Comparison of Nucleic Acid Amplification, Serology, and Microbiologic Culture for Diagnosis ofRhodococcus equiPneumonia in Foals

Abstract

Recently, a technique was described for amplification of Rhodococcus equi-specific chromosomal and vapA DNA from blood and tracheal wash fluids. It was hypothesized that this technique would be more sensitive than standard culture techniques or serology for diagnosis of R. equi pneumonia in foals. Tracheal wash fluid, nasal swabs, whole blood samples, and serum samples from 56 foals with pneumonia were analyzed. Final clinical diagnosis was determined by the attending clinician on the basis of final interpretation of all available information about each foal, including clinical presentation, diagnostic test results, response to therapy, and outcome. Clinical diagnosis was used as a final reference standard for calculation of sensitivity, specificity, and predictive values for PCR, serology using an agar gel immunodiffusion test, and tracheal wash fluid culture. PCR of tracheal wash fluid using primers that recognized the vapA virulence plasmid of R. equi had a diagnostic sensitivity of 100% and specificity of 90.6%. Sensitivity and specificity were 57.1 and 93.8%, respectively, for standard microbiologic culture of tracheal wash fluid and 62.5 and 75.9%, respectively, for serology. PCR of tracheal wash fluid is more sensitive and specific for diagnosis of R. equi pneumonia than are other available diagnostic tests.Rhodococcus equi is a gram-positive, pleomorphic coccobacillus that causes pneumonia and enteritis in foals less than 6 months of age and has been associated with a variety of suppurative processes in immune-suppressed humans (19). The organism is worldwide in distribution and commonly isolated from soil and environmental samples (2,4,23). Strains of R. equi isolated from sick foals uniformly contain an 85-to 90-kb plasmid that carries the gene responsible for expression of a 15-to 17-kDa antigen of undetermined function (vapA) (24,25). Environmental strains of R. equi not associated with equine disease do not contain this plasmid.The onset of clinical signs of R. equi pneumonia in foals is often insidious, and the infection is not recognized until severe abscessation has occurred and prognosis is poor. Culture of the organism from tracheal wash (TW) fluid is currently considered the "gold standard" for diagnosis (7). However, it can be difficult to reliably grow R. equi from a single TW sample, possibly because of prior antibiotic administration (12,14) or the presence of multiple pathogenic bacterial species (5, 21). Hillidge reported that only 7 of 11 foals (62%) with positive R. equi cultures at necropsy, and 57 of 89 foals (64%) with radiographic evidence of lung abscessation, yielded R. equi on culture of TW (10). Other studies reported a 100% positive result on cultures of TW fluid from foals later confirmed to have R. equi pneumonia at necropsy (14,17).Recently, a technique was described for amplification of R. equi-specific chromosomal and vapA DNA from blood and TW fluids (20). It was hypothesized that this technique would be more sensitive than standard culture techniques or serology for dia...

Internally Controlled Real-Time PCR Method for Quantitative Species-Specific Detection and vapA Genotyping of Rhodococcus equi

“…However, these culture-based procedures are lengthy and sometimes lack adequate sensitivity due either to prior antibiotic treatments or, in the case of respiratory specimens (typically bronchoalveolar lavage [BAL] aspirate or sputum), to the presence of multiple bacterial contaminants (28). An added problem is the difficulties posed by the identification due to the micro-and macroscopic morphological variability exhibited by these bacteria and the lack of accuracy of biochemical tests for R. equi species determination (8,15,29,36).…”

Current Understanding of Rhodococcus Equi infection and its Zoonotic Implications

“…PCR assays are employed for detection R. equi infection, which are rapid, sensitive and specific (Sellon et al, 2001;Arriaga et al, 2002;Ladro´n et al, 2003;Pusterla et al, 2007;Letek et al, 2008). The virulence of R. equi is associated with plasmids encoding virulence-associated proteins predominantly protein A (VapA) or protein B (VapB) (Letek et al, 2008;Takai et al, 1991b).…”

Pathogenicity of Rhodococcus equi in mice, isolated from environment, human and horse clinical samples

“…It has been recognized as an important pathogen due to reports of 3% of foal deaths widespread in the world (Takai et al 1993, Sellon et al 2001. R. equi is a major lung pathogen in less than 6-month-old foals with high mortality (Ozgur et al 2000).…”