Search Results (3,060)

Liquid biopsy is moving beyond mutation-centric assays to multimodal frameworks that integrate cell-free DNA (cfDNA) signals with additional analytes such as circulating tumor cells (CTCs) and extracellular vesicles (EVs). In this review, we summarize emerging technologies across analytes for early cancer detection, emphasizing sequencing and error-suppression strategies and the growing evidence for multi-cancer early detection (MCED), tissue-of-origin (TOO) inference, diagnostic triage, and longitudinal surveillance. At low tumor fractions, fragmentomic and methylation features preserve tissue and chromatin context; when combined with radiomics using deep learning, they support blood-first, high-specificity risk stratification, increase positive predictive value (PPV), reduce unnecessary procedures, and enhance early prediction of treatment response and relapse. Building on these findings, we propose a pathway-aware workflow: initial blood-based risk scoring, followed by organ-directed imaging, and targeted secondary testing when indicated. We further recommend that model reports include not only discrimination metrics but also calibration, decision-curve analysis, PPV/negative predictive value (NPV) at fixed specificity, and TOO accuracy, alongside multi-site external validation and blinded dataset splits to improve generalizability. Overall, liquid biopsy is transitioning from signal discovery to deployable multimodal decision systems; standardized pre-analytical and analytical workflows, robust error suppression, and prospective real-world evaluations will be pivotal for clinical implementation. Full article

The phage-resistant mutant (PRM) strains of Escherichia coli (E. coli) exhibited abundant genetic and phenotypic diversity. IS elements played a vital role in creating various genetic divergences and regulating gene functions under phage infection stress. Genetic variations of PRM strains derived from E. coli MG1655 and mutation frequencies of coevolved E. coli populations with phages were explored by high-throughput sequencing and resequencing. Infrequent-restriction-site PCR (IRS-PCR) and carbon utilization test revealed the genetic and phenotypic diversity of the PRM strains. Numerous and discrepant mutation sites (MSs) were observed in the PRM strains and the coevolved populations, and many MSs were related to the synthesis of flagella and LPS, which often serve as receptors in a phage invasion. The insertions of various IS elements in key gene locations were also frequently found in the PRM strains, which indicate for the first time that IS elements played a vital role in generating genetic divergence and regulating gene functions under phage infection stress. Resequencing revealed that the coevolved populations at three evolving stages had discrepant profiles of MSs, and nearly all detected MSs occurred in the coevolved populations, which led to coexisting phages that increased the mutation rates and expedited the occurrence of the defective MSs in E. coli populations. In summary, our results reveal that the widespread and abundant presence of phages may provide one important force driving bacterial genomic evolution and prompt bacterial genetic divergence via accelerated mutation and increased mutation rates in the E. coli genome. Full article

Background/Objectives: Caffeic acid is a hydroxycinnamic acid that has a wide range of applications in the medical field. The synthesis of caffeic acid using microbial fermentation technology is an environmentally friendly method. Methods: By engaging various enzymes, specifically 4-hydroxyphenylacetate 3-monooxygenase (HpaB), sourced from diverse bacterial strains, we successfully engineered a functional version of this enzyme within Escherichia coli, enabling the production of caffeic acid. In addition to the two common tyrosine ammonia lyases (TAL) and HpaC, different combinations of HpaB demonstrated varying abilities in converting the substrate L-tyrosine into the desired product, caffeic acid. Results: Under shake-flask culture conditions, the highest yield of caffeic acid was achieved with an enzyme mixture containing HpaB from Escherichia coli, reaching 75.88 mg/L. Enhancing the activity of the rate-limiting enzyme through engineering could potentially increase caffeic acid titer. This study aims to conduct a semi-rational design of HpaB through structure-based approaches to screen for mutants that can enhance the production of caffeic acid. Initially, the predicted three-dimensional structure of HpaB was generated using AlphaFold2, and subsequent analysis was conducted to pinpoint the critical mutation sites within the substrate-binding pocket. Five key amino acid residues (R113, Y117, H155, S210 and Y461) located in the vicinity of the flavin adenine dinucleotide binding domain in HpaB from Escherichia coli could be instrumental in modulating enzyme activity. Subsequently, the mutant S210G/Y117A was obtained by iterative saturation mutagenesis, which increased the titer of caffeic acid by 1.68-fold. The caffeic acid titer was further improved to 2335.48 mg/L in a 5 L fermenter. The findings show that the yield of caffeic acid was significantly enhanced through the integration of semi-rational design and fermentation process optimization. Full article

The interplay between chromatin structure and phase-separating proteins is an emerging topic in cell biology with implications for understanding disease states. Here, we investigate the functional relationship between bromodomain protein 4 (BRD4) and chromatin architecture. By combining molecular dynamics simulations with live-cell imaging, we demonstrate that BRD4, when mutated at specific N-terminus sites, significantly impacts the organization and dynamics of chromatin nanodomains, known as nucleosome clutches. Our findings reveal that a constitutively phosphorylated mutant of BRD4 condenses nucleosome clutches, while treatment with (+)-JQ1 increases the diffusion dynamics of single nucleosomes and decondenses nucleosome clutches. Simultaneously, we demonstrate that BRD4 mutations can alter localization of BRD4 to chromatin as well as modify single nucleosome dynamics. These results suggest that both chromatin binding and phase separation of BRD4 could co-regulate the nanoscale chromatin architecture and the chromatin microenvironment. Our observations shed light on the nuanced regulation of chromatin structure by BRD4, offering insights into its role in maintaining the nuclear architecture and transcriptional activity. Full article

The rapid growth of electric vehicle (EV) adoption poses significant challenges for the rational planning of charging infrastructure, where economic efficiency and service quality are inherently conflicting. To support scientific decision-making in charging station siting, this study proposes an integrated multi-objective optimization and decision-support framework that combines an improved Non-dominated Sorting Genetic Algorithm II (NSGA-II) with an entropy-weighted TOPSIS method. A bi-objective siting model is developed to simultaneously minimize total operator costs and maximize user satisfaction. User satisfaction is explicitly characterized by a nonlinear charging distance perception function and a queuing-theoretic waiting time model, enabling a more realistic representation of user service experience. To enhance convergence performance and solution diversity, the NSGA-II algorithm is improved through variable-wise random chaotic initialization, opposition-based learning, and adaptive crossover and mutation operators. The resulting Pareto-optimal solutions are further evaluated using an improved entropy-weighted TOPSIS approach to objectively identify representative compromise solutions. Simulation results demonstrate that the proposed framework achieves superior performance compared with the standard NSGA-II algorithm in terms of operating cost reduction, user satisfaction improvement, and multi-objective indicators, including hypervolume, inverted generational distance, and solution diversity. The findings confirm that the proposed NSGA-II–TOPSIS framework provides an effective, robust, and interpretable decision-support tool for EV charging station planning under conflicting objectives. Full article

Nanoparticles interact dynamically with proteins, often leading to adsorption-induced conformational changes that alter protein function and contribute to corona formation. Here we investigated the adsorption and unfolding of a model protein GB1 on latex nanoparticle surfaces using a combination of mutational analysis, equilibrium binding assays, stopped-flow kinetics and Φ-value interpretation. Seven site-directed variants of GB1 were studied to dissect residue-specific contributions to adsorption energetics. Fluorescence binding isotherms revealed that D46A and T53A mutations weakened surface affinity, while kinetic analysis demonstrated that D46A reduced adsorption rate by ~6-fold and produced a dramatic unfolding/refolding shift, identifying Asp46 as a key docking site. Φ-value analysis further highlighted Asp46 and Thr53 as central residues in the adsorption transition state, whereas mutations in the hydrophobic core or distal loops had negligible effects. These results support a dock–loosen–unfold mechanism in which electrostatic recognition initiates binding, followed by hydrophobic exposure and hairpin stabilization. This residue-level sampling of key sites advances mechanistic understanding of protein–nanoparticle interactions and suggests strategies for tuning surface charge to control corona formation. Our approach provides a generalizable method to map adsorption transition states, with implications for designing safer nanomaterials, predicting protein corona composition, and harnessing protein unfolding in biosensing applications. Full article

Background/Objectives: Assigned female at birth (AFAB) individuals who identify as transgender or gender-diverse (TGD) with concurrent breast cancer or high-risk genetic mutations represent a unique population, requiring consideration of oncologic and aesthetic goals. These patients sought chest masculinization with oncologic gender-affirming mastectomy (OGAM) or non-binary reconstruction to alleviate gender dysphoria and treat their breast cancer. There is limited literature on surgical techniques in this patient population. Methods: A retrospective chart review of AFAB TGD adults (>18 years of age) who underwent OGAM or non-binary reconstruction at the University of Washington between 2019 and 2023 was conducted. All patients had a consultation with a plastic surgeon for reconstruction and a minimum of one year follow-up. Demographic data, oncologic status, post-operative complications, and revision surgical history were collected. Results: Eight AFAB TGD individuals met the inclusion criteria. The mean age at the time of mastectomy was 35.13 years (SD = 8.04), and the mean BMI was 29.88 (SD = 6.40). Indications for mastectomy included a breast cancer diagnosis (N = 4) or a strong family history of breast cancer or genetic predisposition (N = 4). Two (25%) patients underwent nipple-sparing mastectomies (NSM), two patients (25%) underwent skin-sparing mastectomy with Goldilocks reconstruction, and four patients (50%) underwent simple mastectomy (oncologic gender-affirming mastectomy), flat closure with free nipple graft (FNG). Two patients had staged nipple mastectomy with secondary nipple reduction and fat grafting. Six patients had immediate reconstruction, four (50%) patients underwent immediate double-incision OGAM with FNG, and two (25%) patients underwent Goldilocks procedures—one with and one without FNG. One patient (12.5%) experienced a surgical site infection, and three patients (37.5%) underwent revision surgery. No patients had positive margins following their mastectomy. Conclusions: This case series highlights the importance of a multidisciplinary and highly personalized approach for AFAB and TGD individuals undergoing oncologic gender-affirming mastectomy or non-binary reconstruction. We reviewed reconstructive options performed at our institution, demonstrating safe oncologic and reconstructive techniques that emphasized collaboration between breast and plastic surgeons. Full article

The sustained growth of electricity demand and the global transition toward low-carbon energy systems have intensified the need for efficient, flexible, and reliable operation of electrical distribution networks. In this context, the coordinated integration of distributed renewable energy resources and demand-side flexibility has emerged as a key strategy to improve technical performance and economic efficiency. This work proposes an integrated optimization framework for active power supply in a radial, distribution-like network through the optimal siting and sizing of photovoltaic (PV) units and wind turbines (WTs), combined with a real-time pricing (RTP)-based demand-side response (DSR) program. The problem is formulated using the branch-flow (DistFlow) model, which explicitly represents voltage drops, branch power flows, and thermal limits in radial feeders. A multiobjective function is defined to jointly minimize annual operating costs, active power losses, and voltage deviations, subject to network operating constraints and inverter capability limits. Uncertainty associated with solar irradiance, wind speed, ambient temperature, load demand, and electricity prices is captured through probabilistic modeling and scenario-based analysis. To solve the resulting nonlinear and constrained optimization problem, an Improved Whale Optimization Algorithm (I-WaOA) is employed. The proposed algorithm enhances the classical Whale Optimization Algorithm by incorporating diversification and feasibility-oriented mechanisms, including Cauchy mutation, Fitness–Distance Balance (FDB), quasi-oppositional-based learning (QOBL), and quadratic penalty functions for constraint handling. These features promote robust convergence toward admissible solutions under stochastic operating conditions. The methodology is validated on a large-scale radialized network derived from the IEEE 118-bus benchmark, enabling a DistFlow-consistent assessment of technical and economic performance under realistic operating scenarios. The results demonstrate that the coordinated integration of PV, WT, and RTP-driven demand response leads to a reduction in feeder losses, an improvement in voltage profiles, and an enhanced voltage stability margin, as quantified through standard voltage deviation and fast voltage stability indices. Overall, the proposed framework provides a practical and scalable tool for supporting planning and operational decisions in modern power distribution networks with high renewable penetration and demand flexibility. Full article

The Na⁺/K⁺/2Cl⁻ cotransporter (NKCC) gene encodes a critical membrane transporter involved in cellular ion homeostasis and plays a pivotal role in osmoregulation and salinity adaptation in aquatic organisms. This study identified and validated SNP markers in the NKCC gene associated with low-salinity tolerance in Scylla paramamosain. Four SNPs (g.196C>A, g.8374T>A, g.8385T>A and g.91143T>A) were screened and genotyped in low-salinity tolerant and intolerant groups. Association analysis revealed that mutant genotypes at all four sites were significantly enriched in the tolerant group (p <0.05), with the values of odds ratios (OR) greater than 1. The tolerant group exhibited significantly higher genetic diversity than the intolerant group. Haplotype analysis showed the wild CTTT haplotype dominated in the intolerant group, while mutant-containing haplotypes were significantly elevated in the tolerant group. A positive correlation was observed between the mutant and NKCC expression. Functional validation by qRT-PCR demonstrated that mutant allele carriers exhibited significantly higher NKCC mRNA expression levels than the wild-type carriers. Moreover, the expression level of homozygous mutations is significantly higher than that of heterozygous mutations. These validated SNPs could provide effective molecular markers for marker-assisted selection breeding of low-salinity tolerant S. paramamosain strains, offering important theoretical and practical implications for sustainable aquaculture development. Full article

Bovine viral diarrhea virus (BVDV) primarily causes bovine viral diarrhea/mucosal disease, an infectious disease having a significant economic impact on the cattle-farming industry globally. Comprehensive monitoring and in-depth studies of the pathological characteristics of viruses are crucial in formulating effective prevention and control strategies. The isolation, identification, molecular characterization, and pathogenicity analysis of a BVDV strain isolated from persistently infected cattle ear tissue samples are reported in this study. This newly isolated strain is a noncytopathogenic BVDV, which we named HB2411. Homology between the HB2411 and U63479 strains was determined to be 96.7%, and the phylogenetic tree indicated that HB2411 belongs to the BVDV-1b subtype. Genetic variation analysis of the E2 protein of the HB2411 strain revealed multiple amino-acid mutation sites. Recombination analysis of the newly isolated HB2411 strain suggested a potential cross-geographical transmission event. BALB/c mice were intraperitoneally inoculated with the BVDV strain to evaluate the pathogenicity and virulence of BVDV-1b HB2411. BVDV was detected in multiple organs of BALB/c mice, with the highest viral load in the liver. BVDV infection promoted the expression of inflammatory cytokines in mice livers, necessitating further studies on the virulence and pathogenic mechanisms of this new strain to reduce economic losses caused to the animal husbandry industry. Full article

Monogenic forms of severe early-onset obesity often involve genetic disruptions in the hypothalamic leptin-melanocortin pathway. Pathogenic variants in the SIM1 gene, a key transcription factor required for the development of the paraventricular nucleus, are a known cause of Prader–Willi-like syndrome, characterized by hyperphagia, severe obesity, and developmental delay. We performed targeted next-generation sequencing of 52 obesity-associated genes on a cohort of pediatric patients with severe early-onset obesity. Identified variants were analyzed for population frequency and predicted pathogenicity using in silico tools. The structural impact of the novel missense variants was assessed using protein domain modeling with AlphaFold3. We identified five rare SIM1 variants in eleven patients. Four were heterozygous nonsynonymous variants: one frameshift in the bHLH domain (p.Ser18Ter), one frameshift in the Per-ARNT-Sim domain (p.His143Ter), and two missense variants, p.Pro30Ala and p.Ser663Leu. Structural modeling suggested that the missense variants are likely to disrupt critical protein–protein interactions. The fifth variant was a synonymous change, c.1173G>A, p.(Ser391Ser), which was detected in five unrelated patients. Bioinformatic analysis predicted that this variant could alter splicing. Structural modeling suggested that the missense variants interfere with SIM1 function. This study expands the mutational spectrum of SIM1-linked monogenic obesity, reporting novel likely pathogenic frameshift variants, a missense variant, and a recurrent synonymous variant with a potential splice-site effect. The majority of the variants are predicted to affect the SIM1 protein. Our findings strengthen the critical role of the SIM1 gene in hypothalamic development and energy homeostasis. The results underscore the importance of including the SIM1 gene in genetic testing panels for children with severe obesity and hyperphagia, enabling precise diagnosis and potential future personalized management. Functional in vitro or in vivo validation of these variants is required to confirm their pathogenicity. Full article

Enterovirus D68 (EV-D68), a neurotropic respiratory pathogen, poses a considerable clinical threat through its link to pediatric acute flaccid myelitis (AFM) and severe respiratory illness. The possibility of recurrent epidemics, evidenced since the 2014 outbreak, remains a major concern. Genomic determinants of virulence are central to this threat. Sequence variations that affect host–receptor interactions, immune evasion, and replication efficiency serve as critical modifiers of pathogenicity. This article systematically reviews the evidence for specific genomic sites that enhance EV-D68 virulence, focusing on three critical regions: the VP1 receptor-binding site, the 2Apro/TRAF3 cleavage site, and the 3Cpro immunoregulatory region. Mutations in the VP1 receptor-binding site can alter affinity for host receptors such as sialic acid, heparan sulfate, and MFSD6, thereby shaping viral entry and tissue tropism. Alterations in the 2Apro/TRAF3 cleavage site may impair proteolytic cleavage of host TRAF3, attenuating immune evasion and reducing viral pathogenicity. Variations in the 3Cpro region affect its efficiency in cleaving host proteins involved in translation and autophagy, ultimately modulating viral replication and antiviral responses. Finally, we propose that monitoring for mutations in these key virulence determinants, particularly within the surface-exposed VP1, is essential for effective outbreak preparedness. Full article

Ene reductases, belonging to the Old Yellow Enzyme (OYE) family, are widely used for biocatalysis. The OYE from Thermus scotoductus SA-01 (TsOYE) gained great attention due to its broad substrate scope, high stereoselectivity, thermostability, and catalytic versatility. Recently, the otherwise noncovalently bound flavin cofactor (FMN) was covalently anchored in several TsOYE mutants using the “flavin-fixing” method. However, the biochemical properties of these mutants remained unexplored. A detailed comparative study of wild-type (WT) TsOYE and the flavin-fixing variant F1 (F1 TsOYE) revealed that F1 TsOYE has a lower stability and poorer catalytic activity. Interestingly, both WT and F1 TsOYE have comparable redox potential values. These results suggest that the decrease in activity and stability is primarily caused by changes in structure and structural dynamics induced by the mutations and the covalent flavin-protein linkage. Replacing residues in the flavinylation recognition site did not result in significant repair of enzyme activity. Our findings highlight the sensitivity of TsOYE activity to covalent FMN incorporation and its associated mutations and underscore the necessity of structural insights for further rational design. This study also provides critical groundwork for optimizing the flavin-fixing strategy. Full article

The global distribution of HIV-1 subtypes exhibits significant regional variations, with evolving epidemiological patterns over time. China currently experiences concurrent circulation of multiple HIV-1 subtypes, and the transmission landscape is becoming increasingly complex and diversified. We performed prospective molecular surveillance and drug-resistance profiling of HIV-1 in Wuhan City to delineate the local genotypic structure and to guide antiretroviral therapy. A total of 149 whole blood samples from HIV-1-infected individuals preserved in 2022 at a hospital in Wuhan were selected. Peripheral-blood mononuclear cells (PBMCs) were isolated, total RNA extracted, and the Gag, Pol, and Env regions were amplified by nested RT-PCR and sequenced. The sequencing and phylogenetic tree results revealed that subtype B constituted the predominant clade (73/149, 49.1%), followed by CRF07_BC (20, 13.4%), CRF01_AE (13, 8.7%), CRF55_01B (2, 1.3%), and subtype C (1, 0.7%). Drug resistance mutations were detected in 36 cases, involving 41 mutation sites across 21 distinct types. Resistance profiles included two protease inhibitor-associated mutation sites (2 types), seven nucleoside reverse transcriptase inhibitor (NRTI)-related mutation sites (6 types), and 32 non-nucleoside reverse transcriptase inhibitor (NNRTI)-associated mutation sites (13 types). Full article

Chinese wolfberry (Lycium barbarum L.), a specialty crop with ecological, medical, and economic value in Ningxia province of China, is subject to severe damage from Aphis gossypii Glover. Currently, A. gossypii populations show extremely high-level resistance to beta-cypermethrin in the major wolfberry planting areas in Ningxia. The specific resistance mechanisms, however, are still not known. In this work, we collected a field A. gossypii strain (HSP) from a wolfberry orchard in Ningxia in 2021 using a single-time sampling method, and its resistance to beta-cypermethrin was determined to be extremely high (994.74-fold) as compared with that of a susceptible strain (SS). Then we explored the potential resistance mechanisms from two aspects, namely, metabolic detoxification and target-site alterations. Bioassays of beta-cypermethrin with or without a synergist showed that piperonyl butoxide (PBO) significantly increased the toxicity of beta-cypermethrin (4.72-fold) to the HSP strain, while triphenyl phosphate (TPP) and diethyl maleate (DEM) exhibited no significant synergistic effects. Correspondingly, the O-demethylase activity of cytochrome P450s in the HSP strain was 1.68-fold higher than that in the susceptive strain (SS), whereas changes in carboxylesterases and glutathione S-transferases activities were unremarkable. Also, fifteen upregulated P450 genes were identified by both RNA-Seq and qRT-PCR technologies, containing eleven CYP6 genes, three CYP4 genes, and one CYP380 gene. Especially, five CYP6 genes with high relative expression levels (>3.00-fold) were intensively expressed by beta-cypermethrin induction in the HSP aphids. These metabolism-related results indicate the key role of P450-mediated metabolic detoxification in HSP resistance to beta-cypermethrin. Sequencing of voltage-gated sodium channel (VGSC) genes identified a prevalent M918L mutation and a new G1012D mutation in HSP A. gossypii. Moreover, heterozygous 918 M/L and 918 M/L + G1012D mutations were the dominant genotypes with frequencies of 60.00% and 36.67% in the HSP population, respectively. Overall, VGSC mutations along with P450-mediated metabolic resistance contributed to the extremely high resistance of the HSP wolfberry aphids to beta-cypermethrin, providing support for A. gossypii control and resistance management in the wolfberry planting areas of Ningxia using insecticides with different modes of action. Full article