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1.
对流行病学调查、临床症状检查和ELISA检测为犬瘟热阳性的自然发病犬,取肠内容物为病料,采用同步培养方法接种于犬肾细胞系(MDCK)进行病毒的分离,并对分离株进行了形态学特征、血凝特性、动物感染及RT-PCR鉴定。结果表明:病料接种MDCK细胞产生明显的细胞病变(CPE),电镜负染观察接毒细胞培养物见有典型的犬瘟热病毒粒子。分离株不凝集鸡及人“O”型红细胞,接种犬出现明显的临床症状和病理变化。用RT-PCR技术检测病毒细胞培养液,扩增出的片段长为760 bp,与预期设计的长度相同,由此确证分离株为犬瘟热病毒,命名为CDV-GZ2株。  相似文献   

2.
A morbillivirus infection of tumour cells is known to exert oncolytic activity, but the mechanism of this inhibitory action has not been well defined. Matrix metalloproteinases (MMPs) are important enzymes degrading the extracellular matrix and are often upregulated in malignant neoplasms. Recent studies have demonstrated that RECK may potently suppress MMP-2 and -9 activity, thus inhibiting angiogenesis and metastasis. In this study, real time quantitative polymerase chain reaction (RT-qPCR) was used to determine the effect of persistent infection with canine distemper virus (CDV) infection on the expression of MMPs and their inhibitors (TIMPS) in a canine macrophage/monocytic tumour cell line (DH82). The activity of proMMP-2 and proMMP-9 was also verified zymographically. Following CDV infection, MMP-2, TIMP-1 and TIMP-2 were down-regulated, while RECK was upregulated. These findings suggest that CDV infection restores RECK expression in tumour cells and may interfere with the intracellular processing of MMPs and TIMPs, thus possibly influencing tumour cell behaviour beneficially for the host. However, this needs to be verified in in vivo studies.  相似文献   

3.
Oncolytic virotherapy is a novel strategy for treatment of cancer in humans and companion animals as well. Canine distemper virus (CDV), a paramyxovirus, has proven to be oncolytic through induction of apoptosis in canine‐derived tumour cells, yet the mechanism behind this inhibitory action is poorly understood. In this study, three human mammary tumour cell lines and one canine‐derived adenofibrosarcoma cell line were tested regarding to their susceptibility to CDV infection, cell proliferation, apoptosis, mitochondrial membrane potential and expression of tumour necrosis factor‐alpha‐induced protein 8 (TNFAIP8). CDV replication‐induced cytopathic effect, decrease of cell proliferation rates, and >45% of infected cells were considered death and/or under late apoptosis/necrosis. TNFAIP8 and CDVM gene expression were positively correlated in all cell lines. In addition, mitochondrial membrane depolarization was associated with increase in virus titres (p < 0.005). Thus, these results strongly suggest that both human and canine mammary tumour cells are potential candidates for studies concerning CDV‐induced cancer therapy.  相似文献   

4.
The Yanaka strain, a field isolate of Canine distemper virus (CDV), caused extensive syncytial cytopathic effects (CPEs) followed by cell death in vitro. Syncytium formation is an important aspect of CDV pathogenicity, but the mechanism of the fusion-induced cell death is still not understood. In this study, the involvement of apoptosis in the CDV-induced CPE was investigated. We also examined apoptosis in cells infected with a persistent strain of CDV, the Yanaka-BP strain derived from the Yanaka strain, because this strain does not cause obvious CPE. DNA laddering together with Terminal transferase dUTP nick endlabeling (TUNEL) assay indicated that the Yanaka strain infection, but not the Yanaka-BP infection induced apoptosis. In addition, flow cytometric analysis similarly indicated that the Yanaka-BP strain induced apoptosis significantly less frequently than the Yanaka strain did. Thus, absence of apoptosis may be implicated in the CPE and establishment of persistent CDV infection.  相似文献   

5.
Infection of canine footpads with canine distemper virus (CDV) can result in so-called hard pad disease characterized by footpad epidermal proliferation and hyperkeratosis. Cultured canine footpad keratinocytes (CFK) were inoculated with a virulent canine distemper virus strain (A75/17-CDV) to study the effects of CDV-infection on keratinocyte proliferation. Infection was analyzed by immunohistochemistry and in situ hybridization for CDV nucleoprotein (N-protein) antigen and mRNA. CDV caused a persistent, non-cytocidal infection with spread from single cells to infection of the confluent cell layer 7 days post infection (p.i.). Absolute cell numbers were significantly higher in infected cultures compared to control cultures from day 4 until day 6 p.i. Infected cultures contained significantly more total DNA on day 5 p.i. compared to controls. Immunohistochemical investigation of proliferation markers Ki67 and BrdU demonstrated a nearly two-fold increase in numbers of positive cells on day 5 p.i. compared to controls. These findings demonstrate that canine distemper virus infection of canine footpad keratinocytes in vitro was associated with proliferation.  相似文献   

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8.
To know growth profiles of canine distemper virus (CDV) on Vero cells stably expressing canine signaling lymphocyte activation molecule (Vero-DogSLAMtag; Vero-DST cells), the propagation of three strains of CDV was tested in Vero-DST cells in comparison with parental Vero cells. Strain MD77 could grow well in both cell lines, but demonstrated no syncytium formation or indistinguishable rounding cytopathic effects (CPE) in Vero cells. Strains Onderstepoort and KDK-1 also grew well in Vero-DST cells with apparent syncytium CPE, while they grew less or no efficiently, respectively, in Vero cells. All three CDV strains demonstrated the peak titers, in Vero-DST cells before reaching to an extensive CPE and drastic decrease of titers at/after full CPE. Immunohistochemistry revealed that viral antigens of all CDV strains were found exclusively in the syncytia in Vero-DST cells, while in Vero cells, viral antigen was identified in their single cells for strain MD77 but none for other strains. Thus, every strain of CDV could grow well in Vero-DST cells and behaved differently against Vero cells. These results would be of practical value for workers of CDV because 1) In Vero-DST cells, by observation of distinct syncytium CPE, the highest titer or the best growth of virus could be identified; 2) In Vero cells, various CDV strains could be readily classified after propagation in Vero-DST cells.  相似文献   

9.
A new canine cell line, named CCT, was established from the cutaneous malignant histiocytosis in a 4-year-old male Borzoi. CCT proliferated with loose adherence and doubling time was approximately 30 hr. When co-cultured with latex beads, CCT phagocytized beads vigorously. Lysozyme and vimentin were positive by immunostaining, and non-specific esterase and acid phosphatase were positive by cytochemical staining. These features indicated the cells had a histiocytic nature. Furthermore, by subcutaneous injection to nude mice CCT could successfully form tumors with the morphological and immunohistochemical features similar to the original tumor.  相似文献   

10.
The relationship between the canine distemper virus (CDV) infection and apoptosis in the canine lymphoid tissues was investigated using immunostaining for single stranded DNA (ssDNA), TdT-mediated dUTP-biotin nick end-labeling (TUNEL) method, and electron microscopy. Twenty-six lymphoid tissues from 8 spontaneously CDV-infected dogs and 1 non-infected dog were used, and lesions were classified into 4 groups according to frequency of the CDV-antigen. Histologically, the degree of lymphoid depletion tended to depend on amount of CDV antigen. The numbers of ssDNA- and TUNEL-labeling cells were significantly high in the lymphoid tissues with abundant viral antigen. However, ssDNA- and TUNEL-positive lymphocytes were also frequently found even in the lymphoid tissues where there was only a small amount of CDV-antigen in sinus histiocytes. The incidence and distribution of apoptotic cells in the CDV-antigens-negative lymphoid tissues from infected dogs were equal to those from a non-infected dog. Double labeling immunostaining using a ssDNA and a CDV nucleocapsid protein (CDV-NP) antibody revealed that there were ssDNA positive but CDV-NP negative cells besides those stained doubly positive. Ultrastructurally, lymphocytes in the CDV-infected lymphoid tissues frequently had characteristic morphological features of apoptosis such as apoptotic bodies. All these results suggest that CDV leads to lymphocytic apoptosis directly or indirectly, resulting in severe lymphoid depletion and immunosuppression in acute or subacute phase of CDV infection.  相似文献   

11.

Backgrounds

The aim of this study was to confirm the propagation of various canine distemper viruses (CDV) in hamster cell lines of HmLu and BHK, since only a little is known about the possibility of propagation of CDV in rodent cells irrespective of their epidemiological importance.

Methods

The growth of CDV in hamster cell lines was monitored by titration using Vero.dogSLAMtag (Vero-DST) cells that had been proven to be susceptible to almost all field isolates of CDV, with the preparations of cell-free and cell-associated virus from the cultures infected with recent Asian isolates of CDV (13 strains) and by observing the development of cytopathic effect (CPE) in infected cultures of hamster cell lines.

Results

Eleven of 13 strains grew in HmLu cells, and 12 of 13 strains grew in BHK cells with apparent CPE of cell fusion in the late stage of infection. Two strains and a strain of Asia 1 group could not grow in HmLu cells and BHK cells, respectively.

Conclusion

The present study demonstrates at the first time that hamster cell lines can propagate the majority of Asian field isolates of CDV. The usage of two hamster cell lines suggested to be useful to characterize the field isolates biologically.  相似文献   

12.
为建立可以同时检测犬瘟热病毒(CDV)和犬细小病毒(CPV)的双重PCR方法,本研究根据GenBank登录的CDV N蛋白序列和CPV NS基因保守序列,设计合成2对特异性引物。通过优化反应条件,对CDV阳性病毒株反转录后的cDNA模板和CPV的DNA模板进行双重PCR扩增,同时得到2条与试验设计相符的669 bp(CDV)和392 bp(CPV)特异性条带,建立了同时检测CDV和CPV的双重PCR方法。实验结果表明:在同一PCR反应体系中可以同时检测这2种病毒,而对犬腺病毒Ⅰ型、犬腺病毒Ⅱ型、狂犬病毒检测均为阴性;CDV和CPV的最低检出限分别为101.8TCID50和101.4TCID50。采用该方法对在黑龙江省不同地区所采集的30份犬病料样品进行检测,CDV阳性率为30%;CPV阳性率为23.33%,表明建立的PCR方法可以用于临床诊断。  相似文献   

13.
Brain tissue from 33 dogs with non-suppurative encephalitis was examined for evidence of canine distemper virus (CDV) encephalitis. Sections were examined for lesions, inclusion bodies, syncytial cells and CDV antigen using a double bridge unlabelled antibody enzyme technique. Histopathological lesions considered to be typical of granulomatous meningoencephalomyelitis were found in seven dogs. They all lacked inclusion bodies, syncytial cells and CDV antigen. The remaining 26 dogs all had histopathological lesions typical of CDV encephalitis. Inclusion bodies were found in 24 dogs, four of which also had syncytial cells and CDV antigen was detected immunocytochemically in 25. One dog had no inclusion bodies or syncytial cells and was immunohistochemically negative. Syncytial cells have been found to be of limited diagnostic value for the diagnosis of CDV encephalitis. While inclusion bodies proved to be a good diagnostic criterion for the confirmation of CDV infection, the immunohistochemical demonstration of CDV antigen proved to be superior. CDV antigen was more prevalent than inclusion bodies in tissue sections and much more easily detectable.  相似文献   

14.
为研制犬副流感特异性诊断试剂,我们以犬副流感病毒(CPIV)免疫8周龄BALB/c小鼠,采用淋巴细胞杂交瘤技术获得4株稳定分泌针对CPIV的单克隆抗体(MAb)细胞株,分别命名为4F386、584C9、4G7F4和4C9D8.4株MAb腹水针对CPIV的间接ELISA抗体效价达1:10~5～1:10~6,与犬瘟热病毒(CDV)和犬细小病毒(CPV)均不发生交叉反应.MAb 4F386和4C9D8为IgG,5B4C9和4G7F4为IgM.Western blot检测表明,4F386与CPIV的F蛋白发生特异性反应,4G7F4与CPW的HN蛋白发生特异性反应,而584C9和4C9D8不与变性的CPIV蛋白发生反应.4株MAb均具有中和病毒活性,间接免疫荧光检测均呈为阳性.本研究为进一步研制CPIV特异性诊断和治疗制剂创造了条件.  相似文献   

15.
Initial non-inflammatory demyelination in canine distemper virus infection (CDV) develops against a background of severe immunosuppression and is therefore, thought to be virus-induced. However, recently we found a marked invasion of T cells throughout the central nervous system (CNS) in dogs with acute distemper despite drastic damage to the immune system. In the present study, this apparent paradox was further investigated by immunophenotyping of lymphocytes, following experimental CDV challenge in vaccinated and non-vaccinated dogs. In contrast to CDV infected, unprotected dogs, vaccinated dogs did not become immunosuppressed and exhibited a strong antiviral immune response following challenge with virulent CDV. In unprotected dogs rapid and drastic lymphopenia was initially due to depletion of T cells. In peripheral blood, CD4(+) T cells were more sensitive and depleted earlier and for a longer time than CD8(+) cells which recovered soon. In the cerebrospinal fluid (CSF) we could observe an increase in the T cell to B cell and CD8(+) to CD4(+) ratios. Thus, partial protection of the CD8(+) cell population could explain why part of the immune function in acute distemper is preserved. As found earlier, T cells invaded the CNS parenchyma in these dogs but also in the protected challenged dogs, which did not develop any CNS disease at all. Since markers of T cell activation were upregulated in both groups of animals, this phenomenon could in part be related to non-specific penetration of activated T cells through the blood brain barrier. However, in diseased animals much larger numbers of T cells were found in the CNS than in the protected dogs, suggesting that massive invasion of T cells in the brain requires CDV expression in the CNS.  相似文献   

16.
Canine distemper virus (CDV) infection induces multifocal demyelination in the central nervous system (CNS). It is thought that the resident macrophages of the CNS, the microglia, as well as invading monocytes associated with the inflammatory reaction may play a central role in the demyelinating process. To evaluate changes in peripheral monocytes in CDV infection their immunophenotype was characterized by flow cytometry during the course of an experimental CDV infection in dogs. The highest number of CDV-infected monocytes was found in dogs developing demyelinating lesions. In CD18, CD45, CD44, and CD14 neither up- nor down-regulation was observed. Marked up-regulation occurred in a number of surface molecules including CD1c, B7-1 and B7-2, MHC I, and CD11b. Peak expression was found at 4-5 weeks post-infection (PI), regardless of clinical outcome. All these molecules play an important role in the host's immune response, notably antigen presentation and cell adhesion. These results demonstrate that CDV infection in vivo may enhance several macrophage functions. This could lead to more effective clearance of the virus but may also increase demyelination through a bystander effect in animals that accumulated significant amounts of CDV in the CNS.  相似文献   

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19.
Between January 1981 and October 1985, brain tissue specimens from 192 skunks that were negative for antibodies against rabies virus were obtained from 2 Illinois Public Health laboratories (A and B). Brain lesions were detected microscopically in specimens from 17 of the 91 (18.7%) skunks from laboratory B and in specimens from 30 of the 101 (29.7%) skunks from laboratory A. Lesions in 3 skunks (1 from laboratory A, 2 from B) were caused by cerebral parasitism. Lesions in the remaining 44 skunks were characterized by perivascular, nonsuppurative, mononuclear cell infiltrates and foci of glial cells of differing severity. The similarity of lesions and the finding of inclusions diagnostic of canine distemper virus (CDV) in some skunks indicated that CDV may be the main cause of neurologic disease in nonrabid skunks. Seventeen of 36 (47.2%) skunks evaluated for antibody against CDV, using an unlabeled antibody-enzyme method, were positive for CDV. Findings in skunks from the 2 laboratories indicated similar annual prevalences of brain lesions in 1982, 1983, and 1984. The highest percentage (40.5%) of nonrabid skunks with encephalitis was found in skunks submitted to laboratory B in 1981, which was concurrent with a rabies epizootic among skunks in Illinois in 1981. The number of skunks from both laboratories with CDV infection peaked during winter-spring. Importance of CDV in skunk population dynamics remains to be elucidated; however, infection with CDV appears to be enzootic and occasionally epizootic in skunks. Because enzootic/epizootic CDV may bias rabies surveillance data, caution in interpretation of surveillance data is necessary.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

20.
Primary cultures of bovine fibroblast (BF) and canine brain cells, persistently infected with virulent R252-canine distemper virus (CDV), were cocultured with African green monkey (Vero) cells. Transfer of persistent CDV from BF to Vero cells varied inversely with the in vitro passage level (age) of the CDV-infected BF cells. Successful transfer of CDV to Vero cells was signaled by the transient appearance of viral syncytia, rapid spread of viral antigen to all Vero cells in the culture, and by recovery of cell-free Vero-infectious virus in culture fluids. With time, viral cytopathic effects in Vero cells containing CDV disappeared, and the infected lines could not be distinguished from noninfected control Vero cells, except by immunoassay for viral antigen.  相似文献   

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