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Open Access Research article

The hyl Efm gene in pHylEfm of Enterococcus faecium is not required in pathogenesis of murine peritonitis

Diana Panesso125, Maria C Montealegre128, Sandra Rincón125, Maria F Mojica128, Louis B Rice67, Kavindra V Singh13, Barbara E Murray134 and Cesar A Arias125*

Author Affiliations

1 Department of Internal Medicine, Division of Infectious Diseases, Center for the Study of Emerging and Reemerging Pathogens, Houston, TX, USA

2 Laboratory for Antimicrobial Research, University of Texas Medical School at Houston, Houston, TX, USA

3 Laboratory of Enterococcal Research, University of Texas Medical School at Houston, Houston, TX, USA

4 Department of Microbiology and Molecular Genetics, University of Texas Medical School at Houston, Houston, TX, USA

5 Molecular Genetics and Antimicrobial Resistance Unit, Universidad El Bosque, Bogotá, Colombia

6 Medical and Research Services, Louis Stokes Cleveland Department of Veterans Medical Center, Cleveland, OH, USA

7 Department of Medicine, Case Western Reserve University, School of Medicine, Cleveland, OH, USA

8 Centro Internacional de Entrenamiento e Investigaciones Médicas (CIDEIM), Cali, Colombia

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BMC Microbiology 2011, 11:20  doi:10.1186/1471-2180-11-20

Published: 25 January 2011

Abstract

Background

Plasmids containing hylEfm (pHylEfm) were previously shown to increase gastrointestinal colonization and lethality of Enterococcus faecium in experimental peritonitis. The hylEfm gene, predicting a glycosyl hydrolase, has been considered as a virulence determinant of hospital-associated E. faecium, although its direct contribution to virulence has not been investigated. Here, we constructed mutants of the hylEfm-region and we evaluated their effect on virulence using a murine peritonitis model.

Results

Five mutants of the hylEfm-region of pHylEfmTX16 from the sequenced endocarditis strain (TX16 [DO]) were obtained using an adaptation of the PheS* system and were evaluated in a commensal strain TX1330RF to which pHylEfmTX16 was transferred by mating; these include i) deletion of hylEfm only; ii) deletion of the gene downstream of hylEfm (down) of unknown function; iii) deletion of hylEfm plus down; iv) deletion of hylEfm-down and two adjacent genes; and v) a 7,534 bp deletion including these four genes plus partial deletion of two others, with replacement by cat. The 7,534 bp deletion did not affect virulence of TX16 in peritonitis but, when pHylEfmTX16Δ7,534 was transferred to the TX1330RF background, the transconjugant was affected in in vitro growth versus TX1330RF(pHylEfmTX16) and was attenuated in virulence; however, neither hylEfm nor hylEfm-down restored wild type function. We did not observe any in vivo effect on virulence of the other deletions of the hylEfm-region

Conclusions

The four genes of the hylEfm region (including hylEfm) do not mediate the increased virulence conferred by pHylEfmTX16 in murine peritonitis. The use of the markerless counterselection system PheS* should facilitate the genetic manipulation of E. faecium in the future.