Biology subjects

de Silva, T.

Publications and source records attributed to de Silva, T..

7 recordsLinked to original sources

bioRxiv2024

Molecular characterisation of Streptococcus pyogenes (StrepA) non-invasive isolates during the 2022-23 UK upsurge

At the end of 2022 into early 2023 the UK Health Security Agency reported unusually high levels of scarlet fever and invasive disease caused by Streptococcus pyogenes (StrepA or group A Streptococcus). During this time, we collected and genome sequenced 341 non-invasive throat and skin S. pyogenes isolates identified during routine clinical diagnostic testing in Sheffield, a large UK city. We compared the data with that obtained from a similar collection of 165 isolates from 2016-17. Numbers of throat-associated isolates collected peaked in early December 2022, reflecting the national scarlet fever upsurge, while skin infections peaked later in December. The most common emm-types in 2022-23 were emm1 (28.7%), emm12 (24.9%), and emm22 (7.7%) in throat; and emm1 (22%), emm12 (10%), emm76 (18%), and emm49 (7%) in skin. Whilst all emm1 isolates were the M1UK lineage, comparison with 2016-17 revealed diverse lineages in other emm-types, including emm12, and emergent lineages within other types including a new acapsular emm75 lineage, demonstrating that the upsurge was not completely driven by a single genotype. Analysis of the capsule locus predicted only 51% of throat isolates would produce capsule compared to 78% of skin isolates. 90% of throat isolates were also predicted to have high NADase and Streptolysin O (SLO) expression, based on the promoter sequence, compared to only 56% of skin isolates. Our study has highlighted the value in analysis of non-invasive isolates to characterise tissue tropisms, as well as changing strain diversity and emerging genomic features which may have implications for spillover into invasive disease and future S. pyogenes upsurges. Data summaryAll new genome sequence data is available on the NCBI short read archive under the bioproject PRJNA1062601 and individual accession numbers are listed in Supplementary Table 1 and Table 2. Impact statementThe human bacterial pathogen Streptococcus pyogenes, also known as group A Streptococcus or StrepA, caused a dramatic and sudden upsurge in scarlet fever in the UK at the end of 2022 into early 2023. We present molecular characterisation of this upsurge, through genome sequence analysis of throat, skin and other types of non-severe infection isolates collected by the microbiology diagnostic lab at the Northern General Hospital in Sheffield, England. We found that, whilst two strain types were the predominant cause of infections during the upsurge, other types had emerged or changed when compared to a similar collection from 2016-17. We also identified differences between throat-associated isolates and skin-associated isolates and highlighted important bacterial factors that might influence infection types. Isolates from non-severe throat/skin types of infections are rarely saved and therefore our knowledge of them is limited. However, here we demonstrate that study of such isolates may be key to understanding upsurges of more severe infections.

microbiology↗

bioRxiv2020

Broad and strong memory CD4+ and CD8+ T cells induced by SARS-CoV-2 in UK convalescent COVID-19 patients.

COVID-19 is an ongoing global crisis in which the development of effective vaccines and therapeutics will depend critically on understanding the natural immunity to the virus, including the role of SARS-CoV-2-specific T cells. We have conducted a study of 42 patients following recovery from COVID-19, including 28 mild and 14 severe cases, comparing their T cell responses to those of 16 control donors. We assessed the immune memory of T cell responses using IFN{gamma} based assays with overlapping peptides spanning SARS-CoV-2 apart from ORF1. We found the breadth, magnitude and frequency of memory T cell responses from COVID-19 were significantly higher in severe compared to mild COVID-19 cases, and this effect was most marked in response to spike, membrane, and ORF3a proteins. Total and spike-specific T cell responses correlated with the anti-Spike, anti-Receptor Binding Domain (RBD) as well as anti-Nucleoprotein (NP) endpoint antibody titre (p<0.001, <0.001 and =0.002). We identified 39 separate peptides containing CD4+ and/or CD8+ epitopes, which strikingly included six immunodominant epitope clusters targeted by T cells in many donors, including 3 clusters in spike (recognised by 29%, 24%, 18% donors), two in the membrane protein (M, 32%, 47%) and one in the nucleoprotein (Np, 35%). CD8+ responses were further defined for their HLA restriction, including B*4001-restricted T cells showing central memory and effector memory phenotype. In mild cases, higher frequencies of multi-cytokine producing M- and NP-specific CD8+ T cells than spike-specific CD8+ T cells were observed. They furthermore showed a higher ratio of SARS-CoV-2-specific CD8+ to CD4+ T cell responses. Immunodominant epitope clusters and peptides containing T cell epitopes identified in this study will provide critical tools to study the role of virus-specific T cells in control and resolution of SARS-CoV-2 infections. The identification of T cell specificity and functionality associated with milder disease, highlights the potential importance of including non-spike proteins within future COVID-19 vaccine design.

immunology↗

bioRxiv2020

Spike mutation pipeline reveals the emergence of a more transmissible form of SARS-CoV-2

We have developed an analysis pipeline to facilitate real-time mutation tracking in SARS-CoV-2, focusing initially on the Spike (S) protein because it mediates infection of human cells and is the target of most vaccine strategies and antibody-based therapeutics. To date we have identified thirteen mutations in Spike that are accumulating. Mutations are considered in a broader phylogenetic context, geographically, and over time, to provide an early warning system to reveal mutations that may confer selective advantages in transmission or resistance to interventions. Each one is evaluated for evidence of positive selection, and the implications of the mutation are explored through structural modeling. The mutation Spike D614G is of urgent concern; it began spreading in Europe in early February, and when introduced to new regions it rapidly becomes the dominant form. Also, we present evidence of recombination between locally circulating strains, indicative of multiple strain infections. These finding have important implications for SARS-CoV-2 transmission, pathogenesis and immune interventions.

evolutionary biology↗

bioRxiv2019

The Consequences of Egg Adaptation in the H3N2 Component to the Immunogenicity of Live Attenuated Influenza Vaccine

Adaptation in egg-passaged vaccine strains may cause reduced vaccine effectiveness due to altered antigenicity of the influenza haemagglutinin. We tested whether egg adaptation modified serum and mucosal antibody responses to the A(H3N2) component in the Live Attenuated Influenza Vaccine (LAIV). Twice as many children seroconverted to an egg-adapted H3N2 than the equivalent wildtype strain. Seroconversion to the wildtype strain was greater in children seronegative pre-LAIV, whereas higher mucosal IgA responses to wildtype antigen were observed if seropositive prior to vaccination. Sequencing of virus from nasopharyngeal swabs from 7 days post-LAIV showed low sequence diversity and no reversion of egg-adaptive mutations.

immunology↗

bioRxiv2019

High burden and seasonal variation of paediatric scabies and pyoderma prevalence in The Gambia: a cross-sectional study

BackgroundScabies is a WHO neglected tropical disease common in children in low-and middle-income countries. Excoriation of scabies lesions can lead to secondary pyoderma infection, most commonly by Staphyloccocus aureus and Streptococcus pyogenes (group A streptococcus, GAS), with the latter linked to acute post-streptococcal glomerulonephritis (APSGN) and potentially rheumatic heart disease (RHD). There is a paucity of data on the prevalence of these skin infections and their bacterial aetiology from Africa.\n\nMaterials/methodsA cross-sectional study, conducted over a four-month period that included the dry and rainy season, was conducted to determine the prevalence of common skin infections in Sukuta, a peri-urban settlement in western Gambia, in children <5 years. Swabs from pyoderma lesions were cultured for S. aureus and GAS.\n\nResultsOf 1441 children examined, 15.9% had scabies (95% CI 12.2-20.4), 17.4% had pyoderma (95% CI 10.4-27.7) and 9.7% had fungal infections (95% CI 6.6-14.0). Scabies were significantly associated with pyoderma (aOR 2.74, 95% CI 1.61-4.67). Of 250 pyoderma swabs, 80.8% were culture-positive for S. aureus, and 50.8% for GAS. Participants examined after the first rains were significantly more likely to have pyoderma than those examined before (aRR 2.42, 95% CI 1.38-4.23), whereas no difference in scabies prevalence was seen (aRR 1.08, 95% CI 0.70-1.67). Swab positivity was not affected by the season.\n\nConclusionsHigh prevalence of scabies and pyoderma were observed. Pyoderma increased significantly during rainy season. Given the high prevalence of GAS pyoderma among children, further research on the association with RHD in West Africa is warranted.\n\nSummaryThis cross-sectional study of skin infections in The Gambia revealed prevalence of scabies, pyoderma and fungal infections in children <5 years of 15.9%, 17.4% and 9.7% respectively, with increased bacterial skin infections in the rainy season.

epidemiology↗

bioRxiv2019

Genomic and epidemiological evidence of a dominant Panton-Valentine leucocidin-positive Methicillin Resistant Staphylococcus aureus lineage in Sri Lanka with spread to the United Kingdom and Australia

ObjectiveTo undertake the first detailed genomic analysis of methicillin-resistant Staphylococcus aureus (MRSA) isolated in Sri Lanka.\n\nMethodsA prospective observational study was performed on 94 MRSA isolates collected over a four month period from the Anuradhapura Teaching Hospital, Sri Lanka. Screening for mecA, mecC and the Panton-Valentine leucocidin (PVL)-associated lukS-PV/lukF-PV genes and molecular characterisation by spa typing was undertaken. Whole genome sequencing (WGS) and phylogenetic analysis was performed on selected multilocus sequence type (MLST) clonal complex 5 (CC5) isolates from Sri Lanka, England, Australia and Argentina.\n\nResultsAll 94 MRSA harboured the mecA gene. Nineteen spa types associated with nine MLST clonal complexes were identified. Most isolates were from skin and soft tissue infections (76.9%), with the remainder causing more invasive disease. Sixty two (65.9%) of isolates were PVL positive with the majority (56 isolates; 90.3%) belonging to a dominant CC5 lineage. This lineage, PVL-positive ST5-MRSA-IVc, was associated with community and hospital-onset infections. Based on WGS, representative PVL-positive ST5-MRSA-IVc isolates from Sri Lanka, England and Australia formed a single phylogenetic clade, suggesting wide geographical circulation.\n\nConclusionsWe present the most detailed genomic analysis of MRSA isolated in Sri Lanka to date. The analysis identified a PVL-positive ST5-MRSA-IVc that dominates MRSA clinical infections in Sri Lanka. Furthermore, transmission of the strain has occurred in the United Kingdom and Australia.

microbiology↗

bioRxiv2018

LOW-BIAS RNA SEQUENCING OF THE HIV-2 GENOME FROM BLOOD PLASMA

Accurate determination of the genetic diversity present in the HIV quasi-species is critical for the development of a preventative vaccine: in particular, little is known about viral genetic diversity for the second strain of HIV, HIV-2. A better understanding of HIV-2 biology is relevant to the HIV vaccine field because a substantial proportion of infected people experience long-term viral control, and prior HIV-2 infection is associated with slower HIV-1 disease progression in co-infected subjects. The majority of traditional and next generation sequencing methods have relied on target amplification prior to sequencing, introducing biases that may obscure the true signals of diversity in the viral population. Additionally, target-enrichment through PCR requires a priori sequence knowledge, which is lacking for HIV-2. Therefore, a target enrichment free method of library preparation would be valuable for the field. We applied an RNA shotgun sequencing (RNA-Seq) method without PCR amplification to cultured viral stocks and patient plasma samples from HIV-2 infected individuals. Libraries generated from total plasma RNA were analysed with a two-step pipeline: (1) de novo genome assembly, followed by (2) read re-mapping. By this approach, whole genome sequences were generated with a 28x-67x mean depth of coverage. Assembled reads showed a low level of GC-bias and comparison of the genome diversity on the intra-host level showed low diversity in the accessory gene vpx in all patients. Our study demonstrates that RNA-Seq is a feasible full-genome de novo sequencing method for blood plasma samples collected from HIV-2 infected individuals.\n\nIMPORTANCEAn accurate picture of viral genetic diversity is critical for the development of a globally effective HIV vaccine. However, sequencing strategies are often complicated by target enrichment prior to sequencing, introducing biases that can distort variant frequencies, which are not easily corrected for in downstream analyses. Additionally, detailed a priori sequence knowledge is needed to inform robust primer design when employing PCR amplification, a factor that is often lacking when working with tropical diseases localised in developing countries. Previous work has demonstrated that direct RNA shotgun sequencing (RNA-Seq) can be used to circumvent these issues for HCV and Norovirus. We applied shotgun RNA sequencing (RNA-Seq) to total RNA extracted from HIV-2 blood plasma samples, demonstrating the applicability of this technique to HIV-2 and allowing us to generate a dynamic picture of genetic diversity over the whole genome of HIV-2 in the context of low-bias sequencing.

microbiology↗