Presence of phage-plasmids in multiple serovars of Salmonella enterica.

Evidence is accumulating in the literature that the horizontal spread of antimicrobial resistance (AMR) genes mediated by bacteriophages and bacteriophage-like plasmid (phage-plasmid) elements is much more common than previously envisioned. For instance, we recently identified and characterized a circular P1-like phage-plasmid harbouring a bla CTX-M-15 gene conferring extended-spectrum beta-lactamase (ESBL) resistance in Salmonella enterica serovar Typhi. As the prevalence and epidemiological relevance of such mechanisms has never been systematically assessed in Enterobacterales, in this study we carried out a follow-up retrospective analysis of UK Salmonella isolates previously sequenced as part of routine surveillance protocols between 2016 and 2021. Using a high-throughput bioinformatics pipeline we screened 47 784 isolates for the presence of the P1 lytic replication gene repL, identifying 226 positive isolates from 25 serovars and demonstrating that phage-plasmid elements are more frequent than previously thought. The affinity for phage-plasmids appears highly serovar-dependent, with several serovars being more likely hosts than others; most of the positive isolates (170/226) belonged to S. Typhimurium ST34 and ST19. The phage-plasmids ranged between 85.8 and 98.2 kb in size, with an average length of 92.1 kb; detailed analysis indicated a high amount of diversity in gene content and genomic architecture. In total, 132 phage-plasmids had the p0111 plasmid replication type, and 94 the IncY type; phylogenetic analysis indicated that both horizontal and vertical gene transmission mechanisms are likely to be involved in phage-plasmid propagation. Finally, phage-plasmids were present in isolates that were resistant and non-resistant to antimicrobials. In addition to providing a first comprehensive view of the presence of phage-plasmids in Salmonella, our work highlights the need for a better surveillance and understanding of phage-plasmids as AMR carriers, especially through their characterization with long-read sequencing.

Genomic epidemiology reveals geographical clustering of multidrug-resistant Escherichia coli ST131 associated with bacteraemia in Wales.

Antibiotic resistance is a significant global public health concern. Uropathogenic Escherichia coli sequence type (ST)131, a widely prevalent multidrug-resistant clone, is frequently associated with bacteraemia. This study investigates third-generation cephalosporin resistance in bloodstream infections caused by E. coli ST131. From 2013-2014 blood culture surveillance in Wales, 142 E. coli ST131 genomes were studied alongside global data. All three major ST131 clades were represented across Wales, with clade C/H30 predominant (n = 102/142, 71.8%). Consistent with global findings, Welsh strains of clade C/H30 contain ß-lactamase genes from the blaCTX-M-1 group (n = 65/102, 63.7%), which confer resistance to third-generation cephalosporins. Most Welsh clade C/H30 genomes belonged to sub-clade C2/H30Rx (58.3%). A Wales-specific sub-lineage, named GB-WLS.C2, diverged around 1996-2000. An introduction to North Wales around 2002 led to a localised cluster by 2009, depicting limited genomic diversity within North Wales. This investigation emphasises the value of genomic epidemiology, allowing the detection of genetically similar strains in local areas, enabling targeted and timely public health interventions.

Genomic epidemiology of the clinically dominant clonal complex 1 in the Listeria monocytogenes population in the UK.

Listeria monocytogenes is a food-borne pathogen, typically affecting the elderly, immunocompromised patients and pregnant women. The aim of this study was to determine the population structure of L. monocytogenes clonal complex 1 (CC1) in the UK and describe the genomic epidemiology of this clinically significant CC. We interrogated a working dataset of 4073 sequences of L. monocytogenes isolated between January 2015 and December 2020 from human clinical specimens, food and/or food-production environments. A minimum spanning tree was reconstructed to determine the population structure of L. monocytogenes in the UK. Subsequent analysis focused on L. monocytogenes CC1, as the cause of the highest proportion of invasive listeriosis in humans. Sequencing data was integrated with metadata on food and environmental isolates, and information from patient questionnaires, including age, sex and clinical outcomes. All isolates either belonged to lineage I (n=1299/4073, 32%) or lineage II (n=2774/4073, 68%), with clinical isolates from human cases more likely to belong to lineage I (n=546/928, 59%) and food isolates more likely to belong to lineage II (n=2352/3067, 77%). Of the four largest CCs, CC1 (n=237) had the highest proportion of isolates from human cases of disease (CC1 n=160/237, 67.5 %; CC121 n=13/843, 2 %; CC9 n=53/360, 15 %; CC2 n=69/339, 20%). Within CC1, most cases were female (n=95/160, 59%, P=0.01771) and the highest proportion of cases were in people >60 years old (39/95, 41%, P=1.314×10-6) with a high number of them aged 20-39 years old (n=35/95, 37%) most linked to pregnancy-related listeriosis (n=29/35, 83%). Most of the male cases were in men aged over 60 years old (40/65, 62%), and most of the fatal cases in both males and females were identified in this age group (42/55, 76%). Phylogenetic analysis revealed 23 5 SNP single linkage clusters comprising 80/237 (34 %) isolates with cluster sizes ranging from 2 to 19. Five 5 SNP clusters comprised isolates from human cases and an implicated food item. Expanding the analysis to 25 SNP single linkage clusters resolved an additional two clusters linking human cases to a potential food vehicle. Analysis of demographic and clinical outcome data identified CC1 as a clinically significant cause of invasive listeriosis in the elderly population and in women of child-bearing age. Phylogenetic analysis revealed the population structure of CC1 in the UK comprised small, sparsely populated genomic clusters. Only clusters containing isolates from an implicated food vehicle, or food processing or farming environments, were resolved, emphasizing the need for clinical, food and animal-health agencies to share sequencing data in real time, and the importance of a One Health approach to public-health surveillance of listeriosis.

Microevolution of Campylobacter jejuni during long-term infection in an immunocompromised host.

Campylobacteriosis typically manifests as a short-lived, self-limiting gastrointestinal infection in humans, however prolonged infection can be seen in cases with underlying immunodeficiency. Public Health England received 25 isolates of Campylobacter jejuni from an individual with combined variable immunodeficiency over a period of 15 years. All isolates were typed and archived at the time of receipt. Whole genome sequencing (WGS) and antimicrobial susceptibility testing were performed to examine the relatedness of the isolates and to investigate the changes in the genome that had taken place over the course of the infection. Genomic typing methods were compared to conventional phenotypic methods, and revealed that the infection was caused by a single, persistent strain of C. jejuni belonging to clonal complex ST-45, with evidence of adaptation and selection in the genome over the course of the infection. Genomic analysis of sequence variants associated with antimicrobial resistance identified the genetic background behind rRNA gene mutations causing variable levels of resistance to erythromycin. This application of WGS to examine a persistent case of campylobacteriosis provides insight into the mutations and selective pressures occurring over the course of an infection, some of which have important clinical relevance.

Antimicrobial resistance profiles of Shigella dysenteriae isolated from travellers returning to the UK, 2004-2017.

PURPOSE: Antimicrobial resistance (AMR) profiles of 754 strains of Shigella dysenteriae isolated between 2004 and 2017 from UK travellers reporting symptoms of gastrointestinal (GI) disease were reviewed to look for evidence of emerging AMR associated with travellers' diarrhoea. METHODOLOGY: A travel history was provided for 72.7 % (548/754) of cases, of which 90.9 % (498/548) reported travel outside the UK within 7 days of onset of symptoms, and 9.1 % (50/498) reported no travel in that time frame. During the course of this study, whole genome sequencing (WGS) was implemented for GI disease surveillance, and we compared phenotypic AMR profiles with those derived from WGS data (n=133).Results/Key findings. The phenotypic and genotypic AMR results correlated well, with 90.1 % (121/133) isolates having concordant results to 10 classes of antimicrobials. Resistance to the first-line drugs commonly used in the treatment of shigellosis was observed throughout the study (ampicillin, 54.1%; chloramphenicol, 33.7 %; sulphonamides, 76.0 %; trimethoprim, 80.0%). Between 2004 and 2017, resistance to all classes of antimicrobials (except the phenicols) increased. The proportion of isolates exhibiting reduced susceptibility to ciprofloxacin increased from 3.8 % in 2004 to 75.7 % in 2017, and this was significantly associated with cases reporting travel to Asia compared to Africa (P<0.001). Of the 201 sequenced isolates, 3.0 % (20/201) had either blaCTX-M-15 or blaCMY-4. CONCLUSIONS: Increasing MDR, along with resistance to the fluroquinolones and the third generation cephalosporins, in Shigella dysenteriae causing travellers' diarrhoea provides further evidence for the need to regulatethe use of antimicrobial agents and continuous monitoring of emerging AMR.

Species-wide whole genome sequencing reveals historical global spread and recent local persistence in Shigella flexneri.

Shigella flexneri is the most common cause of bacterial dysentery in low-income countries. Despite this, S. flexneri remains largely unexplored from a genomic standpoint and is still described using a vocabulary based on serotyping reactions developed over half-a-century ago. Here we combine whole genome sequencing with geographical and temporal data to examine the natural history of the species. Our analysis subdivides S. flexneri into seven phylogenetic groups (PGs); each containing two-or-more serotypes and characterised by distinct virulence gene complement and geographic range. Within the S. flexneri PGs we identify geographically restricted sub-lineages that appear to have persistently colonised regions for many decades to over 100 years. Although we found abundant evidence of antimicrobial resistance (AMR) determinant acquisition, our dataset shows no evidence of subsequent intercontinental spread of antimicrobial resistant strains. The pattern of colonisation and AMR gene acquisition suggest that S. flexneri has a distinct life-cycle involving local persistence.