Cyclospora cayetanensis: a review, focusing on some of the remaining questions about cyclosporiasis.

Cyclosporiasis is a protracted, relapsing gastroenteritis and has a short recorded history. Cyclospora cayetanensis is an enigmatic parasite since its discovery highlights the need for isolation of cases of infection that could be part of widespread outbreaks. It is associated with diarrhoea among children in developing countries in the Americas, where C. cayetanensis is endemic; traveller's diarrhoea and/or food and waterborne outbreaks sometimes occur in the developed countries. In SubSaharan Africa and Egypt, cyclosporiasis has been reported to occur in both immunocompromised and immunocompetent patients. Zoonotic species of Cyclospora have also been identified worldwide in primates, indicating likely endemicity of this underreported disease. This can be attributed to the lack of awareness in the public and medical profession concerning the disease, which is, therefore, not routinely tested at the health centres. The correlation between the density of water contamination and the prevalence of cyclosporiasis among the individuals of each area is significant. No doubt, water is the main vehicle of transmission in the present community. Soil contact and poultry are significant risk factors. All literature on C. cayetanensis, cyclosporiasis worldwide, and endemic cyclosporiasis was searched from libraries, colleagues and internet. Although cyclosporiasis is considered an enigma worth unravelling, with many aspects of this disease and its transmission having been uncovered only recently, the situation has been rapidly changing since the disease first came to medical attention in the 1970s.

Schistosoma mansoni: effect of dietary zinc supplement on egg granuloma in Swiss mice treated with praziqantel.

Schistosomiasis is one of the most important parasitic diseases in Egypt and chemotherapy is considered the most effective method of control. This study was conducted to assess the effectiveness of zinc administration against Schistosoma mansoni infection by evaluating the activities of arylesterase and paraoxonase (PON1) enzymes, and the degree of liver damage. One hundred and twenty albino mice were divided into two groups; one was an infected control and the other a treated group which was further subdivided into three according to the praziquantel and zinc supplementation given. Blood and liver samples, collected 10 weeks post-infection, were subjected to parasitological, histopathological, and enzyme assays, and immunological studies. The results showed that dietary zinc supplementation led to marked reduction in worm load, and egg deposition in the liver and intestine. Histopathological examination showed marked reduction in the number and diameter of hepatic granulomas in the treated groups. The activity of arylesterase and PON1 enzymes were partially restored in infected animals receiving zinc. IL-10 mRNA expression was higher in the treated groups than in the infection control group. In conclusion, zinc administration could be a promising adjuvant therapy for S. mansoni infection.

Real-time PCR/RFLP assay to detect Giardia intestinalis genotypes in human isolates with diarrhea in Egypt.

Two major genotypic assemblages of Giardia intestinalis infect humans; the nested real-time polymerase chain reaction (PCR) was used for targeting the triose phosphate isomerase (tpi) gene to detect and genotype G. intestinalis in human feces in Egypt. Among 97 fecal samples, 30 (31%) were diagnosed as giardiasis by saline wet mount microscopy after staining with Lugol's iodine. The tpi gene was amplified from 41 (42.3%) fecal samples, of which 11 were microscopy-negative specimens. Of the total samples, 24 (58.5%) contained assemblage A group I, and 7 (17.1%) were assemblage A group II from the group of patients complaining of intermittent diarrhea. Eight (19.5%) samples contained assemblage B from patients with persistent diarrhea. Two (5%) samples had a mixture of assemblage A group II and assemblage B. The technique was able to detect as few as 20 trophozoites per PCR on fecal DNA-isolated, microscopy-negative, and quantitative (q)PCR-positive specimens; there was a higher average cycle threshold value than microscopy-positive and qPCR-positive specimens, suggesting that they represented true, low-burden infections. In conclusion, we could genotype G. intestinalis from fresh stool samples in Egypt; in infections commonly presented with intermittent diarrhea, the most prevalent genotype was assemblage A group I. The most vulnerable age group included 10- to 20-yr-old individuals.

Fasciola gigantica: evaluation of the effect of phenyl vinyl sulfone in vitro.

Phenyl vinyl sulfone is a synthetic inhibitor of cysteine protease and has antihelminthic and antiprotozoal properties. Phenyl vinyl sulfone was assayed in vitro for antifasciola activity against adult Fasciola gigantica worms using a well-established culture medium. Worms were treated with phenyl vinyl sulfone for incubation periods ranging from 0 to 12h and its activity was assessed in terms of viability, motility and death of worms. Phenyl vinyl sulfone exhibited a minimum effective concentration of 50 ppm after 12h. Three hundred parts per million concentrations were most potent causing immediate death of adult flukes in vitro. Histopathological studies showed that there was tegumental flattening, rupture of vesicles, and spine loss. Marked reduction in size and number of ova and sperms in the convoluted tubules of the reproductive organs was observed in comparison to the untreated control group. In conclusion, phenyl vinyl sulfone shows potent activity against F. gigantica in vitro, and the authors recommend carrying out more studies to detect its efficacy in vivo.

Detection and differentiation of Entamoeba histolytica and Entamoeba dispar isolates in clinical samples by PCR.

A total of 140 out of 180 outpatients attended MISR University for Science and Technology Hospital complained of abdominal pain, diarrhoea and/or dysentery. Stool examination showed 47 (33.6%) had Entamoeba sp., 36 (25.7%) had cysts and 11 (7.9%) had trophozoites. Of 40 asymptomatic ones, 4 (10%) had cysts. A total of 51 positive stool samples for Entamoeba sp. (40 cysts & 11 trophozoites) were tested by Ne-sted Polymerase Chain Reaction (N-PCR) and Restriction Enzyme Digestion (RED) to clarify true E. histolytica from E. dispar. The results showed that 9/51 (17.6%) had E. dispar, while 31 (60.8%) had E. histolytica and 11 (21.6%) had dual infection with both E. histolytica and E. dispar. All E. histolytica PCR proved cases were from the symptomatic group, 11 had trophozoites and 34 had cysts. Thus, the result showed the potential use of molecular tools in detection of E. histolytica and E. dispar, and is a promising tool for epidemiology, particularly to differentiate pathogenic and non pathogenic Entamoeba sp.

Touchdown PCR, ELISA and stool examination for early diagnosing of Schistosoma mansoni in mice.

A detection assay for Schistosoma mansoni DNA in mouse serum samples based on touchdown PCR was evaluated. Fifty four mice (7 weeks old) were infected with 60 +/- 10 cercariae of S. mansoni (Egyptian strain) through tail skin. Three days and 1-8 weeks post-infection (PI), six mice were randomly chosen from the infected ones and blood was collected by cardiac puncture. Sera and faeces were collected for 8 weeks PI. Antisoluble egg antigen (SEA) IgG for S. mansoni was detected in infected mice sera of at 3 weeks PI by ELISA. The earliest deposition of eggs in the small intestines was at the 5th week PI. The eggs were detected in faeces at the 8th weeks by modified Kato's thick-smear. However, S. mansoni DNA was detected by touchdown PCR in sera at 2 weeks PI. The intensity of S. mansoni DNA bands in infected mice sera detected in polyacrelamide gel electrophoresis increased gradually during the experimental period. Data recommended touchdown PCR potential tool for the early diagnosis of S. mansoni infection.

Co-infection with Cryptosporidium parvum and Cyclospora cayetanensis in immunocompromised patients.

Forty nine stool specimens collected from severe diarrheic patients. Eight were suffering from Hodgkin's lymphoma, and the rest were suffering from acute lymph plastic leukaemia. All were examined microscopically for protozoan parasites mainly, Cryptosporidium parvum and Cyclospora cayetanensis. Of the patients, 34 (69.4%) were positive and 15 (30.6%) were negative by both microscopy and nested PCR. An additional 12 (24.5%) who were negative by microscopy were positive by nested PCR. Stool examination revealed 16 cases with C. parvum, and 6 with C. cayetanensis, and 3 cases showed mixed infection. The results were compared with the established nested PCR assay to detect DNA directly from stool specimens. The patients <3 years old more affected by Cryptosporidium infection, unlike Cyclospora sp. Infection was in older age groups, which reflected the modes of parasite' transmission.. Diarrheal illness was stronger for Cyclospora than for Cryptosporidium. After the extraction of DNA from stool, a 402-bp fragment of C. parvum, and 602 bp fragment of C. cayetanensis was amplified. The amplified products, 194-bp DNA fragment for C. parvum, and 306 bp DNA fragment of C. cayetanensis were used for the second run. This study indicated that primers were specific for DNA of C. parvum and C. cayetanensis. PCR detected a total of 22 (44.9%) positives for C. parvum infection (6 negative by AF stool examination), and 12 (24.5%) positives for C. cavetanensis. Infection (6 negative by AF stool examination), 7 (14.3%) showed mixed infection (4 negative by AF stool examination), all microscopic negative specimens were positive by successive stool examination. Microscopy exhibited sensitivity of 72.7% for C. parvum, 50% for C. cayetanensis and 100% specificity for both parasites compared to 100% sensitivity and specificity with PCR. So, PCR is more sensitive and easier to interpret but required more hands-on time to perform and is more expensive. However, PCR batch analysis reduces the cost considerably.

Degree of symptoms versus copro-antigen levels in Giardia lamblia infection.

A total of 82 out-patients were examined for Giardia copro-antigens and 12 neonate stool samples as control. ELISA had a sensitivity of 100% and specificity of 91.67%. ELISA (O.D.) had neither significant correlation to Giardia cyst count, to stool consistency or presence of blood, mucus or fat in stool, nor to age but positive correlation to the severity of diarrhoea, colic, nausea, anorexia, weight loss, distension and fatigue. Giardia cyst count was higher in cases with loose stool, while ELISA (O.D.) correlated positively with symptoms except constipation and vomiting. The different in clinical outcome of giardiasis can be attributed, partially to strain differences and host resistance.

Molecular characterization of Cryptosporidium parvum isolates obtained from humans.

Fifty stool specimens collected from severe diarrheic patients attending Misr University Hospital, were examined microscopically for protozoan parasites mainly, Cryptosporidium parvum. Stool examination revealed 22 cases with C. parvum, 8 with E. histolytica, 14 with G. intestinalis and six were parasite-free. The results were compared with the established nested PCR assay to detect DNA directly from stool specimens. After the extraction of DNA from stool, a 402-bp fragment of C. parvum DNA was amplified with two 26-mer outer primers. The amplified products, 194-bp DNA fragment, were used for a second run. This study indicated that the used primers are specific for DNA of C. parvum. The PCR detected a total of 28 positives; six of these cases were negative by AF stool examination, which eventually confirmed to be positive by several successive examinations of the stool and/or duodenal aspiration. Microscopy exhibited 78.5% sensitivity and 100% specificity compared to 100% specificity and sensitivity with PCR. Consequently, PCR is more sensitive and easier to interpret but required more hands-on time to perform and is more expensive than microscopy. However, PCR batch analysis reduces the cost considerably.

Human infection with Dicrocoelium dendriticum in Riyadh district (Saudi Arabia).

Dicrocoelium dendriticum (the lancet fluke) is less common liver parasite. Adults inhibit bile duct, while characteristic eggs pass in stools. During one academic year, a total of 1196 patients referred to Parasitology Laboratory from King Abdel Aziz University Hospitals, 121 patients were positive for D. dendriticum. The majority was Saudis, and only 32 of the 121 patients (26.4%) had true dicrocoeliosis, as detected by re-examination after three days of liver free diet. The other patients have false Dicrocoeliasis. The infection was influenced by the patients' behaviours. The Saudis consume more sheep meat during month (Du-Elheja) of Haj and Feast. Examination of the slaughtered sheep at the governmental slaughterhouses showed the majority of them have the liver infected (9-24%) with the lancet liver fluke. It is concluded that rate of infection proportionates with the rate of sheep infection and the consumption of infected livers. For proper estimation of human infection should be on diet free liver before examination.

Human infection with Onchocerca volvulus in Asir District (Saudi Arabia).

During one academic year, three patients were referred to Parasitology Laboratory from Dermatology Outpatients Clinics in King Abdulaziz University Hospitals. They were diagnosed as Sowda (chronic hyperactive form of onchocerciasis volvulus). The patients came from Asir Region in the Southern of the Kingdom. The lesion was characterized by a sever papule dermatitis localized to the lower limbs, with marked skin darkening. There was extensive follicular hyperplasia of the regional lymph nodes in two cases only. The skin snips taken from the three patients were positive microfilariae. On the other hand, the urine sample of one patient was positive. Six months after the onset of treatment by the clinicians in the Specialized Hospital, skin snips and urine samples were negative.

Fasciola immature stages sought in Lymnaea species and Biomphalaria species in the water bodies of Dakahlia Governorate.

Examination of the five different water bodies in Dakahlia governorate, revealed four species of Lymnaea. These were L. natalensis (68.4%). L. truncatula (16%), L. stagnalis (12.2%) and L. columella (3.4%). Also, two species of Biomphalaria were recovered. These were B. alexandrina (54.7%) and B. glabrata (45.3%). Examination of all these snails showed natural infection with immature stages of Fasciola sp. in 5.5% of L. natalensis (= cailliaudi), 3.1% in L. truncatula and 0.67% in B. alexandrina. The importance of these snails in dissemination and spreading of fascioliasis was discussed.