Augmented Innate and Adaptive Immune Responses Under Conditions of Diabetes–Filariasis Comorbidity

Metainflammation, as seen in chronic diabetes subjects, impairs immunity and increases the susceptibility to infections. In the present study, the effect of diabetes on immune response against filariasis was studied. Both toll-like receptor (TLR)-mediated and crude antigen-induced immune responses were quantified, in whole blood cultures from filariasis-infected subjects (LF+), with and without diabetes. Blood cultures were stimulated with TLR ligands (TLR2 and TLR4) or filarial antigen or were left unstimulated (control) for 18 h. Cytokine, chemokine, and defensin secretion was quantified by ELISA. Expression of HLA-DR, B7-1, B7-2, activation marker (CD69), and Th (Th1, Th2, Th17, and Th9) phenotypes was quantified by flow cytometry. Expression of immunomodulatory effectors (Cox-2, HO-1, IDO-1, and p47Phox) and Th-polarizing transcription factors (T-bet, GATA3, and ROR-γt) was quantified by quantitative PCR. Secretion of IL-27, IL-1Ra, IL-12, IL-33, IL-9, and SDF-1 was increased under diabetes conditions with increased Th9 polarization and increased expression of Cox-2 and IDO. Overall, diabetes was found to augment both TLR-mediated and antigen-induced inflammation, which can promote chronic pathology in LF+ subjects.

Evaluation of schistosomula crude antigen (SCA) as a diagnostic tool for Schistosoma mansoni in low endemic human population

The study aimed to determine the potential of schistosomula crude antigen (SCA) as a diagnostic target for anti-S. mansoni antibody detection. Cercariae were transformed into schistosomula, homogenized through sonication, and then centrifuged to obtain the SCA. SCA was evaluated using ELISA and dot blots immunoassays on 30 S. mansoni infected sera samples obtained from chronic patients and 30 non-infected humans’ sera samples. Either Kato-Katz or saline gradient method or both were employed as the diagnostic reference. Dot blots immunoassay was further performed on protein eluted from 10 to 12 kDa immunoreactive band identified by Western blot analysis. The area under the ROC curve was 0.95 (AUC 0.95, CI 0.88–1.01, p < 0.0001). The sensitivity and specificity of SCA-ELISA and dot blots assays were 96.67% and 86.67% respectively. The human IgG-specific response against SCA was significantly higher in S. mansoni infected individuals (OD = 0.678 ± 0.249) compared to the non-infected population (OD = 0.235 ± 0.136) (p < 0.0001). Our study showed that SCA and its 10–12 kDa component could be useful as diagnostic tools for chronic schistosomiasis.

Interferon-γ Response of Mycobacterium avium subsp. paratuberculosis Infected Goats to Recombinant and Synthetic Mycobacterial Antigens

Despite its potential for early diagnosis of Mycobacterium avium subsp. paratuberculosis (MAP) infection, the IFN-γ release assay is not used routinely, because of low specificity of the established crude antigen preparation Johnin (PPDj). Limited data are available assessing the potential of MAP-derived protein and lipopeptide antigens to replace PPDj in assays for goats, while cattle and sheep have been studied more extensively. Furthermore, MAP infection is claimed to interfere with the diagnosis of bovine tuberculosis when other crude antigen preparations (PPDb, PPDa) are applied. In this study, the diagnostic potential of MAP-derived recombinant protein antigens, synthetic MAP lipopentapeptides and of Mycobacterium bovis-specific peptide cocktails was assessed compared to crude mycobacterial antigen preparations in experimentally infected goats. Goats were inoculated with MAP, or Mycobacterium avium subsp. hominissuis (MAH) as surrogate for environmental mycobacteria, non-exposed animals served as controls. Mycobacterium avium Complex-specific antibody and PPDj-induced IFN-γ responses were monitored in vivo. Infection status was assessed by pathomorphological findings and bacteriological tissue culture at necropsy 1 year after inoculation. The IFN-γ response to 13 recombinant protein antigens of MAP, two synthetic MAP lipopentapeptides and three recombinant peptide cocktails of Mycobacterium bovis was investigated at three defined time points after infection. At necropsy, MAP or MAH infection was confirmed in all inoculated goats, no signs of infection were found in the controls. Antibody formation was first detected 3–6 weeks post infection (wpi) in MAH-inoculated and 11–14 wpi in the MAP-inoculated goats. Maximum PPDj-induced IFN-γ levels in MAH and MAP exposed animals were recorded 3–6 and 23–26 wpi, respectively. Positive responses continued with large individual variation. Antigens Map 0210c, Map 1693c, Map 2020, Map 3651cT(it), and Map 3651c stimulated increased whole blood IFN-γ levels in several MAP-inoculated goats compared to MAH inoculated and control animals. These IFN-γ levels correlated with the intensity of the PPDj-induced responses. The two synthetic lipopentapeptides and the other MAP-derived protein antigens had no discriminatory potential. Stimulation with Mycobacterium bovis peptide cocktails ESAT6-CFP10, Rv3020c, and Rv3615c did not elicit IFN-γ production. Further work is required to investigate if test sensitivity will increase when mixtures of the MAP-derived protein antigens are applied.

Investigation of Trypanosoma evansi infection in bullfighting cattle in Southern Thailand

Background and Aim: Trypanosoma evansi infection has been reported in Thai livestock such as beef and dairy cattle. However, there is little information on T. evansi infection in bullfighting cattle in Southern Thailand. The aim of this study was to investigate the infection of T. evansi in bullfighting cattle presented for health checks at the Animal Hospital, Faculty of Veterinary Science, Prince of Songkla University, Thailand. Materials and Methods: Blood and serum samples were collected from 177 bullfighting cattle from April 2016 to February 2017 after bullfighting matches. Animal inspected showed signs of fever, weight loss, or exercise intolerance. Investigation of T. evansi infection was tested using polymerase chain reaction (PCR) with TBR primers and using indirect enzyme-linked immunosorbent assay with T. evansi crude antigen. Results: The seroprevalence of T. evansi in bullfighting cattle was 22.60% (40/177). The PCR results detected no parasite DNA in this study. However, bullfighting cattle may serve as T. evansi reservoirs. Conclusion: Health checking procedures for T. evansi should be promoted for bullfighting events so that infected animals can be quarantined in the preparatory stages of such events.

The Role of Phosphatidylinositol Mannosides in the Serological Diagnosis of Mycobacterial Infections

Accurate diagnosis of mycobacterial infections, such as bovine tuberculosis and paratuberculosis, remains challenging. Available direct diagnostic tests aimed at detecting the pathogen are highly specific but lack sensitivity, depending on the stage of infection and the prevalence of infection in a population. The sensitivity of indirect diagnostic assays that measure the host immune response to infection is similarly affected by disease characteristics. The choice of antigen used to detect a host response to infection has a critical impact on test sensitivity and specificity. Many indirect tests rely on crude antigen preparations and cell-free extracts, of which the production is poorly standardized. Moreover, these preparations contain ample uncharacterized cross-reactive compounds. To enhance serological test specificity, existing assays depend on the pre-treatment of samples and a relatively high cut-off value, that in turn influences test sensitivity. Research therefore focuses on the identification of more specific, defined antigens to improve diagnostics. In the current study, we extracted phosphatidylinositol mannosides (PIMs) and investigated their potential use in antibody-based tests. Our results demonstrate that specific IgG class antibodies are generated against PIMs in cows, but this is unrelated to tuberculosis or paratuberculosis infection status, making these antigens unsuitable for diagnostic applications. In addition, we demonstrate that PIMs are widely present in crude antigen preparations and in serum pre-absorption buffer. Our results indicate that PIMs are cross-reactive compounds with immunodominant B cell epitopes that could impair serological test specificity.

Penentuan Kadar Protein dan Fraksi Protein Crude Antigen Cysticercus cellulosae dari Isolat Lokal

Sistiserkosis merupakan penyakit parasitik yang disebabkan oleh larva cacing pita yang disebut Cysticercus. Cysticercus yang ditemukan pada babi adalah Cysticercus cellulosae yang merupakan larva dari cacing pita Taenia solium. Tujuan penelitian ini adalah untuk mengetahui kadar dan fraksi protein Crude antigen C. cellulosae. C. cellulosae yang digunakan adalah isolat lokal yang diperoleh dari babi terinfeksi yang berasal dari Karangasem – Bali. Penentuan kadar protein larva dilakukan dengan menggunakan alat Invitrogen Quibit Fluorometer dan penentuan fraksi protein larva dengan teknik elektroforesis menggunakan SDS-PAGE. Hasil penelitian menunjukan bahwa kadar protein C. cellulosae isolat lokal sebesar 867 µg/ml. Sedangkan untuk fraksinasi protein larva diperoleh 6 bands protein dengan berat molekul: bands 1 = 120,5 kDa, bands 2 = 99,8 kDa, bands 3 = 60,8 kDa, bands 4 = 47,7 kDa, bands 5 = 35,6 kDa, dan bands 6 = 23,3 kDa.

Seroprevalence of rickettsia spp. and a study of the tick fauna in dogs from the municipality of Seropédica, State of Rio de Janeiro

The aim of this study was to investigate the presence of anti-Rickettsia spp. antibodies, the tick fauna, and the ticks that are carriers of rickettsiae of the spotted fever group (SFG). About 68 (24%) of the 283 serum samples tested by indirect immunofluorescence (IFA) reacted against the R. rickettsii crude antigen. The titers varied between 1:64 and 1:512. At the time of collection, 189 (64.5%) of the 293 dogs included in this study, were infested with ticks. Ticks classified as Rhipicephalus sanguineus and Amblyomma sculptum were identified. None of the ticks examined for SFG rickettsiae using polymerase chain reaction (PCR) were positive. The presence of the anti-R. rickettsii antibodies detected by IFA, albeit at low titers, suggests the circulation of SFG rickettsiae, which requires permanent surveillance because there are records on human fatalities related to spotted fever and to avoid any future threats to the students moving extensively in the areas near of the Rural Federal University of Rio de Janeiro.