Table 1_Development and validation of a reverse transcription-insulated isothermal PCR assay for detection of caprine kobuvirus and its epidemiological and molecular characterization in China.docx

We developed and validated a reverse transcription-insulated isothermal PCR (iiPCR) targeting the conserved 3D gene for detection of CKoV. The assay demonstrated high analytical specificity, a limit of detection of 39.9 copies·μL⁻¹, and good repeatability.

A total of 722 goat fecal samples were collected from five farms in Sichuan, Chongqing, Yunnan, and Hunan in China and tested using the newly developed method. The overall CKoV positivity rate was 40.44% (292/722), and the farm-level positivity rate was 100% (5/5), with the highest rate in Hunan (84.00%, 21/25), followed by Yunnan (42.86%, 48/112), Sichuan (38.58%, 206/534), and Chongqing (33.33%, 17/51). Lambs exhibited substantially higher positivity rates (57.14–83.33%) than goats aged ≥3 months (13.39–15.52%), and chi-square test confirmed a statistically significant association between age and CKoV positivity (χ² = 252.97, df = 6, p < 0.001), confirming age-dependent susceptibility in lambs.