Erdogan Malatyali

Blastocystis in ulcerative colitis patients: Genetic diversity and analysis of laboratory findings.

OBJECTIVEnTo determine Blastocystis frequency and subtypes (ST) in ulcerative colitis (UC) patients and analyse some laboratory findings between Blastocystis positive and negative cases.nnnMETHODSnFaecal samples from 150 UC patients in Adnan Menderes University, Training and Research Hospital were examined by direct microscopy and cultivated in Jones medium. Blastocystis positive cultures were subjected to DNA isolation and subtypes were identified by sequencing of barcode region. A retrospective analysis was conducted on C reactive protein (CRP), leucocyte counts (WBC), neutrophil counts, and sedimentation rates.nnnRESULTSnThe overall positive rate of Blastocystis was 8% (12 patients) and the most abundant subtype was ST3 (eight isolates, 66.7%), followed by ST1, ST2 and ST7. Laboratory findings between Blastocystis infected and non-infected UC patients were not significantly different. Blastocystis frequency was 3.8% among the patients in active stage, while it was 11.8% among the patients in remission stage.nnnCONCLUSIONSnThe present study confirms previous findings that have indicated the predominance of Blastocystis ST3 in humans and contributes additional evidence that suggests the low colonisation of Blastocystis infection in ulcerative colitis patients during active stage.

Blastocystis subtypes in cancer patients: Analysis of possible risk factors and clinical characteristics.

The aim of the present study was to examine the frequency and genetic diversity of Blastocystis in cancer patients of a Medical Oncology Department in Aydın, Turkey. Patients stool samples were examined between January 2013 and February 2014 by both microscopy and culture methods. Culture positive samples were subjected to DNA isolation and Sequence Tagged Site (STS)-PCR analysis. Possible etiological factors and clinical features of Blastocystis infection were also analyzed and compared between Blastocystis infected and non-infected subgroups. Blastocystis was detected in 15 (6.5%) of 232 stool samples by microscopy and in 25 (10.8%) by culture methods. Out of 25 culture positive isolates, the most prevalent subtype was ST3 (59%), followed by ST1 (23%) and ST2 (18%). Blastocystis frequency was higher in the male patients than the females (19% vs. 6.5%, p<0.05) and in the patients living in urban areas than rural (15.3% vs. 6.6%, p<0.05). Interestingly, Blastocystis was more frequent in patients with lung cancer than the other cancer types (χ2=18, p<0.05) and also in the patients who had received at least eight chemotherapy cycles than fewer (21.4% vs. 9.9%, p<0.05). The rate of gastrointestinal symptoms was not significantly different between infected and non-infected cases. The pathogenic and clinical impacts of Blastocystis in cancer patients should be further examined, particularly as relates to treatment, microbiota and cancer type.

Use of Internal Transcribed Spacer Sequence Polymorphisms as a Method for Trichomonas vaginalis Genotyping

OBJECTIVEnTrichomonas vaginalis is the most common non-viral, sexually transmitted pathogen with a worldwide distribution. The aim of the present study was to design a new genotyping tool for T. vaginalis isolates using internal transcribed spacer (ITS) sequences.nnnMETHODSnFirst, a total of 20 cryopreserved T. vaginalis isolates were thawed and genomic DNA was isolated from fresh cultures. A polymerase chain reaction (PCR) was performed to amplify the ITS regions and the amplicons were sequenced. These sequences were aligned with others from Genbank and polymorphisms were detected. At last, each ITS sequence was given a different sequence type.nnnRESULTSnMore than 99% homology was observed among sequences. Of 20 isolates, five had identical ITS sequence to reference (L29561) defined as ITST1. Moreover, 13 had A58 deletion (ITST10), one had C203T mutation (ITST2), and one had both A58 deletion and C203T mutation (ITST11). ITS typing of T. vaginalis sequences on Genbank revealed a total of 11 ITS types with the predominance of ITST1 (44.4%) globally.nnnCONCLUSIONSnITS typing seems to be an applicable and useful tool for a better understanding of molecular epidemiology as well as for the dissemination of T. vaginalis clones.

Determination of Trichomonas vaginalis Genotypes Using PCR-Restriction Fragment Length Polymorphism (RFLP)

OBJECTIVEnTrichomonas vaginalis is an anaerobic protozoon and the most common non-viral sexually transmitted pathogen. The present study was designed to determine the genotypes of T. vaginalis using polymerase chain reaction (PCR)-restriction fragment length polymorphism of actin gene.nnnMETHODSnA total of 20 isolates from symptomatic females isolated and cryopreserved at Adnan Menderes University, Research and Training Hospital Parasitology Laboratory were included. The isolates from liquid nitrogen were thawed and grown in trypticase-yeast extract-maltose medium prior to the study. Following nucleic acid extraction, the actin gene of T. vaginalis was amplified using nested PCR and amplicons were concentrated with phenol-chloroform-isoamyl alcohol precipitation. The final products were digested with HindII, MseI, and RsaI and were visualized using agarose gel electrophoresis.nnnRESULTSnMost isolates were actin genotype E (n=9, 45%). The remaining isolates were genotype G (n=7, 35%), genotype N (n=1, 5%), and genotype H (n=1, 5%); two were mixed genotypes of E and H (10%).nnnCONCLUSIONnTo the best of our knowledge, this study is the first to provide data on T. vaginalis genotypes in Turkey. However, further studies should be conducted to understand the molecular epidemiology of T. vaginalis at the national and global levels.

Investigation of the Applicability of a Rapid Diagnosis Test in the Diagnosis of Cystic Echinococcosis

OBJECTIVEnEchinococcus granulosus, the etiological agent of cystic echinococcosis (CE) in humans and livestock, is a widely distributed zoonotic pathogen tapeworm. The infection is transmitted to humans by the ingestion of E. granulosus eggs released in the feces of definitive hosts such as dogs. The larval stage of the parasite develops a slowly enlarging cyst in the visceral organs, particularly in the liver and/or lung. The aim of the present study was to evaluate the diagnostic value of an immunochromatographic test (ICT) for CE.nnnMETHODSnA total of 50 sera from surgically and/or pathologically confirmed patients with CE were included in the study as the study group; the control group comprised patients who tested negative for enzyme-linked immunosorbent assay (ELISA). Sera were selected from the collection at Adnan Menderes University, Faculty of Medicine, Parasitology Laboratory, by simple random sampling. The collection included sera obtained between 2010 and 2014; antibody titers of each serum sample were determined using in-house ELISA, before storage at -20°C. The presence of E. granulosus antibody in the sera was determined using a commercially available ICT (VIRAPID® HYDATIDOSIS) kit method.nnnRESULTSnIn the study group (E. granulosus-confirmed cases), two (4%) of the 50 sera were negative and 48 (96%) were positive with ICT. In the control group (ELISA-negative), all were negative with ICT.nnnCONCLUSIONnThe rapid diagnostic test has been evaluated as a practical, easy-to-use method for detecting CE, and it can be used as a screening test in routine diagnosis and research.OBJECTIVEnEchinococcus granulosus, the etiological agent of cystic echinococcosis (CE) in humans and livestock, is a widely distributed zoonotic pathogen tapeworm. The infection is transmitted to humans by the ingestion of E. granulosus eggs released in the feces of definitive hosts such as dogs. The larval stage of the parasite develops a slowly enlarging cyst in the visceral organs, particularly in the liver and/or lung. The aim of the present study was to evaluate the diagnostic value of an immunochromatographic test (ICT) for CE.nnnMETHODSnA total of 50 sera from surgically and/or pathologically confirmed patients with CE were included in the study as the study group; the control group comprised patients who tested negative for enzyme-linked immunosorbent assay (ELISA). Sera were selected from the collection at Adnan Menderes University, Faculty of Medicine, Parasitology Laboratory, by simple random sampling. The collection included sera obtained between 2010 and 2014; antibody titers of each serum sample were determined using in-house ELISA, before storage at -20°C. The presence of E. granulosus antibody in the sera was determined using a commercially available ICT (VIRAPID® HYDATIDOSIS) kit method.nnnRESULTSnIn the study group (E. granulosus-confirmed cases), two (4%) of the 50 sera were negative and 48 (96%) were positive with ICT. In the control group (ELISA-negative), all were negative with ICT.nnnCONCLUSIONnThe rapid diagnostic test has been evaluated as a practical, easy-to-use method for detecting CE, and it can be used as a screening test in routine diagnosis and research.

Genotyping of Echinococcus granulosus Isolates by Sequencing of Mitochondrial Cytochrome C Oxidase Subunit 1 (cox1) Gene in Aydin

Cyst hydatid (CH) is a zoonotic infection that is characterized by the development of metacestode form of Echinococcus granulosus primarily in liver of humans and ruminants. With a worldwide distribution, the infection is still considered as an important parasitic disease that threatens the public health in Turkey as in the other developing countries. Morphological and biological features of parasite fail to discriminate isolates for typing so molecular methods should be used for this purpose. Recently, a total of eleven genotypes of E.granulosus (G1-10 and lion strain) have been identified and these genotypes are highly correlated with host specificity of the parasite. The aim of this study, was to determine the genotypes of E.granulosus isolates from human samples in Aydın. Cyst fluids from CH operated cases in Adnan Menderes University Faculty of Medicine, Training and Research Hospital were used in the present study. Samples were washed with phosphate buffered saline (PBS) and stored in 70% ethanol at -20ºC. Mitochondrial cytochrome C oxidase subunit 1 (cox1) gene was amplified partially by polymerase chain reaction (PCR). The PCR products were sequenced initially, compared to other database in Genbank and evolutionary distances were estimated with references. The genotypes of E.granulosus isolates were determined according to the closest or exact matches to the references. A total of 20 E.granulosus isolates were genotyped in the present study, most of them (15 isolates, 75%) were identified as Genotype 1 (G1), that is defined as sheep genotype and the remaining isolates were defined as pig/camel genotype G6/7 (five isolates, 25%). A possible explanation to our results may be related to the geographical position of Turkey. The identification of G6/7 in addition to sheep genotype G1 indicated that pigs and camels in this area have role in the transmission and distribution of E.granulosus to humans. There is still limited information about the molecular epidemiology of E.granulosus in Turkey. This study reveals the first data about the genotype distribution of E.granulosus in our city, therefore the findings may help to design control program for the disease with a combination of epidemiological data.

Multilocus sequence typing of Blastocystis isolates in Aydin, Turkey.

Blastocystis, a stramenopile protozoon of the human gastrointestinal tract, has a worldwide distribution. Multilocus sequence typing (MLST) is described as a portable, universal, and definitive method for accurate strain identification of microorganisms and was recently used for detecting intra-subtype variability of Blastocystis. The present study aimed to determine MLST sequence types in Blastocystis isolates from a routine diagnostic laboratory at Adnan Menderes University, Training and Research Hospital, Turkey. Samples were inoculated into Joness medium after native-Lugol examination. Total genomic DNA was isolated from positive cultures with a commercially available kit. A total of 11 polymerase chain reactions were performed for each isolate (five loci for subtype 3 and six loci for subtype 4) using previously published MLST primers. The amplicons were sequenced and queried against a MLST database. A total of 11 isolates were amplified with subtype 3 MLST primers and could be sequenced at all loci; however none of the isolates were amplified with subtype 4 MLST primers. The isolates in our study population contained ten new alleles and nine sequence types. The present study contributes to existing knowledge of MLST data for Blastocystis isolates and is the first MLST study from Turkey. Our study confirms the extensive genetic diversity in Blastocystis that was reported in previous studies using different methods.