In 2017, diarrheal diseases caused mortality in half a million children under the age of five in the world's poorest countries. Epidemiologic data indicate that besides rotavirus (27%) and adenovirus (11.2%), the most common causes of bacterial diarrheal disease are Shigella (13%), Campylobacter jejuni (9%) and enterotoxigenic E. coli (4%). Shigella is the major group affecting children aged 12-36 months with moderate to severe disease and high mortality risk. There are four major Shigella species that comprise over 50 different serotypes and subtypes based on the conformational epitopes of the O polysaccharide antigen (O-antigen) of bacterial LPS, which is an important antigen.

Shigella must invade epithelial cells, multiply and spread intercellularly and ultimately cause cell death with the help of several proteins encoded mainly on a large virulence plasmid. The entry process requires the invasion plasmid antigens (Ipa antigens), and a gentamicin protection assay in Hela cells is a routine procedure for assessing the invasiveness of a Shigella strain. The infected cells can also be stained with Giemsa (Giemsa staining) and microscopically evaluated for a more qualitative analysis of the fate of the bacteria. The plaque assay measures epithelial cell killing and cell-to-cell spread in tissue culture. The Sereny test in the guinea pig measures the virulence of a strain to cause acute inflammatory disease. During a live or subunit Shigella vaccine trial, serum and mucosal antibody elicitation are evaluated using purified LPS and affinity-purified Ipa antigens. Mucosal antibodies are measured as antibody-secreting cells (ASC) in an ELISpot assay and antibody-in lymphocyte supernatant (ALS) assays with cultured PBMCs. Serum antibodies are evaluated for functionality in a serum bactericidal assay (SBA).

Investigators at the Walter Reed Army Institute of Research in MD, the University of MD at Baltimore, the University of Texas in Austin, Pasteur Institute in Paris, France, and the University of Florida in Gainesville, FL (and others if needed) will be invited to submit their protocols for these assays.