Biology subjects

Khalid, H.

Publications and source records attributed to Khalid, H..

6 recordsLinked to original sources

bioRxiv2026

Molecular analysis of Lancefield group C/G streptococci causing human infections in Sheffield, UK.

Lancefield group C/G streptococci (GCS/GGS) are increasingly recognised as significant human pathogens that cause a disease spectrum similar to Streptococcus pyogenes. Despite their high clinical burden in the UK, their genomic diversity remains poorly understood. We performed whole-genome sequencing (WGS) on a prospective collection of 109 consecutive GCS/GGS isolates from all infection types in Sheffield, UK, over five months in 2020. Streptococcus dysgalactiae subsp. equisimilis (SDSE) accounted for 104 isolates, while five were identified as Streptococcus canis. The SDSE population was highly diverse, comprising 15 genomic clusters and 38 unique emm-ST combinations. We identified the presence of the ST20/stG62647 international lineage (24% of isolates), a cluster globally associated with severe invasive disease. Antimicrobial resistance genes were prevalent (49%), predominantly linked to mobile genetic elements carrying tetracycline and macrolide resistance. Furthermore, a variation in the penicillin-binding protein PBP2X (P601L) was linked to reduced penicillin sensitivity (MIC 0.03 mg/L). There were few or no genetic changes in isolates obtained from the same patient, even when they were collected 8-10 weeks apart, indicating long-term persistence within a host. The unexpected detection of S. canis in human infections and the high diversity of SDSE, persistence and virulence-associated regions underscore the need for enhanced national genomic surveillance to track emerging virulent and antibiotic-resistant SDSE lineages. Impact statementLancefield group C and G streptococci, most often the species Streptococcus dysgalactiae subsp. equisimilis (SDSE), are an increasingly significant human pathogen, often mirroring the severity of infections caused by Lancefield group A Streptococcus (S. pyogenes). Despite its clinical importance, we know little about the population of SDSE circulating in the UK. This study provides the first comprehensive genomic analysis of SDSE isolates from a single UK region, identifying a highly diverse population comprising 15 distinct genomic clusters but with evidence of long-term persistence within a single host. Notably, we confirm the presence of the international stG62647/ST20 lineage in the UK, which is globally associated with severe invasive disease. Our findings also reveal a high prevalence of antimicrobial resistance genes ([~]49%), primarily linked to mobile genetic elements, and the presence of a specific variation in the penicillin-binding protein PBP2X that reduces penicillin sensitivity. Additionally, the unexpected detection of S. canis in human infections rather than animals highlights a need for monitoring. By defining the UKs SDSE population structure and its resistance landscape, this research underscores the critical need for enhanced national genomic surveillance to track emerging high-virulence and antibiotic-resistant lineages Data summarySequence files for isolates from Sheffield used for this study have been uploaded to the sequence read archive with project accession number PRJNA1333937 and accession numbers provided in Supplementary Dataset 1. The completed genome for SDE096 been deposited on GenBank with the accession number JBSXMJ000000000.

microbiology↗

bioRxiv2024

Highly conserved brain vascular receptor ALPL mediates transport of engineered viral vectors across the blood-brain barrier

Delivery of systemically administered therapeutics to the central nervous system (CNS) is restricted by the blood-brain barrier (BBB). Bioengineered Adeno-Associated Virus (AAV) capsids have been shown to penetrate the BBB with great efficacy in mouse and non-human primate models, but their translational potential is often limited by species selectivity and undefined mechanisms of action. Here, we apply our RNA-guided TRACER AAV capsid evolution platform to generate VCAP-102, an AAV9 variant with markedly increased brain tropism following intravenous delivery in both rodents and primates. VCAP-102 demonstrates a similar CNS tropism in cynomolgus macaque, african green monkey, marmoset and mouse, showing 20- to 400-fold increased transgene expression across multiple brain regions relative to AAV9. We demonstrate that the enhanced CNS tropism of VCAP-102 results from direct interaction with alkaline phosphatase (ALPL), a highly conserved membrane-associated protein expressed on the brain vasculature. VCAP-102 interacts with human, primate and murine ALPL isoforms, and ectopic expression of ALPL is sufficient to initiate receptor-mediated transcytosis of VCAP-102 in an in vitro transwell model. Our work identifies VCAP-102 as a cross-species CNS gene delivery vector with a strong potential for clinical translation and establishes ALPL as a brain delivery shuttle capable of efficient BBB transport to maximize CNS delivery of biotherapeutics.

neuroscience↗

bioRxiv2023

CD9 co-operation with syndecan-1 is required for a major staphylococcal adhesion pathway

ObjectivesEpithelial colonisation is a critical first step in bacterial pathogenesis. Staphylococcus aureus can utilise several host factors to associate with cells, including 5{beta}l integrin and heparan sulphate proteoglycans, such as the syndecans. Here, we demonstrate that a partner protein of both integrins and syndecans, the host membrane adapter protein tetraspanin CD9, is essential for syndecan-mediated staphylococcal adhesion. Fibronectin is also essential in this process while integrins are only critical for post-adhesion entry into human epithelial cells. Methods and ResultsTreatment of epithelial cells with CD9-derived peptide or heparin caused significant reductions in staphylococcal adherence, dependent on both CD9 and syndecan-1. Exogenous fibronectin caused a CD9-dependent increase in staphylococcal adhesion whereas blockade of {beta}1 integrins did not affect adhesion but did reduce the subsequent internalisation of adhered bacteria. CD9 disruption or deletion increased {beta}1 integrin-mediated internalisation, suggesting that CD9 coordinates sequential staphylococcal adhesion and internalisation. ConclusionsCD9 controls staphylococcal adhesion through syndecan-1, using a mechanism that likely requires CD9-mediated syndecan organisation to correctly display fibronectin at the host cell surface. We propose that CD9-derived peptides or heparin analogues could be developed as anti-adhesion treatments to inhibit the initial stages of staphylococcal pathogenesis.

microbiology↗

bioRxiv2022

Development of allosteric, selective cyclin-dependent kinase 2 (CDK2) inhibitors that are negatively cooperative with cyclin binding and show potential as contraceptive agents

Compared to most ATP-site kinase inhibitors, small molecules that target an allosteric pocket have the potential for improved selectivity due to the often observed lower structural similarity at these distal sites. Despite their promise, relatively few examples of structurally confirmed, high-affinity allosteric kinase inhibitors exist. Cyclin-dependent kinase 2 (CDK2) is a target for many therapeutic indications, including non-hormonal contraception.1 However, an inhibitor against this kinase with exquisite selectivity has not reached the market because of the structural similarity between CDKs.1-2 In this paper, we describe the development and mechanism of action of new type III inhibitors that bind CDK2 with nanomolar affinity, making them the highest affinity, structurally confirmed allosteric CDK inhibitors reported. Notably, these anthranilic acid inhibitors exhibit a strong negative cooperative relationship with cyclin binding, which remains an underexplored mechanism for CDK2 inhibition. Furthermore, the binding profile of these compounds in both biophysical and cellular assays demonstrate the promise of this series for further development into a therapeutic selective for CDK2 over highly similar kinases like CDK1. The potential of these inhibitors as efficacious contraceptive agents is seen by incubation with mouse testicular explants, where they recapitulate Cdk2-/- and Spdya-/- phenotypes.

pharmacology and toxicology↗

bioRxiv2021

Comparative genomic analysis of skin and soft tissue Streptococcus pyogenes isolates from low- and high-income settings

Streptococcus pyogenes is a leading cause of human morbidity and mortality, especially in resource limited settings. The World Health Organisation has recently made a vaccine for S. pyogenes a global health priority to reduce the burden of the post-infection rheumatic heart disease. For a vaccine to be active against all relevant strains in each region, molecular characterisation of circulating S. pyogenes isolates is needed. We performed extensive comparative whole genome analyses of S. pyogenes isolates from skin and soft tissue infections in The Gambia, West Africa, where there is a high burden of such infections. To act as a comparator to this low-income country (LIC) collection of isolates, we performed genome sequencing of isolates from skin infections in Sheffield, UK, as representative high-income country (HIC) isolates. LIC isolates from The Gambia were genetically more diverse (46 emm-types in 107 isolates) compared to HIC isolates from Sheffield (23 emm-types in 142 isolates), with only 7 overlapping emm-types and with diverse genetic backgrounds. Characterisation of other molecular markers indicated some shared features, including a high prevalence of the skin infection-associated emm-pattern D and the variable fibronectin-collagen-T antigen (FCT) types FCT-3 and FCT-4. A previously unidentified FCT (FCT-10) was identified in the LIC isolates, belonging to two different emm-types. A high proportion (79/107; 73.8%) of LIC isolates carried genes for tetracycline resistance, compared to 53/142 (37.3%) HIC isolates. There was also evidence of different circulating prophages, as very few prophage-associated DNases and lower numbers of superantigens were detected in LIC isolates. Our study provides much needed insight into the genetics of circulating isolates in a LIC (The Gambia), and how they differ from those circulating in HICs (Sheffield, UK). Common molecular features may act as bacterial drivers for specific infection types, regardless of the diverse genetic background.

microbiology↗

bioRxiv2019

Pan-viral protection against arboviruses by targeting inoculation site-based skin macrophages

Arthropod-borne viruses (arboviruses) are important human pathogens for which there are no specific antiviral medicines. The large number of genetically-distinct arbovirus species, coupled with the unpredictable nature of their outbreaks, has made developing virus-specific anti-viral medicines challenging. Instead, we have defined and targeted a key aspect of the host innate immune response to virus at the arthropod bite that is common to all arbovirus infections, potentially circumventing the need for virus-specific therapies at this site. Using mouse models and human skin explants, we identify innate immune responses by dermal macrophages in the skin as a key determinant of disease severity. Post-exposure treatment of the inoculation site by a topical innate immune agonist significantly suppressed both the local and subsequent systemic course of infection and improved clinical outcome in mice to infection with a variety of arboviruses from the Alphavirus, Flavivirus and Orthobunyavirus genuses. In the absence of treatment, anti-viral interferon expression to virus in the skin was restricted to dermal dendritic cells. In contrast, targeting the more populous skin-resident macrophages with an immune agonist elicited protective responses in key cellular targets of virus that otherwise replicated virus to high levels. By defining and targeting a key aspect of the innate immune response to virus at the mosquito bite site, we have shown that it is possible to improve outcome to infection by targeting pathways activated at the site of inoculation, and thereby identified a putative new strategy for limiting disease following infection with a variety of genetically-distinct arboviruses.\n\nOne-sentence summaryWe demonstrate that activation of innate immune responses to arbovirus at the mosquito bite is a limiting factor for preventing efficient systemic dissemination of virus and that therapeutic targeting of skin-resident macrophages can have defining inhibitory effects on the later systemic course.

immunology↗