M. Dolores Bargues

ITS-2 rDNA sequencing of Gnathostoma species (Nematoda) and elucidation of the species causing human gnathostomiasis in the Americas.

From several gnathostome species the complete internal transcribed spacer ITS-2 ribosomal DNA (rDNA) repeat sequence and a fragment of the 5.8S rDNA were obtained by direct polymerase chain reaction cycle-sequencing and silver-staining methods. The size of the complete ITS-1 sequence in agarose gel electrophoresis was also obtained. The ITS-2 enabled the differentiation of Gnathostoma spinigerum from Thailand and Gnathostoma binucleatum from Mexico and Ecuador and confirmed the validity of the latter. Gnathostoma turgidum, Gnathostoma sp. I (=Gnathostoma procyonis sensu Almeyda-Artigas et al., 1994), and Gnathostoma sp. II (=G. turgidum sensu Foster, 1939 pro parte), all from Mexico, proved to be independent species, but Gnathostoma sp. III, also from Mexico, could not be differentiated from G. turgidum. In Mexico and Ecuador, gnathostomes involved in human infection and that had been classified as G. spinigerum belong to G. binucleatum. The 5.8S rDNA sequences of the 6 Gnathostoma species studied were identical. The results of the ITS-1 agreed with those results of ITS-2.

High fascioliasis infection in children linked to a man-made irrigation zone in Peru

We detected 10 protozoan and nine helminth species in surveys of 338 5–15 year‐old Quechua schoolchildren in three communities of the Asillo zone of the Puno region, located at a very high altitude of 3910 m in the Peruvian Altiplano. The area proved to be hyperendemic for human fascioliasis with a 24.3% overall mean prevalence of Fasciola hepatica, local prevalences ranging between 18.8 and 31.3%, and infection intensities of up to 2496 eggs per gram of faeces (epg), with 196–350 epg (mean: 279 epg) and 96–152 epg (123 epg) as arithmetic and geometric means, respectively. Prevalences did not significantly vary between schools and in relation to sex. No statistical differences were found in intensities between schools, nor according to sex or age groups between and within schools, although the highest overall egg counts were detected in girls and in the youngest age group. Asillo zone is a man‐made irrigation area built only recently to which both liver fluke and lymnaeid snails have quickly adapted. The region appears to be isolated from the Northern Bolivian Altiplano natural endemic area. Such man‐made water resources in high altitude areas of Andean countries pose a high fascioliasis risk. Significant positive association of F. hepatica with protozooses following a one host life cycle, such as Giardia intestinalis, suggests that human infection mainly occurs through drinking water. This is supported by additional evidence such as the absence of typical aquatic vegetation in the drainage channels inhabited by lymnaeid snails, the absence of aquatic vegetables in the traditional nutrition habits of the Quechua inhabitants, and the lack of potable water systems inside dwellings, which requires inhabitants to obtain water from irrigation canals and drainage channels.

Hyperendemic human fascioliasis in Andean valleys: an altitudinal transect analysis in children of Cajamarca province, Peru.

A coprological survey including 476 2-18 year old school children from six rural localities between 2627 and 3061 m altitude was performed in Cajamarca province, Peru. Prevalences of fascioliasis ranging from 6.7 to 47.7% (mean 24.4%) proved to be the highest so far recorded in that human hyperendemic area. Higher prevalences in females and in the 2-5 year old group were not significant. Intensities ranged from 24 to 864 eggs per gram (arithmetic mean: 113; geometric mean: 68), the majority shedding less than 100, and without significant differences according to gender or age group. Fasciola hepatica was the most common helminth within a spectrum of 11-12 protozoan and 9-11 helminth species, 97.3% of the children showing infection with at least one parasite. The highest levels corresponded to coinfection with seven different species in females and subjects older than 5 years. Fascioliasis prevalence correlation with altitude appeared significant. An epidemiological characterisation of the valley transmission pattern of fascioliasis in Cajamarca is made by comparison with other better known hyperendemic areas. Results suggest that human fascioliasis may be widespread throughout different parts of Cajamarca province, even far away from the city, and that long-term fascioliasis chronicity and superimposed repetitive infections may be probably frequent.

Genetic Variability and Phylogenetic Relationships within Trypanosoma cruzi I Isolated in Colombia Based on Miniexon Gene Sequences

Phylogenetic studies of Trypanosoma cruzi have identified the existence of two groups: T. cruzi I and T. cruzi II. There are aspects that still remain unknown about the genetic variability within the T. cruzi I group. Given its epidemiological importance, it is necessary to have a better understanding of T. cruzi transmission cycles. Our purpose was to corroborate the existence of haplotypes within the T. cruzi I group and to describe the genetic variability and phylogenetic relationships, based on single nucleotide polymorphisms (SNPs) found in the miniexon gene intergenic region, for the isolates from different hosts and epidemiological transmission cycles in Colombian regions. 31 T. cruzi isolates were molecularly characterized. Phylogenetic relationships within T. cruzi I isolates showed four haplotype groups (Ia–Id), associated with their transmission cycle. In previous studies, we reported that haplotype Ia is mainly associated with the domestic cycle and domiciliated Rhodnius prolixus. Haplotype Ib is associated with the domestic cycle and peridomestic cycle, haplotype Ic is closely related with the peridomestic cycle, and haplotype Id is strongly associated with the sylvatic cycle. The phylogenetic methodologies applied in this study are tools that bolster the associations among isolates and thus shed light on Chagas disease epidemiology.

Schistosomiasis reaches Europe

An outbreak of urogenital schistosomiasis has been detected in Europe, with patients affected in France,1,2 Germany,1,2 and Italy. The infection originated in Corsica, in a river north of Porto-Vecchio, a popular tourist destination (figure). The introduction of schistosomiasis is believed to be associated with infected people arriving from a region endemic for the exclusively African parasite, Schistosoma haematobium and disseminating parasite eggs through their urine into snail breeding sites along the river.

Fascioliasis: a worldwide parasitic disease of importance in travel medicine.

Fascioliasis is a foodborne zoonotic disease caused by the two parasite species Fasciola hepatica and Fasciola gigantica. This trematodiasis has never been claimed special relevance for travellers and migrants. However, the situation has drastically changed in the last two decades, in a way that fascioliasis should today be included in the list of diseases to be enhanced in Travel Medicine. Different kind of travellers have been involved in human infection reports: business travellers, tourists, migrants, expatriated workers, military personnel, religious missionaries, and refugees. Europe is the continent where more imported cases have been reported in many countries. More cases would have been probably reported in Europe if fascioliasis would be a reportable disease. In the Americas, most of the reports concern cases diagnosed in USA. Relative few patients have been diagnosed in studies on travellers performed in Asia. In Africa, most cases were reported in Maghreb countries. Blood eosinophilia and the ingestion of watercress or any other suggestive freshwater plant in anamnesis are extremely useful in guiding towards a fascioliasis diagnosis in a developed country, although may not be so in human endemic areas of developing countries. Several suggestive clinical presentation aspects may be useful, although the clinical polymorphism may be misleading in many cases. Non-invasive techniques are helpful for the diagnosis, although images may lead to confusion. Laparoscopic visualization should assist and facilitate procurement of an accurately guided biopsy. Endoscopic retrograde cholangiopancreatography (ERCP) is the first choice in patients in the chronic phase. ERCP and sphincterotomy are used to extract parasites from the biliary tree. Fluke egg finding continues to be the gold standard and enables for burden quantification and establishing of the drug dose. Many serological and stool antigen detection tests have been developed. Immunological techniques present the advantages of being applicable during all periods of the disease, but fundamentally during the invasive or acute period, as well as to other situations in which coprological techniques may present problems. Triclabendazole is the drug of choice at present, although the spread of resistance to this drug is challenging. Prevention mainly concerns measures to avoid individual infection by considering the different human infection sources.

A new baseline for fascioliasis in Venezuela: lymnaeid vectors ascertained by DNA sequencing and analysis of their relationships with human and animal infection

BackgroundHuman and animal fascioliasis poses serious public health problems in South America. In Venezuela, livestock infection represents an important veterinary problem whereas there appear to be few human cases reported, most of which are passively detected in health centres. However, results of recent surveys suggest that the situation may be underestimated in particular areas. To obtain a baseline for future fascioliasis assessment, studies were undertaken by means of rDNA ITS-2 and ITS-1 and mtDNA cox 1 sequencing to clarify the specific status of Venezuelan lymnaeids, their geographical distribution and fascioliasis transmission capacity, by comparison with other American countries and other continents.ResultsResults obtained completely change the lymnaeid scenario known so far. The relatively rich lymnaeid fauna of Venezuela has been proven to include (i) Lymnaea meridensis and L. neotropica as the only native members, (ii) L. cubensis and Pseudosuccinea columella introduced from the Caribbean area, and (iii) Galba truncatula and L. schirazensis introduced from the Old World. The absence of representatives of the stagnicoline and Radix groups is remarkable. Four species are fascioliasis vectors: G. truncatula, L. cubensis and L. neotropica, which have the capacity to give rise to human endemic areas, and P. columella, which is a source of animal infection and is responsible for the spread of disease. Vector capacity in the apparently highland endemic L. meridensis is to be confimed, although may be expected given its phylogenetic relationships. Similarly as elsewhere, the non-transmitting L. schirazensis has been confused with L. cubensis, also with G. truncatula and possibly with L. neotropica.ConclusionsThe new scenario leads to the re-opening of many disease aspects. In Venezuela, altitude appears to be the main factor influencing fascioliasis distribution. Human infection shows an altitude pattern similar to other Andean countries, although a differing highland/lowland impact on animal infection does not appear evident. The overlap of G. truncatula, L. cubensis and probably also L. neotropica in temperate and cold zones suggests a higher risk for human infection in mid and high altitude areas. A lymnaeid species mapping by means of DNA markers becomes a priority to determine human and animal fascioliasis distribution in Venezuela, owing to the importance of lymnaeid vectors in defining transmission and epidemiological patterns.

Higher physiopathogenicity by Fasciola gigantica than by the genetically close F. hepatica: experimental long-term follow-up of biochemical markers

BACKGROUND Fascioliasis is caused by Fasciola hepatica and F. gigantica. The latter, always considered secondary in human infection, nowadays appears increasingly involved in Africa and Asia. Unfortunately, little is known about its pathogenicity, mainly due to difficulties in assessing the moment a patient first becomes infected and the differential diagnosis with F. hepatica. METHODS A long-term, 24-week, experimental study comparing F. hepatica and F. gigantica was made for the first time in the same animal model host, Guirra sheep. Serum biochemical parameters of liver damage, serum electrolytes, protein metabolism, plasma proteins, carbohydrate metabolism, hepatic lipid metabolism and inflammation were analysed on a biweekly basis as morbidity indicators. Serum anti-Fasciola IgG, coproantigen and egg shedding were simultaneously followed up. RESULTS rDNA and mtDNA sequencing and the morphometric study by computer image analysis system (CIAS) showed that fasciolids used fitted standard species characteristics. Results demonstrated that F. gigantica is more pathogenic, given its bigger size and biomass but not due to genetic differences which are few. Fasciola gigantica shows a delayed development of 1-2 weeks regarding both the biliary phase and the beginning of egg shedding, with respective consequences for biochemical modifications in the acute and chronic periods. CONCLUSIONS The higher F. gigantica pathogenicity contrasts with previous studies which only reflected the faster development of F. hepatica observed in short-term experiments.

An Updated Insight into the Sialotranscriptome of Triatoma infestans: Developmental Stage and Geographic Variations

Background Triatoma infestans is the main vector of Chagas disease in South America. As in all hematophagous arthropods, its saliva contains a complex cocktail that assists blood feeding by preventing platelet aggregation and blood clotting and promoting vasodilation. These salivary components can be immunologically recognized by their vectors hosts and targeted with antibodies that might disrupt blood feeding. These antibodies can be used to detect vector exposure using immunoassays. Antibodies may also contribute to the fast evolution of the salivary cocktail. Methodology Salivary gland cDNA libraries from nymphal and adult T. infestans of breeding colonies originating from different locations (Argentina, Chile, Peru and Bolivia), and cDNA libraries originating from F1 populations of Bolivia, were sequenced using Illumina technology. Coding sequences (CDS) were extracted from the assembled reads, the numbers of reads mapped to these CDS, sequences were functionally annotated and polymorphisms determined. Main findings/Significance Over five thousand CDS, mostly full length or near full length, were publicly deposited on GenBank. Transcripts that were over 10-fold overexpressed from different geographical regions, or from different developmental stages were identified. Polymorphisms were mapped to derived coding sequences, and found to vary between developmental instars and geographic origin of the biological material. This expanded sialome database from T. infestans should be of assistance in future proteomic work attempting to identify salivary proteins that might be used as epidemiological markers of vector exposure, or proteins of pharmacological interest.

DNA sequence characterisation and phylogeography of Lymnaea cousini and related species, vectors of fascioliasis in northern Andean countries, with description of L. meridensis n. sp. (Gastropoda: Lymnaeidae)

BackgroundLivestock fascioliasis is a problem throughout Ecuador, Colombia and Venezuela, mainly in Andean areas where the disease also appears to affect humans. Transmission patterns and epidemiological scenarios of liver fluke infection have shown to differ according to the lymnaeid vector snail species involved. These Andean countries present the vectors Lymnaea cousini, L. bogotensis and L. ubaquensis, unknown in the rest of Latin America. An exhaustive combined haplotype study of these species is performed by means of DNA sequencing of the nuclear ribosomal 18S RNA gene, ITS-2 and ITS-1, and mitochondrial DNA cox 1 gene.ResultsThe conserved 5.8S rDNA sequence corroborated that no pseudogenes are involved in the numerous non-microsatellite/minisatellite-related indels appearing between the ITS-2 and ITS-1 sequences when comparing different L. cousini - L. bogotensis populations. Sequence analyses and phylogenetic reconstruction methods including other lymnaeid vector species show that (i) L. bogotensis is a synonym of L. cousini, (ii) L. ubaquensis is a synonym of Pseudosuccinea columella, and (iii) populations of L. cousini hitherto known from Venezuelan highlands indeed belong to a new species for which the name L. meridensis n. sp. is proposed. This new species is described and a complete phenotypic differentiation provided.ConclusionsITS-2, ITS-1 and cox 1 prove to be good markers for specimen classification and haplotype characterisation of these morphologically similar lymnaeids in endemic areas. Analysis of the 18S gene and phylogenetic reconstructions indicate that L. cousini and L. meridensis n. sp. cluster in an evolutionary line different from the one of P. columella, despite their external resemblance. This suggests an evolutionary phenotypic convergence related to similar environments and which has given rise to frequent specimen misclassification. Body size and phylogenetic relationships of L. meridensis n. sp. with well-known vectors as Lymnaea cousini and P. columella, as well as with Galba/Fossaria species, suggest that the new species may participate in disease transmission to both animals and humans in altitude areas during the yearly window in which temperatures are higher than the F. hepatica minimum development threshold. The involvement of L. cousini and P. columella in the transmission and geographical/altitudinal distribution of fascioliasis in these Andean countries is analysed.