Unpleasant Souvenir: Imported Plasmodium falciparum Malaria in Türkiye.

Objective: Each year, approximately 125 million people visit malaria-endemic countries. This study aimed to investigate the clinical characteristics of imported Plasmodium falciparum malaria infections in Türkiye. Methods: The study included patients diagnosed with P. falciparum malaria between 1996 and 2022. A retrospective evaluation was conducted on whole blood samples and/or blood smears, as well as detailed medical histories, clinical manifestations, and laboratory findings. A total of 131 imported cases of P. falciparum were included in the study. Results: Among the patients, 121 were male. Of these, 101 had traveled to Africa, while 30 had visited Asia. Among the patients, 109 were returned travelers, and 22 were refugees/migrants. Early trophozoites were observed in all patients, while gametocytes were detected in 30 patients. Cerebral malaria developed in 15 patients, resulting in the death of two individuals. Additionally, 10 patients received preventive chemoprophylaxis. Conclusion: Turkey is situated on migration routes that connect two continents to Europe, where more than 95% of the global malaria burden exists. The importation of malaria through returned travelers poses a risk of malaria reintroduction in our country, given the presence of suitable vectors, climate conditions, and environmental factors. Importantly, 30 patients (22.9%) exhibited gametocyte forms of P. falciparum, which have the potential to infect Anopheles species, thus establishing a basis for local malaria transmission.

Investigation of Malaria, Leishmaniasis, and Scabies Risk after Earthquakes and Recommendations for Prevention.

This study examines the risk of malaria, leishmaniasis, and scabies following earthquakes in southeastern Türkiye. The focus is on the impact on the local population and Syrian immigrants. Recommendations for prevention include vector control measures such as indoor residual spraying and distribution of insecticidal nets. Surveillance and early detection through rapid diagnostic tests and active case finding are important. Public awareness campaigns and community engagement are crucial for promoting protective measures. Strengthening healthcare infrastructure, providing essential supplies, and capacity building is essential. For leishmaniasis, early detection and treatment, vector control, health education, and community engagement are vital. Scabies outbreaks affect the socioeconomically depressed local population and Syrian immigrants. Early detection, treatment, contact tracing, health education, hygiene promotion, and improved living conditions are necessary. Implementing these interventions and strategies can effectively prevent, control, and manage these diseases. Tailoring approaches to the specific context and needs of affected communities is crucial. By addressing these challenges, we can protect the health and well-being of the affected population.

Evaluation of Four Adult Visceral Leishmaniasis Cases.

Leishmania infantum is the species responsible for visceral leishmaniasis [(VL), kala-azar], which is observed sporadically mainly in pediatric age groups in the Aegean, Mediterranean and Central Anatolian regions of Türkiye. The aim of this study is to evaluate the diagnosis, clinic, laboratory results and treatments of four adult patients with VL who applied to our hospital. The patients were referred to our hospital to investigate hematological malignancy. In the study, the data of four patients (three men, one woman; age range: 30-40 years) who were diagnosed with VL and treated in the infectious diseases clinic of our hospital between January 2022 and April 2022 were evaluated retrospectively. The diagnosis of VL was made according to appropriate clinical and physical examination findings, biochemical and serological tests (indirect fluorescent antibody test and rK39 rapid antigen test) and polymerase chain reaction (PCR) results, as well as the presence of amastigote forms of the parasite in bone marrow samples. Serology positivity was found in all patients, and bone marrow positivity was found in two patients. According to the results of RT-PCR in all patients, it was determined that the species causing the disease was L. infantum/L. donovani. Initially, the most common symptoms were fever, fatigue, and abdominal distension. None of the patients had an immunosuppressive condition. It was understood that all the patients lived in the rural area of Syria's Idlib province. Hepatosplenomegaly, increased erythrocyte sedimentation rate, anemia, leukopenia and thrombocytopenia were found in all patients. The patients were treated with liposomal amphotericin-B (L-AMB). One patient did not come for follow-ups, the other three patients were found to have completely recovered in their follow-up. No recurrence was observed in any of the patients. In conclusion, VL should be considered in patients who apply to health institutions with complaints of fever, hepatosplenomegaly, increased erythrocyte sedimentation rate, anemia, leukopenia and thrombocytopenia.

Knock, knock, knocking on Europe's door: Threat of leishmaniasis in Europe with a focus on Turkey.

Leishmaniasis epidemiology is currently undergoing substantial transformations in both Turkey and Europe, signifying potential implications for public health. This review analyzes the evolving patterns within Turkey and their potential ramifications for Europe. Within Turkey, the dynamics of leishmaniasis are undergoing noteworthy alterations, manifesting in a rise in cutaneous leishmaniasis (CL) cases and the emergence of Leishmania major and Leishmania donovani. These transformations are predominantly driven by factors such as the distribution of vectors, human activities, climate fluctuations, and migration. Across Europe, particularly in countries within the Mediterranean basin, leishmaniasis is endemic, primarily attributed to Leishmania infantum. Recent evidence suggests a resurgence of the disease even in previously non-endemic areas, propelled by climate change, urbanization, and migration. The changing landscape of leishmaniasis in Turkey carries direct implications for Europe. The presence and distribution of Leishmania tropica, L. major, and L. donovani raise concerns regarding cross-border transmission. Turkey's strategic position along migration routes further compounds the risk, alongside the facilitative effects of climate change and host mobility. Embracing a One Health approach with public awareness campaigns should be a priority. To ensure the protection of public health in Europe, it is imperative to adopt a proactive approach by establishing robust surveillance mechanisms, implementing preventive measures, and cultivating collaboration with Turkey. The invaluable experience, strategic geographical location, and well-established infrastructure of Turkey make this collaboration crucial in effectively addressing the evolving dynamics of leishmaniasis and its potential impacts on Europe.

[Investigation of the Efficacy of Cinnamaldehyde, Cannabidiol and Eravacycline in a Malaria Model]. / Sitma Modelinde Cinnamaldehyde, Cannabidiol ve Eravacycline'in Etkinliginin Arastirilmasi.

In this study, it was aimed to investigate the antimalarial activity of cinnamaldehyde (CIN) and cannabidiol (CBD) which have shown various biological activities such as potent antimicrobial activity and eravacycline (ERA), a new generation tetracycline derivative, in an in vivo malaria model. The cytotoxic activities of the active substances were determined by the MTT method against L929 mouse fibroblasts and their antimalarial activity were determined by the four-day test in an in vivo mouse model. In this study, five groups were formed: the CIN group, the CBD group, the ERA group, the chloroquine group (CQ) and the untreated group (TAG). 2.5 x 107 parasites/mL of P.berghei-infected erythrocyte suspension was administered IP to all mice. The determined doses of active substances were given to the mice by oral gavage in accordance with the four-day test and the parasitemia status in the mice was controlled for 21 days with smear preparations made from the blood taken from the tail end of the mice. The IC50 values, which express the cytotoxic activity values of the active substances were determined as 27.55 µg/mL, 16.40 µM and 48.82 µg/mL for CIN, CBD and ERA, respectively. The mean parasitemia rate in untreated mice was 33% on day nine and all mice died on day 11. On the ninth day, when compared with the TAG group, no parasites were observed in the CIN group, while the average parasitemia was 0.08% in the CBD group and 17.8% in the ERA group. Compared to the mice in the TAG group, the life expectancy of the other groups was prolonged by eight days in the CIN group, 12 days in the CBD group and eight days in the ERA group. It has been determined that all three active subtances tested in this study suppressed the development of Plasmodium parasites in an in vivo mouse model and prolonged the life span of the mice. It is thought that the strong antimalarial activity of CIN and CBD shown in the study and the possible positive effect of ERA on the clinical course can be improved by combining them with the existing and potential antimalarial molecules.

[Comparison of Conventional Methods with Molecular Methods in the Diagnosis of Trichomonas vaginalis and Investigation of Metronidazole Resistance]. / Trichomonas vaginalis'in Tanisinda Konvansiyonel Yöntemlerin Moleküler Yöntemlerle Karsilastirilmasi ve Metronidazol Direncinin Arastirilmasi.

Trichomoniasis is a sexually transmitted parasitic infection caused by Trichomonas vaginalis. In the diagnosis of trichomoniasis, direct microscopy (DM) is preferred, which is a cheap and fast method, although it has low sensitivity. Culture methods, which are accepted as the gold standard, can only be applied in certain centers due to the need for experienced personnel and the ability to get results within 2-7 days, despite their high sensitivity. In this study, it was aimed to compare conventional microscopic and culture methods used in the routine diagnosis of T.vaginalis with polymerase chain reaction (PCR) method and to investigate ntr4 and/or ntr6 gene polymorphism in the nitroreductase gene region, which are thought to be associated with metronidazole resistance in T.vaginalis strains isolated from clinical specimens. Vaginal swab specimens were collected from the posterior fornix of the vagina with two sterile ecuvion sticks during the gynecological examinations of 200 patients who applied to the Balikesir University Health Practice and Research Hospital, Obstetrics and Gynecology Polyclinic between March 2019 and August 2021. The first swab sample was used for direct microscopic examination, Giemsa staining and conventional PCR analysis, while the second swab specimen was taken into trypticase-yeast-extract-maltose (TYM) medium for T.vaginalis culture and followed for eight days at 37 °C. All specimens were screened for the presence of T.vaginalis using primers specific to the ß-tubulin (btub1) gene region and clinical isolates grown in TYM medium were examined for metronidazole resistance using primers specific for the nitroreductase gene region by using conventional PCR. Drug resistance test was also performed for the isolates in which polymorphism associated with metronidazole resistance was detected. Eight (4%) of 200 patient specimens were found positive by both culture/staining and PCR methods. The mean age of the patients included in the study was 39.9, while the mean age of the patients with positive T.vaginalis was 41.8. The most common clinical findings in the patients were foul-smelling vaginal discharge (36%), groin pain (21%), vaginal itching (19%), and burning sensation during urination (18%). In three out of eight T.vaginalis strains isolated from clinical samples, the presence of polymorphism in the ntr6 gene, which is thought to be associated with metronidazole resistance, was demonstrated by PCR. It was observed that three isolates with ntr6 gene polymorphism were phenotypically resistant to metronidazole (MLK= 390 µM). In this study, the fact that three of eight clinical isolates that were resistant to metronidazole by the broth microdilution method and as well as showing ntr6 gene polymorphism supported the thesis that there might be a close relationship between metronidazole resistance and ntr6 gene polymorphism. As a result, the use of culture and molecular methods in the diagnosis of T.vaginalis, in addition to the microscopy method, may contribute to a more accurate laboratory diagnosis of the agent, to detect metronidazole resistance molecularly and phenotypically, to determine metronidazole resistance rates in our country and to update treatment protocols within the framework of these data.

[Autochthonous Case of Malaria Prediagnosed as Leukemia]. / Lösemi Ön Tanili Yerli Sitma Olgusu.

Malaria is a parasitic disease transmitted by infected female Anopheles mosquitoes. There are five species of Plasmodium species that can infect humans. Of these species, especially P.falciparum and P.vivax pose the greatest threat to human health. In the 2014 report of the World Health Organization, it was reported that there were no locally acquired cases of malaria in 16 countries including Türkiye. Malaria cases originating from outside the country and imported due to migration, travel and working abroad are reported as import cases. In this report, a case of non-imported malaria followed with a preliminary diagnosis of leukemia was presented. A 14-year-old female patient who was admitted to a health institution with complaints of high fever, headache, chills, nausea-vomiting, and diarrhea that had been going on for two weeks, was pre-diagnosed as leukemia and was referred to Manisa Celal Bayar University Faculty of Medicine, Hafsa Sultan Hospital, Department of Pediatric Hematology and after pancytopenia was detected in the complete blood count. The anamnesis of the patient revealed that she had no history of international travel and that she had been prescribed medications such as paracetamol, amoxicillin, and metoclopramide for flu-like complaints while working in the Southeastern Anatolia, Aegean, and Mediterranean Regions of Türkiye. Bone marrow aspiration was performed for the etiological examination of pancytopenia. Giemsa-stained blood smears, rapid diagnostics, and real-time quantative polymerase chain reaction (qRt-PCR) analyses were performed in the medical parasitology laboratory and malaria was suspected in both bone marrow and peripheral blood smears. P.vivax erythrocytic forms and gametocytes were present in abundance in smear preparations stained with Giemsa, and rapid diagnosis kit was positive for P.vivax. The strain was genotyped as P.vivax by qRt-PCR analysis. For the treatment of the patient, airalam (artemether + lumefantrine) tablets were provided with 2 x 4 daily posology for three days after the diagnosis, and primaquine was provided after one week of the diagnosis as 1 x 2 tablets (1 x 15 mg) for 14 days, and the patient was discharged without complications following the treatment regimen. The fight against malaria continues uninterruptedly since the establishment of the Republic of Türkiye. Tropical diseases, especially malaria, is of great importance for Türkiye due to numerous reasons such as its location in the subtropical region where Anopheles mosquitoes are capable of malaria transmission, it is situated at the crossroads on the migration routes between continents where human traffic is busy, there are many people who go abroad for work and most importantly rising temperatures due to climate change. For this reason, this case report is important to emphasize the importance of malaria for the country and to increase the awareness of clinicians and laboratories about malaria and the possibility of autochthonous malaria transmission in Türkiye.

Antiprotozoal activity of auranofin on Trypanosoma cruzi, Leishmania tropica and Toxoplasma gondii: in vitro and ex vivo study.

BACKGROUND: Three obligate intracellular protozoan parasite species, which are responsible for significant morbidity and mortality and settle in macrophage cells, affect more than one-half of the world's population, namely, Trypanosoma cruzi, Leishmania tropica and Toxoplasma gondii, which are causative agents of Chagas disease, leishmaniasis and toxoplasmosis, respectively. In the current study, it was aimed to investigate the in vitro and ex vivo antiprotozoal activity of auranofin on T. cruzi, L. tropica and T. gondii. METHODS: The in vitro drug efficacy (IC50) of auranofin was investigated by haemocytometry and the CellTiter-Glo assay methods and the ex vivo drug efficacy (IC50) by light microscopic examination of Giemsa-stained slides. Also, the cytotoxic activity (CC50) of auranofin was examined by the CellTiter-Glo assay. The selectivity index (SI) was calculated for auranofin. RESULTS: According to IC50, CC50 and SI data, auranofin did not exhibit cytotoxic activity on Vero cells, but exhibited antiprotozoal activity on epimastigotes and intracellular amastigotes of T. cruzi, promastigotes and intracellular amastigotes of L. tropica and intracellular tachyzoites of T. gondii (p<0.05). CONCLUSIONS: The detection antiprotozoal activity of auranofin on T. cruzi, L. tropica and T. gondii according to the IC50, CC50 and SI values is considered an important and promising development. This is significant because auranofin may be an effective alternative treatment for Chagas disease, leishmaniasis and toxoplasmosis in the future.

Visceral Leishmaniasis Caused by Leishmania Tropica.

PURPOSE: In Turkey, the main causative agent of visceral leishmaniasis (VL) is Leishmania. infantum and the main causative agent of cutaneous leishmaniasis (CL) is Leishmania tropica. In this study, we aimed to discuss the possible mechanisms, clinical aspects, and threat of visceralizing L. tropica. METHODS: This study includes seven cases of VL caused by L. tropica.Five patients were male (71%) and four were adults (57%). RESULTS: All the VL patients complained of fever and splenomegaly. Fatigue, pancytopenia, and hepatomegaly were present in six patients each (86%), while weight loss and gastrointestinal system (GIS) symptoms were present in 5 patients (71%). CONCLUSIONS: In this study, we have evaluated seven cases of visceralized L. tropica (VLT) in the context of the changing leishmaniasis epidemiology in Turkey. We have evaluated the possible mechanisms of visceralization; inter- and intraspecies genetic exchange with all the old world leishmaniasis agents present in the region, stress induced by inappropriate use of drugs, and possible ongoing adaptation mechanisms of Leishmania spp. The threat posed by VLT is significant as L. tropica is the most widespread and most common cause of leishmaniasis in Turkey. We do not know the vectorial capacity of the sand flies for the transmission of VLT strains or if these strains are in circulation in Turkey. Future studies should be carried out to investigate these issues as the transition of L. tropica from a mild disease-causing agent to a mortal one poses a significant public health concern for Turkey and Europe.

[Evaluation of Ex Vivo Cultivation Potentials of Trypanosoma cruzi, Leishmania tropica ve Toxoplasma gondii Parasites in J774, Vero and HeLa Cell Lines]. / Trypanosoma cruzi, Leishmania tropica ve Toxoplasma gondii Parazitlerinin J774, Vero ve HeLa Hücre Hatlarinda Ex Vivo Kültivasyon Potansiyellerinin Degerlendirilmesi.

Three obligate intracellular protozoan parasite species, namely Trypanosoma cruzi, Leishmania tropica and Toxoplasma gondii, causative agents of Chagas disease, Leishmaniasis and toxoplasmosis, respectively, which are responsible for significant morbidity and mortality and reside in macrophage cells, affect more than half of the world's population in connection with socio-economic and geographical factors and also causes neglected parasitic diseases of increasing importance. This study aimed to evaluate the ex vivo cultivation potential of T.cruzi, L.tropica and T.gondii parasites in J774, Vero and HeLa cells and to reproduce in a short time and in large amounts without losing their virulence properties. Ex vivo experimental models were created by infecting J774, Vero and HeLa cell lines confluently produced in cell culture flasks with T.cruzi, L.tropica and T.gondii parasites. In ex vivo cultivation, one passage was applied for seven days and three times in a row. Cells removed from the surface after each passage were plated on eight-well chamber slides. Giemsa stained slides were prepared and infection rates were evaluated by light microscopic examination. At the end of the study, it was observed that all three cell lines could be infected with T.cruzi, L.tropica and T.gondii parasites, and infection rates increased in all cell lines after consecutive passages. As a result of ex vivo cultivation, the best cell lines from which T.cruzi and L.tropica strains grew, were J774, Vero and HeLa, and HeLa, J774 and Vero cell lines for T.gondii strain, respectively (p<0.05). Trypanosoma cruzi, L.tropica and T.gondii parasites were successfully grown in J774, Vero and HeLa cell lines by ex vivo culture method in a short time and in large amounts without losing their virulence properties. Cell lines with the best ex vivo cultivation potential for T.cruzi and L.tropica parasites were J774, Vero and HeLa, respectively, while HeLa, J774 and Vero for T.gondii. It is thought that the data obtained in this regard will contribute to many studies on the development of vaccines, drugs and new diagnostic kits.

Design, synthesis, in vitro - In vivo biological evaluation of novel thiazolopyrimidine compounds as antileishmanial agent with PTR1 inhibition.

The leishmaniasis are a group of vector-borne diseases caused by a protozoan parasite from the genus Leishmania. In this study, a series of thiazolopyrimidine derivatives were designed and synthesized as novel antileishmanial agents with LmPTR1 inhibitory activity. The final compounds were evaluated for their in vitro antipromastigote activity, LmPTR1 and hDHFR enzyme inhibitory activities, and cytotoxicity on RAW264.7 and L929 cell lines. Based on the bioactivity results, three compounds, namely L24f, L24h and L25c, were selected for evaluation of their in vivo efficacy on CL and VL models in BALB/c mice. Among them, two promising compounds, L24h and L25c, showed in vitro antipromastigote activity against L. tropica with the IC50 values of 0.04 µg/ml and 6.68 µg/ml; against L. infantum with the IC50 values of 0.042 µg/ml and 6.77 µg/ml, respectively. Moreover, the title compounds were found to have low in vitro cytotoxicity on L929 and RAW264.7 cell lines with the IC50 14.08 µg/ml and 21.03 µg/ml, and IC50 15.02 µg/ml and 8.75 µg/ml, respectively. LmPTR1 enzyme inhibitory activity of these compounds was determined as 257.40 µg/ml and 59.12 µg/ml and their selectivity index (SI) over hDHFR was reported as 42.62 and 7.02, respectively. In vivo studies presented that L24h and L25c have a significant antileishmanial activity against footpad lesion development of CL and at weight measurement of VL group in comparison to the reference compound, Glucantime®. Also, docking studies were carried out with selected compounds and other potential Leishmania targets to detect the putative targets of the title compounds. Taken together, all these findings provide an important novel lead structure for the antileishmanial drug development.

Leishmania kinetoplast DNA contributes to parasite burden in infected macrophages: Critical role of the cGAS-STING-TBK1 signaling pathway in macrophage parasitemia.

Leishmania parasites harbor a unique network of circular DNA known as kinetoplast DNA (kDNA). The role of kDNA in leishmania infections is poorly understood. Herein, we show that kDNA delivery to the cytosol of Leishmania major infected THP-1 macrophages provoked increased parasite loads when compared to untreated cells, hinting at the involvement of cytosolic DNA sensors in facilitating parasite evasion from the immune system. Parasite proliferation was significantly hindered in cGAS- STING- and TBK-1 knockout THP-1 macrophages when compared to wild type cells. Nanostring nCounter gene expression analysis on L. major infected wild type versus knockout cells revealed that some of the most upregulated genes including, Granulysin (GNLY), Chitotriosidase-1 (CHIT1), Sialomucin core protein 24 (CD164), SLAM Family Member 7 (SLAMF7), insulin-like growth factor receptor 2 (IGF2R) and apolipoprotein E (APOE) were identical in infected cGAS and TBK1 knockout cells, implying their involvement in parasite control. Amlexanox treatment (a TBK1 inhibitor) of L. major infected wild type cells inhibited both the percentage and the parasite load of infected THP-1 cells and delayed footpad swelling in parasite infected mice. Collectively, these results suggest that leishmania parasites might hijack the cGAS-STING-TBK1 signaling pathway to their own advantage and the TBK1 inhibitor amlexanox could be of interest as a candidate drug in treatment of cutaneous leishmaniasis.

Could RPMI-1640 Medium be Used in the Diagnosis and Isolation of Cutaneous Leishmaniasis Lacking NNN Medium?

Laboratory diagnosis of leishmaniasis is based on culture, microscopic examination, serological and molecular methods. The gold standard method is to see amastigotes in microscopic examination and to grow promastigotes in Novy, MacNeal, Nicolle (NNN) medium. NNN medium is frequently used for culture all over the world. In our study, it was aimed to investigate whether the use of RPMI-1640 medium is an appropriate method in cases where the gold standard NNN medium is not available for the diagnosis of cutaneous leishmaniasis (CL). Smears were prepared from the needle aspiration fluid sample from the patient who applied to Manisa Celal Bayar University Faculty of Medicine and had lesions suspicious of CL, and were stained with Giemsa for the presence of amastigotes. The samples taken were directly inoculated into RPMI-1640 broth and incubated at 26 °C for the presence of promastigotes. On consecutive days after incubation, it was checked for promastigote growth. Genotyping of the grown isolate was performed with primers and probes specific to the internal transcribed spacer-1 (ITS-1) gene region with the help of real-time polymerase chain reaction. The amastigote form was observed in the microscopic examination of the needle aspiration fluid sample smear preparations taken from the patient. On the other hand, promastigote growth was observed in RPMI-1640 broth from the 3rd day. In addition, the isolate obtained from the CL patient was determined to be Leishmania tropica as a result of the species determination made by genotyping. It is thought that this study is important in terms of suggesting an alternative medium for the diagnosis of leishmaniasis in laboratories where the gold standard NNN medium is easily accessible. RPMI-1640 medium, which is easily obtained and prepared in parasitology laboratories, can help in the diagnosis of the disease and treatment follow-up, Leishmania spp. isolation and drug resistance studies.

Origanum Sipyleum Methanol Extract in Combination with Ponatinib Shows Synergistic anti-Leukemic Activities on Chronic Myeloid Leukemia Cells.

Origanum sipyleum is used in folk medicine due to its anti-inflammatory, antimicrobial, and antioxidant properties. Ponatinib, an effective tyrosine kinase inhibitor in the treatment of chronic myeloid leukemia (CML), has severe side effects. Thus, we aimed to determine a novel herbal combination therapy that might not only increase the anti-leukemic efficacy but also reduce the dose of ponatinib in targeting CML cells. Origanum sipyleum was extracted with methanol (OSM), and secondary metabolites were determined by phytochemical screening tests. The cytotoxic effects of OSM on K562 cells were measured by WST-1 assay. Median-effect equation was used to analyze the combination of ponatinib and OSM (p-OSM). Apoptosis, proliferation, and cell-cycle were investigated by flow-cytometry. Cell-cycle-related gene expressions were evaluated by qRT-PCR. OSM that contains terpenoids, flavonoids, tannins, and anthracenes exhibited cytotoxic effects on K562 cells. The median-effect of p-OSM was found as synergistic; OSM reduced the ponatinib dose ∼5-fold. p-OSM elevated the apoptotic and anti-proliferative activity of ponatinib. Consistently, p-OSM blocked cell-cycle progression in G0/G1, S phases accompanied by regulations in TGFB2, ATR, PP2A, p18, CCND1, CCND2, and CCNA1 expressions. OSM enhanced the anti-leukemic activity of ponatinib synergistically via inducing apoptosis, suppressing proliferation, and cell-cycle. As a result, OSM might offer a potential strategy for treating patients with CML.

Increasing the Sensitivity of Leishmania RNA Virus 2 (LRV2) Detection with a Modification in cDNA Synthesis

Objective: Leishmania RNA virus was detected the first time in the New World Leishmania species. Recent studies were also showed the presence of Leishmania RNA virus 2 (LRV2) in Old Word Leishmania species including Turkish L. major and L. tropica isolates. This study aimed to increase the sensitivity of qPCR with a modification in the denaturation step of cDNA preparation protocol. Methods: In this study, LRV2+ three L. major, two L. tropica strains and L. major control strain (MHOM/SU/73/5-ASKH) were included. Total RNA isolation was done using different numbers of Leishmania promastigotes (108, 105 and 103). Before cDNA synthesis, samples were denatured at 95 °C for 2 min, as a modification of the kit procedure. qPCR was undertaken using 0.5 mM primers (LRV F-HR/LRV R-HR) diluted in SYBR Green Master mix. Results: We observed lower Ct values in amplicons with the modified version than with the classical kit protocol for cDNA synthesis, in all of the strains used in the study. The addition of pre-denaturation step at 95 °C showed lower Ct values meaning the sensitivity increased. Different parasite dilutions showed similar results. Conclusion: It is important to increase the sensitivity especially with the aim for detecting LRV in clinical samples obtained from patients probably have less number of parasites. The presence and burden of the virus can help to understand the relationship between the clinical findings and the pathogenicity of the parasite which may lead to changes in the course of treatment.

Investigation of in vitro Efficacy of Miltefosine on Chronic Cutaneous Leishmaniasis

Objective: Leishmaniasis is the second deadliest parasitic disease in the World Health Organisation's list of neglected diseases, following malaria. Cutaneous leishmaniasis (CL) is the most common form of the disease and it is one of the few communicable diseases with increasing incidence rates owing to factors like armed conflicts and climate change. CL can be divided into two major groups: Acute CL (ACL) and chronic CL (CCL). The aim of this study was to compare the in vitro efficacy of miltefosine and pentavalent antimony compounds in the CCL patient samples. Methods: Five isolates previously isolated from 5 CCL patients were included in this study. Genotyping is performed using internal transcribed spacer 1 (ITS 1) gene region real-time PCR. In vitro drug efficacy tests were applied to determine their activity against meglumine antimoniate (MA) and miltefosine. Serial dilutions (512, 256, 128, 64, 32, 16, 8 and 4 µg/mL) prepared from MA and miltefosine were prepared in 96-well flat-bottom cell culture plates and incubated at 24 °C for 48 hours. The efficacy of the drug on Leishmania spp. promastigotes after 24 and 48 hours was evaluated by hemocytometer slide and XTT cell viability test. Results: All of the samples were genotyped as L. tropica. Evaluation of 24 and 48 hours showed, 128 µg/mL and 256 µg/mL and 32 µg/mL and 64 µg/mL concentrations of miltefosine and MA were enough to kill all the promastigotes respectively. The results of the hemocytometer slide and XTT were consistent. Conclusion: There are no studies investigating the in vitro efficacy of miltefosine with the CCL patient group. To overcome the treatment challenges experienced in this special patient group, more studies are needed. According to our results, it is concluded that miltefosine is efficient for the treatment of CCL and further clinical studies with miltefosine will reveal valuable data.

[Investigation of the Anti-Leishmanial Effects of Prangos ferulacea and Ferula orientalis Extracts Collected from Sirnak Province Against Leishmania tropica Isolated in Turkey]. / Sirnak Ili ve Çevresinden Toplanan Prangos ferulacea ve Ferula orientalis Ekstrelerinin Türkiye'den Izole Edilmis Leishmania tropica'ya Karsi Anti-Leishmanial Etkilerinin Arastirilmasi.

Leishmaniasis is a vector-borne disease that is caused by the protozoa of Leishmania genus. Leishmaniasis is endemic in tropical, subtropical, and large areas of the Mediterranean basin, and covers a total of 98 countries worldwide. It is estimated, according to the World Health Organization (WHO) data, that approximately 350 million people are at risk in these areas, and approximately 12 million people are infected. Increased drug resistance has been documented lately, in the treatment of leishmaniasis which causes almost 1.2 million new cases annually. Thus, interest in plant-derived active substances has increased in recent years, and new anti-leishmanial agents are investigated with in vitro studies. The aim of the present study was to investigate the anti-leishmanial effects of Prangos ferulacea and Ferula orientalis plant extracts collected from the rural areas of Sirnak province against Leishmania tropica. The water, chloroform, and ethanol extracts of the roots, stems, and fruits of P.ferulaceae and F.orientalis plants were obtained, and the cytotoxic activity tests of the extracts were performed. L.tropica isolate obtained from the Parasite Bank in Manisa Celal Bayar University in Turkey (MHOM/TR/2012/CBCL-LT) was grown on NNN and RPMI 1640 broth medium. The cytotoxicity of each extract on the L.tropica isolate was evaluated with the XTT test. Amphotericin B (AmpB) was used as the positive control, and the IC50 values were determined. The lowest IC50 values of the plant extracts were found to be as follows: P.ferulaceae root chloroform extract 36 µg/ml and fruit chloroform extract 20 µg/ml, F.orientalis root ethanol extract 2.5 µg/ml, and fruit ethanol extract 48 µg/ml, stem chloroform extract 24 µg/ml, and fruit chloroform extract 3.1 µg/ml. It was also determined in our study that only P.ferulaceae root ethanol extract showed cytotoxic activity on the WI-38 fetal lung fibroblast cell line at 65.19 µg/ml at 72 hours. This is the first study that assessed the anti-leishmanial activities of P.ferulaceae and F.orientalis plants that grow in high altitude areas of our country. It was determined that P.ferulaceae root ethanol extract and fruit chloroform extract had the lowest IC50 values among the 18 plant extracts that we examined for their anti-leishmanial activities. The outcomes of this study will be useful in further studies for the determination of active compounds in P.ferulaceae and F.orientalis plant extracts.

Growth of Trichomonas vaginalis in Basic Media Available in Routine Microbiology Laboratories

Objective: Trichomonas vaginalis is a protozoan parasite with unicellular, flagellate, and anaerobic metabolism. It is the second most prevalent pathogen among sexually transmitted agents after viruses. Microscopic examination, culture, and molecular methods are used in the laboratory diagnosis of T. vaginalis. However, in most routine microbiology laboratories, microscopy is preferred instead of culture and molecular methods for T. vaginalis diagnosis because microscopy is cheaper than other methods. This study aimed to produce T. vaginalis in media that can be detected frequently in microbiology laboratories. Methods: In this study, four media, namely, thioglucholate medium (THIO), brain heart infusion medium (BHI), tryptic soy broth medium (TSB), and Brucella broth medium (BRB) were modified and tested. Trypticase-yeast extract-maltose (TYM) medium was used as a reference medium. Each medium tested was enriched with three different serum additives. T. vaginalis trophozoite at a density of 104 parasites/mL was inoculated into each medium and incubated at 37 °C for 10 days. We determined the number of trophozoites using a hemocytometer, and the viability rates were determined using trypan blue. Results: Trichomonas vaginalis grew extremely well on THIO, BHI, and TSB media but not on BRB media. The time and number of parasites peaked were determined as 100×104 parasites/mL on THIO-ATS and THIO-FCS media on days five and four, respectively, 100×104 parasites/mL on BHI-ATS on day three, 98×104 parasite/mL on BHI-FCS media on day five, 100×104 parasites/mL on TSB-ATS on day four, and 82×104 parasite/mL on TSB-FCS media on day seven. Compared with the reference medium, TYM, T. vaginalis trophozoites survived significantly longer in THIO, BHI, and TSB media. Conclusion: The rich content of THIO, TSB, and BHI media, which are widely available in routine microbiology laboratories, may allow T. vaginalis growth.

Autochthonous transmission of Leishmania donovani and Leishmania major with all the components of infection cycle at Europe's doorstep.

OBJECTIVES: Leishmaniasis is a vector-borne disease and dogs may act as urban reservoirs. Turkey and most of the Mediterranean basin countries are endemic for leishmaniasis. In this study, it is aimed to report the autochthonous leishmaniasis cases, with all the components of the infection cycle (reservoir, vector, and the host) in a region close to Europe. METHODS: Nine human and four canine autochthonous leishmaniasis cases were included in the study. Direct microscopy, culture methods, serological, and molecular tests were applied to the samples obtained from the cases. RESULTS: VL and CL patients consisted of 2 L.infantum, 1 L. donovani, 2 L. tropica, and 2 L. tropica,1 L. major,1 L. infantum infected patients respectively. CanL cases were infected with L. infantum, L. donovani, L. tropica, and L. major. CONCLUSIONS: All the cases were autochthonous cases located in Manisa province. As Greece and all the Mediterranean basin countries in Europe share competent vectors, it is concluded that the detection of all 4 species of Leishmania parasites in such proximity to Europe poses an important public health threat for Europe. This study reports all four species of Leishmania spp., including L. major and L.donovani in close proximity to continental Europe.