Detection of Shigella Species in Wastewater - A Pilot Study for Community and Building Scale Wastewater-Based Surveillance of Bacterial Pathogens - Project Summary / Abstract Bacillary dysentery or shigellosis is caused by bacteria of the Shigella species: S. dysenteriae, S. flexneri, S boydii, and S. sonnei. Infections range from mild or asymptomatic to severe bloody diarrhea, so reported prevalence grossly underestimates actual prevalence. Shigellosis is a global public health concern and antimicrobial resistance has compounded the problem. Moreover, shigellosis is also sexually transmitted in the United States, Europe, and other developed countries. Such outbreaks are driven by the emergence of antibiotic resistant strains of S. flexneri infecting men who have sex with men. The objective of this proof-of-concept study is to demonstrate that wastewater-based epidemiology (WBE) using digital PCR to detect pathogen molecular markers can provide a more accurate picture of community prevalence of bacterial pathogens than traditional case reporting. Wastewater surveillance for viral pathogens has been in use for decades and its use to monitor SARS-CoV-2 is now widely applied globally. Analogous methods for wastewater surveillance of bacterial pathogens by digital PCR has lagged. We will develop and validate methods to detect Shigella, an important bacterial agent of diarrheal disease, in wastewater. We will also field test the methods to assess shigellosis prevalence at community and building scale and link the data to the actual population residing in the wastewater collection area. We will provide actionable data to the local health department which can then develop targeted public health interventions. The independent but complementary aims in demonstrating the proof-of-concept are: 1. Develop and validate sensitive, specific, and reproducible WBE methods for detection of Shigella species in wastewater using digital PCR. Three subaims will: 1.1) optimize methods for extraction and detection of Shigella in wastewater using laboratory-grown strains of Shigella species to “spike” authentic wastewater and laboratory prepared “synthetic” wastewater; 1.2) validate molecular targets for differentiation of S. flexneri from S. sonnei in wastewater; and 1.3) culture of Shigella from wastewater to assess antibiotic resistance genotypes and phenotypes. 2. Assess proof-of-concept of WBE as a public health tool for Shigella at community and building scale with emphasis on high risk congregate populations, i.e., day care centers. Two subaims will: 2.1) measure the prevalence of Shigella species at community scale by sampling at the wastewater treatment plant intake and using wastewater flow rate as a population normalization marker; and 2.2) extend shigellosis surveillance to building scale targeting high risk congregate pediatric populations. Strengths of this proposal are the innovative application of WBE to a bacterial pathogen, our method for population normalization, differentiation of Shigella species, prevalence measurement of Shigella species at community and building scale, and the expertise of our multidisciplinary team.
