Comparative performance of environmental sample types for lumpy skin disease virus surveillance in dairy systems

Lumpy Skin Disease Virus (LSDV) is a transboundary viral disease that causes substantial economic losses to the dairy industry. Although clinical surveillance is the primary approach to disease diagnosis at the time of outbreaks, it often fails to detect virus circulation before the appearance of clinical signs. Environmental surveillance offers a practical, non-invasive, and cost-effective alternative; however, standardized protocols for environmental sample collection and processing for LSDV are currently unavailable. A longitudinal surveillance study was conducted across Gujarat, India, from June 2025 to May 2026. Environmental samples, including drinking water (n = 633), manger swabs (n = 587), slurry (n = 624), leftover feed (n = 610), dung piles (n = 616), farm-pooled milk (n = 563), and Dairy Cooperative Society (DCS)-pooled milk (n = 2,230), were systematically collected and screened using quantitative real-time qPCR. At the time of the outbreak, clinical samples, including nasal swabs (n = 39), blood (n = 22), and individual animal milk (n = 18), were also collected, and data was correlated with corresponding environmental samples, drinking water n = 46, dung pile n = 49, leftover feed n = 10, manger swab n = 47, and farm pooled milk n = 32. In total, 6,598 samples were collected across 53 livestock farms and 3 livestock markets across 7 dairy production zones during the period of June 2025 to May 2026. These samples comprise 5,863 environmental samples and 472 nasal swabs collected through routine surveillance between June 2025- May 2026 and January to May 2026, respectively and 184 environmental samples and 79 clinical samples collected during outbreaks. During routine surveillance, drinking water exhibited the highest odds of LSDV detection (OR = 2.93, 95% CI: 2.21–3.88; q < 0.001), with a positivity rate of 35.23% (223/633). This was followed by leftover feed (OR = 2.86, 95% CI: 2.15–3.79; q < 0.001), slurry (OR = 2.81, 95% CI: 2.12–3.73; q < 0.001), and manger swabs (OR = 2.64, 95% CI: 1.98–3.52; q < 0.001), with positivity rates of 34.26% (209/610), 34.29% (214/624), and 33.05% (194/587), respectively. In comparison, farm-pooled milk, used as the reference category, showed the lowest positivity rate. Among the different samples collected from the affected farms during the outbreak period, manger swabs demonstrated the highest odds of LSDV detection (OR = 5.05, 95% CI: 1.52–16.69; q = 0.020)- 48.94% positivity (23/47), although the clinical samples showed overall higher positivity (70.88%). This study provides a standardized framework for environmental surveillance of LSDV, including optimal sample type and optimized preprocessing protocols for routine environmental surveillance of LSDV. These findings establish environmental surveillance as a practical complement to clinical surveillance for the early detection of LSDV circulation and improved outbreak preparedness in high-density dairy production systems. Not applicable