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Foodborne diseases represent a link between environmental, animal, and human health interfaces from the One Health perspective. Whole genome sequencing (WGS) is becoming the gold standard in foodborne surveillance, worldwide. WGS than provides precision data from pathogens allows laboratories to resolve the genetic relations among all sources from One Health perspective, especially during outbreak investigations, getting insights into their transmission routes and pathogenicity.Salmonellaspp. is the most prevalent foodborne bacteria in Colombia, in 2020, 268 foodborne outbreaks were reported to National health institute (INS) and 3079 invasive and faecal salmonellosis samples from 81% of the Colombian geographical regions. Several studies in different food sources exhibited an increase of contamination withSalmonella, a public health concern due to the steady development of antimicrobial resistance associated to specific serovars. However, integration ofSalmonella spp. data including food-chain supply and clinical interfaces is very scarce in Colombia. This study carried out a national comparison ofSalmonellaisolates collected from food-chain supply and clinical sources. Using an epidemiological and phenotypic approach, we demonstrated the higher resolution of WGS compared with PFGE, routinely used inSalmonellasurveillance in Colombia. For example, the resolution of PFGE allowed the description of two main clusters of foodSalmonellaEnteritidis isolates which were expanded to eight clades by using WGS. Virulence factors and antimicrobial determinant genes observed in the foodborne clades should be considered a public health concern in Colombia. WGS is a technology that provides precise and valid evidence for the establishment of dissemination routes of foodborne high-riskSalmonellaclades, but it requires an integrative and continued collaboration between the stakeholders across the One Health interfaces to promote and support integrated real-time actions in public health.

We have developed an efficient and inexpensive pipeline for streamlining large-scale collection and genome sequencing of bacterial isolates. Evaluation of this method involved a worldwide research collaboration focused on the model organism Salmonella enterica, the 10KSG consortium. By optimising a logistics pipeline that collected isolates as thermolysates, permitting shipment in ambient conditions, the project assembled a diverse collection of 10,419 clinical and environmental isolates from low- and middle-income countries in less than one year. The bacteria were obtained from 51 countries/territories dating from 1949 to 2017, with a focus on Africa and Latin-America. All isolates were collected in barcoded tubes and genome sequenced using an optimised DNA extraction method and the LITE pipeline for library construction. After Illumina sequencing, the total reagent cost was less than USD$10 per genome. Our method can be applied to genome-sequence other large bacterial collections at a relatively low cost, within a limited timeframe, to support global collaborations.

2.AbstractSalmonella Enteritidis is one of the most commonly reported serovars of non-typhoidal Salmonella causing human disease and is responsible for both gastroenteritis and invasive non-typhoidal Salmonella (iNTS) disease worldwide. Whole-genome sequence (WGS) comparison of Salmonella Enteritidis isolates from across the world have identified three distinct clades, named Global Epidemic, Central/East African and West African, all of which have been implicated in epidemics: the Global Epidemic clade was linked to poultry-associated gastroenteritis, while the two African clades were related to iNTS disease. However, the distribution and epidemiology of these clades across Africa is poorly understood because identification of these clades currently requires whole genome sequencing capacity. Here, we report a sensitive, time- and cost-effective real-time PCR assay capable of differentiating between the Salmonella Enteritidis clades to facilitate surveillance and to inform public health responses.3.Impact statementChallenges in the diagnosis and treatment of invasive Salmonella Enteritidis (S. Enteritidis) bloodstream infections in sub–Saharan Africa are responsible for a case fatality rate of approximately 15% (12). It is important to identify distinct clades of S. Enteritidis in diagnostic laboratories in the African setting to determine whether particular outbreaks are associated with different health outcomes. Here, we have described the development of a high-quality molecular classification assay for the clade-typing of S. Enteritidis that is ideal for use in public health laboratories in resource-limited settings.

Background The Global Enteric Multicenter Study (GEMS) determined the etiologic agents of moderate-to-severe diarrhea (MSD) in children under 5 years old in Africa and Asia. Here, we describe the prevalence and antimicrobial susceptibility of nontyphoidal Salmonella (NTS) serovars in GEMS and examine the phylogenetics of Salmonella Typhimurium ST313 isolates. Methods Salmonella isolated from children with MSD or diarrhea-free controls were identified by classical clinical microbiology and serotyped using antisera and/or whole-genome sequence data. We evaluated antimicrobial susceptibility using the Kirby-Bauer disk-diffusion method. Salmonella Typhimurium sequence types were determined using multi-locus sequence typing, and whole-genome sequencing was performed to assess the phylogeny of ST313. Results Of 370 Salmonella-positive individuals, 190 (51.4%) were MSD cases and 180 (48.6%) were diarrhea-free controls. The most frequent Salmonella serovars identified were Salmonella Typhimurium, serogroup O:8 (C2-C3), serogroup O:6,7 (C1), Salmonella Paratyphi B Java, and serogroup O:4 (B). The prevalence of NTS was low but similar across sites, regardless of age, and was similar among both cases and controls except in Kenya, where Salmonella Typhimurium was more commonly associated with cases than controls. Phylogenetic analysis showed that these Salmonella Typhimurium isolates, all ST313, were highly genetically related to isolates from controls. Generally, Salmonella isolates from Asia were resistant to ciprofloxacin and ceftriaxone, but African isolates were susceptible to these antibiotics. Conclusions Our data confirm that NTS is prevalent, albeit at low levels, in Africa and South Asia. Our findings provide further evidence that multidrug-resistant Salmonella Typhimurium ST313 can be carried asymptomatically by humans in sub-Saharan Africa., In agreement with prior studies, we show that nontyphoidal Salmonella is prevalent in stool of children

Bloodstream infections caused by nontyphoidal Salmonella are a major public health concern in Africa, causing ~49,600 deaths every year. The most common Salmonella enterica pathovariant associated with invasive nontyphoidal Salmonella disease is Salmonella Typhimurium sequence type (ST)313. It has been proposed that antimicrobial resistance and genome degradation has contributed to the success of ST313 lineages in Africa, but the evolutionary trajectory of such changes was unclear. Here, to define the evolutionary dynamics of ST313, we sub-sampled from two comprehensive collections of Salmonella isolates from African patients with bloodstream infections, spanning 1966 to 2018. The resulting 680 genome sequences led to the discovery of a pan-susceptible ST313 lineage (ST313 L3), which emerged in Malawi in 2016 and is closely related to ST313 variants that cause gastrointestinal disease in the United Kingdom and Brazil. Genomic analysis revealed degradation events in important virulence genes in ST313 L3, which had not occurred in other ST313 lineages. Despite arising only recently in the clinic, ST313 L3 is a phylogenetic intermediate between ST313 L1 and L2, with a characteristic accessory genome. Our in-depth genotypic and phenotypic characterization identifies the crucial loss-of-function genetic events that occurred during the stepwise evolution of invasive S. Typhimurium across Africa., Stepwise evolution of invasive Salmonella Typhimurium in Africa is defined using genotypic and phenotypic analyses of isolates collected over a 50-yr period.

Invasive non-typhoidal Salmonella disease (iNTS) is a major cause of morbidity and mortality globally, particularly as a cause of bloodstream infection in children and immunocompromised adults in sub-Saharan Africa. Vaccines to prevent non-typhoidal Salmonella (NTS) would represent a valuable public health tool in this setting to avert cases and prevent expansion of antimicrobial resistance. Several NTS and combination typhoidal-NTS vaccine candidates are in early-stage development, although the pathway to licensure is unclear due to challenges in conducting large phase III field trials. Controlled human infection models (CHIM) present an opportunity to accelerate vaccine development for a range of enteric pathogens. Several recent typhoidal Salmonella CHIMs have been conducted safely and have played pivotal roles in progressing vaccine candidates to pre-qualification and licensure. The Challenge Non-Typhoidal Salmonella (CHANTS) consortium has been formed with funding from the Wellcome Trust, to deliver the first NTS CHIM, which can act as a platform for future vaccine evaluation. This paper reports the conclusions of a consultation group workshop convened with key stakeholders. The aims of this meeting were to: (1) define the rationale for an NTS CHIM (2) map the NTS vaccine pipeline (3) refine study design and (4) establish potential future use cases.

We report the complete genome sequencing and annotation of four Salmonella enterica serovar Enteritidis isolates, two that are representative of the Central/Eastern African clade (CP255 and D7795) and two from the Global Epidemic clade (A1636 and P125109)., We report the complete genome sequencing and annotation of four Salmonella enterica serovar Enteritidis isolates, two that are representative of the Central/Eastern African clade (CP255 and D7795) and two of the Global Epidemic clade (A1636 and P125109).

In recent years, novel lineages of invasive non-typhoidalSalmonella(iNTS) serovars Typhimurium and Enteritidis have been identified in patients with bloodstream infection in sub-Saharan Africa. Here, we isolated and characterised 32 phages capable of infectingS. Typhimurium andS. Enteritidis, from water sources in Malawi and the UK. The phages were classified in three major phylogenetic clusters that were geographically distributed. In terms of host range, Cluster 1 phages were able to infect all bacterial hosts tested, whereas Clusters 2 and 3 had a more restricted profile. Cluster 3 contained two sub-clusters, and 3.b contained the most novel isolates. This study represents the first exploration of the potential for phages to target the lineages ofSalmonellathat are responsible for bloodstream infections in sub-Saharan Africa.

Non-typhoidal Salmonella associated with multidrug resistance cause invasive disease in sub-Saharan African. Specific lineages of serovars S. Typhimurium and S. Enteritidis are implicated. We characterised the genomic diversity of 100 clinical Non-typhoidal Salmonella collected from 93 patients in 2001 from the eastern and 2006 to 2018 in the western regions of The Gambia respectively. Phenotypic susceptibility applied Kirby Baur disk diffusion and whole genome sequencing utilized Illumina platforms. The predominant serovars were S. Typhimurium ST19 (31/100) and S. Enteritidis ST11 (18/100) restricted to invasive disease with the notable absence of S. Typhimurium ST313. Phylogenetic analysis performed in the context of 495 African strains from the European Nucleotide Archive confirmed the presence of the S. Enteritidis virulent epidemic invasive multidrug resistant West African clade. Multidrug resistance including chloramphenicol and azithromycin has emerged among the West African S. Enteritidis clade 7/9 (78%) with potential for spread, thus having important implications for patient management warranting systematic surveillance and epidemiologic investigations to inform control.Data summarySequences are deposited in the NCBI sequence reads archive (SRA) under BioProject ID:PRJEB38968. The genomic assemblies are available for download from the European Nucleotide Archive (ENA): http://www.ebi.ac.uk/ena/data/view/. Accession numbers SAMEA6991082 to SAME6991180

Non-typhoidal Salmonella (NTS) usually cause gastroenteritis in humans, but in recent years NTS have begun to cause epidemics of bloodstream infections in Africa. Salmonella Enteritidis is the second most common serovar associated with this invasive form of NTS disease (iNTS) in Africa. To establish a systemic infection, Salmonella must survive and replicate within host cells, with macrophages being a primary target. Genomic characterisation of S. Enteritidis isolates from human bloodstream has identified two new clades that are unique to Africa and distinct from the Global Epidemic clade. The African S. Enteritidis clades exhibit genomic degradation, and possess a distinct prophage repertoire and are multi-drug resistant. However, little is known about the virulence factors that allow African S. Enteritidis to cause systemic infection in susceptible hosts. We screened libraries of random insertion mutants of African and Global S. Enteritidis by transposon insertion sequencing (TIS), and identified about 280 genes belonging to each clade that contribute to bacterial survival and replication in murine macrophages. The genes were associated with 5 pathogenicity-islands, or encoded the global regulators PhoPQ and OmpR-EnvZ. Experiments are ongoing to investigate the role in intra-macrophage replication of genes that are uniquely identified in African Salmonella. It is hoped that our findings will contribute to a greater understanding of African Salmonella infection biology, and that some of the virulence-associated genes could be potential targets for novel therapeutics.

Marine picocyanobacteria of the genus Synechococcus are ubiquitous phototrophs in oceanic systems. Consistent with these organisms occupying vast tracts of the nutrient impoverished ocean, most marine Synechococcus so far studied are monoploid, i.e., contain a single chromosome copy. The exception is the oligoploid strain Synechococcus sp. WH7803, which on average possesses around 4 chromosome copies. Here, we set out to understand the role of resource availability (through nutrient deplete growth) and physical stressors (UV, exposure to low and high temperature) in regulating ploidy level in this strain. Using qPCR to assay ploidy status we demonstrate the relative stability of chromosome copy number in Synechococcus sp. WH7803. Such robustness in maintaining an oligoploid status even under nutrient and physical stress is indicative of a fundamental role, perhaps facilitating recombination of damaged DNA regions as a result of prolonged exposure to oxidative stress, or allowing added flexibility in gene expression via possessing multiple alleles.

Salmonellosis is an endemic human infection, associated with both sporadic cases and outbreaks throughout Colombia. Typhimurium is the most common Colombian serovar ofSalmonella enterica, responsible for 32.5% of theSalmonellainfections. Whole genome sequencing (WGS) is being used increasingly in Europe and the USA to study the epidemiology ofSalmonella, but there has not yet been a WGS-based analysis ofSalmonellaassociated with bloodstream infection in Colombia. Here, we analysed 209 genome sequences of ColombianS. Typhimurium and monophasicS. 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 ofSalmonellain 2016, and resistance to nalidixic acid was associated with a plasmid-encodedqnrB19gene. Our findings suggest multiple transfers ofS. Typhimurium between Europe and Colombia.Author summaryThe large-scale genome sequencing ofSalmonellaTyphimurium and monophasicSalmonella4,[5],12:i:-involved bloodstream isolates from Colombia. The two serovars were responsible for about 1/3 ofSalmonellainfections in Colombia in the past 20 years. To identify the population structure we used Whole Genome Sequencing, performedin silicosequence 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 geneqnrB19. Our findings will inform the ongoing efforts to combat Salmonellosis by Colombian public health departments.

In recent years, novel lineages of invasive non-typhoidal Salmonella (iNTS) serovars Typhimurium and Enteritidis have been identified in patients with bloodstream infection in Sub-Saharan Africa. Here, we isolated and characterised 32 phages capable of infecting S. Typhimurium and S. Enteritidis, from water sources in Malawi and the UK. The phages were classified in three major phylogenetic clusters that were geographically distributed. In terms of host range, Cluster 1 phages were able to infect all bacterial hosts tested, whereas Clusters 2 and 3 had a more restricted profile. Cluster 3 contained two sub-clusters, and 3.b contained the most novel isolates. This study represents the first exploration of the potential for phages to target the lineages of Salmonella that are responsible for bloodstream infections in Sub-Saharan Africa.

Developments in transcriptomic technology and the availability of whole-genome-level expression profiles for many bacterial model organisms have accelerated the assignment of gene function. However, the deluge of transcriptomic data is making the analysis of gene expression a challenging task for biologists. Online resources for global bacterial gene expression analysis are not available for the majority of published data sets, impeding access and hindering data exploration. Here, we show the value of preexisting transcriptomic data sets for hypothesis generation. We describe the use of accessible online resources, such as SalComMac and SalComRegulon, to visualize and analyze expression profiles of coding genes and small RNAs. This approach arms a new generation of “gene detectives” with powerful new tools for understanding the transcriptional networks of Salmonella , a bacterium that has become an important model organism for the study of gene regulation. To demonstrate the value of integrating different online platforms, and to show the simplicity of the approach, we used well-characterized small RNAs that respond to envelope stress, oxidative stress, osmotic stress, or iron limitation as examples. We hope to provide impetus for the development of more online resources to allow the scientific community to work intuitively with transcriptomic data.

Hydrocarbons are ubiquitous in the ocean, where alkanes such as pentadecane and heptadecane can be found even in waters minimally polluted with crude oil. Populations of hydrocarbon-degrading bacteria, which are responsible for the turnover of these compounds, are also found throughout marine systems, including in unpolluted waters. These observations suggest the existence of an unknown and widespread source of hydrocarbons in the oceans. Here, we report that strains of the two most abundant marine cyanobacteria, Prochlorococcus and Synechococcus, produce and accumulate hydrocarbons, predominantly C15 and C17 alkanes, between 0.022 and 0.368% of dry cell weight. Based on global population sizes and turnover rates, we estimate that these species have the capacity to produce 2-540 pg alkanes per mL per day, which translates into a global ocean yield of ∼ 308-771 million tons of hydrocarbons annually. We also demonstrate that both obligate and facultative marine hydrocarbon-degrading bacteria can consume cyanobacterial alkanes, which likely prevents these hydrocarbons from accumulating in the environment. Our findings implicate cyanobacteria and hydrocarbon degraders as key players in a notable internal hydrocarbon cycle within the upper ocean, where alkanes are continually produced and subsequently consumed within days. Furthermore we show that cyanobacterial alkane production is likely sufficient to sustain populations of hydrocarbon-degrading bacteria, whose abundances can rapidly expand upon localized release of crude oil from natural seepage and human activities.