Molecular Characterization of Carbapenem-Resistant Acinetobacter baumannii Blood Culture Isolates from Three Hospitals in Turkey.

We aimed to investigate the clonal relationships, common sequence types, and carbapenemase genes in 177 non-repetitive blood culture isolates of Acinetobacter baumannii collected from patients at three university hospitals in Turkey in 2016. Molecular epidemiological characteristics of the isolates were examined using pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST) (Pasteur scheme-cpn60, fusA, gltA, pyrG, recA, rplB, and rpoB). Multiplex PCR was used to investigate the carbapenemase genes, including blaOXA-23-like, blaOXA-24-like, blaOXA-48-like, blaOXA-58-like, blaIMP, blaVIM, and blaNDM. PFGE genotyping yielded 92 pulsotypes with a clustering ratio of 69.7%. As per a ≥85% similarity coefficient, 159 (90.9%) isolates were found to be clonally related. The blaOXA-23-like and blaOXA-58-like genes were identified in 100% and 28.2% of the isolates, respectively. The blaNDM gene was identified in two isolates. The MLST analysis included 54 isolates with different pulsotypes, and 29 sequence types (STs). Most of the isolates (n = 36) belonged to the clonal complex (CC)2, one isolate belonged to CC1, and one isolate belonged to CC164. Sixteen new STs (ST1235-ST1250) were identified. Identifying both global ST2 and a large number of new STs, revealed high genetic diversity in A. baumannii isolates in the study population.

[The Identification of Acinetobacter baumannii Blood Isolates by MALDI-TOF MS, ARDRA and blaOXA-51-like Gene-Specific Real-Time PCR]. / Acinetobacter baumannii Kan Izolatlarinin MALDI-TOF MS, ARDRA ve blaOXA-51-benzeri Gene Özgül Gerçek Zamanli Polimeraz Zincir Reaksiyonu ile Tanimlanmasi.

The Acinetobacter calcoaceticus-Acinetobacter baumannii (Acb) complex consists of phenotypically very similar nosocomial species; A.baumannii, Acinetobacter nosocomialis, Acinetobacter pittii, Acinetobacter seifertii and Acinetobacter djikshoorniae and one environmental species A.calcoaceticus. The rapid and accurate identification of the members of Acb complex is critical as these nosocomial pathogens can show differences in antimicrobial susceptibility and clinical outcomes. The conventional phenotypic methods are slow, unreliable and less efficient for the differentiation of Acb complex species, including the A.baumannii species within the Acb complex. Although various molecular methods are available, such as amplified ribosomal DNA restriction analysis (ARDRA) and blaOXA-51-like gene specific PCR, they are usually inconvenient for the routine diagnostic laboratories. Recently, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) offers an opportunity for rapid, cost-effective and convenient bacterial identification in routine diagnostic procedures conducted in clinical laboratory. In this study, we aimed to evaluate the diagnosis performance of MALDI-TOF MS system to identify blood isolates of A.baumannii. A total of 180 nonduplicate carbapenem resistant Acb complex (strain numbers; TR1-TR60) and A.baumannii (TR61-TR180) blood isolates were collected from the intensive care units of the three university hospitals in Turkey from January 2016 to December 2016. All isolates were evaluated by using blaOXA-51-like gene specific real time (Rt-PCR) analysis, ARDRA (restriction enzymes-AluI, CfoI, MboI, MspI, RsaI) method and MALDI-TOF MS (VITEK® MS, bioMérieux, France) system. All the strains except TR10, TR31, TR35 and TR52 were identified as A.baumannii by ARDRA method. Out of 177 of all the isolates, presence of blaOXA-51-like gene was found except for TR10, TR31 and TR52 isolates. However, TR31 without the presence of blaOXA-51-like gene was identified as A.pittii using the ARDRA. Totally 176 isolates which were identified as A.baumannii by both of the methods, ARDRA and Rt-PCR- blaOXA-51-like, were accepted as a reference for the evaluation of the diagnosis performance capacity of the MALDI-TOF MS. Overall, for all 176 isolates tested, the sensitivity obtained with the MALDI-TOF MS were 99.4% with 75% specificity. The accuracy value of the method was determined as 98.9% for the identification of A.baumannii to the species level. MALDI-TOF MS is increasingly used in diagnostic microbiology for the routine identification of bacteria to the genus, species or subspecies level with high rates of sensitivity and specificity. In future, by expanding the database, MALDI-TOF MS system would possibly become the ideal method for routine diagnostic laboratories that could potentially identify more species and even determine some characteristics of antimicrobial resistance and virulence determinants.

Assessment of the effectiveness of a ventilator associated pneumonia prevention bundle that contains endotracheal tube with subglottic drainage and cuff pressure monitorization.

The effectiveness of prevention bundles on the occurrence and mortality of ventilator associated pneumonia (VAP) was evaluated in many studies. However, the effectiveness of endotracheal tube with subglottic secretion drainage (ETT-SD) and cuff pressure monitorization in VAP bundles have not been adequately assessed. In this study, we aimed to evaluate the effectiveness of VAP bundle containing ETT-SD and cuff pressure monitorization. This was a prospective, controlled study that was carried out between March 2011 and April 2012 including intubated patients. The study was conducted at the Anesthesiology Intensive Care Unit 1 and 2 (10 beds each) in a 898-bed university hospital. Occurrence of VAP and compliance with the parameters of the VAP prevention bundles were assessed daily. Patients intubated with the standard endotracheal tube were recruited as controls, mainly in the first six months of the study as ETT-SD and cuff pressure monometer had not yet been implemented. In the second term, patients intubated with ETT-SD were included as cases. Occurrence of VAP, mortality, and compliance with VAP prevention bundles were monitored. A total of 133 patients, 37 cases and 96 controls were recruited. VAP incidence declined from 40.82 to 22.16 per 1000 ventilator days among controls and cases, respectively (p<005). On average, VAP occurred 17.33±21.09 days in the case group and 10.43±7.83 days in the control group (p=0.04). However, mortality of cases and controls at the 14th and 30th days was not different. VAP prevention bundles including the utilization of ETT-SD, monitoring cuff pressure, and oral care with chlorhexidine were efficient in reducing the rate of VAP.

Is airborne transmission of Acinetobacter baumannii possible: A prospective molecular epidemiologic study in a tertiary care hospital.

BACKGROUND: Understanding the dynamics of aerial spread of Acinetobacter may provide useful information for production of effective control measurements. We investigated genetic relationships between air and clinical isolates of Acinetobacter baumannii in an intensive care unit (ICU) setting. METHODS: We conducted a prospective surveillance study in a tertiary care hospital for 8 months. A total of 186 air samples were taken from 2 ICUs. Clonal characteristics of air isolates were compared with the prospective clinical strains and the previously isolated strains of ICU patients over a 23-month period. RESULTS: Twenty-six (11.4%) air samples yielded A baumannii, of which 24 (92.3%) isolates were carbapenem-resistant. The Acinetobacter concentration was the highest in bedside sampling areas of infected patients (0.39 CFU/m3). Air isolates were clustered in 13 genotypes, and 7 genotypes (including 18 air strains) were clonally related to the clinical strains of 9 ICU patients. One clone continued to be cultured over 27 days in ICU air, and air isolates could be clonally related to 7-week retrospective and approximately 15-week prospective clinical strains. CONCLUSIONS: The results of this study suggest that infected patients could spread significant amounts of Acinetobacter to ICU air. These strains could survive in air for some weeks and could likely still infect new patients after some months. Special control measurements may be required against the airborne spread of Acinetobacter in ICUs.

[Investigation of beta-lactamase genes and clonal relationship among the extended-spectrum beta-lactamase producing nosocomial Escherichia coli isolates]. / Genislemis spektrumlu beta-laktamaz üreten nozokomiyal Escherichia coli izolatlarinda beta-laktamaz gen oranlari ve klonal iliskinin arastirilmasi

Extended-spectrum beta-lactamase (ESBL) producing microorganisms currently cause a major problem. Among theseCTX-M beta-lactamase producing Escherichia coli has also disseminated worldwide as an important cause of both nosocomial and community-acquired infections. The aims of this study were to determine the prevalence of the beta-lactamase genes, antibiotic susceptibilities and clonal relationships of ESBL-producing nosocomial E.coli isolates. A total of 76 ESBL-producing E.coli strains isolated from urine (n= 26), blood (n= 25) and wound (n= 25) specimens of hospitalized patients identified as nosocomial infection agents according to the CDC criteria between June 2010-June 2011 were included in the study. Antibiotic susceptibilities of the isolates were detected by Kirby-Bauer disc diffusion method according to CLSI recommendations. ESBL production was tested by double disc diffusion method, and cefotaxime/cefotaxime-clavulanic acid E-test strips (AB Biodisk, Sweden) were used for indeterminate results. Presence of TEM, SHV, CTX-M, OXA-2 group, 0XA-10 group, PER, VEB and GES beta-lactamase genes were investigated by polymerase chain reaction (PCR) using specific primers. Pulsed-field gel electrophoresis (PFGE) method was used for the detection of clonal relationships among the strains. Most of the ESBL-producing E.coli strains were isolated from samples of inpatients in intensive care (35%), internal medicine (16%) and general surgery (13%) units. All of the 76 strains were found susceptible to imipenem, meropenem and amikacin; however all were resistant to cefotaxime and ceftriaxone. The susceptibility rates of the isolates to cefoxitin, ertapenem, cefoperazone/sulbactam, piperacillin-tazobactam, gentamicin, ciprofloxacin, cefepime, amoxicillin-clavulanic acid, aztreonam and ceftazidime were 96%, 83%, 63%, 61%, 50%, 41%, 25%, 21%, 20% and 18%, respectively. Among E.coli isolates, the frequency of CTX-M, TEM, OXA-2 group, PER, SHV and OXA-10 group beta-lactamase genes were found as 89.5%, 59.2%, 15.8%, 14.5%, 11.8% and 3.9%, respectively, while none of the isolates were positive for VEB and GES beta-lactamase genes. In 1 (1.3%) strain none of the investigated genes were detected. PCR analyses of the isolates revealed that 25 harbored CTX-M and TEM genes together, while 20 harbored only CTX-M and two harbored only TEM genes. Single SHV gene was not detected in any of the isolates. PFGE demonstrated no major clonal relationship between ESBL-producing isolates. This study indicated that CTX-M type enzymes were highly endemic among ESBL-producing nosocomial E.coli strains in our hospital, with the polyclonal spread of ESBL-producing bacteria without any dominant epidemic clone. In conclusion, it was considered that further studies are needed to explain the relationship between epidemic clones and plasmids with the use of plasmid analysis and multilocus sequence typing methods.

Fungus ball in concha bullosa: an unusual cause of retro-orbital pain.

Fungus ball in the concha bullosa is an extremely rare disease. We described a case of the fungus ball in the concha bullosa in a 22-year-old woman. Preoperative diagnosis was based on nasal endoscopy and computed tomography scanning. The patient was endoscopically operated on. The examination of the removed material was reported as fungal infection. This case was found worth writing because of the location of the concha bullosa and its rare occurrence in this location.

Brucella and peritoneal dialysis related peritonitis: case report and review of literature.

Isolated case reports of peritonitis due to Brucella spp. during peritoneal dialysis (PD) continue to surface in the medical literature. However, the optimal treatment regimen for these patients, in particular with regards to the fate of PD catheter, is still largely unknown. We report a case of brucella peritonitis successfully treated with intraperitoneal administration of amikacin, along with oral rifampicin and doxycycline but without catheter removal. Furthermore, we have reviewed the literature up until present day.

An uncommon case of acute brucellosis presenting with severe thrombocytopenia.

A 49-year-old man was admitted to the hospital with complaints of fatigue, epistaxis and a skin rash. The whole blood count revealed isolated thrombocytopenia (4,000/mL), and the patient was admitted to the hematology department with a diagnosis of immune thrombocytopenia. He did not respond to steroid treatment for 15 days, and a subfebrile fever developed during this period. A diagnosis of acute brucellosis was considered due to positive serological tests and a blood culture positive for Brucella spp. After starting doxycycline and rifampicin therapy, the patient's thrombocyte count increased to 15,000/mL on the third day, to 41,000/mL on the sixth day and was normal on the 21st day of treatment. A diagnosis of brucellosis must be considered in patients presenting with severe and isolated thrombocytopenia in countries where brucellosis is endemic.

[Investigation of the susceptibilities of extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella spp. strains to ertapenem and other carbapenems]. / Genislemis Spektrumlu Beta-Laktamaz Üreten Escherichia coli ve Klebsiella spp. Suslarinin Ertapenem ve Diger Karbapenemlere Karsi Duyarliliklarinin Arastirilmasi

Infections caused by extended-spectrum beta-lactamase (ESBL) producing Escherichia coli and Klebsiella spp. constitute severe problems. Carbapenems are commonly used to treat these infections. However, infections caused by carbapenem-resistant gram-negative bacteria show an increasing trend recently. The aim of this study was to investigate the susceptibilities of ESBL-producing E.coli and Klebsiella spp. to ertapenem and other carbapenems. A total of 239 E.coli, 28 K.pneumoniae and 11 K.oxytoca strains isolated from clinical specimens (208 urine, 16 blood, 26 wound, 17 sterile body fluids, four tracheal aspirates and seven others) of hospitalized patients and outpatients between January 2007-February 2008, were included to the study. The isolates were identified by conventional methods, and antibiotic susceptibility tests were performed by Kirby Bauer disc diffusion method according to Clinical and Laboratory Standards Institute (CLSI) standards. ESBL production was tested by double disk diffusion method. When ESBL production was indeterminate, cefotaxime-clavulanic acid E test (BioMerieux, France) was used. According to the CLSI standards modified Hodge test was performed for carbapenem resistant isolates and minimal inhibitory concentration (MIC) values were detected for ertapenem (Etest®, BioMerieux, France), imipenem and meropenem (M.I.C. Evaluator Strips, Oxoid, UK). All of the isolates were found susceptible to amikacin (278/278; 100%), whereas the susceptibility rates for imipenem/meropenem and ertapenem were 99.3% (276/278) and 98.6% (274/278), respectively. When evaluated individually, ertapenem susceptibilities of E.coli, K.pneumoniae and K.oxytoca strains were 99.2%, 96.4% and 90.9%, respectively, while these rates were 100%, 96.4% and 90.9%, respectively, for imipenem/meropenem. Carbapenem resistance was detected in two E.coli, one K.oxytoca and one K.pneumoniae isolates. While two Klebsiella spp. Isolates were resistant to all of the tested carbapenems (MIC > 32 µg/ml), two E.coli isolates were resistant to ertapenem (MIC > 32 µg/ml) but susceptible to imipenem (MIC= 0.25 µg/ml) and meropenem (MIC= 0.5 µg/ml). Carbapenemase production was demonstrated by modified Hodge test in all of the carbapenem-resistant isolates. In conclusion, ESBL-producing gram-negative isolates should be routinely tested with a screening method for carbapenemase activity and confirmation tests should be performed in suspected cases.

MRSA genotypes in Turkey: persistence over 10 years of a single clone of ST239.

OBJECTIVES: Methicillin-resistant Staphylococcus aureus (MRSA) is a significant cause of life-threatening human infections. The clinical impact of MRSA is mounting, not only due to the ever-increasing prevalence but also due to the occurrence of new, community-acquired MRSA strains. The aim of this prospective, multi-centre study was to determine the prevalence and genetic relatedness of clinically relevant MRSA isolates, in Turkey. METHODS: During a 1-year period, data from 20 successive patients with invasive S. aureus infections were collected from eight university hospitals, geographically distributed over the six main regions of Turkey. Among these S. aureus isolates, the genetic association of MRSA isolates was investigated by pulsed-field gel electrophoresis (PFGE) and spa typing. A selected number of isolates were also analyzed by multilocus sequence typing (MLST). Furthermore, Panton Valentine leucocidin (PVL) genes were examined. RESULTS: In this study, the rate of methicillin resistance in S. aureus in patients with apparent infections (sepsis, meningitis, lung abscess or septic arthritis) ranged from 12 to 75% within the seven participating centres. Typing by pulsed-field gel electrophoresis and spa typing revealed the presence of 22 closely related genotypes. According to the PFGE and spa typing results, 53 out of 54 MRSA isolates were closely related. These isolates were of spa type t030 or a related spa type, contain an SCC mec type III element and belong to sequence type ST239. None of the isolates contained the PVL genes. CONCLUSIONS: Despite the broad surface area of Turkey, a single predominant clone of ST239 circulates in hospitals in different regions and only few new types of MRSA were introduced over the past years. These results place Turkey in the epicenter of ST239 prevalence.

Species distribution, antifungal susceptibility and clonal relatedness of Candida isolates from patients in neonatal and pediatric intensive care units at a medical center in Turkey.

The aim of this study was to assess species distribution, antifungal susceptibility and clonal relationships among Candida strains isolated from a group of pediatric/neonatal intensive care (PICU/NICU) patients that had a very high mortality rate (76%). The cases of 21 patients (19 with candidemia, 2 with Candida meningitides) treated over a 1-year period in a Turkish hospital PICU and NICU were retrospectively analyzed. Twenty-eight Candida isolates were detected from blood (20), cerebrospinal fluid (CSF) (2) and other specimens (6). Candida species were identified using the API ID 32C System. Susceptibility testing was done (all 28 isolates) for amphotericin B, fluconazole and itraconazole using the broth microdilution method. Arbitrarily primed polymerase chain reaction (AP-PCR) was used for molecular typing of the 3 most common ones; C. albicans (15), C. parapsilosis (6), and C. pelliculosa (4). Electrophoretic karyotyping (EK) was done to check clonal identity obtained by AP-PCR. Of the 20 blood isolates, 8 (40%) were C. albicans, 12 (60%) were non-albicans Candida, and one of the 2 CSF isolates was C. albicans. The overall species distribution was as follows: 15 C. albicans isolates, 6 C. parapsilosis isolates, 4 C. pelliculosa isolates, 2 C. famata isolates and 1 C tropicalis isolate. Amphotericin B had the best antifungal activity with a MIC90 of 0.125 microg/ml, and the rates of susceptibility to fluconazole and itraconazole were 93% and 82%, respectively. AP-PCR revealed 11 genotypes (4 were identical pairs, 7 were distinct) among the 15 C. albicans isolates, 2 genotypes (5 were classified in the same type) among the 6 C. parapsilosis isolates, and 4 separate genotypes for the 4 C. pelliculosa isolates. Karyotyping results correlated well with the AP-PCR findings. As indicated in the previous research, our results confirmed that non-albicans Candida species have become more frequently causative agents for invasive fungal infections in the ICU. Transmission of C. albicans and C. pelliculosa was relatively low, but transmission of C. parapsilosis was high, suggesting that more effective control and very strict treatment protocols are needed for patients having high mortality and invasive fungal infection in ICU.

Comparison of the antibacterial activity of lidocaine 1% versus alkalinized lidocaine in vitro.

BACKGROUND: Infections after epidural and spinal blocks are rare. The topical anesthetic liclocaine used in these procedures has been found to have antibacterial effects on various microorganisms. OBJECTIVE: The aim of this study was to assess the antibacterial effects of alkalinized liclocaine on Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. METHODS: Lidocaine 2%, alkalinized lidocaine, and physiologic saline (as a control solution) were added to standard bacterial preparations. The final concentration of the lidocaine was 10 mg/mL (1%). At baseline and 3 and 6 hours after incubation at 37°C, 3-mL aliquots were vortexed and pipetted into sterile polystyrene spectrophotometer cuvettes. Baseline referred to the end of the period of preparation of the solution (≤20 minutes). Growth was measured as the optical density at a wavelength of 540 nm. RESULTS: Compared with the control, lidocaine significantly inhibited the growth of S aureus, E coli, and P aeruginosa at baseline and 3 and 6 hours after incubation (all, P < 0.05). Alkalinized lidocaine significantly inhibited the growth of S aureus at baseline and 3 and 6 hours (all, P < 0.05), while it significantly inhibited the growth of E coli and P aeruginosa only at 6 hours (both, P < 0.05). The growth of E coli was significantly less in lidocaine than in alkalinized lidocaine at 0 and 3 hours (both, P < 0.05). CONCLUSION: The antibacterial effect of lidocaine 1% on S aureus was not changed after alkalinization. The effect of alkalinized lidocaine on E coli and P aeruginosa was significant only at 6 hours. Lidocaine significantly inhibited the growth of these 3 microorganisms at all study periods.

Clinical, microbiologic, and epidemiologic characteristics of Pseudomonas aeruginosa infections in a University Hospital, Malatya, Turkey.

BACKGROUND: Pseudomonas aeruginosa strains are generally resistant to many antibiotics, and nosocomial infections because of this species are one of the major problems in many hospitals. Molecular typing provides very useful information about origin and transmission of the strains. The aims of the present study were to investigate clinical and microbiologic characteristics of the nosocomial infections caused by P aeruginosa strains in a medical center and to bring up the cross-transmission level of this opportunistic pathogen in a university hospital by analyzing the clonal relationship among the isolates. METHODS: A total of 105 P aeruginosa strains had been identified among the 80 inpatients in a 1-year period from August 2003 to August 2004. Demographic, clinical, and epidemiologic data of the patients were prospectively recorded. The standardized disk-diffusion method was used to determine resistance of the strains to imipenem, ceftazidime, aztreonam, amikacin, gentamicin, mezlocillin, cefepime, tobramycin, meropenem, ceftriaxone, and ciprofloxacin. Clonal relatedness of the strains was investigated by pulsed-field gel electrophoresis (PFGE). RESULTS: Of the 105 P aeruginosa strains identified, 45 (43%) were isolated from the patients hospitalized in intensive care units. Thirteen patients had repeated pseudomonas infection (total 38 infections/13 patients); 26 of these repeated infections in 9 patients showed the same localization. Half of the patients had at least 1 underlying disease such as burn (48%), chronic illness (32%), and malignancy (20%). Fifty-seven patients (71%) had urinary and/or other catheterization. Urinary tract infection (35%) was the most frequent infection encountered, followed by respiratory tract infection (34%) and sepsis (13%). Resistance to the antibiotics tested was in the 12% to 88% range; amikacin was the most effective and ceftriaxone was the least effective antibiotic. The PFGE typing method showed that 28 of the 80 patients' isolates were clonally related, including 23 indistinguishable or closely related strains (29%), and 5 possibly related strains (6%). Epidemiologic data of the 16 patients (20% of the patients) confirmed a clonal relationship among the strains. Of the 26 isolates of the 9 patients having repeated infection in the same location, 18 (69%) were in the clonally related groups, whereas 11 of the 12 strains isolated from repeated infections on different body sites were clonally different. CONCLUSION: Our results indicated that P aeruginosa infections in our hospital mainly affected the patients hospitalized in intensive care units and those having catheterization, burn, and/or chronic illness. Amikacin was the best antibiotic as far as bacterial resistance was considered. Although lack of major PFGE type confirmed no P aeruginosa outbreak, typing results showed that cross transmission and treatment failure are the 2 main problems, which should be consider together to prevent this bacterial infection in medical centers.

[Distribution of bacteria isolated from urine cultures in Malatya University Hospital laboratory (letter)]. / Malatya Universite hastanesi Laboratuvarinda Idrar kültürlerinden izole edilen bakterilerin dagillmi.

The aim of this letter was to report the distribution of bacterial species isolated from the urine samples of patients in Malatya, which is located in Estern Anatolia part of Turkey. A total of 3.774 urine cultures were performed in the period of April-June 2006, and 792 (21%) of them yielded bacterial growth. The isolates have been identified by conventional methods and confirmed by Phoenix 100 (Becton-Dickinson) system. As a result, 702 (89%) Gram negative and 90 (11%) Gram positive bacteria were isolated from the samples. The most frequently isolated bacteria were Escherichia coli (58%), followed by Klebsiella spp. (14%), Pseudomonas spp. (6.4%), Enterococcus spp. (5%), Staphylococcus spp. (3.8%) and Streptococcus spp. (1.7%). The species distribution was found as follows; K. pneumoniae ssp pneumoniae (95/110), P. aeruginosa (48/51), E.faecalis (27/40), E. cloacae (19/29), P.mirabilis (19/22), C.freundii (8/12), coagulase negative staphylococci (19/30) and S. aureus (11/30). The first three array were shared by E. coli, Klebsiella spp. and Pseudomonas spp. for the samples of both outpatients and inpatients, while Pseudomonas spp. and E. coli were the most frequently isolated bacteria from the urine samples of intensive care unit patients. Our data was found parallel to the results of other national and international studies.

The prevalence of fecal colonization of enterococci, the resistance of the isolates to ampicillin, vancomycin, and high-level aminoglycosides, and the clonal relationship among isolates.

The gastrointestinal tract carriage of enterococci was searched in 150 hospitalized patients and 100 outpatients, and clonal relatedness of the isolates and their resistance to ampicillin, vancomycin, and high-level streptomycin and gentamicin were investigated. A stool sample or rectal swab collected from each patient was inoculated into appropriate media within an hour. Enterococcus species were identified by using conventional biochemical tests, API-20 Strep assay, and BBL crystal kit. Antibiotic susceptibility tests were performed using Kirby-Bauer disk diffusion method. A polymerase chain reaction (PCR) was used to detect vanA and vanB genes. Pulsed-field gel electrophoresis (PFGE) and arbitrarily primed-polymerase chain reaction (AP-PCR) methods were used for molecular typing of the strains. Enterococci were isolated from 90 (60%) of the specimens collected from 150 inpatients. Of these 90 isolates, 37 (41%) had high-level gentamicin resistance, 36 (40%) had high-level streptomycin resistance, and 50 (55.6%) had ampicillin resistance. Fecal colonization was found in 30% of the outpatients. Resistances to ampicillin, high-level streptomycin, and gentamicin were 13%, 10%, and 3%, in these patients' isolates, respectively. No vancomycin-resistant enterococci were detected by both agar diffusion and PCR assays in our study. Both typing procedures were applied on 78 Enterococcus strains isolated from inpatients. AP-PCR typing showed that 30 (50.8%) of the 59 E. faecium and 5 (50%) of the 10 E. faecalis strains were clonally related. These values were found to be 12 (20.3%) and two (20%) by PFGE, respectively. The typing procedures did not find any clustered strains in the six E. durans and three E. avium isolates. Neither PFGE nor AP-PCR result was significantly different among the sensitive and resistant strains. Our results indicate that the high prevalence of colonization with ampicillin and highlevel aminoglycoside-resistant enterococci is an important problem in our medical center. The high clonal diversity among the isolates indicates limited spread of antibiotic-resistant strains between patients.

Nosocomial Stenotrophomonas maltophilia cross-infection: three cases in newborns.

BACKGROUND: Increased nosocomial Stenotrophomonas maltophilia infection rates in newborns, especially in recent years, are a significant cause for concern. These cases are the second case group in the literature to have been identified as nosocomial cross-infection with S. maltophilia in neonates. OBJECTIVE: To investigate the clinical, microbiological, and epidemiologic features of the outbreak caused by S. maltophilia in the neonatal intensive care unit within a period of 7 days. METHODS: Three cases with nosocomial S. maltophilia infection considered to be the result of cross-transmission were prospectively analyzed. Arbitrarily primed polymerase chain reaction (AP-PCR) performed with M13 primer and pulsed-field gel electrophoresis (PFGE) of genomic DNA after digestion with XbaI were used to determine clonal relationship among the isolates. Results S. maltophilia was isolated from the blood cultures of all 3 patients. Molecular typing confirmed that the 3 cases were epidemiologically linked. CONCLUSIONS: Opportunistic pathogens such as S. maltophilia can lead to major problems in neonates. Molecular typing is helpful to improve effective control programs for preventing the spread of the infection.

Comparison of the semisolid agar antifungal susceptibility test with the NCCLS M38-P broth microdilution test for screening of filamentous fungi.

Antifungal susceptibility testing of pathogenic molds is being developed. A simple screening semisolid agar antifungal susceptibility (SAAS) test accurately measures susceptibilities of yeasts. The performance of the SAAS screening test for filamentous fungi was assessed by comparing MICs of four antifungals (amphotericin B [AMB], AMB lipid complex [ABEL], itraconazole [ITZ], and posaconazole [POS]) for 54 clinical mold isolates with the results of the National Committee for Clinical Laboratory Standards (NCCLS) proposed broth microdilution method (M38-P). The SAAS test utilized inocula stabbed into tubes of 0.5% semisolid heart infusion agar. In both tests MICs were read after incubation at 35 degrees C for 48 h. The isolates tested were Aspergillus fumigatus, Aspergillus niger, Aspergillus flavus, other Aspergillus spp., Fusarium spp., Penicillium sp., Mucor sp., Scedosporium prolificans, Trichophyton sp., and an unidentified dematiaceous mold. Concordance of test results was determined as the percent agreement of MICs +/- 1 dilution. The overall agreement between the tests for each drug was as follows: AMB, 94%; ABEL, 83%; ITZ, 94%; POS, 94%. For the Aspergillus spp., all but one were susceptible to ITZ by SAAS test; all were susceptible to POS (MIC range, 0.25 to 4 micro g/ml). Three of six non-Aspergillus molds that were resistant to AMB and ABEL by SAAS (MIC >/= 2 micro g/ml) were also resistant by the NCCLS test. The SAAS test compared favorably to the NCCLS broth microdilution test for molds, and most of the clinical isolates tested were susceptible to all four drugs.

Analysis of three outbreaks due to Klebsiella species in a neonatal intensive care unit.

OBJECTIVE: To investigate the clinical, microbiological, and epidemiologic features of three outbreaks caused by Klesiella during 3 years. SETTING: Neonatal intensive care unit of a university hospital. PATIENTS: Thirty affected neonates. METHODS: Data were collected through chart reviews and conversations with physicians. Screening samples were obtained from the staff, the neonates, and the environment. Antibiogram typing and arbitrarily primed polymerase chain reaction-based fingerprinting were used to type the strains. RESULTS: The first outbreak had 13 K. pneumoniae strains isolated. The second outbreak had 10 K. oxytoca strains isolated. The third outbreak had 20 K. pneumoniae strains isolated. More than half of the patients had low birth weights, were premature, and underwent mechanical ventilation and intravenous catheterization. Approximately three-fourths of the patients died. The isolates tested were completely susceptible to meropenem, cefoxitin, and ciprofloxacin and were resistant to cephalothin. More than half of these strains were resistant to many beta-lactam antibiotics, amikacin, and trimethoprim/sulfamethoxazole. Typing procedures yielded 3 antibiotypes and 3 genotypes among the isolates of the first outbreak, 3 antibiotypes with 1 subtype and 2 genotypes with 1 subtype in the second outbreak, and 2 antibiotypes and 2 genotypes in the third outbreak. CONCLUSIONS: Klebsiella outbreaks mainly affected premature neonates with intravenous catheters, mechanical ventilation, or both. The high mortality rate (76.7%) was notable. Resistance to multiple antibiotics, but mainly to broad-spectrum beta-lactam antibiotics, was observed, particularly in K. pneumoniae isolates. Molecular typing indicated that the three outbreaks were not related to one other.

[Comparison of methods used to detect methicillin resistance in staphylococci]. / Stafilokoklarda metisilin direncinin saptanmasinda kullanilan yöntemlerin karsilastirilmasi.

In this study, the methicillin resistance of 112 Staphylococcus aureus and 93 coagulase negative Staphylococcus (CNS) strains, which were initially found methicillin resistant by routine disk diffusion method in our laboratory, have been searched by microdilution and oxacillin agar screen test and the results were compared with the results obtained by disk diffusion method. The presence of mecA gene was investigated by polymerase chain reaction in case of discordant results. All S. aureus strains (100%) and 69.9% (65/93) of CNS strains were found resistant to methicillin by three of the methods. Of CNS isolates, 28 strains which were found methicillin resistant by disk diffusion method, were found methicillin susceptible by oxacillin agar screen method, and 27 of these were detected as mecA positive. Our results indicated that, the three methods tested were reliable for the detection of methicillin resistance in S. aureus strains, but oxacillin agar screen revealed to be unsatisfactory for the detection of methicillin resistance in CNS.