disease. For example, CTFV causes mild febrile illness or more severe disease including infection of the central nervous system, and/or hemorrhagic fever (Goodpasture et al. 1978; Attoui et al. 2005). Additionally, EYAV infections have been associated with human neurological disease (Moutailler et al. 2016). Notably, although some coltiviruses (e.g. CTFV, EYAV, TFRV) have been detected in wildlife (such as bats and rodents), there is no clear evidence that these viruses can cause disease in animals (Moutailler et al. 2016; Weiss et al. 2017; Williamson et al. 2019). In this study, a novel coltivirus, named LSPV, was identified in one of the sick pangolins [2-Lishui (M. javanica)]. Given the high abundance of LSPV in the meta-transcriptomic data, combined with the clinical features and pathologic changes appeared in the pangolin (2-Lishui), as well as the detection of LSPV in several organs, our data suggest that LSPV may have caused systemic infection and the death of the pangolin in question, although this will need to be confirmed with additional data.\nIt is generally recognized that the coltivirus genome comprises twelve segments of linear double-stranded RNA (Fujita et al. 2017). Recently, a novel coltivirus, Forest reovirus (TFRV), was identified in free-tailed bats (C. aloysiisabaudiae; Weiss et al. 2017). Interestingly, the virus genome lacks segments 6, 7, and 12, and it was proposed that these missing segments were not identified probably due to low similarities between TFRV and known coltiviruses (Weiss et al. 2017). Strikingly, as with TFRV, we were unable to identify viral segments 6, 7, and 12 in LSPV, despite the use of methods that previously obtained the complete genome of the highly divergent Jingmen tick virus (Qin et al. 2014). It is therefore clear that additional studies are needed to infer whether these segments are indeed absent from LSPV and TFRV. Finally, as pangolin 2-Lishui may have been imported from Indonesia (Fig. 6) it is possible that LSPV, like DYPV, was also imported into China.\nFinally, despite some discussion concerning the role of pangolins in the emergence of the novel coronavirus (severe acute respiratory syndrome coronavirus 2, SARS-CoV-2;Wahba et al. 2020; Wong et al. 2020), the cause of the corona virus disease 2019 ( COVID-19) outbreak (Wu et al. 2020; Zhou et al. 2020), we have not found coronaviruses in these pangolins by meta-transcriptomic and PCR methods. Therefore, more efforts are needed to infer the role of pangolins in the transmission of SARS-CoV-2."}