Ureidopenicillins Are Potent Inhibitors of Penicillin-Binding Protein 2 from Multidrug-Resistant Neisseria gonorrhoeae H041.

Effective treatment of gonorrhea is threatened by the increasing prevalence of Neisseria gonorrhoeae strains resistant to the extended-spectrum cephalosporins (ESCs). Recently, we demonstrated the promise of the third-generation cephalosporin cefoperazone as an antigonococcal agent due to its rapid second-order rate of acylation against penicillin-binding protein 2 (PBP2) from the ESC-resistant strain H041 and robust antimicrobial activity against H041. Noting the presence of a ureido moiety in cefoperazone, we evaluated a subset of structurally similar ureido ß-lactams, including piperacillin, azlocillin, and mezlocillin, for activity against PBP2 from H041 using biochemical and structural analyses. We found that the ureidopenicillin piperacillin has a second-order rate of acylation against PBP2 that is 12-fold higher than cefoperazone and 85-fold higher than ceftriaxone and a lower MIC against H041 than ceftriaxone. Surprisingly, the affinity of ureidopenicillins for PBP2 is minimal, indicating that their inhibitory potency is due to a higher rate of the acylation step of the reaction compared to cephalosporins. Enhanced acylation results from the combination of a penam scaffold with a 2,3-dioxopiperazine-containing R1 group. Crystal structures show that the ureido ß-lactams overcome the effects of resistance mutations present in PBP2 from H041 by eliciting conformational changes that are hindered when PBP2 interacts with the weaker inhibitor ceftriaxone. Overall, our results support the potential of piperacillin as a treatment for gonorrhea and provide a framework for the future design of ß-lactams with improved activity against ESC-resistant N. gonorrhoeae.

Antimicrobial resistance survey and whole-genome analysis of nosocomial P. Aeruginosa isolated from eastern Province of China in 2016-2021.

BACKGROUND: Pseudomonas aeruginosa is a major Gram-negative pathogen that can exacerbate lung infections in the patients with cystic fibrosis, which can ultimately lead to death. METHODS: From 2016 to 2021, 103 strains of P. aeruginosa were isolated from hospitals and 20 antibiotics were used for antimicrobial susceptibility determination. Using next-generation genome sequencing technology, these strains were sequenced and analyzed in terms of serotypes, ST types, and resistance genes for epidemiological investigation. RESULTS: The age distribution of patients ranged from 10 days to 94 years with a median age of 69 years old. The strains were mainly isolated from sputum (72 strains, 69.9%) and blood (14 strains, 13.6%). The size of these genomes ranged from 6.2 Mb to 7.4 Mb, with a mean value of 6.5 Mb. In addition to eight antibiotics that show inherent resistance to P. aeruginosa, the sensitivity rates for colistin, amikacin, gentamicin, ceftazidime, piperacillin, piperacillin-tazobactam, ciprofloxacin, meropenem, aztreonam, imipenem, cefepime and levofloxacin were 100%, 95.15%, 86.41%, 72.82%, 71.84%, 69.90%, 55.34%, 52.43%, 50.49%, 50.49%, 49.51% and 47.57% respectively, and the carriage rate of MDR strains was 30.69% (31/101). Whole-genome analysis showed that a total of 50 ST types were identified, with ST244 (5/103) and ST1076 (4/103) having a more pronounced distribution advantage. Serotype predictions showed that O6 accounted for 29.13% (30/103), O11 for 23.30% (24/103), O2 for 18.45% (19/103), and O1 for 11.65% (12/103) of the highest proportions. Notably, we found a significantly higher proportion of ExoU in P. aeruginosa strains of serotype O11 than in other cytotoxic exoenzyme positive strains. In addition to this, a total of 47 crpP genes that mediate resistance to fluoroquinolones antibiotics were found distributed on 43 P. aeruginosa strains, and 10 new variants of CrpP were identified, named 1.33, 1.34, 1.35, 1.36, 1.37, 1.38, 1.39, 1.40, 1.41 and 7.1. CONCLUSIONS: We investigated the antibiotic susceptibility of clinical isolates of P. aeruginosa and genomically enriched the diversity of P. aeruginosa for its prophylactic and therapeutic value.

Identification of a CTX-M-255 ß-lactamase containing a G239S substitution selectively conferring resistance to penicillin/ß-lactamase inhibitor combinations.

OBJECTIVES: An Escherichia coli isolate, WGS1363, showed resistance to piperacillin/tazobactam but susceptibility to cephalosporins and contained a previously unrecognized ß-lactamase, CTX-M-255, as the only acquired ß-lactamase. CTX-M-255 was identical to CTX-M-27 except for a G239S substitution. Here, we characterize the hydrolytic spectrum of CTX-M-255 and a previously reported ß-lactamase, CTX-M-178, also containing a G239S substitution and compare it to their respective parental enzymes, CTX-M-27 and CTX-M-15. METHODS: All ß-lactamase genes were expressed in E. coli TOP10 and MICs to representative ß-lactam-antibiotics were determined. Furthermore, blaCTX-M-15,  blaCTX-M-27, blaCTX-M-178 and blaCTX-M-255 with C-terminal His-tag fusions were affinity purified for enzyme kinetic assays determining Michaelis-Menten kinetic parameters against representative ß-lactam-antibiotics and IC50s of clavulanate, sulbactam, tazobactam and avibactam. RESULTS: TOP10-transformants expressing blaCTX-M-178 and blaCTX-M-255 showed resistance to penicillin/ß-lactamase combinations and susceptibility to cephalothin and cefotaxime in contrast to transformants expressing blaCTX-M-15 and blaCTX-M-27. Determination of enzyme kinetic parameters showed that CTX-M-178 and CTX-M-255 both lacked hydrolytic activity against cephalosporins and showed impaired hydrolytic efficiency against penicillin antibiotics compared to their parental enzymes. Both enzymes appeared more active against piperacillin compared to benzylpenicillin and ampicillin. Compared to their parental enzymes, IC50s of ß-lactamase-inhibitors were increased more than 1000-fold for CTX-M-178 and CTX-M-255. CONCLUSIONS: CTX-M-178 and CTX-M-255, both containing a G239S substitution, conferred resistance to piperacillin/tazobactam and may be characterized as inhibitor-resistant CTX-M ß-lactamases. Inhibitor resistance was accompanied by loss of activity against cephalosporins and monobactams. These findings add to the necessary knowledge base for predicting antibiotic susceptibility from genotypic data.

Exposure levels and target attainment of piperacillin/tazobactam in adult patients admitted to the intensive care unit: a prospective observational study.

PURPOSE: The objective of this study was to evaluate the exposure and the pharmacodynamic target attainment of piperacillin/tazobactam (PTZ) in adult critically ill patients. METHODS: We conducted a prospective observational study in the intensive care unit (ICU) of the Hôpital du Sacré-Cœur de Montréal (a Level I trauma centre in Montreal, QC, Canada) between January 2021 and June 2022. We included patients aged 18 yr or older admitted to the ICU who received PTZ by intravenous administration. Demographic and clinical characteristics were collected, and clinical scores were calculated. On study day 1 of antimicrobial therapy, three blood samples were collected at the following timepoints: one hour after PTZ dose administration and at the middle and at the end of the dosing interval. The sampling schedule was repeated on days 4 and 7 of therapy if possible. Samples were analyzed by ultra-high performance liquid chromatography with diode array detector to determine the total piperacillin concentration. Middle- and end-of-interval concentrations were used for target attainment analyses, and were defined as a concentration above the minimal inhibitory concentration of 16 mg·L-1, corresponding to the breakpoint of Enterobacteriaceae and Pseudomonas aeruginosa. RESULTS: Forty-three patients were recruited and 202 blood samples were analyzed. The most prevalent dose was 3/0.375 g every six hours (n = 50/73 doses administered, 68%) with a 30-min infusion. We observed marked variability over the three sampling timepoints, and the median [interquartile range] piperacillin concentrations at peak, middle of interval, and end of interval were 109.4 [74.0-152.3], 59.3 [21.1-74.4], and 25.3 [6.8-44.6] mg·L-1, respectively. When assessing target attainment, 37% of patients did not reach the efficacy target of a trough concentration of 16 mg·L-1. The majority of patients who were underexposed were patients with normal to augmented renal clearance. CONCLUSION: In this prospective observational study of adult ICU patients receiving intravenous PTZ, a large proportion had subtherapeutic concentrations of piperacillin. This was most notable in patients with normal to augmented renal clearance. More aggressive dosage regimens may be required for this subpopulation to ensure attainment of efficacy targets.

RéSUMé: OBJECTIF: L'objectif de cette étude était d'évaluer l'exposition et l'atteinte des cibles pharmacodynamiques de la pipéracilline/tazobactam (PTZ) chez la patientèle adulte aux soins intensifs. MéTHODES: Nous avons réalisé une étude observationnelle prospective dans l'unité de soins intensifs (USI) de l'Hôpital du Sacré-Cœur de Montréal (un centre de traumatologie de niveau 1 à Montréal, QC, Canada) entre janvier 2021 et juin 2022. Nous avons inclus les patient·es adultes âgé·es de 18 ans ou plus admis·es à l'USI ayant reçu de la PTZ par administration intraveineuse. Les caractéristiques démographiques et cliniques ont été recueillies, et les scores cliniques ont été calculés. Au jour 1 de la thérapie antimicrobienne, trois échantillons sanguins ont été prélevés aux moments suivants : 1 h après l'administration de la dose de PTZ, au milieu et à la fin de l'intervalle d'administration. Le calendrier d'échantillonnage a été répété aux jours 4 et 7 de la thérapie si possible. Les échantillons ont été analysés par chromatographie liquide à ultra-haute performance avec détecteur à diodes pour déterminer la concentration totale de pipéracilline. Les concentrations du milieu et de fin d'intervalle ont été utilisées pour les analyses d'atteinte de cible, définie comme une concentration supérieure à la concentration minimale inhibitrice de 16 mg·L-1, associée aux Enterobacteriaceae et au Pseudomonas aeruginosa. RéSULTATS: Quarante-trois patient·es ont été recruté·es et 202 échantillons sanguins ont été analysés. La dose la plus prévalente était une dose de 3/0,375 g aux 6 h (n = 50/73 doses administrées, 68 %) avec une perfusion sur 30 min. Nous avons observé une variabilité marquée aux trois temps de prélèvement, et les concentrations médianes [intervalle interquartile] de pipéracilline au pic, au milieu et à la fin de l'intervalle étaient respectivement de 109,4 [74,0-152,3], 59,3 [21,1-74,4] et 25,3 [6,8-44,6] mg·L−1. Lors de l'évaluation de l'atteinte de la cible, 37 % des patient·es n'ont pas atteint la cible d'efficacité d'une concentration de 16 mg·L−1 à la fin de l'intervalle posologique. La majorité des patient·es sous-exposé·es étaient des personnes dont la clairance rénale était normale ou augmentée. CONCLUSION: Dans cette étude observationnelle prospective de patient·es adultes aux soins intensifs recevant de la PTZ par voie intraveineuse, une grande proportion de patient·es présentait des concentrations sous-thérapeutiques de pipéracilline. Ceci était plus marqué chez les patient·es ayant une clairance rénale normale ou augmentée. Des schémas posologiques plus agressifs pourraient être nécessaires pour cette sous-population afin de favoriser l'atteinte des cibles d'efficacité.

High oral carriage of multidrug resistant Gram-negative bacilli in adolescents: the SOPKARD-Junior study.

Introduction: The colonization of the oral cavity by potentially pathogenic antimicrobial-resistant bacteria in adolescents and its consequences is very poorly understood. The present study focused on the occurrence of oral colonization by Gram-negative bacilli (GNB) and their multidrug resistance, including the production of extended-spectrum ß-lactamases (ESBLs) and carbapenemases, among healthy adolescents and risk factors associated with GNB colonization. Materials and methods: This study was conducted as part of "A program for the early detection of risk factors for lifestyle diseases SOPKARD-Junior" (SOPKARD-Junior). Oral samples were collected from 182 adolescents from four public elementary schools in Sopot, Poland, aged 13-14 years. Bacterial strains were identified by the MALDI-TOF MS method. Screening of antimicrobial resistance was performed using a disk diffusion method. The NG-Test® CARBA-5 was used to detect and differentiate the five most widely distributed carbapenemases. Demographic and clinical data were collected and statistical analysis of risk factors was performed. Results: A total of 68 out of 182 (37.4%) healthy adolescents was documented oral colonization with Gram-negative bacilli, including 50/182 (27.5%) multidrug resistant (MDR-GNB) strains. Over 60% of oral carriage concerned three main genera Enterobacter spp., Pseudomonas spp., and Serratia spp., which were detected in 22.1%, 19.1%, and 19.1% of participants, respectively. Citrobacter spp., Escherichia coli, Klebsiella spp., Hafnia spp., Aeromonas spp., Acinetobacter spp., and Stenotrophomonas spp. were also isolated. The antimicrobial resistance to ampicillin (100%), ceftazidime (69.1%), meropenem (60.3%), gentamycin (60.3%), piperacillin/tazobactam (52.9%), and piperacillin (45.6%) were the most common. Among 73.5% GNB strains multidrug resistance was observed, including all Pseudomonas spp. strains. Among MDR-GNB, 30.4% were resistant to four groups of antibiotics, half of the MDR Pseudomonas spp. strains were resistant to 10 groups of antibiotics. Extended-spectrum ß-lactamases were produced by Enterobacter cloacae, Klebsiella spp., and Serratia spp. (7.4%). Colonization by ESBLs-positive GNB strains was significantly associated with recurrent respiratory infections, nasal congestion, and bronchitis (p<0.05). Conclusion: Our study revealed high oral carriage of multi-drug resistant Gram-negative bacilli in healthy adolescents and the association of ESBL-producing strains with respiratory infections. Further studies on oral colonization with GNB are necessary due to the possibility of distinct infections and the acquisition of antibiotic resistance by resident microbiota.

Early treatment response to piperacillin/tazobactam in patients with bloodstream infections caused by non-ESBL ampicillin/sulbactam-resistant Escherichia coli: a binational cohort study.

PURPOSE: This study aimed to compare treatment outcomes for bloodstream infections (BSI) caused by a piperacillin/tazobactam (PIP/TAZ)-susceptible E. coli among three patient groups: BSI caused by ampicillin/sulbactam (AMP/SLB)-resistant isolates treated with PIP/TAZ, BSI caused by AMP/SLB-sensitive isolates treated with PIP/TAZ, and BSI caused by AMP/SLB-resistant isolates treated with another monotherapy. METHODS: This retrospective study was conducted in two academic centres in Europe. Adult patients with E. coli BSI were screened from 2014 to 2020. Inclusion criteria were non-ESBL BSI and initial monotherapy for ≥ 72 h. To reduce the expected bias between the patient groups, propensity score matching was performed. The primary outcome was early treatment response after 72 h and required absence of SOFA score increase in ICU/IMC patients, as well as resolution of fever, leukocytosis, and bacteraemia. RESULTS: Of the 1707 patients screened, 315 (18.5%) were included in the final analysis. Urinary tract infection was the most common source of BSI (54.9%). Monotherapies other than PIP/TAZ were cephalosporins (48.6%), carbapenems (34.3%), and quinolones (17.1%). Enhanced early treatment response rate was detected (p = 0.04) in patients with BSI caused by AMP/SLB-resistant isolates treated with another monotherapy (74.3%) compared to those treated with PIP/TAZ (57.1%), and was mainly driven by the use of cephalosporins and quinolones (p ≤ 0.03). Clinical success, 28-day mortality, and rate of relapsing BSI did not significantly differ between the groups. CONCLUSIONS: Our study suggests that initial use of PIP/TAZ may be associated with reduced early treatment response in E. coli BSI caused by AMP/SLB-resistant isolates compared to alternative monotherapies.

In vitro activity of imipenem/relebactam against non-Morganellaceae Enterobacterales and Pseudomonas aeruginosa in the Asia-Pacific region: SMART 2017-2020.

OBJECTIVES: To describe the in vitro activity of imipenem/relebactam (IMR) against non-Morganellaceae Enterobacterales (NME) and Pseudomonas aeruginosa, including piperacillin/tazobactam-nonsusceptible and meropenem-nonsusceptible isolates, infecting hospitalized patients in the Asia-Pacific region. METHODS: From 2017 to 2020, 49 clinical laboratories in nine countries in the Asia-Pacific region participated in the SMART global surveillance program and contributed 26 783 NME and 6383 P. aeruginosa. Minimum inhibitory concentrations (MICs) were determined using CLSI broth microdilution and interpreted using CLSI M100 (2021) breakpoints. ß-Lactamase genes were identified in selected isolate subsets (2017-2020) and oprD was sequenced in molecularly characterized P. aeruginosa collected in 2020. RESULTS: Amikacin (97.9% susceptible), IMR (95.8%), meropenem (95.4%), and imipenem (92.6%) were the most active agents against NME. Among piperacillin/tazobactam-nonsusceptible NME (n=4070), 76.1% were IMR-susceptible (range by country, 97.5% [New Zealand] to 50.6% [Vietnam]); 22.4% of meropenem-nonsusceptible NME (n=1225) were IMR-susceptible (range by country, 68.8% [South Korea] to 7.6% [Thailand]). A total of 2.7% of NME carried a metallo-ß-lactamase (MBL), 0.9% an OXA-48-like carbapenemase (MBL-negative), and 0.7% a KPC (MBL-negative). Amikacin (94.0% susceptible) and IMR (90.3%) were the most active agents against P. aeruginosa; 71.2% of isolates were imipenem-susceptible. Relebactam increased susceptibility to imipenem by 25.6% (from 40.5% to 66.1%) in piperacillin/tazobactam-nonsusceptible and by 44.8% (from 7.1% to 51.9%) in meropenem-nonsusceptible P. aeruginosa. Only 4.3% of P. aeruginosa were MBL-positive. A total of 70.3% (90/128) of IMR-nonsusceptible P. aeruginosa were oprD-deficient. CONCLUSION: In 2017-2020, 96% of NME and 90% of P. aeruginosa from the Asia-Pacific region were IMR-susceptible. IMR percent susceptible rates were higher in countries with lower MBL carriage.

In Vitro Susceptibility of Achromobacter Species Isolated from Cystic Fibrosis Patients: a 6-Year Survey.

We conducted in vitro antimicrobial susceptibility testing of 267 Achromobacter isolates for 16 antibiotics from 2017 to 2022. The highest susceptibility was found for piperacillin-tazobactam (70%) and ceftazidime-avibactam (62%). Between 30% and 49% of strains were susceptible to tigecycline, ceftazidime, and meropenem. We applied species-specific Achromobacter xylosoxidans breakpoints for piperacillin-tazobactam, meropenem, and trimethoprim-sulfamethoxazole and EUCAST pharmacokinetic/pharmacodynamic (PK/PD) breakpoints for the others. A. xylosoxidans was the most frequently isolated species, followed by Achromobacter insuavis and Achromobacter ruhlandii.

The Role of MexCD-OprJ and MexEF-OprN Efflux Systems in the Multiple Antibiotic Resistance of Pseudomonas aeruginosa Isolated from Clinical Samples.

Increasing antimicrobial resistance and the development of multi-drug resistant (MDR) Pseudomonas aeruginosa is dependent on the expression of efflux pumps. This study aimed to investigate the role of overexpression of MexCD-OprJ and MexEF-OprN efflux pumps in reduced susceptibility to antimicrobial agents among P. aeruginosa strains. Totally, 100 clinical isolates of P. aeruginosa were collected from patients and the strains were identified by standard diagnostic tests. The MDR isolates were detected using the disk agar diffusion method. The expression levels of MexCD-OprJ and MexEF-OprN efflux pumps were evaluated by the real-time PCR. Forty-one isolates showed MDR phenotype, while piperacillin-tazobactam and levofloxacin were the most- and least-effective antibiotics, respectively. Also, all 41 MDR isolates showed a more than tenfold increase in the expression of mexD and mexF genes. In this study, a significant relationship was observed between the rate of antibiotic resistance, the emergence of MDR strains, and increasing the expression levels of MexEF-OprN and MexCD-OprJ efflux pumps (P < 0.05). Efflux systems mediated resistance was a noteworthy mechanism causative to multidrug resistance in P. aeruginosa clinical isolates. The study results demonstrated mexE and mexF overexpression as the primary mechanism conferring in the emergence of MDR phenotypes among P. aeruginosa strains. In addition, we also show that piperacillin/tazobactam exhibited a stronger ability in the management of infections caused by MDR P. aeruginosa in this area.

Geographic patterns of antimicrobial susceptibilities for Bacteroides spp. worldwide: Results from the Antimicrobial Testing Leadership and Surveillance (ATLAS) programme, 2007-2020.

Antimicrobial susceptibilities of 4973 Bacteroides spp. isolates recovered from various sources of patients from 12 countries (99.6% from European countries) in the Antimicrobial Testing Leadership and Surveillance (ATLAS) programme, 2007-2020, were investigated. The minimum inhibitory concentrations (MICs) of the isolates with six commonly used agents were determined using the agar dilution method. Among the isolates, 10 Bacteroides spp. were included: B. fragilis (n=3180, 64.0%) was encountered most frequently, followed by B. thetaiotaomicron (n=675) and B. ovatus (n=409). During the 14 years, the proportion of B. fragilis declined, but the proportion of non-fragilis Bacteroides spp. increased. More than 90% of the isolates tested were susceptible to piperacillin-tazobactam, meropenem and tigecycline. Significantly lower susceptibility rates to cefoxitin (P<0.001), clindamycin (P<0.001), piperacillin/tazobactam (P<0.001) and tigecycline (P=0.006) were observed among non-fragilis Bacteroides spp. isolates than among B. fragilis isolates. Moreover, the susceptibility rates to clindamycin (P=0.003) and tigecycline (P=0.044) decreased significantly among non-fragilis Bacteroides spp. over time. Clindamycin susceptibility rates >80% were found in Greece (100%), Sweden (86.3%) and the UK (80.7%), and the lowest susceptibility rates were found in the USA (42.9%) and Japan (53.9%). In conclusion, the susceptibility of Bacteroides spp. to commonly used antibiotics varied geographically. Empirical antibiotic therapy for suspected anaerobic infections with clindamycin and cefoxitin should be avoided due to high resistance rates. Piperacillin-tazobactam, meropenem, metronidazole and tigecycline could be considered favourable options for the treatment of infections caused by Bacteroides spp.

EUCAST rapid antimicrobial susceptibility testing (RAST) compared to conventional susceptibility testing: implementation and potential added value in a tertiary hospital in Belgium.

OBJECTIVES: EUCAST breakpoints for short incubation disk diffusion allow rapid antimicrobial susceptibility testing (RAST) directly from positive blood cultures. We evaluate the RAST methodology and assess its potential added value in a setting of low prevalence of multidrug-resistant (MDR) organisms. METHODS: In our two-part study, we performed RAST on 127 clinical blood cultures at 6 and 8 h and determined categorical agreement with direct susceptibility testing. We also measure the impact of susceptibility results on antimicrobial therapy compared to empirical treatment. RESULTS: Categorical agreement was 96.2% at 6 h (575/598 isolate-drug combinations) and 96.6% at 8 h (568/588 combinations). Major errors involved piperacillin/tazobactam in 16 of 31 cases. The second part of our study shows that AST reporting proved essential in correcting ineffective empirical therapy in 6.3% of the patients (8/126). CONCLUSION: EUCAST RAST is an inexpensive and reliable method of susceptibility testing, although care must be taken with reporting piperacillin/tazobactam. In support of RAST implementation, we show that AST remains of great importance in providing effective therapy, even in a setting of low MDR prevalence and elaborate antibiotic guidelines.

Prevalence of Multidrug Resistant Pseudomonas aeruginosa Isolated from Clinical Specimens in Tertiary Care Hospital.

BACKGROUND: Pseudomonas aeruginosa is an opportunistic pathogen, which causes nosocomial infections in human. The rapid increase in drug resistance of this pathogen is a global concern. The aim of this study was to determine the clinical burden of P.aeruginosa, its antibiotic susceptibility pattern along with metallo-ß-lactamase (MBL) detection. METHODS: The descriptive cross-sectional study was conducted in Upendra Devkota Memorial National Institute of Neurological and Allied Sciences from January to August 2021. Isolation and identification of P. aeruginosa from clinical specimens was performed by using standard laboratory procedure. All bacterial isolates were phenotypically screened for multidrug resistance using Kirby Bauer disc diffusion method. All the multidrug resistant P.aeruginosa were phenotypically screened for MBL producer by Imipenem-EDTA combined disc diffusion test (CDDT). RESULTS: A total of 770 samples were processed of which 36 isolates of P. aeruginosa were obtained. P.aeruginosa was isolated mainly from tracheal aspirates, sputum, blood and urine. Among 36 isolates, 50% were found to be multidrug resistant (MDR). More percentage of P.aeruginosa isolates were found resistant to aztreonam, ofloxacin and levofloxacin (52.8%). Furthermore, this study reveals antibiotics like piperacillin/tazobactam and carbapenem were found to be good choice for the treatment of infection caused by this organism. Among MDR isolates 66.7% were found to be MBL producer. CONCLUSIONS: The data in this study highlights the prevalence of multidrug resistant, MBL producer, and colistin resistant P.aeruginosa in clinical specimens. In this study, carbapenems and piperacillin/tazobactam were found to be most effective antimicrobial drugs for empirical therapy in P.aeruginosa infections.

[Relationship of Hypervirulent Capsular Genotypes of Klebsiella pneumoniae with Antibiotic Susceptibility and Beta-Lactamase Genes]. / Klebsiella pneumoniae Hipervirülan Kapsül Genotiplerinin Antibiyotik Duyarliligi ve Beta-Laktamaz Genleri ile Iliskisi.

Klebsiella pneumoniae which is a causative agent of nosocomial infections, is protected from phagocytosis and the lethal effect of serum bactericidal proteins owing to its capsular polysaccharides (CPS). The important leading problem in treating infections caused by this bacterium that successfully develops antimicrobial resistance, especially via mobile genetic elements, is that it acquires beta-lactamase genes, which are responsible for the resistance against beta-lactam antibiotics. Due to the increase in studies targeting capsular polysaccharides in developing strategies for vaccination and treatment, we aimed to investigate the possible relationship of the capsular genotypes of K.pneumoniae isolates obtained from various clinical specimens with antibiotic susceptibility and beta-lactamase genes. In K.pneumoniae clinical isolates; K1, K2, K5, K20, K54 and K57, which are known as hypervirulent capsular types, were investigated by polymerase chain reaction (PCR) method. In isolates whose capsular genotypes were determined, antibiotic susceptibility was examined by Kirby-Bauer disc diffusion method. Colistin resistance was investigated by the broth microdilution method. Carbapenem resistance was confirmed with the carbapenem inactivation test. The beta-lactamase genes blaCTX-M1, blaCTX-M2, blaCTX-M9, blaCTX-M8/25, blaKPC, blaNDM-1, and blaOXA-48-like were investigated by using PCR. Of the 38 K.pneumoniae isolates whose capsular genotypes were determined, 15 (39.5%), 12 (31.6%), seven (18.4%), two (5.3%), one (2.6%) and one (2.6%) were K5-CPS, K2-CPS, K20-CPS, K1-CPS, K54-CPS and K57-CPS genotypes, respectively. blaOXA-48-like, blaNDM-1, and blaCTX-M1 were detected in 68.4, 10.5, and 7.9%, whereas coexistence of blaOXA-48-like with blaNDM-1, and blaOXA-48-like with blaCTX-M1 were determined in 7.9, and 5.3% of the isolates, respectively. The relationship of the blaCTX-M1 gene, detected only in three K20-CPS isolates, was found to be statistically significant with this genotype. In addition, of the blaNDM-1-positive four isolates, three were K5-CPS, and one was K2-CPS, while blaOXA-48-like was similarly detected mostly in K5-CPS and K2-CPS (10 and 9 isolates, respectively). Except for the two isolates that were resistant to colistin, one K1-CPS and the other K5-CPS, the highest resistance was detected against cefotaxime (36/38) and the lowest resistance was detected against gentamicin (23/38) as a result of antibiotic susceptibility tests. The resistance relationship of K5-CPS isolates with amoxicillin/clavulanate, piperacillin/tazobactam, cefoxitin and ertapenem and the susceptibility relationship of K20-CPs isolates with amoxicillin/clavulanate, piperacillin/tazobactam, imipenem, tetracycline and trimethoprim/sulfamethoxazole were found as statistically significant. Our study is the first to investigate the relationship between K.pneumoniae capsular genotypes and beta-lactamase genes in Turkey. As a result, it is believed that this research will contribute to the determination of vaccine and treatment options by providing data to wider and regional studies that will examine other capsule genotypes and antibiotic resistance genes to clarify the importance of the capsule in virulence.

Suppressive effect of therapeutic antibiotic regimen on antipneumococcal Th1/Th17 responses in neonatal mice.

BACKGROUND: Antibiotics are commonly used in human neonates, but their impact on neonatal T cell immunity remains poorly understood. The aim of this study was to investigate the impact of the antibiotic piperacillin with the beta-lactamase inhibitor tazobactam on neonatal CD4+ and CD8+ T cell responses to Streptococcus pneumoniae. METHODS: Splenic and lung cells were isolated from the neonatal mice receiving piperacillin and tazobactam or saline (sham) and cultured with S. pneumoniae to analyze T cell cytokine production by ELISA and flow cytometry. RESULTS: Antibiotic exposure to neonatal mice resulted in reduced numbers of CD4+/CD8+ T cells in the spleen and lungs compared to control mice. Upon in vitro stimulation with S. pneumoniae, splenocytes and lung cells from antibiotic-exposed mice produced lower levels of IFN-γ (Th1)/IL-17A (Th17) and IL-17A cytokines, respectively. Flow cytometric analysis revealed that S. pneumoniae-stimulated splenic CD4+ T cells from antibiotic-exposed mice expressed decreased levels of IFN-γ and IL-17A compared to control mice, whereas lung CD4+ T cells produced lower levels of IL-17A. However, no significant difference was observed for IL-4 (Th2) production. CONCLUSIONS: Neonatal mice exposure to piperacillin and tazobactam reduces the number of CD4+ and CD8+ T cells, and suppresses Th1 and Th17, but not Th2, responses to S. pneumoniae. IMPACT: Exposure of neonatal mice with a combination of piperacillin and tazobactam reduces CD4+/CD8+ T cells in the spleen and lungs. Antibiotic exposure suppresses neonatal Th1 and Th17, but not Th2, responses to Streptococcus pneumoniae. Our findings may have important implications for developing better therapeutic strategies in the neonatal intensive care unit.