[Multilocus sequence analysis, biofilm production, antibiotic susceptibility and synergy tests of Burkholderia species in patients with and without cystic fibrosis]. / Kistik fibrozisli ve kistik fibrozisi olmayan hasta grubunda Burkholderia türlerinin multilokus sekans analizi, biyofilm olusturma, antibiyotik duyarlilik ve sinerji testleri.

Burkholderia spp. emerged as important pathogens in the airways of immunocompromised humans, especially those with cystic fibrosis (CF). Failure of identification with conventional techniques, high intrinsic resistance to most antibiotics and biofilm formation can cause difficulties in the treatment of these infections. The aim of this study was to identify Burkholderia spp. strains isolated from CF and non-CF patients with with routine microbiological methods, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and multilocus sequence analysis (MLSA), to determine of the antibiotic susceptibility and synergies, and to evaluate biofilm formation of these isolates. A total of 38 Burkholderia spp. (25 CF, 13 non-CF) from 26 patients were identified by biochemical, phenotypical and matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) and sequence types were revealed by multilocus sequence analysis (MLSA). Sequence types of isolates were identified using the PubMLST database. Characteristics of biofilm formation of clinical isolates were evaluated by microplate method. Antibiotic susceptibilities of ceftazidime, meropenem, trimethoprim-sulfamethoxazole (TMP-SXT) and levofloxacin were determined by broth microdilution method according to CLSI (2017) guidelines. Synergy tests were performed by checkerboard method. Clinical isolates were identified as Burkholderia cenocepacia (n= 16), Burkholderia contaminans (n= 11), Burkholderia gladioli (n= 4), Burkholderia dolosa (n= 4), Burkholderia multivorans (n= 2) and Burkholderia seminalis (n= 1). Sequence types of these isolates were determined as ST19, ST72, ST102, ST180, ST482, ST602, ST629, ST740, ST839 and ST1392. The correct identification at the species-level with MALDI-TOF MS was 94-100% for all isolates except B.contaminans. Biofilm formation among the identified species in the study was determined as 53% (n= 20). There was no statistical difference when the biofilm production was evaluated separately among Burkholderia species and biofilm production rates between CF (56%, 14/25) and non-CF (46%, 6/13) Burkholderia isolates (p> 0.05). Overall rates of resistance to ceftazidime, meropenem, TMP-SXT, and levofloxacin of the isolates were 35%, 66%, 50% and 40%, respectively. The antibiotic resistance against Burkholderia spp., isolates obtained from CF patients were more susceptible to ceftazidime, but no significant difference was found for other antibiotics. Synergy was determined between meropenem and TMP-SXT in two isolates. Antagonism was detected in 15 isolates, 12 of them were between meropenem and ceftazidime, three of them were between ceftazidime and TMP-SXT. Numerous resistance mechanisms may lead to higher resistance in this bacteria, whereas the antagonism between meropenem and ceftazidime in this study might be attributed to the expression of beta-lactamases. In this study, the distinctness of sequence types between Burkholderia spp. isolated from CF and non-CF patient, provided a better understanding about the importance of biofilm formation for the infections with these bacteria and emphasized that the management of therapy should be driven by the antibiotic test results.

Efficacy of new leukocyte parameters versus serum C-reactive protein, procalcitonin, and interleukin-6 in the diagnosis of neonatal sepsis.

BACKGROUND: The aim of this study was to investigate and compare the efficacy of the new leukocyte parameters mean neutrophil and monocyte volume (MNV, MMV), conductivity (MNC, MMC), scattering (MNS, MMS) and volume distribution width (NDW, MDW) with serum C-reactive protein (CRP), procalcitonin (PC) and interleukin (IL)-6 in the diagnosis of neonatal sepsis. METHODS: A total of 227 newborns (132 boys, 95 girls) were analyzed. There were 116 infants in the sepsis group (proven sepsis, n = 40; clinical sepsis, n = 76) and 111 in the control group. Venous blood samples were collected from infants at the time of diagnosis and complete blood count, peripheral blood smear, blood cultures, CRP, PC, IL-6 and MNV, MMV, MNC, MMC, MNS, MMS, NDW, and MDW were analyzed. RESULTS: MNV, NDW, MMV and, MDW were higher in infants with sepsis than in controls (P < 0.05 for all). MNS was lower in the patients with sepsis (P = 0.002). There was no significant difference between the sepsis and control groups in terms of MNC, MMC and MMS. CONCLUSION: Although the predictive value of leukocyte parameters including neutrophil and monocyte volume, conductivity, scattering and volume distribution width in the diagnosis of neonatal sepsis was lower than that of CRP, PC and IL-6, some of these new parameters may be useful in the differential diagnosis of newborn sepsis, along with the other screening tools. In particular, MNV seems to be the most useful parameter with the highest specificity; also, the importance of PC in the diagnosis of early onset sepsis was confirmed.

[Antimicrobial susceptibilities of clinical Nocardia isolates identified by 16S rRNA gene sequence analysis]. / 16S rRNA gen dizi analizi ile tanimlanan klinik Nocardia izolatlarinin antimikrobiyal duyarliliklari.

Nocardia species are ubiquitous in the environment and responsible for various human infections such as pulmonary, cutaneous, central nervous system and disseminated nocardiosis. Since the clinical pictures and antimicrobial susceptibilities of Nocardia species exhibit variability, susceptibility testing is recommended for every Nocardia isolates. The aims of this study was to determine the antimicrobial susceptibilities of Nocardia clinical isolates and to compare the results of broth microdilution and disc diffusion susceptibility tests. A total of 45 clinical Nocardia isolates (isolated from 17 respiratory tract, 8 brain abscess, 7 pus, 3 skin, 3 conjunctiva, 2 blood, 2 tissue, 2 pleural fluid and 1 cerebrospinal fluid samples) were identified by using conventional methods and 16S rRNA gene sequence analysis. Susceptibility testing was performed for amikacin, ciprofloxacin, ceftriaxone, linezolid and trimethoprim-sulfamethoxazole (TMP-SMX) by broth microdilution method according to the Clinical and Laboratory Standards Institute (CLSI) criteria recommended in 2011 approved standard (M24-A2) and disk diffusion method used as an alternative comparative susceptibility testing method. Among the 45 Nocardia strains, N.cyriacigeorgica (n: 26, 57.8%) was the most common species, followed by N.farcinica (n: 12, 26.7%), N.otitiscaviarum (n: 4, 8.9%), N.asteroides (n: 1, 2.2%), N.neocaledoniensis (n: 1, 2.2%) and N.abscessus (n: 1, 2.2%). Amikacin and linezolid were the only two antimicrobials to which all isolates were susceptible for both broth microdilution and disk diffusion tests. In broth microdilution test, resistance rates to TMP-SMX, ceftriaxone and ciprofloxacin were found as 15.6%, 37.8% and 84.4% respectively, whereas in the disk diffusion test, the highest resistance rate was observed against ciprofloxacin (n: 33, 73.3%), followed by TMP-SMX (n: 22, 48.9%) and ceftriaxone (n: 15, 33.3%). In both of these tests, N.cyriacigeorgica was the species with the highest resistance to ciprofloxacin (n: 25, 96.2%). When the susceptibility test results were compared, amikacin (κ= 1), linezolid (κ= 1), and ceftriaxone (κ= 0.903) showed very good agreement, whereas ciprofloxacin showed good agreement (κ= 0.672). For TMP-SMX no agreement was found between the two test methods (κ= 0.092). In conclusion, due to the identification of different species with molecular methods and increased frequency of Nocardia infections in recent years, in vitro susceptibility testing for Nocardia species is important to guide the appropriate antimicrobial treatment.

[Accurate diagnosis of Pseudomonas luteola in routine microbiology laboratory: on the occasion of two isolates]. / Rutin mikrobiyoloji laboratuvarinda Pseudomonas luteola'nin dogru tanisi: Iki sus nedeniyle.

Pseudomonas luteola which was previously known as Chryseomonas luteola; is a gram-negative, non-fermentative, aerobic, motile, non-spore-forming bacillus. It is frequently found as a saprophyte in soil, water and other damp environments and is an opportunistic pathogen in patients with underlying medical disorders or with indwelling catheters. It has been reported as an uncommon cause of bacteremia, sepsis, septic arthritis, meningitis, endocarditis, and peritonitis. Thus, early and accurate identification of this rare species is important for the treatment and also to provide information about the epidemiology of P.luteola infections. This report was aimed to draw attention to the accurate identification of P.luteola in clinical samples, upon the isolation and identification in two cases in the medical microbiology laboratory of a university hospital. In February 2011, a 66-year-old man, with chronic obstructive pulmonary disease, coronary artery disease and aplastic anemia, was admitted to our hospital due to progressive dyspnea. A chest tube was inserted on the 20th day of admission by the reason of recurrent pleural effusion. Staphylococcus aureus and a non-fermentative gram-negative bacillus (NFGNB) with wrinkled, sticky yellow colonies were isolated from the pleural fluid sample obtained on the 9th day following the insertion of the chest tube. In February 2012, a 7-year-old male cystic fibrosis patient who had no signs and symptoms of acute pulmonary exacerbation was admitted to the hospital for a routine control. This patient had chronic colonization with Pseudomonas aeruginosa and S.aureus and his sputum sample obtained at this visit revealed isolation of P.aeruginosa, S.aureus, Aspergillus fumigatus and a wrinkled, sticky yellow NFGNB. Both of these NFGNB were identified as P.luteola by the Phoenix automated microbial identification system (BD Diagnostics, USA). To evaluate the microbiological characteristics of these two isolates, the strains were further analysed by VITEK MS (bioMerieux, France) and Microflex LT mass spectrometer (Bruker Daltonics, Germany). Both of the MALDI-TOF-MS systems identified the isolates as P.luteola and 16S rRNA gene sequencing (ABI PRISM 3100, Applied Biosystems, USA) also confirmed the identification. The strains had wrinkled, sticky yellow colonies which were oxidase-negative, catalase-positive and non-fermentative. The Gram stained smears of the colonies revealed clusters of gram-negative bacilli probably embedded into a biofilm matrix. Since there are no accepted standards for testing the antibiotic susceptibility of P.luteola strains, the standards determined by CLSI for "other non-Enterobacteriaceae" (non-fermentative bacteria excluding P.aeruginosa, Acinetobacter spp., Burkholderia cepacia, B.mallei, B.pseudomallei and Stenotrophomonas maltophilia) were used for the susceptibility testing. Gradient MIC method (E-Test, bioMerieux, France) revealed that the isolates were susceptible to gentamicin, piperacillin-tazobactam, ceftazidime, cefepime, meropenem, colistin and levofloxacin. Accurate and prompt identification of P.luteola which is identified as a rare pathogen in serious cases is of critical importance since it has been suggested that this organism is likely to become more frequent as a nosocomial pathogen since the interventional processes increase in current medical practice. This report supported that Phoenix automated phenotypic identification system (BD Diagnostics, USA) and the two MALDI-TOF-MS based systems (VITEK MS and Bruker Microflex LT mass spectrometer) were successfull in the accurate identification of P.luteola.

Antimicrobial Activity of Copper Alloys Against Invasive Multidrug-Resistant Nosocomial Pathogens.

The emergence and spread of antibiotic resistance demanded novel approaches for the prevention of nosocomial infections, and metallic copper surfaces have been suggested as an alternative for the control of multidrug-resistant (MDR) bacteria in surfaces in the hospital environment. This study aimed to evaluate the antimicrobial activity of copper material for invasive MDR nosocomial pathogens isolated over time, in comparison to stainless steel. Clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) (n:4), OXA-23 and OXA-58 positive, MDR Acinetobacter baumannii (n:6) and Pseudomonas aeruginosa (n:4) were evaluated. The antimicrobial activity of coupons containing 99 % copper and a brass alloy containing 63 % copper was assessed against stainless steel. All the materials demonstrated statistically significant differences within each other for the logarithmic reduction of microorganisms. Among the three materials, the highest reduction of microorganisms was seen in 99 % copper and the least in stainless steel. The result was statistically significant especially for 0, 2, and 4 h (P = 0.05). 99 % copper showed a bactericidal effect at less than 1 h for MRSA and at 2 h for P. aeruginosa. 63 % copper showed a bactericidal effect at 24 h for P. aeruginosa strains only. Stainless steel surfaces exhibited a bacteriostatic effect after 6 h for P. aeruginosa strains only. 99 % copper reduced the number of bacteria used significantly, produced a bactericidal effect and was more effective than 63 % copper. The use of metallic copper material could aid in reducing the concentration of bacteria, especially for invasive nosocomial pathogens on hard surfaces in the hospital environment.

Potassium abnormalities in current clinical practice: frequency, causes, severity and management.

OBJECTIVE: We aimed to investigate the prevalence and etiology of potassium abnormalities (hypokalemia and hyperkalemia) and management approaches for hospitalized patients. SUBJECTS AND METHODS: Over a 4-month period, all hospitalized patients at Hacettepe University Medical Faculty Hospitals who underwent at least one measurement of serum potassium during hospitalization were included. Data on serum levels of electrolytes, demographic characteristics, cause(s) of hospitalization, medications, etiology of potassium abnormality and treatment approaches were obtained from the hospital records. RESULTS: Of the 9,045 hospitalized patients, 1,265 (14.0%) had a serum potassium abnormality; 604 (6.7%) patients had hypokalemia and 661 (7.30%) had hyperkalemia. In the hypokalemic patients, the most important reasons were gastrointestinal losses in 555 (91.8%) patients and renal losses in 252 (41.7%) patients. The most frequent treatment strategies were correcting the underlying cause and replacing the potassium deficit. Of the 604 hypokalemic patients, 319 (52.8%) were normokalemic at hospital discharge. The most common reason for hyperkalemia was treatment with renin-angiotensin-aldosterone system blockers in 228 (34.4%) patients, followed by renal failure in 191 (28.8%). Two hundred and ninety-eight (45.0%) patients were followed without any specific treatment. Of the 661 hyperkalemic patients, 324 (49.0%) were normokalemic at hospital discharge. CONCLUSION: This study showed a high prevalence of potassium imbalance among hospitalized patients. Although most of the potassium abnormalities were mild/moderate, approximately half of the patients treated for hypokalemia or hyperkalemia were discharged from the hospital with ongoing dyskalemia.

Evaluation of multiplex real time polymerase chain reaction and procalcitonin in the diagnosis of sepsis.

BACKGROUND: Sepsis is one of the most serious and life-threatening clinical conditions of childhood. This study has been designed to evaluate how useful multiplex real-time polymerase chain reaction (PCR) is in the early diagnosis of responsible microorganisms of sepsis and to specify how serial procalcitonin level measurement is helpful to support diagnosis of sepsis. METHODS: A total of 79 blood samples from 69 consecutive patients were collected for this prospective study between 01 Sept 2009 - 29 Feb 2012. RESULTS: In the evaluation of patients who are diagnosed with sepsis out of 69 patients with 79 clinical sepsis, 24 (30.37%) had positive blood culture and 19 (24%) samples were positive for PCR. When blood culture and multiplex real-time PCR results were compared, multiplex real-time PCR had a sensitivity of 75% and specificity of 100%. When the 2 microorganisms that multiplex real-time PCR cannot detect are excluded sensitivity increased to 81.8% and specificity did not differ. Procalcitonin levels on the day sepsis is suspected had a mean level of 13.91 ng/mL (+/- 49.26), on the 1st day (after 24 hours) the mean level of procalcitonin was 15.05 ng/mL (+/- 43.95), on the 2nd day (after 48 hours) it was 14.89 ng/mL (+/- 41.57). Mean procalcitonin levels of 50 children admitted with complaints other than infection and systemic inflammation was 0.06 ng/mL (+/- 0.04). CONCLUSIONS: In conclusion, multiplex real-time PCR test would be useful in the early diagnosis of sepsis. Studying procalcitonin levels is helpful in the early diagnosis of sepsis but does not have any correlation with the isolation of microorganisms in blood culture and survival.

[Carbapenem resistance in ESBL positive Enterobacteriaceae isolates causing invasive infections]. / Invazif enfeksiyonlara neden olan GSBL pozitif Enterobacteriaceae izolatlarinda karbapenem direnci.

The aim of this study was to investigate the presence of carbapenem resistance in Enterobacteriaeceae isolates recovered from invasive infections, in Hacettepe University Hospital, Ankara, Turkey, between 2005-2009, by phenotypic and genotypic methods. A total of 210 non-duplicated Escherichia coli (n= 153), Klebsiella pneumoniae (n= 47) and Klebsiella oxytoca (n= 10) isolates which were all determined to be extended-spectrum beta-lactamase (ESBL) positive with the BD Phoenix automated identification and antibiotic susceptibility system (Sparks, USA), were included in the study. The isolates were recovered from patients with bloodstream infections. Susceptibility of the isolates to imipenem, meropenem and ertapenem was detected with microdilution method according to the standards of Clinical and Laboratory Standards Institute (CLSI) minimal inhibitory concentration (MIC) breakpoints. Doripenem susceptibility was detected by the E-test (bioMerieux, Hazelwood, USA). All isolates which were found to be non-susceptible to any of the carbapenem antibiotics tested, were characterized by the phenotypic confirmatory tests and the presence of the resistance genes; blaAmpC, blaCTX-M, blaKPC, blaNDM, blaOXA, blaIMP ve blaVIM were screened by polymerase chain reaction (PCR). Among the 210 ESBL-producing Enterobacteriaceae blood isolates, 23 (11%) were identified as non-susceptible to any of the carbapenems tested. Resistance rates for imipenem, meropenem and ertapenem were 5.7% (n= 12), 1.9% (n= 4) and 2.4% (n= 5), respectively. Doripenem was more active than the other carbapenems, with a resistance rate of 1.0%. Seven of 23 isolates were ESBL negative with cefotaxime/clavulanic acid (CTX/CLA) and ceftazidime/clavulanic acid (CAZ/CLA) combined disk diffusion test, however, six of them were ESBL positive with the addition of boronic acid (BA) to CTX/CLA. Among the three isolates positive for Modifiye Hodge test (MHT) and/or ertapenem-BA tests, blaOXA-48 was detected in one and blaAmpC in the other. Phenotypic pAmpC activity was present in three K.pneumoniae isolates of which one was positive for blaAmpC gene. One K.pneumoniae isolate resistant to all carbapenems with MICs > 256 µg/ml and positive for phenotypic meropenem-BA, MHT, imipenem-EDTA, ceftazidime-CAZ/CLA, cefoxitin-BA production, was found to inhabit blaOXA-48 gene. Five isolates were positive for blaOXA-1 and one for blaOXA-10. Two isolates were positive for blaCTX-M, however blaIMP, blaVIM and blaNDM-1 genes were not detected among the isolates. In conclusion, carbapenem non-susceptibility which was low among the Enterobacteriaceae strains isolated in our center, was mostly attributed to the presence of blaOXA type carbapenemases and no accumulation of blaKPC and blaNDM were detected.

[Epidemiology, clinical and microbiological characteristics of invasive streptococcal infections in Turkey, 2010-2011]. / Türkiye'de invazif streptokok enfeksiyonlarinin epidemiyolojisi, klinik ve mikrobiyolojik özellikleri, 2010-2011.

A one-year active surveillance study was conducted to investigate the epidemiological and microbiological characteristics of invasive group A streptococci (GAS) infections in Turkey and to provide data for the establishment of national preventive strategies related to invasive GAS infections. A total of 46 clinical microbiology laboratories from 12 different regions of Turkey (Istanbul; Eastern and Western Marmara; Eastern and Western Blacksea; Aegean; Mediterranean; Western, Central, Northeastern, Middle-eastern and Southeastern Anatolia) participated in the study. Accordingly, GAS strains isolated from sterile body sites (blood, cerebrospinal, synovial, pleural, peritoneal, pericardial fluids) in the study centers between June 2010-June 2011, were sent to Maltepe University Hospital Clinical Microbiology Laboratory for microbiological confirmation and further analysis. The isolates were identified by conventional methods, and for serotyping, opacity factor (OF) and T protein types were investigated. For genotyping GAS lysate preparation, emm gene amplification and sequencing were performed by using the protocols recommended by Centers for Disease Control and Prevention. A total of 65 invasive GAS strains were isolated in 15 of the participant centers, during the study period. The rate of invasive GAS isolation exhibited regional variation, with the highest rates in the Eastern Blacksea (Trabzon, n= 19), followed by Istanbul (n= 17) and Western Anatolia (Ankara, Konya, n= 14). Of the patients with invasive GAS infection 33 were female, 32 were male, with the age range of 0-89 years. GAS strains were most commonly isolated from soft tissue specimens (n= 18), followed by abscess material (n= 10), sterile body fluids (n= 8) and blood (n= 7) samples. Serotyping revealed that 55% (36/65) of the strains were OF positive, and the majority of T protein was polygroup T (n= 20), followed by U (n= 14), B (n= 5), X (n= 3) and Y (n= 2). T protein was not detected in 22 isolates. The strains were found to have 17 different emm types;emm1 (n= 13), emm4 (n= 6), emm6 (n= 6), emm12 (n= 6), emm24 (n= 4), emm14 (n= 3) and emm28 (n= 3). Nine of the strains could not be typed by sequencing. The correlation between emm typing and serotyping was detected as 58%. It was observed that 26-valent vaccines included 70.5% of the invasive GAS strains included in this study. Our study provided initial data concerning the epidemiological properties of invasive GAS infections and characterization of GAS strains in Turkey. The incidence of invasive GAS infections is low in our country. Although immunization programme by 26-valent GAS vaccine is not currently an urgent public health issue for our country, the results of this study indicated that emm types 4 and 24 should better be included in such a vaccine to be used in Turkey. Additionally, since epidemiological features of GAS infections and the microbiological characteristics of the strains can vary by time, for the diagnosis of invasive streptococcal infections and to take the necessary preventive measures, epidemiological studies should be conducted repeatedly.

The airway microbiota in siblings with primary ciliary dyskinesia: Related factors and correlation with clinical characteristics.

OBJECTIVES-AIM: We aimed to show the composition and structure of and explore affecting factors on airway microbiota in primary ciliary dyskinesia (PCD) patients using culture-independent techniques. METHOD: A cross-sectional observational study was performed. We recruited 14 PCD patients (seven pairs of siblings) and nine parents. Bacterial rDNA was extracted from sputum and nasal samples. Sputum samples were also inoculated on suitable bacteriological media. RESULTS: Thirty-three separate genera were detected in sputum samples of PCD patients, and 41 were in nasal samples of parents. The detected genera were dominated by phyla Proteobacteria in PCD patients and their parents. Culture-dependent analyses could not detect many of the bacterial species detected with culture-independent analyses. There were no significant differences in alpha diversity between the siblings' pairs, and siblings' samples did not cluster together nearly as strongly as nonsiblings' samples. Patients who had no new complaints and no bacterial growth with the culture-dependent method at the time of study and patients who had no Haemophilus influenzae growth in the previous year had a significantly greater diversity (p < .05). Microbiota communities tended to cluster together by age, pulmonary exacerbation status, the existence of at least one H. influenzae growth with culture-dependent analyses in the previous year, and forced expiratory volume in 1 sec z and FEF25-75 z-scores. CONCLUSION: The airway microbiota of patients with PCD have presented more diverse bacterial communities than had been indicated with culture-dependent methods. The study identifies relationships between bacterial airway microbiota composition and the clinical measures of patients. Sibling pairs have no more community similarities than nonsibling PCD patients. Our results may indicate that the patients' clinical characteristics, which determine the disease severity, might affect the PCD microbiome.

Molecular characterization of oxacillinases and genotyping of invasive Acinetobacter baumannii isolates using repetitive extragenic palindromic sequence-based polymerase chain reaction in Ankara between 2004 and 2010.

BACKGROUND: Multidrug-resistant Acinetobacter baumannii (MDRAB) is an increasing problem worldwide. We aimed to determine the antibiotic susceptibility, diversity of oxacillinases, and molecular types of MDRAB. METHODS: A total of 100 non-duplicate A. baumannii blood culture isolates were evaluated. Antimicrobial susceptibilities of the isolates were determined according to the standard Clinical and Laboratory Standards Institute (CLSI) broth microdilution method. Colistin, doripenem, and tigecycline susceptibilities were analyzed by E-test. The presence of bla(OXA-23-like), bla(OXA-24-like), bla(OXA-51-like), and bla(OXA-58-like) genes was investigated by multiplex polymerase chain reaction (PCR). Typing of A. baumannii isolates was performed using repetitive extragenic palindromic sequence-based PCR (rep-PCR; DiversiLab). RESULTS: Most isolates were susceptible to colistin (98% susceptible) and tigecycline (94% susceptible), whereas fewer isolates were susceptible to imipenem, meropenem, and doripenem (17%, 17%, and 18% susceptible, respectively). Carbapenem resistance was associated with the presence of bla(OXA-23-like) (31% of isolates) and bla(OXA-58-like) (23% of isolates) genes. The occurrence of isolates carrying bla(OXA-58-like) genes increased between y 2004 and 2009, but decreased in 2010. In contrast, isolates with bla(OXA-23-like) genes increased during the 2008-2010 period. Out of 100 isolates, 62 were categorized into 13 major rep-PCR patterns, with the highest prevalence in pattern 1 (10 isolates), followed by patterns 2 and 3 (9 isolates each). CONCLUSIONS: Carbapenem-resistant invasive A. baumannii isolates carrying the bla(OXA-23-like) gene became more prevalent and replaced isolates carrying the bla(OXA-58-like) carbapenemase gene through the 7 y. Rep-PCR genotyping of these strains confirmed that ongoing MDRAB resulted from a long-term persistence and mixture of several clusters.

[Evaluation of methicillin-resistant Staphylococcus aureus bacteremia and comparison of prognosis according to vancomycin MIC values: experience of the last ten years]. / Metisiline dirençli Staphylococcus aureus bakteriyemisinin incelenmesi ve vankomisin MIK degerlerine göre prognozun karsilastirilmasi: Son on yillik deneyim.

Hospital acquired methicillin-resistant Staphylococcus aureus (MRSA) infections are important health problems. Mortality and morbidity rates associated with MRSA infections are increasing with mortality rates being higher for MRSA bacteremia than the other clinical presentations of MRSA infections. Initiation of treatment immediately and use of appropriate antibacterial agents may lead to better clinical outcomes in MRSA bacteremia. The aims of this study were to evaluate the treatment and clinical outcomes of patients with MRSA bacteremia in a tertiary care hospital in Ankara, Turkey. Two hundred forty seven MRSA strains isolated from blood cultures at Hacettepe University Faculty of Medicine, Clinical Microbiology Laboratory between January 2000-December 2010, were evaluated retrospectively. Demographic characteristics, duration of bacteremia, types and duration of antibiotic treatment, presence of other pathogens and all other necessary information were collected from patients' registry. One hundred eighty four patients who had clinically significant bacteremia were analyzed. The mean age of the patients was 55 ± 17 years, of them 44.6% were female and 55.4% were male. The median length of hospital stay was 61 days. The median duration for the development of MRSA bacteremia from the time of admission was 23 days. Overall mortality rate was 54.9%, and mortality rate due to MRSA bacteremia was 19%. The rate of treatment success was 81%. There were 3 (1.6%) patients with vancomycin MIC value of 0.5 mg/L, 140 (76.1%) patients with 1 mg/L and 41 (22.3%) patients with 2 mg/L. The median duration from the time of MRSA bacteremia detection to the time of death was shorter in unsuccessfully treated group than successfully treated group (7 days vs. 30 days, p< 0.001). Thirty days mortality rate was higher in unsuccessfully treated group than successfully treated group (94.3% vs. 50.7%, p< 0.001). The median time interval between positive and negative cultures was 9.5 days. Number of patients with MRSA bacteremia had been decreasing for the last five years (36 patients in 2006, 18 in 2007, 16 in 2008, 12 in 2009 and one in 2010). In multivariate logistic regression analysis, it was shown that, intubation (OR: 5.086, 95% CI: 2.094-12.351; p< 0.001) and malignancy (OR: 2.789, 95% CI: 1.185-6.564; p= 0.019) were independent risk factors for mortality. In this study, it was shown that mortality rate was high in MRSA bacteremia and high MIC value was not an independent risk factor for mortality. It was also noted that when there was no clinical response to vancomycin, the therapy should be changed immediately. To decrease MRSA bacteremia rates in the hospital adherence to rules of infection control and prevention proved to be important factors.

[Detection of resistance mutations in chronic hepatitis B patients receiving antiviral therapy for over one year]. / Bir yildan uzun süreli antiviral ilaç kullanan kronik hepatit B hastalarinda direnç mutasyonlarinin saptanmasi

Primary mutations conferring hepatitis B virus (HBV) antiviral resistance and associated secondary/compensatory mutations were investigated in this study by DNA sequencing (SEQ) and two commercial Line Probe Assays (LiPAs) (Inno-Lipa HBV DRv2 and Inno-Lipa HBV DRv3, Innogenetics, Belgium). A total of 71 subjects under follow-up for chronic hepatitis B and receiving lamivudine (LVD) therapy for one year or longer at the Hacettepe University Faculty of Medicine, Department of Internal Medicine, Gastroenterology Unit were included in the study with informed consent. Male to female ratio and mean age was noted as 47/24 and 43 ± 15.8 (age range: 13-71) years, respectively. Twenty samples with low HBV DNA levels (mean: 204.6 IU/ml) could not be interpreted by SEQ due to insufficient amplification. All samples with a positive consensus PCR were further analysed via LiPAs, as directed by the manufacturer. Thus a total of 51 and 56 samples could be evaluated via SEQ and LiPA assays, respectively. In 13 of the 51 (25.5%) samples by SEQ and in 9 of 56 (16%) samples by LiPAs, primary and compensatory mutations associated with resistance were identified. Multiple mutations that comprise L180M + M204I in four and L180M + M204V in one sample and single mutations (M204I) in three samples were identified by SEQ. In one sample which had multiple mutations associated with LVD resistance single mutations (S202G, N236T) associated with entecavir resistance and in two other such samples mutations associated with adefovir resistance were detected by SEQ. Also, in three samples amino acid substitution at position rt215 (QÆS) as alone and in one sample with multiple mutations were observed via SEQ. In five of nine samples primary and compensatory multiple mutations (L180M + M204I in one sample, L80I + L180M + M204I in two samples, L80I/V + M204I in one sample) and primary single mutations associated with LVD resistance (M204I/V) in four samples were detected by Inno-Lipa HBV DRv2. Two different mutations (G202, ILFM184) were observed in two samples with multiple mutations associated LVD resistance via Inno-Lipa HBV DRv3. However, mutation at position rt184 was evaluated as a weak positive. Any mutation associated with adefovir resistance was not detected by LiPA. As a result, SEQ and LiPAs displayed comparable performances for the detection of HBV drug resistance mutations. We suggested that for the evaluation of discordant results, it should be better to test consecutive serum samples.

[Investigation of baseline antiviral resistance in treatment-naive chronic hepatitis B cases]. / Tedavi Naif Kronik Hepatit B Olgularinda Bazal Antiviral Direncin Arastirilmasi

Naturally-occurring mutations associated with resistance to nucleoside/nucleotide analogues (NA) can be detected in a group of treatment-naive individuals chronically infected with hepatitis B virus (HBV). Genotypic resistance testing prior to the initiation of NA therapy may facilitate the selection of optimal drug regime and help to prevent early emergence of clinical resistance. In this study, presence of resistance mutations in treatment-naive individuals with chronic hepatitis B (CHB) was investigated in Hacettepe University Hospital, a referral center in Ankara province, Turkey. A total of 42 patients (17 female, 25 male; age range: 18-62 years) diagnosed as CHB were enrolled in the study with informed consent. All of the patients were negative for hepatitis C and D viruses and human immunodeficiency virus coinfections, and none had a history of interferon or NA treatment. HBV viral load, HBV markers and hepatic enzymes in patients were determined via standardized commercial assays. For the detection of NA resistance mutations, a partial sequence of approximately 250 nucleotides, harboring the frequently-observed sites for NA resistance was amplified via nested PCR and characterized by direct sequencing of the amplicons. The sequences were handled and interpreted for the presence of mutations via various softwares and a web-based virtual phenotyping tool. Well-characterized sequences were obtained in 30 out of 42 samples (71.4%). All circulating HBV strains were observed as genotype D. Nucleotide variations were detected in 19 individuals (63.5%) that comprise silent mutations without amino acid substitution in 8 (26.6%), mutations with undetermined significance in 7 (23.3%) and mutations associated with NA resistance in 3 (10%) patients. Mutations conferring resistance to entecavir + lamivudine (S202G, M204V, L180M, T184N) were identified in one patient whereas L180P, A181Q and A194V substitutions associated with probable lamivudin + adefovir and tenofovir resistance, respectively, were detected in other patients. All patients with resistance mutations were HBsAg and HBeAg positive, anti-HBe negative and had viral loads exceeding 3 x 10(7) IU/ml. In two patients, the route for HBV transmission was vertical. Since no follow-up samples were available from individuals with resistance mutations, alterations in serological markers, viral load and mutation patterns could not be monitored. In conclusion, the presence of NA resistance mutations were revealed in treatment-naive CHB cases in a referral hospital in Turkey. The impact and cost-effectivity of detecting naturally-occurring resistance mutations for clinical follow-up prior to the antiviral therapy need to be elucidated by prospective studies.

Serotype distribution of Streptococcus pneumoniae and pneumococcal vaccine coverage in adults in Turkey between 2015 and 2018.

OBJECTIVE: To evaluate the serotype distribution and antibiotic resistance in pneumococcal infections in adults and to provide a perspective regarding serotype coverage of both current and future pneumococcal vaccines. PATIENTS AND METHODS: This passive surveillance study was conducted with the Streptococcus pneumoniae strains isolated from the specimens of patients with pneumonia (materials isolated from bronchoalveolar lavage), bacteraemia, meningitis, pleuritis and peritonitis between 2015 and 2018. Serogrouping and serotyping were performed by latex particle agglutination and by conventional Quellung reaction using commercial type-specific antisera, respectively. The strains were analysed for penicillin, cefotaxime, erythromycin and moxifloxacin susceptibilities by E-test. RESULTS: In the whole study group (410 samples from adults aged ≥18 years), the most frequent serotypes were 3 (14.1%), 19 F (12%) and 1 (9.3%). The vaccine coverage for PCV13, PCV15, PCV20 and PPV23 was 63.9%, 66.6%, 74.1% and 75.9%, respectively, in all isolates. Penicillin non-susceptibility in invasive pneumococcal disease (IPD) was 70.8% and 57.1% in the patients aged <65 and ≥65 years, respectively. About 21.1% and 4.3% of the patients with and without IPD had cefotaxime resistance. Non-susceptibility to erythromycin and moxifloxacin was 38.2% and 1.2%, respectively. CONCLUSIONS: The results revealed that novel PCV vaccines may provide improved coverage as compared with the currently available vaccine, PCV13. The significant antibiotic resistance rates imply the need to extend the serotype coverage of the vaccines. Continuing the surveillance in pneumococcal diseases is critical to explore the serotype distribution and incidence changes of IPD cases in the population and to inform policy makers to make necessary improvements in the national immunization programmes.Key messagesThis multicentre study demonstrated the most recent serotype distribution and antibiotic resistance in adult population in Turkey.Shifting from PCV13 to novel conjugated vaccines will significantly increase the coverage.Continuing the surveillance in pneumococcal diseases is critical to explore the serotype distribution changes and the incidence of cases with invasive pneumococcal disease in the population.

Effects of freezing on the bactericidal activity of human milk.

OBJECTIVES: Storage of human milk by freezing has been recommended for long-term storage. The present study analyzed the bactericidal activity of human milk on Escherichia coli and Pseudomonas aeruginosa and determined the changes in bactericidal activity following freezing at -20°C and -80°C for 1 month and 3 months. METHODS: Forty-eight milk samples were collected from 48 lactating mothers. Each sample was divided into 10 aliquots. Two of the samples were processed immediately and the others were stored at both -20°C and -80°C until analysis after 1 month and 3 months of freezing. RESULTS: All of the fresh milk samples showed bactericidal activity against E coli and P aeruginosa. Freezing at -20°C for 1 month did not cause statistically significant alteration in bactericidal activity (P > 0.017), whereas storage for 3 months lowered the degree of bactericidal activity significantly (P < 0.017) against E coli. Bactericidal activity was protected when the samples were stored at -80°C. There was no statistically significant difference in the bactericidal activity of human milk against E coli between freezing at -20°C and -80°C for 1 month (P > 0.017); however, when milk was stored for 3 months, -80°C was significantly more protective (P < 0.017). Freezing at -20°C and -80°C for 1 month and 3 months did not cause any significant change in bactericidal activity against P aeruginosa (P > 0.05). CONCLUSIONS: Storage by freezing at -80°C is more appropriate to keep bactericidal capacity of stored human milk >1 month if affordable and available, especially in intensive care settings.

Epidemiologic trends in HBV infections at a reference centre in Turkey: an 11-year retrospective analysis.

OBJECTIVE: To identify changes in hepatitis B epidemiology after the implementation of the nationwide vaccination program in Turkey, hepatitis B virus (HBV) and related tests performed over a period of 11 years (2000-2010) at a reference centre were retrospectively overviewed and statistically analysed for trends. RESULTS: Assay results for Hepatitis B surface antigen (HBsAg) and e antigen (HBeAg), Anti-HBs, Anti-HBe, Anti-HBc immunoglobulins and HBV DNA as well as aspartate aminotransferase (AST), alanine aminotransferase (ALT), gama-glutamyl transpeptidase (GGT) and alkaline phosphatase (AP) levels, obtained via standardized commercial assays were included in the analysis. Overall, a stable anti-HBs incidence (43.6%) and male predominance in infected individuals were noted. Total Anti-HBc was detected in 43.3% of the Anti-HBs reactive population, demonstrating that the immunity against HBV has still been acquired through virus exposure. An intermediate HBsAg seroprevalence of 6.0% was observed with a significant decrease from 12.3% to 5.0% from 2000 to 2010. Anti-HBe positive infections were more frequent than those with HBe antigenemia (77.1% vs. 18.5%) with a notable increase from 2000 to 2003. HBV DNA was detected in 23.6-25.6% with serological markers of viral replication and was more prevalent in HBeAg positive individuals in parallel with AST, ALT and GGT levels. Evidence for horizontal transfer as the major transmission route was revealed with a reduction of childhood HBV infections, attributable to the ongoing vaccination efforts.

Detection and genotyping of cervical HPV with simultaneous cervical cytology in Turkish women: a hospital-based study.

PURPOSE: To investigate the prevalence of cervical human papilloma virus (HPV) with genotyping and simultaneous cervical cytology among Turkish women in a university hospital. A review of literature was done as well to summarize the results of similar Turkish studies based on hospital data. METHODS: Women who were subjected to cervical HPV-DNA testing with simultaneous cervical Pap test were included. Seeplex HPV 18-plex Genotyping Test was used for HPV detection and typing. Liquid-based cytology was used for Pap test and Bethesda system was used for results. RESULTS: Study group included 890 patients with a mean age of 39.5 years. The prevalence of any HPV was 25.7% while high-risk HPV was positive in 23.0%. There were no significant differences in HPV prevalence between younger and older women. Among HPV-positive women, 89.5% had at least one type of high-risk HPV. Most common HPV was type 16 followed by type 31 and 51. Abnormal cervical cytology rate was 11.6%. Rate of HPV positivity was significantly higher in women with abnormal cervical cytology compared to women with normal cytology (54.4 vs. 22.0%). CONCLUSIONS: Cervical HPV infection is a serious and gradually growing problem for Turkish women according to hospital-based data. This may be associated with low age at marriage and more sensitive HPV detection methods.

[Investigation of SCCmec types and Panton-Valentine leukocidin in community-acquired and nosocomial Staphylococcus aureus strains: comparing skin and soft tissue infections to the other infections]. / Toplumdan Kazanilmis ve Nozokomiyal Staphylococcus aureus Suslarinda SCCmec Tiplerinin ve Panton-Valentine Lökosidin Varliginin Arastirilmasi: Deri ve Yumusak Doku Enfeksiyonlari ile Diger Enfeksiyonlarin Karsilastirilmasi

Infections due to methicillin-resistant Staphylococcus aureus (MRSA) are important health care problems since they are usually multidrug resistant. Although MRSA is isolated especially from nosocomial infections, community-acquired MRSA infections are increasing. Methicillin resistance is due to the expression of mecA gene, which is located on SCCmec gene cassette. Different SCCmec types can be detected in hospital-acquired and community-acquired (CA-) MRSA strains. CA-MRSA strains might harbour Panton-Valentine leukocidin (PVL), an important virulence factor in skin and soft tissue infections. Strains carrying PVL has the ability to penetrate undamaged skin and cause more severe infections. The aim of this study was to detect SCCmec types and PVL gene in S.aureus strains isolated from skin and soft tissue infections and to compare with strains isolated from other infections in a university hospital in Ankara, Turkey. S.aureus strains isolated from skin and soft tissue infections (n= 285) and a control group consisting of 161 strains isolated from other infections (53 blood, 48 lower respiratory tract samples, 30 sterile body fluids, 30 genitourinary tract samples) chosen by stratification and random selection method, were included in the study. Among skin and soft tissue infection strains 46.7% were from the hospitalized patients and 48.4% of skin and soft tissue infection strains were from female patients. The mean age of the skin and soft tissue infection patients was 45.5 years. Among the control strains 60.9% were from the hospitalized patients and 41.6% of the control patients were female. The mean age of the control patients was 50.2 years. Strains were identified by the Phoenix system (Becton Dickinson, USA) and identification was confirmed by tube coagulase test. Methicillin resistance was determined by the Phoenix system which determines both oxacillin and cefoxitin minimum inhibitor concentrations and, confirmed by oxacillin agar screening and/or cefoxitin disk diffusion test. All isolates were screened for the presence of mecA and PVL genes and SCCmec types were determined by PCR. MRSA constituted 20.3% (n= 58) of skin and soft tissue infection isolates and 24.2% (n= 39) of the control group. Of the 97 MRSA, 85 had a SCCmec type III-like pattern with an additional dcs region, three had type IV, three had type IIIa, one had type IIIb, one had type II and four could not be typed. The difference between SCCmec type distribution in skin and soft tissue infection and other infections' (control) groups was not statistically significant (p> 0.05). Two of the three SCCmec type IV strains were type IVa. Ten (2.2%) PVL positive strains, three of which were from the control group; were all methicillin susceptible S.aureus (MSSA). Although PVL positive MRSA was not common, detection of SCCmec type IVa, a marker for CAMRSA, and PVL positive MSSA strains which might act as a reservoir for PVL positive MRSA, indicated the importance of ongoing surveillance for MRSA.

[Comparison of culture and real-time PCR methods in the detection of Streptococcus pneumoniae and Haemophilus influenzae in acute otitis media effusion specimens]. / Akut Orta Kulak Iltihabi Örneklerinde Streptococcus pneumoniae ve Haemophilus influenzae'nin Saptanmasinda Kültür ve Gerçek Zamanli PCR Yöntemlerinin Karsilastirilmasi

Streptococcus pneumoniae and Haemophilus influenzae are the major etiologic agents of acute otitis media. This study was aimed to compare the detection rate of S.pneumoniae and H.influenzae by culture and real-time polymerase chain reaction (Rt-PCR) in the middle ear effusions of patients diagnosed as acute otitis media. A total of 60 middle ear effusion samples collected from children with acute otitis media were included in the study. The samples were inoculated and incubated in BACTEC Ped Plus blood culture bottles and BACTEC 9120 system (BD Diagnostic Systems, MD), respectively, and the isolates were identified by conventional methods. For the molecular diagnosis of H.influenzae and S.pneumoniae, ply pneumolysin gene and HIB capsule region, respectively were amplified by Rt-PCR (LightCycler, Roche Diagnostics, Germany). H.influenzae and S.pneumoniae were isolated from 5 (8.3%) and 3 (5%) of the patient samples with conventional culture methods, respectively. In addition in 11.6% of the samples other microorganisms (Staphylococcus epidermidis, Streptococcus intermedius, Streptococcus sanguinis, Moraxella catarrhalis, Pseudomonas aeruginosa, Candida albicans) were also isolated. On the other hand H.influenzae and S.pneumoniae were detected in 38 (63.3%) and 24 (40%) of the samples with Rt-PCR, respectively. There was about eight fold increase in the detection frequency of H.influenzae and S.pneumoniae with Rt-PCR compared to culture methods. When culture was accepted as the gold standard method, the sensitivity, specificity and positive predictive value of Rt-PCR in the detection of H.influenzae and S.pneumoniae were estimated as 80%, 51% and 98.2%, respectively. As a result, Rt-PCR was shown to be a sensitive method and could be preferred for the rapid diagnosis of H.influenzae and S.pneumoniae in the etiological diagnosis of acute otitis media, especially in culture negative cases.