Pascal Cholley

Most Multidrug-Resistant Pseudomonas aeruginosa Isolates from Hospitals in Eastern France Belong to a Few Clonal Types

ABSTRACT This study aimed to determine the genetic diversity of clinical multidrug-resistant Pseudomonas aeruginosa. We used pulsed-field gel electrophoresis and multilocus sequence typing to analyze 187 strains isolated in different French hospitals. To illustrate the diversity of resistance mechanisms to antibiotics in a given clone, we identified β-lactamases with an extended spectrum by using phenotypic and genotypic methods. Typing results showed that the majority of our multidrug-resistant isolates belong to a few clonal types (ST235, ST111, and ST175) that are already spreading worldwide. These successful international clones sporadically produced extended-spectrum β-lactamase-encoding genes but mostly became extensively resistant to β-lactams after derepression of intrinsic resistance mechanisms (i.e., AmpC cephalosporinase). Our results indicate that cross-transmission plays a major role in the spread of multidrug-resistant P. aeruginosa in hospital settings.

The role of water fittings in intensive care rooms as reservoirs for the colonization of patients with Pseudomonas aeruginosa.

ObjectiveTo assess the role of the water environment in the Pseudomonas aeruginosa colonization of patients in intensive care units in the absence of axa0recognized outbreak.Design and settingProspective, single-centre study over an 8-week period in two adult ICUs at axa0university hospital. Environmental samples were taken from the water fittings of rooms once per week, during axa08-week period. Patients were screened weekly for P.xa0aeruginosa carriage. Environmental and humans isolates were genotyped by using pulsed-field gel electrophoresis.ResultsP.xa0aeruginosa was detected in 193 (86.2%) of the 224 U-bend samples and 10 of the 224 samples taken from the tap (4.5%). Seventeen of the 123 patients admitted were colonized with P.xa0aeruginosa. Only one of the 14 patients we were able to evaluate was colonized by axa0clone present in the water environment of his room before the patients first positive sample was obtained.ConclusionThe role of the water environment in the acquisition of P.xa0aeruginosa by intensive care patients remains unclear, but water fittings seem to play axa0smaller role in non-epidemic situations than expected by many operational hospital hygiene teams.

IMP-29, a Novel IMP-Type Metallo-β-Lactamase in Pseudomonas aeruginosa

ABSTRACT Analysis of two clonally related multiresistant Pseudomonas aeruginosa isolates led to the identification of a novel IMP-type metallo-β-lactamase. IMP-29 was significantly different from the other IMP variants (the closest variant being IMP-5 with 93% amino acid identity). The blaIMP-29 gene cassette was carried by a class 1 integron in strain 10.298, while in strain 10.266 it was located in a rearranged DNA region on a 30-kb conjugative plasmid. Biochemical analysis confirmed that IMP-29 efficiently hydrolyzed carbapenems.

Lagooning of wastewaters favors dissemination of clinically relevant Pseudomonas aeruginosa

The significance of wastewater treatment lagoons (WWTLs) as point sources of clinically relevant Pseudomonas aeruginosa that can disseminate through rural and peri-urban catchments was investigated. A panel of P. aeruginosa strains collected over three years from WWTLs and community-acquired infections was compared by pulsed field gel electrophoresis (PFGE) DNA fingerprinting and multilocus sequence typing (MLST). Forty-four distantly related PFGE profiles and four clonal complexes were found among the WWTL strains analyzed. Some genotypes were repeatedly detected from different parts of WWTLs, including the influent, suggesting an ability to migrate and persist over time. MLST showed all investigated lineages to match sequence types described in other countries and strains from major clinical clones such as PA14 of ST253 and C of ST17 were observed. Some of these genotypes matched isolates from community-acquired infections recorded in the WWTL geographic area. Most WWTL strains harbored the main P. aeruginosa virulence genes; 13% harbored exoU-encoded cytoxins, but on at least six different genomic islands, with some of these showing signs of genomic instability. P. aeruginosa appeared to be highly successful opportunistic colonizers of WWTLs. Lagooning of wastewaters was found to favor dissemination of clinically relevant P. aeruginosa among peri-urban watersheds.

A Bundle of Measures to Control an Outbreak of Pseudomonas aeruginosa Associated With P-Trap Contamination

OBJECTIVE To describe an outbreak of multidrug-resistant Pseudomonas aeruginosa in which the hospital waste-pipe system was the likely source of contamination and to report the bundle of measures that facilitated the long-term control of the outbreak. DESIGN Outbreak investigation. SETTING The hematology unit of a tertiary-care referral center. PATIENTS Patients who were colonized or infected with P. aeruginosa belonging to the clonal outbreak. METHODS Patients admitted to our 15-bed stem-cell transplantation hematology unit were screened for P. aeruginosa carriage. Pseudomonas aeruginosa isolates were also obtained from diagnostic samples. We assessed the microbiological contamination of P-traps, water and toilets for 42 months. Extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) were screened and identified by polymerase chain reaction (PCR) and sequencing. Molecular typing of ESBL- or MBL-producing isolates was carried out using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). RESULTS From 2009 to 2013, a biclonal outbreak of IMP-19-producing ST235 (11 cases) and IMP-29-producing ST111 (10 cases) of P. aeruginosa occurred. The environmental investigation strongly suggested that P-traps were the reservoirs for the outbreak strains. A bundle of infection control measures, including engineering interventions on water outlets and disinfection of P-traps, controlled the outbreak. CONCLUSIONS We report a prolonged outbreak of IMP-producing high-risk clones of P. aeruginosa, for which P-traps seems to play a major role in cross-transmission. It appears essential to implement proactive measures to limit the bacterial load in water fittings of high-risk units. Infect Control Hosp Epidemiol 2018;39:164-169.

Clonal Dissemination of Pseudomonas aeruginosa Isolates Producing Extended-Spectrum β-Lactamase SHV-2a

ABSTRACT From January to December 2011, 24 Pseudomonas aeruginosa strains producing the extended-spectrum β-lactamase SHV-2a were identified in 13 hospitals in France. With one exception, all the strains belonged to the same clone. Double-disk synergy tests with cefepime and clavulanate were able to detect all the SHV-2a-positive isolates.

High prevalence and moderate diversity of Pseudomonas aeruginosa in the U-bends of high-risk units in hospital

The presence of P. aeruginosa in water supply is clearly identified as a risk factor for P. aeruginosa infection in critical care units, even if routes of transmission are often unclear and remain a matter of debate. We determined here the frequency of U-bends contaminated with P. aeruginosa in high-risk units and described the population structure of this opportunistic pathogen in a non-outbreak situation. Eighty-seven U-bends from sinks of rooms in five wards were sampled 3 times and P. aeruginosa was detected in 121 of the 261 (46.4%) U-bend samples. We genotyped 123 P. aeruginosa isolates with pulsed-field gel electrophoresis and multilocus sequence typing and found 41 pulsotypes distributed in 21 Sequence Types (STs). Seven major ST (ST111, CC235, CC253, ST520, ST539, ST1216, and ST1725) were overrepresented in the collection, including the high-risk clones ST111, CC253, and CC235. The distribution of the 21 STs in the cladogram of the species was uneven with most major STs clustering into 2 clades. The major STs were found in different units and buildings and could be represented by a high diversity of pulsotypes. Altogether, this suggests a long term presence of P. aeruginosa in the hospital water network, possibly contaminated by the distribution water or by plumbing fittings before putting into service. Analysis of resistance rates showed that the deficiency of porin OprD was very frequent in U-bends isolates that may benefit from this resistance mechanism in hospital water fittings. In conclusion, our study demonstrates that U-bends of high-risk units are very frequently contaminated with P. aeruginosa with a moderate genomic diversity and with an over-representation of adapted clones.

Trends of extended-spectrum β-lactamase-producing Escherichia coli sequence type 131 and its H30 subclone in a French hospital over a 15-year period

Sequence type 131 (ST131) is a predominant lineage among extraintestinal pathogenic Escherichia coli. It plays a major role in the worldwide dissemination of E. coli producing extended-spectrum β-lactamases (ESBLs). Here we describe the long-term epidemiology of this clonal group in a French university hospital, where the incidence of ESBL-producing E. coli has increased from 0.018 case per 1000 patient-days in the year 2000 to 0.50 case per 1000 patient-days in 2014. The first of the 141 ST131 isolates was recovered in 2006, and the ST131 clonal group accounted for 18.1% of total ESBL-producing E. coli over the whole period (2000-2014). Subclonal typing showed that 75.9% (107/141) of ST131 isolates were H30, of which 81.3% (87/107) were H30-Rx. The large majority (137/141) of ESBLs produced were of the CTX-M group, with 94 CTX-M-15, 19 CTX-M-1, 10 CTX-M-27, 8 CTX-M-14 and four other CTX-M types (nu2009=u20096). Pulsed-field gel electrophoresis (PFGE) analysis showed high diversity, which increased during the course of the study. The 141 ST131 isolates clustered in 53 pulsotypes (PTs), with 2 dominant PTs (PT14 and PT13) with 36 and 17 isolates, respectively. These findings showed that ST131 was a predominant clone among ESBL-producing E. coli in our hospital, even though it only accounted for <20%. Moreover, ST131 should be regarded not as a unified entity but as a cluster of distinct clonal subsets even if the increase in resistance within ST131 has a strong clonal basis, being attributable mainly to the spread of C1/H30-R and C2/H30-Rx clades.

Outbreak of IMI-1 carbapenemase-producing colistin-resistant Enterobacter cloacae on the French island of Mayotte (Indian Ocean)

The spread of carbapenemase-producing Enterobacteriaceae in the Southwest Indian Ocean islands is poorly known. Here we describe an outbreak of colistin-resistant Enterobacter cloacae harbouring blaIMI-1 in the French overseas department of Mayotte. Between October 2015 and January 2017, all isolates of imipenem-non-susceptible E. cloacae at Mayotte Medical Center and University Hospital of Reunion Island were screened for carbapenemase production. Positive isolates were typed by pulsed-field gel electrophoresis and whole-genome sequencing (WGS)-based multilocus sequence typing (MLST), and all β-lactamase genes were identified by PCR and sequencing. Resistance profiles were determined by agar diffusion and Etest. Genetic support of the blaIMI-1 gene was determined by WGS. A total of 18 E. cloacae isolates harbouring blaIMI-1 were detected in 17 patients from Mayotte. Pulsed-field gel electrophoresis (PFGE) analysis showed 16 of the 18 strains to be clonally related and belonging to ST820. Based on clinical data, this outbreak most likely had a community origin. The blaIMI-1 gene in the 18 isolates was carried by a new variant of an integrative mobile element involving the Xer recombinases, called EcloIMEX-8. The mcr-1-mcr-5 genes were absent from the collection. The isolates belonged to E. cloacae cluster XI, known to be colistin heteroresistant. Here we report the first outbreak of IMI-1-producing Enterobacteriaceae. IMI-1-producers may be underdetected in microbiology laboratories because of their unusual antimicrobial resistance profile (resistant to imipenem but with intermediate resistance to ertapenem and susceptible to extended-spectrum cephalosporins) and the absence of blaIMI-1 in the panel of genes targeted by molecular diagnostic kits.

Épidémiologie des infections à Pseudomonas aeruginosa

Resume Bacille a Gram negatif, saprophyte de l’environnement hydrique, Pseudomonas aeruginosa apparait aujourd’hui comme un pathogene majeur au sein des etablissements de sante, ou il est responsable de 10 % des infections nosocomiales, ce taux augmentant jusqu’a 15 % dans les services de reanimation. Le portage endogene a l’admission des patients et l’acquisition des souches en cours d’hospitalisation, soit a partir de l’environnement hydrique, soit par transmission croisee de patient a patient, representent les differentes voies de contamination precedant la survenue des infections. La dissemination de clones epidemiques precede le plus souvent l’acquisition d’une multi-resistance aux antibiotiques chez un patient donne sous l’effet de la pression de selection des antibiotiques. L’emergence de souches toto-resistantes a recemment complique la prise en charge des patients infectes et entraine une surmortalite inquietante. L’etude de l’epidemiologie hospitaliere de P. aeruginosa passe par deux techniques d’analyse moleculaire complementaires: le profil de macrorestriction de l’ADN total par electrophorese en champ pulse pour l’epidemiologie locale et la MLST (multilocus sequence typing) pour l’approche macro-epidemiologique. Les donnees epidemiologiques acquises sur P. aeruginosa dans les services de reanimation depuis quelques annees bouleversent l’image univoque ancree dans les esprits. Ainsi la transmission directe de patient a patient et la frequence du portage a l’admission rapprochent cette espece des autres pathogenes hospitaliers multi-resistants.