Epitope-Specific Antibody Responses to a Plasmodium falciparum Subunit Vaccine Target in a Malaria-Endemic Population.

Circumsporozoite protein (CSP) coats the Plasmodium falciparum sporozoite surface and is a major malaria subunit vaccine target. We measured epitope-specific reactivity to field-derived CSP haplotypes in serum samples from Malian adults and children on a custom peptide microarray. Compared to children, adults showed greater antibody responses and responses to more variants in regions proximal to and within the central repeat region. Children acquired short-lived immunity to an epitope proximal to the central repeat region but not to the central repeat region itself. This approach has the potential to differentiate immunodominant from protective epitope-specific responses when combined with longitudinal infection data.

Role of Order in the Mechanism of Charge Transport across Single-Stranded and Double-Stranded DNA Monolayers in Tunnel Junctions.

Deoxyribonucleic acid (DNA) has been hypothesized to act as a molecular wire due to the presence of an extended π-stack between base pairs, but the factors that are detrimental in the mechanism of charge transport (CT) across tunnel junctions with DNA are still unclear. Here we systematically investigate CT across dense DNA monolayers in large-area biomolecular tunnel junctions to determine when intrachain or interchain CT dominates and under which conditions the mechanism of CT becomes thermally activated. In our junctions, double-stranded DNA (dsDNA) is 30-fold more conductive than single-stranded DNA (ssDNA). The main reason for this large change in conductivity is that dsDNA forms ordered monolayers where intrachain tunneling dominates, resulting in high CT rates. By varying the temperature T and the length of the DNA fragments in the junctions, which determines the tunneling distance, we reveal a complex interplay between T, the length of DNA, and structural order on the mechanism of charge transport. Both the increase in the tunneling distance and the decrease in structural order result in a change in the mechanism of CT from coherent tunneling to incoherent tunneling (hopping). Our results highlight the importance of the interplay between structural order, tunneling distance, and temperature on the CT mechanism across DNA in molecular junctions.

Pyridones in drug discovery: Recent advances.

Pyridones have been utilized as privileged scaffolds in drug discovery. Some of the important roles where this class of heterocycles have found utility in medicinal chemistry include the ability to 1) serve both as a hydrogen bond acceptor and/or a donor; 2) act as a bioisostere for amides, phenyls, pyridines and other nitrogen- or oxygen-containing heterocycles; and 3) impact a target drug molecule's lipophilicity, aqueous solubility and metabolic stability. Detailed discussions of recent advances in their utilization as nonpeptidic mimics and as kinase hinge binding motifs are included. Selected literature examples published from the past twenty years where pyridones have been employed as bioisosteres for phenyls, pyridines, pyridine N-oxides and phenol rings are provided. In addition, this review summarizes the current understanding of possible reactive metabolites related to the pyridone structure.

Duplex Healing of Selectively Thiolated Guanosine Mismatches through a Cd2+ Chemical Stimulus.

The on-column selective conversion of guanosine to thioguanosine (tG) yields modified oligomers that exhibit destabilisation over the fully complementary duplex. Restoration to a stabilised duplex is induced through thio-directed Cd2+ coordination; a route for healing DNA damage. Short oligomers are G-specifically thiolated through a modified on-column protocol without the need for costly thioguanosine phosphoramidites. Addition of Cd2+ ions to a duplex containing a highly disrupted tG central mismatch sequence, 3'-A6 tG4 T6 -5', suggests a (tG)8 Cd2 central coordination regime, resulting in increased base stacking and duplex stability. Equilibrium molecular dynamic calculations support the hypothesis of metal-induced healing of the thiolated duplex. The 2 nm displacement of the central tG mismatched region is dramatically reduced after the addition of a chemical stimuli, Cd2+ ions, returning to a minimized fluctuational state comparable to the unmodified fully complementary oligomer.

Self-Priming Enzymatic Fabrication of Multiply Modified DNA.

The self-priming synthesis of multiply modified DNA by the extension of repeating unit duplex "oligoseeds" provides a source of versatile DNA. Sterically-demanding nucleotides 5-Br-dUTP, 7-deaza-7-I-dATP, 6-S-dGTP, 5-I-dCTP as well as 5-(octadiynyl)-dCTP were incorporated into two extending oligoseeds; [GATC]5 /[GATC]5 and [A4 G]4 /[CT4 ]4 . The products contained modifications on one or both strands of DNA, demonstrating their recognition by the polymerase as both template (reading) and substrate (writing). Nucleobase modifications that lie in the major groove were reliably read and written by the polymerase during the extension reaction, even when bulky or in contiguous sequences. Repeat sequence DNA over 500 bp long, bearing four different modified units was produced by this method. The number, position and type of modification, as well as the overall length of the DNA can be controlled to yield designer DNA that offers sequence-determined sites for further chemical adaptations, targeted small molecule binding studies, or sensing and sequencing applications.

Phenomenological vs. biophysical models of thermal stress in aquatic eggs.

Predicting species responses to climate change is a central challenge in ecology. These predictions are often based on lab-derived phenomenological relationships between temperature and fitness metrics. We tested one of these relationships using the embryonic stage of a Chinook salmon population. We parameterised the model with laboratory data, applied it to predict survival in the field, and found that it significantly underestimated field-derived estimates of thermal mortality. We used a biophysical model based on mass transfer theory to show that the discrepancy was due to the differences in water flow velocities between the lab and the field. This mechanistic approach provides testable predictions for how the thermal tolerance of embryos depends on egg size and flow velocity of the surrounding water. We found support for these predictions across more than 180 fish species, suggesting that flow and temperature mediated oxygen limitation is a general mechanism underlying the thermal tolerance of embryos.

Cytoplasmic actin is an extracellular insect immune factor which is secreted upon immune challenge and mediates phagocytosis and direct killing of bacteria, and is a Plasmodium Antagonist.

Actin is a highly versatile, abundant, and conserved protein, with functions in a variety of intracellular processes. Here, we describe a novel role for insect cytoplasmic actin as an extracellular pathogen recognition factor that mediates antibacterial defense. Insect actins are secreted from cells upon immune challenge through an exosome-independent pathway. Anopheles gambiae actin interacts with the extracellular MD2-like immune factor AgMDL1, and binds to the surfaces of bacteria, mediating their phagocytosis and direct killing. Globular and filamentous actins display distinct functions as extracellular immune factors, and mosquito actin is a Plasmodium infection antagonist.

Nanoscale silicon substrate patterns from self-assembly of cylinder forming poly(styrene)-block-poly(dimethylsiloxane) block copolymer on silane functionalized surfaces.

Poly(styrene)-block-poly(dimethylsiloxane) (PS-b-PDMS) is an excellent block copolymer (BCP) system for self-assembly and inorganic template fabrication because of its high Flory-Huggins parameter (χ âˆ¼ 0.26) at room temperature in comparison to other BCPs, and high selective etch contrast between PS and PDMS block for nanopatterning. In this work, self-assembly in PS-b-PDMS BCP is achieved by combining hydroxyl-terminated poly(dimethylsiloxane) (PDMS-OH) brush surfaces with solvent vapor annealing. As an alternative to standard brush chemistry, we report a simple method based on the use of surfaces functionalized with silane-based self-assembled monolayers (SAMs). A solution-based approach to SAM formation was adopted in this investigation. The influence of the SAM-modified surfaces upon BCP films was compared with polymer brush-based surfaces. The cylinder forming PS-b-PDMS BCP and PDMS-OH polymer brush were synthesized by sequential living anionic polymerization. It was observed that silane SAMs provided the appropriate surface chemistry which, when combined with solvent annealing, led to microphase segregation in the BCP. It was also demonstrated that orientation of the PDMS cylinders may be controlled by judicious choice of the appropriate silane. The PDMS patterns were successfully used as an on-chip etch mask to transfer the BCP pattern to underlying silicon substrate with sub-25 nm silicon nanoscale features. This alternative SAM/BCP approach to nanopattern formation shows promising results, pertinent in the field of nanotechnology, and with much potential for application, such as in the fabrication of nanoimprint lithography stamps, nanofluidic devices or in narrow and multilevel interconnected lines.

Click modification of diazido acridine intercalators: a versatile route towards decorated DNA nanostructures.

Diazido derivatives of 3,6-diamino acridine (proflavine) intercalate into DNA and undergo functionalization through click chemistry to form 1D nanostructures with redox active, conductive nanowire, and fluorescent properties. This two-step approach, intercalation followed by click modification allows for the controlled decoration of DNA nanostructures.

Enzymatic Method for the Synthesis of Long DNA Sequences with Multiple Repeat Units.

A polymerase chain reaction (PCR) derived method for preparing long DNA, consisting of multiple repeat units of one to ten base pairs, is described. Two seeding oligodeoxynucleotides, so-called oligoseeds, which encode the repeat unit and produce a duplex with 5'-overhangs, are extended using a thermostable archaeal DNA polymerase. Multiple rounds of heat-cool extension cycles, akin to PCR, rapidly elongate the oligoseed. Twenty cycles produced long DNA with uniformly repeating sequences to over 20 kilobases (kb) in length. The polynucleotides prepared include [A]n /[T]n , [AG]n /[TC]n , [A2 G]n /[T2 C]n , [A3 G]n /[T3 C]n , [A4 G]n /[T4 C]n , [A9 G]n /[T9 C]n , [GATC]n /[CTAG]n , and [ACTGATCAGC]n /[TGACTAGTCG]n , indicating that the method is extremely flexible with regard to the repeat length and base sequence of the initial oligoseeds. DNA of this length (20 kb≈7 µm) with strictly defined base reiterations should find use in nanomaterial applications.

A simple, robust, broadly applicable insertion mutagenesis method to create random fluorescent protein: target protein fusions.

A simple, broadly applicable method was developed using an in vitro transposition reaction followed by transformation into Escherichia coli and screening plates for fluorescent colonies. The transposition reaction catalyzes the random insertion of a fluorescent protein open reading frame into a target gene on a plasmid. The transposition reaction is employed directly in an E. coli transformation with no further procedures. Plating at high colony density yields fluorescent colonies. Plasmids purified from fluorescent colonies contain random, in-frame fusion proteins into the target gene. The plate screen also results in expressed, stable proteins. A large library of chimeric proteins was produced, which was useful for downstream research. The effect of using different fluorescent proteins was investigated as well as the dependence of the linker sequence between the target and fluorescent protein open reading frames. The utility and simplicity of the method were demonstrated by the fact that it has been employed in an undergraduate biology laboratory class without failure over dozens of class sections. This suggests that the method will be useful in high-impact research at small liberal arts colleges with limited resources. However, in-frame fusion proteins were obtained from 8 different targets suggesting that the method is broadly applicable in any research setting.

Coordination chemistry of 6-thioguanine derivatives with cobalt: toward formation of electrical conductive one-dimensional coordination polymers.

In this work we have synthetized and characterized by X-ray diffraction five cobalt complexes with 6-thioguanine (6-ThioGH), 6-thioguanosine (6-ThioGuoH), or 2'-deoxy-6-thioguanosine (2'-d-6-ThioGuoH) ligands. In all cases, these ligands coordinate to cobalt via N7 and S6 forming a chelate ring. However, independently of reagents ratio, 6-ThioGH provided monodimensional cobalt(II) coordination polymers, in which the 6-ThioG(-) acts as bridging ligand. However, for 2'-d-6-ThioGuoH and 6-ThioGuoH, the structure directing effect of the sugar residue gives rise to mononuclear cobalt complexes which form extensive H-bond interactions to generate 3D supramolecular networks. Furthermore, with 2'-d-6-ThioGuoH the cobalt ion remains in the divalent state, whereas with 6-ThioGuoH oxidation occurs and Co(III) is found. The electrical and magnetic properties of the coordination polymers isolated have been studied and the results discussed with the aid of DFT calculations, in the context of molecular wires.

Floodplain rehabilitation as a hedge against hydroclimatic uncertainty in a migration corridor of threatened steelhead.

A strategy for recovering endangered species during climate change is to restore ecosystem processes that moderate effects of climate shifts. In mid-latitudes, storm patterns may shift their intensity, duration, and frequency. These shifts threaten flooding in human communities and reduce migration windows (conditions suitable for migration after a storm) for fish. Rehabilitation of historic floodplains can in principle reduce these threats via transient storage of storm water, but no one has quantified the benefit of floodplain rehabilitation for migrating fish, a widespread biota with conservation and economic value. We used simple models to quantify migration opportunity for a threatened migratory fish, steelhead (Oncorhynchus mykiss), in an episodic rain-fed river system, the Pajaro River in central California. We combined flow models, bioenergetic models, and existing climate projections to estimate the sensitivity of migration windows to altered storm patterns under alternate scenarios of floodplain rehabilitation. Generally, migration opportunities were insensitive to warming, weakly sensitive to duration or intensity of storms, and proportionately sensitive to frequency of storms. The rehabilitation strategy expanded migration windows by 16-28% regardless of climate outcomes. Warmer conditions raised the energy cost of migrating, but not enough to matter biologically. Novel findings were that fewer storms appeared to pose a bigger threat to migrating steelhead than warmer or smaller storms and that floodplain rehabilitation lessened the risk from fewer or smaller storms across all plausible hydroclimatic outcomes. It follows that statistical downscaling methods may mischaracterize risk, depending on how they resolve overall precipitation shifts into changes of storm frequency as opposed to storm size. Moreover, anticipating effects of climate shifts that are irreducibly uncertain (here, rainfall) may be more important than anticipating effects of relatively predictable changes such as warming. This highlights a need to credibly identify strategies of ecosystem rehabilitation that are robust to uncertainty. Rehabilitación de Planicies Inundables como Cerco contra la Incertidumbre Hidroclimática en un Corredor Migratorio de Oncorhynchus mykiss, Especie Amenazada.

Impedimetric Polyaniline-Based Aptasensor for Aflatoxin B1 Determination in Agricultural Products.

An impedimetric aptasensor based on a polyaniline (PAni) support matrix is developed through the surface modification of a screen-printed carbon electrode (SPE) for aflatoxin B1 (AFB1) detection in foodstuffs and feedstuffs for food safety. The PAni is synthesized with the chemical oxidation method and characterized with potentiostat/galvanostat, FTIR, and UV-vis spectroscopy techniques. The stepwise fabrication procedure of the PAni-based aptasensor is characterized by means of cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) methods. The impedimetric aptasensor is optimized using the EIS technique, and its feasibility of detecting AFB1 in real sample matrices is evaluated via a recovery study in spiked foodstuffs and feedstuffs, such as pistachio nuts, cinnamons, cloves, corn, and soybeans, with a good recovery percentage, ranging from 87.9% to 94.7%. The charge transfer resistance (RCT) at the aptasensor interface increases linearly with the AFB1 concentration in the range of 3 × 10-2 nM to 8 × 10-2 nM, with a regression coefficient (R2) value of 0.9991 and detection limit of 0.01 nM. The proposed aptasensor is highly selective towards AFB1 and partially selective to AFB2 and ochratoxin A (OTA) due to their similar structures that differ only at the carbon-carbon double bond located at C8 and C9 and the large molecule size of OTA.

Natural immunity to malaria preferentially targets the endothelial protein C receptor-binding regions of PfEMP1s.

Antibody responses to variant surface antigens (VSAs) produced by the malaria parasite Plasmodium falciparum may contribute to age-related natural immunity to severe malaria. One VSA family, P. falciparum erythrocyte membrane protein-1 (PfEMP1), includes a subset of proteins that binds endothelial protein C receptor (EPCR) in human hosts and potentially disrupts the regulation of inflammatory responses, which may lead to the development of severe malaria. We probed peptide microarrays containing segments spanning five PfEMP1 EPCR-binding domain variants with sera from 10 Malian adults and 10 children to determine the differences between adult and pediatric immune responses. We defined serorecognized peptides and amino acid residues as those that elicited a significantly higher antibody response than malaria-naïve controls. We aimed to identify regions consistently serorecognized among adults but not among children across PfEMP1 variants, potentially indicating regions that drive the development of immunity to severe malaria. Adult sera consistently demonstrated broader and more intense serologic responses to constitutive PfEMP1 peptides than pediatric sera, including peptides in EPCR-binding domains. Both adults and children serorecognized a significantly higher proportion of EPCR-binding peptides than peptides that do not directly participate in receptor binding, indicating a preferential development of serologic responses at functional residues. Over the course of a single malaria transmission season, pediatric serological responses increased between the start and the peak of the season, but waned as the transmission season ended. IMPORTANCE Severe malaria and death related to malaria disproportionately affect sub-Saharan children under 5 years of age, commonly manifesting as cerebral malaria and/or severe malarial anemia. In contrast, adults in malaria-endemic regions tend to experience asymptomatic or mild disease. Our findings indicate that natural immunity to malaria targets specific regions within the EPCR-binding domain, particularly peptides containing EPCR-binding residues. Epitopes containing these residues may be promising targets for vaccines or therapeutics directed against severe malaria. Our approach provides insight into the development of natural immunity to a binding target linked to severe malaria by characterizing an "adult-like" response as recognizing a proportion of epitopes within the PfEMP1 protein, particularly regions that mediate EPCR binding. This "adult-like" response likely requires multiple years of malaria exposure, as increases in pediatric serologic response over a single malaria transmission season do not appear significant.

Online Courses Provide Robust Learning Gains and Improve Learner Confidence in the Foundational Biomedical Sciences.

The early stages of medical school involve education in a number of foundational biomedical sciences including genetics, immunology, and physiology. However, students entering medical school may have widely varying levels of background in these areas due to differences in the availability and quality of prior education on these topics. Even students who have recently taken formal courses in these subjects may not feel confident in their level of preparation, leading to anxiety for early-stage medical students. These differences can make it difficult for instructors to create meaningful learning experiences that are appropriate for all students. Additionally, actual or perceived differences in preparation may lead fewer students from diverse backgrounds to apply to medical school. Therefore, creating an efficient and scalable way to increase students' knowledge and confidence in these topics addresses an important need for many medical schools. We recorded pre- and post-course quiz scores for 9790 individuals who completed HMX online courses, developed in accordance with evidence-based learning practices and covering the fundamentals of biochemistry, genetics, immunology, pharmacology, and physiology. Each question was accompanied by a Likert scale question to assess the learner's confidence in their answer. Learners' median post-course quiz performance and self-assessed confidence significantly increased relative to pre-course quiz performance for each course. Improvements were consistent across US-based medical schools, non-US medical schools, and course runs open to the public. This indicates that online courses created using evidence-based learning practices can lead to significant increases in knowledge and confidence for many learners, helping prepare them for further medical education. Supplementary Information: The online version contains supplementary material available at 10.1007/s40670-022-01660-4.

Increased posterior tibial slope is an independent risk factor of anterior cruciate ligament reconstruction graft rupture irrespective of graft choice.

INTRODUCTION: Anterior cruciate ligament (ACL) reconstruction failure remains a commonly seen complication despite advances in technique and graft options. Recently, several studies have shown that the inclination of the tibial plateau in the sagittal plane affects the stability of the knee joint. The purpose of this study was to determine if an increased posterior slope of the tibia is associated with failure of ACL reconstruction irrespective of the graft used. METHODS: From June 2002 to August 2003, a total of 100 patients with a symptomatic ACL-deficient knee were randomised to receive either a hamstring autograft or posterior tibialis allograft. All allografts were from a single tissue bank, aseptically processed, and fresh-frozen without terminal irradiation. ACL graft failures requiring reoperation with a minimum of 10-year follow-up were identified via telephone survey. Lateral radiographs of the knee of all patients were reviewed, and the slope of the tibia was measured using a standardised technique. Two fellowship-trained orthopaedic sports medicine specialists, one board-certified general orthopaedic surgeon, and two fellowship-trained musculoskeletal radiologists measured the tibial slope in all patients. RESULTS: At a minimum of 10-year follow-up, there were four (8.3%) autograft and 13 (26.5%) allograft failures that required revision reconstruction. The overall average tibial slope of the nonfailure cohort was 9.4°. The overall average tibial slope of the failure cohort was 11.9° (P â€‹= â€‹0.0002). The average slope of the allograft failures was 11.5°compared with an average slope of 9.6° in the nonfailures (P â€‹= â€‹0.01). The average slope of the autograft failures was 13.1° compared with 9.3° in the nonfailures (P â€‹= â€‹0.011). The mean difference in tibial slope measurements was 0.665 (95% confidence interval: 0.569-0.750). The interrater reliability, as measured by the intraclass correlation coefficient, for tibial slope was 0.898 (95% confidence interval: 0.859-0.928). The Cronbach α was 0.904. CONCLUSION: In a prospective, randomised trial of ACL reconstructions using either autograft or allograft, failures were associated with a significantly increased slope of the tibia compared with the nonfailures at 10-year follow-up.

Unlocking the thermoelectric potential of the Ca14AlSb11 structure type.

Yb14MnSb11 and Yb14MgSb11 are among the best p-type high-temperature (>1200 K) thermoelectric materials, yet other compounds of this Ca14AlSb11 structure type have not matched their stability and efficiency. First-principles computations show that the features in the electronic structures that have been identified to lead to high thermoelectric performances are present in Yb14ZnSb11, which has been presumed to be a poor thermoelectric material. We show that the previously reported low power factor of Yb14ZnSb11 is not intrinsic and is due to the presence of a Yb9Zn4+xSb9 impurity uniquely present in the Zn system. Phase-pure Yb14ZnSb11 synthesized through a route avoiding the impurity formation reveals its exceptional high-temperature thermoelectric properties, reaching a peak zT of 1.2 at 1175 K. Beyond Yb14ZnSb11, the favorable band structure features for thermoelectric performance are universal among the Ca14AlSb11 structure type, opening the possibility for high-performance thermoelectric materials.

Temperature-Dependent Coherent Tunneling across Graphene-Ferritin Biomolecular Junctions.

Understanding the mechanisms of charge transport (CT) across biomolecules in solid-state devices is imperative to realize biomolecular electronic devices in a predictive manner. Although it is well-accepted that biomolecule-electrode interactions play an essential role, it is often overlooked. This paper reveals the prominent role of graphene interfaces with Fe-storing proteins in the net CT across their tunnel junctions. Here, ferritin (AfFtn-AA) is adsorbed on the graphene by noncovalent amine-graphene interactions confirmed with Raman spectroscopy. In contrast to junctions with metal electrodes, graphene has a vanishing density of states toward its intrinsic Fermi level ("Dirac point"), which increases away from the Fermi level. Therefore, the amount of charge carriers is highly sensitive to temperature and electrostatic charging (induced doping), as deduced from a detailed analysis of CT as a function of temperature and iron loading. Remarkably, the temperature dependence can be fully explained within the coherent tunneling regime due to excitation of hot carriers. Graphene is not only demonstrated as an alternative platform to study CT across biomolecular tunnel junctions, but it also opens rich possibilities in employing interface electrostatics in tuning CT behavior.

Pyrrolyl-, 2-(2-thienyl)pyrrolyl- and 2,5-bis(2-thienyl)pyrrolyl-nucleosides: synthesis, molecular and electronic structure, and redox behaviour of C5-thymidine derivatives.

A series of modified nucleosides based on thymidine have been prepared by Pd-catalysed cross-coupling between N-alkyl-alkynyl functionalised pyrrolyl- (py), 2-(2-thienyl)pyrrolyl- (tp) or 2,5-bis(2-thienyl)pyrrolyl (tpt) groups with 5-iodo-2'-deoxyuridine. The length of the alkyl chain linking the nucleoside and pyrrolyl-containing unit, N(CH(2))(n)C[triple bond, length as m-dash]C-nucleoside (where n = 1-3) was also varied. The compounds have been characterised by (1)H NMR, ES-MS, UV-vis, cyclic voltammetry (CV) and, in some cases, single-crystal X-ray diffraction. Cyclic voltammetry studies demonstrated that all the py-, tp- and tpt-alkynyl derivatives 1-7 can be electrochemically polymerised to form conductive materials. It was found that increasing the N-alkyl chain length in these cases resulted in only minor changes in the oxidation potential. The same behaviour was observed for the tp- and tpt-modified nucleosides 9-12; however, the py-derivative, 8, produced a poorly conducting material. DFT calculations on the one-electron oxidised cation of the modified nucleosides bearing tp or tpt showed that spin density is located on the pyrrolyl and thienyl units in all cases and that the coplanarity of adjacent rings increases upon oxidation. In contrast, in the corresponding pyrrolyl cases the spin density is distributed over the whole molecule, suggesting that polymerisation does not occur solely at the pyrrolyl-Cα position and the conjugation is interrupted.