Lactate dehydrogenase activity in Bacteroides fragilis group strains with induced resistance to metronidazole.

The aims of this study were to induce in vitro metronidazole resistance in nim-negative Bacteroides fragilis group strains and to determine the lactate dehydrogenase (LDH) activity of the induced strains. A collection of B. fragilis group strains were identified by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF/MS). Minimum inhibitory concentrations (MICs) for metronidazole were determined by the agar dilution technique. The presence of nim genes was screened by PCR. A sample of 52 nim-negative metronidazole-susceptible strains were selected at random and were exposed to metronidazole in the resistance induction experiment. LDH activity was measured by spectrophotometry. Of the 52 selected strains, 12 (23.1%) acquired resistance to metronidazole. MICs ranged from 8mg/L to 96mg/L. Eight of the twelve induced strains displayed decreased LDH activity, whilst only one expressed a significant increase in LDH activity with LDH values of 49.1U/mg and 222.0U/mg, respectively. In conclusion, in vitro induction of metronidazole resistance could be selected in nim-negative B. fragilis group strains. A statistically significant decrease in LDH activity was in contrast to previous findings in which, underlying higher metronidazole MICs, an increase in LDH activity compensated for the decreased activity of pyruvate-ferredoxin oxidoreductase (PFOR). These findings could be explained if the induction caused only physiological and not genetic changes. We believe that genetic mutations in the B. fragilis strain that demonstrated an emergent increase in LDH activity were responsible for the increased activity.

Antimicrobial susceptibility and the in vitro postantibiotic effects of vancomycin and ciprofloxacin against Bacillus cereus isolates.

BACKGROUND: Vancomycin and ciprofloxacin were often used in the therapy of infections associated with Bacillus cereus. METHODS: Four B. cereus food and clinical isolates were chosen for determination of time-kill curves and postantibiotic effects (PAE) of ciprofloxacin and vancomycin. RESULTS: According to the minimum inhibition concentration (MIC), breakpoints defined by CLSI for Staphylococcus spp. were all four strains intermediate for vancomycin (MIC = 4 µg/ml) and sensitive to ciprofloxacin (MIC = 0.2 µg/ml) except the strain Bc63 resistant to the last antimicrobial (MIC = 1.6 µg/ml). The lowest CFU values of tested strains were reached after 3-5 hours of exposure to 4 × MIC of vancomycin, and after 6-7 hours exposure to 10 × MIC of ciprofloxacin. The maximum reduction of the CFU in the presence of vancomycin and ciprofloxacin was about 2.46 log10 and 2.48 log10, respectively. The average duration of the PAE of vancomycin and ciprofloxacin was 0.94 and 1.60 hours, respectively. The statistically significant differences between PAEs induced with 3 × MIC, 4 × MIC and 8 × MIC of vancomycin were observed (P < 0.05). Both antibiotics did not affect the sporulation of tested bacterial strains. CONCLUSIONS: The differences in PAE duration were strain and antimicrobial dependent.

KPC-Producing Klebsiella pneumoniae Isolates in Croatia: A Nationwide Survey.

BACKGROUND: In the last few years, Klebsiella pneumoniae strains producing K. pneumoniae carbapenemase (KPC) enzymes have emerged as important multidrug-resistant pathogens in hospitalized patients. This report describes KPC-producing isolates collected through the Croatian antimicrobial resistance surveillance program in the early stage of their dissemination in Croatia. MATERIALS AND METHODS: Forty-eight KPC-producing K. pneumoniae isolates, collected during a period from February 2011 to August 2013, were analyzed in this study. Antimicrobial susceptibility profiles were determined using disk diffusion and E-test. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were used for epidemiological analysis. Identification of ß-lactamase genes and associated antibiotic resistance mechanisms was performed by polymerase chain reaction and positive products were sequenced. Localization of blaKPC was investigated by S1 PFGE and Southern hybridization. RESULTS: Of 40 participating centers in Croatia, KPC isolates were recorded in 9 of them. They all had multidrug-resistant phenotype, but showed varying levels of resistance to carbapenems. All isolates displayed ST258, and PFGE showed that all but one were closely related. All isolates harbored blaKPC-2. Isolate with a unique PFGE pattern produced TEM-1, while others produced TEM-116. All isolates harbored blaSHV-11, but were negative for blaCTX-M and blaAmpC genes. All isolates contain one KPC-harboring plasmid, ranging in size from ∼60 to ∼210 kb, characterized as FIIs and IncR. CONCLUSION: This report describes that the early stage of KPC-producing K. pneumoniae dissemination in Croatia is associated with a prolific PFGE type belonging to ST258. So far, the spread of an outbreak strain is limited to the northwest region of the country.

Antimicrobial efficacy of photodynamic therapy, Nd:YAG laser and QMiX solution against Enterococcus faecalis biofilm.

BACKGROUND: Lasers have been recommended in final root canal disinfection protocol, however, there is no clear evidence about their efficacy against bacteria in biofilms. The aim of the study was to evaluate and compare the disinfection effect of antimicrobial photodynamic therapy (aPDT), Nd:YAG laser and QMiX solution against Enterococcus faecalis biofilm. METHODS: The study sample consisted of 65 dentine slices, which were inoculated with E. faecalis and incubated anaerobically for three weeks. The dentine discs were randomly allocated to one of the following experimental groups: aPDT (100 mW, 10 mg/ml phenothiazinium chloride, 1 min), Nd:YAG laser (2 W, 15 Hz, 4×5 s), QMiX solution (1 min). Positive controls did not receive any treatment and negative controls were treated with 5.25% NaOCl. To harvest surviving adherent cells, each dentine sample was transffered to a test tube containing of TSB, serial ten-fold dilutions were made and aliquot of 1 ml was plated onto blood agar plates and incubated for 48 h. Colony forming units grown were counted and transformed into actual counts based on the dilution factor. The remaining viable cells after each protocol were analysed by FISH. RESULTS: The aPDT and the QMiX solution were equally effective, with the reduction rate of E. faecalis CFUs of 98.8% and 99.3% respectively (p=1.107). The Nd:YAG laser caused 96% reduction of E. faecalis (P<0.001). CONCLUSION: The aPDT and the QMiX solution showed similar antibacterial efficacy against old E. faecalis biofilm, followed by Nd:YAG irradiation.

The antimicrobial effectiveness of photodynamic therapy used as an addition to the conventional endodontic re-treatment: a clinical study.

BACKGROUND: The purpose of the study was to evaluate the efficacy of antimicrobial photodynamic therapy (aPDT) used as an adjunct to the endodontic re-treatment in the eradication of microorganisms from previously filled root canals. METHODS: The study sample consisted of 21 randomly selected patients with root filled and infected root canal system with chronic apical periodontitis on incisors or canines, who have had previously endodontic treatment. Microbiological samples from the root canals were collected after accessing the canal, following the endodontic re-treatment and after the aPDT procedure. During instrumentation, the root canals were irrigated with 2.5% sodium hypochlorite (NaOCl), and the final irrigation protocol included 17% ethylenediaminetetraacetic acid followed by NaOCl. Root canals were filled with a phenothiazinium chloride and irradiated with a diode laser (λ=660 nm, 100 mW) for 1 min. Microbiological samples from the root canals were cultivated on selective plates, and the identification was done by micromorphology, macromorphology and different API strips as well as bacterial counts (colony forming units). RESULTS: Fourteen bacteria species were isolated from the root canals initially, with a mean value of 4.57 species per canal. Although endodontic re-treatment alone produced a significant reduction in the number of bacteria species (p<0.001), the combination of endodontic treatment and aPDT was statistically more effective (p<0.001). No bacteria were cultivated from the main root canals of 11 teeth. CONCLUSION: The results indicated that the aPDT used as an adjunct to the conventional endodontic therapy achieved a significant further reduction of intracanal microbial load.

Prevalence and molecular characteristics of methicillin-resistant Staphylococcus aureus strains isolated in a multicenter study of nursing home residents in Croatia.

BACKGROUND: Residents of nursing homes (NHs) are often hospitalized and could present a potential reservoir for methicillin-resistant Staphylococcus aureus (MRSA). The aim of the study was to determine the prevalence for MRSA carriage in residents and staff in Croatian NHs and to characterize MRSA strains using genotyping techniques. METHODS: A cross-sectional study was performed among 877 residents and staff of 7 NHs representing 3 major Croatian regions. Nasal swabs from residents and staff and other samples from residents with invasive devices were obtained. Identified isolates were submitted to susceptibility testing and genotyping with SCCmec typing, S aureus protein A (spa) locus typing, and pulsed-field gel electrophoresis (PFGE). RESULTS: The overall prevalence of MRSA colonization was 7.1% (95 confidence interval, 5.4%-8.8%), ranging from 0% to 28.8%. Four MRSA isolates were found in NH staff. All MRSA isolates were negative for Panton-Valentine leukocidin-encoding genes. SCCmec type II was found in 32 MRSA strains; SCCmec IV, in 27 strains; SCCmec I, in 3 strains. The predominant spa type was t008, found in 49 strains; PFGE analysis revealed 2 major clonal groups. CONCLUSIONS: MRSA strains were found to be colonizing residents and staff of 7 NHs in Croatia. Our study demonstrates the spread of 2 clones within and among Croatian NHs. The data presented here provide an important baseline for future surveillance of MRSA in NH.

Antimicrobial efficacy of Er,Cr:YSGG laser-activated irrigation compared with passive ultrasonic irrigation and RinsEndo(®) against intracanal Enterococcus faecalis.

OBJECTIVE: The aim of this study was to evaluate the antibacterial efficacy of active irrigation techniques: Erbium, chromium: yttrium, scandium, gallium, garnet (Er,Cr:YSGG) laser-activated irrigation (LAI), passive ultrasonic irrigation (PUI), RinsEndo(®), and conventional syringe irrigation, against intracanal Enterococcus faecalis. MATERIALS AND METHODS: Root canals of 100 human extracted teeth were instrumented, sterilized in plasma, contaminated with E. faecalis, and incubated for 10 days. The samples were randomly distributed into four experimental groups (n=20 each): I: LAI by Er,Cr:YSGG (1.25 W, 20 Hz) for 4×5 sec; II. PUI for 60 sec; III. RinsEndo(®) system for 60 sec; IV. 30 gauge syringe irrigation for 60 sec; and one positive control group (n=10). In all experimental groups, 2.5% sodium hypochlorite (NaOCl) was used as an irrigant, whereas the positive controls were rinsed with saline. The root canals were sampled by flushing with saline solution before and after the treatments, serially diluted, and cultured. The presence or absence of E. faecalis in the root canals was checked by polymerase chain reaction. RESULTS: There were no differences among the three active irrigation techniques (p>0.05), which were more effective than the conventional syringe irrigation (p<0.001). CONCLUSIONS: LAI, PUI, and RinsEndo were equally effective in the elimination of intracanal 10-day-old E. faecalis. However, LAI generated more negative bacterial samples, which warrants further investigations.

Unusually low prevalence of Mycoplasma genitalium in urine samples from infertile men and healthy controls: a prevalence study.

OBJECTIVE: To detect Mycoplasma genitalium in urine samples of infertile men and men without any signs of infection in order to investigate whether M. genitalium and other genital mycoplasmas (Mycoplasma hominis and Ureaplasma spp) are found more often in urine samples of infertile men than in asymptomatic controls and to determine resistance to macrolides. METHODS: The study included first void urine samples taken from 145 infertile men and 49 men with no symptoms of urethritis. M. genitalium, Chlamydia trachomatis and Neisseria gonorrhoeae were detected by commercial PCR. Trichomonas vaginalis was detected by microscopy and culture. M. hominis and Ureaplasma spp were detected by culture. M. genitalium was detected by in-house conventional and real-time PCR. RESULTS: Two M. genitalium positive samples were found among samples obtained from infertile men. All asymptomatic men were M. genitalium negative. Macrolide resistance was not found in either of the two positive samples. CONCLUSIONS: In comparison with reported data, an unusually low prevalence of M. genitalium was found in infertile men. The reasons for this unexpected result are not known; possibly, local demographic and social characteristics of the population influenced the result. Further studies to investigate M. genitalium in infertile and other groups of patients are needed.

[Croatian guidelines for diagnostics and treatments of Helicobacter pylori infection]. / Hrvatski postupnik za dijagnostiku i terapiju infekcije Helicobacterom pylori.

In the past 30-year period of investigations, the crucial role of Helicobacter pylori in chronic gastritis, gastric and duodenal ulcer development, and subsequently in gastric cancer and MALT lymphoma pathogenesis, has been recognized. During the first meeting of European Helicobacter Study Group in 1996 in Maastricht, the first recommendations for diagnostics and treatments of Helicobacter pylori infection were published, later reviewed in 2000, 2007 and 2010. The first meeting of Croatian doctors focusing on the same topics, but suitable to specific national circumstances, was held as early as 1998. The need for updating the old guidelines has emerged during the last years. The working expert group of gastroenterologists was formed and gathered on Consesus Conference in December 2012 in Zagreb, to arrive to current guidelines for the clinical management of Helicobacter pylori infection in Croatia. The following topics relating to Helicobacter pylori infection were examined: 1. indications and contraindications for diagnostics and treatments; 2. diagnostic methods and 3. treatments applicable in our country.

Antibiotic susceptibility of isolates from paediatric intensive care units in Zagreb.

AIM: Meropenem Yearly Susceptibility Test Information Collection (MYSTIC) Program is a longitudinal global surveillance study to monitor in vitro data on microbial susceptibility in centers that prescribe meropenem. Results of the six years period (2002-2007) for the antimicrobial efficacy of meropenem compared to other broad-spectrum agents against Gram-negative and Gram-positive isolates collected at pediatric intensive care units of the University Hospital Center Zagreb in Croatia were reported. METHODS: A total of 110 Gram-negative and 43 Gram-positive pathogens from pediatric specimens were tested. The minimum-inhibitory concentrations (MICs) were determined by broth microdilution method according to CLSI. RESULTS: There was no resistance to either imipenem or meropenem observed for Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis. High resistance rates of K. pneumoniae to ceftazidime and gentamicin (50%) are a raising concern. Pseudomonas aeruginosa was the most resistant Gram-negative species with two (12%) of the strains resistant to meropenem, three (18%) to imipenem, 10 (47%) to gentamicin and six (35%) to piperacillin/tazobactam and ciprofloxacin. According to our results meropenem remains an appropriate antibiotic for the treatment of severe infections caused by Gram-negative bacteria in pediatric population. CONCLUSIONS: The results indicate that meropenem has excellent potency and spectrum of activity despite being prescribed for a long time for the treatment of seriously ill patients, and still appears to be a reliable option for the initial empirical therapy of serious nosocomial infections in children. However, later studies have shown the emergence of carbapenem-resistant Gram-negative bacteria after 2008.

First Report of NDM-1-Producing Acinetobacter guillouiae.

BACKGROUND: Acinetobacter spp. is an opportunistic pathogen that has demonstrated increasing relevance in nosocomial infections. Carbapenem-resistant strains have been reported worldwide. METHODS: Since 2014, screening for metallo-ß-lactamases (MBLs) in all Acinetobacter spp. isolates using phenotypic methods and PCR has been implemented at the University Hospital Center Zagreb. RESULTS: The bacterial strain was isolated from the drain of a child hospitalized in a paediatric intensive care unit and identified as Acinetobacter guillouiae using a MALDI TOF automated system. The strain was resistant to meropenem, ceftazidime, cefotaxime, ceftriaxone, cefepime, sulbactam/ampicillin, gentamicin and ciprofloxacin, intermediately susceptible to piperacillin/tazobactam and imipenem, and susceptible to amikacin and colistin. The Hodge test and combined disk test with EDTA were positive. The MICs of meropenem and imipenem were not reduced by cloxacillin, but a small reduction of two dilutions was observed following the addition of sodium chloride, which indicated that OXA-58 was produced. PCR and sequencing of chromosomal DNA from boiled colonies revealed blaOXA-58 and blaNDM-1 genes. CONCLUSION: This is the first report of NDM-1 in Acinetobacter spp. in Croatia. The early detection of these genes will aid in the prevention and in the achievement of adequate infection control by limiting the spread of these organisms.

Estimating the burden of pneumococcal pneumonia among adults: a systematic review and meta-analysis of diagnostic techniques.

BACKGROUND: Pneumococcal pneumonia causes significant morbidity and mortality among adults. Given limitations of diagnostic tests for non-bacteremic pneumococcal pneumonia, most studies report the incidence of bacteremic or invasive pneumococcal disease (IPD), and thus, grossly underestimate the pneumococcal pneumonia burden. We aimed to develop a conceptual and quantitative strategy to estimate the non-bacteremic disease burden among adults with community-acquired pneumonia (CAP) using systematic study methods and the availability of a urine antigen assay. METHODS AND FINDINGS: We performed a systematic literature review of studies providing information on the relative yield of various diagnostic assays (BinaxNOW® S. pneumoniae urine antigen test (UAT) with blood and/or sputum culture) in diagnosing pneumococcal pneumonia. We estimated the proportion of pneumococcal pneumonia that is bacteremic, the proportion of CAP attributable to pneumococcus, and the additional contribution of the Binax UAT beyond conventional diagnostic techniques, using random effects meta-analytic methods and bootstrapping. We included 35 studies in the analysis, predominantly from developed countries. The estimated proportion of pneumococcal pneumonia that is bacteremic was 24.8% (95% CI: 21.3%, 28.9%). The estimated proportion of CAP attributable to pneumococcus was 27.3% (95% CI: 23.9%, 31.1%). The Binax UAT diagnosed an additional 11.4% (95% CI: 9.6, 13.6%) of CAP beyond conventional techniques. We were limited by the fact that not all patients underwent all diagnostic tests and by the sensitivity and specificity of the diagnostic tests themselves. We address these resulting biases and provide a range of plausible values in order to estimate the burden of pneumococcal pneumonia among adults. CONCLUSIONS: Estimating the adult burden of pneumococcal disease from bacteremic pneumococcal pneumonia data alone significantly underestimates the true burden of disease in adults. For every case of bacteremic pneumococcal pneumonia, we estimate that there are at least 3 additional cases of non-bacteremic pneumococcal pneumonia.

Surveillance of surgical site infection after cholecystectomy using the hospital in Europe link for infection control through surveillance protocol.

BACKGROUND: The third most common healthcare-associated infection is surgical site infection (SSI), accounting for 14%-16% of infections. These SSIs are associated with high morbidity, numerous deaths, and greater cost. METHODS: A prospective study was conducted to assess the incidence of SSI in a single university hospital in Croatia. We used the Hospital in Europe Link for Infection Control through Surveillance (HELICS) protocol for surveillance. The SSIs were classified using the standard definition of the National Nosocomial Infections Surveillance (NNIS) system. RESULTS: The overall incidence of SSI was 1.44%. The incidence of infection in the open cholecystectomy group was 6.06%, whereas in the laparoscopic group, it was only 0.60%. The incidence density of in-hospital SSIs per 1,000 post-operative days was 5.76. Patients who underwent a laparoscopic cholecystectomy were significantly younger (53.65±14.65 vs. 64.42±14.17 years; p<0.001), spent roughly one-third as many days in the hospital (2.40±1.72 vs. 8.13±4.78; p<0.001), and had significantly shorter operations by nearly 26 min (60.34±28.34 vs. 85.80±37.17 min; p<0.001). Procedures that started as laparoscopic cholecystectomies and were converted to open procedures (n=28) were reviewed separately. The incidence of SSI in this group was 17.9%. The majority of converted procedures (71.4%) were elective, and the operating time was significantly longer than in other two groups (109.64±85.36 min). CONCLUSION: The HELICS protocol has a good concept for the monitoring of SSI, but in the case of cholecystectomy, additional factors such as antibiotic appropriateness, gallbladder entry, empyema of the gallbladder, and obstructive jaundice must be considered.

Mycoplasma genitalium: clinical significance and diagnosis.

Mycoplasma genitalium is considered the smallest self-replicating cell. It was first isolated in 1981, from 2 of 13 men with urethritis. Mycoplasma genitalium causes urethritis, cervicitis and pelvic inflammatory disease. Because of difficulties in cultivation, the diagnosis is based exclusively on PCR methodology. The recommended therapy for Mycoplasma genitalium infections is azithromycin or doxycycline. Development of macrolide resistance was shown to correlate with treatment failure.

First report of KPC-producing Klebsiella pneumoniae in Croatia.

In February 2011, a 78-year-old male patient was admitted to Clinical Hospital Center Zagreb with subdural haematoma. Klebsiella pneumoniae with reduced susceptibility to carbapenems was isolated. PCR revealed the presence of bla(KPC), bla(TEM), and bla(SHV) genes. Sequencing of bla(KPC) gene identified K. pneumoniae carbapenemase (KPC)-2 beta-lactamase. The strain belonged to ST37 clone by multilocus sequence typing. Infection control efforts limited the spread of KPC-producing clone of K. pneumoniae in our hospital so far. To our knowledge, this is the first report of a KPC-producing K. pneumoniae in Croatia.

Value of rapid aetiological diagnosis in optimization of antimicrobial treatment in bacterial community acquired pneumonia.

In 80 adult patients with community acquired pneumonia (CAP) conventional microbiological methods, polymerase chain reaction (PCR) and serum C-reactive protein (CRP) levels were performed and the appropriateness of the empirical antimicrobial treatment was evaluated according to bacterial pathogen detected. The aetiology was determined in 42 (52.5%) patients, with Streptococcus pneumoniae as the most common pathogen. PCR applied to bronchoalveolar lavage (BAL) provided 2 and PCR on sputum samples 1 additional aetiological diagnosis of CAP The mean CRP values in the S. pneumoniae group were not significantly higher than in the group with other aetiological diagnoses (166.89 mg/L vs. 160.11 mg/L, p = 0.457). In 23.8% (10/42) of patients with determined aetiology, the empirical antimicrobial treatment was inappropriate. PCR tests need further investigation, particularly those for the atypical pathogens, as they are predominant in inappropriately treated patients. Our results do not support the use of CRP as a rapid test to guide the antimicrobial treatment in patients with CAP.

[Comparative urinary bactericidal activity of oral antibiotics against gram-positive pathogens]. / Usporedna urinarna baktericidna aktivnost oralnih antibiotika prema gram-pozitivnim urinarnim patogenima.

In routine bacteriological laboratories the antibacterial activity of antibiotics is determined by in vitro testing, usually by disk-diffusion test. However, in vitro testing does not always reflect antibacterial efficiency of antibiotics in vivo. In this investigation, the urine samples obtained in a single oral dose pharmacokinetic study were examined for their bactericidal activity against a range of relevant Gram-positive urinary tract pathogens. Urinary bactericidal activity of linezolid had been previously compared with ciprofloxacin but not with other oral antibiotics such as beta-lactams. Linezolid showed satisfactory urinary bactericidal titres throughout the whole testing period against all Gram-positive cocci. Fluoroquinolones displayed high and persisting levels of urinary bactericidal activity against staphylococci, but their activity against enterococci was weaker. According to the results of ex-vivo testing amoxycillin could be recommended only for infections caused by E. faecalis. Amoxycillin combined with clavulanic acid can be considered as a therapeutic option for infections caused by S. saprophyticus and E. faecalis. Older cephalosporins had high titres only against S. saprophyticus. Their drawback is a short elimination half-time in urine resulting in rapid decrease of urinary bactericidal titers during dosing interval. Furthermore, they do not show activity against enterococci due to their intrinsic resistance to cephalosporins.

Spread of CTX-M-15 positive Providencia spp. causing urinary tract infections at the University Hospital Split in Croatia.

AIM: During 2010-2011, six Providencia spp. (five Providencia stuartii and one Providencia rettgeri) urine isolates with unusual resistance phenotype were collected from various hospital units at the University Hospital Split in Croatia. The aim of the study was to analyze the mechanisms of resistance to expanded-spectrum cephalosporins. METHODS: The antimicrobial susceptibility to a wide range of antibiotics was determined by broth microdilution method according to CLSI guidelines. A double-disk-synergy test (DDST) was performed to detect ESBLs. The transferability of cefotaxime resistance was determined by conjugation. The presence of genes encoding ESBLs was determined by PCR while genotyping of the isolates was performed by PFGE. RESULTS: All strains were positive for ESBL production by DDST. They were uniformly resistant to amoxycillin alone and combined with clavulanate, cefazoline, cefuroxime, ceftazidime, cefotaxime, ceftriaxone, gentamicin and ciprofloxacin. P. stuartii strains transferred cefotaxime resistance to E. coli recipient strain with frequency ranging from 10-5 to 5x10-4. Five P. stuartii strains were positive for TEM and CTX-M ß-lactamases while P. rettgeri was positive only for TEM ß-lactamases. Five CTX-M producing isolates were shown to be clonally related. CONCLUSIONS: Continuous surveillance in tracking CTX-M-15- producing P. stuartii in the hospitals is necessary to prevent their spread to other hospitals and community. Global spread of ESBL positive Providencia spp all over the world is of great clinical concern.

Infrequent finding of metallo-ß-lactamase VIM-2 in carbapenem-resistant Pseudomonas aeruginosa strains from Croatia.

One hundred sixty-nine nonreplicate imipenem-resistant Pseudomonas aeruginosa strains isolated in a large hospital on the coastal region of Croatia were studied. The most active antibiotics were colistin and amikacin. Most of the isolates were multiresistant. The most prevalent serotype was O12, followed by O11. Six strains carried the bla(VIM-2) gene located in a novel class 1 integron composed in its variable part of the bla(VIM-2)-bla(oxa-10)-ΔqacF-aacA4 genes. Metallo-ß-lactamase-producing strains belonged to sequence types ST235 and ST111.