Mikrobiyoloji bülteni (MIKROBIYOL BUL )

Publisher: Ankara Mikrobiyoloji Derneği


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    Mikrobiyoloji bülteni, Bulletin of microbiology
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    ABSTRACT: The aim of this study was to evaluate methicillin-resistant Staphylococcus aureus (MRSA) bacteremia cases who were followed at the Infectious Diseases Unit of Internal Medicine Department, at Hacettepe University Adult and Oncology Hospitals between January 2004-December 2011. A total of 198 patients, of them 99 had positive MRSA blood cultures (case group), and 99 without MRSA bacteremia (control group) who were selected randomly among patients at the same wards during the same time period, were included in the study. Demographic data, risk factors for MRSA bacteremia and antibiotic use of case (60 male, 39 female; mean age: 59.37 ± 16.96 yrs) and control (60 male, 39 female; mean age: 59.11 ± 17.60 yrs) groups were obtained from the patient files and the hospital data system and were compared. Methicillin susceptibility was determined by the cefoxitin (30 µg, BD, USA) disc diffusion method and confirmed by mecA PCR test. Antimicrobial susceptibilities were also determined by disc diffusion and Etest (BioMerieux, France) methods according to CLSI guidelines. There was no statistically significant difference between the two groups according to age, gender, presence of an underlying chronic disease, burn, hemodialysis, malignancy or immunosupression (p> 0.05). The results of the univariate analysis revealed that antibiotic use and parameters most likely to be associated with MRSA bacteremia (obesity, cerebrovascular event, hospitalization history, central/arterial catheter, presence of tracheostomy, invasive/non-invasive mechanical ventilation, use of proton pump inhibitors, H2 receptor blockers, sucralfate, nasogastric or urinary tubes, gastrostomia, total parenteral nutrition, acute organ failure and surgical operation) were found to be statistically higher in the case group (p< 0.05). Median length of hospital stay was also higher in the case group (59 days versus 8 days; p< 0.001). Multivariate regression analysis indicated that obesity (OR= 7.98; p= 0.013), central venous catheterization (OR= 6.65; p= 0.005), nasogastric tube (OR= 16.58; p< 0.001) and use of H2 receptor blockers (OR= 4.41; p= 0.010) were independent risk factors. The number of patient given at least one antibiotic (92 in case group, 51 in control group) was statistically higher than those who were not (48 in case group, 7 in control group) (OR= 14.86; p< 0.001). Use of antibiotics [ampicillin-sulbactam and/or amoxicillin-clavulanate, fluoroquinolones, aminoglycosides, piperacillin-tazobactam (TZP), meropenem (MEM), imipenem (IPM), vancomycin (VAN), cephalosporins and teicoplanin (TEC)] were found to be statistically significantly higher in the case group by univariate analysis (p< 0.05). In multivariate analysis, it was determined that TZP (OR= 6.82; p< 0.001), IPM (OR= 3.97; p= 0.023) and VAN (OR= 8.46; p= 0.001) use were independent risk factors in MRSA bacteremia. The duration of MEM (p= 0.037) and cephalosporin use (p< 0.001) were significantly longer in the case group, however there was no statistically significant difference between the duration of use of other antibiotics (p> 0.05). All MRSA isolates were mecA gene positive (n= 99), the resistance rates for ciprofloxacin, rifampin, gentamicin, tetracyclin, cefoxitin, erythromycin and clindamycin were 95%, 95%, 94%, 96%, 98%, 71% and 36%, respectively. All of the isolates were found to be susceptible to trimethoprim-sulfamethoxazole, VAN, TEC, tigecycline, linezolid and daptomycin. Mortality rates in patients who were infected with MRSA strains exhibiting vancomycin MIC value of ≤ 1.0 µg/ml (n= 49) and with MRSA strains exhibiting MIC > 1.0 µg/ml (n= 50) were 34.6% (17/49) and 60% (30/50), respectively. This difference was found to be statistically significant (p= 0.012). Thus it was concluded that the mortality rate increased in patients infected with MRSA with high (> 1.0 µg/ml) vancomycin MIC value. The results of this study indicated that obesity, presence of central venous catheter and nasogastric tube, and the use of H2 receptor blockers, IPM, TZP and VAN were independent risk factors for MRSA bacteremia. This was the first study showing the relationship between increasing mortality and high vancomycin MIC values in MRSA bacteremia in Turkey.
    Mikrobiyoloji bülteni 10/2014; 48(4):523-37.
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    ABSTRACT: Liver-derived paraoxonase-1 (PON1) enzyme that is found in the circulation is bound to high-density lipoproteins and reduces the amount of oxidized lipids with its antioxidant effect. Humans have at least three different PON gene regions which are adjacent to the other on the 7th chromosome. It has been shown that PON1 gene and its polymorphisms are related with various diseases. It is also known that, hepatitis C virus (HCV) is tightly associated with the cell lipoproteins in each step of its replication cycle leading to modulation of the host lipid metabolism. The aim of this study was to investigate the relationship between the response to chronic hepatitis C (CHC) therapy and aminoacid changes in 55' and 192' regions of PON1 enzyme believed to be involved in the pathophysiology of many chronic diseases. A total of 49 CHC patients (27 male, 22 female; mean age: 52.9 ± 12.6 yrs), all infected with HCV genotype 1b and positive for anti-HCV and HCV-RNA were included in the study. Patients who were HCV-RNA negative at the sixth month following at least once pegilated interferon + ribavirin treatment, were considered as therapy-responders, whereas those who were HCV-RNA positive were considered as non-responders. The genomic DNAs were isolated from patients' blood samples in their routine follow-ups and Q/R192 and L/M55 PON1 polymorphism analysis in 55. and 192. regions was performed by T-ARMS-PCR (Tetra-primer amplification refractory mutation system-polymerase chain reaction) method. In our study, the analysis of PON1 polymorphisms yielded 44.1% of LL, 44.1% of LM and 11.8% of MM genotypes at position 55 and 55.9% of QQ, 41.2% of QR, and 2.9% of RR genotypes at position 192 in therapy-responders. In the evaluation of combined genotype analysis of the patients, there was only one case who was responsive to treatment with LL/RR genotype. Of the patients, eight harbored LL/QQ genotypes and seven of them (87.5%) were responsive to treatment. However, statistical analysis indicated that there was no relationship between PON1 L/M55 and PON Q/R192 polymorphisms and response to CHC treatment (chi-square test, p> 0.05). Our data did not support a relationship between PON1 polymorphisms and response to CHC therapy, in contrast to a few studies pointing out of this correlation. This might be attributed to relatively low number of patients included. In conclusion, since antiviral agents used for CHC therapy are limited and costly, it was thought that further investigations with large numbers of patients should be conducted to establish the presence of any relationship between the response to CHC therapy and genotypes of the PON1 enzyme.
    Mikrobiyoloji bülteni 10/2014; 48(4):596-605.
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    ABSTRACT: West Nile virus (WNV) infection which is asymptomatic or mild in normal population, it may cause serious clinical conditions leading to death in eldery and immunosupressed patients. The virus is mainly transmitted by mosquito bites, however transfusion, transplantation, transplasental and nosocomial ways have also been reported to be responsible for viral transmission. It is known that WNV may cause life-threatining conditions such as central nervous system (CNS) infections especially in bone marrow and solid organ transplant recipients. In this report, the first case of WNV encephalitis in an immunosuppressed patient with renal transplant in Turkey was presented. A 25-year-old male patient admitted to our hospital with the complaints of generalized myalgia, nausea and vomiting, after the 24. day of renal transplantation from a live donor. Since he developed diffuse tonic clonic seizures during his follow up, he was diagnosed as meningoencephalitis with the results of cranial magnetic resonance imaging (MR) and cerebrospinal fluid (CSF) biochemistry. Bacterial and fungal cultures of blood and CSF yielded negative results. CMV antigenemia test and CMV IgM in blood, and nucleic acid tests for CMV, EBV, HSV-1/2, VZV, HHV-6, enterovirus and parvovirus in CSF were also negative. However, WNV RNA was detected in CSF by an in-house reverse transcriptase (RT) nested PCR method. The sequence analysis (GenBank BLAST) of the virus showed that it had 99% similarity with Lineage-1 WNV strains. To define the transmission way of the virus to the recipient, WNV-RNA was searched in the renal biopsy sample and found negative by RT nested PCR. The clinical condition of the patient was improved with supportive therapy and by the de-escalation of immunosuppressive drugs [Mycophenolate mofetil (MMF; 1 g/day), cyclosporin (1 mg/kg/day)]. However WNV meningoencephalitis recurred one month later. The patient presented with fever, myalgia, confusions, leukocytosis, anemia, and repeating WNV-RNA positivity in CSF. This time cyclosporin was stopped, MMF was given in low dose (1 g/day), and high dose parenteral acyclovir and intravenous immunoglobulin (400 mg/kg/day, 7 days) were initiated. The patient recovered completely after 10 days without any neurological abnormalities. In conclusion, especially in endemic areas, WNV should be considered in the differential diagnosis of CNS infections develop in solid organ transplant cases and patients with other immunodeficiencies who present with fever, generalized myalgia, gastrointestinal symptoms and/or neurological disorders.
    Mikrobiyoloji bülteni 10/2014; 48(4):674-82.
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    ABSTRACT: We are grateful to Hatipoğlu and Turhan [Mikrobiyol Bul 2014; 48(1): 188-9] for their interest in our study published in Mikrobiyol Bul 2013; 47(2): 382-4. As Hatipoğlu and Turhan mentioned in their comment, ertapenem is more sensitive than other carbapenem antibiotics for the identification of New Delhi Metallo-beta-lactamase (NDM-1) producers among carbapenem-resistant strains being studied. However, its low specificity [Dortet et al. Biomed Res Int 2014; 2014: 249856] makes it equal with other carbepenems. Since all the isolates in our study were not tested for ertapenem susceptibility, we used the susceptibility data for three carbapenems to increase the sensitivity of our study regarding isolate selection. We agree Hatipoğlu and Turhan about the Modified Hodge Test (MHT) and we did not use MHT at all in our study. However we couldn't understand how they came to a conclusion that we used MHT and didn't mention in Material and Methods section. ZnSO4 supplemented MHT which was recommended by the authors [Dortet et al. Biomed Res Int 2014; 2014: 249856] has a sensitivity rate of about 85%. Thus we used molecular methods instead of MHT not to miss any single isolate. Hatipoğlu and Turhan mentioned about previously reported four NDM-1 positive isolates without any international relation in Turkey. However, since this mentioned study [Alp et al. J Hosp Infect 2013; 84(2): 178-80] was published after the appeal, acceptance and publication of our study, eventually we didn't have the opportunity to discuss the data of Alp's report. In the same study authors stated that NDM-1 producing isolates were isolated from pediatric patients and had no connection with patients from Indian peninsula. At the same time Poirel et al. [Antimicrob Agents Chemother 2014; 58(5): 2929-33] reported in their study that NDM-1 producing isolates from pediatric patients had clonal relation with Enterobacter cloacae strains and subject to an outbreak. The evaluation of the previous reports about NDM-1 indicated that NDM-1 was initially originated from foreign sources before exhibiting endemicity in a country. Thus the situation in our region was not an exception. In conclusion, medical facilities taking care of foreign patients should pay particular attention to identification of NDM-1 isolates and establishment of appropriate control measures.
    Mikrobiyoloji bülteni 10/2014; 48(4):709-10.
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    ABSTRACT: The frequency of fungal infections have increased recently in parallel to prolonged survival of patients with chronical infections, common use of the broad-spectrum antibiotics and cytotoxic drugs and surgical interventions. Fungi such as Trichosporon, Fusarium and Geotrichum that were previously evaluated as contaminant/colonization, become important causes of morbidity and mortality especially in neutropenic patients. The aim of this study was to investigate the presence of virulence factors such as acid proteinase, phospholipase, esterase, coagulase and hemolytic activity among Trichosporon species. A total of 40 Trichosporon strains, of them 24 (60%) were T.asahii, 6 (15%) were T.inkin and 10 (25%) were the other species (one of each of T.aquatile, T.asteroides, T.coremiiforme, T.cutaneum, T.dermatis, T.faecale, T.japonicum, T.montevideense, T.mucoides, T.ovoides) were included in the study. Identification of the isolates was performed according to microscopic morphology (blastospores, arthrospores, pseudohyphae and true hyphae) on corn meal agar media, and carbohydrate assimilation patterns (API ID32C; bioMérieux, France). Secretory acid proteinase, phospholipase and esterase activities of the strains were evaluated by 1% bovine serum albumin containing agar, by egg yolk containing solid medium, and by Tween 80 containing solid medium, respectively. Hemolytic activity of the isolates were evaluated by 5-10% sheep blood Sabouraud dextrose agar. Coagulase enzyme activity was determined by using human and rabbit plasma. In our study, all of the 40 Trichosporon spp. strains were found negative in terms of acid proteinase and phospholipase enzyme activity, however all were positive for esterase enzyme activity. Hemolytic enzyme activity were identified in a total of 15 (37.5%) strains, being "+++" in three strains (2 T.asahii, 1 T.japonicum), and "++" in 12 isolates (9 T.asahii, 1 T.inkin, 1 T.asteroides, 1 T.mentevideense). Although 11 of those 15 positive strains were T.asahii, there was no statistical difference between the species in terms of hemolytic enzyme activity (p> 0.05). Coagulase enzyme activity was detected in 5% (2/40; 1 T.asahii, 1 T.inkin) of the strains with human plasma and in 27.5% (11/40; 9 T.asahii, 1 T.inkin, 1 T.montevideense) with rabbit plasma. In conclusion, our data indicated that esterase, coagulase and hemolytic activities detected in Trichosporon spp. might play role in the pathogenesis of Trichosporon infections, however, further large-scaled clinical and mycological studies are needed to prove this relation.
    Mikrobiyoloji bülteni 10/2014; 48(4):628-38.
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    ABSTRACT: Reactivation of Toxoplasma gondii infections and serious clinical manifestations such as encephalitis may develop in immunocompromised subjects and AIDS patients. Different protocols are used for the treatment of toxoplasmosis in high-risk patient groups, however life-long prophylactic therapy against reactivation risk in AIDS patients may lead to several undesired results. Atovaquone is an effective antiprotozoal agent against toxoplasmosis with minor side effects. On the other hand, Astragalus membranaceus root extract (AmE) has been shown to have immunomodulatory and antimicrobial activities, empowering immunity by enhancing proliferation and activation of phagocytic cells mainly macrophages, and inducing Th1 type immune response. The aim of this study was to investigate the effectiveness of atovaquone alone and in combination with AmE, in the treatment of toxoplasmosis, and on the levels of IL-2, IL-12 and IFN-γ in experimentally infected mice with T.gondii. For this purpose, four experimental groups, each consisting of eight BALB/c mice, were set with the approval of Ethics Committee for the Animal Experiments. All the mice were infected with 0.5 ml of a suspension containing 2 x 104/ml trophozoites prepared from T.gondii RH strain by intraperitoneal injection. Twenty-four hours after the infection, atovaquone (100 mg/kg/day) was given to atovaquone group, AmE (0.075 mg/g) to astragalus group and atovaquone (100 mg/kg/day) plus AmE (0.075 mg/g) to Atovaquone + Astragalus (Ato + Astra) group by oral gavage. The mice in the fourth group, which was the control group, were all infected but untreated. The above administrations were carried out for seven days. On the 8th day peritoneal fluids of mice were collected under anaesthesia and trophozoite numbers per 1 ml were detected by counting on the Thoma slide. In addition, the heart bloods of mice were drawn and IL-2, IL-12, IFN-γ levels were determined in serum samples by using commercial ELISA kits (eBioscience, Austria). The mean number of trophozoites in Ato + Astra group was found significantly lower than the number of trophozoites in the other three groups (p< 0.05). The number of trophozoites in the atovaquone and astragalus groups were found significantly lower than the number of trophozoites in the control group (p< 0.05). There was a significant increase in IL-2 levels of astragalus group compared with the other three groups, in addition when IL-2 levels of Ato + Astra group were compared with ones in other three groups, a significant decrease was noticed (p< 0.05). There was a definite increase in IL-12 levels of atovaquone, astragalus and the control groups compared to those in Ato + Astra group (p< 0.05). A significant increase was found in IFN-γ levels in atovaquone and Ato + Astra groups compared with those in the control group (p< 0.05). Within the reach of our literature survey, this study was the first research in which the effectiveness of the combination of atovaquone and AmE was investigated in the treatment of acute toxoplasmosis. The results of our study suggested that there might be a synergy between atovaquone and AmE in the treatment of acute toxoplasmosis. In case these results are supported by further studies, atovaquone and AmE combination may have a potential to be used for therapy in immunocompromized patients such as AIDS patients who have a risk for toxoplasmosis.
    Mikrobiyoloji bülteni 10/2014; 48(4):639-51.
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    ABSTRACT: Toxoplasmosis is generally asymptomatic in immunocompetent subjects, however serious manifestations of the disease may develop in immunocompromised patients and in pregnant women. The mean Toxoplasma gondii seroprevalence which is approximately 40% in Turkish population, indicates the high risk for the development of acute toxoplasmosis in those cases. One of the transmission ways of T.gondii is the consumption of contaminated water. The aim of this study was to detect the presence of T.gondii in the environmental and drinking water samples by using standard polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) methods. A total of 96 water samples, of them 76 were environmental water samples collected from creeks in Giresun province center and its various districts (Piraziz, Bulancak, Keşap, Espiye) and 20 from drinking water samples, were included in the study. The samples were precipitated by the aluminium sulfate method and DNAs were isolated from the pellets formed with the sucrose gradient method. PCR and LAMP were applied to the isolated DNAs, by the use of primers F3 and B3 specific to B1 gene of the parasite. In our study, T.gondii was detected in none of the drinking water samples by PCR and LAMP, however T.gondii DNAs were positive in 13.2% (10/76) of the environmental water samples. Both of the methods yielded the same results in those samples. The stations that were positive for T.gondii were Aksu creek in Giresun province center; Gelivera creek in Espiye; Yolağzı, Keşap, Keşap Login Bridge creeks in Keşap; Bulancak, Karadere and İncivez creeks in Bulancak; Piraziz and Çayırağzı creeks in Piraziz counties. Resistance of T.gondii to chlorination and the inadequacy of the filtration processes, create a serious threat among people in contact with rivers, sea and drinking waters particularly in areas with high humidity. As far as the national literature was considered, no report about the water-borne T.gondii infections were detected in Giresun province of Black Sea region, Turkey. Thus this study will aid to the literature related to water-borne T.gondii infections. The results of this study emphasizes the essential need for appropriate disinfection procedures and purification processes before the discharge of waste water to prevent the cases of water-borne toxoplasmosis.
    Mikrobiyoloji bülteni 10/2014; 48(4):661-8.
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    ABSTRACT: Febrile neutropenia which is a common complication of cancer treatment, is one of the major causes of morbidity and mortality. Several gram-negative and gram-positive bacteria are responsible for infections in neutropenic patients, however the most common microorganisms are Escherichia coli and coagulase-negative staphylococci, in decreasing order. Although Brucella spp. infections are endemic in Turkey, brucellosis-related febrile neutropenia has only rarely been reported. In this report, a case of brucellosis-related febrile neutropenia in a patient with acute myeloblastic leukemia (AML) was presented. A 56-year-old male patient presenting with fever, petechiae/purpura, leukocytosis, thrombocytopenia, and anemia was admitted to our hospital. Laboratory studies revealed a hemoglobin level of 8.27 g/dl, leukocyte count of 77.100 k/ml, absolute neutrophil count of 200 k/ml, and platelets at 94.200 k/ml. The patient was diagnosed as AML-M1 and piperacillin/tazobactam was started as the first-line antibiotic therapy due to the febrile neutropenia. On admission, blood and urine cultures were negative. Once the fever was controlled, remission/induction chemotherapy was initiated. However, fever developed again on the eight day, and vancomycin was added to the therapy. Since the fever persisted, the antibiotic therapy was gradually replaced with meropenem and linezolid. However, fever continued and the patient's general condition deteriorated. Subsequently performed Brucella tube agglutination test revealed positivity at 1/320 titer and the microorganism grown in blood culture (Bactec 9050; BD, USA) was identified as B.melitensis by conventional methods. Rifampicin and doxycycline therapy was started immediately, however, the patient died due to septic shock. If the tests for brucellosis were performed earlier when response to second step antibiotic therapy lacked in this patient, it was assumed that mortality could be prevented by the prompt initiation of the appropriate treatment. Thus, since brucellosis is endemic in Turkey, it should be considered as a possible agent of febrile neutropenia especially in patients unresponsive to empiric antibiotherapy and appropriate diagnostic tests should be performed.
    Mikrobiyoloji bülteni 10/2014; 48(4):669-73.
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    ABSTRACT: Tuberculosis (TB) is one of those infections with high morbidity and mortality in all around the world. Hundreds of people died from this disease without diagnosed or due to resistant strains in Turkey. Therefore, it is important to identify postmortem cases who have died from tuberculosis. Molecular methods have been widely used as well as conventional methods in the diagnosis of tuberculosis. The aim of this study was to compare the two different real-time polymerase chain reaction (Rt-PCR) system in the postmortem diagnosis of Mycobacterium tuberculosis infections in paraffin-embedded tissues. A total of 40 paraffin-embedded tissue samples [lung (n= 35), brain (n= 2), heart (n= 2), lymph node (n= 1)] in which histopathologic findings consistent with TB (necrotizing granulomatous inflammation, gelatinous caseous pneumonia, necrotic fibrous nodul) obtained from 37 autopsy cases (31 male, 6 female; age range: 25-85 yrs) were included in the study. Paraffin-embedded tissues were deparafinized with xylene and ethyl alcohol and then DNA isolation was done with QIAsymphony DSP Virus/Pathogen Midi kit in the QIAsymphony device. DNA amplification process was performed by Rt-PCR using the kit Artus® M. tuberculosis RG-PCR in the Rotor-Gene® Q device (Qiagen, Germany). Likewise, after deparafinization process, samples placed in the cartridge and isolation and Rt-PCR was performed by Xpert® MTB/RIF (Cepheid, USA) system, simultaneosly. Seventeen and 20 out of the 40 paraffin-embedded tissues yielded positive results with Qiagen and Xpert system, respectively. M.tuberculosis DNA was found positive in 13 (32.5%) and negative in 16 (40%) of the samples by both of the systems, exhibiting 72.5% (29/40) of concordance. On the other hand, seven (17.5%) samples that were positive with Xpert system yielded negative result with the Qiagen, while four (10%) samples that were positive with Qiagen yielded negative result with the Xpert system. Of the 20 positive cases detected with Xpert MTB/RIF system, 15 were found rifampicin-susceptible, and three were rifampicin-resistant. In two samples in which M. tuberculosis DNA was low positive, rifampicin resistance could not be detected. The identification of M.tuberculosis infections in postmortem cases will contribute epidemiological data in Turkey. In these cases, effective sampling and diagnosing of M.tuberculosis infections by acid-fast stain and culture methods are crucial. However, in cases without microbiological sampling the detection of M.tuberculosis DNA in paraffin-embedded tissues with PCR, although there are differences between PCR systems has diagnostic value. In conclusion, our data indicated that Xpert MTB/RIF system is more favourable to detect M.tuberculosis DNA in paraffin-embedded tissues, with the advantages of determination of rifampicin resistance, and detection of more positive results within a shorter time.
    Mikrobiyoloji bülteni 10/2014; 48(4):577-84.
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    ABSTRACT: Following the report of the first AIDS case in Turkey in 1985, the total number of HIV-positive and AIDS cases is 6802 near the end of June 2013, according to the data obtained from the Turkish Ministry of Health. Although combined antiretroviral drug therapy has greatly improved the life-span and the life quality of the patients, HIV-1 drug resistance poses a major obstacle for treatment outcome. The aim of this study was to detect the presence of primary drug resistance in antiretroviral therapy-naive cases. Plasma samples obtained from 190 cases (143 male, 28 female, 19 unknown; age range: 20-72 yrs; mean age: 34.7 yrs) sent to AIDS Confirmation Center of Turkish Public Health Institution, from different provinces of Turkey (Ankara: 64; Adana: 42; Istanbul: 20; Antalya: 13; Gaziantep: 11; Erzurum: 5; Elazig: 3; Malatya: 3; Mersin: 3; and 26 samples from other 21 regions) for routine HIV-1 genotyping test between May 2010-April 2014 period were included in the study. In viral pol gene, complete protease encoding sequence and the first 335 codons of reverse transcriptase (RT) encoding sequence were analyzed by Sanger population-based sequencing method with a commercial test kit (ViroSeq, Abbott/Celera Diagnostics, USA). An alternative in house PCR method with different set of primers was used when the commercial test failed. Primary drug resistance mutations were identified according to the WHO 2009 drug resistance surveillance list. The mean HIV-RNA level at the time of resistance testing was 5.07 (range, 2.09-7.67) log10 copies/ml and the median CD4+ T cell count was 280.3 (range, 0-1000) cells/mm3. The period between the diagnosis of HIV infection and HIV-1 drug resistance test was 18.4 (range, 0-973) weeks. Most prevalent HIV-1 subtypes were subtype B (31%); recombinant B, F1 (24.7%), sub-subtype A1 (16.8%), recombinant B/CRF02_AG (10.5%) and CRF02_G (7.8%). Analysis of our results showed that 10% (19/190) of the samples exhibited resistance mutations. Detected mutations were as follows: M41L, K70E, M184V, L210W and T215C/D/S, responsible for nucleoside RT inhibitor (NRTI) resistance; K103N/S and Y181C, responsible for non-nucleoside RT inhibitor (NNRTI) resistance; M46L and L90M, responsible for protease inhibitor (PI) resistance. NRTI, NNRTI and PI mutation rates in the samples were found as 5.2%, 3.1% and 2.1%, respectively. Both NRTI and NNRTI mutations were detected in one sample. Two mutations leading to NRTI resistance were obtained together in five samples. Seven out of the 19 strains with primary resistance mutation were isolated from samples sent from Ankara, three from Adana, two of each from Istanbul, Erzurum and Mersin, and one of each from Amasya, Samsun and Tokat provinces. There were no statistically significant differences in terms of age, gender, CD4+ T cell count, time from diagnosis to resistance testing between the patient group with primary drug resistance and the group without resistance (p> 0.05). However, the mean HIV-RNA level in patients with primary drug resistance [4.77 (2.95-6.87) log10 copies/ml] was significantly lower than those without primary drug resistance [5.11 (2.09-7.67) log10 copies/ml] (p= 0.043). Our results revealed a relatively high (10%) prevalence of HIV-1 primary drug resistance. We therefore support the need for routine HIV resistance testing so that clinicians can individualize their treatments taking into account the presenting drug resistance.
    Mikrobiyoloji bülteni 10/2014; 48(4):585-95.
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    ABSTRACT: Hepatitis B virus (HBV) infection is a global major health problem. Currently, 10 genotypes (A-J) of hepatitis B virus (HBV) are identified based on the nucleic acid sequence heterogeneity, and these genotypes have been shown to have distinct geographic distribution. Reports of the previous studies indicated that the genotype D is the predominant type among hepatitis B patients in different regions of Turkey. However, recent studies indicated that other HBV genotypes are also seen with an increasing rate. Although epidemiological and clinical information on genotype E infection is currently limited, it is known that genotype E infection is common in West and Central Africa. In this report, the first case of HBV genotype E infection in Turkey was presented. A 22-year-old Nigerian male employee who resided in Manisa for five years was admitted to Celal Bayar University Hospital Manisa, Turkey, for his routine check-up. Since HBsAg was found positive, other HBV markers were tested with a repeated serum sample. Laboratory findings were as follows; HBsAg (+), anti-HBs (-), HBeAg (-), anti-HBe (+), anti-HBc (+), anti-HCV (-), anti-HIV (-), ALT: 44 U/L and AST: 45 U/L. HBV-DNA level was detected as 700 IU/ml by real-time PCR (Artus HBV QS RGQ Qiagen, Germany). HBV-DNA isolated from the serum sample of the patient was amplified by PCR and polymerase gene segment of HBV was directly sequenced. UPGMA method was used for phylogenetic analysis and Inno-LIPA HBV genotyping method (Innogenetics, Belgium) was performed to determine multiple HBV genotype infection. On the basis of those methods the genotype of the virus was identified as genotype E. The partial sequences of the HBV polymerase gene were loaded to the international DNA data bank (GenBank) for contribution to the global HBV surveillance. This report emphasized that besides genotype D the other HBV genotypes could be found in Turkey. Since the patient was an inactive HBsAg carrier before his residence in Turkey, this case was regarded as an imported HBV genotype E case. In conclusion, detection of different HBV genotypes, their epidemiology and molecular characteristics are important for both national and global HBV surveillance and better clinical approach.
    Mikrobiyoloji bülteni 10/2014; 48(4):683-8.
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    ABSTRACT: Acinetobacter baumannii which is a significant cause of nosocomial infections, increases the rate of morbidity and mortality in health care settings especially in intensive care units (ICUs). The aim of this study was to determine the antibiotic resistance profiles of A.baumannii strains isolated from blood cultures of inpatients from different ICUs, wards and hospital environment and evaluate their clonal relationships and epidemiologic features. A total of 54 A.baumannii strains (47 from the blood cultures and 7 from the hospital environment), identified between 01 January 2012-28 December 2012 at the Clinical Microbiology Laboratory of Ankara Numune Training and Research Hospital, Turkey, were included in the study. Identification of A.baumannii isolates and their antimicrobial [sulbactam-ampicillin (SAM), piperacillin (PIP), piperacillin-tazobactam (TZP), ceftazidime (CFZ), cefoperazone-sulbactam (SCF), cefepime (CEF), imipenem (IMP), meropenem (MER), amikacin (AMK), gentamicin (GEN), netilmicin (NT), ciprofloxacin (CIP), levofloxacin (LVF), tetracycline (TET), tigecycline (TG), colistin (COL), trimethoprim-sulfamethoxazole (SXT)] susceptibility testing were performed by Vitek 2 (bioMérieux, France) system. The clonal relationship between the A.baumannii isolates was analysed by pulsed-field gel electrophoresis (PFGE). In our study colistin, tigecycline and netilmicin were found to be the most effective agents against A.baumannii isolates. All of the clinical isolates (n= 47) were found susceptible to COL, however all were resistant to SAM, PIP, TZP, CEF, IPM, CFZ, MER and CIP. While 1.85%, 14.8%, 14.8%, 16.6%, 59.2% and 22.2% of the isolates were susceptible to SCF, AMK, NT, GEN, TG and SXT, respectively; 1.85%, 1.85%, 9.2%, 16.6%, 38.8% and 27.7% of the isolates were intermediate to SCF, TET, AMK, NT, LVF and TG, respectively. Similarly, all of the environmental A.baumannii isolates (n= 7) were resistant to SAM, PIP, TZP, CFZ, CEF, IPM, MER and CIP, and all were susceptible to TG and COL. The resistance rates of the environmental isolates to SCF, AMK, GEN, NT, LVF, TET and SXT were determined as 57.1%, 85.7%, 85.7%, 28.8%, 28.6%, 85.7% and 57.1%, respectively. PFGE analysis done by the use of ApaI enzyme revealed the presence of one major clone. Dendogram analysis indicated that environmental and clinical isolates were in the same clone indicating that the outbreak was possibly originated from the same internal ICUs. Our data emphasized that multidrug resistant A.baumannii isolates were quite common in our hospital, and enviromental cross-contamination throughout the year was confirmed by molecular methods. Despite the precautions such as continous education on effective hand washing, use of gloves and hospital cleaning, established in our hospital, this single clonal spread was attributed to staff shortage and poor adherence to infection control rules. In conclusion, for the prevention of dissemination of multidrug resistant A.baumannii strains and control of nosocomial infections, infection control strategies should be established and strict compliance to these rules should be provided.
    Mikrobiyoloji bülteni 10/2014; 48(4):566-76.
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    ABSTRACT: The increasing rate of antibiotic resistance in Escherichia coli, the most common pathogen of urinary tract infections (UTIs), leads to difficulties in choosing appropriate antibiotic treatment and achieving treatment success. The aim of this study was to investigate in vitro activity of fosfomycin, presented as a favorable choice for the treatment of UTIs caused especially by extended-spectrum beta-lactamase (ESBL)-producing strains. A total of 244 E.coli strains, of them 118 were ESBL positive and 126 were negative, isolated from urine samples of inpatients and outpatients between May 2011-May 2012, were included in the study. Antibiotic susceptibilities of the isolates were determined by disk diffusion method (DDM) and ESBL production was confirmed by double-disc diffusion method according to the CLSI (Clinical and Laboratory Standards Institute) recommendations. Minimum inhibitor concentration (MIC) values for fosfomycin were detected by E-test method. Fosfomycin zone diameters and MIC values of isolates were interpreted according to the breakpoints of both CLSI and EUCAST (European Committee on Antimicrobial Susceptibility Testing). Susceptibilities of ESBL positive and negative isolates to fosfomycin and other antibiotics, and the results of fosfomycin susceptibility tests obtained by different methods were compared. The correlation between fosfomycin zone diameters and MIC values was calculated. In the study, the resistance rates of ESBL-producing isolates to ciprofloxacin, trimethoprim-sulfamethoxazole, gentamicin and amikacin were detected as 67%, 51%, 51% and 19%, respectively, while those rates were as 9%, 21%, 4% and 11%, respectively in non-ESBL producers. The difference between the two groups were found statistically significant (p< 0.001). Fosfomycin resistance of ESBL-producing and non-producing isolates were 3% and 1%, respectively, indicating no significant difference between the two groups (p= 0.356). According to fosfomycin MIC breakpoints defined by CLSI, 98.3% of ESBL-producing isolates and 100% of non-producing isolates were found susceptible to fosfomycin. According to EUCAST recommendations 98.3% of ESBL-producing isolates and 99.2% of non-producing isolates were found susceptible to fosfomycin. There was no significant difference between ESBL-positive and -negative strains according to CLSI and EUCAST recommendations (p= 0.233 and p= 0.611, respectively). When the methods were compared with each other, there were significant differences between DDM and CLSI-MIC or EUCAST-MIC (p= 0.033 and p= 0.049, respectively) and between CLSI-MIC and EUCAST-MIC (p< 0.001). There was a weak reverse linear correlation between fosfomycin zone diameters and MIC values (r= -0.138, p= 0.032). It was concluded that fosfomycin which had a high activity against ESBL-producing isolates was an appropriate alternative antibiotic in the treatment of UTIs.
    Mikrobiyoloji bülteni 10/2014; 48(4):545-55.
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    ABSTRACT: Acinetobacter baumannii is an important opportunistic and multidrug-resistant pathogen leading to nosocomial infections. Over the last 10 years, a significant and threatening increase in resistance to carbapenems, mainly due to the dissemination of class D beta-lactamases, has been reported in A.baumannii worldwide. The most common types of beta-lactamases causing carbapenem resistance in A.baumannii are the OXA-23, OXA-24, OXA-40, OXA-58 and OXA-143 type serine beta-lactamases. The aim of this study was to investigate the presence of OXA type beta-lactamases in carbapenem-resistant A.baumannii strains and the clonal relationship between the strains. A total of 105 non-duplicate carbapenem-resistant A.baumannii strains isolated from various clinical samples (68 blood, 18 bronchoalveolar lavage, 13 drainage, 3 urine, 2 cerebrospinal fluid and 1 catheter samples) in the Microbiology Laboratories of Selcuk University, Meram (2009-2012) and Selcuklu (2007-2008) Medical School Hospitals, were included in the study. The isolates were identified by conventional methods and Phoenix 100 BD (BD Diagnostic, USA) and Vitek II (bioMerieux, France) automated systems. Carbapenem susceptibility test was performed by Kirby-Bauer disk diffusion method according to the CLSI standards. blaOXA 23-like, blaOXA 24-like, blaOXA 58-like and blaOXA 51-like genes were amplified by multiplex PCR assay and clonal relatedness was investigated by pulsed-field gel electrophoresis (PFGE) using ApaI enzyme. The blaOXA 51-like gene was determined in all carbapenem-resistant A.baumannii isolates, while the blaOXA 23-like and blaOXA 58-like genes were detected in 46.6% and 53.3% of isolates, respectively. However blaOXA 24-like gene was not demonstrated in any isolates. blaOXA 23-like gene was determined in both Meram and Selcuklu Medical School hospitals, but blaOXA 58-like gene was detected only in Meram Medical School hospital. PFGE analysis of the isolates revealed 32 different groups in blaOXA 23-like producing A.baumannii strains and 23 different groups determined in blaOXA 58-like producing strains. No common epidemic isolates were detected in the two hospitals, however it was noted that some clones produced small outbreaks in Meram MS hospital. In this study it was shown that blaOXA 23-like and blaOXA 58-like genes together with blaOXA 51-like gene had significant roles in the carbapenem-resistance of A.baumannii strains. Carbapenem-resistant A.baumannii strains producing blaOXA 23-like and blaOXA 58-like enzymes showed the epidemic potential of this nosocomial pathogen and the requirement of molecular typing methods to identify the epidemiologic relationship of the isolates.
    Mikrobiyoloji bülteni 10/2014; 48(4):556-65.
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    ABSTRACT: This study was conducted to investigate the respiratory viruses and subtyping of influenza A virus when positive by multiplex PCR in patients with flu-like symptoms, after the pandemic caused by influenza A (H1N1)pdm09. Nasopharyngeal swab samples collected from 700 patients (313 female, 387 male; age range: 24 days-94 yrs, median age: 1 yr) between December 2010 - January 2013 with flu-like symptoms including fever, headache, sore throat, rhinitis, cough, myalgia as defined by the World Health Organization were included in the study. Nucleic acid extractions (Viral DNA/RNA Extraction Kit, iNtRON, South Korea) and cDNA synthesis (RevertAid First Strand cDNA Synthesis Kits, Fermentas, USA) were performed according to the manufacturer's protocol. Multiplex amplification of nucleic acids was performed using DPO (dual priming oligonucleotide) primers and RV5 ACE Screening Kit (Seegene, South Korea) in terms of the presence of influenza A (INF-A) virus, influenza B (INF-B) virus, respiratory syncytial virus (RSV), and the other respiratory viruses. PCR products were detected by automated polyacrylamide gel electrophoresis using Screen Tape multiple detection system. Specimens which were positive for viral nucleic acids have been further studied by using specific DPO primers, FluA ACE Subtyping and RV15 Screening (Seegene, South Korea) kits. Four INF-A virus subtypes [human H1 (hH1), human H3 (hH3), swine H1 (sH1), avian H5 (aH5)] and 11 other respiratory viruses [Adenovirus, parainfluenza virus (PIV) types 1-4, human bocavirus (HBoV), human metapneumovirus (HMPV), rhinovirus types A and B, human coronaviruses (HCoV) OC43, 229E/NL63] were investigated with those tests. In the study, 53.6% (375/700) of the patients were found to be infected with at least one virus and multiple respiratory virus infections were detected in 15.7% (59/375) of the positive cases, which were mostly (49/59, 83%) in pediatric patients. RSV and rhinovirus coinfections were the most prevalent (18/29, 62.7%) dual infections. The distribution of 436 respiratory viruses identified from 375 patients were as follows; 189 (43.3%) RSV, 93 (21.4%) rhinovirus, 86 (19.8%) INF-A, seven (1.6%) INF-B, 22 (5%) PIV types 1-3, 14 (3.2%) HMPV, 11 (2.5%) HCoV, nine (2%) HBoV, and five (1.2%) adenovirus. Fifty-five (64%) out of 86 INF-A viruses were subtyped as hH3, 24 (27.9%) were sH1 and seven (8.1%) were hH1. Avian H5 was not detected in any samples. The overall prevalence rates of INF-A, INF-B, RSV and other respiratory viruses were 12%, 1%, 27%, and 14.6%, respectively. RSV was the most prevalent respiratory agent in pediatric (161/313, 51%) cases, while INF-A virus in adult (24/62, 38.7%) patients. Influenza viruses were detected as responsible pathogens in 13.3% (93/700) of the patients with flu-like symptoms. Among the cases, a 1-month-old baby was infected with three virus strains (INF-A hH1+INF-A sH1+HCoV OC43) and a 82-year-old patient was infected with two INF-A virus subtypes (hH3 + sH1). INF-A viruses were mostly detected (79/86) in winter period, from December to March. INF-A virus sH1, was the most prevalent subtype in flu cases till February 2011 (22/86), after replaced by INF-A virus hH3. Beginning from February 2012, a significant increase observed in the cases infected with INF-A virus subtype hH3 (39/86). In conclusion, the identification and surveillance of influenza virus types and subtypes circulating in populations have importance both for epidemiological data and selection of vaccine strains.
    Mikrobiyoloji bülteni 10/2014; 48(4):652-60.
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    ABSTRACT: It was proven in the early 1980s that cervical cancer (CC) biopsy specimens and CC cell lines contain human papillomavirus (HPV) DNA sequences. In subsequent years, researchers discovered that the E6 and E7 genes of HPV are expressed in CC tissues and these oncoproteins interact with cellular proteins, including pRb and p53. Establishment of the relationship between HPV infections and CC led to increasing interest on this topic. Comprehensive epidemiological studies determined that specific types of HPV are major risk factors for CC. These HPV types, called high-risk or oncogenic types, are associated with other anogenital cancers and a subset of head and neck cancers. A number of commercial and in-house diagnostic techniques, each of which has a different approach and methodology, have been developed to diagnose HPV infections. HPV testing based on the detection of viral DNA has become an important part of CC screening programs. These screening-typing methods and combined approaches, which are used to diagnose and follow-up (management) HPV infections, have advantages and disadvantages. The choice of method is complicated by several factors and challenges that arise owing to the nature of the virus and the assay methodology. For example, a number of different HPV types can cause genital infections, multiple sequence variations are often observed even in the same HPV genotype, the detection sensitivity of the available methods is variable depending on the HPV genotypes and the viral DNA copy number in the samples examined. The capability for the detection of multiple infections and the ability to perform genotyping differ between the methods. The aim of this review is to summarize the methods used for the diagnosis and follow-up of HPV infections, and to share the current informations that may be helpfull for the choice of appropriate method. For this purpose, molecular-based HPV tests that were used in the past and their development processes were described briefly, then currently accepted and widely used methods for diagnosis, screening, and typing were discussed in detail, along with their advantages and disadvantages.
    Mikrobiyoloji bülteni 10/2014; 48(4):689-706.
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    ABSTRACT: Since hepatitis B virus (HBV) infections may cause serious clinical symptoms and become chronic, the appropriate use and interpretation of laboratory tests in diagnosis is crucial. Implementation of serological tests for all HBV markers leads to high cost, labour force and time loss, so diagnostic algorithms are used for this purpose. The aims of this retrospective study were to detect the number of unnecessary test requests in HBV infection serology and financial burden as its consequence and to find out the reasons and ways of solutions. Three-years records of microbiology laboratory of Dumlupinar University Evliya Celebi Training and Research Hospital, Kutahya, Turkey, between 2011-2013 were analysed for the detection of unnecessary test requests and a total of 179.640 HBV serological test results were evaluated. Serological tests requested more than one on the same day and HBeAg, anti-HBe and anti-HBc IgM tests required from the patients with natural immunity or vaccinated ones were considered as unnecessary test requests. The main reasons of unnecessary test requests were detected as follows; unable to check the previous HBV test results in HIMS (Hospital Information Management System) owing to restricted of time, not being friendly user of HIMS, not trusting the test results, not knowing diagnostics algorithms, requiring of the tests by secretaries or other staff like nurses, not questioning HBV vaccination and requiring screening tests because of the defensive medicine. Survey questions prepared to search these reasons and awareness level of diagnostic algorithms were handed and asked to fill out 146 physicians working at clinics which made over 100 unnecessary requests in 3 years. Total numbers of unnecessary requested serological HBV tests were 5415 (3.01%) and total financial burden for our hospital in three years was 42.256 TL (18.373 USD) according to the notifications of Turkish Social Security Institution. It was determined that the most unnecessary test requests were HBeAg (44.86%), anti-HBe (37.75%) and anti-HBc IgM (37.41%). The most unnecessary test requests were made by internal medicine, obstetrics and gynaecology, general surgery and cardiology clinics, respectively. According to questionnaires filled by the 100 clinicians, the most common causes of unnecessary tests were hepatitis test requests from hospitalized patients for screening (96%), the tests required by secretary or nurse (62%), pre-surgical screening tests (53%) and lack of knowledge about diagnostic algorithms (42%). In conclusion, it is suggested that test requests should be restricted by using screening algorithms and in-service training should be provided, tests required by secretary or nurse should be stopped and previous test results should be conveyed to the clinicians actively through HIMS in order to prevent unnecessary test requests causing economic and workforce loss in HBV infection serology. These measures can provide early and true diagnosis for the patients and also will reduce the economic loss.
    Mikrobiyoloji bülteni 10/2014; 48(4):618-27.
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    ABSTRACT: Staphylococci are one of the most common pathogens isolated from nosocomial and community acquired infections. Antibiotics such as clindamycin and erythromycin have been useful options for treating skin and soft-tissue infections caused by staphylococci. However, expression of macrolide-lincosamide-streptogramin B resistance (MLSB) can limit the effectiveness of these drugs. The aims of this study were to investigate the prevalence and phenotypes of MLSB resistance in staphylococcus strains isolated from clinical samples and to determine the telithromycin activity against these isolates. A total of 218 strains [92 Staphylococcus aureus and 126 coagulase-negative staphylococci (CNS)] isolated from different clinical samples (wound, abscess, blood, sterile body fluids, catheter, upper respiratory tract samples) between February 2011 to December 2012 were included in the study. The isolates were identified by using conventional methods and automated bacterial identification system (BD Phoenix 100™ System, Becton Dickinson, USA). Methicillin resistance of the isolates was determined with the use of cefoxitin (30 µg) disk and telithromycin (15 µg) activity was detected by Kirby-Bauer disk diffusion method. MLSB resistance phenotypes were investigated by the D-test method using erythromycin (15 µg) and clindamycin (2 µg) disks. Of 92 S.aureus isolates, 23 were methicillin-resistant (MRSA) and 69 were methicillin-susceptible (MSSA), whereas 78 of 126 CNS isolates were methicillin-resistant (MRCNS) and 48 were methicillin-susceptible (MSCNS). Hundred and seventy-two (79%) isolates were found as erythromycin-resistant, and the rates of erythromycin resistance in MRSA, MSSA, MRCNS and MSCNS strains were 83%, 71%, 95% and 63%, respectively. Inducible type of MLSB resistance (iMLSB type) was observed in 26%, 6%, 51% and 33%; chromosomal resistance (cMLSB type) in 32%, 27%, 27% and 17% and efflux pump connected resistance (MSB type) in 42%, 67%, 22% and 50% of the MRSA, MSSA, MRCNS and MSCNS, respectively. Forty-four (20%) strains were found susceptible to both clindamycin and erythromycin (S type resistance). Resistance due to enzymatic inactivation (L type) was observed only in two of the CNS strains (0.9%), one was methicillin-resistant and the other was susceptible. Total telithromycin resistance was detected as 26.6% (n= 58), while the resistance rates in MRSA, MSSA, MRCNS and MSKNS isolates were 35%, 35%, 28% and 8%, respectively. Telithromycin resistance rate was 34% (58/172) in erythromycin-resistant isolates. However, all erythromycin-susceptible isolates (n= 46) were also susceptible to telithromycin. Telithromycin-resistant isolates frequently exhibited cMLSB phenotype (39/44; 67.2%), followed by MSB (16/72; 27.6%) and iMLSB (3/56; 5.2%). In conclusion, clindamycin is still an effective antibiotic for the treatment of staphylococcal infections in our hospital, however, 34% resistance rate against telithromycin may limit the use of this agent which is an alternative for the treatment of infections caused by clindamycin and erythromycin-resistant strains.
    Mikrobiyoloji bülteni 07/2014; 48(3):469-476.