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A novel coronavirus (SARS-CoV-2) outbreak, responsible for a pneumonia-associated respiratory disorder (COVID-19), has started in early December 2019 in Wuhan, China, and has rapidly spread around the world. Rapid and accurate diagnostic testing plays a crucial role in tackling the COVID-19 pandemic. In this study, it was aimed to compare 5 primer sets designed to amplify different regions for the detection of SARS-CoV-2 and to perform sequence analysis. Conventional RT-PCR was carried out using primers targeting different regions of the virus genome including ORF1ab, Envelope (E), RNA-dependent RNA polymerase (RdRp), Spike (S) and Nucleocapsid (N) genes for the diagnosis of COVID-19. DNA sequence of ORF1ab gene from each sample were compared with the DNA sequence data of SARS-CoV-2 stored in the GenBank and ORF1ab phylogenetic tree was constructed. The amplicon sizes of ORF1ab, S, E, N and RdRp genes were 588 bp, 440 bp, 145 bp, 323 bp and 196 bp, respectively. The SARS-CoV-2 RNA was detected from 74% of total samples from RdRp gene, 87% for N gene, 74% for S gene, 61% for E gene and 82% for ORF1ab region. The ORF1ab sequences of SARS-CoV-2 from 82 patients were had 100% identity to the sequence of Wuhan isolate and among themselves. The phylogenetic analysis revealed that all isolates formed a cluster. The results of this study suggest that the N region is the best for SARS-CoV-2 identification.

Objectives Appropriate antibiotic therapy and prevention of cross-contamination are the most important subjects in tuberculosis (TB) control. The aim of this study was to investigate the major phylogenetic clades and transmission rate of multidrug-resistant (MDR) Mycobacterium tuberculosis isolates (n = 200) from patients with TB in Sivas and Konya Provinces of Turkey. Methods The phylogenetic relationship among the isolates was investigated by spoligotyping method. In addition, the 24-locus mycobacterial interspersed repetitive unit–variable-number tandem repeat (MIRU-VNTR) typing method was used to reveal cross-contamination. Results Spoligotyping revealed 13 different spoligotypes. A total of 188 strains (94.0%) were included in the cluster. The most prominent spoligofamily was the T family (43.0% of strains), followed by LAM (26.0%), H (8.0%), X and S (both 6.0%) and U (5.0%). Also, 12 strains (6.0%) belonged to the Beijing profile. MIRU-VNTR results showed 176 (88.0%) different genotypes among the isolates. In total, 24 strains (12.0%) were in the cluster. Conclusions According to spoligotyping, there is a heterogeneous M. tuberculosis population in Turkey. MIRU-VNTR results showed that cross-contamination observed between MDR M. tuberculosis isolates in Turkey is controllable.

WOS: 000484847800013, The purpose of this study was to determine the phenotypic antibiotic susceptibility patterns, extended-spectrum beta-lactamase (ESBL) genes and genotypic profiles of ESBL-positive Escherichia coli strains isolated from urine samples obtained from outpatients with urinary tract infection in Turkey. A total of 120 E. coli strains during 2017, 2018, and 2019 (40 patients per year) were examined for antibiotic susceptibility patterns by disc diffusion method, for ESBL genes using PCR and sequencing and for molecular typing by pulsed-field gel electrophoresis (PFGE) method. The isolates were evaluated for their sensitivity to 21 different antibiotics. Four different antimicrobial resistance patterns were determined according to antibiotic susceptibility status of the isolates. The beta-lactamase genes detected in the isolates were CTX-M-15 + OXA-1 (n = 14), CTX-M-15 (n = 24), TEM-1 + CTX-M-15 (n = 52), TEM1 + SHV-12 (n = 6), SHV-12 1 (n = 6), TEM-1 + CTX-M-1 (n = 6), TEM-1 + CTX-M-16 (n = 6) and TEM-1 + CTX-M-9 (n = 6). The CTX-M-15 was the most prevalent ESBL enzyme in the isolates. As a result of PFGE analysis performed by XbaI enzyme restriction process, one major PFGE profile and three main groups (Group I-I-III) were observed. While antibiotic resistance profiles of the strains showed four groups (RI-RII-RIII-RIV), PFGE band profiles showed a major group (90% similarity ratio). High ESBL production and decreased susceptibility to broad-spectrum cephalosporins were observed in E. coli strains. In addition, PFGE analysis showed high clonal similarity among E. coli isolates.

WOS:000623839300001 PMID: 33645410 The anti-proliferative effects of 5-methylquinolinium (5MQ) of nicotinamide N-methyltransferase (NNMT) have not been previously investigated on a cervical cancer cell line. NNMT is a metabolic enzyme that is correlated with tumour progression and metastasis. 5MQ is a small molecule inhibitor of NNMT. 0.1-500 mu M of 5MQ was tested on the HeLa epithelial cervical cancer cell line. Cell viability was assessed with the MTT test. TWIST, ZEB1, SERPIN1, SIRT1, CD16, mRNA and various protein expression levels were analysed with Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) and Western Blotting, respectively. 5MQ significantly inhibited HeLa cell proliferation in a concentration and time-dependent manner. Increased cell shrinkage, loss of cellular adhesions and apoptotic bodies were observed in HeLa cells after 5MQ treatment. Following treatment with 5MQ, ZEB1, SIRT1, CD16 mRNA levels were increased while TWIST and SERPIN1 mRNA levels were reduced. Expressions of oncogenic proteins phospho-Akt and SIRT1 were decreased. 5MQ can effectively inhibit HeLa cell proliferation without apparently affecting HEK-293 cell proliferation. IMPACT STATEMENT What is already known on this subject? NNMT is a cytosolic enzyme involved in tumour progression, metastasis and treatment resistance. It was overexpressed in many human malignancies. 5-amino-1-methylquinolinium (5MQ) is a novel small molecule inhibitor of NNMT that has shown promising results in the treatment of obesity and in senescent muscle regeneration. 5MQ has not been tested on the HeLa cervical cancer cell line, previously. What do the results of this study add? In this study, 5MQ was tested on the HeLa cervical cancer cell line for the first time and the molecular changes associated with 5MQ treatment were analysed. 5MQ demonstrated significant anti-proliferative activity on HeLa cells, which displayed morphological signs of apoptosis. Treatment of HeLa cells with 5MQ led to an increase in ZEB1, SIRT1 mRNA while TWIST mRNA was decreased. Phospho-Akt and Sirtuin1 protein expressions were decreased. What are the implications of these findings for clinical practice and/or further research? 5MQ can effectively inhibit HeLa cell proliferation without apparently affecting HEK-293 cell proliferation. 5MQ treatment was associated with a decrease in the expression of phospho-Akt and Sirtuin1 proteins, both of which have been reported to maintain tumour progression. 5MQ can further be investigated and modified for anti-cancer therapy.

Prevalence of neural tube defect (NTD) has reduced after folic acid intake. However; which mechanisms are effective in NTD are not known exactly. In this study; due to the possible effects on hypoxic pathway and embryonic development, particularly on extracellular matrix components, Hif-1α Pro582Ser and TGF-β3 IVS5+104 A/G SfaN1 polymorphisms were studied by PCR-RFLP method both on children with NTDs and mothers. Statistical differences were seen for Hif-1α and TGF-β3 IVS5+104 A/G SfaN1 polymorphisms in children with NTDs but no difference was seen in mothers. Both genes are effective on many pathways and our results suggest that regulation of extracellular matrix components of children during fetal life is important in neural tube defects formation. The results of this study show that Hif-1α Pro582Ser and TGF-β3 IVS5+104 A/G SfaN1 polymorphisms may play a role in NTDs.

Vancomycin-resistant enterococcus (VRE) is a global threat to public health. Knowledge about the occurrence of vanA-carrying enterococci in broiler and environmental samples is important as antibiotic resistance can be transferred to human bacteria. The aim of this study was to investigate the presence of VRE in broiler cloacal and environmental (house) samples and to genotype the isolates. In this study, 350 swabs were collected from broiler farms. All samples were plated onto enterococcus selective agar containing 6 mg/L vancomycin and 64 mg/L ceftazidime. Minimum inhibitory concentration (MIC) values were determined for vancomycin and teicoplanin. Vancomycin-resistant Enterococcus faecium (VREfm) was isolated from 6 out of 300 (2%) broiler cloacal samples and 13 out of 50 (26%) house samples. All E. faecium isolates had vanA genes. All VREfm isolates (19 isolates) were confirmed to be 95% similar to each other. In conclusion, although 20 years have passed since the ban on avoparcin in Turkey, the present study shows that VREfm isolates are still present in broiler production and especially in broiler houses, and most importantly, a major VREfm clone was isolated from broiler cloacal and house samples.

Tunicamycin is an inhibitor of glycosylation that disturbs protein folding machinery in eukaryotic cells. Tunicamycin causes accumulation of unfolded proteins in cell endoplasmic reticulum (ER) and induces ER stress. ER stress is an essential mechanism for cellular homeostasis has role in cell death via reprogramming of protein processing, regulation of autophagy and apoptosis. In this study we show effect of tunicamycin on subpopulation and invasion of CD44+/CD24- MCF7 breast cancer stem cells. CD44+/CD24- cells were isolated from MCF7 cell line by fluorescence activated cell sorting (FACS) and treated with tunicamycin. ER stress was monitored by evaluation of X-box binding protein 1(XBP-1) mRNA splicing, cleaved activating transcription factor 6 (ATF6) nuclear translocation and CCAAT/enhancer-binding protein homologous protein (CHOP) expression. CD44+/CD24- subpopulation was analyzed using flow cytometry. Invasion was investigated by scratch assay, trypan blue staining, 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) proliferation and in vitro migration assays. Increased level of spliced XBP-1, ATF6 nuclear translocation and CHOP protein expression were detected in CD44+/CD24- and original MCF7 cells treated with tunicamycin. Also, a significant decline in CD44+/CD24- cell subpopulation was determined in the cells treated with tunicamycin. The results also showed inhibited invasion, increased cell death, suppressed proliferation and reduced migration in the CD44+/CD24- and CD44+/CD24- rich MCF7 cell culture, under effect of tunicamycin. Our results indicate that CD44+/CD24- phenotype MCF7 cells are susceptible to tunicamycin. The results showed that tunicamycin-induced ER stress suppresses CD44+/CD24- phenotype cell subpopulation and in vitro invasion and accelerates tumorosphore formation. These results suggest that tunicamycin-induced ER stress inhibits CD44+/CD24- phenotype MCF7 breast cancer stem cells. We conclude that using ER-targeting chemicals like tunicamycin is an interesting approach to target breast cancer stem cells inside tumor.

WOS: 000469890500001, PubMed: 30853450, Many reports have been published on the suspected vertical transmission of Encephalitozoon cuniculi; however, prior to 2003, these reports were based on circumstantial evidence, such as histopathological, immunohistochemical, or serological diagnosis of the infection. In 2003, vertical transmission of the parasite was confirmed by detection of E. cuniculi DNA in fetuses with the nested polymerase chain reaction (PCR) technique. However, the passage of the parasite to eyes of fetus during the intrauterine stage still requires verification. In the current study, natively infected with parasite spores female rabbits were mated with non-infected males. All resulting offspring that died before ten postpartum days were investigated using molecular techniques to confirm the intrauterine transmission of the parasite to the offspring' eyes. In total, 119 DNA samples from rabbit offspring tissues were collected from blood, kidney, brain, eye (both eyes were used as single samples), lung, placenta, liver and heart were used for PCR. Parasitic DNA in the eyes of offspring was detected (54%) 6 of 11 naturally seropositive mother rabbits. PCR results were found to be positive for the eyes of 63% (19/30) of the offsprings from seropositive rabbits. Therefore, mother rabbits naturally infected with E. cuniculi showed the molecular presence of the parasite in their offspring' eyes. Sequence analysis confirmed the partial DNA sequence data of E. cuniculi and blast analysis identified the agent as genotype I. These results confirm transmission of E. cuniculi to rabbit offspring' eyes in the intrauterine period. This is the first molecular evidence to show ocular transmission of the infection via an intrauterine route in rabbits.

Summary Introduction Trichosporon asahii is the most common species that causes trichosporonosis. Materials and methods In the present study, a collection of 68 T. asahii strains recovered from hospitalized patients urine samples between 2011 and 2016 was examined. T. asahii strains were identified by sequencing the intergenic spacer 1 region (IGS1) and genotyped. In addition, proteinase, phospholipase, esterase, haemolytic activity, and biofilm formation of a total of T. asahii strains were investigated. Results The predominant genotype was 1 (79.3%) and followed by 5 (8%), 3 (6.9%), 6 (3.4%), 4 (1.1%), 9 (1.1%). In none of the 68 strains, proteinase and phospholipase activities could be detected, while all were found to be esterase positive. Biofilm production and hemolytic activity were detected in 23.5 and 97% respectively. Discussion Our results indicated that six genotypes were (1, 5, 3, 6, 4, 9) present among T. asahii strains and no property was found to associate with a genotype, in terms of virulence factors.

Lung cancer is among the leading causes of cancer-related deaths, and thusmonitoring, treatment and course of the disease has become a major area of research following diagnosis of the disease. The present study aimed to evaluate the role of HIF pathway in development of non-small-cell lung cancer (NSCLC) in patients diagnosed with NSCLC using their tumor and non-tumor tissue specimens. Both tumor and non-tumor tissue specimens were collected from 10 patients with NSCLC.They were placed in Trizol, and stored at - 80 C ° until processing. RNA was isolated from these tissue specimens using the Trizol/chloroform method. Then, the RNA was converted to cDNA by RT PCR. Amplified cDNA was used for gene expression via specific probes and primers. The mean age of patients (9 male, 1 female)was 62.5 years. The HIF (Hypoxia-Inducible Factor)-1 gene expression was 3 times higher in tumor tissuesthan in non-humor tissues. In relation to this pathway, there was an increased gene expression of 3.72forVEGF (Vascular Endothelial Growth Factor), 3.11forMYC (Avian Myelocytomatosis Viral Oncogene Homolog), 2.28 forTGFA (Transforming Growth Factor) and 2.28 for VEGFR-1 (Vascular Endothelial Growth Factor Receptor-1). The increased HIF expression in the present study results in increased hypoxia in the microenvironment of tumor tissue, while induction of HIF by hypoxia results in expression of genes regulated by HIF (VGF, VEGFR-1,MYC, TGFA). Our results indicate that hypoxia and hypoxia-induced pathways play a role in development and metastasis of tumor in patients with NSCLC. Keywords: Non-small-cell lung cancer, gene expression, HIF pathway.

p53 is a transcription factor that regulates multiple cellular processes that are also important in cellular fates such as cell cycle arrest or programmed cell death. Induction of growth arrest or cell death by p53 prevents the repli­cation of damaged DNA and proliferation of genetically abnormal cells. Therefore, inactivation of p53 by muta­tion or deletion is also important in ensuring the cellular homeostasis. However, studies showed that p53 deficient mice and cells such as Saos-2 cells are maintaining their life. This situation suggests that p53-related proteins might compensate the functions of p53 in p53 deficient organisms. The identification of two p53-related proteins, p63 and p73 revealed the transcription of p53 responsive genes in p53 deficient organisms. Both p63 and p73 pro­teins have high homology with the p53 protein and share some of the functions of p53. In contrast to p53, p63 and p73 rarely mutated in human cancers. Here we studied to summarize the current information about the p53 and other p53-related proteins, p63 and p73 that are included into the p53 gene family., p53, hücre akıbetinin belirlenmesinde önemli rol oynayan hücre döngüsünün durdurulması veya programlı hücre ölümüne yönlendirilmesi gibi hücresel birçok yolağın dü­zenlenmesinde görev alan bir transkripsiyon faktörüdür. p53 aracılığı ile programlı hücre ölümünün başlatılması veya hücre çoğalmasının durdurulması, hasarlı DNA rep­likasyonunu ve genetik olarak normal olmayan hücrelerin çoğalmasını engellemektedir. Bu nedenle p53 işlevlerinin mutasyon ya da delesyon ile engellenmesi hücre home­ostasisin sağlanmasında da ayrıca önemlidir. Bununla birlikte, çalışmalar p53\'ten yoksun farelerin ve Saos-2 gibi hücrelerin yaşamlarını devam ettirebildiklerini göstermek­tedir. Bu durum, p53\'ten yoksun olan organizmalarda p53 işlevini karşılayan p53-ilişkili proteinlerin olabileceğini dü­şündürmüştür. p53 ilişkili p63 ve p73 proteinlerinin keşfi, p53\'ten yoksun hücrelerde p53 bağımlı genlerin transkrip­siyonlarına bir açıklama getirmiştir. p63 ve p73 proteinle­rinin her ikisi de p53 proteini ile yüksek oranda benzerlik göstermekte ve p53\'ün bazı işlevlerini de paylaşmaktadır­lar. p53\'ten farklı olarak p63 ve p73 insan kanserlerinde nadir olarak mutasyona uğramıştır. Burada p53 hakkında­ki güncel bilgilerle beraber p53-ilişkili proteinler olan p63 ve p73 hakkındaki bilgileri de özetlemeye çalıştık.

Homocysteine levels have been investigated in patients with obstructive sleep apnoea syndrome (OSAS), a syndrome associated with a high level of comorbid cardiovascular disease (CVD). While significant increases in homocysteine levels have been observed in OSAS patients with CVD, no increases have been noted in OSAS patients without CVD. This study was designed to investigate the methylenetetrahydrofolate reductase ( MTHFR) gene, which is essential for homocysteine metabolism and has been shown to have a causal role in the development of CVD. Eighty subjects, 30 diagnosed with OSAS by polysomnography and 50 controls (healthy volunteers with no symptoms of OSAS) were enrolled. Two mutations in the MTHFR gene were investigated using polymerase chain reactions and restriction fragment length polymorphisms. No significant differences were found in mean age, body mass index, homocysteine levels, or MTHFR allele or genotype distributions between patient and control groups.

Citation Yenicesu GI, Cetin M, Ozdemir O, Cetin A, Ozen F, Yenicesu C, Yildiz C, Kocak N. A prospective case–control study analyzes 12 thrombophilic gene mutations in Turkish couples with recurrent pregnancy loss. Am J Reprod Immunol 2010; 63: 126–136 Problem Recurrent pregnancy loss (RPL) is a heterogeneous disorder. The contribution of specific thrombophilic genes to the pathophysiology of RPL has remained controversial. We evaluated the prevalences of 12 thrombophilic gene mutations among homogenous Caucasian couples with RPL and fertiles. Method of study This was a prospective case–control study evaluating 272 women with RPL and 152 of their male partners, and a control group of 56 fertile couples. We investigated mutations including FV Leiden, factor V H1299R, factor II prothrombin G20210A, F XIII V34L, β-fibrinogen −455G>A, plasminogen activator inhibitor-1, GPIIIa L33P (HPA-1 a/b L33P), MTHFR C677T, MTHFR A1298C, ACE I/D, Apo B R3500Q, and Apo E. Results Overall, heterozygous mutations of FV Leiden, FXIII V34L, GPIIIa L33P, Apo E4, and prothrombin G20210A and homozygous mutations of PAI-1and MTHFR C677T were associated with RPL. There was no meaningful association between RPL and other studied genes. Conclusion In contrast to the other mutations and polymorphisms, FV Leiden, FXIII V34L, GPIIIa L33P, Apo E, prothrombin G20210A, PAI-1 and MTHFR C677T gene mutations may help to identify the couples at risk for recurrent pregnancy loss.

WOS: 000263540800026, Familial Mediterranean fever (FMF) is the most frequent hereditary inflammatory disease characterized by self-limited recurrent attacks of fever and serositis. The aim of the current study is to determine the frequency of the mutations in 365 suspected FMF patients and to reveal whether there is a correlation between genotype and phenotype of these patients. All patients were clinically examined according to Tell-Hashomer FMF criteria and were screened genetically in terms of common 12 Mediterranean fever gene (MEFV) mutations. Various point mutations were detected in 270 (74%) patients. The most frequent mutation was M694V (26.85% of the alleles) and was followed by E148Q (15.55%), M680I (G/C) (9.62%) and V726A (7.96%). Patients who bear M694V homozygous mutation had most severe disease phenotype and high risk for amyloidosis (P = 0.04). Our results indicate that Sivas population has a wide range of heterozygous mutated carriers of MEFV gene and there is a high frequency of E148Q allele when compared to the other Mediterranean groups.

Bacterial strain typing is an important process for diagnosis, treatment and epidemiological investigations. Current bacterial strain typing methods may be classified into two main categories: phenotyping and genotyping. Phenotypic characters are the reflection of genetic contents. Genotyping, which refers discrimination of bacterial strains based on their genetic content, has recently become widely used for bacterial strain typing. The methods already used in genotyping of bacteria are quite different from each other. In this review we tried to summarize the basic principles of DNAbased methods used in genotyping of bacteria and describe some important DNA regions that are used in genotyping of bacteria. J Microbiol Infect Dis 2011;1(1):42-46.

WOS: 000298011800002, PubMed: 21897766, Objective: The purpose of this study was to evaluate the relationship between 5,10-methylenetetrahydrofolate reductase (MTHFR) polymorphisms and Attention Deficit Hyperactivity Disorder (ADHD) in a sample of Turkish children. Study Design: MTHFR gene polymorphisms were assessed in 40 patients with ADHD and 30 healty controls. Two mutations in the MTHFR gene were investigated using polymerase chain reactions and restriction fragment length polymorphisms. Results: Although there were no statistically significant differences in genotype distributions of the C677T alleles between the ADHD and the control groups (p=0,678) but the genotypic pattern of the distributions of the A1298C alleles was different between the ADHD patients and the controls (p=0,033). Conclusions: Preliminary data imply a possible relationship between A1298C MTHFR polymorphisms and the ADHD.

Growing evidences indicate that tumor regrowth and invasion refer a small subpopulation of tumor cells called cancer stem cells (CSCs). Autophagy is an essential catabolic process for cell homeostasis that involves in cell survival and death and also development of various human diseases including cancer. Here we aimed to investigate the effect of autphagy on the subpopulation of CD44 C /CD24 � /low phenotype MCF7 and Hep-2 cells. CD44 C /CD24 � /low cells were isolated from MCF7 and Hep-2 cell cultures by using fluorescence-activated cell sorting (FACS). The CD44 C /CD24 � /low and the original cells were separately treated with 20 mM of C2 ceramide for 24 hours in order to stimulation of autophagy. LC3 proteins aggregation was monitored to evaluate autophagy in the cells by using LC3 targeted immunofluorescence staining assay. Subpopulation of CD44 C /CD24 � /low cells in total cultures were investigated by flow cytometry analysis and eventually statistics were carried out. Results showed an enhanced aggregation of LC3 proteins in the cells and also CD44 C /CD24 � /low cells under effect of C2 ceramide. Moreover, the subpopulation ofCD44 C /CD24 � /low cells werefoundsignificantlylower in thecellscultures treated with C2 ceramide in comparison with negative controls. The results elucidated that CD44 C /CD24 � /low MCF7 and Hep-2 cells were susceptible to autophagy stimulatory reagent C2 ceramide. Additionally, our results suggest that CD44 C /CD24 � /low cell subpopulation in the cell cultures is reduced under autophagic condition. Therefore, we suppose that autophagy may has a negative impact on the breast and Larynx cancer stem cells self-renewal potency or expression of CD44 and CD24 cell surface markers, however further studies are needed to reveal the exact mechanism.