Emerging and Re-Emerging Parasitic Infections of the Central Nervous System (CNS) in Europe.

In a rapidly evolving global landscape characterized by increased international travel, migration, and ecological shifts, this study sheds light on the emergence of protozoal and helminthic infections targeting the central nervous system (CNS) within Europe. Despite being traditionally associated with tropical regions, these infections are progressively becoming more prevalent in non-endemic areas. By scrutinizing the inherent risks, potential outcomes, and attendant challenges, this study underscores the intricate interplay between diagnostic limitations, susceptibility of specific population subsets, and the profound influence of climate fluctuations. The contemporary interconnectedness of societies serves as a conduit for introducing and establishing these infections, warranting comprehensive assessment. This study emphasizes the pivotal role of heightened clinician vigilance, judicious public health interventions, and synergistic research collaborations to mitigate the potential consequences of these infections. Though rare, their profound impact on morbidity and mortality underscores the collective urgency required to safeguard the neurological well-being of the European populace. Through this multifaceted approach, Europe can effectively navigate the complex terrain posed with these emergent infections.

Determination of Parasitic Contamination in Vegetables Collected from Local Markets in Izmir Province, Türkiye

OBJECTIVE: Fresh vegetables are an important part of a healthy and nutrient-rich diet but the consumption of raw vegetables without proper washing is the main way for transmission of parasites. This study was aimed at determining the rate of parasitic contamination in prewashed fresh vegetables sold at randomly selected 10 retail markets which is the last step to reach the consumer in Izmir, Türkiye. METHODS: A total of 80 samples selected from eight types of vegetables including tomato, spinach, lettuce, rocket, mint, parsley, dill, and cucumber were examined for parasitic agents microscopically by sedimentation method after washing samples with normal saline. Statistical analysis was performed using SPSS software version 20.0. RESULTS: Protozoan cysts, helminth eggs, and larvae were detected in 21 (26.2%) of 80 samples from eight different vegetable species. Rhabditidiform larvae 18.7%, Blastocystis spp. 5%, Toxocara spp. 2.5%; Ascaris spp., Fasciola spp., Entamoeba histolytica/ Entamoeba dispar and hooked worms were found in 1.2%. Spinach and mint samples were contaminated with parasites significantly more than other fresh vegetable samples (p<0.008, odds ratio =80.0; p<0.017, odds ratio =46.6 respectively). Cruznema spp., a plant nematode, was found at the highest rate according to the results of culture, polymerase chain reaction and sequencing, respectively. CONCLUSION: In this study, the parasitic contamination was found in approximately one of the four vegetables sold in randomly selected markets in Izmir. These findings show that vegetables sold in local markets can cause parasitic infections if they are consumed without adequate washing and awareness should be raised on this issue. In addition, it was concluded that morphological examinations should be confirmed by molecular studies and sequencing as much as possible in order to avoid misdiagnosis of rhabditidiform larvae.

A rare case of cardiac alveolar echinococcosis without any complications for eight years / Un caso raro de equinococosis alveolar cardíaca sin complicaciones durante ocho años

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First time identification of subconjunctival Dirofilaria immitis in Turkey: giant episcleral granuloma mimicking scleritis.

Dirofilariasis is a vector-borne disease that is present worldwide. This report describes a giant subconjunctival granuloma which mimics scleritis, caused by D. immitis. A 60-year-old man was referred with the complaints of irritation, redness, and swelling at the medial part of the right eye. He was living in Izmir province located in western Turkey. Slit-lamp examination showed a firm, immobile mass measuring 13.0 × 5.0 × 5.0 mm with yellowish creamy color. The mass was completely removed surgically under local anesthesia mainly for diagnosis. Histopathology revealed typical morphological features of a filarioid nematode in favor of Dirofilaria as characterized by the external smooth cuticular surface, cuticular layer, muscle layer, and intestinal tubule. Molecular study was performed using DNA isolated from paraffin-embedded tissue sections of the worm. PCR amplification and then DNA sequence analysis of cytochrome c oxidase subunit 1 (cox1) gene fragment confirmed that the worm was D. immitis. It is suggested that this may represent the first human case of D. immitis occurring in subconjunctival granuloma in Turkey. Although rare, D. immitis caused by ocular dirofilariasis in humans should be considered.

Acute Toxoplasmosis During Pregnancy: A Hard Call

Toxoplasma gondii is an obligate intracellular parasite that infects all animals, including humans, and causes toxoplasmosis. If toxoplasmosis occurs during pregnancy, it may affect the foetus owing to transplacental transmission. Such transmission may lead to foetal complications, some of which can be very serious, e.g. hydrocephaly and chorioretinitis; however, not all cases of acute toxoplasmosis during pregnancy result in foetal complications. The decision whether to continue or terminate the pregnancy is a difficult problem for families as well as healthcare professionals, thus making it important. Here we present a case of acute toxoplasmosis at 6 weeks of pregnancy. The patient was directly advised to terminate the pregnancy. However, with detailed laboratory analyses, close follow-up and treatment to prevent transplacental transmission, she successfully completed the pregnancy and eventually delivered a healthy baby. By presenting this case, we aimed to review acute toxoplasmosis during pregnancy.

Use of the ELISA (Em2-Em18) and Western Blotting Methods on Diagnosis of Alveolar Echinococcosis

Objective: Alveolar echinococcosis (AE) is one of the most lethal parasitic zoonoses in the Northern Hemisphere, and early serological detection is important to start treatment and to improve survival. A total of 50 sera samples of patients diagnosed as having various diseases were examined for by two different serological diagnostic methods. Methods: Em2-Em18 ELISA (Bordier Affinity Products, Crissier, Switzerland) and Echinococcus Western Blot immünoglobulin G (IgG) (LDBIO Diagnostics, Lyon, France) were used for analyisis. Results: A high titer of antibodies was found in 9 of 10 patients diagnosed as having AE with Em2-Em18 ELISA, in 2 of 21 patients with cystic echinococcosis, in 1 of 2 patients with fascioliasis and in 1 patient with chronic hepatitis. The Echinococcus Western Blot IgG test, used as a confirmatory test, showed IgG antibody in 85.7% (18/21) of patients with CE, while all serum samples of 10 patients with AE were evaluated as positive. This method yielded an incorrect diagnosis in the patient with chronic hepatitis and in the patient with granulomatous inflammation with caseification. Samples taken from patients with liver-related diseases and other parasitic-related diseases were found to be negative. Conclusion: The serological methods used in the study were found to be important in the early diagnosis of alveolar echinococcosis in the endemic areas, since it could be used in sero-epidemiological studies.

Effect of Extracts of the Endemic Plants Centaurea lydia and Phlomis nissolii on Toxoplasma gondii.

OBJECTIVE: This study aimed to investigate the potential antitoxoplasma activities of extracts of the endemic plants Centaurea lydia and Phlomis nissolii in a fibroblast cell culture infected with T. gondii trophozoites. METHODS: WI-38 cell lines treated with plant extracts (55 µg/mL each) and an untreated control were infected with 5×105 T. gondii trophozoites, and the number of parasites in the medium was determined on days 7, 14, and 24. RESULTS: No cytotoxic effects of C. lydia and P. nissolii extracts were detected at concentrations of 0.86-55 µg/mL in the WI-38 cell line, and the absence of the cytotoxicity of these extracts on the fibroblast cell line was considered as a positive effect. C. lydia extract at 55 µg/mL had marked activity against T. gondii trophozoites. A 47.5-fold increase was observed in the number of trophozoites in the control group, while a 84-fold decrease was found in the C. lydia extract group. However, a 36-fold increase was detected in the P. nissolii extract group, indicating no antitoxoplasma activity. CONCLUSION: The extract of C. lydia, an endemic plant, was found to be a good drug candidate for treating toxoplasmosis. The in vitro activity of the extract of this endemic plant should be further investigated in animal models in vivo.

[Transverse myelitis associated with toxocariasis and the importance of locally produced antibodies for diagnosis]. / Toksokariyaza bagli bir transvers miyelit olgusu ve tanida yerel antikor üretiminin önemi.

Toxocariasis caused by Toxocara canis or less frequently by T.catis is a common parasitic infection worldwide. Clinical spectrum in humans can vary from asymptomatic infection to serious organ disfunction depending on the load of parasite, migration target of the larva and the inflammatory response of the host. Transverse myelitis (TM) due to toxocariasis is an uncommon illness identified mainly as case reports in literature. In this report, a case of TM who was diagnosed as neurotoxocariasis by serological findings has been presented. A 44-year-old male patient complained with backache was diagnosed as TM in a medical center in which he has admitted two years ago, and treated with pregabalin and nonsteroidal drugs for six months. Because of the progression of the lesions he readmitted to another center and treated with high dose steroid therapy for three months. After six months of follow up, improvement has been achieved, however, since his symptoms reccurred in the following year he was admitted to our hospital. Magnetic resonance imaging (MRI) examination revealed a TM in a lower segment of spinal cord. He was suffering with weakness and numbness in the left lower extremity. There was no history of rural life or contact with cats or dogs in his anamnesis. Physical examination revealed normal cranial nerve functions, sensory and motor functions. There has been no pathological reflexes, and deep tendon reflexes were also normal. Laboratory findings yielded normal hemogram and biochemical tests, negative PPD and parasitological examination of stool were negative for cysts and ova. Viral hepatitis markers, anti-HIV, toxoplasma-IgM, CMV-IgM, rubella-IgM, EBV-VCA-IgM, VDRL, Brucella tube agglutination, echinococcus antibody, autoantibody tests and neuromyelitis optica test were negative. Examination of CSF showed 20 cells/mm3 (mononuclear cells), 45 mg/dl protein and normal levels of glucose and chlorine. In both serum and CSF samples of the patient Toxocara-IgG antibodies were detected by Western blot (WB) assay. Low molecular weight bands (30-40 kDa) were detected in both of the samples by repeated WB testing. CSF revealed more intense bands suggesting local antibody production. Therefore the patient was diagnosed as neurotoxocariasis, and treated with steroid and mebendazole for six weeks. Clinical improvement was detected in the case and thoracic MRI revealed significant improvement in myelitis signs two months after treatment. In conclusion, toxocariasis should be considered in the differential diagnosis of TM although the involvement of central nervous system is rare and serological testing should be performed properly in the serum and CSF samples for the diagnosis.

[Detection of Echinococcus granulosus and Echinococcus multilocularis in cyst samples using a novel single tube multiplex real-time polymerase chain reaction]. / Kist örneklerinde yeni bir tek tüp multipleks gerçek zamanli polimeraz zincir reaksiyonu ile Echinococcus granulosus ve Echinococcus multilocularis'in saptanmasi.

Cystic echinococcosis (CE) and alveolar echinococcosis (AE) caused by Echinococcus granulosus and Echinococcus multilocularis, respectively, are important helminthic diseases worldwide as well as in our country. Epidemiological studies conducted in Turkey showed that the prevalence of CE is 291-585/100.000. It has also been showed that the seroprevalence of AE is 3.5%. For the diagnosis of CE and AE, radiological (ultrasonography, computed tomography, magnetic resonance) and serological methods, in addition to clinical findings, are being used. The definitive diagnosis relies on pathological examination When the hydatid cysts are sterile or does not contain protoscolex, problems may occur during pathological discrimination of E.granulosus and E.multilocularis species. In this study, we aimed to develop a novel multiplex real-time polymerase chain reaction (M-RT-PCR) targeting mitochondrial 12S rRNA gene of E.granulosus and E.multilocularis using Echi S (5'-TTTATGAATATTGTGACCCTGAGAT-3') and Echi A (5'-GGTCTTAACTCAACTCATGGAG-3') primers and three different probes; Anchor Ech (5'-GTTTGCCACCTCGATGTTGACTTAG-fluoroscein-3'), Granulosus (5'-LC640-CTAAGGTTTTGGTGTAGTAATTGATATTTT-phosphate-3') and Multilocularis (5'-LC705-CTGTGATCTTGGTGTAGTAGTTGAGATT-phosphate-3') that will enable the diagnosis of CE and AE in same assay. During M-RTR-PCR, plasmids containing E.granulosus (GenBank: AF297617.1) and E.multilocularis (GenBank: NC_000928.2) mitochondrial 12S rRNA regions were used as positive controls. Cysts samples of patients which were pathologically confirmed to be CE (n: 10) and AE (n: 15) and healthy human DNA samples (n: 25) as negative control as well as DNA samples of 12 different parasites (Taenia saginata, Hymenolepis nana, Trichuris trichiura, Fasciola hepatica, Enterobius vermicularis, Toxoplasma gondii, Pneumocystis jirovecii, Trichomonas vaginalis, Cryptosporidium hominis, Strongyloides stercoralis, Plasmodium falciparum, Plasmodium vivax) were used to develop M-RT-PCR. E.granulosus and E.multilocularis control plasmids were constructed to detect analytic sensitivity of the test using TOPO cloning. Positive control plasmids were diluted to determine analytical sensitivity and specificity by distilled water at 10(6)-10(5)-10(4)-10(3)-10(2)-10(1)-1 plasmid copy of dilution in each reaction. According to the results, analytical sensitivity of the assay for E.granulosus and E.multilocularis was 1 copy plasmid/µl reaction. The non-existence of cross reactivity with 12 different parasites' DNA samples showed the analytical specificity of the assay. Displaying Echinococcus DNA in cyst samples among 25 patients and species discrimination as well as non-existence of cross reactivity with human DNA samples showed that the clinical sensitivity and specificity of the assay were 100%. As a result, the M-RT-PCR developed in the present study provided a sensitive, specific, rapid, and reliable method in the diagnosis of echinococcosis and the discrimination of E.granulosus and E.multilocularis from cyst samples.

Gastrointestinal Parasitosis: Histopathological Insights to Rare but Intriguing Lesions of the Gastrointestinal Tract.

OBJECTIVE: Gastrointestinal parasitosis is a significant cause of morbidity and mortality. Definitive diagnosis is usually made by stool tests and/or serology but may require tissue evaluation. Although pathologists are usually familiar with common parasites, it is not well established whether the diagnosis could be suspected without seeing the "parasite" itself. MATERIAL AND METHOD: Resection or biopsy specimens of 32 cases with Giardia intestinalis (n=20), Enterobius vermicularis (n=5), Entamoeba histolytica (n=4), Fasciola hepatica (n=1), Strongyloides spp. (n=1) and Taenia saginata (n=1) infections were retrospectively re-evaluated for accompanying mucosal changes, and compared with nonparametric tests. RESULTS: The most common changes were congestion (65.6%) and eosinophilic infiltration (50%). Chronic active mucosal inflammation accompanied 37.5% of the cases. More than 10 eosinophils/HPF were present in 43.8%. Only one case of G. intestinalis, E. vermicularis, E. histolytica, and F. hepatica showed more than 50 eosinophils/HPF. Mucosal architectural abnormalities were present in 34.4%. Granulomas, giant cells and Charcot-Leyden crystals were only seen accompanying F. hepatica. No statistically significant difference was found between parasite subspecies regarding presence of inflammation, lymphoid aggregates, architectural distortion, congestion, ulceration and increase of eosinophils. CONCLUSION: Parasites induce nonspecific inflammation, slight mucosal architectural changes, mild eosinophilic infiltrate or granuloma formation. They may cause ulceration, bowel obstruction or perforation. Parasitosis should also be considered when evaluating cases mimicking inflammatory bowel disease, celiac disease or those that do not fulfill diagnostic criteria.

Seroepidemiological Study of Toxocariasis among Volunteers Animal Husbandry Workers and Veterinary in Southern Anatolia in Turkey in 2014.

BACKGROUND: Human toxocariasis is a parasitic infection caused by the larvae of Toxocara canis. We examine the Toxocara seroprevalance in veterinarians and animal husbandry workers living in the Mugla Province, Turkey to evaluate better the risk factors for Toxocara exposure. METHODS: In 2014, 376 volunteers participated in the study in 2014. All blood specimens were tested using a commercial enzyme immunoassay kit and ELISA positive samples were confirmed by Western Blot (WB) method. RESULTS: The seroprevalence of Toxocara, as determined by WB, was 8%. A statistically significant correlation was evident between patient age and Toxocara positivity among animal husbandry workers (P = 0.029). A strong association was also evident between sex and seropositivity in the animal husbandry group (P=0.024). Veterinarians working in pet clinics did in fact exhibit higher Toxocara seropositivities relative to those of other groups (P = 0.029). A statistically significant difference was detected between the rural geographic areas surveyed (P = 0.04). CONCLUSION: In Mugla Province, seroprevalence of Toxocara is lower than other regions. Despite the low seroprevalence observed, especially in high risk professions toxocariasis remains an important medical concern within the region.

[A rarely seen cause of brain abscess: neurotoxocariasis]. / Nadir görülen bir beyin apsesi etkeni: Nörotoksokariazis.

Toxocariasis in man is associated with three syndromes which are visceral larva migrans, ocular larva migrans and covert toxocariasis. Although neurotoxocariasis is defined as the fourth syndrome of toxocariasis, it is usually considered as a neurological disease which is usually concomitant with visceral larva migrans. In this report, a case of brain abscess caused by toxocariasis was presented. A 56 years-old female patient was admitted to our hospital with headache, pain referring to right side of her face and teeth, numbness of forth and fifth finger of her right hand. Cranial diffusion weighted, dynamic magnetic resonance imaging (MRI) revealed a few non-specific intensities at supratentorial white matter, an approximately 13 x 12 mm lesion without contrast enhancement and a significant edema around the white matter in the left frontal cortex. Histologic examination after stereotactic biopsy of the lesion revealed diffuse histiocyte infiltration. A specific agent could not be detected in the histochemical examination. Western-blot test for toxocariasis in serum and cerebrospinal fluid samples were found positive. She was transferred to the infectious diseases clinic, and albendazole therapy (400 mg, q12h) was started. Albendazole treatment was completed for a total of one month following the regression of the cranial MRI findings on the 14th day of therapy. The patient is recalled for cranial MRI control three months later. However, it was noted that she continued albendazole for three months. Compared to the previous MRI, there were two stabilized T2A hyperintense lesions in left cranial hemisphere and minimally regressed lesions at the level of left frontal centrum semiovale. The patient was successfully treated with albendazole. There was no relapse after six month follow-up. This case was presented to withdraw attention to neurotoxocariasis which may be encountered although rarely in the etiology of encephalitis/ brain abscess.

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.

Isolation of Toxoplasma gondii strains similar to Africa 1 genotype in Turkey.

INTRODUCTION: Toxoplasma gondii is a protozoon parasite that has a worldwide dissemination. It can cause serious clinical problems such as congenital toxoplasmosis, retinochoroiditis, and encephalitis. Currently, T. gondii genotypes are being associated with these clinical presentations which may help clinicians design their treatment strategy. CASE REPORTS: Two T. gondii strains named Ankara and Ege-1 were isolated from newborns with congenital toxoplasmosis in Central and Western Anatolia, respectively. Ankara and Ege-1 strains were isolated from the cerebrospinal fluid of newborns. According to microsatellite analysis, Ankara and Ege-1 strains were sorted as Africa 1 genotype. CONCLUSION: T. gondii strains isolated in Turkey were first time genotyped in this study. Africa 1 genotype has previously been isolated in immunosuppressed patients originating from sub-Saharan Africa. The reason of detecting a strain mainly detected in Africa can be associated with Turkey's specific geographical location. Turkey is like a bridge between Asia, Europe and Africa. Historically, Anatolia was on the Silk Road and other trading routes that ended in Europe. Thus, detecting Africa 1 strain in Anatolia can be anticipated. Consequently, strains detected mainly in Europe and Asia may also be detected in Anatolia and vice versa. Therefore, further studies are required to isolate more strains from Turkey.

Action plan to regain unnecessary deferred blood donors due to malaria risk in Turkey.

Malaria was expected to be a major problem during blood donation in Turkey due to existence of malaria cases in southeastern region of Turkey. The present study aimed for the first time, to investigate malaria in "donors deferred for malaria risk" and to determine the regional rates of malaria deferral in Turkey. Blood samples were collected from several Blood Banks of southeastern provinces where local malaria cases still exist and from Blood Bank of Ege University Medical School (EUMS) located in western Turkey where malaria is eradicated decades ago. Plasmodium spp. and specific antibodies were investigated by stained smears, antigen detection, PCR and ELISA. Among the donors deferred for malaria risk, Plasmodium spp. were not detected by microscopy, PCR or antigen detection. Seroprevalances were 2% and 3.92% in western and southeastern regions, respectively. Rate of donor deferral for malaria risk was 0.9% in EUMS and deferrals were exclusively because of travel to southeastern Turkey. In southeastern provinces, deferrals were mainly due to malaria like fever history. The present study first time assessed regional rates of donor deferral due to malaria risk in Turkey. Previously, malaria was expected to be a major problem during blood donation in Turkey due to existence of malaria cases in southeastern region of Turkey. The results of the study showed that 97% of the deferrals were unnecessary. In conclusion, to reduce unnecessary donor deferrals in Turkey, in addition to comprehensive questioning for malaria history, the usage of a malaria antibody screening method should be initiated prior to deferral decision.

Serological monitoring of paediatric visceral leishmaniasis By IFA and ELISA methods.

OBJECTIVE: Visceral leishmaniasis (VL) is endemic in all Mediterranean countries including Turkey, and children are at greater risk than adults in endemic areas. In VL patients, serological assays are considered to be sensitive for the diagnosis and/or follow up. The aim of this study was to assess the usefulness of serology for following up of childhood VL in Turkey. METHODS: Sera obtained from twenty parasitologically confirmed children with VL were tested using IFAT and ELISA. The patients were monitored clinically and serologically (range: 20-500 days) during and after treatment. All VL patients were treated with meglumine antimonate. RESULTS: Anti-Leishmania antibodies in successfully treated VL patients showed a steep decline but, in three patients who had relapsed, an increase was detected. Significantly lower values were observed after treatment with both serological techniques. Mean ELISA optical density values before and after treatment were: 0.78 ± 0.36 (0.26-1.76) and 0.38 ± 0.24 (0.09-0.83) respectively, (p < 0.001) and mean IFAT values (log10 transformed titers) before and after treatment were: 3.02 ± 0.90 (1.81-4.51) and 2.16 ± 0.75 (1.20-3.90) respectively, (p < 0.001). CONCLUSION: ELISA and IFAT are valuable not only for diagnosis but also for monitoring of drug therapy in childhood visceral leishmaniasis as rapid and non-invasive techniques.

Fasciola hepatica infection in echinococcosis suspected cases.

OBJECTIVE: Fasciola hepatica, a liver fluke of sheep and cattle, may accidentally infect humans. The main signs and symptoms of the fasciolosis are eosinophilia, abdominal pain and hepatomegaly and may also be attributed to Cystic Echinococcosis (CE) caused by Echinococcus granulosus, which is a prevalent infection in Turkey. METHODS: Sera samples of 226 CE suspected cases were tested for anti-F. hepatica antibodies by an excretory secretory ELISA (ES-ELISA) and for anti-E. granulosus antibodies by ELISA and indirect hemagglutination (IHA) tests. Cases which were seropositive for fasciolosis were further evaluated radiologically and examined for F. hepatica eggs. RESULTS: Five (2.2%) and 96 (42.4%) of the 226 CE suspected cases were found seropositive for fasciolosis and CE, respectively. Although the radiological findings strongly suggested that there was fasciolosis in three cases, F. hepatica eggs were detected in two patients only. CONCLUSION: These data suggest that human fasciolosis is not as rare as previously reported in Turkey. F. hepatica infection should be suspected especially in the presence of eosinophilia, abdominal pain and liver lesions.

Identification of potential serodiagnostic and subunit vaccine antigens by antibody profiling of toxoplasmosis cases in Turkey.

Toxoplasmosis, caused by infection of the protozoan parasite Toxoplasma gondii, is associated with mild disease in healthy individuals, whereas individuals with depressed immunity may develop encephalitis, neurologic disorders, and other organ diseases. Women who develop acute toxoplasmosis during pregnancy are at risk of transmitting the infection to the fetus, which may lead to fetal damage. A diagnosis is usually confirmed by measuring IgG, or IgM where it is important to determine the onset of infection. A negative IgM result essentially excludes acute infection, whereas a positive IgM test is largely uninterpretable because IgM can persist for up to 18 months after infection. To identify antigens for improved diagnosis of acute infection, we probed protein microarrays displaying the polypeptide products of 1357 Toxoplasma exons with well-characterized sera from Turkey. The sera were classified according to conventional assays into (1) seronegative individuals with no history of T. gondii infection; (2) acute infections defined by clinical symptoms, high IgM titers, and low avidity IgG; (3) chronic/convalescent cases with high avidity IgG but persisting IgM; (iv) true chronic infections, defined by high avidity IgG and no IgM. We have identified 38 IgG target antigens and 108 IgM target antigens that can discriminate infected patients from healthy controls, one or more of which could form the basis of a 'tier-1' test to determine current or previous exposure. Of these, three IgG antigens and five IgM antigens have the potential to discriminate chronic/IgM persisting or true chronics from recent acutely infected patients (a 'tier-2' test). Our analysis of the antigens revealed several enriched features relative to the whole proteome, which include transmembrane domains, signal peptides, or predicted localization at the outer membrane. This is the first protein microarray survey of the antibody response to T. gondii, and will help in the development of improved serodiagnostics and vaccines.

Toxoplasma gondii RH Ankara: production of evolving tachyzoites using a novel cell culture method.

Toxoplasma gondii is one of the most researched parasite due to its easy growth both in vitro and in vivo. Tachyzoites, derived from mouse or rat peritoneum encounters ethical and economical problems when used for research or diagnostic purposes. Currently, research has focused on determining the most suitable cell culture environment to reach highest amount of viable tachyzoites with least host cell contamination. However, gene expression changes that take place throughout the adaptation of evolving T. gondii strains to continuous cell cultures appear as a problem. The present study aimed to determine a novel cell culture strategy for T. gondii RH Ankara strain tachyzoites to harvest abundant tachyzoites with least host cell contamination and minimal antigenic variation at predetermined dates to use as an antigen source in serological assays that will facilitate reduction in animal use. To achieve this purpose, T. gondii RH Ankara strain tachyzoites were incubated with HeLa cell at different ratios for two or three days. In all flasks incubated for two days, viability rate reached to 100% and HeLa cell contamination decreased to levels between 0.12-0.5×10(6)/ml. In the flasks with HeLa-tachyzoite ratio 1/8, the tachyzoite yield and viability ratio were 3×10(6)/ml and 100%, respectively, with accompanying 10 fold decrease (0.12×10(6)/ml) in HeLa contamination. During continuous production, highest tachyzoite yield was obtained from the first passage (3.55×10(6)/ml) and until the end of third subculture viability rates and HeLa cell contaminations were between 98.2-99.4% and 0.31-0.37×10(6)/ml, respectively. ELISA, IFA and Western blot analyses showed that the quality, specificity and sensitivity of the antigen harvested from the first passage of cell culture performed at two days intervals were comparable to the antigen harvested from mice and decreased in the following subcultures. Overall, these results demonstrated that T. gondii RH Ankara strain is still evolving to adapt to cell culture environment and therefore such strains continuously produced in cell cultures should be avoided for serological assays. However, the two day short interval cell culture method described herein offers a chance to reduce the animal use intended for the preparation of serological assays' antigen from local evolving strains.