Molecular and evolutionary bases of within-patient genotypic and phenotypic diversity in Escherichia coli extraintestinal infections.

Although polymicrobial infections, caused by combinations of viruses, bacteria, fungi and parasites, are being recognised with increasing frequency, little is known about the occurrence of within-species diversity in bacterial infections and the molecular and evolutionary bases of this diversity. We used multiple approaches to study the genomic and phenotypic diversity among 226 Escherichia coli isolates from deep and closed visceral infections occurring in 19 patients. We observed genomic variability among isolates from the same site within 11 patients. This diversity was of two types, as patients were infected either by several distinct E. coli clones (4 patients) or by members of a single clone that exhibit micro-heterogeneity (11 patients); both types of diversity were present in 4 patients. A surprisingly wide continuum of antibiotic resistance, outer membrane permeability, growth rate, stress resistance, red dry and rough morphotype characteristics and virulence properties were present within the isolates of single clones in 8 of the 11 patients showing genomic micro-heterogeneity. Many of the observed phenotypic differences within clones affected the trade-off between self-preservation and nutritional competence (SPANC). We showed in 3 patients that this phenotypic variability was associated with distinct levels of RpoS in co-existing isolates. Genome mutational analysis and global proteomic comparisons in isolates from a patient revealed a star-like relationship of changes amongst clonally diverging isolates. A mathematical model demonstrated that multiple genotypes with distinct RpoS levels can co-exist as a result of the SPANC trade-off. In the cases involving infection by a single clone, we present several lines of evidence to suggest diversification during the infectious process rather than an infection by multiple isolates exhibiting a micro-heterogeneity. Our results suggest that bacteria are subject to trade-offs during an infectious process and that the observed diversity resembled results obtained in experimental evolution studies. Whatever the mechanisms leading to diversity, our results have strong medical implications in terms of the need for more extensive isolate testing before deciding on antibiotic therapies.

Coxiella burnetii endocarditis on bioprosthetic aortic valve, with peripheral arterial embolism.

Acute limb ischemia related to Coxiella burnetii endocarditis is rare. We report an original case of a 68-year-old man hospitalized for recurrent acute left limb ischemia in a context of atrial flutter, which revealed C. burnetii endocarditis. This case illustrates that even if embolic events are uncommon, septic embolisms must be systematically searched for in case of C. burnetii endocarditis. Conversely, extensive etiologic explorations must be performed in case of systemic embolism. New molecular techniques represent a major step forward in infective endocarditis diagnosis. Finally, diagnosis must be suspected in case of unexplained fever, inflammatory syndrome, or embolic event, especially in patients at risk. Conversely, in case of chronic Q fever, an immunodeficiency cause must be researched.

Identification by proteomic tool of atypical anti-liver/kidney microsome autoantibodies targets in de novo autoimmune hepatitis after liver transplantation.

De novo autoimmune hepatitis (AIH) occurs after liver transplantation for nonautoimmune disorders. Autoantibodies so-called atypical anti-liver/kidney microsome antibodies (LKMA) with an unusual liver/kidney cytoplasmic staining as judged by indirect immunofluorescence, can be detected in some patients' sera. Few studies investigated their molecular targets, and the aim of this work was to identify the atypical anti-LKMA targets by proteomic tool. This proteomic approach consisted of (a) two-dimensional gel electrophoresis of cytosolic and microsomal proteins obtained by differential centrifugations of rat liver and rat kidney, followed by (b) two-dimensional immunoblotting with sera of patients with de novo AIH (n = 8, including 2 with anti-LKMA antibodies) and then (c) identifications of interest spots performed by ion trap mass spectrometry. By this way several proteins at 25 kDa were unambiguously identified: isoforms of carbonic anhydrase III, members of different glutathione S-transferase (GST) families, and subunit beta1 of proteasome. This is the first report of proteasome and carbonic anhydrase III as autoantigens in de novo AIH. These results could lead to a better diagnosis of this disease using identified autoantigens in diagnostic tests, and strengthen proteomic approach as a new way of autoantigens investigation.

Genetic background of Escherichia coli and extended-spectrum beta-lactamase type.

To assess the implication of the genetic background of Escherichia coli strains in the emergence of extended-spectrum-Beta -lactamases (ESBL), 55 TEM-, 52 CTX-M-, and 22 SHV-type ESBL-producing clinical isolates involved in various extraintestinal infections or colonization were studied in terms of phylogenetic group, virulence factor (VF) content (pap, sfa/foc, hly, and aer genes), and fluoroquinolone resistance. A factorial analysis of correspondence showed that SHV type, and to a lesser extent TEM type, were preferentially observed in B2 phylogenetic group strains that exhibited numerous VFs but were fluoroquinolone-susceptible, whereas the newly emerged CTX-M type was associated with the D phylogenetic group strains that lacked VF but were fluoroquinolone-resistant. Thus, the emergence of ESBL-producing E. coli seems to be the result of complex interactions between the type of ESBL, genetic background of the strain, and selective pressures in ecologic niches.