[show abstract][hide abstract] ABSTRACT: qnrD is a plasmid mediated quinolone resistance gene from unknown origin, recently described in Enterobacteriaceae. It encodes a pentapeptide repeat protein 36-60% different from the other Qnr (A, B, C, S and VC). Since most qnrD-positive strains were described as strains belonging to Proteus or Providencia genera, we hypothesized that qnrD originated in Proteeae before disseminating to other enterobacterial species. We screened 317 strains of Proteeae for qnrD and its genetic support by PCR. For all the seven qnrD-positive strains (4 Proteus mirabilis, 1 Proteus vulgaris and 2 Providencia rettgeri) the gene was carried onto a small non-transmissible plasmid, contrarily to other qnr genes that are usually carried onto large multi-resistant plasmids. Nucleotide sequences of the qnrD-bearing plasmids were 96% identical. Plasmids contained 3 ORFs apart from qnrD and belonged to an undescribed incompatibility group. Only one plasmid, in P. vulgaris, was slightly different with a 1,568-bp insertion between qnrD and its promoter, leading to absence of quinolone resistance. We sought for similar plasmids in 15 reference strains of Proteeae, but which were tested negative for qnrD, and found a 48% identical plasmid (pVERM) in Providencia vermicola. In order to explain how qnrD could have been inserted into such native plasmid, we sought for gene mobilization structures. qnrD was found to be located within a mobile insertion cassette (mic) element which sequences are similar to one mic also found in pVERM. Our conclusions are that (i) the small non-transmissible qnrD-plasmids described here may result from the recombination between an as-yet-unknown progenitor of qnrD and pVERM, (ii) these plasmids are maintained in Proteeae being a qnrD reservoir (iii) the mic element may explain qnrD mobilization from non-transmissible plasmids to mobilizable or conjugative plasmids from other Enterobacteriaceae, (iv) they can recombined with larger multiresistant plasmids conjugated in Proteeae.
PLoS ONE 01/2014; 9(2):e87801. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: The aim of this study was to describe the presence of plasmid-mediated quinolone resistance (PMQR) determinants in extended spectrum β-lactamase-producing Escherichia coli (ESBL-Ec) in hospitals in eastern France. All ESBL-Ec isolated from May 2008 to April 2009 in nine hospitals in eastern France were collected and screened by PCR for the presence of PMQR genes. bla genes were identified by PCR and sequencing. Randomly chosen PMQR-positive ESBL-Ec isolates were characterised by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and phylogenetic group identification. Of the 447 ESBL-Ec tested, 122 (27.3%) had a PMQR determinant. aac(6′)-Ib-cr was more prevalent (118/122; 96.7%) than qnr [4/122 (3.3%), comprising 2 qnrB2, 1 qnrB1 and 1 qnrA1]. Among 37 PMQR-positive ESBL-Ec isolates selected for typing, 26 (70%) carried blaCTX-M-15 of which 25/26 (96%) co-harboured aac(6′)-Ib-cr. Of the 37 isolates, 14 belonged to the B2:ST131 clone, all of which produced AAC(6’)-Ib-cr. On the other hand, most of the qnr genes (3/4) were observed in strains carrying bla genes other than blaCTX-M (qnrB2 and blaSHV-12; qnrA1 and blaTEM-52). These findings suggest that aac(6′)-Ib-cr, rather than qnr, is the spreading PMQR determinant in ESBL-Ec isolates, and the qnr determinant is not specifically associated with blaCTX-M genes in France. aac(6′)-Ib-cr was the predominant PMQR gene in ESBL-Ec isolated in eastern France hospitals, suggesting that the distribution may be due to clonal spread of ST131 CTX-M-15-producing E. coli isolates.
Journal of Global Antimicrobial Resistance. 01/2014;
[show abstract][hide abstract] ABSTRACT: We previously described a pyrosequencing-based method able to guarantee detection of aac(6')-Ib-cr by characterizing precisely the single nucleotide polymorphisms leading to Trp102Arg and Asp179Tyr. By gaining in simplicity and cost-effectiveness, through the use of real-time PCR, we propose here a cutting-edge evolution of our existing pyrosequencing method.
Journal of microbiological methods 09/2013; · 2.43 Impact Factor
[show abstract][hide abstract] ABSTRACT: AAC(6')-Ib-cr is a plasmid-mediated quinolone resistance mechanism described worldwide in Escherichia coli. Since it confers in vitro only a low level of resistance to ciprofloxacin, we evaluated its impact on the in vivo activity of ciprofloxacin. Isogenic strains were obtained by transferring the plasmid p449 harboring aac(6')-Ib-cr into the quinolone susceptible E. coli CFT073-RR and its D87G gyrA mutant. MICs were 0.015, 0.06, 0.25 and 0.5 μg/ml against E. coli strains CFT073-RR, CFT073-RR/p449, CFT073-RRGyrA(R) and CFT073-RRGyrA(R)/p449 respectively. Bactericidal activity was reduced at 1 x MIC for the three resistant derivatives while at a fixed concentration of 0.5 μg/ml, a 99.9% killing was observed of all strains except E. coli CFT073-RRGyrA(R)/p449. In the murine model of pyelonephritis, an optimal regimen of ciprofloxacin (10mg/kg bid) significantly decreased the bacterial count in the kidneys of mice infected with E. coli CFT073 (1.6 vs 4.3 log10CFU/g of kidney as compared to untreated controls P = 0.0001), while no significant decrease was observed for E. coli CFT073-RR/p449 (2.7 vs 3.1 log10CFU/g; P = 0.84), E. coli CFT073-RRGyrA(R) (4.2 vs 4.1 log10CFU/g; P = 0.35), or E. coli CFT073-RRGyrA(R)/p449 (2.9 vs 3.6 log10CFU/g; P = 0.47). While PK/PD parameters accounted for ciprofloxacin failure against gyrA-containing mutants, this was not the case for the aac(6')-Ib-cr containing strains, suggesting an in situ hydrolysis of ciprofloxacin in the latter case.
Antimicrobial Agents and Chemotherapy 09/2013; · 4.57 Impact Factor
[show abstract][hide abstract] ABSTRACT: The concept of sudden infant death syndrome (SIDS) is defined as the sudden, unexpected death of an infant less than a year old which remains unexplained after in-depth investigations comprising a complete autopsy, biological analyses, and a clinical examination of the circumstances surrounding the death. This definition underlines the importance of finding the cause of this disease in order to improve preventative measures to reduce the number of deaths due to sudden infant death syndrome. Among the causes of SIDS, pediatric infectious diseases may be neglected and must be systematically sought after. We report upon a SIDS death case of a four and a half month-old that occurred during his sleep. Following the absence of an evident cause of death a scientific autopsy was performed. The histological examination of pulmonary tissue revealed broncolitic lesions associated with numerous micro-abscesses. The post mortem microbiological analyses revealed evidence of an infection by the respiratory syncytial virus complicated by a bacterial infection due to Haemophilus influenzae. The case underlines the necessity of a multidisciplinary approach to researching SIDS, involving both clinicians and biologists, in order to determine the causes of these deaths.
Annales de biologie clinique. 06/2013; 71(3):299-304.
[show abstract][hide abstract] ABSTRACT: Nocardia spp. are bacteria often implicated in pulmonary diseases and central nervous system infections, especially in immunocompromised patients. We report here the case of an immunocompromised woman presenting an insidious brain abcess initially treated as a cerebral stroke. Despite a cotrimoxazole and rifampicin treatment she did not improve. She died 3 month later after she stopped her treatment.
Annales de biologie clinique. 06/2013; 71(3):345-348.
[show abstract][hide abstract] ABSTRACT: Diarrhea is a frequent complication after kidney transplantation ascribed to adverse effects of the immunosuppressive therapy in case of negative microbiological examination of the stools. The aim of this study was to improve the microbiological diagnosis by implementing molecular tests. Fifty-four severe diarrhea events occurred in 49 adult kidney transplant recipients from September 2010 to November 2011 were investigated. One or several enteric pathogens were detected in 13 (23%) stool samples using classical microbiological methods versus 39 (72%) for the seven commercially available multiplex PCR assays used retrospectively (P=0.006). Interestingly, molecular diagnosis identified 15 multiple infections compared to none using classical techniques. The primary pathogens detected were enteropathogenic Escherichia coli (EPEC) (n=15; 38%), Campylobacter sp. (n=15; 38%) and Norovirus (n=14; 36%). Specificity for Campylobacter and Norovirus infections diagnosis was 75 and 100%, respectively, by comparison to reference methods. Based on molecular findings, ciclosporin and mycophenolate mofetil combination was identified as risk factor of developing Norovirus diarrhea. Norovirus infections were also responsible for higher weight loss than all the other causes of diarrhea. In samples from asymptomatic immunocompromised and immunocompetent patients, EPEC but not Norovirus and Campylobacter infections were detected at similar frequency than that observed in symptomatic kidney transplant recipients. In conclusion, molecular tools significantly improved the detection of single and multiple enteric infections by comparison to classical techniques and could quickly become the key element of the management of severe acute diarrhea in transplant recipients.
Journal of clinical microbiology 04/2013; · 4.16 Impact Factor
[show abstract][hide abstract] ABSTRACT: An unusual multi-drug-resistant Pseudomonas aeruginosa (MDR-PA) was isolated in four patients whilst hospitalized in a French teaching hospital between May and August 2011. All four patients had undergone an oesophago-gastro-duodenoscopy with the same gastroscope over a five-month period. This endoscope was associated with a culture positive for the MDR-PA. Observations of endoscope reprocessing identified deviations from the agreed processes: insufficient initial cleaning, shortening of the immersion time and brushing time, insufficient channel flushing, and inadequate drying prior to storage. Since withdrawing the gastroscope and institution of strict adherence to the agreed processes, no other MDR-PA cases have been isolated.
The Journal of hospital infection 01/2013; · 3.01 Impact Factor
[show abstract][hide abstract] ABSTRACT: Abstract The purpose of this work was to study the genetic determinants responsible for extended-spectrum cephalosporin (ESC) resistance of Salmonella collected during the period of 1995-2008 at the Hussein Dey hospital in Algiers (Algeria). Fourteen ESC-resistant Salmonella isolates were tested towards 22 antimicrobial agents. Polymerase chain reaction (PCR) and sequencing were used to determine the underlying genetic determinants responsible for the extended-spectrum beta-lactamase (ESBL) phenotypes. Enterobacterial Repetitive Intergenic Consensus PCR was employed to type the isolates. All tested isolates were resistant to ticarcillin, ticarcillin-clavulanate, piperacillin, cefuroxime, aztreonam, ceftazidime, cefotaxime (except two isolates), cefepime, and cefpirome. PCR and DNA sequencing identified these ESBLs as TEM-48 (n=6), TEM-4 (n=3), CTX-M-15 (n=4), and one new TEM, designated TEM-188. Thus, continued surveillance for the presence of ESBL-producing (non-typhoidal) salmonellae in Algeria is essential.
Foodborne Pathogens and Disease 08/2012; 9(9):803-8. · 2.28 Impact Factor
[show abstract][hide abstract] ABSTRACT: qnr genes are plasmid-mediated quinolone resistance genes. Five qnr families have been described with several alleles (7 alleles of qnrA, 53 alleles of qnrB, 1 allele of qnrC, 1 allele of qnrD and 6 alleles of qnrS). Their detection requires a PCR specific for each qnr family and further sequencing for allele characterization.
High-resolution melt curve analysis (HRMA) was coupled to multiplex and simplex real-time PCR assays for detection and characterization of qnrA, qnrB and qnrS alleles. The protocol was set using 27 reference strains harbouring the most frequent alleles and was applied to 55 clinical isolates unknown for qnr positivity.
Out of the 27 reference strains tested, 21 alleles showed distinct profiles using HRMA: 6 qnrA, 12 qnrB and 3 qnrS. For the qnrB alleles showing similar profiles, we gathered them into four groups that were easily distinguished. For the alleles that we could not test, in silico analysis showed that they would be identified using the HRMA protocol set. Among the clinical isolates, 28 qnr-positive isolates were detected and the qnr allele was characterized as 8 qnrA1, 4 qnrB1, 5 qnrB2, 3 qnrB4, 1 qnrB8, 1 qnrB5, 3 qnrS1 and 1 qnrS2, with concordant results with PCR sequencing. Two new qnrB alleles were detected and distinguished using HMRA. They were further designated as qnrB25 and qnrB42.
We developed an HRMA assay for characterizing the qnr alleles in clinical isolates. This high-throughput method can be used to screen a large number of isolates. This method allowed the detection of new qnrB alleles.
Journal of Antimicrobial Chemotherapy 07/2012; 67(11):2635-9. · 5.34 Impact Factor
[show abstract][hide abstract] ABSTRACT: PURPOSE: To describe and to compare species and antibiotics resistance patterns of bacteria involved among bacteriuria from hospital and city laboratories and among health-related and community-acquired bacteriuria. METHODS: Epidemiologic transversal study conducted among Bacteriology laboratories of University Hospital (UH) and the whole city of Reims, during the week 21 to 26 June 2010. A standardized investigation form was completed after telephonical interview with the prescriber. RESULTS: One hundred and eighty-nine strains have been isolated among 179 urocultures. One hundred and seven strains were isolated in city laboratories and 82 in UH laboratory. Strains were community-acquired, health-related and nosocomial in 136, 22 and 31 cases, respectively. More Gram positive bacteria and ofloxacin resistant strains were isolated among UH strains (P=0.001 and P=0.015, respectively) and among health-related strains (P=0.01 and P=0.003, respectively). When analysis was restricted only to Enterobacteriaceae or to Escherichia coli, the ofloxacin resistance rate was no more elevated among health-related or UH strains. Ofloxacin resistant Enterobacteriaceae were more frequently resistant to all other classes of antibiotics except nitrofurans. DISCUSSION: Strains isolated in health-related bacteriuria are more frequently ofloxacin resistant principally because of the greater proportion of Gram positive bacteria and because of a non-significant higher ofloxacin resistance rate among Enterobacteriaceae. Numerous studies only focus on Enterobacteriaceae, and the data from our study need to be confirmed on larger samples, in order to validate the predictive value of health-related bacteriuria for ofloxacin resistance.
[show abstract][hide abstract] ABSTRACT: Nosocomial outbreaks of extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae are an increasing concern in neonatal intensive care units (NICUs). We describe an outbreak of ESBL-producing K. pneumoniae that lasted 5 months and affected 23 neonates in our NICU. Proton pump inhibitor and extended-spectrum cephalosporin exposure were significantly associated with the risk of ESBL-producing K. pneumoniae colonisation and/or infection. Thirty isolates recovered from clinical, screening and environmental samples in the NICU were studied by means of Raman spectroscopy, pulsed-field gel electrophoresis and repetitive extragenic palindromic polymerase chain reaction (rep-PCR). The Raman clustering was in good agreement with the results of the other two molecular methods. Fourteen isolates belonged to the Raman clone 1 and 16 to the Raman clone 3. Molecular analysis showed that all the strains expressed SHV-1 chromosomal resistance, plasmid-encoded TEM-1 and CTX-M-15 β-lactamases. Incompatibility groups of plasmid content identified by PCR-based replicon typing indicated that resistance dissemination was due to the clonal spread of K. pneumoniae and horizontal CTX-M-15 gene transfer between the two clones.
European Journal of Clinical Microbiology 05/2012; 31(10):2827-34. · 3.02 Impact Factor
[show abstract][hide abstract] ABSTRACT: We sought to determine the epidemiological patterns of Staphylococcus bacteraemia, with a focus on the proportion of coagulase-negative Staphylococcus (CoNS) as compared to Staphylococcus aureus bacteraemia, and the prognosis.
All patients with significant Staphylococcus bacteraemia at the university hospital in Reims in 2008 were included in the study. Data were retrieved retrospectively from the patient records using a standardized case investigation form. Quantitative variables were compared using the Mann-Whitney U-test and qualitative variables were compared using Fisher's exact test or Pearson's Chi-square test, as appropriate. Bivariate logistic regression was performed on both S. aureus and CoNS bacteraemia. All variables with a p-value of < 0.15 were entered into a multiple logistic regression model.
CoNS represented 31.6% of all strains isolated. The methicillin resistance rate was higher in CoNS (66.1%) than in S. aureus (19.1%) (p < 0.0001). CoNS were more frequently associated with intravascular catheters and neoplastic disease, whereas S. aureus was associated with chronic renal failure (p < 0.0001) and diabetes mellitus (p = 0.004). Mortality was 30.7% for S. aureus and 19.6% for CoNS bacteraemia (p = 0.12). Methicillin resistance was not associated with mortality (p = 0.99). Factors independently associated with mortality in CoNS and S. aureus bacteraemia were age and acute renal failure. The presence of severe sepsis/septic shock was only associated with mortality in S. aureus bacteraemia.
CoNS represent one third of Staphylococcus bacteraemia. The mortality difference between CoNS and S. aureus bacteraemia was not statistically significant. Acute renal failure is associated with mortality in both S. aureus and CoNS bacteraemia.
[show abstract][hide abstract] ABSTRACT: qnr genes are plasmid-mediated quinolone resistance genes mainly harbored on large conjugative multiresistant plasmids. The qnrD gene was recently observed in Salmonella enterica on a small nonconjugative plasmid (p2007057). We describe two strains of Providencia rettgeri harboring qnrD on nonconjugative plasmids. The plasmids were 99% identical, with 2,683 bp and four open reading frames, including qnrD, but exhibited only 53% identity with the plasmid found in S. enterica.
Antimicrobial Agents and Chemotherapy 01/2012; 56(1):565-8. · 4.57 Impact Factor
[show abstract][hide abstract] ABSTRACT: Here we report a case of sustentorial brain abscess due to Listeria monocytogenes. Blood culture and procalcitonine blood measurement were negative. L. monocytogenes was isolated from CSF after inoculation in Castañeda medium.
[show abstract][hide abstract] ABSTRACT: In the present study, we comparatively assessed the pathophysiological mechanisms developed during lung infection of BALB/C female mice infected by an original wild type Klebsiella pneumoniae subsp. ozaenae strain (CH137) or by a referent subspecies K. pneumoniae. subsp. pneumoniae strain (ATCC10031). The mice infected with 2.10⁶ CFU K. p. subsp. pneumoniae (n = 10) showed transient signs of infection and all of them recovered. All of those infected with 1.10⁶ CFU K. p. subsp. ozaenae (n = 10) developed pneumonia within 24 h and died between 48 and 72 h. Few macrophages, numerous polymorphonuclear cells and lymphocytes were observed in their lungs in opposite to K. p. subsp. pneumoniae. In bronchoalveolar lavage, a significant increase in MIP-2, IL-6, KC and MCP-1 levels was only observed in K. p. subsp. ozaenae infected mice whereas high levels of TNF-α were evidenced with the two subspecies. Our findings indicated a lethal effect of a wild type K. p. subsp. ozaenae strain by acute pneumonia reflecting an insufficient alveolar macrophage response. This model might be of a major interest to comparatively explore the pathogenicity of K. p. subsp ozaenae strains and to further explore the physiopathological mechanisms of gram-negative bacteria induced human pneumonia.
Microbes and Infection 06/2011; 13(12-13):1045-51. · 2.92 Impact Factor
[show abstract][hide abstract] ABSTRACT: Plasmid-mediated quinolone resistance genes in clinical strains cannot be detected by phenotypic traits but require gene detection. We developed a multiplex real-time polymerase chain reaction (PCR) assay using high-resolution melting master mix with ResoLight dye to detect qnr genes and a simplex real-time PCR assay using SYBR Green I to detect qepA genes. Using qnr-positive and qepA1-positive control strains, the ResoLight method was able to rapidly identify qnrA, qnrB, qnrS, qnrC, and qnrD genes; the SYBR Green I method identified qepA genes. Among 118 extended spectrum beta-lactamase-producing Enterobacteriaceae isolates, the 2 new assays efficiently detected and identified qnr in 9 strains, but no qepA gene. To our knowledge, this is the first study describing the detection of all 5 qnr and qepA genes using real-time PCR. The 2 tests constitute a significant step forward for screening for plasmid quinolone resistance genes in clinical strains.