Immunogenicity of Self-Associated Aggregates and Chemically Cross-Linked Conjugates of the 42 kDa Plasmodium falciparum Merozoite Surface Protein-1

Self-associated protein aggregates or cross-linked protein conjugates are, in general, more immunogenic than oligomeric or monomeric forms. In particular, the immunogenicity in mice of a recombinant malaria transmission blocking vaccine candidate, the ookinete specific Plasmodium falciparum 25 kDa protein (Pfs25), was increased more than 1000-fold when evaluated as a chemical cross-linked protein-protein conjugate as compared to a formulated monomer. Whether alternative approaches using protein complexes improve the immunogenicity of other recombinant malaria vaccine candidates is worth assessing. In this work, the immunogenicity of the recombinant 42 kDa processed form of the P. falciparum merozoite surface protein 1 (MSP142) was evaluated as a self-associated, non-covalent aggregate and as a chemical cross-linked protein-protein conjugate to ExoProtein A, which is a recombinant detoxified form of Pseudomonas aeruginosa exotoxin A. MSP142 conjugates were prepared and characterized biochemically and biophysically to determine their molar mass in solution and stoichiometry, when relevant. The immunogenicity of the MSP142 self-associated aggregates, cross-linked chemical conjugates and monomers were compared in BALB/c mice after adsorption to aluminum hydroxide adjuvant, and in one instance in association with the TLR9 agonist CPG7909 with an aluminum hydroxide formulation. Antibody titers were assessed by ELISA. Unlike observations made for Pfs25, no significant enhancement in MSP142 specific antibody titers was observed for any conjugate as compared to the formulated monomer or dimer, except for the addition of the TLR9 agonist CPG7909. Clearly, enhancing the immunogenicity of a recombinant protein vaccine candidate by the formation of protein complexes must be established on an empirical basis

Very Low-mass Stellar and Substellar Companions to Solar-like Stars from MARVELS II: A Short-period Companion Orbiting an F Star with Evidence of a Stellar Tertiary And Significant Mutual Inclination

We report the discovery via radial velocity of a short-period (P = 2.430420 \pm 0.000006 days) companion to the F-type main sequence star TYC 2930-00872-1. A long-term trend in the radial velocities indicates the presence of a tertiary stellar companion with $P > 2000$ days. High-resolution spectroscopy of the host star yields T_eff = 6427 +/- 33 K, log(g) = 4.52 +/- 0.14, and [Fe/H]=-0.04 +/- 0.05. These parameters, combined with the broad-band spectral energy distribution and parallax, allow us to infer a mass and radius of the host star of M_1=1.21 +/- 0.08 M_\odot and R_1=1.09_{-0.13}^{+0.15} R_\odot. We are able to exclude transits of the inner companion with high confidence. The host star's spectrum exhibits clear Ca H and K core emission indicating stellar activity, but a lack of photometric variability and small v*sin(I) suggest the primary's spin axis is oriented in a pole-on configuration. The rotational period of the primary from an activity-rotation relation matches the orbital period of the inner companion to within 1.5 \sigma, suggesting they are tidally locked. If the inner companion's orbital angular momentum vector is aligned with the stellar spin axis, as expected through tidal evolution, then it has a stellar mass of M_2 ~ 0.3-0.4 M_\odot. Direct imaging limits the existence of stellar companions to projected separations < 30 AU. No set of spectral lines and no significant flux contribution to the spectral energy distribution from either companion are detected, which places individual upper mass limits of M < 1.0 M_\odot, provided they are not stellar remnants. If the tertiary is not a stellar remnant, then it likely has a mass of ~0.5-0.6 M_\odot, and its orbit is likely significantly inclined from that of the secondary, suggesting that the Kozai-Lidov mechanism may have driven the dynamical evolution of this system.Comment: 37 pages, 7 tables, 21 figures, Accepted in A

Rapid detection of multidrug-resistant Mycobacterium tuberculosis in Cotonou (Benin) using two low-cost colorimetric methods: resazurin and nitrate reductase assays

We have evaluated two simple, rapid and low-cost colorimetric methods for the detection of multidrug-resistant Mycobacterium tuberculosis. A total of 151 M. tuberculosis strains were tested for resistance to rifampicin (RMP) and isoniazid by resazurin microplate assay (REMA) and nitrate reductase assay (NRA) in comparison with the conventional proportion method (PM) on Löwenstein-Jensen medium. A complete agreement was found between NRA and PM, while one false RMP-susceptible result was found by REMA. REMA and NRA tests are rapid and inexpensive, and could be good alternatives to the conventional PM in low-resource countries

Iatrogenic Botulism Outbreak in Egypt due to a Counterfeit Botulinum Toxin A Preparation - A Descriptive Series of Patient Features and Outcome

International audienc

Addition of CpG ODN to recombinant Pseudomonas aeruginosa ExoProtein A conjugates of AMA1 and Pfs25 greatly increases the number of responders

Both the blood stage protein apical membrane antigen 1 (AMA1) and the 25 kDa sexual stage protein (Pfs25) of Plasmodium falciparum are two leading candidates in malarial vaccine development. We have previously demonstrated that conjugation of these malarial antigens to recombinant Pseudomonas aeruginosa ExoProtein A (rEPA) significantly increased the mean specific functional antibody responses in mice; however, some mice responded poorly and were unable to demonstrate a functional response. We hypothesized that the immunogenicities of these two malarial antigens could be further enhanced by inclusion of a CpG oligodeoxynucleotide in the formulation. Mice were immunized with either rEPA conjugated or unconjugated AMA1 and Pfs25 formulated on Alhydrogel with or without the addition of CPG 7909. Mice received the formulations on days 0 and 28, and mouse sera were collected on day 42. ELISA analyses on these sera showed that the addition of CPG 7909 to AMA1-rEPA and Pfs25-rEPA formulated on Alhydrogel induced significantly higher mean antibody titers than the formulations without CPG 7909, and led to a mixed Th1/Th2 response as demonstrated by the production of mouse IgG1 and IgG2a subclasses. The presence of CPG 7909 in the formulations of both conjugated antigens greatly increased the proportion of responders with antibody titers sufficient to inhibit blood-stage parasite growth in vitro or block transmission of sexual stage parasites to mosquitoes. The results obtained in this study indicate the potential use of a combination strategy to increase the number of responders to malarial antigens in humans

Very low mass stellar and substellar companions to solar-like stars from MARVELS. II. A short-period companios orbiting an F star with evidence of a stellar tertiary and significant mutual inclination

We report the discovery via radial velocity (RV) measurements of a short-period (P = 2.430420±0.000006 days) companion to the F-type main-sequence star TYC 2930-00872-1. A long-term trend in the RV data also suggests the presence of a tertiary stellar companion with P > 2000 days. High-resolution spectroscopy of the host star yields Teff = 6427 ± 33 K, log g = 4.52 ± 0.14, and [Fe/H] = −0.04 ± 0.05. These parameters, combined with the broadband spectral energy distribution (SED) and a parallax, allow us to infer a mass and radius of the host star of M1 = 1.21 ± 0.08 M and R1 = 1.09+0.15 −0.13 R . The minimum mass of the inner companion is below the hydrogen-burning limit; however, the true mass is likely to be substantially higher. We are able to exclude transits of the inner companion with high confidence. Further, the host star spectrum exhibits a clear signature of Ca H and K core emission, indicating stellar activity, but a lack of photometric variability and small v sin I suggest that the primary’s spin axis is oriented in a pole-on configuration. The rotational period of the primary estimated through an activity–rotation relation matches the orbital period of the inner companion to within 1.5 σ, suggesting that the primary and inner companion are tidally locked. If the inner companion’s orbital angular momentum vector is aligned with the stellar spin axis as expected through tidal evolution, then it has a stellar mass of ∼0.3–0.4 M . Direct imaging limits the existence of stellar companions to projected separations <30 AU. No set of spectral lines and no significant flux contribution to the SED from either companion are detected, which places individual upper mass limits of M{2,3} 1.0 M , provided they are not stellar remnants. If the tertiary is not a stellar remnant, then it likely has a mass of ∼0.5–0.6M , and its orbit is likely significantly inclined from that of the secondary, suggesting that the Kozai–Lidov mechanism may have driven the dynamical evolution of this system

Murine Cerebral Malaria Development Is Independent of Toll-Like Receptor Signaling

Malaria pigment hemozoin was reported to activate the innate immunity by Toll-like receptor (TLR)-9 engagement. However, the role of TLR activation for the development of cerebral malaria (CM), a lethal complication of malaria infection in humans, is unknown. Using Plasmodium berghei ANKA (PbA) infection in mice as a model of CM, we report here that TLR9-deficient mice are not protected from CM. To exclude the role of other members of the TLR family in PbA recognition, we infected mice deficient for single TLR1, -2, -3, -4, -6, -7, or -9 and their adapter proteins MyD88, TIRAP, and TRIF. In contrast to lymphotoxin α-deficient mice, which are resistant to CM, all TLR-deficient mice were as sensitive to fatal CM development as wild-type control mice and developed typical microvascular damage with vascular leak and hemorrhage in the brain and lung, together with comparable parasitemia, thrombocytopenia, neutrophilia, and lymphopenia. In conclusion, the present data do not exclude the possibility that malarial molecular motifs may activate the innate immune system. However, TLR-dependent activation of innate immunity is unlikely to contribute significantly to the proinflammatory response to PbA infection and the development of fatal CM