A penicillin- and metronidazole-resistant Clostridium botulinum strain responsible for an infant botulism case.

The clinical course of a case of infant botulism was characterized by several relapses despite therapy with amoxicillin and metronidazole. Botulism was confirmed by identification of botulinum toxin and Clostridium botulinum in stools. A C.

Structural insights into the loss of penicillinase and the gain of ceftazidimase activities by OXA-145 β-lactamase in Pseudomonas aeruginosa.

Objectives: We previously described extended-spectrum oxacillinase OXA-145 from Pseudomonas aeruginosa, which differs from narrow-spectrum OXA-35 by loss of Leu-155. The deletion results in loss of benzylpenicillin hydrolysis and acquisition of activity against ceftazidime. We report the crystal structure of OXA-145 and provide the basis of its switch in substrate specificity.

Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases.

Unlabelled: Vancomycin resistance in Gram-positive bacteria results from the replacement of the D-alanyl-D-alanine target of peptidoglycan precursors with D-alanyl-D-lactate or D-alanyl-D-serine (D-Ala-D-Ser), to which vancomycin has low binding affinity. VanT is one of the proteins required for the production of D-Ala-D-Ser-terminating precursors by converting L-Ser to D-Ser. VanT is composed of two domains, an N-terminal membrane-bound domain, likely involved in L-Ser uptake, and a C-terminal cytoplasmic catalytic domain which is related to bacterial alanine racemases.

Antimicrobial Activity of Plectasin NZ2114 in Combination with Cell Wall Targeting Antibiotics Against VanA-Type Enterococcus faecalis.

Antimicrobial peptide plectasin targeting bacterial cell wall precursor Lipid II has been reported to be active against benzylpenicillin-resistant Streptococcus pneumoniae but less potent against vancomycin-resistant enterococci than their susceptible counterparts. The aim of this work was to test plectasin NZ2114 in combination with cell wall targeting antibiotics on vancomycin-resistant Enterococcus faecalis. The activity of antibiotic combinations was evaluated against VanA-type vancomycin-resistant E.

Structural basis for the evolution of vancomycin resistance D,D-peptidases.

Vancomycin resistance in Gram-positive bacteria is due to production of cell-wall precursors ending in D-Ala-D-Lac or D-Ala-D-Ser, to which vancomycin exhibits low binding affinities, and to the elimination of the high-affinity precursors ending in D-Ala-D-Ala. Depletion of the susceptible high-affinity precursors is catalyzed by the zinc-dependent D,D-peptidases VanX and VanY acting on dipeptide (D-Ala-D-Ala) or pentapeptide (UDP-MurNac-L-Ala-D-Glu-L-Lys-D-Ala-D-Ala), respectively. Some of the vancomycin resistance operons encode VanXY D,D-carboxypeptidase, which hydrolyzes both di- and pentapeptide.

Functional motions modulating VanA ligand binding unraveled by self-organizing maps.

The VanA D-Ala:D-Lac ligase is a key enzyme in the emergence of high level resistance to vancomycin in Enterococcus species and methicillin-resistant Staphylococcus aureus. It catalyzes the formation of D-Ala-D-Lac instead of the vancomycin target, D-Ala-D-Ala, leading to the production of modified, low vancomycin binding affinity peptidoglycan precursors. Therefore, VanA appears as an attractive target for the design of new antibacterials to overcome resistance.

The functional vanGCd cluster of Clostridium difficile does not confer vancomycin resistance.

vanGCd, a cryptic gene cluster highly homologous to the vanG gene cluster of Enterococcus faecalis is largely spread in Clostridium difficile. Since emergence of vancomycin resistance would have dramatic clinical consequences, we have evaluated the capacity of the vanGCd cluster to confer resistance. We showed that expression of vanGCd is inducible by vancomycin and that VanGCd , VanXYCd and VanTCd are functional, exhibiting D-Ala : D-Ser ligase, D,D-dipeptidase and D-Ser racemase activities respectively.

Structural and functional characterization of VanG D-Ala:D-Ser ligase associated with vancomycin resistance in Enterococcus faecalis.

d-Alanyl:d-lactate (d-Ala:d-Lac) and d-alanyl:d-serine ligases are key enzymes in vancomycin resistance of Gram-positive cocci. They catalyze a critical step in the synthesis of modified peptidoglycan precursors that are low binding affinity targets for vancomycin. The structure of the d-Ala:d-Lac ligase VanA led to the understanding of the molecular basis for its specificity, but that of d-Ala:d-Ser ligases had not been determined.

Ceftazidime-hydrolysing β-lactamase OXA-145 with impaired hydrolysis of penicillins in Pseudomonas aeruginosa.

Objectives: To describe a novel extended-spectrum oxacillinase, named OXA-145, differing from narrow-spectrum OXA-35 (from the OXA-10 group) by deletion of residue Leu-165. The genetic environment of bla(OXA-145) and the biochemical properties of OXA-145 are reported. We also assessed the impact of the Leu-165 deletion on the hydrolysis spectrum of the ancestor OXA-10.

Molecular basis of vancomycin dependence in VanA-type Staphylococcus aureus VRSA-9.

The vancomycin-resistant Staphylococcus aureus VRSA-9 clinical isolate was partially dependent on glycopeptide for growth. The responsible vanA operon had the same organization as that of Tn1546 and was located on a plasmid. The chromosomal D-Ala:D-Ala ligase (ddl) gene had two point mutations that led to Q260K and A283E substitutions, resulting in a 200-fold decrease in enzymatic activity compared to that of the wild-type strain VRSA-6.

RAHN-2, a chromosomal extended-spectrum class A beta-lactamase from Rahnella aquatilis.

Objectives: Rahnella aquatilis is an environmental enterobacterial species with a chromosomal bla(RAHN-1) gene encoding extended-spectrum class A beta-lactamase RAHN-1. We describe the diversity of bla(RAHN) genes from two groups of strains, G1 and G2, isolated from raw fruits and vegetables, and the new class A beta-lactamase RAHN-2.

Methods: MICs were determined by Etest.

Crystallization and preliminary X-ray analysis of a D-Ala:D-Ser ligase associated with VanG-type vancomycin resistance.

Acquired VanG-type resistance to vancomycin in Enterococcus faecalis BM4518 arises from inducible synthesis of peptidoglycan precursors ending in D-alanyl-D-serine, to which vancomycin exhibits low binding affinity. VanG, a D-alanine:D-serine ligase, catalyzes the ATP-dependent synthesis of the D-Ala-D-Ser dipeptide, which is incorporated into the peptidoglycan synthesis of VanG-type vancomycin-resistant strains. Here, the purification, crystallization and preliminary crystallographic analysis of VanG in complex with ADP are reported.

VanA-type Staphylococcus aureus strain VRSA-7 is partially dependent on vancomycin for growth.

VanA-type Staphylococcus aureus strain VRSA-7 was partially dependent on glycopeptides for growth. The vanA gene cluster, together with the erm(A) and the ant(9)-Ia resistance genes, was carried by the ca. 35- to 40-kb conjugative plasmid pIP848 present at five copies per cell.

Genetic and biochemical characterization of OXA-63, a new class D beta-lactamase from Brachyspira pilosicoli BM4442.

Brachyspira pilosicoli BM4442, isolated from the feces of a patient with diarrhea at the Hospital Saint-Michel in Paris, was resistant to oxacillin (MIC > 256 microg/ml) but remained susceptible to cephalosporins and to the combination of amoxicillin and clavulanic acid. Cloning and sequencing of the corresponding resistance determinant revealed a coding sequence of 807 bp encoding a new class D beta-lactamase named OXA-63. The bla OXA-63 gene was chromosomally located and not part of a transposon or of an integron.

Genetic and biochemical characterization of CAD-1, a chromosomally encoded new class A penicillinase from Carnobacterium divergens.

Carnobacterium divergens clinical isolates BM4489 and BM4490 were resistant to penicillins but remained susceptible to combinations of amoxicillin-clavulanic acid and piperacillin-tazobactam. Cloning and sequencing of the responsible determinant from BM4489 revealed a coding sequence of 912 bp encoding a class A beta-lactamase named CAD-1. The bla(CAD-1) gene was assigned to a chromosomal location in the two strains that had distinct pulsed-field gel electrophoresis patterns.

Joint effect of nitrogen sources and B vitamin supplementation of date juice on lactic acid production by Lactobacillus casei subsp. rhamnosus.

The use of date juice as a substrate for lactic acid production was investigated. Various nitrogen sources were compared with yeast extract for efficient lactic acid production by Lactobacillus casei subsp. rhamnosus.

Purification and characterization of a trypanothione-glutathione thioltransferase from Trypanosoma cruzi.

Although trypanothione [T(S)2] is the major thiol component in trypanosomatidae, significant amounts of glutathione are present in Trypanosoma cruzi. This could be explained by the existence of enzymes using glutathione or both glutathione and T(S)2 as cofactors. To assess these hypotheses, a cytosolic fraction of T.

Purification and characterization of the VanB ligase associated with type B vancomycin resistance in Enterococcus faecalis V583.

Acquired resistance to glycopeptides in enterococci is associated with the production of D-Alanine:D-Alanine ligase-related proteins. The VanA protein associated with high-level vancomycin and teicoplanin resistance (VanA phenotype) synthesizes a new peptidoglycan precursor, D-alanine-D-lactate, that has reduced glycopeptide affinity. Production of a similar protein, VanB, is induced in strains that display variable levels of vancomycin resistance but remain susceptible to teicoplanin (VanB phenotype).