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1.
Nat Commun ; 11(1): 1297, 2020 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-32157083

RESUMO

Synthetic RNA-based gene circuits enable sophisticated gene regulation without the risk of insertional mutagenesis. While various RNA binding proteins have been used for translational repression in gene circuits, the direct translational activation of synthetic mRNAs has not been achieved. Here we develop Caliciviral VPg-based Translational activator (CaVT), which activates the translation of synthetic mRNAs without the canonical 5'-cap. The level of translation can be modulated by changing the locations, sequences, and modified nucleosides of CaVT-binding motifs in the target mRNAs, enabling the simultaneous translational activation and repression of different mRNAs with RNA-only delivery. We demonstrate the efficient regulation of apoptosis and genome editing by tuning translation levels with CaVT. In addition, we design programmable CaVT that responds to endogenous microRNAs or small molecules, achieving both cell-state-specific and conditional translational activation from synthetic mRNAs. CaVT will become an important tool in synthetic biology for both biological studies and future therapeutic applications.


Assuntos
Caliciviridae/metabolismo , Redes Reguladoras de Genes , Mamíferos/genética , Biossíntese de Proteínas , RNA/metabolismo , Proteínas Virais/metabolismo , Motivos de Aminoácidos , Animais , Apoptose/genética , Edição de Genes , Regulação da Expressão Gênica , Células HeLa , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Conformação de Ácido Nucleico , Multimerização Proteica , RNA/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Virais/química
2.
mSphere ; 4(5)2019 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-31533997

RESUMO

Enteric viruses in the Caliciviridae family cause acute gastroenteritis in humans and animals, but the cellular processes needed for virus replication and disease remain unknown. A common strategy among enteric viruses, including rotaviruses and enteroviruses, is to encode a viral ion channel (i.e., viroporin) that is targeted to the endoplasmic reticulum (ER) and disrupts host calcium (Ca2+) homeostasis. Previous reports have demonstrated genetic and functional similarities between the nonstructural proteins of caliciviruses and enteroviruses, including the calicivirus NS1-2 protein and the 2B viroporin of enteroviruses. However, it is unknown whether caliciviruses alter Ca2+ homeostasis for virus replication or whether the NS1-2 protein has viroporin activity like its enterovirus counterpart. To address these questions, we used Tulane virus (TV), a rhesus enteric calicivirus, to examine Ca2+ signaling during infection and determine whether NS1-2 has viroporin activity that disrupts Ca2+ homeostasis. We found that TV increases Ca2+ signaling during infection and that increased cytoplasmic Ca2+ levels are important for efficient replication. Further, TV NS1-2 localizes to the endoplasmic reticulum, the predominant intracellular Ca2+ store, and the NS2 region has characteristics of a viroporin domain (VPD). NS1-2 had viroporin activity in a classic bacterial functional assay and caused aberrant Ca2+ signaling when expressed in mammalian cells, but truncation of the VPD abrogated these activities. Together, our data provide new mechanistic insights into the function of the NS2 region of NS1-2 and support the premise that enteric viruses, including those within Caliciviridae, exploit host Ca2+ signaling to facilitate their replication.IMPORTANCE Tulane virus is one of many enteric caliciviruses that cause acute gastroenteritis and diarrheal disease. Globally, enteric caliciviruses affect both humans and animals and amass >65 billion dollars per year in treatment and health care-associated costs, thus imposing an enormous economic burden. Recent progress has resulted in several cultivation systems (B cells, enteroids, and zebrafish larvae) to study human noroviruses, but mechanistic insights into the viral factors and host pathways important for enteric calicivirus replication and infection are still largely lacking. Here, we used Tulane virus, a calicivirus that is biologically similar to human noroviruses and can be cultivated by conventional cell culture, to identify and functionally validate NS1-2 as an enteric calicivirus viroporin. Viroporin-mediated calcium signaling may be a broadly utilized pathway for enteric virus replication, and its existence within caliciviruses provides a novel approach to developing antivirals and comprehensive therapeutics for enteric calicivirus diarrheal disease outbreaks.


Assuntos
Sinalização do Cálcio , Caliciviridae/patogenicidade , Canais Iônicos/metabolismo , Proteínas não Estruturais Virais/metabolismo , Replicação Viral , Animais , Cálcio/metabolismo , Caliciviridae/química , Linhagem Celular , Homeostase , Interações entre Hospedeiro e Microrganismos , Canais Iônicos/genética , Macaca mulatta , Proteínas não Estruturais Virais/genética
3.
Vet Microbiol ; 236: 108388, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31500726

RESUMO

Neboviruses (NeVs) is an important causative agent of calf diarrhea. Here, 354 diarrhoeic samples were collected from yak on 55 farms in the Qinghai-Tibet Plateau, China. 22.0% of the diarrhoeic samples were detected as NeVs-positive by RT-PCR assay. Phylogenetic analysis of 78 NeVs RdRp fragments showed that 69 strains were closely related to NB-like strains, and the remaining 9 strains were clustered into an independent branch, which may represent a novel RdRp genotype. Two complete NeVs genomes (YAK/NRG-17/17/CH and YAK/HY1-2/18/CH) were successfully sequenced with 7459 nt and 7460 nt in length, respectively. The genomes of the two strains only shared 68.1%-69.3% nt identity with all six known NeVs genomes, and phylogenetic trees based on its genome, VP1, RdRp, VP2, P34, NTPase, P30, VPg and 3CLpro proteins suggested that the two strains may represent a novel NeVs strain with novel VP1 genotype and novel RdRp genotype. Notably, 11.5% NeVs strains were screened as the novel NeVs strains based VP1 and RdRp sequences. These novel NeVs strains were detected from 6 farms in two counties, indicating that the novel NeVs has spread in local region. To best of our knowledge, this is the first detection of NeVs in yak. Moreover, a novel NeVs strain was identified based on complete genome. These results contribute to further understand the prevalence and genetic evolution of NeVs.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/isolamento & purificação , Doenças dos Bovinos/virologia , Genoma Viral , Animais , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/virologia , Bovinos , China/epidemiologia , Diarreia/veterinária , Diarreia/virologia , Filogenia
4.
Zhonghua Liu Xing Bing Xue Za Zhi ; 40(8): 895-899, 2019 Aug 10.
Artigo em Chinês | MEDLINE | ID: mdl-31484250

RESUMO

Objective: To conduct a viral pathogen surveillance program on pediatric inpatients less than five years old with acute gastroenteritis in Shanghai and to better understand the pathogenic spectrum and molecular features in the target population, for setting up programs on control, prevention, medication and vaccine applications of the diseases. Methods: Fecal samples were collected from inpatients less than 5 years old who were admitted to a pediatric hospital for having acute gastroenteritis. Information related to demographic, clinical and epidemiological features of the patients was also collected. Laboratory assays including ELISA, real-time PCR and nested PCR, were performed to detect the presence of pathogens as rotavirus, calicivirus, astrovirus and adenovirus. Results: A total of 1 018 samples were collected (male 671 and 347 female), with the positive detection rate as 40.57% which peaked from autumn till winter, annually. Calicivirus and rotavirus A presented with the highest detection rates (24.75% and 13.95% respectively). The lowest detection rate was found in the 0-6 month-olds (32.20%). 65% of the patients with positive virus had received antibiotic treatment prior to the hospitalization. However, no statistically significant difference was seen, regarding the rates of antibiotic medication in the virus positive or negative populations (P>0.05). Data from the Rotavirus genotype analysis revealed that G9P[8] genotype was the predominant strain, and causing majority of rotavirus infections in all the age groups. Conclusions: Among the inpatients under 5 years of age in Shanghai, the positive detection rate for Calicivirus was higher than that for rotavirus group A, suggesting the necessity to carefully monitor the changes regarding the pathogenic spectrum and subtypes of the virus. Antibiotics should also be attentively administered, together with the development of suitable vaccine.


Assuntos
Caliciviridae/isolamento & purificação , Fezes/virologia , Gastroenterite/virologia , Pacientes Internados/estatística & dados numéricos , Infecções por Rotavirus/diagnóstico , Rotavirus/isolamento & purificação , Doença Aguda , Pré-Escolar , China/epidemiologia , Feminino , Gastroenterite/diagnóstico , Gastroenterite/epidemiologia , Humanos , Lactente , Masculino , Infecções por Rotavirus/epidemiologia , Infecções por Rotavirus/virologia
5.
Forensic Sci Med Pathol ; 15(3): 399-403, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31250258

RESUMO

Rapid identification of pathogenic agents is important in response to the emergence of biocrime and bioterrorism, to facilitate appropriate confinement and treatment. As the rapid determination system of viral genome sequences (RDV method) using exhaustive gene amplification is useful for rapid identification, we examined whether this method could be applied to forensic samples. To detect pathogenic virus in a cat with suspected viral infections, fluid swab samples were applied to the RDV method. The following steps were performed: viral propagation, extraction of the viral genome, amplification of the first library, fragmentation of the library, amplification of the second library using non-specific primer sets, and direct sequencing of the amplicon. To confirm the viruses detected by this method, we performed conventional PCR using virus-specific primers. We detected pathogenic virus genome sequences from the swab samples and confirmed infection with these viruses. In addition, we directly detected a viral genome sequence from the nasal swab sample without the viral propagation step. The RDV method is infrequently used in forensic analysis. This method is practicable with equipment existing in a normal laboratory and is useful for rapid detection and identification of pathogenic viruses in forensic samples. This method would also be applicable to the detection of bacteria and fungi.


Assuntos
Genoma Viral/genética , Cavidade Nasal/virologia , Faringe/virologia , Animais , Caliciviridae/genética , Gatos , DNA Viral , Herpesviridae/genética , Reação em Cadeia da Polimerase , RNA Viral , Manejo de Espécimes
6.
PLoS One ; 14(4): e0214227, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30969980

RESUMO

Bats are implicated as natural reservoirs for a wide range of zoonotic viruses including SARS and MERS coronaviruses, Ebola, Marburg, Nipah, Hendra, Rabies and other lyssaviruses. Accordingly, many One Health surveillance and viral discovery programs have focused on bats. In this report we present viral metagenomic data from bats collected in the Kingdom of Saudi Arabia [KSA]. Unbiased high throughput sequencing of fecal samples from 72 bat individuals comprising four species; lesser mouse-tailed bat (Rhinopoma hardwickii), Egyptian tomb bat (Taphozous perforatus), straw-colored fruit bat (Eidolon helvum), and Egyptian fruit bat (Rousettus aegyptiacus) revealed molecular evidence of a diverse set of viral families: Picornaviridae (hepatovirus, teschovirus, parechovirus), Reoviridae (rotavirus), Polyomaviridae (polyomavirus), Papillomaviridae (papillomavirus), Astroviridae (astrovirus), Caliciviridae (sapovirus), Coronaviridae (coronavirus), Adenoviridae (adenovirus), Paramyxoviridae (paramyxovirus), and unassigned mononegavirales (chuvirus). Additionally, we discovered a bastro-like virus (Middle East Hepe-Astrovirus), with a genomic organization similar to Hepeviridae. However, since it shared homology with Hepeviridae and Astroviridae at ORF1 and in ORF2, respectively, the newly discovered Hepe-Astrovirus may represent a phylogenetic bridge between Hepeviridae and Astroviridae.


Assuntos
Quirópteros/virologia , Metagenoma/genética , Metagenômica , Filogenia , Animais , Caliciviridae/genética , Caliciviridae/isolamento & purificação , Quirópteros/genética , Egito , Fezes/virologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mamíferos/virologia , Oriente Médio , Coronavírus da Síndrome Respiratória do Oriente Médio , Paramyxoviridae/genética , Paramyxoviridae/isolamento & purificação , Picornaviridae/genética , Picornaviridae/isolamento & purificação , Vírus de RNA/genética , Rotavirus/genética , Rotavirus/isolamento & purificação , Arábia Saudita
7.
Viruses ; 11(3)2019 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-30901945

RESUMO

Besides noroviruses, the Caliciviridae family comprises four other accepted genera: Sapovirus, Lagovirus, Vesivirus, and Nebovirus. There are six new genera proposed: Recovirus, Valovirus, Bavovirus, Nacovirus, Minovirus, and Salovirus. All Caliciviridae have closely related genome structures, but are genetically and antigenically highly diverse and infect a wide range of mammalian host species including humans. Recombination in nature is not infrequent for most of the Caliciviridae, contributing to their diversity. Sapovirus infections cause diarrhoea in pigs, humans and other mammalian hosts. Lagovirus infections cause systemic haemorrhagic disease in rabbits and hares, and vesivirus infections lead to lung disease in cats, vesicular disease in swine, and exanthema and diseases of the reproductive system in large sea mammals. Neboviruses are an enteric pathogen of cattle, differing from bovine norovirus. At present, only a few selected caliciviruses can be propagated in cell culture (permanent cell lines or enteroids), and for most of the cultivatable caliciviruses helper virus-free, plasmid only-based reverse genetics systems have been established. The replication cycles of the caliciviruses are similar as far as they have been explored: viruses interact with a multitude of cell surface attachment factors (glycans) and co-receptors (proteins) for adsorption and penetration, use cellular membranes for the formation of replication complexes and have developed mechanisms to circumvent innate immune responses. Vaccines have been developed against lagoviruses and vesiviruses, and are under development against human noroviruses.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/classificação , Animais , Caliciviridae/patogenicidade , Infecções por Caliciviridae/virologia , Genoma Viral , Humanos , Norovirus , Filogenia , Análise de Sequência de DNA
8.
Arch Virol ; 164(5): 1411-1417, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30850861

RESUMO

To investigate the molecular epidemiology and genetic diversity of bovine enteric caliciviruses, a total of 167 fecal samples from diarrheic calves were screened. Bovine noroviruses (BoNoVs) and neboviruses were detected in 56 (33.5%) and 37 (22.1%) fecal samples, respectively. Sequences of the RdRp and capsid gene of selected BoNoVs showed that the GIII.1 and GIII.2 genotypes were in circulation in Turkey. Two of the BoNoV strains were identified as recombinant strains (GIII.P1/GIII.2). All examined neboviruses possessed a Nebraska-like RdRp gene. The two nebovirus strains were classified into lineage 4 based on phylogenetic analysis of VP1 amino acid sequences. One of them showed evidence of a recombination event within the S domain. This study is thus the first to reveal the presence of the BoNoV GIII.1 genotype and recombinant strains of BoNoV and neboviruses in Turkey.


Assuntos
Infecções por Caliciviridae/epidemiologia , Caliciviridae/genética , Proteínas do Capsídeo/genética , Gastroenterite/veterinária , Norovirus/genética , Vírus Reordenados/genética , Sequência de Aminoácidos , Animais , Caliciviridae/isolamento & purificação , Infecções por Caliciviridae/veterinária , Bovinos , Gastroenterite/virologia , Norovirus/isolamento & purificação , Turquia/epidemiologia
9.
J Gen Virol ; 100(1): 35-45, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30444471

RESUMO

In this study, 98 diarrhoeic and 70 non-diarrhoeic samples were collected from 13 dairy farms located across 5 provinces in China from April 2017 to May 2018. Approximately 41.8 % (41/98) of diarrhoeic samples and 5.7 % (4/70) of non-diarrhoeic samples were nebovirus-positive based on RT-PCR results, and some diarrhoeic samples were co-infected with bovine rotavirus (73.2 %), bovine coronavirus (36.6 %) and/or bovine viral diarrhoea virus (31.7 %). A phylogenetic analysis of 23 nebovirus RdRp fragments showed that these strains were closely related to Nebraska-like (NB-like) strains but were all located in a unique large branch. Moreover, a phylogenetic analysis of the 18 complete VP1 sequences from this study revealed that 14 strains belonged to lineage 1 and 4 strains belonged to lineage 3. Notably, all four lineage 3 strains shared the same recombination event, with a breakpoint located within the P1A domain. The complete genome of one nebovirus strain, Bo/YLA-2/17/CH, which had a recombination event within the P1A domain of its VP1, was successfully sequenced and was found to be 7453 nt in length, and this may represent a novel nebovirus strain based on the phylogenetic analysis of its complete genome sequence. In conclusion, this study reveals that neboviruses circulate widely in dairy cows in China and exhibit a unique evolution of RdRp. To the best of our knowledge, this is the first reported recombination event located within the P1A domain of nebovirus VP1.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/isolamento & purificação , Portador Sadio/veterinária , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/virologia , Diarreia/veterinária , Animais , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/virologia , Portador Sadio/epidemiologia , Portador Sadio/virologia , Bovinos , China/epidemiologia , Diarreia/epidemiologia , Diarreia/virologia , Fezes/virologia , Genótipo , Epidemiologia Molecular , Filogenia , RNA Viral/genética , Recombinação Genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Proteínas Virais/genética
10.
PLoS One ; 13(10): e0205890, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30335814

RESUMO

Pre-weaning diarrhea (PWD) in mink kits is a common multifactorial syndrome on commercial mink farms. Several potential pathogens such as astroviruses, caliciviruses, Escherichia coli and Staphylococcus delphini have been studied, but the etiology of the syndrome seems complex. In pooled samples from 38 diarrheic and 42 non-diarrheic litters, each comprising of intestinal contents from 2-3 mink kits from the same litter, the bacterial populations were studied using Illumina Next Generation Sequencing technology and targeted 16S amplicon sequencing. In addition, we used deep sequencing to determine and compare the viral intestinal content in 31 healthy non-diarrheic and 30 diarrheic pooled samples (2-3 mink kits from the same litter per pool). The results showed high variations in composition of the bacterial species between the pools. Enterococci, staphylococci and streptococci dominated in both diarrheic and non-diarrheic pools. However, enterococci accounted for 70% of the reads in the diarrheic group compared to 50% in the non-diarrheic group and this increase was at the expense of staphylococci and streptococci which together accounted for 45% and 17% of the reads in the non-diarrheic and diarrheic group, respectively. Moreover, in the diarrheic pools there were more reads assigned to Clostridia, Escherichia-Shigella and Enterobacter compared to the non-diarrheic pools. The taxonomically categorized sequences from the virome showed that the most prevalent viruses in all pools were caliciviruses and mamastroviruses (almost exclusively type 10). However, the numbers of reads assigned to caliciviruses were almost 3 times higher in the diarrheic pools compared the non-diarrheic pools and Sapporo-like caliciviruses were more abundant than the Norwalk-like caliciviruses. The results from this study have contributed to the insight into the changes in the intestinal microbiota associated with the PWD syndrome of mink.


Assuntos
Diarreia/veterinária , Microbioma Gastrointestinal/genética , Intestinos/microbiologia , Mustelidae/microbiologia , RNA Ribossômico 16S/genética , Criação de Animais Domésticos , Animais , Astroviridae/classificação , Astroviridae/genética , Astroviridae/isolamento & purificação , Caliciviridae/classificação , Caliciviridae/genética , Caliciviridae/isolamento & purificação , Clostridiaceae/classificação , Clostridiaceae/genética , Clostridiaceae/isolamento & purificação , Diarreia/microbiologia , Diarreia/virologia , Enterobacteriaceae/classificação , Enterobacteriaceae/genética , Enterobacteriaceae/isolamento & purificação , Enterococcaceae/classificação , Enterococcaceae/genética , Enterococcaceae/isolamento & purificação , Fezes/microbiologia , Fezes/virologia , Sequenciamento de Nucleotídeos em Larga Escala , Intestinos/virologia , Mustelidae/virologia , Filogenia , Staphylococcaceae/classificação , Staphylococcaceae/genética , Staphylococcaceae/isolamento & purificação , Streptococcaceae/classificação , Streptococcaceae/genética , Streptococcaceae/isolamento & purificação , Síndrome , Desmame
11.
Arch Virol ; 163(10): 2921-2924, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29978262

RESUMO

We report the full genome sequence of the non-pathogenic rabbit lagovirus Lagovirus europaeus/GI.3/O cun/FR/2006/06-11 (GI.3/06-11), collected from a healthy French domestic rabbit in 2006, and initially described as 06-11 strain. The sequence reveals a genomic organization similar to lagoviruses. It was 7,436 bases long and contained two open reading frames (ORF). A dipeptide variation at the potential p23/2C-like helicase cleavage site (EE instead of ED) was observed, a feature only shared with non-recombinant pathogenic lagoviruses in GI.2 and with two European brown hare syndrome viruses (EBHSV) collected in 1982 in Sweden. GI.3/06-11 has only one initiation codon at the beginning of the ORF2 like the avirulent Italian rabbit calicivirus (RCV) and EBHSV. Previous genetic analyses based on the capsid gene sequences showed that GI.3/06-11 was closer to the RCV and pathogenic lagoviruses GI.1 strains than other lagoviruses. This study, by revealing that GI.3/06-11 genome sequence significantly clustered with pathogenic GI.2 strains, gives prominence of new genetic relationship among lagoviruses and should contribute to understand the emergence of pathogenic strains.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/isolamento & purificação , Genoma Viral , Animais , Caliciviridae/classificação , Caliciviridae/genética , Infecções por Caliciviridae/virologia , Proteínas do Capsídeo/genética , Europa (Continente) , Fases de Leitura Aberta , Filogenia , Coelhos
12.
J Pediatr Gastroenterol Nutr ; 67(6): e117-e122, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-29985875

RESUMO

BACKGROUND AND OBJECTIVE: Norovirus (NoV) infections are known to have high-morbidity and mortality rates and are a major health problem globally. The impact of NoV on child development is, however, poorly understood. We evaluated the distribution of NoV genotypes in children from a low-income Brazilian semiarid region, in relation with their clinical symptoms, nutritional status, and co-pathogens. METHODS: The test population included children aged 2 to 36 months from 6 cities of the Brazilian semiarid region. Fecal samples were collected from each child, along with the information regarding their socioeconomic/clinical conditions using a standardized questionnaire. Detection and quantification of NoV were performed by reverse-transcription quantitative polymerase chain reaction, followed by molecular and phylogenetic analyses. RESULTS: The NoV detection rate was 45.2%. Presence of NoV was associated with lower z scores for weight-for-age (P = 0.03), weight-for-height (P = 0.03), and body mass index-for-age (P = 0.03). NoV infection was associated with more frequent respiratory illnesses (P < 0.01). GII.P7 (polymerase) and GII.3 (capsid) were the most frequent NoV genotypes. Analysis of the open reading frame (ORF)1-2 junction identified recombinant NoV strains in 80% of the sequenced samples. Enteroaggregative Escherichia coli coinfection was the major predictor for diarrhea in NoV-positive samples (P < 0.02). Moreover, Shigella spp was also associated with NoV-positive diagnosis (P = 0.02). CONCLUSIONS: This study highlights the genetic variability of NoV and, associated co-infections and undernutrition in infants from low-income Brazilian semiarid region.


Assuntos
Infecções por Caliciviridae/virologia , Caliciviridae/genética , Transtornos da Nutrição Infantil/virologia , Coinfecção/microbiologia , Variação Genética , Estatura , Índice de Massa Corporal , Peso Corporal , Brasil/epidemiologia , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/fisiopatologia , Proteínas do Capsídeo/análise , Transtornos da Nutrição Infantil/epidemiologia , Pré-Escolar , Coinfecção/epidemiologia , Diarreia/virologia , Escherichia coli , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/virologia , Fezes/virologia , Feminino , Genótipo , Humanos , Lactente , Masculino , Estado Nutricional , Filogenia , Reação em Cadeia da Polimerase em Tempo Real , Shigella , Fatores Socioeconômicos
13.
Virol J ; 15(1): 95, 2018 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-29801460

RESUMO

BACKGROUND: Emergence and re-emergence of porcine epidemic diarrhea virus (PEDV) in North America, Asia and Europe has caused severe economic loss to the global swine industry. However, the virome of PEDV infected pigs and its effect on disease severity remains unknown. The advancements of sequencing technology have made it possible to characterize the entire microbiome of different body sites for any host. METHODS: The objective of this study was to characterize the RNA virome in PEDV-positive pigs using the hypothesis-free metagenomics approach based on next-generation sequencing. Specifically, 217 PEDV-positive swine fecal swab samples collected from diarrheic piglets over 17 US states during 2015-2016 were analyzed. RESULTS: A Kraken algorithm-based bioinformatics analysis revealed the presence of up to 9 different RNA genera besides PEDV (Alphacoronavirus genus), including Mamastrovirus (52%, 113/217), Enterovirus (39%, 85/217), Sapelovirus (31%, 67/217), Posavirus (30%, 66/217), Kobuvirus (23%, 49/217), Sapovirus (13%, 28/217), Teschovirus (10%, 22/217), Pasivirus (9%, 20/217), and Deltacoronavirus (3%, 6/217). There were 58 out of 217 piglets (27%) have PEDV infection alone whereas the remaining 159 (73%) shed 2 up to 9 different viruses. CONCLUSION: These findings demonstrated that PEDV infected diarrheic pigs had an extensive RNA viral flora consisting of four different families: Astroviridae, Picornaviridae, Caliciviridae, and Coronaviridae.


Assuntos
Astroviridae/genética , Caliciviridae/genética , Coronaviridae/genética , Infecções por Coronavirus/veterinária , Picornaviridae/genética , Vírus da Diarreia Epidêmica Suína/genética , Doenças dos Suínos/epidemiologia , Algoritmos , Sequência de Aminoácidos , Animais , Astroviridae/classificação , Astroviridae/isolamento & purificação , Caliciviridae/classificação , Caliciviridae/isolamento & purificação , Coinfecção , Biologia Computacional , Coronaviridae/classificação , Coronaviridae/isolamento & purificação , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Fazendas , Fezes/virologia , Sequenciamento de Nucleotídeos em Larga Escala , Metagenômica/métodos , Filogenia , Picornaviridae/classificação , Picornaviridae/isolamento & purificação , Vírus da Diarreia Epidêmica Suína/classificação , Vírus da Diarreia Epidêmica Suína/isolamento & purificação , RNA Viral/genética , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Suínos , Doenças dos Suínos/virologia , Estados Unidos/epidemiologia
14.
mBio ; 9(3)2018 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-29789360

RESUMO

Emerging zoonotic viral diseases remain a challenge to global public health. Recent surveillance studies have implicated bats as potential reservoirs for a number of viral pathogens, including coronaviruses and Ebola viruses. Caliciviridae represent a major viral family contributing to emerging diseases in both human and animal populations and have been recently identified in bats. In this study, we blended metagenomics, phylogenetics, homology modeling, and in vitro assays to characterize two novel bat calicivirus (BtCalV) capsid sequences, corresponding to strain BtCalV/A10/USA/2009, identified in Perimyotis subflavus near Little Orleans, MD, and bat norovirus. We observed that bat norovirus formed virus-like particles and had epitopes and receptor-binding patterns similar to those of human noroviruses. To determine whether these observations stretch across multiple bat caliciviruses, we characterized a novel bat calicivirus, BtCalV/A10/USA/2009. Phylogenetic analysis revealed that BtCalV/A10/USA/2009 likely represents a novel Caliciviridae genus and is most closely related to "recoviruses." Homology modeling revealed that the capsid sequences of BtCalV/A10/USA/2009 and bat norovirus resembled human norovirus capsid sequences and retained host ligand binding within the receptor-binding domains similar to that seen with human noroviruses. Both caliciviruses bound histo-blood group antigens in patterns that overlapped those seen with human and animal noroviruses. Taken together, our results indicate the potential for bat caliciviruses to bind histo-blood group antigens and overcome a significant barrier to cross-species transmission. Additionally, we have shown that bat norovirus maintains antigenic epitopes similar to those seen with human noroviruses, providing further evidence of evolutionary descent. Our results reiterate the importance of surveillance of wild-animal populations, especially of bats, for novel viral pathogens.IMPORTANCE Caliciviruses are rapidly evolving viruses that cause pandemic outbreaks associated with significant morbidity and mortality globally. The animal reservoirs for human caliciviruses are unknown; bats represent critical reservoir species for several emerging and zoonotic diseases. Recent reports have identified several bat caliciviruses but have not characterized biological functions associated with disease risk, including their potential emergence in other mammalian populations. In this report, we identified a novel bat calicivirus that is most closely related to nonhuman primate caliciviruses. Using this new bat calicivirus and a second norovirus-like bat calicivirus capsid gene sequence, we generated virus-like particles that have host carbohydrate ligand binding patterns similar to those of human and animal noroviruses and that share antigens with human noroviruses. The similarities to human noroviruses with respect to binding patterns and antigenic epitopes illustrate the potential for bat caliciviruses to emerge in other species and the importance of pathogen surveillance in wild-animal populations.


Assuntos
Antígenos Virais/imunologia , Antígenos de Grupos Sanguíneos/imunologia , Caliciviridae/imunologia , Norovirus/imunologia , Animais , Antígenos Virais/química , Antígenos Virais/genética , Antígenos de Grupos Sanguíneos/química , Antígenos de Grupos Sanguíneos/genética , Caliciviridae/química , Caliciviridae/classificação , Caliciviridae/genética , Infecções por Caliciviridae/virologia , Proteínas do Capsídeo/química , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/imunologia , Quirópteros/virologia , Humanos , Norovirus/química , Norovirus/classificação , Norovirus/genética , Filogenia , Domínios Proteicos
15.
Virus Res ; 251: 6-13, 2018 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-29709508

RESUMO

Nebovirus is a new genus within the family Caliciviridae and is a causative agent of calf diarrhea. The limited nebovirus genomic sequences that are currently available has hampered understanding of nebovirus genetic evolution. The aim of the present study was to determine the genomic characterization of strain Bo/LZB-1/17/CH, which was previously identified as being similar to the novel genotype strain Bo/DijonA216/06/FR based on partial capsid sequences. Our results show that the complete RNA genome of strain Bo/LZB-1/17/CH is 7453 nucleotides (nt) in length and shares 79.0%-83.5% nt identity with all available nebovirus genomes in the GenBank database. A phylogenetic analysis based on its complete genome sequence revealed that strain Bo/LZB-1/17/CH clustered into an independent branch. Two interesting characteristics were observed in the genome of strain Bo/LZB-1/17/CH. First, the major capsid protein (VP1) of strain Bo/LZB-1/17/CH shares 96.6% amino acid (aa) identity with strain Bo/DijonA216/06/FR but shares only 75.2%-76.8% aa identity with other nebovirus strains and has an even lower identity in the P2 domain (61.1%-65% aa identity). Second, the RNA-dependent RNA polymerase (RdRp) of strain Bo/LZB-1/17/CH is more closely related to NB-like strains than it is to strain Bo/DijonA216/06/FR, and a recombination event was identified within the 3' end of the RdRp in strain Bo/LZB-1/17/CH. In conclusion, the results in this study indicate that strain Bo/LZB-1/17/CH may represent a novel nebovirus strain. To the best of our knowledge, this is the first description of a recombinant event in nebovirus RdRp.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/isolamento & purificação , Doenças dos Bovinos/virologia , Genoma Viral , Genótipo , RNA Replicase/genética , Proteínas Estruturais Virais/genética , Animais , Caliciviridae/classificação , Caliciviridae/genética , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/virologia , Bovinos , Doenças dos Bovinos/epidemiologia , Análise por Conglomerados , Filogenia , RNA Viral/genética , Homologia de Sequência
16.
J Virol ; 92(9)2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29467317

RESUMO

Some viruses within the Caliciviridae family initiate their replication cycle by attachment to cell surface carbohydrate moieties, histo-blood group antigens (HBGAs), and/or terminal sialic acids (SAs). Although bovine nebovirus (BNeV), one of the enteric caliciviruses, is an important causative agent of acute gastroenteritis in cattle, its attachment factors and possibly other cellular receptors remain unknown. Using a comprehensive series of protein-ligand biochemical assays, we sought to determine whether BNeV recognizes cell surface HBGAs and/or SAs as attachment factors. It was found that BNeV virus-like particles (VLPs) bound to A type/H type 2/Ley HBGAs expressed in the bovine digestive tract and are related to HBGAs expressed in humans and other host species, suggesting a wide spectrum of HBGA recognition by BNeV. BNeV VLPs also bound to a large variety of different bovine and human saliva samples of all ABH and Lewis types, supporting previously obtained results and suggesting a zoonotic potential of BNeV transmission. Removal of α1,2-linked fucose and α1,3/4-linked fucose epitopes of target HBGAs by confirmation-specific enzymes reduced the binding of BNeV VLPs to synthetic HBGAs, bovine and human saliva, cultured cell lines, and bovine small intestine mucosa, further supporting a wide HBGA binding spectrum of BNeV through recognition of α1,2-linked fucose and α1,3/4-linked fucose epitopes of targeted HBGAs. However, removal of terminal α2,3- and α2,6-linked SAs by their specific enzyme had no inhibitory effects on binding of BNeV VLPs, indicating that BNeV does not use terminal SAs as attachment factors. Further details of the binding specificity of BNeV remain to be explored.IMPORTANCE Enteric caliciviruses such as noroviruses, sapoviruses, and recoviruses are the most important etiological agents of severe acute gastroenteritis in humans and many other mammalian host species. They initiate infection by attachment to cell surface carbohydrate moieties, HBGAs, and/or terminal SAs. However, the attachment factor(s) for BNeV, a recently classified enteric calicivirus genus/type species, remains unexplored. Here, we demonstrate that BNeV VLPs have a wide spectrum of binding to synthetic HBGAs, bovine and human saliva samples, and bovine duodenal sections. We further discovered that α1,2-linked fucose and α1,3/4-linked fucose epitopes are essential for binding of BNeV VLPs. However, BNeV VLPs do not bind to terminal SAs on cell carbohydrates. Continued investigation regarding the proteinaceous receptor(s) will be necessary for better understanding of the tropism, pathogenesis, and host range of this important viral genus.


Assuntos
Caliciviridae/metabolismo , Epitopos/metabolismo , Fucose/metabolismo , Mucosa Intestinal/virologia , Receptores Virais/metabolismo , Ligação Viral , Animais , Antígenos de Grupos Sanguíneos/metabolismo , Células CHO , Células CACO-2 , Infecções por Caliciviridae/patologia , Infecções por Caliciviridae/virologia , Gatos , Linhagem Celular Tumoral , Cricetulus , Cães , Gastroenterite/patologia , Gastroenterite/veterinária , Gastroenterite/virologia , Células HeLa , Humanos , Células Madin Darby de Rim Canino , Ligação Proteica , Saliva/química , Ácidos Siálicos/metabolismo , Suínos
17.
Vet J ; 232: 57-64, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29428093

RESUMO

A number of novel viruses have been associated with canine gastroenteritis in recent years, from viral families as diverse as Caliciviridae and Picornaviridae to Parvoviridae and Circoviridae. The ability of many of these viruses to cause disease is uncertain, but epidemiological studies are continually adding to our knowledge of these potential pathogens. This review presents a summary of the latest research and current understanding of novel viruses associated with canine gastroenteritis.


Assuntos
Doenças do Cão/virologia , Gastroenterite/veterinária , Animais , Caliciviridae/isolamento & purificação , Circoviridae/isolamento & purificação , Coronaviridae/isolamento & purificação , Cães , Fezes/virologia , Gastroenterite/epidemiologia , Gastroenterite/virologia , Parvoviridae/isolamento & purificação , Picornaviridae/isolamento & purificação , Rotavirus/isolamento & purificação , Viroses/epidemiologia , Viroses/veterinária , Viroses/virologia
18.
Arch Virol ; 163(5): 1271-1277, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29362930

RESUMO

In this study, bovine enteric caliciviruses (BECs) were detected in 49.4% of a total of 253 stool specimens for diarrheic calves collected from 42 industrial dairy farms from March 2010 to February 2012. Genogroup III norovirus (NoVsGIII) were more prevalent (39.5%) than neboviruses (NBs) (15%), and coinfections were observed in 5.1% of the samples tested. Sequence analysis of the partial polymerase gene from 13 NoVsGIII samples indicated the circulation of both genotype 1 and genotype 2 strains. Among the six NB strains sequenced, five were related to the Bo/Nebraska/80/US strain, while one was related to the Bo/Newbury1/76/UK strain.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/isolamento & purificação , Doenças dos Bovinos/epidemiologia , Norovirus/isolamento & purificação , Animais , Animais Recém-Nascidos/virologia , Caliciviridae/genética , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/virologia , Bovinos , Doenças dos Bovinos/virologia , Indústria de Laticínios , Diarreia/veterinária , Diarreia/virologia , Fezes/virologia , Gastroenterite/epidemiologia , Gastroenterite/veterinária , Gastroenterite/virologia , Variação Genética , Genótipo , Irã (Geográfico)/epidemiologia , Norovirus/genética , Filogenia , Prevalência , Análise de Sequência de DNA
19.
Arch Virol ; 163(2): 475-478, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29058151

RESUMO

Neboviruses and genogroup III noroviruses (NoVsGIII) are causative agents of calf diarrhea. The purpose of this study was to investigate the presence of neboviruses and noroviruses in cattle in China. Twenty-eight diarrhea fecal samples collected from 5 different farms were analyzed by RT-PCR. The results showed that 3 nebovirus positive samples were detected on 2 farms, with two strains being related to Bo/DijonA216/06/FR strain and the other one clustering with NB-like strains. Meanwhile, 3 norovirus positive samples were detected on 3 farms, all of which belonged to genotype 1. Our results confirmed the presence of neboviruses and NoVsGIII in China for the first time, and supported the presence of a novel "DijonA216-like" nebovirus genotype.


Assuntos
Infecções por Caliciviridae/veterinária , Caliciviridae/isolamento & purificação , Doenças dos Bovinos/virologia , Diarreia/veterinária , Norovirus/isolamento & purificação , Animais , Caliciviridae/classificação , Caliciviridae/genética , Infecções por Caliciviridae/virologia , Bovinos , China , Diarreia/virologia , Fezes/virologia , Genótipo , Norovirus/classificação , Norovirus/genética , Filogenia
20.
Food Microbiol ; 69: 25-32, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28941906

RESUMO

Human norovirus (NoV) is a leading cause of fresh produce associated outbreaks. Previous research indicates that the roots of growing leafy greens and berries internalize human NoV. However the effect of plant type and inoculum level on internalization rates has not been directly compared. In this study we compared the internalization and dissemination rates of human NoV and its surrogate, Tulane virus (TV) in green onion, radishes, and Romaine lettuce. We also evaluated the effect inoculum level and plant growth matrix on the rate of viral internalization. In the hydroponic growth system, we detected internalization and dissemination of human NoV RNA in green onions. In hydroponically growing green onions inoculated with high titer TV, we found higher rates of internalization and dissemination compared to green onions inoculated with low titer TV. In soil growth systems, no infectious TV was detected in either green onion or radishes. However, in Romaine lettuce plants grown in soil approximately 4 log10 PFU/g was recovered from all tissues on day 14 p.i. Overall, we found that the type of plant, growth matrix, and the inoculum level influences the internalization and dissemination of human NoV and TV.


Assuntos
Caliciviridae/fisiologia , Contaminação de Alimentos/análise , Alface/virologia , Norovirus/fisiologia , Cebolas/virologia , Raphanus/virologia , Verduras/virologia , Internalização do Vírus , Caliciviridae/genética , Caliciviridae/isolamento & purificação , Humanos , Alface/crescimento & desenvolvimento , Norovirus/genética , Norovirus/isolamento & purificação , Cebolas/crescimento & desenvolvimento , Raphanus/crescimento & desenvolvimento , Microbiologia do Solo , Verduras/crescimento & desenvolvimento
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