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1.
Growing organ cultures prepared from foetuses experimentally infected in utero by the viruses PI-3 and BVD-MD or bovine parvovirus proved to be suitable in tests with the re-isolation of these viruses 7, 14, 21, 35, 42 and 70 days after infection. The attempts at demonstrating them by the traditional method of the inoculation of the primary cell culture of foetal kidneys were successful only at the re-isolation of the PI-3 virus seven days after infection. The PI-3 virus without cytopathic effect was demonstrated in the primary cell culture prepared from the spleen, lungs, kidneys and testes of foetuses delivered seven days from infection. The cytopathic effect manifested itself after cell transplantation in the first passage when further multiplication of the virus occurred.  相似文献   

2.
The Mengeling-Vaughn Porcine Kidney (MVPK-1) cell line, derived in October, 1970, from fetal pig kidneys, is susceptible to all 7 types of foot-and-mouth disease (FMD) virus. A plaque assay was developed for FMD virus that depended on washing MVPK-1 cells in serum-free medium before infection and excluding serum from 0.6% gum tragacanth overlay during plaque formation. The numbers of plaques that formed on MVPK-1 cells by a representative strain of each FMD type were comparable with the numbers of those on primary bovine calf kidney (BK) cells. Virus passaged in BK cell cultures did not have to be adapted to the cell line to obtain these results. The cell line lost susceptibility rapidly at 37 C after confluency was reached but retained susceptibility if maintained at room temperature. The cell line has the potential of replacing BK cells for many diverse purposes.  相似文献   

3.
In this first report of the isolation of reovirus from mink, isolates were obtained from 18 to 26 young mink with viral enteritis, by using cultures of cat kidney cells. The isolated also produced cytopathic changes in cell cultures of mink kidney, dog kidney, piglet kidney, calf kidney, bovine embryonic kidney and calf testis. A characteristic feature was the formation of eosinophilic inclusion bodies in the cytoplasma of culture cells. Haemagglutination tests were negative with erythrocytes from cat, rabbit, pig, horse and cattle. Attempts to infect old and young mink, kittens and ferrets with tissue culture material failed. It was not known to what extent this second infection with reovirus influenced the course of mink viral enteritis.  相似文献   

4.
The effects of a neonatal calf diarrhea virus on cell cultures were investigated. Bovine embryonic kidney cell cultures were the most satisfactory for production of virus. Cytoplasmic changes detected after inoculation with a high multiplicity of virus were: 1) cytoplasmic vacuoles; 2) some eosinophilic cytoplasmic inclusions; and 3) some degeneration of cells and detachment from the monolayer. Cultures stained with fluorescein-labeled antibody showed cytoplasmic fluorescence as early as four hr after infection with the maximum fluorescence at five days. No cross reactions were observed between the neonatal calf diarrhea virus and reovirus type 1 or type 3 by the fluorescent antibody technique. Plaques were small and were not produced consistently. The optimal adsorption time was one to two hr. The maximum titer was reached at 18 hr, with the cell-associated titer remaining higher than the cell-free titer until that time. An interferon was produced by cultures infected with either ultraviolet-inactivated or untreated virus.  相似文献   

5.
Immunofluorescence was used to study the virus of infectious bovine rhinotracheitis and the course of infection in cell cultures of calf kidney. An indirect relationship was found to exist between the magnitude of inoculation and the onset of specific fluorescence. Fluorescent plaques are formed as a result of inoculate dilution. The plaques will grow along with incubation time. Release of virus into the culturing medium will first lead to the formation of secondary plaques, then followed by generalised infection of the cell culture. The time at which the infection will begin to be disseminated was found to depend on both multiplicity of the infection and quality of the cell culture. Therefore, no limitation is possible of the time during which only primary infectious foci are recordable. Antibody present in the culturing medium prevent propagation of the infection, but this does not inhibit the course of primary infection nor intercellular virus transmission. The conditions are defined for the microplaque fluorescence method and its use on quantitative virus assay. While reproducible results are offered by that method, its sensitivity is below that of the tubule method, when it comes to identifying the infection due to the cytopathic effect. The microplaque fluorescence method was used to study the conditions for absorption of virus of infectious bovine rhinotracheitis by calf-kidney cell cultures. Absorption was tested under temperatures of 20 degrees C, 37 degrees C, and 40 degrees C and found to be accelerated by higher temperatures. Yet, the total quantity of virus absorbed in 120 minutes was found to be almost the same in all three temperatures. The degree of virus absorption was found to depend on the kind of medium, with the rate of absorption having been strongly increased by adding to the medium serum of different animal species. About 70 per cent of the virus present in the inoculate were absorbed by the calf-kidney cell cultures under defined experimental conditions.  相似文献   

6.
Primary cells derived from calf thyroid (CTY), calf kidney (CK) and piglet kidney (PK) were immortalised by oncogene transfection and their susceptibility to infection by foot-and-mouth disease (FMD) virus and swine vesicular disease (SVD) virus examined. Eighty-five immortalised cell lines (47 CTY, 20 CK and 18 PK) proved stable upon repeated cell culture passage and many supported the growth of FMD virus and several of the PK cell lines supported SVD virus. However, none of the immortalised lines exhibited either the degree of sensitivity or the specificity for all virus serotypes and strains as shown by primary CTY and IB-RS-2 cell cultures which are routinely employed for vesicular virus diagnosis.  相似文献   

7.
Cell culture propagation of porcine rotavirus (reovirus-like agent).   总被引：8，自引：0，他引：8  
Two isolates of porcine rotavirus (reovirus-like agent) were isolated and passaged in primary procine kidney cell cultures. Viral infectivity for cells was monitored by immunofluorescence because viral cytopathic effect was moderate. Successful passage of virus in cell culture required that viral suspensions obtained from infected cell cultures be treated with pancreatin prior to inoculation onto cell monolayers. Porcine rotavirus passage in cell culture also was accomplished, using trypsin treatments in lieu of pancreatin treatments. Porcine rotavirus passaged 10 times in cell culture infected gnotobiotic pigs and caused diarrhea. Gnotobiotic pigs that recovered from this infection were resistant to challenge exposure with porcine rotavirus but were susceptible to challenge exposure with transmissible gastroenteritis virus. As determined by immunofluorescent cross reactions, porcine rotavirus was found to be antigenically related to the human and bovine rotaviruses but not to reovirus type 3 or to transmissible gastroenteritis virus.  相似文献   

8.
This paper describes the isolation of porcine epidemic diarrhea (PED) virus in Vero and porcine cell cultures, and the influence of age on disease in experimental infection. PED virus was isolated from the small intestine of piglets inoculated with PED samples and cultured in Vero, porcine bladder and kidney cells propagated in collagen-coated tissue culture plates in maintenance medium (MM) containing trypsin. In porcine bladder and kidney cell cultures inoculated with isolated PED virus, cytopathic effects (CPE) including cell fusion were detected. Specific brilliant fluorescence was observed in the cytoplasm of these cells. Two- and 7-day old, and 2-, 4-, 8- and 12-week old specific pathogen-free (SPF) pigs were orally inoculated with PED virus isolated from an outbreak. All 2- and 7-day old pigs inoculated developed severe watery diarrhea from post-inoculation day (PID) 1 and died between PID 3 and 4. Although three of five 2-week old pigs developed diarrhea on PID 1-4, they eventually recovered. In the 4-week old group, three of five pigs had mild diarrhea for 1-2 days. None of the 8- and 12-week old pigs showed any clinical signs. Antibodies against PED virus were detected in all surviving pigs by virus neutralization (VN) test and immunofluorescence assay (IFA). Therefore, there is an age-dependent resistance to pathogenic PED virus infection in pigs.  相似文献   

9.
Three different strains of bluetongue virus were adapted to grow in primary bovine foetal kidney cell cultures. The cytopathic effects observed from the three strains were similar, and were characterized by shrinkage of cells and increased granularity. The specificity of the changes was confirmed by the fluorescent antibody technique. No significant immunological cross-reaction was detected by serum-virus neutralization tests from the strains studied.  相似文献   

10.
The direct immunoperoxidase test has been used to detect rinderpest virus antigens in infected bovine kidney cell cultures to study the multiplication of the virus. Infected bovine kidney coverslip cultures were sequentially tested with peroxidase-labelled, anti-rinderpest globulins at 3, 6, 24, 48, 72, 96, 120 and 144 h post-infection. The progressive virus-specific cytopathic changes compared well with the increase in the number of cells showing the presence of viral antigens when tested by the direct immunoperoxidase test. The specificity of the reaction was confirmed by using negative and antiserum-blocked BK cell cultures on coverslips.  相似文献   

11.
Cell cultures of embryonic calf kidney which had been infected with bovid herpes virus 2 were examined for cytological and histochemical changes. The morphological changes recorded from cells damaged by virus infection included the formation of gigant syncytial cells and intranuclear inclusions of Cowdry Type A. The cytological changes in the infected cells were accompanied by variation in enzyme activity. Recordable were rise in lactate dehydrogenase and alkaline phosphatase as well as decline in succinate dehydrogenase and acid phosphate activity. These phenomena were found to have resulted from impediment of cell metabolism by virus action.  相似文献   

12.
Bovine astrovirus serotype 2 (US2) was adapted to primary neonatal kidney cell (NBK) cultures by the addition of 50 micrograms ml-1 of trypsin in the medium. Infectious virus was released from the cells within 7 days post-infection in early passages and within 3 days in later passages. In the absence of trypsin, neither passage of infected cells nor release of infectious virus occurred. The virus was shown to be similar to the fecal astrovirus by a neutralization test and by ultrastructural studies of infected cells. Primary embryo bovine kidney (EBK) and NBK cell cultures supported infection with both fecal and tissue culture adapted (TCA) astrovirus. The time-related development of infection, as studied by immunofluorescence, was similar for both fecal and TCA astrovirus and for both cell culture types. The first indication of viral infection and expression of viral antigens occurred at 7 h post-infection and was characterized by the appearance of a diffuse faint immunofluorescence (IF) of the cytoplasm. Soon after, two or three brilliant IF granules were observed in the nucleus, which appeared to involve the nucleoli. Subsequently, dense granular IF was seen in the perinuclear region of the cytoplasm, which later extended to involve all the cytoplasmic area. In both EBK and NBK cultures infected with either fecal or tissue culture adapted astrovirus, only a minority of cells became infected, even when the multiplicity of infection exceeded one. Occasionally 10-20% of cells were infected, but in most cultures the proportion did not exceed 2% and in NBK cultures, from 3/9 calves, no infected cells were observed. The virus did not infect bovine cell lines. Infectivity of the virus was not removed by treatment with chloroform, and iododeoxyuridine and actinomycin D when added to the medium, did not block replication. Masses of virions were observed by electron microscopy in discrete areas in the cytoplasm, with similar distributions as the viral antigen foci as seen by IF. The mean diameter of the virions was 34 nm. In conclusion, bovine astrovirus lacks both essential lipids and an envelope, probably has an RNA genome, may have a nuclear phase of replication involving the nucleoli which is not blocked by DNA inhibitors, and has a selective cell tropism.  相似文献   

13.
The growth characteristics were studied in the attenuated strains of canine parvovirus CPVA-BN 80/82, mink enteritis virus MEVA-BN 63/82 and feline panleucopenia virus FPVA-BN 110/83 on the stable feline kidney cell line FE, and in the attenuated canine distemper virus CDV-F-BN 10/83 on chicken embryo cell cultures (KEB) and cultures of the stable cell line VERO. When the FE cultures were infected with different parvoviruses in cell suspension at MOI 2-4 TKID50 per cell, the first multiplication of the intracellular virus was recorded 20 hours p. i. In the canine parvovirus, the content of intracellular and extracellular virus continued increasing parallelly until the fourth day; then, from the fourth to the sixth day, the content of extracellular virus still increased whereas that of intracellular virus fell rapidly. In the case of the mink enteritis virus the release of the virus into the culture medium continued parallelly with the production of the cellular virus until the sixth day. In the case of the feline panleucopenia virus the values concerning free virus and virus bound to cells were lower, starting from the second day p. i. When KEB or VERO cultures were infected in cell suspension with the canine distemper virus at MOI about 0.004 per 1 cell, the replicated intracellular virus was first recorded in the KEB cultures five hours after infection but in the VERO cultures only 20 hours after infection, with a timely release of the virus into the culture medium in both kinds of tissue. In the KEB and VERO cultures the highest values of infection titres were recorded on the fourth day p. i., the course of virus multiplication on the cells being parallel with its release into the culture medium.  相似文献   

14.
Equine herpesviruses (EH viruses) were isolated from 9 horses in three separate outbreaks of respiratory disease. The pattern of disease in the three stables is described and evidence is presented that some of the horses were ill, possibly as a result of recurrent infection, and that reactivation of a persistent, latent infection may have occurred. An ulcerative condition of the pharyngeal region was seen in some of the horses with EH virus infection.
The cytopathogenicity for equine foetal kidney cells of the 9 EH viruses varied considerably. One isolate, EH 39 virus, which was recovered from an acute, upper respiratory tract infection, was rapidly cytopathic for equine foetal kidney cell cultures and was shown in neutralisation tests to be identical with, or closely related to equine rhinopneumonitis virus (EH virus type 1) that is associated with acute respiratory disease and abortion in other countries. More slowly cytopathic isolates were recovered from mild to subclinical upper respiratory tract infections. Evidence is presented that the property of slow cytopathogenicity is probably related to the tendency of these viruses to remain cell associated. Slowly cytopathic isolates were recovered from the nasal cavity of horse 89 on two occasions 79 days apart. One of the eight slowly cytopathic isolates, EH 86 virus, was shown to be antigenically distinct from equine rhinopneumonitis virus (EH 39 virus).  相似文献   

15.
Plaque formation, replication and related cytopathic function of 9 strains of transmissible gastroenteritis (TGE) virus were examined in primary cells and cell lines such as CPK, IB-RS-2, ESK, and PK-15 originated from porcine kidney and the effects of trypsin on the replication of TGE virus were examined in CPK cells. All strains produced a cytopathic effect and grew well in CPK cells as well as in primary porcine kidney cells. The effect of trypsin on the plaque formation was different from strains. The number of plaques produced by strains TO-163, Ukiha and Niigata increased from 2.6 to 3.52 times when trypsin was present in the medium during incubation at 37 degrees C for 1 hr after adsorption of the virus at 4 degrees C for 2 hr. The plaque sizes of TO-163, h-5, Ukiha and Niigata became larger from 1.4 to 1.7 times, when trypsin was present in the agar MEM overlay.  相似文献   

16.
OBJECTIVE: To assess the effect of interferon (IFN)-alpha on viability of feline corneal epithelial cells, replication of feline herpesvirus (FHV)-1, and virus-induced cytopathic changes. SAMPLE POPULATION: Healthy eyes from 10 recently euthanatized cats. PROCEDURE: 4 replicate primary cultures of feline corneal epithelial cells were grown after the addition of 10(2) to 10(6) IU of IFN-alpha/mL. Cultures were examined every 24 hours for evidence of cytotoxic changes. Viable cell counts and percentage of viable cells were determined 48 hours after initiation of culture. In a separate experiment, cultures of corneal cells were inoculated with FHV-1 and cultured for 72 hours with or without 10(5) IU of IFN-alpha/mL. The FHV-1-infected cultures were evaluated for viral-induced cytopathic effects, and viral titers were determined in samples of culture supernatant. RESULTS: Interferon-alpha did not have cytotoxic effects on corneal epithelial cells at concentrations ranging from 10(2) to 10(6) IU of IFN-alpha/mL. Interferon-alpha at a concentration of 10(5) IU/mL significantly reduced the cytopathic changes and FHV-1 titers. CONCLUSIONS AND CLINICAL RELEVANCE: Lack of in vitro cytotoxic effects and efficacy against FHV-1 infection in primary cultures of feline corneal cells suggests that the in vivo therapeutic effect of IFN-a should be assessed in controlled clinical trials.  相似文献   

17.
In vitro studies with organ (oviduct and trachea) and chicken embryo kidney cell cultures were attempted to assess the pathogenicity of locally isolated infectious bronchitis virus (IBV-P:120) initially isolated from the oviduct of young chicks. In oviduct cultures infected with IBV, ciliary movements decreased as early as 24 hours postinoculation (PI), and on the 6th day ciliary movements ceased completely. Cytopathic changes were also noticed. Immunofluorescent antigen was detected from 1 to 6 days PI, the maximum being on the 3rd day. The characteristic microscopic changes in the oviduct explants were reduced by 24 hours PI and had completely ceased on the 5th day. Cytopathic effect and immunofluorescent antigen were present from 1 to 8 days PI, being maximum on the 5th day. Histological changes marked by loss of cilia, rounding of the epithelial cells, degeneration, and sloughing were detected from 2 to 8 days PI. Low-embryo-passaged (EP-7) IBV did not produce cytopathic effect on the chicken embryo kidney cell cultures. On the contrary, high-embryo-passaged (EP-14) virus produced cytopathic effect at the third tissue-culture-passage level.  相似文献   

18.
J Lynch  J Swinton  J Pettit  D Key 《Avian diseases》1984,28(4):1135-1139
Attempts to isolate and identify budgerigar papovavirus (BPV) were made during three separate outbreaks of disease diagnosed on pathological grounds. Direct electron microscopy was successful only when large areas of skin were extensively disrupted to release virus and then extracted with fluorocarbon to remove lipids. Direct inoculation of budgerigar tissue suspensions into chicken embryos or chicken cell cultures failed to produce detectable virus. However, when primary cultures of liver and kidney were prepared from affected budgerigars, BPV could be detected by electron microscopy and by the production of a cytopathic effect at the third or fourth passage in cell cultures. The isolated virus was pathogenic for 10-day-old but not 11- or 12-day-old chicken embryos. Inoculated 11- and 12-day-old embryos produced antibodies to BPV that were detectable 2 weeks after hatching by agar-gel-immunodiffusion tests.  相似文献   

19.
Replication of feline infectious peritonitis virus (FIPV) in feline cell cultures was inhibited after incubation of cells with either human recombinant leukocyte (alpha) interferon (IFN) or feline fibroblastic (beta) IFN for 18 to 24 hours before viral challenge exposure. Compared with virus control cultures, FIPV yields were reduced by ranges of 0.1 to 2.7 log10 or 2 to 5.2 log10 TCID50 in cultures treated with human alpha- or feline beta-IFN, respectively; yield reductions were IFN dose dependent. Sensitivity to the antiviral activities of IFN varied with cell type; feline embryo cells had greater FIPV yield reductions than did similarly treated feline kidney or feline lung cells. Comparison of the virus growth curves in IFN-treated and virus control cultures indicated marked reduction in intracellular and extracellular FIPV in IFN-treated cultures. Compared with virus control cultures, intracellular and extracellular infectivity in IFN-treated cultures was delayed in onset by 12 and 30 hours, respectively, and FIPV titers subsequently were reduced by 3 to 3.5 and 5 log10 TCID50, respectively. Frequently, immunofluorescent and electron microscopy of IFN-treated cells or cell culture fluids did not reveal virus; however, even in cultures without viral cytopathic changes, small amounts of virus occasionally persisted in cells.  相似文献   

20.
The Lincoln strain of bovine rotavirus was found to replicate with cytopathic effects in cultures of GBK cells, a stable cell line derived from bovine kidney, when the cultures were maintained in the presence of trypsin. The virus was readily passaged and the infected cells were shown to contain specific viral antigen by indirect immunofluorescent staining. The virus formed plaques in GBK cell monolayers, when trypsin was incorporated in the agar overlay medium. The plaque count increased about twofold when diethylaminoethyl dextran was further included in the overlay medium. Plaque assay in GBK cells was more sensitive than that in MA-104 cells previously reported by Matsuno et al. The specificity of plaques was confirmed by specific inhibition with antiserum against the Lincoln strain.  相似文献   

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