[Evaluation of In Vitro Efficacy of Ceftazidime-Avibactam, Meropenem, and Colistin Single and Binary Combinations Against Carbapenem Resistant Klebsiella pneumoniae Strains Isolated from Various Clinical Specimens]. / Seftazidim-Avibaktam, Meropenem ve Kolistinin Tek Basina ve Ikili Kombinasyonlarinin Çesitli Klinik Örneklerden Izole Edilen Karbapenem Dirençli Klebsiella pneumoniae Suslarina Karsi In Vitro Etkinliginin Arastirilmasi.

It has been reported that many antibiotics used today, including the carbapenem group, fail to treat Klebsiella pneumoniae infections effectively. Despite many studies in recent years, the definitive treatment for carbapenem-resistant Klebsiella pneumoniae (CRKP) infections is still uncertain. In this study, it was aimed to investigate in vitro activities of colistin (COL) and meropenem (MEM), which are frequently used in the treatment of CRKP infections, and ceftazidime-avibactam (CZA), which is recently used in our country, alone or in combination against different CRKP isolates having different carbapenem resistance mechanisms andto analyze whether the presence of colistin resistance, which is an important problem in CRKP strains, influences the drug interaction results. This study was carried out in 42 K.pneumoniae isolates, which were isolated from various clinical samples as an infectious agent in Süleyman Demirel University Faculty of Medicine, Department of Medical Microbiology, Bacteriology Laboratory and whose carbapenem resistance was confirmed by carbapenemase inactivation test. The carbapenemase genes of the isolates were determined by the polymerase chain reaction (PCR) method. Antimicrobial susceptibilities of CRKP strains to CZA, MEM, and COL were determined by the broth microdilution method and in vitro synergy activities of dual combinations of these drugs were evaluated by checkerboard and time-kill methods. Statistical evaluation of categorical data was performed using Fisher's exact test, and p-value of less than 0.05 was considered statistically significant in terms of difference between the groups. Of the 42 CRKP isolates 34 (81%) were only OXA-48 positive, 5 (11.9%) were OXA-48+NDM and 3 (7.1%) were OXA-48+KPC positive. In the checkerboard test, synergy was detected against 97.6% of the isolates both with CZA+MEM and CZA+COL combinations, whereas this rate was 50% with MEM+COL. In the time-kill test, synergy was detected with CZA+MEM and CZA+COL combinations in the OXA-48 positive isolate and OXA-48+KPC positive isolate, while synergy was detected with CZA+COL and MEM+COL combinations in the OXA-48+NDM positive isolate. There was no significant relationship between whether the isolates were resistant to colistin or not and the checkerboard test results of antibiotic combinations (pCZA+MEM= 0.33, pCZA+COL= 0.11, pMEM+COL= 0.61). Results of our study revealed that the most common carbapenemase type in CRKP isolates was OXA-48 in our hospital, and the combinations of CZA with MEM and COL had high potential for synergism against these isolates.

An Investigation into the Association Between Toxoplasma gondii Infection and Bipolar Disorder

Objective: Studies have implicated Toxoplasma gondii in the etiology of mental disorders because of its neurotropic nature and its ability to modulate neurotransmitter pathways. This study aims to investigate T. gondii seroprevalence in patients with bipolar disorder and in healthy controls living in the Isparta Region of Turkey and to assess the probable relationship between T. gondii and bipolar disorder. Methods: Fourty-eight patients with bipolar disorder and 50 healthy controls were included in the study. Sociodemographic data, possible risk factors for T. gondii infection and clinical characteristics were analyzed. Serum anti-T. gondii IgM and IgG antibody levels were measured by using chemiluminescence immunoassay method (Roche Cobas e601 analyzer, Roche Diagnostics, Mannheim, Germany). Results: Anti-T. gondii IgG seropositivity rates were determined as 18.8% and 20% in the patient group and the control group, respectively. No statistically significant relationship was observed between T. gondii IgG seropositivity and bipolar disorder (p=0.876). In the study population, advanced age, low education level, living in a rural region and consumption of unwashed raw vegetable or fruit were found to be the significant risk factors for T. gondii infection (p<0.05). Conclusion: Our preliminary findings do not support the hypothesis that T. gondii infection is related to bipolar disorder. However, further studies would require larger sample sizes to confirm our results.

Comparison of DNA damage and proliferative capacities in smear samples of HPV positive and negative patients by micronucleus counting and AgNOR staining.

Human papillomavirus (HPV) is believed to cause cervical cancer. Thousands of women develop cancer and other diseases caused by HPV each year. HPV 16 and 18 types are found in approximately 70% of cervical cancers. Micronuclei are small chromosomal fragments that are considered indicators of DNA damage. AgNOR positive dots are useful for assessing proliferation. We investigated the relation between HPV-DNA, micronuclei and AgNOR in smear samples. Three groups were defined: HPV negative, 16/18 positive and other high-risk groups (31, 33, 35, 39, 45, 51, 52, 56, 58, 66 and 68) (HR). After typing, micronuclei were identified by Papanicolaou staining and AgNOR regions were detected by silver staining. Serum reactive protein (CRP) also was measured. We found that the average age of HPV negative patients was significantly greater than for the HPV positive groups. We also found that CRP levels were significantly higher in the HPV 16/18 positive group than HPV negative and other HPV group. We found that the number of micronuclei in the HPV 16/18 group was significantly greater than for the HPV negative group. Also, we found that AgNOR staining for the HPV 16/18 group was significantly greater than for the HPV negative group. We found that CRP level, cell proliferation and genome instability were increased in HPV positive patients. The AgNOR and micronucleus tests were useful for evaluating cell proliferation and DNA damage.

Evaluation of biochemical, hematological, RIBA and PCR assays in predicting viremia in anti-HCV positive patients.

INTRODUCTION: The detection of HCV-RNA by PCR assays is considered to be the gold standard for confirming the presence of HCV viremia. However, high costs, long and laborious procedures limit their widespread usage. This retrospective study was conducted to assess the predictive performances of biochemical and hematological parameters, anti-HCV signal-to-cutoff (S/CO) ratios and RIBA assay for HCV viremia. METHODOLOGY: Medical records of 210 patients with positive anti-HCV results were analyzed. Samples were tested for anti-HCV by the Roche Elecsys assay. RIBA and PCR assays were performed with Inno-Lia HCV Score test, and Roche Cobas TaqMan HCV test, respectively. RESULTS: Anti-HCV positive patients were categorized into two groups: positive HCV-RNA(viremic) group (n = 94) and negative HCV-RNA(non-viremic) group (n = 116). All viremic patients had positive RIBA results, while in the non-viremic group, 80 (69%) patients had negative/indeterminate RIBA results and 36 (31%) patients had positive RIBA results. Compared with the non-viremic group, the viremic group had significantly higher alanine aminotransaminase (ALT), aspartate aminotransferase, gamma-glutamyl transferase, mean platelet volume, platelet distribution width and anti-HCV levels, and significantly lower platelet count and plateletcrit levels (p < 0.05). With multivariate logistic regression analysis, serum ALT and anti-HCV levels were found to be strong predictive factors for HCV viremia. A S/CO ratio of ≥ 12.34 was identified as the optimal anti-HCV level to predict viremia. CONCLUSIONS: An anti-HCV S/CO ratio of 12.34 can determine the necessity for PCR assay, when carefully evaluated together with the biochemical and hematological evidence. This approach may reduce the cost of diagnosis particularly in low-resource settings.

Investigation of Carbapenem resistance mechanisms in Klebsiella pneumoniae by using phenotypic tests and a molecular assay.

INTRODUCTION: The aim of this study was to investigate the presence of carbapenemase production and carbapenem resistance mechanisms in 47 carbapenem resistant Klebsiella pneumoniae isolates by phenotypic confirmatory tests and molecular assay. METHODOLOGY: Carbapenem resistance genes KPC, OXA-48 and NDM were investigated with the BD MAX CRE assay kit in the BD MAX real time PCR instrument. Modified Hodge test, MBL gradient strip test, D70C Carbapenemase Detection Set, Temocillin gradient strip test methods were used as phenotypic confirmatory tests. Clonal relationship between study isolates was investigated with pulsed-field gel electrophoresis. RESULTS: Analysis with BD MAX CRE assay revealed OXA-48 positivity in 17 (36%) strains, NDM positivity in 6 (13%) strains and coexistence of OXA-48 + NDM positivity in 8 (17%) strains. In 16 (34%) strains, none of the KPC, OXA-48 and NDM genes were detected. While MHT was the most sensitive phenotypic confirmatory test, D70C disc set had not been considered as a useful tool to assist the search for carbapenemase production. Temocillin gradient test alone could not be considered as sufficient to detect the presence of OXA-48. PFGE analyses revealed that 23 of 31 carbapenemase producing strains were in three major PFGE genotypes (A, B and C). CONCLUSIONS: This study revealed that carbapenem resistance observed in K. pneumoniae isolates was mainly due to OXA-48 and NDM genes and the increase of carbapenem resistance among K. pneumoniae strains in our hospital was due to the interhospital spread of especially 3 epidemic clones.

Health care-associated infection surveillance in a tertiary neonatal intensive care unit: A prospective clinical study after moving to a new building.

BACKGROUND: There are very few prospective clinical studies on neonatal health care-associated infection (HAI) surveillance. HAI surveillance helps reduce not only mortality, but also morbidity, length of hospital stay, and health care costs. METHODS: This prospective clinical study covered a period of 12 months in a tertiary neonatal intensive care unit (NICU). HAI rates were calculated using different denominators: number of patients hospitalized in the NICU, number of patient-days, and number of specific device-days. RESULTS: The HAI rate was 18%, and the incidence density was 17/1,000 patient-days. The most common HAI was bloodstream infection (n = 34; 50%). The most common pathogen was coagulase-negative staphylococci (CoNS; 54.9%) in gram-positive bacteria and in general. Methicillin resistance was 96.4% for CoNS. Klebsiella spp (13.7%) was the most common gram-negative bacteria. Extended-spectrum ß-lactamase positivity was 14.3% for Klebsiella spp and 25% for Escherichia coli. HAI-related mortality was 0.3%. CONCLUSIONS: NICUs should perform their own HAI surveillance with prospective clinical design. Attention paid to handwashing, disinfection and sanitizing, complying with the terms of asepsis, extending in-service training, increasing the number of medical staff, preventing frequent changes in health care staff positions, and improving physical environmental conditions in NICUs might eventually decrease HAI rates.

Detection of the frequency of PER-1 type extended-spectrum ß-lactamase-producing Acinetobacter baumannii clinical isolates in Turkey: a multicenter study.

BACKGROUND/AIM: ß-Lactamases are an important resistance mechanism in Acinetobacter baumannii. Pseudomonas extended-resistance (PER-1) type ß-lactamase-producing strains have been reported from various geographic locations; however, PER-1 type ß-lactamases from Turkish hospitals have not been investigated extensively. The aim of this study was to determine the prevalence of PER-1 type ß-lactamases in A. baumannii isolates in various regions of Turkey. MATERIALS AND METHODS: A total of 763 clinical A. baumannii isolates were collected from 9 university hospitals and 2 state hospitals between 2008 and 2011. Molecular amplification of the OXA-51 gene from the A. baumannii genome was performed in order to verify identification of the species. Real-time polymerase chain reaction was used to detect blaPER-1 genes. RESULTS: PER-1 was detected in 24.6% of the isolates. The annual frequencies of the PER-1 enzyme were detected as 52.2%, 35.9%, and 8.3% in 2008, 2009, and 2010, respectively. PER-1 prevalence decreased gradually over time. The differences observed in PER-1 prevalence among the regions of Turkey were statistically significant (chi-square test; P < 0.001). CONCLUSION: These data demonstrate that the frequency of detection of PER-1 type ß-lactamases in A. baumannii species has decreased in Turkey. However, the increased carbapenem resistance, together with multidrug resistance, has created a worrisome situation regarding this pathogen.

[Distribution of blaOXA genes in Acinetobacter baumannii strains: a multicenter study]. / Acinetobacter baumannii Izolatlarinda blaOXA Genlerinin Dagilimi: Çok Merkezli Bir Çalisma.

Acinetobacter baumannii is the most important agent of nosocomial infections within the Acinetobacter genus. This gram-negative coccobacillus is intrinsically resistant to many antibiotics used in antimicrobial therapy, and capable of developing resistance including carbapenems. The objective of this study was to develop a multiplex real time polymerase chain reaction (qPCR) kit for OXA subgroups in A.baumannii, and to investigate the distribution of OXA subgroups in A.baumannii strains isolated from geographically different regions of Turkey. A total of 834 A.baumannii clinical isolates collected from different state and university medical centers in 13 provinces (Afyonkarahisar, Ankara, Bolu, Elazig, Erzurum, Isparta, Istanbul, Kahramanmaras, Konya, Sakarya, Van) between 2008-2011, were included in the study. The isolates were identified by conventional methods and automated systems [Vitek2 (bioMerieux, ABD) and Phoenix (BD Diagnostic, MD)]. The susceptibility profiles of the isolates were studied with automated systems and standard disc diffusion method. All samples were subjected to qPCR to detect blaOXA-51-like, blaOXA-23-like and blaOXA-58-like genes. A conventional PCR method was also used to detect blaOXA-24-like gene. The resistance rates observed during the study period were as follows: 96.8% for amoxicillin-clavulanate, 86.8% for ciprofloxacin, 74.7% for gentamicin, 71.7% for amikacin, 73.5% for cefaperozone-sulbactam, 72.1% for imipenem and 73% for meropenem. Six hundred and two (72.2 %) isolates were resistant to both imipenem and meropenem. Colistin was found to be the most effective antibiotic against A.baumannii isolates with 100% susceptibility rate. All isolates were positive for blaOXA-51-like, however blaOXA-24-like gene could not be demonstrated in any isolate. Total positivity rates of blaOXA-23-like and blaOXA-58-like genes were found as 53.7% and 12.5%, respectively, while these rates were 74.4% and 17.3% in carbapenem-resistant isolates, respectively. Twenty-five isolates were positive for both blaOXA-23-like and blaOXA-58-like genes. All of the carbapenem-resistant isolates have OXA type genes with the exception of blaOXA-24-like gene. The positivity rates for blaOXA-23-like and blaOXA-58-like genes varied for each center. In addition, there was a decrease in the frequency of blaOXA-58-like gene, however both blaOXA-23-like gene and carbapenem resistance rates increased during the study period. In conclusion, high rates of resistance to carbapenems were also remarkable but A.baumannii strains keep on sensitivity to colistin. Both blaOXA-23-like and blaOXA-58-like genes were shown to be widespread in carbapenem-resistant A.baumannii clinical isolates. However, blaOXA-23-like gene positive strains were increased throughout the study. Currently, multiplex qPCR is the best way for rapid diagnosis of resistant bacteria for prevention of hospital-acquired infections. The multiplex qPCR kit developed in this study could be useful for rapid diagnosis and identify the frequencies of blaOXA-23-like, blaOXA-51-like and blaOXA-58-like genes in carbapenem-resistant A.baumannii clinical isolates.

[Do different interpretative methods used for evaluation of checkerboard synergy test affect the results?]. / Dama Tahtasi Sinerji Testi Sonuçlarinin Farkli Yöntemlerle Yorumlanmasi Sonuçlarimizi Etkiliyor mu?

In recent years, owing to the presence of multi-drug resistant nosocomial bacteria, combination therapies are more frequently applied. Thus there is more need to investigate the in vitro activity of drug combinations against multi-drug resistant bacteria. Checkerboard synergy testing is among the most widely used standard technique to determine the activity of antibiotic combinations. It is based on microdilution susceptibility testing of antibiotic combinations. Although this test has a standardised procedure, there are many different methods for interpreting the results. In many previous studies carried out with multi-drug resistant bacteria, different rates of synergy have been reported with various antibiotic combinations using checkerboard technique. These differences might be attributed to the different features of the strains. However, different synergy rates detected by checkerboard method have also been reported in other studies using the same drug combinations and same types of bacteria. It was thought that these differences in synergy rates might be due to the different methods of interpretation of synergy test results. In recent years, multi-drug resistant Acinetobacter baumannii has been the most commonly encountered nosocomial pathogen especially in intensive-care units. For this reason, multidrug resistant A.baumannii has been the subject of a considerable amount of research about antimicrobial combinations. In the present study, the in vitro activities of frequently preferred combinations in A.baumannii infections like imipenem plus ampicillin/sulbactam, and meropenem plus ampicillin/sulbactam were tested by checkerboard synergy method against 34 multi-drug resistant A.baumannii isolates. Minimum inhibitory concentration (MIC) values for imipenem, meropenem and ampicillin/sulbactam were determined by the broth microdilution method. Subsequently the activity of two different combinations were tested in the dilution range of 4 x MIC and 0.03 x MIC in 96-well checkerboard plates. The results were obtained separately using the four different interpretation methods frequently preferred by researchers. Thus, it was aimed to detect to what extent the rates of synergistic, indifferent and antagonistic interactions were affected by different interpretation methods. The differences between the interpretation methods were tested by chi-square analysis for each combination used. Statistically significant differences were detected between the four different interpretation methods for the determination of synergistic and indifferent interactions (p< 0.0001). Highest rates of synergy were observed with both combinations by the method that used the lowest fractional inhibitory concentration index of all the non-turbid wells along the turbidity/non-turbidity interface. There was no statistically significant difference between the four methods for the detection of antagonism (p> 0.05). In conclusion although there is a standard procedure for checkerboard synergy testing it fails to exhibit standard results owing to different methods of interpretation of the results. Thus, there is a need to standardise the interpretation method for checkerboard synergy testing. To determine the most appropriate method of interpretation further studies investigating the clinical benefits of synergic combinations and additionally comparing the consistency of the results obtained from the other standard combination tests like time-kill studies, are required.

Investigation of Panton-Valentine leukocidin expressing Staphylococcus aureus colonization among children in a child care center.

The presence of Panton-Valentine leukocidin expressing Staphylococcus aureus colonization was investigated with a qualitative nucleic acid hybridization assay among 122 children and 19 staff in a child care center. Genotyping of 5 Panton-Valentine leukocidin-positive isolates by pulsed-field gel electrophoresis revealed that one child and a teacher from the same class were colonized with the clonally related strains. This finding allowed us to suggest that close contact with colonized people is a risk factor for being colonized.

[Evaluation of eosinophil cationic protein levels with clinical symptoms and laboratory findings of patients with cystic echinococcosis]. / Kistik ekinokokkozisli hastalarda eozinofilik katyonik protein düzeylerinin klinik semptomlar ve laboratuvar bulgulariyla birlikte degerlendirilmesi.

Among proteins secreted from activated eosinophil granulocytes, eosinophil cationic protein (ECP) is the most useful tool for the follow-up of inflammatory diseases. Since ECP level reflects the eosinophil activation, it gives valuable information about disease activity. In this study, we aimed to investigate the possible relation between ECP levels and symptoms and laboratory findings of cystic echinococcosis (CE) and to evaluate the role of this protein in the diagnosis of CE. The study which was conducted at Clinical Microbiology Laboratory of Suleyman Demirel University Medical Faculty, Isparta, Turkey, included 58 patients with a pre-diagnosis of CE and 32 healthy individuals as control group. The diagnosis of CE was established serologically by modified enzyme-linked immunosorbent assay (ELISA) and indirect hemagglutination (IHA) test. The quantitative determination of ECP levels was done by fluoro-enzyme immunoassay (FEIA; Uni-CAP ECP, Pharmacia-Upjohn). The mean ECP level was 31.6 +/- 37 microg/ml in the patient group and 9.1 +/- 2.1 microg/ml in the control group, the difference being statistically significant (p = 0.001). Significant differences were also detected for erythrocyte sedimentation rate (ESR) (p = 0.001), total IgE level (p = 0.001), eosinophile count (p = 0.05) and CRP (p = 0.001) between the patient and the control groups. ECP was detected to be high in 35 (60%), IgE in 37 (63%), CRP in 29 (50%) and eosinophile count in 9 (15.5%) patients. While age, gender, ESR, IgE and CRP levels of patients with high ECP levels were not significantly different from levels of patients with normal ECP levels, significantly different eosinophil counts were detected among patients with high ECP values when compared to patients with normal ECP values. Furthermore, a correlation was detected between ECP levels and eosinophil rate, IgE and CRP levels of patients with CE (p = 0.01), while there was no correlation between ECP and ESR levels. Although high ECP level patients exhibited higher ALT and AST levels, no correlation was determined between liver enzyme levels and ECP levels (p > 0.05). The most common symtoms among CE patients were abdominal pain (41%), other gastrointestinal complaints (38%), shortness of breath (12%) and fever (10%). No statistically significant difference in terms of symptoms was detected between patients with high ECP levels and normal ECP levels. However, statistically significant difference was detected between ECP levels of patients with symptoms (except shortness of breath) and patients without symptoms (p < 0.05). In conclusion, ECP seems to be associated with the symptoms and signs of CE and it can be used as a valuable marker besides the other laboratory tests for the evaluation of patients with CE.

[Investigation of the presence of autoantibodies in patients with toxocariasis]. / Toksokariazisli hastalarda otoantikor varliginin arastirilmasi.

Immunopathologic reactions may occur during toxocariasis due to tissue invasion and destruction by the secretions of larvae containing various enzymes with broad spectrum. The aim of this study was to search for autoantibodies such as anti-nuclear (ANA), anti-mitochondrial (AMA), anti-smooth muscle (ASMA), anti-neutrophil cytoplasmic (ANCA), anti-myeloperoxidase (MPO) and liver-kidney microsomal type 1 (LKM-1) antibodies in patients with toxocariasis, in order to investigate the role of toxocariasis as a trigger factor for autoimmune reactions. Forty patients (22 were male; mean age: 35.6 +/- 10.7 years) diagnosed as toxocariasis by clinical findings (abdominal pain, allergic symptoms and/or eosinophilia, without detection of any other causative agents, and without liver dysfunction, diabetes mellitus, cardiac or renal failure, and autoimmune disease) and in-house ELISA positivity and 32 healthy controls (16 were male; mean age: 40.7 +/- 11.2 years) were included to the study. ANA (screen), dsDNA, SS-A, SS-B, Scl-70, LKM-1, MPO and M2 autoantibodies have been investigated by ELISA (Euroimmun, Germany), while ANCA, AMA and ASMA antibodies by indirect immunofluorescence (IMMCO, NY) methods. Autoantibody positivity was detected in 18 (45%) patients of whom 11 yielded a single type, and 7 yielded > or = 2 types of autoantibodies. This rate was 12.5% for control group (two subjects were positive for ANA-Screen, one for anti-M2 and one for anti-LKM-1). The difference between the total positivity rates in patient and control groups was found statistically significant (chi2 = 5.72, p = 0.004). The most frequent autoantibody type among patients were ASMA (n = 6), followed by anti-dsDNA (n = 5), anti-M2 (n = 5), anti-SS-B (n = 4), anti-LKM-1 (n = 3), anti-SS-A (n = 2), ANCA (n = 2) and anti-MPO (n = 1). Positivity rate for ASMA was found statistically significant in patients' group compared to controls (chi2 = 12.24, p = 0.03), while there was no significant difference between the groups in terms of other autoantibody rates (p> 0.05). These data could be related to the possible release of autoantigens following muscle tissue injury during toxocariasis and/or antigenic mimicry of parasitic products during the infection in which muscle invasion is frequently seen. In conclusion, since autoantibodies are frequently detected in toxocariasis, this situation should be taken into consideration in the presence of autoantibodies.

[The relationship between viral load and malondialdehyde and antioxidant enzymes in patients with hepatitis C virus infection]. / Hepatit C virus enfeksiyonu olan hastalarda viral yük ile malondialdehid ve antioksidan enzimlerin iliskisi.

Oxidative stress is known to participate in the pathogenesis of HCV infection. The aim of this study was to investigate the relationship between antioxidant defence state, malondialdehyde (MDA) and viral load in patients with hepatitis C virus (HCV) infection. Fifty patients who were positive for serological and molecular markers of HCV infection, and 40 healthy volunteers as control group were included in the study. The patients were classified according to their viral loads, and the catalase, superoxide dismutase (SOD) and glutathione peroxidase (GP) activities of erythrocytes and MDA in sera of all groups were measured. These substances were detected by using the methods described by Aebi, Woolliams et al, Paglia and Valentine, Draper and Hadley, respectively. As a result, decrease in SOD and GP levels and increase in MDA and catalase levels have been detected in HCV infected patients when compared with healthy controls, and these differences were statistically significant (p<0.05, t=19.3),except for catalase. However, there were no statistically significant difference among groups classified according to viral load (p>0.05, t=1.6). Although our data in HCV infected patients demonstrate a significant decrease in antioxidant enzyme levels and a significant increase in MDA levels, a marker of oxidative stress, it could not possible to make a correlation between these differences and the viral loads of patients.