Marie-Frédérique Lartigue

Genetic Structures at the Origin of Acquisition of the β-Lactamase blaKPC Gene

ABSTRACT Genetic structures surrounding the carbapenem-hydrolyzing Ambler class A blaKPC gene were characterized in several KPC-positive Klebsiella pneumoniae and Pseudomonas aeruginosa strains isolated from the United States, Colombia, and Greece. The blaKPC genes were associated in all cases with transposon-related structures. In the K. pneumoniae YC isolate from the United States, the β-lactamase blaKPC-2 gene was located on a novel Tn3-based transposon, Tn4401. Tn4401 was 10 kb in size, was delimited by two 39-bp imperfect inverted repeat sequences, and harbored, in addition to the β-lactamase blaKPC-2 gene, a transposase gene, a resolvase gene, and two novel insertion sequences, ISKpn6 and ISKpn7. Tn4401 has been identified in all isolates. However, two isoforms of this transposon were found: Tn4401a was found in K. pneumoniae YC and K. pneumoniae GR from the United States and Greece, respectively, and differed by a 100-bp deletion, located just upstream of the blaKPC-2 gene, compared to the sequence of Tn4401b, which was found in the Colombian isolates. In all isolates tested, Tn4401 was flanked by a 5-bp target site duplication, the signature of a recent transposition event, and was inserted in different open reading frames located on plasmids that varied in size and nature. Tn4401 is likely at the origin of carbapenem-hydrolyzing β-lactamase KPC mobilization to plasmids and its further insertion into various-sized plasmids identified in nonclonally related K. pneumoniae and P. aeruginosa isolates.

ISEcp1B-Mediated Transposition of blaCTX-M in Escherichia coli

ABSTRACT Several expanded-spectrum β-lactamase blaCTX-M genes are associated with ISEcp1-like elements in Enterobacteriaceae. We found that ISEcp1B was able to mobilize the adjacent blaCTX-M-19 gene by a transpositional mechanism in Escherichia coli by recognizing a variety of DNA sequences as right inverted repeats.

Extended-Spectrum β-Lactamase CTX-M-1 in Escherichia coli Isolates from Healthy Poultry in France

ABSTRACT Genes encoding extended-spectrum β-lactamase CTX-M-1 were detected in 12 Escherichia coli isolates recovered over a 7-month period from the ceca of healthy poultry in seven districts in France in 2005. Eleven of those strains were not clonally related and had a blaCTX-M-1 gene located on transferable plasmids of different sizes and structures.

In Vitro Analysis of ISEcp1B-Mediated Mobilization of Naturally Occurring β-Lactamase Gene blaCTX-M of Kluyvera ascorbata

ABSTRACT ISEcp1B has been reported to be associated with and to mobilize the emerging expanded-spectrum β-lactamase blaCTX-M genes in Enterobacteriaceae. Thus, the ability of this insertion sequence to mobilize the blaCTX-M-2 gene was tested from its progenitor, Kluyvera ascorbata. Insertions of ISEcp1B upstream of the blaCTX-M-2 gene in K. ascorbata reference strain CIP7953 were first selected with cefotaxime (0.5 and 2 μg/ml). In those cases, ISEcp1B brought promoter sequences enhancing blaCTX-M-2 expression in K. ascorbata. Then, ISEcp1B-mediated mobilization of the blaCTX-M-2 gene from K. ascorbata to Escherichia coli J53 was attempted. The transposition frequency of ISEcp1B-blaCTX-M-2 occurred at (6.4 ± 0.5) × 10−7 in E. coli. Cefotaxime, ceftazidime, and piperacillin enhanced transposition, whereas amoxicillin, cefuroxime, and nalidixic acid did not. Transposition was also enhanced when studied at 40°C.

Identification of Streptococcus agalactiae Isolates from Various Phylogenetic Lineages by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry

ABSTRACT Variations in proteins related to bacterial diversity may affect species identification performed using matrix-assisted laser desorption ionization (MALDI)-time of flight mass spectrometry. Using this method, we identified 110 Streptococcus agalactiae isolates characterized by serotyping and multilocus sequence typing. Serotype III and sequence type 23 strains expressed the widest variation in molecular weight of putative “species-identifying” biomarker ions. Recognition of the diversity of MALDI patterns observed in strains that represent all major intraspecies lineages assists in the constitution of an optimal reference database.

Corynebacterium ulcerans in an Immunocompromised Patient with Diphtheria and Her Dog

ABSTRACT Corynebacterium ulcerans causes zoonotic infections, such as diphtheria and extrapharyngeal infections. We report here the first case of a diphtheria-like illness caused by C. ulcerans in France and transmitted likely by a dog to an immunocompromised woman.

Extended-Spectrum β-Lactamases of the CTX-M Type Now in Switzerland

ABSTRACT The epidemiology of clavulanic acid-inhibited extended-spectrum β-lactamases (ESBLs) was investigated among infection-associated enterobacterial isolates at the University Hospital in Lausanne, Switzerland, from January 2004 to June 2005. Out of 57 nonrepetitive ESBL producers (prevalence rate of 0.7%), 45 produced CTX-M-like ESBLs. CTX-M enzymes were mostly from clonally nonrelated Escherichia coli isolates, from urinary infections and community-acquired infections. Pediatric patients (20 out of 57) accounted for a large number of CTX-M producers. CTX-M-15 was the most frequent CTX-M-type enzyme. The plasmid-located blaCTX-M genes were associated with either ISEcp1 or ISCR1 insertion sequences. This study is the first published report of CTX-M-type β-lactamases in Switzerland.

Molecular Characterization of Human-Colonizing Streptococcus agalactiae Strains Isolated from Throat, Skin, Anal Margin, and Genital Body Sites

ABSTRACT Streptococcus agalactiae carriage was evaluated by sampling four body sites in a group of 249 healthy individuals including both sexes and a wide range of ages; the aims were to study the population structure of colonizing strains by multilocus sequence typing (MLST) and to evaluate their diversity by serotyping, SmaI macrorestriction analysis, and PCR screening for genetic markers of highly virulent clones for neonates. The prevalences of carriage were 27% in women and 32% in men. The major positive body site was the genital tract (23% in women and 21% in men); skin, throats, and anal margins were also positive in 2%, 4%, and 14%, respectively. These human-colonizing strains belonged mostly to serotypes III (24%), Ia (21%), V (18%), and Ib (17%). Twenty-three sequence types (STs) were identified. The MLST characteristics of the strains isolated from a single anatomic site—genital (vagina [women] or from a sample of the first urination after arising from a nights sleep [men]), throat, skin, or anal margin—suggest a body site colonization specificity for particular STs: strains of STs 2, 10, 19, and 196 were isolated only from genital sites; strains of STs 1, 8, and 23 were isolated more frequently from throat florae; and strains recovered only from anal margin samples were more closely related to strains isolated from throats than to those from genital sites. Most strains of STs 1, 8, and 23—STs that are increasingly described as being responsible for adult infections—did not carry any markers of strains virulent for neonates, suggesting that the virulence of these strains is probably associated with other genetic determinants. In addition, the genetic diversities of the strains varied between STs: STs 2, 8, 10, 23, and 196 were the most diverse; STs 1 and 19 were more homogeneous; and ST 17 strains formed three distant groups.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry for bacterial strain characterization.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has emerged as a new technique for rapid and effective bacterial species identification in clinical microbiology laboratories. Conventional bacterial identification is based essentially on biochemical tests and requires lengthy incubation procedures. By contrast, MALDI-TOF MS can identify bacterial species in a few minutes, directly from a freshly grown colony. Various approaches have been developed for bacterial identification and the efficacy of this method has been demonstrated in several studies. This technique should also be suitable for the classification of organisms to subspecies level and for the phenotypic detection of certain antibiotic resistance mechanisms, such as β-lactamases production.

Molecular characterization of erythromycin-resistant Streptococcus agalactiae strains

OBJECTIVES The aim of this study is to identify the molecular characteristics of erythromycin-resistant (Erm(r)) Streptococcus agalactiae strains and to correlate with the clinical origin of strains. METHODS From 711 S. agalactiae strains, 119 Erm(r) strains (17%) were collected, serotyped and screened for macrolide resistance genes. The genetic relationship between strains was established by the PFGE analysis. Strains were tested for the group II intron GBSi1 downstream of the scpB gene, IS1548 in the hylB gene, four prophage DNA fragments and a lineage defined by multilocus sequence typing as ST-17. RESULTS Erythromycin resistance involved 8% of serotype Ia, 15% of serotype Ib, 9% of serotype II, 16% of serotype III, 31% of serotype IV and 35% of serotype V. The prevalence of Erm(r) strains was higher among strains isolated from the gastric fluid of neonates (33%) than in those isolated from bacteraemia and meningitis during early-onset disease (EOD) or late-onset disease (7% and 11%) (P = 0.001). In serotype III, Erm(r) strains were more frequent in vaginal carriage (22%) and colonized neonates (18%) than in EOD (0%) (P = 0.03). The mef(A) gene was the most common in serotype Ia (55%), the erm(A) gene in serotype Ib (75%) and the erm(B) gene in the other serotypes (56% to 75%). All resistant strains with IS1548 also had the erm(B) gene. Erm(r) strains were not randomly distributed in the different PFGE genogroups, and 11% had the GBSi1 intron, 37% had at least one prophage DNA fragment and 7% belonged to ST-17. CONCLUSIONS Erythromycin resistance varied according to the clinical origin, serotype and molecular characteristics of S. agalactiae strains.