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Aetiology of Neonatal Infection in South Asia (ANISA) is a multicenter study in Bangladesh, India and Pakistan exploring the incidence and etiology of neonatal infections. A periurban site in Karachi was selected for its representativeness of the general population in neonatal health indicators. An established demographic surveillance system and other infrastructure needed for conducting the study already existed at this site. ANISA presents a unique challenge because of the need to capture every birth outcome in the community within a few hours of delivery to reliably estimate the incidence and etiology of early-onset sepsis in a setting where home births and deaths are common.Major challenges at the Karachi site are related to early birth reporting and newborn assessment for births outside the catchment areas, parental refusal to participate, diverse ethnicity of the population, collection of biological specimens from healthy controls, political instability and crime, power outages and blood culture contamination. Some of the remedial actions taken include prolonging working hours; developing counseling skills of field workers; hiring staff with different linguistic abilities from within the study community; liaising with health facilities, key community informants, Lady Health Workers and traditional birth attendants; hiring community mobilizers; enhancing community sensitization; developing contingency plans for field work interruptions and procuring backup generators. The specimen contamination rate has decreased through training, supervision and video monitoring of blood collection procedures with individualized counseling of phlebotomists.ANISA offers lessons for successful implementation of complex study protocols in areas of high child mortality and challenging social environments.