Evaluation and Characterization of Fasciola hepatica Tegument Protein Extract for Serodiagnosis of Human Fascioliasis

Morales, Adelaida; Espino, Ana M.

doi:10.1128/cvi.00487-12

Cited by 30 publications

(25 citation statements)

References 56 publications

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“…Th2-dominated responses have been observed in F. hepatica infection in mice, humans, and ruminant hosts. Antibody production is dominated by the immunoglobulin G1 isotype (24,41,42). In experimentally infected cattle, IFN-␥ production and cellular proliferation are detected immediately after infection with the F. hepatica antigen.…”

Section: Discussionmentioning

confidence: 98%

“…Moreover, our research group recently demonstrated that FABP is one of the most prominent components of an F. hepatica tegumental coat antigen (FhTg) that was found to be highly reactive with sera from patients with chronic fascioliasis (24). Consistent with these results, the native F. hepatica FABP (Fh12) purified in this study was found to be highly reactive against human sera obtained from patients with chronic fascioliasis and against antisera raised against specific tegumental coat antigens and excretory-secretory products of F. hepatica.…”

Section: Discussionmentioning

confidence: 99%

“…Each aliquot was subjected to 12.5% SDS-PAGE, and the proteins were visualized by silver staining. Fractions from the second IEF run with a pH between 4.61 and 5.9 that exhibited a polypeptide band around 12 to 15 kDa were manually excised from the gel, washed twice with double-distilled water, digested with sequencing-grade trypsin (Promega, Madison, WI), and analyzed by matrix-assisted laser desorption ionization (MALDI) and tandem mass spectrometry (MS/MS), as previously described (24). Fractions in which the fatty acid binding protein was identified were pooled, exhaustively desalted against PBS, concentrated by ultrafiltration using an YM-10 membrane (which excludes components with molecular masses lower than 10 kDa), and stored at Ϫ20°C.…”

Section: Methodsmentioning

confidence: 99%

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Fasciola hepatica Fatty Acid Binding Protein Induces the Alternative Activation of Human Macrophages

Figueroa-Santiago

Espino

2014

Infect Immun

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The liver fluke Fasciola hepatica is a highly evolved parasite that uses sophisticated mechanisms to evade the host immune response. The immunosuppressive capabilities of the parasite have been associated with antigens secreted through the parasite's tegument, called excretory-secretory products (ESPs). Proteomic studies have identified the fatty acid binding protein (FABP) as one of molecules present in the parasite ESPs. Although FABP has been investigated for potential use in the development of vaccines against fascioliasis, its direct interaction with cells of immune system has not been studied. In this study, FABP was purified in native form from soluble extracts of F. hepatica adult flukes using a combination of molecular sieving chromatography and preparative isoelectric focusing. The immunological effect of the purified protein, termed Fh12, was assayed in vitro using monocyte-derived macrophages (MDM) obtained from healthy human donors. Results from the assay indicate that Fh12 produced a significantly increased arginase expression and activity and induced the expression of chitinase-3-like protein (CHI3L1). The assay also showed that Fh12 downregulated the production of nitric oxide (NO) and the expression of nitric oxide synthase (NOS2). This indicates that Fh12 induced the production of alternatively activated macrophages (AAM). The results also demonstrated the ability of Fh12 to downregulate the secretion of the proinflammatory and inflammatory cytokines tumor necrosis factor alpha (TNF-␣), interleukin-12 (IL-12), and IL-1␤B, even after stimulation with lipopolysaccharide (LPS), as well as its ability to stimulate the overexpression of IL-10. These results suggest a potent anti-inflammatory role for Fh12, which could occur via targeting of Toll-like receptor 4 (TLR4).

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Section: Discussionmentioning

confidence: 98%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Fasciola hepatica Fatty Acid Binding Protein Induces the Alternative Activation of Human Macrophages

Figueroa-Santiago

Espino

2014

Infect Immun

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“…However, other possible explanations also could be considered, i.e., (i) the persistence of low antibody levels as these persons could have been previously exposed to F. hepatica, although at the time of sample collection they did not have eggs in their feces (we do not know whether these persons lived in regions with endemic fascioliasis prior to sample collection), or (ii) cases of resolved fascioliasis infections (we recorded no history of possible previous treatment with triclabendazole). We further evaluated the falsepositive serum samples with two other ELISA methods that were previously validated in our laboratory, utilizing as antigens recombinant F. hepatica saposin-2 (25) and F. hepatica tegumental proteins (52), which showed diagnostic specificity values greater than 95%. The results from these two additional ELISAs indicated that 5 of the 11 false-positive serum samples (2 samples with schistosomiasis, 1 with paragonimiasis, and 2 with hydatidosis) could be true-positive cases that were misdiagnosed in the microscopic examinations.…”

Section: Discussionmentioning

confidence: 99%

Development of Two Antibody Detection Enzyme-Linked Immunosorbent Assays for Serodiagnosis of Human Chronic Fascioliasis

et al. 2014

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Coprological examination based on egg detection in stool samples is currently used as the gold standard for the diagnosis of human fascioliasis. However, this method is not effective during the acute phase of the disease and has poor sensitivity during the chronic phase. Serodiagnosis has become an excellent alternative to coprological examination in efforts to combat the effects of fascioliasis on human and animal health. Two novel recombinant Fasciola hepatica proteins, i.e., a ferritin (FhFtn-1) and a tegument-associated protein (FhTP16.5), were used as antigens to develop in-house enzyme-linked immunosorbent assay (ELISA) methods. The assays were optimized and validated using 152 serum samples from humans with a known infection status, including healthy subjects, patients with chronic fascioliasis, and patients with other parasitic diseases. The FhFtn-1 ELISA was shown to be 96.6% sensitive and 95.7% specific; the respective parameters for the FhTP16.5 ELISA were 91.4% and 92.4%. The performances of the FhFtn-1 and FhTP16.5 ELISAs were compared with that of an available commercial test (the DRG test) using a subset of serum samples. Our in-house tests were slightly more sensitive than the DRG test in detecting antibodies against F. hepatica, but the differences were not statistically significant. In conclusion, the present study provides evidence for the potential of the FhFtn-1 and FhTP16.5 ELISAs as diagnostic tools for human fascioliasis, as might be implemented in conjunction with standard assays for large-scale screenings in areas where the disease is endemic and for the detection of occasional cases in clinical laboratories.

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“…Different antigens of Fasciola including somatic, ES, recombinant and tegumental antigens have been used for immunodiagnosis of human or animal fasciolosis (Morales and Espino 2012;Rokni et al 2003;Shafiei et al 2015). The present study was carried out to determine common antigens of F. hepatica and F. gigantica somatic and ES antigens, using rabbit polyclonal antibodies raised against these two sets of antigens.…”

Section: Discussionmentioning

confidence: 99%

Detection of Fasciola hepatica and Fasciola gigantica common and uncommon antigens, using rabbit hyper immune serum raised against their excretory–secretory and somatic antigens

Khabisi

Sarkari

2016

J Parasit Dis

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Fasciolosis is an important neglected helminth disease caused by two liver flukes, Fasciola hepatica and Fasciola gigantica. The two species of Fasciola are usually different in their morphological and molecular features. They have also common and uncommon antigens in both their somatic and excretory secretory metabolites. In this study, we compared somatic and excretory-secretory (ES) antigens of F. hepatica and F. gigantica, by using rabbit hyper immune serum raised against these antigens. Adult worms were collected from bile ducts of infected animals and species of the fluke was confirmed by RFLP-PCR. ES and somatic antigens of both species were prepared. Rabbits were subcutaneously immunized with either ES or somatic antigens to produce antibodies against these antigens. SDS-PAGE pattern of F. hepatica and F. gigantica somatic antigens was similar and both of them revealed 30 protein bands, ranging from 18 to 180 kDa. In contrast, SDS-PAGE pattern of ES antigen of the two species was different. While protein bands with molecular weight of 18, 27, 29, 48, and 62 kDa were common in both species, bands of 19, 45, 55 and 58 kDa were only noticed in F. hepatica ES antigen. Rabbit polyclonal antibodies, raised against F. hepatica and F. gigantica ES antigen, reacted with main five protein bands, 25, 27, 29, 62 and 67 kDa and polyclonal antibodies raised against somatic antigens of both species reacted with three protein bands, 25, 27 and 72 kDa. Thus, the 25, 27 and 29 kDa protein bands may serve as immunodominant antigens, which might be considered for serodiagnosis of fasciolosis. Moreover, bands of 62 and 67 kDa in ES antigen and 72 kDa in somatic antigens of both species were immunodominant and might be suitable candidate for development of serological assays for diagnosis of fasciolosis.

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Evaluation and Characterization of Fasciola hepatica Tegument Protein Extract for Serodiagnosis of Human Fascioliasis

Cited by 30 publications

References 56 publications

Fasciola hepatica Fatty Acid Binding Protein Induces the Alternative Activation of Human Macrophages

Fasciola hepatica Fatty Acid Binding Protein Induces the Alternative Activation of Human Macrophages

Development of Two Antibody Detection Enzyme-Linked Immunosorbent Assays for Serodiagnosis of Human Chronic Fascioliasis

Detection of Fasciola hepatica and Fasciola gigantica common and uncommon antigens, using rabbit hyper immune serum raised against their excretory–secretory and somatic antigens

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