Antigen-based diagnosis of Schistosoma infection in travellers: a prospective study

BACKGROUND: Travellers infected with Schistosoma spp. might be pauci- or even asymptomatic on first presentation. Therefore, schistosomiasis may remain undiagnosed in this population. Active infection, as evidenced by the presence of the tissue-dwelling worm, can be demonstrated via the detection of adult worm-derived circulating anodic antigen (CAA) utilising a robust well-described lateral flow-(LF) based test applying background-free up-converting reporter particles (UCP). In this prospective study, we assessed the diagnostic value of serum and urine UCP-LF CAA test in comparison with two Schistosoma-specific serological assays detecting antibodies against adult worm antigen-immuno fluorescence assay (AWA-IFA) and against soluble egg antigen-enzyme-linked immunosorbent assay (SEA-ELISA) antigens in travellers. METHODS: Samples were collected from 106 Dutch travellers who reported freshwater contact in sub-Saharan Africa and who were recruited up to 2 years after return. Subjects were asked to complete a detailed questionnaire on travel history, water contact, signs and symptoms compatible with schistosomiasis. RESULTS: Two travellers were positive by serum CAA and an additional one by urine CAA. A total of 22/106 (21%) samples were antibody positive by AWA-IFA and 9/106 (9%) by SEA-ELISA. At follow-up 6 weeks and 6 months after praziquantel treatment, all seropositives remained antibody positive whereas CAA was cleared. Seropositivity could not be predicted by the type of fresh water-related activity, country visited or symptoms reported. CONCLUSION: The low number of UCP-LF CAA positives suggests that in travellers, active infections often do not establish or have very low worm burden. Based on our high seroconversion rates, we conclude that the AWA-IFA assay is the most sensitive test to detect schistosome exposure. Given the lack of predictive symptoms or risk factors, we recommend schistosomiasis screening at least by serology in all travellers with reported freshwater contact in high-endemic areas

A rapid assay for on-site monitoring of infliximab trough levels: a feasibility study

Monitoring levels of biologicals against tumor necrosis factor (TNF) has been suggested to improve therapeutic outcomes in inflammatory bowel diseases (IBDs). This pilot study describes a rapid lateral flow (LF)-based assay for on-site monitoring of serum trough levels of humanized monoclonal antibody infliximab (IFX). The applied chromatographic method utilizes sequential flows of diluted serum, wash buffer, and an immunoglobulin generic label on LF strips with a Test line comprised of TNF-α. The successive flows permitted enrichment of IFX at the Test line before the label was applied. The label, luminescent upconverting phosphor (UCP) particles coated with protein-A, emits a 550-nm visible light upon excitation with 980-nm infrared light. IFX concentrations were determined through measurement of UCP fluorescence at the Test line. The assay was optimized to detect IFX levels as low as 0.17 μg/mL in serum. For patients with IBD, this limit is appropriate to detect levels associated with loss of response (0.5 μg IFX/mL). The assay was evaluated with clinical samples from patients with Crohn's disease and correlated well within the physiologically relevant range from 0.17 to 10 μg/mL with an IFX-specific ELISA. Performance of the assay was further successfully validated with samples from blood donors, IFX negative IBD patients, and rheumatoid arthritis patients that had developed anti-IFX antibodies. Because of its generic nature, the assay is suited for detecting most therapeutic anti-TNF-α monoclonal antibodie

Effect of schistosomiasis on the outcome of patients infected with HIV-1 starting antiretroviral therapy in rural Tanzania.

BACKGROUND:It has been hypothesized that schistosomiasis negatively influences immune reconstitution in people living with HIV starting antiretroviral therapy (ART). In this study, we investigated the effect of schistosomiasis on the course of HIV infection in patients starting ART in a rural part of Tanzania. METHODOLOGY:Retrospective study including patients prospectively enrolled in a HIV cohort in Ifakara, south-central Tanzania between January 1, 2013 and April 1, 2015. Schistosomal circulating anodic antigen (CAA) was assessed in pre-ART cryopreserved plasma. Regression models were utilized to estimate the effect of CAA positivity on virological and immunological failure and a composite outcome of death/loss to follow-up (LFU). PRINCIPAL FINDINGS:At ART-initiation 19.1% (88/461) of patients were CAA-positive. A tendency of higher CD4 increases was seen in CAA-positive patients (+182 cells/μl, interquartile range (IQR), 87-285 cells/μl) compared to CAA-negative patients (+147 cells/μl, IQR, 55-234 cells/μl, p = 0.09) after 10 months of follow-up. After adjustment for baseline risk factors, CAA-positivity showed no association with virological or immunological failure. In CAA-positive patients, 22.7% (20/88) died or were LFU, compared to 29.5% (110/373) of CAA-negative patients (hazard ratio (HR): 0.76, 95% confidence interval (CI), 0.47-1.22, p = 0.25). After adjustment for age, sex, body mass index, educational attainment, WHO-stage, tuberculosis status, and year of ART initiation, CAA-positivity showed a trend of a decreased hazard of death/LFU (HR: 0.58, 95% CI: 0.32-1.05, p = 0.07), while CD4 count at baseline (HR: 0.86, 95% CI: 0.76-1.00, p = 0.02) and MXD (sum of eosinophils, basophils, and monocytes counts) >1,100 cells/μl (HR: 0.56, 95% CI: 0.34-0.93, p = 0.03) were identified as independently protective factors. CONCLUSIONS/SIGNIFICANCE:Schistosomiasis is prevalent in this HIV cohort and may be beneficial for immunological reconstitution, while no effect on virological failure was apparent. A positive effect of schistosomiasis-induced immunomodulation on survival and retention in care needs confirmation in future studies

Antigen-based diagnosis of Schistosoma infection in travellers: a prospective study

BACKGROUND: Travellers infected with Schistosoma spp. might be pauci- or even asymptomatic on first presentation. Therefore, schistosomiasis may remain undiagnosed in this population. Active infection, as evidenced by the presence of the tissue-dwelling worm, can be demonstrated via the detection of adult worm-derived circulating anodic antigen (CAA) utilising a robust well-described lateral flow-(LF) based test applying background-free up-converting reporter particles (UCP). In this prospective study, we assessed the diagnostic value of serum and urine UCP-LF CAA test in comparison with two Schistosoma-specific serological assays detecting antibodies against adult worm antigen-immuno fluorescence assay (AWA-IFA) and against soluble egg antigen-enzyme-linked immunosorbent assay (SEA-ELISA) antigens in travellers. METHODS: Samples were collected from 106 Dutch travellers who reported freshwater contact in sub-Saharan Africa and who were recruited up to 2 years after return. Subjects were asked to complete a detailed questionnaire on travel history, water contact, signs and symptoms compatible with schistosomiasis. RESULTS: Two travellers were positive by serum CAA and an additional one by urine CAA. A total of 22/106 (21%) samples were antibody positive by AWA-IFA and 9/106 (9%) by SEA-ELISA. At follow-up 6 weeks and 6 months after praziquantel treatment, all seropositives remained antibody positive whereas CAA was cleared. Seropositivity could not be predicted by the type of fresh water-related activity, country visited or symptoms reported. CONCLUSION: The low number of UCP-LF CAA positives suggests that in travellers, active infections often do not establish or have very low worm burden. Based on our high seroconversion rates, we conclude that the AWA-IFA assay is the most sensitive test to detect schistosome exposure. Given the lack of predictive symptoms or risk factors, we recommend schistosomiasis screening at least by serology in all travellers with reported freshwater contact in high-endemic areas

Correlation between ELISAs and UCP-LFAs.

<p>Levels of IP-10 (<b>A</b>) and IL-10 (<b>B</b>) in 24 h whole blood samples of 77 <i>M. leprae</i> (antigen), LPS and PHA stimulated WBA samples of Dutch healthy controls were simultaneously determined by ELISAs and wet-format UCP-LFAs. <b>Left panels</b>: results for ELISAs are indicated in pg/ml (ELISA) or as the ratio of the relative fluorescence units (RFUs) measured at Test and Flow-Control lines (UCP-LFA). <i>R²</i> equals the square of the Pearson correlation coefficient. <b>Right panels</b>: Spearman ranking.</p

Kinetics of IP-10 production in WBA.

<p>(<b>A</b>): IP-10 concentrations produced in stimulated whole blood cultures of leprosy patients (upper panel; LP; n = 10: 5 BL (Ethiopia); 2 BT (Ethiopia); 3 BT (The Netherlands) and healthy endemic controls (lower panel; EC, n = 8) in response to <i>M. leprae</i> WCS (left panel; 10 µg/ml), <i>M. leprae</i> unique protein ML2478 (middle panel; 10 µg/ml) and PHA (right panel; 1 µg/ml). IP-10 concentrations were determined by ELISA after 1 h, 4 h, 6 h and 24 h antigen stimulation. Values on the y-axis are concentrations corrected for background values. (<b>B</b>): Comparison of IP-10 concentrations determined by ELISA after 6 h stimulation with ML2478 (10 µg/ml) of whole blood samples.</p

Performance of the portable lightweight UCP-Quant LF strip reader.

<p>Dry-format UCP-LFAs were performed for single detection of IP-10 and anti-PGL-I antibodies in an Ethiopian field setting (<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002845#pntd-0002845-g003" target="_blank">Figure 3</a>). LF strips were analyzed using a portable reader (UCP-Quant). Subsequently, LF strips were shipped to The Netherlands and re-analysed using a dedicated lab-based FluoroCount microtiterplate reader (Packard) adapted for reading UCP-LF strips. <b>Left panel</b>: results are indicated as the ratio of the relative fluorescence units (RFUs) measured at Test and Flow-Control lines. <i>R²</i> equals the square the Pearson correlation coefficient. <b>Right panel</b>: Spearman ranking. The grey box indicates samples scoring values below the specificity threshold.</p

Combined cytokine profiles in response to <i>M. leprae</i>.

<p>Production of IFN-γ (<b>A</b>), IP-10 (<b>B</b>) and IL-10 (<b>C</b>) determined by ELISA, in response to medium (-), PHA, <i>M. leprae</i> WCS or the <i>M. leprae</i>-unique protein ML2478 in 24 h WBA for Ethiopian leprosy patients (n = 11: 2 BT (○) and 9 BL (•), and healthy endemic controls (EC; n = 12; □). For comparison between BT and BL, significant differences were found for <i>M. leprae</i> WCS (Mlep) induced IFN-γ responses (p = 0.036) and ML2478 induced IL-10 responses (p = 0.035). (<b>D</b>): IP-10/IL-10 ratios are depicted for unstimulated samples after 24 h {LP (•) and EC (□)} or after 1 h WBA {LP (▵) and EC (▾)}. (<b>E</b>): Anti-PGL-I antibodies for BL (○) and BT (•) patients were detected by ELISA using natural disaccharide of PGL-I linked to HSA <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002845#pntd.0002845-Cho2" target="_blank">[31]</a> (ND-O-HSA). Optical density (OD₄₅₀) readings were performed using 1∶800 serum dilutions. Median values per group are indicated by horizontal lines. The cut-off for positivity is indicated by the dashed horizontal line.</p

Comparison between single and multiplex UCP-LFAs.

<p>UCP-LFAs were performed for single or multiplex detection of IP-10 (upper panel; n = 149 samples) and anti-PGL-I (lower panel; n = 115 samples) using <i>M. leprae</i> antigen-stimulated WBA samples of Dutch and Ethiopian leprosy patients. Simultaneous detection of IP-10 and anti-PGL-I IgM was performed following the two phase protocol using the UCP^αIP-10conjugate and the UCP^αIgM conjugate. <b>Left panel</b>: <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002845#s3" target="_blank">Results</a> for UCP-LFAs are displayed as the ratio of the relative fluorescence units (RFUs) measured at Test and Flow-Control lines. <i>R²</i> equals the square of the Pearson correlation coefficient. <b>Right panel</b>: Spearman ranking. The grey box indicates samples scoring values below the specificity threshold.</p