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1.
Dae Sub Song Youn Jeong Lee Ok Mi Jeong Yong Joo Kim Chan Hee Park Jung Eun Yoo Woo Jin Jeon Jun Hun Kwon Gun Woo Ha Bo Kyu Kang Chul Seung Lee Hye Kwon Kim Byeong Yeal Jung Jae Hong Kim Jin Sik Oh 《Journal of veterinary science (Suw?n-si, Korea)》2009,10(4):323-329
Active serologic surveillance is necessary to control the spread of the avian influenza virus (AIV). In this study, we evaluated a commercially-available cELISA in terms of its ability to detect AIV antibodies in the sera of 3,358 animals from twelve species. cELISA detected antibodies against reference H1- through H15-subtype AIV strains without cross reactivity. Furthermore, the cELISA was able to detect antibodies produced following a challenge of the AIV H9N2 subtype in chickens, or following vaccination of the AIV H9 or H5 subtypes in chickens, ducks and geese. Next, we tested the sensitivity and specificity of the cELISA with sera from twelve different animal species, and compared these results with those obtained by the hemagglutination-inhibition (HI) test, the "gold standard" in AIV sera surveillance, a second commercially-available cELISA (IZS ELISA), or the agar gel precipitation (AGP) test. Compared with the HI test, the sensitivities and specificities of cELISA were 95% and 96% in chicken, 86% and 88% in duck, 97% and 100% in turkey, 100% and 87% in goose, and 91% and 97% in swine, respectively. The sensitivities and specificities of the cELISA in this study were higher than those of IZS ELISA for the duck, turkey, goose, and grey partridge sera samples. The results of AGP test against duck and turkey sera also showed significant correlation with the results of cELISA (R-value >0.9). In terms of flock sensitivity, the cELISA correlated better with the HI test than with commercially-available indirect ELISAs, with 100% flock sensitivity. 相似文献
2.
Jeoung HY Shin BH Lee WH Song DS Choi YK Jeong W Song JY An DJ 《Journal of Feline Medicine and Surgery》2012,14(10):746-750
To investigate the potential transmission of subtype H3 influenza virus to cats, a serological survey was carried out in South Korea. Serum samples (n = 1027) were obtained from 809 pet cats and 218 domesticated cats living in urban colonies (D-cats) from 2008 to 2010, and tested using an influenza anti-nucleoprotein (NP)-specific enzyme-linked immunosorbent assay (ELISA) and the haemagglutination inhibition (HI) test, which was recommended by the World Organization for Animal Health. Anti-influenza virus antibodies were detected in 3.12% and 2.43% of cat sera tested using the NP-specific ELISA and HI test, respectively. Anti-H3 antibodies were also identified when the HI assay was used for influenza virus serotyping. These data may indicate the sporadic transmission of subtype H3 influenza virus from other infected species to cats in South Korea. 相似文献
3.
Development of enzyme-linked immunosorbent assay with nucleoprotein as antigen for detection of antibodies to avian influenza virus 总被引：3，自引：0，他引：3
During the avian influenza outbreak of 2003-04 in Southeast Asia, two avian influenza viruses (AIV), one of H5N1 subtype and the other H9N2 subtype, were isolated and identified from local farms. The nudeoprotein (NP) gene of the H5N1 AI isolate was cloned, and the segment encoding amino acid 47-384, which covers its major antigenic domains, was subcloned and expressed in E. coli. Subsequently, the NP (47-384) expression product was purified and used as the diagnostic antigen to develop a NP-based type-specific indirect enzyme-linked immunosorbent assay (ELISA) for detecting antibodies to AI from chicken sera. The ELISA is shown to be specific for AIV and does not cross-react with chicken sera that has antibodies to other avian viruses. The NP(47-384)-ELISA was compared with a hemagglutination inhibition test and a commercial AIV ELISA kit in evaluating 150 sera samples from experimentally AIV-infected or vaccinated specific-pathogen-free (SPF) chickens. Our NP(47-384)-ELISA was more sensitive than the two tests and showed an 82% agreement ratio with the HI test and an 80.67% agreement ratio with the commercial kit. The NP(47-384)-ELISA and the commercial AIV ELISA were used to evaluate 448 field sera samples from diseased chickens or vaccinated chickens during the 2003-04 AI outbreak in China. The two ELISA tests had a 95% agreement ratio. We conclude that the NP(47-384)-ELISA developed in our laboratory was specific and sensitive and it has great application potential in China's long-term prevention and control of AI. 相似文献
4.
Avian Influenza (AI), caused by Alphainfluenzaviruses (AIVs), is a contagious respiratory disease in birds and mammals. AIVs have been reported in poultry worldwide and the impact of AIVs on human health is immense. In this study, a serological survey of AIV subtype H5 and H9 was conducted in a live bird market (LBM) in Yangon, Myanmar during February 2016 to September 2016. A total of 621 serum samples were collected from chickens (n = 489) and ducks (n = 132) from 48 vendors in the LBM. The samples were examined for antibodies against influenza viruses by using NP-ELISA and specific antibodies against AIV-H5N1 (Clade 2.3.4) and AIV-H9N2 (Clade 9.4.2) by using Hemagglutination Inhibition (HI) assay. The result of NP-ELISA assay showed that 12.88 % (80/621) of poultry in LBM was positive for AIV antibodies. In detail, 38.06 % (51/134) of layers, 7.08 % (8/113) of backyard chicken, 2.07 % (5/242) of broilers and 12.12 % (16/132) of ducks were AIV positive. The HI test for specific antibodies against AIV-H5N1 and AIV-H9N2 were 1.77 % (11/621) and 4.51 % (28/621), respectively. Our findings revealed the evidence of AIV-H5N1 and AIV-H9N2 exposure in both chicken and ducks in the LBM in Yangon, Myanmar. Risks of influenza infections and transmission among poultry and humans in the LBMs could not be ignored. 相似文献
5.
A latex agglutination test (LAT) was developed for rapid detection of antibodies to H5 avian influenza viruses (AIVs). The hemagglutinin protein of H5 AIV was covalently linked to carboxylated latex by ethyl-dimethyl-amino-propyl carbodiimide to prepare the sensitized latex beads. The LAT was evaluated with the hemagglutination inhibition (HI) assay as the reference test. The H5-LAT showed a sensitivity of 87.0% and specificity of 88.9% in detecting 126 serum samples from experimentally infected chickens and a sensitivity of 82.5% and specificity of 86% in detecting 587 field chicken serum samples from mostly vaccinated chickens. The agreement ratio between H5-LAT and HI was found to be 87.3% and 83.1% for the two groups of samples, respectively. Difficulty with background agglutination in stored chicken sera was overcome by serum pretreatment with either dried chicken liver powder or dilution buffer containing detergent Tween-20. The H5-LAT has advantages over a previously reported whole-virus LAT in terms of biosafety in preparation, chemical stability, and higher specificity. It is a rapid and simple test suitable for field monitoring of antibodies to H5-type AIV. 相似文献
6.
检测禽流感病毒抗体的重组核蛋白间接ELISA方法的建立 总被引：9，自引：3，他引：9
以大肠杆菌系统表达的H9N2亚型禽流感病毒（AIV）核蛋白（NP）为抗原，建立了禽流感间接酶联免疫吸附试验抗体检测技术（NP—ELISA）。对263份待检血清（包括临床收集的243份血清和20份H9N2亚型AIV免疫鸡阳性血清）进行检测，NP—ELISA与琼脂免疫扩散试验（AGP）的总符合率为83．3％，与血凝抑制试验（HI）的总符合率为92％。特异性试验表明，NP—ELISA方法可以检测H5、H7和H9亚型AIV特异性抗体，检测为阳性的血清样品能够被阳性鸡胚尿囊液阻断。敏感性试验证实，NP—ELISA最早可以检测鸡感染后7d的血清样品，并于感染后10d确定100％血清阳性，而AGP检测直到首免后21～28d才出现部分血清阳性，HI检测直到10～14d才出现部分血清阳性，并且NP-ELISA要比HI敏感4～40倍。试验证明，NP—ELISA是检测AIV血清型特异性抗体的一种特异、敏感、快速、经济的血清学检测技术。 相似文献
7.
S. Karki B. Lupiani C. M. Budke S. Manandhar R. Ivanek 《Zoonoses and public health》2014,61(6):442-448
Kathmandu, Nepal has been classified as a high‐risk area for highly pathogenic avian influenza (HPAI) by the Nepali Government. While ducks have an important role in the transmission of avian influenza viruses (AIV), including HPAI, seroprevalence of antibodies to AIV in domestic ducks of Kathmandu has never been assessed. The objectives of this study were (i) to estimate the prevalence of seroconversion to AIV in domestic ducks in major duck‐raising areas of Kathmandu and (ii) to assess the effect of age, sex, presence of swine and the number of ducks on the farm on the carriage of antibodies to AIV in these ducks. From April through July of 2011, a cross‐sectional study was conducted and a total of 310 ducks in the major duck‐raising areas of Kathmandu were sampled. The estimated prevalence of AIV antibodies was 27.2% [95% confidence interval (CI): 24.6–29.5]. Of 62 enrolled farms, 42% had at least one seropositive duck. Half of the enrolled farms also kept pigs of which 52% had at least one seropositive duck. Bivariate analysis indicated association between ducks' seroconversion to AIV and their age, sex and farm size. However, the final multivariable model, after controlling for clustering of ducks within farms, identified age as the only significant risk factor. Based on this model, ducks older than 1 year of age were more likely to be seropositive compared to ducks <6 months of age [odds ratio = 2.17 (1.07–4.39)]. These results provide baseline information about the AIV seroprevalence in domestic ducks in the major duck‐raising areas of Kathmandu and identify a high‐risk group that can be targeted in surveillance activities. Future studies should be conducted to differentiate the subtypes of AIV present among domestic ducks in Kathmandu, with particular interest in the presence of HPAI viruses. 相似文献
8.
9.
Development of an ELISA using a recombinant 41 kDa partial protein (P45N') for the detection of Riemerella anatipestifer infections in ducks 总被引：5，自引：0，他引：5
Riemerella anatipestifer, a gram-negative bacillus, is the causative agent of duck septicemia, a disease which could incur much economic loss in the duck industry. An indirect enzyme-linked immunosorbent assay (ELISA) has been developed to facilitate early detection of R. anatipestifer infection in ducks. The antigen used was a recombinant 41 kDa N-terminal fragment (rP45N') of a newly characterized R. anatipestifer potential surface protein, P45, which was expressed in Escherichia coli as an N-terminal GST fusion protein. The rP45N'-based ELISA successfully detected P45 antibodies in the sera of 20 ducks immunized with bacterin preparations of R. anatipestifer serotypes 1, 10 15, 19 and the ATCC11845 strain. Antibodies to P45 were also detected in the sera of 25% (75/296) of White Pekin ducks which were imported into Singapore from three different farms. Successful discrimination was obtained between sera from infected ducks and that of specific-pathogen free ducks (p<0.01). The rP45N'-GST antigen did not cross-react with antibodies in sera from guinea pigs which were infected with other gram-negative and gram-positive bacterial pathogens, including Aeromonas hydrophila, Citrobacter freundii, E. coli, Klebsiella pneumoniae, Pastuerella multocida, Proteus mirabilis, Salmonella spp., Serratia maccescens, Shigella sonnei and Yersinia enterocolitica. In addition, the DNA sequence encoding P45 was detected in R. anatipestifer serotypes 1, 2, 3, 4, 5, 6, 7, 8, 9, 11, 14, 15, 16, 17, 18, 19 and the ATCC11845 strain, suggesting that P45 is probably also universally expressed in these R. anatipestifer serotypes. Thus, the ELISA described is applicable to the detection of R. anatipestifer infection in ducks. 相似文献
10.
Development and evaluation of a DAS-ELISA for rapid detection of avian influenza viruses 总被引：4，自引：0，他引：4
Rapid detection of avian influenza virus (AIV) infection is critical for control of avian influenza (AI) and for reducing the risk of pandemic human influenza. A double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was developed for this purpose. The method employed a monoclonal antibody (MAb) as the capture antibody and rabbit polyclonal IgG labeled with horseradish peroxidase as the detector antibody, and both antibodies were against type-specific influenza A nucleoprotein (NP). The DAS-ELISA could detect minimally 2.5 ng of influenza viral protein in virus preparations treated with Triton X-100, which is equvilent to 2.5 x 10(2) EID50 virus particles. This DAS-ELISA could detect all 15n AIV subtypes (H1-H15) and did not cross react with other avian pathogens tested. The DAS-ELISA were directly compared with virus isolation (VI) in embryonated chicken eggs, the current standard of influenza virus detection, for 805 chicken samples. The DAS-ELISA results correlated with VI results for 98.6% of these samples, indicating a sensitivity of 97.4% and specificity of 100%. The method was further tested with H5N1 and H9N2 AIV experimentally infected chickens, ducks, and pigeons, as well as field samples obtained from central China in 2005. The DAS-ELISA method has demonstrated application potential as an AIV screening tool and as a supplement for virus isolation in Asia. 相似文献
11.
RT—PCR扩增鸭呼肠孤病毒（Duckreovirus,DRV）的dB基因,克隆到pET-32a（＋）表达载体,再转化大肠杆菌Transetta（DE3）;含有重组质粒pET—σB的大肠杆菌经IPTG诱导,获得大小为55000的以包涵体形式表达的σB重组蛋白。Westernblotting显示,σB重组蛋白能够与兔抗DRV多抗血清特异性结合。以高亲和NI—NTA树脂在变性条件下纯化、梯度尿素复性的σB重组蛋白为包被抗原,建立了检测DRV抗体的间接ELSIA方法。分别以间接ELSIA方法和血清中和试验对DRV感染鸭血清和SPF鸭血清进行检测,两者的符合率为100％。该间接ELSIA方法对鸭瘟病毒、鸭病毒性肝炎病毒和禽流感病毒阳性血清均无交叉反应。 相似文献
12.
To differentiate avian influenza virus (AIV)-infected chickens vs. chickens immunized with inactivated avian influenza virus, an enzyme-linked immunosorbent assay (ELISA) was developed using a recombinant nonstructural protein (NS1) as the diagnostic antigen, which was cloned from an AIV H9N2 subtype strain isolated during the avian influenza outbreak of 2003-04 and expressed in Escherichia coli. Antibodies to the AIV NS1 protein was only detected in the sera of chickens experimentally infected with AIV but not in the sera of chickens immunized with inactivated vaccine. This ELISA is useful for serological diagnosis to distinguish chickens infected with influenza viruses from those immunized with inactivated vaccine. 相似文献
13.
De Marco MA Campitelli L Foni E Raffini E Barigazzi G Delogu M Guberti V Di Trani L Tollis M Donatelli I 《Veterinary microbiology》2004,98(3-4):197-208
We report the results of a 6-year serological and virological monitoring performed in ducks and coots in Italy, in order to assess the degree of influenza A virus circulation in these birds during wintering. A total of 1039 sera collected from 1992 to 1998 was screened by a double antibody sandwich blocking ELISA (NP-ELISA): seroprevalence of antibodies to influenza A viruses was significantly higher in ducks compared to coots (52.2% vs. 7.1%, respectively). The hemagglutination-inhibition (HI) assay, performed on NP-ELISA positive sera, showed that 16.9% of these duck sera and 33.3% of these coot sera had antibodies to at least one influenza virus HA subtype: ducks showed HI antibodies against most of the HA subtypes, except for the H3, H4, H7, and H12; coots were seropositive to the H3 and H10 subtypes, only. From 1993 to 1998, 22 virus strains were obtained from 802 cloacal swabs, with an overall virus isolation frequency of 2.7%. Viruses belonging to the H1N1 subtype were by far the most commonly circulating strains (18/22) and were isolated mainly from ducks (17/18). The remaining viruses were representative of the H10N8, H5N2 and H3N8 subtypes. Our data indicate some differences between influenza A virus circulation in sympatric ducks and coots and a significant antigenic diversity between some reference strains and viruses recently isolated in Italy. 相似文献
14.
Trine H Jensen Gitte Ajjouri Kurt J Handberg Marek J Slomka Vivien J Coward Martine Cherbonnel Véronique Jestin Peter Lind Poul H J?rgensen 《Acta veterinaria Scandinavica》2013,55(1):84
Background
Avian influenza virus (AIV) subtypes H5 and H7 attracts particular attention because of the risk of their potential pathogenicity in poultry. The haemagglutination inhibition (HI) test is widely used as subtype specific test for serological diagnostics despite the laborious nature of this method. However, enzyme-linked immunosorbent assays (ELISAs) are being explored as an alternative test method.H5 and H7 specific monoclonal antibodies were experimentally raised and used in the development of inhibition ELISAs for detection of serological response specifically directed against AIV subtypes H5 and H7. The ELISAs were evaluated with polyclonal chicken anti-AIV antibodies against AIV subtypes: H1N2, H5N2, H5N7, H7N1, H7N7, H9N9, H10N4 and H16N3.Results
Both the H5 and H7 ELISA proved to have a high sensitivity and specificity and the ELISAs detected H5 and H7 antibodies earlier during experimental infection than the HI test did. The reproducibility of the ELISA’s performed at different times was high with Pearson correlation coefficients of 0.96-0.98.Conclusions
The ELISAs are a potential alternative to the HI test for screening of large amounts of avian sera, although only experimental sera were tested in this study. 相似文献15.
An immunoperoxidase monolayer assay for the detection of antibodies against swine influenza virus. 总被引：3，自引：0，他引：3
Kochakorn Direksin HanSoo Joo Sagar M Goyal 《Journal of veterinary diagnostic investigation》2002,14(2):169-171
An immunoperoxidase monolayer assay (IPMA) has been developed to detect antibodies against swine influenza A virus (SIV) in pig sera. The test was evaluated by using sequential sera from pigs experimentally infected with H1N1 subtype of SIV. Two hundred field serum samples that had been examined by the hemagglutination inhibition (HI) test were also tested. Antibodies specific to SIV were detected as early as 3 days postinoculation (dpi) in the IPMA test as compared with 7 dpi by the HI test. Unlike HI, no serum treatment was required in the IPMA test. Regardless of the virus used in the test, IPMA detected antibodies to both H1N1 and H3N2 subtypes of SIV whereas HI detects antibodies against either H1N1 or H3N2, depending upon the virus used in the test. Results of this study indicate that IPMA is a useful test for screening of pig sera for SIV antibodies. 相似文献
16.
Many commercial enzyme-linked immunosorbent assays (ELISAs) are unable to differentiate antibody responses to different avian influenza virus (AIV) subtypes. Developing an ELISA for specifically detecting the H5 antibody is the purpose of this study. Four monoclonal antibodies (Mabs) were raised using A/duck/Yunlin/04 (H5N2). They were confirmed as being specific to H5. Two of these antibodies showed hemagglutination inhibition (HI) activity using the HI test. Using immunodot blot assays, three Mabs recognized both Eurasian and American H5, whereas the other Mab recognized only the tested Eurasian H5 virus. When testing denatured H5 antigen, one of the Mabs lost its antigen binding activity using Western blotting. For detecting the H5 humoral response in serum, one monoclonal antibody was purified and labeled with horseradish peroxidase to set up a blocking ELISA. Chicken sera that blocked H5 Mab binding by > 29% were considered H5 antibody positive. Inhibition percentages for sera from chickens infected with other AIV subtypes, H1 to H15, were < 29%. This blocking ELISA was used for 478 field chicken serum samples. The results showed that the sensitivity and specificity of this ELISA were 98.3% (232/236) and 95.9% (232/242), respectively. This blocking ELISA could be used specifically for detecting the H5 humoral responses in chickens. 相似文献
17.
Choi KS Nah JJ Choi CU Ko YJ Sohn HJ Libeau G Kang SY Joo YS 《Veterinary microbiology》2003,96(1):1-16
An experimental competitive enzyme-linked immunosorbent assay (morbillivirus cELISA) using a recombinant N antigen (rRPV N) expressed in a baculovirus and a ruminant morbillivirus (RPV and PPRV)-specific monoclonal antibody (P-13A9) was developed for simultaneous detection of rinderpest virus (RPV) and peste des petits ruminants virus (PPRV) antibodies and its diagnostic performance was evaluated. A set of known reference antisera against RPV and PPRV belonging to different lineages, experimental sera from cattle vaccinated for a RPV of Asian lineage, and field sera from cattle and sheep/goat populations known to be positive (West Africa) and negative (Korea) for RPV and PPRV were used for the evaluation. Morbillivirus cELISA results on the panel of experimental RPV and PPRV antisera showed high correlation (r=0.97) between the whole virus and the rRPV N antigens, suggesting that the rRPV N contains a ruminant morbillivirus-specific antigenic determinant recognized by the P-13A9 and it may be suitable as an ELISA antigen in place of the whole virus. Morbillivirus cELISA detected anti-RPV and anti-PPRV antibodies in all reference RPV and PPRV antisera containing VN titers >/=1:8, suggesting that the assay can simultaneously detect antibodies against RPV and PPRV. Anti-RPV antibody was detected by morbillivirus cELISA in vaccinated cattle as early as the VNT and continued to be detectable by both the cELISA and the VNT until termination of the study. When applied to field samples from Africa, morbillivirus cELISA showed good agreement with a RP cELISA kit (kappa value of 0.86) in bovine sera and with a peste des petits ruminant cELISA kit (kappa value of 0.81) in caprine/ovine sera. Usefulness of morbillivirus cELISA using the rRPV N protein was discussed. 相似文献
18.
19.
间接ELISA,HI及AGP检测鸡血清中抗AIV抗体的敏感性比较 总被引：7，自引：1，他引：6
用鸭源Ａ型流感病毒Ｈ９Ｎ？滴鼻、点眼辅以腹腔注射,人工感染３周龄ＳＰＦ来航鸡,分别用ＡＧＰ、ＨＩ试验及间接ＥＬＩＳＡ定期测定其抗ＡＩＶ血清效价。结果,ＨＩ试验及间接ＥＬＩＳＡ于攻毒后第７～７９天显示阳性;ＡＧＰ试验从第１３～７９天呈阳性。根据首次检出血清中抗ＡＩＶ抗体的时间,间接ＥＬＩＳＡ比ＡＧＰ、ＨＩ更灵敏、快速。 相似文献
20.
Kang-Seuk Choi Soo-Jeong Kye Woo-Jin Jeon Mi-Ja Park Saeromi Kim Hee-Jung Seul Jun-Hun Kwon 《Journal of veterinary science (Suw?n-si, Korea)》2013,14(3):291-297
A recombinant hemagglutinin-neuraminidase (rHN) protein from Newcastle disease virus (NDV) with hemagglutination (HA) activity was expressed in Spodoptera frugiperda cells using a baculovirus expression system. The rHN protein extracted from infected cells was used as an antigen in a hemagglutination inhibition (HI) test for the detection and titration of NDV-specific antibodies present in chicken sera. The rHN antigen produced high HA titers of 213 per 25 µL, which were similar to those of the NDV antigen produced using chicken eggs, and it remained stable without significant loss of the HA activity for at least 12 weeks at 4℃. The rHN-based HI assay specifically detected NDV antibodies, but not the sera of other avian pathogens, with a specificity and sensitivity of 100% and 98.0%, respectively, in known positive and negative chicken sera (n = 430). Compared with an NDV-based HI assay, the rHN-based HI assay had a relative sensitivity and specificity of 96.1% and 95.5%, respectively, when applied to field chicken sera. The HI titers of the rHN-based HI assay were highly correlated with those in an NDV-based HI assay (r = 0.927). Overall, these results indicate that rHN protein provides a useful alternative to NDV antigen in HI assays. 相似文献