Multiple Stool Sampling and Specific Parasitological Technique are Crucial to Diagnose Strongyloidiasis in Alcoholic Patients.

PURPOSE: Chronic alcoholism is a well-known risk factor for strongyloidiasis, in these patients the disease is potentially more severe, probably due to the breakdown of local protective barriers and immunosuppression caused by alcohol, which can lead to autoinfection and dissemination. The aim of this study was to evaluate multiple stool sampling and a specific parasitological assay agar plate culture (APC) for the diagnosis of Strongyloides stercoralis in alcoholics. METHODS: APC was compared to sedimentation technique (HPJ; Hoffman, Pons and Janer), as parasitological methods to detect S. stercoralis infection in alcoholic individuals. Three stool samples from 60 alcoholic and 60 non-alcoholic individuals were analyzed. RESULTS: S. stercoralis larvae were observed in 11 (18.3%) alcoholic individuals and 1 (1.7%) nonalcoholic individual (P = 0.0042). In view of the combined results, sensitivity for the APC method was 63.6% (CI 31.6-87.6%) with the first sample reaching 100% (CI 67.8-100%) after analyzing three fecal samples. The HPJ sensitivity was 36.4% (CI 12.4-68.4) in the first sample, reaching 72.7% (CI 39.3-92.7) after three samples analyzed. CONCLUSION: The present results suggest that in alcoholic patients, it is important to repeat stool sampling with specific techniques, especially using the APC method, to avoid misdiagnosis in cases that could evolve to disseminated strongyloidiasis.

Seropositivity to Cysticercosis in School-Age Children Living in a Low-Income Municipality in the Midwest Region of Brazil.

Background: Human cysticercosis (CC) is a global public health problem, especially in Latin America, including Brazil. We aimed to analyze the seroprevalence of CC among school-age children and adolescents. Methods: We analyzed the presence of specific IgG antibodies against Taenia solium metacestodes in 500 serum samples from elementary school children and adolescents in Jataí City, state of Goiás, Brazil. IgG antibodies against the antigenic extract of the parasite were detected and analyzed by ELISA, and specific peptides were identified by confirmatory Western Blotting test. Results: Of the 500 study participants, 205 (41%) were male, and 295 (59%) were female. Participants aged between 4 and 18 years (mean age 8.4 years). The percentage of serum samples reactive by ELISA was 37.2%. These samples were analyzed by Western Blotting, which confirmed that the seropositivity rate was 6.2% (95% CI 2.4-14.7) in 31 samples reactive for CC-specific bands, determined in serum samples from 18 male (5-11 years old) and 13 female (4-12 years old) students. Conclusion: The CC seroprevalence demonstrated in schoolchildren suggests that this parasitosis is endemic in the study area. Further investigations are necessary to clarify the local epidemiology of this parasitosis.

Route of dexamethasone administration influences parasite burden in Strongyloides hyperinfection model.

Rodents infected with Strongyloides venezuelensis are experimental models applied to strongyloidiasis research. This study evaluated oral and subcutaneous dexamethasone (DEX) treatments to establish immunosuppression in an experimental model of Strongyloides hyperinfection. Rattus norvegicus Wistar were divided: G I (-): untreated and uninfected animals, G II (+): untreated and infected, G III (o -) orally treated and uninfected, G IV (o +) orally treated and infected, G V (sc -) subcutaneously treated and uninfected, G VI (sc +) subcutaneously treated and infected. For oral administration, DEX was diluted in sterile water (5 µg/ml) and made available to the animals on intervals in experimental days - 5-0, 8-13 and 21-26. For subcutaneous administration, animals received daily injections of DEX disodium phosphate (2 mg/kg). Infection was established by the subcutaneous inoculation of 3000 S. venezuelensis filarioid larvae. Groups were evaluated by egg per gram of feces and parasite females counts and IgG, IgG1 and IgG2a detection. GIV (o +) had egg peaks count on days 13 and 26 and maintained egg elimination until the last experimental day. Parasitic females recovery at day 30 was significantly higher in G IV (o +) when compared to G VI (sc +). Levels of IgG, IgG1 and IgG2a of all groups, except the positive control GII (+), were below the detection threshold. Pharmacological immunosuppression induced by oral administration of DEX produced high parasitic burden, and is a noninvasive method, useful to establish immunosuppression in strongyloidiasis hyperinfection model in rats.

IgG4 and IgE anti-Strongyloides stercoralis as additional parameters in characterizing patients with diabetes from a hyperendemic area.

We report the detection of IgG, IgG1, IgG4 and IgE anti-Strongyloides stercoralis as complementary tool for screening in patients with diabetes in hyperendemic areas for strongyloidiasis. A panel of 119 serum samples were analyzed: 76 from patients with DM2 and 43 patients with other endocrine diseases and a positive correlation for total IgG levels with IgG4 (rs = 0.559; P = 0.024; n = 16) and IgG and IgE (rs = 0.585; P < 0.0001; n = 76) was found in the diabetes group.

Immune complexes as a tool for strongyloidiasis immunodiagnosis in kidney and liver transplant candidate.

Strongyloidiasis is a chronic and asymptomatic infection in immunocompetent patients. Immunocompromised patients, such as organ transplant candidates, can develop severe forms of this disease, and the best way to prevent progression to these forms is early diagnosis. Serological techniques using specific IgG and immune complexes (IC) detection can help in the diagnosis of these patients. This study aimed to detect specific anti-Strongyloides IC and IgG antibodies in kidney transplant (KT) and liver transplant (LT) candidates. A total of 100 blood samples was collected from transplant candidates (50 blood samples each from KT and LT candidates). Serum was obtained and analysed using enzyme-linked immunosorbent assay for IC and IgG detections. The IC levels showed frequencies of 18% and 2% in the KT and LT groups, respectively, whereas anti-Strongyloides IgG was detected in 34% and 12% of KT and LT candidates, respectively. The correlation between IC and IgG detection is poor in KT candidates, while in LT candidates, there is a significant positive correlation. The detection of IC can be an additional tool for the diagnosis of strongyloidiasis, especially when associated with the detection of specific IgG anti-Strongyloides antibodies.

Diagnosis of human strongyloidiasis: Application in clinical practice.

This review considers the advantages and disadvantages of parasitological techniques, methods of detecting antibodies and antigens, as well as molecular biology techniques in the diagnosis of human strongyloidiasis. In addition, it elucidates the potential of different techniques for rapid and effective detection of clinical cases, thus enabling early treatment and preventing fatal consequences of this helminthiasis.

Corrigendum to "scFv against HSP60 of Strongyloides sp. and Its Application in the Evaluation of Parasite Frequency in the Elderly".

[This corrects the article DOI: 10.1155/2020/4086929.].

Strongyloidiasis Serological Analysis with Three Different Biological Probes and Their Electrochemical Responses in a Screen-Printed Gold Electrode.

(1) Background: The validation of biological antigens is the study's utmost goal in biomedical applications. We evaluated three different probes with single and multiple epitopes through electrochemical detection of specific IgG in serum for human strongyloidiasis diagnosis. (2) Methods: Screen-printed gold electrodes were used and probes consisting of two single-epitope synthetic peptides (D3 and C10) with different sequences, and a multi-epitope antigen [detergent phase (DP)-hydrophobic membrane proteins]. Human serum samples from three populations were used: Strongyloides stercoralis positive, positive for other parasitic infections and negative controls. To test the immobilization of probes onto a screen-printed gold electrode and the serum IgG detection, electrochemical analyses were carried out through differential pulse voltammetry (DPV) and the electrode surface analyses were recorded using atomic force microscopy. (3) Results: The electrochemical response in screen-printed gold electrodes of peptides D3 and C10 when using positive serum was significantly higher than that when using the DP. Our sensor improved sensitivity to detect strongyloidiasis. (4) Conclusions: Probes' sequences are critical factors for differential electrochemical responses, and the D3 peptide presented the best electrochemical performance for strongyloidiasis detection, and may efficiently substitute whole antigen extracts from parasites for strongyloidiasis diagnosis in electrochemical immunosensors.

Innovative approaches to improve serodiagnosis of Strongyloides stercoralis infection.

Strongyloidiasis is a helminthiasis of neglected condition that has no gold standard parasitological diagnosis due to the intermittent release of larvae in feces. This study aimed to use an scFv (single chain variable fragment) obtained by Phage Display, previously validated to detect immune complexes in serum samples from individuals infected with Strongyloides stercoralis by enzyme-linked immunosorbent assay (ELISA). Now the ability of scFv to detect the immune complexes was verified by immunofluorescence, flow cytometry using magnetic beads and surface plasmon resonance (SPR). As ELISA, the SPR, immunofluorescence and flow cytometry demonstrated the ability of scFv to detect immune complexes in sera from individuals with strongyloidiasis and discriminate them from sera of individuals with other parasitic diseases and healthy individuals. Besides de conventional ELISA, the novel approaches can also be promptly applied as auxiliary diagnostic tools to the existing parasitological method for accurate diagnosis of human strongyloidiasis.

Beneficial effects of Strongyloides venezuelensis antigen extract in acute experimental toxoplasmosis.

BACKGROUND: Toxoplasma gondii is a protozoan with worldwide distribution and triggers a strong Th1 immune response in infected susceptible hosts. On the contrary, most helminth infections are characterized by Th2 immune response and the use of helminth-derived antigens to regulate immune response in inflammatory disorders has been broadly investigated. OBJECTIVES: The aim of this study was to investigate whether treatment with Strongyloides venezuelensis antigen extract (SvAg) would alter immune response against T gondii. METHODS: C57BL/6 mice were orally infected with T gondii and treated with SvAg, and parasitological, histological and immunological parameters were investigated. RESULTS: It was observed that SvAg treatment improved survival rates of T gondii-infected mice. At day 7 post-infection, the parasite load was lower in the lung and small intestine of infected SvAg-treated mice than untreated infected mice. Remarkably, SvAg-treated mice infected with T gondii presented reduced inflammatory lesions in the small intestine than infected untreated mice and decreased intestinal and systemic levels of IFN-γ, TNF-α and IL-6. In contrast, SvAg treatment increased T gondii-specific IgA serum levels in infected mice. CONCLUSIONS: S venezuelensis antigen extract has anti-parasitic and anti-inflammatory properties during T gondii infection suggesting as a possible alternative to parasite and inflammation control.

Strongyloides-specific IgA, IgG and IgG immune complex profile in patients with pulmonary tuberculosis.

AIMS: To describe an anti-Strongyloides IgA, IgG and IgG immune complex antibody response profile in patients with pulmonary tuberculosis. METHODS AND RESULTS: Saliva and serum samples were collected from 100 individuals: group I, 50 apparently healthy individuals; and group II, 50 pulmonary tuberculosis patients. The IgA, IgG and IgG immune complex detection were carried out via an ELISA immunoenzymatic test. Optical density medians in saliva samples of IgA antibody (median of 7.21) and IgG-IC (median of 4.95) were significantly higher in tuberculosis group compared to control individuals (median IgA of 3.93 and IgG-IC of 2.38). CONCLUSION: This study presents antibody data to the field of pulmonary tuberculosis and strongyloidiasis coinfection, including saliva samples, and especially IgG immune complex detection.

Mimotope-based antigens as potential vaccine candidates in experimental murine cysticercosis.

Human cysticercosis is a public health problem caused by Taenia solium metacestodes; thus, eradication of T. solium transmission by vaccination is an urgent requirement. The Cc48 mimotope from T. solium cysticerci was tested expressed in phage particles (mCc48) and chemically synthesized (sCc48) as a vaccine candidate in experimental murine cysticercosis. For this, BALB/c mice were immunized with mCc48 (G1; n = 40), sCc48 (G2; n = 40) and phosphate-buffered saline (PBS) (G3; n = 40, positive control) and challenged with Taenia crassiceps metacestodes. Another PBS group without parasite challenge was used as a negative control (G4; n = 40). Mice were sacrificed 15, 30, 45 and 60 days post-infection for cysticerci and serum collection. Immunization efficacy was determined by cysticerci counting. Serum samples were tested by ELISA to verify antibody (IgM, IgG, IgA and IgE) and cytokine (IFNγ and IL-4) levels. The sCc48 achieved the highest rates of protection and efficacy (90 and 98%, respectively). The group immunized with mCc48 presented the highest reactivity for IgM, IgG and IgE. All groups presented IL-4, but IFNγ was quite variable among groups. The protection induced by sCc48 synthetic peptide supports further studies of this mimotope as a potential vaccine candidate against cysticercosis.

Egg yolk immunoglobulin Y as a promising tool to detect immune complexes in neurocysticercosis serum samples.

BACKGROUND: Neurocysticercosis (NCC) is a neglected tropical disease and its diagnosis is still a challenge due to non-specific manifestations. Neuroimaging techniques are used in the diagnosis of NCC, however, due to the high cost of these methods and the advantages presented in the use of immunological tests, such as ease of performance and satisfactory results, immunoassays are commonly used to detect antibodies against Taenia sp. antigens. The aim of the present study was to produce, characterize and apply specific polyclonal immunoglobulin Y (IgY) anti-Taenia crassiceps extracted from egg yolk of hens immunized with T. crassiceps metacestodes. METHODS: Indirect enzyme-linked immunosorbent assay (ELISA), avidity ELISA, immunoblotting and indirect immunofluorescence tests were performed for characterization of IgY antibodies. Diagnostic performance was verified by ELISA for immune complex detection testing 90 serum samples. RESULTS: Values of sensitivity, specificity, positive and negative likelihood ratios (LR+/LR-) and area under the curve (AUC) were calculated and presented the following results: sensitivity 83.3%, specificity 96.7%, AUC 0.966, LR+ 25.0 and LR- 0.17. CONCLUSIONS: Results of this pioneering and innovative study demonstrate that anti-T. crassiceps IgY antibodies present potential applicability and can be used as an efficient tool in human NCC serodiagnosis.

Immunoreactivity of proteins within 30-40 kDa range during the acute and the recovery phases in rats experimentally infected with Strongyloides venezuelensis.

In experimental infection with Strongyloides venezuelensis, the acute and recovery phases can be distinguished, unlike human infections caused by Strongyloides stercoralis. The objective of this study was to evaluate the production of anti-Strongyloides IgG antibodies and the recognition of immunogenic protein bands during the acute and the recovery phases in rats experimentally infected with S. venezuelensis. Rats were infected subcutaneously with 400 or 4,000 S. venezuelensis infective larvae. The acute phase was characterized by elimination of a large number of eggs in the faeces on days 6-14 post infection; the recovery phase was characterized by the resolution of the infection between days 30 and 35 post infection. Differences in IgG levels were observed in the acute and the recovery phases. Different antigenic fractions were recognized in both phases of infection. It is concluded that proteins within the 30-40 kDa range are immunoreactive markers for both the acute and the recovery phases in rats experimentally infected with S. venezuelensis, particularly using membrane antigen.

scFv against HSP60 of Strongyloides sp. and Its Application in the Evaluation of Parasite Frequency in the Elderly.

The present study is aimed at evaluating serological method using scFv anti-Strongyloides sp. and reporting the frequencies of the results with conventional parasitological technique (faeces) in elderly individuals. Among 112 elderly individuals (≥60 years of age), 14.28% were positive for at least one enteroparasite, with one individual positive for S. stercoralis. Sera were evaluated for the presence of anti-Strongyloides sp. antibodies using total or detergent fraction extracts of Strongyloides venezuelensis, which presented positivity rates of 19.64% and 10.71%, respectively. An anti-HSP60 single-chain variable fragment from Strongyloides sp. was used to detect parasite antigens, with 5.36% (6 individuals) of ELISA-positive individuals returning a positive result. While the serological test indicates previous or recent infection and may be limited by antigen purification, the anti-HSP60 method reflects the presence of Strongyloides sp. immune complexes and exhibits greater sensitivity and specificity. Our results demonstrate the variable occurrence of enteroparasites in elderly individuals residing in long-term nursing homes and validate a novel epidemiological tool to describe infection cases by Strongyloides sp.

Use of polyclonal IgY antibodies to detect serum immune complexes in patients with active hookworm infection.

Definitive diagnosis of hookworm infection is usually based on the microscopic detection of eggs in a stool sample; however, several cases display a low or irregular egg output. Serodiagnosis can be a useful tool to identify these cases, but conventional tests do not differentiate past from active infections. The aim of this study was to obtain and apply egg yolk polyclonal immunoglobulin (IgY) antibodies to detect immune complexes (ICs) in serum samples from patients infected with hookworm. Hens were immunized with Ancylostoma ceylanicum saline extract, their eggs were collected and then IgY antibodies were extracted and purified. Antibody purity was tested by 12% sodium dodecyl sulphate polyacrylamide gel electrophoresis and specificity was assessed by immunoblotting and immunofluorescence. IgY production was evaluated by kinetics enzyme-linked immunosorbent assay (ELISA). Sandwich ELISA tested the ability of IgY to detect ICs in serum samples, from which diagnostic parameters were calculated. Antibody responses increased steadily from day 7 to 42. In the immunoblotting assay, IgY recognized two protein complexes. The immunofluorescence assay showed no staining in control samples. The sandwich ELISA presented a very high diagnostic value, with a sensitivity of 90% and a specificity of 86.7%. Our pioneer strategy highlights the potential use of egg yolk IgY as a diagnostic test to detect active hookworm infection.

Parasitological and immunological aspects of oral and subcutaneous prednisolone treatment in rats experimentally infected with Strongyloides venezuelensis.

Strongyloides venezuelensis is a model to study human strongyloidiasis, which infects wild rodents and shares common antigenic epitopes with Strongyloides stercoralis. This study aimed to evaluate parasitological and immunological parameters of prednisolone immunosuppression protocols in rats (Rattus novergicus) infected with S. venezuelensis. Rats were divided into six groups (n = 36): untreated and uninfected (-) or infected (+); oral treatment and uninfected (o-) or infected (o+); subcutaneous treatment and uninfected (sc-) or infected (sc+). For oral immunosuppression, 5 mg/mL of water diluted prednisolone were given five days before infection, and in the days 8 and 21 (for 5 days). For subcutaneous immunosuppression, 10 mg/kg of prednisolone were given daily. The infection was established by the subcutaneous injection of approximately 3,000 S. venezuelensis filarioid larvae per animal. All animals from the (+) and (o+) groups survived, while four rats from the (sc+) died prior to necropsy date. Parasitological analysis showed higher egg elimination in (o+) in comparison to (+) and (sc+) on 7, 13 and 26 days post infection (d.p.i.).The recovery of parasitic females at day 30 was significantly higher in (o+), compared to (+). The (+) and (o+) groups showed a clear increase in anti-S. venezuelensis IgG, IgG1 and IgG2 from 13th d.p.i. Oral immunosuppression led to a higher number of adult females and increased egg output while maintaining IgG and subclasses antibody levels comparable to the positive control.

Shotgun proteomics of Strongyloides venezuelensis infective third stage larvae: Insights into host-parasite interaction and novel targets for diagnostics.

Strongyloides venezuelensis is an important alternative source of antigen for the serologic diagnosis of human strongyloidiasis. Proteomics techniques applied to the analysis of the protein content of infective third stage larvae (iL3) of S. venezuelensis provide a powerful tool for the discovery of new candidates for immunodiagnosis. This study presents an overview of the protein iL3 S. venezuelensis focusing on the diagnosis of strongyloidiasis. A total of 877 proteins were identified by shotgun proteomics. Many of these proteins are involved in different cellular processes, metabolic as well as structural maintenance. Our results point to a catalog of possible diagnostic targets for human strongyloidiasis and highlight the need for evaluation of uncharacterized proteins, especially the proteins within the CAP domain, transthyretin, and BTPI inhibitor domains, as a repertoire as yet unexplored in the context of strongyloidiasis diagnostic markers. We believe that the protein profile presented in this shotgun analysis extends our understanding of the protein composition within the Strongyloides genus, opening up new perspectives for research on biomarkers that may help with the diagnosis of human strongyloidiasis. Data are available via ProteomeXchange with identifier PXD013703.

Evaluation of the Dot-ELISA as a diagnostic test for human strongyloidiasis based on the detection of IgA in saliva.

This study aimed to evaluate the use of saliva samples in the Dot-ELISA test for immunodiagnosis of human strongyloidiasis. The Dot-ELISA presented similar results to the ELISA test, with 70% and 60% sensitivity and 85% and 90% specificity, respectively, for IgA in the saliva. The Dot-ELISA with alternative saliva samples may be a suitable tool for diagnosing human strongyloidiasis, especially in populations with high levels of exposure to helminth.