Metagenomic Analysis of Environmental Samples from Wildlife Rescue Station at Poyang Lake, China / 生物医学与环境科学（英文）

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.

Boletín epidemiológico 12 - 2022. Semana epidemiológica del 20 al 26 de marzo del 2022 / Epidemiological bulletin 12 - 2022. Epidemiological week from March 20 to 26, 2022

La enfermedad de mano, pie y boca (EMPB) es una patología infecciosa pediátrica común, causada por el enterovirus (EV) de la familia Picornaviridae, incluidos EV-A71 y los virus Coxsackie (CV) CV-A2, CV-A6, CV-A10 y CV- A16. Aunque generalmente es autolimitada, puede provocar graves complicaciones asociadas con una infección neurológica (encefalitis, meningitis) o una enfermedad respiratoria fatal. La mayoría de los casos presentan fiebre, erupciones cutáneas en manos y pies y vesículas o úlceras en la mucosa bucal. Los casos afectan principalmente a niños entre los 0 a 5 años, pero también puede afectar a niños mayores y adultos

Perfil dos portadores de hepatite A no Estado do Rio de Janeiro, no período de 2010 a 2018 / Profile of hepatitis carriers in the State of Rio de Janeiro, from 2010 to 2018

Objetivo: Este estudo busca caracterizar o perfil dos indivíduos com hepatite A, no Estado do Rio de Janeiro, no período de 2010 a 2018. Metodologia: Trata-se de uma pesquisa transversal desenvolvida na abordagem quantitativa. Foram utilizados dados secundários provenientes da Ficha de Notificação das Hepatites Virais do banco do Sistema de Informação de Agravos de Notificação, cedida pela Secretaria Estadual de Saúde do Rio de Janeiro. Resultados: Foram analisados 48672 notificações, sendo que 3734 foram notificações confirmadas de portadores exclusivo da hepatite A, 28719 referentes a outras hepatites, 614 de co-infecção por mais de uma hepatite viral e 15605 ignoradas. A maior prevalência da hepatite A foi encontrada nos indivíduos com 4 a 8 anos de estudo, da raça negra, faixa ≤ 24 anos e sendo água e alimentos contaminados a principal fonte de contágio. Conclusão: A notificação dos casos e o preenchimento adequado é importante para definições de ações sanitárias capazes de mitigar os danos à saúde pública. Palavras-chave: Hepatite A; Notificação; Prevalência.

Objective: To characterize the profile of individuals with hepatitis A in the State of Rio de Janeiro, from 2010 to 2018. Methodology: This is a cross-sectional research developed in the quantitative approach. Secondary data from the Viral Hepatitis Notification Form of the Bank of the Notifiable Diseases Information System, provided by the State Health Secretariat of Rio de Janeiro, were used. Results: 48672 notifications were analyzed, of which 3734 were confirmed notifications of hepatitis A carriers only, 28719 were related to other hepatitis, 614 were co-infected with more than one viral hepatitis and 15605 were ignored. The highest prevalence of hepatitis A was found in individuals with 4 to 8 years of study, black, aged ≤ 24 years and with contaminated water and food being the main source of contagion. Conclusion: Notification of cases and proper completion is important for the definition of health actions capable of mitigating damage to public health. Keywords: Hepatitis A; Notification; Prevalence.

Objetivo: Este estudio busca caracterizar el perfil de individuos con hepatitis A, en el estado de Río de Janeiro, de 2010 a 2018. Metodología: Se trata de una investigación transversal desarrollada en el enfoque cuantitativo. Se utilizaron datos secundarios del Formulario de Notificación de Hepatitis Virales del Banco del Sistema de Información de Enfermedades Notificables, proporcionado por la Secretaría de Salud del Estado de Río de Janeiro. Resultados: Se analizaron 48672 notificaciones, de las cuales 3734 fueron notificaciones confirmadas de solo portadores de hepatitis A, 28719 estaban relacionadas con otras hepatitis, 614 estaban coinfectadas con más de una hepatitis viral y 15605 fueron ignoradas. La mayor prevalencia de hepatitis A se encontró en individuos de 4 a 8 años de estudio, negros, ≤ 24 años y con agua y alimentos contaminados como la principal fuente de contagio. Conclusión: La notificación de casos y su debida cumplimentación es importante para la definición de acciones de salud capaces de mitigar los daños a la salud pública. Palabra clave: Hepatitis A; Notificación; Prevalencia.

Generation and immunogenicity evaluation of Senecavirus A virus-like particles / 生物工程学报

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.

Human virome in nasopharynx and tracheal secretion samples

BACKGROUND In Brazil the implementation of the Sentinel Surveillance System of Influenza began in 2000. Central public health laboratories use reverse transcription-quantitative polymerase chain reaction (RT-qPCR) for diagnosis of respiratory viruses, but this protocol identifies only specific targets, resulted in inconclusive diagnosis for many samples. Thus, high-throughput sequencing (HTS) would be complementary method in the identification of pathogens in inconclusive samples for RT-qPCR or other specific detection protocols. OBJECTIVES This study aimed to detect unidentified viruses using HTS approach in negative samples of nasopharynx/tracheal secretions by the standard RT-qPCR collected in the Federal District, Brazil. METHODS Nucleic acids were extracted from samples collected in winter period of 2016 and subjected to HTS. The results were confirmed by the multiplex PR21 RT-qPCR, which identifies 21 respiratory pathogens. FINDINGS The main viruses identified by HTS were of families Herpesviridae, Coronaviridae, Parvoviridae and Picornaviridae, with the emphasis on rhinoviruses. The presence of respiratory viruses in the samples was confirmed by the PR21 multiplex RT-qPCR. Coronavirus, enterovirus, bocavirus and rhinovirus were found by multiplex RT-qPCR as well as by HTS analyses. MAIN CONCLUSIONS Wide virus diversity was found by different methodologies and high frequency of rhinovirus occurrence was confirmed in population in winter, showing its relevance for public health.

The role of microRNAs in enteroviral infections

ABSTRACTThe genus Enterovirus, a member of thePicornavirus family, are RNA viruses that can cause poliomyelitis, hand-food-mouth disease, viral meningitis or meningoencephalitis, viral myocarditis and so on. MicroRNAs are a class of highly conserved, small noncoding RNAs recognized as important regulators of gene expression. Recent studies found that MicroRNAs play a significant role in the infection ofEnterovirus, such as enterovirus 71, coxsackievirus B3 and other Enterovirus. Enteroviral infection can alter the expression of cellular MicroRNAs, and cellular MicroRNAs can modulate viral pathogenesis and replication by regulating the expression level of viral or host's genes. Herein, this review summarizes the role of MicroRNAs in enteroviral infection.

Detection of respiratory viruses by real-time polymerase chain reaction in outpatients with acute respiratory infection

Viruses are the major contributors to the morbidity and mortality of upper and lower acute respiratory infections (ARIs) for all age groups. The aim of this study was to determine the frequencies for a large range of respiratory viruses using a sensitive molecular detection technique in specimens from outpatients of all ages with ARIs. Nasopharyngeal aspirates were obtained from 162 individuals between August 2007-August 2009. Twenty-three pathogenic respiratory agents, 18 respiratory viruses and five bacteria were investigated using multiplex real-time reverse transcriptase polymerase chain reaction (RT-PCR) and indirect immunofluorescence assay (IIF). Through IIF, 33 (20.4%) specimens with respiratory virus were recognised, with influenza virus representing over half of the positive samples. Through a multiplex real-time RT-PCR assay, 88 (54.3%) positive samples were detected; the most prevalent respiratory viral pathogens were influenza, human rhinovirus and respiratory syncytial virus (RSV). Six cases of viral co-detection were observed, mainly involving RSV. The use of multiplex real-time RT-PCR increased the viral detection by 33.9% and revealed a larger number of respiratory viruses implicated in ARI cases, including the most recently described respiratory viruses [human bocavirus, human metapneumovirus, influenza A (H1N1) pdm09 virus, human coronavirus (HCoV) NL63 and HCoV HKU1].

Structure and function of 3'- untranslated region in picornavirus / 病毒学报

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.

Current research on picornavirus 3C protease / 病毒学报

The picornavirus family comprises many small viruses, several of which are important pathogens of humans and livestock. The 3C protease (3Cpro) of different species and genera of picornavirus contains the classic G-X-C-G motif and Cys-His-Asp/Glu catalytic triad. 3Cpro conducts maturation cleavage in the regions of VP2-VP3 and VP3-VP1 in P1, 2A-2B and 2B-2C in P2 and the whole P3. Picornavirus 3Cpro has been shown to have significant substrate preference in Q-G/S/A/V/H/R and E-S/G/R/M as well as species and genera specificity through analyses of the maturation cleavage of picornavirus polyproteins. Innate immune adaptors such as TRIF, MAVS, IRF3, IRF7 and NEMO have various potential cleavage sites in picornavirus 3Cpro (TRIF and NEMO show considerable diversity in their cleavage sites). Useful information will be provided for the development of broad-spectrum antiviral agents as well as evasion mechanisms of the innate immune system against picornavirus 3Cpro through continued research of picornavirus 3Cpro.

Metagenomic analysis of bat virome in several Chinese regions / 生物工程学报

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.

An open conformation determined by a structural switch for 2A protease from coxsackievirus A16

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.

Rhinovirus detection using different PCR-based strategies

Human rhinoviruses (HRVs) are the major cause of the common cold. HRVs were recently reclassified into the Enterovirus genus (HEV) in the Picornaviridae family. HRVs and other members of the HEV genus share many common features, including sense RNA genomes and partial nucleotide sequence identity. The aim of this study was to evaluate different HRV detection strategies. Samples from adults with acute respiratory infection (n = 291) who were treated in Sao Paulo Hospital (2001-2003) were tested using three assays. The first assay detected picornaviruses by RT-PCR and hybridization, the second detected rhinoviruses using RT-PCR/sequencing, and the third differentiated HRV from HEV using duplex semi-nested-RT-PCR. Analysis of the results obtained from the first two strategies revealed 83% concordance. Discordant samples were then evaluated by the third protocol, and 82% were negative. The picornavirus detection protocol was more sensitive but less specific than the rhinovirus detection protocols. The semi-nested protocol utilized in the present study was less sensitive and was not useful in differentiating HRV from HEV. Sequencing assays examining different genes would address the best strategy of confirming rhinovirus and enterovirus infections.

Updates in the Relationship Between Human Rhinovirus and Asthma

Human rhinovirus (HRV) is a nonenveloped, single stranded RNA virus belonging to the family Picornaviridae. HRV infections can cause both upper and lower respiratory illnesses in children and adults. Lower respiratory illnesses are more likely to occur in specific high risk groups, including infants, and children and adults with asthma. The relationships between rates of infection and the risk of clinical illness and exacerbation are not completely understood. Recent studies employing polymerase chain reaction and other molecular techniques indicate that there are new branches on the HRV family tree, and one characteristic of recently detected viruses is that they cannot be detected by standard tissue culture. Here we review the current literature and discuss new advances in understanding the link between HRV and asthma.

Vigilância laboratorial das paralisias flácidas agudas no Brasil, no período de 2007 a 2011: identificação das espécies de enterovírus isoladas / Laboratory surveillance of acute flaccid paralysis in Brazil, in the period 2007-2011: species identification of enteroviruses isolated

Os enterovírus humanos (Picornaviridae) são vírus de transmissão predominantemente entérica, possuem distribuição cosmopolita, estando entre os agentes mais prevalentes como causadores de patogenias em humanos. Atualmente, já foram descritos mais de 100 sorotipos para os enterovírus humanos e em grande parte dos casos as infecções associadas são assintomáticas. Surtos e casos esporádicos de enteroviroses são frequentemente notificados em diversas regiões do mundo causando conjuntivite hemorrágica aguda, meningite asséptica, doença de mão pé e boca e poliomielite. A poliomielite é uma doença infecciosa de caráter agudo, que pode assumir desde formas assintomáticas até formas paralíticas (paralisia flácida aguda ou PFA), causada em geral por um dos três sorotipos de poliovírus (PV). O PV selvagem está eliminado do Brasil desde 1989, atualmente sendo restrito a apenas quatro países (Nigéria, Afeganistão, Paquistão, Índia). Entretanto surtos de PFA associados à PV de origem vacinal e a enterovírus não pólio recombinantes tem sido notificados. A caracterização de EVNP é de extrema importância para a investigação da diversidade de vírus co-circulantes, e para relacionar os sintomas clínicos com o sorotipo viral envolvido, incluindo a investigação de vias de transmissão de enterovírus, durante a ocorrência de surtos, além de contribuir para estudos epidemiológicos e com a evolução de enterovírus. Neste estudo foram analisadas amostras relacionadas à PFAs, utilizando RT-PCR e PCR com o objetivo de identificar quais são as espécies de enterovírus humanos associadas. Os membros da espécie C foram sequenciados para a identificação de sorotipo. De um total de 190 amostras, 79 eram da espécie C, 78 da espécie B, 32 da espécie A e 1 amostra era correspondente as espécies A e C,não sendo encontradas amostras da espécie D. Entre as amostras da espécie C,58 correspondiam a PV.Os dados obtidos apresentam similaridades com estudos similares na Europa e Ásia, cobrindo um aspecto pouco observado na epidemiologia dos enterovírus em território brasileiro.

Foot and Mouth Disease : Etiology, Epidemiology and Control Measures

Foot and mouth disease (FMD) is highly infectious disease of cloven-hoofed animals, particularly cattle, sheep, pigs and goats. Also, it is the most important animal pathogen on the global scale because of the potential for rapid and extensive spread through susceptible animal populations. Outbreak can lead to formidable economic consequence for domestic livestock production and international trade. FMD is caused by FMD virus which is a small, non-enveloped, positive-sense RNA virus belonging to the genus Aphthovirus within the family Picornaviridae . There are seven immunologically distinct serotypes; O, A, C, SAT (Southern African Territories) 1, SAT 2, SAT 3 and Asia 1 and a diverse antigenic spectrum of virus strains within each serotype. Characteristic lesion of FMD is the formation of vesicles in the mucosal membranes of mouth, muzzle, foot, and teats. Nowadays, many developed countries have maintained FMD-free as a result of eradication efforts. However, outbreaks of FMD have occurred in several countries, even in Europe, and it is still endemic in Africa, the Middle East, Asia, and South America. Last year, three outbreaks of FMD occurred in our country. Last outbreak reported in November, 2010 induced the enormous social and economical impacts. Culling of infected animals, movement control, and vaccination are the major control measures of FMD. To control the disease, each country has their own strategies based on the current situation of FMD in their country. Therefore, I would like to discuss the causative agent, epidemiological properties and control measures of FMD in this paper.

[Comparative study on 1D [VP1] region of foot and mouth disease virus [type A strain] among different isolates: Khorasan Razavi isolate and other Iranian and neighboring countries isolates]

Foot-and-mouth disease [FMD] is one of the most important virus disease in farm animals. Types O, A and Asial FMD virus have been endemic in Iran. In this study, samples from suspected livestock were analyzed by RT-PCR experiment. The number of 702 nucleotides determined at 1D- 2B region of type A strain isolated from Khorasan Razavi province sequenced and compared with that of other reported isolates type A from Iran and neighboring countries. The results show that field isolated type A has about 89% similarity with other reported isolates type A from Iran and neighboring countries. Furthermore, this virus shows the most similarity with A/IRN/1/87[Samuel. Phylogenitic analysis revealed that virus was closely related to A22-Iraq/99 and A/IRN/iso/105 that rest in the same lineage. The data showed high similarity between type A viruses involved in the Khorasan Razavi province and A/IRN/87v [vaccine strain]; so that it can be concluded that the vaccine can produce prophylactic antibody against this virus

Recombinant DNA and Protein Vaccines for Foot-and-mouth Disease Induce Humoral and Cellular Immune Responses in Mice

BACKGROUND: Foot-and-mouth disease virus (FMDV) is a small single-stranded RNA virus which belongs to the family Picornaviridae, genus Apthovirus. It is a principal cause of FMD which is highly contagious in livestock. In a wild type virus infection, infected animals usually elicit antibodies against structural and non-structural protein of FMDV. A structural protein, VP1, is involved in neutralization of virus particle, and has both B and T cell epitopes. A RNA-dependent RNA polymerase, 3D, is highly conserved among other serotypes and strongly immunogenic, therefore, we selected VP1 and 3D as vaccine targets. METHODS: VP1 and 3D genes were codon-optimized to enhance protein expression level and cloned into mammalian expression vector. To produce recombinant protein, VP1 and 3D genes were also cloned into pET vector. The VP1 and 3D DNA or proteins were co-immunized into 5 weeks old BALB/C mice. RESULTS: Antigen-specific serum antibody (Ab) responses were detected by Ab ELISA. Cellular immune response against VP1 and 3D was confirmed by ELISpot assay. CONCLUSION: The results showed that all DNA- and protein-immunized groups induced cellular immune responses, suggesting that both DNA and recombinant protein vaccine administration efficiently induced Ag-specific humoral and cellular immune responses.

Hepatitis A Vaccine / 한양의대학술지

Hepatitis A virus (HAV) is a RNA virus classified within Picornaviridae family HAV vaccines containing formalin-inactivated virus produced in cell culture have been licensed in the early 1990s. HAV seroepidemiology in Korea is rapidly changing according to improvement of socioeconomical status. This epidemiologic shift is presented by growing numbers of hepatitis A cases among young adults, who are more symptomatic than children. Recently, about 70% of acute viral hepatitis in Korean adults are attributable to HAV. The seroprevalence of HAV among the young people in their teens and twenties is only about 10%, which suggests that a growing number of young adults are susceptible to HAV infection and development of more cases with severe presentation is expected in the near future. Therefore, active campaign for childhood HAV vaccination should be continued, and catch-up vaccination for the young population should be considered in Korea. Some preliminary data suggest the incidence rate of hepatitis A might be higher than 20/100,000 population, and routine childhood vaccination for HAV could be cost-effective in Korea. To provide reasonable recommendation for HAV vaccine, the continuous survey of HAV seroepidemiology and study on the cost-effectiveness of routine childhood vaccination are urgently warranted.

[Analysis of poly protein processing in human parechovirus Type 1 by gene cloning and expression]

Human parechovirus is a genus of picornaviridae. All of picornaviruses have a 3C protease which has a key role in virus protein processing and replication. The aim of this study was to analyse polyprotein processing in human Parechovirus type1 by cloning and expression of 3C gene. After preparation of cDNA from human parechovirus type 1[HPEV-1] RNA genome the region of cDNA that was encoded for 3C protein was inserted into plasmid pUBS by Ligase enzyme and recombinant plasmid was prepared. After transformation and replication of this plasmid in E.coli MC 10.22, DNA was isolated by phenol extraction and then expressed in prokaryotic [E.coli BL-21] and In vitro systems under T7 promoter. The results were detected by SDS-PAGE and analyzed. The products of expression of recombinant plasmids [with out 3C gene] in both prokaryotic and in vitro systems were analyzed. Just one large band same size as primary translation product was observed, but with plasmid including 3C gene, several small bands were detected. These results indicate that human Parechovirus type1 polyprotein processing occurs by 3C protease. 3C protease was checked by anti protease. Our results showed that HPEV-1 has a processing strategy different from other members of Picornaviruses, and 3C protein seems to be the only virus encoded protease that can catalyze cleavages of all sites in the Parechovirus type1 primary polyprotein

Triatoma patagonica (Hemiptera, Reduviidae), a New Host for Triatoma virus

Previous authors demonstrated that Triatoma virus (TrV) is able to infect several species of triatomines when injected with viral inoculum obtained from its original host, T. infestans. Both vertical (transovarian) and horizontal (faecal-oral) mechanisms of viral transmission were also described. In this paper we report the experimental TrV infection of a wild species from southern Argentina, T. patagonica. The inoculum consisted of clarified gut contents of infected T. infestans rubbed on the chicken skin whereupon T. patagonica individuals were fed. The results demonstrate that this is another potential host for the virus, and that the oral route is also effective for experimental interspecific infections