Comparison of agar dilution, broth dilution, cylinder plate and disk diffusion methods for evaluation of anti-leishmanial drugs on Leishmania promastigotes

Leishmaniases are a group of diseases caused by Leishmania parasites. Growing of drug unresponsiveness in leishmaniasis patients necessitates the development of new drugs and accordingly a suitable assay is needed for evaluation of any modalities. The aim of this study was to compare four drug assays methods, agar dilution, broth dilution, cylinder plate and disk diffusion, for evaluation of anti-leishmanial drugs on Leishmania promastigotes, using glucantime as a currently available drug for treatment of leishmaniasis. For broth dilution method, different concentration of glucantime was added to the parasite culture [promastigotes of Leishmania], while in cylinder plate method wells were punched in agar gel and filled with different concentration of drug and zone of inhibition was measured in each well. In disk diffusion method, the parasites were cultivated on the surface of agar; filter paper disks were enriched with various concentration of glucantime and were placed on the surface of agar. In agar dilution method, various concentrations of drug were incorporated onto blood agar and the parasites were cultivated on the surface of the agar. A direct correlation was found between the drug concentration and size of inhibitory zones in cylinder plate and disk diffusion methods. These two drug assays methods provided much better performance in comparison with broth and agar dilution methods. Cylinder plate and disk diffusion methods seem to be acceptable methods for susceptibility testing of anti-leishmanial compounds on Leishmania promastigotes

Performance of antigen B isolated from different hosts and cyst locations in diagnosis of cystic echinococcosis

The aim of this study was to assess the performance of Antigen B [AgB] isolated from different Echinococcus granulosus intermediate hosts and from different cyst locations for immunodiagnosis of human cystic echinococcosis [CE]. Hydatid cyst fluids were collected from lung and liver cysts of sheep, liver cysts of goats, lung cysts of camels and cattle, and liver cysts of human. AgB was purified from each of these hydatid cysts fluids. Serum samples obtained from 47 pathologically confirmed cases of CE along with 30 sera samples from non-CE patients and 40 sera from healthy individuals were tested by ELISA using AgB prepared from different hosts or cyst locations. The highest sensitivity [97.8%] for diagnosis of CE was seen with AgB prepared from human liver cysts. This maximal sensitivity was followed by AgB isolated from those of sheep liver and lung cysts. The least sensitivity was found with AgB prepared from bovine lung cysts. The highest specificities [97.1%] were observed with AgB isolated from human liver cysts followed by those of sheep and goat liver cysts while the lowest specificity was seen with AgB isolated from bovine lung cysts. In view of the specificities and sensitivities of the different AgB, the best validity was found for AgB prepared from human liver cysts while the least validity was found with AgB prepared from bovine lung cysts. For any AgB-based tests, obtaining of the antigen from one of these sources will significantly increase the diagnostic sensitivity and specificity of the assay

Establishment of a modified in vitro cultivation of protoscoleces to adult echinococcus granulosus; an important way for new investigations on hydatidosis

Echinococcus granulosus, a zoonotic cestode parasite, causative agent of hydatid cyst is endemic in many parts of the world including the Middle East. Study on different aspects of this parasite is very important and valuable. However, working with adult worms which their habitat situated in the small intestine of canids, is dangerous and risky. Achieving such risky situation needs a controlled condition which is cultivation of the organisms in the laboratory. In this regard, cultivation of E. granulosus protoscoleces leading to adult worms was established in the laboratory for the first time in Iran. Under aseptic conditions a number of protoscoleces were cultivated in diphasic S.10E.H medium using CO2 incubator to produce adult worms. Different forms of parasites including pre-segmentation stages [PS1 - PS4] and segmentation stages [S5-S8] and developing stages in segmented worms [S10-S11] were observed and evaluated in these medium. Finally adult worms contained four proglottids with a large and distinct genital pore were observed 50-55 days post cultivation. These parasites do not produce fertile eggs and conclusively do not have risk of hydatid disease transmission to the researchers. The mentioned method for producing E. granulosus adult worms can open a new window for researches and facilitate working on different aspects of hydatidosis especially for diagnosis, protection and treatment studies

Helminthic infections of laboratory animals in animal house of Shiraz University of medical sciences and the potential risks of zoonotic infections for researchers

Different parasitic diseases may be transfered from laboratory animals to human [zoonoses]. The current study was designed to determine the helminthic infections in animal house of Shiraz University of Medical Sciences to prevent the possible transmission of zoonotic ones from laboratory animals to the staff and researchers. Sixty laboratory animals including mouse, rats, Guinea pigs and rabbits were randomly selected and examined for any helminthic infections. 83.33% rats were infected with Syphacia muris and Aspiculuris tetraptera, 100% of Guinea pigs were infected with Paraspidodera uncinata, rabbits were infected with Passalurus ambiguous [40%], inbred BALB/C mice were infected with Hymenolepis nana [50%], Aspiculuris tetraptera [90%] and Syphacia obvelata [90%], outbred BALB/C mice were infected with Hymenolepis nana [50%], Aspiculuris tetraptera [90%], Syphacia obvelata [90%] and C57BL/6 mice were infected with Hymenolepis nana [66%], Aspiculuris tetraptera [100%] and Syphacia obvelata [100%]. Our study was revealed minimum and maximum infection frequency in rabbits and guinea pigs respectively. It seems that low and unsuitable space of infected animals in mentioned research center was the main cause of distribution of infection among rats and mice in Animal House of Shiraz University of Medical Sciences

case of gingival myiasis caused by Wohlfahrtia magnified

A gingival myiasis in a four years old mental retarded boy with anorexia and weight loss is presented from southern part of Iran. Entomological studies on larvae showed the larvae as Wohlfartia magnifica which is a rare causative agent of gingival myiasis

Study on the genomic diversity of hymenolepis nana between rat and mouse isolates by RAPD-PCR

Hymenolepis nana is a common parasite of rodents as well as human intestine. This parasite has been reported from all over the world, including Iran. The infection rate has been reported up to 40% in some areas. The infection has various clinical manifestations. The parasite could establish severe hyperinfection in patients with immune deficiency. Regarding the rodents as hosts of the parasite, the infection may disseminate through these hosts to the nature. As H. nana is a zoonoses, phylogenic study of this parasite is of particular importance. Considering these criteria, the genomic diversity of 16 H. nana with the origin of Shiraz and Tehran were studied among the worms of mice and rats by RAPD-PCR. Genomic DNA extracted from individual worms by proteinase K method and three oligonucleotides primer [ABl-17, UBC-358, UBC-387] were used for RAPD-PCR. Similarity index were calculated by Nei and Li method. Data were analysed using UPGMA analysis and dendrograms were obtained by group average method with 100 bootstrapping analysis. The range of genomic similarity determined among specimens by ABl-17 primer was 48.3-90%, by UBC-358 primer 55-87% and by UBC-387 primer 53-97%. Regarding our data and genomic similarity indexes, various isolates were found in both specimens of rats and mice. However no differences were obtained between H. nana from rat or mouse isolates by these primers. The results showed that it is not possible to divide the isolates into two distinct groups based on their origin as Tehran and Shiraz

Application of western blotting using native antigen B for serodiagnosis of human cystic echinococcosis

Hydatidosis is one of the most important helminthic diseases causing serious health and economic problems in many countries including Iran. Currently available diagnostic approaches for diagnosis of cystic echinococcosis [CE] are not satisfactory since they have not achieved a reasonable validity in diagnosis of CE. This study aimed to assess the performances of Western blotting [WB], using native antigen B, for serological diagnosis of human hydatidosis. Antigen B was prepared from sheep hydatid cyst fluid. Serum samples obtained from 40 surgically proven cases of hydatidosis along with serum samples from non-hydatidosis patients and samples from healthy persons were tested by immunoblotting, using native antigen B. From 40 sera of hydatidosis patients, 32 cases [80%] detected 8 kDa subunit, 29 cases [72.5%] recognized the 16 kDa component and 29 cases [72.5%] detected 24 kDa subunit of antigen B. The highest sensitivity [80%] was achieved with 8 kDa subunit of antigen B while the other components [16 and 24 kDa] showed a lower sensitivity [72.5%]. Samples from healthy controls and non-hydatidosis patients did not yield any detectable band in the Western blotting assay. Accordingly, specificity of the system was found to be 100%. Considering the use of native antigen B, application of western blotting, based on 8 kDa subunit of antigen B, is highly recommended for the confirmatory diagnosis of hydatidosis

Direct agglutination test and enzyme linked immunosorbent assay with urine samples for the diagnosis of visceral leishmaniasis

Visceral leishmaniasis [VL] or Kala azar is an infectious disease caused by various species of Leishmania parasites. The aim of this study was to detect and compare the presence of anti-Leishmania antibodies in the urine of visceral leishmaniasis patients using ELISA and DAT methods. A total of 30 urine samples were collected from VL patients referred to Shiraz [southeast of Iran] hospitals. Moreover 31 urine samples were collected from healthy individuals and patients with other diseases such as malaria, brucellosis, hydatidosis and cutaneous leishmaniasis. Collected samples were examined to detect anti-Leishmania antibodies in urine, using ELISA and DAT. Anti-Leishmania antibody was detected in urine of 18 out of 30 [60%] VL patients by DAT while ELISA detected anti-Leishmania antibodies in urine of 28 out of 30 [93.3%] of VL cases. Sensitivity and specificity of urine-based DAT was 60% and 83.9%, respectively while sensitivity and specificity of urine-based ELISA were 93.3% and 93.5%, correspondingly. Urine-based DAT and ELISA have a reasonable specificity and sensitivity in diagnosis of VL. Accordingly, urine-based ELISA might be a suitable alternative for serum based assays for diagnosis of VL

Nutritional status of preschool children infected with giardia intestinalis

Giardia intestinalis is the most common intestinal parasite in human worldwide. It can produce a wide spectrum of clinical manifestations. In order to assess the nutritional status of preschool children infected with Giardia intestinalis, a cross sectional study was made in Marvdasht city, Fars Province, Southern Iran. A total of 337 preschool boys and girls aged 3-6 years were randomly selected for stool examination of intestinal parasites as well as measurement of height, weight, head and arm circumferences. A total of 77 individuals were infected with G. intestinalis. Seventy-one individuals who had only G. intestinalis and 229 with no parasitic infections were selected as infected and control groups, respectively. Z-Score of -2SD was used as cut off point of malnutrition. A total of 9 [12.7%] of infected children and 18 [7.9%] of non infected individuals had a height for age Z-score [HAZ] below -2SD. Eight [11.3%] of former group had a weight for age Z-score [WAZ] under-2SD. In control group 4.4% of preschool children had WAZ under-2SD. 4.2% of infected children had a weight for height Z-score [WHZ] under-2SD but none of the controls had it. Statistical analysis showed a significant difference in height, weight, head circumference, HAZ, and WAZ between infected and control children [P<0.05]. Also, HAZ and WAZ, was significantly different between these two groups, but not for WHZ. A higher infection with G. intestinalis in the children with lower parents' education was observed. However the distribution of malnutrition was not significantly different between boys and girls. In conclusion the present study indicated that giardiasis retarded growth of preschool children in this region

Diagnosis of cystic echinococcosis: ultrasound imaging or countercurrent immunoelectrophoresis?

Cystic echinococcosis is a major zoonotic diseases in the Islamic Republic of Iran. This study was carried out in 3 general hospitals in Shiraz. We examined the records of all 1227 surgical patients with a surgically-proven diagnosis of cystic echinococcosis for the 20-year period 1978-98. The results of countercurrent immunoelectrophoresis were compared with pathology and ultrasound reports to determine whether serological tests could be helpful for diagnosis. Countercurrent immunoelectrophoresis could detect only 62.0% of cases, whereas the pathology and ultrasound results were positive for 96.3% of cases. This study confirms the usefulness of ultrasound and suggests that only in doubtful cases would countercurrent immunoelectrophoresis be useful for diagnosing cystic echinococcosis

case report of Linguatula serrata in human pharynx from shiraz, southern Iran

A case of linguatulosis due to Linguatula serrata, a zoonotic pentastomid parasite in the pharynx of a 35 year old woman from Shiraz is described. After removal of the parasites the patient was discharged without any further complaints

Study on the latent iron deficiency in patients with trichuriasis

Trichuriasis is an intestinal parasitic disease which is caused by Trichuris trichiura, a soil transmitted helminth. In order to evaluate the iron status of trichuriasis in patients a total of 120 [49 males and 71 females] cases with Trichuris trichiura as well as 54 healthy individuals were evaluated. Blood samples were analyzed for serum iron, transferrin saturation%, haemoglobin and other haematological indices. Anaemia based on haemoglobin level less than 13 mg/dI in males and less than 12 mg/dI in females were found in 10 [7 males and 3 females] patients. Of remaining 110 non anaemic patients 11% [14.3% of males and 8.8% in females] would be classified as iron deficient by serum iron and 6% by transferrin saturation%. These findings suggest that serum iron and% transferrin saturation are valid and sensitive tests for detecting iron deficiency in these patients; and T.trichiura infection is associated with high incidence of iron deficiency