A serological survey of the prevalence of Aujeszky's disease antibodies in Thailand using enzyme-linked immunosorbent assays (ELISA), serum neutralization (SN) and latexagglutination tests (LT).

The presence of Aujeszky's disease (AD) antibodies in eluates of whole blood on filter paper and corresponding sera from Thai pigs was determined by ELISA, SNT and LT. From a total of 800 samples tested by ELISA, 26% of the sera and 18% of the eluates showed positive results. From 640 samples tested by SNT and chosen because they gave negative, suspicious, or weakly positive results by ELISA, 22% were positive. A total of 182 suspicious samples were also tested by LT, and among them 63 (35%) were clearly positive. The investigation demonstrated that the older the animal, the greater the probability that antibodies would be found. Owner surveys tended to state that few animals had been vaccinated. This coupled with the high frequency of antibodies detected, indicates that AD-infection among Thailand's swine population is a common problem.     

Comparison of the enzyme-linked immunosorbent assay (ELISA) and serum neutralisation test for the serodiagnosis of Aujeszky's disease

The serum neutralisation test (SNT) and the indirect enzyme-linked immunosorbent assay (ELISA) for Aujeszky's disease were compared, utilising 3202 sera from Aujeszky's disease free pig herds, and 304 SNT reactor and 245 non-reactor sera from Aujeszky's disease infected piggeries. ELISA was found to give good discrimination between positive and negative sera, results showing 96.9% and 99.7% agreement with the SNT in classifying positive and negative reactor pigs respectively. The ELISA appeared to detect a slightly higher proportion of reactors than did the SNT. Absorbance values obtained with ELISA showed a high degree of overall correlation with SW titres (r = 0.916), though correlations were lower when applied to individual sera. The ELISA was considered to be a rapid and convenient procedure, offering many advantages over the SNT for routine use.     

A Serological Survey for Swine Vesicular Disease in Denmark

Pig sera from Danish breeding centres were examined for occurrence of antibodies to swine vesicular disease virus using the counter immunoetlectro-phoresis test (GIET) and the serum neutralization test (SNT). Nine hundred and ten serum samples from 28 breeding centres were tested using the SNT and the GIET in parallel, whereas in a survey of 2062 samples from 218 herds the GIET was used for initial screening. GIET positive samples were subsequently assayed by the SNT. It was concluded that of the 2962 samples tested none had significant content of specific antibody to swine vesicular disease virus, although cross reacting antibody could be demonstrated in some cases.     

Comparison of whole blood dried on filter paper and serum for measurement of the temporal antibody response to avian infectious bronchitis virus by enzyme-linked immunosorbent assay

Two methods for collecting specimens for measuring sequential antibody activity of infectious bronchitis virus (IBV) were compared. Whole blood was collected on filter-paper strips, dried for 2 hr at 37 C, and then stored in plastic bags at 4 C or eluted overnight and tested immediately. Eluates of whole blood were paired with serum samples and tested for IBV antibody activity by enzyme-linked immunosorbent assay (ELISA) at four weekly intervals. Both sampling methods yielded ELISA antibody levels that were detectable at 7 days postinfection (PI), peaked at 21 days PI, and then began to decline by 28 days PI. The paired samples showed no significant difference (P less than 0.05) between ELISA titers at any time tested. Whole blood dried on filter paper could be stored sealed in plastic bags at 4 C for at least 2 weeks with no appreciable loss of antibody titers. Virus-neutralizing antibodies, measured in serum only, were not detectable until 14 days PI but then continued to rise through 28 days PI. It was concluded that eluates of whole blood dried on filter paper may be used as an alternative to sera in ELISA for measuring IBV antibodies.     

Tick-borne encephalitis (TBE) with special emphasis on infection in horses

The tick-borne encephalitis (TBE), also known as early summer meningo-encephalitis, is a geographically limited virus infection transmitted mainly by ticks. The importance of TBE is largely underestimated. The causative agent TBE-Virus (TBEV) is grouped into the genus Flavivirus of the virus family Flaviviridae. Clinical disease including fatal outcomes has been described for men and dogs. With regard to horses only a limited number of case reports is available. In a study performed at the Institute of Virology, Justus-Liebig-University Giessen serum samples from the German endemic region of Marburg-Biedenkopf were tested for antibodies against TBEV. From 240 sera tested 7 (2.9%) were regarded as positive in a serum neutralization test (SNT). In an ELISA, performed in parallel to confirm the SNT results, 5 out of 7 positive sera from the SNT were also positive. The remaining two samples with low SNT-titres and all sera from horses negative in the SNT were also negative in the ELISA. This article is focussed on TBE of horses. In this context different aspects of TBE are included such as properties of the causative agent, interactions between causative agent, host animals and environment, spread of TBEV, pathogenesis, clinical symptoms, diagnosis and control.     

Use of filter paper for blood specimen collection in poultry for use in the detection of antibodies to Newcastle disease virus

Trials were conducted to verify the possibility of poultry blood sampling with filter papers for subsequent examination of the eluates for the presence (and level) of haemagglutination-inhibition antibodies against the Newcastle disease virus (NDV). Qualitative examination was performed in 294 paired samples of sera and eluates, representing 10 selective sets, coming from three vaccinated poultry flocks. The numbers of positive sera (dilution ratios 1:20 and higher) and positive eluates of filter papers (dilution ratios of 1: 2 and higher) were compared and it was found that there were 238 positive paired samples (81%) and 30 were negative (10.2%), hence, there were like reactions in 268 paired samples (91.2% of the total number of samples examined). It was only in 25 paired samples that positivity was recorded just in the sera: 22 times with a titre of 1:20 and three times with a titre of 1:40. In one case, positivity was recorded just in the eluate. The final titres were compared in 181 paired samples of sera and eluates, all diluted at a ratio of 1:2, and it was found that the concentration of the haemagglutination-inhibition antibodies in the eluates corresponded to serum dilution ratio of 1:20. Under this assumption, the antibodies were found to have the same titre in 164 paired samples (55.8%) during the quantitative evaluation. A lower titre was recorded in 82 eluate samples (27.9%) and a higher antibody titre in 48 eluate samples (16.3%) (in comparison with the antibody titres in the respective sera). The over-all average geometrical titre (GMT) of antibodies was 1:65 in the eluates and 1:75 in the sera.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence of Schmallenberg virus circulation in ruminants in Greece

During March 2013, we investigated the presence and the levels of Schmallenberg virus (SBV) circulation in three dairy cow herds and three sheep flocks in Central Macedonia, Greece. In two cow herds, a high number of abortions had been observed during the winter. Six bulk-tank milk samples and 147 individual sera were screened for SBV-specific antibodies by ELISA. Positive reactions were obtained from 5 out of 6 bulk-tank milk samples, 58 out of 90 sera from the 3 cow herds, and 2 sera from 2 of the 3 sheep flocks. Twenty-two ELISA-positive sera were tested by serum neutralization test (SNT). SNT confirmed the presence of neutralizing antibodies against SBV in all samples tested, with titers ranging between 1:32 and ≥1:256. No neutralizing antibodies against Akabane virus (AKAV) or Shamonda virus (SHAV) were detected, indicating that neutralizing antibodies against SBV do not cross react with AKAV or SHAV in SNT. ELISA testing of bulk-tank milk samples proved to be convenient and reliable. None of the tested sera was found positive for SBV by real-time RT-PCR, indicating that the sampling was conducted past the viremia stage. This is the first report of SBV circulation in Greece.     

A serological survey of selected pathogens in wild boar in Slovenia

Serum samples collected from 178 shot wild boars (Sus scrofa) were tested for the presence of antibodies against classical swine fever virus, Aujeszky's disease virus (ADV), porcine reproductive and respiratory syndrome virus, porcine respiratory coronavirus (PRCV), transmissible gastroenteritis virus, swine influenza virus, porcine parvovirus (PPV), swine vesicular disease virus, Actinobacillus pleuropneumoniae (APP), Mycoplasma hyopneumoniae, Salmonella spp., Brucella spp. and Haemophilus parasuis (HPS) throughout Slovenia during the hunting season 2003/2004. The number of samples corresponds to 3% of the total hunting bag. By enzyme-linked immunosorbent assay (ELISA) antibodies against ADV were detected in 55 sera (31%), against PRCV in five sera (3%), PPV in 87 sera (49%), APP in 93 sera (52%), M. hyopneumoniae in 38 sera (21%), Salmonella spp. in 85 sera (47%) and HPS in 33 sera (18%).     

Evaluation of an enzyme-linked immunosorbent assay (ELISA) for serodiagnosis of infectious bovine rhinotracheitis infection, with results of a preliminary survey

An indirect enzyme linked immunosorbent assay (ELISA) procedure was evaluated against the serum neutralisation test (SNT) for the detection of antibodies to infectious bovine rhinotracheitis virus (bovine herpesvirus type l), using 2028 sera from 166 dairy and 172 beef cattle herds. The results showed the ELISA to give high levels of agreement with the SNT in classifying positive and negative sera (98% and 97% respectively). Such disagreements as did occur involved weakly reactive sera with SNT titres of % or less. A number of sera (n=123) with trace neutralising activity of doubtful diagnostic significance were found to give marginal reactivity with ELISA. ELISA absorbance values were found to be highly correlated with SNT titres (r=0.909) on an overall basis, though agreements were lower with individual sera. The ELISA procedure was quicker, cheaper, and detected more reactors than the SNT. It also allowed results to be obtained with a number of sera which were unsuitable for testing by SNT because of their cytotoxic nature. Analysis of ELISA results showed reactors to be present in 57% of tested sera, representing 81% of cattle herds. Reactor rates for sera and herds in the South Island, (37% and 58%), were significantly lower than for those in the North Island (64% and 88%). Antibody prevalence was also found to be significantly lower in districts having a low annual rainfall (<850 mm), and to be lower in beef cattle than in dairy cattle. A surprising exception to the latter occurred in low rainfall districts, where dairy cattle showed significantly lower reactor rates than local beef animals.     

A sensitive dot immunobinding assay for serodiagnosis of African swine fever virus with application in field conditions.

The present work describes a simple dot immunobinding assay (DIA) for African swine fever virus (ASFV) antibody detection that can be used under field conditions. The assay uses nitrocellulose strips dotted with a cytoplasmic soluble antigen (CS-P) of ASFV. The nitrocellulose strips are adhered to a plastic handle. The test serum samples react with the CS-P, and antibodies are detected using a protein A-peroxidase conjugate. Both incubations are carried out at 20 C. The efficacy of the DIA as a screening test for ASFV was compared to an enzyme-linked immunosorbent assay (ELISA) and an immunoblotting (IB) test using 343 sera collected from natural African swine fever epizootics and from inapparent ASFV carriers. The DIA had comparable sensitivity to both reference techniques, and all samples positive in the ELISA and IB test were also positive in the DIA. False-positive reactions were not detected when whole blood or poorly preserved serum samples were tested by DIA. Some poorly preserved sera that were positive initially by the ELISA were no longer ELISA positive in a later run, although they were positive in IB and DIA. These positive DIA and IB test results could be caused by the differences in antibody epitope binding.     

乳胶凝集试验与血清中和试验检测猪伪狂犬病血清抗体的 …

应用血清中和试验（ＳＮＴ）和伪狂犬病乳胶凝集试验（ＬＡＴ）诊断试剂盒对两种伪狂犬病是性血清、伪狂犬病病毒（ＰＲＶ）高兔血甭及６０份被检猪血清进行了ＰＲＶ抗体效价测定和相关性分析,两种方法测得的抗体效价之间呈强相关性（ｒ＝０．９６）,且ＬＡＴ效价比ＳＮＴ一般高出一个滴度;能干为自３５个猪场的４１４份猪血清进行了ＰＲＶ抗体检测,并与ＳＮＴ检测结果进行了对比,结果在ＳＮＴ检测为阳笥的１７１份血清中,ＬＡ     

Evaluation of serological tests for detecting tick-borne encephalitis virus (TBEV) antibodies in animals

Tick-borne encephalitis (TBE) in animals is not well understood yet. TBE virus (TBEV) serology in several host species could be valuable for epidemiological analyses in the field as well as for the detection of clinical cases. However, performance and suitability of the available test systems are not well assessed. Therefore, we evaluated two commercial TBEV-ELISA kits in a pilot study and compared them for their suitability in veterinary applications. For this purpose, we tested 163 field collected goat sera and evaluated the results by serum neutralization test (SNT) as "gold standard". Twenty-eight SNT positive sera (17.2%) were detected. The best suited ELISA kit was used for determination of a species-specific cutoff for horses, cattle, sheep, goats, pigs, mice, dogs, rabbits and monkeys with defined sera from animals without known or with improbable contact to TBEV. The level of non-specific ELISA results does not only differ between animal species but may also be influenced by the age of the tested animals. The number of sera which tested false positive by ELISA was higher in older than in young sheep. In order to obtain defined polyclonal sera as references, two dogs, cattle, goats, sheep, rabbits and pigs each, as well as one horse and 90 mice were immunized four times with a commercially available TBEV vaccine. In conclusion, our results demonstrated that commercial TBEV-ELISA kits are suitable for application in veterinary medicine for both, verification of clinical TBE cases and epidemiological screening. However, positive ELISA results should be verified by SNT. Only a very low number of false negative ELISA-results were found.     

gIII antibody responses in pigs following vaccination and challenge to Aujeszky's disease.

Serological responses to a genetically engineered Aujeszky's disease "marker" vaccine (dl gIII + dl tk) were monitored using a blocking-ELISA (B-ELISA), a serum neutralisation test (SNT) and an indirect ELISA (I-ELISA). The B-ELISA is capable of differentiating pigs vaccinated with the above vaccine from natural infection. The SNT and the I-ELISA indicated that the pigs responded to vaccination and challenge. All three tests showed that the controls and the in-contact pigs always reacted negative for antibodies. The B-ELISA was able to detect pigs challenged with a field isolate 24 days post-challenge. These pigs remained positive until 110 days post-challenge when last tested. These findings indicate that the B-ELISA could be used successfully with this vaccine in a control eradication programme. This trial also shows that the vaccine virus did not spread to the in-contact pigs and also the vaccinated and challenged pigs did not transmit the disease to other susceptible pigs when they were introduced 14 days after challenge.     

Comparison of filter-paper-eluted whole blood with serum in fowl cholera serology using the enzyme-linked immunosorbent assay

Two methods for collecting blood for measuring antibody activity of Pasteurella multocida were compared. Whole blood was collected on filter-paper strips, dried for 48 hr at room temperature, and then stored in sealed plastic bags at 4 C. Blood was also collected in the usual manner with a needle and syringe, and serum was harvested and stored at -20 C until tested. Eluates of whole blood, obtained by overnight elution of two 4.8-mm discs in 200 microliters of buffered saline at 4 C, were compared with conventionally harvested serum for antibody activity by enzyme-linked immunosorbent assay (ELISA). Paired samples, taken from the same bird at the same time, showed no significant difference (P less than 0.05) in antibody activity as measured by absorbance when the disc-elution process itself was considered to be a 1:20 dilution. It was concluded that eluates of blood, derived from whole blood dried on filter-paper strips, may be used as an alternative to sera in ELISA for measuring P. multocida antibody activity.     

贵州省猪弓形虫病的血清学调查

为全面了解贵州省猪弓形虫感染情况,对采自9个市（州、地）264个养猪场（户）的2 906份血清用酶联免疫吸附试验（ELISA）检测猪弓形虫抗体,总体阳性率为65.83%,变化范围为27.88%~85.42%。采集65份猪血清做ELISA和间接血凝试验（indirect heamagglutination assay,IHA）比较测定,两种方法总体符合率为58.46%。IHA方法补充检测177份猪血清,弓形虫抗体阳性率为27.68%。血清学调查结果与国内部分省（市）报道相符。     

Prevalence of coronavirus antibodies in Iowa swine.

Three hundred and forty-seven serum samples from 22 Iowa swine herds were screened for TGEV/PRCV neutralizing antibody. Ninety-one percent of the sera and all 22 herds were positive. These sera were then tested by the blocking ELISA test to distinguish TGEV and PRCV antibody. The ELISA test confirmed the high percentage of TGEV/PRCV positive sera. By the blocking ELISA test, 12 herds were PRCV positive, 6 herds were TGEV positive and 4 herds were mixed with sera either positive for TGEV or PRCV antibody. The results suggest a recent increase in TGEV/PRCV seroprevalence in Iowa swine most likely due to subclinical PRCV infections.     

Survey of trichinosis in breeding and cull swine, using an enzyme-linked immunosorbent assay

Serum samples obtained from 40,927 swine at various locations in North Carolina between Aug 1, 1987 and July 31, 1988, were tested for antibodies to Trichinella spiralis, using an ELISA based on a larval T spiralis excretory-secretory antigen. In the ELISA, samples were considered to have positive results if the optical density (OD) reading was equal to or 5 times greater than the mean OD value of 4 negative-control sera from trichina-free swine. Of the 40,927 serum samples tested, 154 (0.38%) were positive by ELISA; the rate for breeding swine was 0.35% (105/30,162), and the rate for cull swine was 0.45% (49/10,765). Of the 49 seropositive samples from cull swine, 11 were from out of the state, 22 had no identification, and 16 were known to originate from North Carolina. Seropositivity had a bimodally seasonal distribution, with peaks in March and September. There was no difference between the mean age of seropositive and seronegative swine, but males were at greater risk for seropositivity than were females. Pigs from lots with less than 100 sera tested were at increased risk for seropositivity, as were pigs from the central coastal region of North Carolina.     

Relationship between the anti-FMD virus antibody reaction as measured by different assays, and protection in vivo against challenge infection.

The antibody response of cattle after vaccination against foot-and-mouth disease (FMD) virus was monitored using the serum neutralization test (SNT), the sandwich ELISA, liquid-phase ELISA, sandwich competition ELISA, liquid-phase competition ELISA, and the liquid-phase sandwich competition (blocking) ELISA. The competition ELISAs (in particular the "blocking" ELISA) were the most effective at detecting reactivity in these cattle sera. However, 95% of negative sera also competed in the most sensitive ELISA (the "blocking" ELISA) to titres of 1:32 (4% of the sera competed to a titre of 1:128). Comparisons between the different ELISAs, and between these ELISAs and the SNT, demonstrated that the tests were not measuring exactly the same reaction of antibody with FMD virus. With respect to the capacity of animals to resist FMD virus challenge, neither the SNT nor the competition ELISAs were consistently able to identify such animals. The anti-FMD virus antibody titres obtained could be classified into three zones; the "white zone" wherein antibody titres were high and donor animals likely to be protected; the "black zone" wherein antibody titres were low and donor animals likely to be susceptible to infection; the "grey zone" wherein the antibody titres were intermediary and no interpretation could be made with respect to protection. Assays such as ELISA and SNT cannot and do not measure immunological protection; they are a measure of antibody responses and nothing more, and should be interpreted in terms of the "three zone" phenomenon.     

Evaluation of two commercial enzyme-linked immunosorbent assays for detection of bovine viral diarrhoea virus in serum and skin biopsies of cattle

AIM: To assess the ability of two commercial bovine viral diarrhoea (BVD) virus (BVDV) antigen-capture enzyme-linked immunosorbent assays (ELISAs) to detect virus in serum and skin biopsies. METHODS: Thirty cattle persistently infected (PI) with BVDV were identified using routine diagnostic laboratory testing. Additional ear-notch skin biopsies and blood samples were collected from these animals to confirm the diagnosis, and from 246 cohorts, to determine their BVDV status. Skin biopsies were soaked overnight in buffer and the eluate collected. All sera and eluate were tested using two commercially available ELISAs for detecting BVDV antigen, and a subsample of positive and negative sera was tested using a polymerase chain reaction (PCR) test. A study was also performed to ascertain the risk of cross contamination occurring during the collection and processing of skin biopsies. RESULTS: Both serum and skin samples tested using either ELISA resulted in the detection of all cattle identified as PI and no non-infected cattle were incorrectly classified as infected using either method. Agreement between all assays (ELISAs, whether performed on serum or skin, and PCR) was 100%. No cross-contamination of skin samples between animals was evident using routine biopsy methods. CONCLUSIONS: Viraemic cattle infected with BVDV were accurately identified using either of the two commercial ELISAs evaluated on either serum or skin samples. CLINICAL RELEVANCE: Either skin biopsies or serum samples can be collected from cattle to determine their BVDV status. This should overcome problems in accurately identifying the infection status of young calves in which colostral antibodies might interfere with the antigen-capture ELISA.     

Quantitative and Qualitative Study of Enzyme-linked Immunosorbent Assay to Detect IgG against Japanese Encephalitis Virus in Swine Sera

This study was designed to investigate the application of indirect enzyme-linked immunoassay (ELISA) in detecting IgG against Japanese encephalitis virus in swine sera and the qualitative nature of this test. The attenuated strain SA14-14-2 of Japanese encephalitis virus (JEV) was inoculated into 9-day-old chicken embryos and virus was harvested, purified and suspended in 0.9% saline as JEV antigen. The control antigen was prepared by the same method as for the antigen. In the ELISA, the optimal concentrations of antigen coated and dilution factor were selected using chi2 test. Ninety-two swine sera negative to haemagglutination inhibition (HI) were tested by this assay and the positive threshold was determined. The results of this study indicate that indirect ELISA has high specificity, sensitivity and reproducability. Simultaneous testing of 74 serum samples from nine pig farms was carried out to compare the existing HI test and the indirect ELISA. The coincidence rate of the two assays was 85.1% (63/74) and no significant difference was observed between them (p > 0.05). This ELISA test can detect 46 swine serum samples qualitatively and the titre of eight swine serum samples through endpoint dilution quantitatively within one 96-well plate.