On the genetic basis of tail-loss evolution in humans and apes.

The loss of the tail is among the most notable anatomical changes to have occurred along the evolutionary lineage leading to humans and to the 'anthropomorphous apes'1-3, with a proposed role in contributing to human bipedalism4-6. Yet, the genetic mechanism that facilitated tail-loss evolution in hominoids remains unknown. Here we present evidence that an individual insertion of an Alu element in the genome of the hominoid ancestor may have contributed to tail-loss evolution. We demonstrate that this Alu element-inserted into an intron of the TBXT gene7-9-pairs with a neighbouring ancestral Alu element encoded in the reverse genomic orientation and leads to a hominoid-specific alternative splicing event. To study the effect of this splicing event, we generated multiple mouse models that express both full-length and exon-skipped isoforms of Tbxt, mimicking the expression pattern of its hominoid orthologue TBXT. Mice expressing both Tbxt isoforms exhibit a complete absence of the tail or a shortened tail depending on the relative abundance of Tbxt isoforms expressed at the embryonic tail bud. These results support the notion that the exon-skipped transcript is sufficient to induce a tail-loss phenotype. Moreover, mice expressing the exon-skipped Tbxt isoform develop neural tube defects, a condition that affects approximately 1 in 1,000 neonates in humans10. Thus, tail-loss evolution may have been associated with an adaptive cost of the potential for neural tube defects, which continue to affect human health today.

SARS-CoV-2 monoclonal antibody treatment followed by vaccination shifts human memory B cell epitope recognition suggesting antibody feedback.

Therapeutic anti-SARS-CoV-2 monoclonal antibodies (mAbs) have been extensively studied in humans, but the impact on immune memory of mAb treatment during an ongoing immune response has remained unclear. Here, we evaluated the effect of infusion of the anti-SARS-CoV-2 spike receptor binding domain (RBD) mAb bamlanivimab on memory B cells (MBCs) in SARS-CoV-2-infected individuals. Bamlanivimab treatment skewed the repertoire of memory B cells targeting Spike towards non-RBD epitopes. Furthermore, the relative affinity of RBD memory B cells was weaker in mAb-treated individuals compared to placebo-treated individuals over time. Subsequently, after mRNA COVID-19 vaccination, memory B cell differences persisted and mapped to a specific defect in recognition of the class II RBD site, the same RBD epitope recognized by bamlanivimab. These findings indicate a substantial role of antibody feedback in regulating human memory B cell responses, both to infection and vaccination. These data indicate that mAb administration can promote alterations in the epitopes recognized by the B cell repertoire, and the single administration of mAb can continue to determine the fate of B cells in response to additional antigen exposures months later.

Cervical cancer screening in Brazilian Amazon Indigenous women: Towards the intensification of public policies for prevention.

BACKGROUND: Indigenous women are vulnerable to cervical cancer. Screening is a strategy to reduce the burden of the disease. OBJECTIVE: To evaluate the prevalence profile of cervical cancer screening cytological results in Brazilian indigenous women by age and frequency of tests compared to non-indigenous women. METHODS: A cross-sectional study evaluating the prevalences of screening test results in indigenous women assisted in the Brazilian Amazon from 2007 to 2019 (3,231 tests), compared to non-indigenous women (698,415 tests). The main outcome was the cytological result. Other variables were frequency, age groups, and population. The frequency was categorized as "1st test", the first test performed by the women in their lifetime, or "screening test," tests from women who had previously participated in screening. Analyzes were based on prevalences by age group and population. We used Prevalence Ratios (PR) and 95% Confidence Intervals for risks and linear regression for trends. RESULTS: Data from the 1st test showed a higher prevalence of Low-grade Squamous Intraepithelial Lesion (LSIL) in indigenous women. Peaks were observed in indigenous under 25, 35 to 39, 45 to 49, and 60 to 64. The prevalence of High-grade Squamous Intraepithelial Lesion or more severe (HSIL+) was low in both groups in women younger than 25. The indigenous HSIL+ prevalence curve showed a rapid increase, reaching peaks in women from 25 to 34 years, following a slight decrease and a plateau. In screening tests, HSIL+ was more prevalent in indigenous from 25 to 39 (PR 4.0,2.3;6.8) and 40 to 64 (PR 3.8,1.6;9.0). In indigenous, the PR of HSIL+ results in screening tests over 1st tests showed no screening effect in all age groups. In non-indigenous, there was a significant effect toward protection in the age groups over 25. CONCLUSION: This screening study of indigenous women from diverse ethnicities showed a higher prevalence of cytological LSIL and HSIl+ than in non-indigenous women. The protective screening effect in reducing HSIL+ prevalence was not observed in indigenous.

Implicações no cuidado de um familiar doente: mulheres negras cuidadoras

Introdução: As desvantagens ligadas ao gênero feminino são visualizadas em diversos aspectos da vida, mediante ao patriarcado. É possível compreender que, no caso de mulheres negras, estas se encontram desfavorecidas frente as estruturas sociais, atravessadas por condicionantes de classe, raça e gênero. Além disso, é visto que, em diversos casos, as cuidadoras mulheres, sobretudo, mulheres negras, não escolhem de forma voluntária este papel dentro da conjuntura familiar. Objetivo: Identificar as implicações do cuidado ao familiar adoecido realizado por mulheres negras cuidadoras considerando suas condições socioculturais. Metodologia: Este trabalho é uma revisão integrativa, cuja busca no banco de dados foi realizada em dezembro de 2020, onde 3 resultados foram encontrados na base de dados Web of Science, 48 na base de dados PubMed e 29 na base de dados LILACS. Depois de ler os resumos dos 80 artigos e aplicar os critérios de exclusão, 7 artigos foram selecionados para leitura completa. Finalmente, 4 artigos foram incluídos para análise. Resultados: Apenas um estudo abordou exclusivamente mulheres, e a maioria eram afro-americanos. Dados indicam que, dentro de uma comunidade quilombola, o cuidado com a saúde é repassado do mais velho para o mais novo, como símbolo de respeito aos saberes ancestrais. Além disso, ficou evidente que cuidadores afro-americanos de pessoas com demência precisam de informações de qualidade sobre cuidados e autocuidado, necessitando de recursos em sua comunidade. Conclusão: É possível notar as desigualdades de acordo com as construções históricas, políticas e culturais causadas, para diferenciar homens e mulheres no cuidado familiar. Havendo uma necessidade de debates que tenham como enfoque a população negra, principalmente as cuidadoras, visando estratégias para reduzir sua sobrecarga para cuidar de familiares em adoecimento.

Introducción: Las desventajas vinculadas al género femenino se visualizan en diversos aspectos de la vida, a través del patriarcado. Es posible entender que, en el caso de las mujeres negras, ellas están en desventaja frente a las estructuras sociales, atravesadas por condiciones de clase, raza y género. Además, se observa que, en varios casos, las cuidadoras, especialmente las mujeres negras, no eligen voluntariamente este rol dentro del contexto familiar. Objetivo: Identificar las implicaciones del cuidado a un familiar enfermo realizado por cuidadoras negras considerando sus condiciones socioculturales. Metodología: Este trabajo es una revisión integradora, cuya búsqueda en la base de datos tuvo lugar en diciembre de 2020, donde se encontraron 3 resultados en la base de datos Web of Science, 48 en la base de datos PubMed y 29 en la base de datos LILACS. Después de leer los resúmenes de los 80 artículos. y aplicar los criterios de exclusión, se seleccionaron 7 artículos para su lectura íntegra. Finalmente, 4 artículos fueron incluidos para el análisis. Resultados: Solo un estudio se refería exclusivamente a mujeres, y la mayoría eran afroamericanas. Los datos indican que, en el seno de una comunidad quilombola, los cuidados de salud se transmiten de los más ancianos a los más jóvenes, como símbolo de respeto a los conocimientos ancestrales. Además, se puso de manifiesto que los cuidadores afroamericanos de personas con demencia necesitan información de calidad sobre cuidados y autocuidados y precisan recursos en su comunidad. Conclusión: Es posible percibir las desigualdades según las construcciones históricas, políticas y culturales provocadas, para diferenciar hombres y mujeres en el cuidado de la familia. Hay una necesidad de debates que se centren en la población negra, especialmente en las cuidadoras, con miras a elaborar estrategias para reducir su carga para cuidar a los familiares enfermos.

Introduction: The disadvantages linked to the female gender are viewed in different aspects of life, through patriarchy. It is possible to understand that, in the case of black women, they are disadvantaged in the face of social structures, crossed by class, race and gender conditioning factors. In addition, it is seen that, in several cases, female caregivers, especially black women, do not voluntarily choose this role within the family environment. Objective: To identify the implications of care for a sick family member performed by black women caregivers considering their sociocultural conditions. Methodology: This work is an integrative review, whose search in the database was carried out in December 2020, where 3 results were found in the Web of Science database, 48 in the PubMed database and 29 in the LILACS database. After reading the abstracts of the 80 articles and applying the exclusion criteria, 7 articles were selected for full reading. Finally, 4 articles were included for analysis. Results: Only one study exclusively addressed women, and most were African Americans. Data indicate that, within a quilombola community, health care is passed on from the oldest to the youngest, as a symbol of respect for ancestral knowledge. In addition, it became evident that African-American caregivers of people with dementia need quality information about care and self-care, requiring resources in their community. Conclusion: It is possible to notice the inequalities according to the historical, political and cultural constructions caused, in order to differentiate men and women in family care. There is a need for debates that focus on the black population, especially caregivers, aiming at strategies to reduce their burden to care for sick family members.

Debugging and consolidating multiple synthetic chromosomes reveals combinatorial genetic interactions.

The Sc2.0 project is building a eukaryotic synthetic genome from scratch. A major milestone has been achieved with all individual Sc2.0 chromosomes assembled. Here, we describe the consolidation of multiple synthetic chromosomes using advanced endoreduplication intercrossing with tRNA expression cassettes to generate a strain with 6.5 synthetic chromosomes. The 3D chromosome organization and transcript isoform profiles were evaluated using Hi-C and long-read direct RNA sequencing. We developed CRISPR Directed Biallelic URA3-assisted Genome Scan, or "CRISPR D-BUGS," to map phenotypic variants caused by specific designer modifications, known as "bugs." We first fine-mapped a bug in synthetic chromosome II (synII) and then discovered a combinatorial interaction associated with synIII and synX, revealing an unexpected genetic interaction that links transcriptional regulation, inositol metabolism, and tRNASerCGA abundance. Finally, to expedite consolidation, we employed chromosome substitution to incorporate the largest chromosome (synIV), thereby consolidating >50% of the Sc2.0 genome in one strain.

Synthetic chromosome fusion: Effects on mitotic and meiotic genome structure and function.

We designed and synthesized synI, which is ∼21.6% shorter than native chrI, the smallest chromosome in Saccharomyces cerevisiae. SynI was designed for attachment to another synthetic chromosome due to concerns surrounding potential instability and karyotype imbalance and is now attached to synIII, yielding the first synthetic yeast fusion chromosome. Additional fusion chromosomes were constructed to study nuclear function. ChrIII-I and chrIX-III-I fusion chromosomes have twisted structures, which depend on silencing protein Sir3. As a smaller chromosome, chrI also faces special challenges in assuring meiotic crossovers required for efficient homolog disjunction. Centromere deletions into fusion chromosomes revealed opposing effects of core centromeres and pericentromeres in modulating deposition of the crossover-promoting protein Red1. These effects extend over 100 kb and promote disproportionate Red1 enrichment, and thus crossover potential, on small chromosomes like chrI. These findings reveal the power of synthetic genomics to uncover new biology and deconvolute complex biological systems.

Antibody gene features associated with binding and functional activity in vaccine-derived human mAbs targeting malaria parasites.

Adjuvants have been essential to malaria vaccine development, but their impact on the vaccine-induced antibody repertoire is poorly understood. Here, we used cDNA sequences from antigen-specific single memory B cells to express 132 recombinant human anti-Pfs230 monoclonal antibodies (mAbs). Alhydrogel®-induced mAbs demonstrated higher binding to Pfs230D1, although functional activity was similar between adjuvants. All Alhydrogel® mAbs using IGHV1-69 gene bound to recombinant Pfs230D1, but none blocked parasite transmission to mosquitoes; similarly, no AS01 mAb using IGHV1-69 blocked transmission. Functional mAbs from both Alhydrogel® and AS01 vaccines used IGHV3-21 and IGHV3-30 genes. Antibodies with the longest CDR3 sequences were associated with binding but not functional activity. This study assesses adjuvant effects on antibody clonotype diversity during malaria vaccination.

Identification of an ex vivo inhibitor of the schizophrenia biomarker Ndel1 by high throughput screening.

Ndel1 oligopeptidase activity shows promise as a potential biomarker for diagnosing schizophrenia (SCZ) and monitoring early-stage pharmacotherapy. Ndel1 plays a pivotal role in critical aspects of brain development, such as neurite outgrowth, neuronal migration, and embryonic brain formation, making it particularly relevant to neurodevelopmental disorders like SCZ. Currently, the most specific inhibitor for Ndel1 is the polyclonal anti-Ndel1 antibody (NOAb), known for its high specificity and efficient anti-catalytic activity. NOAb has been vital in measuring Ndel1 activity in humans and animal models, enabling the prediction of pharmacological responses to antipsychotics in studies with patients and animals. To advance our understanding of in vivo Ndel1 function and develop drugs for mental disorders, identifying small chemical compounds capable of specifically inhibiting Ndel1 oligopeptidase is crucial, including within living cells. Due to challenges in obtaining Ndel1's three-dimensional structure and its promiscuous substrate recognition, we conducted a high-throughput screening (HTS) of 2,400 small molecules. Nine compounds with IC50-values ranging from 7 to 56 µM were identified as potent Ndel1 inhibitors. Notably, one compound showed similar efficacy to NOAb and inhibited Ndel1 within living cells, although its in vivo use may pose toxicity concerns. Despite this, all identified compounds hold promise as candidates for further refinement through rational drug design, aiming to enhance their inhibitory efficacy, specificity, stability, and biodistribution. Our ultimate goal is to develop druggable Ndel1 inhibitors that can improve the treatment and support the diagnosis of psychiatric disorders like SCZ.

A human antibody epitope map of the malaria vaccine antigen Pfs25.

Pfs25 is a leading antigen for a malaria transmission-blocking vaccine and shows moderate transmission-blocking activity and induction of rapidly decreasing antibody titers in clinical trials. A comprehensive definition of all transmission-reducing epitopes of Pfs25 will inform structure-guided design to enhance Pfs25-based vaccines, leading to potent transmission-blocking activity. Here, we compiled a detailed human antibody epitope map comprising epitope binning data and structures of multiple human monoclonal antibodies, including three new crystal structures of Pfs25 in complex with transmission-reducing antibodies from Malian volunteers immunized with Pfs25 conjugated to EPA and adjuvanted with AS01. These structures revealed additional epitopes in Pfs25 capable of reducing transmission and expanded this characterization to malaria-exposed humans. This work informs immunogen design to focus the antibody response to transmission-reducing epitopes of Pfs25, enabling development of more potent transmission-blocking vaccines for malaria.

A scoping review on hyposalivation associated with systemic conditions: the role of physical stimulation in the treatment approaches.

BACKGROUND: Several systemic conditions can result in distinct degrees of salivary gland damage and consequent hypofunction. The development of successful management schemes is highly challenging due to the complexity of saliva. This study aimed to systematically map the literature on the physical stimulation of salivary glands for hyposalivation management and the response of individuals according to different systemic conditions causing salivary impairment. METHODS: A systematic search in the literature was performed. Two reviewers independently selected clinical trials, randomized or not, that used physical stimulation to treat hyposalivation caused by systemic conditions. Studies evaluating healthy subjects without hyposalivation were included as controls. Single-arm clinical studies or case series were also included for protocol mapping (PRISMA extension for scoping reviews). RESULTS: Out of 24 included studies, 10 evaluated healthy subjects, from which 9 tested transcutaneous electrical nerve stimulation (TENS) and 1 tested acupuncture and electroacupuncture. Fourteen studies evaluated individuals with hyposalivation: 6 applied TENS, 6 applied low-level laser therapy (LLLT), and 2 applied acupuncture, carried out in post-chemotherapy, medication use, postmenopausal women, hemodialysis patients, smokers, diabetics, Sjögren's syndrome (SS). All showed increased salivation after treatment, except for two LLLT studies in individuals with SS. CONCLUSIONS: Among the different patient groups, individuals with Sjögren's syndrome (SS) exhibited the poorest responses, while those with medication-induced hyposalivation demonstrated the most favorable treatment outcomes, independently of the management strategy for saliva stimulation. It means that physical stimulation of salivary glands holds promise as an alternative for managing hyposalivation in cases of reversible gland damage. However, to make informed decisions in current practice, it is necessary to conduct new well-designed randomized clinical trials with appropriate methodologies.

Mpox vaccine and infection-driven human immune signatures: an immunological analysis of an observational study.

BACKGROUND: Monkeypox virus has recently infected more than 88 000 people, raising concerns about our preparedness against this emerging viral pathogen. Licensed and approved for mpox, the JYNNEOS vaccine has fewer side-effects than previous smallpox vaccines and has shown immunogenicity against monkeypox in animal models. This study aims to elucidate human immune responses to JYNNEOS vaccination compared with mpox-induced immunity. METHODS: Peripheral blood mononuclear cells and sera were obtained from ten individuals vaccinated with one or two doses of JYNNEOS and six individuals diagnosed with monkeypox virus infection. Samples were obtained from seven individuals before vaccination to serve as a baseline. We examined the polyclonal serum (ELISA) and single B-cell (heavy chain gene and transcriptome data) antibody repertoires and T-cell responses (activation-induced marker and intracellular cytokine staining assays) induced by the JYNNEOS vaccine versus monkeypox virus infection. FINDINGS: All participants were men between the ages of 21 and 60 years, except for one woman in the group of mpox-convalescent individuals, and none had previous orthopoxvirus exposure. All mpox cases were mild. Vaccinee samples were collected 6-33 days after the first dose and 5-40 days after the second dose. Mpox-convalescent samples were collected 20-102 days after infection. In vaccine recipients, gene-level plasmablast and antibody responses were negligible and sera displayed moderate binding to recombinant orthopoxviral proteins (A29L, A35R, E8L, A30L, A27L, A33R, B18R, and L1R) and native proteins from the 2022 monkeypox outbreak strain. By contrast, recent monkeypox virus infection (within 20-102 days) induced robust serum antibody responses to monkeypox virus proteins and to native monkeypox virus proteins from a viral isolate obtained during the 2022 outbreak. JYNNEOS vaccine recipients presented robust orthopoxviral CD4+ and CD8+ T-cell responses. INTERPRETATION: Infection with monkeypox virus resulted in robust B-cell and T-cell responses, whereas immunisation with JYNNEOS elicited more robust T-cell responses. These data can help to inform vaccine design and policies for preventing mpox in humans. FUNDING: National Cancer Institute (National Institutes of Health), National Institute of Allergy and Infectious Diseases (National Institutes of Health), and Icahn School of Medicine.

Clinical management of salivary gland hypofunction in patients with head and neck cancer: a scoping review of physical salivary stimulation methods.

OBJECTIVE(S): This scoping review aims to map the scientific literature on the therapies currently available for physical salivary stimulation in individuals with hyposalivation caused by radiotherapy. STUDY DESIGN: Studies were included when they comprised the target population of adult individuals receiving radiotherapy of the head and neck region and who developed or were at risk of developing hyposalivation. Two reviewers selected the studies and extracted data on the type of physical salivary stimulation therapy used, the degree of glandular tissue involvement, and the percentage of salivary flow alteration. Therapies were classified according to either prophylactic application (before/during radiotherapy) or therapeutic application (post-radiotherapy). RESULTS: Sixteen articles were included: 4 tested transcutaneous electrical nerve stimulation (TENS), 3 studied low-level lasers, 7 researched acupunctures, and 2 investigated acupuncture-like TENS. The outcomes of the prophylactic studies indicated beneficial effects (similar salivary flow or reduced salivary flow loss), although most studies did not include a comparable control group. Therapeutic studies presented conflicting results. CONCLUSION(S): Prophylactic therapies of physical salivary stimulation may produce better effects than therapeutic applications. However, the protocols best indicated could not be defined. Well-designed, controlled clinical trials should be researched in the future to support the clinical recommendation of any of these treatments.

Fracture load and failure mode of semi-direct resin composite occlusal veneers: Influence of design and mechanical cycling.

AIMS: To evaluate the influence of the type of preparation and mechanical cycling on the fracture load and failure mode of semi-direct posterior resin composite restorations. METHODS: In total, 70 healthy third molars were used; 10 belonging to the control group (C - unprepared teeth) and 60 teeth prepared and restored with nanoparticle resin composite, divided into 3 groups (n = 20): O - exclusively occlusal preparation (Table Top); OV - occlusal preparation with buccal extension (Veneerlay); OVP - Occlusal preparation with chamfer on the proximal and buccal-lingual/palatal surfaces (Overlay). The preparations were performed with diamond burs with a thickness of 1 mm. The restorations were made with nanoparticulate resin composite (Filtek Z350 XT) and subsequently received additional polymerization (thermoprocessing). Cementation was performed with the use of universal adhesive system (Single Bond Universal) on dental substrate and dual resin cement (RelyX ARC). Half of the teeth in each group (n = 10) were submitted to mechanical cycling, simulating 6 months of clinical service (5 × 105 fatigue cycles), under dynamic loading of 130 N, at a frequency of 2Hz. Fracture load tests were performed in a universal testing machine with a 200 kgf load cell. Failure mode was classified using scores. Generalized linear models and Fisher Exact tests were applied to the data (significance level of 5%). RESULTS: There were no significant differences between the types of preparation (p = 0.9435), or relative to cycling (p = 0.3764). The Fisher Exact Test showed a significant association between the groups and the type of failure (p = 0.0006), with preparations O (with cycling) and OVP (with and without cycling) exhibiting most failures with restoration fractures without involvement of the dental remnant. CONCLUSIONS: Semi-direct restorations showed resistance to fracture load similar to that of healthy teeth and were capable of withstanding functional masticatory loads. The type of preparation influenced the failure mode of teeth. The Table Top and Overlay types of preparations were those had fewer catastrophic failures, suggesting that since they are more conservative preparations, they made it possible for the tooth to receive a new restorative procedure in the future in cases of failures.

CREEPY: CRISPR-mediated editing of synthetic episomes in yeast.

Use of synthetic genomics to design and build 'big' DNA has revolutionized our ability to answer fundamental biological questions by employing a bottom-up approach. Saccharomyces cerevisiae, or budding yeast, has become the major platform to assemble large synthetic constructs thanks to its powerful homologous recombination machinery and the availability of well-established molecular biology techniques. However, introducing designer variations to episomal assemblies with high efficiency and fidelity remains challenging. Here we describe CRISPR Engineering of EPisomes in Yeast, or CREEPY, a method for rapid engineering of large synthetic episomal DNA constructs. We demonstrate that CRISPR editing of circular episomes presents unique challenges compared to modifying native yeast chromosomes. We optimize CREEPY for efficient and precise multiplex editing of >100 kb yeast episomes, providing an expanded toolkit for synthetic genomics.

The potential of conventional and bulk-fill bioactive composites to inhibit the development of caries lesions around restorations.

PURPOSE: To determine caries inhibition potential of conventional and bulk-fill bioactive composites around restorations. METHODS: Enamel and dentin blocks were prepared using a diamond saw under water irrigation, finished (SiC, 600- and 800-grit) and polished (SiC 1,200, final polish= 0.2 µm). Blocks were then selected through enamel surface microhardness, and enamel and dentin standard cavities were restored (n=10/group) with conventional bioactive composite (Beautifil II, BTF), bulk-fill bioactive composite (Activa BioACTIVE, ACT), glass-ionomer cement (Ionofil Plus, ION), conventional composite (GrandioSO, GSO), and bulk-fill composite (Admira Fusion X-TRA, ADM). Afterwards, the blocks were subjected to pH cycling: 4 hours in demineralization and 20 hours in remineralization solutions for 7 days, before being cut in the middle. One half was used to calculate the carious lesion area (ΔS) using values obtained by cross-sectional microhardness (CSMH) testing. The other half was submitted to polarized light microscopy (PLM) and scanning electron microscopy (SEM). The % of internal gap formation (GAP) of restorations' replicas were analyzed under SEM. Data were analyzed by ANOVA and Tukey test (α= 5%). RESULTS: In terms of CSMH, ION group exhibited the lowest ΔS values, with no significant difference to ADM. The composites BTF and ACT were similar to each other (P< 0.05) and to their negative controls (GSO and ADM), respectively. ION showed lower caries formation under PLM, whereas the GSO group presented a greater demineralized area. ION presented the highest % of internal GAP formation. Bioactive composites (BTF and ACT) were similar to their corresponding conventional ones (GSO and ADM) in terms of GAP formation. CLINICAL SIGNIFICANCE: The glass-ionomer cement was more effective in inhibiting the formation of caries lesions around restorations. Because of the glass-ionomer cement's limited application in high load-bearing areas, the conventional bioactive composite would be a promising clinical choice.

Boophilin D1, a Kunitz type protease inhibitor, as a source of inhibitors for the ZIKA virus NS2B-NS3 protease.

Arboviruses are a global concern for a multitude of reasons, including their increased incidence and human mortality. Vectors associated with arboviruses include the mosquito Aedes sp., which is responsible for transmitting the Zika virus. Flaviviruses, like the Zika virus, present only one chymotrypsin-like serine protease (NS3) in their genome. Together with host enzymes, the NS2B co-factor NS3 protease complex are essential for the viral replication cycle by virus polyprotein processing. To search for Zika virus NS2B-NS3 protease (ZIKVPro) inhibitors, a phage display library was constructed using the Boophilin domain 1 (BoophD1), a thrombin inhibitor from the Kunitz family. A BoophilinD1 library mutated at positions P1-P4' was constructed, presenting a titer of 2.9x106 (cfu), and screened utilizing purified ZIKVPro. The results demonstrated at the P1-P4' positions the occurrence of 47% RALHA sequence (mut 12) and 11.8% RASWA sequence (mut14), SMRPT, or KALIP (wt) sequence. BoophD1-wt and mutants 12 and 14 were expressed and purified. The purified BoophD1 wt, mut 12 and 14, presented Ki values for ZIKVPro of 0.103, 0.116, and 0.101 µM, respectively. The BoophD1 mutant inhibitors inhibit the Dengue virus 2 protease (DENV2) with Ki values of 0.298, 0.271, and 0.379 µM, respectively. In conclusion, BoophD1 mut 12 and 14 selected for ZIKVPro demonstrated inhibitory activity like BoophD1-wt, suggesting that these are the strongest Zika inhibitors present in the BoophD1 mutated phage display library. Furthermore, BoophD1 mutants selected for ZIKVPro inhibit both Zika and Dengue 2 proteases making them potential pan-flavivirus inhibitors.

Immunological characterization of a VIR protein family member (VIR-14) in Plasmodium vivax-infected subjects from different epidemiological regions in Africa and South America.

Plasmodium vivax is a major challenge for malaria control due to its wide geographic distribution, high frequency of submicroscopic infections, and ability to induce relapses due to the latent forms present in the liver (hypnozoites). Deepening our knowledge of parasite biology and its molecular components is key to develop new tools for malaria control and elimination. This study aims to investigate and characterize a P. vivax protein (PvVir14) for its role in parasite biology and its interactions with the immune system. We collected sera or plasma from P.vivax-infected subjects in Brazil (n = 121) and Cambodia (n = 55), and from P. falciparum-infected subjects in Mali (n = 28), to assess antibody recognition of PvVir14. Circulating antibodies against PvVir14 appeared in 61% and 34.5% of subjects from Brazil and Cambodia, respectively, versus none (0%) of the P. falciparum-infected subjects from Mali who have no exposure to P. vivax. IgG1 and IgG3 most frequently contributed to anti-PvVir14 responses. PvVir14 antibody levels correlated with those against other well-characterized sporozoite/liver (PvCSP) and blood stage (PvDBP-RII) antigens, which were recognized by 7.6% and 42% of Brazilians, respectively. Concerning the cellular immune profiling of Brazilian subjects, PvVir14 seroreactive individuals displayed significantly higher levels of circulating atypical (CD21- CD27-) B cells, raising the possibility that atypical B cells may be contribute to the PvVir14 antibody response. When analyzed at a single-cell level, the B cell receptor gene hIGHV3-23 was only seen in subjects with active P.vivax infection where it comprised 20% of V gene usage. Among T cells, CD4+ and CD8+ levels differed (lower and higher, respectively) between subjects with versus without antibodies to PvVir14, while NKT cell levels were higher in those without antibodies. Specific B cell subsets, anti-PvVir14 circulating antibodies, and NKT cell levels declined after treatment of P. vivax. This study provides the immunological characterization of PvVir14, a unique P. vivax protein, and possible association with acute host's immune responses, providing new information of specific host-parasite interaction. Trial registration: TrialClinicalTrials.gov Identifier: NCT00663546 & ClinicalTrials.gov NCT02334462.

Modeling the role of charged residues in thermophilic proteins by rotamer and dynamic cross correlation analysis.

Discerning the determinants of protein thermostability is very important both from the theoretical and applied perspective. Different lines of evidence seem to indicate that a dynamical network of salt bridges/charged residues plays a fundamental role in the thermostability of enzymes. In this work, we applied measures of dynamic variance, like the Gini coefficients, Kullback-Leibler (KL) divergence and dynamic cross correlation (DCC) coefficients to compare the behavior of 3 pairs of homologous proteins from the thermophilic bacterium Thermus thermophilus and mesophilic Escherichia coli. Molecular dynamic (MD) simulations of these proteins were performed at 303 K and 363 K. In the characterization of their side chain rotamer distributions, the corresponding Gini coefficients and KL-divergence both revealed significant correlations with temperature. Similarly, a DCC analysis revealed a higher trend to de-correlate the movement of charged residues at higher temperatures in the thermophilic proteins, when compared with their mesophilic homologues. These results highlight the importance of dynamic electrostatic network interactions for the thermostability of enzymes.

Mpox vaccine and infection-driven human immune signatures.

Background: Mpox (formerly known as monkeypox) outbreaks outside endemic areas peaked in July 2022, infecting > 85,000 people and raising concerns about our preparedness against this emerging viral pathogen. Licensed and approved for mpox, the JYNNEOS vaccine has fewer side effects than previous smallpox vaccines and demonstrated efficacy against mpox infection in humans. Comparing JYNNEOS vaccine- and mpox-induced immunity is imperative to evaluate JYNNEOS' immunogenicity and inform vaccine administration and design. Methods: We examined the polyclonal serum (ELISA) and single B cell (heavy chain gene and transcriptome data) antibody repertoires and T cells (AIM and ICS assays) induced by the JYNNEOS vaccine as well as mpox infection. Findings: Gene-level plasmablast and antibody responses were negligible and JYNNEOS vaccinee sera displayed minimal binding to recombinant mpox proteins and native proteins from the 2022 outbreak strain. In contrast, recent mpox infection (within 20-102 days) induced robust serum antibody responses to A29L, A35R, A33R, B18R, and A30L, and to native mpox proteins, compared to vaccinees. JYNNEOS vaccine recipients presented comparable CD4 and CD8 T cell responses against orthopox peptides to those observed after mpox infection. Interpretation: JYNNEOS immunization does not elicit a robust B cell response, and its immunogenicity may be mediated by T cells. Funding: Research reported in this publication was supported, in part, by the National Cancer Institute of the National Institutes of Health under Award Number U54CA267776, U19AI168631(VS), as well as institutional funds from the Icahn School of Medicine.