Leishmaniasis in Turkey: first clinical isolation of Leishmania major from 18 autochthonous cases of cutaneous leishmaniasis in four geographical regions.

OBJECTIVE: To report isolation of Leishmania major strains obtained from 18 Turkish autochthonous cutaneous leishmaniasis (CL) patients infected with L. major between 2011 and 2014. METHODS: Initial diagnosis relied on microscopy and culture in enriched medium, prepared by adding specific amounts of liver extract, protein and lipid sources to NNN medium. Promastigotes were then transferred to RPMI medium including 10% of foetal calf serum for mass culture. Species-specific real-time PCR targeting ITS1 region of Leishmania spp. was performed using both lesion aspiration samples and cultured promastigotes. Two of 18 isolates were identified by isoenzyme analysis in the Leishmaniasis Reference Center in Montpellier, France. Each isolate was inoculated into the footpads of six mice to observe the pathogenicity of L. major. Developing lesions were observed, and the thickening of footpads was measured weekly. RESULTS: Melting curve analyses of 18 isolates showed a peak concordant with L. major, and two of them were confirmed by isoenzyme analyses as L. major zymodeme MON103. In the mouse model, acute lesions seen on day 21 were accepted as an indication of heavy infection. Severe impairments were observed on all mouse footpads over 3 weeks, which even progressed to extremity amputation. CONCLUSION: Cutaneous leishmaniasis-causing L. major was recently identified in Adana province in southern Turkey, with PCR. Our study shows that such CL cases are not limited to Adana but currently present from western to Southeastern Anatolia, and along the Mediterranean coast. The role of small mammals, the main reservoirs of L. major in Anatolia, needs to be elucidated, as do the underlying factors that cause severe clinical manifestations in L. major infections in Turkey, contrary to the infections in neighbouring countries.

Assessment of in vivo antimalarial activities of some selected medicinal plants from Turkey.

Resistant infections lead to increased necessity of searching novel drugs and drug combinations. The purpose of this paper was to investigate antimalarial properties of some selected medicinal plants that have been traditionally used in Turkey for antipyretic and analgesic purposes. Lavandula stoecheas subsp. cariensis, Phlomis nissolii, Phlomis bourgaei, Phlomis leucophracta, Centaurea hierapolitana, Centaurea polyclada, Centaurea lydia, Scrophularia cryptophila, Scrophularia depauperata, Scrophularia floribunda, Rubia davisiana, and Alkanna tinctoria subsp. subleiocarpa were investigated for their in vivo antimalarial activities in mice infected with Plasmodium yoelii. Two hundred fifty to 500 mg/kg doses of plant extracts were given to mice as a single daily dose for 4 days. P. nissolii water extract, C. lydia chloroform extract, S. cryptophila ethanol extract, and C. polyclada methanol extract showed antimalarial activity with reducing parasitaemia. The chemotherapeutic effects of plant extracts ranged between 13.5% and 66.91%. The chemosuppressions exerted by combined plant extracts of P. nissolii, S. cryptophila, and C. lydia with C. polyclada methanol extract were detected as 51.25%, 57.33%, and 58.33%, respectively. Investigation of cytotoxic activities against brine shrimps revealed that methanol extract of C. polycada, chloroform extract of C. lydia, and ethanol extract of S. cryptophila showed cytotoxic activities, while water extract of P. nissolii was not active against brine shrimps.

Leishmaniasis in Turkey: Determination of Leishmania Species by Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF MS).

BACKGROUND: Cutaneous leishmaniasis (CL) is endemic in Southeastern Anatolia, mainly in Sanliurfa and Hatay provinces, and the causative agents are mostly Leishmania tropica and less frequently L. infantum. Here, we report the first MALDI-TOF analyses of Leishmania promastigotes obtained from the cultures of two CL cases from Osmaniye and Hatay provinces who were initially diagnosed by microscopy, culture and identified as L. infantum with Real-Time PCR (RT-PCR). METHODS: Samples obtained from the skin lesions of patients were initially stained with Giemsa and cultivated in NNN medium. Examination of the smears and cultures revealed Leishmania amastigotes and promastigotes, respectively. The promastigotes (MHOM/TR/2012/CBU15 and MHOM/TR/2012/MK05) obtained from the cultures of both patients were used for RT-PCR targeting the ITS-1 region in the SSU of rRNA. The reference strains of four Leishmania species (L. infantum, L. donovani, L. tropica and L. major) were initially assessed with MALDI-TOF and their data were added to MALDI-TOF Biotyper Library. RESULTS: Both RT-PCR and MALDI-TOF analyses indicated that the causative agent in both patient samples was L. infantum. CONCLUSION: Despite disadvantages such as requirement of culture fluid with nothing but promastigotes and high cost, MALDI-TOF analysis may be a fast, sensitive and specific diagnostic tool in especially large-scale research studies, where the cost declines, relatively.

A mass screening survey of cystic echinococcosis by ultrasonography, Western blotting, and ELISA among university students in Manisa, Turkey.

Cystic echinococcosis (CE) is one of the most important zoonotic diseases in a wide geographic area, including Turkey. In the present project, a total of 4275 students from Celal Bayar University, Manisa, Turkey, were screened by ultrasonography (US) and specific antibodies for CE were examined by Western blotting (WB) and ELISA in finger prick blood samples of 2034 of 4275 volunteered students. We aimed to report the apparent prevalence of CE based on different diagnostic procedures and to compare WB and ELISA with US in diagnosis of CE in a mass screening setting. Six new cases were diagnosed as CE by US during the survey. In addition to these cases, three students were also detected to have been previously operated and pathologically confirmed for hepatic CE. US revealed parenchymal changes in these cases in concordance with their operation history; so, the prevalence of CE by US was calculated as 0.21% (9/4275) (95%CI, 0.11-0.39%) among university students in Manisa. Bands were detected at 8, 28, 32, 38, 42, 47, 70 and 90kDa by WB and the cases were considered to be positive for CE when at least three of the bands were seen together. Apparent prevalence of CE by ELISA and WB were found to be 2.11% (43/2034) (95%CI, 1.57-2.83%) and 0.25% (5/2034) (95%CI, 0.10-0.57%), respectively. Of the six US positive cases, WB was positive in only one case with two cysts in the liver. All of four cases with liver involvement were positive by ELISA. The high prevalence of CE among university students in Manisa indicated that CE is a major health problem in this area of Turkey. Our results supported that WB is rather difficult and not feasible as a mass screening test and may not be effective for confirmation especially in asymptomatic cases. As a result, we recommend US to be used initially in mass screening surveys for CE followed by confirmation by ELISA for suspected cases. Further examination primarily by chest X-ray followed by computed tomography and/or magnetic resonance imaging, if needed, should be recommended for US negative, ELISA and WB positive individuals who may have non-abdominal cysts.

[Cryopreservation of plasmodia with malaria models and establishment of a cryobank]. / Sitma modeli etkenleri ile kriyoprezervasyon çalismalari ve kriyobanka olusturulmasi.

OBJECTIVE: Cryopreservation is simply a method of keeping living cells frozen with the chance of regaining cellular viability, functions and antigenic structures whenever required, after heating. METHODS: In the present study, dimethyl sulphoxide (DMSO) was mixed with the red blood cells having 20% of parasitemia obtained from the mice infected with Plasmodium yoelii and Plasmodium berghei at a final concentration of 15%. For cryopreservation: both test tubes containing each Plasmodium species were kept 10 minutes at room temperature, 30 minutes at +4°C, 90 minutes at -20°C and finally at -80°C. Some were left at this temperature, while some were transferred into the liquid nitrogen tank at -196°C after being left at -80°C for three hours. RESULTS: Our observations and assessments demonstrated that both P. yoelii and P. berghei might keep their viability and virulence at -80°C and -196°C between the first and the sixth months of cryopreservation. CONCLUSION: It can be concluded that the cryopreservation of P. yoelii and P. berghei at -80°C and -196°C are successful, indicating the advantage of the establishment of parasite cryobanks in research laboratories.

Investigation of the prevalence of amoebiasis in Izmir province and determination of Entamoeba spp. using PCR and enzyme immunoassay.

Amoebiasis is a common and life-threatening disease. The discrimination of the pathogenic Entamoeba histolytica from the non-pathogenic Entamoeba dispar could be done by advanced methods such as enzyme immunoassay (EIA) and PCR. The aim of this study was to investigate the prevalence of amoebiasis in Izmir province, and differentiate the Entamoeba species by PCR and EIA. Stool samples of 2,047 individuals were examined by direct microscopy, formalin ethyl acetate concentration, trichrome staining and culture, and those found to be positive for E. histolytica/dispar by any of these methods were further analyzed by PCR and EIA for species identification. Fifty-nine of 2,047 (2.9%) stool samples were found to be positive for E. histolytica/dispar with microscopy and/or culture. Among these positive samples, E. histolytica was detected in 14 (23.7%) and 5 (8.5%) samples with PCR and antigen-specific ELISA (EIA), respectively. E. dispar was diagnosed in 31 (52.5%) and 52 (88.1%) of 59 samples with species-specific PCR and EIA, respectively. Risk factors related to infection with Entamoeba spp. and other intestinal parasites included living in shanty houses (p < 0.01), a history of recent immigration to Izmir (p < 0.01), having no social security (p < 0.05) and living with a crowded family (p < 0.01). The results demonstrated the significance of amoebiasis as a public health problem among people with low socio-economic status in Izmir province.

The prevalence of intestinal parasites in the province of Izmir, Turkey.

Stool samples of a total of 2,047 people in Izmir province were examined by wet mount, formalin ethyl acetate concentration, and trichrome staining methods with an aim to reveal the prevalence of intestinal parasites in Izmir together with related personal and environmental risk factors. Geographical mapping showing the density and variation of the species of intestinal parasites in Izmir was done after all maps were scaled, and the coordinates were determined with GeoMedia5.0. The prevalence of the intestinal parasites was found to be 25.6% in Izmir, with a variation between the districts. Blastocystis hominis was the leading parasite, and the prevalence of parasites was higher in children compared to adults; however, the difference was statistically insignificant. There was also no significant difference between the parasite prevalence and sex, marital status, education, income, frequent eating outside, and habitual raw meat eating. Significant differences were found between the parasite prevalence and crowded families, early immigrants, individuals with no social security, and people living in close contact with their livestock. The parasites were found to be less common among individuals who had been drinking bottled water and living in a house with a sewage system. The results demonstrated a correlation between the intestinal parasites and environmental conditions in our study group. We further plan to expand the study group to cover all regions of Turkey.

In vitro and in vivo activities of Haplophyllum myrtifolium against Leishmania tropica.

This study aimed to evaluate the in vitro and in vivo leishmanicidal activity of an endemic Turkish plant and compare its efficacy with a reference drug. In addition to the in vitro activities of the ethanol, acidified and alkaloid extracts and furoquinoline alkaloids skimmianine and gamma-fagarine, in vivo antileishmanial activitiy of the acidified extract of Haplophyllum myrtifolium Boiss. (Rutaceae) were investigated against Leishmania tropica (L. tropica), a causative agent of cutaneous leishmaniasis. All the extracts and pure compounds showed in vitro inhibitory activity against the promastigotes of. L. tropica. The in vitro 50% inhibitory concentrations of y-fagarine, acidified extract, ethanol extract, skimmianine and alkaloid extract against promastigotes were determined as 8.7, 9.4, 10.9, 25.7 and 25.8 microg/ml respectively. In vivo results of Haplophyllum myrtifolium acidified extract showed that this plant has a limited effect on decreasing the lesion size of experimental mice infected with Leishmania tropica. To the best of our knowledge, this is the first time both the in vitro and in vivo antileishmanial activity of Haplophyllum mrytifolium have been reported in the same research.

Health inequities: lower socio-economic conditions and higher incidences of intestinal parasites.

BACKGROUND: Intestinal parasitic infections affect child health and development and slow down growth, while reducing adults' productivity and work capacity. The aim of the present study was to determine and compare the incidences of intestinal parasitic infections and the socio-economic status of two near primary school children in Manisa, a western city of Turkey. METHODS: A total of 352 children were involved a questionnaire study from a private school (Ulkem Primary School - UPS, 116 children) and a community-based school (Sehzadeler Primary School - SPS, 236 children). Of these, stool samples could be obtained from a total of 294 students; 97 (83.6%) from UPS, and 197 (83.5%) from SPS. The wet mount preparations of the stool samples were examined; samples were also fixed in polyvinyl alcohol and examined with modified formalin ethyl acetate sedimentation and trichrome staining techniques. Data were analyzed using SPSS for Windows version 10.0. The chi-squared test was used for the analytic assessment. RESULTS: The percentages of the students found to be infected with intestinal parasites, were 78 (39.6%) and 13 (13.4%) in SPS and UPS, respectively. Totally 91 (31.0%) of the students from both schools were found to be infected with at least one intestinal parasite. Giardia lamblia was found to be the most common pathogenic intestinal parasite and Blastocystis hominis was prevalent independently from the hygienic conditions. The factors which significantly (p < 0.05) increase the incidence of intestinal parasites were uneducated and unemployed mother, lower social status of father, living in crowded houses with insufficient indoor spaces, using the tap water as drinking water, and living at shanty areas. CONCLUSION: Intestinal parasitic infections in school children were found to be a public health problem that increased due to lower socio-economic conditions. We conclude that organization of education seminars including the topics such as prevention of the infectious diseases, improving general hygienic conditions, and application of supportive programs for the parents may be suggested not only to reduce intestinal parasitic infections, but also to elevate the socio-cultural levels.

A province-based study using sampling method to investigate the prevalence of cystic echinococcosis among primary school children in Manisa, Turkey.

A province-based field study using a portable ultrasound scanner (US) was performed for the first time using sampling method to investigate the prevalence of cystic echinococcosis (CE) in primary school children in Manisa, Turkey. A total of 6093 children from 37 primary schools was selected as the representative sample of the total number of 166,766 primary school children, and examined by the US. Prevalence was found to be 0.15%, as nine children were diagnosed with CE, seven new and two previously operated. A questionnaire applied to the children revealed no significant relationship between the risk factors and the infection (P>0.05). In conclusion, it would be advisable to repeat the study at the same schools in eight years' time, in order to evaluate the efficacy of the control programs, since nearly all students involved in this study will have graduated by then. Besides, it is recommended to choose a sampling group to find the prevalence of an infection in a defined region.

[Evaluation of malaria cases in Manisa from 2002 to 2004]. / Manisa ilinde 2002-2004 yillari arasinda saptanan sitma olgularinin degerlendirilmesi.

The province of Manisa is a relatively well developed agricultural and industrial center in western Turkey. There is a regular and extensive influx of workers from malaria-endemic regions of Anatolia to the province of Manisa during certain periods of the year, leading to contact between the local people and incoming workers. Major seasonal elevations can be detected in the prevalence of malaria in the province of Manisa and therefore an active fight against malaria is still carried out. In the present study, malaria cases detected and reported by the Malaria Control Dispensary of the Manisa City Health Centre during the last 3 years in Manisa have been evaluated. Comparison of present and prior data of malaria prevalence in Manisa has shown a remarkable decrease in recent years. A total of 14, 5 and 5 cases were reported in 2002, 2003 and 2004, respectively. Classification of the cases according to their origin (local or immigrant), age and the time diagnosis was also presented.

The value of ultrasonography alone in screening surveys of cystic echinococcosis in children in Turkey.

A total of 1,205 primary school children were examined for cystic echinococcosis in five villages of Manisa, Turkey, to evaluate the efficacy of diagnostic methods of this infection in community-based screening surveys. Six hundred and thirty children from three villages, examined by a portable ultrasound scanner, chest microfilm and serological methods (ELISA, indirect hemagglutination) in our previous study, were designated as Study Group 1; and 575 children, from two adjacent villages, examined by ultrasonography alone in the present study, were designated as Study Group 2. In Study Group 1, hepatic cystic echinococcosis was detected in two cases (0.3%) by ultrasonography, while 43 (8.9%) and 49 (10.1%) cases were found to be positive for cystic echinococcosis by ELISA and indirect hemagglutination, respectively. Three of 575 children (0.5%) were diagnosed with cystic echinococcosis (two hepatic and one renal involvement) by ultrasonography alone in Study Group 2; and lung lesions were later detected in both cases with liver involvement by chest radiography. Our results suggested that serological tests may be beneficial in suspected cases for confirmation and differential diagnosis, but have some drawbacks, such as discrepancy in results and high false seropositivity rates. Chest microfilm is not easy in field studies and exposure to X-ray is undesirable. As a reliable, simple, inexpensive and rapid technique, ultrasonography alone is recommended to be used in community-based screening surveys for cystic echinococcosis with confirmatory tests for suspected cases found during the screening program.

[Incidence in Manisa of Trichomonas vaginalis in women with a vaginal discharge.]. / Manisa'da vajinal akintili kadinlarda Trichomonas vaginalis sikligi

Vaginal discharge caused by Trichomonas vaginalis is one of the most frequently encountered sexually transmitted infections in women. This study was planned to investigate the frequency of T. vaginalis in patients with a vaginal discharge. Vaginal specimens were collected from 233 women with a vaginal discharge and from 100 women without any discharge as a control group. Materials were taken with a speculum and sterile cotton swabs from the posterior fornix of the vagina; and inoculated into Trypticase-yeast extract-maltose (TYM) medium. T. vaginalis was detected in 11 (4.7%) samples out of the 233 specimens from women with a discharge using both diagnostic methods (direct examination and culture). No T. vaginalis trophozoites were detected in the control group.