Evaluation of antibacterial activity of five biocides and the synergistic effect of biocide/EDTA combinations on biofilm-producing and non-producing Stenotrophomonas maltophilia strains isolated from clinical specimens in Iran.

BACKGROUND: The overuse of biocides in healthcare-facilities poses risk for emergence and spread of antibiotic resistance among nosocomial pathogens. Hospital-acquired infections due to S. maltophilia have been increased in the recent years and with its various resistance mechanisms contribute to patient morbidity and mortality in hospitals. The current study aimed to evaluate the susceptibility of biofilm-producing and non-producing S. maltophilia clinical isolates to five commonly used hospital biocides, alone and in combination with EDTA to examine the synergistic effect of combining EDTA on the bactericidal activity of them by microbroth dilution method. As well as the frequency of efflux genes encoding resistance to biocides among isolates. This study also intended to assess the effect of exposure of S. maltophilia isolates to sub-inhibitory concentrations of sodium hypochlorite upon the antimicrobial susceptibility patterns. RESULTS: Based on minimum inhibitory and bactericidal concentrations of biocides sodium hypochlorite 5% (w/v) and ethyl alcohol 70% (v/v) were the strongest and weakest biocides against S. maltophilia isolates, respectively. The combination of EDTA with biocides significantly increased the effectiveness of the studied biocides. Exposure to sub-inhibitory concentration of sodium hypochlorite showed a significant change in the susceptibility of isolates towards ceftazidime (p = 0.019), ticarcillin/clavulanate (p = 0.009), and chloramphenicol (p = 0.028). As well as among the isolates examined, 94 (95%) were able to produce biofilm. The frequency of sugE1 resistance genes was found in 90.7% of our clinical S. maltophilia isolates. None of the isolates carried qacE and qacEΔ1 gene. CONCLUSIONS: The current study recommended that using the mixture of biocides with EDTA can be effective in reducing nosocomial infections. Also, this study demonstrated that exposure to sub-inhibitory concentrations of sodium hypochlorite leads to reduced antibiotic susceptibility and development of multidrug-resistant S. maltophilia strains.

Analysis of drug resistance genes of integrons in clinical isolates of Escherichia coli from elderly bloodstream infections.

This experiment was carried out to provide a basis for the treatment of clinical bloodstream infections by analyzing the drug resistance characteristics and integrated gene distribution of Escherichia coli in bloodstream infections in elderly patients. For this aim, E. coli were collected for bacterial identification and drug sensitivity testing from bloodstream infections in elderly patients in the hospital from January 2016 to December 2019. ESBLs positive strains were assayed for genotypes and their integron carriage rates by PCR amplification. The characteristics and differences of various genotype rates were compared and analyzed. Results showed that a total of 230 E. coli strains were isolated. The detection rate of ESBLs-producing bacteria was 37.39 %. ESBLs-producing E. coli showed a high rate of resistance to cefepime, levofloxacin, cotrimoxazole, and ticarcillin/clavulanic acid (>40%). The resistance rate of 230 strains of E. coli to meropenem, minocycline, amikacin, gentamicin and cefoxitin was less than 20%. Among the ESBLs-producing E. coli in bloodstream infections in elderly patients, CTX-M-9 accounted for 27.91%, CTX-M-2 for 17.44%, and SHV for 13.95%. The detection rate of type I integrated genes was 41.30%, and type II and III integrated genes were not detected. ESBLs-producing genotyping-positive bacteria were detected with more than 50% of type I integrated genes. It was concluded that type I integrated genes in ESBLs-producing E. coli isolated from elderly patients carried resistance genes such as CTX-M-9 and CTX-M-2 aggravating multi-drug resistance in bacteria.

Characteristics of Stenotrophomonas maltophilia isolates from cystic fibrosis patients in Russia.

Stenotrophomonas maltophilia is a common opportunistic microorganism and an important respiratory pathogen in cystic fibrosis (CF). The aim of this study was to determine antimicrobial resistance phenotypes, sequence-types (ST) and genetic determinants of antibiotic resistance in S. maltophilia strains recovered from CF patients in Russia. S. maltophilia isolates recovered from 170 CF patients were analyzed. Minimum inhibitory concentrations of antibacterial agents were determined using Sensititre Gram Negative GNX2F plates and the results were interpreted according to Clinical and Laboratory Standards Institute (CLSI) criteria. Whole-genome sequencing (WGS) was performed on MGISEQ-2000 platform. SPAdes software, Galaxy, ResFinder, Integrall and PubMLST were used for analysis of WGS data. S. maltophilia strains were identified from 24/170 (14%) CF patients. In total, 25 isolates were detected, two strains were isolated from the same patient. The isolates belonged to 17 different STs, including 5 new STs; ST4 was the most prevalent ST. Resistance to ceftazidime was observed in 60% of strains, to ticarcillin-clavulanate - in 32%, to levofloxacin - in 24%, to trimethoprim/sulfamethoxazole - in 12% of strains. All isolates were susceptible to minocycline. All ST4 isolates were resistant or intermediate to ceftazidime and ticarcillin-clavulanate. In two isolates, the sul1 gene was detected. In one isolate, sul1 was part of a class 1 integron. The detected integron also contained the blaGES-7 and aac(6')-Ib-cr genes. The ST4 sequence-type was the most prevalent ST among S. maltophilia strains recovered from CF patients in Russia. Antibiotic resistance genes, including sul1, blaGES-7, aac(6')-Ib-cr, were detected in single strains.

Antibiotic regimens for late-onset neonatal sepsis.

BACKGROUND: Neonatal sepsis is a major cause of morbidity and mortality. It is the third leading cause of neonatal mortality globally constituting 13% of overall neonatal mortality. Despite the high burden of neonatal sepsis, high-quality evidence in diagnosis and treatment is scarce. Due to the diagnostic challenges of sepsis and the relative immunosuppression of the newborn, many neonates receive antibiotics for suspected sepsis. Antibiotics have become the most used therapeutics in neonatal intensive care units, and observational studies in high-income countries suggest that 83% to 94% of newborns treated with antibiotics for suspected sepsis have negative blood cultures. The last Cochrane Review was updated in 2005. There is a need for an updated systematic review assessing the effects of different antibiotic regimens for late-onset neonatal sepsis. OBJECTIVES: To assess the beneficial and harmful effects of different antibiotic regimens for late-onset neonatal sepsis. SEARCH METHODS: We searched the following electronic databases: CENTRAL (2021, Issue 3); Ovid MEDLINE; Embase Ovid; CINAHL; LILACS; Science Citation Index EXPANDED and Conference Proceedings Citation Index - Science on 12 March 2021. We also searched clinical trials databases and the reference lists of retrieved articles for randomised controlled trials (RCTs) and quasi-RCTs. SELECTION CRITERIA: We included RCTs comparing different antibiotic regimens for late-onset neonatal sepsis. We included participants older than 72 hours of life at randomisation, suspected or diagnosed with neonatal sepsis, meningitis, osteomyelitis, endocarditis, or necrotising enterocolitis. We excluded trials that assessed treatment of fungal infections. DATA COLLECTION AND ANALYSIS: Three review authors independently assessed studies for inclusion, extracted data, and assessed risk of bias. We used the GRADE approach to assess the certainty of evidence. Our primary outcome was all-cause mortality, and our secondary outcomes were: serious adverse events, respiratory support, circulatory support, nephrotoxicity, neurological developmental impairment, necrotising enterocolitis, and ototoxicity. Our primary time point of interest was at maximum follow-up. MAIN RESULTS: We included five RCTs (580 participants). All trials were at high risk of bias, and had very low-certainty evidence. The five included trials assessed five different comparisons of antibiotics. We did not conduct a meta-analysis due to lack of relevant data. Of the five included trials one trial compared cefazolin plus amikacin with vancomycin plus amikacin; one trial compared ticarcillin plus clavulanic acid with flucloxacillin plus gentamicin; one trial compared cloxacillin plus amikacin with cefotaxime plus gentamicin; one trial compared meropenem with standard care (ampicillin plus gentamicin or cefotaxime plus gentamicin); and one trial compared vancomycin plus gentamicin with vancomycin plus aztreonam. None of the five comparisons found any evidence of a difference when assessing all-cause mortality, serious adverse events, circulatory support, nephrotoxicity, neurological developmental impairment, or necrotising enterocolitis; however, none of the trials were near an information size that could contribute significantly to the evidence of the comparative benefits and risks of any particular antibiotic regimen. None of the trials assessed respiratory support or ototoxicity. The benefits and harms of different antibiotic regimens remain unclear due to the lack of well-powered trials and the high risk of systematic errors. AUTHORS' CONCLUSIONS: Current evidence is insufficient to support any antibiotic regimen being superior to another. RCTs assessing different antibiotic regimens in late-onset neonatal sepsis with low risks of bias are warranted.

ANTECEDENTES: La sepsis neonatal es una causa importante de morbilidad y mortalidad. Es la tercera causa de mortalidad neonatal a nivel mundial y constituye el 13% de la mortalidad neonatal total. A pesar de la elevada carga de la sepsis neonatal, la evidencia de alta calidad en el diagnóstico y el tratamiento es escasa. Debido a las dificultades de diagnóstico de la sepsis y a la relativa inmunosupresión del neonato, muchos reciben antibióticos por sospecha de sepsis. Los antibióticos se han convertido en el tratamiento más utilizado en las unidades de cuidados intensivos neonatales, y los estudios observacionales realizados en países de ingresos altos indican que entre el 83% y el 94% de los neonatos tratados con antibióticos por sospecha de sepsis tienen hemocultivos negativos. La última revisión Cochrane se actualizó en 2005. Se necesita una revisión sistemática actualizada que evalúe los efectos de los diferentes regímenes de antibióticos para la sepsis neonatal de inicio tardío. OBJETIVOS: Evaluar los efectos beneficiosos y perjudiciales de diferentes regímenes antibióticos para la sepsis neonatal de inicio tardío. MÉTODOS DE BÚSQUEDA: Se hicieron búsquedas en las siguientes bases de datos electrónicas: CENTRAL (2021, número 3); Ovid MEDLINE; Embase Ovid; CINAHL; LILACS; Science Citation Index EXPANDED y Conference Proceedings Citation Index ­ Science el 12 de marzo de 2021. También se buscaron ensayos controlados aleatorizados (ECA) y cuasialeatorizados en las bases de datos de ensayos clínicos y en las listas de referencias de artículos identificados. CRITERIOS DE SELECCIÓN: Se incluyeron ECA que compararon diferentes regímenes de antibióticos para la sepsis neonatal de inicio tardío. Se incluyeron participantes mayores de 72 horas de vida en el momento de la asignación al azar, con sospecha o diagnóstico de sepsis neonatal, meningitis, osteomielitis, endocarditis o enterocolitis necrosante. Se excluyeron los ensayos que evaluaron el tratamiento de las infecciones micóticas. OBTENCIÓN Y ANÁLISIS DE LOS DATOS: Dos autores de la revisión, de forma independiente, evaluaron los estudios para inclusión, extrajeron los datos y evaluaron el riesgo de sesgo. Se utilizó el método GRADE para evaluar la certeza de la evidencia. El desenlace principal fue la mortalidad por todas las causas, y los desenlaces secundarios fueron: eventos adversos graves, asistencia respiratoria, apoyo circulatorio, nefrotoxicidad, deterioro del desarrollo neurológico, enterocolitis necrosante y ototoxicidad. El punto temporal principal de interés fue el seguimiento máximo. RESULTADOS PRINCIPALES: Se incluyeron cinco ECA (580 participantes). Todos los ensayos tuvieron alto riesgo de sesgo y evidencia de certeza muy baja. Los cinco ensayos incluidos evaluaron cinco comparaciones diferentes de antibióticos. No se realizó un metanálisis debido a la falta de datos relevantes. De los cinco ensayos incluidos, un ensayo comparó cefazolina más amikacina con vancomicina más amikacina; un ensayo comparó ticarcilina más ácido clavulánico con flucloxacilina más gentamicina; un ensayo comparó cloxacilina más amikacina con cefotaxima más gentamicina; un ensayo comparó meropenem con atención estándar (ampicilina más gentamicina o cefotaxima más gentamicina); y un ensayo comparó vancomicina más gentamicina con vancomicina más aztreonam. Ninguna de las cinco comparaciones encontró evidencia de una diferencia al evaluar la mortalidad por todas las causas, los eventos adversos graves, el apoyo circulatorio, la nefrotoxicidad, el deterioro del desarrollo neurológico o la enterocolitis necrosante; sin embargo, ninguno de los ensayos se acercó a un tamaño de información que pudiera contribuir significativamente a la evidencia de los beneficios y los riesgos comparativos de cualquier régimen antibiótico en particular. Ninguno de los ensayos evaluó la asistencia respiratoria o la ototoxicidad. Los efectos beneficiosos y perjudiciales de los diferentes regímenes de antibióticos aún no están claros debido a la falta de ensayos con un poder estadístico adecuado y al alto riesgo de errores sistemáticos. CONCLUSIONES DE LOS AUTORES: La evidencia actual no es suficiente para apoyar que un régimen de antibióticos sea superior a otro. Se justifica la realización de ECA con bajo riesgo de sesgo que evalúen diferentes regímenes antibióticos en la sepsis neonatal de inicio tardío.

Resurrecting Old ß-Lactams: Potent Inhibitory Activity of Temocillin against Multidrug-Resistant Burkholderia Species Isolates from the United States.

Burkholderia spp. are opportunistic human pathogens that infect persons with cystic fibrosis and the immunocompromised. Burkholderia spp. express class A and C ß-lactamases, which are transcriptionally regulated by PenRA through linkage to cell wall metabolism and ß-lactam exposure. The potency of temocillin, a 6-methoxy-ß-lactam, was tested against a panel of multidrug-resistant (MDR) Burkholderia spp. In addition, the mechanistic basis of temocillin activity was assessed and compared to that of ticarcillin. Susceptibility testing with temocillin and ticarcillin was conducted, as was biochemical analysis of the PenA1 class A ß-lactamase and AmpC1 class C ß-lactamase. Molecular dynamics simulations (MDS) were performed using PenA1 with temocillin and ticarcillin. The majority (86.7%) of 150 MDR Burkholderia strains were susceptible to temocillin, while only 4% of the strains were susceptible to ticarcillin. Neither temocillin nor ticarcillin induced bla expression. Ticarcillin was hydrolyzed by PenA1 (kcat/Km = 1.7 ± 0.2 µM-1 s-1), while temocillin was slow to form a favorable complex (apparent Ki [Ki app] = ∼2 mM). Ticarcillin and temocillin were both potent inhibitors of AmpC1, with Ki app values of 4.9 ± 1.0 µM and 4.3 ± 0.4 µM, respectively. MDS of PenA revealed that ticarcillin is in an advantageous position for acylation and deacylation. Conversely, with temocillin, active-site residues K73 and S130 are rotated and the catalytic water molecule is displaced, thereby slowing acylation and allowing the 6-methoxy of temocillin to block deacylation. Temocillin is a ß-lactam with potent activity against Burkholderia spp., as it does not induce bla expression and is poorly hydrolyzed by endogenous ß-lactamases.

Influence of the α-Methoxy Group on the Reaction of Temocillin with Pseudomonas aeruginosa PBP3 and CTX-M-14 ß-Lactamase.

The prevalence of multidrug-resistant Pseudomonas aeruginosa has led to the reexamination of older "forgotten" drugs, such as temocillin, for their ability to combat resistant microbes. Temocillin is the 6-α-methoxy analogue of ticarcillin, a carboxypenicillin with well-characterized antipseudomonal properties. The α-methoxy modification confers resistance to serine ß-lactamases, yet temocillin is ineffective against P. aeruginosa growth. The origins of temocillin's inferior antibacterial properties against P. aeruginosa have remained relatively unexplored. Here, we analyze the reaction kinetics, protein stability, and binding conformations of temocillin and ticarcillin with penicillin-binding protein 3 (PBP3), an essential PBP in P. aeruginosa We show that the 6-α-methoxy group perturbs the stability of the PBP3 acyl-enzyme, which manifests in an elevated off-rate constant (koff) in biochemical assays comparing temocillin with ticarcillin. Complex crystal structures with PBP3 reveal similar binding modes of the two drugs but with important differences. Most notably, the 6-α-methoxy group disrupts a high-quality hydrogen bond with a conserved residue important for ligand binding while also being inserted into a crowded active site, possibly destabilizing the active site and enabling water molecule from bulk solvent to access and cleave the acyl-enzyme bond. This hypothesis is supported by the observation that the acyl-enzyme complex of temocillin has reduced thermal stability compared with ticarcillin. Furthermore, we explore temocillin's mechanism of ß-lactamase inhibition with a high-resolution complex structure of CTX-M-14 class A serine ß-lactamase. The results suggest that the α-methoxy group prevents hydrolysis by locking the compound into an unexpected conformation that impedes access of the catalytic water to the acyl-enzyme adduct.

Pseudomonas aeruginosa eradication therapy and risk of acquiring Aspergillus in young children with cystic fibrosis.

BACKGROUND: While Aspergillus detection rates in adults, adolescents and older children with cystic fibrosis (CF) have increased, the risk of acquiring this fungal pathogen in young children is unknown. AIM: To determine the risk and explanatory factors of acquiring Aspergillus in children with CF by age 5 years. METHODS: Cross-sectional analysis of clinical, bronchoalveolar lavage and treatment data from the Australasian Cystic Fibrosis Bronchoalveolar Lavage study was used to identify predictive factors for detecting Aspergillus at age 5 years. A parametric repeated time-to-event model quantitatively described the risk and factors associated with acquiring Aspergillus and Pseudomonas aeruginosa from birth until age 5 years. RESULTS: Cross-sectional analysis found that the number of P. aeruginosa eradication courses increased the odds of detecting Aspergillus at age 5 years (OR 1.61, 95% CI 1.23 to 2.12). The median (IQR) age for the first P. aeruginosa positive culture was 2.38 (1.32-3.79) years and 3.69 (1.68-4.74) years for the first Aspergillus positive culture. The risk of P. aeruginosa and Aspergillus events changes with time after the first year of study entry. It also decreases for P. aeruginosa after completing P. aeruginosa eradication (HR 0.15, 95% CI 0.00 to 0.79), but increases for Aspergillus events (HR 2.75, 95% CI 1.45 to 5.41). The risk of acquiring both types of events increases after having had a previous event. CONCLUSION: In young children with CF, completing P. aeruginosa eradication therapy and previous Aspergillus events are associated with increased risk of acquiring Aspergillus.

Pharmacokinetics of ticarcillin-clavulanate in premature infants.

Ticarcillin-clavulanate covers a broad spectrum of pathogens that are common in premature infants. In infants <30 weeks gestational age, pharmacokinetic data to guide ticarcillin-clavulanate dosing are lacking. We enrolled 15 premature infants <30 weeks gestational age, determined pharmacokinetic parameters, and performed dosing simulations to determine optimal dosing for ticarcillin-clavulanate. The infants had a median (range) postnatal age (PNA) of 18 days (6-44 days) and gestational age of 25 weeks (23-28 weeks). Clearance was lower in infants with a PNA <14 days (0.050 L/kg/h [range 0.043-0.075]) compared with a PNA ≥14-45 days (0.078 L/kg/h [0.047-0.100]), consistent with maturation of renal function. Dosing simulations determined that ticarcillin 75 mg/kg q12h (PNA <14 days) or q8h (PNA ≥ 14-45 days) achieved the target exposure for organisms with a minimum inhibitory concentration ≤16 µ/mL in >90% of simulated infants. For highly resistant organisms (minimum inhibitory concentration 32 µg/mL), increased dosing frequency or extended infusion are necessary.

Impacts of L1 Promoter Variation and L2 Clavulanate Susceptibility on Ticarcillin-Clavulanate Susceptibility of Stenotrophomonas maltophilia.

Inducible expression of L1 and L2 ß-lactamases is the principal mechanism responsible for ß-lactam resistance in Stenotrophomonas maltophilia Ticarcillin-clavulanate (TIM) is one of the few effective ß-lactams for S. maltophilia treatment. Clavulanate (CA) is a ß-lactamase inhibitor that specifically targets class A, C, and D ß-lactamases. In view of the presence of class B L1 ß-lactamase, it is of interest to elucidate why TIM is valid for S. maltophilia treatment. The L1-L2 allelic variation and TIM susceptibilities of 22 clinical isolates were established. Based on L1 and L2 protein sequences and TIM susceptibility, three L1-based phylogenetic clusters (L1-A, L1-B, and L1-C) and three L2-based phylogenetic clusters (L2-A, L2-B1, and L2-B2) were classified. The contribution of each L1- and L2-based phylogenetic cluster to ticarcillin (TIC) and TIM susceptibility was investigated. All the L1s and L2s tested contributed to TIC resistance. The L1s tested were inert to CA; nevertheless, the sensitivities of L2s to CA were widely different. In addition, the genetic organizations upstream of the L1 gene varied greatly in these isolates. At least three different L1 promoter structures (K279a type, D457 type, and none) were found among the 22 isolates assayed. The differences in the L1 promoter structure had a great impact on TIC-induced L1 ß-lactamase activities. Collectively, the L1 promoter activity in response to TIC challenge and L2 susceptibility to CA are critical factors determining TIM susceptibility in S. maltophilia.

Inhaled antibiotics for pulmonary exacerbations in cystic fibrosis.

BACKGROUND: Cystic fibrosis is a genetic disorder in which abnormal mucus in the lungs is associated with susceptibility to persistent infection. Pulmonary exacerbations are when symptoms of infection become more severe. Antibiotics are an essential part of treatment for exacerbations and inhaled antibiotics may be used alone or in conjunction with oral antibiotics for milder exacerbations or with intravenous antibiotics for more severe infections. Inhaled antibiotics do not cause the same adverse effects as intravenous antibiotics and may prove an alternative in people with poor access to their veins. This is an update of a previously published review. OBJECTIVES: To determine if treatment of pulmonary exacerbations with inhaled antibiotics in people with cystic fibrosis improves their quality of life, reduces time off school or work and improves their long-term survival. SEARCH METHODS: We searched the Cochrane Cystic Fibrosis Group's Cystic Fibrosis Trials Register. Date of the last search: 03 October 2018.We searched ClinicalTrials.gov, the Australia and New Zealand Clinical Trials Registry and WHO ICTRP for relevant trials. Date of last search: 09 October 2018. SELECTION CRITERIA: Randomised controlled trials in people with cystic fibrosis with a pulmonary exacerbation in whom treatment with inhaled antibiotics was compared to placebo, standard treatment or another inhaled antibiotic for between one and four weeks. DATA COLLECTION AND ANALYSIS: Two review authors independently selected eligible trials, assessed the risk of bias in each trial and extracted data. They assessed the quality of the evidence using the GRADE criteria. Authors of the included trials were contacted for more information. MAIN RESULTS: Four trials with 167 participants are included in the review. Two trials (77 participants) compared inhaled antibiotics alone to intravenous antibiotics alone and two trials (90 participants) compared a combination of inhaled and intravenous antibiotics to intravenous antibiotics alone. Trials were heterogenous in design and two were only available in abstract form. Risk of bias was difficult to assess in most trials, but for all trials we judged there to be a high risk from lack of blinding and an unclear risk with regards to randomisation. Results were not fully reported and only limited data were available for analysis.Inhaled antibiotics alone versus intravenous antibiotics aloneOnly one trial (n = 18) reported a perceived improvement in lifestyle (quality of life) in both groups (very low-quality of evidence). Neither trial reported on time off work or school. Both trials measured lung function, but there was no difference reported between treatment groups (very low-quality evidence). With regards to our secondary outcomes, one trial (n = 18) reported no difference in the need for additional antibiotics and the second trial (n = 59) reported on the time to next exacerbation. In neither case was a difference between treatments identified (both very low-quality evidence). The single trial (n = 18) measuring adverse events and sputum microbiology did not observe any in either treatment group for either outcome (very low-quality evidence).Inhaled antibiotics plus intravenous antibiotics versus intravenous antibiotics aloneNeither trial reported on quality of life or time off work or school. Both trials measured lung function, but found no difference between groups in forced expiratory volume in one second (one trial, n = 28, very low-quality evidence) or vital capacity (one trial, n = 62). Neither trial reported on the need for additional antibiotics or the time to the next exacerbation; however, one trial (n = 28) reported on hospital admissions and found no difference between groups. Both trials reported no difference between groups in adverse events (very low-quality evidence) and one trial (n = 62) reported no difference in the emergence of antibiotic-resistant organisms (very low-quality evidence). AUTHORS' CONCLUSIONS: There is little useful high-level evidence to judge the effectiveness of inhaled antibiotics for the treatment of pulmonary exacerbations in people with cystic fibrosis. The included trials were not sufficiently powered to achieve their goals. Hence, we are unable to demonstrate whether one treatment was superior to the other or not. Further research is needed to establish whether inhaled tobramycin may be used as an alternative to intravenous tobramycin for some pulmonary exacerbations.

Degradation of ticarcillin by subcritical water oxidation method: Application of response surface methodology and artificial neural network modeling.

This study was performed to investigate the mineralization of ticarcillin in the artificially prepared aqueous solution presenting ticarcillin contaminated waters, which constitute a serious problem for human health. 81.99% of total organic carbon removal, 79.65% of chemical oxygen demand removal, and 94.35% of ticarcillin removal were achieved by using eco-friendly, time-saving, powerful and easy-applying, subcritical water oxidation method in the presence of a safe-to-use oxidizing agent, hydrogen peroxide. Central composite design, which belongs to the response surface methodology, was applied to design the degradation experiments, to optimize the methods, to evaluate the effects of the system variables, namely, temperature, hydrogen peroxide concentration, and treatment time, on the responses. In addition, theoretical equations were proposed in each removal processes. ANOVA tests were utilized to evaluate the reliability of the performed models. F values of 245.79, 88.74, and 48.22 were found for total organic carbon removal, chemical oxygen demand removal, and ticarcillin removal, respectively. Moreover, artificial neural network modeling was applied to estimate the response in each case and its prediction and optimizing performance was statistically examined and compared to the performance of central composite design.

Comparison of Two Phenotypic Algorithms To Detect Carbapenemase-Producing Enterobacteriaceae.

A novel algorithm designed for the screening of carbapenemase-producing Enterobacteriaceae (CPE), based on faropenem and temocillin disks, was compared to that of the Committee of the Antibiogram of the French Society of Microbiology (CA-SFM), which is based on ticarcillin-clavulanate, imipenem, and temocillin disks. The two algorithms presented comparable negative predictive values (98.6% versus 97.5%) for CPE screening among carbapenem-nonsusceptible Enterobacteriaceae However, since 46.2% (n = 49) of the CPE were correctly identified as OXA-48-like producers by the faropenem/temocillin-based algorithm, it significantly decreased the number of complementary tests needed (42.2% versus 62.6% with the CA-SFM algorithm).

Development of an algorithm for phenotypic screening of carbapenemase-producing Enterobacteriaceae in the routine laboratory.

BACKGROUND: Carbapenemase-producing Enterobacteriaceae (CPE) are difficult to identify among carbapenem non-susceptible Enterobacteriaceae (NSE). We designed phenotypic strategies giving priority to high sensitivity for screening putative CPE before further testing. METHODS: Presence of carbapenemase-encoding genes in ertapenem NSE (MIC > 0.5 mg/l) consecutively isolated in 80 French laboratories between November 2011 and April 2012 was determined by the Check-MDR-CT103 array method. Using the Mueller-Hinton (MH) disk diffusion method, clinical diameter breakpoints of carbapenems other than ertapenem, piperazicillin+tazobactam, ticarcillin+clavulanate and cefepime as well as diameter cut-offs for these antibiotics and temocillin were evaluated alone or combined to determine their performances (sensitivity, specificity, positive and negative likelihood ratios) for identifying putative CPE among these ertapenem-NSE isolates. To increase the screening specificity, these antibiotics were also tested on cloxacillin-containing MH when carbapenem NSE isolates belonged to species producing chromosomal cephalosporinase (AmpC) but Escherichia coli. RESULTS: Out of the 349 ertapenem NSE, 52 (14.9%) were CPE, including 39 producing OXA-48 group carbapenemase, eight KPC and five MBL. A screening strategy based on the following diameter cut offs, ticarcillin+clavulanate <15 mm, temocillin <15 mm, meropenem or imipenem <22 mm, and cefepime <26 mm, showed 100% sensitivity and 68.1% specificity with the better likelihood ratios combination. The specificity increased when a diameter cut-off <32 mm for imipenem (76.1%) or meropenem (78.8%) further tested on cloxacillin-containing MH was added to the previous strategy for AmpC-producing isolates. CONCLUSION: The proposed strategies that allowed for increasing the likelihood of CPE among ertapenem-NSE isolates should be considered as a surrogate for carbapenemase production before further CPE confirmatory testing.

Intra- and Interspecies Effects of Outer Membrane Vesicles from Stenotrophomonas maltophilia on ß-Lactam Resistance.

The treatment ofStenotrophomonas maltophiliainfection with ß-lactam antibiotics leads to increased release of outer membrane vesicles (OMVs), which are packed with two chromosomally encoded ß-lactamases. Here, we show that these ß-lactamase-packed OMVs are capable of establishing extracellular ß-lactam degradation. We also show that they dramatically increase the apparent MICs of imipenem and ticarcillin for the cohabituating speciesPseudomonas aeruginosaandBurkholderia cenocepacia.

The characteristics of genetically related Pseudomonas aeruginosa from diverse sources and their interaction with human cell lines.

We investigated a collection of Pseudomonas aeruginosa strains from hospitalised patients (n = 20) and various environmental sources (n = 214) for their genetic relatedness; virulence properties; antibiotic resistance; and interaction with intestinal (Caco-2), renal (A-498), and lung (Calu-3) cell lines. Using RAPD-PCR, we found high diversity among the strains irrespective of their sources, with only 6 common (C) types containing strains from both a clinical and environmental source. Environmental strains belonging to these C-types showed greater adhesion to A-498 cells than did clinical strains (17 ± 13 bacteria/cell versus 13 ± 11 bacteria/cell; p < 0.001), whereas clinical strains showed significantly greater adhesion to Calu-3 and Caco-2 cells than did environmental strains (p < 0.001 for both). The virulence genes and antibiotic resistance profiles of the strains were similar; however, the prevalence of environmental strains carrying both exoS and exoU was significantly (p < 0.0368) higher than clinical strains. While all strains were resistant to ticarcillin and ticarcillin-clavulanic acid, resistance against aztreonam, gentamicin, amikacin, piperacillin, and ceftazidime varied among environmental and clinical strains. These results suggest that environmental strains of P. aeruginosa carry virulence properties similar to clinical strains, including adhesion to various human cell lines, with some strains showing a higher adhesion to specific cell lines, indicating they may have a better ability to cause infection in those sites under predisposing conditions of the host.

Higher risk of neutropenia associated with piperacillin-tazobactam compared with ticarcillin-clavulanate in children.

BACKGROUND: A series of cases of piperacillin-tazobactam (P/T)-associated neutropenia has been observed recently in children in our center. Because neutropenia was seldom observed in children treated with ticarcillin-clavulanic acid (T/C), we conducted a study to determine if there is an increased risk of neutropenia in children exposed to P/T in comparison with T/C. METHODS: Medical records of subjects aged <18 years who received at least 1 dose of P/T or T/C between 1 January 2008 and 30 June 2011 were reviewed. RESULTS: Two hundred ninety-nine episodes of treatment (65 P/T, 234 T/C) met inclusion criteria. Among those episodes, 213 had data allowing complete white blood cell count analysis and were included in the final analysis. Thirteen cases of neutropenia were observed during the study period. The average time to onset was 17.6 days and all patients were aged <13 years. Seven cases (10.8%) occurred in the P/T group and 6 (2.6%) in the T/C group (unadjusted odds ratio, 4.59; 95% confidence interval, 1.48-14.17). Although a statistically significant correlation was observed between age, treatment duration, and total dose and the development of neutropenia (r = -0.121, P = .037; r = 0.267, P < .001; r = 0.260, P < .001, respectively), this was not the case for sex, indications, neutrophil count at initiation, and concomitant drug treatments. CONCLUSIONS: Although our results need to be confirmed, they suggest that children receiving long courses of therapy (>2 weeks) with P/T may be at increased risk of neutropenia, compared with T/C.

Mutations in ß-Lactamase AmpC Increase Resistance of Pseudomonas aeruginosa Isolates to Antipseudomonal Cephalosporins.

Mutation-dependent overproduction of intrinsic ß-lactamase AmpC is considered the main cause of resistance of clinical strains of Pseudomonas aeruginosa to antipseudomonal penicillins and cephalosporins. Analysis of 31 AmpC-overproducing clinical isolates exhibiting a greater resistance to ceftazidime than to piperacillin-tazobactam revealed the presence of 17 mutations in the ß-lactamase, combined with various polymorphic amino acid substitutions. When overexpressed in AmpC-deficient P. aeruginosa 4098, the genes coding for 20/23 of these AmpC variants were found to confer a higher (2-fold to >64-fold) resistance to ceftazidime and ceftolozane-tazobactam than did the gene from reference strain PAO1. The mutations had variable effects on the MICs of ticarcillin, piperacillin-tazobactam, aztreonam, and cefepime. Depending on their location in the AmpC structure and their impact on ß-lactam MICs, they could be assigned to 4 distinct groups. Most of the mutations affecting the omega loop, the R2 domain, and the C-terminal end of the protein were shared with extended-spectrum AmpCs (ESACs) from other Gram-negative species. Interestingly, two new mutations (F121L and P154L) were predicted to enlarge the substrate binding pocket by disrupting the stacking between residues F121 and P154. We also found that the reported ESACs emerged locally in a variety of clones, some of which are epidemic and did not require hypermutability. Taken together, our results show that P. aeruginosa is able to adapt to efficacious ß-lactams, including the newer cephalosporin ceftolozane, through a variety of mutations affecting its intrinsic ß-lactamase, AmpC. Data suggest that the rates of ESAC-producing mutants are ≥1.5% in the clinical setting.

Stenotrophomonas maltophilia infection among young children in a cardiac intensive care unit: a single institution experience.

Stenotrophomonas maltophilia can present as bacteremia, respiratory tract infection, urinary tract infection, soft tissue and wound infections, bone and joint infections, meningitis, and endocarditis especially in immunosuppressed patients and those with underlying medical conditions. The incidence and impact of S. maltophilia in young children with heart disease are poorly defined. A single center retrospective observational study was conducted in infants <180 days of age with positive S. maltophilia cultures over a period of 5 years. The overall incidence for S. maltophilia infection was 0.8 % (n = 32/3656). Among 32 identified infants, there were 47 episodes of S. maltophilia infection 66 % of infants had prior exposure to broad spectrum antibiotics. 97 % of positive isolates were susceptible to trimethoprim/sulfamethoxazole and 91 % to levofloxacin as well as ticarcillin/clavulanate. Ventilator-free days and absolute lymphocyte count prior to acquiring infection were significantly lower in non-survivors than in survivors. 100 % of survivors had clearance of positive cultures compared to 50 % in non-survivors (p < 0.05). The crude all-cause mortality rate was 37.5 %. All non-survivors had increased length of ICU stay and duration of mechanical ventilation and had delayed clearance of infection and required longer duration of treatment.

Agrobacterium-mediated genetic transformation and plant regeneration of the hardwood tree species Fraxinus profunda.

This transformation and regeneration protocol provides an integral framework for the genetic improvement of Fraxinus profunda (pumpkin ash) for future development of plants resistant to the emerald ash borer. Using mature hypocotyls as the initial explants, an Agrobacterium tumefaciens-mediated genetic transformation system was successfully developed for pumpkin ash (Fraxinus profunda). This transformation protocol is an invaluable tool to combat the highly aggressive, non-native emerald ash borer (EAB), which has the potential to eliminate native Fraxinus spp. from the natural landscape. Hypocotyls were successfully transformed with Agrobacterium strain EHA105 harboring the pq35GR vector, containing an enhanced green fluorescent protein (EGFP) as well as a fusion gene between neomycin phosphotransferase (nptII) and gusA. Hypocotyls were cultured for 7 days on Murashige and Skoog (MS) medium with 22.2 µM 6-benzyladenine (BA), 4.5 µM thidiazuron (TDZ), 50 mg L(-1) adenine hemisulfate (AS), and 10 % coconut water (CW) prior to transformation. Hypocotyls were transformed using 90 s sonication plus 10 min vacuum infiltration after Agrobacterium was exposed to 100 µM acetosyringone for 1 h. Adventitious shoots were regenerated on MS medium with 22.2 µM BA, 4.5 µM TDZ, 50 mg L(-1) AS, 10 % CW, 400 mg L(-1) timentin, and 20 mg L(-1) kanamycin. Timentin at 400 and 20 mg L(-1) kanamycin were most effective at controlling Agrobacterium growth and selecting for transformed cells, respectively. The presence of nptII, GUS (ß-glucuronidase), and EGFP in transformed plants was confirmed using polymerase chain reaction (PCR), while the expression of EGFP was also confirmed through fluorescent microscopy and reverse transcription-PCR. This transformation protocol provides an integral foundation for future genetic modifications of F. profunda to provide resistance to EAB.

Accumulation of amikacin in synovial fluid after regional limb perfusion of amikacin sulfate alone and in combination with ticarcillin/clavulanate in horses.

OBJECTIVES: To determine the effect of regional limb perfusion (RLP) with amikacin sulfate alone and in combination with ticarcillin/clavulanate on synovial fluid concentration and antimicrobial activity of amikacin. SAMPLE POPULATION: Experimental study. METHODS: RLP with amikacin alone (A; 2.5 g) or amikacin and ticarcillin/clavulanate (AT; 2.5 g amikacin, 7 g ticarcillin/clavulanate) was performed with a tourniquet placed at mid-antebrachium in standing, sedated horses. Perfusate blood was collected immediately after injection and again before tourniquet release. Blood from the jugular vein was collected before tourniquet release. Synovial fluid from the middle carpal joint was collected 0, 30, and 60 minutes after tourniquet release. Amikacin concentration and antimicrobial activity of synovial fluid against Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, and Pseudomonas aeruginosa were determined. RESULTS: There was significantly lower amikacin concentration in the middle carpal joint synovial fluid of group AT compared with group A at 30 minutes (AT = median 4.4 µg/mL, IQR 3.0-11.2 µg/mL; A = 17.5 µg/mL, 6.6-80.1 µg/mL) and 60 minutes (AT = median 4.6 µg/mL, IQR 3.1-8.1 µg/mL; A = 15.0 µg/mL, 6.7-61.7 µg/mL) after tourniquet release. Zones of inhibition for ticarcillin-resistant Klebsiella pneumoniae from group AT were significantly smaller than group A from synovial fluid at 30 and 60 minutes after tourniquet release and in the perfusate serum before tourniquet release. CONCLUSIONS: The combination of amikacin with ticarcillin/clavulanate during RLP resulted in significantly lower amikacin synovial concentration and antimicrobial activity on amikacin susceptible and ticarcillin resistant cultures compared with amikacin alone.