Anti-brain protein autoantibodies are detectable in extraparenchymal but not parenchymal neurocysticercosis.

Neurocysticercosis (NC) presents a spectrum of clinical manifestations, with two broad clinical entities based on the central nervous system location of the parasite: extraparenchymal (EP-NC) and parenchymal (P-NC). In this work, using quantitative immunoblot methodology, we demonstrate the presence of autoantibodies to brain proteins in CSF from EP-NC, but not P-NC, patients. There was striking correlation between the level of autoantibodies and the levels of the secreted metacestode glycoprotein HP-10, suggesting that the level of stimulation of the autoantibody response may be a function of the number of viable parasites. Nine corresponding proteins autoantigens were provisionally identified by mass spectroscopy.

A modified lateral flow assay, using serum, for the rapid identification of human and bovine cysticercosis in the absence of false positives.

Background: Previously we reported the use of a monoclonal antibody-based (HP10) antigen (Ag) detection lateral flow assay (LFA) for the diagnosis of extraparenchymal neurocysticercosis (EP-NCC). The assay performed well when used with cerebrospinal fluid (CSF) samples but not with their paired serum samples, due to false-positive reactions in some known negative control cases. Methods: Our novel modification involves pretreatment of serum samples using a combination of sodium deoxycholate and dithiothreitol. Results: The modification overcomes the problem of false positives when using negative serum samples from clinically characterized cases of EP-NCC and bovine cysticercosis. In general, there was good agreement between HP10 Ag enzyme-linked immunosorbent assay (ELISA) and the HP10 Ag-LFA, but the HP10 Ag-ELISA was marginally more sensitive than the modified HP10 Ag-LFA. Conclusions: The modified HP10 Ag-LFA provides a field test for the rapid identification of endemic human and bovine cysticercosis.

Comparison of complementary diagnostic tests in cerebrospinal fluid and serum for neurocysticercosis.

The role of immunologic tests in the diagnosis of neurocysticercosis (NC) is controversial and few studies have made comparisons among them. The objective of this study was to compare immunological tests in both serum and cerebrospinal fluid (CSF) for the diagnosis of NC. We conducted a case-control study in Cuenca, Ecuador, enrolling patients with NC (N = 24) and matching them with other neurosurgical patients (N = 18). To detect cysticercal antigen, we used an HP10 antigen assay in serum and CSF ("HP10 Ag -serum -CSF") and a commercial antigen assay in serum (apDia, "ELISA-Ag-serum"), and to detect cysticercal DNA, we used a polymerase chain reaction (PCR) assay in CSF ("PCR-CSF"). Assay sensitivities were: HP10 Ag-serum (41.7%, 95% confidence interval [CI] 22.1-63.4), HP10 Ag-CSF (87.5%, 95% CI: 67.6-97.3), ELISA-Ag-serum (62.5%, 95% CI: 40.6-81.2), and PCR-CSF (79.2%, 95% CI: 57.9-92.9). Sensitivities were higher when limiting to participants with extraparenchymal NC. Specificity was 100% for all assays except ELISA-Ag-serum (72.2%). This preliminary study demonstrated the potential usefulness of the PCR and HP10 Ag assay in CSF, especially for extraparenchymal NC; thus, they could be considered as complementary diagnostic tools when neuroimaging is not conclusive.

Reciprocal contribution of clinical studies and the HP10 antigen ELISA for the diagnosis of extraparenchymal neurocysticercosis.

To evaluate diagnosis of active neurocysticercosis, paired cerebral spinal fluid (CSF) and serum samples from 24 neurocysticercosis (NCC) patients and 17 control neurological patients were assayed in the HP10 Taenia antigen (Ag) ELISA. The CSF samples were also tested with an HP10 Lateral Flow Assay (LFA). The HP10 Ag was detected by ELISA in the CSF of 5/5 patients with Definitive extraparenchymal NCC, and in 4/5 of the corresponding sera. In the Definitive parenchymal group, on the other hand, the HP10 Ag was absent in 2/3 CSF (with a very low value in the one positive sample) and all the corresponding serum samples. Samples of CSF from 4/7 patients in the Probable parenchymal group, were also significantly HP10 Ag positive, suggesting the presence of extraparenchymal cysts not identified by the imaging studies. With the possible exception of one patient, the corresponding serum samples of the Probable parenchymal NCC group, were all HP10 Ag negative. Samples of CSF from 9 NCC patients diagnosed with Mixed parenchymal and extraparenchymal NCC were all significantly HP10 Ag positive, confirming the presence of extraparenchymal cysts, with only 7/9 of the corresponding serum samples being HP10 positive. Thus detection of the HP10 Ag indicates extraparenchymal and not parenchymal cyst localization and is more sensitive with CSF than serum. Three neurological patients clinically diagnosed as subarachnoid cyst, hydrocephalus and tuberculoma, respectively, were clearly positive for HP10 Ag. Of these, two were confirmed as NCC by subsequent imaging; the third died prior to further examination. Thus, a total of 8 patients had their clinical diagnosis questioned. Finally, there was good agreement between the HP10 Ag ELISA and LFA with CSF samples giving an optical density ≥0.4 in the ELISA assay. In conclusion, the HP10 Ag assay should provide a valuable and reciprocal tool in the clinical diagnosis and follow up of extraparenchymal NCC.

Infección por Hymenolepis diminuta en infante del municipio Girardot, Maracay, estado Aragua, Venezuela / Infection by Hymenolepis diminuta in infant from the Girardot municipality, Maracay, Aragua State, Venezuela

Se presenta un caso inusual de infección por Hymenolepis diminuta en infante de un año de edad residente en Maracay, estado Aragua. El hallazgo se realizó de manera fortuita por el personal del Laboratorio de Parasitosis Intestinales y Serología de Esquistosomiasis de la Dirección General de Salud Ambiental del Ministerio del Poder Popular para la Salud, a donde fueron remitidos ejemplares adultos del parásito eliminado de forma espontánea por el paciente. Antes de la expulsión del parásito, el paciente presentó sintomatología digestiva, fiebre y manifestaciones alérgicas. La representante del infante refirió la presencia de ratas y ratones en el jardín de la vivienda. Este caso se constituye en el primer informe de infección por H. diminuta en el estado Aragua. Se sugiere realizar investigación epidemiológica más rigurosa para determinar la prevalencia de himenolepiosis.

Presentation of an unusual case of Hymenolepis diminuta infection in a one-year old infant residing at Maracay, Aragua State. The finding occurred fortuitously by the staff of the Intestinal Parasites and Schistosomiasis Serology Laboratory of the General Direction of Environmental Health of the Ministerio del Poder Popular para La Salud, who received two adult forms of the parasite eliminated spontaneously by the patient. Before the expulsion of the parasite, the patient had digestive symptoms, fever and allergic reactions. The infant’s guardian referred the presence of rats and mice in the garden of the house. This case is the first report of this type of infection by H. diminuta in Aragua State. A more rigorous epidemiological investigation is suggested to determine the prevalence of himenolepiosis.

Diagnostic epitope variability within Taenia solium 8 kDa antigen family: implications for cysticercosis immunodetection.

To study diagnostic epitopes within the Taenia solium 8 kDa antigen family, six overlapping synthetic peptides from an 8 kDa family member (Ts8B2) were synthesized and evaluated by ELISA and MABA with sera from patients with neurocysticercosis (NCC), from infected pigs and from rabbits immunized with recombinant Ts8B2 protein. The pre-immune rabbit sera and the Ts8B2 recombinant protein served as negative and positive controls, respectively. A similar analysis was done with the already described antigenic peptides from another member of the 8 kDa family, highly similar to Ts8B2, the CyDA antigen. Surprisingly, neither the Ts8B2 peptides nor the CyDA peptides were recognized by infected human and porcine sera. However, the entire Ts8B2 recombinant, as well as amino and carboxy-terminal halves were recognized by the positive serum samples. The observed lack of recognition of linear Ts8B2 peptides suggests that the principal serological response to the Ts8B2 family is focused on conformational epitopes in contrast to the previously observed antigenicity of the CyDA peptides. This differential antigenicity of 8 kDa family peptides could be related with parasite antigenic variability. The fact that rabbits experimentally immunized with Ts8B2 did make anti-peptide antibodies to peptides Ts8B2-6 and CyDA-6, located in the carboxy-terminal region demonstrated that the Ts8B2 peptides are not intrinsically non-immunogenic.

Peptide epitopes of the Taenia solium antigen Ts8B2 are immunodominant in human and porcine cysticercosis.

Ts8B2 is a gene which encodes for a member of the Taenia solium metacestode 8kDa antigen family. Since the Ts8B2-GST recombinant protein compares very favourably with other diagnostic antigens, and in order to study the antigenic nature and structure of this molecule, the Ts8B2 was expressed in prokaryotic and eukaryotic systems. The diagnostic potential of the recombinant Ts8B2 proteins was evaluated by enzyme-linked immunosorbent assays (ELISA) using a collection of serum and cerebrospinal fluid (CSF) samples from patients with clinically defined neurocysticercosis (NCC), and also sera from T. solium infected pigs. Despite the predicted glycosylation of the Ts8B2-Bac recombinant protein, there was very little difference in assay sensitivity/specificity when the Ts8B2 reagent was expressed in either prokaryotic or eukaryotic systems, suggesting that peptidic Ts8B2 epitopes are immunodominant in porcine cysticercosis and human neurocysticercosis. Conveniently, production of recombinant Ts8B2 in Escherichia coli is economical and facile, making it a feasible and practical choice as a diagnostic reagent for use in endemic areas. The Ts8B2 ELISA is particularly useful for the diagnosis of active as opposed to inactive cases of NCC and conduct of the assay is also facilitated by the fact that assay sensitivity is significantly greater when serum as opposed to CSF samples are employed.

Situación nutricional y características sociodemográficas de niños en una comunidad rural del estado Cojedes: Año 2005 / Children’s nutritional situacion and socio-demographic characteristics, in a rural comunity of the states of cojedes: Year 2005

La evaluación del estado nutricional, especialmente en grupos vulnerables, permite medir en forma indirecta la calidad de vida de una comunidad. Objetivo: Conocer el estado nutricional de niños que viven una comunidad rural del Estado Cojedes y las condiciones socioeconómicas de su grupo familiar. Metodología: Estudio descriptivo, transversal con 31 niños de 6 a 14 años, de ambos sexos. El diagnóstico nutricional se realizó por los indicadores: Talla-edad. Peso-talla, Peso-edad e Índice de Masa Corporal (IMC). Se utilizó para la clasificación, la referencia nacional y para la estratificación socioeconómica, Graffar - Méndez Castellano. En el análisis estadístico se utilizó distribución de frecuencia con porcentajes y medidas de tendencia central para las diferentes variables. Resultados: 83,9 % de los niños tuvo diagnóstico nutricional normal y 28,1 % presentó talla baja, 83,3% de las familias se encontró en situación de pobreza crítica. Conclusión: pese al estado de pobreza de las familias evaluadas, la mayoría de los escolares tienen IMC normal. Sin embargo, el hallazgo de talla baja refleja los posibles efectos deletéreos de la condición socioeconómica que predomina en la comunidad.

The evaluation of the nutritional state, especially in vulnerable groups, allows to measure in indirect form the quality of life of a community. Objetive: to know the state nutritional children who live a rural community on the Cojedes State and the socioeconomics conditions of their familiar group. Methodology: A descriptive, cross-sectional study was made where 31 children of 6 to 14 years, both sexes evaluated themselves. The nutritional diagnosis was made by the indicators: Stature-age. Weigh-it stature, Weigh-age and Index of Corporal Mass (IMC). Classification was used the national reference, and for the socioeconomic stratification, Graffar - Méndez Castellanos. For the statistical analysis the frequency allocation in percentage and measures of central tendency for the different variables was used. Results: 83,9% of the children had I diagnose normal nutritional and 28,1 % presented/displayed low stature. 83,3% of the families were in situation of critical poverty. Conclusion: in spite of the state of poverty of the evaluated families, most of the students they have a normal nutritional diagnosis, but the present low stature, reflects the possible deleterious effects of the socioeconomic condition that predominates.

Evaluation of recombinant HP6-Tsag, an 18 kDa Taenia saginata oncospheral adhesion protein, for the diagnosis of cysticercosis.

With the objective of providing inexpensive and reproducible assays for the detection of antibodies indicating exposure to Taenia saginata and Taenia solium, we have evaluated the diagnostic utility of the T. saginata oncosphere adhesion protein (HP6-Tsag), expressed in baculovirus (HP6-Bac) and bacteria (HP6-GST [glutathione S-transferase]), employing enzyme-linked immunosorbent assays (ELISAs) and sera from T. saginata infected cattle, T. solium infected pigs and serum and cerebrospinal fluid (CSF) samples from clinically defined T. solium neurocysticercosis (NCC) patients. The two recombinant proteins were antigenic in all three systems, with the signal to background ratio of the HP6-Bac ELISA slightly higher than that for the HP6-GST ELISA. Assay performance in cattle was similar to previously described peptide-based ELISA assays, although NCC sample sensitivity/specificity was marginally better. The sensitivity of the HP6-Bac and HP6-GST ELISAs was close for active human NCC (77.4 and 80.6% for serum and 76.9 and 73.1% for CSF samples, respectively). In inactive human NCC, however, the sensitivity of the HP6-Bac ELISA was almost twice that of the HP6-GST ELISA. Because peptides are relatively expensive and recombinant proteins are simple and economical to produce, the latter may provide useful reagents for antibody detection in countries with endemic cysticercosis/NCC.

Molecular cloning and characterisation of Ts8B1, Ts8B2 and Ts8B3, three new members of the Taenia solium metacestode 8 kDa diagnostic antigen family.

Antibody screening of a lambdaZAP-XR Taenia solium metacestode cDNA library yielded a clone (Ts8B1), with an insert of 345 bp, and an open reading frame of 258 bp, that coded for a protein with 85 amino acid residues. Alignment of the predicted amino acid sequence with sequences from SWISSPROT revealed an 88% identity with TcA5.5, a 10 kDa immunodiagnostic antigen of T. crassiceps, 75% identity with CyDA a T. solium metacestode antigen, 40-50% identity with several variants of the 8 kDa subunit of antigen B of Echinococcus spp. and with members of the T. solium metacestode 8 kDa antigen family. Two other Ts8B1 related molecules, Ts8B2 and Ts8B3, were identified in the metacestode cDNA library by PCR, coding for 85 and 66 amino acid polypeptides, respectively. Both Ts8B1 and Ts8B2 were characterized as E/S antigens through their subcellular localisation in the secretory membrane system when expressed in NRK cells. The three cDNA inserts were expressed, purified and probed in enzyme-linked immunosorbent assays (ELISA) with sera and cerebro-spinal fluid from patients with confirmed neurocysticercosis, and with sera from pigs infected with T. solium. The most promising antigen, Ts8B2, performed with a sensitivity of 96.8% and specificity of 93.1% in the detection of active NCC when using serum samples in the assay and performed similarly in the porcine system. The implications of these findings are discussed.

Taenia solium: characterization of a small heat shock protein (Tsol-sHSP35.6) and its possible relevance to the diagnosis and pathogenesis of neurocysticercosis.

A cDNA encoding for a predicted small heat shock protein (sHSP), Tsol-sfISP35.6, has been isolated by antibody screening of a Taenia solium c-DNA library. The clone was a full-length sequence (1172 bp) with an open reading frame of 945 bp and encoded for a 314 amino acid protein with deduced molecular mass of 35.6 kDa, isoelectric point of 5.6 arid the characteristic HSP20/alpha-crystallin domain duplicated. It was highly conserved, with a high sequence similarity with other platyhelminth sHSPs. Western blot analysis, using serum from neurocysticercosis patients (NCC), indicated that the purified Tsol-sHSP35.6 expression product was immunogenic, while in indirect ELISA, using the purified Tsol-sHSP35.6 expression product as antigen and serum samples from pigs and humans, 80% of T. solium infected pigs and 84% of patients with active, or 71% of patients with inactive NCC were sero-positive. The possible relevance of Tsol-sHSP35.6 in the diagnosis and pathogenesis of NCC is discussed.

Cloning and characterization of Taenia saginata paramyosin cDNA.

A lambdaZAP-express cDNA library of Taenia saginata metacestodes was constructed. Antibody screening yielded a clone with an insert of 3,408 bp, an open reading frame of 2,589 bp, a deduced sequence of 863 amino acid and a molecular mass of 98.89 kDa. Alignments of the predicted amino acid sequence showed identity with paramyosins from several species: 98.8% with Taenia solium, 96.3% with Echinococcus.granulosus and about 70% with Schistosoma spp. The insert was expressed and purified. A collagen binding assay was performed which showed that T. saginata GST-paramyosin retained this property in a dose-dependent manner. Problems were encountered due to high backgrounds in serological assays in the homologous T. saginata system. However, the recombinant paramyosin was recognized by antibodies present in 31.6% of sera from T. solium seropositive cysticercosis patients and 100% of the sera from acute cysticercosis patients. The immunodominant epitope was the carboxyl-terminal fragment of the molecule.