Sulfate Import in <i>Salmonella</i> Typhimurium Impacts Bacterial Aggregation and the Respiratory Burst in Human Neutrophils

Abstract

During enteric salmonellosis, neutrophil-generated reactive oxygen species alter the gut microenvironment, favoring survival of <i>Salmonella</i> Typhimurium. While type 3 secretion system 1 (T3SS-1) and flagellar motility are potent <i>Salmonella</i> Typhimurium agonists of the neutrophil respiratory burst <i>in vitro,</i> neither of these pathways alone is responsible for stimulation of a maximal respiratory burst. To identify <i>Salmonella</i> Typhimurium genes that impact the magnitude of the neutrophil respiratory burst, we performed a two-step screen of defined mutant libraries in coculture with human neutrophils. We first screened <i>Salmonella</i> Typhimurium mutants lacking defined genomic regions and then tested single-gene deletion mutants representing particular regions under selection. A subset of single-gene deletion mutants was selected for further investigation. Mutants in four genes, <i>STM1696</i> (<i>sapF</i>), <i>STM2201</i> (<i>yeiE</i>), <i>STM2112</i> (<i>wcaD</i>), and <i>STM2441</i> (<i>cysA</i>), induced an attenuated respiratory burst. We linked the altered respiratory burst to reduced T3SS-1 expression and/or altered flagellar motility for two mutants (Δ<i>STM1696</i> and Δ<i>STM2201</i>). The Δ<i>STM2441</i> mutant, defective for sulfate transport, formed aggregates in minimal medium and adhered to surfaces in rich medium, suggesting a role for sulfur homeostasis in the regulation of aggregation/adherence. We linked the aggregation/adherence phenotype of the Δ<i>STM2441</i> mutant to biofilm-associated protein A and flagellins and hypothesize that aggregation caused the observed reduction in the magnitude of the neutrophil respiratory burst. Our data demonstrate that <i>Salmonella</i> Typhimurium has numerous mechanisms to limit the magnitude of the neutrophil respiratory burst. These data further inform our understanding of how S<i>almonella</i> may alter human neutrophil antimicrobial defenses.