Comparative proteomics of uropathogenic Escherichia coli during growth in human urine identify UCA-like (UCL) fimbriae as an adherence factor involved in biofilm formation and binding to uroepithelial cells

Uropathogenic Escherichia coli (UPEC) are the primary cause of urinary tract infection (UTI) in humans. For the successful colonisation of the human urinary tract, UPEC employ a diverse collection of secreted or surface-exposed virulence factors including toxins, iron acquisition systems and adhesins. In this study, a comparative proteomic approach was utilised to define the UPEC pan and core surface proteome following growth in pooled human urine. Identified proteins were investigated for subcellular origin, prevalence and homology to characterised virulence factors. Fourteen core surface proteins were identified, as well as eleven iron uptake receptor proteins and four distinct fimbrial types, including type 1, P, F1C/S and a previously uncharacterised fimbrial type, designated UCA-like (UCL) fimbriae in this study. These pathogenicity island (PAI)-associated fimbriae are related to UCA fimbriae of Proteus mirabilis, associated with UPEC and exclusively found in members of the E. coli B2 and D phylogroup. We further demonstrated that UCL fimbriae promote significant biofilm formation on abiotic surfaces and mediate specific attachment to exfoliated human uroepithelial cells. Combined, this study has defined the surface proteomic profiles and core surface proteome of UPEC during growth in human urine and identified a new type of fimbriae that may contribute to UTI

The construct of maternal positivity in mothers of children with intellectual disability

BACKGROUND: Despite the elevated levels of stress, anxiety and depression reported by mothers of children with intellectual disabilities (ID), these mothers also experience positive well‐being and describe positive perceptions of their child. To date, maternal positivity has been operationalised in different ways by using a variety of measures. In the present study, we tested whether a latent construct of maternal positivity could be derived from different measures of positivity. METHOD: One hundred and thirty‐five mothers of 89 boys and 46 girls with ID between 3 and 18 years of age completed measures on parental self‐efficacy, their satisfaction with life, family satisfaction, their positive affect and their positive perceptions of their child with ID. We conducted a confirmatory factor analysis of latent positivity and subsequently tested its association with child social skills and behaviour problems, and maternal mental health. RESULTS: A latent maternal positivity factor achieved a statistically good fit by using the five observed indicators of positivity. Parental self‐efficacy had the strongest loading on the latent factor. Maternal positivity was significantly negatively associated with maternal psychological distress, maternal stress and child problem behaviours and positively associated with child positive social behaviour. CONCLUSIONS: These findings lend support to the importance of examining parental positivity in families raising a child with ID, and using multiple indicators of positivity. Associations with negative psychological outcomes suggest that interventions focused on increasing parental positivity may have beneficial effects for parents. Further research is needed, especially in relation to such interventions

Types and correlates of school non-attendance in students with autism spectrum disorders

School non-attendance in autism spectrum disorders (ASD) has received very little attention to date. The study aimed to provide a comprehensive description of school non-attendance in students with ASD. Through an online survey, parents of 486 children (mean age 11 years) reported on school attendance over one month, and reasons for instances of non-attendance. On average, students missed five days of school of a possible 23 days. Persistent non-attendance (absent on 10%+ of available sessions) occurred among 43% of students. School non-attendance was associated with child older age, not living in a two-parent household, parental unemployment and, especially, attending a mainstream school. School refusal accounted for 43% of non-attendance. School exclusion and school withdrawal each accounted for 9% of absences. Truancy was almost non-existent. Non-problematic absenteeism (mostly related to medical appointments and illness) accounted for 32% of absences. Non-problematic absenteeism was more likely among those with intellectual disability, school refusal was more likely among older students, and school exclusion was more likely among students from single-parent, unemployed, and well educated households. Findings suggest school non-attendance in ASD is a significant issue, and that it is important to capture detail about attendance patterns and reasons for school non-attendance

Measurement invariance of the Positive Gains Scale in families of children with and without disabilities

Aims The present study sought to establish measurement invariance for the Positive Gains Scale (PGS) across 1219 mothers of children with developmental disabilities, 234 mothers of children with spina bifida/hydrocephalus, and 157 mothers of children without disabilities. Methods and procedures A three-step test for measurement invariance across the three groups was conducted using Multigroup Confirmatory Factor Analysis. Outcomes and results Loadings between the three groups were invariant, suggesting the criteria to assume metric invariance was met. However, the assumption of scalar invariance was not met, suggesting that item intercepts differed between the three groups. Conclusions and implications Our findings suggest that the PGS cannot be meaningfully used to compare outcomes between mothers of children with developmental disabilities and other mothers. These findings may have wider implications for research utilising well-being measures to make comparisons with carers of children with developmental disabilities

Molecular characterization of Escherichia coli strains that cause symptomatic and asymptomatic urinary tract infections

The differences between Escherichia coli strains associated with symptomatic and asymptomatic urinary tract infections (UTIs)remain to be properly determined. Here we examined the prevalence of plasmid types and bacteriocins, as well as genetic relatedness, in a defined collection of E. coli strains that cause UTIs. Comparative analysis identified a subgroup of strains with a high number of virulence genes (VGs) and microcins M/H47. We also identified associations between microcin genes, VGs, and specific plasmid types

Identification of genes important for growth of asymptomatic Bacteriuria Escherichia coli in urine

Escherichia coli is the most important etiological agent of urinary tract infections (UTIs). Unlike uropathogenic E. coli, which causes symptomatic infections, asymptomatic bacteriuria (ABU) E. coli strains typically lack essential virulence factors and colonize the bladder in the absence of symptoms. While ABU E. coli can persist in the bladder for long periods of time, little is known about the genetic determinants required for its growth and fitness in urine. To identify such genes, we have employed a transposon mutagenesis approach using the prototypic ABU E. coli strain 83972 and the clinical ABU E. coli strain VR89. Six genes involved in the biosynthesis of various amino acids and nucleobases were identified (carB, argE, argC, purA, metE, and ilvC), and site-specific mutants were subsequently constructed in E. coli 83972 and E. coli VR89 for each of these genes. In all cases, these mutants exhibited reduced growth rates and final cell densities in human urine. The growth defects could be complemented in trans as well as by supplementation with the appropriate amino acid or nucleobase. When assessed in vivo in a mouse model, E. coli 83972carAB and 83972argC showed a significantly reduced competitive advantage in the bladder and/or kidney during coinoculation experiments with the parent strain, whereas 83972metE and 83972ilvC did not. Taken together, our data have identified several biosynthesis pathways as new important fitness factors associated with the growth of ABU E. coli in human urine

O antigen biogenesis sensitises Escherichia coli K-12 to bile salts, providing a plausible explanation for its evolutionary loss

Published: October 4, 2023Escherichia coli K-12 is a model organism for bacteriology and has served as a workhorse for molecular biology and biochemistry for over a century since its first isolation in 1922. However, Escherichia coli K-12 strains are phenotypically devoid of an O antigen (OAg) since early reports in the scientific literature. Recent studies have reported the presence of independent mutations that abolish OAg repeating-unit (RU) biogenesis in E. coli K-12 strains from the same original source, suggesting unknown evolutionary forces have selected for inactivation of OAg biogenesis during the early propagation of K-12. Here, we show for the first time that restoration of OAg in E. coli K-12 strain MG1655 synergistically sensitises bacteria to vancomycin with bile salts (VBS). Suppressor mutants surviving lethal doses of VBS primarily contained disruptions in OAg biogenesis. We present data supporting a model where the transient presence and accumulation of lipid-linked OAg intermediates in the periplasmic leaflet of the inner membrane interfere with peptidoglycan sacculus biosynthesis, causing growth defects that are synergistically enhanced by bile salts. Lastly, we demonstrate that continuous bile salt exposure of OAg-producing MG1655 in the laboratory, can recreate a scenario where OAg disruption is selected for as an evolutionary fitness benefit. Our work thus provides a plausible explanation for the long-held mystery of the selective pressure that may have led to the loss of OAg biogenesis in E. coli K-12; this opens new avenues for exploring long-standing questions on the intricate network coordinating the synthesis of different cell envelope components in Gram-negative bacteria.Jilong Qin, Yaoqin Hong, Renato Morona, Makrina Totsik

Cysteine-Dependent Conformational Heterogeneity of Shigella flexneri Autotransporter IcsA and Implications of Its Function

Published November/December 2022Shigella IcsA is a versatile surface virulence factor required for early and late pathogenesis stages extracellularly and intracellularly. Despite IcsA serving as a model Type V secretion system (T5SS) autotransporter to study host-pathogen interactions, its detailed molecular architecture is poorly understood. Recently, IcsA was found to switch to a different conformation for its adhesin activity upon sensing the host stimuli by Shigella Type III secretion system (T3SS). Here, we reported that the single cysteine residue (C130) near the N terminus of the IcsA passenger had a role in IcsA adhesin activity. We also showed that the IcsA passenger (IcsAp) existed in multiple conformations, and the conformation populations were influenced by a central pair of cysteine residues (C375 and C379), which was not previously reported for any Type V autotransporter passengers. Disruption of either or both central cysteine residues altered the exposure of IcsA epitopes to polyclonal anti-IcsA antibodies previously shown to block Shigella adherence, yet without loss of IcsA intracellular functions in actin-based motility (ABM). Anti-IcsA antibody reactivity was restored when the IcsA-paired cysteine substitution mutants were expressed in an DipaD background with a constitutively active T3SS, highlighting an interplay between T3SS and T5SS. The work here uncovered a novel molecular switch empowered by a centrally localized, short-spaced cysteine pair in the Type V autotransporter IcsA that ensured conformational heterogeneity to aid IcsA evasion of host immunity.Jilong Qin, Yaoqin Hong, Renato Morona, Makrina Totsik

Comprehensive analysis of type 1 fimbriae regulation in fimB -null strains from the multidrug resistant Escherichia coli ST131 clone

Summary\ud \ud Uropathogenic Escherichia coli (UPEC) of sequence type 131 (ST131) are a pandemic multidrug resistant clone associated with urinary tract and bloodstream infections. Type 1 fimbriae, a major UPEC virulence factor, are essential for ST131 bladder colonization. The globally dominant sub-lineage of ST131 strains, clade C/H30-R, possess an ISEc55 insertion in the fimB gene that controls phase-variable type 1 fimbriae expression via the invertible fimS promoter. We report that inactivation of fimB in these strains causes altered regulation of type 1 fimbriae expression. Using a novel read-mapping approach based on Illumina sequencing, we demonstrate that ‘off’ to ‘on’ fimS inversion is reduced in these strains and controlled by recombinases encoded by the fimE and fimX genes. Unlike typical UPEC strains, the nucleoid-associated H-NS protein does not strongly repress fimE transcription in clade C ST131 strains. Using a genetic screen to identify novel regulators of fimE and fimX in the clade C ST131 strain EC958, we defined a new role for the guaB gene in the regulation of type 1 fimbriae and in colonisation of the mouse bladder. Our results provide a comprehensive analysis of type 1 fimbriae regulation in ST131, and highlight important differences in its control compared to non-ST131 UPEC

The co-transcriptome of uropathogenic Escherichia coli-infected mouse macrophages reveals new insights into host-pathogen interactions

© 2014 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd. Urinary tract infections (UTI) are among the most common infections in humans. Uropathogenic Escherichia coli (UPEC) can invade and replicate within bladder epithelial cells, and some UPEC strains can also survive within macrophages. To understand the UPEC transcriptional programme associated with intramacrophage survival, we performed host-pathogen co-transcriptome analyses using RNA sequencing. Mouse bone marrow-derived macrophages (BMMs) were challenged over a 24h time course with two UPEC reference strains that possess contrasting intramacrophage phenotypes: UTI89, which survives in BMMs, and 83972, which is killed by BMMs. Neither of these strains caused significant BMM cell death at the low multiplicity of infection that was used in this study. We developed an effective computational framework that simultaneously separated, annotated and quantified the mammalian and bacterial transcriptomes. Bone marrow-derived macrophages responded to the two UPEC strains with a broadly similar gene expression programme. In contrast, the transcriptional responses of the UPEC strains diverged markedly from each other. We identified UTI89 genes up-regulated at 24h post-infection, and hypothesized that some may contribute to intramacrophage survival. Indeed, we showed that deletion of one such gene (pspA) significantly reduced UTI89 survival within BMMs. Our study provides a technological framework for simultaneously capturing global changes at the transcriptional level in co-cultures, and has generated new insights into the mechanisms that UPEC use to persist within the intramacrophage environment