Genetic diversity of Pneumocystis jirovecii isolates among Turkish population based on mitochondrial large subunit ribosomal RNA and dihydropteroate synthase gene typing.

Pneumocystis jirovecii (P. jirovecii) is an atypical fungus that can cause severe interstitial pneumonia in immunocompromised patients. In this study, mitochondrial large subunit ribosomal RNA (mtLSU-rRNA) and dihydropteroate synthase (DHPS) gene polymorphism in P. jirovecii isolates were investigated in Western Turkey's Izmir province and its surroundings. For this purpose, a total of 157 P. jirovecii isolates obtained from bronchoalveolar lavage samples of hospitalized cases and lung tissue samples of autopsy cases who died outside hospital were examined. Genotypes were identified by direct sequencing of mtLSU-rRNA restriction fragment length polymorphism analysis of the DHPS gene amplicons. The mtLSU-rRNA analysis revealed that genotype 2 was the most common genotype with 58%. The following genotypes were genotype 3 (13%), genotype 1 (11.6%) and genotype 4 (5.1%), while genotype 5 (0.7%) was detected in only one autopsy case. In addition, 16 (11.6%) cases had dual or triple different genotypes (mixed infection). It was observed that the genotype distribution was not affected by characteristics such as age, gender and immune status. However, the predominance of genotype 2 in solid organ tumors and the predominance of mixed infection in patients with chronic pulmonary disease were statistically significant. On the other hand, DHPS gene amplification was positive in 137 (87.3%) of 157 samples. While no mutation was observed in 135 samples, the association of wild-type and 57th codon mutation was detected in two hospitalized cases (1.5%). In this study, important epidemiological data on the distribution of mtLSU-rRNA genotypes were obtained. Also the existence of DHPS gene mutations associated with potential drug resistance in our community was shown for the first time. Further studies are needed to evaluate the possible effects of genotypes on the prognosis of the disease to help with the clinician's treatment decisions. LAY ABSTRACT: Pneumocystis jirovecii (P. jirovecii) is an atypical fungus that can cause life-threatening pneumonia in immunocompromised patients. In this study, we investigated the mtLSU-rRNA and DHPS gene polymorphisms in P. jirovecii isolates from both hospital and autopsy cases.

Prevalence of Pneumocystis jirovecii colonization in autopsy cases in Turkey.

Detection of Pneumocystis jirovecii and its DNA in clinically asymptomatic people is defined as colonization. The aim of this study was to reveal the colonization prevalence of P. jirovecii and affecting factors in an immunocompetent population. The study included 200 cases undergoing forensic autopsy between February 2015 and April 2015. The cause of death was non-medical conditions (group 1) in 111 cases (55.5 %), medical conditions (group 2) in 73 cases (36.5 %) and undetermined (group 3) in 16 cases (group 3). Tissue specimens about 1 g in weight were taken from the right upper pulmonary lobe. After DNA extraction, nested PCR targeting mitochondrial large subunit rRNA was used to detect P. jirovecii. Of 200 cases, 37 (18.5 %) had P. jirovecii DNA. There was not a significant difference in place of living, gender, smoking status and medication use between the cases with P. jirovecii and those without P. jirovecii. A significantly high rate of P. jirovecii colonization was detected in group 2 (χ²=7.674; P=0.022). P. jirovecii-colonized cases also had a chronic disease in 2 of 13 (group 1), 12 of 20 (group 2) and 1 of 4 (group 3) cases (χ²=5.571; P=0.062). A significantly high rate of the cases aged 0-1 year had P. jirovecii (5/11; 45.5 %) (χ²=5.639; P=0.018). The results of the study suggest that infants and patients with chronic diseases like cardiac or pulmonary diseases can be at risk for P. jirovecii colonization.

[Pneumocystis jirovecii Pneumonia in a Patient with Interstitial Lung Disease]. / Interstisyel Akciger Hastaligi Zemininde Gelisen Pneumocystis jirovecii Pnömonisi.

Pneumocystis pneumonia (PCP) caused by Pneumocystis jirovecii (P. jirovecii) is an opportunistic pulmonary infection that occurs in immunocompromised patients. Here, a 49-year-old female patient who was admitted to our hospital with respiratuary distress and whose bronchoalveolar lavage (BAL) fluid specimens had P. jirovecii and Aspegillus fumigatus was presented. She had been treated with corticosteroids because of interstisial lung disease and she was also diabetic. It is important to define the coinfection developed in the presence of immunosuppression.

In vitro effects of resveratrol on Trichinella spiralis.

Resveratrol, a natural phytoalexin found mainly in grapes, possesses antibacterial, antifungal, and antiviral properties. However, there is no information about its effects on helminths such as Trichinella sp. In the present study, we investigated the effects of resveratrol on the viability of Trichinella spiralis life stages in vitro. Adult forms, newborn larvae (NBL), and muscle larvae (ML) were incubated with resveratrol at concentrations varying from 12.5 to 200 microg/ml. Resveratrol showed significant anthelmintic activity against NBL and adult forms of Trichinella, but not against ML. Our results suggest that resveratrol may be useful as a therapeutic agent to treat trichinellosis in early stages and warrant its further assessment in animal models of disease.

Identification of Acanthamoeba genotype T4 and Paravahlkampfia sp. from two clinical samples.

In this study, two free-living amoebae strains, Acanthamoeba genotype T4 and Paravahlkampfia sp., which were isolated from keratitis cases are presented. While the Acanthamoeba strain was isolated as a single agent, the Paravahlkampfia strain was found together with herpes simplex virus. Neither of the patients were contact lens wearers, but they did have a history of minor corneal trauma. Amoebae were detected on non-nutrient agar covered with Escherichia coli. Based on PCR-amplified 18S rRNA-gene analysis the first isolate was identified as Acanthamoeba genotype T4 and the second as Paravahlkampfia sp. In thermotolerance tests, the maximum temperature at which trophozoites continued to divide was determined as 37 degrees C for this Acanthamoeba strain and 35 degrees C for the Paravahlkampfia strain. To the best of our knowledge, the Acanthamoeba strain described herein is the second molecularly identified Acanthamoeba strain in an Acanthamoeba keratitis patient in Turkey. However, the Paravahlkampfia isolate is believed to be the first strain that has been isolated from a keratitis patient and has been molecularly differentiated from Vahlkampfia.

[The evaluation of the techniques used for diagnosis of Entamoeba histolytica in stool specimens]. / Diskida Entamoeba histolytica'nin saptanmasinda kullanilan yöntemlerin birlikte degerlendirilmesi.

In this study, stool samples of 9378 patients from different clinics, who presented at the laboratory of the department of parasitology of the Dokuz Eylul University, Faculty of Medicine with several gastrointestinal complaints from January 2004 to May 2006, were examined. All stool samples were examined with the saline-Lugol method and, in suspicious cases, by trichrome staining, cultivation in Robinson's medium and/or antigen detection in stool with the Entamoeba CELISA Path kit. Forty-one cases (0.44%), in which Entamoeba histolytica/Entamoeba dispar cysts and/or trophozoites were detected by at least one method, were found to be positive. Out of these 41 cases, four methods were used in 24 cases, three methods in 14 cases, whereas only saline-Lugol and trichrome staining methods were used in 3 cases. Even though all 41 positive cases had been examined with the saline-Lugol method, only 25 cases were found to be positive with this method for E. histolytica/E. dispar cysts and/or trophozoites. The remaining 16 cases were diagnosed by the other three methods. Today it is necessary to distinguish E. histolytica from E. dispar because the patient does not need to be treated if E. dispar is identified whereas if E. histolytica is identified the patient needs urgent treatment. That's why it is necessary to get reliable results using diagnostic methods together and, when needed, by ELISA specific for E. histolytica.

[Cellular immune response of patients with anthroponotic cutaneous leishmaniasis in Sanliurfa]. / Sanliurfa'da antroponotik kutanöz leishmaniasis hastalarinin hücresel immun cevabi.

Patients with cutaneous leishmaniasis (CL) have strong delayed-type hypersensitivity and in vitro proliferative responses to leishmanial antigens during active and cured diseases. To define the T cell response in patients with antroponotic CL infected with L. tropica is important to clarify the immunopathologic nature of the disease. T cell responses of acute and of already healed CL patients were defined using IFN-gamma, IL-5, IL-4, IL-10 cytokine measurement assays. In this study, while Th2 cell response was found to be dominant in active CL cases, Th1 cell response was more distinctive in the group of already healed CL cases. Differentiation of cellular response in the different stages of infection might be helpful in understanding the prognosis of leishmaniasis.

Evaluation of antigenic variations between two virulent toxoplasma strains.

Toxoplasma gondii infection in humans is routinely assessed by serological means. Here, the authors attempted to compare the response of different Toxoplasma strains to serological tests and to evaluate the antigenic profiles of the RH and RH Ankara (TRH) strains with Western blotting. Anti-Toxoplasma IgG antibodies of 72 patients were examined with the indirect immunofluorescence antibody (IFA) test, ELISA and Western blotting (WB) by using antigen from both strains. Antigenic variations between strains did not affect IFA and ELISA test results, but qualitative and quantitative differences between the WB patterns were observed. A number of bands with molecular masses varying between 17 and 105 kDa were detected in WB. Fourteen different bands were obtained with the assay performed with RH strain antigen. An additional four bands were observed with TRH strain antigen. Also, an 80 kDa band was observed to stain darker in the blot with TRH strain antigen, whereas with RH strain antigen 30 and 38 kDa bands were darker. The results showed that strain-specific polymorphism in tachyzoite antigens of different Toxoplasma strains is important in the evaluation of WB but not in conventional serological analyses such as ELISA and IFA.

Albendazole: single or combination therapy with permethrin against pediculosis capitis.

Pediculosis capitis is a worldwide problem and a growing concern because of resistance to pediculicides. In the present study, we investigated whether albendazole could be used in the treatment of pediculosis capitis in combination with 1% permethrin or alone. A total of 150 children were randomly divided to five groups of 30 each. Group 1 got albendazole in a single dose (400 mg), group 2 got albendazole at 400 mg for 3 days, group 3 was given 1% permethrin, group 4 took 1% permethrin and albendazole in a single dose (400 mg), and group 5 got 1% permethrin and albendazole in a dose of 400 mg for 3 days. Groups given albendazole were also given another 400 mg dose of albendazole after 1 week. The success rate of treatment at the 2-week follow-up for all groups was 61.5%, 66.6%, 80.0%, 84.6%, and 82.1%, respectively. No statistically significant difference was found between the groups. The results of this study suggest that albendazole is effective against pediculosis capitis and there is no synergistic effect between albendazole and 1% permethrin.

[Evaluation of patients presenting with a suspicion of cystic echinococcosis to the serology laboratory of the Parasitology Department of Dokuz Eylül University Medical Faculty]. / Dokuz Eylül Universitesi Tip Fakültesi Parazitoloji Anabilim Dali Seroloji Laboratuvari'na Kistik Ekinokokkozis süphesiyle basvuran hastalarin degerlendirilmesi.

Cystic echinococcosis (CE) is a zoonosis, caused by the metacestode form of Echinococcus granulosus. The diagnosis of CE is difficult using the clinical features of the disease and it depends on the combination of serological methods aimed at determining the specific antibody response and on imaging techniques that support the serology. In this study, 465 patients who presented with a suspicion of CE to the serology laboratory of the Parasitology Department of Dokuz Eylul University between January 2003 and June 2004 were evaluated. The specific anti-E. granulosus antibodies in the serum samples of patients were examined by an in-house enzymelinked immunosorbent assay (ELISA) test and a commercial indirect hemagglutination (IHA) test. Seventy eight (17%) of the patients were positive with ELISA and 65 (14%) of them were positive with IHA with varying titrations. Fifty six (12%) of the sera were positive with both methods. Fifty-six of the eighty patients who underwent CE surgery at different times before this study were found to be seropositive, while the remaining 24 were found to be seronegative. We concluded that it is very important to combine at least two methods in the serological diagnosis of CE.

[Evaluation of three different ELISA kits for the diagnosis of trichinellosis]. / Trisinellozis tanisinda uç farkli ELISA kitinin degerlendirilmesi.

Human trichinellosis caused by Trichinella spp. is a widely distributed parasitic disease, acquired by ingestion of undercooked or raw meat containing larvae of the parasite. In January 2004, a trichinellosis outbreak following the consumption of raw minced meat ball, occurred in Izmir, Turkey. In the present study, an in-house excretory/secretory (ES) IgG ELISA and two commercial IgG ELISA kits (Kit # I; IBL, Hamburg, kit # II; Cypress, Belgium) have been used to evaluate their diagnostic efficiencies in this outbreak. Serum samples were collected from 100 patients with trichinellosis, from 16 patients with other parasitosis and autoimmune diseases (2 toxocariasis, 8 hydatic cyst, 1 fascioliasis, 5 rheumatoid arthritis) and from 20 healthy subjects. While the sensitivity of in-house ELISA has been found as 100%, the sensitivities of commercial kits number I and II have been found as 75% and 48%, respectively. The specificities of these three tests have been detected as 93.7%, 87.5% and 100%, respectively. The results obtained in this study suggest that in-house ELISA is of significant diagnostic value for the diagnosis of trichinellosis.

[An imported case of mixed malaria caused by Plasmodium falciparum and Plasmodium ovale.]. / Plasmodium falciparum ve Plasmodium ovale'nin etken oldugu importe bir miks sitma olgusu.

Malaria caused by Plasmodium species is an important parasitic infection in Turkey as in the rest of the world. Malaria cases originating in our country are caused by P. vivax; those caused by other Plasmodium spp. are imported cases. In this article, after work-related travel to Cameroon, a patient who acquired specific clinical signs and symptoms of malaria has been evaluated. The major clinical findings of the patient were fever, chills and shaking. After examination of thin and thick blood smears prepared from the peripheral blood of the patient, a 20% rate of Plasmodium parasitemia was obtained and the case was considered to be a mixed P. falciparum and P. ovale infection. In addition, P. falciparum infection was confirmed using the Optimal Malaria Rapid Test and the presence of another Plasmodium species besides P. falciparum was confirmed. Primaquine was added to quinine and doxycycline therapy for P. ovale hypnozoites. No Plasmodium was found in control blood smears after clinical improvement. In this case, it has been emphasized that in differential diagnosis of fever seen after travel to malaria endemic areas, malaria must be considered and prophylaxis must be carried out before travel.

[Use of the Western Blot assay in the diagnosis of trichinosis.]. / Trichinellosis tanisinda Western Blot tekniginin uygulanmasi

Trichinosis is a worldwide parasitic disease acquired by the consumption of raw or uncooked meat containing encysted larva. The western blot (WB) technique has an important role in the serological diagnosis of trichinosis as a confirmatory test. In our study, IgG antibodies specific for Trichinella were found in the sera of 25 patients with trichinosis seropositivity by In house ES ELISA. Of 10 control individuals, 2 were found to be positive for toxocariasis, 2, positive for fascioliasis and 6 were found to be healthy by the WB test. In WB analysis, serum samples from the 25 persons suffering from trichinosis showed common profile bands with the band size ranging from 14 to 123 kDa. These serum samples showed the following important three antigenic bands with different sensitivities of 45 kDa (88.0%), 55 kDa (60%), and 88-90 kDa (64%). As a result of this study, these three antigenic bands in WB test are considered to be valuable in the diagnosis of human trichinosis.

Interleukin (IL) 5 levels and eosinophilia in patients with intestinal parasitic diseases.

AIM: Intestinal parasitic diseases are commonly accompanied with diarrhoeal symptoms and allergic reactions. Eosinophilia occurs as a result of IL-5 synthesized from Th2 cells during allergic reactions. IL-5 acts as a factor activating eosinophils. The aim of this study was to compare the IL-5 cytokine measurements in serum samples and cell cultures. And also to compare eosinophilia observed in helminth infections and protozoon infections accompanied with allergy. METHODS: Twenty-three patients who presented with diarrhoeal symptoms and allergic complaints were tested positive for intestinal parasites, as well as 21 controls with allergic complaints who did not have any intestinal parasites were included in this study. IL-5 production in in vitro cell cultures prepared by using phytohemaglutinin (PHA) to stimulate peripheral blood mononuclear cells (PBMC) obtained from the blood samples taken from these patients were compared with the IL-5 level in serum. Furthermore, the IL-5 production in protozoon and helminth infections was also compared. Absolute eosinophil values in 1 mm(3) of blood were calculated by means of peripheral smear in both groups within the scope of the study. RESULTS: Parasites such as helminth detected in 15 (65.2%) and protozoon in 8 (34.8%) of the patients were included in this study. As regards the values of the sera in both patients with parasite infection and controls, the IL-5 production was found to be higher in the cell culture supernatant (P<0.001 and P<0.05). When the IL-5 level of the patients with helminth parasites was compared with that of those with protozoon, it was determined that the IL-5 level in serum was more significant in the patients with protozoon than in those with helminth (P<0.05). In the study group, the patients were found to have parasites, the percentage of eosinophil was 7.0% compared to 6.5% in the control group. Thus, there was no significant difference between the eosinophil values (P>0.05). CONCLUSION: It was found that IL-5 cytokine levels in serum samples from the patients with helminth and protozoon displayed more measurable values as compared to the IL-5 levels after stimulation with mitogen. It is concluded that IL-5 acts as a triggering factor in the toxiallergic complaints commonly seen in helminth and protozoon infections.