Biology subjects

Lacharme-Lora, L.

Publications and source records attributed to Lacharme-Lora, L..

5 recordsLinked to original sources

bioRxiv2023

Salmonella enterica serovar Typhimurium ST313 sublineage 2.2 has emerged in Malawi with a characteristic gene expression signature and a fitness advantage

Invasive non-typhoidal Salmonella (iNTS) disease is a serious bloodstream infection that targets immune-compromised individuals, and causes significant mortality in sub-Saharan Africa. Salmonella enterica serovar Typhimurium ST313 causes the majority of iNTS in Malawi, and we performed an intensive comparative genomic analysis of 608 isolates obtained from fever surveillance at the Queen Elizabeth Hospital, Blantyre between 1996 and 2018. We discovered that following the upsurge of the well-characterised S. Typhimurium ST313 lineage 2 from 1999 onwards, two new multidrug-resistant sublineages designated 2.2 and 2.3, emerged in Malawi in 2006 and 2008, respectively. The majority of S. Typhimurium isolates from human bloodstream infections in Malawi now belong to sublineage 2.2 or 2.3. To identify factors that characterised the emergence of the prevalent ST313 sublineage 2.2, we performed genomic and functional analysis of two representative strains, D23580 (lineage 2) and D37712 (sublineage 2.2). Comparative genomic analysis showed that the chromosome of ST313 lineage 2 and sublineage 2.2 were broadly similar, only differing by 29 SNPs and small indels and a 3kb deletion in the Gifsy-2 prophage region that spanned the sseI pseudogene. Lineage 2 and sublineage 2.2 have unique plasmid profiles that were verified by long read sequencing. The transcriptome was initially explored in 15 infection-relevant conditions and within macrophages. Differential gene expression was subsequently investigated in depth in the four most important in vitro growth conditions. We identified up-regulation of SPI2 genes in non-inducing conditions, and down-regulation of flagellar genes in D37712, compared to D23580. Following phenotypic confirmation of transcriptional differences, we discovered that sublineage 2.2 had increased fitness compared with lineage 2 during mixed-growth in minimal media. We speculate that this competitive advantage is contributing to the continuing presence of sublineage 2.2 in Malawi.

bioinformatics↗

bioRxiv2022

Genome-wide fitness analysis identifies genes required for in vitro growth and macrophage infection by African and Global Epidemic pathovariants of Salmonella Enteritidis

Salmonella Enteritidis is the second most common serovar associated with invasive non-typhoidal Salmonella (iNTS) disease in sub-Saharan Africa. Previously, genomic and phylogenetic characterisation of S. Enteritidis isolates from human bloodstream led to the discovery of the Central/Eastern African (CEAC) and West African clades, which were distinct from the gastroenteritis-associated Global Epidemic clade (GEC). The African S. Enteritidis clades have unique genetic signatures that include genomic degradation, novel prophage repertoires and multi-drug resistance, but the molecular basis for the enhanced propensity of African S. Enteritidis to cause bloodstream infection is poorly understood. We used transposon insertion sequencing (TIS) to identify the genetic determinants of the GEC representative strain P125109 and the CEAC representative strain D7795 for growth in three in vitro conditions (LB or minimal NonSPI2 and InSPI2 growth media), and for survival and replication in RAW 264.7 murine macrophages. We identified 207 in vitro-required genes that were common to both S. Enteritidis strains and also required by S. Typhimurium, S. Typhi and Escherichia coli, and 63 genes that were only required by individual S. Enteritidis strains. Similar types of genes were required by both P125109 and D7795 for optimal growth in particular media. Screening the transposon libraries during macrophage infection identified 177 P125109 and 201 D7795 genes that contribute to bacterial survival and replication in mammalian cells. The majority of these genes have proven roles in Salmonella virulence. Our analysis also revealed candidate strain-specific macrophage fitness genes, some of which represent potential novel Salmonella virulence factors. IMPACT STATEMENTInvasive non-typhoidal Salmonella (iNTS) disease is a systemic infection that has a high case fatality rate of 15% and is responsible for an estimated 66,500 deaths/year in sub-Saharan Africa. The main causative agents are pathovariants of Salmonella Typhimurium, known as S. Typhimurium ST313, and Salmonella Enteritidis (S. Enteritidis), known as Central/Eastern African (CEAC) and West African S. Enteritidis. Whilst the African S. Typhimurium pathovariant has been an active focus of research over the past decade, studies on African S. Enteritidis have been lacking. We used transposon insertion sequencing (TIS) to identify the genetic requirements of both African and Global Epidemic S. Enteritidis to grow in vitro and to infect murine macrophages. To our knowledge, this is the first genome-wide functional analysis of African S. Enteritidis under conditions relevant to infection of a mammalian host. We show that the gene sets required for growth under laboratory conditions and macrophage infection by African and Global Epidemic S. Enteritidis were broadly similar, and that the majority of the genes that contribute to survival and replication in macrophage already have proven roles in Salmonella virulence. Our analysis did identify candidate strain-specific macrophage fitness genes, some of which could be novel Salmonella virulence factors.

microbiology↗

bioRxiv2018

Campylobacter jejuni transmission and colonisation in broiler chickens is inhibited by Faecal Microbiota Transplantation.

BACKGROUNDCampylobacter jejuni, the most frequent cause of foodborne bacterial infection, is found on around 70% of retail chicken. As such there is a need for effective controls in chicken production. Microbial-based controls such as probiotics are attractive to the poultry industry, but of limited efficacy. Furthermore, as commercially-produced chickens have no maternal contact, their pioneer microbiome is likely to come from the hatchery environment. Early delivery of microbials that lead to a more natural avian microbiome may, therefore, improve bird health and reduce susceptibility to C.jejuni colonisation.\n\nA faecal microbiota transplant (FMT) was used to transfer a mature cecal microbiome to newly-hatched broiler chicks and its effects on C.jejuni challenge assessed. We used both a seeder-bird infection model that mimics natural bird-to-bird infection alongside a direct-challenge model. We used a 16S rRNA gene sequencing-based approach to characterize the transplant material itself alongside changes to the chicken microbiome following FMT.\n\nRESULTSFMT changes the composition of the chicken intestinal microbiome. We observed little change in species richness following FMT compared to untreated samples, but there is an increase in phylogenetic diversity within those species. The most significant difference in the ceca is an increase in Lactobacilli, although not a major component of the transplant material, suggesting the FMT results in a change in the intestinal milieu as much as a direct change to the microbiome.\n\nUpon direct challenge, FMT resulted in lower initial intestinal colonisation with C.jejuni. More significantly, in a seeder-bird challenge of infection transmission, FMT reduced transmission and intestinal colonisation until common UK retail age of slaughter. In a repeat experiment, transmission was completely blocked following FMT treatment. Delayed FMT administration at 7 days of-age had limited effect on colonisation and transmission.\n\nCONCLUSIONSWe show that transfer of a whole mature microbiome to newly-hatched chicks reduces transmission and colonisation of C.jejuni. This indicates that modification of the broiler chick microbiome can reduce intestinal colonisation of C.jejuni to levels projected to lead to lower the human infection rate. We believe these findings offer a way to identify key taxa or consortia that are effective in reducing C.jejuni colonisation and improving broiler gut health.

microbiology↗

bioRxiv2018

Adding function to the genome of African Salmonella ST313

Salmonella Typhimurium ST313 causes invasive nontyphoidal Salmonella (iNTS) disease in sub-Saharan Africa, targeting susceptible HIV+, malarial or malnourished individuals. An in-depth genomic comparison between the ST313 isolate D23580, and the well-characterized ST19 isolate 4/74 that causes gastroenteritis across the globe, revealed extensive synteny. To understand how the 856 nucleotide variations generated phenotypic differences, we devised a large-scale experimental approach that involved the global gene expression analysis of strains D23580 and 4/74 grown in sixteen infection-relevant growth conditions. Comparison of transcriptional patterns identified virulence and metabolic genes that were differentially expressed between D23580 versus 4/74, many of which were validated by proteomics. We also uncovered the S. Typhimurium D23580 and 4/74 genes that showed expression differences during infection of murine macrophages. Our comparative transcriptomic data are presented in a new enhanced version of the Salmonella expression compendium SalComD23580: bioinf.gen.tcd.ie/cgi-bin/salcom_v2.pl. We discovered that the ablation of melibiose utilization was caused by 3 independent SNP mutations in D23580 that are shared across ST313 lineage 2, suggesting that the ability to catabolise this carbon source has been negatively selected during ST313 evolution. The data revealed a novel plasmid maintenance system involving a plasmid-encoded CysS cysteinyl-tRNA synthetase, highlighting the power of large-scale comparative multi-condition analyses to pinpoint key phenotypic differences between bacterial pathovariants.

microbiology↗

bioRxiv2017

The role of a single non-coding nucleotide in the evolution of an epidemic African clade of Salmonella

Introductory ParagraphSalmonella enterica serovar Typhimurium ST313 is a relatively newly emerged sequence type that is causing a devastating epidemic of bloodstream infections across sub-Saharan Africa. Analysis of hundreds of Salmonella genomes has revealed that ST313 is closely-related to the ST19 group of S. Typhimurium that cause gastroenteritis across the world. The core genomes of ST313 and ST19 vary by just 1000 single-nucleotide polymorphisms (SNPs). We hypothesised that the phenotypic differences that distinguish African Salmonella from ST19 are caused by certain SNPs that directly modulate the transcription of virulence genes.\n\nHere we identified 3,597 transcriptional start sites (TSS) of the ST313 strain D23580, and searched for a gene expression signature linked to pathogenesis of Salmonella. We identified a SNP in the promoter of the pgtE gene that caused high expression of the PgtE virulence factor in African S. Typhimurium, increased the degradation of the factor B component of human complement, contributed to serum resistance and modulated virulence in the chicken infection model. The PgtE protease is known to mediate systemic infection in animal models. We propose that high levels of expression PgtE of by African S. Typhimurium ST313 promotes bacterial survival and bacterial dissemination during human infection.\n\nOur finding of a functional role for an extra-genic SNP shows that approaches used to deduce the evolution of virulence in bacterial pathogens should include a focus on non-coding regions of the genome.

molecular biology↗