Identical Penicillin-Binding Domains in Penicillin-Binding Proteins of Streptococcus pneumoniae Clinical Isolates with Different Levels of β-Lactam Resistance

We have sequenced the penicillin-binding domains of the complete repertoire of penicillin-binding proteins and MurM from 22 clinical isolates of Streptococcus pneumoniae that span a wide range of β-lactam resistance levels. Evidence of mosaicism was found in the genes encoding PBP 1a, PBP 2b, PBP 2x, MurM, and, possibly, PBP 2a. Five isolates were found to have identical PBP and MurM sequences, even though the MICs for penicillin G ranged from 0.25 to 2.0 mg/liter. When the sequences encoding PBP 1a, PBP 2b, and PBP 2x from one of these isolates were used to transform laboratory strain R6, the resulting strain had a resistance level higher than that of the less resistant isolates carrying that PBP set but lower than that of the most resistant isolates carrying that PBP set. This result demonstrates that if the R6 strain is arbitrarily defined as the standard genotype, some wild genetic backgrounds can either increase or decrease the PBP-based resistance phenotype

A PBP2x from a clinical isolate of Streptococcus pneumoniae exhibits an alternative mechanism for reduction of susceptibility to beta-lactam antibiotics.

International audienceThe human pathogen Streptococcus pneumoniae is one of the main causative agents of respiratory tract infections. At present, clinical isolates of S. pneumoniae often exhibit decreased susceptibility toward beta-lactams, a phenomenon linked to multiple mutations within the penicillin-binding proteins (PBPs). PBP2x, one of the six PBPs of S. pneumoniae, is the first target to be modified under antibiotic pressure. By comparing 89 S. pneumoniae PBP2x sequences from clinical and public data bases, we have identified one major group of sequences from drug-sensitive strains as well as two distinct groups from drug-resistant strains. The first group includes proteins that display high similarity to PBP2x from the well characterized resistant strain Sp328. The second group includes sequences in which a signature mutation, Q552E, is found adjacent to the third catalytic motif. In this work, a PBP2x from a representative strain from the latter group (S. pneumoniae 5259) was biochemically and structurally characterized. Phenotypical analyses of transformed pneumococci show that the Q552E substitution is responsible for most of the reduction of strain susceptibility toward beta-lactams. The crystal structure of 5259-PBP2x reveals a change in polarity and charge distribution around the active site cavity, as well as rearrangement of strand beta3, emulating structural changes observed for other PBPs that confer drug resistance to Gram-positive pathogens. Interestingly, the active site of 5259-PBP2x is in closed conformation, whereas that of Sp328-PBP2x is open. Consequently, S. pneumoniae has evolved to employ the same protein in two distinct mechanisms of antibiotic resistance

[Epidemiology and clinical characteristics of childhood parapneumonic empyemas]

International audienceSeveral studies have reported an increasing incidence of childhood parapneumonic empyemas in various countries. AIM OF THE STUDY: The aim of our study was to estimate the annual incidence of complicated community-acquired pneumonias in children during a 9-year period in a French area and to describe the epidemiological and clinical characteristics of these complications. POPULATION AND METHODS: We have listed the children from 28 days to 15 years old, hospitalized in the 2 children hospitals of the Isere district for a community-acquired pneumonia complicated with a pleural empyema or a pulmonary abscess from 1995 to 2003. RESULTS: During the study period, 90 children were hospitalized for a complicated pneumonia including 83 pleural empyemas and 7 isolated lung abscess. The average number of cases was 4 per year from 1995 to 1998 then increased since 1999 to reach 34 cases in 2003, according to a linear model (P<0,001). The incidence of the complicated pneumonia, plotted to the paediatric population of the area has gone up from 0.5 per 100000 to 13 per 100000 children between 1995 and 2003. CONCLUSION: The incidence of the complicated pneumonias in children increased since 1999 in a French area. Additional investigations are necessary to identify the causes of this increase

Biochemical Characterization of Streptococcus pneumoniae Penicillin-Binding Protein 2b and Its Implication in β-Lactam Resistance

Extensive use of β-lactam antibiotics has led to the selection of pathogenic streptococci resistant to β-lactams due to modifications of the penicillin-binding proteins (PBPs). PBP2b from Streptococcus pneumoniae is a monofunctional (class B) high-molecular-weight PBP catalyzing the transpeptidation between adjacent stem peptides of peptidoglycan. The transpeptidase domain of PBP2b isolated from seven clinical resistant (CR) strains contains 7 to 44 amino acid changes over the sequence of PBP2b from the R6 β-lactam-sensitive strain. We show that the extracellular soluble domains of recombinant PBP2b proteins (PBP2b*) originating from these CR strains have an in vitro affinity for penicillin G that is reduced by up to 99% from that of the R6 strain. The Thr446Ala mutation is always observed in CR strains and is close to the key conserved motif (S(443)SN). The Thr446Ala mutation in R6 PBP2b* displays a 60% reduction in penicillin G affinity in vitro compared to that for the wild-type protein. A recombinant R6 strain expressing the R6 PBP2b Thr446Ala mutation is twofold less sensitive to piperacillin than the parental S. pneumoniae strain. Analysis of the Thr446Ala mutation in the context of the PBP2b CR sequences revealed that its influence depends upon the presence of other unidentified mutations

Two cases of type A infant botulism in Grenoble, France: no honey for infants.

International audienceWe report two severe cases of infant botulism diagnosed at Grenoble University Hospital, France, respectively in 2006 and 2009. Both cases were characterized by a delay in diagnosis, severe neurological manifestations and extended period of hospitalization in intensive care unit, but a complete recovery. Infant botulism is a rare but life-threatening disease. It primarily affects infants, and the main risk factor is honey ingestion. Diagnosis should be systematically evoked by pediatricians in infants suffering from constipation, fatigue, muscle weakness, difficult feeding and altered cry, but before the onset of generalized flaccid paralysis, so as to administer specific treatment (BabyBIG®, a human derived botulinum antitoxin) at an early stage of the disease when it is most effective. In conclusion, parents should be aware of the role of honey as a source of spores of Clostridium botulinum and therefore infant botulism in the first year of life

[Results of 4 years of microbiological testing of bronchoscopes]

International audienceINTRODUCTION: In the context of reducing endoscopy-related infectious risk and new national guidelines on microbiological samples from bronchoscopy, the results of a surveillance program set up in a hospital center were analyzed. METHODS: Over 4 years, scheduled samples were taken from disinfected bronchoscopes. Bacteriology and mycology tests were used to search for microorganisms. The results were interpreted as falling within three levels: target, alert, and action. Factors that could explain the contamination were studied: age of the bronchoscope, number of uses per year, brand, and model. RESULTS: Out of 96 scheduled samples taken, the compliance rate for the period was 83% and increased (p=0.06) over the 4 years. We identified 15 Pseudomonas (six aeruginosa and nine other species), one Stenotrophomonas, one enterobacterium, and two filamentous fungi. None of the factors studied had a significant effect on sample contamination. CONCLUSION: The microbiological surveillance of bronchoscopes is an indispensable part of the quality assurance of bronchoscope maintenance. It can lead to maintenance of the bronchoscope when a noncompliant result is found