[In Vitro Activity of Ceftazidime-avibactam and Colistin Against Carbapenem-Resistant Klebsiella pneumoniae Clinical Isolates]. / Karbapenem Dirençli Klebsiella pneumoniae Klinik Izolatlarina Karsi Seftazidim-avibaktam ve Kolistinin In Vitro Etkinligi.

Infections caused by multi drug-resistant gram-negative bacilli are increasingly reported worldwide. Colistin, tigecycline and aminoglycosides are almost the only and last choice antibiotics in the treatment of infections caused by carbapenem-resistant Enterobacterales members. Ceftazidime-avibactam is a novel antibiotic combination consisting of a broad-spectrum cephalosporin and avibactam with good antimicrobial activity against carbapenem-resistant Enterobacterales members. The aim of this study was to assess the in vitro activity of ceftazidime-avibactam and colistin against carbapenem-resistant Klebsiella pneumoniae isolates and to obtain local antimicrobial surveillance data. A total of 150 carbapenem-resistant K.pneumoniae isolates obtained from various clinical samples of the patients hospitalized in our hospital between 2018-2021 were included in the study. Duplicate isolates were excluded from the study. The isolates were recovered from blood (n= 72), tracheal aspirate (n= 40), wound (n= 20), biopsy and abscess (n= 10), steril body fluid (n= 5), and peripheral venous catheter (n= 3) samples. Isolates were identified by matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS, Bruker Daltonics, Germany). The minimum inhibitory concentration (MIC) values of the isolates for meropenem, colistin, ceftazidime, and ceftazidime-avibactam were determined by broth microdilution method. Susceptibility of the isolates to the tested antibiotics was evaluated by the European Committee of Antimicrobial Susceptibility Testing (EUCAST) criteria. The presence of carbapenemases (VIM, IMP, NDM, KPC, and OXA-48) was investigated by polymerase chain reaction (PCR) using specific primers. The mcr-1, mcr-2, mcr-3, mcr-4, and mcr-5 genes were evaluated by PCR for plasmid-mediated colistin resistance. All K.pneumoniae isolates were found to be positive for at least one of the carbapenemase genes evaluated in the study. The blaOXA-48 gene was detected in 107 (71.3%), blaKPC gene in 25 (16.7%); blaNDM gene in 7 (4.7%), co-production of blaOXA-48 and blaKPC genes in 10 (6.7%), co-production of blaOXA-48 and blaNDM genes in 1 (0.6%) isolate. None of the isolates harbored the blaVIM and blaIMP genes. None of the mcr genes screened in the study were detected among the isolates. The susceptibility of the isolates to ceftazidime-avibactam and colistin was 92.7% (139/150) and 48% (72/150), respectively. The MIC50 and MIC90 values for meropenem, ceftazidime, ceftazidime-avibactam, and colistin of the isolates were determined as 32/256, > 128/> 128, 1/8, and 4/16 µg/ml, respectively. Of the ceftazidimeavibactam resistant isolates, seven were positive for blaNDM, three for blaKPC, and one for both blaOXA-48 and blaNDM genes. High ceftazidime-avibactam MIC levels (> 128 µg/ml) were detected in metallo-betalactamase producing isolates. Consequently, our data suggested that ceftazidime-avibactam exhibited as a good alternative therapeutic choice for carbapenem-resistant K.pneumoniae isolates. It is noteworthy that high rate of colistin resistance was detected in K.pneumoniae isolates. Another notable finding of this study is the increase in K.pneumoniae isolates producing blaKPC for our country. To prevent the development of resistance which is observed even in last-choice therapeutic antibiotics, the principles of rational antibiotic use should be followed. The appropriate antimicrobial susceptibility testing should be routinely performed for surveillance of ceftazidime-avibactam and colistin.

An Investigation into Bacterial Bloodstream Infections and Antibiotic Resistance Profiles in a Tertiary Hospital for a Ten-Year Period.

BACKGROUND: Bloodstream infections are one of the major causes of healthcare-associated morbidity and mortality. The present study aims to investigate the prevalence of the microorganisms isolated from blood cultures and to evaluate susceptibilities to antimicrobial agents in a tertiary center, Gulhane Training and Research Hospital, Ankara, Turkey. METHODS: Blood cultures (BCs) were incubated in BACTEC/9050 (Becton Dickinson, USA) (2007 - 2015) and BacT/ALERT (bio-Merieux, France) (2014 - 2016) automated systems. PhoenixTM 100 system (Becton Dickinson, USA) (2007 - 2014), MALDI-TOF MS (Bruker, USA) (2015 - 2016) and conventional techniques were used for the identification of isolated microorganisms. According to CLSI (2007 - 2014) and EUCAST (2015 - 2016) criteria, Kirby-Bauer disc diffusion method, PhoenixTM system, and broth microdilution were applied for antimicrobial susceptibility testing. Two five-year periods were statistically compared regarding antibiotic resistance. RESULTS: From the overall evaluated 31,380 BCs, 7,367 cultures (23.5%) were positive, excluding 503 BCs (6.4%), which were interpreted as contamination. Of 7,367 isolated microorganisms, 3,680 (50.0%) were gram-negative, 3,303 (44.8%) were gram-positive bacteria, and 384 (5.2%) were fungi. Coagulase-negative staphylococci (CoNS) were predominantly isolated (n = 2,075; 28.2%) among gram-positives. E.coli (n = 978; 13.3%) was the most frequently isolated gram-negative species. Between the first and the last five-year period, three genera (Enterococcus spp., Acinetobacter spp., Streptococcus spp.) showed significant differences when isolated, and only Enterococcus spp. showed increased isolation rates. In total, 90.3% of CoNS and 32% of S. aureus were methicillin-resistant. Only 75 strains of Enterococcus spp. (12.1%) were vancomycin-resistant. ESBL was detected in 40.6% of E. coli and 30.7% of Klebsiella spp. isolates. Carbapenem resistance showed a significant increase, particularly in K. pneumoniae (> 20%). CONCLUSIONS: The findings suggest that there was a threatening condition in antimicrobial resistance rates, especially for some antimicrobials between two periods. Although antimicrobial resistance is usually associated with MRSA, carbapenem resistance, ESBL, and VRE, the problem is far beyond these definitions, consisting of not just medicine, but also commercial companies, food industry, veterinarians, and other areas.

Intracranial infections: lessons learned from 52 surgically treated cases.

OBJECTIVE: Intracranial infections are serious and life-threatening health problems. They may present as subdural empyemas or intracerebral abscesses. Surgical drainage and subsequent antibiotic treatment is the main technique for a satisfactory clinical outcome. The aims of this study were to present a 10-year intracranial infection series and discuss the surgical characteristics in the light of literature. METHODS: Fifty-two patients with intracranial infection underwent surgical treatment between 2008 and 2018. Eleven patients were female and 41 patients were male. The mean age was 40.46 years (range 10-75 years). Eighteen patients had intracerebral abscesses, and 34 had subdural empyemas. All patients underwent surgical treatment as well as an antibiotic regimen. RESULTS: No etiological agent was isolated in 29 (56%) cases. Bacterial agents were detected in 20 cases, while fungi were observed in 3 cases. Staphylococci species were the most common agents and were isolated in 8 (15%) cases. Endoscopic aspiration was performed in 3 cases, while surgical drainage and capsule resection via craniotomy was performed in 49 cases. An associated intracranial tumor was diagnosed in 2 patients with brain abscesses. Four (8%) patients died despite surgical and medical treatments. CONCLUSIONS: Surgical treatment via craniotomy is an older method, but it is still the best to treat the intracranial infections not only for decompression of the brain but also to attain an accurate diagnosis. The abscess wall should always be histologically examined after surgery to rule out any intracranial tumor.

The effects of different types of hair shaving on the body image and surgical site infection in elective cranial surgery.

AIMS AND OBJECTIVES: To investigate the effects of different types of shaving on body image and surgical site infection in elective cranial surgery. BACKGROUND: Hair shaving before cranial surgery is commonly performed in many countries. However, the impact of shaving on the patients' body image and surgical site infection is not, as yet, well investigated. DESIGN: A randomised-controlled design was used in this study. METHODS: The sample comprised 200 patients who underwent elective cranial surgery between March 2013-August 2014. The Center for Disease Control and Prevention criteria were applied for the preoperative preparation of patients and for the follow-up of surgical site infection. Wound swab cultures were obtained four times from all patients. The Social Appearance Anxiety Scale was used to assess changes in the body image of patients. FINDINGS: The rate of surgical site infection was 1% for each group and for all patients. There was no difference between the groups of surgical site infection. Coagulase-negative staphylococci and Staphylococcus epidermidis were mostly isolated in the swab cultures. The Social Appearance Anxiety Scale score decreased in patients who underwent strip shaving and increased in patients with regional shaving. CONCLUSION: There is no difference between strip shaving and regional shaving in the development of surgical site infection after cranial surgery. In addition, regional hair shaving negatively affects the patients' body image. RELEVANCE TO CLINICAL PRACTICE: Findings of this study provide useful evidence-based information for healthcare professionals. The development and implementation of effective interventions result in the prevention of surgical site infection and improvement of the patients' body image in elective cranial surgery.

Effect of 2 elastomeric ligatures on microbial flora and periodontal status in orthodontic patients.

INTRODUCTION: The aim of this study was to compare the effects of a nonconventional elastomeric ligature (Slide; Leone, Florence, Italy) with those of a conventional elastomeric ligature (Ormco, Orange, Calif) on microbial flora and periodontal status in orthodontic patients. METHODS: A total of 13 orthodontic patients scheduled for fixed orthodontic treatment were selected for this study. The use of Slide and conventional elastomeric ligatures in fixed orthodontic appliances was tested. Microbial and periodontal records were obtained before bonding and 1 and 5 weeks after bonding. For the statistical analysis and calculations, SPSS software (version 15.0; SPSS, Chicago, Ill) was used. In the statistical decisions, P <0.05 values were accepted as significantly different. RESULTS: No significant differences between Slide and conventional elastomeric ligatures were evident at 1 week or 5 weeks after bonding, with regard to gingival index, plaque index, gingival bleeding index, or pocket depth scores (P >0.05). Similarly, aerobic and anaerobic bacteria counts did not differ significantly on the surface or on the elastics (P >0.05). CONCLUSIONS: Although the Slide ligatures cover the total surface of the bracket, they do not cause significantly more plaque accumulation or periodontal problems than do the conventional elastomeric ligatures.

[Investigation of various virulence factors among the hospital and community-acquired Staphylococcus aureus isolates by real-time PCR method]. / Hastane ve Toplum Kaynakli Staphylococcus aureus Izolatlarinda Çesitli Virülans Faktörlerinin Gerçek Zamanli PCR Yöntemiyle Arastirilmasi

Staphylococcus aureus is the most common cause of skin and soft tissue infections in the community and the most important cause of nosocomial infections. In this research, it was aimed to detect the presence of staphylococcal enterotoxin A to D (SEA, SEB, SEC and SED, respectively), toxic shock syndrome toxin-1 (TSST-1), Panton-Valentine leukocidin (PVL) and SCCmec phenotype in methicillin-resistant S.aureus (MRSA) isolates and to demonstrate the genotypic association between hospital acquired and community acquired isolates. In the study the virulence factors of 147 S.aureus strains isolated from various clinical samples at Gulhane Military Medical Academy Hospital Microbiology Laboratory between 2007 and 2010 were investigated by real-time polymerase chain reaction. MLVA (multiple locus variable number of tandem repeat analysis) method was used to demonstrate the genotypic association between hospital-and community-acquired isolates. Seventy-two (%48.9) of 147 S.aureus isolates were determined as community acquired and 75 (%51.1) as hospital acquired. Ninety-three (63.2%) isolates possesed at least one toxin (77 strains harboured one, and 16 strains harboured more than one). Of the isolates in which toxin was detected 59.1% (55/93) were hospital acquired, 40.9% (38/93) were community acquired. SEA was determined in 59 (40.1%), SEB in 8 (5.4%), SEC in 12 (8.1%), SED in 8 (5.4%), TSST- 1 in 17 (11.5%) and PVL in 6 (4.0%) of the isolates. Methicillin resistance was determined in 61.1% (44/72) of the hospital acquired isolates and 6.7% of the (5/75) community acquired isolates. In our study, SCCmec type III was detected in 90.9% (40/44) of hospital acquired MRSA isolates and SCCmec type IV in 40.0% (2/5) of community acquired MRSA isolates. Most of the strains (40/47; 85.1%) carrying SEA were hospital acquired, and they were determined as methicillin-resistant. According to MLVA, hospital and community acquired groups' clustering rates, number of clones, number of unique profile were determined as; 73.6% and 57.3%; 34% and 47%; 19% and 32%, respectively. It was concluded that high prevalence of SEA toxin in hospital acquired MRSA isolates indicated that there was a possible association between the presence of toxin and antimicrobial resistance.

[Comparison of endotracheal aspiration and mini-BAL culture results in the diagnosis of ventilator-associated pneumonia]. / Ventilatörle Iliskili Pnömoni Tanisinda Kullanilan Endotrakeal Aspirat Kültürü ile Mini-BAL Kültürünün Karsilastirilmasi

The objective of this study was to compare the results of cultures obtained by mini-bronchoalveolar lavage (BAL) and endotracheal aspiration (ETA) techniques, used for rapid and accurate determination of pathogens causing ventilator-associated pneumonia (VAP) in intensive care units. Of the 92 patients on mechanical ventilation followed at the emergency intensive care unit of our hospital between June 2010 and June 2011, 30 (32.2%) patients were diagnosed as VAP and they were included in this study. VAP diagnosis were based on the clinical and radiological findings. Clinical pulmonary infection score (CPIS) of > 6 was accepted as the clinical criteria of VAP. Initially ETA samples were collected from the patients followed by mini-BAL sampling 15 minutes later, together with urine and two blood cultures. Microbiological evaluation and identification were performed by conventional methods and Phoenix 100 (BD Diagnostic Systems, ABD) automated system. In quantitative culture analysis, > 10.000 cfu/ml for BAL and > 100.000 cfu/ml for ETA were accepted as the positive result. The mean ages of VAP-developed (n= 30; 18 were male) and nondeveloped (n= 62; 39 were male) patients were 68.23 ± 16.19 and 52.16 ± 10.41 years, respectively, and the mean durations of mechanical ventilation were 29.57 ± 15.78 and 12.11 ± 6.01 days, respectively. Multivariate logistic regression analysis showed that older age (p< 0.001) and duration of mechanical ventilation (p< 0.001) were independent risk factors for VAP development. There was also a statistically significant difference in CPIS values between patients who developed VAP and not (6.8 ± 1.15 and 2.71 ± 1.06, respectively; p< 0.001). The use of CPIS for VAP diagnosis was found to be useful in patients on mechanical ventilation. In our study, a total of 16 strains (six A.baumannii, three P.aeruginosa, one K.pneumoniae, six S.aureus) were isolated from ETA cultures, while 34 strains (16 A.baumannii, six P.aeruginosa, four K.pneumoniae, two E.coli, six S.aureus) were isolated from mini-BAL cultures of 30 VAP patients. The contamination rate for ETA cultures was found as 27% (8/30), however there was no contamination in mini-BAL samples. The rates of negative cultures for ETA and mini-BAL were 20% (6/30) and 7% (2/30), respectively. Seven (87.5%) of the eight contaminated ETA samples, yielded pathogenic bacterial growth (six A.baumannii, one K.pneumoniae) in mini-BAL samples. Similarly, of the six negative ETA samples, 5 (83%) yielded bacterial growth (two E.coli, two K.pneumoniae, one P.aeruginosa) in mini-BAL samples. Statistical analysis with Spearman test indicated no positive correlation between the culture results of mini-BAL and ETA (p= 0.464), and the concordance between the culture results of those methods was found as 50%. It was concluded that the use of mini-BAL instead of ETA samples for the isolation of causative microorganisms of VAP seemed to be more useful due to the high contamination risk in ETA culturing techniques and higher bacterial isolation rates in mini-BAL sampling.

[Investigation of West Nile virus RNA in blood donors by real-time RT-PCR]. / Kan Donörlerinde Gerçek Zamanli RT-PCR ile Bati Nil Virusu RNA'sinin Arastirilmasi

West Nile virus (WNV), a member of Flaviviridae family, is an enveloped, icosahedral symmetric RNA virus. Primary reservoir hosts of WNV are birds, but the virus can cause various infections in humans and other mammals. The most common and natural transmission way of WNV infections is mosquito bites, however, humans can be infected by different routes. The most important non-mosquito transmission route is contaminated blood and blood products. In this study, we aimed to investigate the risk of WNV transmission through blood and blood products in Ankara, Turkey. The presence of WNV RNA was investigated by in house real-time reverse transcriptase-polymerase chain reaction (RT-PCR) in serum samples obtained from 729 healthy blood donors (mean age: 27.7 years; 711 were male), regardless of the donor's seropositivity status since the virus can be transmitted at the early stages of infection when seroconversion has not yet developed. Serum samples were collected in August-September 2009, the period when these infections are more frequent due to mosquito activity. The vast majority of donors (n= 702, 96.3%) have been inhabiting in Ankara and 569 (78%) of donors have had risk factors for arboviral infections (e.g. outdoor activity, mosquito and tick bites). WNV RNA was not detected by real-time RT-PCR analysis in any serum sample included in this study. According to the results of our study, it can be said that the risk of WNV transmission through blood and blood products is low in Ankara. However, WNV seropositivity was detected within the range of 0.56 to 2.4% among blood donors in previous studies and probable and confirmed WNV infections have been reported in our region. In addition, WNV outbreaks have emerged in some countries neighbouring Turkey recently. Thus, the risk of WNV transmission through blood and blood products should not be ignored and blood donor questionnaires should be evaluated in detail.

Ozone therapy prevents renal inflammation and fibrosis in a rat model of acute pyelonephritis.

INTRODUCTION: Not only bacterial characteristics but also oxidative/nitrosative stress could play a significant role in renal parenchymal inflammatory processes in acute pyelonephritis (APN). This study was conducted to evaluate the effect of ozone therapy (OT), as an immunomodulator and antioxidant, on the renal function, morphology and biochemical parameters of oxidative stress in an experimental model of APN in rats. MATERIALS AND METHODS: Forty rats were divided equally into five groups as control, APN, APN + Antibiotic, APN + OT, and APN + Antibiotic + OT. APN was induced by 0.1 ml of freshly prepared Escherichia coli (ATCC 25922) solution containing 10(10) colony-forming unit/ml into the kidney. A control group was administered 0.1 ml of 0.9 % NaCl solution. Treatment was begun 72 h after bacterial inoculation. Control and APN groups were given 0.9% NaCl solution, APN + Antibiotic and APN + OT were given either antibiotic (ciprofloxacine 150 mg/kg intramuscular/twice daily) or OT. APN + Antibiotic + OT group was given both antibiotic and OT for five consecutive days. At the end of the seventh day, animals were killed via decapitation and trunk blood was collected. Both kidneys were harvested and one half of each kidney were immediately stored for antioxidant enzyme activity, tissue lipid peroxidation and protein carbonyl content. The remainder was fixed for histopathologic examination. RESULTS: E. coli-induced APN increased the renal glomerular and tubular dysfunction, oxidative stress parameters and antioxidant enzyme activities. Either antibiotherapy or OT markedly ameliorated renal dysfunction, the antioxidant status of the kidneys and histopathological injuries subjected to E. coli-induced APN. Interestingly, the combination of antibiotherapy and OT was much more effective than either of the treatment modalities alone. CONCLUSION: The combination of antibiotherapy and OT markedly ameliorated renal dysfunction and improved antioxidant status and histopathologic modalities in rats subjected to E. coli-induced APN than either antibiotherapy or OT treatment alone. Therefore, OT may be considered as an adjuvant therapy to classical antibiotherapy to prevent renal inflammation and fibrosis in APN.

Effect of a self-etching adhesive containing an antibacterial monomer on clinical periodontal parameters and subgingival microbiologic composition in orthodontic patients.

INTRODUCTION: The aims of this study were to evaluate the effect of a self-etching adhesive system containing an antibacterial monomer on periodontal health and subgingival microbiologic composition in orthodontic patients and to compare it with a conventional adhesive system. METHODS: A split-mouth design was chosen, and 15 patients were included in the study. Brackets in contralateral quadrants were bonded with either a conventional adhesive system (control) or a self-etching adhesive system that contained an antibacterial monomer. Clinical periodontal parameters including plaque index, gingival index, probing depths, and bleeding on probing were determined. Subgingival plaque samples were collected before bracket placement (T0) and at the 6-month follow-up (T1). The real-time TaqMan polymerase chain reaction assay was used to determine the subgingival counts of Porphyromonas gingivalis, Tannerella forsythensis, Prevotella intermedia, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, and Campylobacter rectus. For clinical periodontal parameters, analysis of covariance (ANCOVA) and, for bacterial counts, Wilcoxon tests were used for statistical comparisons at the P <0.05 level. RESULTS: Clinical periodontal parameters were not changed, and they were not different between the groups from T0 to T1. T forsythensis and F nucleatum increased during the treatment period in both groups (P <0.05). The majority of the bacteria were T nucleatum at T0 and T1 in both groups. Changes in bacterial load from T0 to T1 were not different between groups except for T forsythensis and F nucleatum (P <0.05). CONCLUSIONS: The use of an antibacterial monomer did not have an additional positive effect on clinical periodontal parameters. When used in bonding orthodontic brackets, the antibacterial monomer failed to reduce periodontopathogenic bacteria when compared with the conventional adhesive system during a 6-month treatment period.

Impact of methods and duration of surgical hand scrub on bacterial count: A randomized controlled trial.

BACKGROUND: There is no standard protocol for surgical scrubbing. This study aims to compare the effectiveness of surgical hand scrub duration and method by analyzing their effects on bacterial count. METHODS: The study was conducted on 180 surgical nurses and surgeons. While the duration of surgical hand scrub in Groups I and II was one minute, participants in Group I used a nail brush, whereas Group II did not. Similarly, the duration of surgical hand scrub in Groups III and IV was two minutes, but Group III used a nail brush, whereas Group IV did not. Bacterial count on the hands of all participants was measured before and after the surgical hand scrub and after the surgery by using the glove juice method. RESULTS: Bacterial count on the hands of the participants in Group III after surgical hand scrub was significantly higher than Group IV (P < .001). We did not find any statistically significant difference between Group II and Group IV in terms of bacterial count on the hands immediately after surgical hand scrub and after the surgery (P = .401, P =.658, respectively). CONCLUSIONS: This study found that brushing during surgical hand scrub increased the number of bacteria on the hand. Besides, one-minute surgical hand scrub was equally effective as two-minute scrub to reduce the number of bacteria on the hand.

Stability of hepatitis C virus RNA in blood samples by TaqMan real-time PCR.

The storage conditions of blood samples for reliable results are very important in hepatitis C virus (HCV) RNA amplification tests used in routine HCV analyses. According to many studies, storage conditions could affect the RNA stability for HCV RNA detection. We have studied HCV RNA stability in blood samples stored at 4 degrees C. Nineteen blood samples containing different HCV RNA levels were stored at 4 degrees C and they were then analyzed by TaqMAN real-time PCR method. HCV RNA levels remained almost stable (100%) at least for five weeks at this storage condition. However, among them, the stability period was up to 11 weeks in two of the samples. As with these findings, there was a slightly significant correlation between the positivity time and the beginning HCV RNA levels (r=0.474, P=0.040). We conclude that, blood samples can be stored at 4 degrees C for five weeks without any significant difference in detected HCV RNA level by using TaqMan real-time PCR.

Simultaneous detection and differentiation of pathogenic and nonpathogenic Leptospira spp. by multiplex real-time PCR (TaqMan) assay.

Leptospirosis, caused by pathogenic Leptospira, is one of the most important zoonoses in the world. Several molecular techniques have been developed for detection and differentiation between pathogenic and saprophytic Leptospira spp. The aim of this study was to develop a rapid and simple assay for specific detection and differentiation of pathogenic Leptospira spp. by multiplex real-time PCR (TaqMan) assay using primers and probes targeting Leptospira genus specific 16S ribosomal RNA gene, the pathogen specific lig A/B genes and nonpathogen Leptospira biflexa specific 23S ribosomal RNA gene. Sixteen reference strains of Leptospira spp. including pathogenic and nonpathogenic and ten other negative control bacterial strains were used in the study. While the 16S primers amplified target from both pathogenic and non-pathogenic leptospires, the ligA/B and the 23S primers amplified target DNA from pathogenic and non-pathogenic leptospires, respectively. The multiplex real-time PCR (TaqMan) assay detection limit, that is, the sensitivity was found approximately 1 x 10(2) cells/ml for ligA/B gene and 23S ribosomal RNA gene, and 10 cells/ml 16S ribosomal RNA. The reaction efficiencies were 83-105% with decision coefficients of more than 0.99 in all multiplex assays. The multiplex real-time PCR (TaqMan) assay yielded negative results with the ten other control bacteria. In conclusion, the developed multiplex real-time PCR (TaqMan) assay is highly useful for early diagnosis and differentiation between pathogenic and non-pathogenic leptospires in a reaction tube as having high sensitivity and specificity.

[Expression of hepatitis B virus core antigen gene region in yeast cell]. / Hepatit B virusu kor antijeni gen bölgesinin mayada ekspresyonu.

In this study, the core antigen (HBcAg) gene region of hepatitis B virus (HBV) was transformed and expressed into an eukaryotic expression vector by recombinant DNA technology in order to obtain the protein used in anti-HBc tests which is being one of the most important marker for the serodiagnosis of HBV infections. For this purpose, HBV-DNA positive patient sera were used as the source of viral nucleic acids, and the primers coding HBcAg gene region have been designed. After the amplification of HBcAg gene region by polymerase chain reaction (PCR), the amplicons purified by Invisorb Spin Rapid PCR Kit" (Invitek, Germany), were cloned to pYES2.1 plasmid via the TOPO TA expression kit (Invitrogen, USA) and this plasmid was transformed to competent bacteria (TOPO 10F' Escherichia coli) by CaCl2 method. After competent bacteria were grown on LB (Lysogeny Broth) agar media supplemented with ampicillin, the plasmid "pYES2.1 + HBcAg" were isolated and transformed to Saccaromyces cerevisiae via the "S.c. EasyComp Transformation Kit" (Invitrogen, USA). Finally, the expression of HBcAg by the yeast was confirmed with the use of in house ELISA method. Since the diagnostic kits used in our country for hepatitis B serology are usually imported products, this creates a great economical burden. Thus, the experience and knowledge that builds up following such studies will help to produce our own diagnostic products using our equity.

Mycoplasma pneumoniae infection associated with pancytopenia: a case report.

Immune hemolytic anemia is a rare condition in childhood. Cold agglutinins have been implicated in the etiology of the hemolysis and frequently observed during Mycoplasma pneumoniae infections. We present here a case of cold agglutinin-related hemolytic anemia, thrombocytopenia, and leukopenia secondary to M. pneumoniae associated pneumonia. It is suggested that even though very rare, M. pneumoniae infection should be considered as the underlying disease in a patient presenting with pancytopenia.

[An outbreak of vanA genotype Enterococcus faecium in pediatric clinic of a training hospital]. / Bir egitim hastanesinin çocuk kliniginde vanA genotip Enterococcus faecium salgini.

Vancomycin-resistant enterococci (VRE) have been increasingly reported from most countries around the world following initial isolation from patients in United States and European countries. A vancomycin-resistant Enterococcus faecium (VREF) outbreak was determined by hospital infection control committee in the pediatric unit of Gulhane Military Medical Academy, Ankara, Turkey in the first week of March 2008. While one of the 4 VREF strains was isolated from urine culture of a patient with neuroblastoma, the remaining strains were isolated from cultures of urine and rectal swab samples of a patient with nephrotic syndrome and from the hospital room doorknob of this patient. Aims of this study were to determine antibiotic susceptibilities by E-test, to investigate the presence of vanA, vanB and vanC-2 resistance genes by polymerase chain reaction (PCR), and to genotype the 4 strains by pulsed field gel electrophoresis (PFGE) and repetitive PCR (rep-PCR) (DiversiLab, bioMérieux, France). All isolates conferred high level [minimum inhibitor concentration (MIC) > 256 mg/L] vancomycin and teicoplanin resistance by E-test method. The isolates were also found resistant to gentamicin, streptomycin, ampicillin, erythromycin, penicillin and were susceptible to tetracycline and linezolid. The vanA gene was detected in all strains by PCR. It was demonstrated that the 4 VRE strains belonged to a single clone as shown by both PFGE and rep-PCR methods. Prompt and accurate detection of VRE and determination of the genotypes is of crucial importance to prevent horizontal transfer of the strains in the hospital. When compared with PFGE, the DiversiLab commercial rep-PCR seems to be a reliable and more rapid method to detect the genetic relationship between strains leading to an outbreak.

[Genotypic characteristics of Neisseria meningitidis serogroup W-135 strains isolated as the agent of fatal meningitis in a military hospital]. / Bir askeri hastanede fatal menenjit etkeni olarak izole edilen Neisseria meningitidis serogrup W-135 suslarinin genotipik özellikleri.

The aim of this study was to describe the genetic characterization of a total of 6 Neisseria meningitidis serogroup W-135 strains isolated from patients with meningitis and carriers in a military hospital in 2007-2008. Suspected colonies on modified Thayer-Martin medium plates were screened for oxidase reactivity and Gram stain. If gram-negative diplococci were present, a biochemical profile by the API NH system was used for species confirmation. Pulse field gel electrophoresis typing of Nhel-digested DNA was performed by a previously described method. Multi-locus sequence typing (MLST) was performed using the standard primers as listed on the Neisseria MLST website. Three distinct sequence types (STs) were identified: ST-11, ST-2754, ST-3751. One of the clinical isolates was identified as the same sequence type with Hajj isolate (ST-11) and the isolate with ST-2754 was the same as the first Turkish clinical strain isolated in 2003. These data demonstrated that along with ST-11 which is a known Hajj isolate, the ST-2754 strain causing meningococcal disease in Turkey beginning from the year 2003, should be carefully monitored.

False resistance after artemether-lumefantrine treatment in a falciparum malaria patient in Turkey: A case report.

INTRODUCTION: Artemisinin-based combination therapy (ACT) is recommended by the World Health Organization as first-line treatment of uncomplicated Plasmodium falciparum malaria. ACT treatments failures among travellers returning from Africa to non-endemic countries are considered to be caused by resistance. CASE PRESENTATION: We report on a case of artemether-lumefantrine treatment failure in a Turkish traveller with uncomplicated P. falciparum malaria returning from Bamako, Mali. CONCLUSIONS: Information on returning travellers, includes ensuring that the patients receive supervised treatment with the recommended dose of a quality controlled medicine, routine follow-up of all cases, assessment of adequate absorption of the drug, and/or testing the prevalence of molecular markers of drug resistance if validated, can be an important source of an early warning system for emerging resistance.

Staphylococcal cassette chromosome mec (SCCmec) characterization and panton-valentine leukocidin gene occurrence for methicillin-resistant Staphylococcus aureus in Turkey, from 2003 to 2006.

Methicillin-resistant Staphylococcus aureus (MRSA) cause serious community-acquired and nosocomial diseases all over the world. We determined the SCCmec types and occurrence of the PVL gene by using TaqMan real-time PCR method, and correlated these with phenotypic antibiotic susceptibility patterns for MRSA strains collected from Gulhane Military Medical Academy Hospital (GMMAH) during 4 years study period. To our knowledge, this is the first report from Turkey of molecular SCCmec typing analysis of MRSA stains. A total of 385 clinical MRSA isolates collected in the clinical and Microbiology Laboratory at GMMAH between 2003 and 2006 were included in the study. Overall, SCCmec types-I, II, III, IV, V, nontypeable and PVL occurrence were detected in 11 (2.8%), 3 (0.8%), 316 (82.1%), 20 (5.1%), 20 (5.1%), 15 (3.9%) and 5 (1.3%) isolates, respectively. A total of 330 (85.5%) were SCCmec-I/II/III and 40 (10.3%) were SCCmec IV/V. SCCmec-I/II/III isolates were recovered more from patients with serious infections in surgical departments especially those with intensive care units than the SCCmec-IV/V isolates (chi(2) = 13.560, P < 0.001). SCCmec-I/II/III MRSA strains were predominantly recovered from blood stream (53.0%, P = 0.014), while SCCmec-IV/V strains were predominately isolated from skin and soft tissue and abscess (55.0%, P < 0.001). The PVL gene was detected in 10.0% of SCCmec-IV/V isolates in contrast to 0.3% in SCCmec-I/II/III (chi(2) = 25.164, P < 0.001). SCCmec-I/II/III MRSA strains were more resistant to clindamycin (chi(2) = 5.078, P = 0.024), amoxicillin-clavulanate (chi(2) = 84.912, P < 0.001), erythromycin (chi(2) = 4.651, P = 0.031), gentamicin (chi(2) = 24.869, P < 0.001), and rifampin (chi(2) = 18.878, P < 0.001) than SCCmec-IV/V MRSA strains. This data indicates that SCCmec-III MRSA strains that do not carry the PVL gene are the predominant MRSA strains in our hospital setting in Ankara, capital of Turkey and that SCCmec-I/II/III MRSA strains may cause serious infections in surgical departments especially those with intensive care units.

Molecular characterization of methicillin-resistant Staphylococcus aureus nasal isolates from Turkey.

Methicillin-resistant Staphylococcus aureus (MRSA) colonize most frequently in the anterior nares of the nose and cause serious infections all over the world. The aim of this study was to determine the nasal carriage rate of S. aureus and MRSA strains in Turkish elementary school children. We also analyzed molecular characterizations of MRSA strains by using pulse field gel electrophoresis (PFGE), multi locus sequence typing (MLST), staphylococcal chromosomal cassette mec (SCCmec) typing, and detection of the Panton-valentine leucocidin (PVL) gene. The nasal swabs were obtained from 4,050 children during a 4 month period in Ankara. In vitro antimicrobial susceptibility testing to 1 mug oxacillin and 30 mug cefoxitin was determined by a disk diffusion method. We found that the 1,001 of 4,050 (24.7%) children were colonized with S. aureus. Three S. aureus strains were resistant to oxacillin and cefoxitin. The rate of MRSA among all children was 0.07%. The MRSA strains revealed three different PFGE pattern. All MRSA isolates by harbored the SCCmec type IV element, but not the PVL gene. The two MRSA isolate belonged to sequence type (ST) 30, whereas the other one was a unique type. The results of this study demonstrated that S. aureus nasal carriage rate was consistent with previous studies. However, MRSA carriage rate was low. This study also indicated that the ST30-type IV without PVL gene MRSA clone may be expected to spread in Turkish community.