The aim of this study was to compare the immune responses of C57BL/6 mice immunized with two pathogens, foot-and-mouth disease virus (FMDV) and Senecavirus A (SVA), and to provide clues for revealing the regulatory mechanisms of acquired immunity. Inactivated and purified FMDV and SVA antigens were used to immunize C57BL/6 mice respectively, and the mice immunized with PBS were taken as the control. The percentages of Th1 and Th2 cells in the spleen lymphocytes of mice in each group were analyzed by flow cytometry at 14 and 28 days after immunization. RNA-Seq was performed for the spleen. Mouse macrophages were stimulated with the antigens in vitro to examine the expression of the differentially expressed genes (DEGs) screened out. The results showed that 14 days after immunization, there was no significant difference in the magnitude of the Th1/Th2 immune response elicited by the FMDV and SVA antigens. After 28 days, the magnitudes of the Th1 and Th2 immune responses elicited by the SVA antigen were higher than those elicited by the FMDV antigen. RNA-Seq revealed two common DEGs, Rsad2 and Tspan8, between the two immunization groups, which indicated that the two genes may be involved in the activation of the Th1/Th2 immune responses by FMDV and SVA antigens. FMDV and SVA antigens stimulated macrophages to secrete interleukin (IL)-12 and IL-33 in vitro, and the expression of Tspan8 and Rsad2 was consistent with the RNA-Seq results. The expression of Rsad2 was regulated by type I interferons (IFNα, IFNβ). In this study, we obtained the DEGs involved in the immune responses to the two antigens in mouse spleen, which provides a molecular basis for investigating the immune response mechanisms induced by FMDV and SVA.
Animals ; Foot-and-Mouth Disease Virus/genetics* ; Mice ; Spleen/cytology* ; Mice, Inbred C57BL ; Antigens, Viral/genetics* ; Transcriptome ; Th1 Cells/immunology* ; Immunization ; Viral Vaccines/immunology* ; Th2 Cells/immunology* ; Foot-and-Mouth Disease/immunology* ; Interleukin-33/genetics* ; Female ; Macrophages/immunology* ; Picornaviridae

OBJECTIVE: To improve the understanding of the virome and bacterial microbiome in the wildlife rescue station of Poyang Lake, China. METHODS: Ten smear samples were collected in March 2019. Metagenomic sequencing was performed to delineate bacterial and viral diversity. Taxonomic analysis was performed using the Kraken2 and Bracken methods. A maximum-likelihood tree was constructed based on the RNA-dependent RNA polymerase (RdRp) region of picornavirus. RESULTS: We identified 363 bacterial and 6 viral families. A significant difference in microbial and viral abundance was found between samples S01-S09 and S10. In S01-S09, members of Flavobacteriia and Gammaproteobacteria were the most prevalent, while in S10, the most prevalent bacteria class was Actinomycetia. Among S01-S09, members of Myoviridae and Herelleviridae were the most prevalent, while the dominant virus family of S10 was Picornaviridae. The full genome of the pigeon mesivirus-like virus (NC-BM-233) was recovered from S10 and contained an open reading frame of 8,124 nt. It showed the best hit to the pigeon mesivirus 2 polyprotein, with 84.10% amino acid identity. Phylogenetic analysis showed that RdRp clustered into Megrivirus B. CONCLUSION This study provides an initial assessment of the bacteria and viruses in the cage-smeared samples, broadens our knowledge of viral and bacterial diversity, and is a way to discover potential pathogens in wild birds.
Animals ; Animals, Wild/genetics* ; Lakes ; Phylogeny ; Picornaviridae/genetics* ; Viruses/genetics* ; China ; Metagenomics ; Genome, Viral

To develop Senecavirus A (SVA) virus-like particles (VLPs), a recombinant prokaryotic expression plasmid pET28a-SVA-VP031 was constructed to co-express SVA structural proteins VP0, VP3 and VP1, according to the genomic sequence of the field isolate CH-FJ-2017 after the recombinant proteins were expressed in E .coli system, and purified by Ni+ ion chromatographic method. The SVA VLPs self-assemble with a high yield in vitro buffer. A typical VLPs with an average diameter of 25-30 nm which is similar to native virions by using TEM detection. Animals immunized by SVA VLPs shown that the VLPs induced high titers neutralizing antibodies in Guinea pigs. This study indicated that the VLPs produced with co-expressing SVA structural proteins VP0, VP3 and VP1 in prokaryotic system is a promising candidate and laid an important foundation for the development of a novel SVA VLPs vaccine.
Animals ; Antibodies, Neutralizing ; Escherichia coli/genetics* ; Genomics ; Guinea Pigs ; Picornaviridae/genetics*

Both sides of the picornavirus genome have 5'-untranslated region (5'UTR) and 3'- untranslated region (3'UTR). This study demontrated that both the 5'-and 3'-UTR can form complex structures, such as stem-loop, clover and pseudoknot structure, These structures play an important role in the regulaton of the replication and translation of the viruses. This article reviewed the progress of research on the structure and function of picornavirus' 3'-UTR over recent years.
3' Untranslated Regions ; Animals ; Humans ; Nucleic Acid Conformation ; Picornaviridae ; chemistry ; genetics ; metabolism ; Picornaviridae Infections ; virology ; RNA, Viral ; chemistry ; genetics ; metabolism

OBJECTIVEHuman parechovirus (HPeV) is a single-stranded, positive sense RNA virus in the Parechovirus genus within the large family of Picornaviridae. As a possible new pathogen of neonatal sepsis, meningoencephalitis and other infections in young children, HPeV gets more and more attention. This study aimed to better understand the association of HPeV with central nervous system (CNS) infectious diseases and sepsis among hospitalized children in Beijing.

METHODA total of 577 cerebrospinal fluid (CSF) samples were retrospectively collected from 557 children suspected of CNS infections in 2012. Three hundred and fifty-one of them were male and 206 were female. HPeV was screened by reverse transcription-nested PCR (RT-nPCR) with the universal primers which target the highly conserved 5'UTR. The positive samples were genotyped by amplifying and sequencing for the VP3/VP1 junction region. The sequences were compared with the HPeV sequences from GenBank and performed phylogenetic analysis.Some samples other than CSF from HPeV positive children, including serum, nasopharyngeal aspirate and stool, were collected and carried out screening for HPeV.

RESULTWith the RT-nPCR by universal primers, HPeVs were detected in 18 out of 577 CSF samples obtained from 18 children with a positive rate of 3.1%. The ratio of male and female was 2: 1. There were no statistically significant differences on infection rate between boys (12/351, 3.4%) and girls (6/206, 2.9%). All of 18 positive CSF samples were negative for enterovirus, Epstein-Barr virus (EBV), human cytomegalovirus (HCMV), and herpes simplex virus 1 and 2 (HSV).HPeVs from 10 positive CSF samples were genotyped successfully, consisting of 7 HPeV3 and 3 HPeV1. In addition, 2 of 8 serum samples were positive for HPeV3 and 1 of 2 stool samples were positive for HPeV 1. HPeVs were identified in CSF from children aged from 15 days to 14 years, in which 7 cases were infants younger than 3 months and 5 cases were infants from 3 months to one year. Three children older than the age of 9 years (9, 13 and 14 years) were positive for HPeV. Most of the children (6/8) infected with HPeV3 were younger than 3 months and were diagnosed as sepsis, while the rest of HPeV3 positive children were diagnosed as meningitis and bronchopneumonia. HPeV3 infection clustered in August, while HPeV1 in January.

CONCLUSIONHPeVs were associated with CNS infections and sepsis in hospitalized children in Beijing, especially in children younger than one year.HPeV3 was the predominant type identified in CSF.

Adolescent ; Age Distribution ; Central Nervous System Infections ; cerebrospinal fluid ; epidemiology ; virology ; Cerebrospinal Fluid ; virology ; Child ; Child, Hospitalized ; Child, Preschool ; Female ; Genotype ; Humans ; Infant ; Infant, Newborn ; Male ; Parechovirus ; classification ; genetics ; isolation & purification ; Picornaviridae Infections ; cerebrospinal fluid ; epidemiology ; virology ; RNA, Viral ; genetics ; Retrospective Studies ; Reverse Transcriptase Polymerase Chain Reaction ; Seasons ; Sepsis ; cerebrospinal fluid ; epidemiology ; virology ; Sequence Analysis, DNA

Antibodies, Viral ; analysis ; Cerebrospinal Fluid ; virology ; Child ; Child, Preschool ; Feces ; virology ; Genotype ; Humans ; Infant ; Meningoencephalitis ; diagnosis ; virology ; Parechovirus ; classification ; genetics ; isolation & purification ; Picornaviridae Infections ; diagnosis ; epidemiology ; virology ; RNA, Viral ; genetics ; Respiratory Tract Infections ; diagnosis ; virology ; Sepsis ; diagnosis ; virology ; Sequence Analysis, DNA

Bats are important reservoir animals and more than 60 viruses have been identified in bats with many of them highly pathogenic to human. In order to understand the natural background, genetic diversity of bat viruses in China and discover potential viral pathogens, Solexa sequencing based viral metagenomics focusing on bats tissues was established and to analyze the virome of bats collected from Jilin, Yunnan and Hunan province. By Solexa sequencing, 116 442 324 useful reads were obtained and assembled into 4 872 contigs, of which 8.2% (4 002/4 4872) were annotated to 36 viral families, including 19 vertebrate virus families, 6 plant virus families, 4 insect virus families and 4 phages. Further contigs analyses showed that some adenovirus, bocavirus, picobirnavirus, parvovirus contigs sequences were similar with known viruses. However, part of them shared limited identities to these viruses implying the discovery of new viruses. Moreover, PCR validation of adenovirus and bocavirus confirmed the results obtained by viral metagenomics. This study aimed to understand bat virome in China by viral metagenomics and could be helpful to establish effective surveillance on wildlife-associate zoonoses.
Adenoviridae ; genetics ; isolation & purification ; Animals ; Bunyaviridae ; genetics ; isolation & purification ; China ; Chiroptera ; virology ; Genome, Viral ; genetics ; Metagenome ; genetics ; Metagenomics ; methods ; Picornaviridae ; genetics ; isolation & purification

OBJECTIVETo understand the clinical characteristics of different groups human rhinovirus (HRV)-A, B and C infection in children with acute respiratory tract infections (ARI) in Beijing.

METHODRespiratory tract specimens (n = 1412) collected from children with ARI during Jan. 2011 to Dec. 2012 were tested for HRV by using semi-nested PCR. Gene fragments of VP4/VP2 capsid protein amplified from HRV positive specimens were sequenced for HRV genotype confirmation. Then epidemiological characteristics of these HRV-positive cases were analyzed.

RESULTAmong these 1412 specimens tested, 103 (7.3%) were HRV positive, including 54 (52.4%) positive for HRV-A, 14 (13.6%) for HRV-B, 35 (34.0%) for HRV-C determined by sequence analysis. The positive rates of HRV-A, B and C (2.5%, 16/638; 0.3%, 2/638 and 1.3%, 8/638) in children with acute upper respiratory tract infections (URI) were lower than those (5.8%, 36/623; 1.8%, 11/623 and 3.9%, 24/623) in children with acute lower respiratory tract infections (LRI) (P = 0.003, 0.011, 0.003). In children with LRI, the positive rates of HRV-A, C were similar to each other (P = 0.112), and both were higher than that of HRV-B (P = 0.000, P = 0.026). The severity of ARI among children positive for different groups HRV showed no significant difference evaluated by Kruskal-Wallis H test (Hc = 0.044, P > 0.05), as well as that between children co-infected with HRV and other viruses and those infected with HRV only evaluated by Wilcoxon rank sum test (Zc = 0.872, P > 0.05).

CONCLUSIONHRV is one of important pathogens for children with ARI, especially LRI in Beijing. The positive rates of HRV-A and HRV-C are similar to each other, and both are higher than that of HRV-B. No significant difference was shown among children with different HRV genotypes by evaluation of the severity of ARI, and co-infections of HRV with other viruses do not significantly increase the severity of ARI.

Acute Disease ; Adolescent ; Age Distribution ; Child ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Infant ; Infant, Newborn ; Male ; Phylogeny ; Picornaviridae Infections ; epidemiology ; virology ; Respiratory Tract Infections ; epidemiology ; virology ; Reverse Transcriptase Polymerase Chain Reaction ; Rhinovirus ; classification ; genetics ; isolation & purification ; Sequence Analysis, RNA ; Severity of Illness Index

Coxsackievirus A16 belongs to the family Picornaviridae, and is a major agent of hand-foot-and-mouth disease that infects mostly children, and to date no vaccines or antiviral therapies are available. 2A protease of enterovirus is a nonstructural protein and possesses both self-cleavage activity and the ability to cleave the eukaryotic translation initiation factor 4G. Here we present the crystal structure of coxsackievirus A16 2A protease, which interestingly forms hexamers in crystal as well as in solution. This structure shows an open conformation, with its active site accessible, ready for substrate binding and cleavage activity. In conjunction with a previously reported "closed" state structure of human rhinovirus 2, we were able to develop a detailed hypothesis for the conformational conversion triggered by two "switcher" residues Glu88 and Tyr89 located within the bll2-cII loop. Substrate recognition assays revealed that amino acid residues P1', P2 and P4 are essential for substrate specificity, which was verified by our substrate binding model. In addition, we compared the in vitro cleavage efficiency of 2A proteases from coxsackievirus A16 and enterovirus 71 upon the same substrates by fluorescence resonance energy transfer (FRET), and observed higher protease activity of enterovirus 71 compared to that of coxsackievirus A16. In conclusion, our study shows an open conformation of coxsackievirus A16 2A protease and the underlying mechanisms for conformational conversion and substrate specificity. These new insights should facilitate the future rational design of efficient 2A protease inhibitors.
Coxsackievirus Infections ; virology ; Crystallography, X-Ray ; Cysteine Endopeptidases ; chemistry ; genetics ; Fluorescence Resonance Energy Transfer ; Hand, Foot and Mouth Disease ; enzymology ; pathology ; virology ; Humans ; Picornaviridae ; chemistry ; enzymology ; genetics ; Protein Conformation ; Structure-Activity Relationship ; Substrate Specificity ; Viral Proteins ; chemistry ; genetics

To understand the infections and molecular biological characteristics of different human rhinovirus (HRV) genotypes -A, B, C, especially C in children with acute respiratory tract infections (ARI) in Beijing. Seven hundreds and three respiratory tract specimens were collected from children with ARI during Jan. 2011 to Dec. 2011. Semi-nested PCR was developed for detecting HRVs. Gene fragment of VP4/VP2 capsid protein amplified from HRV positive specimens was sequenced and analyzed by software DNAStar, the phylogenetic tree was then constructed by MEGA 5. 05. Among these 703 specimens tested, 54 (7.7%, 54/703) were HRV positive, including 25 (46.3%, 25/54) positive for HRV-A, 8 (14. 8%, 8/54) for HRV-B, 21 (38. 9%, 21/54) for HRV-C determined by sequence analysis. Most of these children (94. 4%00, 51/54) infected with HRVs were younger than 5 years old, and the highest positive rate was shown in group younger than 1 year (11. 4%). These patients positive for HRVs were diagnosed as bronchiolitis (23.1%), asthma (20.0%), pneumonia (1.0%), bronchitis (4.4%) and upper respiratory tract infections (4. 1%). Sequence analysis of VP4/VP2 gene fragment revealed that 70. 0% to 100. 0% nucleotide identity was shown among the sequences within the same HRV genotype, and 55. 5% to 65. 8% nucleotide identity among the sequences from different HRV genotypes. In conclusion, HRVs, especially HRV-C, are important pathogens for children with ARI in Beijing. The prevalence of HRV-C is similar to that of HRV-A, higher than that of HRV-B. High sequence variation among different HRV genotypes was indicated in this study.
Acute Disease ; epidemiology ; Adolescent ; Child ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Infant ; Male ; Molecular Sequence Data ; Phylogeny ; Picornaviridae Infections ; epidemiology ; virology ; Respiratory Tract Infections ; epidemiology ; virology ; Rhinovirus ; classification ; genetics ; isolation & purification ; Seasons ; Viral Proteins ; genetics