In Vitro Study on Protoscolicidal Effect of Methanolic Extract of Allium hirtifolium on Protoscoleces of Cystic Echinococcosis.

BACKGROUND: Cystic echinococcosis/ hydatidosis is one of the most important parasitic zoonoses worldwide. Usually, surgery is used for the treatment of active cysts. Due to the risk of cyst rupture during surgery, a protoscolicidal agent is usually injected into the cyst before surgery to prevent secondary cyst formation in case of cyst rupture. Due to the side effects of chemical protoscolicidal agents attempts have been focused on herbal medicine. In this regard, Allium sativum has already been used; however, the protoscolicidal effect of Allium hirtifolium from Allium family has not been investigated, so far. OBJECTIVE: The present study was conducted in order to assess the protoscolicidal effect of methanolic extract of Allium hirtifolium in different concentrations, temperature and exposure times. Cetrimide as a positive control and DMSO as a negative control were also used in this study. METHODS: The viability of protoscoleces was determined by 0.1% eosin staining method. RESULTS: The protoscolicidal effect of Allium hirtifolium on protoscoleces was as follows: The mean percent and standard deviation (SD) of protoscolicidal activity of 50 mg/ml concentration of Allium hirtifolium extract in room temperature at 10, 20, 30, 40 and 50 minutes of exposure was 5.88±2.39, 8.59±4.54, 11.82±5.81, 12.31±6.16and 16.96±8.42 respectively; while at 37°C it was 9.10±5.01, 11.52±7.36, 18.64±4.36, 27.07±6.57 and 38.15±12.80, respectively. CONCLUSION: In conclusion, although Allium hirtifolium from Allium family is similar to Allium sativum; the protoscolicidal effect of its methanolic extract is not similar and less than A. sativum extract.

Molecular characterization of Toxocara spp. eggs isolated from public parks and playgrounds in Shiraz, Iran.

Human toxocariasis, a worldwide parasitic disease, is caused by the larval stage of intestinal nematodes of dogs and cats, namely Toxocara canis and Toxocara cati. Human infection occurs by the accidental ingestion of embryonated eggs present in the soil, vegetables or on other contaminated surfaces, as well as via consumption of uncooked paratenic hosts, such as bird meat and giblets. The objective of this study was to evaluate the contamination of soil in public parks and playgrounds in Shiraz using microscopy and molecular methods. A total of 150 soil samples were collected from public parks and playgrounds in various areas of Shiraz, southern Iran. The samples were treated with saturated zinc sulphate solution, and Toxocara spp. eggs were detected by microscopic observation followed by nested polymerase chain reaction (PCR). To differentiate T. canis and T. cati eggs from each other, PCR restriction fragment length polymorphism (RFLP) of the internal transcribed spacer (ITS)-rDNA region by SalI endonuclease enzyme was used. PCR-sequencing was performed to confirm the results of the PCR-RFLP method. Based on the flotation results of the 150 soil samples, six (4%) were found to be positive for Toxocara spp. eggs, whereas nested-PCR showed 24 samples to be positive (16%). Based on the PCR-RFLP method and the sequence of the ITS-rDNA region, a total of 23 out of 24 isolates were confirmed as T. cati and one out of 24 as T. canis. The results showed a higher number of soil samples to be positive for Toxocara by the molecular method than microscopy, and higher T. cati infection in soil samples, which could have an important role in human infection with toxocariasis in this region.

The occurrence of Toxocara species in naturally infected broiler chickens revealed by molecular approaches.

Consuming raw and undercooked meat is known to enhance the risk of human toxocariasis because Toxocara species have a wide range of paratenic hosts, including chickens. The aim of this study was to identify species of Toxocara in naturally infected broiler chickens using molecular approaches. A polymerase chain reaction (PCR) method was used for the differentiation of Toxocara canis and Toxocara cati larvae recovered from tissues and organs, and identified by microscopic observations. Thirty-three 35- to 47-day-old broiler chickens were used for examination of Toxocara larvae. The duodenum, liver, lungs, heart, kidneys, skeletal muscles and brain of each chicken were examined using the pepsin method, and DNA from each tissue was extracted as the template for PCR assay. The findings revealed that 5 of 33 (15.2%) broiler chickens were infected with Toxocara larvae. Larvae were recovered from the liver (n = 19), duodenum (n = 8), skeletal muscles (n = 8) and brain (n = 2) of broiler chickens naturally infected with Toxocara spp. The results showed that the frequencies of the species in the chickens were T. canis larvae (n = 5, 83.3%) and T. cati larvae (n = 1, 16.7%). Our data from the present study demonstrated the importance of broiler chickens as a paratenic host for the parasite's life cycle in the environment. The implementation of DNA amplification as a routine diagnostic technique is a specific and alternative method for identification of Toxocara larvae, and allowed the observation of specific species under field conditions within the locations where broiler chickens are typically raised and exposed to Toxocara spp. eggs or larvae.

Trend of toxocariasis in Iran: a review on human and animal dimensions.

One of the neglected soil and/or food-borne diseases with international public health importance is toxocariasis. Human cases are being increasingly reported from Asian, African, Oceania, European and the American countries. Hence, human toxocariasis (HT) is now considered as a major zoonosis with global and regional importance. In Iran, human and animal toxocariasis is an endemic disease with clinical and epidemiologic health problem aspects. Doubtless, understanding the epidemiology and the trend of this important parasitic disease and its affecting factors will provide the establishment of effective prevention and control programs. To better understand the trend of toxocariasis researches in Iran, this study was performed to analyze different aspects of this zoonotic disease including history, life cycle, species, human animals and environmental studies, diagnostic aspects and treatments to find out the gaps, including different aspects of clinical sings in human patients, new and specific recombinant antigens based on the native antigens, new diagnostic tools, especially rapid diagnostic tests, paratenic hosts status and new treatment procedures which is necessary to be investigated in the future studies on this important zoonotic disease.

Evidence that the Echinococcus granulosus G6 genotype has an affinity for the brain in humans.

The present study investigates the molecular characteristics of cerebral Echinococcus cysts. A total of 10 specimens of cerebral Echinococcus cysts, including six formalin-fixed paraffin blocks and four intact cerebral cysts, were used for this study. The target DNA was successfully amplified from eight samples and sequenced. BLAST analysis indicated that sequenced isolates belong to the Echinococcus granulosus (G6) genotype. All of the eight sampled brain cysts belonged to the G6 genotype, while all of the eight liver cysts belonged to G1. This is a strong indication that G6 has a higher affinity for the human brain than G1.

The serological study of cystic echinococcosis and assessment of surgical cases during 5 years (2007-2011) in Khorram Abad, Iran.

BACKGROUND: Echinococcus granulosus is a cestode whose larval stage causes cystic echinococcosis in wild animals, livestock, and human. OBJECTIVE: The aim of this study is to highlight the seroepidemiology of E. granulosus infection and surgical cases in the general population of Khorram Abad district, southwest of Iran. MATERIALS AND METHODS: Anti-E. granulosus antibodies were tested in 617 inhabitants in Khorram Abad by enzyme-linked immunoassay and antigen B. The surgical cases of cystic echinococcosis were investigated in Shohaday-e-Ashayer Hospital of Khorram Abad (as a referral center) from 2007 to 2011. RESULTS: In total, 95 (15.4%) of the 617 participants (mean age 39.6 ± 17.6 years) had anti-E. granulosus antibodies. Prevalence of infection was more in males (60%) than females (40%), and showed statistical significance (P < 0.001). High-titer antibodies were most prevalent among the subjects aged 20-29 years. There was significant association between the presence of Echinococcus antibodies and the sector of residence, education of volunteers, and occupation (P < 0.05). According to hospital records, 58 cystic echinococcosis cases were referred to the hospital during the 5 years. Among the cases, 28 (48.3%) were men and 30 (51.7%) were women. They were between 4 and 74 years of age (36.6 ± 18.9 years). The liver was the organ where most of the hydatid cysts (51.7%) were located, followed by lungs (20.7%). CONCLUSION: This is the first report of the seroprevalence and contributing factors for E. granulosus infection in the general population in Khorram Abad. The findings confirm the importance of diagnosing human cystic echinococcosis in these regions, given the prevalence rates of surgical cases during the last 5 years, and need further evaluation of the risk factors present.

Laboratory detection of strongyloidiasis: IgG-, IgG4 - and IgE-ELISAs and cross-reactivity with lymphatic filariasis.

Enzyme-linked immunosorbent assays (ELISAs) were developed for the detection of IgG, IgG4 and IgE antibodies against Strongyloides stercoralis. A commercial ELISA (IVD Research, USA) was also used, and the sensitivities and specificities of the four assays were determined. Serum samples from 26 patients with S. stercoralis infection and 55 patients with other infections or no infection were analysed. Sensitivities of the IgG4 , IgG, IgE and IgG (IVD) assays were 76.9%, 84.6%, 7.7% and 84.6%, respectively, while the specificities were 92.7%, 81.8%, 100% and 83.6%, respectively. If filariasis samples were excluded, the specificities of the IgG4 -ELISA and both IgG-ELISAs increased to 100% and 98%, respectively. A significant positive correlation was observed between IgG- and IgG4 -ELISAs (r = 0.4828; P = 0.0125). IgG- and IgG- (IVD) ELISAs (r = 0.309) were positively correlated, but was not significant (P = 0.124). Meanwhile there was no correlation between IgG4 - and IgG- (IVD) ELISAs (r = 0.0042; P = 0.8294). Sera from brugian filariasis patients showed weak, positive correlation between the titres of antifilarial IgG4 and the optical densities of anti-Strongyloides IgG4 -ELISA (r = 0.4544, P = 0.0294). In conclusion, the detection of both anti-Strongyloides IgG4 and IgG antibodies could improve the serodiagnosis of human strongyloidiasis. Furthermore, patients from lymphatic filariasis endemic areas who are serologically diagnosed with strongyloidiasis should also be tested for filariasis.

Seroprevalence of toxocariasis in hypereosinophilic individuals in Ahwaz, south-western Iran.

Eosinophilia in human peripheral blood is caused by different agents, including toxocariasis. The present study aimed to evaluate the prevalence of toxocariasis in hypereosinophilic individuals in the city of Ahwaz, located in south-western Iran, using enzyme-linked immunosorbent assay and Western blot techniques. Serum samples were examined from 100 individuals with peripheral blood eosinophilia and also from another 100 individuals without eosinophilia as the control group. In hypereosinophilic individuals seroprevalence antibodies against Toxocara were found in 19 (19%), of whom 12 (63.15%) were female and 7 (36.85%) were male. Positive sera were subsequently confirmed by Western blot. All of the observed bands ranged from 24 to 100 kDa. Antibodies against Toxocara were found in 1% of the control group, but were not confirmed by Western blot. The results showed significant differences between the frequency of infection within age and gender (P < 0.05); the highest prevalence of infection was observed in adults. Differences between the hypereosinophilic and healthy individuals, in terms of Toxocara infection frequency, also proved significant (P < 0.05).The present study thus confirmed the significant prevalence of toxocariasis as a hygienic problem among hypereosinophilic individuals in this area. It is, therefore, necessary to examine these individuals for toxocariasis.

Human cystic echinococcosis in Yasuj District in Southwest of Iran: an epidemiological study of seroprevalence and surgical cases over a ten-year period.

Cystic echinococcosis is a zoonotic infection of humans and domestic animals caused by the larval stages of the cestode Echinococcus granulosus. Cystic echinococcosis is one of the most important zoonotic diseases in Iran, where human cases are frequently reported from different regions of the country. The objective of this study was to determine the sero-epidemiological and surgical cases of human hydatidosis in the Yasuj district, a region in southwest of Iran. Serum samples were taken from 500 individuals attending Yasuj health centers. Anti-hydatid cyst antibody was checked by enzyme-linked immunosorbent assay (ELISA), using antigen B. Moreover, retrospective studies were carried out using local hospital records of CE patients between 1997 and 2006. Results of the sero-epidemiological study showed that 36 out of 500 cases (7.2%) have antibody against hydatid cyst. Of these 36 cases, 49.6% were women and 50.4% were men. The highest rate of infection was recorded in individuals aged between 30 and 39 years. Hospital records showed that during the 10 years, 105 cases of hydatidosis were admitted in Yasuj hospitals. Of all cases, 70 (66.7%) were women and 35 (33.3%) were men. Hepatic cyst was recorded in 81% of the cases where nephrotic cysts were the second most prevalent ones. Recurrence of the disease was noted in 14.3% of all cases. Results of this study demonstrated that hydatidosis is an important endemic disease, with a nearly constant prevalence rate during the last 10 years, in Yasuj district in Iran. Further studies are needed to investigate the reasons for high rate of surgical cases in housewives and striking prevalence of hydatidosis in a specific (30-40 years) age group.

Histopathologic changes and larval recovery of Toxocara cati in experimentally infected chickens.

This study was made to determine the distribution pattern of Toxocara cati larvae in chickens as a paratenic host and its potential zoonotic risk by consuming infected chickens. Two groups of chickens were fed with 1,000 and 3,000 embryonated eggs of T. cati. The chickens were necropsied 3, 7, 14, and 21 days postinfection. The liver, lungs, kidneys, spleen, small intestine, and half of all the striated muscles were digested for larval recovery. Squash method was used for brain. Larvae were recovered from the liver and brain of infected chickens with 1,000 embryonated eggs. Samples of these tissues were prepared for histopathologic studies. Experimental chickens exhibited hemorrhages in the liver, lungs, and kidneys on all days postinfections (dpi). White spots on the liver surfaces that showed necrotic foci, infiltration of eosinophils, and a few lymphocytes around necrotic areas were seen on 14 and 21 dpi. Remains of larvae were present in the liver on 14 dpi. Pathologic findings showed that larvae migrated in different organs of chickens. We suggest that chickens could be paratenic hosts, and human infection with T. cati might occur after consumption of raw or undercooked meat of infected chicken with T. cati.

The occurrence of severe infections with Rhabditis axei in AIDS patients in Iran.

During an investigation on strongyloidiasis in AIDS patients from Iran, direct smears of stools revealed many eggs and larvae of Rhabditis axei, which were subcultured to confirm identification.

Comparative clinical trial of mebendazole and metronidazole in giardiasis of children.

Giardiasis and soil-transmitted helminthosis are prevalent in many parts of the world. Application of a single drug for treatment of both giardiasis and soil-transmitted helminthosis is highly desirable. One hundred children of both sexes, ranging from 7 to 12 years old with Giardia intestinalis cysts or trophozoites in their stool samples, were randomly separated into two groups of 50 individuals each. Each group received either mebendazole tablets 200 mg thrice daily for 5 days or metronidazole tablet 15 mg/kg/day in three divided doses, for 7 days. The results were evaluated by microscopic examination of stools on two successive occasions: 1 week and 2 weeks after treatment. Each occasion consisted of three stool examination on 3 successive days. The cure rate of mebendazole-treated children was 43 out of 50 (86 per cent), and for metronidazole it was 45 out of 50 (90 per cent) with no statistical difference between the two groups. No side-effects were observed in the mebendazole-treated group, whereas nausea, anorexia and metallic taste were observed in 4.9, 6 and 24 per cent of metronidazole-treated children, respectively. In conclusion, the use of mebendazole, which is less toxic with no side-effects, is available and effective on Giardia intestinalis as well as intestinal nematodes, is recommended especially in the areas where giardiasis and intestinal nematodiasis are prevalent and overlap.

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.

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

Seroprevalence of toxocara infection in school children in Shiraz, southern Iran.

The presence of anti-Toxocara antibodies in the sera of school children of Shiraz, southern Iran, was studied by means of an indirect enzyme linked immunosorbent assay with excretory/secretory antigen of infective stage larva. A total of 519 individuals of both sexes aged 6-13 years were analysed. The total prevalence was 25.6 per cent. A higher rate of infection was observed in urban (30.15 per cent) than rural (20.2 per cent) residents. Most potential risk factors were not related to Toxocara prevalence and no differences existed between socioeconomic classes except for parental education. Neither age or sex was found to be significantly associated with positive serology.

Prevalence of Echinococcus granulosus infection in stray dogs and herbivores in Shiraz, Iran.

One hundred and five stray male and female dogs in different age groups were autopsied and their small intestines examined for Echinococcus granulosus. Thirty eight dogs (36.19%) harbored two-to-several thousand E. granulosus in their intestinal content. The prevalence rate was higher in males (44.83%) than in females (25.53%). Livers and lungs of 3245 sheep, 2576 goats, 756 cattle and 25 buffaloes slaughtered for meat purposes at different Shiraz slaughterhouses were examined for hydatidosis: 2.09% of livers and 2.68% of lungs of sheep, 2.17% and 2.36% of goats, 4.49% and 6.48% of cattle and 4% and 8% of buffaloes, respectively, were infected.