Field evaluation of a new commercially available ELISA based on a recombinant antigen for diagnosing Chlamydophila abortus (Chlamydia psittaci serotype 1) infection

A new commercially available ELISA (ELISAr-Chlamydia) for detecting antibodies against Chlamydophila abortus has been evaluated using sheep field serum samples. The ELISA is based on a recombinant antigen which expresses part of a protein from the 80-90kDa family that is specific to C. abortus. Sera (105) from six flocks with confirmed ovine chlamydial abortion (OEA) outbreaks were used in this study, as well as sera (258) from 18 flocks which had suffered no OEA in the last lambing. The ELISAr-Chlamydia was compared with the complement fixation test (CFT) and with an ELISA using purified C. abortus elementary bodies (ELISA-EB), employing as reference technique a comparative microimmunofluorescence test that differentiates C. abortus infection from Chlamydophila pecorum infection. The results showed that the sensitivity of ELISAr-Chlamydia was 90.9% with a specificity of 85.9%, the sensitivity of CFT was 71.0% with a specificity of 83.6%, while the sensitivity of ELISA-EB was 95.2% and the specificity was 54.2%. Furthermore, ELISAr-Chlamydia was the test with fewer false positives resulting from positive reactivity to C. pecorum, although 15% of the sera positive for C. pecorum but negative for C. abortus antibodies reacted positively. This study demonstrated with field material that ELISAr-Chlamydia provides the most balanced results between sensitivity and specificity, especially in flocks with no clinical OEA but reactivity to C. abortus.     

Abnormalities of serum electrolyte concentrations in dogs with hypoadrenocorticism

BACKGROUND: The sensitivity and specificity of the sodium to potassium ratio (Na:K ratio) as a cutoff for recommendation of an adrenocorticotropic hormone (ACTH) stimulation test in dogs suspected of having hypoadrenocorticism (HA) is unknown. Additionally, abnormalities in plasma ionized calcium (iCa2+) and ionized magnesium (iMg2+) concentrations and venous pH of dogs with HA are incompletely documented. OBJECTIVES: To define the sensitivity and specificity of the Na:K ratio as a diagnostic aid for HA in dogs and to examine for associations between venous pH and the Na:K ratio, iCa2+ concentration, or iMg2+ concentration in dogs with HA. ANIMALS: Seventy-six dogs with HA and 200 dogs randomly selected from the general hospital population. METHODS: Retrospective study. Dogs were included in the study if results of an ACTH stimulation test confirmed a diagnosis of HA, the dog had a serum sodium concentration below the reference range or a serum potassium concentration above the reference range, and the dog was treated with mineralocorticoids. Receiver operating curve analysis was used to determine optimal cutoffs of sensitivity and specificity for the Na:K ratio in diagnosing HA. RESULTS: Use of Na:K ratios of 27 or 28 classified 95% of dogs correctly as diseased or not diseased. The sensitivity of a Na:K ratio of 28 was 93% (CI, 85-98%) and that of 27 was 89% (CI, 80-95%). The specificity of a Na:K ratio of 28 was 96% (CI, 92-98%) and that of 27 was 97% (CI, 93- 99%). The sensitivity and specificity of a Na:K ratio of 24 were 79% (95% CI, 67-86%) and 100% (98%, CI, 97%-100%), respectively. CONCLUSIONS AND CLINICAL IMPORTANCE: Na:K ratios of 27 or 28 identify the highest percentage of dogs with suspected mineralocorticoid and glucocorticoid deficiency correctly. In dogs with a Na:K ratio of 24 or less, the likelihood of confirming a diagnosis of HA with an ACTH stimulation test is high.     

Diagnostic accuracy of PCR for Jaagsiekte sheep retrovirus using field data from 125 Scottish sheep flocks

Using a representative sample of Scottish sheep comprising 125 flocks, the sensitivity and specificity of PCR for Jaagsiekte sheep retrovirus (JSRV) was estimated. By combining and adapting existing methods, the characteristics of the diagnostic test were estimated (in the absence of a gold standard reference) using repeated laboratory replicates. As the results of replicates within the same animal cannot be considered to be independent, the performance of the PCR was calculated at individual replicate level.The median diagnostic specificity of the PCR when applied to individual animals drawn from the Scottish flock was estimated to be 0.997 (95% confidence interval [CI] 0.996–0.999), whereas the median sensitivity was 0.107 (95% CI 0.077–0.152). Considering the diagnostic test as three replicates where a positive result on any one or more replicates results in a positive test, the median sensitivity increased to 0.279. Reasons for the low observed sensitivity were explored by comparing the performance of the test as a function of the concentration of target DNA using spiked positive controls with known concentrations of target DNA. The median sensitivity of the test when used with positive samples with a mean concentration of 1.0 target DNA sequence per 25 μL was estimated to be 0.160, which suggests that the PCR had a high true (analytical) sensitivity and that the low observed (diagnostic) sensitivity in individual samples was due to low concentrations of target DNA in the blood of clinically healthy animals.     

Risk factors and characteristics of H5N1 Highly Pathogenic Avian Influenza (HPAI) post-vaccination outbreaks

Highly pathogenic avian influenza (HPAI) virus H5N1 is now endemic in South-East Asia but HPAI control methods differ between countries. A widespread HPAI vaccination campaign that started at the end of 2005 in Viet Nam resulted in the cessation of poultry and human cases, but in 2006/2007 severe HPAI outbreaks re-emerged. In this study we investigated the pattern of this first post-vaccination epidemic in southern Viet Nam identifying a spatio-temporal cluster of outbreak occurrence and estimating spatially smoothed incidence rates of HPAI. Spatial risk factors associated with HPAI occurrence were identified. Medium-level poultry density resulted in an increased outbreak risk (Odds ratio (OR) = 5.4, 95% confidence interval (CI): 1.6–18.9) but also climate-vegetation factors played an important role: medium-level normalised difference vegetation indices during the rainy season from May to October were associated with higher risk of HPAI outbreaks (OR = 3.7, 95% CI: 1.7–8.1), probably because temporal flooding might have provided suitable conditions for the re-emergence of HPAI by expanding the virus distribution in the environment and by enlarging areas of possible contacts between domestic waterfowl and wild birds. On the other hand, several agricultural production factors, such as sweet potatoes yield, increased buffalo density, as well as increased electricity supply were associated with decreased risk of HPAI outbreaks. This illustrates that preventive control measures for HPAI should include a promotion of low-risk agricultural management practices as well as improvement of the infrastructure in village households. Improved HPAI vaccination efforts and coverage should focus on medium poultry density areas and on the pre-monsoon time period.     

Development and evaluation of a rapid immunomagnetic bead assay for the detection of classical swine fever virus antigen

Classical swine fever (CSF) is a highly contagious and severe viral disease of swine resulting in substantial production losses in different farming systems in many regions of the world. The accurate and rapid detection of CSF outbreaks is reliant on sensitive and specific laboratory testing and is a key component of disease control. Specific detection of CSF virus can be achieved by virus isolation in tissue culture, antigen capture or the detection of viral RNA using molecular techniques. In order to reduce the time taken to achieve a diagnostic result and simplify testing methods, an antigen capture ELISA using immunomagnetic beads (IMB) as the solid phase was developed and compared to a microplate-based antigen capture (AC)-ELISA. The IMB-ELISA has up to 64-fold greater analytical sensitivity than the AC-ELISA and initial estimates of diagnostic sensitivity and specificity are 100%. The IMB-ELISA has a highly robust, rapid and stable test format and is simpler to perform than the AC-ELISA. The IMB-ELISA has the added advantage that a result can be sensitively and specifically determined by eye, lending it to the possibility of adaptation to a near-to-field test with minimal equipment or expertise needed.     

Zero-inflated models for identifying disease risk factors when case detection is imperfect: Application to highly pathogenic avian influenza H5N1 in Thailand

Logistic regression models integrating disease presence/absence data are widely used to identify risk factors for a given disease. However, when data arise from imperfect surveillance systems, the interpretation of results is confusing since explanatory variables can be related either to the occurrence of the disease or to the efficiency of the surveillance system. As an alternative, we present spatial and non-spatial zero-inflated Poisson (ZIP) regressions for modelling the number of highly pathogenic avian influenza (HPAI) H5N1 outbreaks that were reported at subdistrict level in Thailand during the second epidemic wave (July 3rd 2004 to May 5th 2005). The spatial ZIP model fitted the data more effectively than its non-spatial version. This model clarified the role of the different variables: for example, results suggested that human population density was not associated with the disease occurrence but was rather associated with the number of reported outbreaks given disease occurrence. In addition, these models allowed estimating that 902 (95% CI 881–922) subdistricts suffered at least one HPAI H5N1 outbreak in Thailand although only 779 were reported to veterinary authorities, leading to a general surveillance sensitivity of 86.4% (95% CI 84.5–88.4). Finally, the outputs of the spatial ZIP model revealed the spatial distribution of the probability that a subdistrict could have been a false negative. The methodology presented here can easily be adapted to other animal health contexts.     

Prevalence and distribution of paratuberculosis (Johne's disease) in cattle herds in Ireland

A simple random survey was conducted in Ireland during 2005 to estimate the ELISA-prevalence of paratuberculosis, commonly called Johne's disease (JD), in the cattle population. Serum samples were collected from all 20,322 females/breeding bulls over 12 months-of-age in 639 herds. All samples were tested using a commercially available absorbed ELISA. The overall prevalence of infected herds, based on the presence of at least one ELISA-positive animal, was 21.4% (95% CI 18.4%-24.9%). Herd prevalence levels amongst dairy herds (mean 31.5%; 95% CI: 24.6%, 39.3%) was higher than among beef herds (mean 17.9%; 95% CI: 14.6%-21.8%). However, the animal level prevalence was similar. The true prevalence among all animals tested, was calculated to be 2.86% (95%CI: 2.76, 2.97) and for animals >= 2 yrs, it was 3.30% (95%CI: 3.17, 3.43). For animals in beef herds, true prevalence was 3.09% (95%CI: 2.93, 3.24), and for those in dairy herds, 2.74% (95%CI: 2.59, 2.90). The majority of herds had only one ELISA-positive infected animal. Only 6.4% (95% CI 4.7%-8.7%) of all herds had more than one ELISA-positive infected animal; 13.3% (CI 8.7%-19.7%) of dairy herds ranging from two to eight ELISA-positive infected animals; and, 3.9% beef herds (CI 2.4%-6.2%) ranging from two to five ELISA-positive infected animals. The true prevalence of herds infected and shedding Mycobacterium avium subspecies paratuberculosis is estimated to be 9.5% for all herd types; 20.6% for dairy herds; and 7.6% for beef herds. If ELISA positive animals <2-years-of-age are excluded, the true herd prevalene reduces to: 9.3% for all herd types; 19.6% for dairy herds; and 6.3% for beef herds based on a test specificity (Sp) of 99.8% and test sensitivity (Se) (i.e., ability to detect culture-positive, infected animals shedding at any level) of 27.8-28.9%.     

Utility of 2 Immunological Tests for Antemortem Diagnosis of Equine Protozoal Myeloencephalitis (Sarcocystis neurona Infection) in Naturally Occurring Cases

Background: Antemortem diagnosis of equine protozoal myeloencephalitis (EPM) is challenging. Limited information is available regarding a commercial test (surface antigen 1 [SAG‐1] ELISA). Performance of another commercial test (indirect fluorescent antibody test [IFAT]) using samples from an independent group has not been well described. Hypothesis/Objectives: The primary goal was to evaluate the SAG‐1 ELISA and IFAT using naturally occurring EPM cases. A secondary goal was to obtain more information regarding clinical presentation. Animals: Hospital cases were admitted over 20 months and classified into 4 groups. Confirmed positive cases (n = 9) had asymmetric or multifocal neurologic deficits or both and postmortem lesions consistent with EPM. Confirmed negative cases (n = 17) had variable clinical signs and postmortem lesions consistent with another neurologic disease (or no lesions). Suspected positive cases (n = 10) had asymmetric or multifocal deficits or both, marked improvement after treatment for EPM, and other likely diseases excluded. Suspected negative cases (n = 29) had orthopedic disease and no neurologic deficits. Methods: Results of immunological testing (SAG‐1 ELISA and IFAT on serum or cerebrospinal fluid [CSF] or both), neurologic examinations, CSF analyses, and postmortem examinations were analyzed retrospectively. Results: SAG‐1 ELISA sensitivity was 12.5% (95% CI, 1.6–38.4) and specificity was 97.1% (95% CI, 84.7–99.9) using serum. IFAT sensitivity was 94.4% (95% CI, 72.7–99.9) and specificity was 85.2% (95% CI, 66.3–95.8) using serum; sensitivity was 92.3% (95% CI, 64.0–99.8) and specificity was 89.7% (95% CI, 72.7–97.8) using CSF. Conclusions and Clinical Importance: Low sensitivity of the SAG‐1 ELISA limited its usefulness for antemortem diagnosis of EPM in this patient population.     

Performance of clinical signs to detect bluetongue virus serotype 8 outbreaks in cattle and sheep during the 2006-epidemic in The Netherlands

The performance of clinical signs as a diagnostic test for the detection of BTV-8 outbreaks during the 2006-epidemic in The Netherlands was evaluated by constructing and analysing receiver operating characteristic (ROC) curves. The area under the ROC curve of the BT-associated clinical signs in cattle was 0.77. An optimal efficient test (maximising both sensitivity and specificity) in cattle herds combined a sensitivity (Se) of 67% with a specificity (Sp) of 72%, comprising the following clinical signs: ulcerations and/or erosions of oral mucosa or erosions of lips/crusts in or around nostrils or oedema of the nose or hyperaemic/purple coloration of tongue, tongue protrusion or coronitis or apathy/tiredness or muscle necrosis, stiffness of limbs or loathing or refusal to move, prostration or torticollis or anoestrus. The area under the ROC curve of the BT-associated clinical signs in sheep was 0.81. The optimal efficient test in sheep flocks combined a Se of 76% with a Sp of 72%, comprising the following clinical signs: ulcerations of oral mucosa or serous nasal discharge or erosions/ulceration of tongue mucosa or hypersensitivity of the skin or muscle necrosis, stiffness of limbs or coronitis or grinding of teeth or salivation or weakness/paresis.     

Diagnostic sensitivity and specificity of an enzyme-linked immunosorbent assay for the diagnosis of Brucella ovis infection in rams.

An enzyme-linked immunosorbent assay (ELISA) for antibody to Brucella ovis was compared with a standard complement fixation test. Sera of 176 rams from uninfected flocks gave 175 negative and one suspect ELISA reaction (diagnostic specificity 99.4%) whereas the complement fixation test yielded 167 negative, seven suspect and two anticomplementary reactions (diagnostic specificity of 96.0%). Diagnostic sensitivity was evaluated on sera of 79 rams from which B. ovis had been isolated. The ELISA showed 75 positive and four suspect reactions, while complement fixation test revealed 64 positive, 13 suspect and two negative results. Considering both positive and suspect reactions, the diagnostic sensitivity was 100% for ELISA and 97.5% for complement fixation test. The ELISA method was considered more specific, more sensitive and technically more advantageous than complement fixation test as a serodiagnostic test for B. ovis infection in rams.     

Evaluation of an immunofluorescence test on direct smears of conjunctival and urogenital swabs taken from koalas for the detection of Chlamydia psittaci

The Chlamydia-Cel Vet IF Test (CCVIT), a commercially available immunofluorescence test for use on direct smears of clinical specimens, was evaluated in a colony of 43 captive koalas. The test is based on a monoclonal antibody directed against the chlamydial common group specific lipopolysaccharide antigen. Swabs were taken from conjuncitva and penis or urogenital sinus and used for direct smear evaluation and cell culture isolation. Compared with isolation of the organism in cell culture, the CCVIT on direct smears of conjunctival swabs presented a sensitivity of 88%, a specificity of 100%, a positive predictive value (PV+) of 100% and a negative predictive value (PV-) of 97%. The CCVIT on direct smears of urogenital swabs presented a sensitivity of 90%, a specificity of 84%, a PV+ of 86% and a PV- of 89%. The overall sensitivity was 89% (95% confidence interval [CI] of 71% to 97%), the specificity 94% (95% CI of 84% to 98%), the PV+ 89% and the PV- 94%. It was concluded that the CCVIT on direct smears was suitable as a diagnostic screening test for the detection of Chlamydia psittaci in koalas.     

Inspection of wool lots at sales as a diagnostic test for louse infestation

The accuracy of visual inspection of wool lots for lice as a test for louse infestation was estimated using information provided by 178 woolgrowers in Queensland, Australia. The estimated sensitivity of inspection was 36% (95% confidence interval, 19-58%) and the specificity was 95% (95% CI, 88-98%). Accuracy was influenced by timing, after shearing, of pesticide application for louse control and by class of pesticide last applied after shearing. Visual inspection was less sensitive (29%) if pesticides were applied >3 months after shearing and less sensitive (21%) if an insect growth regulator was the class of pesticide last used after shearing. Based on 36% sensitivity, it was estimated that 16 inspections would have to be conducted to reduce the false negative test rate to <20% in the study population. We suggest that visual inspection of wool lots could be used to efficiently monitor the prevalence of louse infestations in Queensland sheep flocks. Positive inspection results are more likely to represent real louse infestations, rather than a false test result, in flocks grazed in the more extensive regions of Queensland.     

Design and Results of an Intensive Monitoring Programme for Avian Influenza in Meat‐Type Turkey Flocks During Four Epidemics in Northern Italy

Surveillance programmes for low pathogenicity (LPAI) and high pathogenicity avian influenza (HPAI) infections in poultry are compulsory for EU Member States; yet, these programmes have rarely been evaluated. In Italy, following a 1999 HPAI epidemic, control measures, including vaccination and monitoring, were implemented in the densely populated poultry area (DPPA) where all epidemics in Italy have been concentrated. We evaluated the monitoring system for its capacity to detect outbreaks rapidly in meat‐type turkey flocks. The evaluation was performed in vaccination areas and high‐risk areas in the DPPA, in 2000–2005, during which four epidemics occurred. Serum samples and cloacal swabs were taken from vaccinated birds and unvaccinated (sentinel) birds. We compared the detection rate of active, passive and targeted surveillance, by vaccination status, using multinomial logistic regression. A total of 13 275 samplings for serological testing and 4889 samplings for virological testing were performed; 6315 production cycles of different bird species were tested. The outbreaks detection rate in meat‐type turkeys was 61% for active surveillance (n = 222/363 outbreaks), 32% for passive surveillance and 7% for targeted surveillance. The maximum likelihood predicted values for the detection rates differed by vaccination status: in unvaccinated flocks, it was 50% for active surveillance, 40% for passive surveillance and 10% for targeted surveillance, compared to respectively 79%, 17% and 4% for vaccinated flocks. Active surveillance seems to be most effective in detecting infection, especially when a vaccination programme is in place. This is the first evaluation of the effectiveness of different types of surveillance in monitoring LPAI infections in vaccinated poultry using field data.     

Evaluation of single and comparative intradermal tuberculin tests for tuberculosis eradication in caprine flocks in Castilla y León (Spain)

Goats can act as reservoirs for tuberculosis (TB) infection. The main etiological agents of TB in goats are Mycobacterium caprae and Mycobacterium bovis and they infect also a wide range of domestic and wild animals and humans. Control programmes based mainly on the application of single and comparative intradermal tuberculin (SIT and SCIT respectively) tests are being implemented in certain regions of Spain with a high density of caprine flocks as Castilla y León, including goats with epidemiological relationship with cattle. The aim of this study was to evaluate the performance of the intradermal tests in naturally TB-infected caprine flocks from this region. The study was performed using data from 17,450 goats in 54 different flocks that were classified as TB-infected in the control programmes executed in 2010 and 2011. Data from 1237 goats from 7 dairy flocks depopulated after the first intradermal testing were used to estimate the sensitivity (Se) using bacteriology as the gold-standard. Overall Se of the SIT test using the severe interpretation was 43.9% (CI 95%, 40.4–47.4) and decreased to 38.8% (CI 95%, 35.5–42.3) using the standard interpretation. Overall Se of the SCIT test ranged between 21.3% (CI 95%, 17.6–25.4) and 7% (CI 95%, 4.9–9.8) depending of the interpretation criteria. A significant weak positive correlation was found between age and skin fold thickness (Spearman’s test p < 0.05). Results from this study yielded, in general, low Se values probably due the systematic detection and slaughter of reactors as a consequence of the eradication programme in previous years or the presence of factors that may interfere in the diagnosis. Therefore, these results suggest the necessity of including ancillary diagnostic tools and/or strict interpretation criteria to maximize detection of positive animals in infected settings.     

Improvements to the hemagglutination inhibition test for serological assessment of recombinant fowlpox-H5-avian-influenza vaccination in chickens and its use along with an agar gel immunodiffusion test for differentiating infected from noninfected vaccinated animals

In general, avian influenza (AI) vaccines protect chickens from morbidity and mortality and reduce, but do not completely prevent, replication of wild AI viruses in the respiratory and intestinal tracts of vaccinated chickens. Therefore, surveillance programs based on serological testing must be developed to differentiate vaccinated flocks infected with wild strains of AI virus from noninfected vaccinated flocks in order to evaluate the success of vaccination in a control program and allow continuation of national and international commerce of poultry and poultry products. In this study, chickens were immunized with a commercial recombinant fowlpox virus vaccine containing an H5 hemagglutinin gene from A/turkey/Ireland/83 (H5N8) avian influenza (AI) virus (rFP-H5) and evaluated for correlation of immunological response by hemagglutination inhibition (HI) or agar gel immunodiffusion (AGID) tests and determination of protection following challenge with a high pathogenicity AI (HPAI) virus. In two different trials, chickens immunized with the rFP-H5 vaccine did not develop AGID antibodies because the vaccine lacks AI nucleoprotein and matrix genes, but 0%-100% had HI antibodies, depending on the AI virus strain used in the HI test, the HI antigen inactivation procedure, and whether the birds had been preimmunized against fowlpox virus. The most consistent and highest HI titers were observed when using A/turkey/Ireland/83 (H5N8) HPAI virus strain as the beta-propiolactone (BPL)-inactivated HI test antigen, which matched the hemagglutinin gene insert in the rFP-H5 vaccine. In addition, higher HI titers were observed if ether or a combination of ether and BPL-inactivated virus was used in place of the BPL-inactivated virus. The rFP-H5 vaccinated chickens survived HPAI challenge and antibodies were detected by both AGID and HI tests. In conclusion, we demonstrated that the rFP-H5 vaccine allowed easy serological differentiation of infected from noninfected birds in vaccinated populations of chickens when using standard AGID and HI tests.     

Comparison of four diagnostic tests for the identification of serum antibodies in small ruminants infected with Mycoplasma agalactiae

AIM: To determine the diagnostic capability of a newly developed Western blot (WB) assay for the detection of serum antibodies against Mycoplasma agalactiae compared with conventional serological tests, and to identify the best test for routine diagnostic use. METHODS: The serological test methods used were: two commercial indirect enzyme-linked immunosorbent assays (ELISA), viz ELISA-1, using a bacterial antigen preparation, and ELISA-2, using a recombinant protein (lipoprotein p48) antigen; the complement fixation test (CFT); and a newly developed WB assay, the latter both using a bacterial antigen preparation. Thirty sera from goats infected with M. agalactiae and 97 sera from non-infected sheep were tested using all four methods. RESULTS: Staining patterns in the WB were quite variable. An immuno-dominant band of 41 kDa was detected in 63% of sera from infected animals. The same band also appeared, although mostly very weakly, in 10% of sera from non-infected animals. When suspicious or very weak reactors were omitted, the diagnostic sensitivity (DSE) and diagnostic specificity (DSP), respectively, for the four assays were: WB=56.7%, 97.9%; ELISA-1=76.7%, 99.0%; ELISA-2=56.7%, 100%; and CFT=40.0%, 94.8%. CONCLUSIONS: ELISA-1 performed best in this comparison. While the WB can be used, it did not have a technical advantage over the ELISA. The CFT should be discouraged as the primary screening method for contagious agalactia and should be replaced by ELISA-1. Results from this study confirm that serological test methods for contagious agalactia are useful for the detection of infected flocks but will not detect every individual infected animal.     

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.