Large-scale screening of a targeted Enterococcus faecalis mutant library identifies envelope fitness factors

Spread of antibiotic resistance among bacteria responsible for nosocomial and community-acquired infections urges for novel therapeutic or prophylactic targets and for innovative pathogen-specific antibacterial compounds. Major challenges are posed by opportunistic pathogens belonging to the low GC% gram-positive bacteria. Among those, Enterococcus faecalis is a leading cause of hospital-acquired infections associated with life-threatening issues and increased hospital costs. To better understand the molecular properties of enterococci that may be required for virulence, and that may explain the emergence of these bacteria in nosocomial infections, we performed the first large-scale functional analysis of E. faecalis V583, the first vancomycin-resistant isolate from a human bloodstream infection. E. faecalis V583 is within the high-risk clonal complex 2 group, which comprises mostly isolates derived from hospital infections worldwide. We conducted broad-range screenings of candidate genes likely involved in host adaptation (e.g., colonization and/or virulence). For this purpose, a library was constructed of targeted insertion mutations in 177 genes encoding putative surface or stress-response factors. Individual mutants were subsequently tested for their i) resistance to oxidative stress, ii) antibiotic resistance, iii) resistance to opsonophagocytosis, iv) adherence to the human colon carcinoma Caco-2 epithelial cells and v) virulence in a surrogate insect model. Our results identified a number of factors that are involved in the interaction between enterococci and their host environments. Their predicted functions highlight the importance of cell envelope glycopolymers in E. faecalis host adaptation. This study provides a valuable genetic database for understanding the steps leading E. faecalis to opportunistic virulence.

Multiresistant Salmonella enterica serovar 4,[5],12:i:- in Europe: a new pandemic strain?

A marked increase in the prevalence of S. enterica serovar 4,[5],12:i:- with resistance to ampicillin, streptomycin, sulphonamides and tetracyclines (R-type ASSuT) has been noted in food-borne infections and in pigs/pig meat in several European countries in the last ten years. One hundred and sixteen strains of S. enterica serovar 4,[5],12:i:- from humans, pigs and pig meat isolated in England and Wales, France, Germany, Italy, Poland, Spain and the Netherlands were further subtyped by phage typing, pulsed-field gel electrophoresis and multilocus variable number tandem repeat analysis to investigate the genetic relationship among strains. PCR was performed to identify the fljB flagellar gene and the genes encoding resistance to ampicillin, streptomycin, sulphonamides and tetracyclines. Class 1 and 2 integrase genes were also sought. Results indicate that genetically related serovar 4,[5],12:i:- strains of definitive phage types DT193 and DT120 with ampicillin, streptomycin, sulphonamide and tetracycline resistance encoded by blaTEM, strA-strB, sul2 and tet(B) have emerged in several European countries, with pigs the likely reservoir of infection. Control measures are urgently needed to reduce spread of infection to humans via the food chain and thereby prevent the possible pandemic spread of serovar 4,[5],12:i:- of R-type ASSuT as occurred with S. Typhimurium DT104 during the 1990s.