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Aleutian mink disease parvovirus (ADV) mRNAs are found in macrophages in lymph nodes and peritoneal exudate cells from ADV-infected mink. Therefore, we developed an in vitro infection system for ADV by using primary cultures of mink macrophages or macrophage cell lines. In peritoneal macrophage cultures from adult mink, virulent ADV-Utah I strain showed nuclear expression of viral antigens with fluorescein isothiocyanate-labeled ADV-infected mink serum, but delineation of specific viral proteins could not be confirmed by immunoblot analysis. Amplification of ADV DNA and production of replicative-form DNA were observed in mink macrophages by Southern blot analysis; however, virus could not be serially propagated. The human macrophage cell line U937 exhibited clear nuclear expression of viral antigens after infection with ADV-Utah I but not with tissue culture-adapted ADV-G. In U937 cells, ADV-Utah I produced mRNA, replicative-form DNA, virion DNA, and structural and nonstructural proteins; however, virus could not be serially passaged nor could [3H]thymidine-labeled virions be observed by density gradient analysis. These findings indicated that ADV-Utah I infection in U937 cells was not fully permissive and that there is another restricted step between gene amplification and/or viral protein expression and production of infectious virions. Treatment with the macrophage activator phorbol 12-myristate 13-acetate after adsorption of virus reduced the frequency of ADV-positive U937 cells but clearly increased that of human macrophage line THP-1 cells. These results suggested that ADV replication may depend on conditions influenced by the differentiation state of macrophages. U937 cells may be useful as an in vitro model system for the analysis of the immune disorder caused by ADV infection of macrophages.  相似文献   

3.
We studied different parameters during the development of acute interstitial pneumonia in mink kits caused by neonatal infection with Aleutian disease virus (ADV). When histological lesions, presence of intranuclear inclusion bodies, and intranuclearly localized ADV antigen were correlated with levels of single-stranded virion and duplex replicative forms of ADV DNA in the different tissues, it was concluded that the lung, probably alveolar type II cells, is the major primary target for viral replication and cytopathology. The presence of the duplex dimeric replicative-form DNA, a strong marker of parvovirus replication, was also observed in low amount in the mesenteric lymph node, suggesting replication of ADV in this organ, although no viral cytopathology could be demonstrated. Moreover, a few intranuclear inclusion bodies were demonstrated in kidney and liver from affected kits, but intranuclearly localized ADV antigen could not be demonstrated in liver sections, and neither could duplex dimer replicative-form DNA, suggesting that these organs are nevertheless not a major site of ADV replication. When the data were compared with results previously reported for ADV-infected adult mink and ADV-infected permissive cell cultures, the data suggested that the pattern of ADV replication in alveolar type II cells is similar to that seen in infected cell cultures but that the replication in the other kit organs resembles the restricted pattern seen in adult mink.  相似文献   

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The transplacental transmission of Aleutian mink disease parvovirus (ADV) was studied in experimental infection of 1-year-old female non-Aleutian mink. The ADV-seronegative female mink were inoculated with ADV prior to mating or after the expected implantation of the embryos during pregnancy. A group of uninfected females served as a control group. Animals from each group were killed prior to or shortly after parturition. The in situ hybridization technique with radiolabeled strand-specific RNA probes was used to determine target cells of virus infection and virus replication. In both infected groups, ADV crossed the endotheliochorial placental barrier, although animals infected before mating already had high antibody titers against ADV at the time of implantation. The percentage of dead and resorbed fetuses was much higher in dams infected before mating. In the placentae of these mink, virus DNA and viral mRNA were detected in cells in the mesenchymal stroma of the placental labyrinth and hematoma but only occasionally in the cytotrophoblast of the placental hematoma. Placentae of animals infected during pregnancy showed in addition very high levels of virus and also viral replication in a large number of cytotrophoblast cells in the placental hematoma, which exhibited distinct inclusion bodies. In both groups, neither virus nor virus replication could be detected in maternal endothelial cells or fetal syncytiotrophoblast of the placental labyrinth. Fetuses were positive for virus and viral replication at high levels in a wide range of tissues. Possible routes of transplacental transmission of ADV and the role of trophoblast cells as targets for viral replication are discussed.  相似文献   

6.
Aleutian disease virus (ADV) infection was analyzed in vivo and in vitro to compare virus replication in cell culture and in mink. Initial experiments compared cultures of Crandell feline kidney (CRFK) cells infected with the avirulent ADV-G strain or the highly virulent Utah I ADV. The number of ADV-infected cells was estimated by calculating the percentage of cells displaying ADV antigen by immunofluorescence (IFA), and several parameters of infection were determined. Infected cells contained large quantities of viral DNA (more than 10(5) genomes per infected cell) as estimated by dot-blot DNA-DNA hybridization, and much of the viral DNA, when analyzed by Southern blot hybridization, was found to be of a 4.8-kilobase-pair duplex monomeric replicative form (DM DNA). Furthermore, the cultures contained 7 to 67 fluorescence-forming units (FFU) per infected cell, and the ADV genome per FFU ratio ranged between 2 X 10(3) and 164 X 10(3). Finally, the pattern of viral antigen detected by IFA was characteristically nuclear, although cytoplasmic fluorescence was often found in the same cells. Because no difference was noted between the two virus strains when cultures containing similar numbers of infected cells were compared, it seemed that both viruses behaved similarly in infected cell culture. These data were used as a basis for the analysis of infection of mink by virulent Utah I ADV. Ten days after infection, the highest levels of viral DNA were detected in spleen (373 genomes per cell), mesenteric lymph node (MLN; 750 genomes per cell), and liver (373 genomes per cell). In marked contrast to infected CRFK cells, the predominant species of ADV DNA in all tissues was single-stranded virion DNA; however, 4.8-kilobase-pair DM DNA was found in MLN and spleen. This observation suggested that MLN and spleen were sites of virus replication, but that the DNA found in liver reflected sequestration of virus produced elsewhere. A final set of experiments examined MLN taken from nine mink 10 days after Utah I ADV infection. All of the nodes contained ADV DNA (46 to 750 genomes per cell), and although single-stranded virion DNA was always the most abundant species, DM DNA was observed. All of the lymph nodes contained virus infectious for CRFK cells, but when the genome per FFU ratio was calculated, virus from the lymph nodes required almost 1,000 times more genomes to produce an FFU than did virus prepared from infected cell cultures.(ABSTRACT TRUNCATED AT 400 WORDS)  相似文献   

7.
Strand-specific hybridization probes were utilized in in situ molecular hybridization specifically to localize replicative form DNA of Aleutian mink disease parvovirus (ADV). Throughout in vitro infection, duplex replicative form DNA of ADV was located in the cell nuclei. Single-stranded virion DNA and capsid proteins were present in the nuclei early in infection, but were later translocated to the cytoplasm. In neonatal mink, ADV causes acute interstitial pneumonia, and replicative forms of viral DNA were found predominantly in alveolar type II cells of the lung. Viral DNA was also found in other organs, but strand-specific probes made it possible to show that most of this DNA represented virus sequestration. In addition, glomerular immune complexes containing intact virions were detected, suggesting that ADV virions may have a role in the genesis of ADV-induced glomerulonephritis.  相似文献   

8.
The genomes of canine parvovirus and mink enteritis virus were compared by restriction enzyme analysis of their replicative-form DNAs. Of 79 mapped sites, 68, or 86%, were found to be common for both types of DNA, indicating that canine parvovirus and mink enteritis virus are closely related viruses. Whether they evolved from a common precursor or whether canine parvovirus is derived from mink enteritis virus, however, cannot be deduced from our present data.  相似文献   

9.
We suppressed the B-cell development and antibody response in mink by using treatment with polyclonal anti-immunoglobulin M (anti-IgM) to study the effects of antiviral antibodies on development of Aleutian mink disease parvovirus (ADV)-induced disease in more detail. Newborn mink kits were injected intraperitoneally with 1 mg of either anti-IgM or a control preparation three times a week for 30 to 34 days. At 21 days after birth, groups of mink kits were infected with the highly virulent United isolate of ADV. At selected time points, i.e., postinfection days 9, 13, 29, and 200, randomly chosen mink kits were sacrificed, and blood and tissues were collected for analyses. The efficacy of immunosuppressive treatment was monitored by electrophoretic techniques and flow cytometry. Effects of treatment on viral replication, on viral mRNA levels, and on development of acute or chronic disease were determined by histopathological, immunoelectrophoretic, and molecular hybridization techniques. Several interesting findings emerged from these studies. First, antiviral antibodies decreased ADV mRNA levels more than DNA replication. Second, suppression of B-cell development and antibody response in mink kits infected at 21 days of age resulted in production of viral inclusion bodies in alveolar type II cells. Some of these kits showed mild clinical signs of respiratory disease, and one kit died of respiratory distress; however, clinical signs were seen only after release of immunosuppression, suggesting that the production of antiviral antibodies, in combination with the massive amounts of free viral antigen present, somehow is involved in the induction of respiratory distress. It is suggested that the antiviral antibody response observed in mink older than approximately 14 days primarily, by a yet unknown mechanism, decreases ADV mRNA levels which, if severe enough, results in restricted levels of DNA replication and virion production. Furthermore, such a restricted ADV infection at low levels paves the way for a persistent infection leading to immunologically mediated disease. The potential mechanisms of antibody-mediated restriction of viral mRNA levels and mechanisms of disease induction are discussed.  相似文献   

10.
Information was sought on the temporal replication of Aleutian disease virus in 27 royal pastel mink. Groups of three were examined 8 to 126 days after they were inoculated subcutaneously with 10(3) 50% lethal doses of the Pullman strain. Much individual variation was noted in the onset of infection, occurrence of viremia, and extent of virus replication in the tissues. Thus, virus was detected in lymph nodes regional to the site of inoculation in only some mink during the first 14 days after inoculation. During this period, virus was often present as well in the mesenteric lymph node and spleen. First detected on day 10, viremia was present in all mink examined on day 28 but occurred irregularly thereafter, even when virus was widespread in the tissues. Except in five mink succumbing to the disease, the tissue distribution of virus after day 28 tended to be more limited, and the titers were generally lower than they had been earlier. Even though present in the lymph nodes and spleen, virus was often absent from the kidney, liver, and intestine after day 28. Specific antibody was detected on day 28 and was present in all mink thereafter, ostensibly without any adverse effect on virus replication. In most mink, the infection was considered subclinical, for it was usually not accompanied by a rise in serum gamma globulin or by morphologic evidence of the disease. The virologic findings in this study have a bearing on the relationship of subclinical infections to both horizontal and vertical transmission of the virus.  相似文献   

11.
Aleutian disease virus (ADV) of mink is a nondefective parvovirus with a single-stranded DNA genome. We characterized the viral DNA forms found in infected cells prepared by a modified Hirt extraction procedure. Double-stranded DNA molecules corresponding in size to 4.8-kilobase-pair duplex monomers and 9.6-kilobase-pair duplex dimers were identified in agarose gels by blot hybridization to 32P-labeled ADV DNA. A rapidly reannealing ADV duplex monomer was isolated on a preparative scale and physically mapped with a series of restriction endonucleases. The map derived was similar to one derived from double-stranded ADV DNA produced by self-primed synthesis on virion DNA, but differed from restriction endonuclease maps reported for other parvovirus DNAs. The purified duplex monomer could be labeled with [32P]dCTP by nick translation and used as a probe in blot hybridization to detect ADV sequences in DNA from small numbers of infected cells. Additional studies indicated that double-stranded ADV DNA could first be detected at 24 h after infection.  相似文献   

12.
By using strand-specific in situ hybridization and immunohistochemistry, evidence for replication of the Aleutian mink disease parvovirus was observed in cells resembling macrophages and cells resembling follicular dendritic cells at 10 days after infection but only in macrophages at 60 days. Sequestration of the Aleutian mink disease parvovirus in larger numbers of macrophages and follicular dendritic cells was noted at both 10 and 60 days.  相似文献   

13.
W J Hadlow  R E Race    R C Kennedy 《Journal of virology》1987,61(10):3235-3240
Information was sought on the temporal distribution of transmissible mink encephalopathy virus in royal pastel mink inoculated subcutaneously with 10(3.0) 50% intracerebral lethal doses of the Idaho strain. As determined by intracerebral assay in mink, extremely little replication of the virus occurred during the preclinical stage of infection. It seemed largely limited to lymph nodes draining the site of inoculation. Virus first appeared in the central nervous system (CNS) at 20 weeks, when all mink were still clinically normal. Early spongiform degeneration, limited to the posterior sigmoid gyrus of the frontal cortex, was first found at 28 weeks, or a few weeks before onset of clinical disease in most of the mink. Once virus reached the CNS, where greater concentrations occurred than elsewhere, it appeared in many extraneural sites (spleen, liver, kidney, intestine, mesenteric lymph node, and submandibular salivary gland). These seemingly anomalous findings, especially the limited extraneural replication of virus as a prelude to infection of the CNS, suggest that mink are not natural hosts of the virus. The results of this study support the generally held view that transmissible mink encephalopathy arises from chance or inadvertent infection of ranch mink with an exogenous virus, most likely feed-borne wild scrapie virus.  相似文献   

14.
Mink enteritis virus (MEV) is one of the most important pathogens in the mink industry. Recent studies have shed light into the role of microRNAs (miRNAs), small noncoding RNAs of length ranging from 18–23 nucleotides (nt), as critical modulators in the host-pathogen interaction networks. We previously showed that miRNA miR-181b can inhibit MEV replication by repression of viral non-structural protein 1 expression. Here, we report that two other miRNAs (miR-320a and miR-140) inhibit MEV entry into feline kidney (F81) cells by downregulating its receptor, transferrin receptor (TfR), by targeting the 3′ untranslated region (UTR) of TfR mRNA, while being themselves upregulated.  相似文献   

15.
Aleutian disease virus (ADV), an autonomous parvovirus, persistently infects mink and induces very high levels of virus-specific antibody. All strains of ADV infect all mink, but only highly virulent strains cause progressive disease in non-Aleutian mink. The development of antibody to individual ADV proteins was evaluated by Western blotting by using the sera of 22 uninfected mink and 163 naturally or experimentally infected mink. ADV has virion proteins of 86,000 and 78,000 daltons that are closely related. A new, possibly nonvirion protein of 143,000 daltons was observed, as well as a known nonvirion protein of 71,000 daltons. Sera from mink experimentally or naturally infected with ADV of high or low virulence generally reacted about equally with all four proteins. The only exceptions noted were that 8 of 15 sera of mink infected transplacentally preferentially reacted with the two virion proteins and sera from mink with the monoclonal gammopathy of Aleutian disease reacted preferentially with either virion (10 of 12) or nonvirion (2 of 12) proteins.  相似文献   

16.
A previously described type virus stock (designated PP-1R), isolated by cocultivating baboon cells with mink cells transformed by Kirsten sarcoma virus (64J1), has been further cloned and characterized. End point-diluted stocks of PP-1R have been obtained that are free of focus-forming activity and lack both Kirsten sarcoma and primate type C viral sequences. Nucleic acid hybridization experiments show that the cloned virus (MiLV) is an endogenous, genetically transmitted virus of the mink (Mustela vison). MiLV replicates in canine, feline, and 64J1 mink cells but not in an untransformed mink cell line. Multiple viral gene copies can be detected in the DNA of normal mink cells in culture and in normal mink tissues; related endogenous viral genes are also detected in several related Mustela species. The virus codes for a p30 protein very closely related antigenically to that of feline leukemia virus but contains p15 and p12 proteins that are antigenically distinct. The mink cell line, Mv1Lu, and its Kirsten sarcoma-transformed derivatives, 64J1, express relatively low levels of type C viral RNA related to MiLV and normally do not produce detectable levels of MiLV p30 protein or complete, infectious viral particles. Infection of sarcoma virus-transformed mink cells with baboon type C virus, however, can augment the level of expression of endogenous mink viral RNA and can result in the synthesis and packaging of mink viral RNA and p30 antigen in extracellular virions. Since the Mv1Lu cell line and its tranformed derivatives have become widely used in studies of retroviruses, the possibility of activating endogenous mink viral genes should be considered by investigators working with these cells.  相似文献   

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【目的】本研究旨在研究水貂肠炎病毒(mink enteritis virus,MEV)的基因组遗传进化特征。【方法】对采自山东境内水貂养殖场的109份水貂腹泻样品进行MEV的分离和鉴定,利用血凝和血凝抑制试验、多步生长曲线绘制以及蛋白的三级结构模拟等,对分离毒株生物学特性进行分析,通过重叠PCR对分离株进行全基因扩增,使用MegAlign进行序列同源性比对分析,利用DNAMANV6对基因组5’末端和3’末端回文结构进行预测,应用MEGAV6进行遗传进化分析。【结果】共分离得到5株病毒,经电镜观察和间接免疫荧光试验鉴定为MEV毒株,分别命名为MUTQS-1-5,GenBank登录号分别为OK275645、OK275646、OK275647、OK275648和OK275649;各分离株5’-和3’-UTR分别由长回文序列组成,具有典型的细小病毒基因组末端的茎环样结构,NS1和VP2基因的推导氨基酸序列存在多个非同义突变位点,其中NS1蛋白的E/Q545V位氨基酸突变,以及VP2蛋白的F267Y、Y324I位氨基酸突变为首次在MEV上发现;生物学特性分析表明,上述突变并未明显改变病毒的血凝及...  相似文献   

19.
Neonatal mink kits infected with Aleutian mink disease parvovirus (ADV) develop an acute interstitial pneumonia with clinical symptoms and pathological lesions that resemble those seen in preterm human infants with respiratory distress syndrome and in human adults with adult respiratory distress syndrome. We have previously suggested that ADV replicates in the alveolar type II epithelial cells of the lung. By using double in situ hybridization, with the simultaneous use of a probe to detect ADV replication and a probe to demonstrate alveolar type II cells, we now confirm this hypothesis. Furthermore, Northern (RNA) blot hybridization showed that the infection caused a significant decrease of surfactant-associated protein C mRNA produced by the alveolar type II cells. We therefore suggest that the severe clinical symptoms and pathological changes characterized by hyaline membrane formation observed in ADV-infected mink kits are caused by a dysfunction of alveolar surfactant similar to that observed in respiratory distress syndrome in preterm infants. However, in the infected mink kits the dysfunction is due to the replication of ADV in the lungs, whereas the dysfunction of surfactant in preterm infants is due to lung immaturity.  相似文献   

20.
Infection of young adult BALB/cByJ mice with mouse parvovirus-1, a newly recognized, lymphocytotropic, nonpathogenic parvovirus, was examined by in situ hybridization. Virus appeared to enter through the small intestine and was disseminated to the liver and lymphoid tissues. Strand-specific probes detected virion DNA in a consistently larger number of cells than replicative forms of viral DNA and/or viral mRNA. The number of signal-positive cells in the intestinal mucosa, lymph nodes, spleen, and thymus increased through day 10 after oral inoculation but decreased after seroconversion. Positive cells were still detected, however, in peripheral lymphoid tissues of mice examined at 9 weeks postinoculation. The results underscore the need to assess potential effects of persistent mouse parvovirus-1 infection on immune function in mice.  相似文献   

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