Sensitivity and specificity of the agar-gel-immunodiffusion test, ELISA and the skin test for detection of paratuberculosis in United States Midwest sheep populations

Our objective was to estimate the sensitivity and specificity of the agar-gel-immunodiffusion test (AGID), the ELISA, and the skin test for the detection of Mycobacterium avium subspecies paratuberculosis (MAP) in sheep using Bayesian methods without a gold standard. Fourteen flocks (2 465 sheep) were used. Five flocks (450 sheep) were considered MAP non-infected and 9 flocks (2 015 sheep) had sheep infected with MAP. Sheep were skin tested and blood was collected for AGID and ELISA testing. Results were analyzed using a Bayesian 3-test in 1-population model fitted in WinBUGS. The model allowed for dependence (correlation) between the two serologic tests, but these two tests were assumed to be conditionally independent of the skin test. The estimated specificity was 99.5% (95% PI of 98.9-99.9%) for the AGID; 99.3% (98.4-99.8%) for the ELISA using an optical density measured cutoff of 0.20; 99.2% (98.1-99.8%) using a cutoff of 0.15; 97.5% (95.8-98.7%) using a cutoff of 0.10; and 98.7% (97.3-99.5%) for the skin test. The estimated sensitivities were 8.3% (6.2-10.7%) for the AGID; 8.0% (6.0-10.4%), 10.6% (8.3-13.1%), and 16.3% (13.5-19.4%) for the ELISA using the cutoffs 0.20, 0.15, and 0.10 respectively; and 73.3% (62.3-85.8%) for the skin test. The skin test was specific in non-infected populations and sensitive in infected populations, although in some cases a positive skin test might represent MAP exposure rather than infection. The AGID and ELISA were specific but lacked sensitivity. The AGID and ELISA consistently identified two different populations of infected sheep with only moderate overlap between positive test results.     

Sensitivity and specificity of two serological tests for the detection of ovine paratuberculosis

OBJECTIVE: To determine the sensitivity and specificity of an absorbed ELISA and an AGID test for the detection of clinical and subclinical paratuberculosis in sheep. DESIGN: By testing a panel of sera from 1257 Australian Merino and crossbred sheep greater than 1 year of age, of which 1137 sheep were not infected with Mycobacterium avium subsp paratuberculosis and 120 sheep had paratuberculosis. PROCEDURE: Sera were collected from 457 sheep in Victoria and 800 sheep in Western Australia. Presence of M a paratuberculosis infection in Victorian sheep was determined by histological examination of intestinal tissues, whereas sheep from Western Australia were presumed to be free of Johne's disease. The ability of an absorbed ELISA to discriminate between infected and uninfected sheep was described by test sensitivity and specificity, the distribution of ELISA OD, and the area under a receiver operating characteristic curve. RESULTS: The absorbed ELISA had a specificity of 98.2 to 99.5% (CI) and a sensitivity of 35 to 54% (CI). In sheep from infected flocks in Victoria, the AGID test had a specificity of 99 to 100% (CI) and a sensitivity of 38 to 56% (CI). The sensitivity of serological tests was higher in sheep with a body condition representative of the lower quintile of their flock of origin. CONCLUSION: The AGID test and absorbed ELISA are useful tests for the detection of ovine paratuberculosis. Although the tests had a similar accuracy, they detected different subpopulations of infected sheep with only moderate overlap. The AGID test had a higher specificity than the absorbed ELISA.     

Comparative efficacy of standard AGID and precipitinogen inhibition test with monoclonal antibodies based competitive ELISA for the serology of <Emphasis Type="Italic">Peste des Petits Ruminants</Emphasis> in sheep and goats

This project was conducted to investigate the comparative efficiency of competitive ELISA (cELISA), standard Agar Gel Immunodiffusion Test (AGID) and Precipitinogen Inhibition Test (PIT) for the diagnosis of Peste des Petits Ruminants (PPR) in Pakistan. To deal with this, serum samples from 198 sheep and 82 goats were collected from three different government livestock farms and all the samples were run simultaneously with the three serological tests. The samples found positive for PPR antibodies through cELISA, AGID and PIT were 96 (34.2%), 60 (21.4%) and 72 (25.7%), respectively. Kappa statistics were applied to evaluate the concordance between the laboratory-based test (cELISA) and field-based tests (AGID and PIT). Kappa statistics scores for cELISA versus AGID and PIT were 0.6343 (95% Confidence Interval CI 0.5231–0.7456) and 0.7134 (95% Confidence Interval CI 0.5987–0.8281), respectively, which indicate a “substantial” agreement between cELISA and AGID and “significant” agreement between cELISA and PIT. AGID and PIT revealed relative diagnostic sensitivities with cELISA of 59.3% and 69.7% and relative diagnostic specificities of 98.3% and 97.2%, respectively. The data suggested that for mass screening and control of PPR, these serological tests proved practical in the absence of cELISA since they have high relative diagnostic specificities and a satisfactory relative diagnostic sensitivities.     

Seroprevalence of, and risk factors for, peste des petits ruminants in sheep and goats in Northern Jordan

Peste des petits ruminants (PPR) is an economically important disease that affect sheep and goat industry in Asia and Africa. In this study, we investigated the seroprevalence, and risk factors, of PPR in sheep and goat flocks from five different governorates (Irbid, Jarash, Ajloun, Mafraq and Zarka) located in Northern Jordan. Serum samples from 929 and 400 sheep and goats, respectively, corresponding to 122 sheep flock and 60 goats flock were collected. Seroprevalence was determined using PPR competitive ELISA. Health status and management information were collected using a semi-structured pre-tested questionnaire. The individual true prevalence of PPR in sheep and goats was 29 and 49%, respectively. The flock level true prevalence of PPR was 60 and 74% in sheep and goats, respectively. In both sheep and goat flocks, large flock size, visiting live animals market and inadequate veterinary services were identified as risk factors for PPR seropositivity. Mixed (sheep and goats) raising was identified as a risk factor for PPR seropositivity in sheep flocks only.     

Sensitivity and specificity of pooled faecal culture and serology as flock-screening tests for detection of ovine paratuberculosis in Australia

The flock-level sensitivity of pooled faecal culture and serological testing using AGID for the detection of ovine Johne’s disease-infected flocks were estimated using non-gold-standard methods. The two tests were compared in an extensive field trial in 296 flocks in New South Wales during 1998. In each flock, a sample of sheep was selected and tested for ovine Johne’s disease using both the AGID and pooled faecal culture. The flock-specificity of pooled faecal culture also was estimated from results of surveillance and market-assurance testing in New South Wales.

The overall flock-sensitivity of pooled faecal culture was 92% (95% CI: 82.4 and 97.4%) compared to 61% (50.5 and 70.9%) for serology (assuming that both tests were 100% specific). In low-prevalence flocks (estimated prevalence <2%), the flock-sensitivities of pooled faecal culture and serology were 82% (57 and 96%) and 33% (19 and 49%), respectively, compared to 96% (85 and 99.5%) and 85% (72 and 93%), respectively, in higher-prevalence flocks (estimated prevalence ≥2%). A Bayesian approach incorporating prior knowledge on flock-specificity of pooled culture produced similar estimates and probability intervals. These estimates assume conditional independence of the two tests, and therefore might have over-estimated the true flock-sensitivities of the tests if the flock-sensitivities of pooled faecal culture and serology were correlated.

The estimated minimum flock-specificity of pooled culture when used for surveillance and assurance testing was 99.1% (96.9 and 99.9%). Surveillance and assurance programs in Australia are designed to provide a flock-sensitivity of 95% for an assumed prevalence of 2%. Pooled faecal culture is performing at close to this level—whereas the flock-sensitivity of serology appears to be lower than expected, particularly in lower prevalence flocks.     

Specificity of absorbed ELISA and agar gel immuno-diffusion tests for paratuberculosis in goats with observations about use of these tests in infected goats

OBJECTIVE: To determine the specificity of serological tests that are currently used in veterinary diagnostic laboratories in Australia for detection of Mycobacterium avium subsp paratuberculosis infection in goats. DESIGN: A laboratory study. PROCEDURE: Four tests were studied, comprising AGID with M. a. paratuberculosis antigen derived from cattle isolates of caprine or bovine origin, the EMAI caprine Johne's disease absorbed ELISA and the CSL PARACHEK Johne's absorbed EIA. The specificities of AGID and ELISA for paratuberculosis (Johne's disease) were estimated after examining a panel of 1000 serum samples collected from goats in Western Australia, a region free of paratuberculosis. In addition a comparison was made of test performance in a small number of paratuberculous goats from New South Wales using sera from two archival collections. RESULTS: The specificity of the AGID tests was 100% while the specificities of the two absorbed ELISA were 99.7 to 99.8% at appropriate positive-negative cut-offs. Based on testing the small sample of sera from infected goats, the absorbed ELISA tests detected about twice as many goats with Johne's disease as the AGID. Each test detected paratuberculous animals regardless of whether infection was caused by cattle or sheep strains of M. a. paratuberculosis. CONCLUSIONS: Both ELISA and AGID tests for paratuberculosis have high specificity and can be used in a market assurance program without risk of generating large proportions of false positive test results. However, the results suggested the ELISA is more sensitive for detection of infected goats and should be used in preference to the AGID. The two formats of ELISA evaluated in this study have similar characteristics and could be used in paratuberculosis control programs for the goat industries, but further data on sensitivity would increase confidence in their application.     

Antibody prevalence of low-pathogenicity avian influenza and evaluation of management practices in Minnesota backyard poultry flocks

Low‐pathogenicity avian influenza (LPAI) viruses have caused illness in poultry and humans with poultry contact. To determine whether there is evidence of exposure to avian influenza viruses (AIV) among backyard poultry in Minnesota and their human caretakers, 150 flocks of backyard birds were sampled for antibodies to AIV from August 2007 through December 2008. One hundred flocks were tested through routine slaughter surveillance by the Minnesota Board of Animal Health and an additional 50 flocks were contacted and sampled by study investigators. Blood was collected from 10 to 13 birds from each flock and a survey of biosecurity and management practices was administered to the flock owner. Blood samples were tested by agar gel immunodiffusion (AGID) for influenza A antibodies. Tested flocks had a median flock size of 100 birds (range: 12–800 birds), and were most commonly owned for meat for personal use (81% of respondents), fun or hobby (58%) and eggs for personal use (56%). Although 7% of flock owners reported that their birds had shown respiratory signs in the previous 3 months, only 1 of 150 flocks tested positive for influenza by AGID. Antibodies to LPAI H6N1 were detected in the positive flock. The owner of the positive flock did not have antibodies to H6 or other common AIV. Based on the findings of this study, the risk of transmission of LPAI viruses from backyard poultry to owners in Minnesota appears to be low under current conditions and management practices.     

Ovine brucellosis in alberta

Two parallel surveys of rams from Alberta sheep flocks were conducted to determine the presence of infection with Brucella ovis. In a retrospective study over a period of 24 months, using complement fixation test, 12 flocks out of 142 tested were considered infected. In another 17-month survey of slaughter rams by serology and culture methods 11 flocks out of 124 were found to be infected. The overall prevalence of ovine brucellosis was 8.6% of the flocks tested which represented 12.5% of the estimated sheep flocks in Alberta. Up to 67% of rams in infected flocks reacted to complement fixation test.

The complement fixation test was evaluated for its efficiency in the diagnosis of ovine brucellosis and compared with a limited number of an enzyme-linked immunosorbent assay (ELISA) results and clinical criteria. The complement fixation test as well as ELISA identified all culture positive rams. Both serological tests appeared satisfactory for the diagnosis of B. ovis epididymitis when the results could be interpreted in the light of flock history and clinical findings.

     

Paratuberculosis in sheep: an emerging disease in South Africa

During a serological survey for ovine paratuberculosis a total of 145934 ovine serum samples from 2019 farms throughout South Africa were tested by means of the AGID assay. Fifty-two infected farms were identified in the Western Cape and Eastern Cape provinces. Links between infected farms in the two provinces were established. Examination of the distribution of infected farms in the Western Cape indicated a positive correlation between acid soils and occurrence of infection. In an attempt to increase the sensitivity and facilitate screening of large numbers of sera two commercial ELISA systems were evaluated for their potential use in a future monitoring program. Sera from histologically positive sheep and known negative sheep flocks were used. The highest sensitivity (50. 9%) was found if both ELISA systems were run concurrently and the results of both systems combined.     

Comparison of IS900 tissue PCR, bacterial culture, johnin and serological tests for diagnosis of naturally occurring paratuberculosis in goats

Comparative efficacy of an IS900 tissue PCR, bacterial culture, johnin, agar-gel immunodiffusion (AGID) and absorbed-ELISA tests was investigated in 43 goats naturally infected with paratuberculosis. On histological examination, tissue sections from all animals showed typical granulomatous inflammatory changes. The lesions were classified as multibacillary (MB) (n=30), which had diffuse granulomatous lesions with abundant acid-fast bacilli (AFB), and paucibacillary (PB) (n=13), which had focal or multifocal granulomatous lesions with few AFB. The sensitivities of johnin test, tissue culture, faecal culture, tissue PCR, AGID and ELISA were 68% (17/25), 100% (30/30), 84.6% (22/26), 100% (30/30), 96.2% (25/26) and 100% (26/26) in MB goats, and 88.8 (8/9), 46.1% (8/13), 40% (4/10), 61.5% (8/13), 50% (5/10), and 70% (7/10) in PB goats, respectively. Except for the johnin test, which showed higher sensitivity in PB goats, all other tests displayed significantly higher sensitivities in MB goats. The results indicate the usefulness of tissue PCR, culture and serological tests in the diagnosis of clinically affected paratuberculous goats, especially with multibacillary pathology.     

Prevalence of Bovine viral diarrhoea virus (BVDV) antibodies among sheep and goats in India

The aim of this study was to determine the prevalence of pestivirus antibodies in sheep and goats in India. A total of 2803 serum samples collected between 2004 and 2008 from 1777 sheep in 92 flocks and 1026 goats in 63 flocks belonging to 13 states were tested by competition ELISA for detection of pestivirus antibodies. In sheep, the true prevalence rate was 23.4% (95% confidence interval: 22.9%–27.0%) and in goats it was 16.9% (95% CI: 16.4%–21.3%). The flock level seroprevalence was 66.3% for sheep and 54.0% for goats. Geographical variation in individual and flock prevalence was highly significant. A significant association (p?<?0.05) was found between sheep and goat flocks having cattle contact and the flock level seroprevalence. The seroprevalence was lower in 6 months–1 year age group compared to the 1–2 year and >2 year age groups in both sheep and goats. Cross neutralization studies on 61 seropositive sheep and 34 seropositive goat samples representing all positive flocks, exhibited > four fold higher titre to bovine viral diarrhoea virus type 1 (BVDV-1) in 41 sheep and 23 goat samples and to BVDV-2 in one sheep and goat each. This study for the first time showed serological evidence of wide spread BVDV infections in Indian sheep and goats, with BVDV-1 predominating and BVDV-2 occasionally besides highlighting the potential risk of infection to other species, which needs to be considered whenever BVD control measures are initiated.     

Comparison of serological tests based on outer membrane or internal antigens for detecting antibodies to Brucella ovis in infected flocks.

The aim of this work was to compare the performance of 6 serological tests using outer or internal antigens from Brucella for the diagnosis of Brucella ovis infection in sheep in an endemic area. Outer membrane antigens included a hot saline extract (HS) and the rough lipopolysaccharide (R-LPS) from B. ovis. Internal antigens were LPS-free total cytosolic proteins (CP) and an 18-kDa cytosolic protein (p18) from Brucella spp. Sera from 200 sheep from naturally infected flocks were assayed by agar gel immunodiffusion test (AGID) and by complement fixation test (CFT), both using HS, and by 4 ELISA using HS, R-LPS, CP, and p18, respectively. The percentage of positive results was 45.5% for ELISA with HS, 42.0% for ELISA with p18, 39.5% for CFT, 33.5% for ELISA with R-LPS, 29.0% for ELISA with CP, and 18.0% for AGID. Taking CFT as the reference test for calculating relative test parameters, the ELISA with HS had the best sensitivity (96.2%), while AGID and the ELISA with R-LPS had the best specificity (96.6%). The ELISA with CP was not more sensitive than the ELISA with p18 (67.1% vs. 79.7%) in spite of the higher number of antigens in CP. The lower relative sensitivity of tests using internal antigens might reflect a lack of antibodies to cytosolic proteins in some infected animals or a shorter persistence of these antibodies relative to antibodies to outer membrane components after recovery from infection.     

A comparison of whole virus and recombinant transmembrane ELISA and immunodiffusion for detection of ovine lentivirus antibodies in Italian sheep flocks

Sera from two sheep experimentally infected with ovine lentivirus (OLV) and from 186 sheep selected from flocks with known high or low prevalence of infection or on the basis of virological or histopathological examination were simultaneously tested by whole virus (WV) ELISA, recombinant transmembrane (r-TM) ELISA and AGID assay. Antigens for both the WV ELISA and AGID were prepared from an Italian field isolate; recombinant antigen was derived from the N-terminal region of the transmembrane envelope protein of strain K1514. The WV ELISA detected the highest number of seropositives, followed by the r-TM ELISA and AGID test. The sensitivity and specificity of the r-TM ELISA relative to the WV ELISA were 0.66 and 0.95, respectively. Immunoblot analysis of 14 WV ELISA-positive and r-TM ELISA-negative sera showed that the major core protein was immunodominant on WV antigen. It is concluded that the r-TM ELISA was more sensitive than the AGID test but less sensitive that the WV ELISA, particularly for detecting antibodies in the early stages of infection.Abbreviations AGID agar gel immunodiffusion - bp base pair - ELISA enzyme-linked immunosorbent assay - FBS fetal bovine serum - IB immunoblot - kD kilodalton - OLV ovine lentivirus - MEM minimal essential medium - p.i. post-infection - r recombinant - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - TM transmembrane - WV whole virus     

Benign footrot – an epidemiological investigation into the occurrence, effects on production, response to treatment and influence of environmental factors

SUMMARY Benign footrot was studied in 1 1/2-years-old Merinos on 2 farms in central Victoria from September 1987 to August 1990, Inclusive. Treatment groups of 100 sheep grazed together with the remaining untreated sheep. Inspections were carried out every 3 weeks during the spring transmission period until the number of lesions greater than score 2 dropped below 3%. At each inspection, each sheep was weighed and lesion scores for each foot and digit were recorded, the treated group of sheep was treated by standing in 20% (w/v) zinc sulphate-sodium lauryl sulphate for 1 hour, and bacteriological samples were randomly collected from 5 sheep with and 5 without lesions. Dichelobacter nodosus organisms were obtained from sheep in both groups.
Laboratory tests indicated benign organisms in flock A and low virulence, intermediate organisms in flock B. During the first 2 years, the number and severity of lesions were greater in flock A than in flock B. However, in the third year, with an early 'autumn break', there was a rapid and severe outbreak of footrot in flock B; 98% of the flock had lesions at the first inspection in July 1989. Flock A had a less dramatic increase in lesions of footrot. Both treated and untreated groups in flock B recovered rapidly between the third and fourth inspections. A later increase in lesions for both flocks coincided with damage caused by barley grass seeds. During this period there was a significant difference (P / 0.001) in body weight between the treated and untreated sheep on farm B. An extra 0.2 kg of skirted fleece was obtained from treated sheep when shorn in 1990, compared with the untreated group (P = 0.04). Treated sheep had fewer tender fleeces, 11% compared with 25%. Wool from the untreated group was 0.5 micron finer but the value was 1.79 a head (7%) less than that from the treated group.     

Use of Brucella canis antigen for detection of ovine serum antibodies against Brucella ovis

Brucella ovis causes a genital disease of sheep manifested by epididymitis in rams and placentitis in ewes producing reduced fertility in the flock. Clinical diagnosis is not sensitive enough and bacteriological testing is not feasible for detection of the disease in large numbers of animals. Indirect methods of serological testing are preferred for routine diagnosis, of which agar gel immunodiffusion (AGID), complement fixation (CF) and ELISA tests are recommended as the most efficient. Since B. ovis shares antigenic components with Brucella canis, it would seem that either strain could be used as antigen with the same results; however, the advantage of the B. canis (M-) strain variant is that it can be used to develop a satisfactory antigen for agglutination tests. We present data on AGID and IELISA tests using B. ovis antigen and rapid screening agglutination test (RSAT), 2-mercapto-ethanol RSAT (2ME-RSAT) and IELISA using B. canis antigen. We tested 225 animals. The cut-off values were adjusted by ROC analysis using 51 negative and 32 positive sera; the IELISA-B. canis cut-off value was 39 (%P) and IELISA-B. ovis, 51 (%P), with 100% sensitivity and specificity. Of the 32 positive sera from the infected flock RSAT detected 32 (100%), 2ME-RSAT 29 (91%) and AGID 31 (97%). Of the 142 sera from suspicious flocks, 46 were negative and 56 positive in all the tests; 16 were positive by RSAT, IELISA-B. canis and IELISA-B. ovis, 20 positive only with RSAT and 2 positive only by both IELISAs. RSAT is a very sensitive screening test that, because of its simplicity and easy interpretation, following a study in larger sample, could replace AGID as a screening test for diagnosis of ovine brucellosis caused by B. ovis. The IELISA-B. canis or IELISA-B. ovis could be used as confirmatory tests, since they show equal specificity and sensitivity.     

Persistence of immunity in commercial egg-laying hens following vaccination with a killed H6N2 avian influenza vaccine

The California poultry industry experienced an outbreak of H6N2 avian influenza beginning in February 2000. The initial infections were detected in three commercial egg-laying flocks and a single noncommercial backyard flock but later spread to new premises. The vaccination of pullet flocks with a commercially prepared, killed autogenous vaccine prior to their placements on farms with infected or previously infected flocks was used as a part of the eradication programs for some multiage, commercial egg production farms. The purpose of this study was to follow three vaccinated flocks on two commercial farms to track the immune responses to vaccination. The antibody-mediated responses of the three flocks followed in this study were markedly different. One flock achieved 100% seroconversion at 12.5 wk of age, but by 32 wk of age, all of the hens were seronegative by agar gel immunodiffusion (AGID). In contrast, at 32 wk of age, flocks from the other farm (flocks 2A and 2B) were 95% and 72% seropositive by AGID, respectively. Of the differences that were identified between the vaccination protocols on the two farms, the distinction that could explain the level of disparity between responses is the delivery of the second dose of vaccine with a bacterin on the first farm, which may have interfered with the persistence of immunity in this flock. Hens from flocks 2A and 2B were experimentally challenged at 25 wk of age with H6N2 avian influenza virus. Hens from flock 2A did not transmit virus to naive contact-exposed hens, but hens from flock 2B did. At 34 wk of age, hens from flock 2A were again challenged and naive contact-exposed hens were infected in this second trial. These challenge experiments served to demonstrate that despite detectable antibody responses in flocks 2A and 2B, the birds were protected from infection for less than 21 wk after the second vaccination.     

Serological survey and epidemiological investigation of maedi-visna in sheep in Finland

A survey for antibodies to maedi-visna virus (MV) in the Finnish sheep surveillance flocks was conducted in 1994. Examination of a total of 12931 serum samples from animals over 1 year of age from 545 flocks (81% of all flocks) revealed eight seropositive flocks and the subsequent epidemiological investigation yielded one additional seropositive flock, indicating a low prevalence of 1.6%. The infection was very probably imported from Sweden in 1981, but it was not detected until the survey was conducted 13 years later. The entire primary infection flock was slaughtered in 1995. 77% of the sheep were seropositive but the animals were clinically healthy and only one (5%) of the contact flocks of the primary infection flock had contracted the infection. This secondary infection flock, 77% of which was seropositive, was slaughtered in 1994; however, animals in this flock had respiratory problems and the lungs of three sheep showed typical MV lesions. Seven (24%) of its contact flocks had contracted the infection and these each had one or two seropositive animals except for one flock which had seven (18%) seropositive animals. The results show that the initial spread of MV can be insidious and wide before infection is revealed in surveys or any clinical cases are encountered.     

An enzyme-linked immunosorbent assay for detection of antibodies to maedi-visna virus in sheep. II. Comparison to conventional agar gel immunodiffusion test.

A study was conducted to compare the indirect enzyme-linked immunosorbent-assay (i-ELISA) test using antigen prepared by a simple technique using sodium dodecyl sulfate (SDS) treatment to the conventional agar gel immunodiffusion test (AGID). Ten specific-pathogen-free (SPF) sheep were inoculated with maedi-visna virus (MVV) and serum antibody titers compared over a period of 14 weeks. All the sheep seroconverted by the i-ELISA compared to 90% by the AGID. The i-ELISA detected antibody at a mean of 2.6 weeks prior to the AGID. In both tests, fluctuations were observed in the serum antibody response of two sheep. The i-ELISA had a specificity of at least 98.8% and an increased relative sensitivity of 15.5% compared to the AGID, based on the analysis of sera from experimental sheep with MVV free status and sera from sheep from various sources. Of the sera from a seronegative flock which had been monitored with the AGID after a "test and remove" eradication program, 10.2% were positive by the i-ELISA. It was concluded that the AGID test may not be adequate to monitor samples for an eradication scheme.     

Evaluation of three serological tests for diagnosis of Maedi-Visna virus infection using latent class analysis

Maedi-Visna virus (MVV) infection in sheep is present in several European countries, including Norway. The current Norwegian surveillance and control programme for MVV infection uses three serological tests: an agar gel immunodiffusion test (AGID) and two commercially available indirect ELISAs (Institut Pourquier, P-ELISA and HYPHEN BioMed, H-ELISA). From 18 flocks with suspected or confirmed MVV infection, sera from naturally infected sheep were obtained, and sensitivity (Se) and specificity (Sp) of the three tests were estimated in absence of a perfect reference test using latent class models in a Bayesian analysis. The AGID had higher Sp (95% posterior credibility interval (PCI) [98.4; 99.9]) than either ELISA (95% PCI: P-ELISA, [95.1; 99.0]; H-ELISA, [91.4; 96.6]), but much lower Se (95% PCI: AGID, [41.4; 59.8]; P-ELISA, [92.7; 100.0]; H-ELISA, [90.9; 99.4]). Currently the P-ELISA is used for screening and positive samples are subsequently confirmed by a setup using all three tests in a serial reading. The Se and Sp of the serial interpretations with and without the H-ELISA were estimated. The results suggested that the H-ELISA could be dropped as a confirmatory test as the Se of the three test serial reading was reduced significantly without adding a significant improvement of the Sp compared to the serial reading of the P-ELISA and AGID alone. However, the perceived cost of false positives versus false negatives will influence this decision. Estimates of the predictive values for the tests and combinations suggested that the P-ELISA is a good choice of screening, but confirmatory tests are needed to achieve acceptable levels of positive predictive values.     

Seroepidemiological survey of sheep in Carinthia for the dissemination of ruminant pestiviruses

In this study serological investigations were performed to determine the prevalence of pestiviral infections in sheep in one Federal State of Austria, namely Carinthia. 1527 blood samples from sheep in 147 flocks were collected and tested by Enzyme-linked immunosorbent assay and virus-neutralisation tests for antibodies to ruminant pestiviruses. The estimated flock prevalence was 47.6%, the individual prevalence 16.3%. Significant geographical variations in the flock as well in the individual prevalence were found. The highest prevalence in sheep and in sheep flocks was established in the region Spittal/Drau with 25.9% and 69.7%.The individual and the flock prevalence was significantly higher on farms where cattle or sheep from other farms were present than on farms with no cattle (p < 0.017). All Enzyme-linked immunosorbent assay positive sera were tested for Bovine viral diarrhea virus-1 (strain NADL), Bovine viral diarrhea virus-2 (strain 125) and for Border disease virus (strain MOREDUN) by virus neutralisation tests. Seventy out of 249 positive samples revealed the highest titres (> or = two-fold) to Bovine viral diarrhea virus-1 and 25 to Border disease virus. The remaining positive samples did not show clear results because of cross reactions.