Efficacy of dual carbapenem treatment in a murine sepsis model of infection due to carbapenemase-producing Acinetobacter baumannii.

OBJECTIVES: To evaluate the in vivo efficacy of a dual carbapenem combination containing imipenem plus meropenem against carbapenem-resistant Acinetobacter baumannii producing carbapenemases OXA-23 or OXA-58. METHODS: An experimental model of peritonitis using C57BL/6J female mice was developed and the minimum lethal doses were calculated for infections due to OXA-23 or OXA-58 producers of A. baumannii clinical isolates. The efficacies of the carbapenems in monotherapy and in combination were tested. RESULTS: Meropenem was better than imipenem in mice infected with either of the carbapenem-resistant A. baumannii (CRAb) strains. The combination of meropenem plus imipenem significantly improved the clearance of CRAbs from spleen compared with non-treated groups. The carbapenem-containing combination was better than imipenem for treating mice infected with both carbapenemase producers. In blood, the carbapenem combination significantly decreased the bacterial load of the OXA-23 producers compared with imipenem or meropenem used in monotherapy. CONCLUSIONS: These results suggest that dual carbapenem combination could be an option for the treatment of infections due to carbapenemase-producing A. baumannii such as OXA-23 and OXA-58 producers.

Outbreak of Pseudomonas aeruginosa producing VIM carbapenemase in an intensive care unit and its termination by implementation of waterless patient care.

BACKGROUND: Long-term outbreaks of multidrug-resistant Gram-negative bacilli related to hospital-building water systems have been described. However, successful mitigation strategies have rarely been reported. In particular, environmental disinfection or replacement of contaminated equipment usually failed to eradicate environmental sources of Pseudomonas aeruginosa. METHODS: We report the investigation and termination of an outbreak of P. aeruginosa producing VIM carbapenemase (PA-VIM) in the adult intensive care unit (ICU) of a Swiss tertiary care hospital with active case finding, environmental sampling and whole genome sequencing (WGS) of patient and environmental strains. We also describe the implemented control strategies and their effectiveness on eradication of the environmental reservoir. RESULTS: Between April 2018 and September 2020, 21 patients became either infected or colonized with a PA-VIM strain. For 16 of them, an acquisition in the ICU was suspected. Among 131 environmental samples collected in the ICU, 13 grew PA-VIM in sink traps and drains. WGS confirmed the epidemiological link between clinical and environmental strains and the monoclonal pattern of the outbreak. After removing sinks from patient rooms and implementation of waterless patient care, no new acquisition was detected in the ICU within 8 months after the intervention. DISCUSSION: Implementation of waterless patient care with removal of the sinks in patient rooms was successful for termination of a PA-VIM ICU outbreak linked to multiple environmental water sources. WGS provides highly discriminatory accuracy to investigate environment-related outbreaks.

Dimercaptosuccinic acid in combination with carbapenems against isogenic strains of Escherichia coli producing or not producing a metallo-ß-lactamase in vitro and in murine peritonitis.

BACKGROUND: Carbapenemase-producing Enterobacterales represent a major therapeutic challenge. MBLs, requiring zinc at their catalytic site, could be inhibited by meso-dimercaptosuccinic acid (DMSA), a heavy metal chelator already widely used for treating lead intoxication. OBJECTIVES: To evaluate the activity of carbapenems alone or combined with DMSA against MBL-producing Escherichia coli in a severe murine peritonitis model. METHODS: Isogenic strains of wild-type E. coli CFT073 producing the MBLs NDM-1, VIM-2 and IMP-1, and the control serine carbapenemases OXA-48 and KPC-3 were constructed. MIC determinations and time-kill assays were performed for imipenem, meropenem and ertapenem alone or in combination with DMSA. Infected mice were treated intraperitoneally for 24 h with imipenem, DMSA or their combination. Bacterial counts in peritoneal fluid and spleen were assessed at 24 h. RESULTS: DMSA in combination with each carbapenem caused a significant decrease in the MICs for all MBL-producing strains, in a concentration-dependent manner, but did not provide benefit against non-MBL strains. In mice infected with the NDM-1-producing strain, the combination of imipenem and DMSA significantly reduced bacterial counts in peritoneal fluid (P = 0.0006) and spleen (P < 0.0001), as compared with imipenem alone, with no benefit against the KPC-3-producing and CFT073 strains. DMSA concentrations in plasma of mice were comparable to those obtained in humans with a standard oral dose. CONCLUSIONS: DMSA restores the activity of carbapenems against MBL-producing strains, and its combination with carbapenems appears to be a promising strategy for the treatment of NDM-producing E. coli infections.

Activity of the novel siderophore cephalosporin cefiderocol against multidrug-resistant Gram-negative pathogens.

The novel siderophore cephalosporin cefiderocol (S-649266) with potent activity against Gram-negative pathogens was recently developed (Shionogi & Co., Ltd.). Here, we evaluated the activity of this new molecule and comparators against a collection of previously characterized Gram-negative isolates using broth microdilution panels. A total of 753 clinical multidrug-resistant Gram-negative isolates collected from hospitals worldwide were tested against cefiderocol and antibiotic comparators (ceftolozane-tazobactam [CT], meropenem [MEM], ceftazidime [CAZ], ceftazidime-avibactam [CZA], colistin [CST], aztreonam [ATM], amikacin [AMK], ciprofloxacin [CIP], cefepime [FEP], and tigecycline [TGC]) for their susceptibility. The collection included Escherichia coli (n = 164), Klebsiella pneumoniae (n = 298), Enterobacter sp. (n = 159), Pseudomonas aeruginosa (n = 45), and Acinetobacter baumannii (n = 87). Resistance mechanisms included producers of carbapenemases and extended-spectrum ß-lactamases (ESBLs). In addition, a series of colistin-resistant enterobacterial isolates (n = 74), including 15 MCR-1 producers, were tested. The MIC90 of cefiderocol was 2 mg/L, while those of comparative drugs were >64 mg/L for CT, MEM, CAZ, CZA, and AMK, >32 mg/L for ATM, >16 mg/L for FEP, 8 mg/L for CST, and 2 mg/L for TGC. The MIC50 of cefiderocol was 0.5 mg/L, while those of other drugs were >64 mg/L for CAZ, 64 mg/L for CT, >32 mg/L for ATM, >16 mg/L for FEP, 8 mg/L for MEM and AMK, >4 mg/L for CIP, 1 mg/L for CZA, 0.5 mg/L for TGC, and <0.5 mg/L for CST. Only 20 out of 753 strains showed MIC values of cefiderocol ≥8 µg/mL. Compared to the other drugs tested, cefiderocol was more active, with the exception of colistin and tigecycline showing equivalent activity against certain subgroups of bacteria.

Antimicrobial activity of octenidine against multidrug-resistant Gram-negative pathogens.

Multidrug-resistant (MR) Gram-negative (GN) pathogens pose a major and growing threat for healthcare systems, as therapy of infections is often limited due to the lack of available systemic antibiotics. Well-tolerated antiseptics, such as octenidine dihydrochloride (OCT), may be a very useful tool in infection control to reduce the dissemination of MRGN. This study aimed to investigate the bactericidal activity of OCT against international epidemic clones of MRGN. A set of five different species (Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Acinetobacter baumannii, and Pseudomonas aeruginosa) was studied to prove OCT efficacy without organic load, under "clean conditions" (0.3 g/L albumin) and under "dirty conditions" (3 g/L albumin + 3 mL/L defibrinated sheep blood), according to an official test norm (EN13727). We used five clonally unrelated isolates per species, including a susceptible wild-type strain, and four MRGN isolates, corresponding to either the 3MRGN or 4MRGN definition of multidrug resistance. A contact time of 1 min was fully effective for all isolates by using different OCT concentrations (0.01% and 0.05%), with a bacterial reduction factor of >5 log10 systematically observed. Growth kinetics were determined with two different wild-type strains (A. baumannii and K. pneumoniae), proving a time-dependent efficacy of OCT. These results highlight that OCT may be extremely useful to eradicate emerging highly resistant Gram-negative pathogens associated with nosocomial infections.

Neonatal infections with multidrug-resistant ESBL-producing E. cloacae and K. pneumoniae in Neonatal Units of two different Hospitals in Antananarivo, Madagascar.

BACKGROUND: We investigated the molecular mechanism of ß-lactam resistance in extended-spectrum ß-lactamase (ESBL)-producing Enterobacterial strains isolated in neonatal units of different hospitals in Anatnanarivo, Madagascar. METHODS: Bacteria were identified by standard biochemical methods, disc diffusion antibiograms and Etest. Resistance genes were sought by PCR. Strains were characterized by Rep-PCR (Diversilab), plasmid analysis and rep-typing. RESULTS: From April 2012 to March 2013, 29 ESBL-producing E. cloacae and 15 K. pneumoniae were isolated from blood culture (n = 32) or gastric samples (n = 12) performed at day 0 or 2 from 39/303 newborns suspected of early neonatal infection. These infants were treated with expanded spectrum cephalosporins, due to lack of carbapenems, leading to a high mortality rate (45 %). Isolates recovered were all, but 4, multidrug resistant, particularly to fluoroquinolones (FQ) except for 21 E. cloacae isolates. Isolates produced TEM-1 and CTX-M-15 ß-lactamases and their genes were located on several self-transferable plasmids of variable sizes sizes that could not be linked to a major plasmid incompatibility group. E. cloacae isolates belonged to 6 Rep-types among which two counted for 11 isolates each. The FQ resistant E. cloacae isolates belonged to one clone, whereas the FQ susceptible E. cloacae isolates belonged to four clones. The K. pneumoniae isolates belonged to 9 Rep-types among which one included five isolates. CONCLUSION: This study is the first molecular characterization of ESBL-producing isolates from neonatology units in Madagascar, a country with limited epidemiological data. It revealed an important multi-clonal dissemination of CTX-M-15-producing isolates reflecting both the high community carriage and the very early nosocomial contamination of the neonates.

[Emerging and important antibiotic resistance in Gram negative bacteria: epidemiology, theory and practice]. / Résistances aux antibiotiques émergentes et importantes chez les bactéries Gram négatif: épidémiologie, aspects théoriques et détection.

Emerging and clinically-relevant antibiotic resistance mechanisms among Gram-negative rods are the extended-spectrum beta-lactamases (ESBL), carbapenemases, and 16S RNA methylases conferring resistance to aminoglycosides. Those resistance determinants do confer multiresistance to antibiotics. They are found in Enterobacteriaceae (especially community-acquired isolates, Pseudomonas aeruginosa and Acinetobacter baumannii). Detection of ESBL-producing and carbapenemase-producing isolates rely on the use of rapid diagnostic techniques that have to be performed when a reduced susceptibility to 3rd/4th generation cephalosporins or to carbapenems is observed, respectively. Only an early detection of those emerging resistance traits may contribute to limit their nosocomial spread and to optimize the antibiotic stewardship.

Intercontinental spread of OXA-48 beta-lactamase-producing Enterobacteriaceae over a 11-year period, 2001 to 2011.

OXA-48 beta-lactamase producers are emerging as an important threat mostly in the Mediterranean area. We report here the molecular epidemiology of a collection of OXA-48 beta-lactamase-positive enterobacterial isolates (n=107) recovered from European and north-African countries between January 2001 and December 2011. This collection included 67 Klebsiella pneumoniae, 24 Escherichia coli and 10 Enterobacter cloacae. Using the EUCAST breakpoints, ninety-eight isolates (91.6%) were of intermediate susceptibility or resistant to ertapenem, whereas 66% remained susceptible to imipenem. Seventy-five per cent of the isolates co-produced an extended-spectrum beta-lactamase, most frequently CTX-M-15 (77.5%). Susceptibility testing to non-beta-lactam antibiotics showed that colistin, tigecycline, amikacin, and fosfomycin remain active against most of the isolates. Multilocus sequence typing indicated that the most common sequence types (ST) were ST101 and ST38 for K. pneumoniae and E. coli, respectively. The bla(OXA-48) gene was located on a 62 kb IncL/M plasmid in 92.5% of the isolates, indicating that a single plasmid was mainly responsible for the spread of that gene. In addition, this study identified multiple cases of importation of OXA-48 beta-lactamase producers at least in Europe, and spread of OXA-48 beta-lactamase producers giving rise to an endemic situation, at least in France.

Outbreak of NDM-1-producing Acinetobacter baumannii in France, January to May 2013.

We report the first outbreak of carbapenem-resistant NDM-1-producing Acinetobacter baumannii in Europe, in a French intensive-care unit in January to May 2013. The index patient was transferred from Algeria and led to the infection/colonisation of five additional patients. Concurrently, another imported case from Algeria was identified. The seven isolates were genetically indistinguishable, belonging to ST85. The bla(NDM-1) carbapenemase gene was part of the chromosomally located composite transposon Tn125. This report underscores the growing concern about the spread of NDM-1-producing A. baumannii in Europe.

Carbapenemase-producing Enterobacteriaceae in Europe: a survey among national experts from 39 countries, February 2013.

The spread of carbapenemase-producing Enterobacteriaceae (CPE) is a threat to healthcare delivery, although its extent differs substantially from country to country. In February 2013, national experts from 39 European countries were invited to self-assess the current epidemiological situation of CPE in their country. Information about national management of CPE was also reported. The results highlight the urgent need for a coordinated European effort on early diagnosis, active surveillance, and guidance on infection control measures.

Carbapenem-hydrolyzing class D ß-lactamase OXA-48 in Klebsiella pneumoniae isolates from Tunisia.

Two carbapenem-resistant Klebsiella pneumoniae isolates producing the plasmid-encoded carbapenem-hydrolyzing OXA-48 were identified. These isolates, recovered from two patients hospitalized in two different hospitals in Tunisia in December 2010, were not clonally related. Molecular investigations showed that both isolates co-produced the narrow-spectrum ß-lactamases TEM-1 and SHV-1, together with the extended-spectrum ß-lactamase CTX--15.

Nosocomial dissemination of extended-spectrum ß-lactamase VEB-1a-producing Providencia stuartii isolates in a Tunisian hospital.

A collection of 20 multidrug-resistant Providencia stuartii isolates recovered from 2005 to 2009 at the Military Hospital of Tunis, Tunisia, was analysed. They all expressed the extended-spectrum ß-lactamase (ESBL) VEB-1a. The bla (VEB-1a) gene was plasmid-located and it was associated with complex genetic structures, including Re elements. Pulsed-field gel electrophoresis (PFGE) revealed a clonal relationship between all of these isolates. This study identified a nosocomial dissemination of an ESBL-producing P. stuartii clone in a Tunisian hospital over a long period of time.

[CTX-M-15-producing Klebsiella pneumoniae: a change in the epidemiology of ESBL]. / Klebsiella pneumoniae CTX-M-15: vers une modification de l'épidémiologie des ß-lactamases à spectre étendu.

Klebsiella pneumoniae is frequently involved in nosocomial outbreaks worldwide. High level of resistance is common for these bacteria leading to reduce antibiotic treatments and prolonged hospital stay for patients. Resistance determinants are often located on plasmids. During the 1980-1990s, ESBL encoding genes belonged to the TEM and SHV type. From the early 2000s, a new trend was observed with ESBL of the CTX-M type being increasingly described in K. pneumoniae, and more particularly CTX-M-15.

Evaluation of the performance of rapid tests for screening carriers of acquired ESBL-producing Enterobacterales and their impact on turnaround time.

BACKGROUND: Extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales constitute a global burden for hospital infection, and the identification of carriers by screening patients at risk is recommended by several guidelines. AIM: To evaluate the impact of rapid ESBL tests on the turnaround time (TAT) of screening. METHODS: Rectal swabs were analysed by culture and synergism tests for identification of non-Esherichia coli Enterobacterales that produce ESBLs (NEcESBL-producing Enterobacterales). The Rapid ESBL NP and NG CTX-M MULTI tests were performed on colonies grown on chromogenic media. The results of polymerase chain reaction and sequencing of ESBL genes were used as the gold standard. RESULTS: Among 473 analysed swabs, 75 (15.9%) grew NEcESBL-producing Enterobacterales, leading to 89 isolates. Sensitivities of the synergism, Rapid ESBL NP and NG CTX-M MULTI tests were 0.97 [95% confidence interval (CI) 0.88-0.99], 0.81 (95% CI 0.69-0.89) and 0.90 (95% CI 0.80-0.96), respectively. Specificities were 0.92 (95% CI 0.73-0.99), 0.85 (95% CI 0.64-0.95) and 0.96 (95% CI 0.78-1.00), respectively. Considering the 473 rectal swabs, ESBL screening using the synergism, Rapid ESBL NP and NG CTX-M MULTI tests was calculated. Sensitivities were 0.96 (95% CI 0.86-0.99), 0.81 (95% CI 0.68-0.90) and 0.91 (95% CI 0.79-0.97); specificities were 1.00 (95% CI 0.98-1.00), 0.99 (95% CI 0.98-1.00) and 1.00 (95% CI 0.99-1.00); positive predictive values were 0.96 (95% CI 0.86-0.99), 0.94 (95% CI 0.81-0.98) and 1.00 (95% CI 0.91-1.00); and negative predictive values were 1.00 (95% CI 0.98-1.00), 0.98 (95% CI 0.96-0.99) and 0.99 (95% CI 0.97-1.00), respectively. When no NEcESBL-producing Enterobacterales were observed, the mean TAT was 30 h. When NEcESBL-producing Enterobacterales were identified, the mean TATs were 74.7, 38.0 and 36.7 h for the synergism, Rapid ESBL NP and NG CTX-M MULTI tests, respectively. CONCLUSION: The two rapid ESBL tests showed good performance and allowed a reduction in TAT for screening protocols to identify patients carrying ESBL-producing Enterobacterales.

Evaluation of a DNA microarray, the check-points ESBL/KPC array, for rapid detection of TEM, SHV, and CTX-M extended-spectrum beta-lactamases and KPC carbapenemases.

Extended-spectrum beta-lactamases (ESBLs) and Klebsiella pneumoniae carbapenemases (KPC carbepenemases) have rapidly emerged worldwide and require rapid identification. The Check-Points ESBL/KPC array, a new commercial system based on genetic profiling for the direct identification of ESBL producers (SHV, TEM, and CTX-M) and of KPC producers, was evaluated. Well-characterized Gram-negative rods (Enterobacteriaceae, Pseudomonas aeruginosa, Acinetobacter baumannii) expressing various ss-lactamases (KPC-2, SHV, TEM, and CTX-M types) were used as well as wild-type reference strains and isolates harboring ss-lactamase genes not detected by the assay. In addition, phenotypically confirmed ESBL producers isolated in clinical samples over a 3-month period at the Bicetre hospital were analyzed using the Check-Points ESBL/KPC array and by standard PCR. The Check-Points ESBL/KPC array allowed fast detection of all TEM, SHV, and CTX-M ESBL genes and of the KPC-2 gene. The assay allowed easy differentiation between non-ESBL TEM and SHV and their ESBL derivatives. None of the other tested ss-lactamase genes were detected, underlining its high specificity. The technique is suited for Enterobacteriaceae but also for P. aeruginosa and A. baumannii. However, for nonfermenters, especially P. aeruginosa, a 1:10 dilution of the total DNA was necessary to detect KPC-2 and SHV-2a genes reliably. The Check-Points ESBL/KPC array is a powerful high-throughput tool for rapid identification of ESBLs and KPC producers in cultures. It provided definitive results within the same working day, allowing rapid implementation of isolation measures and appropriate antibiotic treatment. It showed an interesting potential for routine laboratory testing.

Intrafamilial transmission of extended-spectrum-beta-lactamase-producing Escherichia coli and Salmonella enterica Babelsberg among the families of internationally adopted children.

OBJECTIVES: International adoption from developing countries has become an increasing phenomenon in recent years. Given the high prevalence of multidrug-resistant (MDR) bacteria in these countries, the adopted children represent a group at risk for both carriage and infection with MDR bacteria. The dynamics of intrafamilial transmission of MDR bacteria after adoption was studied in a prospective study from January 2002 to January 2005. METHODS: Stool samples, taken at the first visit to the outpatient adoption practice and subsequently every month, from the adopted children of an orphanage of Bamako (Mali) and from all the members of their adoptive families were screened for MDR bacteria and bacterial pathogens. Bacteria were characterized by standard biochemical methods, disc diffusion antibiograms, PFGE and plasmid analysis. beta-Lactamase genes were sought by PCR. RESULTS: Over the study period, 52 ESBL-producing Enterobacteriaceae (E-ESBL), with Escherichia coli (56%) being the most prevalent, were isolated from 24/25 adoptees at arrival in France. During follow-up, the transmission of ESBL-producing E. coli and Salmonella enterica Babelsberg between the adoptees and their adoptive family members has clearly been demonstrated for 5/22 families (23%). The mean duration of the carriage for the adopted children was 9 months (1-15 months). CTX-M-15 was the most prevalent resistance gene among the E-ESBLs (93%), while SHV-12 was found among the S. enterica Babelsberg studied. CONCLUSIONS: International travellers, transfer of patients and now adoption may contribute to the global emergence of MDR bacteria. Thus, in addition to the usual screening of adopted children for infectious diseases, additional screening for MDR bacteria should be recommended, at least for children coming from countries with a high prevalence of MDR bacteria.

[KPC carbapenemases: what is at stake in clinical microbiology?]. / Carbapénèmases de type KPC: quel enjeu en microbiologie clinique?

Emergence and dissemination of carbapenem resistance in the world represent a significant threat for management of hospital-acquired infections. From the early 2000s, Enterobacteriaceae that produce Klebsiella pneumoniae carbapenemases (KPC) have initially been reported from the USA and now worldwide, becoming the most important carbapenemase. These KPC producing-bacteria are mostly involved in nosocomial and systemic infections. They are mostly Enterobacteriaceae and more rarely Pseudomonas aeruginosa. KPC beta-lactamases confer decreased susceptibility or resistance to virtually all beta-lactams. Therefore, carbapenems (imipenem, meropenem, ertapenem) may become inefficient for treating enterobacterial infections with KPC-producing bacteria, which are in addition resistant to many other non beta-lactam molecules, leaving only few available therapeutic options. Detection of KPC-producing bacteria may be difficult based on routine antibiotic susceptibility testing. Several phenotypic tests have been proposed, but until now, only molecular methods are reliable techniques for their identification. It is therefore critical to implement efficient infection control measures to detect patients who are colonized or infected with these pathogens in order to limit their spread.

Molecular characterization of multidrug-resistance in Gram-negative bacteria from the Peshawar teaching hospital, Pakistan.

Extended-spectrum ß-lactamases, carbapenemases, 16S rRNA methylases conferring pan-drug aminoglycoside resistance and colistin resistance were investigated among Gram-negative bacteria recovered from clinical samples (infections) from 200 individuals hospitalized at the Khyber Teaching Hospital of Peshawar, north Pakistan, from December 2017 to March 2018. Out of 65 isolates recovered, 19% were carbapenem resistant and 16% carried a bla NDM-1 gene, confirming the widespread distribution of NDM producers in this country. The association of the NDM carbapenem-resistance determinant, together with the extended-spectrum ß-lactamase CTX-M-15 and 16S rRNA methylases, was frequent, explaining the multidrug-resistance pattern observed. All isolates remained susceptible to colistin.