- Volume 102, Issue 2, 2021

Crimean–Congo haemorrhagic fever virus (CCHFV) is a tick-borne virus causing Crimean–Congo haemorrhagic fever (CCHF), a disease reported to have a high fatality rate in numerous countries. The virus is geographically widespread due to its vector, and numerous wild and domestic animals can develop asymptomatic infection. Serological and limited molecular evidence of CCHFV has previously been reported in Camelus dromedarius (the dromedary, or one-humped camel) in the United Arab Emirates (UAE). In this study, 238 camel samples were screened for CCHFV RNA where 16 camel samples were positive for CCHFV by RT-PCR. Analysis of full-length CCHFV genome sequences revealed a novel lineage in camels from the UAE, and potential reassortment of the M segment of the genome.

Bunyamwera (BUNV), Batai (BATV) and Ngari (NRIV) are mosquito-borne viruses that are members of the genus Orthobunyavirus in the order Bunyavirales. These three viruses are enveloped with single-stranded, negative-sense RNA genomes consiting of three segments, denoted as Small (S), Medium (M) and Large (L). Ngari is thought to be the natural reassortant progeny of Bunyamwera and Batai viruses. The relationship between these ‘parental’ viruses and the ‘progeny’ poses an interesting question, especially given that there is overlap in their respective transmission ecologies, but differences in their infection host ranges and pathogenesis. We compared the in vivo kinetics of these three viruses in a common laboratory system and found no significant difference in growth kinetics. There was, however, a tendency of BATV to have smaller plaques than either BUNV or NRIV. Furthermore, we determined that all three viruses are stable in extracellular conditions and retain infectivity for a week in non-cellular media, which has public health and biosafety implications. The study of this understudied group of viruses addresses a need for basic characterization of viruses that have not yet reached epidemic transmission intensity, but that have the potential due to their infectivity to both human and animal hosts. These results lay the groundwork for future studies of these neglected viruses of potential public and One Health importance.

The zoonotic emerging Rift Valley fever virus (RVFV) causes sporadic disease in livestock and humans throughout Africa and the Saudi Arabian peninsula. Infection of people with RVFV can occur through mosquito bite or mucosal exposure during butchering or milking of infected livestock. Disease typically presents as a self-limiting fever; however, in rare cases, hepatitis, encephalitis and ocular disease may occur. Recent studies have illuminated the neuropathogenic mechanisms of RVFV in a rat aerosol infection model. Neurological disease in rats is characterized by breakdown of the blood–brain barrier late in infection, infiltration of leukocytes to the central nervous system (CNS) and massive viral replication in the brain. However, the route of RVFV entry into the CNS after inhalational exposure remains unknown. Here, we visualized the entire nasal olfactory route from snout to brain after RVFV infection using RNA in situ hybridization and immunofluorescence microscopy. We found widespread RVFV-infected cells within the olfactory epithelium, across the cribriform plate, and in the glomerular region of the olfactory bulb within 2 days of infection. These results indicate that the olfactory tract is a major route of infection of the brain after inhalational exposure. A better understanding of potential neuroinvasion pathways can support the design of more effective therapeutic regiments for the treatment of neurological disease caused by RVFV.

Host IFNL4 haplotype status contributes to the development of chronic hepatitis C virus (HCV) infection in individuals who are acutely infected with the virus. In silico studies revealed that specific amino acid variants at multiple sites on the HCV polyprotein correlate with functional single-nucleotide polymorphisms (SNPs) in the IFNL4 locus. Thus, SNPs at the IFNL4 locus may select variants that influence virus replication and thereby the outcome of infection. Here, we examine the most significantly IFNL4-associated amino acid variants that lie in the ‘lambda (L) 2 loop’ of the HCV NS5B RNA polymerase. L2 loop variants were introduced into both sub-genomic replicon and full-length infectious clones of HCV and viral replication was examined in the presence and absence of exogenous IFNλ4. Our data demonstrate that while mutation of the NS5B L2 loop affects replication, individual IFNL4-associated variants have modest but consistent effects on replication in both the presence and absence of IFNλ4. Given the strong genetic association between these variants and IFNL4, these data suggest a nuanced effect of each individual position on viral replication, the combined effect of which might mediate resistance to the effects of IFNλ4.

Astroviruses are non-enveloped, positive-sense, ssRNA viruses and often associated with gastrointestinal diseases. Murine astrovirus (MuAstV) was first confirmed in a laboratory mouse colony in 2011. Although infected mice do not present significant clinical symptoms, the virus might interfere with research results. A recent surveillance has shown that MuAstV is highly prevalent in laboratory mice. The aims of the present study were to identify and characterize MuAstV strains as well as to investigate the prevalence rate of viral RNA in laboratory mice in Taiwan, and to estimate the origin and past population demography of MuAstVs. Based on molecular surveillance, MuAstV RNA was detected in 45.7 % of laboratory mice (48/105) from seven of nine colonies. Three fully sequenced MuAstV strains, MuAstV TW1, TW2 and TW3, exhibited 89.1−94.4 % and 89.1–90.0 % nucleotide identities with the reference strains MuAstV STL1 and STL2, respectively. Phylogenetic analyses of the partial regions of the RNA-dependent RNA polymerase (RdRp) and capsid protein (CP) genes of 18 Taiwan strains along with other astroviruses revealed that there are three distinct lineages of mouse astrovirus, MuAstV1, MuAstV2 and mouse astrovirus JF755422. The mutation rates of MuAstV1 were 2.6×10−4 and 6.2×10−4 substitutions/site/year for the RdRp and CP regions, respectively. Based on the above molecular clock, the colonization of MuAstV1 in laboratory mice was between 1897 and 1912, in good agreement with the establishment of ‘modern’ laboratory mouse facilities. Since its initial infection, the population size of MuAstV1 has increased 15–60-fold, probably consistent with the increased use of laboratory mice. In conclusion, MuAstV1 has been associated with modern laboratory mice since the beginning, and its influence on research results may require further investigation.

Marek’s disease virus (MDV) is a highly cell-associated oncogenic alphaherpesvirus that causes T cell lymphoma in chickens. MDV-encoded Meq and vIL8 proteins play important roles in transformation and early cytolytic infection, respectively. Previous studies identified a spliced transcript, meq-vIL8, formed by alternative splicing of meq and vIL8 genes in MDV lymphoblastoid tumour cells. To determine the role of Meq-vIL8 in MDV pathogenesis, we generated a recombinant MDV (MDV-meqΔSD) by mutating the splice donor site in the meq gene to abrogate the expression of Meq-vIL8. As expected, our results show that MDV-meqΔSD virus grows similarly to the parental and revertant viruses in cell culture, suggesting that Meq-vIL8 is dispensable for MDV growth in vitro. We further characterized the pathogenic properties of MDV-meqΔSD virus in chickens. Our results show that lack of Meq-vIL8 did not affect virus replication during the early cytolytic phase, as determined by immunohistochemistry analysis and/or viral genome copy number, but significantly enhanced viral DNA load in the late phase of infection in the spleen and brain of infected chickens. In addition, we observed that abrogation of Meq-vIL8 expression reduced the mean death time and increased the prevalence of persistent neurological disease, common features of highly virulent strains of MDV, in inoculated chickens. In conclusion, our study shows that Meq-vIL8 is an important virulence factor of MDV.

In Japan, tulip-growing areas have been plagued by viral diseases for decades, but the viruses causing the damage remain undescribed. In this study, Nicotiana benthamiana and Chenopodium quinoa plants mechanically inoculated with crude sap from a symptomatic tulip flower exhibited necrosis symptoms. Additionally, flexuous and filamentous virus particles were detected by electron microscopy analysis. Moreover, we determined the complete sequences of two genomic segments of the tulip streak virus (TuSV), which is a new virus associated with streaking symptoms, on the basis of a next-generation sequencing analysis. Homology analyses of the amino acid sequence of RNA-dependent RNA polymerase and the terminal sequence of the genomic RNA indicated that TuSV is associated with viruses in the family Phenuiviridae, but differs substantially from other reported viruses.

Tobamoviruses are often referred to as the most notorious viral pathogens of pepper crops. These viruses are not transmitted by invertebrate vectors, but rather by physical contact and seeds. In this study, pepper plants displaying mild mottle and mosaic symptoms were sampled in four different regions of Peru. Upon double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) tests, seven samples cross-reacted weakly with antibodies against pepper mild mottle virus (PMMoV), suggesting the presence of tobamoviruses. When employing RT-PCR, conserved primers amplified cDNA fragments of viruses from two putative new tobamovirus species in the samples. The complete genome of two representative isolates were, therefore, sequenced and analysed in silico. These viruses, which were tentatively named yellow pepper mild mottle virus (YPMMoV) and chilli pepper mild mottle virus (CPMMoV), shared highest nucleotide genome sequence identities of 83 and 85 % with bell pepper mottle virus (BpeMV), respectively. Mechanical inoculation of indicator plants with YPMMoV and CPMMoV isolates did not show any obvious differences in host ranges. These viruses were also inoculated mechanically on pepper plants harbouring different resistance L alleles to determine their pathotypes. Pepper plants carrying unfunctional L alleles (L 0) to tobamoviruses were infected by all isolates and presented differential symptomatology for YPMMoV and CPMMoV. On the other hand, pepper plants carrying L 1, L 2, L 3 and L 4 alleles were resistant to all isolates, indicating that these viruses belong to pathotype P0.

Beet soil-borne virus (BSBV) is a sugar beet pomovirus frequently associated with Beet necrotic yellow veins virus, the causal agent of the rhizomania disease. BSBV has been detected in most of the major beet-growing regions worldwide, yet its impact on this crop remains unclear. With the aim to understand the life cycle of this virus and clarify its putative pathogenicity, agroinfectious clones have been engineered for each segment of its tripartite genome. The biological properties of these clones were then studied on different plant species. Local infection was obtained on agroinfiltrated leaves of Beta macrocarpa. On leaves of Nicotiana benthamiana, similar results were obtained, but only when heterologous viral suppressors of RNA silencing were co-expressed or in a transgenic line down regulated for both dicer-like protein 2 and 4. On sugar beet, local infection following agroinoculation was obtained on cotyledons, but not on other tested plant parts. Nevertheless, leaf symptoms were observed on this host via sap inoculation. Likewise, roots were efficiently mechanically infected, highlighting low frequency of root necrosis and constriction, and enabling the demonstration of transmission by the vector Polymyxa betae. Altogether, the entire viral cycle was reproduced, validating the constructed agroclones as efficient inoculation tools, paving the way for further studies on BSBV and its related pathosystem.