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Leishmaniasis is an infectious disease in which different clinical manifestations are classified into three primary forms: visceral, cutaneous and mucocutaneous. These disease forms are associated with parasite species of the protozoan genus Leishmania. For instance, Leishmania infantum and Leishmania tropica are typically linked with visceral (VL) and cutaneous (CL) leishmaniasis, respectively; however, these two species can also cause other form to a lesser extent. What is more alarming is this characteristic, which threatens current medical diagnosis and treatment, is started to be acquired by other species. Our purpose was to address this issue; therefore, gel-based and gel-free proteomic analyses were carried out on the species L. infantum to determine the proteins differentiating between the parasites caused VL and CL. In addition, L. tropica parasites representing the typical cases for CL were included. According to our results, electrophoresis gels of parasites caused to VL were distinguishable regarding the repetitive down-regulation on some specific locations. In addition, a distinct spot of an antioxidant enzyme, superoxide dismutase, was shown up only on the gels of CL samples regardless of the species. In the gel-free approach, 37 proteins that were verified with a second database search using a different search engine, were recognized from the comparison between VL and CL samples. Among them, 31 proteins for the CL group and six proteins for the VL group were determined differentially abundant. Two proteins from the gel-based analysis, pyruvate kinase and succinyl-coA:3-ketoacid-coenzyme A transferase analysis were encountered in the protein list of the CL group.

WOS:000555872700008 PMID: 32755520 Dünya Sağlık Örgütü, yaklaşık bir milyar insanın endemik bölgelerde risk altında olduğunu, son beş yıl içinde bir milyon kutanöz leyşmanyazis (KL) olgusunun ve yılda yaklaşık 300.000 viseral leyşmanyazis (VL) olgusunun olduğunu bildirmektedir. Her yıl yaklaşık 20.000 kişinin VL’ye bağlı öldüğü bilinmektedir. Türkiye’de Leishmania tropica’nın ve Leishmania infantum’un neden olduğu KL’de yılda 2500 civarında olgu bildirilmektedir. Başta Akdeniz ve Ege Bölgesi illerinde olmak üzere diğer birçok ilde son yıllarda ortaya çıkan olgu ve odaklarda önemli oranda artış görülmesi önümüzdeki yıllarda enfeksiyon hızının yükseleceğini göstermektedir. Ülkemizdeki KL’nin ana etkeni L.tropica olup tedavide meglumin antimonat kullanılmaktadır. Bu çalışmada, antimona dirençli ve dirençli olmayan L.tropica izolatlarının gen ve protein ekspresyonları karşılaştırılarak L.tropica’ya özgü antimon direnç genlerinin saptanması amaçlanmıştır. Ülkemizin Ege, Akdeniz ve Güneydoğu bölgelerinden antimonat direnci bulunmayan 3 KL hastasından elde edilmiş L.tropica izolatlarında, laboratuvar ortamında meglumin antimonata karşı 3 dirençli izolat geliştirilmiştir. Bu izolatların mikroarray yöntemi ile gen ekspresyon değişimleri, 2 boyutlu jel elektroforezi ile protein profilleri ve MALDI-TOF/TOF MS ile ilgili proteinleri tanımlanarak birbirleriyle karşılaştırma yapılmıştır. Antimon tedavisine yanıt vermemiş 10 KL hastasından elde edilmiş L.tropica izolatlarına antimon bileşiklerine yönelik direnç testleri uygulanmış ve direnç gelişiminden sorumlu genlerin ekspresyonlarını saptamak amacıyla kantitatif gerçek zamanlı polimeraz zincir reaksiyonu uygulanmıştır. Ayrıca, protein profilleri karşılaştırılarak antimon direnci olan ve olmayan izolatlardaki protein ekspresyon düzeylerindeki farklılıklar belirlenmiş ve farklılık saptanan proteinlerin tanımlanması gerçekleştirilmiştir. Bu çalışmalar sonucunda, L.tropica izolatlarının antimon bileşiklerine karşı direnç geliştirilen izolatlarında, direnç geliştirmesinde enolaz, “Elongation factor-2 (EF-2)”, “Heat shock protein 70 (HSP 70)”, tripanotyon redüktaz, protein kinaz C ve metalo-peptidaz proteinlerinin rol oynadığı saptanmış ve hastalardan alınan doğal dirençli izolatlarda da benzer ekspresyon değişimi gösterilmiştir. Sonuç olarak, ülkemizdeki L.tropica izolatlarının deneysel olarak çok kısa sürede meglumin antimonata (Glucantime®) karşı direnç kazandığı saptanmıştır. Ülkemizde yaşayan ve yurt dışından ülkemize giriş yapan KL hastalarının yetersiz ve eksik tedavi görmesi durumunda, dirençli suşların ve olgu sayısının hızla artabileceği ve dirençli leyşmanyazis odaklarının oluşabileceği öngörülmektedir. 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. L.tropica 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.

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/TOFMS 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 L.infantum/tropica were also shown to be present.

Objective: Visceral Leishmaniasis (VL) is a widespread infectious disease of the reticuloendothelial system which is mostly caused by Leishmania infantum and L.donovani. VL is a zoonotic infection and dogs act as the main reservoirs of the disease while the vectors are the female Phlebotomus, that is to say, sandflies. Although Leishmania spp. are attributed to reproduce asexually, the genetic exchange between different strains has recently been demonstrated and causes interspecific hybrids. We aimed to present the complete infection cycle of a hybrid strain of Leishmania infantum/donovani with the host, reservoir, and vector. Materials and Methods: A patient presenting with VL proved to have a dog with canine leishmaniasis. Bone marrow aspirate from the patient and lymph node aspirates from the dog were inspected for Leishmania spp. amastigotes under light microscopy. Samples were cultured in enriched NNN medium. Serological examinations were made using IFAT for the patient serum and rK39 test for both human and dog samples. Light traps were placed to collect Phlebotomus. Leishmania molecular identification was executed by primers and probes specific to ITS-1 and cytochrome B gene region of Leishmania parasites. Results: Leishmania spp. amastigotes were detected in the slides while promastigotes were present in the enriched NNN medium. For patient serum, IFAT IgG was 1/256 positive, and rK39 was positive for both dog and human samples. Fifty-two Phlebotomus were caught (24 male, 28 female): 35 of them were P. similis, ten were P. neglectus/syriacus, six were P .papatasi, and one was P. tobbi. Molecular analysis showed that the Leishmania spp. isolates from the patient, dog, and P.tobbi were identical. Gel image revealed two bands in all three samples, thus leading to the conclusion that the infecting strain was a hybrid of L. infantum and L. donovani. Conclusion: We found identical isolates that are putative hybrids of L. donovani and L. infantum in human, dog reservoir and sandfly.. Keywords: Leishmaniasis, Phlebotomus, Hybrid, Epidemiology, Migration, Turkey

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 theFive 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.All of the samples were genotyped asThere are no studies investigating theLeishmaniasis, Dünya Sağlık Örgütü’nün ihmal edilen hastalıklar sıralamasında sıtmadan sonra dünyada en fazla ölüme sebep olan ikinci paraziter hastalıktır. Kutanöz leishmaniasis (KL), Leishmaniasis’in en yaygın görülen formu olup, çatışma ve çevresel faktörlerden dolayı, insidansı artmakta olan az sayıdaki bulaşıcı hastalıklardan biri olarak bilinmektedir. KL iki ana gruba ayrılabilir: Akut KL (AKL) ve kronik KL (KKL). Bu çalışmada, oral yol ile kullanılan bir alkilfosfolipid analoğu olan miltefosinin, standart tedavide kullanılan beş değerli antimon bileşiklerine göre tedavi etkinliğininÇalışmada, daha önce 5 KKL hastasından elde edilen hasta örnekleri kullanılmıştır. Bu izolatlara internal transcribed spacer-1 problu gerçek zamanlı polimeraz zincir reaksiyonu testi uygulanarak genotipleme yapılmıştır. Genotipleme sonrası, meglumin antimoniat (MA) ve miltefosine karşı etkinliklerini değerlendirmek amacıylaTüm örneklerLiteratürde, KKL hasta örneklerinde miltefosin etkinliğinin değerlendirildiği bir çalışma bulunmamaktadır. Bu özel hasta grubunda karşılaşılan tedavi zorluklarının giderilmesi için daha fazla çalışma yapılması gerekmektedir. Elde ettiğimiz sonuçlara göre, miltefosinin KKL tedavisinde etkili bir ajan olduğu görülmüş ve miltefosin ile yapılacak klinik çalışmaların değerli veriler sunacağı sonucuna varılmıştır.

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 Şırnak 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.

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 autoch-thonous 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., Scientific Research Pro-jects Office of Manisa Celal Bayar University [2010-057, 2013-002, 2014-010, 2015-090], This study was supported by the Scientific Research Pro-jects Office of Manisa Celal Bayar University (No: 2010-057; 2013-002; 2014-010 and 2015-090) .

Leishmaniasis is an infectious disease in which different clinical manifestations are classified into three primary forms: visceral, cutaneous and mucocutaneous. These disease forms are associated with parasite species of the protozoan genus Leishmania. For instance, Leishmania infantum and Leishmania tropica are typically linked with visceral (VL) and cutaneous (CL) leishmaniasis, respectively; however, these two species can also cause other form to a lesser extent. What is more alarming is this characteristic, which threatens current medical diagnosis and treatment, is started to be acquired by other species. Our purpose was to address this issue; therefore, gel-based and gel-free proteomic analyses were carried out on the species L. infantum to determine the proteins differentiating between the parasites caused VL and CL. In addition, L. tropica parasites representing the typical cases for CL were included. According to our results, electrophoresis gels of parasites caused to VL were distinguishable regarding the repetitive down-regulation on some specific locations. In addition, a distinct spot of an antioxidant enzyme, superoxide dismutase, was shown up only on the gels of CL samples regardless of the species. In the gel-free approach, 37 proteins that were verified with a second database search using a different search engine, were recognized from the comparison between VL and CL samples. Among them, 31 proteins for the CL group and six proteins for the VL group were determined differentially abundant. Two proteins from the gel-based analysis, pyruvate kinase and succinyl-coA:3-ketoacid-coenzyme A transferase analysis were encountered in the protein list of the CL group.

Microscopic methods are accepted as the gold standard in the diagnosis of malaria and in the followup of treatment. However, as the microscopical methods require experienced personnel, it is important to confirm the diagnosis with a different method for accurate diagnosis and treatment follow-up. In our study, we aimed to investigate the utility of the use of real time reverse transcriptase polymerase chain reaction (rRT-PCR), as well as microscopic methods for malaria treatment follow-up. In our study, we formed five groups each consisting of five male Balb/c mice. Each mouse was injected intraperitoneally with 107/ml Plasmodium berghei parasites. After 48 hours following the injection, the mice in the first, second and third groups received 50 mg/kg/day of chloroquine treatment for one, two and three days, respectively. The fourth group was not treated and the fifth group of mice received saline for three days. The parasitemia was monitored for 21 days by blood smears prepared from the end of tail of the mice and searching the presence of the target gene region of the parasite by rRT-PCR. Both the blood smears and rRT-PCR results were positive for groups I, II, IV and V. Both blood smears and rRT-PCR results of mice in groups other than the third group were found to be positive. Blood smears of the mice in third group were found to be positive on the 5th and 7th days of the infection, and the subsequent preparations were evaluated as negative. rRT-PCR results showed positivity on day seven, but no presence of the target gene region of the parasite was detected on the other days. The comparison of microscopy and rRT-PCR methods, had shown parallel results. Apart from the microscopic examination method, it was concluded that the rRT-PCR method is important in the diagnosis of malaria and in the follow-up of the patient during the treatment process, and that different methods that support each other should be used.

Purpose Due to mass population movements driven by internal conflicts and wars, cutaneous leishmaniasis (CL) is becoming increasingly important in Turkey. This study is aimed at determining the clinical aspects, diagnosis and genotyping of CL patients coming to Turkey from abroad. Methods in our study, the clinical materials obtained from the patients or sent for diagnostic purposes from other centers to our laboratory between years 2012 and 2016 were assessed retrospectively. in total, there were 38 patients from Syria, Iraq, Afghanistan, Iran, and Turkmenistan. Results 29 (76%), 28 (73%) and 33 (87%) samples were positive by light microscopy, Novy-McNeal-Nicolle(NNN), and enriched medium, respectively. By ITS-1 gene region PCR, 31 (81%) of the cases were positive. 35 of the patients were tested positive by at least one of the diagnostic methods. By genotyping, 21 Leishmania tropica, 8 Leishmania major, 3 Leismania infantum, 2 Leishmania donovani, and 1 Leishmania aethopica were detected. Conclusion This study is aimed at informing the clinicians working in the field for the import CL cases and recording the changing epidemiological features of CL in the region as well as discussing the possible focus for L. aethiopica infection which has not been shown in the region before.

Plasmodium falciparum malaria causes about 450.000 deaths every year, mostly in children around the world. The infection is seen in cases coming from abroad and may lead to deaths in Turkey. Many native P.falciparum malaria cases and deaths due to this infection were observed in Turkey during mid 1900's when malaria was epidemic. But only two native cases were reported in the last 50 years, both from Manisa. First case was a one-year old baby who has come to Manisa from Urfa with his family and has never been abroad. He has diagnosed with Plasmodium vivax malaria and treated with chloroquine and primaquine. A previously obtained thin blood film was examined and characteristic P.falciparum rings in red blood cells were observed and the case was published together with photographs as probable P.falciparum and P.vivax mixed infection. After this case, microscopists working in Malaria Control Unit of Manisa were informed about the differentiation of malaria species in thin blood samples. Soon afterward, another case who have never been abroad before were also diagnosed with P.falciparum and P.vivax mixed infection and this case was also published with photographs taken from thin blood samples. As molecular diagnostic methods were not improved and widespread in those years, it could not be applied in both cases. A Giemsa stained thin blood sample of the baby case was incidentally found 22 years afterwards and with the aim of molecular diagnosis, the blood sample on the slide previously processed for DNA isolation, then analysed with "FTD Malaria Differentiation (Fast Track Diagnostics, Luxembourg)" multiplex kit with real-time polymerase chain reaction by using probes special for P.falciparum, P.ovale, P.malariae, P.vivax species. DNA's belonging to P.falciparum and P.vivax were found to be positive, the case is molecularly proved to have P.falciparum and P.vivax mixed infection. This case indicated that Turkey is convenient for the expansion of P.falciparum malaria in terms of the climate and vectors and suggested that the potential danger may increase with the effects of global warming, wars and migrations and may jump to Europe over Turkey. The case which molecularly proved the existence of native P.falciparum malaria in the near future in Turkey, was presented to draw attention to the danger of this infection for Turkey and Europe.

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. L.tropica 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.

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/TOFMS 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 L.infantum/tropica were also shown to be present.

Cutaneous Leishmaniasis (CL) is the most common form of leishmaniasis. CL can be divided into two major groups: acute CL (ACL) and chronic CL (CCL). The aim of this study is to compare the efficacy of miltefosin and pentavalent antimony compounds in vivo with the CCL patient samples.Three study groups were formed, each consisting of five male Mus musculus (Balb/C) mice. In this model, promastigotes from the culture of a CCL patient were utilized. 100 μL L. tropica promastigote suspension with a density of 10The footpad measurements of the miltefosine group were lower than the control group statistically. Between the MA group and the miltefosine group and MA group and the control group, there was no statistically significant difference. Giemsa stained slides revealed amastigotes in one, two and all of the slides for the miltefosine, MA and control group, respectively. Molecular tests were performed with the Rotor-Gene device and L. tropica consistent peaks were obtained in one of the miltefosine group, four in the MA group and all mice in the control group.Demonstration of both clinical and laboratory improvement in four of the five experimental animals provides strong evidence that miltefosine is an effective drug in the treatment of CCL. In the literature, no clinical or laboratory studies using miltefosine have been performed with CCL patients only.

Human African trypanosomiasis, also known as sleeping sickness, is a parasitic disease in which Glossina is transmitted by human intervention andIn this study,As a result of this study, it has been concluded that the best medium, to produce abundantlyThis study is one of the first studies related to the production of

Objective This study aims to determine the optimum liquid medium for the reproduction of Trypanosoma cruzi strains and provide cryopreservation. Methods The reproduction density of T. cruzi strain was evaluated in the following five different commercial liquid culture media: RPMI 1640, Medium 199 (M199), Schneider's Insect Medium (SIM), Nutrient Broth (NB), Brain Heart Infusion Broth (BHIB). Cultures were monitored on every other day for a period of 24 days. Cryopreservation of T. cruzi was also performed and viability was tested after six months. Results Epimastigotes of T. cruzi were not found to be produced in NB and BHIB media. Significant difference was not observed among the reproduction potential of RPMI-1640, M199, and SIM after evaluating the data for the first 10 days. Between days 12 and 24, RPMI-1640 was found to be the best reproduction medium. From the 18th day onwards, parasites transformed amastigotes. On the 24th day, the highest level of amastigote amount was observed, and reproduction was determined to have stopped. As a result of cryopreservation, it was determined that the survival of T.cruzi continued after six months.. Conclusion Thus, the selection of RPMI-1640 medium, followed by M199 and SIM media would be appropriate when studying T. cruzi epimastigotes. Studies using epimastigotes should be planned for up to 18 days and for those using amastigotes, it would be appropriate to plan the studies after the 18th day. Moreover, T. cruzi can be cryopreserved with 15% DMSO and stored for a long time in liquid nitrogen.

Leishmaniasis, seen in tropical and subtropical regions, is an infectious disease caused by the protozoan parasite Leishmania species. There are three main forms of leishmaniasis: cutaneous, mucocutaneous and visceral leishmaniasis. Cutaneous leishmaniasis (CL) has become an increasing problem as the number of travels around the world increases and people go to work in endemic areas. Turkey has received great numbers of immigrants in recent years, from its neighboring countries like Iraq, Islamic Republic of Iran, Afghanistan, Turkmenistan and the Syrian Arab Republic because of the political instabilities in these countries as well as the job opportunities caused by large-scale development projects undertaken by Turkey. In this report, imported CL cases detected in five truck drivers transporting from Hatay to Turkmenistan, Syria, Saudi Arabia, Iran and Georgia, Uzbekistan and Azarbaijan countries were presented. The patients admitted to Mustafa Kemal University, Faculty of Medicine Dermatology Policlinic, with wound complaints on their bodies were directed to the Department of Parasitology to obtain smear samples from their wounds. The age range of the patients were 38 to 43 years. Patients with wound trail for a period ranging from one month to one year had a number of lesions varying between 2-7 and in all cases, a smear preparation was prepared from the lesions for diagnostic purposes. Clinical material obtained from five patients with pre-diagnosis of CL was firstly examined with Giemsa stain. Samples taken from the patients were inoculated into modified NNN (Novy-MacNeal-Nicolle) medium for the evaluation of the presence of the promastigotes. Promastigotes obtained from the inoculated medium were also genotyped using the ITS1 region. In all of the slides prepared from the clinical material taken from the patients amastigotes were determined. The growth of promastigotes were observed in only three of the clinical specimen inoculated media. The genotyped three species were Leishmania tropica, Leishmania infantum/donovani and Leishmania major. In this study, the importance of support for the diagnosis of different microbiological methods used in the diagnosis of leishmaniasis infection which occurred during the outbreaks of the disease has been put forward. In addition, it was aimed to draw attention to the importance of imported CL cases in our country diagnosed in five truck drivers making transportation from Hatay to Turkmenistan, Syria, Saudi Arabia, Iran, Georgia, Uzbekistan and Azerbaijan.

Objective The Wolrd Health Organization (WHO) encourages all countries to investigate antimalarial drug substances derived from herbal sources with the slogan "Hunt of the Next Artemisinin" due to the emergence of resistant strains of Plasmodium species to artemisinin. In the broad and simple sense, it was planned to help guide the young researchers set in-vitro and in-vivo models of malaria in order to be used in drug research and active ingredient studies. Methods In-vitro study, young Plasmodium berghei trophozoites were removed from the liquid nitrogen tank and resuspended in appropriate conditions, followed by incubation with chloroquine and tetracycline at concentrations of 0.1, 0.4, 0.8, 1.6, 6.4, 12.8 μg/mL for 24 hours at +37°C in a shaking incubator. In-vivo studies, Tetracycline group (TG) and Chloroquine group (KG) were administered 50 mg/kg of tetracycline and chloroquine by intragastric lavage and untreated control group (TACG) were administered the same amount of saline via the same route. The suppression of parasitemia in mice was followed for 24 days. Results In our in-vitro study it was observed that 0.8 μg/mL of chloroquine and 1.6 μg/mL of tetracycline was enough to suppress parasitemia. In our in-vivo drug study, all of the mice in the TG group died at day 24, and all of the mice in the TAKG group died at day 12, with no parasitemia observed in the mice in the KG group. Conclusion Our study suggests that if tetracycline therapy is administered when the induction of chloroquine therapy is delayed, the exacerbation of the parasitemia may be prevented and when chloroquine is obtained chloroquine therapy can be commenced thus preventing the loss of the patient.

Amaç: Kan ve dokularda yerleşen paraziter hastalıklar insan sağlığı için dünya çapında büyük tehlike oluşturmaktadır. Sıtma, leishmaniasis ve toksoplazmoz her yıl çok sayıda kişinin yaşamını kaybetmesine neden olmakta, tedavi maliyetleri ve iş gücü kaybı nedeniyle ülke ekonomilerine büyük yük getirmektedir. Bu çalışmada, Plasmodium spp., Leishmania spp. ve Toxoplasma gondii’nin Gerçek Zamanlı Polimeraz Zincir Reaksiyonu (GZ-PZR) ile klinik örneğin mikrolitresinde belirlenebilecek en düşük parazit sayısının belirlenmesi, test validasyonlarının yapılması ve GZ-PZR testinin rutin laboratuvarlara dâhil edilmesi amaçlanmıştır. Yöntem: Leishmania tropica (promastigot ve amastigot), L. infantum (promastigot ve amastigot), T. gondii Thoma lamı ile mikroskop altında sayılarak P. falciparum ve P. vivax ise yayma preparatlar hazırlanıp Giemsa ile boyandıktan sonra mikroskop altında incelenerek parazitemi yoğunluğu saptanmıştır. Daha sonra bu parazitlerin PBS solüsyonu ile mikrolitrede 1, 5, 10, 25, 50, 100, 500, 1.000, 2.500, 5.000 ve 10.000 parazit olacak şekilde dilüsyonları yapılmıştır. Tüm parazit süspansiyonlarından DNA izolasyonu yapılarak GZ-PZR ile pozitif sonuç alınan en düşük parazit süspansiyonu belirlenmiştir. Daha sonra da negatif sonuç alınan süspansiyon ile bir önceki pozitif süspansiyon arası yine dilüsyon yapılarak testin saptayabildiği en düşük parazit sayısı belirlenmiştir. Bulgular: GZ-PZR testinde P. falciparum ile L. tropica ve L. infantum’un promastigot ve amastigot formlarında en düşük parazit sayısı 10, P. vivax ve T. gondii parazitlerinde ise 12 olarak bulunmuştur. Tartışma: Leishmania spp., Plasmodium spp. ve T. gondii’nin GZ-PZR ile tanısında çok düşük sayıdaki parazitlerin başarı ile saptanması, bu testin her üç parazitin moleküler tanısında güvenle kullanılabileceğini düşündürmüştür. Objective: Parasitic diseases that involve blood and tissues pose a significant threat to human health worldwide. Malaria, leishmaniasis and toxoplasmosis kill thousands of people worldwide annually, and impose a great burden on national economies due to treatment costs and loss of labour. The aim of this study are to determine the lowest number of parasites detectable by Real Time Polymerase Chain Reaction (RT-PCR) for Plasmodium spp., Leishmania spp. and T. gondii and to validate the RT-PCR test, and integrate RT-PCR in routine laboratories applications. Methods: Leishmania tropica (both promastigotes and amastigotes), L. infantum (both promastigotes and amastigotes) and T. gondii were counted using a hemocytometer, while Giemsa-stained smears of P. falciparum and P. vivax were examined under the microscope to define their parasitic load. Then, dilutions as 1, 5, 10, 25, 50, 100, 500, 1.000, 2.500, 5.000 and 10.000 parasites/ul were prepared in PBS solutions. DNA isolation was performed from all parasite suspensions and the lowest parasite suspension which was positively detected by RT-PCR method was identified. This was followed by further dilutions between the previous negative and last positive dilutions, to further determine the lowest number of parasites that RT-PCR could eventually identify. Results: Our assessments showed that the lowest number of parasites necessary for positive RT-PCR were 10 for P. falciparum and amastigotes and promastigote forms of L. tropica/L. infantum, while 12 for P. vivax and T. gondii. Conclusion: These results show that RT-PCR is a reliable diagnostic option to be used successfully for the detection of Leishmania spp., Plasmodium spp. and T. gondii, even for infections with low parasitemia.

A series of thiazolopyrimidine derivatives was designed and synthesized as aLeishmania majorpteridine reductase 1 (LmPTR1) enzyme inhibitor. TheirLmPTR1 inhibitor activities were evaluated using the enzyme produced byEscherichia coliin a recombinant way. the antileishmanial activity of the selected compounds was tested in vitro againstLeishmaniasp. Additionally, the compounds were evaluated for cytotoxic activity against the murine macrophage cell line RAW 264.7. According to the results, four compounds displayed not only a potent in vitro antileishmanial activity against promastigote forms but also low cytotoxicity. Among them, compoundL16exhibited an antileishmanial activity for both the promastigote and amastigote forms ofL. tropica, with IC(50)values of 7.5 and 2.69 mu M, respectively. in addition, molecular docking studies and molecular dynamics simulations were also carried out in this study. in light of these findings, the compounds provide a new potential scaffold for antileishmanial drug discovery., Turkiye Bilimsel ve Teknolojik Arastirma KurumuTurkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [SBAG-213-S026]; Ege Universitesi [2014BIL003], Turkiye Bilimsel ve Teknolojik Arastirma Kurumu, Grant/Award Number: SBAG-213-S026; Ege Universitesi, Grant/Award Number: 2014BIL003

Amaç: Dünyada yaklaşık 1 milyar insanın leishmaniasis riski altında olduğu ve leishmaniasis bildirimi yapan 200 ülke olduğu Dünya Sağlık Örgütü tarafından bildirilmektedir. En sık görülen formu olan kutanöz leishmaniasis tedavisinde ülkemizde meglumin antimonat, sodyum stiboglukonat ve son yıllarda amfoterisin B ilaçları kullanılmaktadır. Kullanılan ilaçların toksik etkisinin yüksek ve maliyetli olması yanı sıra son yıllarda meglumin antimonat ve sodyum stiboglukonat ilaçlarına karşı direnç geliştiği bildirilmektedir. Leishmaniasis tedavisinde kullanılmak üzere yeni ilaç adaylarına gereksinim vardır. Çalışmamızda, Olea europaea bitkisinin yaprak ve meyve kısımlarının su, sulu etanol ve kloroform çözücülerinde hazırlanmış ekstrelerinin Leishmania tropica parazitine karşı duyarlılığının in vitro araştırılması amaçlanmıştır. Yöntem: Manisa Celal Bayar Üniversitesi Tıp Fakültesi Parazit Bankası’nda MHOM/TR/2012/CBCL-LT kodu verilerek korunan Leishmania tropica izolatının Olea europaea bitkisinin ekstrelerine karşı in vitro olarak antileishmanial aktivitesi XTT ve hemositometre yöntemiyle araştırılmıştır. Ayrıca bitki ekstrelerinin sitotoksik aktiviteleri incelenmiştir. Bulgular: Antileishmanial aktivitenin değerlendirilmesinde yaprak su ekstresinin IC50 değeri 6.77 mg/kg, yaprak sulu etanol ekstresinin IC50 değeri 2.45 mg/kg, yaprak kloroform ekstresinin IC50 değeri 6.50 mg/ kg, meyve sulu etanol ekstresinin IC50 değeri 4.16 mg/kg ve meyve kloroform ekstresinin IC50 değeri 0.23 mg/kg olarak belirlenmiş ve meyve su ekstresinin antileishmanial etkisinin istatiksel olarak anlamlı diğerlerinin ise anlamsız olduğu görülmüştür. Sonuçların ilaçsız kontrol grubu ile istatistiksel olarak karşılaştırılmasına göre meyve klorofm ekstresinin (p0.05). Olea europaea bitkisinin yaprak su ekstresi (LC50 58.45 µg/ml) dışındaki ekstreler sitotoksik aktivite göstermemiştir. Sonuç: Bu çalışmamızın sonucunda, Olea europaea bitkisinin meyve kloroform ekstresinin antileishmanial aktivite göstermesi projenin amacı olan antileishmanial etken maddeler saptanması konusunda önemli ve ümit verici bir gelişme olarak değerlendirilmektedir, Objective: Meglumine antimonate, sodium stibogluconate and amfoterisin B are used in the treatment of cutaneous leishmaniasis which is the most common form in Turkey. Beside being highly toxic and costly, Leishmania parasites also have been reported to have developed resistance against meglumine antimonate and sodium stibogluconate in recent years. in this study, we aimed to investigate the in vitro susceptibility of water, aqueous ethanol and chloroform extracts of the leaves and fruits of Olea europaea against Leishmania tropica parasites. Method: in vitro antileishmanial activities of Olea europaea plant extracts against Leishmania tropica isolates obtained from Manisa Celal Bayar University Faculty of Medicine with MHOM/TR/2012/CBCL-LT code, were investigated by using XTT and hemocytometer methods. Cytotoxic activities of plant extracts were also determined. Results: Evaluation of antileishmanial activity showed that water of the aqueous ethanol and chloroform extracts of the leaves, and aqueous ethanol and chloroform extracts of the fruits had IC50 values as 6.77 mg/kg, 2.45 mg/kg, 6.50 mg/kg, 4.16 mg/kg, 0.23 mg/kg respectively. the water extract of fruits did not show any antileishmanial activity. Based on statistical comparison of these results with the control group, it was observed that fruit extract (p0,05). Extracts other than leaf water extract (LC50 58.45 µg/ml) of Olea europaea showed no cytotoxic activity. Conclusion: As a result of this study chloroform extract of the fruits of Olea europaea ws found to have antileishmanial activity and this can be considered as an important and promising development in determination of antileishmanial active substances which was the main aim of this study

Background: Cutaneous Leishmaniasis (CL) is the most common form of leishmaniasis. CL can be divided into two major groups: acute CL (ACL) and chronic CL (CCL). The aim of this study is to compare the efficacy of miltefosin and pentavalent antimony compounds in vivo with the CCL patient samples. Materials: Three study groups were formed, each consisting of five male Mus musculus (Balb/C) mice. In this model, promastigotes from the culture of a CCL patient were utilized. 100 ?L L. tropica promastigote suspension with a density of 108 promastigotes/ml were injected into the hint-right footpad of each experimental animal intradermally. Footpads of the mice were measured every two weeks until 24th week. From the 13th week, miltefosin 50 mg/kg/day was administered orally using gavage for 21 days, Meglumin antimoniate (MA) was administered by intramuscular (IM) injection daily for 21 days at 50 mg/kg/day and saline was administered IM for 21 days for the miltefosine, MA and control group, respectively. Results: The footpad measurements of the miltefosine group were lower than the control group statistically. Between the MA group and the miltefosine group and MA group and the control group, there was no statistically significant difference. Giemsa stained slides revealed amastigotes in one, two and all of the slides for the miltefosine, MA and control group, respectively. Molecular tests were performed with the Rotor-Gene device and L. tropica consistent peaks were obtained in one of the miltefosine group, four in the MA group and all mice in the control group. Conclusions: Demonstration of both clinical and laboratory improvement in four of the five experimental animals provides strong evidence that miltefosine is an effective drug in the treatment of CCL. In the literature, no clinical or laboratory studies using miltefosine have been performed with CCL patients only. © 2020, Witold Stefa?ski Institute of Parasitology, Polish Academy of Sciences., Ege Üniversitesi: 17-TIP-017, This project was supported by Research Funds of Ege University with the Project no: 17-TIP-017. Acknowledgements

Objective: Leishmaniasis is endemic in 98 countries, and poses risk to 1 billion people in the world. The infection caused by obligatory intracellular parasite Leishmania spp. in dogs is called Canine Leishmaniasis (CanL). In this study, it was aimed to detect the resistance status of Leishmania tropica strains isolated from dogs in our country against amphotericin B, meglumine antimoniate and sodium stibogluconate applied in leishmaniasis treatment. Method: Leishmania spp. promastigotes, taken out from the liquid nitrogen were first cultured in NNN media, then the growing isolates were transferred to RPMI-1640 medium and abundant amount of promastigotes were obtained. Isolates were genotyped using real-time polymerase chain reaction method with primers and probes specific to the ITS-1 region of Leishmania spp. and in five isolates that were found to be L. tropica causing CanL, resistance status against amphotericin B, meglumine antimoniate and sodium stibogluconate was investigated by hemocytometer and XTT methods. Results: The mean IC50 values were determined as 10.60 mg/ml for meglumine antimoniate, 0.1471 mg/ml for sodium stibogluconate, 0.0328 μM/ml for amphotericin B by hemocytometer method. Average IC50 values determined by XTT method were 10.48 mg/ml for meglumine antimoniate, 0.1470 mg/ml for sodium stibogluconate, 0.0326 μM/ml for amphotericin B. Conclusion: According to our data, while L. tropica strains which were isolated from CanL cases which are very rarely found causative agents in dogs, were not found to be resistant to amphoreticin B and sodium stibogluconate, parasite developed drug resistance against meglumine antimoniate. In future, this situation is thought to be a problem during treatment in human cutaneous leishmaniasis cases.

Objective: In the present study it was aimed to determine the in vitro antileishmanial activity of Berberis vulgaris L. against Leishmania tropica promastigotes. The aerial parts of Berberis vulgaris were collected from Spil Mountain, Manisa. Method: The ethanolic extract of the plant material was prepared. The consecutive concentrations of the plant extract (25-100µg/ml) were set for in vitro antileishmanial assays. In addition to in vitro inhibitory activities against Leishmania tropica promastigotes, the cytotoxicity of the plant extract was also measured by WST-1 Cell proliferation assay. The percentages of parasite inhibition in the presence of B. vulgaris ethanol extract in comparison with glucantime reference group at time interval of 12-72 hours were observed between 88.0 and 100.0 %. The plant extract was found to have cytotoxic activity with 444, 81±2, 12 µg/ml IC50 value. Conclusion: This is the first study that involves the in vitro antileishmanial activity of B. vulgaris which is wildly growing in Manisa, Turkey. Initial results demonstrated that the ethanolic extract of B. vulgaris gave promising results and it could be used as an antileishmanial agent in future. Peer Review History: Received 1 August 2019; Revised 15 August; Accepted 3 September, Available online 15 September 2019 Academic Editor: Dr. Amany Mohamed Alboghdadly, Princess Nourah bint abdulrahman university, Riyadh, amalbgadley@pnu.edu.sa UJPR follows the most transparent and toughest ‘Advanced OPEN peer review’ system. The identity of the authors and, reviewers will be known to each other. This transparent process will help to eradicate any possible malicious/purposeful interference by any person (publishing staff, reviewer, editor, author, etc) during peer review. As a result of this unique system, all reviewers will get their due recognition and respect, once their names are published in the papers. We expect that, by publishing peer review reports with published papers, will be helpful to many authors for drafting their article according to the specifications. Auhors will remove any error of their article and they will improve their article(s) according to the previous reports displayed with published article(s). The main of it is ‘to improve the quality of a candidate manuscript’. Our reviewers check the ‘strength and weakness of a manuscript honestly’. There will increase in the perfection, and transparency. Received file: Reviewer's Comments: Average Peer review marks at initial stage: 6.5/10 Average Peer review marks at publication stage: 9.5/10 Reviewer(s) detail: Dr. Gehan Fawzy Abdel Raoof Kandeel, Pharmacognosy Department, National Research Centre, Dokki, 12622, Giza, Egypt, gehankandeel9@yahoo.com Dr. George Zhu, Tehran University of Medical Sciences, Tehran, Iran, sansan4240732@163.com Similar Articles: PHYTOCHEMICAL SCREENING AND IN VITRO ANTIOXIDANT AND ANTI-DIABETIC POTENTIALS OF PERSEA AMERICANA MILL. (LAURACEAE) FRUIT EXTRACT

Plasmodium falciparum malaria causes about 450.000 deaths every year, mostly in children around the world. The infection is seen in cases coming from abroad and may lead to deaths in Turkey. Many native P.falciparum malaria cases and deaths due to this infection were observed in Turkey during mid 1900's when malaria was epidemic. But only two native cases were reported in the last 50 years, both from Manisa. First case was a one-year old baby who has come to Manisa from Urfa with his family and has never been abroad. He has diagnosed with Plasmodium vivax malaria and treated with chloroquine and primaquine. A previously obtained thin blood film was examined and characteristic P.falciparum rings in red blood cells were observed and the case was published together with photographs as probable P.falciparum and P.vivax mixed infection. After this case, microscopists working in Malaria Control Unit of Manisa were informed about the differentiation of malaria species in thin blood samples. Soon afterward, another case who have never been abroad before were also diagnosed with P.falciparum and P.vivax mixed infection and this case was also published with photographs taken from thin blood samples. As molecular diagnostic methods were not improved and widespread in those years, it could not be applied in both cases. A Giemsa stained thin blood sample of the baby case was incidentally found 22 years afterwards and with the aim of molecular diagnosis, the blood sample on the slide previously processed for DNA isolation, then analysed with "FTD Malaria Differentiation (Fast Track Diagnostics, Luxembourg)" multiplex kit with real-time polymerase chain reaction by using probes special for P.falciparum, P.ovale, P.malariae, P.vivax species. DNA's belonging to P.falciparum and P.vivax were found to be positive, the case is molecularly proved to have P.falciparum and P.vivax mixed infection. This case indicated that Turkey is convenient for the expansion of P.falciparum malaria in terms of the climate and vectors and suggested that the potential danger may increase with the effects of global warming, wars and migrations and may jump to Europe over Turkey. The case which molecularly proved the existence of native P.falciparum malaria in the near future in Turkey, was presented to draw attention to the danger of this infection for Turkey and Europe.

Leishmania virus (LRV) has previously been identified in different Leishmania species. Host-LRV interaction is associated with exacerbated clinical manifestations of cutaneous leishmaniasis (CL) and may cause poor therapeutic response. CL cases due to L. major with large skin lesions resistant to routine therapy were recently identified in Turkey. Here, we report the first autochthonous case of cutaneous leishmaniasis caused by LRV-positive Leishmania major, using conventional PCR targeting the viral capsid protein of LRV. The lesion of the case was 6 months old, relatively large (4 cm), and did not recover despite three consecutive intralesional applications of glucantime. Assessment of LRV's influence on prognosis and clinical outcomes of leishmaniasis, based on additional studies, is required.

Amaç: Bu çalışma, Türkiye’den elde edilen Leishmania spp. izolatlarının kanlı agar ve çikolata agardaki koloni oluşumlarının izlenmesi ve farklılıklarının karşılaştırılması, tek bir hücreden oluşan koloni elde edilerek gerektiğinde genetik çalışmalarda kullanılması amacıyla planlanmıştır. Yöntem: Çalışmamızda kullanılan kanlı agar ve çikolata agarın gerekli ön hazırlık aşamaları yapıldıktan sonra steril petri kaplarına dökülmesi ile besiyerleri hazırlanmıştır. Kullanılan Leishmania spp. izolatları üniversitemiz bünyesinde bulunan Parazit Bankası’ndan sağlanmıştır. Leishmania spp izolatlarının, ITS1 ve hsp70 bölgesine özgü primer ve problar kullanılarak genotiplendirilmesi yapılmıştır. Türlere göre ayrılan izolatların NNN besiyerinde kültürü yapılmış ve üreyen promastigotlar RPMI 1640 sıvı besiyerlerine aktarılarak logaritmik faza girmeleri beklenmiştir. Logaritmik faza giren promastigotların çikolata agar ve kanlı agara ekimleri yapılmış ve 25°C’lik etüve kaldırılarak gün aşırı takip edilmiştir. Bulgular: Etüvden çıkarılarak incelenen plaklarda inkübasyonun 7. gününde çikolata agarda herhangi bir koloni oluşumu gözlenmezken, kanlı agarda Leishmania tropica, Leishmania infantum ve Leishmania donovani de koloni büyüklüklerinin ortalama 0.9 mm olduğu, Leishmania major’de ise ortalama 0.8 mm olduğu saptanmıştır. Plaklara yapılan ekimin 28. gününde ise çikolata agarda L. tropica, L. major, L. infantum ve L. donovani’deki ortalama koloni büyüklükleri sırasıyla 1.0 mm, 0.9 mm, 1.0 mm ve 0.8 mm olarak gözlemlenirken, kanlı agardaki koloni büyüklükleri sırasıyla 3.1 mm, 3.1 mm, 3.3 mm ve 3.0 mm olduğu belirlenmiştir. Sonuç: Plaklara yapılan ekimlerin takibi sonucunda tüm suşların 7., 14., 21. ve 28. günlerdeki koloni oluşumlarının kanlı agarda daha hızlı olduğu saptanmıştır. Kanlı agarın Leishmania spp. ile ilgili yapılması planlanan genetik çalışmalar ve diğer çalışmalarda kullanılmasının uygun olabileceği sonucuna varılmıştır. Objective: This study was planned with the aim of comparing the reproduction differences of Leishmania isolates, obtained from Turkey, on chocolate and blood agars, colony growth views on agar plates and to attain colonies from single cell to utilize for genetic studies. Method: After preparation of blood agar and chocolate agar used in our study, the media were prepared by pouring into sterile petri dishes. The Leishmania isolates, used, were obtained from the ParasiteBank, located within our university. Genotyping was done by using primers and probes specific to ITS1 and hsp70. The isolates, separated according to species, were cultured in NNN medium and promastigotes were transferred to RPMI-1640 and were expected to enter into logarithmic-phase. Promastigotes entering logarithmic-phase were inoculated on chocolate and blood agar. Plates were transferred to 25°C and were controlled every other day. Results: No growth was observed on chocolate agar on 7th day of incubation on plates, while colony size of Leishmania tropica, Leishmania infantum and Leishmania donovani were found to have an average of 0.9 mm and mean colony size of Leishmania major was 0.8 mm on blood agar, On 28th day of cultivation, average colony size of L. tropica, L. major, L. infantum and L. donovani on chocolate agar was observed as 1.0 mm, 0.9 mm, 1.0 mm and 0.8 mm, respectively, while colony size on blood agar was determined as 3.1 mm, 3.1 mm, 3.3 mm and 3.0 mm, respectively. Conclusion: On 7,14,21,28th days, colony size of all the strains were found to be better and their growth were faster on blood agar. It has been concluded that blood agar may be suitable for reproduction of Leishmania for genetic and other studies planned.

Amaç: Bu çalışma, Türkiye’den elde edilen Leishmania spp. izolatlarının kanlı agar ve çikolata agardaki koloni oluşumlarının izlenmesi ve farklılıklarının karşılaştırılması, tek bir hücreden oluşan koloni elde edilerek gerektiğinde genetik çalışmalarda kullanılması amacıyla planlanmıştır. Yöntem: Çalışmamızda kullanılan kanlı agar ve çikolata agarın gerekli ön hazırlık aşamaları yapıldıktan sonra steril petri kaplarına dökülmesi ile besiyerleri hazırlanmıştır. Kullanılan Leishmania spp. izolatları üniversitemiz bünyesinde bulunan Parazit Bankası’ndan sağlanmıştır. Leishmania spp izolatlarının, ITS1 ve hsp70 bölgesine özgü primer ve problar kullanılarak genotiplendirilmesi yapılmıştır. Türlere göre ayrılan izolatların NNN besiyerinde kültürü yapılmış ve üreyen promastigotlar RPMI 1640 sıvı besiyerlerine aktarılarak logaritmik faza girmeleri beklenmiştir. Logaritmik faza giren promastigotların çikolata agar ve kanlı agara ekimleri yapılmış ve 25°C’lik etüve kaldırılarak gün aşırı takip edilmiştir. Bulgular: Etüvden çıkarılarak incelenen plaklarda inkübasyonun 7. gününde çikolata agarda herhangi bir koloni oluşumu gözlenmezken, kanlı agarda Leishmania tropica, Leishmania infantum ve Leishmania donovani de koloni büyüklüklerinin ortalama 0.9 mm olduğu, Leishmania major’de ise ortalama 0.8 mm olduğu saptanmıştır. Plaklara yapılan ekimin 28. gününde ise çikolata agarda L. tropica, L. major, L. infantum ve L. donovani’deki ortalama koloni büyüklükleri sırasıyla 1.0 mm, 0.9 mm, 1.0 mm ve 0.8 mm olarak gözlemlenirken, kanlı agardaki koloni büyüklükleri sırasıyla 3.1 mm, 3.1 mm, 3.3 mm ve 3.0 mm olduğu belirlenmiştir. Sonuç: Plaklara yapılan ekimlerin takibi sonucunda tüm suşların 7., 14., 21. ve 28. günlerdeki koloni oluşumlarının kanlı agarda daha hızlı olduğu saptanmıştır. Kanlı agarın Leishmania spp. ile ilgili yapılması planlanan genetik çalışmalar ve diğer çalışmalarda kullanılmasının uygun olabileceği sonucuna varılmıştır., Objective: This study was planned with the aim of comparing the reproduction differences of Leishmania isolates, obtained from Turkey, on chocolate and blood agars, colony growth views on agar plates and to attain colonies from single cell to utilize for genetic studies. Method: After preparation of blood agar and chocolate agar used in our study, the media were prepared by pouring into sterile petri dishes. The Leishmania isolates, used, were obtained from the ParasiteBank, located within our university. Genotyping was done by using primers and probes specific to ITS1 and hsp70. The isolates, separated according to species, were cultured in NNN medium and promastigotes were transferred to RPMI-1640 and were expected to enter into logarithmic-phase. Promastigotes entering logarithmic-phase were inoculated on chocolate and blood agar. Plates were transferred to 25°C and were controlled every other day. Results: No growth was observed on chocolate agar on 7th day of incubation on plates, while colony size of Leishmania tropica, Leishmania infantum and Leishmania donovani were found to have an average of 0.9 mm and mean colony size of Leishmania major was 0.8 mm on blood agar, On 28th day of cultivation, average colony size of L. tropica, L. major, L. infantum and L. donovani on chocolate agar was observed as 1.0 mm, 0.9 mm, 1.0 mm and 0.8 mm, respectively, while colony size on blood agar was determined as 3.1 mm, 3.1 mm, 3.3 mm and 3.0 mm, respectively. Conclusion: On 7,14,21,28th days, colony size of all the strains were found to be better and their growth were faster on blood agar. It has been concluded that blood agar may be suitable for reproduction of Leishmania for genetic and other studies planned.

EgeUn###, Leishmania virus (LRV) has previously been identified in different Leishmania species. Host-LRV interaction is associated with exacerbated clinical manifestations of cutaneous leishmaniasis (CL) and may cause poor therapeutic response. CL cases due to L. major with large skin lesions resistant to routine therapy were recently identified in Turkey. Here, we report the first autochthonous case of cutaneous leishmaniasis caused by LRV-positive Leishmania major, using conventional PCR targeting the viral capsid protein of LRV. The lesion of the case was 6 months old, relatively large (4 cm), and did not recover despite three consecutive intralesional applications of glucantime. Assessment of LRV’s influence on prognosis and clinical outcomes of leishmaniasis, based on additional studies, is required. ©2019 by EDIMES - Edizioni Internazionali Srl. All rights reserved

Leishmaniasis, seen in tropical and subtropical regions, is an infectious disease caused by the protozoan parasite Leishmania species. There are three main forms of leishmaniasis: cutaneous, mucocutaneous and visceral leishmaniasis. Cutaneous leishmaniasis (CL) has become an increasing problem as the number of travels around the world increases and people go to work in endemic areas. Turkey has received great numbers of immigrants in recent years, from its neighboring countries like Iraq, Islamic Republic of Iran, Afghanistan, Turkmenistan and the Syrian Arab Republic because of the political instabilities in these countries as well as the job opportunities caused by large-scale development projects undertaken by Turkey. In this report, imported CL cases detected in five truck drivers transporting from Hatay to Turkmenistan, Syria, Saudi Arabia, Iran and Georgia, Uzbekistan and Azarbaijan countries were presented. The patients admitted to Mustafa Kemal University, Faculty of Medicine Dermatology Policlinic, with wound complaints on their bodies were directed to the Department of Parasitology to obtain smear samples from their wounds. The age range of the patients were 38 to 43 years. Patients with wound trail for a period ranging from one month to one year had a number of lesions varying between 2-7 and in all cases, a smear preparation was prepared from the lesions for diagnostic purposes. Clinical material obtained from five patients with pre-diagnosis of CL was firstly examined with Giemsa stain. Samples taken from the patients were inoculated into modified NNN (Novy-MacNeal-Nicolle) medium for the evaluation of the presence of the promastigotes. Promastigotes obtained from the inoculated medium were also genotyped using the ITS1 region. In all of the slides prepared from the clinical material taken from the patients amastigotes were determined. The growth of promastigotes were observed in only three of the clinical specimen inoculated media. The genotyped three species were Leishmania tropica, Leishmania infantum/donovani and Leishmania major. In this study, the importance of support for the diagnosis of different microbiological methods used in the diagnosis of leishmaniasis infection which occurred during the outbreaks of the disease has been put forward. In addition, it was aimed to draw attention to the importance of imported CL cases in our country diagnosed in five truck drivers making transportation from Hatay to Turkmenistan, Syria, Saudi Arabia, Iran, Georgia, Uzbekistan and Azerbaijan.

Leishmaniasis is a zoonotic/anthroponotic vector borne parasitic infection which is caused by Leishmania species and transmitted by sand flies (Phlebotomus spp.) The reservoirs of Leishmania species in nature are various wild and domestic carnivores, rodents and human. The aim of this study was to investigate whether the rodents in genera Meriones, Mesocricetus, Rattus and Mus which inhabit in the natural habitat of our country could be natural reservoirs of Leishmania tropica, Leishmania infantum, Leishmania major and Leishmania donovani for cutaneous Leishmaniasis (CL)., The rodents Mus musculus (Balb/C mouse), Mesocricetus auratus (hamster), Meriones unguiculatus (gerbil) and Rattus norvegicus (rat) which are part of the natural habitat in Turkey were used in the study. L.tropica, L.infantum, L.major and L.donovani promastigote isolates obtained from CL patients and cultured in enriched media were injected in the footpads of the animals intradermally using the density of 10

Objective Two pentavalent antimonials, meglumine antimoniate (Glucantime®, France) and sodium stibogluconate (Pentostam®, England), are used to treat cutaneous leishmaniasis (CL) in Turkey. The present study, serving as a guidebook for young researchers, aims to provide basis for conducting drug resistance tests and active ingredient scanning in in vitro and in vivo models. Methods A CL isolate kept in liquid nitrogen was initially thawed and genotyped by real-time polymerase chain reaction (PCR) using ITS1 prob. In vitro and in vivo tests were conducted to determine drug resistance against meglumine antimoniate and sodium stibogluconate. Hemocytometry and XTT (sodium 3,39-[1-(phenylaminocarbonyl)-3,4-tetrazolium]-bis (4-methoxy-6-nitro) benzenesulfonic acid hydrate) methods were used to investigate in vitro drug resistance. CL mouse models were used to analyze in vivo drug resistance. Results The isolate was determined as Leishmania tropica by genotyping by PCR on the internal transcribed spacer 1 (ITS1) gene region. In in vitro drug resistance tests, sodium stibogluconate was observed to be more effective than meglumine antimoniate, but there was no statistically significant difference between the two (p > 0.05). It was observed that the footpad lesions of the animals started to shrink afterward the 5th week of infection following treatment with these agents, and parasitologic recovery was observed at the end of 3 months. Conclusions With an aim to be used as a guidebook for young researchers, active ingredient scanning and drug resistance tests in both in vitro and in vivo models were presented in the current study.

Leishmaniasis is a vector-borne zoonotic disease that shows different clinical features like cutaneous, mucocutaneous, visceral and viscerotropic forms. The protocols used in the treatment of leishmaniasis are toxic and have many limitations during administration. One of the limitations of treatment is the resistance against the protocols in practice. There is also a need to define new treatment options especially for resistant patients. Ex-vivo models using primary cell cultures may be a good source for evaluating new drug options in patients with antimony resistance, in addition to in-vitro and in-vivo studies. In this study, it was aimed to define a new ex-vivo culture model to evaluate treatment options in patients with cutaneous leishmaniasis who did not respond to treatment. In our experimental model of ex-vivo infection, Leishmania tropica promastigotes isolated from a case previously diagnosed with cutaneous leishmaniasis were used. The primary astroglial cell culture used for the ex-vivo model was prepared from 2-3 days old neonatal Sprague Dawley rat brains under sterile conditions by the modification McCarthy's method. The astroglia cells, which reached sufficient density, were infected with antimony resistant L.tropica promastigotes. After 24 hours of incubation, the supernatant on the cells were collected, the cell culture plate was dried at room temperature, then fixed with methyl alcohol and stained with Giemsa to search for L.tropica amastigotes. Amastigotes were intensely observed in glia cells in primary cell cultures infected with L.tropica promastigotes. No promastigotes were seen on Giemsa stained preparations of the precipitates prepared from the bottom sediment after the centrifugation of the liquid medium removed from the infected plates. In this study, promastigotes from a cutaneous leishmaniasis patient unable to respond to pentavalent antimony therapy were shown to infect rat glia cells and converted to amastigote form. This amastigote glial cell model, as far as we know, is the first model in the literature produced by L.tropica. The occurrence of L.tropica amastigote forms in glia cells may be indicative of the ability of Leishmania species to infect the central nervous system. The central nervous system may be an area for the Leishmania amastigotes to escape from the immune system in cases of leishmaniasis without a treatment response. Our study is important because it is the first study to show the infection of glia cells with L.tropica amastigotes.