Effects of lng Mutations on LngA Expression, Processing, and CS21 Assembly in Enterotoxigenic Escherichia coli E9034A

Enterotoxigenic Escherichia coli (ETEC) is a major cause of morbidity in children under 5 years of age in low- and middle-income countries and a leading cause of traveler's diarrhea worldwide. The ability of ETEC to colonize the intestinal epithelium is mediated by fimbrial adhesins, such as CS21 (Longus). This adhesin is a type IVb pilus involved in adherence to intestinal cells in vitro and bacterial self-aggregation. Fourteen open reading frames have been proposed to be involved in CS21 assembly, hitherto only the lngA and lngB genes, coding for the major (LngA) and minor (LngB) structural subunit, have been characterized. In this study, we investigated the role of the LngA, LngB, LngC, LngD, LngH, and LngP proteins in the assembly of CS21 in ETEC strain E9034A. The deletion of the lngA, lngB, lngC, lngD, lngH, or lngP genes, abolished CS21 assembly in ETEC strain E9034A and adherence to HT-29 cells was reduced 90%, compared to wild-type strain. Subcellular localization prediction of CS21 proteins was similar to other well-known type IV pili homologues. We showed that LngP is the prepilin peptidase of LngA, and that ETEC strain E9034A has another peptidase capable of processing LngA, although with less efficiency. Additionally, we present immuno-electron microscopy imagens to show that the LngB protein could be localized at the tip of CS21, and probably helps to control CS21 length. In conclusion, our results demonstrate that the LngA, LngB, LngC, LngD, LngH, and LngP proteins are essential for CS21 assembly, as well as for bacterial aggregation and adherence to HT-29 cells

Role of Sex Steroid Hormones in Bacterial-Host Interactions

Sex steroid hormones play important physiological roles in reproductive and nonreproductive tissues, including immune cells. These hormones exert their functions by binding to either specific intracellular receptors that act as ligand-dependent transcription factors or membrane receptors that stimulate several signal transduction pathways. The elevated susceptibility of males to bacterial infections can be related to the usually lower immune responses presented in males as compared to females. This dimorphic sex difference is mainly due to the differential modulation of the immune system by sex steroid hormones through the control of proinflammatory and anti-inflammatory cytokines expression, as well as Toll-like receptors (TLRs) expression and antibody production. Besides, sex hormones can also affect the metabolism, growth, or virulence of pathogenic bacteria. In turn, pathogenic, microbiota, and environmental bacteria are able to metabolize and degrade steroid hormones and their related compounds. All these data suggest that sex steroid hormones play a key role in the modulation of bacterial-host interactions

The ATPase FliI Can Interact with the Type III Flagellar Protein Export Apparatus in the Absence of Its Regulator, FliH

Salmonella FliI is the ATPase that drives flagellar protein export. It normally exists as a complex together with the regulatory protein FliH. A fliH null mutant was slightly motile, with overproduction of FliI resulting in substantial improvement of its motility. Mutations in the cytoplasmic domains of FlhA and FlhB, which are integral membrane components of the type III flagellar export apparatus, also resulted in substantially improved motility, even at normal FliI levels. Thus, FliH, though undoubtedly important, is not essential

Type Three Secretion System in Attaching and Effacing Pathogens

Enteropathogenic Escherichia coli and enterohemorrhagic E. coli are diarrheagenic bacterial human pathogens that cause severe gastroenteritis. These enteric pathotypes, together with the mouse pathogen Citrobacter rodentium, belong to the family of attaching and effacing pathogens that form a distinctive histological lesion in the intestinal epithelium. The virulence of these bacteria depends on a type III secretion system (T3SS), which mediates the translocation of effector proteins from the bacterial cytosol into the infected cells. The core architecture of the T3SS consists of a multi-ring basal body embedded in the bacterial membranes, a periplasmic inner rod, a transmembrane export apparatus in the inner membrane, and cytosolic components including an ATPase complex and the C-ring. In addition, two distinct hollow appendages are assembled on the extracellular face of the basal body creating a channel for protein secretion: an approximately 23 nm needle, and a filament that extends up to 600 nm. This filamentous structure allows these pathogens to get through the host cells mucus barrier. Upon contact with the target cell, a translocation pore is assembled in the host membrane through which the effector proteins are injected. Assembly of the T3SS is strictly regulated to ensure proper timing of substrate secretion. The different type III substrates coexist in the bacterial cytoplasm, and their hierarchical secretion is determined by specialized chaperones in coordination with two molecular switches and the so-called sorting platform. In this review, we present recent advances in the understanding of the T3SS in attaching and effacing pathogens

Sexual dimorphism in bacterial infections

Abstract Background Sex differences are important epidemiological factors that impact in the frequency and severity of infectious diseases. A clear sexual dimorphism in bacterial infections has been reported in both humans and animal models. Nevertheless, the molecular mechanisms involved in this gender bias are just starting to be elucidated. In the present article, we aim to review the available data in the literature that report bacterial infections presenting a clear sexual dimorphism, without considering behavioral and social factors. Main body The sexual dimorphism in bacterial infections has been mainly attributed to the differential levels of sex hormones between males and females, as well as to genetic factors. In general, males are more susceptible to gastrointestinal and respiratory bacterial diseases and sepsis, while females are more susceptible to genitourinary tract bacterial infections. However, these incidences depend on the population evaluated, animal model and the bacterial species. Female protection against bacterial infections and the associated complications is assumed to be due to the pro-inflammatory effect of estradiol, while male susceptibility to those infections is associated with the testosterone-mediated immune suppression, probably via their specific receptors. Recent studies indicate that the protective effect of estradiol depends on the estrogen receptor subtype and the specific tissue compartment involved in the bacterial insult, suggesting that tissue-specific expression of particular sex steroid receptors contributes to the susceptibility to bacterial infections. Furthermore, this gender bias also depends on the effects of sex hormones on specific bacterial species. Finally, since a large number of genes related to immune functions are located on the X chromosome, X-linked mosaicism confers a highly polymorphic gene expression program that allows women to respond with a more expanded immune repertoire as compared with men. Conclusion Notwithstanding there is increasing evidence that confirms the sexual dimorphism in certain bacterial infections and the molecular mechanisms associated, further studies are required to clarify conflicting data and to determine the role of specific hormone receptors involved in the gender bias of bacterial infections, as well as their potential as therapeutic targets

The Flagellar Muramidase from the Photosynthetic Bacterium Rhodobacter sphaeroides▿

We have characterized open reading frame RSP0072, which is located within the flgG operon in Rhodobacter sphaeroides. The amino acid sequence analysis of this gene product showed the presence of a soluble lytic transglycosylase domain. The deletion of the N-terminal region (90 amino acids) of the product of RSP0072 yields a leaky nonmotile phenotype, as determined by swarm assays in soft agar. Electron micrographs revealed the lack of flagella in mutant cells. The purified wild-type protein showed lytic activity on extracts of Micrococcus luteus. In contrast, no lytic activity was observed when the residues E57 or E83 were replaced by alanine. Affinity blotting suggests that the protein encoded by RSP0072 interacts with the flagellar rod-scaffolding protein FlgJ, which lacks the muramidase domain present in FlgJ of many bacteria. We propose that the product of RSP0072 is a flagellar muramidase that is exported to the periplasm via the Sec pathway, where it interacts with FlgJ to open a gap in the peptidoglycan layer for the subsequent penetration of the nascent flagellar structure

CesL Regulates Type III Secretion Substrate Specificity of the Enteropathogenic E. coli Injectisome

The type III secretion system (T3SS) is a complex molecular device used by several pathogenic bacteria to translocate effector proteins directly into eukaryotic host cells. One remarkable feature of the T3SS is its ability to secrete different categories of proteins in a hierarchical manner, to ensure proper assembly and timely delivery of effectors into target cells. In enteropathogenic Escherichia coli, the substrate specificity switch from translocator to effector secretion is regulated by a gatekeeper complex composed of SepL, SepD, and CesL proteins. Here, we report a characterization of the CesL protein using biochemical and genetic approaches. We investigated discrepancies in the phenotype among different cesL deletion mutants and showed that CesL is indeed essential for translocator secretion and to prevent premature effector secretion. We also demonstrated that CesL engages in pairwise interactions with both SepL and SepD. Furthermore, while association of SepL to the membrane does not depended on CesL, the absence of any of the proteins forming the heterotrimeric complex compromised the intracellular stability of each component. In addition, we found that CesL interacts with the cytoplasmic domains of the export gate components EscU and EscV. We propose a mechanism for substrate secretion regulation governed by the SepL/SepD/CesL complex