Emergence of a ST307 clone carrying a novel insertion element MITEKpn1 in the mgrB gene among carbapenem-resistant Klebsiella pneumoniae from Moscow,Russia

Carbapenem-resistant Klebsiella pneumoniae (CRKP) represents a major nosocomial pathogen with only a few antimicrobial agents, including colistin, remaining active. However, the emergence of colistin-resistant (Col-R) isolates is compromising the activity of colistin. In this study, a collection of 159 CRKP recovered from three hospitals in Moscow (Russian Federation) was examined. The isolates demonstrated resistance to cephalosporins (100%), ciprofloxacin (92.5%), fosfomycin (90.1%), netilmicin (81.1%), gentamicin (84.3%) and amikacin (49.7%). The rate of colistin resistance (MIC > 2 mg/L by broth microdilution) was 44.7%; moreover, 6.7% of isolates were tigecycline-resistant. Among 18 sequence types (STs) discovered, isolates of five lineages including ST307 (n = 46; 28.9%), ST395 (n = 40; 25.2%), ST377 (n = 17; 10.7%), ST48 (n = 17; 10.7%) and ST23 (n = 16; 10.1%) dominated. Carriage of a bla_OXA-48-like carbapenemase gene was detected in 146 CRKP (91.8%); 11 (6.9%) and 2 (1.3%) isolates harboured bla_NDM-1 and bla_KPC-3, respectively. Among 71 Col-R isolates, colistin MICs ranged from 4 mg/L to >1024 mg/L (MIC_50/90, 2/512 mg/L). All Col-R isolates were mcr-1-negative. In 19 (26.8%) Col-R isolates, inactivation of mgrB by insertion sequences IS1A, IS1R, ISKpn14 and ISKpn26 and a novel miniature inverted-repeat transposable element (MITE) Kpn1 was observed. Carriage of MITEKpn1 was restricted to six ST307 isolates and affected mgrB at nucleotide position 75. mgrB deletion was observed in four (5.6%) Col-R isolates. Moreover, PmrA and/or PmrB were altered in three (4.5%) Col-R isolates with wild-type mgrB. Thus, bla_OXA-48-like-carrying Col-R ST307 K. pneumoniae is emerging as a dominant clone in Moscow.     

Carbapenemase-producing Enterobacteriaceae in a tertiary hospital in Madrid,Spain: high percentage of colistin resistance among VIM-1-producing Klebsiella pneumoniae ST11 isolates

Here we describe the carbapenemase genes, genetic relatedness and antimicrobial susceptibility data of 123 carbapenemase-producing Enterobacteriaceae (CPE) clinical isolates recovered from 2010 to 2012, comprising Klebsiella pneumoniae (n = 79), Klebsiella oxytoca (n = 13), Serratia marcescens (n = 14), Enterobacter cloacae (n = 12), Enterobacter asburiae (n = 4) and Enterobacter aerogenes (n = 1). VIM-1 was the most common carbapenemase (n = 101) followed by KPC-2 (n = 19), OXA-48 (n = 2) and IMP-22 (n = 1). Among the K. pneumoniae isolates, nine sequence types (STs) were identified but two clones were dominant: ST11 (54/79) containing mainly VIM-1-producing isolates; and ST101 (13/79) constituted by KPC-2-producing strains. Pulsed-field gel electrophoresis (PFGE) showed a higher genetic diversity among the remaining Enterobacteriaceae. Amikacin and fosfomycin were the most active agents with 82.9% and 80.5% susceptibility, respectively. Non-susceptibility to tigecycline was detected in 36.5% of strains. Overall, colistin resistance was 24.7% and was as high as 47% in Enterobacter spp. An increase in colistin resistance from 13.5% to 31.7% was observed among K. pneumoniae isolates during the study period. Resistance was focused on ST11 since 83.3% of colistin-resistant strains belonged to this clone. The high level of colistin resistance observed in this study is worrying with respect to the already limited therapeutic options for infections caused by multidrug-resistant Gram-negative bacteria.     

Risk factors and outcomes associated with the isolation of polymyxin B and carbapenem-resistant Enterobacteriaceae spp.: A case–control study

Increasing resistance to polymyxin, a last-line antibiotic, is a growing public health concern worldwide. The primary objective of this study was to identify predictors for the isolation of polymyxin-resistant (PR) carbapenem-resistant Enterobacteriaceae (CRE) among hospitalized patients. The secondary objective was to describe the clinical outcomes of patients with PR-CRE infections. A retrospective case–control study including patients admitted to Singapore General Hospital between June 2012 and June 2016 was conducted. Cases were defined as patients who had clinical cultures from which a PR-CRE was isolated. Controls were randomly selected from patients with polymyxin-susceptible (PS) CRE admitted during the same period, and frequency-matched to site of isolation. We included 37 PR cases and 111 PS controls. Polymyxin resistance was detected predominantly in Enterobacter spp. (54.1%) and Klebsiella pneumoniae (43.2%). Multilocus sequence typing showed little clonal relatedness among the isolates. mcr-1 was detected in two PR-CRE isolates. Multivariable analyses showed that PR-CRE isolation was associated with prior polymyxins (adjusted odds ratio (OR), 21.31; 95% confidence interval (CI), 3.04–150.96) and carbapenem exposures (OR 3.74; CI 1.13–12.44), when adjusted for time at risk and bacteria species. In PR-CRE patients with infections, the 30-day all-cause in-hospital mortality was 50.0% as compared to 38.1% in patients with PS-CRE (P?=?0.346). Prior polymyxin and carbapenem exposures were independent risk factors for isolation of PR-CRE. Outcomes of PR-CRE and PS-CRE infections were similar in this study.     

Temporal evolution of polymyxin B-resistant Klebsiella pneumoniae clones recovered from blood cultures in a teaching hospital during a 7-year period

The polymyxins have become one of the last resorts to treat serious infections caused by KPC-2-producing Klebsiella pneumoniae worldwide. However, the increase of polymyxin consumption has favored the emergence of resistance to these compounds. In this study, we observed an increase in polymyxin B resistance rates from 0 to 30.6% among 224?K. pneumoniae isolates recovered from blood cultures between 2009 and 2015. Only gentamicin, tigecycline and fosfomycin remained active against the polymyxin B-resistant K. pneumoniae (PMB-R-KPN) isolates, which were classified as extensively drug-resistant (XDR; 83.3%), multidrug-resistant (MDR; 13.9%), or pan-drug resistant (2.8%). Most PMB-R-KPN clones belonged to CC258 (ST11, ST258, ST340, and ST437). A C7/ST258 XDR clone carrying distinct resistance determinants (bla_SHV-11, bla_TEM-1, bla_CTX-M-15, bla_CTX-M-14, bla_KPC-2, and rmtB-1) was introduced in 2014. Twelve of 36 PMB-R-KPN isolates showed disruption of mgrB. No mcr–1–positive isolate was found. The rapid detection of PMB-R-KPN isolates allied to implementation of effective infection control measures are of crucial importance to avoid the dissemination of high-risk PMB-R-KPN clones.     

碳青霉烯耐药肠杆菌科细菌临床分布及耐药特征分析

目的 分析综合性三甲医院住院患者临床标本分离碳青霉烯耐药肠杆菌科细菌（CRE）的临床分布及耐药特征,为临床抗感染治疗及院感防控提供参考。方法 收集2015年7月至2017年6月安徽医科大学第二附属医院住院患者临床标本分离CRE药敏数据及相关临床资料,应用WHONET 5.6软件统计分析CRE临床分布及耐药率,药敏折点参照CLSI 2016版标准。结果 2015年7月至2017年6月期间,自住院患者临床标本共分离CRE 82株,由肺炎克雷伯菌（42株,51.22%）、大肠埃希菌（23株,28.05%）、阴沟肠杆菌（9株,10.98%）、黏质沙雷菌（5株,6.10%）、弗劳地枸橼酸杆菌（2株,2.44%）及产酸克雷伯菌（1株,1.22%）构成。临床标本以痰标本（29株,35.37%）,分泌物/脓液标本（20株,24.39%）及尿液标本（14株,17.07%）为主。CRE分布广泛,检出的临床科室包括ICU（14株,17.07%）、呼吸内科（10株,12.20%）等。CRE菌株对临床常用抗菌药物耐药性较高,仅对阿米卡星较为敏感,耐药率为23.17%。结论 CRE菌株临床分布广泛且对常用抗菌药物耐药率高,临床科室应积极送检病原学检验并根据药敏结果合理选择抗菌药物。医院感控部门应联合微生物实验加强对CRE菌株的监测与防控。     

Occurrence of CTX-M-15- and MCR-1-producing Enterobacterales in pigs in Portugal: Evidence of direct links with antibiotic selective pressure

AimsTo undertake a prospective analysis of the occurrence of colistin-resistant and extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales colonizing pigs at two farms in Portugal, and to evaluate the putative correlations with usage of different antibiotics.Materials and methodsOne hundred and two faecal samples recovered from two different Portuguese pig farms were screened for polymyxin-resistant and ESBL-positive Enterobacterales. The authors had undertaken a study at one of the farms previously, but the use of colistin has since been banned; zinc oxide and amoxicillin are used as prophylactic and curative drugs, respectively, at this farm. The other farm included in this study used zinc oxide alone.ResultsNinety-three ESBL-producing isolates (62 Escherichia coli, 29 Klebsiella pneumoniae, one Enterobacter aerogenes and one Enterobacter cloacae) and 17 colistin-resistant isolates (12 E. coli, four K. pneumoniae and one E. cloacae) were recovered. Among the ESBL producers, the majority (84%) produced CTX-M-15, while the others produced CTX-M-1 or CTX-M-9. Many different strain and plasmid backgrounds were identified, ruling out a massive dissemination of one major clone. In total, 17 colistin-resistant isolates were recovered, all from the first farm. All produced MCR-1, corresponding to 12 E. coli (10 clones) and three K. pneumoniae (two clones). The MCR-1 producers were all recovered from the farm where colistin had been used 2 years previously.ConclusionThis study showed a surprisingly high rate of CTX-M-15 producers at two Portuguese pig farms. A link was found between antibiotic selective pressure (ß-lactam or polymyxin) and the corresponding resistance rate.     

Insertion sequence transpositions and point mutations in mgrB causing colistin resistance in a clinical strain of carbapenem-resistant Klebsiella pneumoniae from Vietnam

Resistance among Klebsiella pneumoniae to the last-resort antibiotics carbapenems and colistin is increasing worldwide. In this study, whole-genome sequencing was used to determine the colistin resistance mechanisms in clinical isolates of carbapenem- and colistin-resistant K. pneumoniae from Vietnam. Alterations in the regulatory gene mgrB, via mutations and insertion sequence transpositions, were found in 30 of 31 isolates, emphasising the importance of this resistance mechanism in colistin-resistant K. pneumoniae.     

Activity of ceftazidime-avibactam alone and in combination with polymyxin B against carbapenem-resistant Klebsiella pneumoniae in a tandem in vitro time-kill/in vivo Galleria mellonella survival model analysis

Ceftazidime-avibactam is used clinically in combination with a polymyxin for the treatment of carbapenem-resistant Gram-negative infections; however, there are limited data to support this practice. The objective of this study was to evaluate the activity of ceftazidime-avibactam and polymyxin B alone and in combination against Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae in a tandem in vitro time-kill/in vivo Galleria mellonella survival model assay. Three KPC-3-producing K. pneumoniae clinical isolates were used for all experiments. All isolates harbored mutations in ompk35 and one isolate in ompk36; two isolates were susceptible to both ceftazidime-avibactam and polymyxin B, and one was resistant to both. Ceftazidime-avibactam was bactericidal against 2 of 3 strains at ≥2x minimum inhibitory concentration (MIC) whereas polymyxin B was not bactericidal against any strain at any concentration. Combinations at 1/4x or 1/2x MIC were not bactericidal or synergistic against any of the 3 isolates. In survival experiments, ceftazidime-avibactam at 4x MIC significantly improved larval survival over the untreated control strain whereas polymyxin B at 4x MIC did not. Combining polymyxin B with ceftazidime-avibactam at 4x MIC did not improve survival compared to ceftazidime-avibactam alone. This work indicates there is no improvement in in vitro bactericidal activity or in vivo efficacy when polymyxin B is combined with ceftazidime-avibactam against KPC-producing K. pneumoniae. This combination should be avoided in lieu of ceftazidime-avibactam alone or other potentially more efficacious, less toxic combination regimens.     

Expansion of KPC-producing Klebsiella pneumoniae with various mgrB mutations giving rise to colistin resistance: the role of ISL3 on plasmids

mcr-1 has been reported as the first plasmid-encoded gene conferring colistin resistance. In KPC-producing Klebsiella pneumoniae (KPC-KP), however, colistin resistance is rapidly emerging through other mechanisms. Resistance is frequently due to disruption of the mgrB gene by insertion sequences, e.g. ISL3. The aim of this study was to investigate the expansion of mgrB-mutated KPC-KP isolates. In addition, the localisation and targets of ISL3 sequences within the core and accessory genome of common KPC-KP lineages were identified. A total of 29 clinical K. pneumoniae isolates collected from Italian patients were randomly selected. Whole genome sequences were analysed for resistance genes, plasmids and insertion sequences. In addition, 27 colistin-resistant KPC-KP isolates from a previous study from Crete (Greece) were assessed. Clonal expansion of KPC-KP isolates with various mutations in mgrB among all lineages was observed. In two Italian MLST ST512 isolates and eight Greek ST258 isolates, an identical copy of ISL3 was inserted in mgrB nucleotide position 133. ISL3, a transposable restriction–modification system of 8154 nucleotides, was located on pKpQIL-like plasmids and may transpose into the chromosome. In four isolates, chromosomal integration of ISL3 in diverse inner membrane proteins other than mgrB was identified. Colistin resistance is most often explained by clonal expansion of isolates with mutated mgrB. pKpQIL-like plasmids, which are omnipresent in KPC-KP, carry insertion sequences such as ISL3 that have mgrB as a target hotspot for transposition. Transposition of insertion sequences from plasmids and subsequent clonal expansion may contribute to the emerging colistin resistance in KPC-KP.     

Combination of polymyxin B and minocycline against multidrug-resistant Klebsiella pneumoniae: interaction quantified by pharmacokinetic/pharmacodynamic modelling from in vitro data

Lack of effective treatment for multidrug-resistant Klebsiella pneumoniae (MDR-Kp) necessitates finding and optimising combination therapies of old antibiotics. The aims of this study were to quantify the combined effect of polymyxin B and minocycline by building an in silico semi-mechanistic pharmacokinetic/pharmacodynamic (PKPD) model and to predict bacterial kinetics when exposed to the drugs alone and in combination at clinically achievable unbound drug concentration–time profiles. A clinical K. pneumoniae strain resistant to polymyxin B [minimum inhibitory concentration (MIC) = 16 mg/L] and minocycline (MIC = 16 mg/L) was selected for extensive in vitro static time–kill experiments. The strain was exposed to concentrations of 0.0625–48 × MIC, with seven samples taken per experiment for viable counts during 0–28 h. These observations allowed the development of the PKPD model. The final PKPD model included drug-induced adaptive resistance for both drugs. Both the minocycline-induced bacterial killing and resistance onset rate constants were increased when polymyxin B was co-administered, whereas polymyxin B parameters were unaffected. Predictions at clinically used dosages from the developed PKPD model showed no or limited antibacterial effect with monotherapy, whilst combination therapy kept bacteria below the starting inoculum for >20 h at high dosages [polymyxin B 2.5 mg/kg + 1.5 mg/kg every 12 h (q12h); minocycline 400 mg + 200 mg q12h, loading + maintenance doses]. This study suggests that polymyxin B and minocycline in combination may be of clinical benefit in the treatment of infections by MDR-Kp and for isolates that are non-susceptible to either drug alone.     

Inactivation of mgrB gene regulator and resistance to colistin is becoming endemic in carbapenem-resistant Klebsiella pneumoniae in Greece: A nationwide study from 2014 to 2017

IntroductionIn Greece, the spread of carbapenem-resistant Enterobacteriaceae in humans has led to the reintroduction of colistin as a therapeutic agent. Unfortunately, colistin resistance with different mechanisms has emerged. The present work aims to determine the prevalence of carbapenem and colistin resistance and the corresponding mechanisms in Klebsiella pneumoniae clinical isolates from Greece.MethodsFrom 2014 to 2017, 288 carbapenem-resistant K. pneumoniae clinical strains were gathered from a collection of 973 isolates from eight different hospitals in Greece. Antibiotic susceptibility testing was performed using three different methods. Screening of carbapenem and colistin resistance genes was conducted using polymerase chain reaction (PCR) amplification and sequencing.ResultsAmong the 288 (29.6 %) carbapenem-resistant isolates, 213 (73.9%) were colistin-resistant (minimum inhibitory concentration [MIC] >2 mg/L). The KPC type was the most common carbapenemase gene (116; 40.3%), followed by VIM (41; 14.2%), NDM (33; 11.5%) and OXA-48 (22; 7.6%). Moreover, 44 (15.3%) strains co-produced two types of carbapenemases. No mcr genes were detected for colistin resistance but mutations in chromosomal genes were found. These included inactivation of the mgrB gene for 148 (69.5%) strains, including insertion sequences for 94 (44.1%), nonsense mutations for 4 (1.9%) and missense mutations for 24 (11.3%). Moreover, PCR amplification of mgrB gene was negative for 26 (12.2%) strains. Finally, 65 (30.5%) colistin-resistant strains exhibited a wild-type mgrB, the mechanisms of which remain to be elucidated.ConclusionThis study shows that K. pneumoniae clinical strains in Greece are resistant to both carbapenems and colistin and this is endemic and is likely chromosomally encoded.     

In vitro activity of avibactam (NXL104) in combination with β-lactams against Gram-negative bacteria, including OXA-48 β-lactamase-producing Klebsiella pneumoniae

The objective of this study was to investigate the in vitro antibacterial activity of avibactam (formerly NXL104) in combination with imipenem, cefepime or ceftazidime against Gram-negative bacteria. Bacterial isolates included: Pseudomonas aeruginosa harbouring PER-1 β-lactamase (n = 14); Acinetobacter baumannii harbouring PER-1, OXA-51 and OXA-58 (n = 20); carbapenem-non-susceptible Klebsiella pneumoniae (n = 25) and Escherichia coli (n = 1) harbouring OXA-48; carbapenem-non-susceptible E. coli (n = 1) harbouring both IMP-1 metallo-β-lactamase and extended-spectrum β-lactamase (ESBL); carbapenem-non-susceptible Serratia marcescens (n = 1); and carbapenem-susceptible E. coli (n = 20) and K. pneumoniae isolates (n = 12) with CTX-M-15 ESBL. Minimum inhibitory concentrations (MICs) of imipenem, cefepime and ceftazidime were determined in combination with 4 mg/L avibactam by the Clinical and Laboratory Standards Institute (CLSI) method on Mueller-Hinton agar. Imipenem/avibactam and ceftazidime/avibactam displayed limited potency against A. baumannii isolates, whereas cefepime/avibactam and ceftazidime/avibactam were active against P. aeruginosa. Klebsiella pneumoniae isolates with OXA-48 β-lactamase were resistant to imipenem [MIC for 90% of the organisms (MIC₉₀) ≥4 mg/L]. MIC₉₀ values for the combination of avibactam 4 mg/L with imipenem, cefepime and ceftazidime were in the susceptible range for all strains (MIC₉₀ ≤ 0.5 mg/L). All E. coli and K. pneumoniae isolates with CTX-M-15 β-lactamase were inhibited at ≤1 mg/L for combinations with avibactam and 100% were susceptible by CLSI breakpoint criteria to imipenem, cefepime and ceftazidime. In conclusion, combinations of imipenem, cefepime and ceftazidime with avibactam may present a promising therapeutic strategy to treat infections due to K. pneumoniae with OXA-48 enzyme as well as K. pneumoniae and E. coli with CTX-M-15 enzyme.     

Contributions of insertion sequences conferring colistin resistance in Klebsiella pneumoniae

BackgroundIncreasing colistin consumption is leading to expanding colistin resistance in Klebsiella pneumoniae worldwide, but particularly in Asia. Epidemiological studies indicate a link between specific insertion sequences (ISs) and colistin resistance; however, proof of a colistin-IS correlation is lacking.ObjectivesColistin-resistant mechanisms, and in vitro and in vivo efficacies of colistin against K. pneumoniae with ISs were investigated.MethodsColistin-resistant genes, including mcr-1 gene, were detected in 49 colistin- and carbapenem-resistant K. pneumoniae isolates. crrCAB genetic environments were analysed using whole-genome sequencing and polymerase chain reaction (PCR) mapping. Identified ISs were cloned into pRK415 vectors and investigated for potential contributions to colistin resistance. A Caenorhabditis elegans model was employed for in vivo analysis.ResultsmgrB gene alterations (32/49, 65.3%) were identified as the major colistin-resistant mechanism, followed by variations in crrB (57.1%), pmrB (32.7%), phoQ (20.9%), pmrA (16.3%) and phoP (8.2%) genes. Furthermore, 21 of the 49 tested isolates (42.9%) contained the IS elements, ISKpn26, ISEcp1, IS10R, IS903B or ISKpn14 in mgrB or in the surrounding region of crrCAB, indicating an association between these ISs and colistin resistance. The frequencies of colistin resistance significantly increased in colistin-susceptible K. pneumoniae laboratory strains, with plasmids carrying different ISs from clinical strains. In vivo analysis revealed that K. pneumoniae harboring ISKpn26 was associated with decreased lifespan during colistin treatment, leading to an increased risk for colistin treatment failure.ConclusionsThese findings indicate a correlation between diverse ISs and colistin resistance in K. pneumoniae and confirm a role for ISs in colistin treatment.     

Systematic review and meta-analysis of in vitro efficacy of antibiotic combination therapy against carbapenem-resistant Gram-negative bacilli

The superiority of combination therapy for carbapenem-resistant Gram-negative bacilli (CR-GNB) infections remains controversial. In vitro models may predict the efficacy of antibiotic regimens against CR-GNB. A systematic review and meta-analysis was performed including pharmacokinetic/pharmacodynamic (PK/PD) and time–kill (TK) studies examining the in vitro efficacy of antibiotic combinations against CR-GNB [PROSPERO registration no. CRD42019128104]. The primary outcome was in vitro synergy based on the effect size (ES): high, ES ≥ 0.75, moderate, 0.35 < ES < 0.75; low, ES ≤ 0.35; and absent, ES = 0). A network meta-analysis assessed the bactericidal effect and re-growth rate (secondary outcomes). An adapted version of the ToxRTool was used for risk-of-bias assessment. Over 180 combination regimens from 136 studies were included. The most frequently analysed classes were polymyxins and carbapenems. Limited data were available for ceftazidime/avibactam, ceftolozane/tazobactam and imipenem/relebactam. High or moderate synergism was shown for polymyxin/rifampicin against Acinetobacter baumannii [ES = 0.91, 95% confidence interval (CI) 0.44–1.00], polymyxin/fosfomycin against Klebsiella pneumoniae (ES = 1.00, 95% CI 0.66–1.00) and imipenem/amikacin against Pseudomonas aeruginosa (ES = 1.00, 95% CI 0.21–1.00). Compared with monotherapy, increased bactericidal activity and lower re-growth rates were reported for colistin/fosfomycin and polymyxin/rifampicin in K. pneumoniae and for imipenem/amikacin or imipenem/tobramycin against P. aeruginosa. High quality was documented for 65% and 53% of PK/PD and TK studies, respectively. Well-designed in vitro studies should be encouraged to guide the selection of combination therapies in clinical trials and to improve the armamentarium against carbapenem-resistant bacteria.     

Clinical impact of cefepime breakpoint in patients with carbapenem-resistant Klebsiella pneumoniae bacteraemia

The application of cefepime breakpoint for carbapenem-resistant Klebsiella pneumoniae (CRKP) bacteraemia has not been explored. Adult cases of monomicrobial bloodstream infection (BSI) caused by cefepime-susceptible [minimum inhibitory concentration (MIC) ≤8 mg/L] K. pneumoniae isolates with carbapenem resistance between 2010 and 2015 were reviewed. Patients treated with cefepime were compared with those treated by other active agents using a propensity score-matched analysis to assess therapeutic effectiveness. The primary endpoint was 30-day crude mortality. A total of 114 patients experienced cefepime-susceptible CRKP bacteraemia and 40 (35.1%) died during hospitalisation. A total of 33 patients (28.9%) received cefepime therapy. Fifteen patients (13.2%) had BSI due to carbapenemase-producing isolates, and 86.7% (13/15) of carbapenemase-producing isolates were classified as cefepime susceptible dose-dependent (SDD). In the multivariate logistic regression analysis, 30-day mortality was independently associated with the presence of a critical illness [adjusted odds ratio (aOR) = 12.89, 95% confidence interval (CI) 3.88–42.83; P < 0.001], pneumonia (aOR = 5.97, 95% CI 1.65–21.76; P = 0.007) and rapidly fatal underlying disease (aOR = 6.43, 95% CI 1.30–31.09; P = 0.02). In contrast, cefepime-based therapy (aOR = 0.03, 95% CI 0.003–0.38; P = 0.006) and combination therapy (aOR = 0.09, 95% CI 0.02–0.36; P = 0.001) were protective against a fatal outcome. Based on current breakpoints for Enterobacterales, cefepime therapy was not associated with an unfavourable outcome for CRKP BSI with MIC-based dosing strategies. However, the susceptibility result of SDD to cefepime should alert clinicians for possible therapeutic failure.     

Activity of ceftolozane-tazobactam against Gram-negative pathogens isolated from lower respiratory tract infections in the Asia-Pacific region: SMART 2015-2016

The aim of this study was to investigate the susceptibility of respiratory Gram-negative bacteria to ceftolozane/tazobactam and other antibiotics in the Asia-Pacific region during 2015-2016. MICs were determined using the CLSI standard broth microdilution method and interpreted accordingly. Pseudomonas aeruginosa (1574 isolates), Klebsiella pneumoniae (1226), Acinetobacter baumannii (627) and Escherichia coli (476) accounted for 73.1% of 5342 Gram-negative respiratory pathogens. Susceptibility to ceftolozane/tazobactam of individual Enterobacteriaceae was >80%, except for Enterobacter cloacae (76.6%). Ceftolozane/tazobactam inhibited 81.9% of K. pneumoniae and 91.9% of E. coli, with respective MIC₅₀/MIC₉₀ values of 0.5/>32 and 0.25/2 mg/L. For carbapenem-susceptible, ESBL-producing K. pneumoniae and E. coli, susceptibility was 65.5% and 93.3%, respectively, and respective MIC₅₀/MIC₉₀ values were 2/>32 and 0.5/2 mg/L. Bla_{CTX-M-1 group} was most prevalent in selected ESBL-producing K. pneumoniae (40 of 54 isolates) and E. coli (15 of 22 isolates), with ceftolozane/tazobactam susceptibility rates of 50% and 80%, respectively. Bla_SHV-ESBL was the second most prevalent, and ceftolozane/tazobactam inhibited 20% of 20 K. pneumoniae isolates with bla_SHV-ESBL. The only effective antibiotics for carbapenem-non-susceptible K. pneumoniae (111 isolates) and E. coli (24 isolates) were amikacin and colistin. Ceftolozane/tazobactam was effective against almost all tested P. aeruginosa and carbapenem-non-susceptible strains, with susceptibility of 92.3% and 72.8%, respectively; the respective MIC₅₀/MIC₉₀ values were 1/4 and 2/>32 mg/L. The high susceptibility of ceftolozane/tazobactam remained in different age groups, patient locations, recovery times and countries, except Vietnam. In conclusion, ceftolozane/tazobactam was effective against most respiratory Gram-negative pathogens in the Asia-Pacific region; however, the emergence of carbapenem resistance mandates ongoing surveillance.     

Prevalence and mechanisms of fosfomycin resistance among KPC-producing Klebsiella pneumoniae clinical isolates in China

The threat of antibiotic resistance has increased dramatically in recent years. Fosfomycin, an old antibiotic agent, has been re-introduced to fight infections caused by Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-KP). However, the trend of fosfomycin resistance among KPC-KP strains is increasing. In this study, 80 KPC-KP clinical isolates were collected from three teaching hospitals during 2014–2017 in China and were subjected to whole-genome sequencing (WGS). The fosfomycin resistance phenotype and resistance mechanisms were investigated by antimicrobial susceptibility testing and carbon source growth test, respectively. Among all KPC-KP strains, 80.0% (64/80) were resistant to fosfomycin and 36.3% (29/80) were positive for the mobile fosfomycin resistance gene fosA3. Among the 63 strains that were unable to grow in M9 basic medium with glycerol-3-phosphate (G3P) as the sole carbon source (mediated by mutation of the target gene glpT), there was no significant difference regarding the MIC distribution of fosfomycin between fosA3-positive and fosA3-negative strains (P = 0.577). Among the 50 strains that were negative for fosA3 but positive for fosA, the fosfomycin MICs of strains unable to grow in M9 basic medium with G3P as the sole carbon source were significantly higher (P < 0.001) than in strains that were able to grow in M9 basic medium with G3P as the sole carbon source. Our findings indicate that fosfomycin resistance among KPC-KP in China is an emerging problem and the two major mechanisms of resistance identified were plasmid-mediated fosfomycin resistance gene fosA3 and mutation of the target gene glpT.     

Risk factors for polymyxin-resistant carbapenemase-producing Enterobacteriaceae in critically ill patients: An epidemiological and clinical study

This study aimed to assess the clinical impact and potential risk factors associated with polymyxin-resistant Enterobacteriaceae strains isolated from patients hospitalized in adult and neonatal intensive care units. A case–control study was conducted from September 2015 to January 2017. Antimicrobial susceptibility of polymyxin-resistant Enterobacteriaceae strains was determined by broth microdilution. The presence of resistance genes was evaluated by polymerase chain reaction and DNA sequencing. Renal failure [P=0.02, odds ratio (OR) 11.37, 95% confidence interval (CI) 1.0–128.63], use of a urinary catheter (P<0.01, OR 4.16, 95% CI 38.82–366.07), transfer between hospital units (P=0.03, OR 9.98, 95% CI 1.01–98.42), carbapenem use (P<0.01, OR 45.49, 95% CI 6.93–298.62) and surgical procedure (P<0.01, OR 16.52, 95% CI 2.83–96.32) were found to be risk factors for the acquisition of polymyxin-resistant strains in adult patients. For neonatal patients, use of a central venous catheter (P<0.01, OR 69.59, 95% CI 7.33–660.30) was the only risk factor associated with the acquisition of polymyxin-resistant strains. Analysis of the outcomes revealed that the mortality rate was significantly higher in adult (66.6%) and neonatal (23.5%) patients with polymyxin-resistant strains than in those with polymyxin-susceptible strains. In addition, carbapenem exposure (P<0.01, OR 50.93, 95% CI 2.26–>999.999) was strongly associated with mortality. On the other hand, aminoglycoside use (P<0.03, OR 0.06, 95% CI 0.004–0.97) was a protective factor against mortality from polymyxin-resistant strains. Several risk factors were associated with polymyxin-resistant strains. The high mortality rates showed that acquisition of these strains is a predictor for unfavourable outcomes. Combination treatment with an aminoglycoside and polymyxin might be a better combination to improve patient outcomes.     

Comparison of two DNA microarrays for detection of plasmid-mediated antimicrobial resistance and virulence factor genes in clinical isolates of Enterobacteriaceae and non-Enterobacteriaceae

A DNA microarray was developed to detect plasmid-mediated antimicrobial resistance (AR) and virulence factor (VF) genes in clinical isolates of Enterobacteriaceae and non-Enterobacteriaceae. The array was validated with the following bacterial species: Escherichiacoli (n = 17); Klebsiellapneumoniae (n = 3); Enterobacter spp. (n = 6); Acinetobacter genospecies 3 (n = 1); Acinetobacterbaumannii (n = 1); Pseudomonasaeruginosa (n = 2); and Stenotrophomonasmaltophilia (n = 2). The AR gene profiles of these isolates were identified by polymerase chain reaction (PCR). The DNA microarray consisted of 155 and 133 AR and VF gene probes, respectively. Results were compared with the commercially available Identibac AMR-ve Array Tube. Hybridisation results indicated that there was excellent correlation between PCR and array results for AR and VF genes. Genes conferring resistance to each antibiotic class were identified by the DNA array. Unusual resistance genes were also identified, such as bla_SHV-5 in a bla_OXA-23-positive carbapenem-resistant A. baumannii. The phylogenetic group of each E. coli isolate was verified by the array. These data demonstrate that it is possible to screen simultaneously for all important classes of mobile AR and VF genes in Enterobacteriaceae and non-Enterobacteriaceae whilst also assigning a correct phylogenetic group to E. coli isolates. Therefore, it is feasible to test clinical Gram-negative bacteria for all known AR genes and to provide important information regarding pathogenicity simultaneously.