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The most advanced monoclonal antibodies (mAbs) and vaccines against malaria target the central repeat region or closely related sequences within the Plasmodium falciparum circumsporozoite protein (PfCSP). Here, using an antigen-agnostic strategy to investigate human antibody responses to whole sporozoites, we identified a class of mAbs that target a cryptic PfCSP epitope that is only exposed after cleavage and subsequent pyroglutamylation (pGlu) of the newly formed N terminus. This pGlu-CSP epitope is not targeted by current anti-PfCSP mAbs and is not included in the licensed malaria vaccines. MAD21-101, the most potent mAb in this class, confers sterile protection against Pf infection in a human liver-chimeric mouse model. These findings reveal a site of vulnerability on the sporozoite surface that can be targeted by next-generation antimalarial interventions.

Aims: To investigate the associations between baseline/changes in serum uric acid (sUA) and the risks for cardiovascular/microvascular outcomes and mortality in a type 2 diabetes cohort., Methods: Baseline sUA was measured in 685 individuals, and 463 had a second sUA measurement during follow-up; sUA was analyzed as a continuous variable and categorized into sex-specific tertile subgroups and low/high levels (>4.5 mg/dl women; >5.5 mg/dl men). The risks associated with baseline sUA and its changes were examined by Cox analyses for all outcomes., Results: Median follow up was 10.7 years, there were 173 major cardiovascular events (MACEs), 268 all-cause deaths, 127 microalbuminuria, 104 renal failure, 160 retinopathy and 178 peripheral neuropathy outcomes. Baseline sUA was predictor of all outcomes, except all-cause mortality and retinopathy. In tertile and high/low sUA analyses, the hazard ratios (HRs) varied from 1.6 (microalbuminuria development) to 2.4 (MACEs; cardiovascular mortality). There was interaction with sex for MACEs, an increased risk was observed in women (HR: 2.6), but not in men (HR: 1.2). Changes in sUA were associated with the renal failure (HR: 2.4)., Conclusions: In a prospective cohort, high baseline sUA was a predictor of cardiovascular, renal and peripheral neuropathy. However, sUA changes were only predictor of renal failure., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Human monoclonal antibodies (hmAbs) targeting the Plasmodium falciparum circumsporozoite protein (PfCSP) on the sporozoite surface are a promising tool for preventing malaria infection. However, their mechanisms of protection remain unclear. Here, using 13 distinctive PfCSP hmAbs, we provide a comprehensive view of how PfCSP hmAbs neutralize sporozoites in host tissues. Sporozoites are most vulnerable to hmAb-mediated neutralization in the skin. However, rare but potent hmAbs additionally neutralize sporozoites in the blood and liver. Efficient protection in tissues mainly associates with high-affinity and high-cytotoxicity hmAbs inducing rapid parasite loss-of-fitness in the absence of complement and host cells in vitro. A 3D-substrate assay greatly enhances hmAb cytotoxicity and mimics the skin-dependent protection, indicating that the physical stress imposed on motile sporozoites by the skin is crucial for unfolding the protective potential of hmAbs. This functional 3D cytotoxicity assay can thus be useful for downselecting potent anti-PfCSP hmAbs and vaccines., Competing Interests: Declaration of interests A.N. is a founder of Scians Ltd. R.A.S., A.H.I., and B.J.F. hold a patent describing antibody CIS43, issued on June 1, 2021; International Application No. PCT/US2018/017826. R.A.S., L.T.W., and J.R.F. hold a patent describing antibody L9, issued on May 4, 2020; International Application No. PCT/US2020/031345., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Monoclonal antibody L9 recognizes the Plasmodium falciparum circumsporozoite protein (PfCSP) and is highly protective following controlled human malaria challenge. To gain insight into its function, we determined cryoelectron microscopy (cryo-EM) structures of L9 in complex with full-length PfCSP and assessed how this recognition influenced protection by wild-type and mutant L9s. Cryo-EM reconstructions at 3.6- and 3.7-Å resolution revealed L9 to recognize PfCSP as an atypical trimer. Each of the three L9s in the trimer directly recognized an Asn-Pro-Asn-Val (NPNV) tetrapeptide on PfCSP and interacted homotypically to facilitate L9-trimer assembly. We analyzed peptides containing different repeat tetrapeptides for binding to wild-type and mutant L9s to delineate epitope and homotypic components of L9 recognition; we found both components necessary for potent malaria protection. Last, we found the 27-residue stretch recognized by L9 to be highly conserved in P. falciparum isolates, suggesting the newly revealed complete L9 epitope to be an attractive vaccine target., Competing Interests: Declaration of interests L.T.W. and R.A.S. have submitted a US provisional patent application 62/842,590, filed May 3, 2019, describing antibody L9., (Published by Elsevier Ltd.)

Background: New approaches for the prevention and elimination of malaria, a leading cause of illness and death among infants and young children globally, are needed., Methods: We conducted a phase 1 clinical trial to assess the safety and pharmacokinetics of L9LS, a next-generation antimalarial monoclonal antibody, and its protective efficacy against controlled human malaria infection in healthy adults who had never had malaria or received a vaccine for malaria. The participants received L9LS either intravenously or subcutaneously at a dose of 1 mg, 5 mg, or 20 mg per kilogram of body weight. Within 2 to 6 weeks after the administration of L9LS, both the participants who received L9LS and the control participants underwent controlled human malaria infection in which they were exposed to mosquitoes carrying Plasmodium falciparum (3D7 strain)., Results: No safety concerns were identified. L9LS had an estimated half-life of 56 days, and it had dose linearity, with the highest mean (±SD) maximum serum concentration (C max ) of 914.2±146.5 μg per milliliter observed in participants who had received 20 mg per kilogram intravenously and the lowest mean C max of 41.5±4.7 μg per milliliter observed in those who had received 1 mg per kilogram intravenously; the mean C max was 164.8±31.1 in the participants who had received 5 mg per kilogram intravenously and 68.9±22.3 in those who had received 5 mg per kilogram subcutaneously. A total of 17 L9LS recipients and 6 control participants underwent controlled human malaria infection. Of the 17 participants who received a single dose of L9LS, 15 (88%) were protected after controlled human malaria infection. Parasitemia did not develop in any of the participants who received 5 or 20 mg per kilogram of intravenous L9LS. Parasitemia developed in 1 of 5 participants who received 1 mg per kilogram intravenously, 1 of 5 participants who received 5 mg per kilogram subcutaneously, and all 6 control participants through 21 days after the controlled human malaria infection. Protection conferred by L9LS was seen at serum concentrations as low as 9.2 μg per milliliter., Conclusions: In this small trial, L9LS administered intravenously or subcutaneously protected recipients against malaria after controlled infection, without evident safety concerns. (Funded by the National Institute of Allergy and Infectious Diseases; VRC 614 ClinicalTrials.gov number, NCT05019729.)., (Copyright © 2022 Massachusetts Medical Society.)

The monoclonal antibody CIS43 targets the Plasmodium falciparum circumsporozoite protein (PfCSP) and prevents malaria infection in humans for up to 9 mo following a single intravenous administration. To enhance the potency and clinical utility of CIS43, we used iterative site-saturation mutagenesis and DNA shuffling to screen precise gene-variant yeast display libraries for improved PfCSP antigen recognition. We identified several mutations that improved recognition, predominately in framework regions, and combined these to produce a panel of antibody variants. The most improved antibody, CIS43_Var10, had three mutations and showed approximately sixfold enhanced protective potency in vivo compared to CIS43. Co-crystal and cryo-electron microscopy structures of CIS43_Var10 with the peptide epitope or with PfCSP, respectively, revealed functional roles for each of these mutations. The unbiased site-directed mutagenesis and screening pipeline described here represent a powerful approach to enhance protective potency and to enable broader clinical use of antimalarial antibodies., (© 2022 Banach et al.)

Methyl acetate is considered a prototype molecule to study biodiesel ignition and combustion and is seen as a possible fuel or fuel additive. For this reason, methyl acetate decomposition paths have been investigated in recent years. In the present study, hydrogen abstraction reactions on methyl acetate by OH( 2 Π), HO 2 ( 2 A'), H( 2 Σ g ), O( 3 P), and O 2 ( 3 Σ g ) were conducted, and the effect of methodology and anharmonic corrections on the rate coefficients were evaluated. The M06-2X and B3LYP-D3 functionals with the cc-pVDZ, cc-pVTZ, aug-cc-pVDZ, and aug-cc-pVTZ basis set were used for methodology evaluation and the CCSD(T)/cc-pVTZ//M06-2X/aug-cc-pVTZ for the rate coefficients calculation and literature comparisons. The rate coefficients were calculated in the range of 250-3500 K, including tunneling corrections, methyl-hindered rotations, and anharmonic effects calculated by the VPT2 method. The methodology analysis showed that the B3LYP-D3 functional leads to lower activation energies for all elementary reactions studied, and the double-zeta basis is insufficient to calculate precise rate coefficients. The inclusion of anharmonic corrections consistently lowered the rate coefficients of all elementary reactions studied and changed the Arrhenius plot profile with the temperature. Significant anharmonic effects were observed at higher temperatures, being the reaction with O 2 ( 3 Σ g ) the most affected by this correction. Differences superior to 10 5 cm 3 molec -1  s -1 in the rate coefficients were observed in some cases., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Plant defensins and lipid transfer proteins (LTPs) constitute a large and evolutionarily diverse family of antimicrobial peptides. Defensins and LTPs are two pathogenesis-related proteins (PR proteins) whose characterization may help to uncover aspects about the sugarcane response to pathogens attack. LTPs have also been investigated for their participation in the response to different types of stress. Despite the important roles of defensins and LTPs in biotic and abiotic stresses, scarce knowledge is found about these proteins in sugarcane. By using bioinformatics approaches, we characterized defensins and LTPs in the sugarcane wild species and modern cultivar genomes. The identification of defensins and LTPs showed that all five defensins groups and eight of the nine LTPs have their respective genes loci, although some was only identified in the cultivar genome. Phylogenetic analysis showed that defensins appear to be more conserved among groups of plants than LTPs. Some defensins and LTPs showed opposite expression during pathogenic and benefic bacterial interactions. Interestingly, the expression of defensins and LTPs in shoots and roots was completely different in plants submitted to benefic bacteria or water depletion. Finally, the modeling and comparison of isoforms of LTPs and defensins in wild species and cultivars revealed a high conservation of tertiary structures, with variation of amino acids in different regions of proteins, which could impact their antimicrobial activity. Our data contributed to the characterization of defensins and LTPs in sugarcane and provided new elements for understanding the involvement of these proteins in sugarcane response to different types of stress., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Repeat antigens, such as the Plasmodium falciparum circumsporozoite protein (PfCSP), use both sequence degeneracy and structural diversity to evade the immune response. A few PfCSP-directed antibodies have been identified that are effective at preventing malaria infection, including CIS43, but how these repeat-targeting antibodies might be improved has been unclear. Here, we engineered a humanized mouse model in which B cells expressed inferred human germline CIS43 (iGL-CIS43) B cell receptors and used both vaccination and bioinformatic analysis to obtain variant CIS43 antibodies with improved protective capacity. One such antibody, iGL-CIS43.D3, was significantly more potent than the current best-in-class PfCSP-directed antibody. We found that vaccination with a junctional epitope peptide was more effective than full-length PfCSP at recruiting iGL-CIS43 B cells to germinal centers. Structure-function analysis revealed multiple somatic hypermutations that combinatorically improved protection. This mouse model can thus be used to understand vaccine immunogens and to develop highly potent anti-malarial antibodies., Competing Interests: Declaration of interests S.K., P.T., R.R., M.R., P.D.K., R.A.S. and F.D.B. have submitted a US Provisional Patent Application describing improved CIS43 antibodies (filed November 5, 2021). B.J.F., R.A.S., A.H.I., and N.K.K. hold patents on CIS43 (International Application No. PCT/US2018/017826; US Patent Application No. 16/485,354; issued June 1, 2021). L.T.W., R.A.S., and J.R.F. have submitted a US Provisional Patent Application describing mAb L9 (62/842,590; filed May 3, 2019). The remaining authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

The most advanced malaria vaccine, RTS,S, includes the central repeat and C-terminal domains of the Plasmodium falciparum circumsporozoite protein (PfCSP). We have recently isolated human antibodies that target the junctional region between the N-terminal and repeat domains that are not included in RTS,S. Due to the fact that these antibodies protect against malaria challenge in mice, their epitopes could be effective vaccine targets. Here, we developed immunogens displaying PfCSP junctional epitopes by genetic fusion to either the N-terminus or B domain loop of the E2 protein from chikungunya (CHIK) alphavirus and produced CHIK virus-like particles (CHIK-VLPs). The structural integrity of these junctional-epitope-CHIK-VLP immunogens was confirmed by negative-stain electron microscopy. Immunization of these CHIK-VLP immunogens reduced parasite liver load by up to 95% in a mouse model of malaria infection and elicited better protection than when displayed on keyhole limpet hemocyanin, a commonly used immunogenic carrier. Protection correlated with PfCSP serum titer. Of note, different junctional sequences elicited qualitatively different reactivities to overlapping PfCSP peptides. Overall, these results show that the junctional epitopes of PfCSP can induce protective responses when displayed on CHIK-VLP immunogens and provide a basis for the development of a next generation malaria vaccine to expand the breadth of anti-PfCSP immunity.

In recent years, the antimicrobial activity of peptides isolated from a wide variety of organs from plant species has been reported. However, a few studies have investigated the potential of antimicrobial peptides (AMPs) found in fruits, especially Capsicum chinense (pepper). The present study aimed to purify and characterize peptides from Capsicum chinense fruits and evaluate their inhibitory activities against different phytopathogenic fungi and also analyze the possible mechanisms of action involved in microbial inhibition. After fruit protein extraction and high-performance liquid chromatography (HPLC), different fractions were obtained, named F1 to F10. Peptides in the F4 and F5 fractions were sequenced and revealed similarity with the plant antimicrobial peptides like non-specific lipid transfer proteins and defensin-like peptide. The F4 and F5 fractions presented strong antimicrobial activity against the fungus Fusarium solani and Fusarium oxysporum, causing toxic effects on these fungi, leading to membrane permeabilization, endogenous reactive oxygen species increase, activation of metacaspase and loss of mitochondrial function., (© 2020 The Author(s).)

Growing production and utilization of titanium dioxide nanoparticles (NpTiO 2 ) invariably lead to their accumulation in oceans, rivers and other water bodies, thus increasing the risk to the welfare of this ecosystem. The progressive launch of these nanoparticles in the environment has been accompanied by concern in understanding the dynamics and the toxic effect of these xenobiotic in different ecosystems, either on their own or in tandem with different contaminants (such as organic compounds and heavy metals), possibly altering their toxicity. Nevertheless, it remains unknown if these combined effects may induce damage in freshwater organisms. Therefore, this study aimed to analyze the consequences caused by NpTiO 2 , after a waterborne exposure of 96 h to a Neotropical fish species Hoplias intermedius , as well as after a co-exposure with lead, whose effects for fish have already been well described in the literature. The characterization of NpTiO 2 stock suspension was carried out in order to provide additional information and revealed a stable colloidal suspension. As a result, NpTiO 2 showed some genotoxic effects which were observed by comet assay in gill, kidney and brain cells. Also, the activity of brain acetylcholinesterase (AChE) has not changed, but the activity of muscle AChE decreased in the group exposed only to PbII. Regarding the hepatic antioxidant system, catalase (CAT) did not show any change in its activity, whereas that of superoxide dismutase (SOD) intensified in the groups submitted only to PbII and NpTiO 2 alone. As for lipid peroxidation, there was a decrease in the group exposed to the NpTiO 2 alone and to the co-exposed group (NpTiO 2 +PbII). As far as metallothionein is concerned, its concentration rose for the co-exposed group (NpTiO 2 +PbII) and for the group exposed to PbII alone. Overall, we may conclude that NpTiO 2 alone caused DNA damage to vital tissues. Also, some impairment related to the antioxidant mechanism was described but it is probably not related to the DNA damage observed, suggesting that the genotoxic effect observed may be due to a different mechanism instead of ROS production.

Background: Plant defensins were discovered at beginning of the 90s'; however, their precise mechanism of action is still unknown. Herein, we studied ApDef 1 -Saccharomyces cerevisiae interaction., Methods: ApDef 1 -S. cerevisiae interaction was studied by determining the MIC, viability and death kinetic assays. Viability assay was repeated with hydroxyurea synchronized-yeast and pretreated with CCCP. Plasma membrane permeabilization, ROS induction, chromatin condensation, and caspase activation analyses were assessed through Sytox green, DAB, DAPI and FITC-VAD-FMK, respectively. Viability assay was done in presence of ascorbic acid and Z-VAD-FMK. Ultrastructural analysis was done by electron microscopy., Results: ApDef 1 caused S. cerevisiae cell death and MIC was 7.8μM. Whole cell population died after 18h of ApDef 1 interaction. After 3h, 98.76% of synchronized cell population died. Pretreatment with CCCP protected yeast from ApDef 1 induced death. ApDef 1 -S. cerevisiae interaction resulted in membrane permeabilization, H 2 O 2 increased production, chromatin condensation and caspase activation. Ascorbic acid prevented yeast cell death and membrane permeabilization. Z-VAD-FMK prevented yeast cell death., Conclusions: ApDef 1 -S. cerevisiae interaction caused cell death through cell cycle dependentprocess which requires preserved membrane potential. After interaction, yeast went through uncontrolled ROS production and accumulation, which led to plasma membrane permeabilization, chromatin condensation and, ultimately, cell death by activation of caspase-dependent apoptosis via., General Significance: We show novel requirements for the interaction between plant defensin and fungi cells, i.e. cell cycle phase and membrane potential, and we indicate that membrane permeabilization is probably caused by ROS and therefore, it would be an indirect event of the ApDef 1 -S. cerevisiae interaction., (Copyright © 2016 Elsevier B.V. All rights reserved.)