Serine protease autotransporters of Enterobacteriaceae (SPATEs) are secreted proteins that contribute to virulence and function as proteases, toxins, adhesins, and/or immunomodulators. An extra-intestinal pathogenic E. coli (ExPEC) O1:K1 strain, QT598, isolated from a turkey, was shown to contain vat, tsh , and three uncharacterized SPATE-encoding genes. Uncharacterized SPATEs: Sha ( S erine-protease h emagglutinin a utotransporter), TagB and TagC ( t andem a utotransporter g enes B and C ) were tested for activities including hemagglutination, autoaggregation, and cytotoxicity when expressed in E. coli K-12. Sha and TagB conferred autoaggregation and hemagglutination activities. TagB, TagC, and Sha all exhibited cytopathic effects on a bladder epithelial cell line. In QT598, tagB and tagC are tandemly encoded on a genomic island, and were present in 10% of UTI isolates and 4.7% of avian E. coli . Sha is encoded on a virulence plasmid and was present in 1% of UTI isolates and 20% of avian E. coli . To specifically examine the role of SPATEs for infection, the 5 SPATE genes were deleted from strain QT598 and tested for cytotoxicity. Loss of all five SPATEs abrogated the cytopathic effect on bladder epithelial cells, although derivatives producing any of the 5 SPATEs retained cytopathic activity. In mouse infections, sha gene-expression was up-regulated a mean of sixfold in the bladder compared to growth in vitro . Loss of either tagBC or sha did not reduce urinary tract colonization. Deletion of all 5 SPATEs, however, significantly reduced competitive colonization of the kidney supporting a cumulative role of SPATEs for QT598 in the mouse UTI model.

Autotransporters are secreted proteins with multiple functions produced by a variety of Gram-negative bacteria. In Enterobacteriaceae, a subgroup of these autotransporters are the SPATEs (serine protease autotransporters of Enterobacteriaceae). SPATEs play a crucial role in survival and virulence of pathogens such as Escherichia coli and Shigella spp. and contribute to intestinal and extra-intestinal infections. These high molecular weight proteases are transported to the external milieu by the type Va secretion system and function as proteases with diverse substrate specificities and biological functions including adherence and cytotoxicity. Herein, we provide an overview of SPATEs and discuss recent findings on the biological roles of these secreted proteins, including proteolysis of substrates, adherence to cells, modulation of the immune response, and virulence in host models. In closing, we highlight recent insights into the regulation of expression of SPATEs that could be exploited to understand fundamental SPATE biology.

Urinary tract infections (UTIs) are common bacterial infections and the vast majority of UTIs are caused by extraintestinal pathogenic Escherichia coli (ExPEC) strains referred to as uropathogenic E. coli (UPEC). Successful colonization of the human urinary tract by UPEC is mediated by secreted or surface exposed virulence factors—toxins, iron transport systems, and adhesins, such as type 1 fimbriae (pili). To identify factors involved in the expression of type 1 fimbriae, we constructed a chromosomal transcriptional reporter consisting of lux under the control of the fimbrial promoter region, fimS and this construct was inserted into the reference UPEC strain CFT073 genome at the att Tn7 site. This fimS reporter strain was used to generate a Tn 10 transposon mutant library, coupled with high-throughput sequencing to identify genes that affect the expression of type 1 fimbriae. Transposon insertion sites were linked to genes involved in protein fate and synthesis, energy metabolism, adherence, transcriptional regulation, and transport. We showed that YqhG, a predicted periplasmic protein, is one of the important mediators that contribute to the decreased expression of type 1 fimbriae in UPEC strain CFT073. The Δ yqhG mutant had reduced expression of type 1 fimbriae and a decreased capacity to colonize the murine urinary tract. Reduced expression of type 1 fimbriae correlated with an increased bias for orientation of the fim switch in the OFF position. Interestingly, the Δ yqhG mutant was more motile than the WT strain and was also significantly more sensitive to hydrogen peroxide. Taken together, loss of yqhG may decrease virulence in the urinary tract due to a decrease in production of type 1 fimbriae and a greater sensitivity to oxidative stress.

Urinary tract infections (UTIs) are a common bacterial infectious disease in humans, and strains of uropathogenic Escherichia coli (UPEC) are the most frequent cause of UTIs. During infection, UPEC must cope with a variety of stressful conditions in the urinary tract. Here, we demonstrate that the small RNA (sRNA) RyfA of UPEC strains is required for resistance to oxidative and osmotic stresses. Transcriptomic analysis of the ryfA mutant showed changes in expression of genes associated with general stress responses, metabolism, biofilm formation and genes coding for cell surface proteins. Inactivation of ryfA in UPEC strain CFT073 decreased urinary tract colonization in mice and the ryfA mutant also had reduced production of type 1 and P fimbriae (pili), adhesins which are known to be important for UTI. Furthermore, loss of ryfA also reduced UPEC survival in human macrophages. Thus, ryfA plays a key regulatory role in UPEC adaptation to stress, which contributes to UTI and survival in macrophages.