MCBÜ Açıkerişim Sistemi :: Browsing by Author "Özbilgin, A"

Browsing by Author "Özbilgin, A"

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Evaluation of three new culture media for the cultivation and isolation of Leishmania parasites
Limoncu, M; Özbilgin, A; Balcioglu, I; Özbel, Y
The aim of this study was to establish novel culture media for Leishmania parasites with a potential of obtaining high amounts of promastigotes with long-term viability, and consisting of ingredients that were available in microbiology or parasitology laboratories. Other features of these media included no requirement for blood, FCS (Fetal calf serum) or erythrocyte lysate, inexpensiveness and easiness in preparation. In addition, aspiration samples obtained from cutaneous leishmaniasis (CL) suspected patients were cultivated in these media. Three culture media were prepared; trypticase beef extract hemoglobine (TBH) medium, including trypticase, beef extract and yeast extract as the protein source, glucose as the carbohydrate source, FeNH4 and bovine hemoglobine; Peptone-Yeast extract medium (PY), found to be effective in our previous studies for cultuvation of on Leishmania parasites, with bovine hemoglobine (PYH) and Brain Heart medium, containing bovine hemoglobine (BKH). The number of promastigotes were the highest on day 8 and 13 in RPMI 1640 and BKH medium, respectively. In TBH and PYH, the peak level of reproduction was between day 16 and 19, and it was found to be higher in TBH medium after the day 20. The number of promastigotes were found to be close in BKH, TBH and RPMI-1640 media and lower in PYH medium. Examination of the cultivation of the aqueous lesion specimens of the 10 CL-suspected cases in media revealed reproduction in 9 flasks of RPMI-1640 containing 10% FCS, 7 TBH, 6 BKH and 4 PYH. The differences between the culture media were not found to be statistically significant. These results suggested that, three liquid culture media, assessed in this study, with no requirement for FCS or erythrocyte lysate, were effective in the reproduction of promastigotes, and could be used effectively in the patient isolation and field studies, as well.
Knock, knock, knocking on Europe's door: Threat of leishmaniasis in Europe with a focus on Turkey
Tunali, V; Özbilgin, A
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.
A Native Mixed Plasmodium falciparum and Plasmodium vivax Malaria Case Molecularly Proven After 22 Years in Manisa, Turkey
Ok, ÜZ; Çavus, I; Sidal, U; Lmoncu, E; Özbilgin, A
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.
Putative Bronchopulmonary Flagellated Protozoa in Immunosuppressed Patients
Kilimcioglu, AA; Havlucu, Y; Girginkardesler, N; Çelik, P; Yereli, K; Özbilgin, A
Flagellated protozoa that cause bronchopulmonary symptoms in humans are commonly neglected. These protozoal forms which were presumed to be flagellated protozoa have been previously identified in immunosuppressed patients in a number of studies, but have not been certainly classified so far. Since no human cases of bronchopulmonary flagellated protozoa were reported from Turkey, we aimed to investigate these putative protozoa in immunosuppressed patients who are particularly at risk of infectious diseases. Bronchoalveolar lavage fluid samples of 110 immunosuppressed adult patients who were admitted to the Department of Chest Diseases, Hafsa Sultan Hospital of Celal Bayar University, Manisa, Turkey, were examined in terms of parasites by light microscopy. Flagellated protozoal forms were detected in nine (8.2%) of 110 cases. Metronidazole (500 mg b.i.d. for 30 days) was given to all positive cases and a second bronchoscopy was performed at the end of the treatment, which revealed no parasites. In conclusion, immunosuppressed patients with bronchopulmonary symptoms should attentively be examined with regard to flagellated protozoa which can easily be misidentified as epithelial cells.
Rural life, lower socioeconomic status and parasitic infections
Balcioglu, IC; Kurt, Ö; Limoncu, ME; Dinç, G; Gümüs, M; Kilimcioglu, AA; Kayran, E; Özbilgin, A
This study was conducted both to assess the prevalence of parasitic infections in two neighboring villages of Manisa province in Turkey, with different socioeconomic levels and investigate the influences of some individual and environmental factors on these infections. A total of 100 school children were enrolled and their cellophane tape and stool samples were examined for parasites. Pediculus capitis infestation was diagnosed by visual inspection of the hair. Although the prevalences of both intestinal parasitosis and pediculosis were high in both villages, there was a statistically significant difference for pediculosis, but not for intestinal parasitosis. Lack of social security and father's regular job were found to be highly correlated with all parasitic infections. Pediculosis was more common in younger children, girls and those in crowded families. These results indicate not only the high prevalence of parasitic infections, but also the urgent need for the improvement of basic health services and infrastructure in the rural areas, crowded with poorer immigrant population. (c) 2007 Elsevier Ireland Ltd. All rights reserved.
The Importance of the Contribution of Rapid Test, Serological and Molecular Methods in the Diagnosis of Two Imported Malaria Cases with Atypical Microscopy
Zorbozan, O; Pullukçu, H; Sahar, EA; Karakavuk, M; Can, H; Tunali, V; Döskaya, M; Turgay, N; Töz, S; Özbilgin, A
Malaria is a widespread and life-threatening disease in tropical and subtropical regions. In patients with typical clinical symptoms, malaria is considered as a preliminary diagnosis if there is a travel history to malaria-endemic areas. The basis of the laboratory diagnosis of malaria is the microscopic examination of Giemsa stained smears. On the other hand, the diagnosis and differentiation of Plasmodium species with microscopic examination may have some difficulties. In the first case, adifferent appearance from the classical Plasmodium vivax erythrocytic forms in infected erythrocytes were detected in 1% of all erythrocytes in thin smear blood preparations of a 26-year-old male with complaints of fever and chills and a story of travel to Nigeria. It was observed that parasitic nuclei were not prominent, and were located in the cytoplasm irregularly as chromatin or dye particles, nucleus fragments similar to Schuffner's granules in the form of scattered and granular spots were present in some erythrocytes, the cytoplasm of some Plasmodium erythrocytic forms were irregular and nuclei were not seen. There were no Schuffner's granules in any of the infected erythrocytes. P. vivax was detected by the rapid diagnostic test (OptiMAL, DiaMed GmbH, Switzerland), which searches for the antigens of Plasmodium species, in the peripheral blood sample of the patients. The P. vivax 18S rRNA gene was also detected by the multiplex real-time polymerase chain reaction. Antibodies against Plasmodium species were searched by using the Pan Malaria Antibody CELISA (CeLLabs Pty Ltd, Brookvale, Australia) kit in the patient's serum sample and the optical density (OD) value of the patient sample was measured five times the OD value of the positive control. In the second case, adifferent appearance from the classical P. falciparum erythrocytic forms in infected erythrocytes were detected in 12% of all erythrocytes in thin smear blood preparations of a 31-year-old male who has been suffering from persistent fever, severe headache, pain in the eyes and was known to be working in Nigeria. It was observed that some Plasmodium trophozoites have 1/3 of the size of erythrocytes such as P. vivax and have non-granular cytoplasm, some erythrocytic forms were round and the nucleus and cytoplasm were hardly distinguished, some of them were seen as crescent and close to the nucleus of the cytoplasm and some erythrocytic forms had characteristically a single nucleus and a scattered cytoplasm, similar to mature trophozoites of P. vivax. Although the Plasmodium young trophozoites were similar to P. vivax in means of magnitude, the forms in which the nuclei adhered to the erythrocyte wall were common. There were no P. falciparum gametocyte forms. P. falciparum like young trophozoite was observedonly in one of the four smears. P. falciparum was detected by the commercial rapid diagnostic test and P. falciparum 18S rRNA gene was also detected by the multiplex real-time polymerase chain reaction. Antibody formation against Plasmodium species was not detected in the ELISA test. In these case reports, the importance of the support of rapid diagnostic tests, serological and molecular methods to microscopic diagnosis and species determination of two imported malaria cases were demonstrated.
Detection of Leishmania RNA virus 2 in Leishmania species from Turkey
Nalçaci, M; Karakus, M; Yilmaz, B; Demir, S; Özbilgin, A; Özbel, Y; Töz, S
Background: Leishmania RNA virus (LRV) is a double-stranded RNA (dsRNA) virus infecting some Leishmania strains and triggering a destructive hyperinflammatory response in mammalian hosts in the New World. There is limited knowledge of the presence of this virus in Old World Leishmania species and its role in the outcome of the disease. We aimed to investigate the presence of LRV in Leishmania species/strains from Turkey. Methods: Twenty-nine previously identified Leishmania isolates (24 L. tropica, 2 L. infantum, 3 L. major) were examined for LRV positivity using dsRNA visualization in agarose gel after total nucleic acid extraction and RQ-deoxyribonuclease treatment and amplification of a 526 bp fragment of the LRV2-specific RNA-dependent RNA polymerase gene by reverse transcription polymerase chain reaction. Results: Ten (7 L. tropica [24.13%], 3 L. major [10.34%]) of the 29 Leishmania strains gave positive results for LRV. Basic Local Alignment Search Tool analysis showed that all these viruses are LRV2-1. LRV2 was detected for the first time in L. tropica strains in the present study. Conclusions: The clinical manifestation and resistance status of the disease can be different depending on the host and parasite species/strains. The presence of LRV2 may be one of the factors contributing the course of disease. Further studies are needed to elucidate the specific role of LRV2, as it may be a potential target for effective treatment strategies.
Infecting Glial Cells with Antimony Resistant Leishmania tropica: A New ex-vivo Model
Zorbozan, O; Harman, M; Evren, V; Erdogan, MA; Kilavuz, A; Tunali, V; Çavus, I; Yilmaz, Ö; Özbilgin, A; Turgay, N
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.
An Endemic Plant of Cyprus, Origanum majorana: Is It A New Alternative Natural Product for Malaria Treatment?
Güler, E; Özbilgin, A; Becer, E; Hanoglu, A; Sanlidag, T
Malaria still remains to be a public health threat and one of the most important infectious diseases to get attention from World Health Organization. No domestic malaria cases have been reported on the island of Cyprus since 1948, as a result of successful elimination process. All of the malaria cases detected in recent years are imported cases. As known, hundreds of medicines are obtained from plants and traditional medicine are used in endemic places of malaria. The cause of malaria - Plasmodium parasites, are developing resistance to antimalarial drugs. Hence, research on plant extracts and essential oils have gained great interest in recent years to obtain new and safe agents/substances. In our study, it was aimed to investigate the in vivo antimalarial activities of essential oils obtained from Origanum dubium, Origanum majorana, Salvia fruticosa and Laurus nobilis plants which grows in Northern Cyprus against Plasmodium berghei - the rodent malaria agent. Plants were collected in appropriate seasons and were dried to obtain and analyze essential oils via Clevenger Apparatus system. L929 mouse fibroblast cell line and MTT [3-(4.5-dimethylthiazole-2-yl) -2.5-diphenyltetrazolium bromide] kit were used to determine the cytotoxic activities of the essential oils obtained. In our study, total of 36 mice (Balb/c) of 6 groups (6 mice in each group) were formed: chloroquine group (CG) (50 mg/kg) as malaria reference group, untreated control group (UTCG), O. dubium (OD) (20 mg/kg), O.majorana (OM) (20 mg/kg), S.frutikosa (SF) (20 mg/kg) and L.nobilis (LN) (20 mg/kg). The essential oils were given to mice infected with P.berghei strain orally on 0, 1, 2 and 3rd days (4 times in total). Blood was taken from the tail end of each mouse 24 hours after the last treatment and blood collection was continued every two days until the mice died. Withdrawn blood taken from the mice were prepared as a thin smear and stained with Giemsa. Then, parasitemia percentages in each smear were calculated. As a result of the cytotoxicity tests, cytotoxic activity was not found at 100 mu g/ml (20 mg/kg) in all oils except OD essential oil. While the mice receiving chloroquine continued their lives with the disappearance of the parasite on the 6th day, the mice in the UTCG died on the 9th day. The parasitemia rate reached 35% in the OM group on the 23rd day, in the OD group on the 21st day and in the other groups (SF and LN) on the 14th day and the mice have died. In our study, the difference between the life span in all groups was found statistically significant (ps <= 0.001). As a result, the essential oils O.majorana (14 days increase according to UTCG) an endemic plant of Cyprus and O.dubium (12 days increase according to UTCG) which had an antimalarial effect, decreased parasitemia and increased the life span of mice more than two times, indicated that they could be a source for the acquisition of new antimalarial molecules.
In vitro Susceptibility of Trichomonas vaginalis to Metronidazole, Ornidazole and Proton Pump Inhibitors Pantoprazole and Esomeprazole
Aksoy Gökmen, A; Girginkardesler, N; Kilimcioglu, AA; Sirin, MC; Özbilgin, A
The current treatment of trichomoniasis is based on the use of 5-nitroimidazole derivatives. Although metronidazole is reliable, inexpensive and highly effective against anaerobic microorganisms and protozoa, the development of metronidazole-resistant T.vaginalis strains pose to an increasing problem. Nitroimidazoles are compounds having azomycin (2-nitroimidazole) chemical structure and are obtained from Streptomyces strains. Benzimidazole, which is found in the structure of proton pump inhibitors, is also present in the other components that have antiprotozoal activity. In this study, the in vitro susceptibility of T.vaginalis against metronidazole, ornidazole, and the proton pump inhibitors which are tested recently as antiprotozoal agents; pantoprazole and esomeprazole was investigated. For this purpose a clinical T.vaginalis strain which was formerly isolated and stored after cryopreservation process in our laboratory was used. Minimum inhibitory concentration (MIC) and minimum lethal concentration (MLC) values of those agents against to this strain were determined in vitro by dilution method in 24-well cell culture plates. Trypticase yeast extract maltose medium, horse serum and antibiotic (penicillin + streptomycin) were distributed to each well of cell culture plates and after metronidazole, ornidazole, pantoprazole and esomeprazole solutions were added to two wells for each as 800, 400, 200, 100, 50 and 25 mu g/ml, followed by the addition of 1 ml 5x10(3) T.vaginalis trophozoites into each well. Plates were incubated at 37 degrees C, and viability and motility of the trophozoites were evaluated under light microscope at 24, 48 and 72 hours after incubation. MIC and MLC values of metronidazole/ornidazole in the 72th hour were found as 50 mu g/ml and 100 mu g/ml, respectively. MIC and MLC values for pantoprazole in the 72th hour were 200 mu g/ml and 400 mu g/ml, while the values for esomeprazole were 400 mu g/ml ve 800 mu g/ml, respectively. As a result, T.vaginalis strain used in the study was susceptible to metronidazole and ornidazole, besides, it was considered that pantoprazole and esomeprazole were also effective to the parasite and could be used as alternative drugs. However, further in vitro and clinical studies are clearly needed on the antiprotozoal effects of proton pump inhibitors. To our knowledge, this study was the first in literature, which esomeprazole's susceptibility on T.vaginalis was investigated in vitro.
In vitro Efficacy of Quercus infectoria Oliv. and Achillea millefolium L. Extracts Against Blastocystis spp. Isolates
Özbilgin, A; Durmuskahya, C; Kilimcioglu, AA; Kayalar, H; Kurt, Ö; Ermis, VÖ; Tabak, T; Östan, I
Blastocystis is a common intestinal parasite that can inhabit the intestinal tract of humans and many animals. Despite it was firstly described almost 100 years ago; many subjects are still under debate about Blastocystis, including its life-cycle, pathogenic potential and treatment of infected individuals. Historically, local plant species have been used for therapeutic purposes by the local people of Anatolia. Here, hexane and methanol extracts of two local plants, Quercus infectoria (Fagaceae) and Achillea millefolium, which have been used against diarrhea in Anatolia, were examined for their in vitro efficacies against Blastocystis. LC50 and EC50 values of the plant extracts were determined by Brine Shrimp and Graphpad Prism 5 (R) methods, respectively. The results showed that LC50 (500 mu g/ml) and EC50 (198.8 mu g/ml) concentrations of the methanol extract of A. millefolium were lowest compared to other extracts, its anti-Blastocystis activity was found to be comparable to metronidazole and it showed no cytotoxic activity. These initial results suggest that the methanol extract of A. millefolium may be a novel option for the treatment of Blastocystis infections in humans in future, if confirmed by further, larger-scale studies.
Comparison of culture media in the isolation and diagnosis of cutaneous leishmaniasis
Özbilgin, A; Zeyrek, F; Limoncu, ME; Östan, I; Tabak, T; Asar, K; Türkmen, H; Kilimcioglu, AA
Leishmaniasis is a common and significant parasitic disease among the Middle East and Mediterranean countries. Cutaneous and visceral forms of leishmaniasis are seen in Turkey. Direct microscopic examination of Giemsa-stained smears and inoculation into a culture medium are gold standards in diagnosis. In the present study, three diffferent culture media (pepton-yeast extract, brain heart and nutrient broth) were used in comparison with Novy-MacNeal-Nicolle and RPMI 1640 as control media to assess the effectiveness of new culture media both in diagnosis in patients and after isolation from the mice. In addition, smears prepared from the infectious material were stained with Giemsa and examined under a light microscope. Samples obtained from the lesions of cutaneous leishmaniasis produced on the sole of mice were inoculated into all culture media; growth were detected in pepton-yeast extract and nutrient broth media. Smears prepared from the lesions of these mice were stained with Giemsa and amastigotes were detected on them. Growth was also seen in control media. Five of 12 cultivations of the human samples suspected of having cutaneous leishmaniasis revealed growth in pepton-yeast extract medium and nutrient broth medium. Microscopic examination of these five patients showed the presence of amastigotes and growth in control media, while none was observed in the remaining seven cases. These results indicated that pepton-yeast extract and nutrient broth media could be preferred in both isolation of the parasites and diagnosis of cutaneous leishmaniasis.
Feconomics®: A Simple, Novel and Fast Technique for Stool Concentration in Parasitology Laboratory
Kurt, Ö; Akyar, I; Görgün, S; Kocagöz, T; Özbilgin, A
Feconomics (R) is a new ready-to-use kit for fecal concentration that eliminates the need for centrifugation and floatation by using absorbent beads. To assess its efficacy in the diagnosis of intestinal parasites, a comparative, double-blind study was conducted in the Parasitology Laboratory of Celal Bayar University Medical School. Stools (Group I, n=251) submitted for routine ova and parasite examination were concentrated with both routine formalin ethyl acetate concentration (FEAC) technique and Feconomics (R). Since the number of helminthes identified in the stool samples of patients were very low, helminthes obtained from the animal models in the laboratory were included (Group II, n=11). The iodine-stained samples of all stools and some of the positive samples stained with Gomori's trichrome and Kinyoun's acid fast stain were read by specialists. In Group I, 103 of 251 (41.04%) samples were found to be positive for one or more intestinal parasites; among them, 76(30.28%) and 96(38.25%) stools were found to be positive with FEAC and Feconomics (R), respectively, and the difference was significant (P=0.000). Same parasites were identified with both methods among all 11 samples in Group II. There was no difference between the methods for the morphological integrity and visual appearances of the parasites having cyst or egg forms; yet, it was noticed that the vegetative forms of the parasites were only identified with Feconomics (R). Review of our data indicated that Feconomics (R) may be suggested as a fast and effective fecal concentration method for Parasitology laboratories owing to the identification of higher number of parasites compared to FEAC, and parasites with only vegetative forms such as Dientamoeba fragilis.
Imported Cutaneous Leishmaniasis Cases Detected in Truck Drivers in Hatay
Çulha, G; Çigdem Dogramaci, AC; Kaya, T; Çavus, I; Gülkan, B; Özbilgin, A
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.
Investigation of the Efficacy of Cinnamaldehyde, Cannabidiol and Eravacycline in a Malaria Model
Özel, Y; Çavus, I; Ünlü, M; Özbilgin, A
In this study, it was aimed to investigate the antimalarial activity of cinnamaldehyde (CIN) and can- nabidiol (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 mu g/mL, 16.40 mu M and 48.82 mu g/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.
An Alternative Biphasic Nutrient Medium for the Diagnosis of Cutaneous Leishmaniasis
Gökmen, AA; Öncel, K; Özdemir, OA; Pektas, B; Çavus, I; Güngör, S; Uzun, B; Kaya, S; Karaca, S; Yula, E; Demirci, M; Özbilgin, A
Cutaneous leishmaniasis (CL) caused by the Leishmania spp. parasites, is a disease characterized by nodulo-ulcerative lesions in the skin. CL is transmitted to humans by infected sandflies during blood sucking, and is endemic in about 98 countries over the world. The demonstration of amastigotes via microscopic examination, and the growth of promastigotes in NNN (Novy-MacNeal-Nicolle) medium are gold standard methods for laboratory diagnosis. The aim of this study was to compare the biphasic NNN medium that is frequently used in routine laboratories with the biphasic nutrient medium that can be prepared easily in microbiology laboratories, for the growth of promastigotes. In the study, the aspiration fluid sample was used as clinical sample which was obtained from the skin lesion of a 47-year-old female patient admitted to Izmir Katip Celebi Ataturk Education and Research Hospital dermatology outpatient clinic and pre-diagnosed as CL. The aspirate sample taken from the lesion was evaluated with microscopy, cultivation in two different media and real-time polymerase chain reaction (Rt-PCR) methods. In microscopic examination Leishmania amastigotes were observed in Giemsa-stained smears prepared from the aspiration fluid. In Rt-PCR performed by using specific primers and probes targeting ITS1 region of Leishmania parasite, a melting-curve compatible with L.tropica was detected. For cultivation, triple inoculations of the aspirate sample into NNN (NNN + RPMI 1640 + 10% fetal calf serum) and nutrient media (nutrient agar + nutrient broth + 10% fetal calf serum) were used. The cultures were incubated at 27 degrees C for 10 days, and the number of propagated promastigotes were counted on the third, seventh and tenth days. The growth of Leishmania promastigotes was detected in both media on the third day. The number of promastigotes grown in NNN medium on the third, seventh and tenth days were 10(5)/ml, 10(6)/ml and 10(8)/ml, respectively. Those values in nutrient medium were 10(6)/ml, 10(7)/ml and 10(8)/ml on the third, seventh and tenth days, respectively. Although the number of promastigotes on the third and seventh days were higher in nutrient medium than NNN medium, the number of cultivated promastigotes were equal on the tenth day. As a result, nutrient medium is considered to have an impact in the diagnosis of CL, by providing an alternative to the routine medium used and can readily be available in microbiology and parasitology laboratories with long shelf-life. It was concluded that biphasic nutrient medium could be used as a supplementary medium for diagnosis in laboratories in the absence of NNN medium or can not be provided.
The First Monkey Malaria in Turkey: A Case of Plasmodium knowlesi
Özbilgin, A; Çavus, I; Yildirim, A; Gündüz, C
Plasmodium knowlesi is now added to the known four Plasmodium species (P. vivax, P. falciparum, P. malariae, P. ovale) as a cause of malaria in humans because of the recent increasing rate of cases reported from countries of southeastern Asia. P. knowlesi which infects macaque monkeys (Macaca fascicularis and M. nemestrina) is transmitted to humans especially by Anopheles leucosphyrus and An. hackeri mosquitos. First human cases of P. knowlesi malaria have been detected in Malaysia which have reached high numbers in recent years and also have been reported from countries of Southeast Asia such as Thailand, Philippines, Myanmar, Singapore and Vietnam. However the number of cases reported from western countries are rare and limited only within voyagers. This report is the first presentation of an imported case of P. knowlesi malaria in Turkey and aims to draw attention to the point that it could also be detected in future. A 33-year-old male patient from Myanmar who has migrated to Turkey as a refugee, was admitted to a health center with the complaints of fever with a periodicity of 24 hours, headache, fatigue, cough, sore throat, anorexia, myalgia and arthralgia. He was prediagnosed as upper respiratory tract infection, however because of his periodical fever and background in Myanmar, thick and thin blood films were prepared and sent to our laboratory for further examinations. Microscopic examination of the thin blood films revealed erythrocytic stages compatible with P. knowlesi (three large early trophozoites in an erythrocyte, three late trophozoites with compact view, and three late band-form trophozoites). Upon this, both real-time polymerase chain reaction (Rt-PCR) targeting the small subunit ribosomal RNA (SSU-rRNA) genes of Plasmodium genus and DNA sequence analysis targeting P. knowlesi rRNA gene were performed. As a result, the suspected identification of P. knowlesi by microscopy was confirmed by Rt-PCR and DNA sequencing. The patient was treated with chloroquine and primaquine combination and in the follow-up on the seventh day after the treatment, his parasitemia and symptoms had ceased. Although there were some previous reports concerning about imported patients infected with different Plasmodium species in our country, no cases of P. knowlesi have been reported. This first case presented here emphasizes the occurence of P. knowlesi malaria in Turkey hereinafter due to the increasing number of refugees.
Cutaneous leishmaniasis infection in Balb/c mice using a Leishmania tropica strain isolated from Turkey
Girginkardesler, N; Balcioglu, IC; Yereli, K; Özbilgin, A; Özbel, Y
Leishmania tropica, which is endemic in Turkey, is the causative agent of cutaneous leishmaniasis. Leishmania tropica promastigotes (2 x 10(7)) isolated from a patient with dermal leishmaniasis and reproduced in NNN medium were inoculated subcutaneously into the footpads of 10 Balb/c mice. Cutaneous leishmaniasis developed on the footpads of 4 mice approximately 45 days later. Leishmania tropica amastigotes were observed in smear slides and then cultivated in NNN medium. Balb/c mice are a suitable laboratory model for this isolate of L. tropica and thus a source of amastigotes for studies on the immunology, chemotherapy, and pathogenicity of cutaneous leishmaniasis.
Autochthonous Case of Malaria Prediagnosed as Leukemia
Gülen, H; Türedi Yildirim, A; Çavus, I; Türkmen, H; Özbilgin, A
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 Turkiye. 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 Turkiye. 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 Turkiye. Tropical diseases, especially malaria, is of great importance for Turkiye 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 Turkiye.
A comparison of metronidazole and single-dose ornidazole for the treatment of dientamoebiasis
Kurt, Ö; Girginkardesler, N; Balcioglu, IC; Özbilgin, A; Ok, ÜZ
Recent reports of the pathogenic potential of Dientamoeba fragilis have underlined the need for an effective treatment against this colon-dwelling protozoan. Metronidazole is a well-known and commonly used anti-protozoal agent, but another 5-nitroimidazole derivative, ornidazole, may be preferable, where available, because of its longer half-life and fewer side-effects. This study compared the efficacies of metronidazole and ornidazole in a group of 112 patients with dientamoebiasis. Patients were randomised into two treatment groups: group 1 (n = 56) received metronidazole for 5 days, 20 mg/kg/day for children and 1.5 g/day for adults, in three oral doses, while group 2 (n = 56) received a single oral dose of ornidazole, 30 mg/kg for children and 2 g for adults. Stool samples were examined on the seventh and 14th days after treatment, and clinical symptoms were recorded to evaluate the efficacy of treatment. A statistically significant difference was recorded between the efficacies of ornidazole and metronidazole, both parasitologically (92.9% vs. 69.6%, p 0.001) and clinically (96.4% vs. 76.8%, p 0.001). Patients in the metronidazole group reported more side-effects than patients in the ornidazole group, none of whom required termination of treatment. These results suggest that single-dose ornidazole may be an important alternative agent for the treatment of dientamoebiasis.

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