Electron transport chain capacity expands yellow fever vaccine immunogenicity.

Vaccination has successfully controlled several infectious diseases although better vaccines remain desirable. Host response to vaccination studies have identified correlates of vaccine immunogenicity that could be useful to guide development and selection of future vaccines. However, it remains unclear whether these findings represent mere statistical correlations or reflect functional associations with vaccine immunogenicity. Functional associations, rather than statistical correlates, would offer mechanistic insights into vaccine-induced adaptive immunity. Through a human experimental study to test the immunomodulatory properties of metformin, an anti-diabetic drug, we chanced upon a functional determinant of neutralizing antibodies. Although vaccine viremia is a known correlate of antibody response, we found that in healthy volunteers with no detectable or low yellow fever 17D viremia, metformin-treated volunteers elicited higher neutralizing antibody titers than placebo-treated volunteers. Transcriptional and metabolomic analyses collectively showed that a brief course of metformin, started 3 days prior to YF17D vaccination and stopped at 3 days after vaccination, expanded oxidative phosphorylation and protein translation capacities. These increased capacities directly correlated with YF17D neutralizing antibody titers, with reduced reactive oxygen species response compared to placebo-treated volunteers. Our findings thus demonstrate a functional association between cellular respiration and vaccine-induced humoral immunity and suggest potential approaches to enhancing vaccine immunogenicity.

In Vitro Susceptibility to Ceftazidime-Avibactam and Comparator Antimicrobial Agents of Carbapenem-Resistant Enterobacterales Isolates.

The emergence of carbapenem-resistant Enterobacterales (CRE) has been recognized as a significant concern globally. Ceftazidime/avibactam (CZA) is a novel ß-lactam/ß-lactamase inhibitor that has demonstrated activity against isolates producing class A, C, and D ß-lactamases. Here-in, we evaluated the in vitro activity of CZA and comparator antimicrobial agents against 858 CRE isolates, arising from the Southeast Asian region, collected from a large tertiary hospital in Singapore. These CRE isolates mainly comprised Klebsiella pneumoniae (50.5%), Escherichia coli (29.4%), and Enterobacter cloacae complex (17.1%). Susceptibility rates to levofloxacin, imipenem, meropenem, doripenem, aztreonam, piperacillin/tazobactam, cefepime, tigecycline, and polymyxin B were low. CZA was the most active ß-lactam agent against 68.9% of the studied isolates, while amikacin was the most active agent among all comparator antibiotics (80% susceptibility). More than half of the studied isolates (51.4%) identified were Klebsiella pneumoniae carbapenemase (KPC)-2 producers, 25.9% were New Delhi metallo-ß-lactamase (NDM) producers, and Oxacillinase (OXA)-48-like producers made up 10.7%. CZA was the most active ß-lactam agent against KPC-2, OXA-48-like, and Imipenemase (IMI) producers (99.3% susceptible; MIC50/90: ≤1/2 mg/L). CZA had excellent activity against the non-carbapenemase-producing CRE (91.4% susceptible; MIC50/90: ≤1/8 mg/L). Expectedly, CZA had no activity against the metallo-ß-lactamases (MBL)-producing CRE (NDM- and Imipenemase MBL (IMP) producers; 27.2% isolates), and the carbapenemase co-producing CRE (NDM + KPC, NDM + OXA-48-like, NDM + IMP; 3.0% isolates). CZA is a promising addition to our limited armamentarium against CRE infections, given the reasonably high susceptibility rates against these CRE isolates. Careful stewardship and rational dosing regimens are required to preserve CZA's utility against CRE infections.

LipidA-IDER to Explore the Global Lipid A Repertoire of Drug-Resistant Gram-Negative Bacteria.

With the global emergence of drug-resistant bacteria causing difficult-to-treat infections, there is an urgent need for a tool to facilitate studies on key virulence and antimicrobial resistant factors. Mass spectrometry (MS) has contributed substantially to the elucidation of the structure-function relationships of lipid A, the endotoxic component of lipopolysaccharide which also serves as an important protective barrier against antimicrobials. Here, we present LipidA-IDER, an automated structure annotation tool for system-level scale identification of lipid A from high-resolution tandem mass spectrometry (MS2) data. LipidA-IDER was validated against previously reported structures of lipid A in the reference bacteria, Escherichia coli and Pseudomonas aeruginosa. Using MS2 data of variable quality, we demonstrated LipidA-IDER annotated lipid A with a performance of 71.2% specificity and 70.9% sensitivity, offering greater accuracy than existing lipidomics software. The organism-independent workflow was further applied to a panel of six bacterial species: E. coli and Gram-negative members of ESKAPE pathogens. A comprehensive atlas comprising 188 distinct lipid A species, including remodeling intermediates, was generated and can be integrated with software including MS-DIAL and Metabokit for identification and semiquantitation. Systematic comparison of a pair of polymyxin-sensitive and polymyxin-resistant Acinetobacter baumannii isolated from a human patient unraveled multiple key lipid A structural features of polymyxin resistance within a single analysis. Probing the lipid A landscape of bacteria using LipidA-IDER thus holds immense potential for advancing our understanding of the vast diversity and structural complexity of a key lipid virulence and antimicrobial-resistant factor. LipidA-IDER is freely available at https://github.com/Systems-Biology-Of-Lipid-Metabolism-Lab/LipidA-IDER.

Predictors and Outcomes of Healthcare-Associated Infections Caused by Carbapenem-Nonsusceptible Enterobacterales: A Parallel Matched Case-Control Study.

Objectives: The increasing incidence of carbapenem-nonsusceptible Enterobacterales as major pathogens in healthcare associated infections (HAIs) is of paramount concern. To implement effective prevention strategies against carbapenem-nonsusceptible Enterobacterales (CnSE) HAIs, it is crucial to identify modifiable factors associated with these infections. We identified risk factors for CnSE-HAIs, and compared clinical outcomes of CnSE-HAI and carbapenem-sensitive Enterobacterales (CSE)-HAI patients. Methods: We conducted a multi-centre parallel matched case-control study in two 1700-bedded Singapore acute-care hospitals from 2014-2016. Patients with CnSE-HAIs and CSE-HAIs were compared to a common control group without HAIs (1:1:3 ratio), matched by time-at-risk and patient ward. Carbapenem nonsusceptible was defined as non-susceptibility to either meropenem or imipenem. Presence of healthcare associated infections were defined by the criteria provided by the European Centre for Disease Prevention and Control. Outcomes of CnSE-HAI and CSE-HAI patients were compared using multivariable logistic and cox regression; the models were adjusted for infection and treatment characteristics. Results: Eighty CnSE-HAI and 80 CSE-HAI patients were matched to 240 patients without HAIs. All CRE-HAIs patients had prior antibiotic exposure, with 44 (55.0%) with prior carbapenem exposure. The most common CnSE-HAIs were intra-abdominal infections (28.8%) and pneumonia (23.8%). The most common CnSE species was Klebsiella spp. (63.8%). In the risk factor analysis, presence of drainage devices [adjusted odds ratio (aOR), 2.19; 95% CI, 1.29 - 3.70] and prior carbapenem exposure (aOR,17.09; 95% CI, 3.06 - 95.43) independently predicted CnSE-HAIs. In the crude outcomes analysis, CnSE-HAI patients had higher all-cause in-hospital mortality and longer time to discharge compared to CSE-HAI patients. After adjusting for differences in receipt of antibiotics with reported susceptibility to the Enterobacterales, there was no significant difference in all-cause in-hospital mortality between the two groups (aOR, 1.76; 95% CI, 0.86-3.58). Time to discharge remained significantly longer in patients with CnSE-HAI (adjusted hazard ratio, 0.71; 95% CI, 0.51 - 0.98) after adjusting for disease severity, receipt of antibiotics with reported susceptibility and receipt of appropriate source control. Conclusion: Appropriate management of deep-seated Enterobacterales infections and reducing exposure to carbapenems may reduce risk of CnSE-HAIs in Singapore. Efforts to improve antimicrobial therapy in CnSE-HAI patients may improve patient outcomes.

Therapeutic drug monitoring of meropenem and piperacillin-tazobactam in the Singapore critically ill population - A prospective, multi-center, observational study (BLAST 1).

PURPOSE: To determine percentage of patients with sub-therapeutic beta-lactam exposure in our intensive care units (ICU) and to correlate target attainment with clinical outcomes. MATERIALS AND METHODS: Multi-centre, prospective, observational study was conducted in ICUs from three hospitals in Singapore from July 2016 to May 2018. Adult patients (≥21 years) receiving meropenem or piperacillin-tazobactam were included. Four blood samples were obtained during a dosing interval to measure and determine attainment of therapeutic targets: unbound beta-lactam concentration above (i) minimum inhibitory concentration (MIC) at 40% (meropenem) or 50% (piperacillin) of dosing interval (40-50%fT > MIC) and (ii) 5 × MIC at 100% of dosing interval (100%fT > 5 × MIC). Correlation to clinical outcomes was evaluated using Cox regression. RESULTS: Beta-lactam levels were highly variable among 61 patients, with trough meropenem and piperacillin levels at 21.5 ± 16.8 mg/L and 101.6 ± 81.1 mg/L respectively. Among 85 sets of blood samples, current dosing practices were able to achieve 94% success for 40-50%fT > MIC and 44% for 100%fT > 5 × MIC. Failure to achieve 40-50%fT > MIC within 48 h was significantly associated with all-cause mortality (HR: 9.0, 95% CI: 1.8-45.0), after adjustment for APACHE II score. Achievement of 100%fT > 5 × MIC within 48 h was significantly associated with shorter length of hospital stay. CONCLUSION: Current dosing practices may be suboptimal for ICU patients. Beta-lactam TDM may be useful.

Quantification of Fosfomycin in Combination with Nine Antibiotics in Human Plasma and Cation-Adjusted Mueller-Hinton II Broth via LCMS.

Fosfomycin-based combination therapy has emerged as an attractive option in our armamentarium due to its synergistic activity against carbapenem-resistant Gram-negative bacteria (CRGNB). The ability to simultaneously measure fosfomycin and other antibiotic drug levels will support in vitro and clinical investigations to develop rational antibiotic combination dosing regimens against CRGNB infections. We developed an analytical assay to measure fosfomycin with nine important antibiotics in human plasma and cation-adjusted Mueller-Hinton II broth (CAMHB). We employed a liquid-chromatography tandem mass spectrometry method and validated the method based on accuracy, precision, matrix effect, limit-of-detection, limit-of-quantification, specificity, carryover, and short-term and long-term stability on U.S. Food & Drug Administration (FDA) guidelines. Assay feasibility was assessed in a pilot clinical study in four patients on antibiotic combination therapy. Simultaneous quantification of fosfomycin, levofloxacin, meropenem, doripenem, aztreonam, piperacillin/tazobactam, ceftolozane/tazobactam, ceftazidime/avibactam, cefepime, and tigecycline in plasma and CAMHB were achieved within 4.5 min. Precision, accuracy, specificity, and carryover were within FDA guidelines. Fosfomycin combined with any of the nine antibiotics were stable in plasma and CAMHB up to 4 weeks at -80 °C. The assay identified and quantified the respective antibiotics administered in the four subjects. Our assay can be a valuable tool for in vitro and clinical applications.

In vitro Bactericidal Activities of Combination Antibiotic Therapies Against Carbapenem-Resistant Klebsiella pneumoniae With Different Carbapenemases and Sequence Types.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is becoming increasingly problematic due to the limited effectiveness of new antimicrobials or other factors such as treatment cost. Thus, combination therapy remains a suitable treatment option. We aimed to evaluate the in vitro bactericidal activity of various antibiotic combinations against CRKP with different carbapenemase genotypes and sequence types (STs). Thirty-seven CRKP with various STs and carbapenemases were exposed to 11 antibiotic combinations (polymyxin B or tigecycline in combination with ß-lactams including aztreonam, cefepime, piperacillin/tazobactam, doripenem, meropenem, and polymyxin B with tigecycline) in static time-kill studies (TKS) using clinically achievable concentrations. Out of the 407 isolate-combination pairs, only 146 (35.8%) were bactericidal (≥3 log10CFU/mL decrease from initial inoculum). Polymyxin B in combination with doripenem, meropenem, or cefepime was the most active, each demonstrating bactericidal activity in 27, 24, and 24 out of 37 isolates, respectively. Tigecycline in combination with ß-lactams was rarely bactericidal. Aside from the lower frequency of bactericidal activity in the dual-carbapenemase producers, there was no apparent difference in combination activity among the strains with other carbapenemase types. In addition, bactericidal combinations were varied even in strains with similar STs, carbapenemases, and other genomic characteristics. Our findings demonstrate that the bactericidal activity of antibiotic combinations is highly strain-specific likely owing to the complex interplay of carbapenem-resistance mechanisms, i.e., carbapenemase genotype alone cannot predict in vitro bactericidal activity. The availability of WGS information can help rationalize the activity of certain combinations. Further studies should explore the use of genomic markers with phenotypic information to predict combination activity.

Pharmacokinetics of levetiracetam in neurosurgical ICU patients.

BACKGROUND/OBJECTIVES: The pharmacokinetics (PK) of drugs is dramatically altered in critical illness. Augmented renal clearance (ARC), a phenomenon characterized by creatinine clearance (CrCl) greater than 130 ml/min/1.73m2, is commonly described in critically ill patients. Levetiracetam, an antiepileptic drug commonly prescribed for seizure prophylaxis in the neurosurgical ICU, undergoes predominant elimination via the kidneys. Hence, we hypothesize that current dosing practice of intravenous (IV) levetiracetam 500 mg twice daily is inadequate for critically ill patients due to enhanced drug elimination. The objectives of our study were to describe the population PK of levetiractam using a nonparametric approach to design an optimal dosing regimen for critically ill neurosurgical patients. METHODS: This was a prospective, observational, population PK study. Serial blood samples were obtained from neurosurgical ICU patients who received at least one dose of IV levetiracetam. We used uHPLC to analyze these samples and Pmetrics™ software to perform PK analysis. RESULTS: Twenty subjects were included, with a median age of 54 years and CrCl of 104 ml/min. A two-compartmental model with linear elimination adequately described the profile of levetiracetam. Mean clearance (CL) was 3.55 L/h and volume of distribution (V) was 18.8 L. No covariates were included in the final model. Monte Carlo simulations showed a low probability of target attainment (PTA, trough at steady state of ≥6 mg/L) with a standard dose of 500 mg twice daily. A dose of at least 1000 mg twice daily was required to achieve 80% PTA. Two subjects, both with subtherapeutic trough levels, developed early onset seizures. CONCLUSION: Our study examined the population PK of levetiracetam in a critically ill neurosurgical population. We found that this population displayed higher clearance and required higher doses to achieve target levels.

Performance of Population Pharmacokinetic Models in Predicting Polymyxin B Exposures.

Polymyxin B is the last line of defense in treating multidrug-resistant gram-negative bacterial infections. Dosing of polymyxin B is currently based on total body weight, and a substantial intersubject variability has been reported. We evaluated the performance of different population pharmacokinetic models to predict polymyxin B exposures observed in individual patients. In a prospective observational study, standard dosing (mean 2.5 mg/kg daily) was administered in 13 adult patients. Serial blood samples were obtained at steady state, and plasma polymyxin B concentrations were determined by a validated liquid chromatography tandem mass spectrometry (LC-MS/MS) method. The best-fit estimates of clearance and daily doses were used to derive the observed area under the curve (AUC) in concentration-time profiles. For comparison, 5 different population pharmacokinetic models of polymyxin B were conditioned using patient-specific dosing and demographic (if applicable) variables to predict polymyxin B AUC of the same patient. The predictive performance of the models was assessed by the coefficient of correlation, bias, and precision. The correlations between observed and predicted AUC in all 5 models examined were poor (r2 < 0.2). Nonetheless, the models were reasonable in capturing AUC variability in the patient population. Therapeutic drug monitoring currently remains the only viable approach to individualized dosing.

Elimination of Extracellular Adenosine Triphosphate for the Rapid Prediction of Quantitative Plate Counts in 24 h Time-Kill Studies against Carbapenem-Resistant Gram-Negative Bacteria.

Traditional in vitro time-kill studies (TKSs) require viable plating, which is tedious and time-consuming. We used ATP bioluminescence, with the removal of extracellular ATP (EC-ATP), as a surrogate for viable plating in TKSs against carbapenem-resistant Gram-negative bacteria (CR-GNB). Twenty-four-hour TKSs were conducted using eight clinical CR-GNB (two Escherichia coli, two Klebsiella spp., two Acinetobacter baumannii, two Pseudomonas aeruginosa) with multiple single and two-antibiotic combinations. ATP bioluminescence and viable counts were determined at each timepoint (0, 2, 4, 8, 24 h), with and without apyrase treatment. Correlation between ATP bioluminescence and viable counts was determined for apyrase-treated and non-apyrase-treated samples. Receiver operator characteristic curves were plotted to determine the optimal luminescence threshold to discriminate between inhibitory/non-inhibitory and bactericidal/non-bactericidal combinations, compared to viable counts. After treatment of bacteria with 2 U/mL apyrase for 15 min at 37 °C, correlation to viable counts was significantly higher compared to untreated samples (p < 0.01). Predictive accuracies of ATP bioluminescence were also significantly higher for apyrase-treated samples in distinguishing inhibitory (p < 0.01) and bactericidal (p = 0.03) combinations against CR-GNB compared to untreated samples, when all species were collectively analyzed. We found that ATP bioluminescence can potentially replace viable plating in TKS. Our assay also has applications in in vitro and in vivo infection models.

Clinical Experience with High-Dose Polymyxin B against Carbapenem-Resistant Gram-Negative Bacterial Infections-A Cohort Study.

Population pharmacokinetic studies have suggested that high polymyxin B (PMB) doses (≥30,000 IU/kg/day) can improve bacterial kill in carbapenem-resistant Gram-negative bacteria (CR-GNB). We aim to describe the efficacy and nephrotoxicity of patients with CR-GNB infections prescribed high-dose PMB. A single-centre cohort study was conducted from 2013 to 2016 on septic patients with CR-GNB infection and prescribed high-dose PMB (~30,000 IU/kg/day) for ≥72 h. Study outcomes included 30-day mortality and acute kidney injury (AKI) development. Factors associated with AKI were identified using multivariable regression. Forty-three patients with 58 CR-GNB received high-dose PMB; 57/58 (98.3%) CR-GNB were susceptible to PMB. The median daily dose and duration of high-dose PMB were 32,051 IU/kg/day (IQR, 29,340-34,884 IU/kg/day) and 14 days (IQR, 7-28 days), respectively. Thirty-day mortality was observed in 7 (16.3%) patients. AKI was observed in 25 (58.1%) patients with a median onset of 8 days (IQR, 6-13 days). Higher daily PMB dose (aOR,1.01; 95% CI, 1.00-1.02) and higher number of concurrent nephrotoxins (aOR, 2.14; 95% CI; 1.03-4.45) were independently associated with AKI. We observed that a sizable proportion developed AKI in CR-GNB patients described high-dose PMB; hence, the potential benefits must be weighed against increased AKI risk. Concurrent nephrotoxins should be avoided to reduce nephrotoxicity.

In Vitro Pharmacodynamics of Fosfomycin against Carbapenem-Resistant Enterobacter cloacae and Klebsiella aerogenes.

The increase of carbapenem-resistant Enterobacterales (CRE) and lack of therapeutic options due to the scarcity of new antibiotics has sparked interest toward the use of intravenous fosfomycin against systemic CRE infections. We aimed to investigate the in vitro pharmacodynamics of fosfomycin against carbapenem-resistant Enterobacter cloacae and Klebsiella aerogenes Time-kill studies and population analysis profiles were performed with eight clinical CRE isolates, which were exposed to fosfomycin concentrations ranging from 0.25 to 2,048 mg/liter. The 24-h mean killing effect was characterized by an inhibitory sigmoid maximum effect (Emax) model. Whole-genome sequencing was performed to elucidate known fosfomycin resistance mechanisms. Fosfomycin MICs ranged from 0.5 to 64 mg/liter. The isolates harbored a variety of carbapenemase genes including blaIMP, blaKPC, and blaNDM Five out of eight isolates harbored the fosA gene, while none harbored the recently discovered fosL-like gene. Heteroresistant subpopulations were detected in all isolates, with two out of eight isolates harboring heteroresistant subpopulations at up to 2,048 mg/liter. In time-kill studies, fosfomycin exhibited bactericidal activity at 2 to 4 h at several fosfomycin concentrations (one isolate at ≥16 mg/liter, two at ≥32 mg/liter, two at ≥64 mg/liter, two at ≥128 mg/liter, and one at ≥512 mg/liter). At 24 h, bactericidal activity was only observed in two isolates (MICs, 0.5 and 4 mg/liter) at 2,048 mg/liter. From the Emax model, no significant bacterial killing was observed beyond 500 mg/liter. Our findings suggest that the use of fosfomycin monotherapy may be limited against CRE due to heteroresistance and rapid bacterial regrowth. Further optimization of intravenous fosfomycin dosing regimens is required to increase efficacy against such infections.

The utility of voriconazole therapeutic drug monitoring in a multi-racial cohort in Southeast Asia.

OBJECTIVES: Voriconazole serum concentration, which is affected by several factors, is associated with treatment response and toxicity. There is paucity of data on voriconazole therapeutic drug monitoring (TDM) among Southeast Asians, who exhibit a higher prevalence of CYP2C19-poor metabolisers compared with Caucasians and East Asians. Hence, there are concerns for higher risk of voriconazole accumulation and toxicity. We aim to determine the utility of voriconazole TDM through establishing: (1) proportion of patients achieving therapeutic troughs without dose adjustments; (2) characterisation of patients with sub-therapeutic, therapeutic and supra-therapeutic levels; (3) appropriate dose titrations/dose required for therapeutic troughs; (4) correlation between troughs and adverse events, treatment response/fungal breakthrough. PATIENTS AND METHODS: A single-centre retrospective analysis of data from adults (≥21 years old) with ≥1 voriconazole trough measured at Singapore General Hospital from 2015 to 2017 was performed. RESULTS: Thirty-two patients (45.7%) among 70 patients achieved therapeutic troughs (defined as 2.0-5.5 mg/L) without dose adjustments. Eleven patients (15.7%) experienced hepatotoxicity (troughs 0.5 to >7.5 mg/L). Neurotoxicity occurred in three patients (4.3%) (troughs ≥6.7 mg/L) and all patients had symptom resolution upon dose reduction. Treatment failure of invasive fungal infection appeared less in patients with therapeutic troughs compared with sub-therapeutic troughs (11.4% vs. 14.2%). Two patients experienced treatment failure despite supra-therapeutic voriconazole troughs. CONCLUSIONS: TDM should be implemented due to significant unpredictability in dose exposure. TDM can reduce unnecessary switches to alternatives due to intolerability and rule in the possibility of resistant organisms in the event of treatment failure despite therapeutic troughs, alerting clinicians to switch to alternatives promptly.

Determining the Development of Persisters in Extensively Drug-Resistant Acinetobacter baumannii upon Exposure to Polymyxin B-Based Antibiotic Combinations Using Flow Cytometry.

Polymyxin B-based combinations are increasingly prescribed as a last-line option against extensively drug-resistant (XDR) Acinetobacter baumannii It is unknown if such combinations can result in the development of nondividing persister cells in XDR A. baumannii We investigated persister development upon exposure of XDR A. baumannii to polymyxin B-based antibiotic combinations using flow cytometry. Time-kill studies (TKSs) were conducted in three nonclonal XDR A. baumannii strains with 5 log10 CFU/ml bacteria against polymyxin B alone and polymyxin B-based two-drug combinations over 24 h. At different time points, samples were obtained and enumerated by viable plating and flow cytometry. Propidium iodide and carboxyfluorescein succinimidyl ester dyes were used to differentiate between live and dead cells and between dividing and nondividing cells, respectively, at the single-cell level, and nondividing live cells were resuscitated and characterized phenotypically. Our results from viable plating showed that polymyxin B plus meropenem and polymyxin B plus rifampin were each bactericidal (>99.9% kill compared to the initial inoculum) against 2/3 XDR A. baumannii strains at 24 h. By flow cytometry, however, none of the combinations were bactericidal against XDR A. baumannii at 24 h. Further analysis using cellular dyes in flow cytometry revealed that upon exposure to polymyxin B-based combinations, XDR A. baumannii entered a viable but nondividing persister state. These bacterial cells reinitiated division upon the removal of antibiotic pressure and did not have a growth deficit compared to the parent strain. We conclude that persister cells develop in XDR A. baumannii upon exposure to polymyxin B-based combinations and that nonplating methods appear to complement viable-plating methods in describing the killing activity of polymyxin B-based combinations against XDR A. baumannii.

In vitro Pharmacodynamics and PK/PD in Animals.

In the last decade, considerable advancements have been made to identify the pharmacokinetic/pharmacodynamic (PK/PD) index that defines the antimicrobial activity of polymyxins. Dose-fractionation studies performed in hollow-fiber models found that altering the dosing schedule had little impact on the killing or suppression of resistance emergence, alluding to AUC/MIC as the pharmacodynamic index that best describes polymyxin's activity. For in vivo efficacy, the PK/PD index that was the most predictive of the antibacterial effect of colistin against P. aeruginosa and A. baumannii was ƒAUC/MIC.

Molecular mechanisms of azole resistance in Candida bloodstream isolates.

BACKGROUND: Antifungal resistance rates are increasing. We investigated the mechanisms of azole resistance of Candida spp. bloodstream isolates obtained from a surveillance study conducted between 2012 and 2015. METHODS: Twenty-six azole non-susceptible Candida spp. clinical isolates were investigated. Antifungal susceptibilities were determined using the Sensititre YeastOne® YO10 panel. The ERG11 gene was amplified and sequenced to identify amino acid polymorphisms, while real-time PCR was utilised to investigate the expression levels of ERG11, CDR1, CDR2 and MDR1. RESULTS: Azole cross-resistance was detected in all except two isolates. Amino acid substitutions (A114S, Y257H, E266D, and V488I) were observed in all four C. albicans tested. Of the 17 C. tropicalis isolates, eight (47%) had ERG11 substitutions, of which concurrent observation of Y132F and S154F was the most common. A novel substitution (I166S) was detected in two of the five C. glabrata isolates. Expression levels of the various genes differed between the species but CDR1 and CDR2 overexpression appeared to be more prominent in C. glabrata. CONCLUSIONS: There was interplay of various different mechanisms, including mechanisms which were not studied here, responsible for azole resistance in Candida spp in our study.