Biology subjects

Hinton, J. C. D.

Publications and source records attributed to Hinton, J. C. D..

27 records · Page 2Linked to original sources

bioRxiv2020

Prophage-encoded phage defence proteins with cognate self-immunity

Temperate phages are pervasive in bacterial genomes, existing as vertically-inherited islands called prophages. Prophages are vulnerable to the predation of their host bacterium by exogenous phages. Here we identify BstA, a novel family of prophage-encoded phage defense proteins found in diverse Gram-negative bacteria. BstA drives potent suppression of phage epidemics through abortive infection. During lytic replication, the bstA-encoding prophage is not itself inhibited by BstA due to a self-immunity mechanism conferred by the anti-BstA (aba) element, a short stretch of DNA within the bstA locus. Inhibition of phage replication by distinct BstA proteins from Salmonella, Klebsiella and Escherichia prophages is functionally interchangeable, but each possesses a cognate aba element. The specificity of the aba element ensures that immunity is exclusive to the replicating prophage, and cannot be exploited by heterologous BstA-encoding phages. BstA allows prophages to defend host cells against exogenous phage attack, without sacrificing their own lytic autonomy.

microbiology↗

bioRxiv2019

A window into lysogeny: Revealing temperate phage biology with transcriptomics

Integrated phage elements, known as prophages, are a pervasive feature of bacterial genomes. Prophages can enhance the fitness of their bacterial hosts by conferring beneficial functions, such as virulence, stress tolerance or phage resistance, which are encoded by accessory loci. Whilst the majority of phage-encoded genes are repressed during lysogeny, accessory loci are often highly expressed. However, novel prophage accessory loci are challenging to identify based on DNA sequence data alone. Here, we use bacterial RNA-seq data to examine the transcriptional landscapes of five Salmonella prophages. We show that transcriptomic data can be used to heuristically enrich for prophage features that are highly expressed within bacterial cells and often represent functionally-important accessory loci. Using this approach we identify a novel anti-sense RNA species in prophage BTP1, STnc6030, which mediates superinfection exclusion of phage BTP1 and immunity to closely-related phages. Bacterial transcriptomic datasets are a powerful tool to explore the molecular biology of temperate phages.

microbiology↗

bioRxiv2019

African Salmonella Typhimurium sequence type 313 lineage 2 evades MAIT cell recognition by overexpressing RibB

Mucosal-associated invariant T (MAIT) cells are a subset of innate T lymphocytes activated by bacteria that produce vitamin B2 metabolites. Mouse models of infection have demonstrated a role for MAIT cells in antimicrobial defence. However, proposed protective roles of MAIT cells in human infections remain unproven and clinical conditions associated with a selective absence of MAIT cells have not been identified. We report that typhoidal and non-typhoidal S. enterica strains generally activate MAIT cells. However, African invasive disease-associated multidrug-resistant S. Typhimurium sequence type 313 lineage 2 strains escape MAIT cell recognition through overexpression of ribB, a bacterial gene that encodes the 4-dihydroxy-2-butanone 4-phosphate synthase enzyme of the riboflavin biosynthetic pathway. This MAIT cell-specific phenotype did not extend to other innate lymphocytes. We propose that ribB overexpression is an evolved trait that facilitates evasion from immune recognition by MAIT cells and contributes to the invasive pathogenesis of S. Typhimurium sequence type 313 lineage 2 in vivo.

immunology↗

bioRxiv2019

The fitness landscape of the African Salmonella Typhimurium ST313 strain D23580 reveals unique properties of the pBT1 plasmid

We have used a transposon insertion sequencing (TIS) approach to establish the fitness landscape of the African Salmonella enterica serovar Typhimurium ST313 strain D23580, to complement our previous comparative genomic and functional transcriptomic studies. We used a genome-wide transposon library with insertions every 10 nucleotides to identify genes required for survival and growth in vitro and during infection of murine macrophages. The analysis revealed genomic regions important for fitness under two in vitro growth conditions. Overall, 724 coding genes were required for optimal growth in LB medium, and 851 coding genes were required for growth in SPI-2-inducing minimal medium. These findings were consistent with the essentiality analyses of other S. Typhimurium ST19 and S. Typhi strains. The global mutagenesis approach also identified 60 sRNAs and 413 intergenic regions required for growth in at least one in vitro growth condition. By infecting murine macrophages with the transposon library, we identified 68 genes that were required for intra-macrophage replication but did not impact fitness in vitro. None of these genes were unique to S. Typhimurium D23580, consistent with a high conservation of gene function between S. Typhimurium ST313 and ST19 and suggesting that novel virulence factors are not involved in the interaction of strain D23580 with murine macrophages. We discovered that transposon insertions rarely occurred in many pBT1 plasmid-encoded genes (36), compared with genes carried by the pSLT-BT virulence plasmid and other bacterial plasmids. The key essential protein encoded by pBT1 is a cysteinyl-tRNA synthetase, and our enzymological analysis revealed that the plasmid-encoded CysRSpBT1 had a lower ability to charge tRNA than the chromosomally-encoded CysRSchr enzyme. The presence of aminoacyl-tRNA synthetases in plasmids from a range of gram-negative and gram-positive bacteria suggests that plasmid-encoded essential genes are more common than had been appreciated.

microbiology↗

bioRxiv2019

Genomic and phylogenetic analysis of Salmonella Typhimurium and its monophasic variants responsible for invasive endemic infections in Colombia

Salmonellosis is an endemic human infection, associated with both sporadic cases and outbreaks throughout Colombia. Typhimurium is the most common Colombian serovar of Salmonella enterica, responsible for 32.5% of the Salmonella infections. Whole genome sequencing (WGS) is being used increasingly in Europe and the USA to study the epidemiology of Salmonella, but there has not yet been a WGS-based analysis of Salmonella associated with bloodstream infection in Colombia. Here, we analysed 209 genome sequences of Colombian S. Typhimurium and monophasic S. 4,[5],12:i:-isolates from Colombia from 1999 to 2017. We used a core genome-based maximum likelihood tree to define seven distinct clusters which were predominantly Sequence Type (ST) 19 isolates. We also identified the first ST313 and monophasic ST34 isolates to be reported in Colombia. The history of each cluster was reconstructed with a Bayesian tree to reveal a timeline of evolution. Cluster 7 was closely related to European multidrug-resistant (MDR) DT104. Cluster 4 became the dominant variant of Salmonella in 2016, and resistance to nalidixic acid was associated with a plasmid-encoded qnrB19 gene. Our findings suggest multiple transfers of S. Typhimurium between Europe and Colombia.\n\nAuthor summaryThe large-scale genome sequencing of Salmonella Typhimurium and monophasic Salmonella 4,[5],12:i:-involved bloodstream isolates from Colombia. The two serovars were responsible for about 1/3 of Salmonella infections in Colombia in the past 20 years. To identify the population structure we used Whole Genome Sequencing, performed in silico sequence typing, obtained phylogenetic trees, inferred the evolutionary history, detected the plasmids and prophages, and associated the antibiotic resistance (AMR) genotype with phenotype. Different clusters showed temporal replacement. The Colombian sequence type 313 was distinct from African lineages due to the absence of a key virulence-related gene, bstA. One of the Colombian clusters is likely to belong to the global epidemic of DT104, according to the evolutionary history and the AMR profile. The most common cluster in recent years was resistant to nalidixic acid and carried a plasmid-mediated antibiotic resistant gene qnrB19. Our findings will inform the ongoing efforts to combat Salmonellosis by Colombian public health departments.

genomics↗

bioRxiv2019

The diversity, evolution and ecology of Salmonella in venomous snakes

BackgroundReptile-associated Salmonella are a major, but often neglected cause of both gastrointestinal and bloodstream infection globally. The diversity of Salmonella enterica has not yet been determined in venomous snakes, however other cold-blooded animals have been reported to carry a broad range of Salmonella bacteria. We investigated the prevalence and assortment of Salmonella in a collection of venomous snakes in comparison with non-venomous reptiles.\n\nMethodology/Principle FindingsWe used a combination of selective enrichment techniques and whole-genome sequencing. We established a unique dataset of reptilian isolates to study Salmonella enterica species-level evolution and ecology and investigated differences between phylogenetic groups. We observed that 91% of venomous snakes carried Salmonella, and found substantial diversity between the serovars (n=58) carried by reptiles. The Salmonella serovars belonged to four of the six Salmonella enterica subspecies: diarizonae, enterica, houtanae and salamae. Subspecies enterica isolates were distributed among two distinct phylogenetic clusters, previously described as clade A (52%) and clade B (48%). We identified metabolic differences between S. diarizonae, S. enterica clade A and clade B involving growth on lactose, tartaric acid, dulcitol, myo-inositol and allantoin.\n\nSignificanceWe present the first whole genome-based comparative study of the Salmonella bacteria that colonise venomous and non-venomous reptiles and shed new light on Salmonella evolution. The findings raise the possibility that venomous snakes are a reservoir for human Salmonellosis in Africa. The proximity of venomous snakes to human dwellings in rural Africa may result in contaminated faecal matter being shed on surfaces and in water sources used for human homes and to irrigate salad crops. Because most of the venomous snakes had been captured in Africa, we conclude that the high level of Salmonella diversity reflects the African environmental niches where the snakes have inhabited.\n\nAuthor SummarySalmonella enterica is a remarkable bacterial species that causes Neglected Tropical Diseases globally. The burden of disease is greatest in some of the most poverty-afflicted regions of Africa, where salmonellosis frequently causes bloodstream infection with fatal consequences. The bacteria have the ability to colonise the gastrointestinal tract of a wide range of animals including reptiles. Direct or indirect contact between reptiles and humans can cause Salmonellosis. In this study, we determined the prevalence and diversity of Salmonella in a collection of African venomous snakes for the first time. Using the power of genomics, we showed that the majority of venomous snakes (91%) carry Salmonella, two thirds of which belonged to a subspecies of S. enterica called enterica, which is associated with most cases of human salmonellosis. Within the S. enterica subspecies we identified two evolutionary groups which display distinct growth patterns on infection relevant carbon sources. Our findings could have particular significance in Africa where venomous snakes wander freely around human dwellings and potentially shed contaminated faecal matter in water sources and on surfaces in rural homes.

microbiology↗

bioRxiv2017

The primary transcriptome, small RNAs, and regulation of antimicrobial resistance in Acinetobacter baumannii

AO_SCPCAPBSTRACTC_SCPCAPWe present the first high-resolution determination of transcriptome architecture in the priority pathogen Acinetobacter baumannii. Pooled RNA from 16 laboratory conditions was used for differential RNA-seq (dRNA-seq) to identify 3731 transcriptional start sites (TSS) and 110 small RNAs, including the first identification in A. baumannii of sRNAs encoded at the 3 end of coding genes. Most sRNAs were conserved among sequenced A. baumannii genomes, but were only weakly conserved or absent in other Acinetobacter species. Single nucleotide mapping of TSS enabled prediction of -10 and -35 RNA polymerase binding sites and revealed an unprecedented base preference at position +2 that hints at an unrecognized transcriptional regulatory mechanism. To apply functional genomics to the problem of antimicrobial resistance, we dissected the transcriptional regulation of the drug efflux pump responsible for chloramphenicol resistance, craA. The two craA promoters were both down-regulated >1000-fold when cells were shifted to nutrient limited medium. This conditional down-regulation of craA expression renders cells sensitive to chloramphenicol, a highly effective antibiotic for the treatment of multidrug resistant infections. An online interface that facilitates open data access and visualization is provided as \"AcinetoCom\" (http://bioinf.gen.tcd.ie/acinetocom/).

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↗

bioRxiv2017

Salmonella enterica Serovar Typhimurium ST313 Responsible For Gastroenteritis In The UK Are Genetically Distinct From Isolates Causing Bloodstream Infections In Africa

The ST313 sequence type of Salmonella enterica serovar Typhimurium causes invasive non-typhoidal salmonellosis amongst immunocompromised people in sub-Saharan Africa (sSA). Previously, two distinct phylogenetic lineages of ST313 have been described which have rarely been found outside sSA. Following the introduction of routine whole genome sequencing of Salmonella enterica by Public Health England in 2014, we have discovered that 2.7% (79/2888) of S. Typhimurium from patients in England and Wales are ST313. Of these isolates, 59/72 originated from stool and 13/72 were from extra-intestinal sites. The isolation of ST313 from extra-intestinal sites was significantly associated with travel to Africa (OR 12 [95% CI: 3,53]). Phylogenetic analysis revealed previously unsampled diversity of ST313, and distinguished UK-linked isolates causing gastroenteritis from African-associated isolates causing invasive disease. Bayesian evolutionary investigation suggested that the two African lineages diverged from their most recent common ancestors independently, circa 1796 and 1903. The majority of genome degradation of African ST313 lineage 2 is conserved in the UK ST313 lineages and only 10/44 pseudogenes were lineage 2-specific. The African lineages carried a characteristic prophage and antibiotic resistance gene repertoire, suggesting a strong selection pressure for these horizontally-acquired genetic elements in the sSA setting. We identified an ST313 isolate associated with travel to Kenya that carried a chromosomally-located blaCTX-M-15, demonstrating the continual evolution of this sequence type in Africa in response to selection pressure exerted by antibiotic usage.\n\nThe S. Typhimurium ST313 sequence type has been primarily associated with invasive disease in Africa. Here, we highlight the power of routine whole-genome-sequencing by public health agencies to make epidemiologically-significant deductions that would be missed by conventional microbiological methods. The discovery of ST313 isolates responsible for gastroenteritis in the UK reveals new diversity in this important sequence type. We speculate that the niche specialization of sub-Saharan African ST313 lineages is driven in part by the acquisition of accessory genome elements.

microbiology↗