Biology subjects

Chaguza, C.

Publications and source records attributed to Chaguza, C..

13 recordsLinked to original sources

bioRxiv2026

The global evolutionary dynamics of natural capsule switching in a human-restricted bacterial pathogen

Exchange of genetic loci encoding extracellular capsular polysaccharides in bacteria can change capsular antigens or serotype, which, in turn, can lead to evasion of vaccines targeting the capsule. However, the evolutionary dynamics and drivers of bacterial capsule switching remain poorly understood. Here, we investigate the evolutionary dynamics and correlates of capsule switching in bacteria, using the human-restricted bacterial pathogen Streptococcus pneumoniae as an example, using 10,558 genetically and geographically diverse whole-genome sequences. Our phylogenomic and ancestral state reconstruction analysis revealed that the propensity for capsule switching varies by capsule type and genetic background. We found capsule switches only between ~4% of all possible capsule-type pairs, varying by specific pairs and switching direction. We also found associations between capsule switching patterns and genetic and ecological factors, including nasopharyngeal carriage duration, capsule biosynthesis locus size, and capsule thickness. Furthermore, the genetic similarity of the replacing and replaced capsule loci was primarily associated with the frequency of switching between specific capsule type pairs. Although a few specific non-capsule genomic loci were associated with the propensity for capsule switching across distinct lineages, the collective effect of the genetic background appears to influence capsule switching more strongly. These findings shed light on how bacterial capsule types and lineages undergo capsule switching influenced by genetic and ecological factors, informing our understanding of the emergence of novel capsule-genotype combinations that may impact the design of capsule-targeting glycoconjugate vaccines.

microbiology↗

bioRxiv2026

MxSure: a mixture model for inferring within-host substitution rates and transmission SNP thresholds

Quantifying short-term evolutionary rates of microbial genomes is essential for understanding the processes that shape within-host evolution and for establishing thresholds needed to track transmission. In studies of short-term evolutionary rates, samples are often collected from closely related clusters (e.g. longitudinally from the same host or from transmission pairs), with substantial time intervals separating genomes between clusters. Distinguishing strain replacement from persistence presents is also difficult in these studies. In addition, many public health and metagenomic bacterial strain tracking pipelines output pairwise SNP distances rather than the multiple sequence alignments required by common substitution rate estimation pipelines. This makes it hard to estimate within-host evolutionary rates in many commensal bacterial species that are difficult to culture and isolate. To address these challenges, we introduce MxSure, a tool for estimating substitution rates and transmission thresholds while accounting for strain replacement from pairwise SNP distance data, as commonly generated by transmission tracking and metagenomic analysis pipelines. We demonstrate the accuracy of MxSure through extensive simulations and by analysing species with previously estimated substitution rates from longitudinal metagenomic datasets. Using MxSure, we estimated within-host substitution rates and transmission SNP thresholds for multiple commensal bacterial species including Bifidobacterium longum and Bifidobacterium bifidum from a longitudinal study of the infant gut microbiome.

bioinformatics↗

bioRxiv2026

RNA Quality Control Enables Antibiotic Tolerance

The mechanisms underlying adaptation to antibiotic pressure within complex host environments remain incompletely understood. By experimentally evolving Streptococcus pneumoniae subjected to various antibiotics and immune states, we demonstrate populations adopting distinct adaptive strategies depending on specific selective context. General antibiotic stress drives convergent mutations in rny, encoding the RNA degradosome scaffold RNase Y, that exhibit broad-spectrum antibiotic tolerance and accelerated recovery. Single-cell transcriptomics revealed antibiotic-induced death is driven by transcriptional collapse, a catastrophic loss of RNA quantity and integrity. In contrast, rny mutants avert this via a bet-hedging strategy: a resilient minority maintains a baseline transcriptional profile, while a quiescent majority undergoes selective RNA degradation to preserve transcript fidelity. Upon stress removal, these populations execute a prioritized transcriptional ribosomal reboot, facilitating accelerated recovery. These findings identify RNA turnover as a tunable master regulator to survive combined pressures of antibiotics and immunity.

microbiology↗

bioRxiv2025

Streptococcus pneumoniae accessory capsular genes modulate fitness, pathogenicity and immune evasion

Globally, Streptococcus pneumoniae disproportionally affects children in resource-poor settings, older adults and people living with HIV. Frequently found as an asymptomatic colonizer of the nasopharynx, this versatile pathogen is a prominent cause of pneumonia, meningitis, bacteraemia and otitis media. Recently, a serotype 3 capsule variant (GPSC10-ST700) has expanded in Malawi with enhanced vaccine escape potential. Here, using a mutational and complementation approach, we show that loss of accessory capsular genes in GPSC10-ST700 contribute to increased opsonophagocytic resistance in this lineage. Although originally thought to be nonfunctional pseudogenes, we show that these genes modulate fitness and the global phosphoproteome in serotype 3 strains. These findings highlight that vaccine escape may be mediated through variations in the pneumococcal capsular locus that enhance fitness, pathogenicity and immune evasion, without capsule switching. IMPORTANCEPneumococcal polysaccharide-conjugate vaccines (PCV) target the polysaccharide capsule (CPS), which is a dominant virulence factor. However, current PCVs induce suboptimal protection against serotype 3 strains, which produce a thicker capsule that when released from the bacterial surface, interferes with antibody-mediated bacterial killing and protection. We recently described the clonal expansion of a sequence type (ST) 700-GPSC10 serotype 3 lineage in Malawi post-PCV13 introduction. This lineage is characterized by the absence of at least 6 genes in its cps locus and a distinct antimicrobial resistance (AMR) profile compared to other serotype 3 strains. Here we uncovered a functional role for the accessory capsular genes (acl) in serotype 3, previously considered to be pseudogenes, which modulate capsule production, shedding, serum tolerance, and bacterial fitness. By linking genotype to phenotype, our work provides new insights into the molecular basis of serotype 3 immune evasion, informing the design of more effective pneumococcal vaccines.

microbiology↗

bioRxiv2024

Tiled Amplicon Sequencing Enables Culture-free Whole-Genome Sequencing of Pathogenic Bacteria From Clinical Specimens

Pathogen sequencing is an important tool for disease surveillance and demonstrated its high value during the COVID-19 pandemic. Viral sequencing during the pandemic allowed us to track disease spread, quickly identify new variants, and guide the development of vaccines. Tiled amplicon sequencing, in which a panel of primers is used for multiplex amplification of fragments across an entire genome, was the cornerstone of SARS-CoV-2 sequencing. The speed, reliability, and cost-effectiveness of this method led to its implementation in academic and public health laboratories across the world and adaptation to a broad range of viral pathogens. However, similar methods are not available for larger bacterial genomes, for which whole-genome sequencing typically requires in vitro culture. This increases costs, error rates and turnaround times. The need to culture poses particular problems for medically important bacteria such as Mycobacterium tuberculosis, which are slow to grow and challenging to culture. As a proof of concept, we developed two novel whole-genome amplicon panels for M. tuberculosis and Streptococcus pneumoniae. Applying our amplicon panels to clinical samples, we show the ability to classify pathogen subgroups and to reliably identify markers of drug resistance without culturing. Development of this work in clinical settings has the potential to dramatically reduce the time of diagnosis of drug resistance for multiple drugs in parallel, enabling earlier intervention for high priority pathogens.

genomics↗

bioRxiv2024

The capsule and genetic background, rather than specific loci, strongly influence pneumococcal growth kinetics

Bacterial growth characteristics intrinsic to each strain can impact and influence gene expression, antibiotic susceptibility, and disease pathogenesis. However, little is known about specific genomic variations that influence these bacterial growth features. Here, we investigate the impact of Streptococcus pneumoniae genetics on its in vitro growth features to shed light on genes that may be important targets in the development of vaccines and therapeutics. We use statistical models to estimate growth features and demonstrate that they varied significantly across capsular serotypes and lineages, were strongly correlated with phylogeny, and showed high heritability, highlighting a strong genetic basis. Despite this, genome-wide association studies revealed no specific genomic loci statistically associated with the growth features independently of the genetic background, including those in the locus responsible for capsular polysaccharide synthesis. Our findings suggest that the serotype and lineage, as well as a combination of genomic loci, influence intrinsic pneumococcal growth kinetics, which may have implications for pneumococcal disease pathogenesis.

microbiology↗

bioRxiv2024

Large-scale genomic analysis of SARS-CoV-2 Omicron BA.5 emergence in the United States

The COVID-19 pandemic is marked by the continuing emergence of novel SARS-CoV-2 variants. Questions remain about the mechanisms with which these lineages establish themselves in new geographical areas. In this study, we performed a discrete phylogeographic analysis on [~]19,000 SARS-CoV-2 sequences of Omicron BA.5 lineages between February and June 2022 to better understand how it emerged in different regions of the United States (U.S.). We found that the earliest introductions came from Africa, the putative origin of the variant, but the majority were from Europe, correlating with the high volume of air travelers. Additionally, the analysis revealed extensive domestic transmission between different regions of the U.S. driven by population size and cross-country transmission. Results suggest that most of the within-U.S. spread was between three regions that include California, New York, and Florida. Our results form a framework for understanding novel SARS-CoV-2 variant emergence in the U.S.

genomics↗

bioRxiv2024

Expansion of pneumococcal serotype 23F and 14 lineages with genotypic changes in capsule polysaccharide locus and virulence gene profiles post introduction of pneumococcal conjugate vaccine in Blantyre, Malawi

Since the introduction of the 13-valent pneumococcal conjugate vaccine (PCV13) in Malawi in 2011, there has been persistent carriage of vaccine serotype (VT) Streptococcus pneumoniae, despite high vaccine coverage. To determine if there has been a genetic change within the VT capsule polysaccharide (cps) loci since the vaccines introduction, we compared 1,022 whole-genome-sequenced VT isolates from 1998 to 2019. We identified the clonal expansion of a multidrug-resistant, penicillin non-susceptible serotype 23F GPSC14-ST2059 lineage, a serotype 14 GPSC9-ST782 lineage and a novel serotype 14 sequence type GPSC9-ST18728 lineage. Serotype 23F GPSC14-ST2059 had an I253T mutation within the capsule oligosaccharide repeat unit polymerase Wzy protein, which is predicted in silico to alter the protein pocket cavity. Moreover, serotype 23F GPSC14-ST2059 had SNPs in the DNA binding sites for the cps transcriptional repressors CspR and SpxR. Serotype 14 GPSC9-ST782 harbour a non-truncated version of the large repetitive protein (Lrp), containing a Cna protein B-type domain which is also present in proteins associated with infection and colonisation. These emergent lineages also harboured genes associated with antibiotic resistance, and the promotion of colonisation and infection which were absent in other lineages of the same serotype. Together these data suggest that in addition to serotype replacement, modifications of the capsule locus associated with changes in virulence factor expression and antibiotic resistance may promote vaccine escape. In summary, the study highlights that the persistence of vaccine serotype carriage despite high vaccine coverage in Malawi may be partly caused by expansion of VT lineages post PCV13 rollout. Impact StatementOur findings highlight the potential for clonal expansion of multidrug-resistant, penicillin-non-susceptible vaccine serotype lineages with capsule locus modifications, within a high carriage and disease burden population. This shift has occurred among young children where there has been high vaccine coverage, posing challenges for effective vaccine scheduling and design. Furthermore, this study emphasises the importance of ongoing Streptococcus pneumoniae genomic surveillance as new or modified pneumococcal vaccines are implemented. 2. Data summaryWhole genome sequencing assemblies for the PCVPA survey have been deposited in the BioProject PRJNA1011974.

microbiology↗

bioRxiv2023

Pan-genome study underling the extent of genomic variation of invasive Streptococcus pneumoniae in Malawi

AbstractStreptococcus pneumoniae is a common cause of acute bacterial infections in Malawi. Understanding the molecular mechanisms underlying its invasive behavior is crucial for designing new therapeutic strategies. We conducted a pan-genome analysis to identify potential virulence genes in S. pneumoniae by comparing the gene pool of isolates from carriers nasopharyngeal secretions to isolates from the blood and cerebrospinal fluid of patients. Our analysis involved 1,477 pneumococcal isolates from Malawi, comprising 825 samples from carriers (nasopharyngeal swab) and 652 from patients (368 from blood and 284 from cerebrospinal fluid). We identified 56 serotypes in the cohort. While most serotypes exhibited a similar prevalence in both carriage and disease groups, serotypes 1 and 5, the most abundant serotypes in the entire cohort, were significantly more commonly detected in specimens from patients compared to the carriage group. This difference is presumably due to their shorter nasopharyngeal colonization period. Furthermore, these serotypes displayed genetic distinctiveness from other serotypes. A magnificent genetic difference was observed in the absence of genes from the RD8a genomic island in serotypes 1 and 5 compared to significantly prevalent serotypes in the nasopharynx. RD8a genes play pivotal roles in binding to epithelial cells and performing aerobic respiration to synthesize ATP through oxidative phosphorylation. The absence of RD8a from serotypes 1 and 5 may be associated with a shorter duration in the nasopharynx, theoretically due to a reduced capacity to bind to epithelial cells and access free oxygen molecules required for aerobic respiration (essential to maintain the carriage state). Serotypes 1 and 5, significantly harbor operons that encode phosphoenolpyruvate phosphotransferase systems, which might relate to transporting carbohydrates, relying on phosphoenolpyruvate as the energy source instead of ATP. In conclusion, serotypes 1 and 5 as the most prevalent invasive pneumococcal strains in Malawi, displayed considerable genetic divergence from other strains, which may offer insights into their invasiveness and potential avenues for further research. Author summaryDespite introducing the pneumococcal conjugate vaccine in 2011, Streptococcus pneumoniae remains a major cause of bacterial infection in Malawi. Whilst some pneumococcal strains harmlessly colonize the nasopharynx, others find their way into normally sterile sites, such as lungs, blood, and nervous system, resulting in serious illness. Our study identified specific pneumococcal serotypes as the most invasive in Malawi, characterized by a short colonization period and significant genetic distinctiveness from other strains. This genetic divergence notably included the absence of several genes associated with aerobic respiration and the presence of genes facilitating ATP-independent carbohydrate transport. The presence or absence of these genes may underlie their heightened invasiveness and shorter colonization period. This hypothesis positions these genes as potential candidates for future therapeutic research. We propose that the specific gene gain and/or loss in invasive versus other serotypes may be linked to the development of invasive pneumococcal diseases. Impact StatementOur research applied pan-genomics principles to comprehensively assess diversity within the pneumococcus genome, with the primary objective of identifying pneumococcal virulence genes for advancing vaccine design and drug development. Within this study, we identified Serotypes 1 and 5 as the predominant and highly invasive pneumococcal strains in Malawi, characterized by a short nasopharyngeal colonization period, suggesting their potential for rapid infection of sterile sites within the human body such as blood and the central nervous system. These serotypes exhibited significant genetic divergence from other serotypes in Malawi, notably lacking key genes within the RD8a operon while harboring transporters functioning independently of ATP. Its important to note that these findings are based on computational analysis, and further validation through laboratory experiments is essential to confirm their biological significance and potential clinical applications. The implications of our research offer potential avenues for more effective pneumococcal disease prevention and treatment, not only in Malawi but also in regions facing similar challenges.

microbiology↗

bioRxiv2022

The population-level impact of Enterococcus faecalis genetics on intestinal colonisation and extraintestinal infection

Enterococcus faecalis is a commensal pathogenic bacterium commonly found in the human gastrointestinal tract and a cause of opportunistic infections typically associated with multidrug resistance. The E. faecalis genetic changes associated with pathogenicity and extraintestinal infection, particularly through gut-to-bloodstream translocation, are poorly understood. Here, we investigate the E. faecalis genetic signatures associated with intestinal colonisation and extraintestinal infection and infection of hospitalised and non-hospitalised individuals using heritability estimation and a genome-wide association study (GWAS). We analysed 750 whole-genome sequences of faecal and bloodstream E. faecalis isolates from hospitalised patients and non-hospitalised individuals, respectively, predominantly in Europe. We found that E. faecalis infection of individuals depending on their hospitalisation status and extraintestinal infection are heritable traits and that [~]24% and [~]34% of their variation is explained by the considered genetic effects, respectively. Further, a GWAS using linear mixed models did not pinpoint any clear enrichment of individual genetic changes in isolates from different isolation sites and individuals with varying hospitalisation statuses, suggesting that these traits are highly polygenic. Altogether, our findings indicate that E. faecalis infection and extraintestinal infection are influenced by variation in genetic, host, and environmental factors, and ultimately the opportunistic pathogenic lifestyle of this versatile host generalist bacterium.

microbiology↗

bioRxiv2022

Pneumococcal within-host diversity during colonisation, transmission and treatment

Characterising the genetic diversity of pathogens within the host promises to greatly improve surveillance and reconstruction of transmission chains. For bacteria, it also informs our understanding of inter-strain competition, and how this shapes the distribution of resistant and sensitive bacteria. Here we study the genetic diversity of Streptococcus pneumoniae within individual infants and their mothers by deep sequencing whole pneumococcal populations from longitudinal nasopharyngeal samples. We demonstrate deep sequencing has unsurpassed sensitivity for detecting multiple colonisation, doubling the rate at which highly invasive serotype 1 bacteria were detected in carriage compared to gold-standard methods. The greater resolution identified an elevated rate of transmission from mothers to their children in the first year of the childs life. Comprehensive treatment data demonstrated infants were at an elevated risk of both the acquisition, and persistent colonisation, of a multidrug resistant bacterium following antimicrobial treatment. Some alleles were enriched after antimicrobial treatment, suggesting they aided persistence, but generally purifying selection dominated within-host evolution. Rates of co-colonisation imply that in the absence of treatment, susceptible lineages outcompeted resistant lineages within the host. These results demonstrate the many benefits of deep sequencing for the genomic surveillance of bacterial pathogens.

genomics↗

bioRxiv2020

Identification of colonisation and virulence determinants of Streptococcus pneumoniae via experimental evolution in mouse infection models

Streptococcus pneumoniae is a commensal of the human nasopharynx and a major cause of respiratory and invasive disease. Pneumococcus stimulates upper respiratory tract inflammation that promotes shedding from mucosal surfaces and transmission to new hosts. Colonisation and transmission are partially antagonistic processes. Adhesion to surfaces and evasion of host responses favours the former, whilst detachment, promoted by inflammation, is necessary for the latter. We sought to determine how adaptation and evolution of pneumococcus within its nasopharyngeal niche might progress when selective pressures associated with transmission were removed. This was achieved by serial passage of pneumococci in mouse models of nasopharyngeal carriage, manually transferring bacteria between mice. To assess the role of host environmental factors on pneumococcal evolution, we also performed analogous experimental evolution in a mouse pneumonia model, passaging pneumococci through lungs. Nasopharynx-passaged pneumococci became more effective colonisers, whilst those evolved within lungs showed reduced virulence. We observed selection of mutations in genes associated with cell wall biogenesis and metabolism in both nasopharynx and lung lineages, but identified prominent examples of parallel evolution that were niche specific. We focussed on gpsA, a gene in which the same single nucleotide polymorphism arose in two independently evolved nasopharynx-passaged lineages. We identified a single nucleotide change conferring resistance to oxidative stress and enhanced nasopharyngeal colonisation potential. We show that gpsA is also a frequent target of mutation during human colonisation. These findings highlight the role played by the host environment in determining trajectories of bacterial evolution and the potential of experimental evolution in animal infection models for identification of novel pathogen virulence and colonisation factors.

microbiology↗

bioRxiv2018

Early signals of vaccine driven perturbation seen in pneumococcal carriage population genomic data

Pneumococcal conjugate vaccines (PCV) have reduced pneumococcal diseases globally. Despite this, much remains to be learned about their effect on pathogen population structure. Here we undertook whole genome sequencing of 660 pneumococcal strains from asymptomatic carriers to investigate population restructuring in pneumococcal strains sampled before and after PCV13 introduction in a previously vaccine-naive setting. We show substantial decreasing frequency of vaccine-type (VT) strains and their strain diversity post-vaccination in the vaccinated but not unvaccinated age groups indicative of direct but limited or delayed indirect effect of vaccination. Clearance of identical VT serotypes associated with multiple lineages occurred regardless of their genetic background. Interestingly, despite the increasing frequency of non-vaccine type (NVT) strains through serotype replacement, the serotype diversity was not fully restored to the levels observed prior to vaccination implying limited serotype replacement. The frequency of antibiotic resistant strains was low and remained largely unchanged post-vaccination but intermediate-penicillin-resistant lineages were reduced in the post vaccine population. Significant perturbations marked by changing frequency of accessory genes associated with diverse functions especially mobile genetic elements and bacteriocin activity were detected. This phylogenomic analysis demonstrates early vaccine-induced pneumococcal population restructuring not only at serotype but also accessory genome level.\n\nAuthor summaryDifferent formulations of PCVs have been effective in reducing the invasive pneumococcal disease burden globally. Clinical trials have started to indicate high impact and effectiveness of PCV13 in Sub Saharan Africa (SSA) but there is limited understanding of how the introduction of PCVs alters the population structure of pneumococcal strains at serotype and genomic level. Here we investigated this using pneumococcal strains sampled pre- and post-PCV13 introduction from a previously vaccine naive setting in Northern Malawi. Our findings reveal decrease in frequency of VT serotypes and their associated lineages in the largely vaccinated under-five population but not older individuals indicating a direct but limited or delayed indirect protection. The diversity of serotypes also decreased post-vaccination in VT strains in the under-fives but there was no change in NVT strains suggesting incomplete serotype replacement. At the genomic level, logistic regression revealed changing frequency of accessory genes largely associated with mobile genetic elements but such changes did not include any antibiotic resistance genes. These findings show significant perturbations at serotype and accessory genome level in carried pneumococcal population after two years from PCV13 introduction but the pneumococcal population was still perturbed and had not returned to a new equilibrium state.

genomics↗