Investigation of the presence of Capnophilic bacteria in routine urine cultures.

Capnophilic Escherichia coli (CEC) strains are rarely isolated from urinary tract infections (UTIs). The purpose of this research was to look into the incidence and traits of the CEC strains that cause UTIs. Nine (0.11%) epidemiologically unrelated CEC isolates with varying antibiotic susceptibility patterns were identified from patients with various co-morbidities after the evaluation of 8500 urine samples. Three of these strains belonged to the O25b-ST131 clone, and none of them possessed the yadF gene. Due to adverse incubation conditions, CEC isolation is difficult. Although rare, capnophilic incubation of urine cultures may be considered particularly for patients with underlying predisposing conditions.

[Investigation of Virulence Factors, Phylogenetic Group Features, and the Presence of ST131 Clone in Escherichia coli Isolates, a Urinary Tract Infection Agent in Children]. / Çocuklarda Üriner Sistem Enfeksiyonu Etkeni Olan Escherichia coli Izolatlarinda Virülans Faktörleri, Filogenetik Grup Özellikleri ve ST131 Klonu Varliginin Arastirilmasi.

Urinary tract infection (UTI) caused by Escherichia coli is a significant health issue in children. Today especially E.coli O25b/ST131, defined as a pandemic clone, is a serious public health problem due to its high virulence and antimicrobial resistance rates. In this study, a total of 200 (100 first and 100 recurrent UTI-causing) E.coli isolates from urine samples sent to the Ankara University School of Medicine Cebeci Training and Research Hospital Central Laboratory between January and September 2021 with the preliminary diagnosis of UTI in pediatric patients aged three to 18 years were analyzed for antimicrobial resistance rates, phylogenetic group distributions, virulence factor frequencies and whether they belong to the O25b/ST131 clone. It is aimed in this study that, the obtained data will shed light on new studies for diagnosis, treatment and prophylaxis options that can be developed for more effective UTI management by contributing to the surveillance studies in our country. Antimicrobial susceptibility of E.coli isolates identified by conventional methods was evaluated by Kirby-Bauer disc diffusion method and extended spectrum beta-lactamase (ESBL) production was evaluated by double disc synergy test. Polymerase chain reaction (PCR) was used for the investigation of phylogenetic grouping, the O25b/ST131 clone, virulence genes and the molecular level classification of the isolates detected as uropathogenic E.coli (UPEC). Pulsed-field gel electrophoresis (PFGE) was performed with the isolates collected at different times from the same patient. The highest antimicrobial resistance rates observed were against ampicillin (n= 100, 50%), cefazolin (n= 99, 49.5%), trimethoprim-sulfamethoxazole (n= 55, 27.5%), amoxicillin-clavulanic acid (n= 43, 21.5%) and cefotaxime (n= 43, 21.5%). In recurrent UTI agents, resistance rates were higher for cefotaxime (n= 29, 29%), trimethoprim-sulfamethoxazole (n= 35, 35%) and cefepime (n= 25, 25%) and in O25b/ST131 isolates (n= 67) the rates were higher for amikacin (n= 3, 4.5%), gentamicin (n= 10, 14.9%) and ciprofloxacin (n= 17, 25.4%) when compared to the first UTI agents and non-O25b/ ST131 isolates (p< 0.05). It was found that 29% (n = 58) of the isolates were multidrug resistant (MDR) and 19% (n = 38) produced ESBL.The rate of recurrent UTI agents was found to be higher among ESBL producing isolates and/or MDR isolates (n= 36, 62% and n= 27, 71%, respectively, p< 0.05). It was found that 45.5% (n= 91) of the isolates were in D, 37.5% (n= 75) in B2, 12.5% (n= 25) in A, and 4.5% (n= 9) in B1 phylogenetic groups and isolates belonging to B2 and D phylogenetic groups had higher antibiotic resistance rates and carried more virulence genes (p< 0.05). Of the isolates, 33.5% (n= 67) were found to belong to the O25b/ST131 clone, no significant difference was found between the O25b/ST131 rates among the first and recurrent UTI agents (p> 0.05). It was determined that the isolates most frequently carry virulence genes for adhesion [fimH 97% (n= 194), papA 57% (n= 114), yfcV 49.5% (n= 99)] and iron uptake systems [fyuA 85.5% (n= 171), chuA 78% (n= 156), iutA 73% (n= 146)]. All virulence factors were detected more frequently in isolates belonging to the O25b/ST131 clone (p< 0.05). Of the isolates, 97% (n= 65) belonging to the O25b/ST131 clone and 27.1% (n= 36) not belonging to this clone were defined as UPEC with molecular analysis (p< 0.0001). Thirty-three isolates belonging to 15 patients were evaluated with PFGE, and it was observed that the latter isolate and the first isolate of eight patients (53%) had the same band profile. Focusing on surveillance, diagnostic testing, treatment algorithms, and preventive measures for E.coli and especially for ST131 clone, which is frequently observed as causative agent in childhood UTIs, will help to manage challenging E.coli infections.

Characteristics and outcomes of carbapenemase harbouring carbapenem-resistant Klebsiella spp. bloodstream infections: a multicentre prospective cohort study in an OXA-48 endemic setting.

A prospective, multicentre observational cohort study of carbapenem-resistant Klebsiella spp. (CRK) bloodstream infections was conducted in Turkey from June 2018 to June 2019. One hundred eighty-seven patients were recruited. Single OXA-48-like carbapenemases predominated (75%), followed by OXA-48-like/NDM coproducers (16%). OXA-232 constituted 31% of all OXA-48-like carbapenemases and was mainly carried on ST2096. Thirty-day mortality was 44% overall and 51% for ST2096. In the multivariate cox regression analysis, SOFA score and immunosuppression were significant predictors of 30-day mortality and ST2096 had a non-significant effect. All OXA-48-like producers remained susceptible to ceftazidime-avibactam.

[Investigation of Biofilm Formation Properties of Coagulase Negative Staphylococci Isolated from Catheter-Related Bloodstream Infections]. / Kateter Iliskili Kan Dolasimi Enfeksiyonlarindan Izole Edilen Koagülaz Negatif Stafilokoklarin Biyofilm Olusturma Özelliklerinin Arastirilmasi.

In view of the significant negative impact of biofilm-mediated infection on patient health and the necessity of a reliable phenotypic method to detect biofilm producers, this study aimed to demonstrate phenotypic and molecular biofilm formation in coagulase-negative staphylococci (CoNS) isolated from catheter related infections and to compare the methods used with each other. The study was also aimed to determine the biofilm eradication effect of vancomycin in order to guide for the treatment. For the detection of biofilm formation, a total of 154 CoNS clinical isolates of which 30 being causative agents of catheter related bloodstream infection (CRBSI) (isolated from both the catheter tip and blood cultures of 15 patients), 89 being isolated from peripheral blood cultures of patients without a central venous catheter (CVC) (13 of them were bloodstream infection agents, 76 of them were contaminant), and 35 being isolated as catheter colonizer, were screened by tissue culture plate (TCP), Congo red agar (CRA) method and polymerase chain reaction (icaA, icaD and IS256). Vancomycin minimum inhibitory concentration (MIC) and minimum biofilm eradicating concentration (MBEC) values were determined. The pulsed field gel electrophoresis (PFGE) method was used to show the clonal relationship between CoNS isolated from the catheter tips and peripheral blood of patients with CRBSI. Of the 154 CoNS isolates included in the study, 38.9% were Staphylococcus epidermidis (n= 60), 34.4% were Staphylococcus haemolyticus (n= 53), 20.7% were Staphylococcus hominis (n= 32), and 3.8% were detected as Staphylococcus capitis (n= 6). In our study, biofilm formation was shown in 31.8% with the CRA method and in 68.1% with the TCP method. By TCP method, 22% (n= 34) were determined as weak, 31.2% (n= 48) medium and 14.9% (n= 23) strong biofilm producers. While the sensitivity of the CRA method was found to be low for isolates that were determined as weak positive in the microplate method, the high sensitivity of the CRA method for isolates with medium and strong positivity was found remarkable. The positivity rates of icaA, icaD and IS256 genes in a total of 154 CoNS isolates were found to be 40 (25.9%), 57 (37%) and 77 (50%), respectively. In total, at least one gene positivity was detected in 107 (69.5%) isolates. Single gene positivity was detected in 55 (35.7%), two gene positivity in 35 (22.7%) and three gene positivity was detected in 17 (11%) of the included CoNS. Biofilm formation (four weak, four medium, two strong) was detected by microplate method in 10 of 47 CoNS isolates (five S.epidermidis, three S.hominis, one S.haemolyticus and one S.capitis) in which no genes were detected. Vancomycin MBEC/ MIC values were found to be high and it was observed that as the biofilm forming power of the isolates increased, the MBEC/MIC ratio also increased. The CoNS isolated from the catheter samples and blood of patients diagnosed with CRBSI had a 100% similar profile with PFGE except for one unevaluable isolate. The tissue culture plate (TCP) method was found to be most sensitive, accurate and reproducible screening method for detection of biofilm formation by staphylococci and has the advantage of being a quantitative model to study the adherence of staphylococci. The presence of the icaAD and IS256 gene is not always associated with in vitro biofilm formation. For this reason, it is more appropriate to use more than one method together for the evaluation of biofilm formation. It was thought that the use of reliable methods to specifically detect biofilms could be helpful in diseases that are difficult to treat. Considering the high rates of biofilm and antimicrobial resistance of biofilm-forming isolates in biomedical device associated infections, it was determined that it would not be sufficient to evaluate only the MIC results for susceptibility results.

A reliable method to avoid contamination during cartilage graft preparation in septorhinoplasty.

PURPOSE: The aim of the study is to determine the risk of contamination in the cartilage graft materials prepared on the swester table and those prepared in a sterile package, and to reveal a more reliable method by performing the microbiological examination of these materials. METHODS: Cartilages removed from the nasal septum were divided into four pieces. The first part (Sample A) was directly placed into the medium. Sample B was prepared by being crushed in a sterile package. Sample C was prepared on the auxiliary swester table, and Sample D was prepared on the main swester table actively used by surgery team. All samples were transferred in a 1 ml brain heart(BH) liquid medium. From each BH medium, 100 µl culture was performed on blood agar, eosin-methylene blue-lactose-sucrose agar and chocolate agar. RESULTS: Bacterial growth was detected in 2 of the samples A, in 4 of the samples B, in 24 of the samples C, and in 36 of the samples D. The number of patients with bacterial growth in the samples C and/or D despite no growth in the sample B was 35. When the samples A/B and C/D were compared in terms of bacterial growth, a significant difference was found in all matchings (p < 0.001 for all comparisons).  CONCLUSION: These findings showed that preparation of the cartilage grafts on the swester table was extremely risky for microbiological contamination. Arslan and his colleagues suggest that preparing a graft material in a sterile package is extremely simple, cheap, and it also reduces contamination risk significantly.

[In vitro effect of vancomycin and daptomycin on biofilm formation of coagulase-negative staphylococci strains]. / Vankomisin ve daptomisinin koagülaz-negatif stafilokok izolatlarinin olusturdugu biyofilm üzerine in vitro etkisi.

Coagulase-negative staphylococci (CNS) are one of the primer agents of blood stream infections (BSI) and catheter-related bloodstream infections (CR-BSI) which are associated mostly with the usage of central venous catheters and, important causes of morbidity and mortality despite the usage of antibacterial and supportive treatment. It is important to determine the properties of these causative microorganisms in order to make appropriate treatment of such infections. The aims of our study were to evaluate the biofilm formation of coagulase negative staphylococci (CNS) which were causative agents of bloodstream (BSI) and catheter related bloodstream infections (CR-BSI), to determine the minimum inhibitory concentration (MIC) of planktonic forms and minimal biofilm eradication concentration (MBEC) of sessile forms for vancomycin and daptomycin and to evaluate the efficacy of these antibiotics in infections with biofilm-forming isolates in vitro. A total of 65 CoNS (n= 26 catheter colonizers, n= 28 CR-BSI, n= 11 BSI agents) were identified by conventional methods and also with BD Phoenix (Becton Dickinson, USA) and Bruker Microflex MS (Bruker Daltonics, Germany) systems. Methicillin resistance was determined by the presence of mecA gene with PCR. MIC values of vancomycin and daptomycin were investigated by broth microdilution, for daptomycin medium containing 25 and 50 µg/ml Ca++ were used. Assessment of biofilm formation and detection of MBEC were determined by microplate method. The clonal relationship was investigated by the PFGE method. A total of 65 isolates; 26 catheter colonizers, 28 CR-BSI agents and 11 BSI agents were evaluated and identified as Staphylococcus epidermidis (n= 33), Staphylococcus haemolyticus (n= 16), Staphylococcus hominis (n= 15), and Staphylococcus capitis (n= 1). 81.5% of the isolates were found to be methicillin resistant and all of them were found to be sensitive to vancomycin (MIC= 0.125-4 µg/ml) and daptomycin (MIC= 0.062-0.25 µg/ml in 25 µg/ml Ca++ and MIC= 0.031-0.50 µg/ml in 50 µg/ml Ca++ containing medium). MIC values were lower in medium containing 50 µg/ml Ca++ for daptomycin. As it is known that the efficacy of daptomycin depends on the physiological levels of Ca++, which causes conformational changes in the structure of these antibacterials. Our findings also suggested that high levels of Ca++ are needed to ensure the efficacy of daptomycin. All of the isolates produced biofilm at different strengths of positivity (n= 12/18.5% weak, n= 35/%53.8 moderate, n= 18/%27.7 strong). MBEC and MBEC/MIC values for vancomycin were found to be higher than daptomycin (p< 0.001). Strong biofilm producers had higher MBEC and MBEC/MIC, MBEC50/MIC50 ve MBEC90/MIC90 values (p< 0.05). Especially in infections with biofilm forming isolates, the detection of only MIC values are not always sufficient in the treatment of biofilm-related infections as they reflect the sensitivity of planktonic bacteria. The inconsistency between the MIC and MBEC values and the high rates of MBEC/MIC found in our study supported this prediction.The lower detection of MBEC and MBEC/MIC values of daptomycin compared to the same values of vancomycin suggested that daptomycin might be effective at lower doses than vancomycin in the treatment of biofilm infections.

[Investigation of biofilm formation properties of staphylococcus isolates]. / Stafilokok izolatlarinin biyofilm olusturma özelliklerinin arastirilmasi.

Biofilm production is an important virulence factor which allows staphylococci to adhere to medical devices. The principal component of biofilm is a "polysaccharide intercellular adhesin (PIA)" which is composed of a beta-1,6-N-acetylglucosamine polymer synthesized by an enzyme (N-acetylglucosamine transferase) encoded by the ica operon found on the bacterial chromosome. This operon is composed of four genes (A, B, C, and D), and a transposable element IS256. In this study, we aimed to determine the biofilm production characteristics of invasive/non-invasive staphylococcus isolates and different staphylococcus species. Biofilm production of 166 staphylococci was phenotypically investigated on Congo Red Agar (CRA); the presence of icaA, icaD and IS256 genes were investigated by polymerase chain reaction (PCR). 74 of the isolates (44.6%) were identified as methicillin resistant Staphylococcus aureus (MRSA), 25 (15.1%) as methicillin sensitive S.aureus (MSSA), 25 (37.3%) as Staphylococcus hominis, 20 (12%) as S.epidermidis, ten (15%) as Staphylococcus haemolyticus, nine (13.4%) as Staphylococcus capitis, two (3%) Staphylococcus saprophyticus and one (1.5%) as Staphylococcus warnerii. Of the MRSA strains, 52 were isolated from blood and 22 from nose; all MSSA strains were isolated from nose cultures. Coagulase-negative staphylococci (CoNS) strains were composed of invasive and non-invasive strains isolated from nose, catheter tip and blood cultures from patients with catheter. Production with CRA method was found to be statistically significant in invasive isolates (p< 0.001). It is concluded that; as the biofilm formation capacity of invasive isolates can cause refractory infections and the importance of carriage and hospital infections of these bacteria, it is important to prevent the spread of these isolates. A combination of phenotypic and genotypic tests is recommended for the investigation of biofilm formation in staphylococci. 40.3% of the CoNS isolates, and 85.8% of S.aureus isolates produced biofilm on CRA (p< 0.001) and with PCR method the ratio of carrying three genes was found to be statistically important in S.aureus when compared with CoNS. Carriage of three genes and biofilm formation capacity of invasive isolates can cause refractory infections and the importance of carriage and hospital infections of these bacteria, it is important to prevent the spread of these isolates. A combination of phenotypic and genotypic tests is recommended for the investigation of biofilm formation in staphylococci.

[Molecular characterization of Salmonella enterica serovar Typhimurium strains isolated from humans in Turkey]. / Türkiye'de insanlardan izole edilen Salmonella enterica serovar Typhimurium suslarinin moleküler özellikleri.

Multidrug resistant (MDR) Salmonella infections, especially infections due to Salmonella Typhimurium DT104 phage type strains are an important public health issue in many parts of the world. S.Typhimurium is the most common serotype isolated from clinical samples in Turkey but we have limited data about the phage types of these isolates. The aims of this study were to find out whether these MDR S.Typhimurium isolates are DT104 phage type isolates and have class 1 integrons and to investigate the relationships of these characteristics between plasmid and pulsed field gel electrophoresis (PFGE) profiles. A total of 66 S.Typhimurium stock strains selected from Enterobacteria Laboratory culture collections of Ankara University School of Medicine, Department of Medical Microbiology were investigated by plasmid profile analysis (PPA) and PFGE with the use of XbaI and SpeI enzymes. The presence of class 1 integrons and the phage type 104 were investigated by polymerase chain reaction (PCR). The strains used in the study were sporadically isolated cases from seven provinces after year 2000 with ACSSuT (63), ACGSSuTT/S (1), ACSSuTT/S (1) and ASSuTT/S (1) resistance types [ampicillin (A), chloramphenicol (C), gentamicin (G), streptomycin (S), sulphonamide (Su), tetracycline (T), trimethoprim/sulfamethoxazole (T/S)]. Of the isolates 65 were found as DT104 phage type. Forty-three S.Typhimurium DT104 isolates that carry class 1 integrons had five different bands between 350-1600 base pairs (bp); all of the isolates harbored 1-4 plasmids with sizes ranging from 1.0-180 kbp and 62 isolates had 90 kbp plasmid which was serotype specific and virulence related. S.Typhimurium DT104 isolates were grouped into five (X1-X5) and seven (S1-S7) profiles with XbaI and SpeI enzymes, respectively. When the profiles of the two enzymes were evaluated, 58 of the 65 (89.2%) isolates showed similar (X1.S1) profile. The molecular characteristics of the most S.Typhimurium isolates were clustered in similar groups when class 1 integron, plasmid and PFGE types were analyzed together. In this study we showed that nearly all S.Typhimurium isolates with five drug resistance pattern (ACSSuT) were DT104 isolates. PFGE profiles of these sporadic isolates suggested that they were epidemiologically related.

[Molecular characterization of beta-lactamase-associated resistance in Acinetobacter baumannii strains isolated from clinical samples]. / Klinik örneklerden izole edilen Acinetobacter baumannii suslarinda beta-laktamaz kaynakli direncin moleküler karakterizasyonu.

Acinetobacter baumannii is an important cause of nosocomial infections that particularly increase the mortality and the morbidity at the intensive care units of the hospitals. The aims of this study were to evaluate the resistance genes, antibiotic susceptibility and the clonal relations among Acinetobacter strains isolated from clinical samples and to determine the resistance mechanisms related to these bacteria in our hospital. A total of 201 A.baumannii strains isolated from different clinical samples (35.3% from tracheal aspirate, 27.3% from blood, 18.4% from abscess material, 19% from other samples) of 160 inpatients evaluated at the Ibni Sina Hospital Central Bacteriology Laboratory, Ankara University School of Medicine, Turkey from April 2010 to December 2011, were included in the study. Identification of the isolates and their susceptibility testing against amikacin, ciprofloxacin, tetracycline, sulbactam/ampicillin, trimethoprim/sulfametoxazole (SXT), ceftazidime, gentamicin, imipenem, levofloxacin, meropenem, piperacillin/tazobactam, cefoperazone/sulbactam, cefepime and colistin were performed by the automated systems, namely Vitek 2 (bioMérieux, France) and BD Phoenix (Becton Dickinson, USA). The molecular mechanisms of beta-lactamase resistance and the presence of integrons were analyzed by polymerase chain reaction (PCR). Moreover, since blaPER-1 gene is of high frequency in Turkey, it was also investigated in the isolates. Pulsed-field gel electrophoresis (PFGE) was performed to examine the clonal relations between isolates. Our results indicated that multidrug resistance rate of A.baumannii was 94.5% (190/201), while 94% (189/201) of the isolates were susceptible to colistin thus making it the most potent antimicrobial agent, followed by amikacin and SXT with a susceptibility rate of 32%. Twelve colistin-resistant isolates were further investigated with the E-test method (AB Biodisk, Sweden) and found to be colistin-resistant. While the results were negative for the genes responsible from metallo-beta-lactamase production, positive results were obtained for blaOXA genes at various rates (OXA-51 100%; OXA-23 91.5%; OXA-58 7%; OXA-24 2%). PFGE results revealed four different main clones (29 isolates in genotype A, 23 in genotype B, 18 in genotype C and 7 in genotype D) in the study population. No common epidemic isolate was detected. Class 1 integrons which take part in the transfer of resistance genes were detected in 112 (55.7%) isolates. There was no statistically significant difference between the genotype distributions of class 1 integron positive strains (p> 0.05). The relationship between the presence of integron in multidrug resistant isolates and resistance to tetracyclin, SXT, imipenem, meropenem, cefoperazone/sulbactam and cefepime were found to be statistically significant (p< 0.05). Of the isolates 42 (21%) were positive for blaPER- 1 gene and all were resistant to ceftazidime. This study indicated that blaOXA genes found together with blaOXA-51 genes play an important role in carbapenem resistance of A.baumannii strains. Moreover, multidrug resistance is still an important problem in infections caused by A.baumannii and integrons play a role in the transfer of the resistance genes. In conclusion, multidrug resistant A.baumannii strains were common in our hospital and our epidemiologic data would be helpful for further investigations and in therapeutical approaches.

The emergence of Klebsiella pneumoniae sequence type 395 non-susceptible to carbapenems and colistin from Turkey.

PURPOSE: OXA-48 producing Klebsiella pneumoniae is an emerging threat and outbreaks due to specific sequence types have been commonly reported. Here, we report an outbreak due to multidrug-resistant ST395 K. pneumoniae ST395. To the best of our knowledge, this is the first outbreak of K. pneumoniae ST395 harbouring blaOXA-48 genes in our country. METHODS: The strains were characterized by antimicrobial susceptibility, extended-spectrum ß-lactamase (ESBL) and carbapenemase production, plasmid-mediated colistin, high-level aminoglycoside, and quinolone resistance. Also multidrug efflux pumps and porin coding genes were investigated. Pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), wzi typing and plasmid analysis were used for the epidemiological relationships. RESULTS: All strains were positive for blaOXA-48 with at least one of the ß-lactamase genes (blaCTX-M, blaTEM, blaSHV) and harboured IncL plasmids. 16 of 20 (80%) isolates carried qnrA. All isolates were positive for aac(6')-1b, acrAB-tolC, ompK35, and ompK36 genes but none of them harboured 16s rRNA methyltransferase, mcr-1-5, qepA, oqxAB, and mdtK genes. All strains had the same PFGE pattern, that is, wzi type K2 and found to be ST395 with MLST. CONCLUSION: The association of ST395 with OXA-48-producers could be an emerging threat for Turkey and continuous monitoring is crucial to prevent the spread of these powerful strains.

miR-216b-5p promotes late apoptosis/necroptosis in trastuzumab-resistant SK-BR-3 cells.

Breast cancer is the most common cancer in women. The human epidermal growth factor receptor 2 (HER2) overexpressing subtype is related to poor prognosis with an aggressive phenotype and is reported as one of the most commonly seen subtypes. Trastuzumab is prevalently used as a treatment method for HER2+ breast cancer however, resistance to the drug frequently occurs following the treatment. MicroRNAs (miRNAs) are 19-23 nucleotide long small RNAs, which regulate gene expression at post-transcriptional level and studies show that there are differentially expressed miRNAs between drug sensitive and resistant groups, indicating that they might have some key roles in drug effectiveness. In this study, the aim is to find out the role of miR-216b-5p in trastuzumab resistance. SK-BR-3 cells developed resistance to trastuzumab after continuous treatment with increasing concentrations of the drug for 6 months. To investigate the effect of miR-216b-5p on cancer cell behavior in resistance state, proliferation, motility, and invasion capacities of these resistant cells were analyzed by xCELLigence real-time cell analyzer. To further understand the molecular mechanisms underlying the regulation of resistant SK-BR-3 cells by miR-216b-5p, microarray analysis was performed. Apoptosis analysis was also performed since the pathway enrichment analysis pointed out cell death related pathways. The proliferation, motility, and invasion capacities of the miR-216b-5p transfected resistant cells were diminished compared to sensitive cells. We identified the necroptosis signaling pathway as the result of microarray and pathway enrichment analyses. STAT1, IRF9, and PKR were validated as the significant elements of the pathway, which are also the putative targets of miR-216b-5p. Our apoptosis analysis showed that a significant amount of trastuzumab resistant SK-BR-3 cells entered to late apoptosis/necrosis stage upon miR-216b-5p overexpression, it could be concluded that reexpression of miR-216b-5p sensitizes trastuzumab resistance through necroptosis in breast cancer.

[Molecular epidemiology of carbapenem-resistant Klebsiella pneumoniae strains isolated between 2004-2007 in Ankara University Hospital, Turkey]. / Ankara Universitesi tip fakültesinde 2004-2007 yillari arasinda izole edilen karbapeneme dirençli Klebsiella pneumoniae suslarinin moleküler epidemiyolojisi.

The prevalence of carbapenem-resistant gram-negative bacteria in the hospital setting is in an increasing trend worldwide. Since most of the carbapenem-resistant Enterobacteriaceae are resistant to all antimicrobial agents except polymyxins and tigecycline, the emergence of carbapenem resistance in Klebsiella pneumoniae strains requires careful monitoring. This study was conducted to analyse the epidemiological relatedness between the carbapenem-resistant isolates of K. pneumoniae collected from different wards (intensive-care, surgery, hematology, neurology, internal medicine, emergency services) of Ankara University Hospital. A total of 26 carbapenem-resistant K. pneumoniae isolates (13 blood, 6 urine, 2 bronchoalveolar lavage, 1 abscess, 1 tissue, 1 catheter tip, 1 drainage fluid, 1 tracheal lavage fluid) were identified and antibiotic susceptibility tests were performed with API 20E System or VITEK 2 Compact (Bio-Merieux, France) at the Central Laboratories of Ankara University Hospital between February 2004 and April 2007. MICs of imipenem and meropenem were also confirmed using E-test (AB Biodisk, Sweden). The clonal relationship between the isolates was studied by pulsed-field gel electrophoresis (PFGE). After digestion of total genomic DNA with restriction endonuclease Xbal, the 26 isolates generated 7 PFGE profiles. PFGE pattern B consisting of different antibiotic susceptibility profile was seen only in 2006. Carbapenem-sensitive strains isolated at the same time from the same wards which carbapenem-resistant isolates were recovered, generated different PFGE patterns. The predominant carbapenem-resistant isolates in our hospital were found clonally related. Interhospital transmission of carbapenem-resistant K. pneumoniae strains which have a particular epidemic potential, is likely to occur during patient transfer between wards. It is likely that intensive efforts, similar to those used to control vancomycin resistant enterococci, are needed to identify and control the spread of resistant Klebsiella species. Therefore, active surveillance and strict infection control measures for this multidrug-resistant microorganism should be implemented at local and national basis.

[The performance of two different chromogenic media for the diagnosis of extended spectrum beta-lactamase producing strains]. / Genislemis spektrumlu beta-laktamaz ureten suslarin tanisinda iki farkli kromojenik agarin performansi.

The aim of this study was to evaluate the performances of 2 different chromogenic media for the detection of extended spectrum beta-lactamase (ESBL) producing Escherichia coli and Klebsiella spp. isolated from different clinical samples of patients who were admitted to Baskent University Faculty of Medicine, Adana Application and Research Hospital between September to November 2007. A total of 365 strains [251 were ESBL positive and 114 were negative by double disc synergy (DDS) test] of which 255 were E. coli and 110 were Klebsiella spp. were included to the study. All the isolates have been inoculated onto Drigalski agar (prepared following the formula described by Stürenburg et al.) and chromID ESBL agar (bioMérieux, France) and the production of ESBL were evaluated at 4th and 24th hours for Drigalski agar, and at 24th hours for chromID ESBL agar. The strains which yielded contradictory results by DDS test and chromogenic media, were tested for the presence of TEM, SHV and CTX-M genes by polymerase chain reaction (PCR). The number of ESBL positive strains on Drigalski agar at 4th and 24th hours were 238 and 235, respectively, while chromID ESBL agar detected 259 ESBL positive strains at 24th hours. All (100%) of the 159 ESBL positive E. coli strains by DDS test were also found positive on chromID ESBL agar, and 150 (94.3%) were found positive on Drigalski agar. These rates were detected as 100% (92/92) and 92.3% (85/92) for Klebsiella spp. isolates. Eight of the strains (2 E. coli, 6 Klebsiella spp.) which yielded negative results by DDS test but positive on chromlD ESBL agar, harboured SHV (n = 1), CTX-M (n = 6) and TEM + CTX-M (n = 1) genes detected by PCR. As a result, the consistency of the results obtained by Drigalski agar at 4th and 24th hours has indicated that this medium may provide advantages in rapid diagnosis of ESBL producing bacteria in routine laboratories. The data obtained for chromogenic media seemed to be favourable for the rapid diagnosis of ESBL production, however comparative studies with the use of standard reference methods are needed in order to determine diagnostic sensitivity and specificity of these media.

[Investigation of presence of class 1 integrons in clinical isolates of carbapenem-resistant Acinetobacter baumannii]. / Klinik örneklerden izole edilen karbapeneme dirençli Acinetobacter baumannii suslarinda sinif 1 integron tasiyiciliginin arastirilmasi

Acinetobacter species, particularly Acinetobacter baumannii, are important opportunistic pathogens responsible for nosocomial infections. They are often resistant to a wide range of antibiotics, including broad-spectrum beta-lactams, aminoglycosides and quinolones. This study was aimed to investigate the presence of class 1 integrons in nosocomial A.baumannii isolates. Eighty-nine carbapenem resistant nosocomial A.baumannii strains recovered from various clinical samples at Ankara Numune Teaching and Research Hospital during September 2006-August 2007, were included in the study. To determine the presence of integrons in Acinetobacter isolates, a chromosomal DNA region that consists of internal variable gene sequences restricted to two conserved regions, was amplified by using 5'CS and 3'CS primers. Class 1 integrons were demonstrated in 93.3% (83/89) of the strains. The range of inserted gene cassette sizes detected varied from 100 to 3000 base pairs. Recent studies have shown that the majority of integrons belong to class 1 among Acinetobacter species. This study also indicated that class I integrons were present in 93.3% of the A.baumannii isolates. The isolates were genotyped by pulsed-field gel electrophoresis (PFGE) and found to be distributed into 13 different groups, two of the groups predominated the isolates (group A: 29, group C: 21 isolates). Five of 6 isolates that did not have the class 1 integron (6/89; 6.7%) exhibited the same PFGE pattern (group C). Since integrons are important for the dissemination of antibiotic-resistance genes among nosocomial Acinetobacter species, the investigation of integrons by polymerase chain reaction method seems to be a rapid and simple technique for revealing the epidemic potential of A.baumannii isolates.

Detection of sasX Gene and Distribution of SCCmec Types in Invasive and Non-invasive Coagulase-negative Staphylococci

Background: Coagulase-negative staphylococci, which belong to the normal microbiota of the skin and mucous membranes, are opportunistic pathogens. sasX, a newly described protein, is thought to play an important role in nasal colonization and methicillin-resistant Staphylococcus aureus virulence, and it may be acquired from coagulase-negative staphylococci by horizontal gene transfer. It has been considered that understanding the function of sasX gene may help clarify the relevance of the different adhesion mechanisms in the pathogenesis of infections associated with biofilm. Aims: To investigate the sasX gene presence, staphylococcal cassette chromosome mec types, and antimicrobial resistance patterns of invasive and noninvasive coagulase-negative staphylococci isolates. Study Design: Cross-sectional study. Methods: The study included a total of 180 coagulase-negative staphylococci strains. Non-invasive isolates (n=91) were obtained from the hands of healthy volunteers who do not work at the hospital (n=30), the nasal vestibule of healthy volunteer hospital workers (n=26), and central venous catheter (n=35). Invasive isolates (n=89) were isolated from peripheral blood cultures of inpatients who do not have catheters. All isolates were identified by conventional microbiological methods, automated systems, and, if needed, with matrix-assisted laser desorption/ionization-time of flight. Staphylococcal cassette chromosome mec typing, sasX and mec gene detection, antibiotic susceptibility, and sasX gene sequence analysis were performed. Results: Peripheral blood, central venous catheter colonization, and nasal vestibule isolates were positive for the sasX gene, whereas hand isolates were negative. sasX gene was present in 17 isolates, and no statistical significance was found between invasive and noninvasive isolates (p=0.173). Sequence analysis of the sasX genes showed high homology to related proteins of Staphylococcus phage SPbeta-like and Staphylococcus epidermidis RP62A. staphylococcal cassette chromosome mec type V was the most prevalent regardless of species. staphylococcal cassette chromosome mec type II was more frequent in invasive isolates and found to be statistically important for invasive and noninvasive S. epidermidis isolates (p=0.029). Staphylococcus haemolyticus isolates had the overall highest resistance rates. Resistance to ciprofloxacin, trimethoprim-sulfamethoxazole, and erythromycin was found to be higher in isolates from catheter and blood culture. Staphylococcus hominis isolates had the highest rate for inducible clindamycin resistance. None of the isolates were resistant to vancomycin, teicoplanin, and linezolid. Conclusion: The sasX gene is detected in 9.44% of the isolates. There is no statistical difference between the sasX-positive and -negative isolates in terms of antibacterial resistance and the presence of sasX and SCCmec types. Further studies about the role of sasX at virulence in coagulase-negative staphylococci, especially from clinical samples such as tracheal aspirate and abscess isolates, and distribution of staphylococcal cassette chromosome mec types are needed.

Characterization of Klebsiella pneumoniae Coproducing KPC and NDM-1 Carbapenemases from Turkey.

Aims: The emergence of multidrug-resistant and carbapenem-resistant Klebsiella pneumoniae has became a major public health threat. In this study, we describe the characteristics of isolates coproducing KPC and NDM-1 carbapenemases from patients hospitalized at an emergency unit in Ankara, Turkey, between January and August 2018. The isolates were characterized by antibiogram susceptibility, carbapenemase and extended-spectrum beta-lactamase production, plasmid-mediated colistin (COL) resistance, and high-level aminoglycoside resistance. Pulsed field gel electrophoresis (PFGE), sequencing, wzi typing, multilocus sequence typing, and plasmid analysis were used to investigate the epidemiological relationship between the isolates. Results: All isolates were found to be resistant to amoxicillin-clavulanic acid, piperacillin-tazobactam, cefotaxime, cefoxitin, cefuroxime, ceftazidime, cefepime, imipenem, meropenem, ertapenem, amikacin, gentamicin, ciprofloxacin, levofloxacin, and trimethoprim-sulfamethoxazole. The minimum inhibitory concentration values for imipenem, meropenem, and ertapenem were >32 µg/mL, and >256 µg/mL for amikacin and gentamicin, and two isolates were found to be susceptible to both tigecycline and COL. All strains were positive for SHV, CTX-M, and rmtC, and negative for mcr-1 genes. A/C and FIIAS plasmids were found in all isolates. All isolates had the same PFGE pattern: wzi type 93 and ST15. Conclusion: Here, we have documented the characteristics of KPC- and NDM-1-coproducing isolates that harbored SHV, CTX-M, and rmtC and were typed as wzi 93 and ST15. We conclude that continuous monitoring of carbapenemases for unusual carbapenemase production is crucial to prevent the spread of these powerful isolates.

Prospective Analysis of Hemorrhagic Cystitis and BK Viremia in Allogeneic Hematopoietic Stem Cell Transplantation

Objective: BK virus (BKV) infection has been shown to be related to hemorrhagic cystitis (HC) in allogeneic hematopoietic stem cell transplantation (allo-HSCT). There are conflicting data regarding the association between BKV titers in plasma and clinical disease as well as the risk factors for BKV-related HC. Our aim is to study the risk factors and relationship with plasma BK viral load for development of HC in a prospective analysis. Materials and Methods: We prospectively evaluated 59 patients who received allo-HSCT between 2014 and 2016 by quantitative BK virus polymerase chain reaction (PCR) (Altona Diagnostics, Germany) from blood samples at days 0, 30, 60, and 90 after allo-HSCT. The patients were monitored for signs and symptoms of HC. Results: HC was diagnosed in 22 patients (37%) at a mean of 100 days (range: 0-367 days). In multivariate analysis, the usage of cyclophosphamide (sub-distribution hazard ratio [sdHR]: 7.82, confidence interval [CI]: 1.375-39.645, p=0.02), reactivated CMV (sdHR: 6.105, CI: 1.614-23.094, p=0.008), and positive BKV viremia (sdHR: 2.15, CI: 1.456-22.065, p=0.01) significantly increased the risk of developing HC. Patients with higher viral loads at day 30 and day 60 were diagnosed with more severe HC (p<0.001). Median BK viral loads of >101.5 copies/mL at day 0 (sensitivity 0.727, specificity 0.875), >98.5 copies/mL at day 30 (sensitivity 0.909, specificity 0.875), and >90.0 copies/mL at day 60 (sensitivity 0.909, specificity 0.875) were indicative of HC. Conclusion: Our study showed that administration of cyclophosphamide, CMV reactivation, and BK virus positivity were associated with HC. Plasma BK virus PCR titers at days 0, 30, and 60 after transplant were sensitive tools for predicting clinically proven HC.

Molecular investigation of salmonella choleraesuis and salmonella hadar strains isolated from humans in Turkey.

Eleven Salmonella Choleraesuis and seven Salmonella Hadar strains isolated from various clinical humand samples were investigated by plasmid profile analysis, enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) and pulsed-field gel electrophoresis (PFGE) in order to obtain information at a molecular level on the epidemiology of S. Choleraesuis and S. Hadar, which are significantly present in Turkey. Plasmid profile analysis showed that 10 (90.9%) of 11 S. Choleraesuis isolates harbored one to two plasmids with sizes of 2.0, 5.0 or 6.5 kb; and 5 (71.4%) of 7 S. Hadar isolates harbored one to three plasmids ranging from 2.5 to 70 kb. ERIC-PCR was performed using ERIC-2 primers; since isolates within each serotype showed similar band models, we concluded that ERIC-PCR is not suitable for differentiating isolates within the same serotype and for grouping into clusters. In PFGE using the AvrII enzyme, S. Choleraesuis isolates formed three clusters, and S. Hadar isolates formed three clusters; using the XbaI enzyme, S. Choleraesuis isolates formed two clusters, and S. Hadar isolates formed four clusters. These results showed that plasmid profile analysis and PFGE are reliable and discriminative methods that would complement antibiograms, and could contribute to the investigation of outbreak epidemiology. This is the first report on S. Choleraesuis and S. Hadar isolates from Turkey investigated by plasmid profile analysis, ERIC-PCR and PFGE methods.

Acute pancreatitis, bacterial translocation, and different octreotide regimens: an experimental study.

PURPOSE: To determine the effect of octreotide, octreotide with zinc, levamisole, and misoprostol on the bacterial translocation that develops in rats with acute pancreatitis (AP). METHODS: A total of 36 rats were divided into six groups, each consisting of six rats. Only laparotomy was performed on the first group. Acute pancreatitis was performed on the second group. Octreotide was given to the third, fourth, fifth, and sixth groups. Octreotide, octreotide with zinc, levamisole, and misoprostol were given to groups III, IV, V, VI, respectively. Rats were euthanized 48 h after the occurrence of AP. Blood and mesenteric lymph node samples were collected for polymerase chain reaction (PCR). Pancreatic tissue and terminal ileum were obtained for histopathological examinations. RESULTS: The severity of pancreatitis and mucosal damage of the terminal ileum was higher in group II than groups I, III, IV, V, and VI, histopathologically (P < 0.05). There wasn't a significant difference with respect to OA with Zn or L or M and OA group (P > 0.05). A significant difference was found in PCR positivity in blood and mesenteric lymph node between groups I and II (P < 0.05). CONCLUSIONS: In AP, administering octreotide alone significantly prevented the bacterial translocation by preventing mucosal damage. The zinc, levamisole, or misoprostol with octreotide did not influence the results.

[Influence of reaction optimization on the results of PCR amplification of Panton-Valentine leukocidin genes among Staphylococcus aureus isolates]. / Staphylococcus aureus izolatlarinda Panton-Valentin lökosidin genlerinin polimeraz zincir reaksiyonu ile amplifikasyon sonuçlari üzerine reaksiyon kosullarinin etkisi.

Panton-Valentine leukocidin (PVL) is an important virulence determinant of Staphylococcus aureus. Polymerase chain reaction (PCR) amplification of the genes encoding PVL is the most widely used method for determining PVL-positivity. In this study, we used two different primer sets and different annealing temperatures for each set to investigate the effect of optimization of PCR parameters on the amplification results. A total of 321 S. aureus clinical isolates (84.4% methicillin-resistant S. aureus, 76.9% nosocomial) were included to the study. Two different primer sets and two different annealing temperatures were applied for the amplification of PVL gene. For this purpose while luk-PV-1 and luk-PV-2 primers and 55 degrees C and 58 degrees C annealing temperatures were used to amplify the 433 bp region inhabiting the luk-S PV and luk-F PV genes, PVLup and PVLdn primers and 50 degrees C and 48 degrees C annealing temperatures were used to amplify the 1918 bp region inhabiting the same genes. luk-PV-1 and luk-PV-2 primers yielded amplicons at 55 degrees C in 50.2% (161/321) and at 58 degrees C in 1.6% (5/321) of the isolates. To discriminate the positive amplicons from the crossly amplified PCR products, restriction endonuclease analysis was performed and it was observed that the five amplicons generated by luk-PV-1 and luk-PV-2 primers at 58 degrees C were cut by BspH1 enzyme as expected for the positive amplicons. None of the isolates yielded amplicons by PVLup and PVLdn primers at 50 degrees C, however, only 1.6% of the isolates yielded amplicons at 48 degrees C. These isolates were the same with the ones that were PVL positive with luk-PV-1 and luk-PV-2 primers at 58 degrees C. These data revealed that only 1.6% of the study isolates were PVL positive. These results showed that inappropriate cycling conditions may lead to false-negative or false-positive results in PVL-gene amplification. Restriction endonuclease or sequence analysis may be used to differentiate crossly-amplified sequences from PVL-positive amplicons. We suggest using different primers and different annealing temperatures in PVL gene amplification before concluding on the study results. Using positive and negative control strains in each run and optimization of the study protocol not only according to the literature data but also due view to the laboratory experience seems critical for obtaining reliable data.