Performance of Fecal Inflammatory Biomarkers to Identify Watery Shigellosis: Findings from the Enterics for Global Health (EFGH) Shigella Surveillance Study

BackgroundCurrent syndromic guidelines for diarrhea treatment miss watery Shigella cases, leading to undertreatment of children who may benefit. Incorporating fecal inflammatory biomarkers into diagnosis may improve case identification. MethodsWe conducted an ancillary analysis using samples from six sites (The Gambia, Kenya, Malawi, Bangladesh, Pakistan, and Peru) from the Enterics for Global Health (EFGH)-Shigella surveillance study, a facility-based hybrid study of children aged 6-35 months with diarrhea. Four fecal biomarkers were quantified by enzyme-linked immunosorbent assays at enrollment: myeloperoxidase, calprotectin, neutrophil gelatinase-associated lipocalin (lipocalin-2), and hemoglobin. An ensemble model with leave-one-site-out cross-validation was used to predict watery shigellosis, incorporating biomarkers and nine clinical and socio-economic predictors. We compared the predictive performance of the algorithm using: a) all predictors (including biomarkers); b) all non-biomarker predictors; c) all predictors (with selected biomarkers). ResultsBetween June 2022 and August 2024, a total of 4,191/9,476 (44.2%) children presented with watery diarrhea and had their whole stool tested for the biomarkers and 4,083 stool samples or rectal swabs were tested by qPCR; 735 (18.0%) had Shigella-attributable diarrhea by qPCR. The full model incorporating all 13 predictors achieved an area under the curve (AUC) of 0.75 [95% CI: 0.67-0.78], with a sensitivity of 0.67 and specificity of 0.75. Excluding biomarkers reduced model performance by 8% (AUC 0.67, 95% CI: 0.61-0.70). Adding hemoglobin alone improved the models discriminatory ability by 7%, while further adding myeloperoxidase had marginal contribution (1%), and lipocalin-2 (0%) and calprotectin none (0%). ConclusionFecal hemoglobin substantially improved prediction scores for watery shigellosis. Consequently, implementation of point-of-care assays for hemoglobin could improve clinical diagnosis in these settings and inform appropriate antibiotic treatment. Author SummaryDiarrhea remains a leading cause of illness in young children, with the bacteria Shigella being a major contributor. Current treatment guidelines recommend antibiotics only when diarrhea is visibly bloody. However, many children with Shigella infection have non-bloody, watery diarrhea and are therefore not treated, despite needing care. In this study, we explored whether simple markers of gut inflammation found in stool could improve the identification of Shigella infection. We analyzed stool samples from over 4,000 children across six countries in Africa, Asia, and South America and combined these markers with basic clinical information. We found that detecting small, invisible amounts of blood in stool (fecal hemoglobin) improved identification of non-bloody Shigella diarrhea by 7%. Other inflammation markers added little benefit. By combining fecal hemoglobin with childs age and stool frequency, we developed a simple score that matched performance of the full model enhancing its practical use. This approach outperformed current guidelines, which only treat bloody diarrhea, and was more accurate than treating most children without testing. Importantly, fecal hemoglobin tests are inexpensive and commercially available. Using them at the point of care could help health workers better target antibiotics, improve child outcomes, and promote responsible antibiotic use in low-resource settings.