Research note: occurrence, molecular characterization, and clinical associations of avian intestinal spirochetosis in laying hens in India.

Avian intestinal spirochetosis (AIS) is an enteric disease of laying hens caused by pathogenic Brachyspira species and is likely underrecognized in typical on-farm conditions. Despite India being a major egg-producing country, organized data on AIS in Indian poultry remain limited. This targeted investigation described Polymerase Chain Reaction (PCR) detection of Brachyspira spp., associated field observations, and preliminary species-level evidence in clinically suspected commercial layer flocks in South and North India. Cecal mucosal scrapings were collected from 59 recently deceased laying hens from 12 commercial farms (33 sheds; 25 to 105 weeks of age) and applied to FTA cards. Samples were screened by PCR targeting the Brachyspira 16S rRNA gene and the NADH oxidase (nox) gene; selected amplicons were sequenced and analyzed phylogenetically. Brachyspira spp. were detected in 83.3% of farms, 82% of sheds, and 67.8% of samples in this targeted investigation of clinically suspected flocks. Detection was numerically higher in older flocks, ranging from 37.5% in birds aged 25 to 40 weeks to 84.6% in birds aged 71 to 105 weeks; however, this age related trend was not statistically significant. PCR-positive flocks commonly exhibited loose droppings, wet litter, vent pasting, dirty, fecal-stained eggs (approximately 1 to 5%) of total eggs, and only modest reduction (3 to 5%) in egg production in about one-third of affected flocks. nox sequencing of selected PCR-positive samples identified sequences consistent with Brachyspira pilosicoli, Brachyspira intermedia, Brachyspira pulli, and Brachyspira alvinipulli, providing preliminary evidence of multiple Brachyspira spp. among sequenced samples.