Virus Isolation Research Articles

Stabilized trimeric peptide immunogens of the complete HIV-1 gp41 N-heptad repeat and their use as HIV-1 vaccine candidates

Efforts to develop an HIV-1 vaccine include those focusing on conserved structural elements as the target of broadly neutralizing monoclonal antibodies. MAb D5 binds to a highly conserved hydrophobic pocket on the gp41 N-heptad repeat (NHR) coiled coil and neutralizes through prevention of viral fusion and entry. Assessment of 17-mer and 36-mer NHR peptides presenting the D5 epitope in rodent immunogenicity studies showed that the longer peptide elicited higher titers of neutralizing antibodies, suggesting that neutralizing epitopes outside of the D5 pocket may exist. Although the magnitude and breadth of neutralization elicited by NHR-targeting antigens are lower than that observed for antibodies directed to other epitopes on the envelope glycoprotein complex, it has been shown that NHR-directed antibodies are potentiated in TZM-bl cells containing the FcγRI receptor. Herein, we report the design and evaluation of covalently stabilized trimeric 51-mer peptides encompassing the complete gp41 NHR. We demonstrate that these peptide trimers function as effective antiviral entry inhibitors and retain the ability to present the D5 epitope. We further demonstrate in rodent and nonhuman primate immunization studies that our 51-mer constructs elicit a broader repertoire of neutralizing antibody and improved cross-clade neutralization of primary HIV-1 isolates relative to 17-mer and 36-mer NHR peptides in A3R5 and FcγR1-enhanced TZM-bl assays. These results demonstrate that sensitive neutralization assays can be used for structural enhancement of moderately potent neutralizing epitopes. Finally, we present expanded trimeric peptide designs which include unique low-molecular-weight scaffolds that provide versatility in our immunogen presentation strategy.

First Report of Watermelon Crinkle Leaf-Associated Virus-2 (WCLaV-2) in Watermelon (Citrullus lanatus) in Georgia, USA.

Watermelon (Citrullus lanatus) is a key horticultural crop in Georgia with farmgate value of 142 million USD (2022 Farm Gate Value Report), yet faces challenges from whitefly-transmitted viruses, especially during periods of elevated whitefly populations in the fall. Foliar symptoms on watermelon plants including yellow mottling and chlorosis, wrinkling, bunching, and upward curling, were observed in experimental fields at UGA Tifton and commercial fields in Colquitt County in the fall of 2023. These were similar to those described for watermelon crinkle leaf-associated viruses (WCLaV-1 and WCLaV-2) from Florida (Hendrick et al, 2021) and Texas (Hernandez et al., 2021). The disease incidence reached 100% in both locations. WCLaV-1 was previously identified in Georgia (Adeleke et al., 2022a); however, WCLaV-2 remained undetected in further surveys (Adeleke et al., 2022b). Total nucleic acid was extracted from symptomatic leaf tissues with the MagMAXTM 96 Viral RNA isolation kit (ThermoFisher Scientific, USA), following the manufacturer's guidelines, with the omission of DNAse treatment. The presence of WCLaV-1 was identified through reverse transcription-polymerase chain reaction (RT-PCR) (Hernandez et al., 2021) in 17 out of 24 samples in Tift County, and 12 out of 15 samples from Colquitt County. Within the same set of samples, WCLaV-2 was identified in three samples from Tift County and four samples from Colquitt County by RT-PCR directed at the RdRp gene (Hernandez et al., 2021). WCLaV-1 was not detected in the three samples from Tift County that had WCLaV-2, while mixed infections of WCLaV-2 and WCLaV-1 were observed in the samples form Colquitt County. Two whitefly transmitted viruses, previously reported in Georgia were also identified as mixed infection in these samples (Table S1). The presence of WCLaV-2 was further confirmed by amplifying the movement protein (MP) gene of WCLaV-2 by RT-PCR assays (Hernandez et al., 2021). The amplicons, with expected sizes of 968bp for the RdRp gene and 562bp for the MP gene of WCLaV-2, located on RNA 1 and RNA 2 segments respectively, were directly sequenced from both directions (Genewiz, USA) from a sample collected in Tift County. The resulting data were analyzed via BLASTn search. The MP gene fragment (PP178543) shared 100% identity with isolates from Brazil (LC636074.1), Texas (MW559086.1), and Florida (MZ325858.1). RdRp gene (PP178542) shared >99.7% identity with isolates from Brazil (LC636073.1), Texas (MW559083.1) and Florida (MZ325855.1). WCLaV-1 and WCLaV-2, initially discovered in Asia (Xin et al., 2017), have been assigned to the genus Coguvirus, in the family Phenuiviridae (Walker et al., 2022). Subsequent reports from the USA (Hendrick et al., 2021; Hernandez et al., 2021), Australia (Mulholland et al., 2023), and Brazil (Maeda et al., 2022) indicate the global spread of these viruses. Watermelon is the primary host of WCLaV-2. Despite these findings, biological information, including vector relations, for both viruses and other members of the genus Coguvirus remains elusive. The impact of these viruses on watermelon production and yield in the identified regions remains largely unknown, underscoring the need for further investigations.

Molecular Characterization of Isolates of the Banana Bunchy Top Virus (BBTV) from the District of Chókwè, Mozambique

Banana bunchy top virus (BBTV) was recently detected in Mozambique and appears to be limited to the provinces of Gaza, Maputo and Zambezia, but it has great potential to spread to other provinces. Despite its importance, nothing is known about the BBTV isolates that occur in Mozambique. In this study, the sequences of the S and R genes of forty isolates chosen as representatives of samples collected previously from eleven farms of the four administrative posts of the district of Chóckwè, province of Gaza, were sequenced and analyzed. The S-DNA nucleotide sequences of the analyzed isolates were highly conserved, with identity ranging from 97% to 100%. The same was observed for the R-DNA sequences, with most identities ranging between 98% and 100% among the isolates from Chókwè and above 90% when compared to the isolates from GenBank. The phylogenetic analysis showed that the Mozambican BBTV isolates belong to the Pacific–Indian Oceans (PIO) group, showing greater proximity to the isolate JQ820453 from Malawi than to the isolates from sub-Saharan countries, which were grouped in a distinct subclade. This is the first study conducted to determine the molecular characteristics of BBTV isolates present in Mozambique.

Equine herpesvirus type 1 (EHV-1) replication at the upper respiratory entry site is inhibited by neutralizing EHV-1-specific IgG1 and IgG4/7 mucosal antibodies.

Equine herpesvirus type 1 (EHV-1) is a contagious respiratory pathogen that infects the mucosa of the upper respiratory tract (URT). Mucosal immune responses at the URT provide the first line of defense against EHV-1 and are crucial for orchestrating immunity. To define host-pathogen interactions, we characterized B-cell responses, antibody isotype functions, and EHV-1 replication of susceptible (non-immune) and clinically protected (immune) horses after experimental EHV-1 infection. Nasal secretion and nasal wash samples were collected and used for the isolation of DNA, RNA, and mucosal antibodies. Shedding of infectious virus, EHV-1 copy numbers, viral RNA expression, and host B-cell activation in the URT were compared based on host immune status. Mucosal EHV-1-specific antibody responses were associated with EHV-1 shedding and viral RNA transcription. Finally, mucosal immunoglobulin G (IgG) and IgA isotypes were purified and tested for neutralizing capabilities. IgG1 and IgG4/7 neutralized EHV-1, while IgG3/5, IgG6, and IgA did not. Immune horses secreted high amounts of mucosal EHV-1-specific IgG4/7 antibodies and quickly upregulated B-cell pathway genes, while EHV-1 was undetected by virus isolation and PCR. RNA transcription analysis reinforced incomplete viral replication in immune horses. In contrast, complete viral replication with high viral copy numbers and shedding of infectious viruses was characteristic for non-immune horses, together with low or absent EHV-1-specific neutralizing antibodies during viral replication. These data confirm that pre-existing mucosal IgG1 and IgG4/7 and rapid B-cell activation upon EHV-1 infection are essential for virus neutralization, regulation of viral replication, and mucosal immunity against EHV-1.IMPORTANCEEquine herpesvirus type 1 (EHV-1) causes respiratory disease, abortion storms, and neurologic outbreaks known as equine herpes myeloencephalopathy (EHM). EHV-1 is transmitted with respiratory secretions by nose-to-nose contact or via fomites. The virus initially infects the epithelium of the upper respiratory tract (URT). Host-pathogen interactions and mucosal immunity at the viral entry site provide the first line of defense against the EHV-1. Robust mucosal immunity can be essential in protecting against EHV-1 and to reduce EHM outbreaks. It has previously been shown that immune horses do not establish cell-associated viremia, the prerequisite for EHM. Here, we demonstrate how mucosal antibodies can prevent the replication of EHV-1 at the epithelium of the URT and, thereby, the progression of the virus to the peripheral blood. The findings improve the mechanistic understanding of mucosal immunity against EHV-1 and can support the development of enhanced diagnostic tools, vaccines against EHM, and the management of EHV-1 outbreaks.

Generation and application of immortalized sheep fetal fibroblast cell line

BackgroundPrimary sheep fetal fibroblasts (SFFCs) have emerged as a valuable resource for investigating the molecular and pathogenic mechanisms of orf viruses (ORFV). However, their utilization is considerably restricted due to the exorbitant expenses associated with their isolation and culture, their abbreviated lifespan, and the laborious procedure.ResultsIn our investigation, the primary SFFCs were obtained and immortalized by introducing a lentiviral recombinant plasmid containing the large T antigen from simian virus 40 (SV40). The expression of fibronectin and vimentin proteins, activity of SV40 large T antigen, cell proliferation assays, and analysis of programmed cell death revealed that the immortalized large T antigen SFFCs (TSFFCs) maintained the same physiological characteristics and biological functions as the primary SFFCs. Moreover, TSFFCs demonstrated robust resistance to apoptosis, extended lifespan, and enhanced proliferative activity compared to primary SFFCs. Notably, the primary SFFCs did not undergo in vitro transformation or exhibit any indications of malignancy in nude mice. Furthermore, the immortalized TSFFCs displayed live ORFV vaccine susceptibility.ConclusionsImmortalized TSFFCs present valuable in vitro models for exploring the characteristics of ORFV using various techniques. This indicates their potential for secure utilization in future studies involving virus isolation, vaccine development, and drug screening.

Genome comparison of long-circulating field CnmeGV isolates from the same region

Cnaphalocrocis medinalis granulovirus (CnmeGV), belonging to Betabaculovirus cnamedinalis, can infect the rice pest, the rice leaf roller. In 1979, a CnmeGV isolate, CnmeGV-EP, was collected from Enping County, China. In 2014, we collected another CnmeGV isolate, CnmeGV-EPDH3, at the same location and obtained the complete virus genome sequence using Illumina and ONT sequencing technologies. By combining these two virus isolates, we updated the genome annotation of CnmeGV and conducted an in-depth analysis of its genome features. CnmeGV genome contains abundant tandem repeat sequences, and the repeating units in the homologous regions (hrs) exhibit overlapping and nested patterns. The genetic variations within EPDH3 population show the high stability of CnmeGV genome, and tandem repeats are the only region of high genetic variation in CnmeGV genome replication. Some defective viral genomes formed by recombination were found within the population. Comparison analysis of the two virus isolates collected from Enping showed that the proteins encoded by the CnmeGV-specific genes were less conserved relative to the baculovirus core genes. At the genomic level, there are a large number of SNPs and InDels between the two virus isolates, especially in and around the bro genes and hrs. Additionally, we discovered that CnmeGV acquired a segment of non-ORF sequence from its host, which does not provide any new proteins but rather serves as redundant genetic material integrated into the viral genome. Furthermore, we observed that the host's transposon piggyBac has inserted into some virus genes. Together, dsDNA viruses could acquire non-coding genetic material from their hosts to expand the size of their genomes. These findings provide new insights into the evolution of dsDNA viruses.

H6N2 reassortant avian influenza virus isolate in wild birds in Jiangxi Province, China.

H6 avian influenza virus is widely prevalent in wild birds and poultry and has caused human infection in 2013 in Taiwan, China. During our active influenza surveillance program in wild waterfowl at Poyang Lake, Jiangxi Province, an H6N2 AIV was isolated and named A/bean goose/JiangXi/452-4/2013(H6N2). The isolate was characterized as a typical low pathogenic avian influenza virus (LPAIV) due to the presence of the amino acid sequence PQIETR↓GLFGAI at the cleavage site of the hemagglutinin (HA) protein. The genetic evolution analysis revealed that the NA gene of the isolate originated from North America and exhibited the highest nucleotide identity (99.29%) with a virus recovered from wild bird samples in North America, specifically A/bufflehead/California/4935/2012(H11N2). Additionally, while the HA and PB1 genes belonged to the Eurasian lineage, they displayed frequent genetic interactions with the North American lineage. The remaining genes showed close genetic relationships with Eurasian viruses. The H6N2 isolate possessed a complex genome, indicating it is a multi-gene recombinant virus with genetic material from both Eurasian and North American lineages.

Molecular evidence of cherry leaf roll virus isolate from the phylogenetic group D1 infecting black elderberry (Sambucus nigra) in Ukraine

Molecular evidence of cherry leaf roll virus isolate from the phylogenetic group D1 infecting black elderberry (Sambucus nigra) in Ukraine

Genome sequence of a vesicular stomatitis Indiana virus isolate collected in 1988 from a naturally infected bovine in Mexico.

We report the near-full genome sequence of a vesicular stomatitis Indiana virus (VSIV) originally collected from a naturally infected bovine in south-central Mexico. This sequence represents a coding-complete genome sequence of a VSIV from Mexico, a country where vesicular stomatitis is endemic.

Uncertainties Surrounding Madariaga Virus, a Member of the Eastern Equine Encephalitis Virus Complex.

Background: Madariaga virus (MADV), a member of the eastern equine encephalitis virus (EEEV) complex, circulates in Latin America and exhibits distinct evolutionary and ecological features compared to the North American EEEV. While published data have shed light on MADV ecology, several key aspects remain unknown. Methods: In this study, we compiled data on virus isolation, vector competence, and animal serology collected over six decades in Latin America to identify critical knowledge gaps on MADV transmission and ecology. Results: Specific vertebrate animals serving as amplifying hosts and the mosquito species acting as enzootic and epizootic vectors have not yet been identified. Other aspects that remain unclear are the virus current geographic distribution, the role of equines as hosts in epizootic cycles, and the full impact of MADV on human health in endemic regions. Conclusions: The numerous knowledge gaps surrounding MADV, its widespread distribution in Latin America, and its potential to cause severe disease in animals and humans emphasize the urgent need for increased research efforts, heightened awareness, and intensified surveillance towards this potential emerging threat.

Seasonal antigenic prediction of influenza A H3N2 using machine learning

Antigenic characterization of circulating influenza A virus (IAV) isolates is routinely assessed by using the hemagglutination inhibition (HI) assays for surveillance purposes. It is also used to determine the need for annual influenza vaccine updates as well as for pandemic preparedness. Performing antigenic characterization of IAV on a global scale is confronted with high costs, animal availability, and other practical challenges. Here we present a machine learning model that accurately predicts (normalized) outputs of HI assays involving circulating human IAV H3N2 viruses, using their hemagglutinin subunit 1 (HA1) sequences and associated metadata. Each season, the model learns an updated nonlinear mapping of genetic to antigenic changes using data from past seasons only. The model accurately distinguishes antigenic variants from non-variants and adaptively characterizes seasonal dynamics of HA1 sites having the strongest influence on antigenic change. Antigenic predictions produced by the model can aid influenza surveillance, public health management, and vaccine strain selection activities.

Increased Diversity of Citrus Tristeza Virus in Europe.

This study investigated the genetic diversity of Citrus tristeza virus (CTV) isolates from Montenegro and Croatia, European countries with the northernmost citrus-growing regions situated on the Eastern Adriatic coast. Fifteen complete or nearly complete CTV genomes were reconstructed by high-throughput sequencing of samples collected in distinct municipalities in Montenegro and Opuzen municipality in Croatia. Phylogenetic analyses assigned some of the sequences to VT and T30 strains, previously recorded in Europe, while remarkably other isolates were placed in S1 and RB groups, which have not been reported in Europe so far. In addition, a new phylogenetic lineage comprising only isolates from Montenegro was delineated and tentatively proposed as the MNE cluster. Recombination analysis revealed evidence of 11 recombination events in the sequences obtained in this study, between isolates of related strains, within isolates of the same strain, and between distant strains. These findings show that CTV diversity in Europe is higher than reported before and calls for the reevaluation of management strategies.[Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Molecular characterization of Japanese isolates of soil-borne wheat mosaic virus in the genus Furovirus

Molecular characterization of Japanese isolates of soil-borne wheat mosaic virus in the genus Furovirus

Prevalence and genetic diversity of the <i>Alongshan virus (Flaviviridae)</i> circulating in ticks in the south of Eastern Siberia

Introduction. Tick-borne infections are of great importance for many regions of Russia, including Eastern Siberia. This unfavorable epidemiological situation can be characterized not only by the circulation of well-known tick-borne infections, but also by the identification of new pathogens, the role of which remains little or generally unexplored. Multicomponent flavi-like viruses can cause infectious diseases in humans and pose a threat to public health. The purpose of the study was the identification and molecular genetic characterization of the Alongshan virus (Flaviviridae, ALSV) isolates, transmitted by ticks in the south of Eastern Siberia. Materials and methods. Total 1060 ticks were collected and analyzed from the territory of the Republics of Khakassia, Tuva, Buryatia, Irkutsk Region and Transbaikal Territory (Zabaykalsky Krai) in the spring-summer period 2023. ALSV RNA was detected by RT-PCR followed by nucleotide sequence determination and phylogenetic analysis for each segment of the genome. Results. The ALSV infection rate in Ixodes persulcatus ticks collected in the Republic of Khakassia was 3.3% (95% CI: 1.4–7.5); in Irkutsk Oblast – 1.0% (95% CI: 0.3–3.7); in the Republic of Tuva – 0.9% (95% CI: 0.3–3.4) and in Transbaikal Krai – 0.7% (95% CI: 0.2–3.6). Sequences of all four segments of ALSV genetic variants circulating in I. persulcatus ticks in the south of Eastern Siberia are grouped with sequences found in China and clustered into the Asian subgroup transmitted by taiga ticks. The level of difference in the nucleotide sequences of genome fragments among the identified genetic variants of ALSV ranged from 2 to 3%. Conclusion. The article shows the widespread distribution of ALSV in I. persulcatus ticks in the Republics of Khakassia and Tyva, Irkutsk Oblast and Transbaikal Territory. The obtained data actualize monitoring of changes in the area of distribution of potentially dangerous for humans flavi-like viruses and their vectors.

Biology, phylogenetic and evolutionary relations of Tradescantia mild mosaic virus isolates from Hungary

Biology, phylogenetic and evolutionary relations of Tradescantia mild mosaic virus isolates from Hungary

High parasite diversity maintained after an alga-virus coevolutionary arms race.

Arms race dynamics are a common outcome of host-parasite coevolution. While they can theoretically be maintained indefinitely, realistic arms races are expected to be finite. Once an arms race has ended, for example due to the evolution of a generalist resistant host, the system may transition into coevolutionary dynamics that favor long-term diversity. In microbial experiments, host-parasite arms races often transition into a stable coexistence of generalist resistant hosts, (semi-)susceptible hosts, and parasites. While long-term host diversity is implicit in these cases, parasite diversity is usually overlooked. In this study, we examined parasite diversity after the end of an experimental arms race between a unicellular alga (Chlorella variabilis) and its lytic virus (PBCV-1). First, we isolated virus genotypes from multiple time points from two replicate microcosms. A time-shift experiment confirmed that the virus isolates had escalating host ranges, i.e. that the arms races had occurred. We then examined the phenotypic and genetic diversity of virus isolates from the post-arms race phase. Post-arms race virus isolates had diverse host ranges, survival probabilities, and growth rates; they also clustered into distinct genetic groups. Importantly, host range diversity was maintained throughout the post-arms race phase, and the frequency of host range phenotypes fluctuated over time. We hypothesize that this dynamic polymorphism was maintained by a combination of fluctuating selection and demographic stochasticity. Together with previous work in prokaryotic systems, our results link experimental observations of arms races to natural observations of long-term host and parasite diversity.

Cytochrome c oxidase IV isoform 1 (COX4-1) regulates the proliferation, migration and invasion of trophoblast cells via modulating mitochondrial function

IntroductionSpontaneous miscarriage is a common complication of early pregnancy. Previous studies have shown that mitochondrial function plays an important role in establishment of a successful pregnancy. Cytochrome c oxidase subunit 4 isoform 1 (COX4I1), a component of electron transport chain complex Ⅳ, is required for coupling the rate of ATP production to energetic requirements. However, there is very limited research on its role in trophoblast biology and how its dysfunction may contribute to spontaneous miscarriage. MethodsPlacental villi (7–10 weeks gestational age) collected from either induced termination of pregnancy or after spontaneous miscarriage were examined for expression of COX4I1. COX4I1 was knocked down by siRNA transfection of primary isolates of EVT cells. Real-time cell analysis (RTCA) and 5-Ethynyl-2′-deoxyuridine (EdU) were used to detect changes in proliferation ability after COX4I1 knockdown of EVT cells. Migration and invasion indices were determined by RTCA. Mitochondrial morphology was observed via MitoTracker staining. Oxidative phosphorylation, ATP production, and glycolysis in COX4I1-deficient cells and controls were assessed by a cellular energy metabolism analyzer (Seahorse). ResultsIn placental villous tissue, COX4I1 expression was significantly decreased in the spontaneous miscarriage group. Knockdown of COX4I1 inhibited EVT cell proliferation, increased the migration and invasion ability and mitochondrial fusion of EVT cells. Mitochondrial respiration and glycolysis were impaired in COX4I1-deficient EVT cells. Knockdown of MMP1 could rescue the increased migration and invasion induced by COX4I1 silencing. DiscussionLow expression of COX4I1 leads to mitochondrial dysfunction in EVT, resulting in altered trophoblast function, and ultimately to pregnancy loss.

Continuous year-round isolation of giant viruses from brackish shoreline soils.

Giant viruses, categorized under Nucleocytoviricota, are believed to exist ubiquitously in natural environments. However, comprehensive reports on isolated giant viruses remain scarce, with limited information available on unrecoverable strains, viral proliferation sites, and natural hosts. Previously, the author highlighted Pandoravirus hades, Pandoravirus persephone, and Mimivirus sp. styx, isolated from brackish water soil, as potential hotspots for giant virus multiplication. This study presents findings from nearly a year of monthly sampling within the same brackish water region after isolating the three aforementioned strains. This report details the recurrent isolation of a wide range of giant viruses. Each month, four soil samples were randomly collected from an approximately 5 × 10 m plot, comprising three soil samples and one water sample containing sediment from the riverbed. Acanthamoeba castellanii was used as a host for virus isolation. These efforts consistently yielded at least one viral species per month, culminating in a total of 55 giant virus isolates. The most frequently isolated species was Mimiviridae (24 isolates), followed by Marseilleviridae (23 isolates), Pandoravirus (6 isolates), and singular isolates of Pithovirus and Cedratvirus. Notably, viruses were not consistently isolated from any of the four samples every month, with certain sites yielding no viruses. Cluster analysis based on isolate numbers revealed that soil samples from May and water and sediment samples from January produced the highest number of viral strains. These findings underscore brackish coastal soil as a significant site for isolating numerous giant viruses, highlighting the non-uniform distribution along coastlines.

Dual structure-switching aptamer-mediated signal amplification cascade for SARS-CoV-2 detection

Since the outbreak of the novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) at the end of 2019, the spread of the virus has posed a significant threat to public health and the global economy. This work proposed a one-step, dual-structure-switching aptamer-mediated signal amplification cascade for rapid and sensitive detection of the SARS-CoV-2 nucleocapsid protein. This system consisted of two DNA aptamers with structure-switching functionality and fuel DNA, where a cascade of strand hybridization and displacement triggered fluorescence generation and signal amplification. This aptamer-based amplification cascade required neither an amplification stage using enzymes nor pre-processing steps such as washing, viral isolation, and gene extraction. The assay could distinguish SARS-CoV-2 from other respiratory viruses and detect up to 1.0 PFU/assay of SARS-CoV-2 within 30 min at room temperature. In 35 nasopharyngeal clinical samples, the assay accurately assessed 25 positive and 10 negative clinical swab samples, which were confirmed using quantitative polymerase chain reaction. The strategy reported herein can help detect newly emerging pathogens and biomarkers of various diseases in liquid samples. In addition, the developed detection system consisting of only DNA and fluorophores can be widely integrated into liquid biopsy platforms for disease diagnosis.

Conyza canadensis, a new host of Tobacco leaf curl Yunnan virus in China

AbstractIn April 2023, leaf curl, crinkling symptoms were found on Conyza canadensis plants around tobacco fields located at Yuxi, Yunan province, China. PCR detection with genus‐specific primers showed that symptomatic C. canadensis plants was infected by begomovirus. The full‐length viral components and satellite molecules from the diseased C. canadensis (YN‐2023) were obtained by enrichment of circular DNA with rolling circle amplification, restriction enzyme digestion, cloning, and DNA sequencing. The viral component (2, 750 nt, OR509733) shared the highest nt identity at 92.4% to tobacco leaf curl Yunnan virus isolate YN5741 (TbLCYNV‐YN5741 [CN:Yn:Age:16], MN218671). A satellite molecule (1, 339 nt, OR509734) shared the highest nt identity at 86.5% to tobacco curly shoot betasatellite isolate Y115 (TbCSB‐Y115 [CN:Yn:03], AJ457822). Another satellite molecule (1, 357 nt, OR509735) shared the highest nt identity at 94.2% to ageratum yellow vein Singapore alphasatellite (AYVSGA‐[Singapore:1998], NC003414). Based on the threshold for classification of Begomovirus, Betasatellite, Alphasatellite, YN‐2023 was a new strain of TbLCYNV and associated with TbCSB and AYVSGA. To our best knowledge, this is the first report that C. canadensis is a natural host of TbLCYNV.