Browsing by Author "Akyar, I"

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    Feconomics®: A Simple, Novel and Fast Technique for Stool Concentration in Parasitology Laboratory
    Kurt, Ö; Akyar, I; Görgün, S; Kocagöz, T; Özbilgin, A
    Feconomics (R) is a new ready-to-use kit for fecal concentration that eliminates the need for centrifugation and floatation by using absorbent beads. To assess its efficacy in the diagnosis of intestinal parasites, a comparative, double-blind study was conducted in the Parasitology Laboratory of Celal Bayar University Medical School. Stools (Group I, n=251) submitted for routine ova and parasite examination were concentrated with both routine formalin ethyl acetate concentration (FEAC) technique and Feconomics (R). Since the number of helminthes identified in the stool samples of patients were very low, helminthes obtained from the animal models in the laboratory were included (Group II, n=11). The iodine-stained samples of all stools and some of the positive samples stained with Gomori's trichrome and Kinyoun's acid fast stain were read by specialists. In Group I, 103 of 251 (41.04%) samples were found to be positive for one or more intestinal parasites; among them, 76(30.28%) and 96(38.25%) stools were found to be positive with FEAC and Feconomics (R), respectively, and the difference was significant (P=0.000). Same parasites were identified with both methods among all 11 samples in Group II. There was no difference between the methods for the morphological integrity and visual appearances of the parasites having cyst or egg forms; yet, it was noticed that the vegetative forms of the parasites were only identified with Feconomics (R). Review of our data indicated that Feconomics (R) may be suggested as a fast and effective fecal concentration method for Parasitology laboratories owing to the identification of higher number of parasites compared to FEAC, and parasites with only vegetative forms such as Dientamoeba fragilis.
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    Evaluation of the performance of MALDI-TOF MS and DNA sequence analysis in the identification of mycobacteria species
    Akyar, I; Çavusoglu, C; Ayas, M; Sürücüoglu, S; Ilki, A; Kaya, DE; Besli, Y
    Background/aim: Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry is an alternative way of identifying mycobacteria via the analysis of biomolecules. It is being increasingly used in routine microbiology practice since it permits early, rapid, and cost-effective identification of pathogens of clinical importance. In this study, we aimed to evaluate the efficacy of phenotypic identification of mycobacteria by the MALDI-TOF MS MBT Mycobacteria Library (ML) 4.0 (Bruker, Daltonics) compared to standard sequence analysis. Materials and methods: A total of 155 Mycobacterium clinical and external quality control isolates, comprising nontuberculous mycobacteria (NTM) (n = 95) and the Mycobacterium tuberculosis complex (MTC) (n = 60), were included in the study. Results: Identification by MBT ML4.0 was correctly performed in 100% of MTC and in 91% of NTM isolates. All of the MTC isolates were correctly differentiated from NTM isolates. Conclusion: Based on our results, MBT ML4.0 may be used reliably to identify both NTM and MTC.
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    Cutaneous leishmaniasis caused by Leishmania infantum in Turkey: reports of two cases diagnosed with genotyping and protein fingerprinting
    Culha, G; Akyar, I; Zeyrek, FY; Gündüz, C; Kurt, Ö; Ostan, I; Töz, S; Kocagoz, T; Ozbel, Y; Ozbilgin, A
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    Evaluation of 2015-2016 MOTAKK HBV DNA and HCV RNA External Quality Assessment National Program Results
    Karatayli, E; Soydemir, E; Aksoy, ZB; Kizilpinar, M; Altay Koçak, A; Karatayli, SC; Yurdcu, E; Yildirim, U; Güriz, H; Bozdayi, G; Yurdaydin, C; Ilhan, O; Yildirim, Y; Bozdayi, AM; Oguz, AY; Baris, A; Alp, A; Aksözek, A; Sayiner, A; Karagul, A; Ordu, A; Istanbullu, A; Otlu, B; Aridogan, B; Aksu, B; Buruk, CK; Karahan, C; Güney, Ç; Toksöz, D; Yildirim, D; Çolak, D; Daglar, DE; Findik, D; Kas, E; Çaliskan, E; Zeyrek, FY; Arslan, F; Demir, F; Milletli, F; Kibar, F; Özdinçer, F; Dündar, G; Arslan, H; Agca, H; Aliskan, HE; Güdücüoglu, H; Fidan, I; Akyar, I; Afsar, I; Kaleli, I; Dönmez, I; Yanik, K; Midilli, K; Çubukçu, K; Özdemir, M; Acar, M; Yalinay, M; Kuskucu, MA; Bakici, MZ; Aydin, N; Yilmaz, N; Çeken, N; Ziyade, N; Yilmaz, N; Özgümüs, OB; Gitmisoglu, Ö; Demirgan, R; Kesli, R; Güçkan, R; Sertöz, R; Akgün, S; Aksaray, S; Tezcan, S; Kaygusuz, S; Gökahmetoglu, S; Mese, S; Bayik, SA; Akçali, S; Gürcan, S; Karsligil, T; Us, T; Özekinci, T; Pilgir, T; Aslan, U; Dinç, U; Coskun, USS; Çetinkol, Y; Keskin, Y; Ayaydin, Z; Toraman, ZA
    MOTAKK, as a national external quality control program has been launched to evaluate the molecular detection of viral infections including HBV DNA and HCV RNA in molecular microbiology diagnostic laboratories in Turkey. This program is prepared in compliance with ISO 17043:2010 (Conformity assessment general requirements for proficiency testing) standards, and aims to take the place of external quality control programs from abroad, contributing to standardization and accuracy of molecular diagnostic tests in our country. The aim of this study was to evaluate 2015 and 2016 results of the MOTAKK External Quality Control Program for HBV DNA and HCV RNA viral load. The calls were announced on the web page of MOTAKK (www.motakk.org). The quality control samples were sent to participating laboratories in 2015 and 2016. Main stocks were prepared from patients with chronic hepatitis B and C who had viral load detection with reference methods according to WHO reference materials for viral load studies to improve quality control sera. From these main stocks, samples with different viral loads were prepared from dilutions of plasma with HBV, HCV, HAV, HIV, Parvovirus B19 and CMV negative serologic markers. Quality control samples were sent to the participating laboratories along with the negative samples in the cold chain. The laboratories accomplished the related tests within 2-3 weeks and entered their results on the MOTAKK web page. These results were analysed according to ISO 13528 (Statistical methods for use in proficiency testing by interlaboratory comparison) and scoring reports were created by a software developed by MOTAKK and sent to participating labs. Each laboratory evaluated their own results in comparison with the other laboratory results, reassessed the tests via observing the distance from the mean result and the reference values. The number of laboratories participating in the HBV DNA and HCV RNA external quality control program was 70-73 in 2015-2016. Participants were able to comply with the program tools, registering, entering results and receiving the results reports problem. In HBV panel, 72.6-89.1% and 84.7-90.3% of the participant laboratories were in 1 standard deviation (SD) in 2015-2016, respectively. In HCV panel, 70.8-89.1% and 84.7-90.3% of the participant laboratories were in 1 SD in 2015-2016, respectively. A national external quality control program for HBV DNA and HCV RNA in Turkey has been prepared for the first time with this project and implemented successfully. All the data provided in the MOTAKK external quality control program final report, compensate all the data provided by the quality control program final reports from abroad; additionally, the report allows comparison of used technologies and commercial products.
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    Leishmaniasis in Turkey: first clinical isolation of Leishmania major from 18 autochthonous cases of cutaneous leishmaniasis in four geographical regions
    Özbilgin, A; Çulha, G; Uzun, S; Harman, M; Topal, SG; Okudan, F; Zeyrek, F; Gündüz, C; Östan, I; Karakus, M; Töz, S; Kurt, Ö; Akyar, I; Erat, A; Güngör, D; Kayabasi, Ç; Çavus, I; Bastien, P; Pratlong, F; Kocagöz, T; Özbel, Y
    ObjectiveTo report isolation of Leishmaniamajor strains obtained from 18 Turkish autochthonous cutaneous leishmaniasis (CL) patients infected with L.major between 2011 and 2014. MethodsInitial diagnosis relied on microscopy and culture in enriched medium, prepared by adding specific amounts of liver extract, protein and lipid sources to NNN medium. Promastigotes were then transferred to RPMI medium including 10% of foetal calf serum for mass culture. Species-specific real-time PCR targeting ITS1 region of Leishmania spp. was performed using both lesion aspiration samples and cultured promastigotes. Two of 18 isolates were identified by isoenzyme analysis in the Leishmaniasis Reference Center in Montpellier, France. Each isolate was inoculated into the footpads of six mice to observe the pathogenicity of L.major. Developing lesions were observed, and the thickening of footpads was measured weekly. ResultsMelting curve analyses of 18 isolates showed a peak concordant with L.major, and two of them were confirmed by isoenzyme analyses as L.major zymodeme MON103. In the mouse model, acute lesions seen on day 21 were accepted as an indication of heavy infection. Severe impairments were observed on all mouse footpads over 3weeks, which even progressed to extremity amputation. ConclusionCutaneous leishmaniasis-causing L.major was recently identified in Adana province in southern Turkey, with PCR. Our study shows that such CL cases are not limited to Adana but currently present from western to Southeastern Anatolia, and along the Mediterranean coast. The role of small mammals, the main reservoirs of L.major in Anatolia, needs to be elucidated, as do the underlying factors that cause severe clinical manifestations in L.major infections in Turkey, contrary to the infections in neighbouring countries. ObjectifRapporter sur l'isolement de souches de Leishmania major obtenues a partir de 18 cas de leishmaniose cutanee (LC) de patients turcs autochtones infectes par L. major entre 2011 et 2014. MethodesLe diagnostic initial a porte sur la microscopie et la culture sur un milieu enrichi, prepare en ajoutant des quantites specifiques d'extrait de foie, de proteines et de sources de lipides au milieu NNN. Les promastigotes ont ensuite ete transferes dans le milieu RPMI contenant 10% de serum fOEtal de veau pour la culture de masse. La PCR en temps reel specifique de l'espece et ciblant la region ITS1 de Leishmania spp. a ete realisee en utilisant a la fois les echantillons d'aspiration de la lesion et de promastigotes cultives. Deux des 18 isolats ont ete identifies par analyse des isoenzymes dans le Centre de reference de la leishmaniose a Montpellier, en France. Chaque isolat a ete inocule dans les coussinets plantaires de six souris pour observer la pathogenicite de L. major. Les lesions en developpement ont ete observees et l'epaississement des coussinets plantaires ont ete mesures chaque semaine. ResultatsLes analyses de courbe de fusion des 18 isolats ont montre un pic concordant avec L. major et deux d'entre eux ont ete confirmes par des analyses d'isoenzyme comme L. major de zymodeme MON103. Dans le modele murin, des lesions aigues observees au jour 21 ont ete acceptees comme une indication de forte infection. Des deficiences severes ont ete observees sur tous les coussinets plantaires des souris pendant plus de trois semaines, qui ont meme progresse jusqu'a l'amputation de l'extremite. ConclusionL. major causant la LC a ete recemment identifie dans la province d'Adana dans le sud de la Turquie par la PCR. Notre etude montre que de tels cas de LC ne sont pas limites a Adana, mais sont actuellement presents dans l'ouest et dans le sud-est de l'Anatolie, et le long de la cote mediterraneenne. Le role des petits mammiferes, principaux reservoirs de L. major en Anatolie, devrait etre elucide, de meme que les facteurs sous-jacents qui causent les manifestations cliniques severes dans les infections a L. major en Turquie, contrairement aux infections dans les pays voisins. ObjetivoReportar el aislamiento de cepas de L. major obtenidas de 18 pacientes turcos con leishmaniosis cutanea (LC) autoctona, infectados con Leishmania major entre 2011 y 2014. MetodosEl diagnostico inicial se realizo mediante microscopia y cultivo en medio enriquecido, preparado mediante la adicion de cantidades especificas de extracto de higado, proteina y fuentes de lipido al medio NNN. Los promastigotes fueron transferidos al medio RPMI con un 10% de suero fetal para su cultivo masivo. Se realizo PCR en tiempo real, especie-especifica, que detecta la region ITS1 de Leishmania spp., utilizando tanto muestras aspiradas de las lesiones como promastigotes de cultivo. Dos de los 18 aislados se identificaron mediante analisis isoenzimatico en el Centro de Referencia de la Leishmaniosis en Montpellier, Francia. Cada aislado fue inoculado en las almohadillas de las patas de seis ratones para observar la patogenicidad de L. major. Se observo el desarrollo de las lesiones y se midio semanalmente el engrosamiento de las almohadillas. ResultadosEl analisis de la curva de fusion de los 18 aislados mostro un pico de concordancia con Leishmania major, y dos de ellos fueron confirmados mediante un analisis isoenzimatico como L. major zymodeme MON103. En el modelo de raton, las lesiones agudas observadas en el dia 21 se aceptaron como indicativas de una infeccion masiva. Se observaron danos graves en todas las almohadillas de los ratones a lo largo de tres semanas, que progresaron hasta la amputacion de las extremidades. ConclusionRecientemente se ha identificado, mediante PCR, LC causada por L. major en la provincia de Adana al sur de Turquia. Nuestro estudio muestra que estos casos de LC no estan limitados a Adana y que actualmente existen desde el oeste al sudeste de Anatolia y a lo largo de la costa Mediterranea. Es necesario aclarar el papel que en Anatolia juegan los pequenos mamiferos, principales reservorios de L. major, al igual que el de los factores que hay detras de las manifestaciones clinicas severas en infecciones por L. major en Turquia, al contrario del de las infecciones presentes en paises vecinos.
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    Determination of Antimony Resistance Mechanism of Leishmania tropica Causing Cutaneous Leishmaniasis in Turkey
    Özbilgin, A; Zeyrek, FY; Güray, MZ; Çulha, G; Akyar, I; Harman, M; Özbel, Y; Ertabaklar, H; Çavus, I; Gündüz, C
    World Health Organization reported that approximately one billion people are at risk in endemic areas, one million cases of cutaneous leishmaniasis (CL) and approximately 300,000 cases of visceral leishmaniasis (VL) were reported per year in the last five years. The number of deaths due to VL is reported to be approximately 20,000 per year. Approximately 2500 cases/year have been reported as CL, caused by Leishmania tropica and Leishmania infantum, in Turkey. The significant increase observed in many cities mainly in the provinces of Mediterranean and Aegean regions in cases and foci in recent years, suggests that there may be an increase in this infections in the following years as well. In Turkey, the causative agent of CL is L.tropica and meglumine antimoniate is used in the treatment of CL. We aimed to determine antimony resistance genes specific for L.tropica by comparing the gene and protein expressions of antimony-resistant and non-resistant L.tropica strains. Ltropica isolates obtained from 3 CL patients without antimonate resistance from Aegean, Mediterranean and Southeastern regions of Turkey were provided to transform into 3 resistant isolates against meglumine antimony in the laboratory conditions. Gene expression alterations by microarray method; protein profiles by two-dimensional gel electrophoresis (2D-PAGE) and relevant proteins by MALDI-TOF/TOF MS of these isolates were accomplished and compared. L.tropica isolates from 10 CL patients who did not respond to antimony therapy were analyzed for resistance to antimonial compounds and quantitative real-time polymerase chain reaction was performed to detect the expression of genes responsible for resistance development. Moreover, differences in protein expression levels in isolates with and without antimony resistance were determined by comparing protein profiles and identification of proteins with different expression levels was carried out. Enolase, elongation factor-2, heat shock protein 70, tripanthione reductase, protein kinase C and metallo-peptidase proteins have been shown to play roles in L.tropica isolates developing resistance to antimonial compounds and similar expression changes have also been demonstrated in naturally resistant isolates from patients. In conclusion, it was revealed that L.tropica strains in our country may gain resistance to meglumine antimoniate in a short time. It is foreseen that if the patients living in our country or entering the country are treated inadequately and incompletely, there may be new, resistant leishmaniasis foci that may increase the number of resistant strains and cases rapidly.
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    Diversity of Leishmania Strains Isolated from Cutaneous Leishmaniasis Patients in Turkey and its Reflection to Clinics in Mice Model
    Özbilgin, A; Çulha, G; Güray, MZ; Zeyrek, FY; Akyar, I; Töz, S; Ural, IÖ; Kurt, O; Kocagöz, T; Çavus, I; Gündüz, C
    Although asexual reproduction has been attributed to Leishmania species, genetic exchange has recently been demonstrated, which helped emerging of hybrid isolates. Situated on the crossroads between three continents, Leishmania hybrids may be present in Turkey. In Turkey, visceral leishmaniasis caused by Leishmania infantum is less common, while cutaneous leishmaniasis (CL) caused by Leishmania tropica and L.infantum could reach 2500 reported cases a year. Our aim was to investigate genetic variability of local Leishmania species and presence of hybrid Leishmania strains in Turkey. Twenty CL patients from Sanliurfa and Hatay, where only L.tropica and both L.tropica and L.infantum cause CL, respectively, were registered equally. All isolates were assessed with real-time polymerase chain reaction (Rt-PCR), isoenzyme analysis, gene sequencing, two-dimensional gel electrophoresis (2D-PAGE) and MALDI-TOF/TOF-MS followed by in vivo analyses on mouse model. Identification of differentially expressed proteins was performed. These proteins were confirmed by sequence analysis. All isolates from Sanliurfa were found to be L.tropica which caused cutaneous infection in mice. However, one of 10 isolates from Hatay was found as Leishmania major which caused cutaneous infection. Five isolates were found as L.tropica with Rt-PCR and gene sequencing, one of which had one different protein from the reference L.tropica strain and caused cutaneous infection. Four of the five isolates had five different proteins compared to reference strain and caused both cutaneous and visceral infections. Remaining four isolates showed double melting curves in Rt-PCR, which were concordant with L.tropica and L.infantum. Their sequencing and isoenzyme analyses indicated them as L.infantum. They had six different proteins compared to reference L.infantum strain and caused cutaneous and visceral infections. It is concluded that the isolates with different proteins were hybrid Leishmania species. In the present study, outcomes of the proteomics, genomics, clinical manifestations and tissue tropism on animal models were evaluated together for the first time. In addition to L. tropica and L.infantum, L.major was identified as a causative agent for CL and hybrids of Linfantum/tropica were also shown to be present.
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    Leishmaniasis in Turkey: Determination of Leishmania Species by Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF MS)
    Culha, G; Akyar, I; Zeyrek, FY; Kurt, Ö; Gündüz, C; Töz, SÖ; Östan, I; Cavus, I; Gülkan, B; Kocagöz, T; Özbel, Y; Özbilgin, A
    Background: Cutaneous leishmaniasis (CL) is endemic in Southeastern Anatolia, mainly in Sanliurfa and Hatay provinces, and the causative agents are mostly Leishmania tropica and less frequently L. infantum. Here, we report the first MALDI-TOF analyses of Leishmania promastigotes obtained from the cultures of two CL cases from Osmaniye and Hatay provinces who were initially diagnosed by microscopy, culture and identified as L. infantum with Real-Time PCR (RT-PCR). Methods: Samples obtained from the skin lesions of patients were initially stained with Giemsa and cultivated in NNN medium. Examination of the smears and cultures revealed Leishmania amastigotes and promastigotes, respectively. The promastigotes (MHOM/TR/2012/CBU15 and MHOM/TR/2012/MK05) obtained from the cultures of both patients were used for RT-PCR targeting the ITS-1 region in the SSU of rRNA. The reference strains of four Leishmania species (L. infantum, L. donovani, L. tropica and L. major) were initially assessed with MALDI-TOF and their data were added to MALDI-TOF Biotyper Library. Results: Both RT-PCR and MALDI-TOF analyses indicated that the causative agent in both patient samples was L. infantum. Conclusion: Despite disadvantages such as requirement of culture fluid with nothing but promastigotes and high cost, MALDI-TOF analysis may be a fast, sensitive and specific diagnostic tool in especially large-scale research studies, where the cost declines, relatively.
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    Progressive Multifocal Leukoencephalopathy Among Ibrutinib Treatment In Chronic Lymphocytic Leukemia
    Çetintepe, T; Gediz, F; Akyar, I; Çetintepe, L; Koç, AM
    Introduction: Both chronic lymphocytic leukemia (CLL) itself and the drugs used for its treatment, pose a risk for progressive multifocal leukoencephalopathy (PML). Although the relationship between Rituximab and PML is well known, case reports that have been recently published, suggest that ibrutinib; which is used in the treatment of CLL, may increase the risk of PML. Case report: Here, we report a case of 64 year-old female patient with CLL who was previously treated with rituximab, fludarabine and bendamustin but developed PML after receiving monotherapy with ibrutinib. According to Naranjo's algorithm, the causality relationship with the drug is possible with a score of 3. The patient initially exhibited neurological symptoms. Magnetic resonance of the brain revealed a bilateral asymmetric hyperintensity in the white matter involving the parietal and occipital lobules, and there was no mass effect, edema, hemorrhagic or iscemic lesions. No enhancement of contrast media was observed. The findings were consistent with demyelination and suggestive of PML. Management and outcome: Mirtazapine treatment was initiated. However, neurological sympthoms continuously progressed over the following weeks and the patient, aged 64, died six weeks after diagnosis of PML. Discussion: PML is a rare and often fatal demyelinating disease of the central nervous system (CNS) that is exclusively seen in immunocompromised patients and there is no specific agent to treat PML. The case discussed here, highlights that the use of ibrutinib in chronic lymphocytic leukemia (CLL) therapy may result in PML.