China Virus Research Articles

Genomic characteristics and recombination patterns of swine hepatitis E virus in China.

Zoonotic hepatitis E, mainly caused by swine hepatitis E virus (sHEV), is endemic in China, causing great economic disruption and public health threats. Although recombination is critical for the evolution of viruses, there is a limited assessment of its occurrence among sHEVs. Herein, we analysed all available sHEV full-length genomes isolated in China during the past two decades (40 isolates) compared to 72 other sHEV strains isolated in different countries and determined that sHEV genotype 4 (sHEV4) dominates China. Eight potential natural recombination events were identified, four of which occurred in China and were mainly between sHEV4 strains, indicating the distinct character of China sHEV. One intergenotype recombination event was found in China, alarming the emergence of a new sHEV lineage that could become a critical threat to human health.

COVID-19 is a “Yellow Peril” Redux: Immigration and Health Policy and the Construction of the Chinese as Disease

During the COVID-19 pandemic, Asian Americans have endured a stark rise in discrimination, harassment, and violence. Public discourse regarding COVID-19 has also been filled with anti-Asian and xenophobic rhetoric, including former President Donald Trump’s usage of racially charged epithets like “China Virus” and “Kung Flu.” However, this is not the first time that Asian Americans, and specifically Chinese Americans, have been condemned as a public health threat. In the late 1800s, Chinese immigrants were stereotyped as the “Yellow Peril” and dirty disease carriers amidst growing anti-Chinese sentiment, culminating in the 1882 Chinese Exclusion Act. Additionally, San Francisco Chinatown was intentionally and unfairly targeted by public health officials in attempts to purge the bubonic plague at the turn of the century. While court cases like Wong Wai v. Williamson (1900) and Jew Ho v. Williamson (1900) determined that such public health campaigns violated the Fourteenth Amendment’s Equal Protection Clause by singling out Chinese individuals, their legal outcomes do not represent the overall social consensus, both historically and today. Thus, our current moment of surging anti-Asian rhetoric and racism must be contextualized within America’s long history of branding Asian and Chinese people as unwanted, filthy vectors of contagion to be excluded. This paper argues that the uptick in anti-Asian racial violence during the COVID-19 pandemic is a direct extension of xenophobic scapegoating, racial formation, and sociocultural representation of Chinese immigrants as harbingers of disease that rationalized the 1882 Chinese Exclusion Act and San Francisco’s racist public health measures in 1900. Ultimately, I argue that America's historical intersection of exclusionary immigration law and discriminatory health policy set the stage for COVID-19 to play out as a “Yellow Peril” redux.

The Genetic Characterization of a Novel Natural Recombinant Pseudorabies Virus in China.

We sequenced the complete genome of the pseudorabies virus (PRV) FJ epidemic strain, and we studied the characteristics and the differences compared with the classical Chinese strain and that of other countries. Third-generation sequencing and second-generation sequencing technology were used to construct, sequence, and annotate an efficient, accurate PRV library. The complete FJ genome was 143,703 bp, the G+C content was 73.67%, and it encoded a total of 70 genes. The genetic evolution of the complete genome and some key gene sequences of the FJ strain and PRV reference strains were analyzed by the maximum likelihood (ML) method of MEGA 7.0 software. According to the ML tree based on the full-length genome sequences, PRV FJ strain was assigned to the branch of genotype II, and it showed a close evolutionary relationship with PRV epidemic variants isolated in China after 2011. The gB, gC, gD, gH, gL, gM, gN, TK, gI, and PK genes of the FJ strain were assigned to the same branch with other Chinese epidemic mutants; its gG gene was assigned to the same branch with the classic Chinese Fa and Ea strains; and its gE gene was assigned to a relatively independent branch. Potential recombination events were predicted by the RDP4 software, which showed that the predicted recombination sites were between 1694 and 1936 bp, 101,113 and 102,660 bp, and 107,964 and 111,481 bp in the non-coding region. This result broke the previously reported general rule that pseudorabies virus recombination events occur in the gene coding region. The major backbone strain of the recombination event was HLJ8 and the minor backbone strain was Ea. Our results allowed us to track and to grasp the recent molecular epidemiological changes of PRV. They also provide background materials for the development of new PRV vaccines, and they lay a foundation for further study of PRV.

The Overlooked Tragedy of the Pandemic: How Media Coverage of the COVID-19 Pandemic has Led to an Increase in Anti-Asian Bias and Xenophobia

The media is a vital source of information, especially in times of crisis. Since the 2019 novel coronavirus became a worldwide pandemic, it has become a frequent topic of discussion in the media, and its mysterious origin has caused lots of speculation regarding its roots. Historically, scientists have named novel diseases based on the country or region in which it was thought to have originated; however, in 2015, the World Health Organization (WHO) publicly discouraged this practice because of the stigma it tended to attach to people and places. Despite this warning and the WHO's deliberate name selection for the 2019 novel coronavirus — "COVID-19" — the media and many public figures, including former United States President Donald Trump, have consistently used stigmatizing language, such as "China virus," associating the virus with China because the first confirmed cases of the virus were discovered in Wuhan, China. Just as the WHO warned, this stigmatizing language has created a wave of violence and discrimination towards Asian Americans and caused considerable damage to U.S.-China relations. This Article explores the correlation between the presence of such stigmatizing rhetoric in the media and the damage to U.S.-China relations. It concludes by providing suggestions to combat the crisis of anti-Asian bias and xenophobia and encourages the U.S. and China to set aside their differences and work together to alleviate tensions and end the pandemic.

The Overlooked Tragedy of the Pandemic: How Media Coverage of the COVID-19 Pandemic has Led to an Increase in Anti-Asian Bias and Xenophobia

The media is a vital source of information, especially in times of crisis. Since the 2019 novel coronavirus became a worldwide pandemic, it has become a frequent topic of discussion in the media, and its mysterious origin has caused lots of speculation regarding its roots. Historically, scientists have named novel diseases based on the country or region in which it was thought to have originated; however, in 2015, the World Health Organization (WHO) publicly discouraged this practice because of the stigma it tended to attach to people and places. Despite this warning and the WHO's deliberate name selection for the 2019 novel coronavirus — "COVID-19" — the media and many public figures, including former United States President Donald Trump, have consistently used stigmatizing language, such as "China virus," associating the virus with China because the first confirmed cases of the virus were discovered in Wuhan, China. Just as the WHO warned, this stigmatizing language has created a wave of violence and discrimination towards Asian Americans and caused considerable damage to U.S.-China relations. This Article explores the correlation between the presence of such stigmatizing rhetoric in the media and the damage to U.S.-China relations. It concludes by providing suggestions to combat the crisis of anti-Asian bias and xenophobia and encourages the U.S. and China to set aside their differences and work together to alleviate tensions and end the pandemic.

Molecular characterization of a novel Conyza canadensis-infecting begomovirus in China

A novel monopartite begomovirus was found in naturally infected Conyza canadensis plants exhibiting typical begomovirus-induced yellow vein symptoms in Yunnan Province of China. Analysis of two obtained full-length viral genome sequences showed that they shared 99.8% nucleotide (nt) sequence similarity, and both consisted of 2733 nts (GenBank accession no. OK120264 and OK120266, respectively). Further analysis showed that these two sequences shared the highest nt sequence similarity (78.9%) with tomato yellow leaf curl Vietnam virus (TYLCVV) (EU189150). In the assayed C. canadensis plants, a betasatellite of 1,341 nts (Accession no. OK120265 and OK120267, respectively) was also found, and it shared 98.1% sequence similarity with malvastrum yellow vein betasatellite (MaYVB) (JX679254). Based on the current classification criteria for begomoviruses, we consider that the two obtained viral isolates are a novel begomovirus, and named it as conyza yellow vein virus (CoYVV). Our further analysis result showed that CoYVV is likely originated from a recombination event between tomato yellow leaf curl Yunnan virus (TYLCYnV) and tomato yellow leaf curl China virus (TYLCCNV). To investigate the effect of CoYVV infection in plant, we constructed two infectious clones (i.e., pCoYVV and pMaYVB), and inoculated them individually or together to Nicotiana benthamiana plants through agro-infiltration. The result showed that the plants co-inoculated with CoYVV and MaYVB developed yellow vein and downward leaf-curling symptoms, whereas the plants inoculated with CoYVV alone showed no clear virus-like symptoms. Virus infection in the inoculated N. benthamiana plants was confirmed through polymerase chain reaction (PCR). The result of quantitative PCR (qPCR) showed that in the presence of MaYVB, the accumulation level of CoYVV DNA was significantly increased compared to that in the plants infected with CoYVV alone.

Epidemiological and Genetic Characteristics of Porcine Reproductive and Respiratory Syndrome Virus in South China Between 2017 and 2021.

Porcine reproductive and respiratory syndrome virus (PRRSV) remains a major threat to the swine industry in China and has caused enormous losses every year. To monitor the epidemiological and genetic characteristics of PRRSV in South China, 6,795 clinical samples from diseased pigs were collected between 2017 and 2021, and 1,279 (18.82%) of them were positive for PRRSV by RT-PCR detecting the ORF5 gene. Phylogenetic analysis based on 479 ORF5 sequences revealed that a large proportion of them were highly-pathogenic PRRSVs (409, 85.39%) and PRRSV NADC30-like strains (66, 13.78%). Furthermore, 93.15% of these highly-pathogenic strains were found to be MLV-derived. We next recovered 11 PRRSV isolates from the positive samples and generated the whole genome sequences of them. Bioinformatic analysis showed that seven isolates were MLV-derived. Besides, six isolates were found to be recombinant strains. These eleven isolates contained different types of amino acid mutations in their GP5 and Nsp2 proteins compared to those of the PRRSVs with genome sequences publicly available in GenBank. Taken together, our findings contribute to understanding the prevalent status of PRRSV in South China and provide useful information for PRRS control especially the use of PRRSV MLV vaccines.

Phylogenetic and Spatiotemporal Analyses of the Complete Genome Sequences of Avian Coronavirus Infectious Bronchitis Virus in China During 1985-2020: Revealing Coexistence of Multiple Transmission Chains and the Origin of LX4-Type Virus.

Infectious bronchitis (IB) virus (IBV) causes considerable economic losses to poultry production. The data on transmission dynamics of IBV in China are limited. The complete genome sequences of 212 IBV isolates in China during 1985–2020 were analyzed as well as the characteristics of the phylogenetic tree, recombination events, dN/dS ratios, temporal dynamics, and phylogeographic relationships. The LX4 type (GI-19) was found to have the highest dN/dS ratios and has been the most dominant genotype since 1999, and the Taiwan-I type (GI-7) and New type (GVI-1) showed an increasing trend. A total of 59 recombinants were identified, multiple recombination events between the field and vaccine strains were found in 24 isolates, and the 4/91-type (GI-13) isolates were found to be more prone to being involved in the recombination. Bayesian phylogeographic analyses indicated that the Chinese IBVs originated from Liaoning province in the early 1900s. The LX4-type viruses were traced back to Liaoning province in the late 1950s and had multiple transmission routes in China and two major transmission routes in the world. Viral phylogeography identified three spread regions for IBVs (including LX4 type) in China: Northeastern China (Heilongjiang, Liaoning, and Jilin), north and central China (Beijing, Hebei, Shanxi, Shandong, and Jiangsu), and Southern China (Guangxi and Guangdong). Shandong has been the epidemiological center of IBVs (including LX4 type) in China. Overall, our study highlighted the reasons why the LX4-type viruses had become the dominant genotype and its origin and transmission routes, providing more targeted strategies for the prevention and control of IB in China.

Molecular Detection and Genetic Characterization of Potential Zoonotic Swine Enteric Viruses in Northern China.

Despite significant economic and public health implications, swine enteric viruses that do not manifest clinical symptoms are often overlooked, and data on their epidemiology and pathogenesis are still scarce. Here, an epidemiological study was carried out by using reverse transcription-polymerase chain reaction (RT-PCR) and sequence analysis in order to better understand the distribution and genetic diversity of porcine astrovirus (PAstV), porcine encephalomyocarditis virus (EMCV), porcine kobuvirus (PKV), and porcine sapovirus (PSaV) in healthy pigs reared under specific pathogen-free (SPF) or conventional farms. PKV was the most prevalent virus (51.1%, 247/483), followed by PAstV (35.4%, 171/483), then PSaV (18.4%, 89/483), and EMCV (8.7%, 42/483). Overall, at least one viral agent was detected in 300 out of 483 samples. Out of the 300 samples, 54.0% (162/300), 13.0% (39/300), or 1.0% (3/300) were found coinfected by two, three, or four viruses, respectively. To our knowledge, this is the first report of EMCV detection from porcine fecal samples in China. Phylogenetic analysis revealed genetically diverse strains of PAstV, PKV, and PSaV circulating in conventional and SPF farms. Detection of swine enteric viruses with a high coinfection rate in healthy pigs highlights the importance of continuous viral surveillance to minimize future economic and public health risks.

Continuous surveillance revealing a wide distribution of class I Newcastle disease viruses in China from 2011 to 2020.

The risk-based active surveillance for Newcastle disease virus (NDV) was carried out in China from 2011 to 2020. A total of 110,018 swabs were collected from 28 provinces. 2,389 class I NDVs were isolated and identified by RT-PCR and sequencing. The average annual positivity rate of class I NDVs from 2011 to 2020 was 2.17%. In the last 10 years, the positivity rate was highest in 2011 (4.76%), and has since decreased. Most viruses were isolated from chickens, while others were collected from ducks, geese and pigeons, as well as from the environment. The positivity rates for class I NDVs in poultry ranged from 0.55% to 2.40%. The viruses were isolated from 373 sampling sites in 24 provinces, mainly in East, Central, South and Southwest China. The positivity rates of NDVs in wholesale markets (51.58%) and retail markets (42.83%) were much higher than those in poultry farms (7.14%) and slaughterhouses (3.85%). Phylogenetic analyses showed that most isolates belonged to sub-genotype 1.1.2, while only 22 viruses belonged to sub-genotype 1.2, indicating the viruses in sub-genotype 1.1.2 were the predominant strains in China. The F and HN genes of six strains in the two sub-genotypes were sequenced and analyzed. The cleavage sites of F protein in the six viruses were 112ERQER/L117, 112ERQGR/L117 or 112GRQERL117, which were typical of low virulence NDV. Several mutations were identified in the functional domains of F and HN proteins, including fusion peptide, heptad repeat region, transmembrane domains and neutralizing epitopes. This study revealed the distribution, genetic and phylogenetic characteristics of class I NDVs in China, and could help us to better understand the epidemiological context of class I NDVs in China.

First Report of Natural Infection of Zucchini by Tomato Chlorosis Virus and Cucurbit Chlorotic Yellows Virus in China.

Zucchini (Cucurbita pepo) is an extensively cultivated and important economic cucurbit crop in China. In September 2018 and 2019, interveinal chlorosis and yellowing symptoms, suspected to be caused by either tomato chlorosis virus (ToCV; genus Crinivirus) or cucurbit chlorotic yellows virus (CCYV; genus Crinivirus) or by their co-infection, were observed on zucchini plants in a greenhouse in Shandong Province, China. The incidence of the disease in the greenhouse was 20-30%. To identify the causal agent(s) of the disease, leaf samples from 66 zucchini plants were collected in 14 greenhouses in the cities of Shouguang (n = 12), Dezhou (n = 36), Qingzhou (n = 12), and Zibo (n = 6) in Shandong. Four whitefly (Bemisia tabaci) samples and four symptomatic tomato samples were also collected from these sampling sites (one each for each site) because numerous whiteflies were observed in the sampling greenhouses and ToCV was previously reported in greenhouse tomato plants from these regions (Zhao et al. 2014). To determine whether the symptoms were associated with Crinivirus infection, reverse transcription polymerase chain reaction (RT-PCR) using Crinivirus-specific degenerate primers (CriniRdRp251F/CriniRdRp995R) (Wintermantel and Hladky 2010) was performed first on total RNA extracted using the TRIzol protocol (Jordon-Thaden et al. 2015). Thereafter, the RNA samples were subjected to RT-PCR with ToCV- or CCYV-specific primers (Sun et al. 2016; Gan et al. 2019). Of the 66 zucchini samples, 54 tested positive by the degenerate crinivirus primer pair; and among them, 10 tested positive for ToCV only, 40 positive for CCYV only, and 4 positive for both viruses. Interestingly, while both viruses were detected in all B. tabaci samples, only ToCV was detected in the tomato samples (n = 4). To confirm the identity of the viruses, the amplicons of ToCV (four samples each of tomato, B. tabaci and zucchini) and CCYV (four samples each of B. tabaci and zucchini) were Sanger sequenced (Tsingke Biotechnology Co., Ltd., Beijing, China) after cloning into pMD18-T vectors (Takara, Shiga, Japan). BLASTn analysis demonstrated that all sequences were identical to their respective amplicons. The ToCV sequences (GenBank accession numbers: tomato, MN944406; B. tabaci, MN944404; zucchini, MN944405) shared 100% sequence identity with isolates from Beijing (KT751008, KC887999, KR184675, and KP335046), Hebei (KP217196), and Shandong (KX900412). The CCYV sequence (GenBank accession number MT396249) shared 99.9% sequence identity with isolates China (JN126046, JQ904629, KP896506, KX118632, KY400633, and MK568545), Greece (LT716000, LT716001, LT716002, LT716005, and LT716006), and Cyprus (LT992909, LT992910, and LT992911). To assess the transmissibility of ToCV and CCYV, virus-free B. tabaci (n = 30) were placed in ToCV or CCYV-infected zucchini plants for one day for virus acquisition. Thereafter, the whiteflies were transferred into virus-free zucchini seedlings (cv. 'Zaoqingyidai', 4-leaf-stage, n = 6 for each of the control, ToCV and CCYV treatment) for one day. Three weeks after inoculation, all plants that were inoculated with either ToCV or CCYV displayed same symptoms as those observed in the greenhouses, whereas plants in the control group remained symptom free. RT-PCR analysis using ToCV- and CCYV-specific primers confirmed the infection of the plants with the respective virus, whereas control plants were free from the viruses. CCYV has been previously reported on zucchini in Algeria (Kheireddine et al. 2020), Iran (LR585225), and Cyprus (LT992910). To our knowledge, this is the first report of CCYV infection in zucchini in China, and moreover the first report of ToCV infection in zucchini in the world. Clearly, stringent management is needed to minimize the losses caused by these viruses in greenhouse operations in the region.

Genetic characterization and pathogenicity of a novel recombinant PRRSV from lineage 1, 8 and 3 in China failed to infect MARC-145 cells

The diversity of porcine reproductive and respiratory syndrome virus (PRRSV) in China is increasing rapidly along with mutation and recombination. Recombination could occur between inter- and intra-lineage of PRRSV, which accelerated the complexity of pathogenicity and cell tropism of the recombinant strain. In the present study, a novel PRRSV strain named HN-YL1711 was isolated from a pig farm suffering from severe respiratory difficulty in Henan province, China. The whole genomic sequence analysis indicated that the genome of HN-YL1711 was 15018 nt. It shared 86%, 87.3%, 88.1%, 91.1%, 84.2%, and 84.1% nucleotide similarities with PRRSVs VR2332, CH1a, JXA1, NADC30, QYYZ, and GM2, respectively. Based on phylogenetic analysis of Nsp2, ORF5 and complete genomes, HN-YL1711 was classified into lineage 1 of PRRSV. However, seven genomic break points were detected in recombination analysis, which indicated that the HN-YL1711 originated from multiple recombination among NADC30-like (major parent, lineage 1), JXA1-like (minor parent, lineage 8), and QYYZ-like (minor parent, lineage 3) PRRSV. Porcine alveolar macrophages (PAMs), 3D4/21-CD163 and MARC-145 cells were used to explore the viral adaptation of HN-YL1711. The results indicated that it could infect the PAMs but failed to infect MARC-145 cells. Challenge experiments showed that HN-YL1711 exhibits intermediate virulence in pigs, compared with HP-PRRSV JXA1 and LP-PRRSV CH1a. Taken together, our findings suggest that recombination remains an important factor in PRRSV evolution and that recombination further complicates the cell tropism and pathogenicity of PRRSV.

Through Our Eyes, Hear Our Stories: A Virtual Photovoice Project to Document and Archive Asian American and Pacific Islander Community Experiences During COVID-19.

Through Our Eyes, Hear Our Voices is a virtual photovoice project that documents the impact of COVID-19 on Asian American and Pacific Islander (AAPI) communities. Quantitative studies on the disproportionate impact of COVID-19 are still emerging, but they do not reveal qualitative experiences of a racialized pandemic exacerbated by political leaders labeling it "China virus." As a qualitative participatory action research approach, photovoice is an ideal archival and pedagogical tool to capture the lived experience of AAPI communities. However, we had to adapt photovoice to a virtual research environment. We did so by adopting a variety of digital learning and information sharing platforms. In addition, we enlisted community-based organizations who are providing essential services for underrepresented communities to serve as research mentors for university student researchers. Finally, given the historic nature of the pandemic and the underrepresentation of AAPI experiences in mainstream archives, we emphasized the importance of students as co-producers of archival knowledge.

Genomic characterization of lumpy skin disease virus in southern China.

Lumpy skin disease virus (LSDV) is of high economic importance and has spread rapidly to many European and Asian countries in recent years. LSDV was introduced to China in 2019 and have caused severe outbreaks in several provinces. Here, we detected an LSDV strain (GD01/2020) from a cattle farm with typical LSD symptoms in Guangdong, southern China using a novel quantitative real-time PCR assay targeting the viral GPCR gene. We obtained the whole genomic sequence of GD01/2020 through metagenomic analysis. The GD01/2020 was highly homologous to the LSDVs isolated in Xinjiang, China in 2019, and distinct from all the LSDVs identified in other countries, in their sequences of GPCR and RPO30 genes. The GD01/2020 was a vaccine-recombinant strain, but distinct from two recombinant LSDVs identified in Russia. At least 25 putative recombination events between a vaccine strain and a field strain were identified in the genome of GD01/2020, which could affect the virulence and transmissibility of the virus. These results suggested that a virulent vaccine-recombinant LSDV from an unknown origin was introduced into Xinjiang, China in 2019 and spread to Guangdong, China in 2020.

Circulation, genomic characteristics, and evolutionary dynamics of class I Newcastle disease virus in China

ABSTRACT Newcastle disease caused by Newcastle disease virus (NDV) is one of the most serious threats to chickens and has two clinical forms, typical and atypical, caused by velogenic and lentogenic strains, respectively. To control the epidemic, many vaccines against velogenic class II NDVs have been introduced worldwide, but this has led to accelerated mutation of class II viruses under immune pressure and, on the other hand, to non-vaccine targeting class I NDVs becoming the dominant population in poultry. In this context, this study provided the first large-scale genomic epidemiological and quasispecies dynamic analysis of class I NDVs in China, and found that class I viruses that first appeared in East and South China have spread to central China and become the dominant class with an average evolutionary rate of 1.797 × 10−3. In addition, single nucleotide polymorphism and intra-host single nucleotide variation analyses show that HN and P genes have high mutation rates and may act as front-runners for NDV to expand their host range and enhance their virulence. This study also found that the class I NDV population has accumulated a number of mutations under positive selection and that six isolates with shortened C-terminal extensions of the HN protein are evolving toward increased virulence. These results not only enrich the research resources but also help us to better understand the dynamic evolution and mutational trends of NDV at the genomic level, which is crucial for monitoring, early warning, and controlling the outbreak of Newcastle disease.

Diverse positional 14C labeling-assisted metabolic analysis of pesticides in rats: The case of vanisulfane, a novel vanillin-derived pesticide

Information on pesticide metabolites is crucial for accurate environmental risk assessment. However, identifying the various metabolites of a novel pesticide is challenging since the potential metabolic pathways are unknown. In this study, we coupled diverse positional 14C labeling with high-resolution mass spectrometry to quantitatively and qualitatively study pesticide metabolism in rats. With the unique M/(M + 2) ratios derived from 14C, precursor compounds of metabolites could be better distinguished from impurity ions. Additionally, the use of diverse 14C labeling positions is a powerful tool to elucidate the complete metabolic fate of novel contaminants. Vanisulfane is a novel vanillin-derived antiviral agent with encouraging prospects for the efficient control of cucumber mosaic virus in China, but its metabolic pathways in mammals are still poorly understood. Thus, the metabolism of vanisulfane was studied in rats of both sexes by this strategy. The results showed that phase I and phase II metabolism occurred in both sexes. The former included mainly oxidation reactions, and the latter involved binding reactions that formed glucuronide, sulfate and amino acid conjugates. Sex-related differences were observed in the experiment, with earlier appearance of downstream metabolites and a preference for sulfate conjugate formation in males compared to females. This research facilitates the risk evaluation of vanisulfane, and offers an effective framework for screening unknown pesticide metabolic pathways, which could be applied to establish the metabolic profiles of other novel contaminants with limited information.

Strawberry, a New Natural Host of Brassica Yellows Virus in China.

Brassica yellows virus (BrYV; genus Polerovirus, family Solemoviridae) has an icosahedral spherical virion with a positive-sense single-stranded RNA genome and it is distinguished from turnip yellows virus (TuYV) based on differences in ORF0 and ORF5 (Xiang et al., 2011). To investigate the occurrence and distribution of viruses infecting strawberry (Fragaria ananassa) in the main production areas in China, a survey of nine greenhouses (667 m2 each) was conducted in the cities of Yantai and Beijing, China in August 2020. About 1% of strawberry plants in each greenhouse showed virus-like symptoms of chlorotic spots; 89 symptomatic leaf samples were randomly collected for virus testing. Total RNA was extracted from a pool of eight samples of four different cultivars (Hokowase: 2, Mibao: 2, Sagahonoka: 2, Monterey: 2) from Yantai using RNAprep Pure Plant Plus Kit (TianGen, China). A cDNA library was constructed by NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (NEB, USA) after ribosomal RNA-depletion using an Epicentre Ribo-Zero™ rRNA Removal Kit (Epicentre, USA). High-throughput sequencing was done on Illumina Hiseq 4000, generating 70,931,850 high-quality 150 bp paired-end reads. Clean reads were de novo assembled by Trinity (v2.2.0) and the resulting contigs were screened by BLASTn and BLASTx against GenBank database as described previously (Grabherr et al., 2013). A total of 1,432,164 high-quality reads unmapped to the strawberry genome were obtained and assembled into 93 contigs (ranging from 33 to 8,031 nt). Seven of these contigs (277 to 1,254 nt) shared 98.2 to 100% nt identities with BrYV-A (accession no. HQ388348) and covered 89.5% of the genome of BrYV-A. Subsequent analyses indicated the presence of Strawberry pallidosis-associated virus and Strawberry mottle virus in the analyzed sample, both have been reported in strawberry in China (Shi et al., 2018; Fan et al., 2021). To confirm BrYV infection, total RNA was isolated from the eight samples used for HTS and reverse transcription polymerase chain reaction (RT-PCR) was conducted with two pairs of specific primers (CP and rtp, Supplementary Table 1) designed based on the assembled contigs. PCR products with expected sizes (587 and 609 bp) were observed in one sample (cv. Mibao). BLASTn analysis indicated that the amplicons (accession no. MW548437 and MW548438) shared 98.6% and 99.3% nt identity with BrYV-A, respectively. To obtain the complete sequence of the putative BrYV isolate, the gaps were bridged and the terminal sequences were determined using 5' and 3' RACE kits (Clontech, China) based on the assembled contigs. The complete genome sequence of the putative BrYV isolate has a length of 5,666 nt (accession no. MZ666129) and shares more than 94.3% nt identities with other BrYV isolates. Phylogenetic analysis indicated that the isolate grouped closely with BrYV and further from TuYV (Figure S1). In addition, 11 samples (cv. Benihoppe) of the remaining 81 symptomatic strawberry samples tested positive for BrYV by RT-PCR with the two pairs of primers mentioned above. The sequences (accession no. MZ407232 and MZ407233) revealed 99.5% and 99.3% nt identities with MW548437 and MW548438. To the best of our knowledge, this is the first report of natural infection of BrYV in strawberry plants. Our findings expand the host range of BrYV, but disease association is difficult to establish due to presence of mixed infection and non-fulfillment of Koch's postulates.

Molecular characterization of echovirus 9 strains isolated from hand-foot-and-mouth disease in Kunming, Yunnan Province, China

Echovirus 9 (E9) belongs to the species Enterovirus B. So far, 12 whole genome sequences of E9 are available in GenBank. In this study, we determined the whole genomic sequences of five E9 strains isolated from the stools of patients with hand-foot-and-mouth disease in Kunming, Yunnan Province, China, in 2019. Their nucleotide and amino acid sequences shared 80.8–80.9% and 96.4–96.8% identity with the prototype Hill strain, respectively, and shared 99.3–99.9% and 99.1–99.8% mutual identity, respectively. Recombination analyses revealed that intertype recombination had occurred in the 2C and 3D regions of the five Yunnan E9 strains with coxsackieviruses B5 and B4, respectively. This study augmented the whole genome sequences of E9 in the GenBank database and extended the molecular characterization of this virus in China.

Whole genome analysis identifies intra-serotype recombinants and positive selection sites of dengue virus in mainland China from 2015 to 2020

Immune pressure can select for escape mutants that can become epidemiologically relevant. Thus, surveillance of recombinants and positively selected mutants of the dengue virus (DENV) are vital for preventing and controlling the dengue fever outbreak. However, little is known about recombinants and positively selected mutants of circulating DENV strains in mainland China. In this study, those variants with recombination and adaptive evolutionary sites of circulating DENV strains were identified during 2015–2020. Phylogenetic analysis showed that the DENV-2 was the dominant epidemic serotype, and the dengue epidemic in China was closely related to the imported virus from Southeast Asian countries. Recombination analysis based on 291 complete genomes of naturally circulating DENV identified 10 new intra-serotype recombinant variants. Two or three recombination regions in a single dengue isolate were also observed. The breakpoints of recombinants were distributed in different regions of the genome. In particular, two recombinant strains (strain DENV-4/China/YN/15DGR394 (2015) and XLLM10666) with extremely large exchange fragments were detected. This large-scale gene fragment exchange (eight genomic regions) of strain DENV-4/China/YN/15DGR394 (2015) with substitutions at both the 5′ and 3′ ends of the genome, had never been described before. Moreover, selection pressure analyses revealed seven positive selection sites located in regions encoding the NS1, NS3 and NS5 proteins. Overall, this study is the first to report ten specific intra-serotype recombinants and seven positive selection sites of Chinese epidemic strains of DENV, which highlight their significance for DENV surveillance and effective control.

Geminiviruses employ host DNA glycosylases to subvert DNA methylation-mediated defense

DNA methylation is an epigenetic mechanism that plays important roles in gene regulation and transposon silencing. Active DNA demethylation has evolved to counterbalance DNA methylation at many endogenous loci. Here, we report that active DNA demethylation also targets viral DNAs, tomato yellow leaf curl China virus (TYLCCNV) and its satellite tomato yellow leaf curl China betasatellite (TYLCCNB), to promote their virulence. We demonstrate that the βC1 protein, encoded by TYLCCNB, interacts with a ROS1-like DNA glycosylase in Nicotiana benthamiana and with the DEMETER (DME) DNA glycosylase in Arabidopsis thaliana. The interaction between βC1 and DME facilitates the DNA glycosylase activity to decrease viral DNA methylation and promote viral virulence. These findings reveal that active DNA demethylation can be regulated by a viral protein to subvert DNA methylation-mediated defense.