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Objectives In some countries, third-generation cephalosporins (3GCs) serve as first-line therapy in children with urinary tract infections (UTIs). However, their use may contribute to the emergence of antibiotic resistance, notably among Gram-negative bacteria (GNB). Integrons are bacterial genetic elements involved in antibiotic resistance in GNB. Their absence is associated with >97% susceptibility to trimethoprim/sulfamethoxazole in adults infected with GNB. The objective of this study was to examine the value of integron detection directly from urine samples as a predictive marker of resistance to trimethoprim/sulfamethoxazole in children with GNB-related UTIs. Methods Children admitted to the Limoges University Hospital’s paediatric emergency department between February 2018 and March 2019 with a suspicion of UTI were eligible for the study. Only confirmed cases presenting a positive urine culture with unique GNB were retained for further study analyses. Integrons were detected directly from urines using real-time PCR. Results The data of 72 patients were analysed and integrons were detected in 15 urine samples. The negative predictive value of integron detection for resistance to trimethoprim/sulfamethoxazole was 100% as all of the GNB (all were Enterobacterales) isolated from patients with no integrons detected in their urine samples were susceptible to trimethoprim/sulfamethoxazole. Conclusions The detection of integrons in cases of paediatric patients with suspected UTI could help limit 3GC empirical use and empower an empirical first-line strategy better tailored to the needs of each patient.

Background Considering the increase in MDR Gram-negative bacteria (GNB), the choice of empirical antibiotic therapy is challenging. In parallel, use of broad-spectrum antibiotics should be avoided to decrease antibiotic selection pressure. Accordingly, clinicians need rapid diagnostic tools to narrow antibiotic therapy. Class 1–3 integrons, identified by intI1–3 genes, are genetic elements that play a major role in antibiotic resistance in GNB. Objectives The objective of the IRIS study was to evaluate the negative and positive predictive values (NPVs and PPVs, respectively) of intI1–3 as markers of antibiotic resistance. Methods The IRIS study was an observational cross-sectional multicentre study that enrolled adult subjects with suspected urinary tract or intra-abdominal infections. intI1–3 were detected directly from routinely collected biological samples (blood, urine or intra-abdominal fluid) using real-time PCR. A patient was considered ‘MDR positive’ if at least one GNB, expressing acquired resistance to at least two antibiotic families among β-lactams, aminoglycosides, fluoroquinolones and/or co-trimoxazole, was isolated from at least one biological sample. Results Over a 2 year period, 513 subjects were enrolled and 409 had GNB documentation, mostly Enterobacterales. intI1 and/or intI2 were detected in 31.8% of patients and 24.4% of patients were considered ‘MDR positive’. The NPV of intI1 and/or intI2 as a marker of acquired antibiotic resistances was estimated at 92.8% (89.1%–95.5%). The NPVs for first-line antibiotics were all above 92%, notably >96% for resistance to third-generation cephalosporins. Conclusions The IRIS study strongly suggests that the absence of intI1 and intI2 in biological samples from patients with GNB-related infections is predictive of the absence of acquired resistances.

OBJECTIVES Our aim was to confirm with a large panel of clinical isolates that class 2 integrons are highly prevalent in Proteae and to analyse their genetic characteristics. METHODS Proteae (Proteus spp., Morganella spp. and Providencia spp.) isolates were collected from clinical samples during 2013 at Limoges University Hospital, France. The presence of class 1, 2 and 3 integrons was investigated by quantitative PCR. The presence of a stop codon in the intI2 gene was determined by Sanger sequencing. The gene cassette arrays of class 2 integrons were determined by PCR-RFLP and Sanger sequencing or next-generation sequencing when needed. RESULTS Of the 327 Proteae collected, 103 (31.5%) harboured a class 2 integron and 45 (13.8%) a class 1 integron. No class 3 integrons were detected. One functional IntI2 integrase was detected in a Morganella morganii isolate. Six different gene cassette arrays were detected. Four had already been described in the literature: dfrA1-sat2-aadA1 (72 isolates), dfrA1-catB2-sat2-aadA1 (17), sat2-aadA1 (6) and lnu(F), dfrA1, aadA1 (1). We identified two new gene cassette arrays: (i) a new variant of the dfrA1 gene cassette (one isolate; the one with the functional IntI2); and (ii) the array dfrA1-gcu115-sat2 harbouring the new gcu115 gene cassette with two ORFs encoding proteins of unknown functions (five isolates). CONCLUSIONS We showed a high frequency of class 2 integrons, as well as a diversity of gene cassette arrays, among Proteae. This work highlights that the Proteae tribe plays an important role as a reservoir of class 2 integrons.

Totally implanted venous access ports (TIVAPs) are commonly used implants for the management of acute or chronic pathologies. Although their use improves the patient’s health care and quality of life, they are associated with a risk of infection and subsequent clinical complications, often leading to implant removal. While all TIVAPs appear to be colonized, only a fraction become infected, and the relationship between nonpathogenic organisms colonizing TIVAPs and the potential risk of infection is unknown. We explored bacteria present on TIVAPs implanted in patients with or without signs of TIVAP infection and identified differences in phylum composition and community structure. Our data suggest that the microbial ecology of intravascular devices could be predictive of TIVAP infection status and that ultimately a microbial ecological signature could be identified as a tool to predict TIVAP infection susceptibility and improve clinical management., Totally implanted venous access ports (TIVAPs) are commonly used catheters for the management of acute or chronic pathologies. Although these devices improve health care, repeated use of this type of device for venous access over long periods of time is also associated with risk of colonization and infection by pathogenic bacteria, often originating from skin. However, although the skin microbiota is composed of both pathogenic and nonpathogenic bacteria, the extent and the consequences of TIVAP colonization by nonpathogenic bacteria have rarely been studied. Here, we used culture-dependent and 16S rRNA gene-based culture-independent approaches to identify differences in bacterial colonization of TIVAPs obtained from two French hospitals. To explore the relationships between nonpathogenic organisms colonizing TIVAPs and the potential risk of infection, we analyzed the bacterial community parameters between TIVAPs suspected (symptomatic) or not (asymptomatic) of infection. Although we did not find a particular species assemblage or community marker to distinguish infection risk on an individual sample level, we identified differences in bacterial community composition, diversity, and structure between clinically symptomatic and asymptomatic TIVAPs that could be explored further. This study therefore provides a new view of bacterial communities and colonization patterns in intravascular TIVAPs and suggests that microbial ecology approaches could improve our understanding of device-associated infections and could be a prognostic tool to monitor the evolution of bacterial communities in implants and their potential susceptibility to infections. IMPORTANCE Totally implanted venous access ports (TIVAPs) are commonly used implants for the management of acute or chronic pathologies. Although their use improves the patient’s health care and quality of life, they are associated with a risk of infection and subsequent clinical complications, often leading to implant removal. While all TIVAPs appear to be colonized, only a fraction become infected, and the relationship between nonpathogenic organisms colonizing TIVAPs and the potential risk of infection is unknown. We explored bacteria present on TIVAPs implanted in patients with or without signs of TIVAP infection and identified differences in phylum composition and community structure. Our data suggest that the microbial ecology of intravascular devices could be predictive of TIVAP infection status and that ultimately a microbial ecological signature could be identified as a tool to predict TIVAP infection susceptibility and improve clinical management.

Objectives: Dissemination of antimicrobial resistance (AMR) is a global issue that requires the adoption of a “One-Health” approach promoting integration of human and animal health. Besides culture-dependent techniques frequently used for AMR surveillance, cultivation-independent methods can give additional insights into the diversity and reservoir of AMR genetic determinants. Integrons are molecular markers that can provide overall and reliable estimation of AMR dissemination. In this study, considering the “One-Health” approach, we have analyzed the integron digestive carriage from stools of humans and cattle living in a same area and exposed to different antibiotic selection pressures. Methods: Three collections of human [general population (GP) and intensive care unit patients (ICUs)] and bovine (BOV) stool samples were analyzed. The three main classes of integrons were detected using a multiplex qPCR both from total DNA extracted from stools, and from Gram-negative bacteria obtained by culture after an enrichment step. Results: With the cultivation-independent approach, integron carriage was 43.8, 52.7, and 65.6% for GP, ICU, and BOV respectively, percentages being at least twofold higher to those obtained with the cultivation-dependent approach. Class 1 integrons were the most prevalent; class 2 integrons seemed more associated to cattle than to humans; no class 3 integron was detected. The integron carriage was not significantly different between GP and ICU populations according to the antibiotic consumption, whatever the approach. Conclusion: The cultivation-independent approach constitutes a complementary exploratory method to investigate the integron digestive carriage of humans and bovines, notably within subjects under antibiotic treatment. The high frequency of carriage of integrons in the gut is of clinical significance, integrons being able to easily acquire and exchange resistant genes under antibiotic selective pressure and so leading to the dissemination of resistant bacteria.

Background Staphylococcus aureus (SA) is a leading cause of infectious diseases in sports teams. In recent decades, community-associated SA (CA-SA) strains have emerged worldwide and have been responsible for outbreaks in sports teams. There are very few data on the prevalence of these strains in France, and none on the carriage among athletes. Methods We conducted a cross-sectional study to determine the SA carriage proportion among athletes practicing sports at risk for CA-SA infection in a French county, and determined the methicillin-resistant and/or CA-SA proportion. We also analyzed SA carriage according to risks factors and studied the SA clonality in a sample of our population. Results We included 300 athletes; SA carriage proportion was 61% (n = 183) and one was MRSA carrier (0.33%). The MRSA strain belonged to the clonal complex ST5. None of the strain produced Panton Valentine Leucocidin, and we did not find clonal distribution within the teams. Interestingly, we found a high throat-only carriage (n = 57), 31.1% of the SA carriers. Conclusion We found a high SA carriage with a local epidemiology quite different than that reported in a similar population in the USA. Further studies on SA carriage should include throat sampling. Trial registration The approved protocol was registered on ClinicalTrial.gov , NCT01148485.

To assess any differences between the initial and midstream urine samples from nontoilet-trained, uncircumcised boys. Contamination during urine collection makes the diagnosis of urinary tract infections (UTIs) difficult in nontoilet-trained children, especially in uncircumcised boys. Whether the contamination comes mainly from the initial stream or the contact between urine and perineal skin is not known in this population.A prospective diagnostic study between early and midstream urine samples was conducted on asymptomatic patients with no suspicion of UTI. The clean void method was performed in nontoilet-trained boys under general anesthesia. The exclusion criteria were circumcision, older than 3 years of age, recent antibiotics treatment, and recent UTI. Urinalysis and urine culture were performed, allowing a comparison between early and midstream urine samples.Forty-four patients were enrolled in the study, and 31 satisfactory samples were obtained. A higher contamination rate was found in the early stream (n = 16; 51%) than in the midstream (n = 5; 16%) (P .01). The positive culture from the early stream sample was statistically associated with a lower age (P = .02). The contamination rate of the first stream is 3-fold higher than for the midstream when collecting urine for urine culture.The clean void method in nontoilet-trained, uncircumcised boys provides low-quality urine samples for both early and midstream urine samples. The benefit of catching midstream urine samples for the diagnosis of UTI in this population is even more important when the children are young.

Five consecutive cases of endophthalmitis that developed after cataract extraction by a single surgeon using the same operating room during one morning session are described. Following preoperative topical administration of ciprofloxacin, surgery consisted of phacoemulsification with peristaltic pump and fluid venting, polymethylmethacrylate intraocular lens implantation, and corneal suture. No complications occurred during surgery. All five patients developed endophthalmitis caused by infection with Alcaligenes xylosoxidans in less than 24 hours. Pulsed-field gel electrophoresis was used to prove similarity between strains. Bacterial inquiry on contamination of the operating room environment revealed massive colonization of phacoemulsifier irrigation channels by Pseudomonas fluorescens bacteria from an unestablished source. Four of the five patients ultimately recovered visual acuity better than 20/60.

International audience; OBJECTIVES: Glycopeptides are the drugs of choice to treat infections due to methicillin-resistant Staphylococcus aureus, but since 1995, glycopeptide-intermediate S. aureus (GISA) and heterogeneous GISA (hGISA) have been reported worldwide. Detection of reduced susceptibility to glycopeptides in S. aureus is very difficult in a routine clinical laboratory. The aim of this study was to investigate the prevalence of hGISA/GISA strains using a three-step approach during a 1 year period. METHODS: The following algorithm was adopted: (i) brain heart infusion agar with 4 mg/L teicoplanin was used to screen S. aureus strains for reduced susceptibility to glycopeptides; (ii) for each agar screen-positive strain, an Etest macromethod using modified cut-off values (vancomycin and teicoplanin > or =4 mg/L) was used to detect potential hGISA/GISA; and (iii) the population analysis profile (PAP) method was finally used to confirm the hGISA/GISA phenotype. RESULTS: In total, 2300 strains of S. aureus were screened and 255 (11%) were categorized as hGISA with the PAP method, whereas no GISA strains were detected. Standard MIC values and current MIC breakpoints could not discriminate the hGISA/GISA phenotype from glycopeptide-susceptible S. aureus. Thus laboratories using currently standardized MIC methods cannot be expected to detect S. aureus strains that may exhibit reduced susceptibility to glycopeptides. Molecular typing by PFGE revealed that 238 strains belonged to the same clone. CONCLUSIONS: A clonal hGISA strain has disseminated within our hospital. The method described in this study has to be further investigated to see if it is applicable to other S. aureus strains.

Multidrug-resistant Enterobacteriaceae are a major public health threat and complicate the choice of drugs for empirical antibiotic therapy, especially in sepsis patients who require rapid, appropriate treatment. The objective of this study was to examine the value of integrons as a global predictive marker of acquired antibiotic resistance in septicaemia-causing Enterobacteriaceae by direct detection in positive blood cultures. The integron genetic marker can be detected in a single test, whereas multiple PCRs are needed to detect the hundreds of known antibiotic resistance genes. A total of 166 positive blood cultures were included in the study, and integrons were detected with a quantitative PCR method both in positive blood cultures and isolated Enterobacteriaceae. The results of integron detection directly on positive blood cultures were consistent in 98.8% of cases with integron detection in isolated Enterobacteriaceae. Negative predictive values (NPVs) were >90% for resistance to third-generation cephalosporins, aminoglycosides, ciprofloxacin and trimethoprim/sulfamethoxazole. In the current context of antibiotic stewardship, these good NPVs indicate that this method might be useful for preserving broad-spectrum antibiotics. The results of this proof-of-concept study must be confirmed in order to demonstrate the clinical relevance of integron detection, not only in positive blood cultures but also, to gain time, in raw biological samples.

Ventilator-associated pneumonia (VAP) is the most common infection in critically ill patients. Initial antibiotic therapy is often broad spectrum, which promotes antibiotic resistance so new techniques are under investigation to obtain early microbiological identification and quantification. This trial compares the performance of a new real-time quantitative molecular-based method with conventional culture in patients with suspected VAP. Patients with suspected VAP who were ventilated for at least 48 h were eligible. An endotracheal aspirate (ETA) and a bronchoalveolar lavage (BAL) were performed at each suspected VAP episode. Both samples were analysed by conventional culture and molecular analysis. For the latter, bacterial DNA was extracted from each sample and real-time PCR were run. In all, 120 patients were finally included; 76% (91) were men; median age was 65 years, and clinical pulmonary infection score was ≥6 for 73.5% (86) of patients. A total of 120 BAL and 103 ETA could be processed and culture results above the agreed threshold were obtained for 75.0% (90/120) of BAL and 60.2% (62/103) of ETA. The main isolated bacteria were Staphylococcus aureus, Pseudomonas aeruginosa and Haemophilus influenzae. Performance was 89.2% (83.2%-93.6%) sensitivity and 97.1% (96.1%-97.9%) specificity for BAL samples and 71.8% (61.0%-81.0%) sensitivity and 96.6% (95.4%-97.5%) specificity for ETA samples when the molecular biology method was compared with conventional culture method (chosen as reference standard). This new molecular method can provide reliable quantitative microbiological data and is highly specific with good sensitivity for common pathogens involved in VAP.

International audience; PURPOSE: The purpose of this study is to evaluate the feasibility and interest of real-time polymerase chain reaction (RT-PCR) testing for bacterial genomes in body fluids other than blood in patients with acute severe sepsis. METHODS: Twenty-six consecutive patients admitted for severe sepsis or septic shock were prospectively studied. Body fluids were sampled as clinically indicated and tested using standard microbiological methods and modified RT-PCR methods (universal PCR and specific PCRs). Results of standard microbiological tests were compared with those of PCR tests. RESULTS: Direct RT-PCR testing was successfully performed on all nonblood body fluids. Of 29 body fluids collected, 23 were positive for at least 1 microorganism with conventional tests. Of 18 microbiological tests positive for a single microorganism, 15 fully agreed with RT-PCR assays, and the remaining 3 samples were infected with bacteria not screened by PCR testing. Among the 5 polymicrobial results obtained with conventional tests, RT-PCR agreed in 4 patients. The RT-PCR tests allowed additional clinically relevant bacterial identification in 3 of 6 samples with negative microbiological culture. CONCLUSIONS: Our results indicate that direct PCR testing may improve the detection of bacteria in body fluids other than blood in patients with acute severe sepsis.

Ten enteroinvasive (EIEC) and 25 enteroaggregative (EaggEC) E. coli strains isolated from Senegalese patients were analyzed for their integron content. All strains were resistant to at least two antibiotics. Four EIEC and 15 EaggEC were found to carry a class 1 integron. An identical integron carrying a single dfrA5 cassette, conferring resistance to trimethoprim, was identified in all four EIEC strains. Five EaggEC strains harbored an integron with a single cassette, dfrA7, while the remaining 10 strains carried two integrons, one with a single cassette, aadA1a conferring resistance to streptomycin and spectinomycin, and the second one bearing two cassettes, dfrA13 and oxa5, the later being a beta-lactam resistance cassette. The presence of these integrons is worrying, because trimethoprim is largely used for diarrheal disease therapy in Africa. Thus, the presence of integrons in diarrheagenic strains is of public health importance because a limited number of antibiotics are available in developing countries.

Eighteen of 25 isolates of Salmonella enterica serovar Typhi were multidrug resistant and contained class 1 integrons with a single cassette, dfrVII or aadA1 . The dfrVII -containing integron was likely borne on an IncHI1 plasmid. Salmonella serovar Typhi could become resistant to broad-spectrum cephalosporins by integrating cassettes, such as veb-1 , a common cassette in Asia.