An extensive malacological survey was carried out between 2005-2009 in order to clarify the exact number of lymnaeid species which may be intermediate hosts of Fasciola hepatica in Venezuela. Four species were discovered during this survey, including two local species: Lymnaea cubensis and Lymnaea cousini and two exotic species: Lymnaea truncatula and Lymnaea columella. The most common local species was L. cubensis which was found at 16 out of the 298 sampling sites. This species has a large distribution area throughout the Northern part of Venezuela and was encountered from sea level to an altitude of 1,802 m in state of Trujillo. The second local species L. cousini was collected at only two sites of the Andean Region at altitudes of 3,550 m and 4,040 m, respectively. The European L. truncatula was found at 24 sites all located in the states of Mérida and Táchira at an altitude varying between 1,540-4,000 m. The respective distribution areas of L. cubensis and L. truncatula do not appear to overlap, but more detailed malacological surveys are needed. The fourth lymnaeid species, L. columella was collected in a canal from Mérida at an altitude of 1,929 m and in an irrigation canal from the state of Guárico, at an altitude of 63 m. The role of these four lymnaeid species in the transmission of fascioliasis in Venezuela is discussed.

Lymnaea cousini Jousseaume, 1887 was collected in Mucubaji, Merida State, Venezuela, from a permanent pond located at a very high altitude (3760 m). Identification of the collected specimens was made by comparison with the original description of the shell by Jousseaume and the description of the renal organ and reproductive system of topotypic specimens by Paraense.

In endemic areas with low prevalence and low intensity of infection, the diagnosis of hepatic pathology due to the Schistosoma mansoni infection is very difficult. In order to establish the hepatic morbidity, a double-blind study was achieved in Venezuelan endemic areas, with one group of patients with schistosomiasis and the other one of non-infected people, that were evaluated clinically and by abdominal ultrasound using the Cairo classification. Schistosomiasis diagnosis was established based on parasitologic and serological tests. The increase of the hepatic size at midclavicular and midsternal lines (in hepatometry) and the hard liver consistency were the clinical parameters able to differentiate infected persons from non infected ones, as well as the presence of left lobe hepatomegaly detected by abdominal ultrasound. The periportal thickening, especially the mild form, was frequent in all age groups in both infected and uninfected patients. There was not correlation between the intensity of infection and ultrasound under the current circumstances. Our data suggest that in Venezuela, a low endemic area of transmission of schistosomiasis, the hepatic morbidity is mild and uncommon. The Cairo classification seems to overestimate the prevalence of periportal pathology. The specificity of the method must be improved, especially for the recognition of precocious pathology. Other causes of hepatopathies must be investigated.

We have previously confirmed the presence of common antigens between Schistosoma mansoni and its vector, Biomphalaria glabrata. Cross-reactive antigens may be important as possible candidates for vaccine and diagnosis of schistosomiasis. Sera from outbred mice immunized with a soluble Biomphalaria glabrata antigen (SBgA) of non-infected B. glabrata snails recognized molecules of SBgA itself and S. mansoni AWA by Western blot. Recognition of several molecules of the SBgA were inhibited by pre-incubation with AWA (16, 30, 36, 60 and 155 kDa). The only specific molecule of AWA, inhibited by SBgA, was a 120 kDa protein. In order to determine which epitopes of SBgA were glycoproteins, the antigen was treated with sodium metaperiodate and compared with non-treated antigen. Molecules of 140, 60 and 24 kDa in the SBgA appear to be glycoproteins. Possible protective effects of the SBgA were evaluated immunizing outbred mice in two different experiments using Freund's Adjuvant. In the first one (12 mice/group), we obtained a significant level of protection (46%) in the total worm load, with a high variability in worm recovery. In the second experiment (22 mice/group), no significant protection was observed, neither in worm load nor in egg production per female. Our results suggest that SBgA constitutes a rich source of candidate antigens for diagnosis and prophylactic studies.

After 57 years of successful control of schistosomiasis in Venezuela, the prevalence and intensity of infection have declined. Approximately 80% of the individuals eliminate less than 100 eggs/g of stools, therefore morbidity is mild and the majority are asymptomatic. The sensitivity of Kato-Katz decreases to approximately 60%. Available serological methods for the detection of circulating antigens only reach a 70% of sensitivity. Tests based on the detection of antibodies by immunoenzymatic assays have been improved. The circumoval precipitine test has shown a high sensitivity (97%), specificity (100%), and correlation with oviposition, being considered the best confirmatory diagnostic test. Additionally to the classical immunoenzymatic assays, the development of the alkaline phosphatase immunoassay, allowed to reach a 100% specificity with an 89% sensitivity. Recently, we have developed a modified ELISA in which the soluble egg antigen is treated with sodium metaperiodate (SMP-ELISA) in order to eliminate the glycosilated epitopes responsible for the false positive reactions. The specificity and sensitivity reaches 97% and 99%, respectively. Synthetic peptides from the excretory-secretory enzymes, cathepsin B (Sm31) legumain (Sm32) and cathepsin D (Sm45), have been synthesized. The combination of two peptides derived from the Sm31 have been evaluated, reaching a sensitivity of 96% when analyzed independently and with a 100% specificity. Antibodies raised in rabbits against peptides derived from the Sm31 and Sm32 are currently evaluated in two different antigen-capture-based assays. The development of a simple, cheap and reliable test that correlates with parasite activity is a major goal.

Severe schistosomiasis is a rare event in Venezuela nowadays, after a successful national campaign by the Schistosomiasis Control Program. Unfortunately, this program has practically disappeared, and snail surveillance in field is not a priority, anymore. Thus, schistosomiasis has become a neglected disease in this country. However, surveys in different populations from the endemic area have shown particular epidemiological features described herein. In five communities we evaluated 2,175 persons and searched for the presence of Biomphalaria glabrata snails. Some markers were used for classifying schistosomiasis foci: mean age of the persons with Schistosoma mansoni eggs in the stools, serological tests, presence of B. glabrata snails, and intensity of infection. Places without B. glabrata snails and with few schistosomiasis cases were defined as "past transmission sites"; a site with abundant snails but few cases was defined as "potential risk"; "new transmission" foci were characterized by the presence of infected snails and young people passing eggs in the stools. A "re-emergent" focus has shared these last features, showing in addition a place where schistosomiasis had been reported before. Recent evidences of active transmission with the increasing dispersion of B. glabrata snails, point out the necessity for the re-establishment of the Schistosomiasis Control Program in Venezuela.

Las condiciones geográficas, ambientales, culturales y económicas facilitan en algunas áreas las infecciones simultaneas por varios agentes. Parásitos de un mismo género o de diferentes familias o phyla, entre parásitos y sus hospederos, comparten moléculas capaces de despertar respuestas inmunes comunes. Este fenómeno permitiría la reactividad cruzada entre parásitos, que tendría como ventaja la eficacia de un producto de amplio espectro (vacuna o medicamento) al poder atacar varios agentes parasitarios en una población poliparasitada. Por el contrario, como desventaja se destaca la inespecificidad del inmunodiagnóstico reflejada como falsos positivos por reactividad cruzada, detectándose anticuerpos o antígenos compartidos que no pertenecen propiamente al agente sospechado. Adicionalmente, la respuesta inmune a un agente patógeno puede dirigirse contra tejidos del propio huésped, desencadenando fenómenos de autoinmunidad. En esquistosomiasis se han desarrollado técnicas de inmunoensayo de mejor especificidad que las tradicionales. Sin embargo, siendo Venezuela un país en el cual coexisten áreas endémicas donde se solapan infecciones parasitarias tales como esquistosomiasis, nematodes intestinales y cisticercosis, se presentan inconvenientes de sobreestimación de prevalencias ocasionando limitaciones en el tratamiento masivo con praziquantel y confusión en la definición de verdaderos casos de esquistosomiasis o cisticercosis. Investigaciones tendientes a mejorar el inmunodiagnóstico disminuyendo los falsos positivos, mediante ensayos de captura de anticuerpos y antígenos para ambas parasitosis, se desarrollan en la actualidad.

Geographical, environmental, cultural and economical conditions favor polyparasitism of several agents in tropical areas. Different species of parasites or from different genera, families or phyla, or between parasites and their vectors, usually share molecules that are able to elicit common immune responses, that are defined as antigenic community, responsible for cross reactivity. This phenomenon may have the advantage of the possible application of an effective product (vaccine or drug) simultaneous against different species of parasites. One of the most deleterious consequences is the inespecific reactions in immunodiagnostic test. Additionally, immune response against some pathogens could be inespecifically directed against host tissues, being responsible for autoimmune reactions. In certain parasitic diseases like schistosomiasis, there are still some problems of inespecifity with other helminths. In Venezuela where intestinal parasites, schistosomiasis and cysticercosis can overlap, classic immunoenzymatic assays overestimate prevalences, limiting the capacity of the control programme for the application of massive treatment with praziquantel. Research directed to improve immunodiagnosis, reducing false positives with tests based on the demonstration of specific circulating antibodies or antigens, are currently being investigated.

Schistosomiasis control seems to be different in countries were low parasitic burden and asymptomatic clinical patients are the features of majority of cases. Immunological methods must substitute the traditional coprologic techniques used for some decades in the Control Program. Circumoval Precipitin Test (COPT), intradermal test and ELISA with soluble egg antigen (SEA) are evaluated for using as tools for seroepidemiologic studies. COPT and ELISA were performed after treatment to known their utility when impact of chemotherapy must be assessed. One hundred sixty five persons were followed up 3, 6, 9 and 12 months after treatment. The mean sensitivity of CPT studied by age groups was 95.6% which is very important considering that 88.4% of the studied population excreted less than 100 egg/gr of feces, while sensitivity of intradermal test was 58.2%. Children showed the highest ractivity to COPT. When treatment is effective, COPT reactivity progressively disminish until become negative one year later. In the non cure group, the COPT reactivity disminished but never below 20%. ELISA-SEA did not modify one year after treatment. Effort should be made to isolate fractions of eggs Schistosoma mansoni whose antibodies disappear after treatment.

Dezesseis doentes infectados e não tratados com S. mansoni (5 com infecção recente e 11 com doença crônica), foram submetidos à avaliação de atividade de células exterminadoras na turais (NK) "in vitro" frente a células alvo de linhagem K562. Os níveis de atividade das céluIas NK em 9 de 11 doentes (827r) com a infecção crônica foram significativamente menores (média = 15 ± 6%) quando comparados aos pacientes com infecção recente (média =41 ± 9%, p < 0,001) e aos indivíduos do grupo controle (média = 38 ± 13%, p < 0,001). Porém, tanto nos doentes como nos controles, a atividade de céluIas NK foi estimulada pelo antígeno solúvel do parasito adulto (SAWA), indicando que as céluIas NK, mesmo na fase crônica da infecção, têm capacidade de responder ao antígeno dos parasitos. Estes resultados sugerem a possível participação das células NK no mecanismo efetor de defesa contra o S. mansoni.

Sixteen S. mansoni infected and untreated patients (5 with recent infection and 11 with chronic disease) were evaluated for their in vitro natural killer (NK) activity against the NK sensitive target K562 cell line. NK levels in 9 out of 11 patients (82%) with chronic disease were significantly lower (mean = 15 ± 6%),compared with patients recently infected (mean = 41 ± 9% p < 0.001) and with the control group (mean = 38 ± 13% p < 0.001). However, both patients and controls NK activity was stimulated by soluble adult worm antigens (SAWA), indicating that NK function even in the chronic stage of the infection is able to respond to the parasite antigens. These results suggest the possibility of NK cell participation as effector mechanism against S. mansoni.

A resposta imune celular foi estudada em pacientes com infecção recente ou crônica por Schistosoma mansoni. Células mononucleares do sangue periférico pré cultivadas reagiram significantemente a antígenos do verme adulto (SAWA) do S. mansoni quando comparadas à preparação contendo células frescas. A adição de soro autólogo às células pré-cultivadas resultou em inibição da reação frente a SAWA ou antígenos de memória; o mesmo efeito foi notado quando os soros de pacientes foram adicionados a culturas de células alogênicas obtidas de indivíduos normais. A subpopulação CD4 foi a principal população celular respondedora a SAWA, sendo que esta reatividade foi intensamente suprimida na presença de preparações purificadas contendo monócitos-macrófagos. Estes resultados sugerem a ação de fatores inibidores, tanto humorais como celulares, sobre a resposta imune celular específica ao S. mansoni.

Cell mediated immune response was studied in patients with recent and chronic Schistosoma mansoni infection. Precultured peripheral mononuclear cells showed significantly higher responses to S. mansoni adult worm antigen (SAWA) when compared to fresh cell preparations. The addition of each patient serum to the precultured cells reactions to SAWA or recall antigens demonstrated a strong inhibitory serum action, which was also noted on allogeneic cells derived from healthy subjects. The CD4 subset was the main responding cell to SAWA being this reactivity highly suppressed by the presence of the monocyte macrophage accessory cells. We stressed the simultaneous inhibitory action of humoral and cellular factors on the specific cell response to S. mansoni.