In Vitro Antibiofilm Activity of Fosfomycin Alone and in Combination with Other Antibiotics against Multidrug-Resistant and Extensively Drug-Resistant Pseudomonas aeruginosa.

BACKGROUND: Due to its rapid resistance development and ability to form biofilms, treatment of Pseudomonas aeruginosa infections is becoming more complicated by the day. Drug combinations may help reduce both resistance and biofilm formation. METHODS: Using the microtiter plate assay, we investigated the in vitro inhibition of biofilm formation and the disruption of preformed biofilms in multidrug-resistant and extensively drug-resistant clinical isolates of P. aeruginosa in the presence of peak plasma levels of eight antipseudomonal antibiotics alone and in combination with fosfomycin: ceftazidime, piperacillin/tazobactam, cefepime, imipenem, gentamicin, amikacin, ciprofloxacin and colistin. RESULTS: Combination therapy was significantly superior to monotherapy in its inhibition of biofilm formation. The highest inhibition rates were observed for combinations with colistin, cefepime and ceftazidime. CONCLUSION: Our results support fosfomycin combination therapy as an enhanced prophylactic option. Moreover, combinations with ß-lactam antibiotics and colistin demonstrated a more potent inhibition effect on biofilm formation than protein synthesis inhibitors.

In vitro killing of multidrug/extensively drug-resistant Pseudomonas aeruginosa by fosfomycin alone or in combination with antipseudomonal antibiotics.

Pseudomonas aeruginosa is a leading cause of nosocomial infections. Given the constant rise in resistance, adequate therapy is increasingly demanding. Fosfomycin recently became an appealing treatment option of bacterial infections due to multidrug-resistant bacteria (MDR). So far, fosfomycin synergy with other antibiotics has been assessed in studies, but only a limited number focused on MDR P. aeruginosa and on the effect of these combinations on the duration of the postantibiotic effect (PAE). We investigated synergy of fosfomycin with an array of antipseudomonal antibiotics using gradient diffusion strip cross method and time-kill method, and their effect on the duration of PAE against 51 variously resistant P. aeruginosa isolates. The highest rate of synergy was observed for combination with ceftazidime (23.4%) and gentamicin (19.1%). The PAE of antibiotic combinations was superior to that of the drugs alone. Our findings indicate that fosfomycin combination therapy may be a valuable treatment alternative.

Self-poisoning as a cause of admission in a medical intensive care unit and a question of misuse of prescription medications.

BACKGROUND: Self-poisoning accounts for between 3 and 17% of intensive care unit (ICU) admissions. OBJECTIVES: To display the misused substances, intention, and outcome of patients admitted to the ICU due to self-poisoning. METHODS: A retrospective review of ICU admissions using an electronic patient database was performed. RESULTS: A total of 149 patients were admitted, accounting for 4.6% of ICU admissions. Self-poisoning with self-harm was more frequent than without self-harm intent (62.4% vs. 37.6%). 64.4% of ingested substances were misused prescribed drugs. Females used antipsychotics (X2 = 15, p< 0.05) and benzodiazepines (X2 =11.52, p< 0.05), and males ingested antipsychotics (X2 = 12, p< 0.05) with a self-harm intent. Antipsychotics were always used with a self-harm intent. Illicit drugs (X2 =9.14, p< 0.05) and ethanol (X2 =7.34, p< 0.05) were mostly used without self-harm intent. 102 patients (68.5%) continued treatment in a psychiatric clinic, more often women (X2 (1,N = 145)=7.94, p< 0.005). The readmission rate was low. Mortality was 2.6%. CONCLUSION: Self-poisoning is an infrequent cause of ICU admission. Prescribed psychotropic medications, including benzodiazepines, antipsychotics, and antidepressants were the most frequently misused substances. Psychiatric consultation before the ICU discharge and subsequent psychiatric treatment might lower the repetition and readmission rate.

Gram-negative bacteria as causative agents of ventilator-associated pneumonia and their respective resistance mechanisms.

Ventilator-associated pneumonia (VAP) is a serious and common complication in patients admitted to intensive care unit (ICU) and contributes to mortality. Multidrug Gram-negative bacteria such as Acinetobacter baumannii, Pseudomonas aeruginosa, and Klebsiella pneumoniae are frequently associated with VAP in ICU. A prospective study was set up in three ICUs of the University Hospital Center Zagreb and one ICU in General Hospital Pula from September 2017 to March 2018. Antibiotic susceptibility was determined by broth microdilution method. Production of extended-spectrum ß-lactamases (ESBLs) was determined by double-disk synergy test and carbapenemases by Hodge and carbapenem inactivation method (CIM). The genes encoding ESBLs, carbapenemases of class A, B and D and qnr genes were determined by PCR. In total 97 Gram-negative bacteria isolates were analyzed. P. aeruginosa demonstrated high resistance rates for imipenem and meropenem with 74% and 68% of resistant strains, respectively. Moderate resistance rates were observed for ceftazidime andpiperacillin/tazobactam, ciprofloxacin and gentamicin (44%). All except three A. baumannii isolates, were resistant to carbapenems and to all other antibiotics apart from colistin and amikacin. Eight A. baumannii isolates were positive for blaOXA-23 and 12 for blaOXA-24 genes. Four K. pneumoniae and two E. cloacae strains were ESBL positive and harboured group 1 of CTX-M ß-lactamases. Three P. mirabilis strains were positive for plasmid-mediated ampC ß-lactamase of CMY family. Two carbapenem-resistant K. pneumoniae harboured OXA-48 and one carbapenem-resistant E. cloacae VIM-1. A high proportion of multidrug-resistant P. aeruginosa, K. pneumoniae and extensively resistant A. baumannii was reported. Acquired resistance mechanisms, mainly production of carbapenemases and ESBLs were dominant in A. baumannii and K. pneumoniae, respectively. Resistance of P. aeruginosa isolates was more likely due to upregulation of efflux pumps or porin loss. A marked diversity of ß-lactamases was identified in Enterobacteriaceae.

FALSE POSITIVE PHENOTYPIC DETECTION OF METALLO-BETA-LACTAMASES IN ACINETOBACTER BAUMANNII.

Phenotypic detection of metallo-ß-lactamases (MBLs) in Acinetobacter (A.) baumannii is a serious challenge to clinical microbiologists. MBLs are inhibited by metal chelators such as ethylenediaminetetraacetic acid) (EDTA). Production of MBLs cannot be recognized based on resistance phenotype. Therefore, phenotypic tests using EDTA are recommended. The aim of this study was to investigate the sensitivity and specificity of inhibitor based tests (EDTA) for detection of MBL. A total of 172 A. baumannii strains (123 carbapenemase positive and 49 carbapenemase negative) were analyzed. Phenotypic detection of MBLs was performed by the combined disk test with EDTA (CDT-EDTA) and EPI-dilution test (EPI-DT). Both tests were positive in all 11 isolates possessing VIM-1 MBL, showing 100% sensitivity. However, false positive results were observed in strains with class D carbapenemases using both tests, i.e. all OXA-23 and OXA-24/40 producing organisms and most OXA-58 positive strains (77% with CDT-EDTA vs. 65% with EPI-DT). False positive results can occur because oxacillinases are converted to a less active state in the presence of EDTA, leading to augmentation of the inhibition zone around the carbapenem disk or reduction of carbapenem minimum inhibitory concentrations. This study showed high sensitivity but low specificity of phenotypic methods in the detection of MBLs.

Experience with fosfomycin in the treatment of complicated urinary tract infections caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae.

BACKGROUND: The aim of this study was to evaluate the efficacy of fosfomycin in the treatment of complicated urinary tract infections (cUTIs) caused by extended-spectrum beta-lactamase (ESBL) producing Enterobacteriaceae. METHODS: We retrospectively evaluated 42 ambulatory patients with cUTIs caused by ESBL-producing Enterobacteriaceae at the Outpatient Internal Medicine Clinic of the University Clinical Hospital Centre Zagreb in the period from June 2012 to June 2014. ESBL production was confirmed by double disk synergy test according to Jarlier. In vitro susceptibility to fosfomycin of ESBL-producing Escherichia coli, Klebsiella pneumoniae and Citrobacter freundii isolates was tested according to the European Committee on Antimicrobial Susceptibility Testing methodology. RESULTS: In 42 patients with cUTIs, 43 urinary pathogens susceptible to fosfomycin were isolated in the urine cultures, including 34 E. coli ESBL, seven K. pneumoniae ESBL and two C. freundii ESBL isolates. On average, patients had 2.2 complicating factors (CFs) and received 3.6 fosfomycin doses per treatment course. The overall microbiological cure was 50%, clinical cure was 71% and ESBL eradication rate was 74%. Patients with between zero and one CFs received significantly fewer fosfomycin doses than patients with two or more CFs (p = 0.022). Three kidney transplant patients achieved microbiological cure following prolonged fosfomycin administration. No statistically significant correlation was found between the presence of individual CFs and treatment outcome. CONCLUSIONS: Fosfomycin may be a valid option for oral treatment of cUTIs caused by ESBL-producing pathogens. The optimal duration of fosfomycin treatment for cUTIs remains to be determined.

Incidence, etiology, predictors and outcomes of suspected drug hypersensitivity reactions in a tertiary care university hospital's emergency department : A retrospective study.

BACKGROUND: Allergic drug reactions are adverse drug reactions that result from a specific immunologic response to a medication. Considering the epidemiological and clinical importance of drug allergy, this retrospective analysis focused on drug hypersensitivity in a tertiary care university hospital emergency department (ED). METHODS: In this study 74,929 ED records obtained from March 2012 to March 2015 were reviewed to determine the incidence, etiology, predictors and clinical features of drug hypersensitivity. RESULTS: The observed incidence of drug hypersensitivity was 0.87% of all ED admissions. It was significantly higher in female patients aged 18-29 years (2.26%; P < 0.0001) and during winter months (1.09%; P = 0.0058). Most patients had mild to moderate symptoms which regressed following ED treatment. Only five patients (7 per 100,000 ED visits) were diagnosed with drug-induced anaphylaxis, and only five patients were provisionally diagnosed with severe non-immediate reactions with systemic involvement. No patient died of drug hypersensitivity in the ED, and only a small proportion required subsequent hospitalization. The most common causes of drug hypersensitivity reactions were amoxicillin and paracetamol. CONCLUSION: Drug hypersensitivity is a common reason for tertiary centre emergency admissions. This is the largest analysis of ED drug hypersensitivity admissions so far. Beta-lactams were identified as the leading cause of drug hypersensitivity requiring ED evaluation, which also explains the peak of drug hypersensitivity cases during winter months when the use of these medications is highest.

NDM-1-producing Enterobacter aerogenes isolated from a patient with a JJ ureteric stent in situ.

Urinary tract infections after JJ stent insertion are among the most common complications, and the associated microorganisms carry more antibiotic resistance determinants than those found in urine prior to stent insertion. In line with the trends in healthcare epidemiology which implicate multi-resistant microorganisms in a plethora of healthcare-associated infections, prosthetic stent material also represents an ideal milieu for biofilm formation and subsequent infection development with resistant bacterial agents. Here we describe a case of a 73-year-old Caucasian woman presenting with urinary tract infection after JJ ureteric stent insertion due to ureteric obstruction and hydronephrosis of her left kidney. Extensive microbiological work-up and comprehensive molecular analysis identified the putative microorganism as carbapenem-resistant Enterobacter aerogenes carrying New Delhi metallo-beta-lactamase 1 (NDM-1). This is a first literature report implicating such extensively resistant strain of this species in early indwelling ureteric stent complications, and also the first report of NDM-1 in Enterobacter aerogenes in Croatia and Europe.

Activity of fosfomycin against nosocomial multiresistant bacterial pathogens from Croatia: a multicentric study.

AIM: To determine in vitro susceptibility of multiresistant bacterial isolates to fosfomycin. METHODS: In this prospective in vitro study (local non-random sample, level of evidence 3), 288 consecutively collected multiresistant bacterial isolates from seven medical centers in Croatia were tested from February 2014 until October 2016 for susceptibility to fosfomycin and other antibiotics according to Clinical and Laboratory Standards Institute methodology. Susceptibility to fosfomycin was determined by agar dilution method, while disc diffusion was performed for in vitro testing of other antibiotics. Polymerase chain reaction and sequencing were performed for the majority of extended spectrum ß-lactamase (ESBL)-producing Klebsiella pneumoniae (K. pneumoniae) and carbapenem-resistant isolates. RESULTS: The majority of 288 multiresistant bacterial isolates (82.6%) were susceptible to fosfomycin. The 236 multiresistant Gram-negative isolates showed excellent susceptibility to fosfomycin. Susceptibility rates were as follows: Escherichia coli ESBL 97%, K. pneumoniae ESBL 80%, Enterobacter species 85.7%, Citrobacter freundii 100%, Proteus mirabilis 93%, and Pseudomonas aeruginosa 60%. Of the 52 multiresistant Gram-positive isolates, methicillin-resistant Staphylococcus aureus showed excellent susceptibility to fosfomycin (94.4%) and vancomycin-resistant enterococcus showed low susceptibility to fosfomycin (31%). Polymerase chain reaction analysis of 36/50 ESBL-producing K. pneumoniae isolates showed that majority of isolates had CTX-M-15 beta lactamase (27/36) preceded by ISEcp insertion sequence. All carbapenem-resistant Enterobacter and Citrobacter isolates had blaVIM-1 metallo-beta-lactamase gene. CONCLUSION: With the best in vitro activity among the tested antibiotics, fosfomycin could be an effective treatment option for infections caused by multiresistant Gram-negative and Gram-positive bacterial strains in the hospital setting.

Online learning applied to a course on rational therapeutics: an international comparison between final year students of two medical schools.

INTRODUCTION: Poor prescribing is probably the most common cause of preventable medication errors and many of these events involve junior doctors. In 2009, an electronic problem-based therapeutics course developed at the University of Michigan Medical School (UMMS) was translated and adapted for use at the University of Zagreb Medical School (UZMS). METHODS: After students from both schools took the course in 2010, we compared their responses with an online questionnaire addressing the course quality and its effectiveness. RESULTS: There were no statistically significant differences in the overall average grades awarded for the course (UZMS 4.11 ± 0.86 vs. UMMS 3.96 ± 0.93; 95% CI mean difference (MD) -0.36, 0.07; P = 0.175) with both student groups expressing high satisfaction rates with its quality, accessibility and overall design. UZMS students reported spending less time working through the course than their American colleagues (2.14 ± 1.01 vs. 2.89 ± 1.02 on a five point Likert scale; 95% CI MD 0.51, 0.99; P < 0.05). Furthermore, Croatian students indicated greater difficulty with course materials (3.54 ± 0.59 vs. 3.25 ± 0.59; 95% CI MD -0.42, -0.15; P < 0,05) and weekly multiple choice questions (3.83 ± 0.62 vs. 3.4 ± 0.61; 95% CI MD -0.58, -0.29; P < 0,05) compared with the UMMS students. CONCLUSION: It is possible to adapt and translate successfully whole online teaching resources and implement them internationally in different countries and health care systems, achieving similar, high student satisfaction rates while decreasing administrative and cost burdens. Web based learning may have great potential to offer a cost effective and safe environment in which prescribing skills can be improved.

Distributed lags time series analysis versus linear correlation analysis (Pearson's r) in identifying the relationship between antipseudomonal antibiotic consumption and the susceptibility of Pseudomonas aeruginosa isolates in a single Intensive Care Unit of a tertiary hospital.

The relationship between antibiotic consumption and selection of resistant strains has been studied mainly by employing conventional statistical methods. A time delay in effect must be anticipated and this has rarely been taken into account in previous studies. Therefore, distributed lags time series analysis and simple linear correlation were compared in their ability to evaluate this relationship. Data on monthly antibiotic consumption for ciprofloxacin, piperacillin/tazobactam, carbapenems and cefepime as well as Pseudomonas aeruginosa susceptibility were retrospectively collected for the period April 2006 to July 2007. Using distributed lags analysis, a significant temporal relationship was identified between ciprofloxacin, meropenem and cefepime consumption and the resistance rates of P. aeruginosa isolates to these antibiotics. This effect was lagged for ciprofloxacin and cefepime [1 month (R=0.827, P=0.039) and 2 months (R=0.962, P=0.001), respectively] and was simultaneous for meropenem (lag 0, R=0.876, P=0.002). Furthermore, a significant concomitant effect of meropenem consumption on the appearance of multidrug-resistant P. aeruginosa strains (resistant to three or more representatives of classes of antibiotics) was identified (lag 0, R=0.992, P<0.001). This effect was not delayed and it was therefore identified both by distributed lags analysis and the Pearson's correlation coefficient. Correlation coefficient analysis was not able to identify relationships between antibiotic consumption and bacterial resistance when the effect was delayed. These results indicate that the use of diverse statistical methods can yield significantly different results, thus leading to the introduction of possibly inappropriate infection control measures.

Risk behaviors for getting HIV infection among the Croatian men who have sex with men.

This study was conducted with the aim of obtaining the very first information on the sexual risk behavior of Croatian men who have sex with men (MSM). There were 1127 respondents recruited at four venues: three physical meeting places of the Croatian MSM population (disco club, bar and sauna) in Zagreb and one virtual (gay oriented web-site) meeting place of the Croatian MSM population. The overall response rate was only 19%. The rate of condom use during last anal intercourse was 59% and 56% of those who used a lubricant had chosen an incorrect product to use with latex condoms. There was no connection between drug-use and unprotected anal intercourse (UAI). Only a fifth of those who had also had sex with women (MSM/MSW) during last 12 months regularly used protection. The obtained findings will serve for focused and effective prevention activities and a basis for comparison in future research.