Design and expression of a chimeric recombinant antigen (SsIR-Ss1a) for the serodiagnosis of human strongyloidiasis: Evaluation of performance, sensitivity, and specificity.

BACKGROUND: The sensitivity of parasitological and molecular methods is unsatisfactory for the diagnosis of strongyloidiasis, and serological techniques are remaining as the most effective diagnostic approach. The present study aimed to design and produce a chimeric recombinant antigen from Strongyloides stercoralis immunoreactive antigen (SsIR) and Ss1a antigens, using immune-informatics approaches, and evaluated its diagnostic performance in an ELISA system for the diagnosis of human strongyloidiasis. METHODOLOGY/PRINCIPAL FINDINGS: The coding sequences for SsIR and Ss1a were selected from GenBank and were gene-optimized. Using bioinformatics analysis, the regions with the highest antigenicity that did not overlap with other parasite antigens were selected. The chimeric recombinant antigen SsIR- Ss1a, was constructed. The solubility and physicochemical properties of the designed construct were analyzed and its tertiary structures were built and evaluated. The construct was expressed into the pET-23a (+) expression vector and the optimized DNA sequences of SsIR-Ss1a (873 bp) were cloned into competent E. coli DH5α cells. Diagnostic performances of the produced recombinant antigen, along with a commercial kit were evaluated in an indirect ELISA system, using a panel of sera from strongyloidiasis patients and controls. The physicochemical and bioinformatics evaluations revealed that the designed chimeric construct is soluble, has a molecular with of 35 KDa, and is antigenic. Western blotting confirmed the immunoreactivity of the produced chimeric recombinant antigen with the sera of strongyloidiasis patients. The sensitivity and specificity of the indirect ELISA system, using the produced SsIR-Ss1a chimeric antigen, were found to be 93.94% (95% CI, 0.803 to 0.989) and 97.22% (95% CI, 0.921 to 0.992) respectively. CONCLUSIONS/SIGNIFICANCE: The preliminary findings of this study suggest that the produced SsIR-Ss1a chimeric antigen shows promise in the diagnosis of human strongyloidiasis. However, these results are based on a limited panel of samples, and further research with a larger sample size is necessary to confirm its accuracy. The construct has potential as an antigen in the ELISA system for the serological diagnosis of this neglected parasitic infection, but additional validation is required.

Detection of Anti-Strongyloides Antibodies in the Serum of Liver Transplant Recipients: Need of Screening for This Neglected Helminthiasis.

Immunosuppressed patients, particularly transplant recipients, can develop severe strongyloidiasis. This study aimed to detect anti-Strongyloides IgG antibodies in a panel of sera from liver transplant patients. Two techniques were used: ELISA as the initial screening test and Western blotting as a confirmatory test. ELISA reactivity of 10.9% (32/294) was observed. The 40-30 kDa fraction was recognised in 93.7% (30/32) of the patients, resulting in a positivity rate of 10.2%. These data highlight the importance of serological screening for Strongyloides stercoralis infection in liver transplant recipients.

Serological diagnosis of strongyloidiasis: An evaluation of three commercial assays.

BACKGROUND: Strongyloidiasis is caused by a neglected nematode, manifesting as chronic intestinal infection with potentially severe manifestations. The disease is an emerging problem in non-endemic countries affecting travelers and migrants. Diagnosis of strongyloidiasis is hampered by the lack of standardization and absence of a gold standard. Since adequate direct methods to detect the motile larvae in stool samples are not widely available, other techniques such as serology have been developed. METHODS: We evaluated three commercial ELISA kits (DRG Instruments, IVD Research, and Bordier Affinity Products) to detect IgG antibodies against Strongyloides stercoralis assays utilizing serum samples from travelers with microscopically confirmed strongyloidiasis (n = 50) and other imported helminthic infections (n = 159) as well as healthy controls (n = 50). RESULTS: The DRG, IVD, and Bordier assays showed sensitivities of 58.0%, 64.0%, and 56.0%, respectively. Specificity values were 96.0%, 96.0%, and 92.0% in healthy controls, and 67.3%, 62.9%, and 76.7% in cases with other helminth infections, respectively. Cross-reactions were mostly observed in cases with other nematodes (37.5%, 42.5%, and 20.0%, respectively), but also in trematode (33.3%, 38.1%, and 19.0%, respectively) and in cestode infections (25.0%, 30.0%, and 32.5%, respectively). CONCLUSION: The study demonstrates the diagnostic limitations of serological assays to detect or exclude cases of strongyloidiasis in returning travelers, who frequently present with recent or acute infections.

Evaluation of Somatic Antigens of Adult Toxocara helminthes for Detection of Human Toxocariasis.

We aimed to develop an indirect enzyme-linked immunosorbent assay (ELISA) to evaluate the presence of specific IgG against Toxocara canis and Toxocara cati somatic antigens on the serum of patients with toxocariasis. The sensitivity, specificity, positive and negative predictive values for indirect-ELISA were calculated by receiver operating characteristic curve (ROC) analysis and Youden's J using Likelihood ratio. All statistics were analysed and graphs are plotted using GraphPad Prism version 8.4.3 (Graph Pad Software, La Jolla, CA, USA), with 95% confidence interval (CI). The sensitivity, specificity, positive and negative predictive values for T. canis were 100%, 82%, 79% and 100%, respectively. The mentioned variables for T. cati were 97%, 82%, 78% and 98%, respectively. Five immune reactive bands of 38, 40, 72, 100 and 250 kDa were common in both species. Toxocara crude antigens were highly immunogenic in human sera. Immunoreactive bands against T. canis compared to T. cati somatic antigen were about two times more. Unlike Toxocara excretory-secretory antigen, that was homologue in two species, somatic antigens of T. canis and T. cati showed different immunoreactive bands in our western blot.

A silent threat behind bars: Serosurvey and associated risk factors for strongyloidiasis in female inmates and correctional officers.

Strongyloidiasis has been a neglected parasitic infection caused by Strongyloides genus parasites. Despite assessment of S. stercoralis exposure in different vulnerable populations, seroprevalence in inmates worldwide remains to be fully established. Due to poor sanitation and lack of personal hygienic practices, incarcerated individuals have been considered prone to spread infectious illnesses. Accordingly, the present study has assessed exposure and associated risk factors for strongyloidiasis in women inmates and correctional officers at the Women's State Penitentiary of Parana, part of the third largest incarceration complex in Brazil at the time. Blood samplings were performed in 2020 and 2021from a total of 503 women inmates and 92 correctional officers. Participants voluntarily responded to an epidemiological questionnaire to assess associated risk factors to strongyloidiasis. Serological analysis was performed by ELISA for anti-S. stercoralis IgG detection. Statistical analysis was performed using R software, adopting a 5% level of significance. The data were submitted to univariate analysis by chi-square or Fisher´s Exact test for assessing the association among seropositivity and the variables. The variables with p-value < 0.2 in the univariate analysis were considered fit to be included in the logistic regression. In overall, 356/503 (70.8%; 95% CI: 66.7-74.6) inmates were seropositive for anti-S. stercoralis antibodies, with no statistically associated risk factor to seropositivity. A total of 57/92 (62.0%; 95% CI: 51.8-71.2) correctional officers were seropositive, and logistic regression revealed that individuals older than 50 years were more likely seropositive. In conclusion, the high endemicity observed herein has indicated a history of previous exposure to S. stercoralis and warned for a systematic strongyloidiasis screening for inmates, to prevent long term morbidity and disseminated infection during incarceration.

Atypical Toxocara canis-Induced Hepatic Visceral Larva Migrans: Diagnostic Challenges and Literature Review.

Toxocariasis, a zoonotic infection transmitted by Toxocara canis (from dogs) and Toxocara cati (from cats) larvae, poses rare but severe risks to humans. We present a case of hepatic visceral larva migrans (VLM) caused by Toxocara canis in a 21-year-old male with a history of close contact with a pet dog. Initial symptoms and imaging findings mimicked a pyogenic liver abscess. The initial laboratory investigations revealed neutrophilia and elevated levels of IgE. Despite broad-spectrum antibiotics, persistent fever prompted further investigation. Subsequent serological testing for Toxocara antibodies and histopathological analysis of liver tissue demonstrating eosinophil infiltrates and Charcot-Leyden crystals led to a confirmed diagnosis of a liver abscess caused by Toxocara canis. Serological testing for Toxocara antibodies and histopathological analysis of liver tissue confirmed a Toxocara canis-induced liver abscess. Albendazole treatment yielded significant clinical improvement. This case highlights the necessity of considering toxocariasis in liver abscess differentials, particularly in high-seroprevalence regions like Vietnam. Relying solely on serological tests may be insufficient, emphasizing the need for corroborative evidence, including invasive procedures like liver biopsy, for accurate hepatic toxocariasis diagnosis.

Molecular insights and antibody response to Dr20/22 in dogs naturally infected with Dirofilaria repens.

Subcutaneous dirofilariasis, caused by the parasitic nematode Dirofilaria repens, is a growing concern in Europe, affecting both dogs and humans. This study focused on D. repens Dr20/22, a protein encoded by an alt (abundant larval transcript) gene family. While well-documented in L3 larvae of other filariae species, this gene family had not been explored in dirofilariasis. The research involved cloning Dr20/22 cDNA, molecular characterization, and evaluating its potential application in the diagnosis of dirofilariasis. Although Real-Time analysis revealed mRNA expression in both adult worms and microfilariae, the native protein remained undetected in lysates from both developmental stages. This suggests the protein's specificity for L3 larvae and may be related to a process called SLTS (spliced leader trans-splicing), contributing to stage-specific gene expression. The specificity of the antigen for invasive larvae positions it as a promising early marker for dirofilariasis. However, ELISA tests using sera from infected and uninfected dogs indicated limited diagnostic utility. While further research is required, our findings contribute to a deeper understanding of the molecular and immunological aspects of host-parasite interactions and could offer insights into the parasite's strategies for evading the immune system.

Divergent humoral responses between males and females against 24 kDa excretory-secretory protein of Haemonchus contortus and influence of ovine ß-globin polymorphism.

Lambs harboring the Hb-AA ß-globin haplotype present improved cell-mediated responses and increased resistance against Haemonchus contortus infection. The aim of the present study was to compare the effect of sex and ß-globin haplotypes on specific humoral responses and phenotypes of resistance during H. contortus infection in Morada Nova sheep. As expected, females displayed stronger resistance during the first and second experimental challenges. Differential systemic humoral immune responses were observed comparing sex groups, in which higher levels of specific antibodies targeting 24 kDa excretory-secretory (ES24) protein of H. contortus of IgG and IgM antibodies were respectively observed as predominant isotypes in males and females. The IgM levels were significantly correlated with phenotypes of resistance, evaluated by packed cell volume and fecal egg counts. To our knowledge this is the first study reporting divergent humoral responses profiles to H. contortus infection between male and female sheep. The impact of ß-globin haplotypes was less pronounced in females compared to males. Notably, only males showed significant weight differences across haplotypes, with Hb-AA lambs being the heaviest. Additionally, Hb-AA males had significantly higher PCV (indicating better red blood cell health) and lower FEC (indicating lower parasite burden). These findings suggest a more pronounced effect of ß-globin polymorphisms on H. contortus infection in males, potentially due to their generally weaker resistance compared to females. This study highlights the importance of sex and ß-globin haplotypes in shaping immune responses to H. contortus infection. Specifically, IgM antibodies targeting the ES24 protein appear to play a crucial role in host-parasite interactions and may hold promise for therapeutic development.

Evaluation of copromicroscopy, multiplex-qPCR and antibody serology for monitoring of human ascariasis in endemic settings.

BACKGROUND: The standard diagnosis of Ascaris lumbricoides and other soil-transmitted helminth (STH) infections relies on the detection of worm eggs by copromicroscopy. However, this method is dependent on worm patency and shows only limited accuracy in low-intensity infection settings. We aimed to decipher the diagnostic accuracy of different antibodies using various Ascaris antigens in reference to copromicroscopy and quantitative PCR (qPCR), four months after national STH preventative chemotherapy among school children in western Kenya. METHODOLOGY: STH infection status of 390 school children was evaluated via copromicroscopy (Kato-Katz and mini-FLOTAC) and qPCR. In parallel, Ascaris-specific antibody profiles against larval and adult worm lysates, and adult worm excretory-secretory (ES) products were determined by enzyme-linked immunosorbent assay. Antibody cross-reactivity was evaluated using the closely related zoonotic roundworm species Toxocara cati and Toxocara canis. The diagnostic accuracy of each antibody was evaluated using receiver operating curve analysis and the correspondent area under the curve (AUC). PRINCIPAL FINDINGS: Ascaris was the predominant helminth infection with an overall prevalence of 14.9% (58/390). The sensitivity of mini-FLOTAC and Kato-Katz for Ascaris diagnosis reached only 53.5% and 63.8%, respectively compared to qPCR. Although being more sensitive, qPCR values correlated with microscopic egg counts (R = -0.71, P<0.001), in contrast to antibody levels. Strikingly, IgG antibodies recognizing the ES products of adult Ascaris worms reliably diagnosed active Ascaris infection as determined by qPCR and microscopy, with IgG1 displaying the highest accuracy (AUC = 0.83, 95% CI: 0.75-0.91). CONCLUSION: IgG1 antibody responses against adult Ascaris-ES products hold a promising potential for complementing the standard fecal and molecular techniques employed for monitoring Ascaris infections. This is of particular importance in the context of deworming programs as the antibody diagnostic accuracy was independent of egg counts.

High prevalence of anti-Strongyloides antibody in SARS-CoV-2-infected human sera in a Thai hospital: Rapid serological screening.

COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), can stimulate a systemic inflammatory response with severe lung involvement, multisystem dysfunction, and death in some cases. Immunosuppressive treatments have been proposed for management of COVID-19 patients, but these bring the risk of flare-up of pre-existing infections. Strongyloidiasis can become severe or fatal in immunocompromised individuals. This cross-sectional study determined the prevalence of anti-Strongyloides IgG antibody in sera collected from SARS-CoV-2 infected persons in a tertiary-care Thai hospital from January 2021 to January 2022. The survey was conducted using a rapid immunochromatographic test (ICT) kit based on a recombinant antigen of Strongyloides stercoralis known to be IgG-immunoreactive. High prevalence of anti-Strongyloides IgG antibody was found. Out of 297 SARS-CoV-2-infected patients 117 (39.4 %, 95 % CI 33.8-45.2 %) were positive for S. stercoralis according to the ICT kit. In areas where strongyloidiasis is endemic, we suggest using this point-of-care ICT kit for routine rapid screening in seriously ill COVID-19 patients who will be subjected to immunosuppressive treatment. Prompt anthelminthic treatment should be administered to prevent serious systemic strongyloidiasis in at-risk patients.

Immunochromatography Lateral Flow Strip Enhancement Based on Passive Gold Nanoparticles Conjugation to Detect Schistosma haematobium Antigens in Human Serum.

PURPOSE: This study aimed to develop and evaluate a lateral flow card for the detection of active Schistosoma haematobium infection. METHODS: In order to prepare the immunochromatography lateral flow strip (ICLFS), antibodies purified from schistosomiasis were conjugated passively with gold nanoparticles using a potassium carbonate buffer. RESULTS: The novel ICLFS was able to correctly identify 64 out of 67 samples of schistosomiasis, 6 out of 90 samples of other parasites, and 0 out of 27 control samples. Sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV) were 95.5%, 93.3%, 90%, and 91.4% respectively. Comparatively, the sensitivity, specificity, NPV, and PPV of sandwich enzyme-linked immunosorbent assays (ELISA) conjugated with gold nanoparticles (AuNPs) were 91.1%, 88.8%, 85.9%, and 84.4% respectively. The increased sensitivity and specificity of ICLFS produced superior results to those of sandwich ELISA. CONCLUSION: In conclusion, ICLFS is more beneficial and precise than sandwich ELISA for detection of S. haematobium infection at early stage.

Characterization of antigenic proteins of the Taenia solium postoncospheral form.

Neurocysticercosis is the leading cause for acquired epilepsy worldwide, and it is caused by the larval stage of the parasite Taenia solium. Several proteins of this stage have been characterized and studied to understand the parasite-host interaction, however, the proteins from the early cysticercus stages (the postoncospheral form) have not yet been characterized. The study of the postoncospheral form proteins is important to understand the host-parasite relationship in the early stages of infection. The aim of this work was to identify postoncospheral form antigenic proteins using sera from neurocysticercosis patients. T. solium activated oncospheres were cultured in HCT-8 cells to obtain the postoncospheral form. Soluble total and excretory/secretory proteins were obtained from the postoncospheral form and were incubated with both pool sera and individual serum of neurocysticercosis positive human patients. Immunoblotting showed target antigenic proteins with apparent molecular weights of 23 kDa and 46-48 kDa. The 46-48 kDa antigen bands present in soluble total and excretory/secretory postoncospheral form proteins were analyzed by LC-MS/MS; proteins identified were: nuclear elongation factor 1 alpha, enolase, unnamed protein product/antigen diagnostic GP50, calcium binding protein calreticulin precursor and annexin. The postoncospheral form expresses proteins related to interaction with the host, some of these proteins are predicted to be exosomal proteins. In conclusion, postoncospheral proteins are consistent targets of the humoral immune response in human and may serve as targets for diagnosis and vaccines.

Estimation of diagnostic sensitivity and specificity of abattoir registrations and bulk tank milk ELISA as herd-level tests for Fasciola hepatica using Bayesian latent class modelling.

The common liver fluke, Fasciola hepatica, is a trematode parasite found worldwide, typically with a focal distribution due to its requirement for suitable climatic and environmental conditions to complete its lifecycle. Bovine fasciolosis causes suboptimal production and economic losses, including liver condemnation at slaughter. The lack of reliable diagnostic methods is a disadvantage to the increasing demand for surveillance and control. The aim of this study was to evaluate the diagnostic accuracy of bulk tank milk (BTM) antibody testing and aggregated abattoir registrations (AAR) of liver fluke as herd-level tests for F. hepatica infection using Bayesian latent class models. Data from the abattoirs in 2019-2021 and BTM, sampled in the winter of 2020/2021, were collected from 437 herds on the southwest coast of Norway. The BTM samples were analysed with the SVANOVIR® F. hepatica-Ab ELISA test, with results given as an optical density ratio (ODR), and later dichotomized using the recommended cut-off value from the test manufacturer (ODR ≥0.3). Based on the BTM ELISA test, 47.8% of the herds tested positive. The AAR test was defined as the herd-level proportion of female slaughtered animals registered with liver fluke infection during the study period. For this test, three cut-offs were used (a proportion of 0.05, 0.1 and 0.2). The herds were split into two subpopulations ("Coastal" and "Inland"), which were expected to differ in true prevalence of F. hepatica infection based on climate-related and geographical factors. The diagnostic accuracies of both tests were estimated using Bayesian latent class models with minimally informative priors. Post-hoc analysis revealed that the maximum sum of sensitivity (Se) and specificity (Sp) of the tests was achieved with a herd-level proportion of ≥0.1 registered with liver fluke as the AAR test. Using this cut-off, the median estimate for the diagnostic accuracy of the BTM ELISA was 90.4% (84.0-96.2 95% Posterior Credible Interval (PCI)) for Se and 95.3% (90.6-100% PCI) for Sp, while the median estimate of Se for AAR was 87.5% (81.4-93.1% PCI) and the median estimate of Sp for AAR was 91.0% (85.2-96.5% PCI). The cut-off evaluation of the SVANOVIR® F. hepatica-Ab ELISA test for BTM confirmed the manufacturer's recommended cut-off of ODR ≥0.3 to denote positive and negative herds. This study suggests that AAR and BTM ELISA test can be used as herd-level tools to monitor liver fluke infection, so that appropriate interventions against infection can be implemented as necessary.

Development and evaluation of time-resolved rapid immunofluorescence test for detection of TSOL18 specific antibody in porcine cysticercosis infections.

BACKGROUND: Porcine cysticercosis, a serious zoonotic parasitic disease, is caused by the larvae of Taenia solium and has been acknowledged by the World Organization for Animal Health. The current detection methods of Cysticercus cellulosae cannot meet the needs of large-scale and rapid detection in the field. We hypothesized that the immunofluorescence chromatography test strip (ICS) for detecting Cysticercus cellulosae, according to optimization of a series of reaction systems was conducted, and sensitivity, specificity, and stability testing, and was finally compared with ELISA. This method utilizes Eu3+-labeled time-resolved fluorescent microspheres (TRFM) coupled with TSOL18 antigen to detect TSOL18 antibodies in infected pig sera. RESULTS: ICS and autopsy have highly consistent diagnostic results (n = 133), as determined by Cohen's κ analysis (κ = 0.925). And the results showed that the proposed ICS are high sensitivity (0.9459) with specificity (0.9792). The ICS was unable to detect positive samples of other parasites. It can be stored for at least six months at 4℃. CONCLUSIONS: In summary, we established a TRFM-ICS method with higher sensitivity and specificity than indirect ELISA. Results obtained from serum samples can be read within 10 min, indicating a rapid, user-friendly test suitable for large-scale field detection.

Serology survey of Ascaris suum and Trichinella spiralis in rural pigs in Southwestern Mexico.

BACKGROUND: Parasitic diseases of pigs are a public and veterinary health problem. Helminths influence pork production, whereas backyard pigs can transmit these parasites. OBJECTIVES: This work aimed to investigate the prevalence of antibodies against Ascaris suum and Trichinella spiralis in backyard pigs from Jamiltepec, Region de la Costa, Oaxaca, in Southwestern Mexico. METHODS: Six hundred sixty-four serum samples were obtained from backyard pigs from 23 rural villages distributed in 5 municipalities; samples were taken in a non-probabilistic manner with the owner's consent. The presence of serum antibodies against a total extract of A. suum adult worm was determined by ELISA. In contrast, antibodies to the excretion-secretion products of the T. spiralis muscle larva were determined by Western blot. RESULTS: The global seroprevalence for A. suum was 5.12% and 2.41% for T. spiralis; however, antibodies were only found in 8 villages and distributed in 3 municipalities. The highest frequency of positivity for Ascaris was found in the municipality of Santa Catarina Mechoacán (13.01%), whereas, in Santa María Huazalotitlán, the highest frequency of positivity for Trichinella was found (5.75%). In San Andrés, frequencies were 7.23% and 4.82%, respectively. No statistical differences were observed between populations. CONCLUSIONS: Our data suggest that helminth transmission is restricted by locality. However, further studies must be conducted to understand the factors limiting this transmission to promote pork meat production in parasite-free zones.

Multi-epitope protein production and its application in the diagnosis of opisthorchiasis.

BACKGROUND: Opisthorchiasis and cholangiocarcinoma (CCA) continue to be public health concerns in many Southeast Asian countries. Although the prevalence of opisthorchiasis is declining, reported cases tend to have a light-intensity infection. Therefore, early detection by using sensitive methods is necessary. Several sensitive methods have been developed to detect opisthorchiasis. The immunological detection of antigenic proteins has been proposed as a sensitive method for examining opisthorchiasis. METHODS: The Opisthorchis viverrini antigenic proteins, including cathepsin B (OvCB), asparaginyl endopeptidase (OvAEP), and cathepsin F (OvCF), were used to construct multi-antigenic proteins. The protein sequences of OvCB, OvAEP, and OvCF, with a high probability of B cell epitopes, were selected using BepiPred 1.0 and the IEDB Analysis Resource. These protein fragments were combined to form OvCB_OvAEP_OvCF recombinant DNA, which was then used to produce a recombinant protein in Escherichia coli strain BL21(DE3). The potency of the recombinant protein as a diagnostic target for opisthorchiasis was assessed using immunoblotting and compared with that of the gold standard method, the modified formalin-ether concentration technique. RESULTS: The recombinant OvCB_OvAEP_OvCF protein showed strong reactivity with total immunoglobulin G (IgG) antibodies against light-intensity O. viverrini infections in the endemic areas. Consequently, a high sensitivity (100%) for diagnosing opisthorchiasis was reported. However, cross-reactivity with sera from other helminth and protozoan infections (including taeniasis, strongyloidiasis, giardiasis, E. coli infection, enterobiasis, and mixed infection of Echinostome spp. and Taenia spp.) and no reactivity with sera from patients with non-parasitic infections led to a reduced specificity of 78.4%. In addition, the false negative rate (FNR), false positive rate (FPR), positive predictive value (PPV), negative predictive value (NPV), and diagnostic accuracy were 0%, 21.6%, 81.4%, 100%, and 88.9%, respectively. CONCLUSIONS: The high sensitivity of the recombinant OvCB_OvAEP_OvCF protein in detecting opisthorchiasis demonstrates its potential as an opisthorchiasis screening target. Nonetheless, research on reducing cross-reactivity should be undertaken by detecting other antibodies in other sample types, such as saliva, urine, and feces.

Immunization with a Mu-class glutathione transferase from Echinococcus granulosus induces efficient antibody responses and confers long-term protection against secondary cystic echinococcosis.

Cystic echinococcosis, a zoonosis caused by cestodes belonging to the Echinococcus granulosus sensu lato (s.l.) genetic complex, affects humans and diverse livestock species. Although a veterinary vaccine exhibiting high levels of antibody-mediated protection has successfully reached the market, the large genetic diversity among parasite isolates and their particular host preferences, makes still necessary the search for novel vaccine candidates. Glutathione transferases (GSTs) constitute attractive targets for immunoprophylaxis due to their outstanding relevance in helminth detoxification processes, against both exogenous and endogenous stressors. Among the six GSTs known to be expressed in E. granulosus s.l., EgGST1 (Mu-class), EgGST2 (Sigma-class), and EgGST3 (a still non-classifiable isoenzyme), show the highest proteomic expression. Therefore, their recombinant forms -rEgGST1, rEgGST2 and rEgGST3- were herein analyzed regarding their potential to induce long-term antiparasite protection in mice. Only immunization with rEgGST1 induced long-lasting protection; and accordingly, rEgGST1-specific antibodies enhanced the parasite killing through both the classical activation of the host complement system and the antibody-dependent cellular cytotoxicity by macrophages. These results support further testing of rEgGST1 as a vaccine candidate in diverse hosts due to the broad expression of EgGST1 in different parasite stages and tissues.

A point-of-care testing assay for clonorchiasis using a EuNPs-CsTR1 fluorescent probe-based immunoassay.

Clonorchis sinensis is one of the most important fish-borne zoonotic parasitic worms in humans, and is distributed in several countries with more than 15 million people infected globally. However, the lack of a point-of-care testing (POCT) method is still the critical barrier to effectively prevent clonorchiasis. With the application of novel fluorescent nanomaterials, the development of on-site testing methods with high signal enhancement can provide a simple, precise and inexpensive tool for disease detection. In this study, Eu-(III) nanoparticles (EuNPs) were used as indicative probes, combined with C. sinensis tandem repeat sequence 1 (CSTR1) antigen to capture specific antibodies. Afterward, the complex binds to mouse anti-human IgG immobilized on the test line (T-line) producing a fluorescent signal under UV light. The EuNPs-fluorescent immunoassay (EuNPs-FIA) was successfully constructed, allowing sample detection within 10 min. It enabled both qualitative determination with the naked eye under UV light and quantitative detection by scanning the fluorescence intensity on the test line and control line (C-line). A total of 133 clinical human sera (74 negative, 59 clonorchiasis, confirmed by conventional Kato-Katz (KK) methods and PCR via testing fecal samples corresponding to each serum sample) were used in this study. For qualitative analysis, the cut-off value of fluorescence for positive serum was 31.57 by testing 74 known negative human samples. The assay had no cross-reaction with other 9 parasite-infected sera, and could recognize the mixed infection sera of C. sinensis and other parasites. The sensitivity and specificity of EuNPs-FIA were both 100% compared with KK smear method. Taking advantage of its high precision and user-friendly procedure, the established EuNPs-FIA provides a powerful tool for the diagnosis and epidemiological survey of clonorchiasis.

Helminth-induced impairment of humoral immunity differently contribute to their anti-arthritic effects in mice: Comparison of Schistosoma mansoni and Trichinella spiralis.

AIMS: We have previously reported reduction of anti-type II collagen (IIC) IgG levels in collagen-induced arthritis (CIA) by Schistosoma mansoni (Sm) and Trichinella spiralis (Ts). To clarify the contribution of the impairment of humoral immunity to their anti-arthritic activities, we herein investigated the relationship between anti-IIC IgG levels and arthritic swelling in Sm- or Ts-infected mice. METHODS AND RESULTS: Male DBA/1J mice were infected with Sm cercariae or Ts muscle larvae prior to the IIC immunization. In the Sm-infected mice, paw swelling and anti-IIC IgG levels were continuously lower than those of non-infected control group. In contrast, arthritic swelling in the Ts-infected mice only decreased in the early phase of CIA progression, despite the continued impairment of anti-IIC IgG production throughout the experimental period. Correlation coefficients between residual paw swelling and anti-IIC IgG titers were similar or higher in the Sm group than in the control group, but were similar or lower in the Ts group than in the control group. CONCLUSION: The down-modulations of anti-IIC IgG levels by the two parasitic infections and the correlation analyses suggest that the anti-arthritic activity of Sm was primarily attributed to the modulation of IgG-independent arthritogenic mechanisms and secondarily to the impairment of anti-IIC IgG production. In contrast, Ts could alleviate CIA mainly via the impairment of antibody production.

Anti-Anisakis antibodies in colon cancer patients and their relationship with γδ T-cells.

Many pathogens are related to carcinogenesis. Chronic inflammation, as a result of persistent infection, leads to DNA damage, higher expression of oncogenes, decreased apoptosis and immunosuppression, which are some of the reasons for cancer induction. Among parasites, Schistosoma, Opistorchis and Clonorchis are recognised as infectious agents which contribute to cancer. A relationship between Anisakis and cancer was hypothesised because cellular responses to Anisakis products could result in inflammation and DNA damage. Previous research has shown a decrease in CD8+ γδ T-cells and an increase in αß and γδ T-cell apoptosis in colon cancer (CC) samples. Ninety-two CC patients and 60 healthy subjects were recruited. γδ and αß T-cells were analysed, and their apoptosis was evaluated. Anti-Anisakis antibodies were tested in sera from CC patients and controls. Anti-Anisakis IgG, IgM, IgA and IgE antibodies were significantly higher in CC patients. A significant increase in anti-Anisakis IgA levels was observed in patients with angiolymphatic invasion. The number of all γδ T-cells, as well as CD3+ CD4+ αß T-cells, was significantly lower in CC patients. The apoptosis of all T-cells was significantly increased in patients with CC. We observed a significantly higher percentage of anti-Anisakis IgE positive patients having a deficit of CD3+ γδ T-cells. Our results suggest a relationship between Anisakis and CC.