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The emergence of the circular replication-associated protein (Rep) encoding single-stranded (CRESS) DNA viruses in different hosts has been associated with serious diseases, such as porcine diarrhea. The prevalence and pathogenicity of porcine circovirus-like virus (Po-Circo-like virus (PCLV)), a member of CRESS DNA virus, has not been fully illustrated. In order to understand the frequency of PCLV in pigs with respiratory disease, 519 healthy tissues (268 lungs, 201 lymph nodes and 50 hearts) and 380 tissues (212 lungs, 124 lymph nodes and 44 hearts) diagnosed with respiratory disease were collected for analyzing the prevalence of the PCLV infection using the Tag-Man qPCR assay. In addition, the complete genome of 43 PCLV strains were then sequenced, which were subsequently to analyze their characteristics. We found that 31.7 % (285/899) samples were tested positive for PCL virus. It is interesting to note that just 9.6 % (50/519) of the healthy samples were tested positive for PCLV, 61.8 % (235/380) of the diseased samples were PCLV positive. Analysis of the full genome of 43 PCLV strains showed that the genome of 42 PCLV strains included two distinct stem-loop structures, but the genome of PCLV FJ5-2020 strain contained no stem-loop structures. A phylogenetic tree analysis based on the Rep protein, PCLV could be classified into four genotypes: PCLVa, PCLVb, PCLVc, and PCLVd. In conclusion, this is the first report that the high frequency of PCLV in association in respiratory diseased pigs. PCLV strains were divided into four new genotypes of PCLV.
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Infecções por Circoviridae , Circovirus , Doenças dos Suínos , Suínos , Animais , Filogenia , Infecções por Circoviridae/veterinária , Vírus de DNARESUMO
Enterocytozoon bieneusi is an important pathogen that is responsible for over 90% of documented cases of human microsporidiosis worldwide, causing a threat to public health and husbandry development. In immunocompromised patients, it can cause persistent diarrhoea, wasting diathesis and malabsorption and developing life-threatening chronic diarrhoea. However, there was little information on the prevalence and multilocus genotypes of E. bieneusi in diarrheic pigs in three provinces of southern China. In this study, 1254 faecal samples of diarrheic pigs were collected from 37 pig farms in Hunan, Jiangxi, and Fujian provinces in southern China, and were investigated the prevalence and genotypes of E. bieneusi by nested polymerase chain reaction (PCR) based on the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA gene. The overall prevalence of E. bieneusi was 5.7% (72/1254) in three provinces. Furthermore, the difference was statistically significant (p < 0.001) in the prevalence of E. bieneusi in age groups. ITS sequence analysis revealed that 13 E. bieneusi genotypes were identified, including 8 known genotypes (EbpC, n = 30; Henan-IV, n = 21; CH5, n = 6; EbpA, n = 3; KIN-1, n = 2; O, n = 1; GX3, n = 1; CHS5, n = 1) and 5 novel genotypes (JX1, n = 2; JX2, n = 1; JX3, n = 2; FJ1, n = 1; FJ2, n = 1), and the genotype EbpC was the preponderant genotype. Phylogenetic analysis indicated that all genotypes of E. bieneusi were clustered as the zoonotic group 1. Moreover, a high genetic diversity of E. bieneusi were identified in this study, which the 64, 57, 52 and 64 samples were identified by multilocus sequence typing (MLST) at MS1, MS3, MS4 and MS7 loci, respectively. Then, 45 samples were successfully amplified and sequenced at four loci, forming 41 distinct multilocus genotypes (MLGs). These findings suggest that diarrheic pigs may potentially threaten to transmit E. bieneusi to humans, revealing E. bieneusi genetic variability in diarrheic pigs in three provinces of southern China.
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Enterocytozoon , Animais , Humanos , Suínos , Tipagem de Sequências Multilocus/veterinária , Enterocytozoon/genética , Filogenia , China/epidemiologia , DNA Espaçador Ribossômico/genética , Genótipo , Prevalência , Fezes , Variação Genética , Diarreia/epidemiologia , Diarreia/veterináriaRESUMO
Entamoeba spp. is a common zoonotic intestinal protozoan that can parasitize most vertebrates, including humans and pigs, causing severe intestinal diseases and posing a serious threat to public health. However, the available data on Entamoeba spp. infection in pigs are relatively limited in China. To characterize the infection of Entamoeba spp. within pigs in southern China, 1254 fecal samples of diarrheic pigs were collected from 37 intensive pig farms in Hunan, Jiangxi and Fujian provinces and the infection of Entamoeba spp. was investigated based on the small subunit rRNA (SSU rRNA) gene. The overall infection rate of Entamoeba spp. was 58.4% (732/1254), including 38.4% (118/307) in suckling piglets, 51.2% (153/299) in weaned piglets, 57.9% (55/95) in fattening pigs and 73.4% (406/553) in sows, respectively. Moreover, age and the sampling cities in Jiangxi and Fujian provinces were found to be the key factors influencing the infection of Entamoeba spp. (p < 0.05). Two subtypes (ST1 and ST3) with a zoonotic potential of Entamoeba polecki and Entamoeba suis were detected in all age groups of pigs and all sampling areas, with the predominant species and predominant subtype being E. polecki (91.3%, 668/732) and E. polecki ST1 (573/668), respectively, and E. polecki ST1 + E. polecki ST3 (78.6%, 239/304) being the most frequently detected form of mixed infection. Severe Entamoeba spp. infection and zoonotic subtypes were found in this study, exposing a large public health problem in the study area, and strategies need to be implemented to eliminate the risk in the future.
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Cryptosporidium spp. is recognized as an opportunistic zoonotic parasite that infects humans, wild and domestic animals, and is also a major cause of diarrhea-related disease in immunocompromised individuals, considered a global public health concern. Pig is considered as one of the reservoir hosts of Cryptosporidium spp. can transmit cryptosporidiosis to humans and other animals. However, limited studies on the distribution of Cryptosporidium spp. in diarrheic pigs have been published. Objective of the current study was to investigate the infection and species/genotypes of Cryptosporidium spp. from feces of diarrheic pigs in southern China. A total of 1254 fresh fecal samples were collected from 37 intensive pig farms in Jiangxi, Hunan and Fujian provinces, and were screened for Cryptosporidium spp. infection using a nested PCR assay targeted the small subunit rRNA (SSU rRNA) genes. The overall infection rate of Cryptosporidium spp. was 4.5% (57/1254), including 5.5% (17/307) in suckling piglets, 2.7% (8/299) in weaned piglets, 7.4% (7/95) in fattening pigs and 4.5% (25/553) in sows, respectively. In addition, two human-pathogenic species Cryptosporidium scrofarum (80.7%, 46/57) and Cryptosporidium suis (19.3%, 11/57) were identified. C. scrofarum and C. suis were observed in pigs tested in all age groups. Interestingly, a high colonization incidence of C. scrofarum (16/57) was observed in suckling piglets. This study revealed the prevalence and species of Cryptosporidium spp. in diarrheic pigs in three provinces of southern China, which suggested that diarrhea maybe not a direct factor affecting the prevalence of Cryptosporidium spp. in pigs. More prevention and control of this parasite in pigs should receive greater attention from farmers in investigated provinces.
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Criptosporidiose , Cryptosporidium , Doenças dos Suínos , Animais , China/epidemiologia , Criptosporidiose/epidemiologia , Criptosporidiose/parasitologia , Cryptosporidium/genética , Diarreia/epidemiologia , Diarreia/veterinária , Fezes/parasitologia , Feminino , Genótipo , Prevalência , Suínos , Doenças dos Suínos/epidemiologiaRESUMO
BACKGROUND: The benefits of probiotics being used in animals are well-documented via evidenced growth performance improvement and positive modulations of gut microbiota (GM). Thus, a combination of effective microorganisms (EM) has been frequently used in animal production, including broilers. However, there are only very limited reports of EM on the growth performance and the modulation in GM of partridge shank broiler chicks. METHODS: We attempted to evaluate the effects of a basal diet with the addition of an EM mixture on the growth performance and gut microbiome of the chicks. A total of 100 ten-day-old female partridge shank broiler chicks were randomly divided into two groups of 50 chicks each, of which, one group fed with EM supplementation in the basal diet (designated as EM-treated group), the other group just fed with a basal diet (referred as to non-EM treated group or control group). The body weight, daily feed intake, daily gain, feed conversion ratio and other growth parameters were observed and compared between EM-treated and non-EM-treated chicks, and the gut microbiota was profiled by 16S rRNA-based next generation sequencing (NGS). RESULTS: EM-treated chicks showed significantly increased performances in body weight (BW), average daily gain (ADG) and reduced feed conversion ratio (FCR). Histological observation indicated that dietary supplementation of EM significantly increased the villus heights (VH) and the ratio of villus height to crypt depth (VH/CD), while decreased the CD of jejunum, ilea, and ceca. The results of 16S rRNA-based gut microbiota analyses showed that Firmicutes accounted for the most of the relative abundance (63.24%â¼92.63%), followed by Proteobacteria (0.62%â¼23.94%), Bacteroidetes (0.80%â¼7.85%), Actinobacteria (0.06%â¼13.69%) and others in both EM-treated and non-EM-treated broiler chicks. The addition of EM could not alter the alpha diversity of gut microbiota. Compared with the non-EM-treated chicks, the abundances of bad bacteria in the phyla of Firmicutes, Euryarchaeota, and Ruminococcus were dramatically decreased in that of EM-treated chicks, while the abundances of good bacteria in the phyla of Actinobacteria and WPS-2 were significantly increased. CONCLUSIONS: The supplementation of EM in feed could improve the growth performance and positively influence the morphological characteristics of the intestine, and ameliorate the community and structure of the intestinal microbiota of partridge shank broiler chicks.
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Porcine epidemic diarrhea virus (PEDV), an enteric coronavirus, causes neonatal pig acute gastrointestinal infection with a characterization of severe diarrhea, vomiting, high morbidity, and high mortality, resulting in tremendous damages to the swine industry. Neither specific antiviral drugs nor effective vaccines are available, posing a high priority to screen antiviral drugs. The aim of this study is to investigate anti-PEDV effects of carbazole alkaloid derivatives. Eighteen carbazole derivatives (No.1 to No.18) were synthesized, and No.5, No.7, and No.18 were identified to markedly reduce the replication of enhanced green fluorescent protein (EGFP) inserted-PEDV, and the mRNA level of PEDV N. Flow cytometry assay, coupled with CCK8 assay, confirmed No.7 and No.18 carbazole derivatives displayed high inhibition effects with low cell toxicity. Furthermore, time course analysis indicated No.7 and No.18 carbazole derivatives exerted inhibition at the early stage of the viral life cycle. Collectively, the analysis underlines the benefit of carbazole derivatives as potential inhibitors of PEDV, and provides candidates for the development of novel therapeutic agents.
Assuntos
Antivirais/farmacologia , Carbazóis/farmacologia , Vírus da Diarreia Epidêmica Suína/efeitos dos fármacos , Animais , Antivirais/química , Carbazóis/química , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Relação Dose-Resposta a Droga , Estrutura Molecular , Células Vero , Ligação Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacosRESUMO
Blastocystis sp. is a common pathogen that infects the intestines of humans and animals, causing a threat to public health. However, little information on the prevalence and subtypes of Blastocystis sp. in diarrheic pigs in China is available. Herein, 1254 fecal samples were collected from diarrheic pigs in 37 intensive pig farms in Hunan, Jiangxi, and Fujian provinces in southern China, and the prevalence and subtypes of Blastocystis sp. were investigated. Blastocystis sp. was detected by PCR assay, which amplified the small subunit rRNA (SSU rRNA) gene. Overall prevalence of Blastocystis sp. was 31.4% (394/1254), including 21.5% (66/307), 33.1% (99/299), 58.9% (56/95), and 31.3% (173/553) in suckling piglets, weaned piglets, fattening pigs, and sows, respectively. Moreover, age and region factors were significantly related to prevalence of Blastocystis sp. (p < 0.05). Four Blastocystis sp. subtypes were identified, including ST1, ST3, ST5, and ST14. The preponderant subtype was ST5 (76.9%, 303/394). To our knowledge, ST14 was firstly found in pigs in China. The human-pathogenic subtypes (ST1, ST3, ST5, and ST14) that were observed in this study indicate a potential threat to public health. These findings provided a new sight for studying the genetic structure of Blastocystis sp.
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Porcine deltacoronavirus (PDCoV) is a novel enteric coronavirus and is becoming one of the major causative agents of diarrhea in pig herds in recent years. To date, there are no commercial vaccines or antiviral pharmaceutical agents available to control PDCoV infection. Therefore, developing a reliable strategy against PDCoV is urgently needed. In this study, to observe the antiviral activity of RNA interference (RNAi), four short hairpin RNAs (shRNAs) specific to the nucleocapsid (N) gene of PDCoV were designed and tested in vitro. Of these, a double-shRNA-expression vector, designated as pSil-double-shRNA-N1, was the most effectively expressed, and the inhibition of PDCoV replication was then further evaluated in neonatal piglets. Our preliminary results reveal that plasmid-based double-shRNA-expression targeting the N gene of PDCoV can significantly protect LLC-PK1 cells and piglets from pathological lesions induced by PDCoV. Our study could benefit the investigation of the specific functions of viral genes related to PDCoV infection and offer a possible methodology of RNAi-based therapeutics for PDCoV infection.
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Porcine circovirus 3 (PCV3) infections have been reported in different clinical presentations. However, the prevalence and pathogenicity of PCV3 associated with diarrhea in piglets have been limited. Herein, we present an investigation and genome analyses of PCV3 in piglets experiencing diarrhea, and observed clinical signs, gross lesions, and histological changes in pigs negative for all known pathogens associated with diarrhea but positive for PCV3 alone. Among the feces (n = 141) tested, 16.31% (23/141) were positive for PCV3. Of which, 27.28% (15/55) and 14.29% (5/35) were present in diarrheal samples from suckling and weaned piglets, respectively. Moderate to severe atrophic villi was confined in duodenum, jejunum, and ileum, and significantly decreased average heights of villi, and the depths of crypt were observed in PCV3-infected piglets. The complete genome of a representative strain of PCV3, designated as JX/CH/2018, was determined. Multialignment analysis indicated that JX/CH/2018 had 97.7-99.7% nucleotide identity at the complete genome level, and 97.2-100% at the amino acid level of the capsid protein when compared with reference PCV3 strains. Phylogenetic analysis showed that the PCV3 strain identified in this study belonged to PCV3a lineage. The present study demonstrated that PCV3 is a common virus in diarrheal suckling and weaned piglets.
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Porcine deltacoronavirus (PDCoV) has been recently identified as an emerging enteropathogenic coronavirus that mainly infects newborn piglets and causes enteritis, diarrhea and high mortality. Although coronavirus N proteins have multifarious activities, the subcellular localization of the PDCoV N protein is still unknown. Here, we produced mouse monoclonal antibodies against the PDCoV N protein. Experiments using anti-haemagglutinin antibodies and these monoclonal antibodies revealed that the PDCoV N protein is shuttled into the nucleolus in both ectopic PDCoV N-expressing cells and PDCoV-infected cells. The results of deletion mutagenesis experiments demonstrated that the predicted nucleolar localization signal at amino acids 295-318 is critical for nucleolar localization. Cumulatively, our study yielded a monoclonal antibody against the PDCoV N protein and revealed a mechanism by which the PDCoV N protein translocated into the nucleolus. The tolls and findings from this work will facilitate further investigations on the functions of the PDCoV N protein.
Assuntos
Nucléolo Celular/genética , Infecções por Coronavirus/virologia , Proteínas do Nucleocapsídeo de Coronavírus/genética , Deltacoronavirus/genética , Gastroenterite Suína Transmissível/virologia , Interações Hospedeiro-Patógeno/genética , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/química , Anticorpos Antivirais/biossíntese , Anticorpos Antivirais/química , Linhagem Celular , Nucléolo Celular/metabolismo , Infecções por Coronavirus/patologia , Proteínas do Nucleocapsídeo de Coronavírus/metabolismo , Deltacoronavirus/crescimento & desenvolvimento , Deltacoronavirus/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Células Epiteliais/virologia , Gastroenterite Suína Transmissível/patologia , Expressão Gênica , Hemaglutininas Virais/genética , Hemaglutininas Virais/metabolismo , Rim/patologia , Rim/virologia , Camundongos , Sinais de Localização Nuclear , Transporte Proteico , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Deleção de Sequência , SuínosRESUMO
To date, two genotypes, i.e., genotype 1 (G1) and genotype 2 (G2), of porcine epidemic diarrhea virus (PEDV) have been identified in swine, while the cross protection between the G2a and G1a subgenotypes is undetermined. Hence, in the present study, we attempted to observe a comparative pathogenicity and cross protection of G1a (CV777) and G2a (CH/JX/01) PEDVs. Initially pregnant sows were vaccinated twice with the two kinds of inactivated G1a- and G2a-based PEDV vaccines, respectively and the delivered neonatal piglets were challenged with prototype isolates of G1a and G2a PEDVs, and then the pathogenicity and cross-protection in neonatal piglets were observed. The results showed that CH/JX/01, a highly virulent and dominant G2a PEDV strain currently circulating in China had more severe pathogenicity in vitro and in vivo, and induced more strong immune responses, including higher titers of sIgA in maternal milk than that induced by CV777 PEDV, a prototype of G1a PEDV strain. All piglets from the sows immunized with CH/JX/01 could not only survive when challenged with the homologous PEDV, but also be fully protected when challenged with heterogenous G1a PEDV. In contrast, the piglets from the sows immunized with CV777 could be protected when challenged with homologous PEDV and only partially protected when challenged with heterologous G2a strain of PEDV (CH/JX/01). The findings of this study provide new insights into the pathogenicity, antigenicity, and immunogenicity of currently circulating wild type G2a PEDV, which might be valuable for the development of novel PEDV vaccine candidates with improved efficacy.
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The new emergence of swine acute diarrhea syndrome coronavirus (SADS-CoV) has resulted in high mortality in suckling pigs in China. To date, the transcriptional expression of host cells during SADS-CoV infection has not been documented. In this study, by means of RNA-Seq technology, we investigated the whole genomic expression profiles of intestinal porcine epithelial cells (IPEC-J2) infected with a SADS-CoV strain SADS-CoV-CH-FJWT-2018. A total of 24,676 genes were identified: 23,677 were known genes, and 999 were novel genes. A total of 1,897 differentially expressed genes (DEGs) were identified between SADS-CoV-infected and uninfected cells at 6, 24, and 48 h post infection (hpi). Of these, 1,260 genes were upregulated and 637 downregulated. A Gene Ontology enrichment analysis revealed that DEGs in samples from 6, 24, and 48 hpi were enriched in 79, 383, and 233 GO terms, respectively, which were mainly involved in immune system process, response to stimulus, signal transduction, and cytokine-cytokine receptor interactions. The 1,897 DEGs were mapped to 109 KEGG Ontology (KO) pathways classified into four main categories. Most of the DEGs annotated in the KEGG pathways were related to the immune system, infectious viral disease, and signal transduction. The mRNA of porcine serum amyloid A-3 protein (SAA3), an acute phase response protein, was significantly upregulated during the infection. Over-expressed SAA3 in IPEC-J2 cells drastically inhibited the replication of SADS-CoV, while under-expressed SAA3 promoted virus replication. To our knowledge, this is the first report on the profiles of gene expression of IPEC-J2 cells infected by SADS-CoV by means of RNA-Seq technology. Our results indicate that SADS-CoV infection significantly modified the host cell gene expression patterns, and the host cells responded in highly specific manners, including immune response, signal and cytokine transduction, and antiviral response. The findings provide important insights into the transcriptome of IPEC-J2 in SADS-CoV infection.
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BACKGROUND: In China, large-scale outbreaks of severe diarrhea caused by viruses have occurred in pigs since late 2010. To investigate the prevalence and genetic evolution of diarrhea-associated viruses responsible for the outbreaks, a total of 2987 field diarrheal samples collected from 168 pig farms in five provinces in Southern China during 2012-2018 were tested. RESULTS: Porcine epidemic diarrhea virus (PEDV) was most frequently detected virus with prevalence rates between 50.21 and 62.10% in samples, and 96.43% (162/168) in premises, respectively. Porcine deltacoronavirus (PDCoV) was the second prevalent virus with prevalence rates ranging from 19.62 to 29.19% in samples, and 70.24% (118/168) in premises, respectively. Both transmissible gastroenteritis virus (TGEV) and porcine rotavirus (PoRV) were detected at low prevalence rates of < 3% in samples and 10.12% in premises. In this study, we identified a newly emerged swine acute diarrhea syndrome coronavirus (SADS-CoV) in diarrheal samples of piglets from Fujian province in Southern China, and the prevalence rate of SADS-CoV was 10.29% (7/68). Co-infections of these diarrhea-associated viruses were common. The most frequent co-infection was PEDV with PDCoV, with an average detection rate of 12.72% (380/2987, ranging from 8.26-17.33%). Phylogenetic analysis revealed that PEDVs circulating in Southern China during the last 7 years were clustered with the variant strains of PEDV in genotype IIa. The most frequent mutations were present in the collagenase equivalent (COE) and epitope regions of the spike gene of the PEDVs currently circulating in the field. Genetic relationships of PDCoVs were closely related with Chinese strains, other than those present in the USA, South Korea, Thailand and Lao's public. CONCLUSIONS: The findings of this study indicated that variant PEDV, PDCoV, and SADS-CoV were leading etiologic agents of porcine diarrhea, and either mono-infections or co-infections of pathogenic enteric CoVs were common in pigs in Southern China during 2012-2018. Thus, significant attention should be paid in order to effectively prevent and control porcine viral diarrhea.
Assuntos
Diarreia/veterinária , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/virologia , Alphacoronavirus/isolamento & purificação , Animais , China/epidemiologia , Coinfecção/veterinária , Coinfecção/virologia , Coronavirus/isolamento & purificação , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Diarreia/epidemiologia , Diarreia/virologia , Fezes/virologia , Filogenia , Vírus da Diarreia Epidêmica Suína/classificação , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/isolamento & purificação , Prevalência , Rotavirus/isolamento & purificação , SuínosRESUMO
A simple and accurate reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay was developed and evaluated for the detection of porcine pegivirus (PPgV). The specific RT-LAMP primers targeting the conserved regions of NS5A genes were designed and used to detect PPgV. The optimal reaction parameter for RT-LAMP assay was 63â for 60 min. The detection limit of the RT-LAMP assay was 10 copies of PPgV genome, which was 100 times more sensitive than that of the conventional RT-PCR and comparable to nested RT-PCR and quantitative RT-PCR (qRT-PCR). There was no cross amplification with other related RNA viruses. In the clinical evaluation, the RT-LAMP assay exhibited a similar sensitivity with nested RT-PCR and qRT-PCR. The results indicated that RT-LAMP assay developed in this study could be a highly specific, sensitive, and cost-effective alternative for a rapid detection of PPgV in field settings.
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Infecções por Flaviviridae/veterinária , Flaviviridae/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Doenças dos Suínos/virologia , Animais , Primers do DNA , Flaviviridae/genética , Infecções por Flaviviridae/diagnóstico , Limite de Detecção , RNA Viral/isolamento & purificação , Transcrição Reversa , Sensibilidade e Especificidade , SuínosRESUMO
Swine enteric coronaviruses (SECoVs), including porcine epidemic diarrhea virus (PEDV), swine acute diarrhea syndrome coronavirus (SADS-CoV), and porcine deltacoronavirus (PDCoV) have emerged and been prevalent in pig populations in China for the last several years. However, current traditional inactivated and attenuated PEDV vaccines are of limited efficacy against circulating PEDV variants, and there are no commercial vaccines for prevention of PDCoV and SADS-CoV. RNA interference (RNAi) is a powerful tool in therapeutic applications to inhibit viral replication in vitro. In this study, we developed a small interfering RNA generation system that expressed two different short hairpin RNAs (shRNAs) targeting the M gene of PEDV and SADS-CoV and the N gene of PDCoV, respectively. Our results demonstrated that simultaneous expression of these specific shRNA molecules inhibited expression of PEDV M gene, SADS-CoV M gene, and PDCoV N gene RNA by 99.7%, 99.4%, and 98.8%, respectively, in infected cell cultures. In addition, shRNA molecules significantly restricted the expression of M and N protein, and impaired the replication of PEDV, SADS-CoV, and PDCoV simultaneously. Taken together, this shRNAs expression system not only is proved to be a novel approach for studying functions of various genes synchronously, but also developed to test aspects of a potential therapeutic option for treatment and prevention of multiple SECoV infections.
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Alphacoronavirus/genética , Infecções por Coronavirus/veterinária , Coronavirus/genética , Vírus da Diarreia Epidêmica Suína/genética , RNA Interferente Pequeno , Alphacoronavirus/efeitos dos fármacos , Animais , China , Coronavirus/efeitos dos fármacos , Infecções por Coronavirus/genética , Infecções por Coronavirus/terapia , Genes Virais , Terapia Genética , Vírus da Diarreia Epidêmica Suína/efeitos dos fármacos , Interferência de RNA , RNA Interferente Pequeno/biossíntese , RNA Interferente Pequeno/uso terapêutico , Suínos , Doenças dos Suínos/terapia , Doenças dos Suínos/virologiaRESUMO
Porcine pegiviruses (PPgV) have been first discovered in serum samples from domestic pigs in Germany in 2016 and then in the USA in 2018. To date, there is no documentation with respect to the presence of PPgVs in domestic pigs in China. Herein, we attempted to determine the presence and prevalence of PPgV in China and its genetic characterization. In this study, 469 sera were tested and 34 (7.25%) were positive for PPgV. An ascending trend of the positive rate for PPgV was observed from suckling piglets (1.61%) to nursing piglets (1.85%), finishing pigs (6.56%), and sows (11.34%). The complete genome sequence of a representative strain of PPgV, PPgV_GDCH2017, and the complete E2 gene of 17 PPgV isolates discovered in this study was determined. Sequence analysis indicated that PPgV_GDCH2017 was highly related to other PPgVs with nucleotide and amino acid identities ranging from 87.3 to 97.4% and 94.6-99.3%, respectively, in the complete coding region. Phylogenetic analyses demonstrated that the PPgV_GDCH2017 discovered in this study was closely related to the PPgVs from the USA and clustered in the same genus with pegiviruses from other hosts. The topology of the phylogenetic tree based on the complete E2 gene was consistent with that based on the complete genome of PPgV. Further studies on pathogenicity and pathogenesis of PPgVs are needed.
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Infecções por Flaviviridae/virologia , Flaviviridae/genética , Genoma Viral/genética , Doenças dos Suínos/genética , Animais , China , Flaviviridae/isolamento & purificação , Flaviviridae/patogenicidade , Infecções por Flaviviridae/genética , Alemanha , Filogenia , Suínos/virologia , Doenças dos Suínos/virologia , Estados Unidos , Sequenciamento Completo do GenomaRESUMO
The full-length genome sequence of a novel swine acute diarrhea syndrome coronavirus (SADS-CoV), CH/FJWT/2018, was determined, which was genetically most closely related to CN/GDWT/2017, recently discovered in Fujian, China. The indel sites of the spike (S) gene of CH/FJWT/2018 were most similar to those of bat-origin SADS-related coronaviruses.
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The full-length genome sequence of a variant of porcine epidemic diarrhea virus (PEDV), that of strain CH/JXJA/2017, was highly homologous to CH/ZMDZY/11, a highly virulent Chinese PEDV strain. CH/JXJA/2017 had a distant relationship with the attenuated CV777 vaccine strain, but the insertion sites of the S1 gene were similar to those of the recombinant strain of CH/ZMDZY/11.
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Porcine epidemic diarrhea virus (PEDV) is a devastating cause of diarrhea in pigs worldwide. Most of studies have focused on molecular and pathogenic characterization of PEDV, whereas there were limited studies in understanding the role of gut microbiota (GM) in viral-associated diarrhea. Here, using the Illumina MiSeq platform, we examined and compared the impact of PEDV infection on the GM of sows and their piglets less than 10 days old. Our results showed that PEDV caused alternations in the structure and abundance of GM from levels of phylum to genus, and even species. For sows, a significant decrease of observed species was found in diarrheal sows than that in healthy sows (p < 0.05). The unweighted and weighted UniFrac distances also revealed considerable segregations of GM structure among healthy, asymptomatic, and diarrheal sows. For piglets, Bacteroidetes, the dominant bacteria in healthy piglets, were replaced by Firmicutes in asymptomatic and diarrheal piglets. The abundances of Fusobacteria and Proteobacteria were also remarkably increased in asymptomatic piglets and diarrheal piglets when compared to those of the healthy piglets. Our findings demonstrated that PEDV infection caused severe perturbations of GM, reduced probiotic bacteria, and enriched pathogenic bacteria.