[Evaluation of the Respiratory Viral Panel PCR Test Results Before and After COVID-19 Pandemic]. / COVID-19 Pandemisi Öncesi ve Sonrasinda Solunum Yolu Virüs Paneli PCR Test Sonuçlarinin Degerlendirilmesi.

During the ongoing Coronavirus disease-2019 (COVID-19) pandemic, infections caused by other respiratory viruses continue to be seen and constitute an important health problem. In this study, it was aimed to evaluate the frequencies of respiratory tract viruses detected by respiratory tract virus panel (Allplex Respiratory Panel, Seegene, South Korea) multiplex real-time PCR test in severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pre-pandemic period, and in the first and second year of the pandemic. The distribution of viral agents between these three periods was also investigated. In addition, it was planned to investigate the frequency of coinfection with SARS-CoV-2 and other respiratory tract viruses during the pandemic. When the sum of the three periods were evaluated together, it was observed that at least one respiratory tract virus was detected in 13 802 (32.7%) of 42 174 samples. While at least one respiratory tract virus was detected in 8740 (54.6%) of 16 002 samples in the pre-pandemic period, at least one respiratory tract virus was detected in 1638 (9.4%) of 17 510 samples in the first year of the pandemic, and in 3424 (39.5%) of 8662 samples in the second year of the pandemic. In the first year of the pandemic, a statistically significant difference was observed that the number of viruses detected decreased due to closure measures and the use of personal protective equipment (p<0.001). It was determined that the frequency of the detection of respiratory tract viruses other than SARS-CoV-2 started to increase again and a statistically significant difference occurred in the third period when vaccination started and the transition to normalization began by gradually loosening the closure measures (p<0.001). Rhinovirus was the most frequently detected virus in all three periods of the study (First period: 16.5%; second period: 5.9%; third period: 16.5). More than one respiratory tract virus was detected simultaneously in 2061 (14.9%) of 13 802 samples, in which at least one respiratory tract virus was detected within the scope of the study. Rhinovirus (7.3%) took the first place among the viruses found in coinfection. In the second and third periods covering the pandemic period, it was observed that the SARS-CoV-2 PCR result was also positive in 177 (4.2%) of 4219 samples whose respiratory tract virus panel PCR results were positive and simultaneously had a SARS-CoV-2 PCR test request. Therefore, it was concluded that SARS-CoV-2 coinfection can be observed in the same patient with other respiratory tract viruses in respiratory tract samples. The addition of SARS-CoV-2 to the respiratory tract virus multiplex PCR panels currently in use will enable faster detection of such coinfections. It is thought that both the incidence of respiratory tract virus infections other than SARS-CoV-2 and the rate of coinfection with other respiratory tract viruses in SARS-CoV-2 infection may increase with the relaxation of the measures taken for the control of the pandemic. For this reason, the detection of viruses that cause respiratory tract infections from clinical samples with reliable and rapid methods will ensure the measures to be taken to protect public health without delay and thus contribute to the prevention of the spread of infections.

[Existence of Biofilm and Biofilm-Associated Virulence Genes in Multi-Drug Resistant Invasive Acinetobacter baumannii Isolates]. / Çok Ilaca Dirençli Invaziv Acinetobacter baumannii Izolatlarinda Biyofilm ve Biyofilm Iliskili Virülans Genlerinin Varligi.

Acinetobacter baumannii is a multi-drug resistant (MDR) gram-negative pathogen leading to nosocomial infections. Hospital-acquired infections due to A.baumannii occur especially in patients hospitalized in intensive care units. Important infections related to this bacterium are pneumonia, bacteremia, endocarditis, skin and soft tissue, urinary tract infections and meningitis. Human transmission is usually through the hospital environment or through medical personnel. A.baumannii isolates increases their virulence not only being multiple resistance to antibiotics but as well as the ability to form biofilm. The biofilm formation of A.baumannii isolates were mostly related with genes encoding curli fiber (csgA), the chaperone-usher fimbria (csuE) and the outer membrane (ompA). The aim of this study was to demonstrate biofilm production and virulence genes in MDR invasive A.baumannii isolates. MDR and similarity status previously known invasive A.baumannii (n= 156) isolates were included in the study. Biofilm production was determined by quantitative microplate biofilm method. Virulence genes csgA, csuE, fimH, ompA and blaPER-1 were investigated by polymerase chain reaction (PCR). It was determined that 60.3% (94/156) of all the isolates formed biofilm. Of these 94 isolates, 17 were weak, 33 were medium and 44 were strong. The mean biomass forming capacity of the isolates was found to be 2.23 ± 0.0033. Among the isolates included in the study (n= 156) the frequency of csgA, csuE, ompA, fimH and blaPER-1 genes were 71.2%, 32.1%, 21.8%, 7.1% and 3.2% respectively. The frequency of csgA, ompA, bap, csuE, fimH virulence genes were found to be 41.5%, 24.5%, 20.2% and 5.3% among biofilm positive isolates respectively. Biofilm-forming isolates were most commonly found in pulsotype II 19.1% (18/94), pulsotype IX 17.0% (16/94) and pulsotype VI 12.8% (12/94). In this study, when the distribution of virulence genes were compAred with the isolates that have weak, medium and strong biofilm, all of the studied genes were found to be more abundant in isolates with strong and medium positive biofilm production. This has shown that excluding fimH gene, csgA, csuE and ompA genes have contributed to the biofilm formation in invasive A.baumannii isolates, respectively.

A prospective multicenter study on the evaluation of antimicrobial resistance and molecular epidemiology of multidrug-resistant Acinetobacter baumannii infections in intensive care units with clinical and environmental features.

BACKGROUND: Multidrug-resistant (MDR) Acinetobacter baumannii infections are considered as emerging nosocomial infections particularly in patients hospitalized in intensive care units (ICUs). Therefore, reliable detection of MDR strains is crucial for management of treatment but also for epidemiological data collections. The purpose of this study was to compare antimicrobial resistance and the clonal distribution of MDR clinical and environmental A. baumannii isolates obtained from the ICUs of 10 different hospitals from five geographical regions of Turkey in the context of the demographic and clinical characteristics of the patients. METHODS: A multicenter-prospective study was conducted in 10 medical centers of Turkey over a 6 month period. A total of 164 clinical and 12 environmental MDR A. baumannii isolates were included in the study. Antimicrobial susceptibility testing was performed for amikacin (AN), ampicillin-sulbactam (SAM), ceftazidime (CAZ), ciprofloxacin (CIP), imipenem (IMP) and colistin (COL) by microdilution method and by antibiotic gradient test for tigecycline (TIG). Pulsed-field gel electrophoresis (PFGE) was performed to determine the clonal relationship between the isolates. The detection of the resistance genes, blaOXA-23, blaOXA-24, blaOXA-51, blaOXA-58, blaIMP, blaNDM, blaKPC, blaOXA-48 and blaPER-1 was carried out using the PCR method. RESULTS: The mortality rate of the 164 patients was 58.5%. The risk factors for mortality included diabetes mellitus, liv1er failure, the use of chemotherapy and previous use of quinolones. Antimicrobial resistance rates for AN, SAM, CAZ, CIP, IMP, COL and TIG were 91.8%, 99.4%, 99.4%, 100%, 99.4%, 1.2% and 1.7% respectively. Colistin showed the highest susceptibility rate. Four isolates did not grow on the culture and were excluded from the analyses. Of 172 isolates, 166 (96.5%) carried blaOXA-23, 5 (2.9%) blaOXA-58 and one isolate (0.6%) was positive for both genes. The frequency of blaPER-1 was found to be 2.9%. None of the isolates had blaIMP, blaKPC, blaNDM and blaOXA-48 genes. PFGE analysis showed 88 pulsotypes. Fifteen isolates were clonally unrelated. One hundred fifty-seven (91.2%) of the isolates were involved in 14 different clusters. CONCLUSIONS: Colistin is still the most effective antibiotic for A. baumannii infections. The gene blaOXA-23 has become the most prevalent carbapenemase in Turkey. The distribution of invasive A. baumannii isolates from different regions of Turkey is not diverse so, infection control measures at medical centers should be revised to decrease the MDR A. baumannii infections across the country. The results of this study are expected to provide an important baseline to assess the future prophylactic and therapeutic options.

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.

[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.

Effect of biofilm formation, and biocorrosion on denture base fractures.

PURPOSE: The aim of this study was to investigate the destructive effects of biofilm formation and/or biocorrosive activity of 6 different oral microorganisms. MATERIALS AND METHODS: Three different heat polymerized acrylic resins (Ivocap Plus, Lucitone 550, QC 20) were used to prepare three different types of samples. Type "A" samples with "V" type notch was used to measure the fracture strength, "B" type to evaluate the surfaces with scanning electron microscopy and "C" type for quantitative biofilm assay. Development and calculation of biofilm covered surfaces on denture base materials were accomplished by SEM and quantitative biofilm assay. According to normality assumptions ANOVA or Kruskal-Wallis was selected for statistical analysis (α=0.05). RESULTS: Significant differences were obtained among the adhesion potential of 6 different microorganisms and there were significant differences among their adhesion onto 3 different denture base materials. Compared to the control groups after contamination with the microorganisms, the three point bending test values of denture base materials decreased significantly (P<.05); microorganisms diffused at least 52% of the denture base surface. The highest median quantitative biofilm value within all the denture base materials was obtained with P. aeruginosa on Lucitone 550. The type of denture base material did not alter the diffusion potential of the microorganisms significantly (P>.05). CONCLUSION: All the tested microorganisms had destructive effect over the structure and composition of the denture base materials.

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.

[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.

Erythromycin and penicillin resistance mechanisms among viridans group streptococci isolated from blood cultures of adult patients with underlying diseases.

The aim of the study was to evaluate the species distribution, antimicrobial susceptibility and erythromycin-penicillin resistance mechanisms of viridans streptococci (VGS) isolates from blood cultures of adult patients with underlying diseases. Fifty VGS blood culture isolates were screened for their antibiotic susceptibilities against penicillin G, erythromycin and tetracycline by E-test. Clindamycin, cefotaxime, chloramphenicol, levofloxacin, linezolid and vancomycin susceptibility were performed by broth microdilution method. Erythromycin and penicillin resistance genotypes, ermB and mefA/E, pbp1a, pbp2b and pbp2x are amplified using PCR method. The clinical isolates included Streptococcus mitis (n. 19), S.oralis (n. 13), S.sanguinis, S.parasanguinis (n. 6, each), S.salivarius, S.vestibularis (n. 2, each), S.constellatus, S.sobrinus (n. 1, each). The percentage resistance against erythromycin and penicillin was 36% and 30%, respectively. The genotypic carriage rate of erythromycin resistance genes were: 56% ermB, 28% mefE, 8% ermB+mefE. Penicillin-resistant isolates carried pbp2b (33.3%) and pbp2x (20%) genes. Twenty-four VGS isolates were recovered from patients with cancer. S.mitis and S.oralis predominated among patients with cancer who had erythromycin and penicillin resistance isolates. The importance of classical antimicrobial agents like penicillin and erythromycin warrants the continuous surveillance of invasive VGS isolates and can guide better treatment options especially in patients with underlying diseases.

[Classical and new approaches in laboratory diagnosis of viridans streptococci]. / Viridans streptokoklarin laboratuvar tanisinda klasik ve yeni yaklasimlar.

Viridans group streptococci (VGS) are gram-positive microorganisms that can form alpha-hemolytic colonies on sheep blood agar. They reside as normal flora in oral cavity, respiratory, gastrointestinal, urogenital tract and on skin. They can cause bacteremia, endocarditis, meningitis and septicemia following dental procedures. The diagnosis of VGS are difficult since the taxonomic classification and species na-mes may change due in time. Viridans group streptococci are classified into 5 groups (Sanguinis, Mitis, Mutans, Salivarius, Anginosus) according to biochemical reactions and 16S rRNA sequencing. Since Streptococcus pneumoniae is a member of the Mitis group, the other important species in this group deserves investigation. Genetic exchange between Streptococcus mitis, Streptococcus oralis and S.pneumoniae by transformation and lysis mechanisms occur continously as they share the same anatomical region. These mechanisms play role in exchanging capsular and antibiotic resistance genes between these species. The cultivation of VGS usually starts with the inoculation of various patient specimens into sheep blood agar and the detection of alpha-hemolytic colonies. Observation of gram-positive cocci microscopically, the detection of optochin-resistant and bile insoluble colonies with few exceptions are the further important steps in laboratory diagnosis. VGS are then identified at species level by using biochemical reactions, automated diagnostic systems and molecular methods. The last step in the laboratory diagnosis of VGS is antibiotic susceptibility testing which is of outmost importance as penicillin and erythromycin resistance are on rise. In this review article, classification of VGS, similarities between S.pneumoniae and Mitis group streptococci and the laboratory diagnosis of VGS have been discussed.

[Antimicrobial resistance and existence of metallo-beta-lactamase in Acinetobacter species isolated from adult patients]. / Eriskin hastalardan izole edilen Acinetobacter türlerinde antimikrobiyal direnç ve metallo-beta-laktamaz varligi.

Acinetobacter spp. are the frequent causes of nosocomial infections which are difficult to treat due to multidrug resistance. The aim of this study was to determine the antibiotic susceptibilities and the presence of metallo-beta-lactamases in Acinetobacter spp. isolated from patients admitted to Hacettepe University Adult Hospital. A total of 124 Acinetobacter spp. isolates were included in the study. Antibiotic susceptibilities against imipenem (IMP), meropenem (MER), ceftazidime (CAZ), ciprofloxacin (CIP) and aztreonam (AZT) were studied by microdilution susceptibility testing according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. Multidrug-resistant isolates (MDR) were further tested for susceptibility against colistin by microdilution, and against amikacin (AN), piperacillin-tazobactam (PIP-TAZ), cefepime (FEP), ceftriaxone (CRO), tetracycline (TET), trimetoprim-sulfomethoxazole (SXT) and mezlocillin (MEZ) by disk diffusion method according to CLSI guidelines. Each isolate was also tested for metallo-beta-lactamase (MBL) production by using IMP and EDTA combined disk diffusion test and molecular analysis for bla(IMP-1) and bla(VIM-2) genes was done by polymerase chain reaction (PCR). Among 124 non-duplicate isolates, 72 were identified as Acinetobacter baumannii and 52 as Acinetobacter lwoffii. Minimum inhibitor concentration 50 (MIC50) and minimum inhibitor concentration 90 (MIC90) values of the isolates were 32 and 128 microg/ml for IMP, 16 and 32 microg/ml for MER, 128 and 256 microg/ml for CIP, 64 and 256 microg/ml for CAZ, 128 and 256 microg/ml for AZT, respectively. Forty-three (34.7%) isolates were susceptible to IMP. Overall, 51 (41%) Acinetobacter spp. were found to be resistant to > or = 3 antibiotics belonging to different antimicrobial classes and defined as MDR. Colistin MIC50 and MIC90 values were 2 and 8 microg/ml, respectively and the rate of colistin resistance was 27.5% in MDR isolates. The resistance rates for AN, PIP-TAZ, FEP, CRO, TET, SXT and MEZ were 80.4%, 98%, 92.2%, 100%, 100%, 86.3% and 86.3%, respectively. Among 124 isolates, 64 (51.6%) yielded positive result by IMP-EDTA combined disk test, and all the isolates were negative in terms of the tested MBL genes by PCR. These data revealed high level resistance among the Acinetobacter population in our hospital. The high rate of carbapenem resistance and increasing colistin resistance among Acinetobacter isolates should be surveyed cautiously. The increasing incidence of multidrug resistant Acinetobacter spp. emphasizes the need for new and effective treatment options.

[Value of demonstration of pneumococcal surface antigen A and autolysin genes for the identification of Streptococcus pneumoniae clinical isolates]. / Streptococcus pneumoniae klinik izolatlarinin tanimlanmasinda pnömokokal yüzey antijen A ve otolizin genlerinin gösterilmesinin yeri.

Streptococcus pneumoniae is one of the most frequent causative agents of community acquired pneumoniae, meningitis, sinusitis, bronchitis and otitis media both in children and adults. Conventional laboratory methods may sometimes fail to identify S. pneumoniae. The aims of this study were i) to compare the conventional methods and molecular methods which detected pneumococcal surface antigen A (psaA) and autolysin (lytA) genes; ii) to determine the serotype distribution of S. pneumoniae isolated from the respiratory samples. Randomly chosen 62 S. pneumoniae strains isolated from respiratory samples of patients with clinically proven pneumococcal pneumonia (age range: 1-79 years) between years 2000-2006, were included in the study. Classical microbiological analysis for the isolates included Gram staining, optochin sensitivity test performed in 5% CO2 and ambient air and bile solubility test. Capsular serotyping was performed by using latex particles sensitized with mono-specific typing sera (Statens Serum Institut, Denmark). Quellung reaction (Statens Serum Institut, Denmark) was used for serotyping the isolates that gave equivocal results using latex agglutination. Pneumococcal surface antigen A and autolysin genes were detected by in-house polymerase chain reaction (PCR) using psaA1 (5'-CTT TCT GCA ATC ATT CTT G), psaA2 (5'-GCC TTC TTT ACC TTG TTC TGC), lytAF (5'-ACG CAA TCT AGC AGA TGA AGC) and lytAR (5'-TGT TTG GTT GGT TAT TCG TGC) primers. Twenty six different serotypes were detected in 62 S. pneumoniae isolates. The most prevalent capsule serotype was 14 (n= 6), followed by 19A (n= 5). Four isolates could not be typed by the available antisera. All the isolates were optochin sensitive with or without carbondioxide incubation and were bile soluble. All the isolates included in the study have harboured (100%) psaA and lytA genes. No difference was found between the classical and molecular methods for the identification of S. pneumoniae isolates. In conclusion, detection of psaA and/or lytA genes by molecular methods is of value especially in "nonserotypeable strains" when they are performed with conventional methods in clinically proven S. pneumoniae isolates.

In vitro activity of tigecycline as a therapeutic option against multidrug-resistant Acinetobacter spp.

The study was performed to detect the in vitro activity of tigecycline in multidrug-resistant Acinetobacter isolates from patients in Hacettepe University Adult Hospital, Turkey. The microorganisms were isolated from clinical specimens of patients with respiratory and bloodstream infections. Thirty (66.7%) of the 45 inpatients were in ICUs. In vitro activity of imipenem, meropenem, ceftazidime, ciprofloxacin and aztreonam in 124 Acinetobacter species isolated was evaluated by microdilution test. Overall, 51 (41%) Acinetobacter spp. were found to be resistant to > or = 3 antibiotics belonging to different antimicrobial classes and defined as multidrug-resistant (MDR). Among the MDR Acinetobacter spp. 32 (62.7%) were Acinetobacter baumannii and 19 (37.3%) Acinetobacter lwoffii. In vitro activity of tigecycline against MDR isolates were studied by E-test. Each MDR isolate was also tested for metallo-beta-lactamase (MBL) production using CLSI guidelines. Forty-five (88.2%) of the isolates were found to produce MBL. The MIC90s of all antimicrobial agents tested except tigecycline were > or = 64 microg/ml whereas the MIC50, and MIC90 of tigecycline were found 1 microg/ml and 1.5 microg/ml, respectively. ERIC-PCR results revealed that bloodstream and respiratory isolates had nine and six different patterns, respectively. In conclusion, tigecycline has been shown to have potent in vitro activity against MDR Acinetobacter spp. and might be of therapeutic value in the treatment of infections due to MDR Acinetobacter spp., including those harbouring MBLs. Further clinical trials are needed to confirm the efficacy of tigecycline in the management of MDR Acinetobacter infections.

Validation of urinary antigen test for Streptococcus pneumoniae in patients with pneumococcal pneumonia.

The study was undertaken to prospectively evaluate a Streptococcus pneumoniae urinary antigen test for diagnosis of pneumococcal pneumonia among patient and control groups between 2004 and 2006. Microbiological analysis for these patients included Gram staining for sputum, sputum and blood culture. Nonconcentrated urine samples were tested using an immunochromatographic assay, the NOW S.pneumoniae antigen test. The urinary antigen test was positive in 9 (15.3%) of 59 patients enrolled in the study and in 8 (73%) of 11 patients with pneumococcal pneumonia confirmed by conventional methods. The test revealed a sensitivity of 72.7% and a specificity of 97.6% with conventional microbiological criteria used as the reference standard. The positive predictive value was 88.9% and the negative predictive value was 93%. We concluded that the urinary antigen test can supplement conventional microbiological tests in the diagnosis of pneumococcal pneumonia.

Macrolide resistance mechanisms and in vitro susceptibility patterns of viridans group streptococci isolated from blood cultures.

OBJECTIVES: Our aim was to study the macrolide resistance mechanisms and antimicrobial susceptibilities of viridans group streptococci (VGS) isolated from blood cultures. METHODS: In vitro susceptibilities to nine antimicrobials were studied for 85 VGS isolated from blood cultures by agar dilution. Pheno- and genotyping of erythromycin-resistant isolates were studied by the double disc test and PCR. RESULTS: Resistance to erythromycin was found in 27% (n = 23) of the isolates. Erythromycin-resistant Streptococcus oralis (n = 13) predominated among the other erythromycin-resistant species isolated. The phenotypes among 23 erythromycin-resistant isolates were as follows: 12 constitutive macrolide-lincosamide-streptogramin (cMLS(B)) resistance phenotype and 11 macrolide (M) resistance phenotype. Of the cMLS(B) isolates 11 had erm(B) genes and 11 of the M phenotype isolates had mef(A) genes. Four of the cMLS(B) isolates had both erm(B) and mef(A) genes. None of the isolates had erm(TR) genes. Combined resistance to erythromycin with penicillin, clindamycin, chloramphenicol, tetracycline and quinupristin/dalfopristin was found in 100, 61, 74, 100 and 100% of the isolates, respectively. No resistance was found for vancomycin, linezolid and levofloxacin. CONCLUSIONS: The macrolide resistance mechanisms of our VGS isolates revealed that the cMLS(B) phenotype associated with erm(B) and the M phenotype associated with mef(A) genes are found with similar frequencies.

[Laboratory diagnosis and antibiotic susceptibilities of nontuberculous mycobacteria]. / Tüberküloz disi mikobakterilerin laboratuvar tanisi ve antibiyotik duyarliliklari.

Nontuberculous mycobacteria (NTM) are found widespread in nature. They can live in soil, water supplies, plants and various environmental conditions. Today there are nearly 100 known NTM species and more than 50 of them are associated with human diseases. NTM can cause pulmonary diseases, lymphadenitis, skin, soft tissue and skeletal infections, catheter related bloodstream infections and disseminated infections in patients with underlying diseases like AIDS, chronic obstructive pulmonary diseases, emphysema, bronchiectasis, cystic fibrosis and chronic alcoholism. The laboratory diagnosis of NTM has improved in parallel to the improvements in molecular biology. The new DNA sequencing and microarray technologies have been started to use in laboratories. Antibiotic susceptibility tests can be used both for taxonomic and clinical purposes. In this review article, the importance of laboratory diagnosis and antibiotic susceptibilities of NTM have been discussed.

Evaluation of restriction endonuclease analysis of BRO beta-lactamases in clinical and carrier isolates of Moraxella catarrhalis.

A rapid increase in the prevalance of beta-lactamase producing M. catarrhalis isolates has highlighted its pathogenic potential. In this study, we aimed to detect the BRO beta-lactamases of our clinical (n = 32) and carrier (n =32) strains of Moraxella catarrhalis and compare the relationship of the enzyme type in assesment of MIC results of the antibiotics tested. BRO beta-lactamases were differentiated by restriction endonuclease analysis. Antibiotic susceptibility was performed by the agar dilution method recommended by NCCLS (M7A5). The clinical isolates produced 96.9%, whereas the carrier strains produced 90.6% beta-lactamase positivity by the restriction enzyme analysis. BRO-1 was isolated as 90.6% (n =29) while the BRO-2 and non-beta-lactamase producers (NBLP) were isolated as 6.3% (n =2) and 3.1% (n =1) respectively among clinical isolates. The rate of BRO-1 in the carrier strains was 75.0% (n =24), BRO-2 was 15.6% (n =5) and NBLP was 9.4%, (n =3). The beta-lactamase production with nitrocefin test was 96.9% (31/32) in clinical isolates and 90.6% (29/32) in carrier strains. M. catarrhalis needs a continous monitoring of antibiotic susceptibility; in this era restriction endonuclease analysis could be useful to screen BRO beta-lactamase genes.

[Six years evaluation of Clostridium difficile associated diarrhea]. / Clostridium difficile'ye bagli ishal olgularinin 6 yillik degerlendirmesi.

This study was aimed to detect the presence of Clostridium difficile toxin in the stool samples of patients with antibiotic-associated diarrhea or pseudomembranous colitis, and to relate its presence with the clinical findings of the patients. Between January 1997-April 2003, a total of 726 stool samples were investigated for C. difficile toxin A and/or B by enzyme immunoassay. Of them, 68 (9.4%) were found positive for C. difficile toxin (62 were toxin A, 6 were toxin B). C. difficile associated diarrhea were found to be related mostly with the use of beta-lactam/beta-lactamase inhibitor combinations (32/68), followed by aminoglycosides (12/68), and cephalosporins (8/68). The ages of the patients were between 1-86 years old (mean: 43.3 years), and 36 (52.9%) of them had an underlying conditions. Chronic obstructive pulmonary disease and chronic renal failure were the underlying disease in 18, malignancy in 11, and others (diabetes, hepatitis, transplantation, multiple sclerosis) in 7 of the patients. In conclusion, toxin detection and knowledge of the risk factors are the beneficial guidelines for the diagnosis of C. difficile associated diarrhea in the routine setting.

The vitro efficacy of beta-lactam and beta-lactamase inhibitors against multidrug resistant clinical strains of Mycobacterium tuberculosis.

In vitro activity of beta-lactam antibiotics and their beta-lactamase inhibitor combinations were evaluated for their activity against clinical isolates of Mycobacterium tuberculosis and M. tuberculosis H37Ra. Agar dilution, the BACTEC 460 system and beta-lactamase activity tests were used. Results using the BACTEC 460 and enzyme activity tests showed the best beta-lactamase inhibitor for M. tuberculosis H37Ra to be clavulanic acid. Cefazolin-clavulanic acid gave the lowest minimal inhibitory concentration (MIC) values using dilution tests and M. tuberculosis H37Ra. beta-Lactam antibiotics were combined with clavulanic acid and tested for in vitro activity against 50 selected multidrug-resistant (MDR) and 50 susceptible clinical isolates. Seventy-four percent of the isolates were inhibited by cefazolin-clavulanic acid combination. These results suggested that an appropriate combination of beta-lactam and beta-lactamase inhibitors might be useful in the treatment of tuberculosis due to multidrug-resistant strains.