Comparative Genomics and Transduction Potential of Enterococcus faecalis Temperate Bacteriophages

Abstract

To determine the relative importance of temperate bacteriophage in the horizontal gene transfer of fitness and virulence determinants of Enterococcus faecalis, a panel of 47 bacteremia isolates were treated with the inducing agents mitomycin C, norfloxacin, and UV radiation. Thirty-four phages were purified from culture supernatants and discriminated using pulsed-field gel electrophoresis (PFGE) and restriction mapping. From these analyses the genomes of eight representative phages were pyrosequenced, revealing four distinct groups of phages. Three groups of phages, ⌽FL1 to 3, were found to be sequence related, with ⌽FL1A to C and ⌽FL2A and B sharing the greatest identity (87 to 88%), while ⌽FL3A and B share 37 to 41% identity with ⌽FL1 and 2. ⌽FL4A shares 3 to 12% identity with the phages ⌽FL1 to 3. The ⌽FL3A and B phages possess a high DNA sequence identity with the morphogenesis and lysis modules of Lactococcus lactis subsp. cremoris prophages. Homologs of the Streptococcus mitis platelet binding phage tail proteins, PblA and PblB, are encoded on each sequenced E. faecalis phage. Few other phage genes encoding potential virulence functions were identified, and there was little evidence of carriage of lysogenic conversion genes distal to endolysin, as has been observed with genomes of many temperate phages from the opportunist pathogens Staphylococcus aureus and Streptococcus pyogenes. E. faecalis JH2-2 lysogens were generated using the eight phages, and these were examined for their relative fitness in Galleria mellonella. Several lysogens exhibited different effects upon survival of G. mellonella compared to their isogenic parent. The eight phages were tested for their ability to package host DNA, and three were shown to be very effective for generalized transduction of naive host cells of the laboratory strains OG1RF and JH2-2.Enterococcus faecalis is a member of the natural flora of humans and colonizes the gastrointestinal and vaginal tracts and the oral cavity. In recent years it has emerged as an important opportunistic nosocomial pathogen and is a causative agent of bacteremia, infective endocarditis, and surgical wound and urinary tract infections. The accumulation of acquired antibiotic resistance determinants, in addition to its intrinsic resistance and tenacity, has given rise to the evolution of clinical isolates of E. faecalis that are therapeutically problematic (19). Greater notoriety was afforded to this species following the observed transfer of the conjugative transposon Tn1546 to Staphylococcus aureus, imparting vancomycin resistance (11). Subsequent analysis has revealed that multiple independent E. faecalis-dependent vanA transfers had occurred in the United States prior to 2007 (50). This places enterococci in an important and dynamic position within the health care system, warranting their increased study.The specific determinants that are proposed to contribute to the virulence of E. faecalis are not universally present, and expression of the cognate genes is variable (21, 37). For exa...

Genetic modifications to temperate Enterococcus faecalis phage ϕEf11 that abolish the establishment of lysogeny and sensitivity to repressor, and increase host range and productivity of lytic infection

“…In addition, the source of the wFL1C genes (i.e., the E. faecalis JH2-2 chromosome) was unexpected, as previous studies reported that this E. faecalis strain was susceptible to wFL1C infection, and in fact could form wFL1C lysogens following wFL1C infection, suggesting that this strain did not initially harbour a wFL1C prophage (Yasmin et al, 2010). This conundrum was solved when we used PCR to attempt to detect other regions of the wFL1C genome in JH2-2.…”

“…JH2-2 is a Fus r , Rif r mutant of a clinical E. faecalis isolate (Jacob & Hobbs, 1974) that was generously provided to us by Dr Nathan Shankar. In the course of this study, it was found that this strain harboured a wFL1C-type prophage element (Yasmin et al, 2010), indicating that this strain was a lysogen with a defective prophage. Other E. faecalis strains used in this study are listed in Table 1.…”

A composite bacteriophage alters colonization by an intestinal commensal bacterium

“…At least two of these prophage elements seem to encode cryptic or satellite phage genomes (10) that, by themselves, do not produce functional phage particles but may encode accessory components that aid in the lytic cycle of other integrated prophages found in the V583 chromosome. Several homologs of V583 prophage sequences have been identified in the genomes of other E. faecalis strains, and the total number of integrated prophages varies among strains (8,11). Although prophages are common in E. faecalis, their biological roles are poorly understood.…”

SalB Inactivation Modulates Culture Supernatant Exoproteins and Affects Autolysis and Viability in Enterococcus faecalis OG1RF

“…E. faecalis is the most virulent species in the genus and the cause of approximately 80% of all enterococcal infections, with virulence factors that include adhesins, proteases, and cytolysin (10,11,33). The accumulation of resistance genes is important medically, since intergeneric horizontal transfer of antibiotic resistance genes involving Staphylococcus aureus and Listeria species has been reported (3,16,20,30,38).…”