Biology subjects

Ulhuq, F. R.

Publications and source records attributed to Ulhuq, F. R..

4 recordsLinked to original sources

bioRxiv2025

The Staphylococcus aureus LXG-domain toxins EsxX and SAR0287 do not promote virulence in a zebrafish larval infection model

The Staphylococcus aureus type VIIb secretion system (T7SSb) is a multiprotein secretion system that secretes toxins with antibacterial activity, but which is also required for full virulence in animal models of infection. S. aureus strains carry one of four T7SSb locus types, named essC1 to essC4, each of which encodes a characteristic LXG-family substrate at the T7SS locus. In essC2 strains this LXG-domain protein is EsxX, which has a glycine zipper sequence in its C-terminus and has potent antibacterial membrane-depolarising activity. In this work we recognise conserved features of the essC2 and essC3 systems, identifying the LXG protein SAR0287 as structurally and functionally similar to EsxX. Using a zebrafish larval hindbrain ventricle infection model we demonstrate that the T7SSb of essC2 and essC3 representative strains contribute to bacterial replication and zebrafish mortality. However, there is no significant loss of virulence in the model system if EsxX or SAR0287 are absent. These findings indicate there is no discernible role for either toxin in this virulence model.

microbiology↗

bioRxiv2023

An interbacterial lipase toxin with an unprecedented reverse domain arrangement defines a new class of type VII secretion system effector

The type VII protein secretion system (T7SS) is found in many Gram-positive bacteria and in pathogenic mycobacteria. All T7SS substrate proteins described to date share a common helical domain architecture at the N-terminus that typically interacts with other helical partner proteins, forming a composite signal sequence for targeting to the T7SS. The C-terminal domains are functionally diverse and in Gram-positive bacteria such as Staphylococcus aureus often specify toxic anti-bacterial activity. Here we describe the first example of a new class of T7 substrate, TslA, that has an unexpected reverse domain organisation. TslA is widely found across Bacillota including Staphylococcus, Enterococcus and Listeria. We show that the S. aureus TslA N-terminal domain is a phospholipase A with anti-staphylococcal activity that is neutralised by the immunity lipoprotein TilA. Two small helical partner proteins, TlaA1 and TlaA2 are essential for T7-dependent secretion of TslA and at least one of these interacts with the TslA C-terminal domain to form a helical stack. Cryo-EM analysis of purified TslA complexes indicate that they share structural similarity with canonical T7 substrates. Our findings suggest that the T7SS has the extraordinary feature of recognising a secretion signal present at either end of a substrate.

microbiology↗

bioRxiv2022

Characterisation of key genotypic and phenotypic traits of clinical cystic fibrosis Staphylococcus aureus isolates

IntroductionOne third of people with CF in the UK are co-infected by both Staphylococcus aureus and Pseudomonas aeruginosa. Chronic bacterial infection in CF contributes to the gradual destruction of lung tissue, and eventually respiratory failure in this group. Gap StatementThe contribution of S. aureus to cystic fibrosis (CF) lung decline in the presence or absence of P. aeruginosa is unclear. Defining the molecular and phenotypic characteristics of a range of S. aureus clinical isolates will help further understand its pathogenic capabilities. AimOur objective was to use molecular and phenotypic tools to characterise twenty-five clinical S. aureus isolates collected from mono- and coinfection with P. aeruginosa from people with CF at the Royal Victoria Infirmary, Newcastle upon Tyne. MethodologyGenomic DNA was extracted and sequenced. Multilocus sequence typing was used to construct phylogeny from the seven housekeeping genes. A pangenome was calculated using Roary. and cluster of Orthologous groups were assigned using eggNOG-mapper which were used to determine differences within core, accessory, and unique genomes. Characterisation of sequence type, clonal complex, agr and spa types was carried out using PubMLST, eBURST, AgrVATE and spaTyper, respectively. Antibiotic resistance was determined using Kirby Bauer disk diffusion tests. Phenotypic testing of haemolysis was carried out using ovine red blood cell agar plates and mucoid phenotypes visualised using Congo red agar. ResultsClinical strains clustered closely based on agr type, sequence type and clonal complex. COG analysis revealed statistically significant enrichment of COG families between core, accessory and unique pangenome groups. The unique genome was significantly enriched for replication, recombination and repair, and defence mechanisms. The presence of known virulence genes and toxins were high within this group, and unique genes were identified in 11 strains. Strains which were isolated from the same patient all surpassed average nucleotide identity thresholds, however, differed in phenotypic traits. Antimicrobial resistance to macrolides was significantly higher in the coinfection group. ConclusionThere is huge variation in genetic and phenotypic capabilities of S. aureus strains. Further studies on how these may differ in relation to other species in the CF lung may give insight into inter-species interactions. Data summaryThe assembled GenBank (gbk) files for all clinical isolates in this study have been deposited in ENA under the study accession PRJEB56184, accession numbers for each of the twenty-five clinical isolates have been provided in Table S1. The reference strains were collected from the NCBI BioSample database (www.ncbi.nlm.nih.gov/biosample): MRSA_252 (NC_002952.2), HO 5096 0412 (NC_017763.1), ST398 (NC_017333.1) and NCTC8325 (NC_007795.1).

microbiology↗

bioRxiv2018

A membrane-depolarising toxin substrate of the Staphylococcus aureus Type VII protein secretion system targets eukaryotes and bacteria

The type VII protein secretion system (T7SS) is conserved across Staphylococcus aureus strains and plays important roles in virulence and interbacterial competition. To date only one T7SS substrate protein, encoded in a subset of S. aureus genomes, has been functionally characterized. Here, using an unbiased proteomic approach, we identify TspA as a further T7SS substrate. TspA is encoded distantly from the T7SS gene cluster and is found across all S. aureus strains as well as in Listeria and Enterococci. Heterologous expression of TspA from S. aureus strain RN6390 indicates its C-terminal domain is toxic when targeted to the Escherichia coli periplasm and that it depolarizes the cytoplasmic membrane. The membrane depolarizing activity is alleviated by co-production of the membrane-bound TsaI immunity protein, which is encoded adjacent to tspA on the S. aureus chromosome. Using a zebrafish hindbrain ventricle infection model, we demonstrate that the T7SS of strain RN6390 promotes bacterial replication in vivo, and deletion of tspA leads to increased bacterial clearance. The toxin domain of TspA is highly polymorphic and S. aureus strains encode multiple tsaI homologues at the tspA locus, suggestive of additional roles in intra-species competition. In agreement, we demonstrate TspA-dependent growth inhibition of RN6390 by strain COL in the zebrafish infection model that is alleviated by the presence of TsaI homologues. Significance statementStaphylococcus aureus, a human commensal organism that asymptomatically colonizes the nares, is capable of causing serious disease following breach of the mucosal barrier. S. aureus strains encode a Type VII secretion system (T7SS) that is required for virulence in mouse infection models, and some strains also secrete a nuclease toxin by this route that has antibacterial activity. Here we identify TspA, widely found in Staphylococci and other pathogenic bacteria, as a T7 substrate. We show that TspA has membrane-depolarizing activity and that S. aureus uses TspA to inhibit the growth of a bacterial competitor in vivo.

microbiology↗