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Extra-intestinal pathogenic Escherichia coli (ExPEC) belong to a critical priority group of antibiotic resistant pathogens. ExPEC establish gut reservoirs that seed infection of the urinary tract and bloodstream, but the mechanisms of gut colonisation remain to be properly understood. Ucl fimbriae are attachment organelles that facilitate ExPEC adherence. Here, we investigated cellular receptors for Ucl fimbriae and Ucl expression to define molecular mechanisms of Ucl-mediated ExPEC colonisation of the gut. We demonstrate differential expression of Ucl fimbriae in ExPEC sequence types associated with disseminated infection. Genome editing of strains from two common sequence types, F11 (ST127) and UTI89 (ST95), identified a single nucleotide polymorphism in the ucl promoter that changes fimbriae expression via activation by the global stress-response regulator OxyR, leading to altered gut colonisation. Structure-function analysis of the Ucl fimbriae tip-adhesin (UclD) identified high-affinity glycan receptor targets, with highest affinity for sialyllacto-N-fucopentose VI, a structure likely to be expressed on the gut epithelium. Comparison of the UclD adhesin to the homologous UcaD tip-adhesin from Proteus mirabilis revealed that although they possess a similar tertiary structure, apart from lacto-N-fucopentose VI that bound to both adhesins at low-micromolar affinity, they recognize different fucose- and glucose-containing oligosaccharides. Competitive surface plasmon resonance analysis together with co-structural investigation of UcaD in complex with monosaccharides revealed a broad-specificity glycan binding pocket shared between UcaD and UclD that could accommodate these interactions. Overall, our study describes a mechanism of adaptation that augments establishment of an ExPEC gut reservoir to seed disseminated infections, providing a pathway for the development of targeted anti-adhesion therapeutics. Author summary: ExPEC infection of the urinary tract and bloodstream is frequently seeded from an intestinal reservoir, necessitating an understanding of the mechanisms that promote gut colonisation. Here we employed molecular and structural approaches to define the regulation and function of ExPEC Ucl fimbriae as a gut colonisation factor. We describe how mutations in the non-coding regulatory region of the ucl promoter cause increased Ucl fimbriae expression and promote enhanced gut colonisation via tuned induction by a global regulator that senses oxygen stress. We further define the glycan receptor targets of Ucl fimbriae and characterise the structural features of the Ucl adhesin that facilitate these interactions. These findings explain how ExPEC can adapt to survival in the gut to seed extra-intestinal infection. [ABSTRACT FROM AUTHOR]

Persistent and intermittent fecal shedding, hallmarks of Salmonella infections, are important for fecal-oral transmission. In the intestine, Salmonella enterica serovar Typhimurium (STm) actively invades intestinal epithelial cells (IECs) and survives in the Salmonella -containing vacuole (SCV) and the cell cytosol. Cytosolic STm replicate rapidly, express invasion factors, and induce extrusion of infected epithelial cells into the intestinal lumen. Here, we engineered STm that self-destruct in the cytosol (STmCytoKill), but replicates normally in the SCV, to examine the role of cytosolic STm in infection. Intestinal expansion and fecal shedding of STmCytoKill are impaired in mouse models of infection. We propose a model whereby repeated rounds of invasion, cytosolic replication, and release of invasive STm from extruded IECs fuels the high luminal density required for fecal shedding. [Display omitted] • A lethal biosensor restricts S. Typhimurium intracytosolic growth in epithelial cells • Intracytosolic replication facilitates the expansion of S. Typhimurium loads in the gut • Sustained fecal shedding of S. Typhimurium is fueled by intracytosolic replication Fecal shedding is crucial for transmission of Salmonella among hosts. Using genetic engineering to selectively deplete cytosolic bacteria from enterocytes, Chong et al. show that intracytosolic replication feeds the expansion of intestinal S. Typhimurium needed for efficient shedding. [ABSTRACT FROM AUTHOR]

To determine the value of pre-decannulation capped overnight ICU monitoring for assessing decannulation-readiness in pediatric patients. This study included all pediatric patients, age 18 and under, with a tracheostomy attempting decannulation at the University of Michigan between 2013 and 2018. Patients who underwent major airway reconstruction immediately prior to decannulation were excluded. Descriptive and comparative statistics were calculated to compare the sub-group of patients who underwent pre-decannulation capped overnight ICU monitoring to those who did not. 125 pediatric patients attempted decannulation for a total of 126 attempts with 105 attempts being eligible for inclusion. 75 eligible attempts included pre-decannulation capped overnight ICU monitoring, while 30 did not. Subsequent rates of successful decannulation were 97.33% (73/75) and 100.00% (30/30), respectively (P = 0.366; 95% CI -8.818-9.260). The pre-decannulation capped overnight ICU monitoring passing rate was 98.67% (74/75) despite a complication rate of 5.33% (4/75). Post-decannulation, 98.08% (102/104) of decannulated patients were monitored inpatient for a minimum of 24 h With similar rates of successful decannulation among both sub-groups and previous research demonstrating sufficient ambulatory testing accurately predicts successful decannulation, pre-decannulation capped overnight ICU monitoring is a low-value, high-cost test that can be safely discontinued without compromising patient care. Notably, our study excluded patients undergoing open airway reconstruction immediately prior to decannulation. The 24-h monitoring post-decannulation serves as a safety net for individuals who ultimately fail decannulation. [ABSTRACT FROM AUTHOR]

Post-transcriptional modifications can impact the stability and functionality of many different classes of RNA molecules and are an especially important aspect of tRNA regulation. It is hypothesized that cells can orchestrate rapid responses to changing environmental conditions by adjusting the specific types and levels of tRNA modifications. We uncovered strong evidence in support of this tRNA global regulation hypothesis by examining effects of the well-conserved tRNA modifying enzyme MiaA in extraintestinal pathogenic Escherichia coli (ExPEC), a major cause of urinary tract and bloodstream infections. MiaA mediates the prenylation of adenosine-37 within tRNAs that decode UNN codons, and we found it to be crucial to the fitness and virulence of ExPEC. MiaA levels shifted in response to stress via a post-transcriptional mechanism, resulting in marked changes in the amounts of fully modified MiaA substrates. Both ablation and forced overproduction of MiaA stimulated translational frameshifting and profoundly altered the ExPEC proteome, with variable effects attributable to UNN content, changes in the catalytic activity of MiaA, or availability of metabolic precursors. Cumulatively, these data indicate that balanced input from MiaA is critical for optimizing cellular responses, with MiaA acting much like a rheostat that can be used to realign global protein expression patterns., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Commensal bacteria can interfere with colonization of the host by infiltrating pathogens. In this issue of Cell Host & Microbe, Kim et al. (2019) describe an intriguing mechanism of colonization resistance driven by the mismatching of methylation patterns following uptake of commensal-derived DNA by pathogenic strains of Neisseria., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Extraintestinal pathogenic Escherichia coli (ExPEC) acts as a commensal within the mammalian gut but can induce pathology upon dissemination to other host environments such as the urinary tract and bloodstream. ExPEC genomes are likely shaped by evolutionary forces encountered within the gut, where the bacteria spend much of their time, provoking the question of how their extraintestinal virulence traits arose. The principle of coincidental evolution, in which a gene that evolved in one niche happens to be advantageous in another, has been used to argue that ExPEC virulence factors originated in response to selective pressures within the gut ecosystem. As a test of this hypothesis, the fitness of ExPEC mutants lacking canonical virulence factors was assessed within the intact murine gut in the absence of antibiotic treatment. We found that most of the tested factors, including cytotoxic necrotizing factor type 1 (CNF1), Usp, colibactin, flagella, and plasmid pUTI89, were dispensable for gut colonization. The deletion of genes encoding the adhesin PapG or the toxin HlyA had transient effects but did not interfere with longer-term persistence. In contrast, a mutant missing the type 1 pilus-associated adhesin FimH displayed somewhat reduced persistence within the gut. However, this phenotype varied dependent on the presence of specific competing strains and was partially attributable to aberrant flagellin expression in the absence of fimH These data indicate that FimH and other key ExPEC-associated factors are not strictly required for gut colonization, suggesting that the development of extraintestinal virulence traits is not driven solely by selective pressures within the gut., (Copyright © 2018 Russell et al.)

Hypoxia is an environmental stress encountered by Aspergillus fumigatus during invasive pulmonary aspergillosis (IPA). The ability of this mold to adapt to hypoxia is important for fungal virulence and genetically regulated in part by the sterol regulatory element binding protein (SREBP) SrbA. SrbA is required for fungal growth in the murine lung and to ultimately cause lethal disease in murine models of IPA. Here we identified and partially characterized four genes (dscA, dscB, dscC, and dscD, here referred to as dscA-D) with previously unknown functions in A. fumigatus that are orthologs of the Schizosaccharomyces pombe genes dsc1, dsc2, dsc3, and dsc4 (dsc1-4), which encode a Golgi E3 ligase complex critical for SREBP activation by proteolytic cleavage. A. fumigatus null dscA-D mutants displayed remarkable defects in hypoxic growth and increased susceptibility to triazole antifungal drugs. Consistent with the confirmed role of these genes in S. pombe, both ΔdscA and ΔdscC resulted in reduced cleavage of the SrbA precursor protein in A. fumigatus. Inoculation of corticosteroid immunosuppressed mice with ΔdscA and ΔdscC strains revealed that these genes are critical for A. fumigatus virulence. Reintroduction of SrbA amino acids 1 to 425, encompassing the N terminus DNA binding domain, into the ΔdscA strain was able to partially restore virulence, further supporting a mechanistic link between DscA and SrbA function. Thus, we have shown for the first time the importance of a previously uncharacterized group of genes in A. fumigatus that mediate hypoxia adaptation, fungal virulence, and triazole drug susceptibility and that are likely linked to regulation of SrbA function.