RESUMO
Patients with chronic hepatitis C who achieve a sustained viral response after pegylated interferon therapy have a reduced risk of hepatocellular carcinoma, but the risk after treatment with direct-acting antivirals is unclear. We compared the rates of early development of hepatocellular carcinoma after direct-acting antivirals and after pegylated interferon therapy. We retrospectively analysed 785 patients with chronic hepatitis C who had no history of hepatocellular carcinoma (211 treated with pegylated interferon, 574 with direct-acting antivirals) and were followed up for at least 24 weeks after antiviral treatment. De novo hepatocellular carcinoma developed in 6 of 574 patients receiving direct-acting antivirals and in 1 of 211 patients receiving pegylated interferon. The cumulative incidence of early hepatocellular carcinoma development did not differ between the treatment groups either for the whole cohort (1.05% vs 0.47%, P = .298) or for those patients with Child-Pugh Class A cirrhosis (3.73% vs 2.94%, P = .827). Multivariate analysis indicated that alpha-fetoprotein level >9.5 ng/mL at the time of end-of-treatment response was the only independent risk factor for early development of hepatocellular carcinoma in all patients (P < .0001, hazard ratio 176.174, 95% confidence interval 10.768-2882.473) and in patients treated with direct-acting agents (P < .0001, hazard ratio 128.402, 95% confidence interval 8.417-1958.680). In conclusion, the rate of early development of hepatocellular carcinoma did not differ between patients treated with pegylated interferon and those treated with direct-acting antivirals and was associated with the serum alpha-fetoprotein level at the time of end-of-treatment response.
Assuntos
Antivirais/uso terapêutico , Carcinoma Hepatocelular/epidemiologia , Hepatite C Crônica/tratamento farmacológico , Interferon-alfa/uso terapêutico , Neoplasias Hepáticas/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Hepatite C Crônica/epidemiologia , Humanos , Incidência , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/epidemiologia , Masculino , Pessoa de Meia-Idade , República da Coreia/epidemiologia , Estudos Retrospectivos , Fatores de Risco , Adulto JovemRESUMO
In the past 4 years, incidences of endemic or epidemic respiratory diseases associated with canine influenza H3N2 virus in Asian dogs have been reported in countries such as South Korea and China. Canine species were considered to be the new natural hosts for this virus. However, at the beginning of 2010, influenza-like respiratory signs, such as dyspnoea, were also observed among cats as well as in dogs in an animal shelter located in Seoul, South Korea. The affected cats showed 100â% morbidity and 40â% mortality. We were able to isolate a virus from a lung specimen of a dead cat, which had suffered from the respiratory disease, in embryonated-chicken eggs. The eight viral genes isolated were almost identical to those of the canine influenza H3N2 virus, suggesting interspecies transmission of canine influenza H3N2 virus to the cat. Moreover, three domestic cats infected with intranasal canine/Korea/GCVP01/07 (H3N2) all showed elevated rectal temperatures, nasal virus shedding and severe pulmonary lesions, such as suppurative bronchopneumonia. Our study shows, for the first time, that cats are susceptible to canine influenza H3N2 infection, suggesting that cats may play an intermediate host role in transmitting the H3N2 virus among feline and canine species, which could lead to the endemic establishment of the virus in companion animals. Such a scenario raises a public health concern, as the possibility of the emergence of new recombinant feline or canine influenza viruses in companion animals with the potential to act as a zoonotic infection cannot be excluded.
Assuntos
Doenças do Gato/epidemiologia , Doenças do Gato/virologia , Doenças do Cão/transmissão , Doenças do Cão/virologia , Vírus da Influenza A Subtipo H3N2/isolamento & purificação , Infecções por Orthomyxoviridae/veterinária , Animais , Temperatura Corporal , Doenças do Gato/mortalidade , Doenças do Gato/patologia , Gatos , Análise por Conglomerados , Doenças do Cão/epidemiologia , Doenças do Cão/patologia , Cães , Fezes/virologia , Vírus da Influenza A Subtipo H3N2/classificação , Vírus da Influenza A Subtipo H3N2/genética , Pulmão/virologia , Dados de Sequência Molecular , Infecções por Orthomyxoviridae/epidemiologia , Infecções por Orthomyxoviridae/virologia , Filogenia , RNA Viral/genética , República da Coreia/epidemiologia , Análise de Sequência de DNA , Eliminação de Partículas ViraisRESUMO
AIMS: Atopic dermatitis (AD) is marked by elevated levels of immunoglobulin E and skin lesions such as oedema and haemorrhage. Kimchi is a Korean fermented food that contains beneficial bacteria for human health. In this study, Lactobacillus plantarum CJLP55, CJLP56, CJLP133 and CJLP136 isolated from Kimchi were investigated for their capacity to inhibit AD. METHODS AND RESULTS: The three strains, CJLP55, CJLP133 and CJLP136, suppressed AD-like skin lesions, high serum IgE levels and epidermal thickening. The three strains diminished the accumulation of eosinophils and mast cells into topical inflammatory sites and the enlargement of axillary lymph nodes, which are responsible for the dorsal dermatitis. CJLP55, CJLP133 and CJLP136 decreased production of type 2 cytokines such as IL-4 and IL-5 in lymph node cell culture. CJLP133 and CJLP136 increased IFN-γ secretion, while CJLP55 enhanced IL-10 production. CONCLUSIONS: The three strains isolated from Kimchi suppress house-dust mite-induced dermatitis in NC/Nga mouse, a representative animal model of human AD. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings suggest that lactobacilli isolated from Kimchi inhibit AD, probably by altering the balance of Th1/Th2 ratio or inducing IL-10 production.
Assuntos
Dermatite Atópica/terapia , Microbiologia de Alimentos , Lactobacillus plantarum/isolamento & purificação , Probióticos , Pele/patologia , Administração Oral , Animais , Brassica/microbiologia , Dermatite Atópica/imunologia , Dermatite Atópica/patologia , Modelos Animais de Doenças , Eosinófilos/imunologia , Feminino , Fermentação , Imunoglobulina E/sangue , Inflamação/patologia , Interferon gama/biossíntese , Interleucina-10/biossíntese , Interleucina-4/biossíntese , Interleucina-5/biossíntese , Linfonodos/patologia , Mastócitos/imunologia , Camundongos , PyroglyphidaeRESUMO
BACKGROUND AND OBJECTIVE: Recombinant human bone morphogenetic protein-2 (rhBMP-2) is a potent inducer for the regeneration of mineralized tissue, but has a limited effect on the regeneration of cementum and periodontal ligament (PDL). The aim of the present study was to determine the effects of rhBMP-2 on the in vitro and in vivo biologic activity of well-characterized human PDL stem cells (hPDLSCs) and to elucidate the underlying mechanism of minimal periodontal regeneration by rhBMP-2. MATERIAL AND METHODS: hPDLSCs were isolated and cultured, and then transplanted into an ectopic subcutaneous mouse model using a carrier treated either with or without rhBMP-2. Comprehensive histologic, histometric and immunohistochemical analyses were performed after an 8-wk healing period. The effects of rhBMP-2 on the adipogenic and osteogenic/cementogenic differentiation of hPDLSCs were also evaluated. The effect of rhBMP-2 on both soluble and insoluble collagen synthesis was analyzed, and the expression of mRNA and protein for collagen types I, II, III and V was assessed. RESULTS: In the present study, rhBMP-2 promoted both adipogenic and osteogenic/cementogenic differentiation of hPDLSCs in vitro, and the in vivo potential of hPDLSCs to form mineralized cementum and organized PDL tissue was down-regulated following treatment with rhBMP-2. Collagen synthesis, which plays a crucial role in the regeneration of cementum and the periodontal attachment, was significantly reduced, with associated modification of the relevant mRNA and protein expression profiles. CONCLUSION: In summary, the findings of the present study suggest that enhanced adipogenic differentiation and inhibition of collagen synthesis by hPDLSCs appear to be partly responsible for the minimal effect of rhBMP-2 on cementum and PDL tissue regeneration by hPDLSCs.
Assuntos
Adipogenia/efeitos dos fármacos , Proteínas Morfogenéticas Ósseas/farmacologia , Colágeno/efeitos dos fármacos , Ligamento Periodontal/citologia , Proteínas Recombinantes/farmacologia , Células-Tronco/efeitos dos fármacos , Fator de Crescimento Transformador beta/farmacologia , Adipogenia/fisiologia , Adolescente , Animais , Proteína Morfogenética Óssea 2 , Osso e Ossos/anatomia & histologia , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Cementogênese/efeitos dos fármacos , Colágeno/biossíntese , Colágeno Tipo I/efeitos dos fármacos , Colágeno Tipo II/efeitos dos fármacos , Colágeno Tipo III/efeitos dos fármacos , Colágeno Tipo V/efeitos dos fármacos , Cemento Dentário/anatomia & histologia , Relação Dose-Resposta a Droga , Regulação para Baixo , Humanos , Camundongos , Osteogênese/efeitos dos fármacos , Ligamento Periodontal/efeitos dos fármacos , Regeneração/efeitos dos fármacos , Transplante de Células-Tronco , Células-Tronco/fisiologia , Tela Subcutânea/cirurgia , Fatores de Tempo , Alicerces Teciduais , Adulto JovemRESUMO
Avian-lineage H3N2 canine influenza virus (CIV)-associated respiratory disease, which can be fatal, emerged in South Korean dogs in 2007. We show here that dogs experimentally infected with CIV only developed respiratory tract diseases, as no extrapulmonary lesions and virus antigens were detected. This differs from the multiorgan diseases that avian influenza H5N1 induces in small experimental animals. However, the CIV-infected dogs developed a distinctively severe, long-persistent bronchointerstitial pneumonia, which differs from the acute but short-term bronchopneumonia that human (H1N1 and H3N2) influenza cause in rodents and ferrets. Histopathology and in situ TUNEL assays revealed that the neutrophils infiltrating the lesions were undergoing apoptosis, which probably reflects the attempts by the body to maintain appropriate numbers of neutrophils for defense against secondary bacterial infections. Our observations suggest that neutrophils along with the related chemoattractant cytokines (TNF-alpha, IL-1 and IL-8, etc.) may play a key role in the pathogenesis of H3N2 CIV in dogs.
Assuntos
Doenças do Cão/virologia , Vírus da Influenza A Subtipo H3N2/patogenicidade , Infecções por Orthomyxoviridae/veterinária , Animais , Apoptose , Doenças do Cão/patologia , Cães , Humanos , Vírus da Influenza A Subtipo H1N1 , Virus da Influenza A Subtipo H5N1 , Influenza Humana/virologia , Interleucina-1/sangue , Interleucina-1/fisiologia , Interleucina-8/sangue , Interleucina-8/fisiologia , Pulmão/patologia , Pulmão/virologia , Neutrófilos/patologia , Infecções por Orthomyxoviridae/patologia , Traqueia/patologia , Traqueia/virologia , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
A miniarray system was developed for the simultaneous detection of porcine circovirus type 1 (PCV1) and type 2 (PCV2) in pigs. The system consists of a polymerase chain reaction (PCR) step to amplify target viral DNA, followed by detection of the amplified DNA using a membrane-anchored probe array and an avidin-alkaline phosphatase (Av-AP) indicator system. The lower limit of detection of PCV using the miniarray was 10(1.9) tissue culture infectious dose 50 (TCID(50))/ml and 10(2.08)TCID(50)/ml for PCV1 and PCV2, respectively, and 100 viral copies/microl for both PCV1 and PCV2. We validated the miniarray system using 141 lymph node specimens from pigs with suspected postweaning multisystemic wasting syndrome or porcine dermatitis and nephropathy syndrome. Of the 141 samples evaluated, 55 were identified as positive for PCV by the miniarray. Relative to in situ hybridization, the sensitivity and specificity of the miniarray was 100% and 98.9%, respectively. In contrast to other microarray systems, the miniarray does not require a DNA chip reader, since the results can be determined by visual inspection of colorized spots on a nylon membrane. This system represents an effective alternative method for the differential detection of PCV1 and PCV2 in pigs, as well as the maintenance of PCV-free cell lines and pre-screening of commercial vaccines for possible contamination.
Assuntos
Infecções por Circoviridae/veterinária , Circovirus/isolamento & purificação , Análise de Sequência com Séries de Oligonucleotídeos/veterinária , Síndrome Definhante Multissistêmico de Suínos Desmamados/virologia , Animais , Infecções por Circoviridae/diagnóstico , Infecções por Circoviridae/virologia , Circovirus/genética , DNA Viral/química , DNA Viral/genética , Hibridização In Situ/veterinária , Linfonodos/virologia , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Síndrome Definhante Multissistêmico de Suínos Desmamados/diagnóstico , Sensibilidade e Especificidade , SuínosRESUMO
Several influenza A viral subtypes were isolated from pigs during a severe outbreak of respiratory disease in Korea during 2005 and 2006. They included a classical swine H1N1 subtype, two swine-human-avian triple-recombinant H1N2 subtypes, and a swine-human-avian triple-recombinant H3N2 subtype. In the current study, genetic characterization to determine the probable origin of these recent isolates was carried out for the first time. Phylogenetic analysis indicated that all the recent Korean isolates of H1N1, H1N2, and H3N2 influenza are closely related to viruses from the United States. Serologic and genetic analysis indicated that the Korean H1N2 viral subtypes were introduced directly from the United States, and did not arise from recombination between Korean H1N1 and H3N2. We suggest that the H1N1, H1N2, and H3N2 viral subtypes that were isolated from the Korean swine population originated in North America, and that these viruses are currently circulating in the Korean swine population.
Assuntos
Vírus da Influenza A/classificação , Vírus da Influenza A/isolamento & purificação , Infecções por Orthomyxoviridae/veterinária , Filogenia , Doenças dos Suínos/virologia , Animais , Anticorpos Antivirais/sangue , Sequência de Bases , Linhagem Celular , Embrião de Galinha , Cobaias , Vírus da Influenza A/genética , Vírus da Influenza A/imunologia , Coreia (Geográfico) , Dados de Sequência Molecular , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/transmissão , Infecções por Orthomyxoviridae/virologia , Alinhamento de Sequência , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/transmissão , Proteínas Virais/genéticaRESUMO
The swine influenza virus (SIV) H1N1, H1N2, and H3N2 subtypes circulate in Korean farm. A novel multiplex RT-PCR (m-RT-PCR) was developed to detect and subtype swine influenza viruses. This m-RT-PCR assay could identify H1, H3, N1 and N2 from clinical samples in single tube reaction using DPO system. Korean SIVs are closely related to the United States influenza viruses, and primers were developed for SIV from North American viruses and recently Korean isolates. The sensitivity of the m-RT-PCR was 10TCID(50)/ml for H1N1, H1N2 or H3N2. The lowest viral concentrations detected by single PCR were 1TCID(50)/ml for each subtype. Non-specific reactions were not observed when other viruses and bacteria were used to assess the m-RT-PCR. The results of m-RT-PCR were more effective than virus isolation or hemagglutination (HA) test. This assay using a DPO system provides a rapid, sensitive, and cost-effective laboratory diagnosis for detecting and subtyping of SIV in pigs.
Assuntos
Primers do DNA/genética , Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A Subtipo H3N2 , Vírus da Influenza A , Infecções por Orthomyxoviridae/veterinária , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Doenças dos Suínos , Suínos/virologia , Animais , Vírus da Influenza A Subtipo H1N1/classificação , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Vírus da Influenza A Subtipo H3N2/classificação , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/isolamento & purificação , Vírus da Influenza A/classificação , Vírus da Influenza A/genética , Vírus da Influenza A/isolamento & purificação , Coreia (Geográfico) , Infecções por Orthomyxoviridae/diagnóstico , Infecções por Orthomyxoviridae/virologia , RNA Viral/análise , RNA Viral/isolamento & purificação , Sensibilidade e Especificidade , Doenças dos Suínos/diagnóstico , Doenças dos Suínos/virologiaRESUMO
To eliminate porcine reproductive and respiratory syndrome virus (PRRSV) from a supplying boar stud, samples of serum and semen from 118 boars were assessed three times a month by an indirect fluorescent antibody (IFA) test to detect antibodies, and by a nested reverse transcriptase-PCR (nRT-PCR) to detect the genome of PRRSV. The boars detected as persistently infected carriers were culled. A PRRSV-negative population of boars was established after three months and no semen positive for the virus was detected for six months. Subsequently, a three-step plan was introduced to eliminate PRRSV from the seedstock breeding farm during three parity cycles on the farm over 15 months, each step taking five months. In step 1, umbilical cords taken from piglets at birth and serum samples taken from their dams at the start of weaning were subjected to ifa and nRT-pcr analysis. The sows with PRRSV detected in serum by nRT-pcr were regarded as carrier sows and culled. The rates of detection of PRRSV were reduced from 5 percent to 2.5 percent in the sera of the sows, and from 14.8 percent to 1.8 percent in the umbilical cords of the piglets. In step 2, the sows that had farrowed the piglets with PRRSV detected by nRT-PCR in their cords were considered to have transmitted the infection and removed. During step 2, the virus detection rates in umbilical cords by nRT-pcr were reduced, but not completely eliminated. In step 3, 10-week-old nursery pigs with antibodies to PRRSV in their serum by ifa and elisa were culled. The three steps established the PRRSV-negative state of the multisite farm containing the breeding and nursery farm, and the PRRSV-negative state of both the multisite farm and the supplying boar stud was evaluated by monthly monitoring over at least one parity cycle of the farm for five months.
Assuntos
Cruzamento , Síndrome Respiratória e Reprodutiva Suína/prevenção & controle , Síndrome Respiratória e Reprodutiva Suína/transmissão , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Animais , Anticorpos Antivirais/sangue , Portador Sadio/veterinária , Ensaio de Imunoadsorção Enzimática/métodos , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Sangue Fetal/virologia , Imunofluorescência/métodos , Imunofluorescência/veterinária , Masculino , Paridade , Vírus da Síndrome Respiratória e Reprodutiva Suína/metabolismo , Gravidez , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Sêmen/virologia , Suínos , Fatores de TempoAssuntos
Infecções por Orthomyxoviridae/veterinária , Doenças dos Suínos/epidemiologia , Animais , Primers do DNA , DNA Viral/análise , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/isolamento & purificação , Vírus da Influenza A/genética , Vírus da Influenza A/isolamento & purificação , Coreia (Geográfico)/epidemiologia , Infecções por Orthomyxoviridae/epidemiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Suínos , Doenças dos Suínos/etiologia , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/virologiaRESUMO
Hepatitis E virus (HEV) infections have been reported in pigs throughout the world but have only recently been recorded in Korean pigs. The aim of this study was to investigate whether HEV was present in archived porcine hepatic tissues collected between 1995 and 2004 using RT-PCR and immunohistochemistry and, if so, to determine the genotype of the isolates. Swine HEV was identified in the liver tissue of 42 pigs of 388 submissions (four pigs every year on average). The isolates showed genetic homology with swine and human HEV isolates identified in the United States and Japan (92.5-97%) and phylogenetic tree analysis indicated they belonged to genotype III. The study indicates that HEV is not a newly emerging virus in Korean pigs, but a pathogen that has existed in the country since at least 1995.
Assuntos
Vírus da Hepatite E/isolamento & purificação , Hepatite E/veterinária , Filogenia , Doenças dos Suínos/epidemiologia , Animais , Genótipo , Hepatite E/epidemiologia , Hepatite E/virologia , Vírus da Hepatite E/classificação , Imuno-Histoquímica/veterinária , Coreia (Geográfico)/epidemiologia , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Estudos Soroepidemiológicos , Suínos , Doenças dos Suínos/virologiaRESUMO
A Vero cell attenuated porcine epidemic diarrhea virus (PEDV) strain, DR13, was distinguished from wild-type PEDV using restriction enzyme fragment length polymorphism (RFLP). Cell attenuated DR13 was orally or intramuscularly (IM) administered to late-term pregnant sows, and mortality resulting from the highly virulent PEDV challenge was investigated in passively immunized suckling piglets of the two different groups. The mortality rate of the oral group (13%) was lower than that of the IM group (60%). In particular, the concentration of IgA against PEDV was higher in piglets of sows in the oral group, compared to the IM group. The attenuated DR13 virus remained safe, even after three backpassages in piglets. The findings of this study support the theory that the Vero cell attenuated DR13 virus may be applied as an oral vaccine for inducing specific immunity in late-term pregnant sows with a high margin of protection against PEDV infection.
Assuntos
Infecções por Coronavirus/veterinária , Vírus da Diarreia Epidêmica Suína/classificação , Vírus da Diarreia Epidêmica Suína/patogenicidade , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/virologia , Vacinas Virais , Administração Oral , Animais , Anticorpos Antivirais/sangue , Chlorocebus aethiops , Infecções por Coronavirus/sangue , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/virologia , Feminino , Vírus da Diarreia Epidêmica Suína/imunologia , Vírus da Diarreia Epidêmica Suína/fisiologia , Gravidez , Inoculações Seriadas , Suínos , Doenças dos Suínos/sangue , Fatores de Tempo , Células Vero , Vacinas Virais/administração & dosagem , Vacinas Virais/efeitos adversos , Eliminação de Partículas ViraisRESUMO
A swine influenza H1N1 virus was isolated from a pig during a severe outbreak of respiratory disease in Korea. All genes of the H1N1 isolate, including hemagglutinin (HA), neuraminidase (NA), matrix (M), nucleoprotein (NP), non-structural (NS), PA, PB1 and PB2, were of swine origin. Also, all these genes showed a close phylogenic relationship with those of H1N1 viruses previously isolated from pigs in the United States. These results suggest that North American swine influenza virus has actually been transmitted to pigs in Korea.
Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H1N1/classificação , Doenças dos Suínos/virologia , Animais , Vírus da Influenza A Subtipo H1N1/enzimologia , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Coreia (Geográfico) , FilogeniaRESUMO
Phylogenetic relationships of commercial Chinese truffles and their allies were investigated mainly by morphological studies and analyses of the sequences of ITS regions of nuclear ribosomal DNA. Two species, Tuber indicum and T. himalayense, closely related to the European T. melanosporum (the Perigord Truffle), are recognized among commercial Chinese black truffles. Both T. pseudohimalayense and T. sinense should be regarded as synonyms of T. indicum. Tuber species producing excavated ascomata are not monophyletic, suggesting that excavation of ascomata may have evolved more than once, or evolved once during the evolution of truffle species and then was lost once during the evolution of Tuber species.
Assuntos
Ascomicetos/classificação , Ascomicetos/genética , Filogenia , Ascomicetos/fisiologia , China , DNA Fúngico/análise , DNA Espaçador Ribossômico/análise , Dados de Sequência Molecular , Técnicas de Tipagem Micológica , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
A porcine epidemic diarrhea virus (PEDV) designated DR13 was isolated in Vero cells and serially passaged by level 100. The virus was titrated at regular intervals of the passage level. Open reading frame (ORF) 3 sequences of the virus at passage levels 20, 40, 60, 80, and 100 were aligned and compared using a computer software program. Suitability of the restriction fragment length polymorphism (RFLP) analysis for differentiating the virus from other Korean field strains was investigated. The DR13 field isolate was successively adapted in Vero cells as observed through polymerase chain reaction (PCR) and titration of the virus. RFLP analysis identified change in cleavage sites of HindIII and Xho II from passage levels 75 and 90, respectively; these RFLP patterns of ORF 3 differentiated the Vero cell-adapted virus from its parent strain, DR13, and 12 other strains of PEDV studied. The cell adapted DR13 was tested for its pathogenicity and immunogenicity in piglets and pregnant sows. The results indicated that cell adapted DR13 revealed reduced pathogenicity and induced protective immune response in pigs. Differentiation between highly Vero cell-adapted virus and wild-type virus could be the marker of adaptation to cell culture and a valuable tool for epidemiologic studies of PEDV infections. The results of this study supported that the cell attenuated virus could be applied as a marker vaccine candidate against PEDV infection.
Assuntos
Fases de Leitura Aberta , Polimorfismo de Fragmento de Restrição , Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Animais , Sequência de Bases , Chlorocebus aethiops , Feminino , Coreia (Geográfico) , Camundongos , Reação em Cadeia da Polimerase , Síndrome Respiratória e Reprodutiva Suína/imunologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/crescimento & desenvolvimento , Vírus da Síndrome Respiratória e Reprodutiva Suína/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , SuínosRESUMO
Transmissible gastroenteritis (TGE) and porcine epidemic diarrhea (PED) are highly contagious enteric diseases of piglets. The clinical signs of these diseases are very similar and include watery, yellowish diarrhea. Thus, the effective differential detection of TGE virus and PED virus is required. In the present study, a duplex reverse transcription-polymerase chain reaction (RT-PCR) was established for the differential detection of TGE and PED viruses. The primers were designed for the S gene of each virus. RNA was extracted from the intestines and stool samples that were collected from the swine with diarrhea. The RT-PCR test could detect both TGE and PED viruses with 2 TCID50/200 microl. Among 90 clinical samples, 7 TGE viruses and 2 PED viruses were detected by the duplex RT-PCR. This duplex RT-PCR may be a useful diagnostic method for the rapid, specific, and sensitive differential detection of TGE and PED viruses using clinical samples.
Assuntos
Coronaviridae/genética , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/veterinária , DNA Viral/análise , Diarreia/veterinária , Diarreia/virologia , Gastroenterite Suína Transmissível/diagnóstico , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Vírus da Gastroenterite Transmissível/genética , Animais , Coronaviridae/patogenicidade , Infecções por Coronavirus/genética , Primers do DNA , Diagnóstico Diferencial , Gastroenterite Suína Transmissível/genética , Sensibilidade e Especificidade , Suínos , Vírus da Gastroenterite Transmissível/patogenicidadeRESUMO
Heat-stable antigen (HSA) is a murine differentiating antigen that is expressed on both CD4-CD8- double-negative and CD4+CD8+ double-positive thymocytes but not CD4+ or CD8+ single-positive thymocytes. Effects of anti-HSA monoclonal antibody, R13, on thymocyte apoptosis induced by various stimulations were investigated by a single-cell suspension culture system. Immobilized R13 enhanced the CD3-mediated DNA fragmentation and killing of thymocytes but not the dexamethasone-induced or phorbol myristate acetate-induced killing of thymocytes. Immobilized R13 by itself could not induce thymocyte apoptosis. Soluble R13 enhanced CD3-mediated apoptosis when HSA and T-cell receptor (TCR)/CD3 were co-cross-linked by a cross-reactive secondary antibody. Even without the cross-reactive secondary antibody, soluble R13 enhanced CD3-mediated apoptosis, although a greater than 100-fold increase in the amount of R13 was needed to give a similar enhancement compared with immobilized R13. Neither R13 by itself nor R13 plus secondary antibody induced cytosolic calcium influx, whereas R13 enhanced CD3-mediated cytosolic calcium increase. These results suggest a functional role of HSA in promoting the activation-induced apoptosis of thymocytes and the involvement of HSA in negative selection.