Antibody specific to 43 kDa excretory-secretory antigenic peptide of Taenia solium metacestode as a potential diagnostic marker in human neurocysticercosis

Recent studies suggest excretory-secretory (ES) antigen specific antibody detection tests to be of promising utility in laboratory diagnosis of many parasitic diseases in human including neurocysticercosis (NCC). The objective of the present study was to characterize the ES antigens collected from in vitro culture of Taenia solium metacestode larvae, and to identify specific ES peptides as diagnostic markers. Three ES peptides viz., 67 kDa, 43 kDa and 32 kDa, were found to be diagnostic for NCC based on high sensitivity and specificity of their reactivity to either serum or cerebrospinal fluid (CSF) specimens. More remarkably, the 43 kDa ES peptide was found reactive with CSF and serum specimens from confirmed NCC patients with absolute specificity and a high sensitivity (88.23% in serum and 89.28% in CSF). This peptide was also detected by sera and CSF from clinically suspected NCC patients but with a decreased sensitivity correlating with the decreasing order of the certainty of diagnosis as per a criteria proposed earlier. The 43 kDa ES peptide is suggested to be an important peptide of diagnostic utility in NCC.     

Neurocysticercosis immunodiagnosis using Taenia solium cysticerci crude soluble extract, excretory secretory and lower molecular mass antigens in serum and urine samples of Indian children

Neurocysticercosis (NCC), the most common neurological disorder of parasite etiology, results from lodgment of Taenia solium cysticerci in the central nervous system and is now increasingly being recognized in children. The confirmed diagnosis is based collectively on radiological findings and serodiagnostic techniques. The serodiagnostic techniques have variable sensitivity and specificity depending upon the technique, antigens used, location and number of cysts. Crude soluble extract (CSE), excretory secretory (ES) and lower molecular mass (LMM) (10-30 kDa) antigenic fraction of T. solium cysticerci were evaluated for antibody detection in serum and urine samples by ELISA. Serum and urine samples were collected each from 125 clinically suspected and radiologically proven NCC (111 with single Computed Tomography (CT) lesions and 14 with multiple CT lesions) and 125 control subjects (60 with neurological disorders other than NCC, 40 with other parasitic diseases and 25 apparently healthy subjects). The sensitivity of the ELISA with the use of CSE, ES and LMM antigenic fractions was 38.4%, 63.2% and 30.4% with serum (cut off dilution 400), 46.4%, 44% and 47.2% with neat urine and the specificity was 88%, 76.8% and 85.6% with serum (cut off dilution 400), 66.4%, 65.2% and 58.4% with neat urine samples, respectively. The study suggests that detection of antibody to ES antigen in serum samples may serve useful purpose for the serodiagnosis of human NCC.     

2D-PAGE analysis of Taenia solium metacestode 10-30 kDa antigens for the serodiagnosis of neurocysticercosis in children

Neurocysticercosis (NCC) caused by T. solium metacestode is an increasingly important health issue in Indian children. The sensitivity and specificity of available serological techniques were low in case of single cysticercus granuloma cases which is a more common feature in Indian patients who are children. Serum samples were collected from 13 clinically and radiologically suggestive NCC children and seropositive by ELISA, 25 clinically and radiologically suggestive NCC children and seronegative by ELISA and 25 control subjects. The 10–30 kDa antigens of T. solium metacestode were subjected to 2-dimensional gel electrophoresis (2D-PAGE) followed by enzyme-linked immunoelectrotransfer blot (EITB) assay to detect antibody in serum. Analysis of 10–30 kDa antigenic fraction 2D-PAGE map showed 31 proteins between 10 and ≤28 kDa and innumerable proteins between >28 and 30 kDa with the Isoelectric point of 3–10. All the 13 (100%) NCC seropositive and 15 (60%) out of 25 NCC seronegative samples were reactive with 2D fraction antigens. In the control group, none of the serum was reactive except 2 hydatid samples (92% specificity). The sensitivity and specificity of 2D-PAGE EITB assay were significantly higher than the ELISA which is the routine diagnostic method used in the endemic countries for the serodiagnosis of neurocysticercosis.     

Tear IgA-ELISA: a novel and sensitive method for diagnosis of ophthalmic cysticercosis

For the first time, presence of locally secreted specific IgA antibodies in tear specimen from human with ophthalmic cysticercosis is documented in the present study. The ELISA using Taenia solium metacestode excretory secretory (ES) antigen demonstrated a diagnostic level of IgA antibodies in tears with 100% sensitivity (6 out of 6 confirmed cases of ophthalmic cysticercosis) whereas, 25 of 34 (73.52%) clinically suspected cases were diagnosed positive. The ELISA using T. solium metacestode somatic antigen detected a diagnostic titre of IgA antibody in tears with a sensitivity of 50% (3 out of 6 confirmed cases). The specificity of the tear IgAELISA using T. solium metacestode somatic and ES antigens is observed to be 94.87% and 92.3%, respectively. Overall in tears, the ELISA using T. solium metacestode ES antigens for detection of IgA antibodies shows a higher diagnostic efficiency (93.33%) compared to that using T. solium metacestode somatic antigen (88.88%). The sensitivities of the ELISA for detection of IgA antibodies in tears is observed to be higher than that for detection of IgG antibodies in serum using either somatic or ES antigens of the parasite.     

Evaluation of an antigen from Taenia crassiceps cysticercus for the serodiagnosis of neurocysticercosis

We report here the evaluation of an antigen from Taenia crassiceps cysticercus as a potential reagent in an enzyme-immunoelectrotransfer blotting assay (EITB) and an enzyme-linked immunosorbent assay (ELISA) for the serodiagnosis of neurocysticercosis (NC) using clinical specimens obtained from patients in different phases of the disease. Serum and cerebrospinal fluid (CSF) samples from 64 patients suspected of having NC according to clinical manifestation and brain computed tomography were tested by ELISA with Taenia solium total saline antigen (ELISA-Tso) and by immunoblotting with T. crassiceps glycoproteins antigen (EITB-gpTcra). Forty-five serum samples were also tested immunoblotting with T. solium glycoproteins antigen (EITB-gpTso) and 30 were tested by ELISA with T. crassiceps 14 kDa glycoprotein (ELISA-gp14Tcra). Serum samples from apparently healthy individuals without any parasitic disease and from patients with other parasitic diseases were included as controls. The results of ELISA-Tso analysis with CSF obtained from 64 patients with NC showed that 53 (83%) were reactive. EITB-gpTcra analysis with serum from the same group of patients showed a sensitivity of 91%. Results of EITB-gpTso and EITB-gpTcra analysis with serum samples demonstrated an agreement of 100% between both tests. ELISA-gp14Tcra was positive in 23 (77%) sera, 22 with paired CSF positive. When ELISA-gp14Tcra results were compared to EITB-Tso results, a relative sensitivity of 95% was observed. All serum samples from the control group were negative in ELISA-gp14Tcra and only one serum from an individual with Taenia saginata was reactive in this assay, showing a specificity of 99% for ELISA-gp14Tcra. This fraction was purified in only one step with a good yield for use in immunoassays. We suggest that the gp14Tcra antigen can be used for detecting anti-cysticercus antibodies in serum samples for epidemiological investigation purposes and also for diagnostic screening of NC patients.     

Identification and functional characterization of alpha-enolase from Taenia pisiformis metacestode

Enolase belongs to glycolytic enzymes with moonlighting functions. The role of enolase in Taenia species is still poorly understood. In this study, the full length of cDNA encoding for Taenia pisiformis alpha-enolase (Tpeno) was cloned from larval parasites and soluble recombinant Tpeno protein (rTpeno) was produced. Western blot indicated that both rTpeno and the native protein in excretion–secretion antigens from the larvae were recognized by anti-rTpeno monoclonal antibodies (MAbs). The primary structure of Tpeno showed the presence of a highly conserved catalytic site for substrate binding and an enolase signature motif. rTpeno enzymatic activities of catalyzing the reversible dehydration of 2-phosphoglycerate (2-PGA) to phosphoenolpyruvate (PEP) and vice versa were shown to be 30.71 ± 2.15 U/mg (2-PGA to PEP) and 11.29 ± 2.38 U/mg (PEP to 2-PGA), respectively. Far-Western blotting showed that rTpeno could bind to plasminogen, however its binding ability was inhibited by ?-aminocaproic acid (?ACA) in a competitive ELISA test. Plasminogen activation assay showed that plasminogen bound to rTpeno could be converted into active plasmin using host-derived activators. Immunohistochemistry and immunofluorescence indicated that Tpeno was distributed in the bladder wall of the metacestode and the periphery of calcareous corpuscles. In addition, a vaccine trial showed that the enzyme could produce a 36.4% protection rate in vaccinated rabbits against experimental challenges from T. pisiformis eggs. These results suggest that Tpeno with multiple functions may play significant roles in the migration, growth, development and adaptation of T. pisiformis for survival in the host environment.     

The current status of neurocysticercosis in Eastern and Southern Africa

Some information has been documented on the epidemiology of neurocysticercosis in Eastern and Southern Africa through the monitoring of hospital-based patients with neurocysticercosis, community-based serological surveys of particular socio-economic groups of people and surveys of porcine cysticercosis. Studies have revealed that non-pork eaters have as great a chance of infection as a pork eater, the Xhosa-speaking people of the Eastern Cape Province have the highest prevalence of cysticercosis/taeniosis in South Africa probably due to the common practice of free-range pig farming and the lack of sanitation in these areas. Several studies have revealed high prevalence rates in children and interestingly, patients with active cysts suffering from epilepsy. A startling mode of transmission is where self-trained healers use Taenia segments either for benevolent (e.g. in the treatment of severe intestinal tapeworm infections) or malevolent (evil) purposes (e.g. women "poisoning" an unfaithful husband or lover by adding the contents of Taenia solium segments to beer).     

Taenia crassiceps infection disrupts estrous cycle and reproductive behavior in BALB/c female mice

Previously, it has been shown that parasitic infections are able to alter the normal mammal physiology, at several extents. Thus, we investigated the effects on estrous cycle and sexual behavior induced by intraperitoneal infection with Taenia crassiceps in female host mice. Along the weeks of infection, parasites were collected from the peritoneal cavity of female mice, showing the maximum parasite load at 16 weeks. No parasites were found outside peritoneal cavity. Vaginal estrous cycle was monitored daily for 4, 8, 12 and 16 weeks of infection, and results compared against age-matched female mice. Female sexual behavior (FSB) tests were performed, one test per week. Immediately after the last behavioral test, blood was collected by cardiac puncture for steroid determinations. First of all, there was a strong tissular damage in the female reproductive tract in all infected females. The phases of the estrous cycle were interrupted at 12 and 16 weeks, with increased leukocytes and the presence of a few cornified epithelial cells and nucleated epithelial cells. The FSB decreased starting 6 weeks post infection. On the 16th week, all infected female mice ceased to exhibit sexual responses, and estradiol levels showed a significant decrease. Control mice continued showing FSB and the different phases of the estrous cycle throughout the observation period. Our results strength the notion that parasites may be considered as an evolutionary force in the reproductive ability of mammals.     

Comparison of adult somatic and cysteine proteinase antigens of Fasciola gigantica in enzyme linked immunosorbent assay for serodiagnosis of human fasciolosis

Fasciolosis caused by Fasciola hepatica and Fasciola gigantica is one of the major public health problems in the world and in Iran. Considering that stool examination for Fasciola eggs is not a sensitive method and immunodiagnosis methods are more applicable for this purpose, so the present study was conducted to compare the somatic (S) and cysteine proteinase (CP) antigens of F. gigantica in IgG-ELISA to diagnose human fasciolosis. Serum samples obtained from 100 individuals collected during the fasciolosis outbreak in 1999 in the Gilan province of Northern Iran that were coprologically positive for fasciolosis were analyzed by IgG-ELISA. Sera from healthy control individuals, not infected with any parasitic diseases (n=50) and from others with different parasitic infections including hydatidosis (n=40), toxocariosis (n=20), amoebiosis (n=10), and malaria (n=5) were examined as well. The cut-off point for (S) and CP was 0.40 and 0.35, respectively. All 100 individuals that showed clinical manifestations of fasciolosis, were also seropositive using both antigens, whereas all 50 non-infected controls were seronegative, therefore the sensitivity of the test was 100% for both antigens. The specificity of (S) and CP antigens were calculated as 96.9 and 98.4%, respectively. The positive and negative predictive values of the test regarding (S) antigen were 96 and 100%, whereas these values as for CP antigen were 98 and 100% correspondingly. Two individuals with hydatidosis and two with toxocariasis had antibodies that were reactive against (S) antigen, whereas concerning CP antigen, one individual with hydatidosis and another with toxocariasis showed cross-reactivity against it. We have demonstrated that altogether CP antigen provides a more conclusive diagnosis as possessing lower cut-off and enabling better to discriminate between seronegative and seropositive subpopulations.     

Multiple genotypes of Taenia solium--ramifications for diagnosis,treatment and control

Mitochondrial DNA sequences of Taenia solium have fully been analyzed. Analysis of the full length of cytochrome c oxidase subunit 1 (1620 bp) and cytochrome b (1068 bp) genes of T. solium, isolated from Asia (China, Thailand, Indonesia and India), from Latin America (Mexico, Ecuador, Bolivia, Peru and Brazil) and from Africa (Tanzania, Mozambique and Cameroon), has revealed that the two phylogenies obtained were similar to each other regardless of the genes examined. The isolates from Asia formed a single cluster, whereas those from Latin America combined with those from Africa to form an additional cluster. It was estimated that these two genotypes emerged approximately 4-8 x 10(5) years ago. These results together with recent study of the ancient of human taeniid cestodes emerged several MYA in Africa, historical data on swine domestication, distribution of pigs and colonization patterns suggest that T. solium was introduced recently into Latin America and Africa from different regions of Europe during the colonial age, which started 500 years ago, and that T. solium of another origin independently spread in Asian countries, perhaps from China. Why did not T. solium of European origin invade or spread into Asia during the colonial age? Analysis of T. solium distribution must include other Taenia species, especially T. saginata and T. asiatica, which can not be differentiated from each other morphologically. BESS T-base analysis for differentiation of all human Taenia species including the two genotypes of T. solium, and T. saginata and T. asiatica has also been characterized. BESS T-base analysis differentiates African isolates from Latin American isolates as well but more samples should be analyzed for obtaining conclusive evidence for the latter. Serological analysis of cyst fluid of T. solium cysticerci obtained in China and Indonesia and from Mozambique and Ecuador indicates geographical differences in their banding patterns. These differences are discussed in the light of possible differences in pathology of T. solium worldwide. As it has been speculated that the ancient T. solium emerged several million years ago in Africa, it is necessary to analyze more isolates from Africa. Such working hypothesis may be evaluated combined with symptomatology and serology when we get additional DNA data from such areas, since there are some varieties of manifestation of neurocysticercosis with or without subcutaneous cysticercosis and of antigens of cyst fluid of T. solium from Asia and from Africa and/or America. Transfer of techniques of molecular identification and sero- and immuno-diagnoses between researchers and technicians from endemic countries using their own materials should be promoted with the aim of better international cooperation for the control of cysticercosis.     

Evaluation of excretory secretory and 10–30 kDa antigens of Taenia solium Cysticerci by EITB assay for the diagnosis of neurocysticercosis

Neurocysticercosis (NCC), caused by the presence of Taenia solium Cysticerci in the Central Nervous System is the most common neurological disease of parasite aetiology. The serodiagnostic methods available at present have variable sensitivity and specificity depending upon the antigen and technique used. The present study was aimed to assess the efficacy of T. solium Cysticerci excretory secretory (ES) and lower molecular mass (LMM) 10–30 kDa antigenic fractions for antibody detection in serum and urine samples by enzyme-linked immunoelectrotransfer blot (EITB) for the diagnosis of NCC. Serum and urine samples were collected from 125 clinically suspected and radiologically proven NCC children (111 patients with single lesion and 14 with multiple lesions) and 125 control subjects. With the use of ES and LMM antigenic fractions, the sensitivity of the EITB assay was 85·6% and 80·8% with serum and 76·8% and 50·4% with urine, respectively. The specificity was 64% and 61·6% with serum and 48% and 33·6% with urine samples, respectively. The study suggests that antibody detection to ES antigen in serum by EITB assay may serve better purpose for the serodiagnosis of human NCC.     

Evaluation of three recombinant Leishmania infantum antigens in human and canine visceral leishmaniasis diagnosis

Visceral leishmaniasis (VL) is a neglected disease and is fatal if untreated. Dogs serve as reservoirs for Leishmania infantum (syn. L. chagasi) due to their susceptibility to infection and high skin parasitism. Therefore, VL control in Brazil involves the elimination of seropositive dogs, among other actions. However, the most frequently used serological tests have limitations regarding sensitivity and specificity. In this study, we have selected three Leishmania antigens (C1, C8 and C9) and have produced them as recombinant proteins using pET-28a-TEV vector and Escherichia coli BL-21 as expression system. When tested in ELISA with human samples, the C9 antigen was the one showing the most promising results, with 68% sensitivity and 78% specificity. When testing canine samples, the C1, C8 and C9 antigens showed a sensitivity range from 70% to 80% and specificity range from 60% to 90%. The C1 antigen presented higher sensitivity (80%) and the C8 antigen presented higher specificity (90%). Due to it, we decided to mix and test C1 and C8 antigens together, resulting in the C18 antigen. The mix also yielded high percentages of detected symptomatic and asymptomatic dogs however it did not improve the performance of the diagnostic. Comparison of our tests with the tests recommended by the Brazilian Ministry of Health revealed that our antigens’ sensitivities and the percentage of detected asymptomatic dogs were much higher. Our results suggest that the C1, C8, C18 and C9 recombinant proteins are good antigens to diagnose canine visceral leishmaniasis and could potentially be used in screening tests. To diagnose human visceral leishmaniasis, the C9 antigen presented reasonable results, but more optimization must be performed for this antigen to provide better performance.     

Anisakis/Ascaris IgE ratio improves specificity for the diagnosis of Anisakis simplex sensitization in travellers and immigrants

Anisakis simplex is a fish parasite responsible for human infection and is able to induce IgE-mediated reactions with several clinical manifestations. Laboratory diagnosis of Anisakis allergy is based on the detection of specific IgE using parasite whole antigen. Unfortunately, these diagnostic tools detect cross-reactivities with other nematodes and micro-organisms leading to low specificity of the diagnostic tests. The aim of this retrospective study was to assess the diagnostic value of specific IgE to Anisakis for diagnosis of A. simplex-sensitization in native Spanish residents (IMM, n = 766) and subjects coming from tropical and sub-tropical geographic areas (TRO, n = 233). Since Ascaris is the human parasite most closely related to Anisakis, specific IgE to Ascaris was also determined to assess Anisakis cross-reaction with other nematodes and the diagnostic value of Anisakis/Ascaris IgE ratio for Anisakis allergy was examined. IMM and TRO groups showed similar specific IgE to Anisakis levels, while TRO had higher levels of specific IgE to Ascaris than IMM group (p = 0.001). ROC curve analysis determined that an Anisakis specific IgE threshold of 0.71 kU/L yielded 93% and 82% specificities in IMM and TRO groups, respectively. A cut-off value ≥4.4 for Anisakis/Ascaris IgE ratio increased specificity to 95% for samples having IgE to Ascaris ≥0.35. In conclusion, the ratio of specific IgE to Anisakis and Ascaris improved remarkably the specificity and this parameter easily obtained from the commercially available system could be useful in the diagnosis of hypersensitivity to A. simplex.     

Anthelmintic activity of benzimidazole derivatives against Toxocara canis second-stage larvae and Hymenolepis nana adults

The anthelmintic activity of 11 benzimidazole derivatives (A1-A11) and 2 thioureides N,N′-disubstituted (B1-B2) was determined. Each compound and albendazole was tested in vitro against Toxocara canis larvae and in vivo against Hymenolepis nana adult. Compounds A1-A6 and B1-B2 were designed as albendazole prodrugs. Compounds A8-A11 were designed as direct analogues of A7, which had previously proved to be an effective agent against Fasciola hepatica. Results of the in vitro screening showed that A6 was more active than albendazole at 0.18 μM (relative mobility 40% and 80%, respectively). Whereas that the in vivo evaluation against H. nana, compounds A7-A11 demonstrated significant activity in terms of removing cestode adults in the range of 88-97%, displaying better efficacy than albendazole (83%).     

Occurrence and prevalence of fish-borne Anisakis larvae in the spotted mackerel Scomber australasicus from Taiwanese waters

Anisakid nematodes have been found in a variety of marine fishes throughout the world and they are known to cause anisakiasis in human hosts. The present study investigated the prevalence of potentially zoonotic anisakid larvae in spotted mackerel caught from Taiwanese waters where fish represents an important food sources. Anisakis third-stage larvae (L3, n = 502) were isolated from 250 spotted mackerel Scomber australasicus. Anisakis L3 larvae were divided morphologically into two types, Anisakis type I larvae had a longer ventriculus and mucron while type II larvae had a shorter ventriculus and no mucron. Anisakis species were identified by polymerase chain reaction-restriction fragment length polymorphism (PCR–RFLP) of the internal transcribed spacer (ITS) regions of ribosomal DNA and direct sequencing. A simple molecular taxonomic key, utilizing RFLP by two restriction enzymes HinfI and HhaI, enabled the differentiation of the genus Anisakis. The prevalence, mean intensity and mean abundance of Anisakis nematodes recorded for the total specimens were 72.8%, 2.8 (1–15) and 2.0 (0–15), respectively. Anisakis pegreffii was determined to be the dominant species (prevalence = 57.2%) and important agent of human anisakiasis. A recombinant genotype (Anisakis simplex sensu stricto × A. pegreffii) was identified as the subdominant species (25.3%) followed by Anisakis typica (10%), Anisakis physeteris (4.0%), Anisakis paggiae (3.0%) and Anisakis brevispiculata (0.5%). The topology of the maximum likelihood and neighbor-joining trees show two well supported clades: one includes the species of A. pegreffii and the other includes A. paggiae, A. physeteris and A. brevispiculata, while A. typica has basal position to all other Anisakis spp. analyzed. This study advances our knowledge of the prevalence of different Anisakis spp. in the spotted mackerel from Taiwanese waters, which is helpful for monitoring the fish populations throughout a diverse array of aquatic ecosystems. More importantly, we provide the concise characterization of multiple Anisakis spp. by PCR–RFLP, which could also be applicable for the rapid diagnosis of human anisakiasis.     

Comparison of the effects of extracts from three Vitex plant species on Anopheles gambiae s.s. (Diptera: Culicidae) larvae

Acetone and methanol extracts of different parts of three Vitex species (leaves and stem bark of Vitex trifolia, leaves, stem bark and root bark of Vitex schiliebenii and stem and root bark of Vitex payos) were evaluated for their potential to control Anopheles gambiae Giles s.s. larvae (Diptera: Culicidae). The extracts gave different levels and rate of mortality of the larvae. Some (methanol extract of V. trifolia leaves, acetone extracts of stem bark and leaves of V. schiliebenii, acetone extract of root bark of V. payos) caused 100% mortality at 100 ppm in 72 h, with those of V. schiliebenii and V. payos showing faster rate of mortality (LT₅₀ = 8 h) than that of V. trifolia (LT₅₀ = 14 h). At lower doses of these extracts (≤50 ppm), most of the larvae failed to transform to normal pupae but gave larval–pupal intermediates between 4 and 14 days of exposure. Some pupated normally but the adults that emerged appeared to be weak and died within 48 h. Extracts of the stem bark of V. payos showed interesting effects on the larvae. Initially, the larvae were relatively hyperactive compared to those in control treatments. Later, the ones that did not transform to larval–pupal intermediates became stretched and inactive and died and floated in clusters on the surface. These observations suggest some interesting growth-disrupting constituents in the plants, with possible application in the practical control of mosquito larvae in aquatic ecosystems.     

A hundred years of controversy about the taxonomic status of Echinococcus species

The parasitic diseases which we know today as cystic and alveolar echinococcosis are zoonoses known since antique times, and 1855, respectively. Whether the two clinically and morphologically distinct diseases were caused, according to a “unicistic” and a “dualistic” theory, by only one or two different cestode species was the subject of a fierce, 100 years long debate involving scientists from many countries. The natural life cycle of Echinococcus granulosus was fully clarified in 1855 after successful animal experiments. In contrast, the natural final and intermediate hosts of Echinococcus multilocularis remained unknown, and the advocates of either theory had to draw on a number of surrogate arguments to defend their positions. The seesaw of reasoning and mutual defeats of the two theories, and the final recognition of E. multilocularis as an independent species in the 1950s are described in this article.     

Fluke egg characteristics for the diagnosis of human and animal fascioliasis by Fasciola hepatica and F. gigantica

In trematodiases, shape and size of the fluke eggs shed with faeces are crucial diagnostic features because of their typically reduced intraspecific variability. In fascioliasis, the usual diagnosis during the biliary stage of infection is based on the classification of eggs found in stools, duodenal contents or bile. The aim of the present study is to validate the identification of Fasciola species based on the shape and size of eggs shed by humans, characterizing their morphometric traits using a computer image analysis system (CIAS). The influence of both the geographical location and of the host (human and livestock) has been analysed. Coprological studies were carried out in fascioliasis human endemic areas, where only F. hepatica is present (the northern Bolivian Altiplano and the Cajamarca valley in Peru), and where F. hepatica and F. gigantica coexist (the Kutaisi region of Georgia, the Nile Delta in Egypt, and the Quy Nhon province in Vietnam). Classically, it is considered that at the abopercular end of the shell of Fasciola eggs there is often a roughened or irregular area. Nevertheless, results show that the frequency of the presence of this feature in F. hepatica is population-dependent, and therefore is not a pathognomonic criterion in diagnosis. The study reveals that eggs shed by humans show morphological traits different from eggs shed by animals. In humans, F. hepatica eggs are bigger and F. gigantica eggs are smaller than reported to date from livestock, and their measurements overlap when compared. The material analysed in this study shows that the size of eggs shed by humans from Georgia and Egypt corresponds to the F. hepatica morph, while the size of eggs shed by humans from Vietnam corresponds to the F. gigantica morph. Measurements of F. hepatica and F. gigantica eggs originating from humans and animals from sympatric areas overlap, and, therefore, they do not allow differential diagnosis when within this overlapping range. In this sense, the new results should aid clinicians since the application of the classic egg size range in human samples may lead to erroneous conclusions. Fasciolid egg size in human stool samples ought to be corrected in books and monographs related to medical parasitology and/or tropical medicine as well as in guides for clinicians and parasitic disease diagnosis analysts.     

First report of the activity of predatory fungi on Angiostrongylus cantonensis (Nematoda: Angiostrongylidae) first-stage larvae

The nematode Angiostrongylus cantonensis causes eosinophilic meningoencephalitis in humans and thus alternative methods of control should be studied. The objective of this work was to evaluate the predatory capacity of eight fungal isolates of the species Duddingtonia flagrans (AC001, CG768 and CG722), Monacrosporium thaumasium (NF34), M. sinense (SF53) and Arthrobotrys robusta (I31), A. cladodes (CG719) and A. conoides (I40) on first-stage larvae (L₁) of A. cantonensis under laboratory conditions. The treated groups contained 1000 conidia of the fungal isolates and 1000 A. cantonensis L₁ in Petri dishes containing 2% water-agar medium (2% WA). The control group (without fungi) contained only 1000 A. cantonensis L₁ in 2% WA. Evidence of predation was observed at the end of 7 days. Percentage reductions in L₁ were: AC001, 82.8%; CG768, 71.0%; CG722, 72.8%; NF34, 86.7%; SF53, 89.7%; I40, 48.3%; CG719, 84.7%; and I31, 80.4%. No significant difference was observed (p > 0.01) between the actions of the isolates used; however, a difference was noted (p < 0.01) in relation to the control group. The results of the present work, confirm previous reports of the effectiveness of the fungi D. flagrans, M. thaumasium, M. sinense and A. robusta in controlling larvae of potentially zoonotic nematodes, this being the first report on A. cantonensis L₁.