Environmental surveillance of soil-transmitted helminths and other enteric pathogens in settings without networked wastewater infrastructure

Soil-transmitted helminths (STH) are one of the most prevalent enteric infections world-wide. To control STH-related morbidity, the World Health Organization recommends targeted deworming and improvements in water, sanitation and hygiene. Current surveillance strategies for STH focus on identifying and quantifying eggs in stool samples via microscopy, which exhibits poor specificity and sensitivity, especially in settings with low-intensity infections. Wastewater-based epidemiology is a surveillance tool used to monitor pathogen circulation and could replace stool- based approaches for STH detection. However, sampling strategies for settings lacking networked sanitation outside large urban settlements are not well developed. Here, we report evaluation of sampling strategies for soil and wastewater STH surveillance in rural and peri-urban settings without networked sanitation. We used multi-parallel qPCR assays to detect STH DNA in soil collected from high foot-traffic locations and three types of wastewater samples (passive Moore swabs, grab samples, and sediment from drainage ditches) in Come, Benin and Timiri and Jawadhu Hills in Tamil Nadu, India. We detected STH in soil (India = 32/95, Benin = 39/121) and wastewater (India = 24/60, Benin = 8/64) with a detection frequency across all sample types of 36% in India and 25% in Benin. We evaluated which sample locations and types allowed for more sensitive detection of STH DNA and determined that STH prevalence varied by sample site but did not vary significantly within a given sample site location (e.g., samples collected from multiple locations within one market). Further, we determined that wastewater sediment samples outperformed grab and Moore swab sample types for STH detection. Finally, we expanded our methods to include detection of other enteric pathogens using multiplexed qPCR for wastewater samples. Our results establish sampling strategies for environmental and wastewater surveillance of a wide range of enteric pathogens in settings without networked sanitation.