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.

Dissemination of antimicrobial resistance (AMR) genes by horizontal gene transfer (HGT) mediated through plasmids is a major global concern. Genomic epidemiology studies have shown varying success of different AMR plasmids during outbreaks, but the underlying reasons for these differences are unclear. Here, we investigated two Shigella plasmids (pKSR100 and pAPR100) that circulated in the same transmission network but had starkly contrasting epidemiological outcomes to identify plasmid features that may have contributed to the differences. We used plasmid comparative genomics to reveal divergence between the two plasmids in genes encoding AMR, SOS response alleviation and conjugation. Experimental analyses revealed that these genomic differences corresponded with reduced conjugation efficiencies for the epidemiologically successful pKSR100, but more extensive AMR, reduced fitness costs, and a reduced SOS response in the presence of antimicrobials, compared with the less successful pAPR100. The discrepant phenotypes between the two plasmids are consistent with the hypothesis that plasmid-associated phenotypes contribute to determining the epidemiological outcome of AMR HGT and suggest that phenotypes relevant in responding to antimicrobial pressure and fitness impact may be more important than those around conjugation in this setting. Plasmid phenotypes could thus be valuable tools in conjunction with genomic epidemiology for predicting AMR dissemination.

Shigella spp. are Gram-negative bacteria that are the etiological agent of shigellosis, the second most common cause of diarrheal illness among children under the age of five in low-income countries. In high-income countries, shigellosis is also a sexually transmissible disease among men who have sex with men.

Shigellosis is a diarrhoeal disease caused mainly by Shigella flexneri and Shigella sonnei. Infection from Shigella is thought to be largely self-limiting, with short- to medium- term and serotype-specific immunity provided following clearance. However, cases of men who have sex with men (MSM) associated shigellosis have been reported where Shigella of the same serotype were serially sampled from individuals between 1 to 1862 days apart, possibly due to persistent carriage or reinfection with the same serotype. Here, we investigate the accessory genome dynamics of MSM associated S. flexneri and S. sonnei isolates serially sampled from individual patients at various days apart. We find that pairs likely associated with persistent carriage infection and with smaller single nucleotide polymorphism (SNP) distance, demonstrated significantly less variation in accessory genome content than pairs likely associated with reinfection and with greater SNP-distance. We also observed evidence of antimicrobial resistance (AMR) acquisition during persistent Shigella infection, specifically the gain of extended spectrum beta-lactamase genes in two pairs associated with persistent carriage. Finally, we explored chromosomal structural variations and rearrangements in seven (5 chronic and 2 reinfection associated) pairs of S. flexneri 3a isolates from a MSM-associated epidemic sublineage, which revealed variations at several common regions across pairs. These variations were mediated by insertion sequence (IS) elements which facilitated plasticity of genetic material with a distinct predicted functional profile. This study provides insight on the variation of accessory genome dynamics and large structural genomic changes in Shigella during persistent infection.ImportanceShigella spp are Gram-negative bacteria that are the etiological agent of shigellosis, the second most common cause of diarrhoeal illness globally, particularly among children under the age of 5 in low-income countries. In high-income countries, an alternative transmission pathway of sexually transmissible disease among men who have sex with men (MSM) is emerging as the dominant presentation of the disease. Within MSM we have captured prolonged infection and/or recurrent infection with shigellae of the same serotype, challenging the belief that Shigella infection is short-lived, and confers homologous serotypic immunity. Using this recently-emerged transmission scenario we comprehensively characterise the genomic changes that occur over the course of individual infection with Shigella and uncover a distinct functional profile of variable genome regions in these globally important pathogens.

Increasing antimicrobial resistance and limited alternative treatments have led to fluoroquinolone-resistant Shigella strain inclusion on the WHO global priority pathogens list. In this study we characterized multiple Shigella isolates from Malawi with whole genome sequence analysis, identifying the acquirable fluoroquinolone resistance determinant qnrS1.