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1.
A plasmid, pG3BS, containing a cDNA clone from the 5' coding region of the peplomer glycoprotein gene appears to be specific for enteric transmissible gastroenteritis virus (TGEV) strains and for live-attenuated TGEV vaccines. This cDNA probe is used to differentiate porcine respiratory coronavirus (PRCV) isolates from TGEV field and vaccine strains by a slot blot hybridization assay. Probe pG3BS also hybridizes to canine coronavirus (CCV) RNA but does not hybridize to antigenically related feline infectious peritonitis virus (FIPV) RNA. The RNAs of 13 enteric TGEV isolates from the United States, Japan, and England, 4 US-licensed live-attenuated TGEV vaccines, and antigenically closely related CCV were detected by pG3BS. The RNAs of FIPV and 3 US isolates of PRCV did not react with pG3BS but were detected by a TGEV-derived plasmid, pRP3. Pigs infected with either PRCV or TGEV test serologically positive for TGEV antibody by the serum neutralization test. Characterization of the virus circulating in a swine herd by the pG3BS probe will differentiate between an enteric TGEV and a respiratory PRCV infection.  相似文献   

2.
We detected transmissible gastroenteritis virus (TGEV) antibodies in pig farms in Tochigi prefecture, although the farms had no past record of TGEV vaccination or TGE. Among the farms, Farm A showed a high antibody incidence. We could not confirm if either TGEV or porcine respiratory coronavirus (PRCV) induced the antibodies, since conventional tests failed to discriminate PRCV from TGEV. Therefore, we conducted virological and serological examinations of this farm for 4 years to establish the etiology - TGEV or PRCV. Although no TGEV was detected, PRCVs were isolated from the nasal samples of pigs. Using a commercial ELISA kit, it was found that the antibodies detected in pigs of all the raising stages and sows were raised against PRCV but not TGEV. The phylogenetic analysis of the nucleotide sequences of the isolates showed that they were closely related to each other, and formed a separate cluster apart from the U.S.A. and European strains. In Cesarean-derived, colostrums-deprived piglets inoculated with a PRCV isolate, no clinical signs were seen, and the viruses were mainly isolated from the nasal samples. Moreover, viral genes were detected from the nasal sample of the contact pig. The result suggested that PRCV infection was located in the nasal cavity of pigs, and horizontal transmission easily occurs. From these results, PRCVs with different origins from the exotic PRCVs might be prevalent in pig farms in Japan.  相似文献   

3.
A commercially available blocking ELISA was analyzed for its ability to identify antibodies to porcine coronaviruses (transmissible gastroenteritis virus [TGEV] or porcine respiratory coronavirus [PRCV]), to differentiate antibodies to TGEV and PRCV, and to identify TGEV-infected herds. Nine sera from uninfected pigs, 34 sera from 16 pigs experimentally infected with TGEV, and sera from 10 pigs experimentally infected with PRCV were evaluated using both the TGEV/PRCV blocking ELISA and a virus neutralization (VN) assay. The ELISA was not consistently effective in identifying pigs experimentally infected with TGEV until 21 days postinfection. Sera from 100 commercial swine herds (1,783 sera; median 15 per herd) were similarly evaluated using both tests. Thirty of these commercial herds had a clinical history of TGEV infection and a positive TGEV fluorescent antibody test recorded at necropsy within the last 35 months, while 70 herds had no history of clinical TGEV infection. The blocking ELISA and the VN showed good agreement (kappa 0.84) for the detection of porcine coronavirus antibody (TGEV or PRCV). The sensitivity (0.933) of the ELISA to identify TGEV-infected herds was good when considered on a herd basis. The ELISA was also highly specific (0.943) for the detection of TGEV-infected herds when the test results were evaluated on a herd basis. When sera from specific age groups were compared, the ELISA identified a greater proportion (0.83) of pigs in herds with TGEV antibody when suckling piglets were used. In repeatability experiments, the ELISA gave consistent results when the same sera were evaluated on different days (kappa 0.889) and when sera were evaluated before and after heating (kappa 0.888). The blocking ELISA was determined to be useful for herd monitoring programs and could be used alone without parallel use of the VN assay for the assessment of large swine populations for the detection of TGEV-infected herds.  相似文献   

4.
A competitive inhibition ELISA was developed to detect non-neutralizing antibodies to the peplomer protein of transmissible gastroenteritis virus (TGEV) in porcine sera using a monoclonal antibody as an indicator. It was demonstrated that field strains of the TGEV-related porcine respiratory coronavirus (PRCV) did not induce this antibody, whereas the Miller strain and field strains of TGEV did. The sensitivity of the competitive inhibition ELISA appeared to be similar to that of the virus neutralization (VN) test. The test enables differentiation of pigs which were previously infected with TGEV or PRCV and which cannot be distinguished by the classical anti-TGEV neutralization test. The present test is useful for selective serodiagnosis.  相似文献   

5.
Eight nine-week-old specific-pathogen-free pigs which had been infected with the transmissible gastroenteritis virus (TGEV)-related porcine respiratory coronavirus (PRCV) and four uninfected littermates were challenged with TGEV. The previous PRCV infection failed to protect them against the enteric TGEV infection. Virus excretion in faeces was detected by an ELISA in all the pigs for three to six consecutive days after inoculation. Although little diarrhoea was observed, the infection extended through much of the small intestine of one of the previously infected pigs four days after inoculation. Challenge with TGEV caused a secondary neutralising antibody response. By using a peroxidase conjugate of a monoclonal antibody which recognises a specific antigenic site on TGEV, antibodies against TGEV could be distinguished from antibodies against PRCV in an ELISA blocking test.  相似文献   

6.
Five nonneutralizing monoclonal antibodies (MAb) generated to the virulent Miller strain of transmissible gastroenteritis virus (TGEV) and specific for the S protein were characterized. Competition assays between purified and biotinylated MAb indicated that MAb 75B10 and 8G11 mapped near a new subsite, designated V and 2 MAb, 44C11 and 45A8, mapped to a previously designated subsite D. A fifth MAb mapped between subsites V and E. These MAb were tested with 3 previously characterized MAb to subsites A, E, and F in fixed-cell ELISA and cell culture immunofluorescent assays against 5 reference and 9 field strains of TGEV and 2 US strains (ISU-1 and ISU-3) of porcine respiratory coronavirus (PRCV). Subsites A, E, and F were conserved on all TGEV and PRCV strains examined. The 2 MAb to subsite V, 8G11 and 75B10, reacted only with the Miller TGEV strains (M5C, M6, and M60), except that 75B10 also recognized field strain U328. The MAb 11H8 did not react with 4 field strains or the Purdue strains of TGEV. The 2 MAb to subsite D reacted with all TGEV strains examined, but not with 2 US PRCV strains, 2 European PRCV strains, 1 feline infectious peritonitis virus strain, and 1 canine coronavirus strain. Because of this specificity for TGEV, but not PRCV, these latter 2 subsite D MAb may be useful for the development of competition ELISA to differentiate serologically between TGEV and PRCV infections in swine, similar to the currently used European subsite D MAb.  相似文献   

7.
用固定细胞阻断酶联免疫吸附试验（ＥＬＩＳＡ）对来自丹麦猪的１８０份血清进行了猪传染性胃肠炎病毒（ＴＧＥＶ）和猪呼吸道冠状病毒（ＰＲＣＶ）感染的鉴别诊断，共检出了ＰＲＣＶ抗体阳性血清１０７份（５９．４％），ＴＧＥＶ抗体阳性血清０份。同时也检测了一些来自国内不同ＴＧＥＶ感染类型的猪场血清。该鉴别诊断方法在我国的建立和应用为从ＰＲＣＶ阳性国家进口猪的ＴＧＥ的检疫提供了一条有效途径。  相似文献   

8.
The spike (S) glycoprotein of the Miller strain of transmissible gastroenteritis virus (TGEV) was recently cloned and expressed in baculovirus. The recombinant S protein was used as the coating antigen in a competition (blocking) enzyme-linked immunosorbent assay (ELISA) in combination with monoclonal antibodies to the S protein epitope A (conserved on TGEV and porcine respiratory coronavirus [PRCV]) or epitope D (present on TGEV only) to differentiate PRCV- from TGEV-induced antibodies. One set (set A) of 125 serum samples were collected at different times after inoculation of caesarean-derived, colostrum-deprived (n = 52) and conventional young pigs (n = 73) with 1 of the 2 porcine coronaviruses or uninoculated negative controls (TGEV/PRCV/negative = 75/30/20). A second set (set B) of 63 serum samples originated from adult sows inoculated with PRCV and the recombinant TGEV S protein or with mock-protein control and then exposed to virulent TGEV after challenge of their litters. Sera from set A were used to assess the accuracy indicators (sensitivity, specificity, accuracy) of the fixed-cell blocking ELISA, which uses swine testicular cells infected with the M6 strain of TGEV as the antigen source (ELISA 1) and the newly developed ELISA based on the recombinant S protein as antigen (ELISA 2). The sera from set B (adults) were tested for comparison. The plaque reduction virus neutralization test was used as a confirmatory test for the presence of antibodies to TGEV/PRCV in the test sera. The accuracy indicators for both ELISAs suggest that differential diagnosis can be of practical use at least 3 weeks after inoculation by testing the dual (acute/convalescent) samples from each individual in conjunction with another confirmatory (virus neutralization) antibody assay to provide valid and complete differentiation information. Moreover, whereas ELISA 1 had 10-20% false positive results to epitope D for PRCV-infected pigs (set A samples), no false-positive results to epitope D occurred using ELISA 2, indicating its greater specificity. The progression of seroresponses to the TGEV S protein epitopes A or D, as measured by the 2 ELISAs, was similar for both sets (A and B) of samples. Differentiation between TGEV and PRCV antibodies (based on seroresponses to epitope D) was consistently measured after the third week of inoculation.  相似文献   

9.
根据猪传染性胃肠炎病毒和猪呼吸道冠状病毒的S基因核苷酸序列，设计合成了2对引物，以猪传染性胃肠炎病毒和猪呼吸道冠状病毒细胞培养物为模板，进行PCR特异性片段扩增，猪传染性胃肠炎病毒扩增片段大小为886bp，猪呼吸道冠状病毒扩增片段大小为214bp。建立的套式PCR经过特异性、敏感性试验及对临床送检样品检测，证明本法具有快速、特异和高度敏感的特点。  相似文献   

10.
为了比较冠状病毒基因相关性,获得特异基因克隆制备冠状病毒基因芯片,根据发布的基因序列,每种病毒设计4~17对引物,利用火鸡冠状病毒(TCV)原毒和蔗糖密度梯度离心纯化浓缩的犬冠状病毒(CCV)、猫冠状病毒(FCV)、猫传染性腹膜炎病毒(FIPV)、猪传染性胃肠炎病毒(TGEV)、猪呼吸道冠状病毒(PRCV)、牛冠状病毒(BCV)细胞毒,提取总RNA并反转录和PCR扩增。回收PCR产物连接pGEM-T-easy载体并转化大肠杆菌TGI,经PCR鉴定后测序。将所有基因片段的核苷酸序列和推导的氨基酸序列,分别与GenBank有关病毒相关基因片段的核苷酸序列进行分析比较,确定它们的同源性。通过对不同冠状病毒不同基因片段的克隆和测序,发现同一群冠状病毒核苷酸序列间具有较高的同源性。  相似文献   

11.
A respiratory variant of transmissible gastroenteritis virus (TGEV), designated PRCV-Ind/89, was isolated from a swine breeding stock herd in Indiana. The virus was readily isolated from nasal swabs of pigs of different ages and induced cytopathology on primary porcine kidney cells and and on a swine testicular (ST) cell line. An 8-week-old pig infected oral/nasally with the respiratory variant and a contact pig showed no signs of respiratory or enteric disease. These pigs did not shed virus in feces but did shed the agent from the upper respiratory tract for approximately 2 weeks. Baby pigs from 2 separate litters (2 and 3 days old) also showed no clinical signs following oral/nasal inoculation with PRCV-Ind/89. In a third litter, 5 of 7 piglets (5 days old) infected either oral/nasally or by stomach tube developed a transient mild diarrhea with villous atrophy. However, virus was not isolated from rectal swabs or ileal homogenates of these piglets, and viral antigen was not detected in the ileum by fluorescent antibody staining even though the virus was easily recovered from nasal swabs and lung tissue homogenates. Swine antisera produced against PRCV-Ind/89 or enteric TGEV cross-neutralized either virus. In addition, an anti-peplomer monoclonal antibody, 4F6, that neutralizes TGEV also neutralized the PRCV-Ind/89 isolate. Radioimmunoassays with a panel of monoclonal antibodies indicated that the Indiana respiratory variant and the European PRCV are antigenically similar.  相似文献   

12.
Pigs were inoculated with various strains of transmissible gastroenteritis virus (TGEV) or with porcine respiratory coronavirus (PRCV), and antigenic site-specific antibody responses were compared. A blocking-ELISA was used to study to what extent antibodies in convalescent sera interfered with the binding of monoclonal antibodies (MAB) 57.16 or 57.110 to the attenuated TGEV/Purdue virus. Monoclonal antibody 57.16 is directed against the A site on the peplomer, neutralizes virus, and recognizes TGEV and PRCV. Monoclonal antibody 57.110 is directed against the X site on the peplomer, but does not neutralize virus, and recognizes only TGEV. Antibodies directed against TGEV and PRCV could be detected in a blocking ELISA, using MAB 57.16 as a conjugate. Antibodies directed against both viruses were detectable as early as 1 week after inoculation. Antibody titers correlated well with those in a virus-neutralization test. Antibodies against TGEV could be detected in a blocking ELISA, using MAB 57.110 as a conjugate. Such antibodies were not induced by a PRCV infection. In the blocking ELISA, using MAB 57.110 as a conjugate, antibodies were detectable as early as 2 weeks after inoculation. There was a significant difference between antibody titers reached after infection with various TGEV strains, however. This difference is ascribed to a variation of the antigenic site defined by MAB 57.110 in TGEV strains. Conditions for a differential test for TGE serodiagnosis, and for serologic discrimination between TGEV- and PRCV-infected pigs, are discussed.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

13.
采用蔗糖密度梯度离心,纯化浓缩犬冠状病毒(CCV)、猫冠状病毒(FCV)、猫传染性腹膜炎病毒(FIPV)、猪传染性胃肠炎病毒(TGEV)、猪呼吸道冠状病毒(PRCV)的细胞培养物,分别设计7,17,11,10和4对引物,构建了49个基因片段的克隆。煮沸裂解法制备质粒DNA,回收PCR扩增产物,点制冠状病毒基因芯片。抽提病毒总RNA,利用Cy3-dCTP随机渗入反转录PCR标记,与芯片进行杂交检测,淘汰交叉的克隆片段。结果表明:克隆CCV1,CCV2,CCV5和CCV7可特异诊断CCV,克隆FCV6,FCV7,FCV8和FCV9可特异诊断FCV,克隆FIPV2,FIPV7,FIPV8和FIPV9可特异诊断FIPV,克隆PRCV1,PRCV2和PRCV3可特异诊断PRCV,克隆TGEV3,TGEV4,TGEV5和TGEV6可特异诊断TGEV。将这些特异克隆扩增片段重新点制基因芯片,与病毒PCR产物杂交,未发现交叉现象。基因芯片检测比传统PCR敏感1000倍,可有效应用于这5种动物冠状病毒的检测与区分。  相似文献   

14.
Eighteen litters of sucking piglets were challenged with one of two strains of transmissible gastroenteritis virus (TGEV). During pregnancy, their seronegative dams had been either inoculated intranasally with porcine respiratory coronavirus (PRCV), inoculated orally with TGEV or left untreated. On the basis of weight gain, clinical signs and survival, no differences in response to challenge was detected when piglets suckled by PRCV inoculated sows were compared with those suckled by uninoculated sows. Such a difference was evident when the litters of sows successfully pre-immunized with TGEV were compared with those of unicoculated or PRCV-inoculated sows. The possibility of transplacental transmission of PRCV was investigated in two litters born to sows that had been inoculated with this virus in late pregnancy. All sixteen live-born piglets were seronegative for the virus at birth and PRCV was not isolated from tissues taken from two stillborn piglets.  相似文献   

15.
用酶联免疫吸附试验对采自贵州省内88个县(市)的2906份血清进行了猪传染性胃肠炎病毒及呼吸冠状病毒抗体检测,结果检出猪传染性胃肠炎病毒抗体阳性血清12份,总体阳性率为0.41%;猪呼吸冠状病毒抗体阳性血清44份,总体阳性率为1.51%;2818份血清两种抗体均呈阴性(阴性率为96.97%),32份血清检测无效或无结论(无效率为1.10%)。  相似文献   

16.
The objective of this study was to evaluate the seroprevalence and identify the strains of swine influenza virus (SwIV), as well as the seroprevalence of porcine parvovirus (PPV), transmissible gastroenteritis virus (TGEV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine respiratory coronavirus (PRCV), porcine circovirus type 2 (PCV-2), and classical swine fever virus (CSFV) in pigs in Trinidad and Tobago (T&T). Blood samples (309) were randomly collected from pigs at farms throughout T&T. Serum samples were tested for the presence of antibodies to the aforementioned viruses using commercial ELISA kits, and the circulating strains of SwIV were identified by the hemagglutination inhibition test (HIT). Antibodies against SwIV were detected in 114 out of the 309 samples (37%). Out of a total of 26 farms, 14 tested positive for SwIV antibodies. HI testing revealed high titers against the A/sw/Minnesota/593/99 H3N2 strain and the pH1N1 2009 pandemic strain. Antibodies against PPV were detected in 87 out of the 309 samples (28%), with 11 out of 26 farms testing positive for PPV antibodies. Antibodies against PCV-2 were detected in 205 out of the 309 samples tested (66%), with 25 out of the 26 farms testing positive for PCV-2 antibodies. No antibodies were detected in any of the tested pigs to PRRSV, TGEV, PRCV, or CSFV.  相似文献   

17.
A porcine respiratory, non-enteric virus which is related to the coronavirus transmissible gastroenteritis virus (TGEV) has been isolated in pigs and in cell culture. The isolate was designated TLM 83. It has become very widespread and enzootic among the swine population in Belgium and in other swine raising countries. It causes an infection of the lungs and appears to spread by aerogenic route. It does not replicate in the enteric tract. The experimental infection in conventional and gnotobiotic pigs in isolation remains subclinical. The infection, either experimental or in the field, results in the formation of antibodies which neutralise the classical enteric TGEV. Based on this relationship, this virus is assumed to be a new TGEV-related porcine respiratory coronavirus or TGEV itself which has totally lost its tropism for the enteric tract.  相似文献   

18.
Porcine respiratory coronavirus (PRCV) was identified for the first time in Quebec, using a blocking enzyme-linked immunosorbent assay (ELISA). Unlike the virus neutralization test (VNT), this ELISA was able to distinguish transmissible gastroenteritis virus (TGEV) from PRCV. Among the 15 seropositive fattening herds from group A, sera containing PRCV antibodies represented 74.8%, whereas those with TGEV antibodies represented only 7.2%. In group B, which consisted of 15 sow herds, nine herds expressed only PRCV-specific antibodies while the other herds had animals positive for TGEV-specific antibodies.  相似文献   

19.
Combinations of porcine respiratory coronavirus (PRCV) and either of two swine influenza viruses (H1N1 or H3N2) were administered intranasally and by aerosol to six- to eight-week-old specific pathogen-free pigs. The clinical responses, gross respiratory lesions and growth performances of these pigs were studied and compared with those of single (PRCV, H1N1 or H3N2) and mock-infected animals. PRCV infection caused fever, growth retardation and lung lesions, but no respiratory symptoms. Infection with swine influenza viruses caused rather similar, mild symptoms of disease, with H1N1 infection being the least severe. Combined infections with influenza viruses and PRCV did not appear to enhance the pathogenicity of these viruses. Furthermore, viruses were isolated more frequently from tissues and nasal swabs taken from 'single' than 'dual' infected animals, suggesting a possible in vivo interference between replication of PRCV and swine influenza virus.  相似文献   

20.
本研究根据Genbank登录的猪传染性胃肠炎病毒(TGEV)N基因和S基因保守区序列设计合成了两对引物和探针,通过对荧光定量RT-PCR反应条件的优化,建立了TaqMan荧光定量RT-PCR快速检测TGEV的方法。该方法能有效地鉴别序列密切相关的猪传染性胃肠炎病毒与呼吸道冠状病毒。与常规RT-PCR试验比较表明,所建立的荧光RT-PCR检测技术快速、敏感,检测时限3个小时以内,具有很好的特异性和重复性。通过对39份临床样品进行检测,结果表明所建立的检测方法直接检测样品中猪传染性胃肠炎病毒。  相似文献   

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