Evaluation of two herd-level diagnostic tests for Streptococcus agalactiae using a latent class approach

Streptococcus agalactiae mastitis persists as a significant economic problem for the dairy industry in many countries. In Denmark, the annual surveillance programme for this mastitis pathogen initially based only on bacteriological culture of bulk tank milk (BTM) samples, has recently incorporated the use of the real-time PathoProof Mastitis PCR assay with the goal of improving detection of infected herds. The objective of our study was to estimate the herd sensitivity (Se) and specificity (Sp) of both tests of BTM samples using latent class models in a Bayesian analysis while evaluating the effect of herd-level covariates on the Se and Sp of the tests. BTM samples were collected from all 4258 Danish dairy herds in 2009 and screened for the presence of S. agalactiae using both tests. The highest Se of PCR was realized at a cycle threshold (Ct) cut-off value of 40. At this cut-off, the Se of the PCR was significantly higher (95.2; 95% posterior credibility interval [PCI] [88.2; 99.8]) than that of bacteriological culture (68.0; 95% PCI [55.1; 90.0]). However, culture had higher Sp (99.7; 95% PCI [99.3; 100.0]) compared to PCR (98.8; 95% PCI [97.2; 99.9]). The accuracy of the tests was unaffected by the herd-level covariates. We propose that screenings of BTM samples for S. agalactiae be based on the PCR assay with Ct readings of <40 considered as positive. However, for higher Ct values, confirmation of PCR test positive herds by bacteriological culture is advisable especially when the between-herd prevalence of S. agalactiae is low.     

Comparison of diagnostic tests for the detection of equine infectious anemia antibody

Two diagnostic tests are approved for detecting antibody to equine infectious anemia virus: the agar-gel immunodiffusion (AGID) test and the competitive enzyme-linked immunosorbent assay (ELISA). A total of 420 sera from National Veterinary Services Laboratories check sets were tested with the AGID and competitive ELISA. A 100% correlation was obtained. The AGID and competitive ELISA were further used to test difficult samples with low levels of equine infectious anemia antibody (weak positives). A third test (Western blot) was also used with these weak positive samples to resolve any discordant results.     

Bayesian latent class evaluation of three tests for the screening of subclinical caprine mastitis in Bangladesh

Routine monitoring for subclinical infection is one of the key mastitis control approaches. However, the accuracy of the most commonly used screening tests has not yet been established. The aim of the present study was therefore to evaluate the accuracy of three screening tests, namely California mastitis test (CMT), white side test (WST), and surf field mastitis test (SFMT) for the screening of subclinical caprine mastitis. A cross-sectional study based on 484 randomly collected milk (242 goats) samples from three districts of Bangladesh was conducted for the screening of subclinical mastitis by the aforementioned tests. The Bayesian latent class model was implemented in WinBUGS to estimate the tests’ characteristics and true prevalence of subclinical mastitis. The Bayesian posterior estimates of sensitivities with a 95% credible intervals (CrIs) were 98.60% (95.18–99.95%), 98.28% (94.56–99.92%), and 89.98% (83.39–95.03%), and specificities with 95% CrIs were 99.19% (98.11–99.96%), 99.27% (97.34–99.98%), and 99.28% (97.35–99.98%), respectively for CMT, WST, and SFMT. The true prevalence of subclinical caprine mastitis was estimated to be 43.49% (95% CrI 37.46–48.98%). The positive predictive values (PPV) of the three tests were similar. The serial and parallel interpretation of any test pairs increased the PPV and negative predictive value respectively close to 100%. Based on the simplicity, cost and performance as well WST and SFMT simultaneously could be recommended for the screening of caprine subclinical mastitis in Bangladesh.

     

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.     

Use of a constant-virus diluting-serum microneutralization technique for the detection and quantification of infectious bronchitis virus antibodies

A constant-virus diluting-serum microneutralization test (CVMNT) for avian infectious bronchits virus (IBV) was evaluated for both reliability and repeatability. The virus used in the assay was a chick kidney (CK) cell-adapted strain, the Beaudette strain (IBV 42). Sera tested were from 24-week-old broiler-breeder chickens that had been vaccinated 3 times from a combination vaccine of Newcastle disease virus (NDV) and IBV. Test results were not repeatable or comparable when the same sera were tested on different days, but test results were repeatable and comparable when the sera were tested on the same day. Differences in virus titer at the different times that tests were performed appeared to cause the variation in test results. A comparison was made between the CVMNT and a constant-serum diluting-virus microneutralization test (CSMNT). The CVMNT was able to detect differences in flock antibody titers that the CSMNT could not.     

Comparison and evaluation of three serological tests for detection of antibodies to infectious bovine rhinotracheitis

Abstract

AIMS: To retrospectively describe clinical features of dogs that were presented to a small animal clinic between 2003–10 with macroscopic haematuria, and investigate whether signalment of the dog and severity and duration of the haematuria at admission were associated with specific aetiologies.

METHODS: Medical records were evaluated of 162 dogs with macroscopic haematuria admitted to a University-based small animal clinic in Thessaloniki, Greece, from January 2003 to December 2010. The inclusion criteria were discolouration of the urine sediment combined with abnormal numbers of erythrocytes, when examined microscopically. Data collected from the medical records included signalment, severity, frequency and duration of haematuria, and diagnosis.

RESULTS: Between January 2007 and December 2010, 8,893 dogs were admitted to the clinic; of these 99 (1.1%) were admitted with haematuria. Of the 162 dogs with records of haematuria, 80 (49.4%) were aged between 5.1–10 years, presented with acute (96/162; 59.3%), constant (99/162; 61.1%) and mild/moderate (150/162; 92.6%) haematuria. Of 147 dogs with a recorded diagnosis, the commonest diagnoses were urinary tract infection (UTI, 42/147; 28.6%), urolithiasis (38/147; 25.9%), prostatic disease (25/147; 17.0%) and urinary tumours (13/147; 8.8%). The prevalence of UTI was higher in female (22/56; 39%) than male (20/91; 22%) dogs, and in medium sized (22/52; 42%) than small (6/40; 15%) dogs. Urolithiasis was most prevalent in small (21/40; 52.5%) dogs, and all dogs with urolithiasis presented with mild/moderate haematuria. The prevalence of prostatic disease was highest in large (11/46; 24%) and giant (3/9; 33%) sized dogs and in dogs aged >10 years (8/30; 27%).

CONCLUSIONS AND CLINICAL RELEVANCE: In this retrospective study from one small animal clinic, UTI, urolithiasis, prostatic disease and urinary tumours predominated among the causes of canine haematuria. The consideration of sex, age, and size of the dog and characteristics of haematuria were found to be useful parameters when forming the list of differential diagnoses.     

Comparison of a microneutralisation test with ELISA and precipitin tests for detection of antibodies to infectious bronchitis virus in chickens

A microneutralisation test for infectious bronchitis virus using virus antigens available in Australia, cell culture medium containing low concentrations of serum and an elevated incubation temperature is described. The technique was economical on reagents and of comparable sensitivity to the enzyme-linked immunosorbent assay (ELISA). The value of the microneutralisation, ELISA and precipitin tests in assessing the serological response of a flock of commercial chickens to vaccine and natural virus challenge was determined.     

Development and evaluation of an ELISA for the detection of antibody to infectious laryngotracheitis virus in chickens

An enzyme-linked immunosorbent assay (ELISA) for the detection of antibody to infectious laryngotracheitis (ILT) virus in chickens was developed and compared with the serum-neutralization assay. The ELISA routinely yielded 16-to-32-fold higher titers than the serum-neutralization test. To overcome the requirement for large amounts of purified viral antigen, the microtiter trays were initially coated with an antibody prepared against purified ILT virus. A relatively crude viral preparation could then be used to coat the trays. Sera from specific-pathogen-free chickens less than 12 weeks of age did not show nonspecific binding, although 2.7% of all sera from chickens between 13 and 64 weeks of age had nonspecific activity. The majority of nonspecific reactors came from one highly inbred flock of specific-pathogen-free chickens. A number of modifications of ELISA procedures reported to reduce the nonspecific binding of chicken sera were investigated. Treatment of the serum or the plate and changes in the composition of the diluent did not increase the relative sensitivity of the anti-ILT assay.     

The persistence of infectious pancreatic necrosis virus in Atlantic salmon

Atlantic salmon leucocytes from Infectious Pancreatic Necrosis Virus (IPNV) carriers showed a suppressed response to phytohemagglutinin stimulation compared with uninfected controls. A significant degree of inhibition of DNA synthesis was observed using 3H-thymidine incorporation. IPNV was isolated from 41% of the stimulated leucocyte cultures supernatants, while only 6% of the unstimulated cultures were found to be positive.     

Correlation of virus replication in tissues with histologic lesions in Atlantic salmon experimentally infected with infectious salmon anemia virus

We have studied the replication of virus in tissues and development of lesions associated with infectious salmon anemia virus (ISAV) infection in Atlantic salmon using in situ hybridization (ISH) with a riboprobe targeting ISAV RNA segment 7 messenger RNA. Fish were infected with three ISAV isolates (U5575-1, RPC-01-0593-1, Norway 810/9/99) and then euthanatized sequentially at 3, 6, 10, and 13 days postinoculation (dpi) and thereafter once a week for 8 weeks. Severe histopathologic lesions were observed in tissues from all groups beginning at the onset of mortality. The severe histopathologic lesions correlated with maximum intensity and frequency of ISH signals (P < 0.001). There was a strong association between the hybridization signals and severity of lesions in the liver, kidney, and heart (R = 0.81, 0.70, and 0.78, respectively; P < 0.001). The distribution of ISH signals indicated the presence of a viremia because signals were observed predominantly in individual blood cells and endothelial cells, and possibly hematopoietic cells of head kidney, but not in the necrotic hepatocytes and renal epithelium. Of the organs sampled, the heart was the first and last to show ISH signals, possibly because of increased activity of the endocardial endothelial cells and the underlining macrophages, which continuously trap and remove circulating virus, and therefore represents the best tissue sample for screening of suspected infected fish. On the basis of mortality, severity of lesions, and intensity and frequency of ISH signals, ISAV isolate Norway 810/9/99 was the most virulent and U5575-1 the least virulent isolate studied.     

Performances of different diagnostic tests for feline infectious peritonitis in challenging clinical cases

Objectives : Feline infectious peritonitis (FIP) can be difficult to diagnose. Histopathology is considered the gold standard test but immunohistochemistry (IHC) is mandatory to confirm/exclude the disease. This study aimed to assess the performances of tests carried out in vivo or at postmortem examination in challenging cases in which FIP was confirmed or excluded based on IHC or on adequate follow‐up. Methods : Twelve cases (four without FIP, eight with FIP) were retrospectively studied. Clinical findings, serum protein electrophoresis (SPE), analysis of the effusions (AE), antifeline coronavirus serology, serum concentration of α1‐acid glycoprotein (AGP) and histopathology were classified as consistent, doubtful or non‐consistent with FIP. Sensitivity, specificity and concordance (κ) with the final diagnosis were calculated. Results : Concordance was absent for serology (κ=?0·08) and AE (κ=?0·52), poor for histopathology (κ=0·09), fair for SPE (κ=0·25) and perfect for AGP (κ=1·00). Sensitivity was high for AGP (100%) and low for AE (50%), SPE (37·5%) and histopathology (37·5%). Specificity was high for AGP or histopathology (100%) and low for SPE (50%) and AE (0%). Clinical Significance : IHC must always be performed to confirm FIP. If this is not possible, when histopathology is controversial, elevated AGP concentrations may support the diagnosis of FIP.     

Epidemiological investigation of infectious salmon anaemia (ISA) outbreaks in Norway 2003-2005

Epidemiological information was summarized from 32 outbreaks of infectious salmon anaemia (ISA) on salmon farming sites in Norway in 2003–2005. Virus isolates from the outbreak sites were genotyped, and the genotyping was used to assess possible associations between outbreak sites due to adjacent location, sharing fish farming authorisation, sharing smolt suppliers or sharing broodfish origin of the fish. The ISA outbreaks were distributed along most of the Norwegian coast and showed a variable clinical picture. The virus genotypes clustered into three genogroups. Pairs of outbreak sites matched for adjacent location or registered under the same authorisation, all shared genogroup, which was a significantly higher number of corresponding genogroups than expected by chance. For outbreak sites sharing smolt suppliers, corresponding genogroups appeared in 7 out of 12 matched pairs, which was not significant. An evaluation of broodfish origin associated with genogroups did not support transmission linked to broodfish origin. In conclusion, genotyping of virus isolates from ISA outbreaks supports associations between adjacent outbreaks. This is consistent with horizontal transmission. The present study failed to find evidence for vertical transmission (patterns of genogroups related to smolt suppliers or broodfish companies were not identified).