Improving noninvasive diagnosis of cutaneous and gastrointestinal forms of Pythium insidiosum infections in dogs: complementary roles of serum-based ELISA and polymerase chain reaction.

Abstract

OBJECTIVE: To evaluate the diagnostic performance of ELISA and PCR for detecting pathogenic oomycete infections and to explore associations with clinical presentation and treatment status. METHODS: Canine sera (n = 796) submitted to Pythium Serology Diagnostic Laboratory (from 2023 through 2025) were analyzed by ELISA to detect anti-Pythium antibodies and a PCR targeting Pythium insidiosum, Lagenidium spp, and Paralagenidium spp. Clinical records were reviewed for a subset of submissions. Analyses included χ2 tests, a linear mixed-effects model, and Cohen κ for ELISA-PCR agreement. RESULTS: Across all 697 samples tested by both methods, 69 (9.9%) were positive and 495 (71.0%) were negative by both assays, yielding an overall concordance of 81%. Polymerase chain reaction identified 5 cases, including cutaneous infections and Lagenidium infections tested to be negative by ELISA. Among confirmed positive submissions, ELISA detected anti-Pythium antibodies in 99.4% (182 of 183), whereas PCR detected Pythium DNA in 45.6% (72 of 158). CONCLUSIONS: While ELISA is well suited for first-line screening due to its high sensitivity, PCR provides critical specificity and broader detection, especially in challenging cutaneous cases and when alternative pathogenic oomycetes are suspected. Enzyme-linked immunosorbent assay was highly effective for initial diagnosis and monitoring treatment response, and PCR was most useful for identifying active infections or relapses. CLINICAL RELEVANCE: The combined use of ELISA and PCR enhances diagnostic accuracy, improves detection of misdiagnosed Lagenidium infections, and guides clinical decision-making and epidemiological surveillance of this emerging and serious disease.