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Recently, we observed a 15% decrease in the incidence of type 1 diabetes (T1D) in Australian 0-4-year-old children following the introduction of RV vaccination [[2], [3]], suggesting that RV vaccination could contribute to the primary prevention of this autoimmune disease. Australian surveillance data [[11]] show that the prevalence of RV G3 strains increased slightly along with an increase in strain diversity in the post-RV vaccine era, but G3 remains a minor component of disease-causing RV strains. RV was prevalent in nurseries, and the change to rooming-in would have altered the timing of exposure to RV, delaying it until later in the first year of life when, based on NOD mouse studies [[17]-[19]], RV might promote rather than retard development of diabetes. [Extracted from the article]

Network coupled cells, such as astrocytes, regulate their cellular homeostasis via Ca2+ signals spread between the cells through gap junctions. Intracellular Ca2+ release is controlled by different signaling pathways that can be stimulated by ATP, glutamate and serotonin (5-HT). Based on our findings, all these pathways are influenced by inflammatory agents and must be restored to fully recover the Ca2+ signaling network. An ultralow concentration of the local anesthetic agent bupivacaine reduced 5-HT-evoked intracellular Ca2+ release, and an ultralow concentration of the phosphodiesterase-5 inhibitor sildenafil in combination with vitamin D3 reduced ATP-evoked intracellular Ca2+ release. Combinations of these three substances downregulated 5-HT-, glutamate- and ATP-evoked intracellular Ca2+ release to a more normal Ca2+ signaling state. Furthermore, inflammatory Toll-like receptor 4 expression decreased with a combination of these three substances. Substance P receptor neurokinin (NK)-1 expression was reduced by ultralow concentrations of bupivacaine. Here, bupivacaine and sildenafil (at extremely low concentrations) combined with vitamin D3 have potential anti-inflammatory properties. According to the present study, drug combinations at the right concentrations, especially extremely low concentrations of bupivacaine and sildenafil, affect different cellular biochemical mechanisms and represent a potential solution for downregulating inflammatory parameters, thereby restoring cells or networks to normal physiological homeostasis. [ABSTRACT FROM AUTHOR]

Middle East respiratory syndrome-coronavirus (MERS-CoV) is an emerging virus that causes severe disease with fatal outcomes; however, there are currently no approved vaccines or specific treatments against MERS-CoV. Here, we developed a novel bivalent vaccine against MERS-CoV and rabies virus (RV) using the replication-incompetent P-gene-deficient RV (RVΔP), which has been previously established as a promising and safe viral vector. MERS-CoV spike glycoprotein comprises S1 and S2 subunits, with the S1 subunit being a primary target of neutralizing antibodies. Recombinant RVΔP, which expresses S1 fused with transmembrane and cytoplasmic domains together with 14 amino acids from the ectodomains of the RV-glycoprotein (RV-G), was developed using a reverse genetics method and named RVΔP-MERS/S1. Following generation of RVΔP-MERS/S1 and RVΔP, our analysis revealed that they shared similar growth properties, with the expression of S1 in RVΔP-MERS/S1-infected cells confirmed by immunofluorescence and western blot, and the immunogenicity and pathogenicity evaluated using mouse infection experiments. We observed no rabies-associated signs or symptoms in mice inoculated with RVΔP-MERS/S1. Moreover, virus-specific neutralizing antibodies against both MERS-CoV and RV were induced in mice inoculated intraperitoneally with RVΔP-MERS/S1. These findings indicate that RVΔP-MERS/S1 is a promising and safe bivalent-vaccine candidate against both MERS-CoV and RV. [ABSTRACT FROM AUTHOR]

Deficiency in several of the classical human RAD51 paralogs [RAD51B, RAD51C, RAD51D, XRCC2 and XRCC3] is associated with cancer predisposition and Fanconi anemia. To investigate their functions, isogenic disruption mutants for each were generated in non-transformed MCF10A mammary epithelial cells and in transformed U2OS and HEK293 cells. In U2OS and HEK293 cells, viable ablated clones were readily isolated for each RAD51 paralog; in contrast, with the exception of RAD51B, RAD51 paralogs are cell-essential in MCF10A cells. Underlining their importance for genomic stability, mutant cell lines display variable growth defects, impaired sister chromatid recombination, reduced levels of stable RAD51 nuclear foci, and hyper-sensitivity to mitomycin C and olaparib. Altogether these observations underscore the contributions of RAD51 paralogs in diverse DNA repair processes, and demonstrate essential differences in different cell types. Finally, this study will provide useful reagents to analyze patient-derived mutations and to investigate mechanisms of chemotherapeutic resistance deployed by cancers. [ABSTRACT FROM AUTHOR]

Background: The processes that result in progression of idiopathic pulmonary fibrosis (IPF) remain enigmatic. Moreover, the course of this disease can be highly variable and difficult to accurately predict. We hypothesized analyses of body mass index (BMI), a simple, routine clinical measure, may also have prognostic value in these patients, and might provide mechanistic insights. We investigated the associations of BMI changes with outcome, plasma adipokines, and adaptive immune activation among IPF patients. Methods: Data were analyzed in an IPF discovery cohort (n = 131) from the University of Pittsburgh, and findings confirmed in patients from the University of Alabama at Birmingham (n = 148). Plasma adipokines were measured by ELISA and T-cell phenotypes determined by flow cytometry. Results: Transplant-free one-year survivals in subjects with the greatest rates of BMI decrements, as percentages of initial BMI (>0.68%/month), were worse than among those with more stable BMI in both discovery (HR = 1.8, 95%CI = 1.1–3.2, p = 0.038) and replication cohorts (HR = 2.5, 95%CI = 1.2–5.2, p = 0.02), when adjusted for age, baseline BMI, and pulmonary function. BMI decrements >0.68%/month were also associated with greater mortality after later lung transplantations (HR = 4.6, 95%CI = 1.7–12.5, p = 0.003). Circulating leptin and adiponectin levels correlated with BMI, but neither adipokine was prognostic per se. BMI decrements were significantly associated with increased proportions of circulating end-differentiated (CD28null) CD4 T-cells (CD28%), a validated marker of repetitive T-cell activation and IPF prognoses. Conclusions: IPF patients with greatest BMI decrements had worse outcomes, and this effect persisted after lung transplantation. Weight loss in these patients is a harbinger of poor prognoses, and may reflect an underlying systemic process, such as adaptive immune activation. [ABSTRACT FROM AUTHOR]

Type I interferons (IFNs) play a critical role in host defense against influenza virus infection, and the mechanism of influenza virus to evade type I IFNs responses remains to be fully understood. Here, we found that progranulin (PGRN) was significantly increased both in vitro and in vivo during influenza virus infection. Using a PGRN knockdown assay and PGRN-deficient mice model, we demonstrated that influenza virus-inducing PGRN negatively regulated type I IFNs production by inhibiting the activation of NF-κB and IRF3 signaling. Furthermore, we showed that PGRN directly interacted with NF-κB essential modulator (NEMO) via its Grn CDE domains. We also verified that PGRN recruited A20 to deubiquitinate K63-linked polyubiquitin chains on NEMO at K264. In addition, we found that macrophage played a major source of PGRN during influenza virus infection, and PGRN neutralizing antibodies could protect against influenza virus-induced lethality in mice. Our data identify a PGRN-mediated IFN evasion pathway exploited by influenza virus with implication in antiviral applications. These findings also provide insights into the functions and crosstalk of PGRN in innate immunity. [ABSTRACT FROM AUTHOR]

Oral herpes is a highly prevalent infection caused by herpes simplex virus 1 (HSV-1). After an initial infection of the oral cavity, HSV-1 remains latent in sensory neurons of the trigeminal ganglia. Episodic reactivation of the virus leads to the formation of mucocutaneous lesions (cold sores), but asymptomatic reactivation accompanied by viral shedding is more frequent and allows virus spread to new hosts. HSV-1 DNA has been detected in many oral tissues. In particular, HSV-1 can be found in periodontal lesions and several studies associated its presence with more severe periodontitis pathologies. Since gingival fibroblasts may become exposed to salivary components in periodontitis lesions, we analyzed the effect of saliva on HSV-1 and -2 infection of these cells. We observed that human gingival fibroblasts can be infected by HSV-1. However, pre-treatment of these cells with saliva extracts from some but not all individuals led to an increased susceptibility to infection. Furthermore, the active saliva could expand HSV-1 tropism to cells that are normally resistant to infection due to the absence of HSV entry receptors. The active factor in saliva was partially purified and comprised high molecular weight complexes of glycoproteins that included secretory Immunoglobulin A. Interestingly, we observed a broad variation in the activity of saliva between donors suggesting that this activity is selectively present in the population. The active saliva factor, has not been isolated, but may lead to the identification of a relevant biomarker for susceptibility to oral herpes. The presence of a salivary factor that enhances HSV-1 infection may influence the risk of oral herpes and/or the severity of associated oral pathologies. [ABSTRACT FROM AUTHOR]

Evolving systems, be it an antibody repertoire in the face of mutating pathogens or a microbial population exposed to varied antibiotics, constantly search for adaptive solutions in time-varying fitness landscapes. Generalists refer to genotypes that remain fit across diverse selective pressures; while multi-drug resistant microbes are undesired yet prevalent, broadly-neutralizing antibodies are much wanted but rare. However, little is known about under what conditions such generalists with a high capacity to adapt can be efficiently discovered by evolution. In addition, can epistasis—the source of landscape ruggedness and path constraints—play a different role, if the environment varies in a non-random way? We present a generative model to estimate the propensity of evolving generalists in rugged landscapes that are tunably related and alternating relatively slowly. We find that environmental cycling can substantially facilitate the search for fit generalists by dynamically enlarging their effective basins of attraction. Importantly, these high performers are most likely to emerge at intermediate levels of ruggedness and environmental relatedness. Our approach allows one to estimate correlations across environments from the topography of experimental fitness landscapes. Our work provides a conceptual framework to study evolution in time-correlated complex environments, and offers statistical understanding that suggests general strategies for eliciting broadly neutralizing antibodies or preventing microbes from evolving multi-drug resistance. [ABSTRACT FROM AUTHOR]

[Fam-] trastuzumab deruxtecan (DS-8201a) is a HER2 (ERBB2)-targeting antibody-drug conjugate, composed of a HER2-targeting antibody and a topoisomerase I inhibitor, exatecan derivative, that has antitumor effects in preclinical xenograft models and clinical trials. Recently, [fam-] trastuzumab deruxtecan was reported to enhance antitumor immunity and was beneficial in combination with an anti–PD-1 antibody in a mouse model. In this study, the antitumor effect of [fam-] trastuzumab deruxtecan in combination with an anti–CTLA-4 antibody was evaluated. [Fam-] trastuzumab deruxtecan monotherapy had antitumor activity in an immunocompetent mouse model with EMT6 human HER2-expressing mouse breast cancer cells (EMT6-hHER2). [Fam-] trastuzumab deruxtecan in combination with the anti–CTLA-4 antibody induced more potent antitumor activity than that by monotherapy with either agent. The combination therapy increased tumor-infiltrating CD4+ and CD8+ T cells in vivo. Mechanistically, cured mice with treatment of [fam-] trastuzumab deruxtecan and an anti–CTLA-4 antibody completely rejected EMT6-mock cells similar to EMT6-hHER2 cells, and splenocytes from the cured mice responded to both EMT6-hHER2 and EMT6-mock cells as measured by interferon-gamma release. Taken together, these results indicate that antitumor immunity is induced by [fam-] trastuzumab deruxtecan and is facilitated in combination with anti–CTLA-4 antibody. [ABSTRACT FROM AUTHOR]

RNA contains various chemical modifications, among which N6-methyladenosine (m6A) is the most prevalent modified nucleotide in eukaryotic mRNA. Emerging evidence suggests that m6A plays an important role in regulating a variety of cellular functions by controlling mRNA processing, translation and degradation. Because m6A is not detectable by standard chemical modification-based approaches, immunological methods, such as ELISA, immunoblotting, immunohistochemistry, m6A RNA immunoprecipitation sequencing and m6A individual-nucleotide resolution cross-linking and immunoprecipitation, have been employed to detect m6A in RNA. Although the most important factor determining the success of these methods is the integrity of highly specific antibodies against m6A, the development of m6A-specific monoclonal antibodies has been challenging. We developed anti-m6A monoclonal antibodies using our recently developed single cell-based monoclonal antibody production system. The binding of one selected antibody, #B1-3, to RNA oligoribonucleotide containing a single m6A had an equilibrium dissociation constant of 6.5 nM, and this antibody exhibited negligible binding to oligoribonucleotides containing a single N1-methyladenosine and unmodified adenosine. The binding was competed by the addition of increasing concentrations of N6-methyl-ATP but not N1-methyl-ATP or ATP. Furthermore, this mAb specifically crosslinked m6A-containing oligoribonucleotide by ultraviolet light, resulting in the induction of cDNA truncation at m6A position. These results show the feasibility of using the validated m6A monoclonal antibody for the specific detection of m6A in RNA. [ABSTRACT FROM AUTHOR]

In this study, we analyzed PRRS virus (PRRSv) specific lymphocyte function in piglets vaccinated with Ingelvac PRRSFLEX EU® at two and three weeks of age in the presence of homologous maternal immunity. Complete analysis of maternal immunity to PRRSv was evaluated postpartum, as well as passive transfer of antibodies and T cells to the piglet through colostrum intake and before and after challenge with a heterologous PRRSv at ten weeks of age. Maternal-derived antibodies were detected in piglets but declined quickly after weaning. However, vaccinated animals restored PRRSv-specific antibody levels by anamnestic response to vaccination. Cell analysis in colostrum and milk revealed presence of PRRSv-specific immune cells at suckling with higher concentrations found in colostrum than in milk. In addition, colostrum and milk contained PRRSv-specific IgA and IgG that may contribute to protection of newborn piglets. Despite the presence of PRRSv-specific Peripheral Blood Mononuclear cells (PBMCs) in colostrum and milk, no PRRSv-specific cells could be detected from blood of the piglets at one or two weeks of life. Nevertheless, cellular immunity was detectable in pre-challenged piglets up to 7 weeks after vaccination while the non-vaccinated control group showed no interferon (IFN) γ response to PRRSv stimulation. After challenge, all piglets developed a PRRSv-specific IFNγ-response, which was more robust at significantly higher levels in vaccinated animals compared to the primary response to PRRSv in non-vaccinated animals. Cytokine analysis in the lung lumen showed a reduction of pro-inflammatory responses to PRRSv challenge in vaccinated animals, especially reduced interferon (IFN) α levels. In conclusion, vaccination of maternally positive piglets at 2 and 3 weeks of age with Ingelvac PRRSFLEX EU induced a humoral and cellular immune response to PRRSv and provided protection against virulent, heterologous PRRSv challenge. [ABSTRACT FROM AUTHOR]

Background: Delirium is the most common postoperative complication of the central nervous system (CNS) that can trigger long-term cognitive impairment. Its underlying mechanism is not fully understood, but the dysfunction of the blood-brain barrier (BBB) has been implicated. The serum levels of the axonal damage biomarker, phosphorylated neurofilament heavy subunit (pNF-H) increase in moderate to severe delirium patients, indicating that postoperative delirium can induce irreversible CNS damage. Here, we investigated the relationship among postoperative delirium, CNS damage and BBB dysfunction, using pNF-H as reference. Methods: Blood samples were collected from 117 patients within 3 postoperative days. These patients were clinically diagnosed with postoperative delirium using the Confusion Assessment Method for the Intensive Care Unit. We measured intercellular adhesion molecule-1, platelet and endothelial cell adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin, and P-selectin as biomarkers for BBB disruption, pro-inflammatory cytokines (tumor necrosis factor-alpha, interleukin-1 beta, interleukin-6), and pNF-H. We conducted logistic regression analysis including all participants to identify independent biomarkers contributing to serum pNF-H detection. Next, by multiple regression analysis with a stepwise method we sought to determine which biomarkers influence serum pNF-H levels, in pNF-H positive patients. Results: Of the 117 subjects, 41 were clinically diagnosed with postoperative delirium, and 30 were positive for serum pNF-H. Sensitivity and specificity of serum pNF-H detection in the patients with postoperative delirium were 56% and 90%, respectively. P-selectin was the only independent variable to associate with pNF-H detection (P < 0.0001) in all 117 patients. In pNF-H positive patients, only PECAM-1 was associated with serum pNF-H levels (P = 0.02). Conclusions: Serum pNF-H could be an objective delirium biomarker, superior to conventional tools in clinical settings. In reference to pNF-H, P-selectin may be involved in the development of delirium-related CNS damage and PECAM-1 may contribute to the progression of delirium- related CNS damage. [ABSTRACT FROM AUTHOR]

Immunization vectors based on cytomegalovirus (CMV) have attracted a lot of interest in recent years because of their high efficacy in the simian immunodeficiency virus (SIV) macaque model, which has been attributed to their ability to induce strong, unusually broad, and unconventionally restricted CD8+ T cell responses. To evaluate the ability of CMV-based vectors to mediate protection by other immune mechanisms, we evaluated a mouse CMV (MCMV)-based vector encoding Friend virus (FV) envelope (Env), which lacks any known CD8+ T cell epitopes, for its protective efficacy in the FV mouse model. When we immunized highly FV-susceptible mice with the Env-encoding MCMV vector (MCMV.env), we could detect high frequencies of Env-specific CD4+ T cells after a single immunization. While the control of an early FV challenge infection was highly variable, an FV infection applied later after immunization was tightly controlled by almost all immunized mice. Protection of mice correlated with their ability to mount a robust anamnestic neutralizing antibody response upon FV infection, but Env-specific CD4+ T cells also produced appreciable levels of interferon γ. Depletion and transfer experiments underlined the important role of antibodies for control of FV infection but also showed that while no Env-specific CD8+ T cells were induced by the MCMV.env vaccine, the presence of CD8+ T cells at the time of FV challenge was required. The immunity induced by MCMV.env immunization was long-lasting, but was restricted to MCMV naïve animals. Taken together, our results demonstrate a novel mode of action of a CMV-based vaccine for anti-retrovirus immunization that confers strong protection from retrovirus challenge, which is conferred by CD4+ T cells and antibodies. [ABSTRACT FROM AUTHOR]

Intracellular malaria parasites grow in a vacuole delimited by the parasitophorous vacuolar membrane (PVM). This membrane fulfils critical roles for survival of the parasite in its intracellular niche such as in protein export and nutrient acquisition. Using a conditional knockout (KO), we here demonstrate that the abundant integral PVM protein exported protein 1 (EXP1) is essential for parasite survival but that this is independent of its previously postulated function as a glutathione S-transferase (GST). Patch-clamp experiments indicated that EXP1 is critical for the nutrient-permeable channel activity at the PVM. Loss of EXP1 abolished the correct localisation of EXP2, a pore-forming protein required for the nutrient-permeable channel activity and protein export at the PVM. Unexpectedly, loss of EXP1 affected only the nutrient-permeable channel activity of the PVM but not protein export. Parasites with low levels of EXP1 became hypersensitive to low nutrient conditions, indicating that EXP1 indeed is needed for nutrient uptake and experimentally confirming the long-standing hypothesis that the channel activity measured at the PVM is required for parasite nutrient acquisition. Hence, EXP1 is specifically required for the functional expression of EXP2 as the nutrient-permeable channel and is critical for the metabolite supply of malaria parasites. [ABSTRACT FROM AUTHOR]

Open data science and algorithm development competitions offer a unique avenue for rapid discovery of better computational strategies. We highlight three examples in computational biology and bioinformatics research in which the use of competitions has yielded significant performance gains over established algorithms. These include algorithms for antibody clustering, imputing gene expression data, and querying the Connectivity Map (CMap). Performance gains are evaluated quantitatively using realistic, albeit sanitized, data sets. The solutions produced through these competitions are then examined with respect to their utility and the prospects for implementation in the field. We present the decision process and competition design considerations that lead to these successful outcomes as a model for researchers who want to use competitions and non-domain crowds as collaborators to further their research. [ABSTRACT FROM AUTHOR]

The Warburg Effect has emerged as a potential drug target because, in some cancer cell lines, it is sufficient to subvert it in order to kill cancer cells. It has also been shown that the Warburg Effect occurs in innate immune cells upon infection. Innate immune cells play critical roles in the tumour microenvironment but the Warburg Effect is not fully understood in monocytes. Furthermore, it is important to understand the impact of infections on key players in the tumour microenvironment because inflammatory conditions often precede carcinogenesis and mutated oncogenes induce inflammation. We investigated the metabolic programme in the acute monocytic leukaemia cell line, THP-1 in the presence and absence of lipopolysaccharide, mimicking bacterial infections. We found that stimulation of THP-1 cells by LPS induces a subset of pro-inflammatory chemokines and enhances the Warburg Effect. Surprisingly, perturbation of the Warburg Effect in these cells does not lead to cell death in contrast to what was observed in non-myeloid cancer cell lines in a previous study. These findings indicate that the Warburg Effect and inflammation are activated by bacterial lipopolysaccharide and may have a profound influence on the microenvironment. [ABSTRACT FROM AUTHOR]

Monkeypox virus (MPXV) is a member of the genus Orthopoxvirus, endemic in Central and West Africa. This viral zoonosis was introduced into the United States in 2003 via African rodents imported for the pet trade and caused 37 human cases, all linked to exposure to MPXV-infected black-tailed prairie dogs (Cynomys ludovicianus). Prairie dogs have since become a useful model of MPXV disease, utilized for testing of potential medical countermeasures. In this study, we used recombinant MPXV containing the firefly luciferase gene (luc) and in vivo imaging technology to characterize MPXV pathogenesis in the black-tailed prairie dog in real time. West African (WA) MPXV could be visualized using in vivo imaging in the nose, lymph nodes, intestines, heart, lung, kidneys, and liver as early as day 6 post infection (p.i.). By day 9 p.i., lesions became visible on the skin and in some cases in the spleen. After day 9 p.i., luminescent signal representing MPXV replication either increased, indicating a progression to what would be a fatal infection, or decreased as infection was resolved. Use of recombinant luc+ MPXV allowed for a greater understanding of how MPXV disseminates throughout the body in prairie dogs during the course of infection. This technology will be used to reduce the number of animals required in future pathogenesis studies as well as aid in determining the effectiveness of potential medical countermeasures. [ABSTRACT FROM AUTHOR]

Swine influenza virus (SIVs) infections cause a significant economic impact to the pork industry. Moreover, pigs may act as mixing vessel favoring genome reassortment of diverse influenza viruses. Such an example is the pandemic H1N1 (pH1N1) virus that appeared in 2009, harboring a combination of gene segments from avian, pig and human lineages, which rapidly reached pandemic proportions. In order to confront and prevent these possible emergences as well as antigenic drift phenomena, vaccination remains of vital importance. The present work aimed to evaluate a new DNA influenza vaccine based on distinct conserved HA-peptides fused with flagellin and applied together with Diluvac Forte as adjuvant using a needle-free device (IntraDermal Application of Liquids, IDAL®). Two experimental pig studies were performed to test DNA-vaccine efficacy against SIVs in pigs. In the first experiment, SIV-seronegative pigs were vaccinated with VC4-flagellin DNA and intranasally challenged with a pH1N1. In the second study, VC4-flagellin DNA vaccine was employed in SIV-seropositive animals and challenged intranasally with an H3N2 SIV-isolate. Both experiments demonstrated a reduction in the viral shedding after challenge, suggesting vaccine efficacy against both the H1 and H3 influenza virus subtypes. In addition, the results proved that maternally derived antibodies (MDA) did not constitute an obstacle to the vaccine approach used. Moreover, elevated titers in antibodies both against H1 and H3 proteins in serum and in bronchoalveolar lavage fluids (BALFs) was detected in the vaccinated animals along with a markedly increased mucosal IgA response. Additionally, vaccinated animals developed stronger neutralizing antibodies in BALFs and higher inhibiting hemagglutination titers in sera against both the pH1N1 and H3N2 influenza viruses compared to unvaccinated, challenged-pigs. It is proposed that the described DNA-vaccine formulation could potentially be used as a multivalent vaccine against SIV infections. [ABSTRACT FROM AUTHOR]

Introduction: Studying the role of the immune system in the interaction between mental and physical health is challenging. To study individuals with an intensive, longitudinal study design that requires repetitive sampling in their daily life, non-invasive sampling techniques are a necessity. Urine can be collected in a non-invasive way, but this may be demanding for participants and little is known about fluctuation of inflammatory markers in urine over time. The aim of this study was to investigate the feasibility of non-invasive sampling, and to explore intra-individual differences in inflammatory markers in urine. Materials & methods: Ten healthy individuals collected 24-hour urine for 63 consecutive days. In a pilot analysis, 39 inflammatory markers were examined for detectability in urine, stability over time and under storage conditions, and daily fluctuations. Multiplex analyses were used to quantify levels of eight selected markers: C-reactive protein (CRP), Fractalkine, Interleukin-1 receptor-antagonist (IL-1RA), interferon-α (IFNα), interferon-γ (IFNγ), Interferon gamma-induced protein 10 (IP10), Macrophage inflammatory protein-1β (MIP-1β), and Vascular Endothelial Growth Factor (VEGF). Cross-correlations were calculated between the overnight and 24-hour samples were calculated, to examine whether 24-hour urine could be replaced by the overnight portion for better feasibility. We examined intra- and interindividual differences in the levels of inflammatory markers in urine and the fluctuations thereof. Results: This study showed that levels of selected inflammatory markers can be detected in urine. Cross-correlation analyses showed that correlations between levels of inflammatory markers in the night portion and the 24-hour urine sample varied widely between individuals. In addition, analyses of time series revealed striking inter- and intra-individual variation in levels of inflammatory markers and their fluctuations. Conclusion: We show that the assessment of urinary inflammatory markers is feasible in an intensive day-to-day study in healthy individuals. However, 24-hour urine cannot be replaced by an overnight portion to alleviate the protocol burden. Levels of inflammatory markers show substantial variation between and within persons. [ABSTRACT FROM AUTHOR]

The equine hoof inner epithelium is folded into primary and secondary epidermal lamellae which increase the dermo-epidermal junction surface area of the hoof and can be affected by laminitis, a common disease of equids. Two keratin proteins (K), K42 and K124, are the most abundant keratins in the hoof lamellar tissue of Equus caballus. We hypothesize that these keratins are lamellar tissue-specific and could serve as differentiation- and disease-specific markers. Our objective was to characterize the expression of K42 and K124 in equine stratified epithelia and to generate monoclonal antibodies against K42 and K124. By RT-PCR analysis, keratin gene (KRT) KRT42 and KRT124 expression was present in lamellar tissue, but not cornea, haired skin, or hoof coronet. In situ hybridization studies showed that KRT124 localized to the suprabasal and, to a lesser extent, basal cells of the lamellae, was absent from haired skin and hoof coronet, and abruptly transitions from KRT124-negative coronet to KRT124-positive proximal lamellae. A monoclonal antibody generated against full-length recombinant equine K42 detected a lamellar keratin of the appropriate size, but also cross-reacted with other epidermal keratins. Three monoclonal antibodies generated against N- and C-terminal K124 peptides detected a band of the appropriate size in lamellar tissue and did not cross-react with proteins from haired skin, corneal limbus, hoof coronet, tongue, glabrous skin, oral mucosa, or chestnut on immunoblots. K124 localized to lamellar cells by indirect immunofluorescence. This is the first study to demonstrate the localization and expression of a hoof lamellar-specific keratin, K124, and to validate anti-K124 monoclonal antibodies. [ABSTRACT FROM AUTHOR]

Introduction: Mucosal immune activation, in the context of sexual transmission of HIV-1 infection, is crucial, as the increased presence of activated T cells enhance susceptibility to infection. In this regard, it has been proposed that immunomodulatory compounds capable of modulating immune activation, such as Vitamin D (VitD) may reduce HIV-1 transmission and might be used as a safe and cost-effective strategy for prevention. Considering this, we examined the in vitro effect of the treatment of peripheral blood mononuclear cells (PBMCs) with the active form of VitD, calcitriol, on cellular activation, function and susceptibility of CD4+ T cells to HIV-1 infection. Methods: We treated PBMCs from healthy HIV unexposed individuals (Co-HC) and frequently exposed, HIV-1 seronegative individuals (HESNs) from Colombia and from healthy non-exposed individuals from Canada (Ca-HC) with calcitriol and performed in vitro HIV-1 infection assays using X4- and R5-tropic HIV-1 strains respectively. In addition, we evaluated the activation and function of T cells and the expression of viral co-receptors, and select antiviral genes following calcitriol treatment. Results: Calcitriol reduced the frequency of infected CD4+ T cells and the number of viral particles per cell, for both, X4- and R5-tropic viruses tested in the Co-HC and the Ca-HC, respectively, but not in HESNs. Furthermore, in the Co-HC, calcitriol reduced the frequency of polyclonally activated T cells expressing the activation markers HLA-DR and CD38, and those HLA-DR+CD38-, whereas increased the subpopulation HLA-DR-CD38+. Calcitriol treatment also decreased production of granzyme, IL-2 and MIP-1β by T cells and increased the transcriptional expression of the inhibitor of NF-kB and the antiviral genes cathelicidin (CAMP) and APOBEC3G in PBMCs from Co-HC. Conclusion: Our in vitro findings suggest that VitD treatment could reduce HIV-1 transmission through a specific modulation of the activation levels and function of T cells, and the production of antiviral factors. In conclusion, VitD remains as an interesting potential strategy to prevent HIV-1 transmission that should be further explored. [ABSTRACT FROM AUTHOR]

Dendritic cells (DCs) are professional antigen presenting cells involved in the induction of T cell-mediated adaptive immunity. Plasmacytoid DCs (pDCs) originate from lymphoid precursors and produce type I interferons (IFNs) in response to pathogens. A20 is considered as a negative regulator of toll-like receptor (TLR) signaling pathways, in which Toxoplasma gondii- derived profilin (TgPRF) is a TLR11/12 ligand recognised by DCs to stimulate their maturation/activation. Little is known about contributions of A20 to changes in biological properties of pDCs. The present study, therefore, explored whether pDC functions are influenced by A20. To this end, bone marrow cells were isolated and cultured with Flt3L to attain CD8DCs, CD11bDCs and pDCs and followed by challenge with TgPRP in the presence or absence of A20 siRNA. Expression of maturation markers were analysed by flow cytometry, and secretion of inflammatory cytokines by ELISA, cell migration by a transwell migration assay and expression of signalling molecules by western blotting. As a result, treatment with A20 siRNA enhanced activations of IκB-α and STAT-1, leading to increases in expressions of maturation markers and cytokine productions as well as migration of TgPRP-treated pDCs, while mature CD11bDCs produced at higher levels of TNF-α and IL-6 only. In addition, functions of CD8DCs remained unaltered following A20 silencing. The effects of A20 on pDC maturation and activation were completely abolished by IKK inhibitor and partially blunted by fludarabine. In conclusion, the inhibitory effects of A20 on pDC functions are expected to affect the immune response in T. gondii infection. [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]

Recent studies have begun to elucidate a role for E3 ubiquitin ligases as important mediators of the innate immune response. Our previous work defined a role for the ubiquitin ligase natural killer lytic-associated molecule (NKLAM/RNF19b) in mouse and human innate immunity. Here, we present novel data describing a role for NKLAM in regulating the immune response to Sendai virus (SeV), a murine model of paramyxoviral pneumonia. NKLAM expression was significantly upregulated by SeV infection. SeV-infected mice that are deficient in NKLAM demonstrated significantly less weight loss than wild type mice. In vivo, Sendai virus replication was attenuated in NKLAM-/- mice. Autophagic flux and the expression of autophagy markers LC3 and p62/SQSTM1 were also less in NKLAM-/- mice. Using flow cytometry, we observed less neutrophils and macrophages in the lungs of NKLAM-/- mice during SeV infection. Additionally, phosphorylation of STAT1 and NFκB p65 was lower in NKLAM-/- than wild type mice. The dysregulated phosphorylation profile of STAT1 and NFκB in NKLAM-/- mice correlated with decreased expression of numerous proinflammatory cytokines that are regulated by STAT1 and/or NFκB. The lack of NKLAM and the resulting attenuated immune response is favorable to NKLAM-/- mice receiving a low dose of SeV; however, at a high dose of virus, NKLAM-/- mice succumbed to the infection faster than wild type mice. In conclusion, our novel results indicate that NKLAM plays a role in regulating the production of pro-inflammatory cytokines during viral infection. [ABSTRACT FROM AUTHOR]

Respiratory infection can be exacerbated by the high glucose concentration in the airway surface liquid (ASL). We investigated the effects of salbutamol and phlorizin on the pulmonary function, oxidative stress levels and SGLT1 activity in lung, pulmonary histopathological damages and survival rates of rats with sepsis. Sepsis was induced by cecal ligation and puncture surgery (CLP). Twenty-four hours after surgery, CLP rats were intranasally treated with saline, salbutamol or phlorizin. After 2 hours, animals were anesthetized and sacrificed. Sepsis promoted atelectasis and bronchial inflammation, and led to increased expression of SGLT1 on cytoplasm of pneumocytes. Salbutamol treatment reduced bronchial inflammation and promoted hyperinsuflation in CLP rats. The interferon-ɤ and Interleucin-1β concentrations in bronchoalveolar lavage (BAL) were closely related to the bronchial inflammation regulation. Salbutamol stimulated SGLT1 in plasma membrane; whereas, phlorizin promoted the increase of SGLT1 in cytoplasm. Phlorizin reduced catalase activity and induced a significant decrease in the survival rate of CLP rats. Taken together, sepsis promoted atelectasis and lung inflammation, which can be associated with SGLT1 inhibition. The loss of function of SGLT1 by phlorizin are related to the augmented disease severity, increased atelectasis, bronchial inflammation and a significant reduction of survival rate of CLP rats. Alternatively salbutamol reduced BAL inflammatory cytokines, bronchial inflammation, atelectasis, and airway damage in sepsis. These data suggest that this selective β2-adrenergic agonist may protect lung of septic acute effects. [ABSTRACT FROM AUTHOR]

The ability of DENV2 to display different morphologies (hence different antigenic properties) complicates vaccine and therapeutics development. Previous studies showed most strains of laboratory adapted DENV2 particles changed from smooth to “bumpy” surfaced morphology when the temperature is switched from 29°C at 37°C. Here we identified five envelope (E) protein residues different between two alternative passage history DENV2 NGC strains exhibiting smooth or bumpy surface morphologies. Several mutations performed on the smooth DENV2 infectious clone destabilized the surface, as observed by cryoEM. Molecular dynamics simulations demonstrated how chemically subtle substitution at various positions destabilized dimeric interactions between E proteins. In contrast, three out of four DENV2 clinical isolates showed a smooth surface morphology at 37°C, and only at high fever temperature (40°C) did they become “bumpy”. These results imply vaccines should contain particles representing both morphologies. For prophylactic and therapeutic treatments, this study also informs on which types of antibodies should be used at different stages of an infection, i.e., those that bind to monomeric E proteins on the bumpy surface or across multiple E proteins on the smooth surfaced virus. [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]

Background: The current strategy for the elimination of onchocerciasis is based on annual or bi-annual mass drug administration with ivermectin. However, due to several limiting factors there is a growing concern that elimination of onchocerciasis cannot be achieved solely through the current strategy. Additional tools are critically needed including a prophylactic vaccine. Presently Ov-103 and Ov-RAL-2 are the most promising vaccine candidates against an Onchocerca volvulus infection. Methodology/Principal findings: Protection induced by immunization of mice with the alum-adjuvanted Ov-103 or Ov-RAL-2 vaccines appeared to be antibody dependent since AID-/- mice that could not mount antigen-specific IgG antibody responses were not protected from an Onchocerca volvulus challenge. To determine a possible association between antigen-specific antibody responses and anti-larvae protective immunity in humans, we analyzed the presence of anti-Ov-103 and anti-Ov-RAL-2 cytophilic antibody responses (IgG1 and IgG3) in individuals classified as putatively immune, and in infected individuals who developed concomitant immunity with age. It was determined that 86% of putatively immune individuals and 95% individuals with concomitant immunity had elevated IgG1 and IgG3 responses to Ov-103 and Ov-RAL-2. Based on the elevated chemokine levels associated with protection in the Ov-103 or Ov-RAL-2 immunized mice, the profile of these chemokines was also analyzed in putatively immune and infected individuals; both groups contained significantly higher levels of KC, IP-10, MCP-1 and MIP-1β in comparison to normal human sera. Moreover, human monospecific anti-Ov-103 antibodies but not anti-Ov-RAL-2 significantly inhibited the molting of third-stage larvae (L3) in vitro by 46% in the presence of naïve human neutrophils, while both anti-Ov-103 and anti-Ov-RAL-2 antibodies significantly inhibited the molting by 70–80% when cultured in the presence of naive human monocytes. Interestingly, inhibition of molting by Ov-103 antibodies and monocytes was only in part dependent on contact with the cells, while inhibition of molting with Ov-RAL-2 antibodies was completely dependent on contact with the monocytes. In comparison, significant levels of parasite killing in Ov-103 and Ov-RAL-2 vaccinated mice only occurred when cells enter the parasite microenvironment. Taken together, antibodies to Ov-103 and Ov-RAL-2 and cells are required for protection in mice as well as for the development of immunity in humans. Conclusions/Significance: Alum-adjuvanted Ov-103 and Ov-RAL-2 vaccines have the potential of reducing infection and thus morbidity associated with onchocerciasis also in humans. The development of cytophilic antibodies, that function in antibody-dependent cellular cytotoxicity, is essential for a successful prophylactic vaccine against this infection. [ABSTRACT FROM AUTHOR]

Background: Seasonal influenza results in significant morbidity and mortality worldwide, but the currently licensed inactivated vaccines generally have low vaccine efficacies and could be improved. In this phase 1 clinical trial, we compared seasonal influenza vaccine regimens with different priming strategies, prime-boost intervals, and administration routes to determine the impact of these variables on the resulting antibody response. Methods: Between August 17, 2012 and January 25, 2013, four sites enrolled healthy adults 18–70 years of age. Subjects were randomized to receive one of the following vaccination regimens: trivalent hemagglutinin (HA) DNA prime followed by trivalent inactivated influenza vaccine (IIV3) boost with a 3.5 month interval (DNA-IIV3), IIV3 prime followed by IIV3 boost with a 10 month interval (IIV3-IIV3), or concurrent DNA and IIV3 prime followed by IIV3 boost with a 10 month interval (DNA/IIV3-IIV3). Each regimen was additionally stratified by an IIV3 administration route of either intramuscular (IM) or intradermal (ID). DNA vaccines were administered by a needle-free jet injector (Biojector). Study objectives included evaluating the safety and tolerability of each regimen and measuring the antibody response by hemagglutination inhibition (HAI). Results: Three hundred and sixteen subjects enrolled. Local reactogenicity was mild to moderate in severity, with higher frequencies recorded following DNA vaccine administered by Biojector compared to IIV3 by either route (p <0.02 for pain, swelling, and redness) and following IIV3 by ID route compared to IM route (p <0.001 for swelling and redness). Systemic reactogenicity was similar between regimens. Though no overall differences were observed between regimens, the highest titers post boost were observed in the DNA-IIV3 group by ID route and in the IIV3-IIV3 group by IM route. Conclusions: All vaccination regimens were found to be safe and tolerable. While there were no overall differences between regimens, the DNA-IIV3 group by ID route, and the IIV3-IIV3 group by IM route, showed higher responses compared to the other same-route regimens. [ABSTRACT FROM AUTHOR]

Neutrophils play a critical role in host defense against Pseudomonas aeruginosa infection. Mechanisms underlying the negative regulation of neutrophil function in bacterial clearance remain incompletely defined. Here, we demonstrate that protein tyrosine phosphatase-1B (PTP1B) is a negative regulator of P. aeruginosa clearance by neutrophils. PTP1B-deficient neutrophils display greatly enhanced bacterial phagocytosis and killing, which are accompanied by increased Toll-like receptor 4 (TLR4) signaling activation and nitric oxide (NO) production following P. aeruginosa infection. Interestingly, PTP1B deficiency mainly upregulates the production of IL-6 and IFN-β, leads to enhanced TLR4-dependent STAT1 activation and iNOS expression by neutrophils following P. aeruginosa infection. Further studies reveal that PTP1B and STAT1 are physically associated. These findings demonstrate a negative regulatory mechanism in neutrophil underlying the elimination of P. aeruginosa infection though a PTP1B-STAT1 interaction. [ABSTRACT FROM AUTHOR]

Rhesus macaque is an important animal model in biomedical research, especially human disease, developmental, translational, and pre-clinical research. Blood physiological and biochemical parameters are important markers for physiology, pathology, and toxicology research. However, these parameters have not been systematically reported for Chinese rhesus macaques. To characterize the reference for these parameters, this study collected 1805 Chinese rhesus macaques living in Southwestern China. A total of 24 blood physiological indexes and 27 biochemical parameters were determined. Sex and age were found to affect these parameters. In conclusion, a comprehensive and systematic reference of hematological and biochemical parameters for Chinese rhesus macaque was established in this work on the basis of a large cohort. Such reference will benefit biomedical research employing rhesus macaques as animal models. [ABSTRACT FROM AUTHOR]

Mycobacterium bovis is the causative agent of bovine tuberculosis (TB), a cattle disease of global importance. M. bovis infects bovine macrophages (Mø) and subverts the host cell response to generate a suitable niche for survival and replication. We investigated the role of the anti-inflammatory cytokine interleukin (IL) 10 during in vitro infection of bovine monocyte-derived Mø (bMDM) with two divergent UK strains of M. bovis, which differentially modulate expression of IL10. The use of IL10-targeting siRNA revealed that IL10 inhibited the production of IL1B, IL6, tumour necrosis factor (TNF) and interferon gamma (IFNG) during infection of bMDM with the M. bovis strain G18. In contrast, IL10 only regulated a subset of these genes; TNF and IFNG, during infection with the M. bovis reference strain AF2122/97. Furthermore, nitric oxide (NO) production was modulated by IL10 during AF2122/97 infection, but not at the nitric oxide synthase 2 (NOS2) mRNA level, as observed during G18 infection. However, IL10 was found to promote survival of both M. bovis strains during early bMDM infection, but this effect disappeared after 24 h. The role of IL10-induced modulation of TNF, IFNG and NO production in M. bovis survival was investigated using siRNA targeting TNF, IFNG receptor 1 (IFNGR1) and NOS2. Knock-down of these genes individually did not promote survival of either M. bovis strain and therefore modulation of these genes does not account for the effect of IL10 on M. bovis survival. However, TNF knock-down was found to be detrimental to the survival of the M. bovis strain G18 during early infection. The results provide further evidence for the importance of IL10 during M. bovis infection of Mø. Furthermore, they highlight M. bovis strain specific differences in the interaction with the infected bMDM, which may influence the course of infection and progression of bovine TB. [ABSTRACT FROM AUTHOR]

This study aimed to assess the renopreventive effect of enalapril and/or paricalcitol on streptozotocin (STZ) diabetes-induced nephropathy and to elucidate their mechanisms of action through investigation of the effects on renal oxidative stress, antioxidant defense system and expressions of TNF-α, p53, caspase-3, and Bcl-2. Diabetes mellitus was induced in fasting male Wistar rats by single intraperitoneal injection of STZ (45 mg /kg b.w.) dissolved in citrate buffer (pH 4.5). Ten days after STZ injection, the diabetic rats were treated with enalapril (25 mg/l of drinking water) and/or paricalcitol (8 μg/kg b.w. per os) dissolved in 5% DMSO daily for 4 weeks. The obtained data revealed that the treatment of diabetic Wistar rats with enalapril and/or paricalcitol led to significant decreases in the elevated serum urea, uric acid, creatinine, sodium and potassium levels; thereby reflecting the improvement of the impaired kidney function. The deteriorated kidney lipid peroxidation, GSH content and GST and catalase activities in diabetic rats were significantly ameliorated as a result of treatment with enalapril and/or paricalcitol. The elevated fasting and post-prandial serum glucose levels and the lowered serum insulin and C-peptide levels were also improved. The treatment with enalapril and paricalcitol in combination was the most potent in decreasing the elevated serum glucose levels. Moreover, the treatment of diabetic rats successfully prevented the diabetes-induced histopathological deleterious changes of kidney and islets of Langerhans of pancreas. In association, the immunohistochemically detected pro-inflammatory cytokine, TNF-α, and apoptotic mediators, p53 and caspase-3, were remarkably decreased in kidney of diabetic rats as a result of treatment while the expression of anti-apoptotic protein Bcl-2 was increased. Based on these findings, it can be concluded that enalapril and paricalcitol alone or in combination can prevent STZ diabetes-induced nephropathy through amelioration of the glycemic state and antioxidant defense system together with the suppression of oxidative stress, inflammation and apoptosis. However, the treatment of diabetic rats with enalapril and paricalcitol in combination has no further significant improvement effects on renal function and damage when compared with enalapril or paclitaxel treated diabetic groups. [ABSTRACT FROM AUTHOR]

This study investigated whether stress responsiveness (in one context) can be used to predict dog behavior in daily life. On two occasions (NT1 = 32 puppies; NT2 = 16 young adults), dogs’ physiological stress response after a behavioral test at home was measured in terms of reactivity (10 min post-test) and recovery (40 min post-test) for three salivary markers: cortisol, chromogranin A (CgA) and secretory immunoglobulin A (sIgA). For each marker, it was determined whether dogs with a strong physiological response displayed different behavior in daily life compared to dogs with a weaker physiological response. The results revealed three main findings: first, for CgA and cortisol, different patterns were identified according to sample time. High reactivity related to desirable traits, whereas slow recovery after the behavioral test related to undesirable traits. The findings suggest that increased levels of CgA and cortisol 10 minutes after the behavioral test reflected an adaptive stress response, whereas elevated levels 40 minutes after the test reflected unsuccessful coping. Second, patterns for sIgA differed from CgA and cortisol: significant associations were only found with behavioral traits at T2, mostly considered desirable and related to Trainability. Possibly, the delayed reaction pattern of sIgA caused this difference between markers, as sIgA reflects the (secondary) immune response to stress, due to immunosuppressive effects of cortisol. Third, predictive capacity of puppies’ physiological stress response (T1) was inconclusive, and contrary relations were found with behavioral traits at T2, suggesting that developmental factors play an important role. This study provides new insights about the relation between stress physiology and behavioral traits, and methodological advice is given to study these patterns further. In conclusion, physiological markers could provide additional insights in dogs’ tendencies to display certain behaviors, especially at the young adult stage. Further studies are needed to confirm these patterns. [ABSTRACT FROM AUTHOR]

Laboratory assays for identifying recent HIV-1 infections are widely used for estimating incidence in cross-sectional population-level surveys in global HIV-1surveillance. Adequate assay and laboratory performance are required to ensure accurate incidence estimates. The NIAID-supported External Quality Assurance Program Oversight Laboratory (EQAPOL) established a proficiency testing program for the most widely-used incidence assay, the HIV-1 Limiting Antigen Avidity EIA (LAg), with US Centers for Disease Control and Prevention (CDC)-approved kits manufactured by Sedia Biosciences Corporation and Maxim Biomedical. The objective of this program is to monitor the performance of participating laboratories. Four rounds of blinded external proficiency (EP) panels were distributed to up to twenty testing sites (7 North American, 5 African, 4 Asian, 2 South American and 2 European). These panels consisted of ten plasma samples: three blinded well-characterized HIV-1-seropositive samples that were included as replicates and an HIV-negative control. The seropositive samples spanned the dynamic range of the assay and are categorized as either recent or long-term infection. Participating sites performed the assay according to manufacturers’ instructions and completed an online survey to gather information on kit manufacturer, lot of kit used, laboratory procedures and the experience of technicians. On average, fifteen sites participated in each round of testing, with an average of four sites testing with only the Maxim assay, seven testing with only the Sedia assay and five sites utilizing both assays. Overall, the Sedia and Maxim assays yielded similar infection status categorization across the laboratories; however, for most of the nine HIV+ samples tested, there were significant differences in the optical density readouts, ODn (N = 8) and OD (N = 7), between LAg kit manufacturers (p < 0.05 based on mixed effects models. The EQAPOL LAg program is important for monitoring laboratory performance as well as detecting variations between manufacturers of HIV-1incidence assays. [ABSTRACT FROM AUTHOR]

Cytomegalovirus (CMV) is a ubiquitous β-herpesvirus that establishes life-long latent infection in a high percentage of the population worldwide. CMV induces the strongest and most durable CD8+ T cell response known in human clinical medicine. Due to its unique properties, the virus represents a promising candidate vaccine vector for the induction of persistent cellular immunity. To take advantage of this, we constructed a recombinant murine CMV (MCMV) expressing an MHC-I restricted epitope from influenza A virus (IAV) H1N1 within the immediate early 2 (ie2) gene. Only mice that were immunized intranasally (i.n.) were capable of controlling IAV infection, despite the greater potency of the intraperitoneally (i.p.) vaccination in inducing a systemic IAV-specific CD8+ T cell response. The protective capacity of the i.n. immunization was associated with its ability to induce IAV-specific tissue-resident memory CD8+ T (CD8TRM) cells in the lungs. Our data demonstrate that the protective effect exerted by the i.n. immunization was critically mediated by antigen-specific CD8+ T cells. CD8TRM cells promoted the induction of IFNγ and chemokines that facilitate the recruitment of antigen-specific CD8+ T cells to the lungs. Overall, our results showed that locally applied MCMV vectors could induce mucosal immunity at sites of entry, providing superior immune protection against respiratory infections. [ABSTRACT FROM AUTHOR]

The CD4 binding site (CD4bs) of the HIV-1 envelope glycoprotein is susceptible to multiple lineages of broadly neutralizing antibodies (bnAbs) that are attractive to elicit with vaccines. The CH235 lineage (VH1-46) of CD4bs bnAbs is particularly attractive because the most mature members neutralize 90% of circulating strains, do not possess long HCDR3 regions, and do not contain insertions and deletions that may be difficult to induce. We used virus neutralization to measure the interaction of CH235 unmutated common ancestor (CH235 UCA) with functional Env trimers on infectious virions to guide immunogen design for this bnAb lineage. Two Env mutations were identified, one in loop D (N279K) and another in V5 (G458Y), that acted synergistically to render autologous CH505 transmitted/founder virus susceptible to neutralization by CH235 UCA. Man5-enriched N-glycans provided additional synergy for neutralization. CH235 UCA bound with nanomolar affinity to corresponding soluble native-like Env trimers as candidate immunogens. A cryo-EM structure of CH235 UCA bound to Man5-enriched CH505.N279K.G458Y.SOSIP.664 revealed interactions of the antibody light chain complementarity determining region 3 (CDR L3) with the engineered Env loops D and V5. These results demonstrate that virus neutralization can directly inform vaccine design and suggest a germline targeting and reverse engineering strategy to initiate and mature the CH235 bnAb lineage. [ABSTRACT FROM AUTHOR]

The nucleotide-binding-domain (NBD)–and leucine-rich-repeat (LRR)–containing (NLR) family, pyrin-domain–containing 3 (NLRP3) inflammasome drives pathological inflammation in a suite of autoimmune, metabolic, malignant, and neurodegenerative diseases. Additionally, NLRP3 gain-of-function point mutations cause systemic periodic fever syndromes that are collectively known as cryopyrin-associated periodic syndrome (CAPS). There is significant interest in the discovery and development of diarylsulfonylurea Cytokine Release Inhibitory Drugs (CRIDs) such as MCC950/CRID3, a potent and selective inhibitor of the NLRP3 inflammasome pathway, for the treatment of CAPS and other diseases. However, drug discovery efforts have been constrained by the lack of insight into the molecular target and mechanism by which these CRIDs inhibit the NLRP3 inflammasome pathway. Here, we show that the domain conserved in NAIP, CIITA, HET-E, and TP1 (NACHT) domain of NLRP3 is the molecular target of diarylsulfonylurea inhibitors. Interestingly, we find photoaffinity labeling (PAL) of the NACHT domain requires an intact (d)ATP-binding pocket and is substantially reduced for most CAPS-associated NLRP3 mutants. In concordance with this finding, MCC950/CRID3 failed to inhibit NLRP3-driven inflammatory pathology in two mouse models of CAPS. Moreover, it abolished circulating levels of interleukin (IL)-1β and IL-18 in lipopolysaccharide (LPS)-challenged wild-type mice but not in Nlrp3L351P knock-in mice and ex vivo-stimulated mutant macrophages. These results identify wild-type NLRP3 as the molecular target of MCC950/CRID3 and show that CAPS-related NLRP3 mutants escape efficient MCC950/CRID3 inhibition. Collectively, this work suggests that MCC950/CRID3-based therapies may effectively treat inflammation driven by wild-type NLRP3 but not CAPS-associated mutants. [ABSTRACT FROM AUTHOR]

Introduction: End-stage renal disease (ESRD) strongly associates with cardiovascular disease (CVD) risk. This risk is not completely mitigated by renal replacement therapy. Endothelial dysfunction (ED) and low-grade inflammation (LGI) may contribute to the increased CVD risk. However, data on serum biomarkers of ED and LGI during the transition to renal replacement therapy (dialysis and kidney transplantation) are scarce. Methods: We compared serum biomarkers of ED and LGI between 36 controls, 43 participants with chronic kidney disease (CKD) stage 5 non-dialysis (CKD5-ND), 20 participants with CKD stage 5 hemodialysis (CKD5-HD) and 14 participants with CKD stage 5 peritoneal dialysis (CKD5-PD). Further, in 34 and 15 participants repeated measurements were available during the first six months following dialysis initiation and kidney transplantation, respectively. Serum biomarkers of ED (sVCAM-1, E-selectin, P-selectin, thrombomodulin, sICAM-1, sICAM-3) and LGI (hs-CRP, SAA, IL-6, IL-8, TNF-α) were measured with a single- or multiplex array detection system based on electro-chemiluminescence technology. Results: In linear regression analyses adjusted for potential confounders, participants with ESRD had higher levels of most serum biomarkers of ED and LGI than controls. In addition, in CKD5-HD levels of serum biomarkers of ED and LGI were largely similar to those in CKD5-ND. In contrast, in CKD5-PD levels of biomarkers of ED were higher than in CKD5-ND and CKD5-HD. Similarly, in linear mixed model analyses sVCAM-1, thrombomodulin, sICAM-1 and sICAM-3 increased after PD initiation. In contrast, incident HD patients showed an increase in sVCAM-1, P-selectin and TNF-α, but a decline of hs-CRP, SAA and IL-6. Further, following kidney transplantation sVCAM-1, thrombomodulin, sICAM-3 and TNF-α were lower at three months post-transplantation and remained stable in the three months thereafter. Conclusions: Levels of serum biomarkers of ED and LGI were higher in ESRD as compared with controls. In addition, PD initiation and, less convincingly, HD initiation may increase levels of selected serum biomarkers of ED and LGI on top of uremia per se. In contrast to dialysis, several serum biomarkers of ED and LGI markedly declined following kidney transplantation. [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]

Lymphatic filariasis (LF), a morbid disease caused by the tissue-invasive nematodes Wuchereria bancrofti, Brugia malayi, and Brugia timori, affects millions of people worldwide. Global eradication efforts have significantly reduced worldwide prevalence, but complete elimination has been hampered by limitations of current anti-filarial drugs and the lack of a vaccine. The goal of this study was to evaluate B. malayi intestinal UDP-glucuronosyltransferase (Bm-UGT) as a potential therapeutic target. To evaluate whether Bm-UGT is essential for adult filarial worms, we inhibited its expression using siRNA. This resulted in a 75% knockdown of Bm-ugt mRNA for 6 days and almost complete suppression of detectable Bm-UGT by immunoblot. Reduction in Bm-UGT expression resulted in decreased worm motility for 6 days, 70% reduction in microfilaria release from adult worms, and significant reduction in adult worm metabolism as detected by MTT assays. Because prior allergic-sensitization to a filarial antigen would be a contraindication for its use as a vaccine candidate, we tested plasma from infected and endemic normal populations for Bm-UGT-specific IgE using a luciferase immunoprecipitation assay. All samples (n = 35) tested negative. We then tested two commercially available medicines known to be broad inhibitors of UGTs, sulfinpyrazone and probenecid, for in vitro activity against B. malayi. There were marked macrofilaricidal effects at concentrations achievable in humans and very little effect on microfilariae. In addition, we observed that probenecid and sulfinpyrazone exhibit a synergistic macrofilaricidal effect when used in combination with albendazole. The results of this study demonstrate that Bm-UGT is an essential protein for adult worm survival. Lack of prior IgE sensitization in infected and endemic populations suggest it may be a feasible vaccine candidate. The finding that sulfinpyrazone and probenecid have in vitro effects against adult B. malayi worms suggests that these medications have promise as potential macrofilaricides in humans. [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: The inflammatory contribution to type 2 diabetes (T2D) has suggested new therapeutic targets using biologic drugs designed for rheumatoid arthritis (RA). On this basis, we aimed at investigating whether interleukin-1 (IL-1) inhibition with anakinra, a recombinant human IL-1 receptor antagonist, could improve both glycaemic and inflammatory parameters in participants with RA and T2D compared with tumour necrosis factor (TNF) inhibitors (TNFis).Methods and Findings: This study, designed as a multicentre, open-label, randomised controlled trial, enrolled participants, followed up for 6 months, with RA and T2D in 12 Italian rheumatologic units between 2013 and 2016. Participants were randomised to anakinra or to a TNFi (i.e., adalimumab, certolizumab pegol, etanercept, infliximab, or golimumab), and the primary end point was the change in percentage of glycated haemoglobin (HbA1c%) (EudraCT: 2012-005370-62 ClinicalTrial.gov: NCT02236481). In total, 41 participants with RA and T2D were randomised, and 39 eligible participants were treated (age 62.72 ± 9.97 years, 74.4% female sex). The majority of participants had seropositive RA disease (rheumatoid factor and/or anticyclic citrullinated peptide antibody [ACPA] 70.2%) with active disease (Disease Activity Score-28 [DAS28]: 5.54 ± 1.03; C-reactive protein 11.84 ± 9.67 mg/L, respectively). All participants had T2D (HbA1c%: 7.77 ± 0.70, fasting plasma glucose: 139.13 ± 42.17 mg). When all the enrolled participants reached 6 months of follow-up, the important crude difference in the main end point, confirmed by an unplanned ad interim analysis showing the significant effects of anakinra, which were not observed in the other group, led to the study being stopped for early benefit. Participants in the anakinra group had a significant reduction of HbA1c%, in an unadjusted linear mixed model, after 3 months (β: -0.85, p < 0.001, 95% CI -1.28 to -0.42) and 6 months (β: -1.05, p < 0.001, 95% CI -1.50 to -0.59). Similar results were observed adjusting the model for relevant RA and T2D clinical confounders (male sex, age, ACPA positivity, use of corticosteroids, RA duration, T2D duration, use of oral antidiabetic drug, body mass index [BMI]) after 3 months (β: -1.04, p < 0.001, 95% CI -1.52 to -0.55) and 6 months (β: -1.24, p < 0.001, 95% CI -1.75 to -0.72). Participants in the TNFi group had a nonsignificant slight decrease of HbA1c%. Assuming the success threshold to be HbA1c% ≤ 7, we considered an absolute risk reduction (ARR) = 0.42 (experimental event rate = 0.54, control event rate = 0.12); thus, we estimated, rounding up, a number needed to treat (NNT) = 3. Concerning RA, a progressive reduction of disease activity was observed in both groups. No severe adverse events, hypoglycaemic episodes, or deaths were observed. Urticarial lesions at the injection site led to discontinuation in 4 (18%) anakinra-treated participants. Additionally, we observed nonsevere infections, including influenza, nasopharyngitis, upper respiratory tract infection, urinary tract infection, and diarrhoea in both groups. Our study has some limitations, including open-label design and previously unplanned ad interim analysis, small size, lack of some laboratory evaluations, and ongoing use of other drugs.Conclusions: In this study, we observed an apparent benefit of IL-1 inhibition in participants with RA and T2D, reaching the therapeutic targets of both diseases. Our results suggest the concept that IL-1 inhibition may be considered a targeted treatment for RA and T2D.Trial Registration: The trial is registered with EU Clinical Trials Register, EudraCT Number: 2012-005370-62 and with ClinicalTrial.gov, number NCT02236481. [ABSTRACT FROM AUTHOR]

Although doxorubicin (DOX)-induced cardiomyopathy causes lethal heart failure (HF), no early detection or effective treatment methods are available. The principal mechanisms of cardiotoxicity are considered to involve oxidative stress and apoptosis of cardiomyocytes. However, the effect of DOX on cardiac fibroblasts at non-lethal concentrations remains unknown. The aim of this study was to investigate the direct effect of doxorubicin on the activation of cardiac fibroblasts independent of cell death pathways. We first found that DOX induced α-SMA expression (marker of trans-differentiation) at a low concentration range, which did not inhibit cell viability. DOX also increased MMP1, IL-6, TGF-β and collagen expression in human cardiac fibroblasts (HCFs). In addition, DOX promoted Akt and Smad phosphorylation. A Smad inhibitor prevented DOX-induced α-SMA and IL-6 protein expression. An PI3K inhibitor also prevented MMP1 mRNA expression in HCFs. These findings suggest that DOX directly induces fibrotic changes in HCFs via cell death-independent pathways. Furthermore, we confirmed that these responses are organ- and species-specific for HCFs based on experiments using different types of human and murine fibroblast cell lines. These results suggest potentially new mechanisms of DOX-induced cardiotoxicity from the viewpoint of fibrotic changes in cardiac fibroblasts. [ABSTRACT FROM AUTHOR]

Introduction: The primary rational for using mesenchymal stromal cells (MSCs) to rejuvenate damaged tissue is mostly based on their capacity to trans-differentiate and repair injured organs. However, previous studies have demonstrated that MSCs are beneficial even at very early stages, before differentiation and proliferation can be expected. The aim of the current study was to investigate the multifaceted immunological effects of systemically administrating MSCs in the setting of acute kidney injury (AKI) induced by ischemic-reperfusion (I/R). Methods: A rat model of I/R induced AKI was used. The rats underwent a unilateral nephrectomy with simultaneously clamping the contralateral kidney for 60 minutes. Four treatment groups received intravenously, increasing doses of human MSCs and after 48 hours, the rats were sacrificed. Blood was taken to evaluate renal functions and to measure systemic inflammatory markers. Kidneys were taken for histopathologic examinations and evaluations of intra-renal complement activation and inflammatory mediators. Results: Renal functions improved in U shaped dose dependent manner. Mean serum creatinine levels were 4.5, 2.9, 2.6, 1.7 and 4.1 mg/dL in I/R + placebo, I/R + 150x103 cells, I/R + 250x103 cells, I/R + 500x103 cells and I/R + 1,000x103 cells respectfully (p-values<0.05). Urea demonstrated consistent results with the same U shape improvement manner. The extensive activation of the complement system was ameliorated in the MSCs treatment groups. In addition, MSCs significantly decreased intra-renal levels of IL-1β and TNF-α. It should be noted that the highest doses of MSCs induced renal hypoxia, marked by the Hypoxy-probe staining. Conclusions: The early beneficial effect of MSCs in the setting of AKI may be attributed to their immunomodulatory effects. Safe treatment with MSCs can block the deleterious activation of the complement cascade and alleviate the hazardous inflammatory mediator-related cascade. [ABSTRACT FROM AUTHOR]

Multi-marker tests hold promise for identifying symptomatic women at risk of imminent preterm delivery (PTD, <37 week’s gestation). This study sought to determine the relationship of inflammatory mediators and metabolites in cervicovaginal fluid (CVF) with spontaneous PTD (sPTD) and delivery within 14 days of presentation with symptoms of preterm labour (PTL). CVF samples from 94 (preterm = 19, term = 75) singleton women with symptoms of PTL studied between 19+0–36+6 weeks’ gestation were analysed for cytokines/chemokines by multiplexed bead-based immunoassay, while metabolites were quantified by enzyme-based spectrophotometry in a subset of 61 women (preterm = 16, term = 45). Prevalence of targeted vaginal bacterial species was determined for 70 women (preterm = 14, term = 66) by PCR. Overall, 10 women delivered within 14 days of sampling. Predictive capacities of individual biomarkers and cytokine-metabolite combinations for sPTD and delivery within 14 days of sampling were analysed by logistic regression models and area under the receiver operating characteristic curve. Fusobacterium sp., Mubiluncus mulieris and Mycoplasma hominis were detected in more preterm-delivered than term women (P<0.0001), while, M. curtisii was found in more term-delivered than preterm women (P<0.0001). RANTES (0.91, 0.65–1.0), IL-6 (0.79, 0.67–0.88), and Acetate/Glutamate ratio (0.74, 0.61–0.85) were associated with delivery within 14 days of sampling (AUC, 95% CI). There were significant correlations between cytokines and metabolites, and several cytokine-metabolite combinations were associated with sPTD or delivery within 14 days of sampling (e.g. L/D-lactate ratio+Acetate/Glutamate ratio+IL-6: 0.84, 0.67–0.94). Symptomatic women destined to deliver preterm and within 14 days of sampling express significantly higher pro-inflammatory mediators at mid to late gestation. In this cohort, IL-6, Acetate/Glutamate ratio and RANTES were associated with delivery within 14 days of sampling, consistent with their roles in modulating infection-inflammation-associated preterm labour in women presenting with symptoms of preterm birth. Replication of these observations in larger cohorts of women could show potential clinical utility. [ABSTRACT FROM AUTHOR]

Vaccines that elicit broadly cross-neutralizing antibodies, including antibodies that target the conserved stem of hemagglutinin (HA), are being developed as a strategy for next-generation influenza vaccines that protect against influenza across multiple years. However, efficient induction of cross-neutralizing antibodies remains a challenge, and potential escape mutations have not been well characterized. Here we elicited cross-neutralizing antibodies by immunizing animals with the hemagglutinins from H5 and H9 subtype influenza A viruses that are sensitive to neutralization by stem antibodies. We further isolated and characterized an HA stem monoclonal antibody 4C2 that broadly neutralizes group 1 influenza viruses and identified HA mutations that reduced sensitivity to stem antibodies. Our results offer insights for next-generation influenza vaccine strategies for inducing cross-neutralizing antibodies. [ABSTRACT FROM AUTHOR]

Background: Different psychiatric diagnostic groups have been reported to have cytokine levels deviating from healthy controls. In acute clinical settings however, the specific challenging symptoms and signs are more important than a diagnostic group. Thus, exploration of cytokines and immune activity and their role in specific symptoms is important. Reports in this field so far are sparse. Objective: In the present study, we aimed to examine the association between immune activity measured as levels of cytokines and agitation (independent of diagnostic group) in patients admitted to an acute psychiatric inpatient department. Methods: A total of 316 patients admitted to an acute psychiatric inpatient department were included. Thirty-nine patients with psychosis were subject to subgroup analyses. Agitation was assessed by the Positive and Negative Syndrome Scale, Excitement Component (PANSS-EC). Based on PANNS-EC patients were stratified into two groups: 67 agitated patients and 249 non-agitated patients. Serum concentrations of the following immune markers were measured: interleukin (IL) -1β, IL-4, IL-6, IL-10, tumor necrosis factor (TNF) -α, interferon (IFN) -γ and transforming growth factor (TGF) -β. Results: Serum levels of TNF-α were significantly higher in patients with agitation compared to those without, both when all patients were included in the analyses (p = 0.004) and in the psychosis group (p = 0.027). After correcting for multiple testing, only the findings in the total population remained significant. Conclusions: Our findings suggest an association between TNF-α and agitation in an acute psychiatric population. A similar trend was reproduced to the psychosis subgroup. This suggests that agitation might be an independent entity associated with cytokines across different diagnostic groups. [ABSTRACT FROM AUTHOR]

This study aimed to examine the macrophage phenotype and its relationship to renal function and histological changes in human DN and the effect of TREM-1 on high-glucose-induced macrophage activation. We observed that in renal tissue biopsies, the expression of CD68 and M1 was apparent in the glomeruli and interstitium, while accumulation of M2 and TREM-1 was primarily observed in the interstitium. The numbers of CD68, M1, and M2 macrophages infiltrating in the DN group were increased in a process-dependent manner compared with the control group, and the intensities of the infiltrates were proportional to the rate of subsequent decline in renal function. M1 macrophages were recruited into the kidney at an early stage (I+IIa) of DN. The M1-to-M2 macrophage ratio peaked at this time, whereas M2 macrophages predominated at later time points (III) when the percentage of M1/M2 macrophages was at its lowest level. In an in vitro study, we showed that under high glucose conditions, macrophages began to up-regulate their expression of TREM-1, M1, and marker iNOS and decreased the M2 marker MR. However, the above effects of high-glucose were abolished when TREM-1 expression was inhibited by TREM-1 siRNA. In conclusion, our study demonstrated that there was a positive correlation between the M1/M2 activation state and the progress of DN, and TREM-1 played an important role in high-glucose-induced macrophage phenotype transformation. [ABSTRACT FROM AUTHOR]

The development of effective malaria vaccines remains a global health priority. Currently, the most advanced vaccine, known as RTS,S, has only shown modest efficacy in clinical trials. Thus, the development of more efficacious vaccines by improving the formulation of RTS,S for increased efficacy or to interrupt malaria transmission are urgently needed. The RTS,S vaccine is based on the presentation of a fragment of the sporozoite antigen on the surface of virus-like particles (VLPs) based on human hepatitis B virus (HBV). In this study, we have developed and evaluated a novel VLP platform based on duck HBV (known as Metavax) for malaria vaccine development. This platform can incorporate large and complex proteins into VLPs and is produced in a Hansenula cell line compatible with cGMP vaccine production. Here, we have established the expression of leading P. falciparum malaria vaccine candidates as VLPs. This includes Pfs230 and Pfs25, which are candidate transmission-blocking vaccine antigens. We demonstrated that the VLPs effectively induce antibodies to malaria vaccine candidates with minimal induction of antibodies to the duck-HBV scaffold antigen. Antibodies to Pfs230 also recognised native protein on the surface of gametocytes, and antibodies to both Pfs230 and Pfs25 demonstrated transmission-reducing activity in standard membrane feeding assays. These results establish the potential utility of this VLP platform for malaria vaccines, which may be suitable for the development of multi-component vaccines that achieve high vaccine efficacy and transmission-blocking immunity. [ABSTRACT FROM AUTHOR]