Escherichia coli O25b:H4-ST131
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ABSTRACT:
PROVIDER: PRJNA339173 | ENA |
REPOSITORIES: ENA
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Project description:A total of 1,021 extended-spectrum-?-lactamase-producing Escherichia coli (ESBLEC) isolates obtained in 2006 during a Spanish national survey conducted in 44 hospitals were analyzed for the presence of the O25b:H4-B2-ST131 (sequence type 131) clonal group. Overall, 195 (19%) O25b-ST131 isolates were detected, with prevalence rates ranging from 0% to 52% per hospital. Molecular characterization of 130 representative O25b-ST131 isolates showed that 96 (74%) were positive for CTX-M-15, 15 (12%) for CTX-M-14, 9 (7%) for SHV-12, 6 (5%) for CTX-M-9, 5 (4%) for CTX-M-32, and 1 (0.7%) each for CTX-M-3 and the new ESBL enzyme CTX-M-103. The 130 O25b-ST131 isolates exhibited relatively high virulence scores (mean, 14.4 virulence genes). Although the virulence profiles of the O25b-ST131 isolates were fairly homogeneous, they could be classified into four main virotypes based on the presence or absence of four distinctive virulence genes: virotypes A (22%) (afa FM955459 positive, iroN negative, ibeA negative, sat positive or negative), B (31%) (afa FM955459 negative, iroN positive, ibeA negative, sat positive or negative), C (32%) (afa FM955459 negative, iroN negative, ibeA negative, sat positive), and D (13%) (afa FM955459 negative, iroN positive or negative, ibeA positive, sat positive or negative). The four virotypes were also identified in other countries, with virotype C being overrepresented internationally. Correspondingly, an analysis of XbaI macrorestriction profiles revealed four major clusters, which were largely virotype specific. Certain epidemiological and clinical features corresponded with the virotype. Statistically significant virotype-specific associations included, for virotype B, older age and a lower frequency of infection (versus colonization), for virotype C, a higher frequency of infection, and for virotype D, younger age and community-acquired infections. In isolates of the O25b:H4-B2-ST131 clonal group, these findings uniquely define four main virotypes, which are internationally distributed, correspond with pulsed-field gel electrophoresis (PFGE) profiles, and exhibit distinctive clinical-epidemiological associations.
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Project description:The Escherichia coli lineage sequence type 131 (ST131)-O25b:H4 is a globally spread multidrug-resistant clone responsible for a great proportion of extraintestinal infections. Driven by the significant medical needs associated with this successful pathogenic lineage, we generated murine monoclonal antibodies (MAbs) against its lipopolysaccharide (LPS) O25b antigen in order to develop quick diagnostic tests. Murine monoclonal antibodies were generated by immunizing mice with whole killed nonencapsulated ST131-O25b E. coli cells and screening hybridoma supernatants for binding to purified LPS molecules obtained from an E. coli ST131-O25b clinical isolate. The MAbs selected for further study bound to the surface of live E. coli O25b strains irrespective of the capsular type expressed, while they did not bind to bacteria or purified LPS from other serotypes, including the related classical O25 antigen (O25a). Using these specific MAbs, we developed a latex bead-based agglutination assay that has greater specificity and is quicker and simpler than the currently available typing methods. The high specificities of these MAbs can be explained by the novel structure of the O25b repeating unit elucidated in this article. Based on comparative analysis by nuclear magnetic resonance (NMR) and mass spectrometry, the N-acetyl-fucose in the O25a O-antigen had been replaced by O-acetyl-rhamnose in the O25b repeating unit. The genetic determinants responsible for this structural variation were identified by aligning the corresponding genetic loci and were confirmed by trans-complementation of a rough mutant by the subserotype-specific fragments of the rfb operons.
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Project description:The Escherichia coli sequence type 131 (ST131)-O25b:H4 clone has spread worldwide and become responsible for a significant proportion of multidrug-resistant extraintestinal infections. We generated humanized monoclonal antibodies (MAbs) that target the lipopolysaccharide O25b antigen conserved within this lineage. These MAbs bound to the surface of live bacterial cells irrespective of the capsular type expressed. In a serum bactericidal assay in vitro, MAbs induced >95% bacterial killing in the presence of human serum as the complement source. Protective efficacy at low antibody doses was observed in a murine model of bacteremia. The mode of action in vivo was investigated by using aglycosylated derivatives of the protective MAbs. The significant binding to live E. coli cells and the in vitro and in vivo efficacy were corroborated in assays using bacteria grown in human serum to mimic relevant clinical conditions. Given the dry pipeline of novel antibiotics against multidrug-resistant Gram-negative pathogens, passive immunization with bactericidal antibodies offers a therapeutic alternative to control infections caused by E. coli ST131-O25b:H4.
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