Evidence of Sexual Transmission of Extended-Spectrum ß-Lactamase-Producing Enterobacterales: A Cross-sectional and Prospective Study.

BACKGROUND: Extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-E) represent a major threat to public health. Little is known on their potential for sexual transmission. METHODS: We recruited individuals at a sexually transmitted infection and human immunodeficiency virus (HIV) outpatient clinic in Paris, France, in whom we evaluated the prevalence of ESBL-E intestinal carriage and, among those testing positive, the proportion with clearance 6 months thereafter. We compared carriage prevalence between groups using logistic regression adjusted for age, geographic origin, travel outside Europe, and antibiotic use in the past 6 months. RESULTS: A total of 2157 individuals participated, of whom 226 (10.5%) were ESBL-E carriers. The proportions of ESBL-E carriers varied across sexual groups and were as follows: HIV-negative men who have sex with men (MSM) and who were on preexposure prophylaxis (PrEP), 16.3% (41 of 251); HIV-negative MSM not on PrEP, 9.7% (47 of 487); HIV-positive MSM, 12.2% (61 of 500); HIV-negative men who have sex exclusively with women, 10.0% (44 of 439); and HIV-negative women who have sex with men, 6.9% (n = 33 of 480). After adjustment, ESBL-E prevalence was significantly higher in HIV-negative MSM on PrEP (P < .001) and HIV-positive MSM (P = .01) than in women who have sex with men. A higher number of sexual partners in the past 6 months was associated with ESBL-E carriage after adjustment (P = .004). Escherichia coli sequence type 14 and blaSHV-12-producing ESBL-E were observed only in MSM. Of 102 individuals with ESBL-E returning for testing, 26 (25%) had carriage at 6 months. CONCLUSION: ESBL-E carriage is more frequent in MSM undergoing PrEP or living with HIV and with increasing number of sexual partners. More research is warranted to understand the consequences of ESBL-E carriage in these populations and how transmission can be reduced.

Prevalence of Antimicrobial Resistance and Infectious Diseases in a Hospitalised Migrant Population in Paris, France, a Retrospective Study.

Objectives: The aim of this study was to estimate the prevalence of anti-microbial resistance (AMR) carriage and its risk factors in hospitalized migrants. Additionally, the prevalence of infectious diseases was evaluated, as well as symptoms of psychological trauma. Methods: We conducted a retrospective monocentric cross-sectional study including all migrant patients recently arrived and hospitalised over a one-year period. Results: Among 101 patients, seventy-nine percent originated from Sub-Saharan Africa. The overall AMR carriage rate was 20.7% [95% CI: 12.4; 28.9%]. We isolated 5/92 methicillin-resistant Staphylococcus aureus strains (5.4%) and 15/92 extended-spectrum beta-lactamase-producing Enterobacteriaceae (16.4%). AMR carriage was associated with older age, region of origin and length of migration. Rates of HIV, HBV, and HCV infection were 39.6%, 32.7%, and 5%, reflecting sampling bias linked to reasons for hospitalization. Eleven percent had serological evidence of treponemasis and 7.8% had Chlamydia trachomatis infection. Symptoms of depression or post-traumatic stress disorder were observed for more than half the patients. Conclusion: It appears essential to offer a systematic and comprehensive post-arrival screening of AMR carriage, infectious diseases and psychological trauma to subjects who experienced migration.

Spontaneous and postsurgical/traumatic Klebsiella pneumoniae meningitis: two distinct clinico-microbiological entities.

OBJECTIVES: To analyze and compare the characteristics and outcomes of spontaneous meningitis (SM) versus postsurgical/traumatic meningitis (PSTM) due to Klebsiella pneumoniae. METHODS: A retrospective multicentric cohort study of all K. pneumoniae meningitis cases managed between January 2007 and May 2018 was carried out in seven university hospitals in the Paris area. The microbiological characteristics of 16 available K. pneumoniae isolates were further analyzed, and the genomes of seven of those isolated from SM were sequenced. RESULTS: Among 35 cases, 10 were SM and 25 were PSTM. SM cases more severe than PSTM cases, with higher septic shock (p = 0.004) and in-hospital mortality rates (p = 0.004). In contrast, relapse occurred in five patients from the PSTM group versus no patients from the SM group. All K. pneumoniae strains recovered from SM but none of those recovered from PSTM displayed hypervirulent phenotypic (positive string test) and genotypic (genes corresponding to capsular serotypes K1 or K2; virulence genes rmpA and iutA) characteristics (p < 0.0001). PSTM tended to be more frequently polymicrobial (p = 0.08) and caused by an extended-spectrum ß-lactamase producing strain (p = 0.08) than SM. CONCLUSIONS: SM and PSTM are two entities differing both from a clinical and a microbiological standpoint. SM appears to be a more serious infection, induced by hypervirulent K. pneumoniae strains.

The Diversity of Lipopolysaccharide (O) and Capsular Polysaccharide (K) Antigens of Invasive Klebsiella pneumoniae in a Multi-Country Collection.

Klebsiella pneumoniae is a common cause of sepsis and is particularly associated with healthcare-associated infections. New strategies are needed to prevent or treat infections due to the emergence of multi-drug resistant K. pneumoniae. The goal of this study was to determine the diversity and distribution of O (lipopolysaccharide) and K (capsular polysaccharide) antigens on a large (>500) global collection of K. pneumoniae strains isolated from blood to inform vaccine development efforts. A total of 645 K. pneumoniae isolates were collected from the blood of patients in 13 countries during 2005-2017. Antibiotic susceptibility was determined using the Kirby-Bauer disk diffusion method. O antigen types including the presence of modified O galactan types were determined by PCR. K types were determined by multiplex PCR and wzi capsular typing. Sequence types of isolates were determined by multilocus sequence typing (MLST) targeting seven housekeeping genes. Among 591 isolates tested for antimicrobial resistance, we observed that 19.3% of isolates were non-susceptible to carbapenems and 62.1% of isolates were multidrug resistant (from as low as 16% in Sweden to 94% in Pakistan). Among 645 isolates, four serotypes, O1, O2, O3, and O5, accounted for 90.1% of K. pneumoniae strains. Serotype O1 was associated with multidrug resistance. Fifty percent of 199 tested O1 and O2 strains were gmlABC-positive, indicating the presence of the modified polysaccharide subunit D-galactan III. The most common K type was K2 by both multiplex PCR and wzi capsular typing. Of 39 strains tested by MLST, 36 strains were assigned to 26 known sequence types of which ST14, ST25, and ST258 were the most common. Given the limited number of O antigen types, diverse K antigen types and the high multidrug resistance, we believe that an O antigen-based vaccine would offer an excellent prophylactic strategy to prevent K. pneumoniae invasive infection.

Spread of NDM-5 and OXA-181 Carbapenemase-Producing Escherichia coli in Chad.

We detected for the first time blaNDM-5 and blaOXA-181 in Escherichia coli isolates from hospitalized patients and healthy volunteers in Chad. These resistance genes were located on IncX3 and IncF plasmids. Despite the large diversity of E. coli clones, the identified resistant intestinal isolates belonged mainly to the same sequence type.

Synergistic in vitro activity between aztreonam and amoxicillin-clavulanate against Enterobacteriaceae-producing class B and/or class D carbapenemases with or without extended-spectrum ß-lactamases.

Introduction. Carbapenemase-producing Enterobacteriaceae have become a major public health concern over the last decade and treatment options are limited.Aims. We evaluated the synergistic activity of the combination of aztreonam (ATM) and clavulanate for 41 ß-lactam-resistant clinical isolates harbouring class B or/and class D carbapenemases combined or not with extended-spectrum ß-lactamases (ESBLs).Methodology. The MICs of ATM, with and without amoxicillin-clavulanate (AMC), were determined. Time-kill assays were performed for three representative strains.Results. The ATM-AMC combination had a synergistic effect on 34/41 (83 %) isolates. The MIC of ATM, in the presence of clavulanate, was ≤1 mg l-1 for 15/41 (37 %) isolates and ≤4 mg l-1 for 29/41 (71 %) isolates. Synergistic activity was observed for 34/37 (92 %) isolates producing ESBLs and carbapenemases, compared to 0/4 (0 %) for ESBL-negative strains. Complete or partial bactericidal activity was obtained when the MIC of the combination was 0.5 mg l-1 and 1.5 mg l-1 or 8 mg l-1, respectively.Conclusion. The combination of ATM and AMC could be an attractive unconventional treatment for infections due to carbapenemase- and ESBL-producing Enterobacteriaceae.

Novel patterns in the molecular epidemiology of KPC-producing Klebsiella pneumoniae in Tucumán, Argentina.

BACKGROUND: In Argentina, there has been an abrupt increase in KPC-2-producing Klebsiella pneumoniae (K. pneumoniae). Tucumán is a multi-border area, so the rapid dissemination of carbapenem-resistant K. pneumoniae is a clinically relevant problem for the region. OBJECTIVES: This study aimed to investigate the epidemiological and molecular patterns of KPC-producing K. pneumoniae clinical isolates collected from different hospitals in Tucumán. METHODS: Carbapenem-resistant K. pneumoniae strains were sequentially and uniquely collected during two time periods. Antibiotic susceptibility was determined by the automated Vitex 2® system and using the standard agar dilution test. Multilocus sequence typing and pulsed-field electrophoresis were used for epidemiological analysis. The genetic structures around blaKPC and the encoding genes of extended-spectrum ß-lactamases were detected by polymerase chain reaction and sequencing. Plasmids were analysed by conjugation and using the plasmid relaxase gene-typing method. RESULTS: All 37 isolates were multidrug resistant, and theblaKPC-2 gene was confirmed in all of them. In 17 isolates (45.9%), the blaCTX-M-2 gene was also amplified, as well as blaSHV-2 in five isolates (13.5%) and blaCTX-M-2/blaSHV-2 in four isolates (10.8%). The molecular epidemiology of the blaKPC-2 gene has resulted in it being associated with an IncL/M transferable plasmid disseminating in various sequence types (STs) (ST17, ST556, ST342, ST147, ST461, ST65, ST15 and ST70), and in a new genetic environment with a 764-bp deletion in the ISKpn7-blaKPC region. CONCLUSIONS: These findings contribute to the understanding of the great diversity of the blaKPC-2-carrying genetic platforms.

Molecular epidemiology of ESBL-producing E. coli and K. pneumoniae: establishing virulence clusters.

OBJECTIVE: To genetically characterize clusters of virulence factors (VFs) among extended spectrum ß-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae and assess whether these clusters are associated with genetic determinants or clinical outcomes. METHODS: One hundred forty-eight E. coli and 82 K. pneumoniae clinical isolates were obtained from 213 patients in Paris, France. Isolates underwent ESBL characterization, MultiLocus Sequence Typing (MLST) typing and phylogenetic group identification. Detection of ten E. coli and seven K. pneumoniae VF-encoding genes were assessed, from which a k-medians partition algorithm with Jaccard similarity measure was used to construct clusters. RESULTS: CTX-M was the predominant ESBL and susceptibility to trimethoprim-sulfamethoxazole (32%), ciprofloxacin (22%) and aminoglycosides (32%) was low. In E. coli, there were five identified clusters, with significantly different distributions of ESBL-sequence type (P<0.001), ST131 (P<0.001) and phylogenetic group (P=0.001) between clusters. "Siderophore exclusive", "siderophore exclusive with iroN " and "adhesin sfa/papGIII-rich" clusters had higher 12-month mortality rates compared to others (49% vs 22%, respectively, P=0.02). In K. pneumoniae, three different clusters, with significantly different distributions of aminoglycoside-sensitivity (P<0.004), MLST-type (P<0.001) and relaxase plasmids (P=0.001) were described. CONCLUSION: Distinct clusters of E. coli and K. pneumoniae VFs are observed within ESBL-producing isolates and are strongly associated with several genetic determinants. Their association with overall morbidity and mortality requires further evidence.

Chlamydia trachomatis screening in urine among asymptomatic men attending an STI clinic in Paris: a cross-sectional study.

BACKGROUND: The incidence of Chlamydia trachomatis (Ct) urethritis has been increasing for the past 10 years. There is little data regarding the screening of Ct infection in asymptomatic men in France, despite the national recommendation to screen at-risk asymptomatic men under 30 attending Sexually Transmitted Infections (STI) clinics. Recent data from the French surveillance network Rénachla show indeed that systematic screening is still focused on women. The objective of our study was to determine the prevalence and risk factors for Ct infection in asymptomatic men under 30 attending an STI clinic located in Paris, France. METHODS: We performed a cross-sectional study between April 4, and December 31, 2016 in the database of the software DIAMM Client V8 used in our STI clinic. We extracted the demographic characteristics, sexual behavior and result of STI screening of all asymptomatic men who had consulted and given their consent for the use of their personal data. Those data were collected in usual care through a standardized questionnaire filled in during an appointment with a trained physician. STI screening was performed using PCR kit CT/NG Abbott Realtime® on first void urines. For MSM, a rectal swab was also collected. Risk factors for Ct infection were analyzed by univariate and multivariate modeling using STATA software 8.2. RESULTS: Among 872 men who had attended the clinic, 647 were included and 37 (5.7, 95% CI 4.2 to 7.8) were positive for Ct in urine. In univariate analysis, men who had unprotected sex in the last 6 weeks (OR 2.40 (95%CI 1.16 to 4.94), p = 0.02), and those who had an infected partner (OR 7.6 (95%CI 3.03 to 20), p = 0.0001) were more likely to be infected. In the multivariate analysis having an infected partner was the only risk factor (OR 11.1(95% CI 3.7 to 33.3), p = 0.0001) that remained significant. CONCLUSION: Prevalence of Ct infection is high among asymptomatic men of 30 years or less attending our urban STI clinic especially among those with an infected partner. The Ct screening among this population associated with partner notification, as recommended by the French national guidelines, should be more widely implemented.

Biofilm formation by ESBL-producing strains of Escherichia coli and Klebsiella pneumoniae.

OBJECTIVES: Biofilm production in extended spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae provides a favourable environment for the exchange of antibiotic-resistance genes and could facilitate widespread dissemination. We aimed to assess biofilm development in ESBL-producing E. coli and K. pneumoniae isolates and determine how development relates to microbiological characteristics and clinical outcomes. METHODS: 147 ESBL-producing E. coli and 82 K. pneumoniae were genetically characterized. Biofilm formation was measured at 1.5, 4, 6, and 24 h during culture in blood heart infusion using a microbead immobilization assay (BioFilm Ring test®). Results were given as biofilm formation index (BFI) with lower values indicating increased presence of biofilm (range = 0-21). RESULTS: In total, 57.1% of strains were strong producers of biofilm (BFI < 2), whereas 13.4% lacked biofilm production (BFI > 18). Standard biofilm production (BFI < 7) was common in E. coli isolates (61.9%). For E. coli, biofilm production was less frequently observed in ST131 clones (p = 0.03) but more frequently in strains harbouring toxin (p = 0.008) or adhesin (p = 0.008) virulence factor genes. Despite almost all K. pneumoniae having standard biofilm production (90.2%), there was a 2.4-times higher odds of observing biofilm in ST29/147/323 versus other ST-types (p = 0.13). Patients with standard biofilm producing isolates were not at increased risk of transfer to intensive-care (odds-ratio=2.80, 95%CI=0.59-13.21) or death within 12-months (odds-ratio=1.61, 95%CI=0.75-3.43). CONCLUSION: In these ESBL-producing strains, biofilm production is linked to certain virulence factors in E. coli and is common in K. pneumoniae. Further exploration of whether biofilm production increases dissemination and risk of severe clinical outcomes is needed in larger collections of isolates.

Extended-spectrum ß-lactamase-encoding genes are spreading on a wide range of Escherichia coli plasmids existing prior to the use of third-generation cephalosporins.

To understand the evolutionary dynamics of extended-spectrum ß-lactamase (ESBL)-encoding genes in Escherichia coli, we undertook a comparative genomic analysis of 116 whole plasmid sequences of human or animal origin isolated over a period spanning before and after the use of third-generation cephalosporins (3GCs) using a gene-sharing network approach. The plasmids included 82 conjugative, 22 mobilizable and 9 non-transferable plasmids and 3 P-like bacteriophages. ESBL-encoding genes were found on 64 conjugative, 6 mobilizable, 2 non-transferable plasmids and 2 P1-like bacteriophages, indicating that these last three types of mobile elements also play a role, albeit modest, in the diffusion of the ESBLs. The network analysis showed that the plasmids clustered according to their genome backbone type, but not by origin or period of isolation or by antibiotic-resistance type, including type of ESBL-encoding gene. There was no association between the type of plasmid and the phylogenetic history of the parental strains. Finer scale analysis of the more abundant clusters IncF and IncI1 showed that ESBL-encoding plasmids and plasmids isolated before the use of 3GCs had the same diversity and phylogenetic history, and that acquisition of ESBL-encoding genes had occurred during multiple independent events. Moreover, the blaCTX-M-15 gene, unlike other CTX-M genes, was inserted at a hot spot in a blaTEM-1-Tn2 transposon. These findings showed that ESBL-encoding genes have arrived on wide range of pre-existing plasmids and that the successful spread of blaCTX-M-15 seems to be favoured by the presence of well-adapted IncF plasmids that carry a Tn2-blaTEM-1 transposon.

Hypervirulent Klebsiella pneumoniae, a 5-year study in a French ICU.

PURPOSE: Hypervirulent Klebsiella pneumoniae (hvKp) has emerged as a leading cause of severe community-acquired pneumonia, liver abscess and disseminated infection in the Far East. Data regarding the incidence, clinical features and microbiological characteristics related to hvKp infections in the Western world are scarce. METHODOLOGY: The incidence, clinical features and microbiological characteristics of hvKp infections were investigated through a 5-year survey conducted in a single French intensive care unit. K. pneumoniae strains were screened for hypermucoviscosity based on a string test. Multilocus sequence typing and multiplex PCR analysis targeting virulence genes were performed on string test-positive strains. RESULTS: Over a 53-month period, a total of 59 infections due to K. pneumoniae were identified including 26 community-onset infections. Twelve hvKp infections were documented, accounting for 46.1 % of community-acquired K. pneumoniae. Community-acquired pneumonia (n=6), aspiration pneumonia (n=4) and liver abscess (n=2) represented initial sites and mode of infection. Compared to non-hvKp infections, patients with hvKp infections displayed higher rates of multi-organ failure (83.3 % vs 35.7 %; P=0.04), but mortality rates were not different (50 % vs 35 %; P=0.71). Strains K1/ST23 (n=5) and K2/ST86 (n=5) predominated. All hvKp strains displayed wild-type susceptibility. CONCLUSION: hvKp represent a potentially underestimated cause of fatal infections in the Western world.

Within-a-Day Detection and Rapid Characterization of Carbapenemase by Use of a New Carbapenem Inactivation Method-Based Test, CIMplus.

The dissemination of carbapenemase-producing Enterobacteriaceae (CPE) is a major threat to public health. Rapid and accurate detection of CPE is essential for initiating appropriate antimicrobial treatment and establishing infection control measures. The carbapenem inactivation method (CIM), which has good sensitivity and specificity but a detection time of 20 h, was recently described. In this study, we evaluated the performances of a new version, the CIMplus test, which allows detection of carbapenemases in 8 h and characterization of carbapenemase classes, according to the Ambler classification, in 20 h. A panel of 110 carbapenem-resistant Enterobacteriaceae strains, including 92 CPE strains (with NDM, VIM, IMP, KPC, GES, OXA-48, and OXA-48-like enzymes), was used to evaluate test performance. Carbapenemase activity was detected at 8 h and 20 h. Characterization of carbapenemase classes, using specific inhibitors, was possible in 20 h. The CIMplus test had sensitivities of 95.7% and 97.8% at 8 h and 20 h, respectively, and a specificity of 94.4%, independent of the culture duration. Using a decision algorithm, this test was successful in identifying the carbapenemase class for 98.9% of tested CPE isolates (87/88 isolates). In total, the characterization was correct for 100%, 96.9%, and 100% of Ambler class A, B, and D isolates, respectively. Therefore, this test allows detection of carbapenemase activity in 8 h and characterization of carbapenemase classes, according to the Ambler classification, in 20 h. The CIMplus test represents a simple, affordable, easy-to-read, and accurate tool that can be used without any specific equipment.

A Long-Term Study of the Diversity of OXA-48-Like Carbapenemase-Producing Bacterial Strains in Infected Patients and Carriers.

PURPOSE: Since their emergence at the beginning of the century, OXA-48 carbapenemases have spread in the community and in hospitals. To assess the diversity of OXA-48-producing bacterial strains and plasmids in the hospital setting, we studied the strains isolated from patients in three hospitals in the Paris area. MATERIALS AND METHODS: All possible OXA-48-like strains were included in the study. OXA-48-like and extended-spectrum beta-lactamase-encoding genes were identified, and fingerprinting analysis was performed for all Escherichia coli and Klebsiella pneumoniae strains. The backbones and close genetic environments of blaOXA-48 were assessed by amplifying genes that were specific to the pOXA-48a plasmid and PCR, encompassing the junctions between blaOXA-48 and its direct genetic environment. RESULTS: Overall, 68 strains from 30 patients were studied. These strains belonged to seven different enterobacterial species. OXA-48, OXA-204, and OXA-401 were identified in 62, 3, and 3 isolates, respectively. Additional broad-spectrum beta-lactamases were identified in 34% (23/68) of the strains. The strain diversity was high between and within patients. Identical patterns were observed only within individual patients or among epidemiologically related patients. Plasmid mapping was performed in the 62 OXA-48-producing strains and the 3 OXA-405-producing strains, resulting in the identification of 5 different patterns. CONCLUSION: Because of their ability to transfer between strains, OXA-48 carbapenemases have a high risk of dissemination and may become endemic in France.

Emergence of Plasmid-Mediated Fosfomycin-Resistance Genes among Escherichia coli Isolates, France.

FosA, a glutathione S-transferase that inactivates fosfomycin, has been reported as the cause of enzymatic resistance to fosfomycin. We show that multiple lineages of FosA-producing extended spectrum ß-lactamase Escherichia coli have circulated in France since 2012, potentially reducing the efficacy of fosfomycin in treating infections with antimicrobial drug-resistant gram-negative bacilli.