Enzyme-linked Immunoelectrotransfer Blot Assay Research Articles

Multiantigen print immunoassay (MAPIA) for the diagnosis of neurocysticercosis: a single-center diagnostic optimization and accuracy study in Lima, Peru.

Neurocysticercosis (NCC) is the most common helminthic infection of the human central nervous system. The antibody detection assay of choice is the enzyme-linked immunoelectrotransfer blot assay using lentil-lectin purified parasite antigens (LLGP-EITB, Western blot), an immunoassay with exceptional performance in clinical samples. However, its use is mainly restricted to a few research laboratories because the assay is labor-intensive and requires sophisticated equipment, expertise, and large amounts of parasite material for preparation of reagents. We report a new immunoprint assay (MAPIA) that overcomes most of these barriers. We initially compared the performance of five different antigen combinations in a subset of defined samples in the MAPIA format. After selecting the best-performing assay format (a combination of rGP50 + rT24H + sTs14 antigens), 148 archived serum samples were tested, including 40 from individuals with parenchymal NCC, 40 with subarachnoid NCC, and 68 healthy controls with no evidence of neurologic disease. MAPIA using three antigens (rGP50 + rT24H + sTs14) was highly sensitive and specific for detecting antibodies in NCC. It detected 39 out of 40 (97.5%) parenchymal NCC cases and 40/40 (100%) subarachnoid cases and was negative in 67 out of 68 (98.53%) negative samples. MAPIA using three recombinant and synthetic antigens is a simple and economical tool with a performance equivalent to the LLGP-EITB assay for the detection of specific antibodies to NCC. The MAPIA overcomes existing barriers to adoption of the EITG LLGP and is a candidate for worldwide use.

Serological and molecular detection of neurocysticercosis among epileptic patients in Nagpur, Maharashtra state (India).

Neurocysticercosis (NCC), one of the most important neuroparasitic diseases in humans, is caused by Cysticercus cellulosae, the metacestode stage of digenetic zoonotic cestode Taenia solium. The present study aims at the detection of anti-cysticercus antibodies in the sera of epileptic patients (n=26) visiting a tertiary care hospital in Nagpur, Maharashtra state, India, by an in-house developed indirect IgG-ELISA and enzyme-linked immunoelectro transfer blot (EITB) assay using different antigens (namely, Whole Cyst Antigen (WCA), Cystic Fluid Antigen (CFA), Scolex Antigen (SA), Excretory-Secretory Antigen (ESA) and Membrane-Body Antigen (MBA)) prepared from T. solium metacestodes to find out the status of NCC. An attempt has also been made for molecular detection of NCC from blood samples of those patients by Polymerase Chain Reaction (PCR) assay targeted at large subunit rRNA gene of T. solium. The IgG ELISA level of anti-cysticercus antibodies against WCA, CFA, SA, ESA and MBA antigens were as follows: 19.23 %, 23.07 %, 38.46 %, 30.76 % and 15.38 %. The seroreactivity to CFA, SA and ESA was found in equal proportions in patients with ring-enhancing lesions. In the EITB assay, the lower and medium molecular weight protein bands of SA and ESA were immunodominant compared to the higher WCA and CFA peptides. PCR positivity could be observed in 34.6 % (9/26) of the patients under study. It is the first report of detecting NCC among epileptic patients of the Nagpur region of Maharashtra state in India using serological and molecular tools.

Improvement of Enzyme-Linked Immunoelectrotransfer Blot Assay by Quantitative Approach for Foot-and-Mouth Disease diagnosis

Foot-and-mouth disease (FMD) is a highly contagious animal disease that causes devastating economic losses. The trade of live animals and derived products is only possible if the exporting country is free from disease, according to the World Organization for Animal Health (WOAH) code for FMD. One of the most important ways to prove disease-free is to measure the levels of non-structural protein antibodies (NPS) of FMD virus in the target population sampled. For detection of the disease status of a herd, mass screening and assays such as Enzyme-linked immunosorbent assay (ELISA) and Enzyme-linked immunoelectrotransfer blot assay (EITB) were developed and described in the WOAH diagnostic manual. In this study, recombinant FMDV NS proteins were produced and tested with sera panels collected from uninfected and naturally infected animals using a quantitative Western blot assay as an improved EITB, which enables numerical documentation and statistical analysis. NSP band intensities were used to determine the cut-off values, differentiating infected from non-infected animals and revealing variable sensitivity among the different NSPs. The quantitative EITB results also showed a correlation with the NSP-ELISA results.

Characterization of somatic and metabolic antigens of Cysticercus cellulosae and determination of immunodominant proteins

The present study shows the characterization of four somatic antigens (Whole Cyst Antigen (WCA), Cystic Fluid Antigen (CFA), Scolex Antigen (SA) and Membrane-Body Antigen (MBA)) and one metabolic antigen (Excretory-Secretory Antigen (ESA)) prepared from Cysticercus cellulosae originated from a naturally infected pig. Immunodominat proteins were determined using hyper immune rabbit sera. The Sodium Dodecyl SulphatePolyacrylamide Gel Electrophoresis (SDS-PAGE) profile of the antigens revealed different numbers and patterns of protein bands in the range of 11.80 to 176.74 kDa. The electrophoretic pattern of ESA also revealed a number of closely placed indistinct bands in the range of 20-34 kDa.. The proteins in the lower molecular weight range (< 30 kDa) in the case of WCA, CFA, ESA, and the medium molecular weight range (30-60 kDa) in the case of SA and MBA were observed to be immunodominant in the Enzyme-Linked Immunoelectrotransfer Blot (EITB) assay. These immunodominant proteins could be further investigated as potential diagnostic antigens in the serodiagnosis of cysticercosis in pigs, and neurocysticercosis in human patients.

Evidence for Transmission of Taenia solium Taeniasis/Cysticercosis in a Rural Area of Northern Rwanda.

Cysticercosis is a parasitic infection caused by the metacestode larval stage (cysticercus) of Taenia solium. In humans, cysticercosis may infect the central nervous system and cause neurocysticercosis, which is responsible for over 50,000 deaths per year worldwide and is the major cause of preventable epilepsy cases, especially in low-income countries. Cysticercosis infection is endemic in many less developed countries where poor hygiene conditions and free-range pig management favor their transmission. A cross-sectional study was conducted in 680 children from a rural primary school in Gakenke district (Northern province of Rwanda). Stool samples were collected from participants and analyzed using the Kato-Katz method (KK), formol-ether concentration (FEC), and/or copro-antigen enzyme-linked immunosorbent assay (CoAg-ELISA) to detect taeniasis. Blood samples were collected and analyzed using enzyme-linked immunoelectrotransfer blot (EITB) and antigen enzyme-linked immunosorbent assay (Ag-ELISA) to detect human cysticercosis. The overall proportion of taeniasis positivity was 0.3% (2/680), and both cases were also confirmed by CoAg-ELISA. A total of 13.3% (76/572) of the children studied were positive to cysticercosis (T. solium-specific serum antibodies detected by EITB), of whom 38.0% (27/71) had viable cysticercus (T. solium antigens by Ag-ELISA). This study provides evidence of the highest cysticercosis prevalence reported in Rwanda in children to date. Systematic investigations into porcine and human cysticercosis as well as health education and hygiene measures for T. solium control are needed in Gakenke district.

Animal hydatid cyst genotypes as a potential substitute for human hydatid cyst as a source of antigen for diagnosis of zoonotichydatidosis.

The Diagnosis of hydatidosis is still an unsolved issue due to difficulties in obtaining of patient's hydatid cyst appropriate for antigen extraction. This study evaluated the suitability of HC protoscolices somatic antigens (HCPsS-Ag) fractions from animal origin to substitute that extracted from HC of patients in diagnosis of hydatidosis using enzyme-linked immunoelectro-transfer blot and Enzyme-linked immunosorbent assay (ELISA). Eight fractions in HC-G6 from patients react specifically versus HC-G6 infected patient's sera. Five of them (28, 32, 38, 59 and 89 Kilo Dalton (KDa) and two of them (28KDa and 45KDa) reacted versus HC-G1 and HC-G4 infected sheep and equine sera, respectively. Six fractions in HCPsS-Ag-G1 of sheep react versus HC-G1 sheep infected sera, four (28, 32, 52 and 58 KDa) and two of them reacted versus HC-G6 and HC-G4 infected patient and equine sera, respectively. Two fractions only in HCPsS-Ag-G4 of equine reacted versus infected human and sheep sera. This fraction displayed the same degree of ELISA value versus different infected sera with a significantly perfect classification for kappa agreement and non-statistically significant difference (p ≥ 0.05) for ELISA Optical density values of the positive samples without cross-reaction versus other parasites antibodies in sera. HCPsS-Ag from HC genotypes that developed in humans and animals as HC-G6 and HC-G1 can substitute each other for diagnosis of infection than antigens extracted from non-zoonotic HC-G4. The fraction at 28KDa is the only fraction that can be extracted from any animals HC and used in diagnosis of zoonotic hydatidosis.

Further evaluation of recombinant Tsol-p27 by enzyme-linked immunoelectrotransfer blot for the serodiagnosis of cysticercosis in pigs from Mozambique

BackgroundPorcine cysticercosis has a negative impact on human health and the meat industry, as it makes infected meat unaproprieted for consuption and it is the main etiology of epileptic seizures in developing countries. There are multiple serological assays that use crude antigens with high sensitivity and specificity for the diagnosis of both porcine and human cysticercosis. Nonetheless, antigen preparation is time-consuming, needs a well-equipped laboratory and trained personnel and places those manipulating the meat at great risk for infection. New serodiagnostic approaches to the diagnosis of porcine and human cysticercosis have been directed towards the development of recombinant deoxyribonucleic acid technology for the generation of synthetic proteins that can serve as simplified, low-cost and harmless substitutes for native antigens. The aim of the present study was to further evaluate the recombinant Tsol-p27 protein as a target molecule in immunoassays for the serodiagnosis of porcine cysticercosis. From these data, we hoped to develop recommendations regarding its use in the serodiagnosis of porcine cysticercosis.ResultsWe studied a panel of 83 naturally infected pig sera from Angónia District, Mozambique, an endemic area for porcine and human cysticercosis. These sera were previously tested by antigen enzyme-linked immunosorbent assay (Ag-ELISA) to detect antigens of T. solium. The serum panel was processed by enzyme-linked immunoelectrotransfer blot (EITB) assay against the recombinant Tsol-p27 protein and the Ag-ELISA assay results were used to compare and evaluate the performance of Tsol-p27 for the diagnosis of cysticercosis. Out of 83 sera, 24 (29.0%) were positive for Tsol-p27 and 59 (71%) were negative in the same assay. From the 37 sera that tested positive to Ag-ELISA, 11 (13.3%) were positive to Tsol-p27, while from 46 sera that tested negative to Ag-ELISA, 33 (39.7%) also tested negative to Tsol-p27. The sensitivity and specificity of Tsol-p27 was 29.7% and 71.7%, respectively, while the positive predictive value and negative predictive value were 45.8% and 55.9%, respectively, as calculated using Medcalc® version 15.0 software (MedCalc Software, Ostend, Belgium).ConclusionWhile Tsol-p27 recombinant protein might be suitable for testing human sera, its performance in pigs is not acceptable, so other recombinant proteins should be evaluated alone or multiplexed.

Evaluation of cross-reactivity to Taenia hydatigena and Echinococcus granulosus in the enzyme-linked immunoelectrotransfer blot assay for the diagnosis of porcine cysticercosis

BackgroundTaenia solium is an important zoonotic parasite that infects humans as definitive host (taeniasis) and pigs as intermediate host (cysticercosis). Serological diagnosis of porcine cysticercosis is limited to antigen detection using ELISA, which is known to cross-react with other Taenia species, and antibody detection using the lentil-lectin glycoprotein enzyme-linked immunoelectrotransfer blot (LLGP EITB), which has not been adequately evaluated for cross-reactivity to other parasites. Field studies suggest that the GP50 diagnostic band of the LLGP EITB may cross-react to Taenia hydatigena, a common non-zoonotic parasitic infection of pigs. The objective of this study was to evaluate the specificity of the LLGP EITB assay in pigs infected experimentally with T. hydatigena and Echinococcus granulosus.ResultsTwelve three-month-old seronegative were divided into two groups; six were each given an oral challenge with a single gravid proglottid of T. hydatigena and the other six were each given an oral challenge with 50 gravid proglottids of E. granulosus. Serum samples were collected biweekly until 14 weeks when all pigs underwent a detailed necropsy. Taenia hydatigena cysticerci were found in two of six pigs from the first group. Four T. hydatigena-exposed pigs were seropositive at the GP50-band only on EITB LLGP; two of these had cysts at necropsy while no seronegative pigs had cysts. One E. granulosus-exposed pig was positive to EITB LLGP, again with reactivity only to GP50; all six pigs had hepatic echinococcosis on necropsy.ConclusionThese results provide definitive evidence that the GP50 diagnostic band in pigs cross-reacts with T. hydatigena. Evidence of cross-reaction with E. granulosus was not conclusive.

The epidemiology of Taenia spp. infection and Taenia solium cysticerci exposure in humans in the Central Highlands of Vietnam

BackgroundVietnam is endemic for taeniasis and T. solium cysticercosis. Despite this, information on the epidemiological characteristics of the diseases in the Central Highlands of Vietnam are poorly described. The aims of this study were to determine the epidemiological characteristics of taeniasis (Taenia spp.) and T. solium cysticerci exposure in humans in Dak Lak province in the Central Highlands, Vietnam.MethodsThis cross-sectional study was carried out in six villages in three districts of Dak Lak. A total of 190 households were visited. From each household, between one and five individuals were asked to donate a single faecal and blood sample and respond to a questionnaire. Serum samples were subjected to lentil lectin purified glycoprotein enzyme-linked immunoelectrotransfer blot assay to detect antibodies against T. solium cysticerci. Multiplex real-time PCR was used to detect Taenia spp. infection in faecal samples. A fixed-effects logistic regression model was developed to identify factors associated with the probability of Taenia spp. infection or T. solium cysticerci exposure risk. The contribution of each of identified factor was quantified using population attributable fractions.ResultsThe prevalence of seroexposure to T. solium in Dak Lak was 5% (95% CI 3% to 8%). Consumption of raw vegetables, sourcing drinking water from lakes, streams or ponds and the practice of outdoor defaecation were identified as primary risk factors for the prevalence of T. solium cysticerci exposure, while consuming undercooked pork and beef, pork tongue and observing Taenia proglottids in stool were associated with Taenia spp. infection. Consumption of raw vegetables attributed to 74% of T. solium cysticerci exposure-positive cases and consumption of undercooked beef attributed to 77% of taeniasis cases in these communities.ConclusionsThe prevalence of T. solium seroexposure in Dak Lak is consistent with those reported in other regions of Vietnam. The identified risk factors associated with the prevalence of T. solium seroexposure and taeniasis infection in Dak Lak are modifiable and thus advocate for targeted community intervention programs to mitigating these risks.

The epidemiology of porcine Taenia solium cysticercosis in communities of the Central Highlands in Vietnam

BackgroundTaenia solium cysticercosis, recognized as a neglected tropical disease by the WHO, is distributed mostly in developing countries of Latin America, sub-Saharan Africa and Asia. Pigs and humans act as intermediate hosts, acquiring T. solium cysticerci (larval stage) in their tissue, through the ingestion of T. solium eggs shed in the faeces of humans infected with adult tapeworms. The disease has a negative impact on rural economies due to losses in productivity arising from human disease, pork carcass condemnations and loss of market access. The aim of this study was to estimate the prevalence of T. solium cysticercosis in pigs in Dak Lak Province in the Central Highlands of Vietnam and to identify household level characteristics associated with T. solium porcine cysticercosis.MethodsThis was a cross-sectional study of household pigs in three districts of Dak Lak Province. A total of 408 households in six villages in three districts were visited between June and October 2015. A questionnaire was administered to the head of each household, and within each household, serum samples were collected from three pigs. Serum samples were analyzed using the recombinant T24H antigen in enzyme-linked immunoelectrotransfer blot assay and lentil lectin purified glycoprotein in EITB assay. A Bayesian, mixed-effects logistic regression model was developed to identify management factors associated with the probability of a household having at least one cysticercosis-positive pig.ResultsThe prevalence of porcine T. solium cysticercosis in this study was low at 0.94 [95% confidence interval (CI) 0.51–1.68] cases per 100 pigs at risk, in agreement with other studies conducted throughout Vietnam. Scavenging of food and coprophagy were associated with T. solium cysticercosis [odds ratios 1.98 (95% CrI: 0.55–4.74) and 2.57 (95% CrI: 1.22–4.66), respectively].ConclusionsThis study proves that the seroprevalence of porcine cysticercosis in Dak Lak Province was as low as that of other studies conducted throughout Vietnam. Scavenging of food and coprophagy are modifiable factors, providing the opportunity to decrease the prevalence of porcine cysticercosis further in the province.

Evaluating the Recombinant T24H Enzyme-Linked Immunoelectrotransfer Blot Assay for the Diagnosis of Neurocysticercosis in a Panel of Samples from a Large Community-Based Randomized Control Trial in 60 Villages in Burkina Faso.

Current guidelines for the diagnosis of neurocysticercosis (NCC) recommend the use of the lentil lectin-bound glycoprotein enzyme-linked immunoelectrotransfer blot assay (LLGP-EITB) as the reference standard for serological testing. In response to the drawbacks involved with the use of the LLGP-EITB, a recombinant T24H antigen (rT24H) EITB assay was developed, with promising results. However, the test has yet to be evaluated among individuals from sub-Saharan Africa (SSA). The aim of the present study was to investigate the performance of the rT24H EITB assay for the detection of NCC cases in a panel of serum samples (N = 366, of which 173 patients presented with epileptic seizures and/or severe chronic headaches, and 193 matched manifestation-free participants) collected as part of a large community-based trial in Burkina Faso. A perfect agreement between the rT24H EITB and the native gp24 (and its homodimer, gp42) LLGP-EITB was found (kappa value of 1.0). Furthermore, among patients with the neurological manifestations of interest who underwent a computed tomography scan, the rT24H EITB and native antigen LLGP-EITB had a comparable ability to correctly identify NCC cases with multiple viable (rT24H: sensitivity: 80.0%), single viable (66.7%), and calcified/degenerating cysts only (25.0%), albeit for multiple viable and calcified cysts, the rT24H estimated sensitivity seemed lower, but more uncertain, than previously reported. The rT24H EITB specificity was high (98.2%) and in line with previous studies. This study confirms the value of the recombinant rT24H EITB as an alternative to the native antigen LLGP-EITB for the diagnosis of NCC in a SSA community setting.

Antibody Banding Patterns of the Enzyme-Linked Immunoelectrotransfer Blot and Brain Imaging Findings in Patients With Neurocysticercosis.

The enzyme-linked immunoelectrotransfer blot (EITB) assay is the reference serological test for neurocysticercosis (NCC). A positive result on EITB does not always correlate with the presence of active infections in the central nervous system (CNS), and patients with a single viable brain cyst may be EITB negative. Nonetheless, EITB antibody banding patterns appears to be related with the expression of 3 protein families of Taenia solium, and in turn with the characteristics of NCC in the CNS (type, stage, and burden of viable cysts). We evaluated EITB antibody banding patterns and brain imaging findings of 548 NCC cases. Similar banding patterns were grouped into homogeneous classes using latent class analysis. The association between classes and brain imaging findings was assessed. Four classes were identified. Class 1 (patients negative or only positive to the GP50 band, related to the protein family of the same name) was associated with nonviable or single viable parenchymal cysticerci; class 2 (patients positive to bands GP42-39 and GP24, related to the T24-42 protein family, with or without anti-GP50 antibodies) was associated with intraparenchymal viable and nonviable infections; classes 3 and 4 (positive to GP50, GP42-39, and GP24 but also responding to low molecular weight bands GP21, GP18, GP14, and GP13, related to the 8 kDa protein family) were associated with extraparenchymal and intraparenchymal multiple viable cysticerci. EITB antibody banding patterns correlate with brain imaging findings and complement imaging information for the diagnosis of NCC and for staging NCC patients.

Within herd transmission and evaluation of the performance of clinical and serological diagnosis of foot-and-mouth disease in partially immune cattle herds

The control of foot-and-mouth disease (FMD) in vaccinated populations relies upon surveillance activities such as clinical inspections (CI) and serological monitoring. New evidence to refine current surveillance guidelines has been provided by evaluating (1) the diagnostic performance of CI and serological tests for detection of FMD virus (FMDV) non-structural proteins (NSP), and (2) the within-herd transmission of the virus in partially immune cattle. Data came from 23 affected herds during an epidemic of FMDV type O in Bolivia, in 2007. All cattle (n=957) in these herds were clinically inspected and serum samples were collected one month after the last animal with clinical signs was detected. Samples were tested for the presence of antibodies against NSP using the PANAFTOSA 3ABC-ELISA test and a subset of samples were tested using the enzyme-linked immunoelectrotransfer blot assay (EITB). Data from clinical and serological diagnoses were analysed using a Bayesian model. The sensitivity Se and specificity Sp of the tests, as well as the prevalence and the within-herd reproduction ratio R of FMDV were estimated. In addition, risk factors for infection were identified. The Se of CI, the 3ABC-ELISA and the EITB tests were estimated to be 0.30, 0.88 and 0.96 respectively. The estimated Sp, in the same order, were 0.88, 0.93 and 0.97. The within-herd prevalence of infected animals ranged from 0.04 to 0.91 and R ranged from 1.02 to 2.68. It was observed that cattle coming from areas with high vaccination coverage had a lower risk of becoming infected than home-bred cattle from the affected herds, where vaccination coverage was thought to be low. Although these estimates come from herds kept under specific conditions, they provide a reference for future surveillance design and can inform simulation models for surveillance and control of FMD in similar cattle populations.

Recombinant Protein- and Synthetic Peptide-Based Immunoblot Test for Diagnosis of Neurocysticercosis

One of the most well-characterized tests for diagnosing neurocysticercosis (NCC) is the enzyme-linked immunoelectrotransfer blot (EITB) assay developed at the CDC, which uses lentil lectin-bound glycoproteins (LLGP) extracted from Taenia solium cysticerci. Although the test is very reliable, the purification process for the LLGP antigens has been difficult to transfer to other laboratories because of the need for expensive equipment and technical expertise. To develop a simpler assay, we previously purified and cloned the diagnostic glycoproteins in the LLGP fraction. In this study, we evaluated three representative recombinant or synthetic antigens from the LLGP fraction, individually and in different combinations, using an immunoblot assay (recombinant EITB). Using a panel of 249 confirmed NCC-positive and 401 negative blood serum samples, the sensitivity of the recombinant EITB assay was determined to be 99% and the specificity was 99% for diagnosing NCC. We also tested a panel of 239 confirmed NCC-positive serum samples in Lima, Peru, and found similar results. Overall, our data show that the performance characteristics of the recombinant EITB assay are comparable to those of the LLGP-EITB assay. This new recombinant- and synthetic antigen-based assay is sustainable and can be easily transferred to other laboratories in the United States and throughout the world.

Validity of the Enzyme-linked Immunoelectrotransfer Blot (EITB) for naturally acquired porcine cysticercosis

The Enzyme-linked Immunoelectrotransfer Blot (EITB) has been used widely as a screening test for Taenia solium cysticercosis in swine. However, the relation between seropositivity and infection in pig populations from endemic areas has not been well defined. The aim of this study is to relate EITB seropositivity with infection and infection burden, analyse the trade-off between sensitivity and specificity with various cut-off points for the EITB assay, and finally describe the serology changes in a cohort of rural pigs raised under natural conditions. A group of 107 pigs that were used as controls during a vaccination field trial in Peru was our study population. The prevalence of porcine cysticercosis determined by necropsy examination was 16.82% (18/107) in these animals. Using EITB reactivity to ≥1 band as a cut-off point for the assay, the sensitivity was 88.89% (65.29–98.62, 95% CI) and the specificity was 48.31% (37.59–59.16, 95% CI). Comparing other cut-off points, involving up to as many as 7 reactive bands, a reactivity of ≥3 bands provided the best trade-offs in sensitivity and specificity. Using this cut-off point for the assay, the sensitivity was 77.77% (52.36–93.59, 95% CI) and the specificity was 76.40% (66.22–84.76, 95% CI). A significant association was found between cyst counts over 100 cysts and reactivity to ≥3 bands in the EITB assay (Fisher's exact test, p<0.05). The results of this study suggest that the use of the EITB assay to study porcine cysticercosis may require setting different cut-offs under field and experimental conditions, and depending upon the objective of the screening process.

Parasite antigen in serum predicts the presence of viable brain parasites in patients with apparently calcified cysticercosis only.

Computed tomography (CT) remains the standard neuroimaging screening exam for neurocysticercosis, and residual brain calcifications are the commonest finding. Magnetic resonance imaging (MRI) is more sensitive than CT but is rarely available in endemic regions. Enzyme-linked immunoelectrotransfer blot (EITB) assay uses antibody detection for diagnosis confirmation; by contrast, enzyme-linked immunosorbent assay (ELISA) antigen detection (Ag-ELISA) detects circulating parasite antigen. This study evaluated whether these assays predict undetected viable cysts in patients with only calcified lesions on brain CT. Serum samples from 39 patients with calcified neurocysticercosis and no viable parasites on CT were processed by Ag-ELISA and EITB. MRI was performed for each patient within 2 months of serologic testing. Conservatively high ELISA and EITB cutoffs were used to predict the finding of viable brain cysts on MRI. Using receiver operating characteristic-optimized cutoffs, 7 patients were Ag-ELISA positive, and 8 had strong antibody reactions on EITB. MRI showed viable brain cysts in 7 (18.0%) patients. Patients with positive Ag-ELISA were more likely to have viable cysts than Ag-ELISA negatives (6/7 vs 1/32; odds ratio, 186 [95% confidence interval, 1-34 470.0], P < .001; sensitivity 85.7%, specificity 96.9%, positive likelihood ratio of 27 to detect viable cysts). Similar but weaker associations were also found between a strong antibody reaction on EITB and undetected viable brain cysts. Antigen detection, and in a lesser degree strong antibody reactions, can predict viable neurocysticercosis. Serological diagnostic methods could identify viable lesions missed by CT in patients with apparently only calcified cysticercosis and could be considered for diagnosis workup and further therapy.

The Incidence of Human Cysticercosis in a Rural Community of Eastern Zambia

A community-based longitudinal study was performed in the Eastern Province of Zambia, in which repeated serological samplings were done to determine the incidence of human cysticercosis. Three sampling rounds were carried out at six months intervals. A total of 867 participants presented for all three samplings. All samples were tested for the presence of cysticercus antigens using a monoclonal antibody-based enzyme-linked immunosorbent assay (sero-Ag-ELISA), while a randomly selected sub-sample of 161 samples from each sampling round was tested for specific antibodies using a commercial enzyme-linked immunoelectrotransfer blot (EITB) assay. Stool samples (n = 226) were also collected during the final round of sampling for taeniosis diagnosis by coprology and coproantigen ELISA. Cysticercosis seroprevalence varied from 12.2% to 14.5% (sero-Ag) and from 33.5% to 38.5% (sero-Ab) during the study period. A taeniosis prevalence of 11.9% was determined. Incidence rates of 6300 (sero-Ag, per 100000 persons-year) and 23600 (sero-Ab, per 100000 persons-year) were determined. Seroreversion rates of 44% for sero-Ag and 38.7% for sero-Ab were recorded over the whole period. In conclusion, this study has shown the dynamic nature of T. solium infections; many of the people at risk become (re)infected due to the high environmental contamination, with a high number turning seronegative within a year after infection. An important number of infections probably never fully establish, leading to transient antibody responses and short-term antigen presence.

Sero-prevalence of cystic echinococcosis in small ruminants from hypoendemic Northern Italy

Reliable figures of local distribution and prevalence of cystic echinococcosis (CE) in intermediate hosts are a fundamental prerequisite for implementation of control strategies against cystic echinococcosis (CE), caused by Echinococcus granulosus. With the aim to assess the “true” prevalence of CE in a mountain area of Piedmont region (North-Western Italy), two methods alternative to use of official abattoir data were compared: (i) the necroscopic examination of 117 sheep and goats, killed during wolf attacks while on summer pastures, for presence of hydatid cysts; (ii) the serological examination with an enzyme-linked immuno-electro transfer blot assay (EITB) of 1217 sheep from 9 transhumant flocks for presence of anti-Echinococcus antibodies. EITB was first performed on pooled samples, then each serum sample from positive pools was individually tested. Prevalences were 15.4 and 10.8% with method (i) and method (ii), respectively, and they resulted not statistically different (P>0.05). Seroreactors (from 4.4 to 46%) were found in 7 of 9 tested flocks. Adult Echinococcus tapeworms were found in 2 of 4 necropsied dogs from 3 flocks. On the basis of the results, CE prevalence in small ruminants from Piedmont seems to be higher than that reported by official abattoir statistics in the last decade.

Seroprevalence and Risk Factors for Taenia solium Cysticercosis in Rural Pigs of Northern Peru

Taenia solium is a cestode parasite that causes cysticercosis in both humans and pigs. A serological survey was undertaken to assess the seroprevalence and risk factors associated with porcine cysticercosis in the rural district of Morropon, Peru. Pigs aged between 2 and 60 months were assessed by the Enzyme-linked Immunoelectrotransfer blot (EITB) assay to determine their serological status against porcine cysticercosis in a cross-sectional study. A total of 1,153 pigs were sampled. Porcine seroprevalence was 45.19% (42.31–48.06). The information about the animals and households was analyzed and risk factors associated with seroprevalence were determined by a multivariate logistic regression analysis. In the porcine population, the risk of being seropositive increased by 7% with every month of age (OR 1.07, 95% CI 1.05–1.09), and by 148% for pigs living in East Morropon (OR 2.48, 95% CI 1.82–3.37). Whereas, the presence of latrines in a household decreased the risk of being seropositive by 49% (OR 0.51; 95% CI 0.39–0.67). Sex and rearing system did not represent either risk or protective factors associated with the seroprevalence of porcine cysticercosis. The findings of this study could be used for further development of control programs that might focus on similar population groups within rural communities of developing countries where cysticercosis is endemic.

Evaluation of Dot-ELISA and enzyme-linked immuno-electrotransfer blot assays for detection of a urinary hydatid antigen in the diagnosis of cystic echinococcosis

Background:Several serological assays are used for detection of a hydatid antigen in serum for diagnosis of cystic echinococcosis (CE). However, it requires technical expertise and is associated with the risk of acquiring blood-borne infections. Of late, interests have been shifted to other body fluids like urine, saliva, tear drops as alternate specimens in the diagnosis of CE.Aim:The aim of the study was to evaluate the dot enzyme-linked immunosorbent assay (Dot-ELISA) and electro-immunotransfer blot (EITB) for detection of a hydatid antigen in the urine for diagnosis of CE.Materials and Methods:100 ml of urine samples were collected from the patients with confirmed CE (n=30), patients with suspected CE (n=30), patients with other diseases (n=30) and healthy controls (n=30). A hydatid antigen in urine was detected by Dot-ELISA and EITB using only polyclonal antibodies raised against a complete homogenate hydatid (CHH) antigen in rabbits.Results and Conclusions:The Dot-ELISA using polyclonal antibodies showed a sensitivity of 53.33% and specificity of 96.66%, whereas EITB showed a sensitivity of 46.66%. The Dot-ELISA and EITB employing polyclonal antibodies showed no significant difference in sensitivity (P=0.426). Hence, the Dot-ELISA being a simple procedure can be used for detection of a hydatid antigen in urine for diagnosis of CE.