Genomic profiling of Escherichia coli isolates from bacteraemia patients: a 3-year cohort study of isolates collected at a Sydney teaching hospital

This study sought to assess the genetic variability of Escherichia coli isolated from bloodstream infections (BSIs) presenting at Concord Hospital, Sydney during 2013–2016. Whole-genome sequencing was used to characterize 81 E. coli isolates sourced from community-onset (CO) and hospital-onset (HO) BSIs. The cohort comprised 64 CO and 17 HO isolates, including 35 multidrug-resistant (MDR) isolates exhibiting phenotypic resistance to three or more antibiotic classes. Phylogenetic analysis identified two major ancestral clades. One was genetically diverse with 25 isolates distributed in 16 different sequence types (STs) representing phylogroups A, B1, B2, C and F, while the other comprised phylogroup B2 isolates in subclades representing the ST131, ST73 and ST95 lineages. Forty-seven isolates contained a class 1 integron, of which 14 carried blaCTX -M-gene. Isolates with a class 1 integron carried more antibiotic resistance genes than isolates without an integron and, in most instances, resistance genes were localized within complex resistance loci (CRL). Resistance to fluoroquinolones could be attributed to point mutations in chromosomal parC and gyrB genes and, in addition, two isolates carried a plasmid-associated qnrB4 gene. Co-resistance to fluoroquinolone and broad-spectrum beta-lactam antibiotics was associated with ST131 (HO and CO), ST38 (HO), ST393 (CO), ST2003 (CO) and ST8196 (CO and HO), a novel ST identified in this study. Notably, 10/81 (12.3 %) isolates with ST95 (5 isolates), ST131 (2 isolates), ST88 (2 isolates) and a ST540 likely carry IncFII–IncFIB plasmid replicons with a full spectrum of virulence genes consistent with the carriage of ColV-like plasmids. Our data indicate that IncF plasmids play an important role in shaping virulence and resistance gene carriage in BSI E. coli in Australia

Escherichia coli ST8196 is a novel, locally evolved, and extensively drug resistant pathogenic lineage within the ST131 clonal complex

The H30Rx subclade of Escherichia coli ST131 is a clinically important, globally dispersed pathogenic lineage that typically displays resistance to fluoroquinolones and extended spectrum β-lactams. Isolates EC233 and EC234, variants of ST131-H30Rx with a novel sequence type (ST) 8196, isolated from unrelated patients presenting with bacteraemia at a Sydney Hospital in 2014 are characterised here. EC233 and EC234 are phylogroup B2, serotype O25:H4A, and resistant to ampicillin, amoxicillin, cefoxitin, ceftazidime, ceftriaxone, ciprofloxacin, norfloxacin and gentamicin and are likely clonal. Both harbour an IncFII_2 plasmid (pSPRC_Ec234-FII) that carries most of the resistance genes on an IS26 associated translocatable unit, two small plasmids and a novel IncI1 plasmid (pSPRC_Ec234-I). SNP-based phylogenetic analysis of the core genome of representatives within the ST131 clonal complex places both isolates in a subclade with three clinical Australian ST131-H30Rx clade-C isolates. A MrBayes phylogeny analysis of EC233 and EC234 indicates ST8196 share a most recent common ancestor with ST131-H30Rx strain EC70 isolated from the same hospital in 2013. Our study identified genomic hallmarks that define the ST131-H30Rx subclade in the ST8196 isolates and highlights a need for unbiased genomic surveillance approaches to identify novel high-risk MDR E. coli pathogens that impact healthcare facilities

Phylogenomic analysis of a global collection of Escherichia coli ST38: evidence of interspecies and environmental transmission?

We performed a comprehensive phylogenomic analysis of 925 extraintestinal pathogenic Escherichia coli (ExPEC) ST38 genomes from 38 countries and diverse hosts and sources. The phylogeny resolved two broad clades: A (593 strains; 91% human) and B (332 isolates; 42% human), each with distinct ST38 clusters linked to the carriage of specific bla CTX-M alleles, often in association with other antibiotic resistance genes, class 1 integrons and specific plasmid replicon types. Co-carriage of fyuA and irp2 virulence genes, a reliable proxy for carriage of the Yersinia high-pathogenicity island, featured in 580 (62.7%) genomes. ST38 lineages carrying combinations of ExPEC and intestinal pathogenic Escherichia coli virulence factors were also identified. The F plasmid replicon was identified in 536 (58%) genomes, and 112 of these (21%) carry cjrABC-senB, a virulence operon frequently identified in pandemic ExPEC sequence types. Most (108; 96.4%) cjrABC-senB+ ST38 isolates were from human and other sources, except food animals, and were associated with F5:A-:B10 (41 isolates), F1:A2:B20 (20 isolates), and F24:A-:B1 (15 isolates) F replicon types. ST38 genomes that were inferred to carry a ColV-F virulence plasmid (69; 7.4%) were mostly from human (12; 17.4%), avian (26; 37.7%), or poultry (10; 6.9%) sources. We identified multiple examples of putative inter-host and host-environment transmission events, where genomes differed by <35 SNPs. This work emphasizes the importance of adopting a One Health approach for phylogenomic studies that seek to improve understanding of antimicrobial resistance and pathogen evolution

Genomic Epidemiology of Escherichia coli in Human Blood-Stream Infections

University of Technology Sydney. Faculty of Science.Blood-stream infections (BSI) are associated with high mortality and morbidity world-wide. The most common aetiological agent of these infections is . The rise of multidrug resistant (MDR) . is a major concern in clinical medicine and needs to be monitored for implementation of infection control strategies. The misuse of antimicrobials in clinical treatment, and as growth promotors in food-producing animals, is a key component of the rapid evolution of MDR . . MDR organisms spread antimicrobial resistance and virulence factors by lateral gene transfer via mobile genetic elements (MGEs). Despite this, there is limited knowledge of the origins and underlying mechanisms of BSI MDR . infections in Australia. We conducted genomic epidemiological analyses of . from human blood-cultures collected from a Sydney teaching hospital. The collection was dominated by . causing community onset BSI, with clones carrying a plethora of virulence and antimicrobial resistance genes. Clinical class 1 integrons associated with IS26 were identified in a number of sequence types (ST), indicating that they are important drivers of evolution and spread of clinically important antimicrobial resistance genes. We identified IncFII-IncFIB plasmids in the majority of our collection. A number of STs harboured ColV like IncFII plasmids, carry a number of important virulence traits, were also identified. We observed a novel ST, ST8196, that clustered with globally disseminated . ST131 isolates. We also identified an emerging ST, ST38, that exhibited resistance to a broad range of clinical beta-lactamases that have been identified in a number of pandemic . isolates. Our study provides evidence that . BSIs in Australia have a reservoir of antimicrobial resistance and virulence determinants that are potentially circulating not only in the community and hospital settings but across agricultural settings. These genes are not only present to clinical settings but can circulate within different ecosystems potentially via plasmids or other MGEs. Our findings also reveal that there are specific antimicrobial resistance genes such as ESBLs that are found in dominant . clones in both humans and food producing animals. This is likely to occur due the overuse of antimicrobials in human and animal settings that is driving the increase in antimicrobial resistance among bacteria that cause diseases. It is therefore essential to rigorously monitor these infections to help manage the global problem of antimicrobial resistance, and to reduce or prevent disease outbreaks

Immune response to COVID‐19 vaccination in patients with Waldenström macroglobulinaemia who pause their BTKi therapy

Abstract Patients with Waldenström macroglobulinaemia (WM) are at increased risk of severe COVID‐19 infection and have poor immune responses to COVID‐19 vaccination. This study assessed whether a closely monitored pause in Bruton's Tyrosine Kinase inhibitor (BTKi) therapy might result in an improved humoral response to a 3rd COVID‐19 vaccine dose. Improved response was observed in WM patients who paused their BTKi, compared to a group who did not pause their BTKi. However, the response was attenuated after BTKi recommencement. This data contributes to our understanding of vaccination strategies in this patient group and may help inform consensus approaches in the future

Patients with treated indolent lymphomas immunized with BNT162b2 have reduced anti‐spike neutralizing IgG to SARS‐CoV‐2 variants, but preserved antigen‐specific T cell responses

Patients with indolent lymphoma undertaking recurrent or continuous B cell suppression are at risk of severe COVID-19. Patients and healthy controls (HC; N=13) received 2 doses of BNT162b2: Follicular Lymphoma (FL; N=35) who were treatment naïve (TN; N=11) or received immunochemotherapy (ICT; N=23) and Waldenström's Macroglobulinemia (WM; N=37) including TN (N=9), ICT (N=14), or treated with Bruton's Tyrosine Kinase inhibitors (BTKi; N=12). Anti-spike IgG was determined by a high-sensitivity flow-cytometric assay, in addition to live-virus neutralization. Antigen-specific T cells were identified by co-expression of CD69/CD137 and CD25/CD134 on T cells. A subgroup (N=29) were assessed for third mRNA vaccine response, including omicron neutralization. One month after second BNT162b2, median anti-spike IgG mean fluorescence intensity (MFI) in FL ICT patients (9977) was 25-fold lower than TN (245898) and HC (228255, p=0.0002 for both). Anti-spike IgG correlated with lymphocyte count (r=0.63; p=0.002), and time from treatment, (r=0.56; p=0.007) on univariate analysis, but only with lymphocyte count on multivariate analysis (p=0.03). In the WM cohort, median anti-spike IgG MFI in BTKi patients (39039) was reduced compared to TN (220645, p=0.0008) and HC (p<0.0001). Anti-spike IgG correlated with neutralization of the delta variant (r=0.62, p<0.0001). Median neutralization titer for WM BTKi (0) was lower than HC (40, p<0.0001) for early-clade and delta. All cohorts had functional T cell responses. Median anti-spike IgG decreased 4-fold from second to third dose (p=0.004). Only 5/29 poor initial responders assessed after third vaccination demonstrated seroconversion and improvement in neutralization activity, including to the omicron variant. This article is protected by copyright. All rights reserved