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

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.