Polymorphism, genetic exchange and intragenic recombination of the aureolysin gene among Staphylococcus aureus strains

<p>Abstract</p> <p>Background</p> <p><it>Staphylococcus aureus </it>expresses several proteases, which are thought to contribute to the virulence of this bacterium. Here we focus on aureolysin, the major thermolysin-like metalloprotease. Despite the importance of aureolysin in the physiology and pathogenesis of <it>S. aureus</it>, relatively little information was so far available concerning the <it>aur </it>gene diversity and mobility within and between the major subdivisions of the <it>S. aureus </it>population. Therefore, an epidemiologically and genetically diverse collection of <it>S. aureus </it>strains was used to determine the range of aureolysin (<it>aur</it>) gene polymorphism.</p> <p>Results</p> <p>Sequence analyses support the conclusion that the <it>aur </it>gene occurs in two distinct types of related sequences. The <it>aur </it>gene was much more polymorphic but, at the same time, showed higher purifying selection than genes utilized for multilocus sequence typing (MLST). Gene trees constructed from <it>aur </it>and concatenated MLST genes revealed several putative assortative recombination events (<it>i.e</it>. entire <it>aur </it>gene exchanges) between divergent lineages of <it>S. aureus</it>. Evidence for intragenic recombination events (<it>i.e</it>. exchanges of internal <it>aur </it>segments) across <it>aur </it>genes was also found. The biochemical properties and substrate specificity of the two types of aureolysin purified to homogeneity were studied, revealing minor differences in their affinity to low molecular weight synthetic substrates.</p> <p>Conclusion</p> <p>Although numerous nucleotide differences were identified between the <it>aur </it>alleles studied, our findings showed that a strong purifying selection is acting on the <it>aur </it>gene. Moreover, our study distinguishes between homologous exchanges of the entire <it>aur </it>gene (assortative recombination) between divergent <it>S. aureus </it>lineages and recombination events within <it>aur </it>genes.</p

Mutant Prevention Concentrations of Four Carbapenems against Gram-Negative Rods▿ †

We tested the propensities of four carbapenems to select for resistant Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii mutants by determining the mutant prevention concentrations (MPCs) for 100 clinical strains with various ß-lactam phenotypes. Among the members of the Enterobacteriaceae family and A. baumannii strains, the MPC/MIC ratios were mostly 2 to 4. In contrast, for P. aeruginosa the MPC/MIC ratios were 4 to ≥16. The MPC/MIC ratios for β-lactamase-positive K. pneumoniae and E. coli isolates were much higher (range, 4 to >16 μg/ml) than those for ß-lactamase-negative strains

Binding of Faropenem and Other β-Lactam Agents to Penicillin-Binding Proteins of Pneumococci with Various β-Lactam Susceptibilities▿

Faropenem demonstrated low MICs (≤1 μg/ml) for all penicillin-susceptible and nonsusceptible pneumococci and exhibited very strong abilities to bind to Streptococcus pneumoniae penicillin-binding proteins (PBPs), except for PBP2X. The lower faropenem affinity for PBP2X did not affect MICs for any strains tested, and only imipenem had lower MICs, with much lower binding affinities for all PBPs tested, than faropenem

Affinity of Ceftaroline and Other β-Lactams for Penicillin-Binding Proteins from Staphylococcus aureus and Streptococcus pneumoniae▿

We compared the affinities of ceftaroline for all penicillin-binding proteins (PBPs) with those of ceftriaxone and cefotaxime in 6 Staphylococcus aureus and 7 Streptococcus pneumoniae isolates with various resistance phenotypes. Ceftaroline MICs were ≤1 μg/ml against all S. aureus isolates and were ≤0.25 μg/ml for 4 of 7 isolates of S. pneumoniae. Ceftaroline affinities for penicillin-susceptible S. pneumoniae strains were in the order PBP2X and -3 > PBP1A, -1B, and -2A > PBP2B, and ceftaroline had ≥4-fold higher 50% inhibitory concentrations (IC50s) (0.1 to 4 μg/ml) for PBP2X, -2A, -2B, and -3 than those for the other cephalosporins tested. Among 3 penicillin-resistant S. pneumoniae strains, ceftaroline had a high affinity for PBP2X (IC50, 0.1 to 1 μg/ml), a primary target for cephalosporin PBP binding activity, and high affinities for PBP2B (IC50, 0.5 to 4 μg/ml) and PBP1A (IC50, 0.125 to 0.25 μg/ml) as well, both of which are also known as major targets for PBP binding activity of cephalosporins. Ceftaroline PBP affinities in methicillin-susceptible S. aureus strains were greater than or equal to those of the 3 other β-lactams tested. Ceftaroline bound to PBP2a in methicillin-resistant S. aureus (IC50, 0.01 to 1 μg/ml) with up to 256-fold-higher affinity than those of other agents. Ceftaroline demonstrated very good PBP affinity against all S. aureus and S. pneumoniae strains tested, including resistant isolates

Comparative Antipneumococcal Activities of Sulopenem and Other Drugs▿

For 297 penicillin-susceptible, -intermediate, and -resistant pneumococcal strains, the sulopenem MIC50s were 0.008, 0.06, and 0.25, respectively, and the sulopenem MIC90s were 0.016, 0.25, and 0.5 μg/ml, respectively. The MIC50s of amoxicillin for the corresponding strains were 0.03, 0.25, and 2.0 μg/ml, respectively, and the MIC90s were 0.03, 1.0, and 8.0 μg/ml, respectively. The combination of amoxicillin and clavulanate gave MICs similar to those obtained with amoxicillin alone. The sulopenem MICs were similar to those of imipenem and meropenem. The MICs of ß-lactams increased with those of penicillin G, and among the quinolones tested, moxifloxacin had the lowest MICs. Additionally, 45 strains of drug-resistant type 19A pneumococci were tested by agar dilution and gave sulopenem MIC50s and MIC90s of 1.0 and 2.0 μg/ml, respectively. Both sulopenem and amoxicillin (with and without clavulanate) were bactericidal against all 12 strains tested at 2× MIC after 24 h. Thirty-one strains from 10 countries with various penicillin, amoxicillin, and carbapenems MICs, including those with the highest sulopenem MICs, were selected for sequencing analysis of the pbp1a, pbp2x, and pbp2b regions encoding the transpeptidase active site and MurM. We did not find any correlations between specific penicillin-binding protein-MurM patterns and changes in the MICs

Capability of 11 Antipneumococcal Antibiotics To Select for Resistance by Multistep and Single-Step Methodologies▿ †

Testing of 12 pneumococcal strains with differing resistotypes [including tet(M) positive] showed that tigecycline, amoxicillin-clavulanate, imipenem, and ceftriaxone did not select for resistant clones after 50 sequential subcultures. By comparison, azithromycin, clarithromycin, clindamycin, telithromycin, levofloxacin, moxifloxacin, and gemifloxacin did show resistant clones. Tigecycline also yielded a low frequency of resistance in single-step tests compared to all β-lactams, macrolides/ketolides, and quinolones tested

Comparative Study of the Mutant Prevention Concentrations of Moxifloxacin, Levofloxacin, and Gemifloxacin against Pneumococci▿ †

We tested the propensity of three quinolones to select for resistant Streptococcus pneumoniae mutants by determining the mutant prevention concentration (MPC) against 100 clinical strains, some of which harbored mutations in type II topoisomerases. Compared with levofloxacin and gemifloxacin, moxifloxacin had the lowest number of strains with MPCs above the susceptibility breakpoint (P < 0.001), thus representing a lower selective pressure for proliferation of resistant mutants. Only moxifloxacin gave a 50% MPC (MPC50) value (1 μg/ml) within the susceptible range