The role of farmed animals in the viral spillover from wild animals to humans is of growing importance. Between July and September of 2023 infectious disease outbreaks were reported on six Arctic fox (Vulpes lagopus) farms in Shandong and Liaoning provinces, China, which lasted for 2-3 months and resulted in tens to hundreds of fatalities per farm. Severe Fever with Thrombocytopenia Syndrome Virus (SFTSV) was identified in tissue/organ and swab samples from all the 13 foxes collected from these farms. These animals exhibited loss of appetite and weight loss, finally resulting in death. In autopsy and histopathology, prominently enlarged spleens and extensive multi-organ hemorrhage were observed, respectively, indicating severe systemic effects. Viral loads were detected in various tissues/organs, including brains from 9 of the 10 foxes. SFTSV was also detected in serum, anal swabs, as well as in environmental samples, including residual food in troughs used by dying foxes in follow-up studies at two farms. The 13 newly sequenced SFTSV genomes shared >99.43% nucleotide identity with human strains from China. Phylogenetic analyses showed that the 13 sequences belonged to three genotypes, and that two sequences from Liaoning were genomic reassortants, indicative of multiple sources and introduction events. This study provides the first evidence of SFTSV infection, multi-tissue tropism, and pathogenicity in farmed foxes, representing an expanded virus host range. However, the widespread circulation of different genotypes of SFTSV in farmed animals from different provinces and the diverse transmission routes, highlight its increasing and noticeable public health risk in China.

RNA viruses are characterized by a broad host range and high levels of genetic diversity. Despite a recent expansion in the known virosphere following metagenomic sequencing, our knowledge of the species rank genetic diversity of RNA viruses, and how often they are misassigned and misclassified, is limited. We performed a clustering analysis of 7801 RNA-directed RNA polymerase (RdRp) sequences representing 1897 established RNA virus species. From this, we identified substantial genetic divergence within some virus species and inconsistency in RNA virus assignment between the GenBank database and The International Committee on Taxonomy of Viruses (ICTV). In particular, 27.57% virus species comprised multiple virus operational taxonomic units (vOTUs), including Alphainfluenzavirus influenzae, Mammarenavirus lassaense, Apple stem pitting virus, and Rotavirus A, with each having over 100 vOTUs. In addition, the distribution of average amino acid identity between vOTUs within single assigned species showed a relatively low threshold: <90% and sometimes <50%. However, when only exemplar sequences from virus species were analyzed, 1889 of the ICTV-designated RNA virus species (99.58%) were clustered into a single vOTU. Clustering of the RdRp sequences from different virus species also revealed that 17 vOTUs contained two distinct virus species. These potential misassignments were confirmed by phylogenetic analysis. A further analysis of average nucleotide identity (ANI) values ranging from 70% to 97.5% revealed that at an ANI of 82.5%, 1559 (82.18%) of the 1897 virus species could be correctly clustered into one single vOTU. However, at ANI values >82.5%, an increasing number of species were clustered into two or more vOTUs. In sum, we have identified some inconsistency and misassignment of the RNA virus species based on the analysis of RdRp sequences alone, which has important implications for the development of an automated RNA virus classification system.

The discovery of alphacoronaviruses and betacoronaviruses in plateau pikas (Ochotona curzoniae) expanded the host range of mammalian coronavirus (CoV) to a new order - Lagomorpha. However, the diversity and evolutionary relationships of CoVs in these plateau-region-specific animal population remains uncertain. We conducted a five-year longitudinal surveillance of CoVs harboured by pikas around Qinghai Lake, China. CoVs were identified in 33 of 236 plateau pikas and 2 of 6 Gansu pikas (Ochotona cansus), with a total positivity rate of 14.5%, and exhibiting a wide spatiotemporal distribution across seven sampling sites and six time points. Through meta-transcriptomic sequencing and RT-PCR, we recovered 16 near-complete viral genome sequences. Phylogenetic analyses classified the viruses as variants of either pika alphacoronaviruses or betacoronaviruses endemic to plateau pikas from the Qinghai-Tibet Plateau region. Of particular note, the pika-associated betacoronaviruses may represent a novel subgenus within the genus Betacoronavirus. Tissue tropism, evaluated using quantitative real-time PCR, revealed the presence of CoV in the rectal and/or lung tissues, with the highest viral loads at 10(3.55) or 10(2.80) RNA copies/mu L. Surface plasmon resonance (SPR) assays indicated that the newly identified betacoronavirus did not bind to human or pika Angiotensin-converting enzyme 2 (ACE2) or Dipeptidyl peptidase 4 (DPP4). The findings highlight the ongoing circulation and broadening host spectrum of CoVs among pikas, emphasizing the necessity for further investigation to evaluate their potential public health risks.

Perinereis species are essential benthonic animals in coastal ecosystems and have significant roles as live feed in aquaculture, owing to their high-protein and low-fat nutritional profile. Despite their ecological importance, the viral communities associated with these organisms need to be better understood. In this study, we generated 2.6 × 108 reads using meta-transcriptomic sequencing and de novo assembled 5.3 × 103 virus-associated contigs. We identified 12 novel RNA viruses from two species, Perinereis aibuhitensis and P. wilsoni, which were classified into four major viral groups: Picobirnaviridae, Marnaviridae, unclassified Picornavirales, and unclassified Bunyavirales. Our findings revealed the hidden diversity of viruses and genome structures in Perinereis, enriching the RNA virosphere and expanding the host range of Picobirnaviridae, Marnaviridae, and Bunyavirales. This study also highlighted the potential biosecurity risk of the novel viruses carried by Perinereis to aquaculture.

Hepeviruses have been identified in a broad range of animal hosts, including mammals, birds, and fish. In this study, rodents (n=91) from seven different species and ten pikas (Ochotona curzoniae) were collected in Qinghai Province, China. Using transcriptomic sequencing and confirmatory molecular testing, hepeviruses were detected in 27 of 45 (60 %) long-tailed dwarf hamsters (Cricetulus longicaudatus) and were undetected in other rodents and pika. The complete genome sequences from 14 representative strains were subsequently obtained, and phylogenetic analyses suggested that they represent a novel species within the genus Rocahepevirus, which we tentatively designated as Cl-2018QH. The virus was successfully isolated in human hepatoma (Huh-7) and murine fibroblast (17 Cl-1) cell lines, though both exhibited limited replication as assayed by detection of negative-sense RNA intermediates. A129 immunodeficient mice were inoculated with Cl-2018QH and the virus was consistently detected in multiple organs, despite relatively low viral loads. In summary, this study has described a novel rodent hepevirus, which enhances our knowledge of the genetic diversity of rodent hepeviruses and highlights its potential for cross-species transmission.

Lined seahorse, Hippocampus erectus, is an important aquatic animal due to its medicinal and ornamental purposes. However, our understanding of the viral spectrum in H. erectus is still limited. Here, we studied the viruses in H. erectus using meta-transcriptomic sequencing. A total of 213,770,166 reads were generated and assembled de novo into 539 virus-associated contigs. Three novel RNA viruses from the Astroviridae, Paramyxoviridae, and Picornaviridae families were finally identified. In addition, we identified a strain of nervous necrosis virus from H. erectus. In particular, the unhealthy group showed a higher viral diversity and abundance than the normal group. These results revealed the diversity and cross-species transmission of viruses in H. erectus and highlighted the threat of viral infections to H. erectus.

Following the outbreak of coronavirus disease 2019 (COVID-19), several severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-related coronaviruses have been discovered. Previous research has identified a novel lineage of SARS-CoV-2-related CoVs in bats, including RsYN04, which recognizes human angiotensin-converting enzyme 2 (ACE2) and thus poses a potential threat to humans. Here, we screened the binding of the RsYN04 receptor-binding domain (RBD) to ACE2 orthologs from 52 animal species and found that the virus showed a narrower ACE2-binding spectrum than SARS-CoV-2. However, the presence of the T484W mutation in the RsYN04 RBD broadened its range. We also evaluated 44 SARS-CoV-2 antibodies targeting seven epitope communities in the SARS-CoV-2 RBD, together with serum obtained from COVID-19 convalescents and vaccinees, to determine their cross-reaction against RsYN04. Results showed that no antibodies, except for the RBD-6 and RBD-7 classes, bound to the RsYN04 RBD, indicating substantial immune differences from SARS-CoV-2. Furthermore, the structure of the RsYN04 RBD in complex with cross-reactive antibody S43 in RBD-7 revealed a potently broad epitope for the development of therapeutics and vaccines. Our findings suggest RsYN04 and other viruses belonging to the same clade have the potential to infect several species, including humans, highlighting the necessity for viral surveillance and development of broad anti-coronavirus countermeasures.

SARS-CoV-2, the causative agent of COVID-19, emerged in December 2019. Its origins remain uncertain. It has been reported that a number of the early human cases had a history of contact with the Huanan Seafood Market. Here we present the results of surveillance for SARS-CoV-2 within the market. From January 1st 2020, after closure of the market, 923 samples were collected from the environment. From 18th January, 457 samples were collected from 18 species of animals, comprising of unsold contents of refrigerators and freezers, swabs from stray animals, and the contents of a fish tank. Using RT-qPCR, SARS-CoV-2 was detected in 73 environmental samples, but none of the animal samples. Three live viruses were successfully isolated. The viruses from the market shared nucleotide identity of 99.99% to 100% with the human isolate HCoV-19/Wuhan/IVDC-HB-01/2019. SARS-CoV-2 lineage A (8782T and 28144C) was found in an environmental sample. RNA-seq analysis of SARS-CoV-2 positive and negative environmental samples showed an abundance of different vertebrate genera at the market. In summary, this study provides information about the distribution and prevalence of SARS-CoV-2 in the Huanan Seafood Market during the early stages of the COVID-19 outbreak.

Influenza C virus (ICV) was identified in five pediatric acute respiratory cases in Shandong. Co-infection with other respiratory viruses was detected in four of these cases. Two ICV genomes were obtained and clustered in the S1-sublineage of C/Sao Paulo/378/82, indicating that genetically diverse ICV strains have been circulating in mainland China.

Influenza C virus (ICV) was identified in five pediatric acute respiratory cases in Shandong. Co-infection with other respiratory viruses was detected in four of these cases. Two ICV genomes were obtained and clustered in the S1-sublineage of C/Sao Paulo/378/82, indicating that genetically diverse ICV strains have been circulating in mainland China.

Two novel reassortant highly pathogenic avian influenza viruses (H5N1) clade 2.3.4.4b.2 were identified in dead migratory birds in China in November 2021. The viruses probably evolved among wild birds through different flyways connecting Europe and Asia. Their low antigenic reaction to vaccine antiserum indicates high risks to poultry and to public health.

Dear Editor,Hepaciviruses, members of the family Flaviviridae, are enveloped viruses containing a single-stranded positive-sense RNA genome of approximately 8.9–10.5 kb in size（Simmonds et al., 2017）. To date, 15species（Hepacivirus A–N, and P） have been documented within the Hepacivirus genus that show distinct host ranges, including primates,bats, horses, donkeys, cows, and various rodents（Hartlage et al., 2016）.

Ticks are important vector hosts of pathogens which cause human and animal diseases worldwide. Diverse viruses have been discovered in ticks; however, little is known about the ecological factors that affect the tick virome composition and evolution. Herein, we employed RNA sequencing to study the virome diversity of the Haemaphysalis longicornis and Rhipicephalus microplus ticks sampled in Hubei Province in China. Twelve RNA viruses with complete genomes were identified, which belonged to six viral families: Flaviviridae, Matonaviridae, Peribunyaviridae, Nairoviridae, Phenuiviridae, and Rhabdoviridae. These viruses showed great diversity in their genome organization and evolution, four of which were proposed to be novel species. The virome diversity and abundance of R. microplus ticks fed on cattle were evidently high. Further ecological analyses suggested that host species and feeding status may be key factors affecting the tick virome structure. This study described a number of novel viral species and variants from ticks and, more importantly, provided insights into the ecological factors shaping the virome structures of ticks, although it clearly warrants further investigation.

Despite the discovery of animal coronaviruses related to SARS-CoV-2, the evolutionary origins of this virus are elusive. We describe a meta-transcriptomic study of 411 bat samples collected from a small geographical region in Yunnan province, China, between May 2019 and November 2020. We identified 24 full-length coronavirus genomes, including four novel SARS-CoV-2-related and three SARS-CoV-related viruses. Rhinolophus pusillus virus RpYN06 was the closest relative of SARS-CoV-2 in most of the genome, although it possessed a more divergent spike gene. The other three SARS-CoV-2-related coronaviruses carried a genetically distinct spike gene that could weakly bind to the hACE2 receptor in vitro. Ecological modeling predicted the co-existence of up to 23 Rhinolophus bat species, with the largest contiguous hot-spots extending from South Laos and Vietnam to southern China. Our study highlights the remarkable diversity of bat coronaviruses at the local scale, including close relatives of both SARS-CoV-2 and SARS-CoV.

Abstract Background The longitudinal antigen-specific immunity in COVID-19 convalescents is crucial for long-term protection upon individual re-exposure to SARS-CoV-2, and even more pivotal for ultimately achieving population-level immunity. To better understand the features of immune memory in individuals with different disease severities at one year post-disease onset we conducted this cohort study. Methods We conducted a systematic antigen-specific immune evaluation in 101 COVID-19 convalescents, who had asymptomatic, mild, moderate, or severe disease, through two visits at months 6 and 12 post-disease onset. The SARS-CoV-2-specific antibodies, comprising NAb, IgG, and IgM, were assessed by mutually corroborated assays, i.e. neutralization, enzyme-linked immunosorbent assay (ELISA), and microparticle chemiluminescence immunoassay (MCLIA). Meanwhile, the T-cell memory against SARS-CoV-2 spike, membrane and nucleocapsid proteins was tested through enzyme-linked immunospot assay (ELISpot), intracellular cytokine staining (ICS), and tetramer staining-based flow cytometry, respectively. Results SARS-CoV-2-specific IgG antibodies, and also NAb can persist among over 95% COVID-19 convalescents from 6 months to 12 months after disease onset. At least 19/71 (26%) of COVID-19 convalescents (double positive in ELISA and MCLIA) had detectable circulating IgM antibody against SARS-CoV-2 at 12m post-disease onset. Notably, the percentages of convalescents with positive SARS-CoV-2-specific T-cell responses (at least one of the SARS-CoV-2 antigen S1, S2, M and N protein) were 71/76 (93%) and 67/73 (92%) at 6m and 12m, respectively. Furthermore, both antibody and T-cell memory levels of the convalescents were positively associated with their disease severity. Conclusions SARS-CoV-2-specific cellular and humoral immunities are durable at least until one year after disease onset.

In early January 2020, the novel coronavirus (SARS-CoV-2) responsible for a pneumonia outbreak in Wuhan, China, was identified using next-generation sequencing (NGS) and readily available bioinformatics pipelines. In addition to virus discovery, these NGS technologies and bioinformatics resources are currently being employed for ongoing genomic surveillance of SARS-CoV-2 worldwide, tracking its spread, evolution and patterns of variation on a global scale. In this review, we summarize the bioinformatics resources used for the discovery and surveillance of SARS-CoV-2. We also discuss the advantages and disadvantages of these bioinformatics resources and highlight areas where additional technical developments are urgently needed. Solutions to these problems will be beneficial not only to the prevention and control of the current COVID-19 pandemic but also to infectious disease outbreaks of the future.

Dear editor, The World Health Organization (WHO) declared the Coronavirus disease 2019 (COVID-19) a pandemic on March 11th, 2020. At the time of writing (Nov 30, 2020), the causative virus SARS-CoV-2 has resulted in >62 million confirmed cases worldwide, with over 1453,000 fatalities (https://covid19.who.int/). Although detection of SARS-CoV-2 has become the top priority for patients with pneumonia, co-infection with other common pathogens should not be neglected as co-infection is often associated with a greater risk of complications, particularly secondary bacterial pneumonia. Several case reports and a large-scale surveillance study on various respiratory pathogens via PCR-based methods targeting a limited number of known pathogens have revealed high rates of microbial co-infection, such as co-infection of SARS-CoV-2 and Influenza virus, Dengue virus or Mycobacterium tuberculosis1Cuadrado-Payán E. et al.SARS-CoV-2 and influenza virus co-infection.Lancet. 2020; 395: e84Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar, 2Saddique Arbab et al. Emergence of co-infection of COVID-19 and dengue: a serious public health threat.J Infect. 2020; https://doi.org/10.1016/j.jinf.2020.08.009Abstract Full Text Full Text PDF PubMed Scopus (41) Google Scholar, 3Kumar Rohit et al. COVID-19 and TB co-infection - 'Finishing touch" in perfect recipe to 'severity' or 'death'-journal of infection.J Infect. 2020; 81: e39-e40Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar. The more informative metagenomic next-generation sequencing (mNGS) approach, particularly meta-transcriptome sequencing, is unbiased for characterizing the total infectome within patients. In this journal, researchers previously reported SARS-CoV-2 and co-infections detected in 8 COVID-19 patients by metagenomics4Thanh Tran Tan et al.SARS-CoV-2 and co-infections detection in nasopharyngeal throat swabs of COVID-19 patients by metagenomics.J Infect. 2020; 81: e175-e177Abstract Full Text Full Text PDF PubMed Scopus (18) Google Scholar. Besides that, only few studies with limited sample size have been reported about using metagenomics to analyze co-infections in COVID-19 cases5Chen L. et al. RNA based mNGS approach identifies a novel human coronavirus from two individual pneumonia cases in 2019 Wuhan outbreak.Emerg Microbes Infect. 2020; 9: 313-319Crossref PubMed Scopus (415) Google Scholar,6Peddu V. et al.Metagenomic analysis reveals clinical SARS-CoV-2 infection and bacterial or viral superinfection and colonization.Clin Chem. 2020; 66: 966-972Crossref PubMed Scopus (48) Google Scholar. Herein, we performed the total transcriptome sequencing of upper respiratory tract samples (nasopharynx swabs and sputum) from 162 PCR-confirmed COVID-19 cases from 12 cities in Shandong province, China. RNA sequencing libraries were constructed using MGIEasy mRNA Library Preparation protocol (MGI) after ribosomal RNA (rRNA) was depleted. 162 libraries were obtained in total and subsequently 100 bp and 150 bp paired-end sequencing of the RNA libraries were performed on the MGISEQ-2000RS platform (MGI). The remaining sequencing reads after mapping to the human genome were compared against the non-redundant nucleotide (nt) database using blastn to identify potential viruses. For a broader microbe discovery, we utilized MetaPhlan27Truong D.T. et al. MetaPhlAn2 for enhanced metagenomic taxonomic profiling.Nat Methods. 2015; 12: 902-903Crossref PubMed Scopus (1299) Google Scholar, which covers ∼1 million unique clade-specific marker genes from bacterial, archaeal and eukaryotic reference genomes. Reads mapping to SARS-CoV-2 were found in all samples, with the absolute read numbers from 14 to 115,299,054, and the number of Reads Per Million (RPM) ranging from 0.07 to 717,538.52. Overall, 82 out of the 162 SARS-CoV-2 cases (50.62%) were co-infected by at least one additional potentially pathogenic microbe (Table 1). Among these, 42 cases (25.93%) were co-infected with one pathogen, including viruses (n = 9), bacteria (n = 33). The remaining 40 cases (24.69%) were co-infected with two or more pathogens, including multiple virus co-infection (n = 1), virus and bacteria co-infections (n = 4), virus, bacteria and fungi co-infections (n = 1), and multiple bacterial co-infections (n = 34). Meanwhile, the abundance of SARS-CoV-2 was lower in samples co-infected with at least one microbe (p < 0.05) compared to that of the samples without co-infections (Fig S1c). Further analysis also showed a significantly lower abundance of SARS-CoV-2 in samples with Streptococcus pneumoniae, Haemophilus parainfluenzae and Neisseria meningitidis (p < 0.05) (Fig S2a, 2d, 2f). However, we did not find a positive correlation between associated factors (age or sex of the patients, variety or abundance of the co-infected microbes) and the abundance of SARS-CoV-2 (Fig S1a-b and Fig S2g-l).Table 1Summary of co-infections of SARS-CoV-2 and other microbes.SARS-CoV-2Co-infecting microbesNo. of casesPercent+Virus91.85%+Multiple viruses10.62%+Virus + Bacteria42.47%+Virus + Bacteria + Fungi10.62%+One bacterium3320.37%+Multiple bacteria3420.99%Total8250.62% Open table in a new tab We identified 7 viruses with potential pathogenicity in 15 of the 162 (9.26%) COVID-19 cases. Human alphaherpesvirus 2 was the most frequently detected virus (n = 6), followed by Human H3N2 influenza virus (n = 3), Human coronavirus 229E (n = 2), Human metapneumovirus (n = 2), and Human coronavirus NL63 (n = 1) (Fig 1a, Table S1). In addition, two rare respiratory viruses, Human rhinovirus C11 (n = 1, RPM: 6.84) first reported in 2020 and Human enterovirus C105 (n = 1, RPM: 6969.31) first reported in 2019 in China, were also identified (Fig 1a, Table S1), sharing 91% and 96% nucleotide identity to known reference viruses, respectively. Human enterovirus C105 has been associated with acute flaccid paralysis in children and respiratory tract infection in teenagers8Horner L.M. et al. Acute Flaccid Paralysis Associated with Novel Enterovirus C105.Emerg Infect Dis. 2015; 21: 1858-1860Crossref PubMed Scopus (26) Google Scholar. Our results revealed a relative low co-infection rate of other respiratory viruses with SARS-CoV-2: 9.26% (15/162) versus 20.7% (24/116) in a previous report9Kim D. et al. Rates of Co-infection between SARS-CoV-2 and other respiratory pathogens.JAMA. 2020; 323: 2085-2086Crossref PubMed Scopus (491) Google Scholar. High rates of co-infection of common, but important pathogenic or opportunistic bacteria were also detected (Fig 1a), including S. pneumoniae (n = 37), Stenotrophomonas maltophilia (n = 31), Pseudomonas putida (n = 21), H. parainfluenzae (n = 19), Haemophilus influenzae (n = 14), N. meningitidis (n = 11), Moraxella catarrhalis (n = 3), Streptococcus pyogenes (n = 1), Streptococcus epidermidis (n = 1), as well as two species of mycoplasma: Mycoplasma hyorhinis (n = 3) and Mycoplasma pneumoniae (n = 1) (Fig 1a). Importantly, all bacterial infections were confirmed by PCR using species-specific primers. As a commensal microbe in the respiratory tract of pigs and rarely seen in humans, M. hyorhinis was unexpected identified in three cases, although it was present at a relatively low abundance (39–79 RPM). Overall, 72 of the 162 SARS-CoV-2 patients (44.44%) possessed at least one additional bacterium. One species of fungi, Candida albicans (n = 1, RPM: 600) was identified (Fig 1a). Our results highlighted more potential co-infections with bacteria (44.44%, 72/162) than with viruses (9.26%, 15/162) or fungi (0.62, 1/162), in contrast to a recent study10Zhang H.C. et al. Metatranscriptomic characterization of COVID-19 identified a host transcriptional classifier associated with immune signaling.Clin Infect Dis. 2020; (ciaa663)https://doi.org/10.1093/cid/ciaa663Crossref Scopus (64) Google Scholar. We further analyzed the expression of 52 unique antibiotic resistance genes (ARGs) associated with phenotypic resistance to seven classes of antibiotics: aminoglycoside, nitroimidazole, sulphonamide, phenicol, tetracycline, beta-lactam and macrolide (Fig. 1b). The number of ARGs varied sharply from 0 to 28 among the 162 SARS-CoV-2 cases. Regarding those with concurrent infections (n = 82), most presented resistance to aminoglycosides (66/82), beta-lactam (78/82), macrolide (74/82) and tetracycline (63/82), whereas a small number of samples possessed genes resistant to nitroimidazole (1/82), phenical (26/82) and sulphonamide (24/82) (Fig 1b). Unsurprisingly, a greater diversity of resistance genes was identified in samples with more bacteria (p < 0.05) (Fig S1d). In sum, our mNGS analysis did not reveal high co-infection rates of other respiratory viruses (such as influenza viruses), mycoplasma and fungi with SARS-CoV-2. However, we did document high rates of co-infection between SARS-CoV-2 and multiple bacteria with numerous ARGs. Although the potential influence of co-infections of SARS-CoV-2 and other microbes in disease progress and severity remains poorly understood, the treatment of bacterial infection might provide clinical benefit for COVID-19 in cases where therapeutics are available. This study was approved by the ethics committee of the Shandong First Medical University & Shandong Academy of Medical Sciences. The research-related information was used anonymously. The whole research was supervised by the Health Commission of Shandong Province. All human-related sample processing and sequencing were performed in accordance with relevant guidelines and regulations of the Shandong Provincial CDC. Conceived the project: W.S., D.K., P.H.; collected samples: T.L., N.N., H.Z., S.R., P.H.; performed laboratory work: H.Z., C.L., T.H., J.L.; analysed the data: H.Z., C.L., T.H., W.C., H.W., S.F., O.G.P., W.S.; wrote the paper: H.Z., C.L., E.C.H., W.S. All authors have read and approved the final manuscript. Fig. S1, Fig. S2Fig. S2Comparisons of the abundance of SARS-CoV-2 between samples co-infected with different bacteria and samples without the counterpart bacterium. (a)S. pneumoniae, (b) S. maltophilia, (c) P. putida, (d) H. parainfluenzae, (e) H. influenzae and (f) N. meningitidis. Differences between groups were assessed with a Wilcoxon test. (g) S. pneumoniae, (h) S. maltophilia, (i) P. putida, (j) H. parainfluenzae, (k) H. influenzae and (l) N. meningitidis. Correlation coefficient between groups were assessed with Spearman's rank correlation test.View Large Image Figure ViewerDownload Hi-res image Download (PPT) The authors declare no conflicts of interest. This work was supported by Key research and development project of Shandong province (Grant No. 2020SFXGFY01 and 2020SFXGFY08), and the Academic Promotion Programme of Shandong First Medical University (2019QL006). W.F.S is supported by the Taishan Scholars Programme of Shandong Province (ts201511056). E.C.H. is supported by an Australian Research Council Australian Laureate Fellowship (FL170100022). 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Background. The longitudinal antigen-specific immunity in COVID-19 convalescents is crucial for long-term protection upon individual re-exposure to SARS-CoV-2, and even more pivotal for ultimately achieving population-level immunity. We conducted this cohort study to better understand the features of immune memory in individuals with different disease severities at 1 year post-disease onset. Methods. We conducted a systematic antigen-specific immune evaluation in 101 COVID-19 convalescents, who had asymptomatic, mild, moderate, or severe disease, through 2 visits at months 6 and 12 after disease onset. The SARS-CoV-2-specific antibodies, comprising neutralizing antibody (NAb), immunoglobulin (Ig) G, and IgM, were assessed by mutually corroborated assays (ie, neutralization, enzyme-linked immunosorbent assay [ELISA], and microparticle chemiluminescence immunoassay [MCLIA]). Meanwhile, T-cell memory against SARS-CoV-2 spike, membrane, and nucleocapsid proteins was tested through enzyme-linked immunospot assay (ELISpot), intracellular cytokine staining, and tetramer staining-based flow cytometry, respectively. Results. SARS-CoV-2-specific IgG antibodies, and NAb, can persist among >95% of COVID-19 convalescents from 6 to 12 months after disease onset. At least 19/71 (26%) of COVID-19 convalescents (double positive in ELISA and MCLIA) had detectable circulating IgM antibody against SARS-CoV-2 at 12 months post-disease onset. Notably, numbers of convalescents with positive SARS-CoV-2-specific T-cell responses (>= 1 of the SARS-CoV-2 antigen S1, S2, M, and N proteins) were 71/76 (93%) and 67/73 (92%) at 6 and 12 months, respectively. Furthermore, both antibody and T-cell memory levels in the convalescents were positively associated with disease severity. Conclusions. SARS-CoV-2-specific cellular and humoral immunities are durable at least until 1 year after disease onset.

The live poultry trade is thought to play an important role in the spread and maintenance of highly pathogenic avian influenza A viruses (HP AIVs) in Asia. Despite an abundance of small-scale observational studies, the role of the poultry trade in disseminating AIV over large geographic areas is still unclear, especially for developing countries with complex poultry production systems. Here we combine virus genomes and reconstructed poultry transportation data to measure and compare the spatial spread in China of three key subtypes of AIV: H5N1, H7N9, and H5N6. Although it is difficult to disentangle the contribution of confounding factors, such as bird migration and spatial distance, we find evidence that the dissemination of these subtypes among domestic poultry is geographically continuous and likely associated with the intensity of the live poultry trade in China. Using two independent data sources and network analysis methods, we report a regional-scale community structure in China that might explain the spread of AIV subtypes in the country. The identification of this structure has the potential to inform more targeted strategies for the prevention and control of AIV in China.

Coronavirus disease 2019 (COVID-19) was designated a global pandemic by the World Health Organization (WHO) on March 11, 2020 (1).After great effort, COVID-19 has been well-controlled in China, but new challenges have emerged due to increasing numbers of imported cases from outside of China.On May 8, the Health Commission of Jilin Province reported a confirmed COVID-19 case of a 45-year-old laundry woman from Shulan City in the northeast of China.This case was suspected to be associated with a possible importation event.On May 10, genome sequencing was performed at China CDC using the Illumina MiSeq platform.Due to the low viral load in the samples, 98.1% of the virus genome sequence was obtained after sequencing and assembly, which included the key sites of the virus, and the analysis of the potential source of the virus was not affected.Compared with the reference sequence EPI_ISL_402119 (2), which was isolated from Wuhan on January 7, 2020, the virus had mutations at C241T, C3037T, C14408T, and A23403G, and the GGG mutated to AAC at position 28881-28883 of nucleotide of COVID-19 virus, which shows gene mutation characteristics of the L-lineage from Europe and does not have characteristic mutations of the North America branch of the European L-lineage (mutants at C1059T and G25563T) according to the study of Tang et al (Figure 1) (3).Additionally, the genome sequence of such a case also had a mutation at T19839C.The result of viral genome sequencing provides strong evidence that the first COVID-19 case in Shulan City is related to COVID-19 virus imported from Europe.Although isolation of the imported cases and close contacts are the top priority strategies to interrupt the spread of the virus in China, asymptomatic infected persons may be a potential source of infection for this case or an unknown source of sporadic cases.We suggest that COVID-19 virus nucleic acid testing for respiratory and stool specimens of the imported cases, close contacts, and inbound passengers whom come from high risk countries, territories, and areas should continue to be strengthened.Only when the nucleic acid tests of respiratory and stool specimens are negative at the same time, the isolation can be ended.Epidemiological investigation, case treatment of asymptomatic infections, and close contact tracing management should be continuously accelerated.