Incidence of resistance to erythromycin at our institution reached 53% in 122 Staphylococcus aureus isolates obtained from patients with cystic fibrosis (CF) from 1997 to 1999. Macrolide-resistance genes were sought for in 20 erythromycin-resistant isolates from 9 patients with CF by use of polymerase chain reaction; 13 strains did not contain any known macrolide-resistance genes. Sequence of ribosomal genes rrl (23S rRNA), rplD (L4 protein), and rplV (L22 protein) revealed the presence of mutations in the target site of macrolides in 15 of the 20 isolates. A higher proportion of hypermutator strains was observed in a group of 89 CF staphylococcal isolates, compared with that in the 74 non-CF control isolates (13/89 vs. 1/74 with resistance to rifampin [P=.0045]; 9/89 vs. 1/74 with resistance to streptomycin [P=.04]). Various mutations or deletions of the mutator mutS gene were found not only in 5 of 11 hypermutable strains but also in 3 nonhypermutable strains harboring a large number of ribosomal mutations. The presence of a high proportion of hypermutable strains might explain the adaptation of certain strains in the patients, as well as the emergence of macrolide resistance as a result of antibiotic selective pressure in CF.

The enteropathogen Shigella flexneri invades epithelial cells, leading to inflammation and tissue destruction. We report that Shigella infection of epithelial cells induces an early genotoxic stress, but the resulting p53 response and cell death are impaired due to the bacterium's ability to promote p53 degradation, mainly through calpain protease activation. Calpain activation is promoted by the Shigella virulence effector VirA and dependent on calcium flux and the depletion of the endogenous calpain inhibitor calpastatin. Further, although VirA-induced calpain activity is critical for regulating cytoskeletal events driving bacterial uptake, calpain activation ultimately leads to necrotic cell death, thereby restricting Shigella intracellular growth. Therefore, calpains work at multiple steps in regulating Shigella pathogenesis by disrupting the p53-dependent DNA repair response early during infection and regulating both formation and ultimate death of the Shigella epithelial replicative niche.

The mutator phenotype has been linked in several bacterial genera to a defect in the methyl-mismatch repair system, in which the major components are MutS and MutL. This system is involved both in mismatch repair and in prevention of recombination between homeologous fragments in Escherichia coli and has been shown to play an important role in the adaptation of bacterial populations in changing and stressful environments. In this report we describe the molecular analysis of the mutS and mutL genes of Staphylococcus aureus. A genetic analysis of the mutSL region was performed in S. aureus RN4220. Reverse transcriptase PCR experiments confirmed the operon structure already reported in other gram-positive organisms. Insertional inactivation of mutS and mutL genes and complementation showed the role of both genes in hypermutability in this species. We also designed an in vitro model to study the role of MutS and MutL in homeologous recombination in S. aureus. For this purpose, we constructed a bank of S. aureus RN4220 and mutS and mutL mutants containing the integrative thermosensitive vector pBT1 in which fragments with various levels of identity (74% to 100%) to the S. aureus sodA gene were cloned. MutS and MutL proteins seemed to have a limited effect on the control of homeologous recombination. Sequence of mutS and mutL genes was analyzed in 11 hypermutable S. aureus clinical isolates. In four of five isolates with mutated or deleted mutS or mutL genes, a relationship between alterations and mutator phenotypes could be established by negative complementation of the mutS or mutL mutants.

Six strains of Staphylococcus aureus isolated from cystic fibrosis patients after treatment with azithromycin were cross-resistant to azithromycin and erythromycin. None of the isolates contained erm or msr(A) genes, but they all carried either A2058G/U or A2059G mutations within the rrl genes, with a majority of the rRNA copies bearing the mutation. One strain displayed an additional mutation in the rplV gene, encoding the L22 ribosomal protein