Gut Inflammatory Proteins Associated with Environmental Enteric Dysfunction Can Be Measured in Wastewater

Abstract

Introduction

Inhabitants of low-income countries in areas with poor sanitation suffer from high rates of Environmental Enteric Dysfunction (EED), a subclinical syndrome hallmarked by chronic inflammation of the small intestine driven by repeat and chronic enteric pathogen exposure. EED has been associated with poor linear growth and adverse neurodevelopmental outcomes in children. In adults, it has been hypothesized that EED plays a pathogenic role in obesity and metabolic syndrome. While small intestinal histology is the gold standard for EED diagnosis, protein biomarkers of enteric inflammation have been widely used as a proxy measure of EED. One of the major challenges in using such biomarkers for research is that these biomarkers are dynamic, responding to repeat pathogen exposures and clearance. This makes understanding the cumulative burden of EED and its effects on long-term outcomes difficult. Further, understanding the effect of interventions on EED burden is difficult as repeated measures in individuals may not give an accurate picture. We hypothesized that we could measure the community level of EED by quantifying levels of known fecal biomarkers of EED in wastewater.

Methods

Using commercially available ELISA kits, we attempted to measure alpha-1-antitrypsin, calprotectin, neopterin, myeloperoxidase, and neutrophil gelatinase-associated lipocalin in wastewater. The sewage samples were collected between 7 AM-9 AM from established sites in an existing sewage surveillance study in Dhaka, Bangladesh, where the EED burden is known to be high. The catchment population ranged from 2,317 to 606,332. The samples were processed at our icddr,b lab that same day. 50-100 µl of neat sewage samples were tested using ELISA kits for each EED biomarker.

Results

We detected antitrypsin (range 10.2-44.4 ng/l; mean 29.42±12.15 SD ng/l), neopterin (0-0.64 ng/µl; 0.03±0.03 ng/µl), and NGAL (0.6-1.1 ng/µl; 0.59±0.35 ng/µl) from neat samples. Calprotectin and MPO were not detected from the neat sample. We plan to concentrate the samples to detect these biomarkers in the future.

Discussion

These results demonstrate that biomarkers of EED can be measured in wastewater in a community known to have a high EED burden. The next steps involve comparing these results with those from a low EED burden catchment area (i.e. high-income country) and determining the variation in these wastewater biomarkers over time.

Conclusion

In the long term, community-level EED measures can be correlated to population health outcomes and be used to measure response to both sanitation and pathogen-targeted interventions such as vaccines.