共查询到20条相似文献，搜索用时 281 毫秒
1.
Kidney BA Haines DM Ellis JA Burnham ML Jackson ML 《American journal of veterinary research》2002,63(1):60-63
OBJECTIVE: To evaluate a group of vaccine site-associated sarcomas (VSS) for the presence of feline foamy virus (FeFV) DNA, using polymerase chain reaction (PCR) methods. SAMPLE POPULATION: 50 formalin-fixed paraffin-embedded (FFPE) tissue blocks from VSS of cats. PROCEDURE: DNA was extracted from FFPE sections of each tumor, and regions of the gag and pol genes of FeFV were amplified by use of PCR methods, using 1 primer set for each region. Sensitivity of the method was compared between fresh and FFPE cells, using mouse kidney tissue that was injected with FeFV-infected cultured cells and using agarose-cell pellets. Results-Feline foamy virus DNA was not detected in VSS tissues. Sensitivity of the method was 10 times greater in fresh versus FFPE mouse tissues. Sensitivity of the method in fresh FeFV-infected cultured cells versus FFPE agarose-cell pellets was equal when fixation was 24 or 48 hours and 10 times greater when fixation was 72 hours or 1 week. CONCLUSIONS AND CLINICAL RELEVANCE: A PCR-based method can be successfully applied to FFPE tissues for FeFV DNA detection. Results suggest there is no direct FeFV involvement in the pathogenesis of VSS in cats. 相似文献
2.
3.
Kidney BA Haines DM Ellis JA Burnham M Jackson ML 《American journal of veterinary research》2001,62(6):828-832
OBJECTIVE: To determine whether vaccine site-associated sarcomas (VSS) from cats contain polyomavirus antigen or DNA. SAMPLE POPULATION: 50 formalin-fixed paraffin-embedded tissue blocks of VSS from cats. PROCEDURE: Sections from each tissue block were evaluated for polyomavirus antigen by use of an avidin-biotin-complex immunohistochemical staining method, using rabbit anti-murine polyomavirus polyclonal antiserum as the primary antibody. The DNA was extracted from sections of each tissue block, and a polymerase chain reaction assay was performed, using primers designed to amplify regions of the bovine polyomavirus genome and consensus polyomavirus primers designed to detect unknown polyomaviruses. RESULTS: Polyomavirus antigen and DNA were not detected in any of the VSS. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that polyomaviruses likely do not have any direct involvement in the pathogenesis of VSS in cats. 相似文献
4.
Kidney BA Haines DM Ellis JA Burnham ML Teifke JP Czerwinski G Jackson ML 《American journal of veterinary research》2001,62(6):833-839
OBJECTIVE: To determine whether vaccine site-associated sarcomas (VSS) from cats contain papillomavirus antigen or DNA. SAMPLE POPULATION: 50 formalin-fixed paraffin-embedded tissue blocks of VSS from cats. PROCEDURE: Sections from each tissue block were evaluated for papillomavirus antigen by use of an avidin-biotin-complex immunohistochemical staining method, using rabbit anti-bovine papillomavirus type-1 antibody. The DNA was extracted from sections of each tissue block, and polymerase chain reaction assays were performed, using primers designed to amplify regions of the E5 gene of bovine papillomavirus and consensus primers designed to amplify a region of the L1 gene of animal papillomaviruses. Sections from 20 of the tissue blocks were evaluated by use of nonradioactive in situ hybridization for bovine papillomavirus DNA. RESULTS: Papillomavirus antigen and DNA were not detected in any of the VSS. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that papillomaviruses likely do not have any direct involvement in the pathogenesis of VSS in cats. 相似文献
5.
6.
T Hohdatsu M Yamada M Okada M Fukasawa K Watanabe T Ogasawara M Takagi C Aizawa M Hayami H Koyama 《Veterinary microbiology》1992,30(2-3):113-123
Feline immunodeficiency virus (FIV) proviral DNA was detected by the polymerase chain reaction method (PCR). PCR products were detected by gel electrophoresis and ethidium bromide staining. The P-10, P-15 and P-24 regions of the gag gene of FIV were chosen as the target sequences for amplification, and three primer pairs were prepared. The PCR products subjected to amplification with each primer pair were found to possess sites of digestion by a restriction enzyme, as hypothesized. They did not react with feline leukemia virus (FeLV)-infected or feline syncytium-forming virus (FeSFV)-infected cell-derived DNA, and specifically amplified FIV-infected cell-derived DNA. FIV proviral DNA was detected by the PCR method with either primer pair (one-step amplification: single PCR) in DNA derived from peripheral blood lymphocytes (PBL) from 7 of 12 FIV antibody-positive cats. When PCR products in each of the 12 cats were subjected to a second amplification using the same primer pair (two-step amplification: double PCR), FIV proviral DNA was detected in all of the cats. When PBL samples collected from three cats that were negative and three that were positive in the single PCR were cultured for a few weeks in the presence of interleukin 2, FIV proviral DNA was detected in all six cats by the single PCR method. The results suggest that either the use of cultured PBL as the sample or the performance of the double PCR method enables simple and specific detection of FIV proviral DNA in PBL. 相似文献
7.
Crawford PC Slater MR Levy JK 《Journal of the American Veterinary Medical Association》2005,226(9):1503-1507
OBJECTIVE: To determine the sensitivity, specificity, and overall diagnostic accuracy of polymerase chain reaction (PCR) assays offered by commercial diagnostic laboratories for diagnosis of FIV infection in cats. DESIGN: Prospective clinical trial. ANIMALS: 124 cats. PROCEDURE: Blood was collected from cats that were neither infected with nor vaccinated against FIV, uninfected cats that were vaccinated with a licensed FIV vaccine, and cats experimentally and naturally infected with FIV representing subtypes A, B, and C. Coded blood samples were submitted to 3 laboratories in the United States and Canada offering PCR assays for diagnosis of FIV infection to veterinary practitioners. All laboratories tested fresh blood samples, and 1 laboratory also tested samples submitted as dried blood smears. The FIV infection status in all cats was confirmed by virus isolation. Sensitivity, specificity, and correct results were calculated for each PCR assay. RESULTS: Sensitivity ranged from 41% to 93%. Specificity ranged from 81% to 100% in unvaccinated cats and 44% to 95% in cats vaccinated against FIV. Correct results were obtained in 58% to 90% of 124 cats tested. All tests misidentified both uninfected and infected cats. False-positive results by all laboratories were higher in cats vaccinated against FIV than in unvaccinated cats, suggesting that vaccination interferes with the performance or interpretation of PCR assays used for diagnosis of FIV infection. CONCLUSIONS AND CLINICAL RELEVANCE: PCR assays used for diagnosis of FIV infection presently marketed to veterinary practitioners in North America vary significantly in diagnostic accuracy and did not resolve the diagnostic dilemma resulting from vaccination of cats against FIV. 相似文献
8.
We have previously shown an absence of detectable systemic or local infection in cats exposed to an infectious (100 TCID(50)) feline immunodeficiency virus (FIV) plasma inoculum via either the rectal or vaginal mucosa. In contrast, this same plasma inoculum was infectious via parenteral inoculation. Moreover an equivalent dose of cell-free tissue culture-origin virus inoculum infected 100% of cats by either the rectal or vaginal exposure route. To evaluate this phenomena, we used a tissue culture system to identify a heat-stable factor in the plasma of cats acutely (3 weeks) infected with FIV that blocked infection of naive peripheral blood mononuclear cells (PBMC) by either cell-free or cell-associated FIV in vitro. A single application of as little as a 1:200 dilution of either heparinized or Alsevier's anticoagulated plasma effectively inhibited production of FIV p26 in culture over a 21-day co-culture period. Depletion of antibody using a protein A column abrogated the inhibitory effect of FIV plasma against in vitro FIV infection. Co-inoculation of heat-inactivated plasma with 400 TCID(50) FIV-B-2542 cell-free supernatant virus onto the vaginal mucosa of two cats resulted in complete inhibition of infection in one cat and increased time to infection in the second. Thus, antibody found in the plasma of cats acutely infected with FIV blocks cell-associated and cell-free infection, inhibits virus production in previously infected cells, and reduces mucosal transmission efficiency in vivo. Extrapolation may help explain the relatively inefficient mucosal transmission of human immunodeficiency virus-1 (HIV) and other lentiviruses. 相似文献
9.
10.
OBJECTIVE: To determine the FIV status of Australian cats with lymphosarcoma and relate this to patient characteristics, tumour characteristics (tissue involvement, histological grade and immunophenotype), haematological and serum biochemical values and FeLV status of affected cats. DESIGN: Prospective study of 101 client-owned cats with naturally-occurring lymphosarcoma. PROCEDURE: Western blot analysis, ELISA and immunochromatography were used to detect FIV antibodies in serum from cats with lymphosarcoma. RESULTS: On the basis of Western blot analysis (which was considered the most accurate method for determining FIV status), 50/101 (50%) of cats with naturally-occurring lymphosarcoma were positive for FIV antibodies. Of these 50 cats, 35 had tumours of B-cell phenotype, 13 had T-cell tumours and 2 had tumours classified as non-B/non-T. Tumours from eight of these FIV-positive cats contained FeLV gene sequences, including a 9-month-old cat with FeLV antigenaemia. Compared with FlV-negative cats with lymphosarcoma, FIV-positive cats were more likely to be domestic crossbreds (P = 0.004), male (P = 0.048) and have atypical (especially nasal) forms of lymphosarcoma (P = 0.09). Only 39 of 107 (36%) blood or sera tested using ELISA were positive for FIV antibodies (including 5 false-positives). CONCLUSIONS: The prevalence of FIV infection was considerably higher in our cohort of cats compared with series of lymphosarcoma cases from the Northern hemisphere. A positive FIV status was strongly associated with lymphosarcoma in Australian cats and it is possible that this infection may predispose to the development of lymphoid neoplasia. The presence of FIV infection would have been underestimated if commercial kits alone had been used for serology. 相似文献
11.
Nambiar PR Haines DM Ellis JA Kidney BA Jackson ML 《American journal of veterinary research》2000,61(10):1277-1281
OBJECTIVES: To investigate the role of tumor suppressor gene p53 mutation in feline vaccine site-associated sarcoma (VSS) development and to evaluate the relationship between p53 nucleotide sequence and protein expression. SAMPLE POPULATION: Formalin-fixed paraffin-embedded tissues of 8 feline VSS with dark p53 immunostaining (high p53 expression) and 13 feline VSS with faint or no staining (normal p53 expression). PROCEDURE: DNA was extracted from neoplastic and normal tissue from each paraffin block. The following 3 regions of the p53 gene were amplified by polymerase chain reaction: 379 base pair (bp) region of exon 5, intron 5, and exon 6, 108 bp region of exon 7, and 140 bp region of exon 8. Amplified p53 products were sequenced and compared with published feline p53. The p53 mutations identified were correlated with p53 mutations predicted by immunostaining. RESULTS: Neoplastic cells of 5 of 8 (62.5%) VSS that had high p53 expression harbored single missense mutations within the p53 gene regions examined.The p53 gene mutations were not detected in the 13 tumors with normal p53 immunostaining. Nonneoplastic tissues adjacent to all 21 VSS lacked mutations of these p53 gene regions. CONCLUSIONS: The p53 gene mutations were restricted to neoplastic tissue and, therefore, were unlikely to predispose to VSS. However, p53 mutations may have contributed to cancer progression in 5 of the 21 VSS. There was very good (kappa quotient = 0.67 with a confidence limit of 0.3 to 1.0), although not complete, agreement between prediction of mutation by p53 immunostaining and identification of mutations by sequencing of key p53 gene regions. 相似文献
12.
OBJECTIVE: To examine shedding of cell-free and cell-associated feline immunodeficiency virus (FIV) in semen of domestic cats during acute infection. ANIMALS: 7 specific-pathogen-free sexually intact male cats. PROCEDURE: 6 cats were inoculated IV with 5 x 10(6) 50% tissue culture infective doses of FIV-NCSU1, and 1 cat served as an uninfected (control) cat. Infection was confirmed in the 6 cats. Periodically for up to 16 weeks after inoculation, cats were anesthetized and ejaculates obtained by use of electroejaculation. Virus was isolated from filtered seminal plasma and washed seminal cells by co-cultivation with a feline CD4+ T-cell line. Seminal cell lysates were also examined for a 582-base pair segment of FIV gag provirus DNA, using a nested polymerase chain reaction amplification. RESULTS: During the acute phase of FIV infection, virus was evident in semen of 5 inoculated cats. Five cats had virus-positive seminal plasma and 3 had virus-positive cellular constituents during the study. Virus was isolated from 8/22 (36%) seminal plasma samples and 2/17 (18%) seminal cell specimens. Provirus DNA was detected in 5/24 (21%) seminal cell lysates. Cell-free virus was isolated as early as 6 weeks after inoculation, whereas cell-associated virus was isolated as early as 12 weeks after inoculation. Provirus DNA was detected in seminal cells from one cat as early as 1 week after inoculation. CONCLUSIONS AND CLINICAL RELEVANCE: Cell-free and cell-associated FIV are shed in semen of cats early during the course of infection. Samples obtained before seroconversion may contain virus. Virus shedding in ejaculates varies between and within cats during acute infection. 相似文献
13.
OBJECTIVE: To determine the subtypes of feline immunodeficiency virus (FIV) present in the domestic cat population in Melbourne. METHODS: Blood samples were collected from 42 cats that had serum antibodies against FIV. DNA was extracted and subjected to polymerase chain reaction (PCR) to amplify variable regions of the envelope (env) and group specific antigen (gag) genes of FIV. PCR products were directly sequenced or sequenced after cloning when direct sequencing yielded ambiguous results. Phylogenetic analysis was performed and comparisons made with representative sequences of different subtypes. RESULTS: The variable region of the env gene was successfully amplified by PCR from 41 of the 42 cats. All 41 were found to cluster with subtype A env sequences. The variable region of the gag gene was successfully amplified by PCR from all 42 cats. Forty-one were found to cluster with subtype A gag genes and one was found to cluster with subtype B sequences, suggesting that it may be derived from a recombinant env A/gag B virus. CONCLUSIONS: Subtype A is the predominant FIV type in Melbourne, although a subtype A/B recombinant was identified in the population of FIV positive cats. These results of env gene analysis were similar to those in a previous Australian study, suggesting that subtype A predominates in Australia. The results of the gag gene analysis show the importance of analysing multiple areas of the FIV genome when assigning FIV subtypes. Comparison with other major urban centres may provide useful information about the phylogenic diversity of FIV in Australia. 相似文献
14.
OBJECTIVES: To evaluate and compare methods for DNA extraction from formalin-fixed, paraffin-embedded tissues and methods for detection of Mycobacterium avium subsp paratuberculosis by IS900 PCR for confirmation of Johne's disease in ruminants. DESIGN: A laboratory study. PROCEDURE: Three methods of DNA extraction of differing complexity and two PCR protocols using different pairs of IS900 primers were compared. Sensitivity and specificity were assessed using samples from ruminants with and without histological evidence of Johne's disease. RESULTS: The simplest method of DNA extraction, which involved two cycles of boiling and freezing followed by centrifugation, gave more consistent results than two methods that required solvent extraction of paraffin, proteinase digestion and DNA purification. The sensitivity of detection of M avium subsp paratuberculosis in paraffin blocks stored for 1 to 6 years from 34 cases of Johne's disease in sheep, cattle and goats was 88% for a 229 bp IS900 PCR assay and 71% for a 413 bp assay, using the detection of acid-fast bacilli by Ziehl Neelsen staining of histological sections from the same blocks as the gold standard test. PCR results correlated with the abundance of acid-fast organisms in the tissues. No false positive reactions were detected. CONCLUSION: PCR for identification of M avium subsp paratuberculosis in formalin-fixed, paraffin-embedded intestinal tissues from ruminants is a rapid and useful method. A simple method of sample preparation is effective. Amplification of short fragments of IS900 is more effective than amplification of longer fragments. 相似文献
15.
16.
Arjona A Barquero N Doménech A Tejerizo G Collado VM Toural C Martín D Gomez-Lucia E 《Journal of Feline Medicine and Surgery》2007,9(1):14-22
Laboratory diagnosis of feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV) usually involves both viruses, as the clinical signs are similar and coinfection may occur. Serological methods may not represent an accurate diagnosis: maternal antibodies or cross-reactions may give false positive results to FIV, and false negative results may occur in latent FeLV status, or in certain FIV infection stages. A nested polymerase chain reaction (PCR) technique was designed to detect FeLV, FIV and feline endogenous retrovirus simultaneously. The detection of endogenous sequences was considered indicative of successful DNA extraction. The technique was used to diagnose FIV and FeLV in the blood cells of 179 cats. The kappa value with the serological data was 0.69 for FeLV and 0.87 for FIV. The joint detection of FeLV and FIV by this novel nested PCR is sensitive, specific, fast and convenient, and its applicability for clinical diagnosis is promising, as the direct evidence of the presence of the virus is more realistic than the indirect data provided by the serological detection. 相似文献
17.
Levy JK Crawford PC Slater MR 《Journal of the American Veterinary Medical Association》2004,225(10):1558-1561
OBJECTIVE: To determine the effect of vaccination against FIV on results of serologic assays for FIV infection. DESIGN: Prospective clinical trial. ANIMALS: 26 specific-pathogen-free cats, 102 laboratory-reared cats (42 unvaccinated and uninfected, 41 vaccinated and uninfected, and 19 infected with FIV), and 22 client-owned cats infected with FIV. PROCEDURE: To determine the onset and duration of anti-FIV antibody production in cats following vaccination with a whole-virus vaccine, serum was obtained from the 26 specific-pathogen-free cats prior to vaccination and weekly for 10 weeks, then monthly for 52 weeks, after vaccination; serum was tested for anti-FIV antibodies with lateral flow and microwell plate ELISAs. To determine the diagnostic performance of serologic assays for FIV infection, plasma from uninfected, unvaccinated cats; uninfected, vaccinated cats; and FIV-infected cats was tested for FIV antibodies with the 2 ELISAs, a western blot assay, and an immunofluorescence antibody assay and for FIV antigen with an ELISA. RESULTS: Anti-FIV antibodies were detected in all 26 vaccinated cats 1 year after vaccination. Sensitivity of the antibody assays for FIV infection was high (98% to 100%). Specificity was high in unvaccinated cats (90% to 100%) but poor in vaccinated cats (0% to 54%). None of the vaccinated or infected cats had detectable FIV antigen in plasma. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that vaccination against FIV causes false-positive results for at least 1 year with currently available serologic assays for FIV infection. Negative FIV antibody assay results are highly reliable for detection of uninfected cats, but positive results should be interpreted with caution. 相似文献
18.
The clinical significance of Helicobacter spp. in feline digestive organs needs to be evaluated and formalin-fixed and paraffin-embedded (FFPE) tissue samples provide an invaluable source for molecular studies. In this study, we performed a PCR assay to investigate the presence of Helicobacter DNA in digestive organs from seven cats and compared this occurrence in fresh and formalin-fixed and paraffin-embedded (FFPE) tissue samples from the same organs. The present study identified Helicobacter DNA in the pancreas, liver, stomach, and duodenum in fresh tissue samples but only in the stomach in FFPE samples. To our knowledge this is the first time that Helicobacter DNA have been identified in the feline pancreas. This study indicates that it is important to be aware of differences between results when analyzing FFPE samples compared to fresh tissue samples, especially regarding longer DNA fragments (>200 bp (base pairs)). 相似文献
19.
Pinches MD Diesel G Helps CR Tasker S Egan K Gruffydd-Jones TJ 《Veterinary clinical pathology / American Society for Veterinary Clinical Pathology》2007,36(2):141-147
BACKGROUND: Screening tests for feline retroviruses are thought to have high sensitivity and specificity, although previous studies that evaluated these tests have limitations. Novel statistical approaches have been developed that allow the estimation of sensitivity and specificity in situations where the true state of the disease in individual animals cannot be assured. OBJECTIVE: The purpose of this study was to evaluate the sensitivity and specificity of a variety of retrovirus tests, including some screening tests, in a population of cats potentially infected with either feline leukemia virus (FeLV) and/or feline immunodeficiency virus (FIV) by using a Bayesian statistical approach. METHODS: Four hundred and ninety blood samples from cats being evaluated for FIV infection were tested by 2 rapid immunomigration tests (Witness single [WS], Witness combi [WC]) and a plate-based ELISA (Petcheck) for FIV antibody, and by a newly designed real-time polymerase chain reaction (PCR) assay for FIV provirus. Four hundred and ninety-five blood samples from cats being evaluated for FeLV infection were tested by 2 rapid immunomigration tests (WS, WC) and a plate-based ELISA (Petcheck) for FeLV antigen, and by a FeLV virus isolation technique. Results were then analyzed by using a Bayesian statistical method. RESULTS: For FIV tests, median sensitivity estimates were 0.98 for WS, 0.97 for WC, 0.98 for ELISA, and 0.92 for PCR. Median specificity estimates were 0.96 for WS, 0.96 for WC, 0.93 for ELISA, and 0.99 for PCR. For FeLV tests, median sensitivity estimates were 0.97 for WS, 0.97 for WC, 0.98 for ELISA, and 0.91 for virus isolation. Median specificity estimates were 0.96 for WS, 0.96 for WC, 0.98 for ELISA, and 0.99 for virus isolation. CONCLUSIONS: The use of Bayesian statistical methods overcomes a variety of methodologic problems associated with diagnostic test evaluations, including the lack of a definitive reference test. The sensitivity and the specificity of all 6 evaluated screening tests was high: however, specificity estimates were slightly lower than those reported by most recent studies. 相似文献
20.
With the commercial release in Australia in 2004 of a vaccine against feline immunodeficiency virus (FIV; Fel‐O‐Vax FIV®), the landscape for FIV diagnostics shifted substantially. Point‐of‐care (PoC) antibody detection kits, which had been the mainstay for diagnosing FIV infection since the early 1990s, were no longer considered accurate to use in FIV‐vaccinated cats, because of the production of vaccine‐induced antibodies that were considered indistinguishable from those produced in natural FIV infections. Consequently, attention shifted to alternative diagnostic methods such as nucleic acid detection. However, over the past 5 years we have published a series of studies emphasising that FIV PoC test kits vary in their methodology, resulting in differing accuracy in FIV‐vaccinated cats. Importantly, we demonstrated that two commercially available FIV antibody test kits (Witness? and Anigen Rapid?) were able to accurately distinguish between FIV‐vaccinated and FIV‐infected cats, concluding that testing with either kit offers an alternative to PCR testing. This review summarises pertinent findings from our work published in a variety of peer‐reviewed research journals to inform veterinarians (particularly veterinarians in Australia, New Zealand and Japan, where the FIV vaccine is currently commercially available) about how the approach to the diagnosis of FIV infection has shifted. Included in this review is our work investigating the performance of three commercially available FIV PoC test kits in FIV‐vaccinated cats and our recommendations for the diagnosis of FIV infection; the effect of primary FIV vaccination (three FIV vaccines, 4 weeks apart) on PoC test kit performance; our recommendations regarding annual testing of FIV‐vaccinated cats to detect ‘vaccine breakthroughs’; and the potential off‐label use of saliva for the diagnosis of FIV infection using some FIV PoC test kits. We also investigated the accuracy of the same three brands of test kits for feline leukaemia virus (FeLV) diagnosis, using both blood and saliva as diagnostic specimens. Based on these results, we discuss our recommendations for confirmatory testing when veterinarians are presented with a positive FeLV PoC test kit result. Finally, we conclude with our results from the largest and most recent FIV and FeLV seroprevalence study conducted in Australia to date. 相似文献