Iranian Journal of Parasitology
Volume:8 Issue: 1, Jan-Mar 2013

The children aged under 5 years from vast African areas badly suffer from falciparum malaria and many of them die of this disease. Therapeutic efficacy of anti-malaria drugs, especially pyrimethamine-sulfadoxine (PS) and chloroquine (CQ) to falciparum malaria is frequently evaluated and reported in recent 10 years. Unfortunately, to date, these widespread materials and researches have not been systematically collected and analyzed. In our study, two investigators were employed to widely and independently gather researches on efficacy of PS vs. CQ mono-therapy of falciparum malaria in children aged below 5 years in unpublished and published databases. Meta-analyses were conducted in categories of PS group and CQ group respectively. Pooled OR of PS vs. CQ was 0.11 (95%CI, 0.05-0.24). PS showed higher therapeutic efficacy to falciparum malaria in less-than-5-year children than CQ. Random model was chosen to analyze for the heterogeneity existence between different studies. Subgroup analyses were performed, but heterogeneity was still presented. Heterogeneity might be caused by different resistance of falciparum malaria to PS and CQ in different settings. Malaria type associated with parasite species, basic information of PS and CQ, and PS & CQ resistant malaria control measures were demonstrated and discussed respectively in detail in this article.

Many parasitic helminthes of veterinary importance have genetic features that favor development of anthelmintic resistance, this becoming a major worldwide constrain in livestock production. The develop­ment of anthelmintic resistance poses a large threat to future production and welfare of graz­ing animals. Development of variable degrees of resistance among different species of gastrointes­tinal nematodes has been reported for all the major groups of anthelmintic drugs. It has been ob­served that frequent usage of the same group of anthelmintic; use of anthelmintics in sub-optimal doses, prophylactic mass treatment of domestic animals and frequent and continuous use of a single drug have contributed to the widespread development of anthelmintic resistance in helminthes. The degree and extent of this problem especially with respect to multidrug resistance in nematode popula­tions is likely to increase. Maintaining parasites in refugia and not exposed to anthelmintics, seems to be a key point in controlling and delaying the development of resistance, because the suscepti­ble genes are preserved. Targeted selective treatments attract the interest of scientists to­wards this direction. Additionally, adoption of strict quarantine measures and a combination drug strategy are two important methods of preventing of anthelmintic resistance. Experience from the development of anthelmintic resistance suggests that modern control schemes should not rely on sole use of anthelmintics, but employ other, more complex and sustainable recipes, including parasite resistant breeds, nutrition, pasture management, nematode-trapping fungi, antiparasitic vaccines and botanical dewormers. Most of them reduce reliance on the use of chemicals and are environmental friendly. Finally, if new anthelmintic products are released, an important question will be raised about how they should be used. It is suggested that slowing the development of resistance to a new class are likely to be gained by releasing it in combination with one or more of the older anthelmintic classes, especially where efficacy of the older active(s) remains high.

Malaria remains a serious public health problem with significant morbidity and mortal­ity. This study was conducted to identify whether ficolin-A could play an active role of against mala­ria infection. The function of ficolin-A was analyzed in mouse model. The open reading frame of fico­lin-A was cloned from the liver of new born C57BL/6 mice by RT-PCR and then inserted into the expression vector of eukaryon to construct pVAX1-ficolin-A plasmid. Meanwhile, the open reading frame of the 19-kDa fragment of merozoite surface protein-1 of Plasmodium berghei (MSP119) was cloned and then the expression vector of eukaryon, pVAX1- MSP119 was constructed. Both recombi­nant vectors were used in the mouse model of infection by Plasmodium berghei. pVAX1-ficolin-A alone could not significantly suppress parasite density and prolong sur­vival time of infection mice; however, when injected pVAX1-ficolin-A and pVAX1- MSP119 together, the percent of invasion by Plasmodium was decreased (from 43.78% to 22.23% at 10 day after infec­tion, compared to vector) and the survival time was prolonged significantly in the infection mouse model (P=0.01). Ficolin-A can enhance the immunoprotection of MSP119, it implies ficolin-A may be used as immunoenhancer in the study of vaccine defending malaria.

Toxoplasma gondii is an obligate intracellular parasite that infects humans at high preva­lence rates. The virulent RH strain of T. gondii is generally considered to have lost its cyst forming capacity. This study performed to obtain tissue cysts in mice infected with tachyzoites of RH strain treated with sulfadiazine (SDZ). It provides the opportunity to analyze the conversion of tachyzoite to bradyzoite stage of the RH strain, followed by stage-specific gene-expression analyzing. Two groups of Swiss-Webster and BALB/c mice were infected subcutaneously with 104 tachyzoites of T. gondii, RH strain and given SDZ (300 mg/l) with NaHCO3 (5 g-1) in drinking water from day1 to day 14 post infection (p.i). The infected mice were sacrificed on day 50 post infection. Their brains were removed and the numbers of tissue cysts were microscopically counted. Total RNA was extracted from brains and cDNA synthesis was carried out. Finally, RT-PCR (Reverse transcrip­tion PCR) was used to detect the expression of bradyzoite (BAG1) and tachyzoite (SAG1) specific genes during tachyzoite / bradyzoite stage conversion. Sixty five percent of all infected mice were survived. Cysts were detectable in mice brain (45%) on day 50 p.i. Also RT-PCR of the brain samples was positive for SAG1 and BAG1. It seems that conversion of tachyzoites to bradyzoites in brain of mice undergoing SDZ was not completed until 50 days after inoculation.

Hydatidosis or cystic hydatid disease is one of the most important diseases in human and animals. Identification of strains is important for improvement of control and prevention of dis­ease. The aim of this study was to determine the strains isolated from human and domestic animals in Ilam Province, Iran, using PCR-RFLP method. Respectively, 30 and 4 animal and human hydatid cysts were collected from different slaughter­houses and hospitals of the province. Protoscolices were separated and their DNA genome was extracted by extraction kit. rDNA-ITS1 of each isolated samples was duplicated by BD1(Forward) and 4s (Reverse) Primers. PCR products were studied by electrophoresis and then were digested using TaqI, HpaII, RsaI and AluI restriction enzymes. RFLP products were studied using electrophoresis on 1% agar gel. A fragment of 1000bp was produced from amplification of rDNA-ITS1 of protoscolices using PCR method. After digestion of PCR product by AluI enzyme, 200bp and 800bp, by RsaI, 655bp and 345bp and by HpaII 700bp and 300bp sizes were obtained. TaqI enzyme had no change in fragment size and it remained 1000bp. Considering the method, Ilam strains was specified as E. granulosus sensu stricto (G1-G3). Although sheep strain (G1) is dominated in human and different animal in Iran and the world, but more efforts should be done to clarify the true genotype of Ilam strains specified as E. granulosus sensu stricto (G1-G3).

The sole published data on feline heartworm infection in the Philippines was reported four decades ago. The study therefore endeavoured to assess and provide an update on the current status of heartworm infection in domesticated feline species using serologic and parasitological examina­tion methods. A total of 46 males and 54 females cats showing clinical signs of dirofilariosis from Makati City, Philippines were subjected to two antigen-based test kits and a microfilaria concentration method. The most commonly observed clinical sign was coughing while exercise intolerance was seldom seen. Age groups ranging from 1 to 4 years old exhibited majority of the clinical signs whereas the 8.1 to 12 years category had the least. The results from the different detection methods employed revealed that none of the animals were positive for circulating microfilaria and no detect­able levels of heartworm antigens were obtained. The presence of associated clinical signs is not an outright indicator of feline dirofilariosis and may be indicative of the rarity of heartworm infection in cats in Makati, Philippines.

Trichomonas vaginalis is a pathogenic protozoon and may be contaminated with dsRNA virus called Trichomonas vaginalis virus (TVV). The viral infection is an important factor for its pathogene­sis and sensitivity to metronidazole. The presence of TVV is associated with qualitative and quantitative expression of cysteine proteinases and surface immunogenic; P270. The purpose of this study was to determine TVV frequency in T. vaginalis clinical isolates in Tehran, Iran. The 46 T. vaginalis isolates were collected from Tehran Province and cultured in TYI-S-33 culture medium. Viral RNA was extracted and RT-PCR was done. Of 46 T. vaginalis isolates, 8 isolates (17.39%) were infected with TVV-1. There was not any association between patient age and TVV- infected T. vaginalis. There were 17.39% viral infection in T. vaginalis isolates which was lower than that reported by other researchers. This is the first report on T. vaginalis isolates infection by TVV-1 in Iran.

The aim of this study was to evaluate the possible use of Biomphalaria alexandrina snail antigens in diagnosis of schistosomiasis mansoni using enzyme linked immunolectrotransfere blot (EITB). S. mansoni adult worm crude antigens (AWA), feet and visceral humps of B. alexandrina and Bulinus truncatus were used. Hyperimmune mice sera (HIS) versus each antigen were prepared for diagno­sis of S. mansoni using western blot (WB). Snail foot antigens were more specific in antibodies detection than visceral hump antigens. Three of five polypeptides of B. alexandrina foot antigen identified by S. mansoni HIS showed specific positive reactivity. These polypeptides were at MW of 31/32 and 43 kDa. While, only one of the six polypeptides of B. alexandrina hepatopancrease antigen identified by S. mansoni HIS, at a MW of 43 kDa was specific. Similarly, 2 polypeptides at MW of 44 and 55 kDa were specific in detection of anti- S. haematobium antibodies. However, the antigenically active polypeptide of B. truncatus hepatopan­crease antigen had no specific reactivity towards anti-S. haematobium antibodies. B. alexandrina foot antigens were the most specific of the tested snail antigens in diagno­sis of schistosomiasis mansoni.

Toxoplasmosis is one of the most common parasitic infections of humans and other mammals. This study was aimed to understand the mechanism of action of veterinary medicine-sulfachlo­ropyrazine (SPZ, 99.97%) against Toxoplasma gondii. T. gondii tachyzoites were soaked in PBS (as a control) or SPZ (250 mg/mL) for 2 h at 37 °C. After being processed, any ultrastructural changes of the tachyzoites that had occurred were observed by Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM). The tachyzoites from control groups with a uniform size had a smooth surface and intact cell or nuclear membranes. In addition, an oval-shaped nucleus, conoids and micronemes were also observed. By contrast, many parasites from the SPZ-treated groups were detrimentally affected by the treatment. Some appeared to be of the vacuolization in their cytoplasm, with the substantial reduc­tion in the number of dense granules and the blur of some organelles. The morphology and ultrastructure of tachyzoites can be affected significantly by SPZ, which might kill the parasite by inhibiting its energy metabolism, inducing apoptosis and damaging its structure. The study provides an experimental basis for further study on the mechanism of SPZ against T. gondii.

The aim of the study was to determine the helminthic species occurring in great gerbil Rhombomys opimus collected from Maraveh Tappeh, Golestan Province, northeast Iran. During 2010-2011, a total of 77 R. opimus were captured from rural areas of Maraveh Tap­peh, Golestan Province, using Sherman live traps and examined for infectivity with any larva or adult stages of helminthic parasites. Overall, 63 R. opimus (81.8%) were found infected with different helminthic species. The rate of infectivity with each species was as follows: Trichuris rhombomidis 31.2%, Trichuris muris 32.5%, Trichuris spp. 10.4%, Syphacia muris 2.6%, Dipetalonema viteae (Acanthocheilonema viteae) 37.7%, Skrjabinotae­nia lobata 15.6%, Hymenolepis (=Rodentolepis) nana fraterna 5.2%, and Taenia endothoracicus larva 1.3%. R. opimus is host for several species of cestodes and nematodes in the study area. The high rate of infectivity with D. viteae indicates the susceptibility of these gerbils to this filarial nema­tode. Synchronous infections occurred up to four species of helminthes in one host.

The goal of this study was to know the identity of Leishmania species responsible of cutaneous leishmaniasis (CL) in Fars Province, southern Iran. Five counties of Shiraz, Firouz Abad, Ghir-Karzin, Farashband and Larestan were pros­pected. Forty-four patients exhibiting cutaneous lesions were selected. Samples collected on skin lesions were examined both microscopically (after Giemsa staining) and molecularly (after PCR-RFLP). On the 44 examined patients, 39 exhibit Leishmania sp. by microscopical examination, all confirmed by PCR. For five patients with negative microscopical examination, PCR was positive for three of them. Among these 42 positive samples, 3 (7%) were infected by L. tropica and 39 (93%) by L. major. Leishmania major is the most prevalent species in prospected area and L. tropica occurs in Shiraz and Ghir-Karzin counties.

The aim of this study was to evaluate the functional capacity of the liver based on the activity of specific enzymes and bilirubin in serum and also to investigate the influence of mechanical and toxic effects of Fasciola hepatica on the structures of the blood vessels and biliary tract in the sheep liver. Blood samples and liver of 63 indigenous sheep of Pramenka breed, slaughtered in the period from March to December 2009 were used. Based on parasitological findings in the liver, all animals were divided into two groups: control (n=34) and infected group (n=29). For investigation and description of pathological changes in sheep liver, naturally infected with F. hepatica, corrosion cast technique was used. Biochemical analysis of tested parameters showed a significant elevation (P≤0.05) of serum gamma-glutamyl transferase (GGT), total bilirubin (TBIL) and direct bilirubin (DBIL) in infected sheep group comparing with the control group. No significant differences were observed for activity of aspartate aminotranferase (AST) between groups. Vascular and biliary systems of the liver were found to be affected. Results of biochemical analysis are consistent with pathological findings and measuring of tested parameters could be used in early diagnosis of sheep fasciolosis and to test the effectiveness of anthelmintic therapy. Corrosion cast technique is very useful for investigation of pathological changes and neoangiogenesis of vascular and biliary system in sheep liver, caused by mechanical and toxic effects of F. hepatica.

Caprine besnoitiosis is an economically important disease of goats. Neospora caninum, another coccidian parasite of worldwide distribution, infects several animal species and is a major cause of abortion in cattle. Combined infections of N. caninum and Besnoitia caprae can occur in geographi­cal areas endemic for both species of parasite in goats. This experiment was conducted to investigate the possible cross-immunity between these two infections in experimentally infected BALB/c mice. Forty BALB/c mice were divided into four equal groups. The mice of Groups 1 and 4 were inoculated with 1×106 live virulent tachyzoites of N. caninum (NC-1), while animals of Groups 2 and 3 were inoculated with sterile tissue culture medium. Each mouse in Groups 1 and 2 was chal­lenged 28 days later with 1×106 live virulent bradyzoites of B. Caprae (BC-1). Following the challenge, the mice in Groups 1 and 2 showed 100% morbidity and 100% mortality within 9 days post infection, while all the animals of Groups 3 and 4 remained alive. The dead animals were necropsied. The survivors (mice in Group 3 and 4) were euthanized 9 days after inoculation and the gross and histopathological lesions in different organs were investigated. Immunization and challenge experiments with lethal dose of B. caprae in the highly suscepti­ble BALB/c mice showed no cross-protection between N. caninum and B. caprae.

Despite Echinococcus granulosus, there are merely two old reports of E. multilocularis infection among Iranian canids of Moghan Plain, the only area known endemic for the species. We detected specific DNA markers in fecal samples by PCR (Copro-PCR) for differential diagnosis of Echinococcus species in living canids. Totally 144 fecal samples from domestic dogs, red foxes and a golden jackal were examined for genus-specific Echinococcus coproantigens using ELISA. Forty two positive or ambiguous samples were further examined for Echinococcus species-specific DNA markers by two different set of nested-PCR. Twenty five out of 144 (17.4%) animals were contaminated with E. granulosus including 14 (23.7%) domestic dogs, 10 (11.9%) red foxes and one (100%) golden jackal. But none of them harboured E. multilocularis species-specific Copro-DNA. The overall prevalence of E. granulosus and E. multilocularis infections in canids of the area was estimated to be 17.4% and 0.0%, respectively. There was a significant relation between the results of Copro-PCR and CA-ELISA. The lack of E. multilocularis infection, compared to previous reports may be due to the differences in used diagnostic methods and/or recently limited territories of wild canids and altered their food resources in this particular area.

Neosporosis is caused by an obligate intracellular parasitic protozoa Neospora caninum which infect variety of hosts. NcSRS2 is an immuno-dominant antigen of N. caninum which is consi­dered as one of the most promising targets for a recombinant or DNA vaccine against neosporosis. As no study has been carried out to identify the molecular structure of N. caninum in Iran, as first step, we prepared a scheme to identify this gene in this parasite in Iran. Tachyzoite total RNA was extracted and cDNA was synthesized and NcSRS2 gene was amplified using cDNA as template. Then the PCR product was cloned into pTZ57R/T vector and transformed into E. coli (DH5α strain). Finally, the recombinant plasmid was extracted from trans­formed E. coli and sequenced. Bioinformatics analysis also carried out. The PCR product of NcSRS2 gene was sequenced and recorded in GenBank. The deduced amino acid sequence of NcSRS2 in current study was compared with other N. caninum NcSRS2 and showed some identities and differences. NcSRS2 gene of N. caninum successfully cloned in pTZ57R/T. Recombinant plasmid was confirmed by sequencing, colony PCR and enzymatic digestion. It is ready to express recombi­nant protein for further studies.

The leishmanicidal and cytotoxic activity of ten essential oils obtained from ten plant specimens were evaluated. Essential oils were obtained by the steam distillation of plant leaves without any prior processing. Cytotoxicity was tested on J774 macrophages and leishmanicidal activity was assessed against four species of Leishmania associated with cutaneous leishmaniasis. Seven essential oils exhibited activity against Leishmania parasites, five of which were toxic against J774 macrophages. Selectivity indices of >6 and 13 were calculated for the essential oils of Ocimum basilicum and Origanum vulgare, respectively. The essential oil of Ocimum basilicum was active against promastigotes of Leishmania and innocuous to J774 macrophages at concentrations up to 1600 µg/mL and should be further investi­gated for leishmanicidal activity in others in vitro and in vivo experimental models.

Like several other parasites, Teladorsagia circumcincta secretes or excretes urea, but nei­ther the rate of efflux nor the possible metabolic sources of the urea has been considered. Parasites were maintained by passage through sheep. Urea efflux was measured using phe­nol/hypochlorite after treatment with urea aminohydrolase. The kinetics of creatine amidinohy­drolase and arginine amidinohydrolase were characterised by coupling the reactions with urea aminohy­drolase and glutamate dehydrogenase. Infective L3 T. circumcincta secreted or excreted urea at 25% of the rate of NH3/NH­4+. The rate of urea efflux was about 84 pmol h‑1 (103 larvae)‑1 over 4 hours, corresponding to about 11 nmol h-1 mg-1 protein. We could not detect urea aminohydrolase activity, but urea production by both creatine amidinohydrolase and arginine amidinohydrolase could be detected. The apparent Km and Vmax of creatine amidinohydrolase were 1.1 mM and 48 nmol h-1 mg-1 protein, respectively, and the activity was greatest at pH 8. The apparent Km and Vmax of arginine amidinohydrolase were 0.7 mM and 62 nmol h-1 mg-1 protein, respectively, and the activity was greatest at pH 7.9. The activity of creatine amidinohydrolase and arginine amidinohydrolase was sufficient to account for the rate of urea secretion or excretion.

This study was carried out in Opi-Agu a tropical semi-urban autonomous community comprising of three villages in Enugu State, Nigeria, between the months of April and June 2010. It was designed to determine the prevalence of Onchocerca volvulus infection and assess the perception of the disease among the inhabitants of this community. A total number of 305 individuals comprising of 148 males and 157 females were ex­amined for various manifestations of onchocerciasis symptoms using rapid epidemiological assess­ment (REA) method. Out of this number, 119 (39.02%) individuals were infected. Prevalence of infection among age groups and villages varied. Age group 41 yr and above had the highest (31.00%) prevalence, while among the villages, Ogbozalla village ranked higher (45.71%) than the other villages. Overall the prevalence of infection among the sexes revealed that males were more infected (43.24%) than the females (35.03%). Lichenified onchodermatitis (LOD) was the most prevalent (35.29%) onchocercia­sis symptom among others identified in the area, while leopard skin (LS) had the lowest (20.17%) occurrence and blindness (0.00%) which is the most devastating effect of O. volvulus infec­tion was not observed. Questionnaire responses from 410 individuals revealed that 34.8% respon­dent from Idi village and 28.1% from Ibeku village believed that O. volvulus infection occurs through poor personal hygiene. Bite of blackfly ranked least (10.6%) among the respondent’s knowledge of the causes of onchocerciasis in Opi-Agu community. Opi-Agu community members had poor knowledge of onchocerciasis, the vector and of its etiologic organism. There is need for integration of community health education with mass chemo­therapy.

The aim of present study was to determine the seroprevalence of canine visceral leishmania­sis (CVL) among stray and owned dogs in Kouhsar district of Alborz Province, central Iran. The study was performed from March 2011 to July 2011 using Direct Agglutination Test (DAT). Three hundred and thirty seven dogs including 257 stary and 80 owned dogs were selected by random sampling. The agreement between serological data and sex, age, life style of dogs and clini­cal signs were assessed by Chi-square. DAT showed that from 337 serum samples collected from owned and stray dogs, 12sera (3.6%) were positive. The seroprevalance was 10% (8/80) among owned dogs and 1.6% (4/257) among stray dogs. A significant difference in seroplevalance was seen between owned and stray dogs (P = 0.01). The highest seroprevalence rate (14%) was observed among the ownership dogs of 5 years old and above. Statistical analysis revealed significant relation between seroprelvalence and age (P= 0.02). There was no statistically significant relation between male (6.3%) and female (2.2%) seropreva­lence (P= 0.085). This survey indicates the importance and necessity of serologic screening of visceral leishma­niasis in human and dogs in Kouhsar district.

Cytoadherence of Plasmodium falciparum- infected erythrocytes to host cells is an impor­tant trait for parasite survival and has a major role in pathology of malaria disease. Infections with P. falciparum usually consist of several subpopulations of parasites with different adhesive proper­ties. This study aimed to compare relative sizes of various binding subpopulations of different P. falciparum isolates. It also investigated the adhesive phenotype of a laboratory P. falciparum line, A4, using different binding techniques. Seven different P. falciparum isolates (ITG, A4, 3D7 and four field isolates) were cultivated to late trophozoite and schizont and then cytoadherence to cell differentiation 36 (CD36), intercellu­lar cell adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule (V-CAM) and E-selectin were examined. The relative binding sizes of parasite subpopulations to human receptors were meas­ured by mini-column cytoadherence method. The adhesion phenotype of P. falciparum-A4 line was evaluated by in vitro static, flow-based and mini-column binding assays. The relative binding size of ITG, A4 and 3D7 clones to a column made with CHO/ICAM-1 was 68%, 54% and 0%, respectively. The relative binding sizes of these lines to CHO/CD36 were 59.7%, 28.7% and 0%, respectively. Different field isolates had variable sizes of respective CD36 and ICAM1-binding subpopulations. A4 line had five different subpopulations each with different binding sizes. This study provided further evidence that P. falciparum isolates have different binding subpopulations sizes in an infection. Furthermore, measurement of ICAM-1 or CD36 binding subpopula­tions may practical to study the cytoadherence phenotypes of P. falciparum field isolates at the molecular level.

The aim of this study was to compare superoxide dismutase (SOD) activity in Fasciola hepatica and Fasciola gigantica parasites. Materials: F. gigantica and F. hepatica helminths were collected from abattoir and cultured in buffer media for 4 h at 37 °C. Excretory-Secretory (ES) products were collected, centrifuged and stored at -20◦C. E-S protein concentration was measured by Bradford method and SOD activity was detected using RANSOD kit (Randox Lab. Crumlin, UK). Statistical t-test was conducted for analysis of results. Protein concentration for F. hepatica and F. gigantica were obtained 7.293 ug/ml and 19.65 ug/ml respectively and SOD activity as 0.721 U/ml and 1.189 U/ml, in that order. ES protein concentration of two species was significantly different (P<0.05), however the difference of SOD activity of two species was not significant. Two species of Fasciola have comparable SOD biochemical defense enzyme and can help us explain the parasite survival in host tissue.

Leucocytozoonosis is a disease of birds caused by obligate intracellular protozoa of the genus Leucocytozoon. We determined the prevalence of Leucocytozoon spp. using light and transmission electron microscopy in domestic birds in southwest of Iran. A total of 825 blood smears from 275 birds were examined for presence of infection. The structure morphology of Leucocytozoon spp. was studied using light and electron microscopy. Forty-four (16.0%) of the birds were positives for Leucocytozoon. The detected parasite were found in 14 chickens (5.1%), 12 geese (4.3 %), 10 ducks (3.6%), and 8 turkeys (2.9%). The majority of the records were from the northeastern regions. Leucocytozoonosis are distributed in the Lorestan province bird population and electron microscopy can resolve the problem to distinguish between similar species of Leucocytozoon.

Diagnosis of some diseases is difficult due to invasive sampling. Urine has been candi­date as a non-invasive and convenient alternative. It has many advantages and easy accessibility but some technical ills should be removed. Finding a suitable extraction method for improving urine DNA quantity and quality in altering invasive specimens for molecular diagnosis of some infectious diseases, was the main object of present research. Toxoplasmosis was selected as an experimental model, regarding the congenital and ocular forms, its abundance and requirement to invasive sample for diagnosis. Samples prepared by adding some defined Toxoplasma gondii (RH strain) tachyzoites to normal urine. Several urine DNA extrac­tion and purification methods comparatively were tested for finding the best one. The amount of extracted DNA assessed using Nanodrope spectrophotometer and a multiplex semi-nested PCR were designed for evaluating the results. Urine samples with known number of tachyzoites were purified comparatively by five bet­ter methods. The results reviled that Cinnagen kit performed with more efficacies. It works well up to 1-5tachyzoites µl-1 of urine. The amount and quality of extracted DNA of more than 100 urine samples with defined tachyzoites were analyzed using a nested PCR method. Finally methods were enough sensitive to detect one tachyzoite DNA in final PCR reaction. This method was enough eligible and sensitive to perform molecular tests for different purposes of instance detecting toxoplasmosis by urine sam­ple as a convenience and non invasive method; although it is better to perform some more experi­ments using patients samples comparing gold methods.

Visceral Leishmaniasis or Kala Azar is endemic in certain regions of India. In endemic areas, the constella­tion of fever, progressive weight loss, weakness, pronounced splenomegaly, anemia, leukope­nia, and hypergammaglobulinemia is highly suggestive of visceral leishmaniasis. Demonstra­tion of the parasite in liver, splenic or bone marrow aspirates is confirmatory. We present a case in which Leishmania donovani (LD) bodies were demonstrated on splenic aspirate. We were unable to demon­strate LD bodies on bone marrow aspiration.