rRNA Operon Copy Number Can Explain the Distinct Epidemiology of Hospital-Associated Methicillin-Resistant Staphylococcus aureus.

The distinct epidemiology of original hospital-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) and early community-associated MRSA (CA-MRSA) is largely unexplained. S. aureus carries either five or six rRNA operon copies. Evidence is provided for a scenario in which MRSA has adapted to the hospital environment by rRNA operon loss (six to five copies) due to antibiotic pressure. Early CA-MRSA, in contrast, results from wild-type methicillin-susceptible S. aureus (MSSA) that acquired mecA without loss of an rRNA operon. Of the HA-MRSA isolates (n = 77), 67.5% had five rRNA operon copies, compared to 23.2% of the CA-MRSA isolates (n = 69) and 7.7% of MSSA isolates (n = 195) (P < 0.001). In addition, 105 MSSA isolates from cystic fibrosis patients were tested, because these patients are repeatedly treated with antibiotics; 32.4% of these isolates had five rRNA operon copies. For all subsets, a correlation between resistance profile and rRNA copy number was found. Furthermore, we showed that in vitro antibiotic pressure may result in rRNA operon copy loss. We also showed that without antibiotic pressure, S. aureus isolates containing six rRNA copies are more fit than isolates with five copies. We conclude that HA-MRSA and cystic fibrosis isolates most likely have adapted to an environment with high antibiotic pressure by the loss of an rRNA operon copy. This loss has facilitated resistance development, which promoted survival in these niches. However, strain fitness decreased, which explains their lack of success in the community. In contrast, CA-MRSA isolates retained six rRNA operon copies, rendering them fitter and thereby able to survive and spread in the community.

Surveillance study of the susceptibility of Haemophilus influenzae to various antibacterial agents in Europe and Canada.

BACKGROUND: Haemophilus influenzae is a major respiratory tract pathogen that is becoming increasingly resistant to beta-lactam antibiotics. MATERIALS AND METHODS: Using a microdilution method performed to Clinical and Laboratory Standards Institute (CLSI) guidelines, we determined the minimal inhibitory concentrations (MICs) of various antibacterial agents against 536 isolates of H. influenzae. The isolates were obtained from patients with respiratory tract infections being treated in 18 European and two Canadian centres between 2006 and 2007. RESULTS: Levofloxacin, moxifloxacin, cefixime and cefpodoxime with MIC(90) values of < or = 0.03, < or = 0.03, 0.03 and 0.06 g/mL, respectively, were the four most active agents tested. Overall, amoxicillin resistance was observed in 25.0% of the strains, but was generally reversed with the addition of clavulanic acid. In 73 strains (13.6%) resistance was due to beta-lactamase (BL) production while the remainder (n = 61; 11.4%) were BL-negative, amoxicillin-resistant (BLNAR) strains. Comparison of penicillin binding protein 3B sequences in BLNAR isolates revealed that only mutations at amino acids 502 (alanine [Ala] --> threonine [Thr]/valine [Val]) and 526 (asparagine [Asn] --> lysine [Lys]) were significantly associated with amoxicillin resistance among European H. influenzae isolates (p < 0.0001 for both). CONCLUSIONS: This surveillance study highlights an increased prevalence of amoxicillin-resistant strains of H. influenzae compared with a previous study that we performed in 2004/2005. The third-generation cephalosporins cefixime and cefpodoxime, as well as amoxicillin plus clavulanic acid, continue to be very active against both BL-positive and BLNAR strains of H. influenzae, and thus remain useful treatment options for patients with respiratory tract infections.

In vitro activity of AR-709 against Streptococcus pneumoniae.

We investigated the in vitro activity of AR-709, a novel diaminopyrimidine antibiotic currently in development for treatment of community-acquired upper and lower respiratory tract infections, against 151 Streptococcus pneumoniae strains from various European countries. AR-709 showed excellent activity against both drug-susceptible and multidrug-resistant pneumococci.

In vitro activity of telavancin against gram-positive clinical isolates recently obtained in Europe.

The in vitro activity of telavancin was tested against 620 gram-positive isolates. For staphylococci, MICs at which 50 and 90% of isolates were inhibited (MIC(50) and MIC(90)) were both 0.25 microg/ml, irrespective of methicillin resistance. MIC(50) and MIC(90) were 0.25 and 0.5 microg/ml for vancomycin-susceptible enterococci and 1 and 2 microg/ml for vancomycin-resistant enterococci, respectively. Streptococcus pneumoniae, group A and B beta-hemolytic streptococci, and viridans streptococci were inhibited by < or =0.12 microg/ml.

Dependence of correlations between antibody titres and opsonophagocytosis on pneumococcal serotype and patient morbidity in pre- and post-pneumococcal vaccination states.

Pre- vs. post-vaccination changes in correlations between IgG concentrations (ELISA titres) and opsonophagocytic activity (OPA) against Streptococcus pneumoniae serotypes 6B, 14 and 23F induced by the 23-valent polysaccharide vaccine were studied in paired serum samples received from elderly individuals, haemodialysed patients and kidney transplant recipients by the Spanish Pneumococcal Reference Laboratory. The pre- and post-vaccination parameters considered were: ELISA and OPA titres and the percentage of subjects with post-vaccination OPA values above the cut-off levels; the correlations between OPA and ELISA (Spearman correlation coefficient, r); and the amount of IgG needed to obtain OPA (beta coefficient). Non-significant pre-vaccination correlations between OPA and ELISA were found. Vaccination increased the correlation coefficient between OPA and ELISA to a statistically significant level for serotypes 6B, 14 and 23F in samples from haemodialysed patients, for serotypes 14 and 23F in samples from elderly individuals, and for none of the serotypes in samples from transplant recipients. In all cases, except for serotype 23 in transplant recipients, vaccination increased the beta coefficient, indicating that lower amounts of IgG were needed to obtain high OPA titres. A globally lower response was obtained for serotype 23 and/or transplant recipients.

Longitudinal European surveillance study of antibiotic resistance of Haemophilus influenzae.

OBJECTIVES: We assessed the current resistance rates of Haemophilus influenzae against beta-lactams and other agents in Europe and compared the results with those of our previously performed surveillance study. METHODS: MICs of the antibiotics were determined using broth microdilution. The penicillin-binding domain of PBP3 of beta-lactamase (BL)-negative, amoxicillin-resistant (BLNAR) isolates was sequenced. RESULTS: The percentage of BL-positive and BLNAR strains ranged from 0% to 17.6% and 0% to 33.9%, respectively. Compared with 1997/98 and 2002/03, the overall percentage of strains non-susceptible to amoxicillin decreased from 19.8% and 23.3%, respectively, to 16.4% in 2004/05. The percentage of BL-producing strains decreased from 11.0% and 13.7%, respectively, to 7.6%, whereas the number of BLNAR strains remained stable (8.8% and 9.6%, respectively, versus 8.8% in 2004/05). Comparison of penicillin binding protein (PBP) 3B gene sequences between BLNAR and susceptible strains revealed novel amino acid mutations. CONCLUSIONS: In spite of large inter-regional differences, the overall resistance of H. influenzae to amoxicillin in Europe seems to decline due to a decreasing number of BL-producing strains, whereas the overall percentage of BLNAR strains seems relatively constant.

Bacterial resistance: a sensitive issue complexity of the challenge and containment strategy in Europe.

The development of antimicrobial agents has been a key achievement of modern medicine. However, their overuse has led to an increasing incidence of infections due to antibiotic-resistant microorganisms. Quantitative figures on the current economic and health impact of antimicrobial resistance are scant, but it is clearly a growing challenge that requires timely action. That action should be at the educational, ethical, economic and political level. An important first step would be to increase public awareness and willingness to take the necessary measures to curb resistance. Hence, studies are needed that would provide solid, quantitative data on the societal impact of antibiotic resistance. This review discusses the complexity of resistance, identifies its main drivers and proposes measures to contain it on a European scale.

Novel mobile variants of staphylococcal cassette chromosome mec in Staphylococcus aureus.

Staphylococcus aureus staphylococcal cassette chromosome mec type IV (SSCmec IV) is associated with virulent community-acquired methicillin-resistant Staphylococcus aureus (MRSA) and frequent horizontal transfer among staphylococci. To gain insight into the mechanism of transfer, we studied the ccrA/B type 2 recombinase-mediated excision of SCCmec IV (n = 5 strains) and SCCmec II (n = 2). In SCCmec IV- but not SCCmec II-containing strains, spontaneous excision of the cassette was observed. Introduction of ccrA/B type 2 recombinase genes under control of an S. aureus bacterial phage promoter in the different strains yielded excision of SCCmec II and multiple excision variants of SCCmec IV. Sequencing of the alternatively excised products in SCCmec IV strains identified a 100-bp shortened SCCmec' variant and a 5,877-bp, conserved SCC-like element that lacks mecA and ccrA/B recombinases. Excision of the SCC-like element in wild-type S. aureus was dependent on the presence of SCCmec. The element could be excised separately or as part of a novel composite cassette together with SCCmec. The relative abundance of and variety in SCCmec IV excisions may contribute to the frequency of horizontal transfer and genetic plasticity in SCCmec IV MRSA strains.

Priorities for antibiotic resistance surveillance in Europe.

Antibiotic resistance is an increasing global problem. Surveillance studies are needed to monitor resistance development, to guide local empirical therapy, and to implement timely and adequate countermeasures. To achieve this, surveillance studies must have standardised methodologies, be longitudinal, and cover a sufficiently large and representative population. However, many fall short of these requirements that define good surveillance studies. Moreover, current efforts are dispersed among many, mostly small, initiatives with different objectives. These studies must be tailored to the various reservoirs of antibiotic-resistant bacteria, such as hospitalised patients, nursing homes, the community, animals and food. Two studies that could serve as examples of tailored programmes are the European Antimicrobial Resistance Surveillance System (EARSS), which collects resistance data during the diagnosis of hospitalised patients, and the DANMAP programme, which collects data in the veterinary sector. As already noted by the WHO, genetic studies that include both the typing of isolates and the characterisation of resistance determinants are necessary to understand fully the spread and development of antibiotic resistance.

Short-chain oligosaccharide protein conjugates as experimental pneumococcal vaccines.

Streptococcus pneumoniae remains a major cause of acute respiratory infections worldwide and is responsible for approximately 1 million childhood deaths each year. Despite the widespread use of antibiotics, the mortality and morbidity of pneumococcal disease remains high. Therefore, effective vaccines to prevent pneumococcal disease are needed. Bacterial vaccine development in general follows a similar track starting from large, rather crude vaccines (whole live, attenuated pathogen) towards smaller, better defined subunit vaccines. Ultimately, this track leads to the development and evaluation of minimal, highly defined subunit vaccines, based on a collection of single protective epitopes. This mini-review deals with capsular saccharide based vaccines. After a short overview of the development of pneumococcal vaccines from the 23 - valent polysaccharide vaccines to polysaccharide-protein conjugate vaccines, it focuses on the vaccine potential of synthetic oligosaccharides, conjugated to carrier proteins.

Hydrogen peroxide-mediated killing of Caenorhabditis elegans by Streptococcus pyogenes.

Caenorhabditis elegans is currently introduced as a new, facile, and cheap model organism to study the pathogenesis of gram-negative bacteria such as Pseudomonas aeruginosa and Salmonella enterica serovar Typhimurium. The mechanisms of killing involve either diffusible exotoxins or infection-like processes. Recently, it was shown that also some gram-positive bacteria kill C. elegans, although the precise mechanisms of killing remained open. We examined C. elegans as a pathogenesis model for the gram-positive bacterium Streptococcus pyogenes, a major human pathogen capable of causing a wide spectrum of diseases. We demonstrate that S. pyogenes kills C. elegans, both on solid and in liquid medium. Unlike P. aeruginosa and S. enterica serovar Typhimurium, the killing by S. pyogenes is solely mediated by hydrogen peroxide. Killing required live streptococci; the killing capacity depends on the amount of hydrogen peroxide produced, and killing can be inhibited by catalase. Major exotoxins of S. pyogenes are not involved in the killing process as confirmed by using specific toxin inhibitors and knockout mutants. Moreover, no accumulation of S. pyogenes in C. elegans is observed, which excludes the involvement of infection-like processes. Preliminary results show that S. pneumoniae can also kill C. elegans by hydrogen peroxide production. Hydrogen peroxide-mediated killing might represent a common mechanism by which gram-positive, catalase-negative pathogens kill C. elegans.