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Background: The National Program for Chagas disease was implemented in Bolivia in 2006, and it greatly decreased the number of infections through vector control. Subsequently, a treatment regimen of benznidazole (BNZ) was started in seropositive school-age children living in certified vector control areas. Methods and findings: We conducted a 12-month follow-up study and seven blood samples were taken during and after the treatment. Serology, conventional diagnostic PCR (cPCR) and quantitative Real-time PCR (qPCR) were performed. Plasma Th1/Th2/Th17 cytokines levels were also determined. Approximately 73 of 103 seropositive children complied with BNZ, with three interruptions due to side effects. To evaluate each individual’s treatment efficacy, the cPCR and qPCR values during the final 6 months of the follow-up period were observed. Among 57 children who completed follow-up, 6 individuals (11%) showed both cPCR(+) and qPCR(+) (non reactive), 24 (42%) cPCR(-) but qPCR(+) (ambiguous) and 27 (47%) cPCR(-) and qPCR(-) (reactive). Within 14 Th1/Th2/Th17 cytokines, IL-17A showed significantly higher levels in seropositive children before the treatment compared to age-matched seronegative children and significantly decreased to the normal level one-year after. Moreover, throughout the follow-up study, IL-17A levels were positively co-related to parasite counts detected by qPCR. At the 12 months’ time point, IL-17A levels of non-reactive subjects were significantly higher than either those of reactive or ambiguous subjects suggesting that IL-17A might be useful to determine the reactivity to BNZ treatment. Conclusions: Plasma levels of IL-17A might be a bio-marker for detecting persistent infection of T. cruzi and its chronic inflammation. [ABSTRACT FROM AUTHOR]

Background: Zika virus (ZIKV) and Dengue virus (DENV) are often co-endemic. The high protein-sequence homology of flaviviruses renders IgG induced by and directed against them highly cross-reactive against their antigen(s), as observed on a large set of sera, leading to poorly reliable sero-diagnosis. Methods: We selected Domain III of the ZIKV Envelope (ZEDIII) sequence, which is virus specific. This recombinant domain was expressed and purified for the specific detection of ZEDIII-induced IgG by ELISA from ZIKV-RT-PCR-positive, ZIKV-IgM-positive, flavivirus-positive but ZIKV-negative, or flavivirus-negative sera. We also assessed the reactivity of ZEDIII-specific human antibodies against EDIII of DENV serotype 4 (D4EDIII) as a specific control. Sera from ZEDIII-immunized mice were also tested. Results: Cross-reactivity of IgG from 5,600 sera against total inactivated DENV or ZIKV was high (71.0% [69.1; 72.2]), whereas the specificity and sensitivity calculated using a representative cohort (242 sera) reached 90% [84.0; 95.8] and 92% [84.5; 99.5], respectively, using a ZEDIII-based ELISA. Moreover, purified human IgG against D2EDIII or D4EDIII did not bind to ZEDIII and we observed no D4EDIII reactivity with ZIKV-induced mouse polyclonal IgGs. Conclusions: We developed a ZEDIII-based ELISA that can discriminate between past or current DENV and ZIKV infections, allowing the detection of a serological scar from other flaviviruses. This could be used to confirm exposure of pregnant women or to follow the spread of an endemic disease. [ABSTRACT FROM AUTHOR]

Zika virus infection is associated with the development of Guillain-Barré syndrome (GBS), a neurological autoimmune disorder caused by immune recognition of gangliosides and other components at nerve membranes. Using a high-throughput ELISA, we have analyzed the anti-glycolipid antibody profile, including gangliosides, of plasma samples from patients with Zika infections associated or not with GBS in Salvador, Brazil. We have observed that Zika patients that develop GBS present higher levels of anti-ganglioside antibodies when compared to Zika patients without GBS. We also observed that a broad repertoire of gangliosides was targeted by both IgM and IgG anti-self antibodies in these patients. Since Zika virus infects neurons, which contain membrane gangliosides, antigen presentation of these infected cells may trigger the observed autoimmune anti-ganglioside antibodies suggesting direct infection-induced autoantibodies as a cause leading to GBS development. Collectively, our results establish a link between anti-ganglioside antibodies and Zika-associated GBS in patients. [ABSTRACT FROM AUTHOR]

Mother-to-child-transmission (MTCT) of human T-cell lymphotropic virus type-1(HTLV-1) contributes disproportionately to the burden of HTLV-1 associated diseases. All preventive measures to avoid MTCT rely on the identification of infected mothers. However, the impact of pregnancy on HTLV-1 diagnosis has not been clearly assessed. Paired samples from 21 HTLV-1 infected women taken during pregnancy and while not pregnant were analysed by CMIA and PCR. The signal-to-cut-off values (S/CO) were higher during pregnancy than in the paired non-pregnant samples. HTLV-1 proviral load did not alter significantly by pregnant state. S/CO positively correlated with HTLV proviral load. Pregnancy does not impair the diagnosis of HTLV-1/2, by either immunological (CMIA) or molecular (qPCR/nPCR) tests. [ABSTRACT FROM AUTHOR]

Background: Rift Valley fever (RVF) is one of the main vector borne zoonotic diseases that affects a wide range of ruminants and human beings in Africa and the Arabian Peninsula. A rapid and specific test for RVF diagnosis at the site of a suspected outbreak is crucial for the implementation of control measures. Methodology/Principal findings: A first-line lateral flow immunochromatographic strip test (LFT) was developed for the detection of the nucleoprotein (N) of the RVF virus (RVFV). Its diagnostic performance characteristics were evaluated using reference stocks isolates recovered from different hosts and in geographic regions mimicking clinical specimens and from known RVF negative serum samples. A high level of diagnostic accuracy (DSe (35/35), DSp (167/169)) was observed, including the absence of cross-reactivity with viruses belonging to different genera. Conclusion/Significance: The fact no specialized reagents and laboratory equipment are needed, make this assay a valuable, first-line diagnostic tool in resource-poor diagnostic territories for on-site RVFV detection, however the staff require training. [ABSTRACT FROM AUTHOR]

Background: Pathogens causing acute fever, with the exception of malaria, remain largely unidentified in sub-Saharan Africa, given the local unavailability of diagnostic tests and the broad differential diagnosis. Methodology: We conducted a cross-sectional study including outpatient acute undifferentiated fever in both children and adults, between November 2015 and June 2016 in Kinshasa, Democratic Republic of Congo. Serological and molecular diagnostic tests for selected arboviral infections were performed on blood, including PCR, NS1-RDT, ELISA and IFA for acute, and ELISA and IFA for past infections. Results: Investigation among 342 patients, aged 2 to 68 years (mean age of 21 years), with acute undifferentiated fever (having no clear focus of infection) revealed 19 (8.1%) acute dengue–caused by DENV-1 and/or DENV-2 –and 2 (0.9%) acute chikungunya infections. Furthermore, 30.2% and 26.4% of participants had been infected in the past with dengue and chikungunya, respectively. We found no evidence of acute Zika nor yellow fever virus infections. 45.3% of patients tested positive on malaria Rapid Diagnostic Test, 87.7% received antimalarial treatment and 64.3% received antibacterial treatment. Discussion: Chikungunya outbreaks have been reported in the study area in the past, so the high seroprevalence is not surprising. However, scarce evidence exists on dengue transmission in Kinshasa and based on our data, circulation is more important than previously reported. Furthermore, our study shows that the prescription of antibiotics, both antibacterial and antimalarial drugs, is rampant. Studies like this one, elucidating the causes of acute fever, may lead to a more considerate and rigorous use of antibiotics. This will not only stem the ever-increasing problem of antimicrobial resistance, but will–ultimately and hopefully–improve the clinical care of outpatients in low-resource settings. Trial registration: ClinicalTrials.gov . [ABSTRACT FROM AUTHOR]

Background: Evaluation of therapeutic response in chronic Chagas disease is a major challenge, due to prolonged persistence of Trypanosoma cruzi-specific antibodies, lack of sensitivity of parasitological tests, and need for long-term follow-up to observe negative seroconversion of conventional serological tests (CS). The objective of this study was to evaluate F2/3-ELISA serology, a promising early biomarker of therapeutic response, and T.cruzi Polymerase chain reaction (PCR) for T. cruzi Deoxyribonucleic acid (DNA), for neonatal diagnosis and evaluation of parasitemia after treatment. Methods: Prospective cohort study, with three-year clinical, serological and parasitological follow-up of pediatric Chagas disease patients treated with benznidazole. Serology was evaluated by Enzyme-Linked ImmunoSorbent Assay (ELISA), Indirect hemagglutination (IHA) and F2/3-ELISA; Parasitemia by microhematocrit (MH) and PCR. Results: A cohort of 107 pediatric patients treated with benznidazole was enrolled in the study. ELISA and IHA were initially reactive in 100% of patients, F2/3-ELISA serology was reactive in 80% (86/107) and 91% (97/107) had detectable parasitemia. Seventy-six (71%) patients completed at least 36 months of serological follow up after treatment. Although a similar decreasing linear trend was observed for all serological tests, F2/3-ELISA presented earlier, age dependent, negative seroconversion compared to CS. All patients reaching undetectable CS titers had previously seroreverted by F2/3-ELISA. All patients with persistently decreasing antibody titers had negative PCRs throughout the follow up period. No new cardiological lesions were observed during the 3 years follow-up period. Conclusions: The data reported here, using CS, F2/3 ELISA and PCR provide support for the efficacy of benznidazole in congenital Chagas diseases. These results provide support for scaling up of screening, diagnosis and access to benznidazole treatment. Trial registration: ClinicalTrials.gov 0028/04 in the Research Council, Secretary of Health Buenos Aires city Goberment. [ABSTRACT FROM AUTHOR]

The explosive spread of Zika virus (ZIKV) and associated complications in flavivirus-endemic regions underscore the need for sensitive and specific serodiagnostic tests to distinguish ZIKV, dengue virus (DENV) and other flavivirus infections. Compared with traditional envelope protein-based assays, several nonstructural protein 1 (NS1)-based assays showed improved specificity, however, none can detect and discriminate three flaviviruses in a single assay. Moreover, secondary DENV infection and ZIKV infection with previous DENV infection, both common in endemic regions, cannot be discriminated. In this study, we developed a high-throughput and multiplex IgG microsphere immunoassay (MIA) using the NS1 proteins of DENV1-DENV4, ZIKV and West Nile virus (WNV) to test samples from reverse-transcription-polymerase-chain reaction-confirmed cases, including primary DENV1, DENV2, DENV3, WNV and ZIKV infections, secondary DENV infection, and ZIKV infection with previous DENV infection. Combination of four DENV NS1 IgG MIAs revealed a sensitivity of 94.3% and specificity of 97.2% to detect DENV infection. The ZIKV and WNV NS1 IgG MIAs had a sensitivity/specificity of 100%/87.9% and 86.1%/78.4%, respectively. A positive correlation was found between the readouts of enzyme-linked immunosorbent assay and MIA for different NS1 tested. Based on the ratio of relative median fluorescence intensity of ZIKV NS1 to DENV1 NS1, the IgG MIA can distinguish ZIKV infection with previous DENV infection and secondary DENV infection with a sensitivity of 88.9–90.0% and specificity of 91.7–100.0%. The multiplex and high-throughput assay could be applied to serodiagnosis and serosurveillance of DENV, ZIKV and WNV infections in endemic regions. [ABSTRACT FROM AUTHOR]

Bacillus anthracis and Yersinia pestis are zoonotic bacteria capable of causing severe and sometimes fatal infections in animals and humans. Although considered as diseases of antiquity in industrialized countries due to animal and public health improvements, they remain endemic in vast regions of the world disproportionally affecting the poor. These pathogens also remain a serious threat if deployed in biological warfare. A single vaccine capable of stimulating rapid protection against both pathogens would be an extremely advantageous public health tool. We produced multiple-antigen fusion proteins (MaF1 and MaF2) containing protective regions from B. anthracis protective antigen (PA) and lethal factor (LF), and from Y. pestis V antigen (LcrV) and fraction 1 (F1) capsule. The MaF2 sequence was also expressed from a plasmid construct (pDNA-MaF2). Immunogenicity and protective efficacy were investigated in mice following homologous and heterologous prime-boost immunization. Antibody responses were determined by ELISA and anthrax toxin neutralization assay. Vaccine efficacy was determined against lethal challenge with either anthrax toxin or Y. pestis. Both constructs elicited LcrV and LF-specific serum IgG, and MaF2 elicited toxin-neutralizing antibodies. Immunizations with MaF2 conferred 100% and 88% protection against Y. pestis and anthrax toxin, respectively. In contrast, pDNA-MaF2 conferred only 63% protection against Y. pestis and no protection against anthrax toxin challenge. pDNA-MaF2-prime MaF2-boost induced 75% protection against Y. pestis and 25% protection against anthrax toxin. Protection was increased by the molecular adjuvant CARDif. In conclusion, MaF2 is a promising multi-antigen vaccine candidate against anthrax and plague that warrants further investigation. [ABSTRACT FROM AUTHOR]

Background: Accurate and accessible diagnosis is key for the control of visceral leishmaniasis (VL). Yet, current diagnostic tests for VL have severe limitations: they are invasive or not suitable as point of care (POC) test or their performance is suboptimal in East Africa. We analysed the antigens in the VL serodiagnostics development pipeline to identify shortcomings and to propose strategies in the development of an alternative POC test for VL in East Africa. Objectives: The objective of this study was to identify and to analyse all antigens for VL serodiagnosis that have been published before 2018 in order to identify candidates and gaps in the pipeline for a new POC test in East Africa. Methods: A systematic literature search was performed on PubMed for original research articles on Leishmania-specific antigens for antibody detection of VL in humans. From each article, the following information was extracted: the antigen name, test format and characteristics, its reported sensitivity and specificity and study cohort specifications. Results: One hundred and seven articles containing information about 96 tests based on 89 different antigens were included in this study. Eighty six of these tests, comprising 80 antigens, were evaluated in phase I and II studies only. Only 20 antigens, all of which are native, contain a carbohydrate and/or lipid moiety. Twenty-four antigens, of which 7 are non-native, are composed of antigen mixtures. Nineteen tests, comprising 18 antigens, have been evaluated on East African specimens, of which only 2 (rK28 based immunochromatographic test and intact promastigote based indirect fluorescent antibody technique) consistently showed sensitivities above 94 and specificities above 97% in a phase III study and one in a phase II study (dot blot with SLA). Only rK28 is a non-native mixture of antigens which we consider suitable for further evaluation and implementation. Conclusions: The development pipeline for an alternative serodiagnostic test for VL is almost empty. Most antigens are not sufficiently evaluated. Non-protein antigens and antigen mixtures are being neglected. We propose to expand the evaluation of existing antigen candidates and to investigate the diagnostic potential of defined non-native carbohydrate and lipid antigens for VL serodiagnosis in East Africa. [ABSTRACT FROM AUTHOR]

Background: Babesia, a tick-borne genus of intraerythrocytic parasites, is understudied in humans outside of established high-endemic areas. There is a paucity of data on Babesia in Africa, despite evidence that it is regionally present. A pilot study suggested that Babesia was present in a rural district of Tanzania. Methodology/Principal findings: A cross-sectional study was conducted July-August 2017: residents in a case hamlet that had clustering of subjects with high signal-to-cut off (S/CO) ratios for antibodies against B. microti in the pilot study, and a control hamlet that had lacked significant signal, were evaluated for B. microti. Subjects aged ≥15yrs (n = 299) underwent clinical evaluation and household inspections; 10ml whole blood was drawn for Babesia transcription mediated amplification (TMA), B. microti indirect fluorescent antibody testing (IFA) and rapid diagnostic testing (RDT) for Plasmodium spp. Subjects aged <15yrs (n = 266) underwent a RDT for Plasmodium and assessment by ELISA for B. microti antibodies. A total of 570 subjects participated (mean age 22 [<1 to 90yrs]) of whom 50.7% were female and 145 (25.5%) subjects were Plasmodium RDT positive (+). In those <15yrs, the median ELISA S/CO was 1.11 (IQR 0.80–1.48); the median S/CO in the case (n = 120) and control (n = 146) hamlets was 1.19 (IQR 0.81–1.48) and 1.06 (IQR 0.80–1.50) respectively (p = 0.4). Children ≥5yrs old were more likely to have a higher S/CO ratio than those <5yrs old (p<0.001). One hundred (38%) subjects <15yrs were Plasmodium RDT+. The median S/CO ratio (children <15yrs) did not differ by RDT status (p = 0.15). In subjects ≥15yrs, no molecular test was positive for Babesia, but four subjects (1.4%) were IFA reactive (two each at titers of 128 and 256). Conclusions/Significance: The findings offer further support for Babesia in rural Tanzania. However, low prevalence of seroreactivity questions its clinical significance. [ABSTRACT FROM AUTHOR]

The 2013–2016 Ebola virus outbreak in West Africa was the largest and deadliest outbreak to date. Here we conducted a serological study to examine the antibody levels in survivors and the seroconversion in close contacts who took care of Ebola-infected individuals, but did not develop symptoms of Ebola virus disease. In March 2017, we collected blood samples from 481 individuals in Makeni, Sierra Leone: 214 survivors and 267 close contacts. Using commercial, quantitative ELISAs, we tested the plasma for IgG-specific antibodies against three major viral antigens: GP, the only viral glycoprotein expressed on the virus surface; NP, the most abundant viral protein; and VP40, a major structural protein of Zaire ebolavirus. We also determined neutralizing antibody titers. In the cohort of Ebola survivors, 97.7% of samples (209/214) had measurable antibody levels against GP, NP, and/or VP40. Of these positive samples, all but one had measurable neutralizing antibody titers against Ebola virus. For the close contacts, up to 12.7% (34/267) may have experienced a subclinical virus infection as indicated by detectable antibodies against GP. Further investigation is warranted to determine whether these close contacts truly experienced subclinical infections and whether these asymptomatic infections played a role in the dynamics of transmission. [ABSTRACT FROM AUTHOR]

Background: Oral cholera vaccine (OCV) containing killed Vibrio cholerae O1 and O139 organisms (Bivalent-OCV; Biv-OCV) are playing a central role in global cholera control strategies. OCV is currently administered in a 2-dose regimen (day 0 and 14). There is a growing body of evidence that immune responses targeting the O-specific polysaccharide (OSP) of V. cholerae mediate protection against cholera. There are limited data on anti-OSP responses in recipients of Biv-OCV. We assessed serum antibody responses against O1 OSP, as well as antibody secreting cell (ASC) responses (a surrogate marker for mucosal immunity) and memory B cell responses in blood of adult recipients of Biv-OCV in Dhaka, Bangladesh. Methodology/Principal findings: We enrolled 30 healthy adults in this study and administered two doses of OCV (Shanchol) at days 0 and 14. Blood samples were collected before vaccination (day 0) and 7 days after each vaccination (day 7 and day 21), as well as on day 44. Serum responses were largely IgA with minimal IgG and IgM responses in this population. There was no appreciable boosting following day 14 vaccination. There were significant anti-OSP IgA ASC responses on day 7 following the first vaccination, but none after the second immunization. Anti-OSP IgA memory B cell responses were detectable 30 days after completion of the vaccination series, with no evident induction of IgG memory responses. In this population, anti-Ogawa OSP responses were more prominent than anti-Inaba responses, perhaps reflecting impact of previous exposure. Serum anti-OSP responses returned to baseline within 30 days of completing the vaccine series. Conclusion: Our results call into question the utility of the 2-dose regimen separated by 14 days in adults in cholera endemic areas, and also suggest that Biv-OCV-induced immune responses targeting OSP are largely IgA in this highly endemic cholera area. Studies in children in cholera-endemic areas need to be performed. Protective efficacy that extends for more than a month after vaccination presumably is mediated by direct mucosal immune response which is not assessed in this study. Our results suggest a single dose of OCV in adults in a cholera endemic zone may be sufficient to mediate at least short-term protection. [ABSTRACT FROM AUTHOR]

Background: Visceral Leishmaniasis in humans presents with fever, anemia, and splenomegaly and can be lethal if not treated. Nevertheless, the majority of Leishmania infantum-infected individuals does not manifest symptoms and remain so provided they are not immunosuppressed. In this work, the performance of different tests was evaluated to detect asymptomatic individuals who were living in Teresina, Piauí state, Brazil, an endemic area for VL. Methodology: L. infantum-specific antibodies were detected by ELISA and two different rapid immunochromatographic (IC) diagnostic tests, Kalazar Detect and OnSite, and parasitic loads were detected by real time PCR [qPCR]. Additionally, we measured levels of the biomarkers monokine induced by IFN-γ (MIG) and IFN-γ-induced protein 10 (IP-10) before and after stimulation of whole blood with soluble Leishmania antigen [SLA]. Principal findings: Kalazar Detect and OnSite detected, respectively, 76% and 64% of patients presenting with active Visceral Leishmaniasis; 50% and 57% of patients remained positive in these tests, respectively, after treatment. Of the healthy participants in the study who were living in the endemic area, only 1.7% were positive with both of the IC tests. On the other hand, reactivity in ELISA tests revealed that 13% of these individuals presented asymptomatic infections; among VL patients, 84% presenting with active disease were reactive in ELISA, and after treatment, 55.5% were seropositive. L. infantum DNA was present in the blood of 37.9% of infected individuals living in the endemic area, while IP-10 and MIG biomarkers were detected in 26.7% of them. The greatest concordance of positivity occurred between ELISA and qPCR. Conclusion: The association of different techniques can detect asymptomatic infections, however, more research is necessary to develop ideal biomarkers that are simple to use in the clinic and in field studies in areas endemic for Visceral Leishmaniasis. [ABSTRACT FROM AUTHOR]

Nipah virus (NiV) is a highly pathogenic re-emerging virus that causes outbreaks in South East Asia. Currently, no approved and licensed vaccine or antivirals exist. Here, we investigated the efficacy of ChAdOx1 NiVB, a simian adenovirus-based vaccine encoding NiV glycoprotein (G) Bangladesh, in Syrian hamsters. Prime-only as well as prime-boost vaccination resulted in uniform protection against a lethal challenge with NiV Bangladesh: all animals survived challenge and we were unable to find infectious virus either in oral swabs, lung or brain tissue. Furthermore, no pathological lung damage was observed. A single-dose of ChAdOx1 NiVB also prevented disease and lethality from heterologous challenge with NiV Malaysia. While we were unable to detect infectious virus in swabs or tissue of animals challenged with the heterologous strain, a very limited amount of viral RNA could be found in lung tissue by in situ hybridization. A single dose of ChAdOx1 NiVB also provided partial protection against Hendra virus and passive transfer of antibodies elicited by ChAdOx1 NiVB vaccination partially protected Syrian hamsters against NiV Bangladesh. From these data, we conclude that ChAdOx1 NiVB is a suitable candidate for further NiV vaccine pre-clinical development. [ABSTRACT FROM AUTHOR]

Background: Typhoid fevers are infections caused by the bacteria Salmonella enterica serovar Typhi (Salmonella Typhi) and Paratyphi A, B and C (Salmonella Paratyphi). Approximately 17.8 million incident cases of typhoid fever occur annually, and incidence is highest in children. The accuracy of current diagnostic tests of typhoid fever is poorly understood. We aimed to determine the comparative accuracy of available tests for the pediatric population. Methods: We first conducted a systematic literature review to identify studies that compared diagnostic tests for typhoid fever in children (aged ≤15 years) to blood culture results. We applied a Bayesian latent-class extension to a network meta-analysis model. We modelled known diagnostic properties of bone marrow culture and the relationship between bone marrow and blood culture as informative priors in a Bayesian framework. We tested sensitivities for the proportion of negative blood samples that were false as well as bone marrow sensitivity and specificity. Results: We found 510 comparisons from 196 studies and 57 specific to the pediatric population. IgM-based tests outperformed their IgG-based counterparts for ELISA and Typhidot tests. The lateral flow IgG test performed comparatively well with 92% sensitivity (72% to 98% across scenario analyses) and 94% specificity. The most sensitive test of those investigated for the South Asian pediatric population was the Reverse Passive Hemagglutination Assay with 96% sensitivity (98% - 100% across scenario analyses). Adding a Widal slide test to other typhoid diagnostics did not substantially improve diagnostic performance beyond the single test alone, however, a lateral flow-based IgG rapid test combined with the typhoid/paratyphoid (TPT) assay yielded improvements in sensitivity without substantial declines in specificity and was the best performing combination test in this setting. Conclusion: In the pediatric population, lateral-flow IgG, TPT and Reverse Passive Hemagglutination tests had high diagnostic accuracy compared to other diagnostics. Combinations of tests may provide a feasible option to increase diagnostic sensitivity. South Asia has the most informed set of data on typhoid diagnostic testing accuracy, and the evidence base in other important regions needs to be expanded. [ABSTRACT FROM AUTHOR]

Fever is a regulated increase of the body temperature resulting from both infectious and non-infectious causes. Fever is known to play a role in modulating immune responses to infection, but the potential of febrile temperatures in regulating antigen binding affinity to antibodies has not been explored. Here we investigated this process under in vitro conditions using Isothermal titration calorimetry and ELISA. We used selected malarial and dengue antigens against specific monoclonal antibodies, and observed a marked increase in the affinity of these antibody-antigen complexes at 40°C, compared to physiological (37°C) or pathophysiological temperatures (42°C). Induced thermal equilibration of the protein partners at these temperatures in vitro, prior to measurements, further increased their binding affinity. These results suggest another positive and adaptive role for fever in vivo, and highlight the favourable role of thermal priming in enhancing protein-protein affinity for samples with limited availability. [ABSTRACT FROM AUTHOR]

Background: Asymptomatic Leishmania donovani infections outnumber clinical presentations, however the predictors for development of active disease are not well known. We aimed to identify serological, immunological and genetic markers for progression from L. donovani infection to clinical Visceral Leishmaniasis (VL). Methods: We enrolled all residents >2 years of age in 27 VL endemic villages in Bihar (India). Blood samples collected on filter paper on two occasions 6–12 months apart, were tested for antibodies against L. donovani with rK39-ELISA and DAT. Sero converters, (negative for both tests in the first round but positive on either of the two during the second round) and controls (negative on both tests on both occasions) were followed for three years. At the start of follow-up venous blood was collected for the following tests: DAT, rK39- ELISA, Quantiferon assay, SNP/HLA genotyping and L.donovani specific quantitative PCR. Results: Among 1,606 subjects enrolled,17 (8/476 seroconverters and 9/1,130 controls) developed VL (OR 3.1; 95% CI 1.1–8.3). High DAT and rK39 ELISA antibody titers as well as positive qPCR were strongly and significantly associated with progression from seroconversion to VL with odds ratios of 19.1, 30.3 and 20.9 respectively. Most VL cases arose early (median 5 months) during follow-up. Conclusion: We confirmed the strong association between high DAT and/or rK39 titers and progression to disease among asymptomatic subjects and identified qPCR as an additional predictor. Low predictive values do not warrant prophylactic treatment but as most progressed to VL early during follow-up, careful oberservation of these subjects for at least 6 months is indicated. [ABSTRACT FROM AUTHOR]

Background: Despite decades of use of control programs, schistosomiasis remains a global public health problem. To further reduce prevalence and intensity of infection, or to achieve the goal of elimination in low-endemic areas, there needs to be better diagnostic tools to detect low-intensity infections in low-endemic areas in Brazil. The rationale for development of new diagnostic tools is that the current standard test Kato-Katz (KK) is not sensitive enough to detect low-intensity infections in low-endemic areas. In order to develop new diagnostic tools, we employed a proteomics approach to identify biomarkers associated with schistosome-specific immune responses in hopes of developing sensitive and specific new methods for immunodiagnosis. Methods and findings: Immunoproteomic analyses were performed on egg extracts of Schistosoma mansoni using pooled sera from infected or non-infected individuals from a low-endemic area of Brazil. Cross reactivity with other soil-transmitted helminths (STH) was determined using pooled sera from individuals uniquely infected with different helminths. Using this approach, we identified 23 targets recognized by schistosome acute and chronic sera samples. To identify immunoreactive targets that were likely glycan epitopes, we compared these targets to the immunoreactivity of spots treated with sodium metaperiodate oxidation of egg extract. This treatment yielded 12/23 spots maintaining immunoreactivity, suggesting that they were protein epitopes. From these 12 spots, 11 spots cross-reacted with sera from individuals infected with other STH and 10 spots cross-reacted with the negative control group. Spot number 5 was exclusively immunoreactive with sera from S. mansoni-infected groups in native and deglycosylated conditions and corresponds to Major Egg Antigen (MEA). We expressed MEA as a recombinant protein and showed a similar recognition pattern to that of the native protein via western blot. IgG-ELISA gave a sensitivity of 87.10% and specificity of 89.09% represented by area under the ROC curve of 0.95. IgG-ELISA performed better than the conventional KK (2 slides), identifying 56/64 cases harboring 1–10 eggs per gram of feces that were undiagnosed by KK parasitological technique. Conclusions: The serological proteome approach was able to identify a new diagnostic candidate. The recombinant egg antigen provided good performance in IgG-ELISA to detect individuals with extreme low-intensity infections (1 egg per gram of feces). Therefore, the IgG-ELISA using this newly identified recombinant MEA can be a useful tool combined with other techniques in low-endemic areas to determine the true prevalence of schistosome infection that is underestimated by the KK method. Further, to overcome the complexity of ELISA in the field, a second generation of antibody-based rapid diagnostic tests (RDT) can be developed. [ABSTRACT FROM AUTHOR]

Laboratory confirmation of Zika virus (ZIKV) infection during pregnancy is challenging due to cross-reactivity with dengue virus (DENV) and limited knowledge about the kinetics of anti-Zika antibody responses during pregnancy. We described ZIKV and DENV serological markers and the maternal-fetal transfer of antibodies among mothers and neonates after the ZIKV microcephaly outbreak in Northeast Brazil (2016). We included 89 microcephaly cases and 173 neonate controls at time of birth and their mothers. Microcephaly cases were defined as newborns with a particular head circumference (2 SD below the mean). Two controls without microcephaly were matched by the expected date of delivery and area of residence. We tested maternal serum for recent (ZIKV genome, IgM and IgG3 anti-NS1) and previous (ZIKV and DENV neutralizing antibodies [NAbs]) markers of infection. Multiple markers of recent or previous ZIKV and DENV infection in mothers were analyzed using principal component analysis (PCA). At delivery, 5.6% of microcephaly case mothers and 1.7% of control mothers were positive for ZIKV IgM. Positivity for ZIKV IgG3 anti-NS1 was 8.0% for case mothers and 3.5% for control mothers. ZIKV NAbs was slightly higher among mothers of cases (69.6%) than that of mothers of controls (57.2%; p = 0.054). DENV exposure was detected in 85.8% of all mothers. PCA discriminated two distinct components related to recent or previous ZIKV infection and DENV exposure. ZIKV NAbs were higher in newborns than in their corresponding mothers (p<0.001). We detected a high frequency of ZIKV exposure among mothers after the first wave of the ZIKV outbreak in Northeast Brazil. However, we found low sensitivity of the serological markers to recent infection (IgM and IgG3 anti-NS1) in perinatal samples of mothers of microcephaly cases. Since the neutralization test cannot precisely determine the time of infection, testing for ZIKV immune status should be performed as early as possible and throughout pregnancy to monitor acute Zika infection in endemic areas. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) is a mosquito-borne positive sense RNA virus. Recently, ZIKV emerged into the Western hemisphere as a human health threat, with severe disease associated with developmental and neurological complications. The structural envelope protein of ZIKV and other neurotropic flaviviruses contains an extended CD-loop relative to non-neurotropic flaviviruses, and has been shown to augment ZIKV stability and pathogenesis. Here we show that shortening the CD-loop in ZIKV attenuates the virus in mice, by reducing the ability to invade and replicate in the central nervous system. The CD-loop mutation was genetically stable following infection in mice, though secondary site mutations arise adjacent to the CD-loop. Importantly, while shortening of the CD-loop attenuates the virus, the CD-loop mutant maintains antigenicity in immunocompetent mice, eliciting an antibody response that similarly neutralizes both the mutant and wildtype ZIKV. These findings suggest that the extended CD-loop in ZIKV is a determinant of neurotropism and may be a target in live-attenuated vaccine design, for not only ZIKV, but for other neurotropic flaviviruses. [ABSTRACT FROM AUTHOR]

Burkholderia pseudomallei causes melioidosis, a common source of pneumonia and sepsis in Southeast Asia and Northern Australia that results in high mortality rates. A caprine melioidosis model of aerosol infection that leads to a systemic infection has the potential to characterize the humoral immune response. This could help identify immunogenic proteins for new diagnostics and vaccine candidates. Outbred goats may more accurately mimic human infection, in contrast to the inbred mouse models used to date. B. pseudomallei infection was delivered as an intratracheal aerosol. Antigenic protein profiling was generated from the infecting strain MSHR511. Humoral immune responses were analyzed by ELISA and western blot, and the antigenic proteins were identified by mass spectrometry. Throughout the course of the infection the assay results demonstrated a much greater humoral response with IgG antibodies, in both breadth and quantity, compared to IgM antibodies. Pre-infection sera showed multiple immunogenic proteins already reactive for IgG (7–20) and IgM (0–12) in most of the goats despite no previous exposure to B. pseudomallei. After infection, the number of IgG reactive proteins showed a marked increase as the disease progressed. Early stage infection (day 7) showed immune reaction to chaperone proteins (GroEL, EF-Tu, and DnaK). These three proteins were detected in all serum samples after infection, with GroEL immunogenically dominant. Seven common reactive antigens were selected for further analysis using ELISA. The heat shock protein GroEL1 elicited the strongest goat antibody immune response compared to the other six antigens. Most of the six antigens showed the peak IgM reactivity at day 14, whereas the IgG reactivity increased further as the disease progressed. An overall MSHR511 proteomic comparison between the goat model and human sera showed that many immune reactive proteins are common between humans and goats with melioidosis. [ABSTRACT FROM AUTHOR]

Background: The burden of scrub typhus in endemic areas is poorly understood. This study aimed at estimating the proportion of hospitalisations and outpatient visits for undifferentiated fever in the community that may be attributable to scrub typhus. Methodology and principal findings: The study was a retrospective cohort with a nested case-control study conducted in the South Indian state of Tamil Nadu. We conducted house-to-house screening in 48 villages (42965 people, 11964 households) to identify hospitalised or outpatient cases due to undifferentiated fever during the preceding scrub typhus season. We used scrub typhus IgG to determine past infection. We calculated adjusted odds ratios for the association between IgG positivity and case status. Odds ratios were used to estimate population attributable fractions (PAF) indicating the proportion of hospitalised and outpatient fever cases attributable to scrub typhus. We identified 58 cases of hospitalisation and 236 outpatient treatments. 562 people were enrolled as control group to estimate the background IgG sero-prevalence. IgG prevalence was 20.3% in controls, 26.3% in outpatient cases and 43.1% in hospitalised cases. The PAFs suggested that 29.5% of hospitalisations and 6.1% of outpatient cases may have been due to scrub typhus. In villages with a high IgG prevalence (defined as ≥15% among controls), the corresponding PAFs were 43.4% for hospitalisations and 5.6% for outpatients. The estimated annual incidence of scrub typhus was 0.8/1000 people (0.3/1000 in low, and 1.3/1000 in high prevalence villages). Evidence for recall error suggested that the true incidences may be about twice as high as these figures. Conclusions: The study suggests scrub typhus as an important cause for febrile hospitalisations in the community. The results confirm the adequacy of empirical treatment for scrub typhus in hospitalised cases with undifferentiated fever. Since scrub typhus may be rare among stable outpatients, the use of empirical treatment remains doubtful in these. [ABSTRACT FROM AUTHOR]

Background: Mammarenaviruses are associated with human hemorrhagic fever diseases in Africa and America. Recently, a rodent mammarenavirus, Wēnzhōu virus (WENV) and related viruses, have been reported in China, Cambodia, and Thailand. Moreover, in Cambodia, these viruses were suspected to be associated with human disease. In China, Yunnan Province is famous for its abundant animal and plant diversity and is adjacent to several South-eastern Asia countries. Therefore, it is necessary to know whether WENV-related viruses, or other mammarenaviruses, are prevalent in this province. Methodology/Principal findings: Small mammals were trapped, euthanized, and sampled. Mammarenavirus RNA was detected using a nested reverse transcription polymerase chain reaction (RT-PCR) and quantified by real-time RT-PCR. A total of 1040 small mammals belonging to 13 genera and 26 species were trapped in Yunnan Province. WENV-related mammarenaviruses were detected in 41 rodent liver samples, mainly in brown rats (Rattus norvegicus) and oriental house rats (R. tanezumi).Viral nucleocapsid protein was detected in liver sections by indirect immunofluorescence assay. Full-length-genomes were amplified by RT-PCR and used for phylogenetic analysis with the MEGA package. Recombination analysis was performed using the SimPlot and Recombination Detection Program. Conclusions/Significance: WENV related viruses circulated in small mammals in Yunnan Province. Whole genome sequence analysis of five selected viral strains showed that these viruses are closely related to WENVs discovered in Asia and form an independent branch in the phylogenetic tree in the WENV clade. Paying attention to investigate the influence of these viruses to public health is essential in the epidemic regions. [ABSTRACT FROM AUTHOR]

Background: The Plasmodium vivax Duffy Binding Protein (PvDBP) is a key target of naturally acquired immunity. However, region II of PvDBP, which contains the receptor-binding site, is highly polymorphic. The natural acquisition of antibodies to different variants of PvDBP region II (PvDBPII), including the AH, O, P and Sal1 alleles, the central region III-V (PvDBPIII-V), and P. vivax Erythrocyte Binding Protein region II (PvEBPII) and their associations with risk of clinical P. vivax malaria are not well understood. Methodology: Total IgG and IgG subclasses 1, 2, and 3 that recognize four alleles of PvDBPII (AH, O, P, and Sal1), PvDBPIII-V and PvEBPII were measured in samples collected from a cohort of 1 to 3 year old Papua New Guinean (PNG) children living in a highly endemic area of PNG. The levels of binding inhibitory antibodies (BIAbs) to PvDBPII (AH, O, and Sal1) were also tested in a subset of children. The association of presence of IgG with age, cumulative exposure (measured as the product of age and malaria infections during follow-up) and prospective risk of clinical malaria were evaluated. Results: The increase in antigen-specific total IgG, IgG1, and IgG3 with age and cumulative exposure was only observed for PvDBPII AH and PvEBPII. High levels of total IgG and predominant subclass IgG3 specific for PvDBPII AH were associated with decreased incidence of clinical P. vivax episodes (aIRR = 0.56–0.68, P≤0.001–0.021). High levels of total IgG and IgG1 to PvEBPII correlated strongly with protection against clinical vivax malaria compared with IgGs against all PvDBPII variants (aIRR = 0.38, P<0.001). Antibodies to PvDBPII AH and PvEBPII showed evidence of an additive effect, with a joint protective association of 70%. Conclusion: Antibodies to the key parasite invasion ligands PvDBPII and PvEBPII are good correlates of protection against P. vivax malaria in PNG. This further strengthens the rationale for inclusion of PvDBPII in a recombinant subunit vaccine for P. vivax malaria and highlights the need for further functional studies to determine the potential of PvEBPII as a component of a subunit vaccine for P. vivax malaria. [ABSTRACT FROM AUTHOR]

Mayaro virus (MAYV) of the genus alphavirus is a mosquito-transmitted emerging infectious disease that causes an acute febrile illness, rash, headaches, and nausea that may turn into incapacitating, persistent arthralgias in some victims. Since its discovery in Trinidad in 1954, cases of MAYV infection have largely been confined there and to the northern countries of South America, but recently, MAYV cases have been reported in some island nations in the Caribbean Sea. Accompanying these reports is evidence that new vectors, including Aedes spp. mosquitos, recently implicated in the global spread of Zika and chikungunya viruses, are competent for MAYV transmission, which, if true, could facilitate the spread of MAYV beyond its current range. Despite its status as an emerging virus, there are no licensed vaccines to prevent MAYV infection nor therapeutics to treat it. Here, we describe the development and testing of a novel DNA vaccine, scMAYV-E, that encodes a synthetically-designed consensus MAYV envelope sequence. In vivo electroporation-enhanced immunization of mice with this vaccine induced potent humoral responses including neutralizing antibodies as well as robust T-cell responses to multiple epitopes in the MAYV envelope. Importantly, these scMAYV-E-induced immune responses protected susceptible mice from morbidity and mortality following a MAYV challenge. [ABSTRACT FROM AUTHOR]

The bivalent killed whole-cell oral cholera vaccine (BivWC) is being increasingly used to prevent cholera. The presence of O-antigen-specific memory B cells (MBC) has been associated with protective immunity against cholera, yet MBC responses have not been evaluated after BivWC vaccination. To address this knowledge gap, we measured V. cholerae O1-antigen MBC responses following BivWC vaccination. Adults in St. Marc, Haiti, received 2 doses of the BivWC vaccine, Shanchol, two weeks apart. Participants were invited to return at days 7, 21, 44, 90, 180 and 360 after the initial vaccination. Serum antibody and MBC responses were assessed at each time-point before and following vaccination. We observed that vaccination with BivWC resulted in significant O-antigen specific MBC responses to both Ogawa and Inaba serotypes that were detected by day 21 and remained significantly elevated over baseline for up to 12 months following vaccination. The BivWC oral cholera vaccine induces durable MBC responses to the V. cholerae O1-antigen. This suggests that long-term protection observed following vaccination with BivWC could be mediated or maintained by MBC responses. [ABSTRACT FROM AUTHOR]

Background: Leprosy is a treatable infectious disease caused by Mycobacterium leprae. However, there is additional morbidity from leprosy-associated pathologic immune reactions, reversal reaction (RR) and erythema nodosum leprosum (ENL), which occur in 1 in 3 people with leprosy, even with effective treatment of M. leprae. There is currently no predictive marker in use to indicate which people with leprosy will develop these debilitating immune reactions. Our peripheral blood mononuclear cell (PBMC) transcriptome analysis revealed that activation of the classical complement pathway is common to both RR and ENL. Additionally, differential expression of immunoglobulin receptors and B cell receptors during RR and ENL support a role for the antibody-mediated immune response during both RR and ENL. In this study, we investigated B-cell immunophenotypes, total and M. leprae-specific antibodies, and complement levels in leprosy patients with and without RR or ENL. The objective was to determine the role of these immune mediators in pathogenesis and assess their potential as biomarkers of risk for immune reactions in people with leprosy. Methodology/findings: We followed newly diagnosed multibacillary leprosy cases (n = 96) for two years for development of RR or ENL. They were compared with active RR (n = 35), active ENL (n = 29), and healthy household contacts (n = 14). People with leprosy who subsequently developed ENL had increased IgM, IgG1, and C3d-associated immune complexes with decreased complement 4 (C4) at leprosy diagnosis. People who developed RR also had decreased C4 at leprosy diagnosis. Additionally, elevated anti-M. leprae antibody levels were associated with subsequent RR or ENL. Conclusions: Differential co-receptor expression and immunoglobulin levels before and during immune reactions intimate a central role for humoral immunity in RR and ENL. Decreased C4 and elevated anti-M. leprae antibodies in people with new diagnosis of leprosy may be risk factors for subsequent development of leprosy immune reactions. [ABSTRACT FROM AUTHOR]

Background: Hosts repeatedly bitten by sand flies develop antibodies against sand fly saliva and screening of these immunoglobulins can be employed to estimate the risk of Leishmania transmission, to indicate the feeding preferences of sand flies, or to evaluate the effectiveness of vector control campaigns. Previously, antibodies to sand fly saliva were detected using whole salivary gland homogenate (SGH) or recombinant proteins, both of which also have their disadvantages. This is the first study on sand flies where short peptides designed based on salivary antigens were successfully utilized for antibody screening. Methodology/Principal findings: Specific IgG was studied in hosts naturally exposed to Phlebotomus orientalis, the main vector of Leishmania donovani in East Africa. Four peptides were designed by the commercial program EpiQuest-B, based on the sequences of the two most promising salivary antigens, yellow-related protein and ParSP25-like protein. Short amino acid peptides were synthesised and modified for ELISA experiments. Specific anti-P. orientalis IgG was detected in sera of dogs, goats, and sheep from Ethiopia. The peptide OR24 P2 was shown to be suitable for antibody screening; it correlated positively with SGH and its specificity and sensitivity were comparable or even better than that of previously published recombinant proteins. Conclusions/Significance: OR24 P2, the peptide based on salivary antigen of P. orientalis, was shown to be a valuable tool for antibody screening of domestic animals naturally exposed to P. orientalis. We suggest the application of this promising methodology using species-specific short peptides to other sand fly-host combinations. [ABSTRACT FROM AUTHOR]

Background: Validation of elimination of trachoma as a public health problem is based on clinical indicators, using the WHO simplified grading system. Chlamydia trachomatis (Ct) infection and anti-Ct antibody responses (anti-Pgp3) have both been evaluated as alternative indicators in settings with varying levels of trachoma. There is a need to evaluate the feasibility of using tests for Ct infection and anti-Pgp3 antibodies at scale in a trachoma-endemic country and to establish the added value of the data generated for understanding transmission dynamics in the peri-elimination setting. Methodology/Principal findings: Dried blood spots for serological testing and ocular swabs for Ct infection testing (taken from children aged 1–9 years) were integrated into the pre-validation trachoma surveys conducted in the Northern and Upper West regions of Ghana in 2015 and 2016. Ct infection was detected using the GeneXpert PCR platform and the presence of anti-Pgp3 antibodies was detected using both the ELISA assay and multiplex bead array (MBA). The overall mean cluster-summarised TF prevalence (the clinical indicator) was 0.8% (95% CI: 0.6–1.0) and Ct infection prevalence was 0.04% (95%CI: 0.00–0.12). Anti-Pgp3 seroprevalence using the ELISA was 5.5% (95% CI: 4.8–6.3) compared to 4.3% (95%CI: 3.7–4.9) using the MBA. There was strong evidence from both assays that seropositivity increased with age (p<0.001), although the seroconversion rate was estimated to be very low (between 1.2 to 1.3 yearly events per 100 children). Conclusions/Significance: Infection and serological data provide useful information to aid in understanding Ct transmission dynamics. Elimination of trachoma as a public health problem does not equate to the absence of ocular Ct infection nor cessation in acquisition of anti-Ct antibodies. [ABSTRACT FROM AUTHOR]

Background: We examined relationships between schistosome infection, HIV transmission or acquisition, and all-cause death. Methods: We retrospectively tested baseline sera from a heterosexual HIV-discordant couple cohort in Lusaka, Zambia with follow-up from 1994–2012 in a nested case-control design. Schistosome-specific antibody levels were measured by ELISA. Associations between baseline antibody response to schistosome antigens and incident HIV transmission, acquisition, and all-cause death stratified by gender and HIV status were assessed. In a subset of HIV- women and HIV+ men, we performed immunoblots to evaluate associations between Schistosoma haematobium or Schistosoma mansoni infection history and HIV incidence. Results: Of 2,145 individuals, 59% had positive baseline schistosome-specific antibody responses. In HIV+ women and men, baseline schistosome-specific antibodies were associated with HIV transmission to partners (adjusted hazard ratio [aHR] = 1.8, p<0.005 and aHR = 1.4, p<0.05, respectively) and death in HIV+ women (aHR = 2.2, p<0.001). In 250 HIV- women, presence of S. haematobium-specific antibodies was associated with increased risk of HIV acquisition (aHR = 1.4, p<0.05). Conclusion: Schistosome infections were associated with increased transmission of HIV from both sexes, acquisition of HIV in women, and increased progression to death in HIV+ women. Establishing effective prevention and treatment strategies for schistosomiasis, including in urban adults, may reduce HIV incidence and death in HIV+ persons living in endemic areas. [ABSTRACT FROM AUTHOR]

Background: Age patterns in asymptomatic and symptomatic infection with Leishmania donovani, the causative agent of visceral leishmaniasis (VL) in the Indian subcontinent (ISC), are currently poorly understood. Age-stratified serology and infection incidence have been used to assess transmission levels of other diseases, which suggests that they may also be of use for monitoring and targeting control programmes to achieve elimination of VL and should be included in VL transmission dynamic models. We therefore analysed available age-stratified data on both disease incidence and prevalence of immune markers with the aim of collating the currently available data, estimating rates of infection, and informing modelling and future data collection. Methodology/Principal findings: A systematic literature search yielded 13 infection prevalence and 7 VL incidence studies meeting the inclusion criteria. Statistical tests were performed to identify trends by age, and according to diagnostic cut-off. Simple reversible catalytic models with age-independent and age-dependent infection rates were fitted to the prevalence data to estimate infection and reversion rates, and to test different hypotheses about the origin of variation in these rates. Most of the studies showed an increase in infection prevalence with age: from ≲10% seroprevalence (<20% Leishmanin skin test (LST) positivity) for 0-10-year-olds to >10% seroprevalence (>20% LST-positivity) for 30-40-year-olds, but overall prevalence varied considerably between studies. VL incidence was lower amongst 0-5-year-olds than older age groups in most studies; most showing a peak in incidence between ages 5 and 20. The age-independent catalytic model provided the best overall fit to the infection prevalence data, but the estimated rates for the less parsimonious age-dependent model were much closer to estimates from longitudinal studies, suggesting that infection rates may increase with age. Conclusions/Significance: Age patterns in asymptomatic infection prevalence and VL incidence in the ISC vary considerably with geographical location and time period. The increase in infection prevalence with age and peaked age-VL-incidence distribution may be due to lower exposure to infectious sandfly bites in young children, but also suggest that acquired immunity to the parasite increases with age. However, poor standardisation of serological tests makes it difficult to compare data from different studies and draw firm conclusions about drivers of variation in observed age patterns. [ABSTRACT FROM AUTHOR]

Background: The severity of cardiac disease in chronic Chagas disease patients is associated with different features of T-cell exhaustion. Here, we assessed whether the ability of T cells to secrete IFN-γ in response to T. cruzi was linked to disruption in immune homeostasis and inflammation in patients with chronic Chagas disease. Methodology/Principal findings: PBMCs from chronic Chagas disease patients and uninfected controls were examined for frequencies of T. cruzi-responsive IFN-γ-producing cells by ELISPOT and cellular expression and function of IL-7R using flow cytometry. Serum levels of IL-7, IL-21, IL-27, soluble IL-7R, and inflammatory cytokines were also evaluated by ELISA or CBA techniques. Patients possessing T. cruzi-specific IFN-γ-producing cells (i.e. IFN-γ producers) had higher levels of memory T cells capable of modulating the alpha chain of IL-7R and an efficient response to IL-7 compared to that in patients lacking (i.e. IFN-γ nonproducers) parasite-specific T-cell responses. IFN-γ producers also showed low levels of soluble IL-7R, high basal expression of Bcl-2 in T cells and low basal frequencies of activated CD25+ T cells. Modulation of IL-7R was inversely associated with serum IL-6 levels and positively associated with serum IL-8 levels. Circulating IL-21 and IL-27 levels were not associated with the frequency of IFN-γ producing cells but were reduced in less severe clinical forms of the disease. In vitro stimulation of PBMCs with IL-7 or IL-27 enhanced IFN-γ production in IFN-γ producers but not in IFN-γ nonproducers. Conclusions/Significance: Alterations of the IL-7/IL-7R axis and in the levels of inflammatory cytokines were linked to impaired T. cruzi-specific IFN-γ production. These alterations might be responsible of the process of immune exhaustion observed in chronic Chagas disease. [ABSTRACT FROM AUTHOR]

Taiwan is an island located in the south Pacific, a subtropical region that is home to 61 species of snakes. Of these snakes, four species—Trimeresurus stejnegeri, Protobothrops mucrosquamatus, Bungarus multicinctus and Naja atra—account for more than 90% of clinical envenomation cases. Currently, there are two types of bivalent antivenom: hemorrhagic antivenom against the venom of T. stejnegeri and P. mucrosquamatus, and neurotoxic antivenom for treatment of envenomation by B. multicinctus and N. atra. However, no suitable detection kits are available to precisely guide physicians in the use of antivenoms. Here, we sought to develop diagnostic assays for improving the clinical management of snakebite in Taiwan. A two-step affinity purification procedure was used to generate neurotoxic species-specific antibodies (NSS-Abs) and hemorrhagic species-specific antibodies (HSS-Abs) from antivenoms. These two SSAbs were then used to develop a sandwich ELISA (enzyme-linked immunosorbent assay) and a lateral flow assay comprising two test lines. The resulting ELISAs and lateral flow strip assays could successfully discriminate between neurotoxic and hemorrhagic venoms. The limits of quantification (LOQ) of the ELISA for neurotoxic venoms and hemorrhagic venoms were determined to be 0.39 and 0.78 ng/ml, respectively, and the lateral flow strips were capable of detecting neurotoxic and hemorrhagic venoms at concentrations lower than 5 and 50 ng/ml, respectively, in 10–15 min. Tests of lateral flow strips in 21 clinical snakebite cases showed 100% specificity and 100% sensitivity for neurotoxic envenomation, whereas the sensitivity for detecting hemorrhagic envenomation samples was 36.4%. We herein presented a feasible strategy for developing a sensitive sandwich ELISA and lateral flow strip assay for detecting and differentiating venom proteins from hemorrhagic and neurotoxic snakes. A useful snakebite diagnostic guideline according to the lateral flow strip results and clinical symptoms was proposed to help physicians to use antivenoms appropriately. The two-test-line lateral flow strip assay could potentially be applied in an emergency room setting to help physicians diagnose and manage snakebite victims. [ABSTRACT FROM AUTHOR]

The Global Program to Eliminate Lymphatic Filariasis (LF) relies on rapid diagnostic tests (RDTs) to determine where annual mass drug administration for LF is required and when it can be stopped. These tests detect a Wuchereria bancrofti glycoprotein in the blood of infected persons via a carbohydrate moiety recognized by the monoclonal antibodies AD12 and DH6.5. Loiasis cross-reactivity with LF RDTs has recently been recognized as a serious obstacle to LF elimination in loiasis-endemic areas. To better understand the nature of this cross-reactivity, we used the DH6.5 antibody to immunoaffinity purify Loa loa antigens from the sera of individuals with a positive RDT due to loiasis. Immunoblot analysis revealed many circulating AD12/DH6.5-reactive antigens, and proteomic analysis identified multiple L. loa proteins in LF RDT-positive loiasis sera. These included both secreted and somatic proteins, suggesting that they may be released by dying L. loa adult worms and/or microfilariae. Unlike the single high molecular weight W. bancrofti circulating filarial antigen that is reliably present in the blood of persons with bancroftian filariasis, reactive L. loa antigens appeared to be only transiently present in the blood of a subset of persons with loiasis. These key differences between the circulating antigens of W. bancrofti and L. loa can be used to differentiate positive results generated by both species and may lead to improved diagnostic tests for LF and loiasis. [ABSTRACT FROM AUTHOR]

Background: Tanzania is among the Rift Valley fever (RVF) epizootic/endemic countries in sub Saharan Africa, where RVF disease outbreaks occur within a range of 3 to 17-year intervals. Detection of Rift Valley fever virus (RVFV) antibodies in animals in regions with no previous history of outbreaks raises the question of whether the disease is overlooked due to lack-of effective surveillance systems, or if there are strains of RVFV with low pathogenicity. Furthermore, which vertebrate hosts are involved in the inter-epidemic and inter-epizootic maintenance of RVFV? In our study region, the Kyela and Morogoro districts in Tanzania, no previous RVF outbreaks have been reported. Methodology: The study was conducted from June 2014 to October 2015 in the Kyela and Morogoro districts, Tanzania. Samples (n = 356) were retrieved from both the local breed of zebu cattle (Bos indicus) and Bos indicus/Bos Taurus cross breed. RVFV antibodies were analyzed by two enzyme-linked immunosorbent assay (ELISA) approaches. Initially, samples were analyzed by a RVFV multi-species competition ELISA (cELISA), which detected both RVFV IgG and IgM antibodies. All serum samples that were positive with the cELISA method were specifically analysed for the presence of RVFV IgM antibodies to trace recent infection. A plaque reduction neutralization assay (PRNT80) was performed to determine presence of RVFV neutralizing antibodies in all cELISA positive samples. Findings: Overall RVFV seroprevalence rate in cattle by cELISA in both districts was 29.2% (104 of 356) with seroprevalence rates of 33% (47/147) in the Kyela district and 27% (57/209) in the Morogoro district. In total, 8.4% (30/356) of all cattle sampled had RVFV IgM antibodies, indicating current disease transmission. When segregated by districts, the IgM antibody seroprevalence was 2.0% (3/147) and 12.9% (27/209) in Kyela and Morogoro districts respectively. When the 104 cELISA positive samples were analyzed by PRNT80 to confirm that RVFV-specific antibodies were present, the majority (89%, 93/104) had RVFV neutralising antibodies. Conclusion: The results provided evidence of widespread prevalence of RVFV antibody among cattle during an inter-epizootic/inter-epidemic period in Tanzania in regions with no previous history of outbreaks. There is a need for further investigations of RVFV maintenance and transmission in vertebrates and vectors during the long inter-epizootic/inter-epidemic periods. [ABSTRACT FROM AUTHOR]

Background: Estimates of current global rabies mortality range from 26,000 to 59,000 deaths per annum. Although pre-exposure prophylaxis using inactivated rabies virus vaccines (IRVs) is effective, it requires two to three doses and is regarded as being too expensive and impractical for inclusion in routine childhood immunization programmes. Methodology/ Principal findings: Here we report the development of a simian-adenovirus-vectored rabies vaccine intended to enable cost-effective population-wide pre-exposure prophylaxis against rabies. ChAdOx2 RabG uses the chimpanzee adenovirus serotype 68 (AdC68) backbone previously shown to achieve pre-exposure protection against rabies in non-human primates. ChAdOx2 differs from AdC68 in that it contains the human adenovirus serotype 5 (AdHu5) E4 orf6/7 region in place of the AdC68 equivalents, enhancing ease of manufacturing in cell lines which provide AdHu5 E1 proteins in trans. We show that immunogenicity of ChAdOx2 RabG in mice is comparable to that of AdC68 RabG and other adenovirus serotypes expressing rabies virus glycoprotein. High titers of rabies virus neutralizing antibody (VNA) are elicited after a single dose. The relationship between levels of VNA activity and rabies virus glycoprotein monomer-binding antibody differs after immunization with adenovirus-vectored vaccines and IRV vaccines, suggesting routes to further enhancement of the efficacy of the adenovirus-vectored candidates. We also demonstrate that ChAdOx2 RabG can be thermostabilised using a low-cost method suitable for clinical bio-manufacture and ambient-temperature distribution in tropical climates. Finally, we show that a dose-sparing effect can be achieved by formulating ChAdOx2 RabG with a simple chemical adjuvant. This approach could lower the cost of ChAdOx2 RabG and other adenovirus-vectored vaccines. Conclusions/ Significance: ChAdOx2 RabG may prove to be a useful tool to reduce the human rabies death toll. We have secured funding for Good Manufacturing Practice- compliant bio-manufacture and Phase I clinical trial of this candidate. [ABSTRACT FROM AUTHOR]

The gold-standard method for quantifying neutralizing antibody responses to many viruses, including dengue virus (DENV), is the plaque reduction neutralization test (PRNT, also called the immunofocus reduction neutralization test). The PRNT conducted on 96-well plates is high-throughput and requires a smaller volume of antiserum than on 6- or 24-well plates, but manual plaque counting is challenging and existing automated plaque counters are expensive or difficult to optimize. We have developed Viridot (Viridot package), a program for R with a user interface in shiny, that counts viral plaques of a variety of phenotypes, estimates neutralizing antibody titers, and performs other calculations of use to virologists. The Viridot plaque counter includes an automatic parameter identification mode (misses <10 plaques/well for 87% of diverse DENV strains [n = 1521]) and a mode that allows the user to fine-tune the parameters used for counting plaques. We compared standardized manual and Viridot plaque counting methods applied to the same wells by two analyses and found that Viridot plaque counts were as similar to the same analyst's manual count (Lin’s concordance correlation coefficient, ρc = 0.99 [95% confidence interval: 0.99–1.00]) as manual counts between analysts (ρc = 0.99 [95% CI: 0.98–0.99]). The average ratio of neutralizing antibody titers based on manual counted plaques to Viridot counted plaques was 1.05 (95% CI: 0.98–1.14), similar to the average ratio of antibody titers based on manual plaque counts by the two analysts (1.06 [95% CI: 0.84–1.34]). Across diverse DENV and ZIKV strains (n = 14), manual and Viridot plaque counts were mostly consistent (range of ρc = 0.74 to 1.00) and the average ratio of antibody titers based on manual and Viridot counted plaques was close to 1 (0.94 [0.86–1.02]). Thus, Viridot can be used for plaque counting and neutralizing antibody titer estimation of diverse DENV strains and potentially other viruses on 96-well plates as well as for formalization of plaque-counting rules for standardization across experiments and analysts. [ABSTRACT FROM AUTHOR]

Cystic echinococcosis (CE) is one of the most widespread helminthic zoonoses and is caused by the tapeworm Echinococcus granulosus complex. CE diagnosis and monitoring primarily rely on imaging techniques, complemented by serology. This is usually approached by the detection of IgG antibodies against hydatid fluid (HF), but the use of this heterogeneous antigenic mixture results in a variable percentage of false positive and negative results, and has shown to be useless for follow-up due to the long persistence of anti-HF antibodies in cured patients. To improve test performances and standardization, a number of recombinant antigens mainly derived from HF have been described, among them the B2t and 2B2t antigens. The performance of these antigens in the diagnosis and follow up of patients with CE has been so far evaluated on a limited number of samples. Here, we evaluated the performances of tests based on B2t and 2B2t recombinant antigens compared to HF in IgG-ELISA and immunochromatography (IC) for the diagnosis and follow-up of patients with CE in a retrospective cohort study. A total of 721 serum samples were collected: 587 from 253 patients with CE diagnosed by ultrasonography (US), 42 from patients with alveolar echinococcosis and 92 from healthy donors from Salamanca (Spain). The highest overall sensitivity was obtained with HF in ELISA (85.5%), followed by IC containing HF and 2B2t-HF (83.0% and 78.2%, respectively). The lowest sensitivity was obtained with B2t and 2B2t in ELISA (51.8%). The highest specificity was obtained with IC containing 2B2t-HF (100%), and the lowest with HF-ELISA (78.0%). The lowest cross-reactivity with sera from patients with alveolar echinococcosis was detected with the recombinant antigens in ELISA (9.5% - 16.7%) and the highest with the HF-IC (64.3%). The results of B2t and 2B2t-ELISA were influenced by cyst stage, as classified by US according to the WHO-Informal Working Group on Echinococcosis (WHO-IWGE), with low sensitivity for inactive (CE4 and CE5) cysts, and by the drug treatment, with higher sensitivity in patients after drug treatment compared with patients not subjected to drug treatment. The two recombinant antigens in ELISA provided promising results for monitoring patients in follow-up, although their use is limited to patients with positive serology against them at the beginning of the follow-up. Potential biological reasons behind the low sensitivity of the recombinant antigens and possible strategies to enhance the performance of CE serology are discussed. [ABSTRACT FROM AUTHOR]

Japanese encephalitis is mainly considered a rural disease, but there is growing evidence of a peri-urban and urban transmission in several countries, including Cambodia. We, therefore, compared the epidemiologic dynamic of Japanese encephalitis between a rural and a peri-urban setting in Cambodia. We monitored two cohorts of 15 pigs and determined the force of infection–rate at which seronegative pigs become positive–in two study farms located in a peri-urban and rural area, respectively. We also studied the mosquito abundance and diversity in proximity of the pigs, as well as the host densities in both areas. All the pigs seroconverted before the age of 6 months. The force of infection was 0.061 per day (95% confidence interval = 0.034–0.098) in the peri-urban cohort and 0.069 per day (95% confidence interval = 0.047–0.099) in the rural cohort. Several differences in the epidemiologic dynamic of Japanese encephalitis between both study sites were highlighted. The later virus amplification in the rural cohort may be linked to the later waning of maternal antibodies, but also to the higher pig density in direct proximity of the studied pigs, which could have led to a dilution of mosquito bites at the farm level. The force of infection was almost identical in both the peri-urban and the rural farms studied, which shifts the classic epidemiologic cycle of the virus. This study is a first step in improving our understanding of Japanese encephalitis virus ecology in different environments with distinct landscapes, human and animal densities. [ABSTRACT FROM AUTHOR]

Background: In India, dengue disease is emerging as the most important vector borne public health problem due to rapid and unplanned urbanization, high human density and week management of the disease. Clinical cases are grossly underreported and not much information is available on prevalence and incidence of the disease. Methodology: A cross sectional, stratified, facility based, multistage cluster sampling was conducted between May 4 and June 27, 2017 in Pune city. A total of 1,434 participants were enrolled. The serum samples were tested for detection of historical dengue IgG antibodies by ELISA using the commercial Panbio Dengue IgG Indirect ELISA kit. Anti-dengue IgG-capture Panbio ELISA was used for detection of high titered antibodies to detect recent secondary infection. We used this data to estimate key transmission parameters like force of infection and basic reproductive number. A subset of 120 indirect ELISA positive samples was also tested for Plaque Reduction Neutralizing Antibodies for determining serotype-specific prevalence. Findings: Overall, 81% participants were infected with dengue virus (DENV) at least once if not more. The positivity was significantly different in different age groups. All the adults above 70 years were positive for DENV antibodies. Over 69% participants were positive for neutralizing antibodies against all 4 serotypes suggesting intense transmission of all DENV serotypes in Pune. Age-specific seroprevalence was consistent with long-term, endemic circulation of DENV. There was an increasing trend with age, from 21.6% among <36 months to 59.4% in age group 10–12 years. We estimate that 8.68% of the susceptible population gets infected by DENV each year resulting into more than 3,00,000 infections and about 47,000 to 59,000 cases per year. This transmission intensity is similar to that reported from other known hyper-endemic settings in Southeast Asia and the Americas but significantly lower than report from Chennai. Conclusions: Our study suggests that Pune city has high disease burden, all 4 serotypes are circulating, significant spatial heterogeneity in seroprevalence and suboptimal immunity in younger age groups. This would allow informed decisions to be made on management of dengue and introduction of upcoming dengue vaccines in the city. [ABSTRACT FROM AUTHOR]

Background: Canine leishmaniasis (CanL) is a zoonotic disease, caused by Leishmania infantum and transmitted by Phlebotomus perniciosus in the Mediterranean basin. Previously, an ELISA based on the P. perniciosus salivary protein SP03B was proposed as a valid tool to screen for canine exposure to sand fly bites across regions endemic for CanL. Although this approach is useful in laboratory settings, a practical tool for immediate application in the field is needed. In this study we propose the rSP03B sero-strip, the first immunochromatographic test (ICT) in the field of vector exposure able to rapidly screen dogs living in endemic areas for the presence of P. perniciosus and to aid in the evaluation of vector control programs. Methodology/Principal findings: The ICT was prepared using the bacterially expressed recombinant protein rSP03B as antigen. For test optimization, pre-immune sera from non-bitten laboratory-bred Beagles were used as negative controls. In order to validate the test, sera from laboratory-bred Beagles experimentally exposed to P. perniciosus bites were used as positive controls. Additionally, all samples were tested by ELISA using whole salivary gland homogenate (SGH) and the rSP03B protein as antigen. An almost perfect degree of agreement was found between the ICT and the SGH-ELISA. Furthermore, the newly proposed rSP03B sero-strip showed a sensitivity of 100% and a specificity of 86.79%. Conclusions/Significance: We developed a simple and rapid ICT based on the P. perniciosus rSP03B salivary protein, able to replace the standard ELISA used in previous studies. Our rSP03B sero-strip showed to be highly sensitive and specific in the detection of antibodies (IgG) against P. perniciosus saliva. In the future, this test can be employed during large-scale epidemiological studies of CanL in the Mediterranean area to evaluate the efficacy of vector control programs. [ABSTRACT FROM AUTHOR]

Background: Chikungunya virus (CHIKV) is the most common alphavirus infecting humans worldwide, causing acute and chronically debilitating arthralgia at a great economic expense. Methodology/Principal findings: To facilitate our study of CHIKV, we generated a mCherry tagged replication-competent chimeric virus, CHIKV 37997-mCherry. Single particle cryoEM demonstrated icosahedral organization of the chimeric virus and the display of mCherry proteins on virus surface. CHIKV 37997-mCherry is attenuated in both IFNαR knockout and wild-type mice. Strong anti-CHIKV and anti-mCherry antibody responses were induced in CHIKV 37997-mCherry infected mice. Conclusions/Significance: Our work suggests that chimeric alphaviruses displaying foreign antigen can serve as vaccines against both aphaviruses and other pathogens and diseases. [ABSTRACT FROM AUTHOR]

Background: Aedes mosquitoes severely affect the health and wellbeing of human populations by transmitting infectious diseases. In French Polynesia, Aedes aegypti is the main vector of dengue, chikungunya and Zika, and Aedes polynesiensis the primary vector of Bancroftian filariasis and a secondary vector of arboviruses. Tools for assessing the risk of disease transmission or for measuring the efficacy of vector control programmes are scarce. A promising approach to quantify the human-vector contact relies on the detection and the quantification of antibodies directed against mosquito salivary proteins. Methodology/Principal findings: An ELISA test was developed to detect and quantify the presence of immunoglobulin G (IgG) directed against proteins from salivary gland extracts (SGE) of Ae. aegypti and Ae. polynesiensis in human populations exposed to either species, through a cross-sectional study. In Tahiti and Moorea islands where Ae. aegypti and Ae. polynesiensis are present, the test revealed that 98% and 68% of individuals have developed IgG directed against Ae. aegypti and Ae. polynesiensis SGE, respectively. By comparison, ELISA tests conducted on a cohort of people from metropolitan France, not exposed to these Aedes mosquitoes, indicated that 97% of individuals had no IgG directed against SGE of either mosquito species. The analysis of additional cohorts representing different entomological Aedes contexts showed no ELISA IgG cross-reactivity between Ae. aegypti and Ae. polynesiensis SGE. Conclusions/Significance: The IgG response to salivary gland extracts seems to be a valid and specific biomarker of human exposure to the bites of Ae. aegypti and Ae. polynesiensis. This new immuno-epidemiological tool will enhance our understanding of people exposure to mosquito bites, facilitate the identification of areas where disease transmission risk is high and permit to evaluate the efficacy of novel vector control strategies in Pacific islands and other tropical settings. [ABSTRACT FROM AUTHOR]

Objective: To gain further insight on the association between human toxocariasis and epilepsy in light of the new evidence in the last years. Methods: A systematic review was conducted without date and language restriction in the following electronic databases: MEDLINE (PubMed), Ingenta Connect, Science Direct (Elsevier), RefDoc, Scopus, HighWire, Scielo and the database of the Institute of Neuroepidemiology and Tropical Neurology of the Limoges University (IENT). Two investigators independently conducted the search up to November 2017. A pooled odds ratio (OR) was estimated using a random effects model. Meta-regression was conducted to investigate potential sources of heterogeneity. Results: Database search produced 204 publications. Eleven case-control studies were included that were carried out in 13 countries worldwide. A total number of 4740 subjects were considered (2159 people with epilepsy and 2581 people without epilepsy). The overall pooled OR was 1.69 (95% CI 1.42–2.01) for the association between epilepsy and Toxocara spp. seropositivity. A positive association was constantly reported in the restricted analysis (WB as confirmatory or diagnostic test, younger population, and population–based studies). Meta-regression showed no statistically significant association between covariates and outcome. Conclusion: The updated meta-analysis provides epidemiological evidence of a positive association between Toxocara seropositivity and epilepsy. New surveys supported the association, mainly population-based studies. On this basis, health strategies to reduce the impact of Toxocara spp are strongly advised. Further research should be performed to understand the physiopathological mechanisms of toxocara-associated epileptogenesis. [ABSTRACT FROM AUTHOR]

Crimean-Congo hemorrhagic fever virus (CCHFV) causes severe acute human disease with lethal outcome. The knowledge about the immune response for this human health threat is highly limited. In this study, we have screened the glycoprotein of CCHFV for novel linear B-cell epitopic regions using a microarray approach. The peptide library consisted of 168 synthesized 20mer peptides with 10 amino acid overlap covering the entire glycoprotein. Using both pooled and individual human sera from survivors of CCHF disease in Turkey five peptide epitopes situated in the mucin-like region and GP 38 (G15-515) and GN G516-1037 region of the glycoprotein were identified as epitopes for a CCHF immune response. An epitope walk of the five peptides revealed a peptide sequence located in the GN region with high specificity and sensitivity. This peptide sequence, and a sequence downstream, reacted also against sera from survivors of CCHF disease in South Africa. The cross reactivity of these peptides with samples from a geographically distinct region where genetically diverse strains of the virus circulate, enabled the identification of unique peptide epitopes from the CCHF glycoprotein that could have application in development of diagnostic tools. In this study clinical samples from geographically distinct regions were used to identify conserved linear epitopic regions of the glycoprotein of CCHF. [ABSTRACT FROM AUTHOR]

Background: Dengue is a febrile illness transmitted by mosquitoes, causing disease across the tropical and sub-tropical world. Antibody prevalence data and serotype distributions describe population-level risk and inform public health decision-making. Methodology/Principal findings: In this cross-sectional study we used data from a pediatric dengue seroprevalence study to describe historical dengue serotype circulation, according to age and geographic location. A sub-sample of 780 dengue IgG-positive sera, collected from 30 sites across urban Indonesia in 2014, were tested by the plaque reduction neutralization test (PRNT) to measure the prevalence and concentration of serotype-specific neutralizing antibodies according to subject age and geography. PRNT results were obtained from 776 subjects with mean age of 9.6 years. 765 (98.6%) neutralized one or more dengue serotype at a threshold of >10 (1/dil). Multitypic profiles were observed in 50.9% of the samples; a proportion which increased to 63.1% in subjects aged 15–18 years. Amongst monotypic samples, the highest proportion was reactive against DENV-2, followed by DENV-1, and DENV-3, with some variation across the country. DENV-4 was the least common serotype. The highest anti-dengue antibody titers were recorded against DENV-2, and increased with age to a geometric mean of 516.5 [1/dil] in the oldest age group. Conclusions/Significance: We found that all four dengue serotypes have been widely circulating in most of urban Indonesia, and more than half of children had already been exposed to >1 dengue serotype, demonstrating intense transmission often associated with more severe clinical episodes. These data will help inform policymakers and highlight the importance of dengue surveillance, prevention and control. [ABSTRACT FROM AUTHOR]

Background: A cluster of eleven patients, including eight family members and three healthcare workers with fever and thrombocytopenia occurred in Yixing County, Jiangsu Province, China, from October to November 1996. However, the initial investigation failed to identify its etiology. Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by SFTS bunyavirus (SFTSV), which was first discovered in 2009. The discovery of novel SFTSV resulted in our consideration to test SFTSV on the remaining samples of this cluster in September 2010. Methodology/Principal findings: We retrospectively analyzed the epidemiological and clinical data of this cluster. The first case, one 55-year-old man with fulminant hemorrhagic diseases, died on October 14, 1996. His younger brother (the second case) developed similar hemorrhagic diseases after nursing him and then died on November 3. From November 4 to November 15, nine other patients, including six family members and three medical staffs, developed fever and thrombocytopenia after exposure to the second case. The sera of six patients were collected on November 24, 1996. IgM antibodies against SFTSV were detected in all of the six patients’ sera using enzyme-linked immunosorbent assay (ELISA), while IgG antibodies were detected in one patient’s serum using an indirect immunofluorescence assay (IFA). We also found that IgG antibodies against SFTSV were still detected in four surviving patients’ sera 14 years after illness onset. Conclusions and significance: The mysterious pathogen of the cluster in 1996 was proved to be SFTSV on the basis of its epidemiological data, clinical data and serological results. It suggests that SFTSV has been circulating in China for more than 10 years before being identified in 2009, and SFTSV IgG antibodies can persist for up to 14 years. [ABSTRACT FROM AUTHOR]

Background: Plasmodium knowlesi is the most common cause of malaria in Malaysian Borneo, with reporting limited to clinical cases presenting to health facilities and scarce data on the true extent of transmission. Serological estimations of transmission have been used with other malaria species to garner information about epidemiological patterns. However, there are a distinct lack of suitable serosurveillance tools for this neglected disease. Methodology/Principal findings: Using in silico tools, we designed and expressed four novel P. knowlesi protein products to address the distinct lack of suitable serosurveillance tools: PkSERA3 antigens 1 and 2, PkSSP2/TRAP and PkTSERA2 antigen 1. Antibody prevalence to these antigens was determined by ELISA for three time-points post-treatment from a hospital-based clinical treatment trial in Sabah, East Malaysia (n = 97 individuals; 241 total samples for all time points). Higher responses were observed for the PkSERA3 antigen 2 (67%, 65/97) across all time-points (day 0: 36.9% 34/92; day 7: 63.8% 46/72; day 28: 58.4% 45/77) with significant differences between the clinical cases and controls (n = 55, mean plus 3 SD) (day 0 p<0.0001; day 7 p<0.0001; day 28 p<0.0001). Using boosted regression trees, we developed models to classify P. knowlesi exposure (cross-validated AUC 88.9%; IQR 86.1–91.3%) and identified the most predictive antibody responses. Conclusions/Significance: The PkSERA3 antigen 2 had the highest relative variable importance in all models. Further validation of these antigens is underway to determine the specificity of these tools in the context of multi-species infections at the population level. [ABSTRACT FROM AUTHOR]

Background: To establish correlates of human immunity to the live plague vaccine (LPV), we analyzed parameters of cellular and antibody response to the plasminogen activator Pla of Y. pestis. This outer membrane protease is an essential virulence factor that is steadily expressed by Y. pestis. Methodology/Principal findings: PBMCs and sera were obtained from a cohort of naïve (n = 17) and LPV-vaccinated (n = 34) donors. Anti-Pla antibodies of different classes and IgG subclasses were determined by ELISA and immunoblotting. The analysis of antibody response was complicated with a strong reactivity of Pla with normal human sera. The linear Pla B-cell epitopes were mapped using a library of 15-mer overlapping peptides. Twelve peptides that reacted specifically with sera of vaccinated donors were found together with a major cross-reacting peptide IPNISPDSFTVAAST located at the N-terminus. PBMCs were stimulated with recombinant Pla followed by proliferative analysis and cytokine profiling. The T-cell recall response was pronounced in vaccinees less than a year post-immunization, and became Th17-polarized over time after many rounds of vaccination. Conclusions/Significance: The Pla protein can serve as a biomarker of successful vaccination with LPV. The diagnostic use of Pla will require elimination of cross-reactive parts of the antigen. [ABSTRACT FROM AUTHOR]