Concordance in diagnostic testing for respiratory pathogens of bighorn sheep

Abstract

Reliable diagnostic tests are essential for disease investigation and management. This is particularly true for diseases of free-ranging wildlife where sampling is logistically difficult precluding retesting. Clinical assays for wildlife diseases frequently vary among laboratories because of lack of appropriate standardized commercial kits. Results of diagnostic testing may also be called into question when investigators report different etiologies for disease outbreaks, despite similar clinical and pathologic findings. To evaluate reliability of diagnostic testing for respiratory pathogens of bighorn sheep (Ovis canadensis), we conducted a series of ring tests across 6 laboratories routinely involved in detection of Mycoplasma ovipneumoniae, Pasteurellaceae, lktA (the Pasteurellaceae gene encoding leukotoxin), and 3 reference laboratories. Consistency of results for replicate samples within laboratories was high (median agreement ¼ 1.0). Agreement between laboratories was high for polymerase chain reaction (PCR) detection of M. ovipneumoniae and culture isolation of Mannheimia spp. and Bibersteinia trehalosi (median agreement ¼ 0.89-0.95, Kappa ¼ 0.65-0.74), and lower for PCR detection of Mannheimia spp. lktA (median agreement ¼ 0.58, Kappa ¼ 0.12). Most errors on defined status samples were false negatives, suggesting test sensitivity was a greater problem than specificity. However, tests for M. haemolytica and lktA yielded some false positive results. Despite differences in testing protocols, median agreement among laboratories and correct classification of controls for most agents was !0.80, meeting or exceeding the standard required by federal proficiency testing programs. This information is valuable for interpreting test results, laboratory quality assessments, and advancing diagnosis of respiratory disease in wild sheep. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

Respiratory pathogens and their association with population performance in Montana and Wyoming bighorn sheep populations

“…This is most likely to have occurred with M . haemolytica , which tests often fail to distinguish from Mannheimia glucosida [ 41 , 42 ], but false positives could have occurred for other Pasteurellaceae or M . ovipneumoniae .…”

“…The primers and protocol for this PCR were described by Walsh et al . 2016 [ 42 ]. Our sampling approach did not allow evaluation of detection probability for this test.…”

Evidence for strain‐specific immunity to pneumonia in bighorn sheep

“…(Besser et al ) and performed PCR for lktA , the gene encoding leukotoxin A, the major virulence factor of Mannheimia spp. and B. trehalosi (Walsh et al ). If an agent was detected by either PCR or culture on any sample, we classified the animal as positive for that agent.…”

Pneumonia in bighorn sheep: Risk and resilience

“…to leukotoxin positive Pasteurellaceae, to a multi‐factorial respiratory disease complex (Besser et al ). Much attention has focused on virulent Pasteurellaceae bacteria where problems with accurate detection and classification have also complicated efforts to establish an association with pneumonia outbreaks in wild sheep (Angen et al , Walsh et al , Miller et al , Shanthalingam et al , Walsh et al ).…”

“…Nasal swabs from ≥18 animals should be adequate (85% probability) to detect shedding by PCR in bighorn sheep populations with M. ovipneumoniae prevalence of ≥0.10. Larger sample sizes may be required to account for non‐detection error associated with field sampling and diagnostic testing (Walsh et al ). Strain‐type can be identified in PCR‐positive samples.…”