Despite advances in algorithms and computational platforms, de-novo genome assembly remains a challenging process. Due to the constant innovation in sequencing technologies (Sanger, SOLiD, Illumina, 454, PacBio and Oxford Nanopore), genome assembly has evolved to respond to the changes in input data type. This paper includes a broad and comparative review of the most recent short-read, long-read and hybrid assembly techniques. In this review, we provide (1) an algorithmic description of the important processes in the workflow that introduces fundamental concepts and improvements; (2) a review of existing software that explains possible options for genome assembly; and (3) a comparison of the accuracy and the performance of existing methods executed on the same computer using the same processing capabilities and using the same set of real and synthetic datasets. Such evaluation allows a fair and precise comparison of accuracy in all aspects. As a result, this paper identifies both the strengths and weaknesses of each method. This comparative review is unique in providing a detailed comparison of a broad spectrum of cutting-edge algorithms and methods.

The origin of SARS-CoV-2 is debated, even after 18 months into the COVID-19 pandemic and a special investigation conducted by the World Health Organization. The RaTG13 sequence has been a highlight of discussions surrounding the origin of SARS-CoV-2. Here we express our opinion about the need for better reporting standards for information about sequencing data, especially for pathogens, citing our findings with the reported RaTG13 genome.

Leishmania major is the main causative agent of cutaneous leishmaniasis in humans. The Friedlin strain of this species (LmjF) was chosen when a multi-laboratory consortium undertook the objective of deciphering the first genome sequence for a parasite of the genus Leishmania. The objective was successfully attained in 2005, and this represented a milestone for Leishmania molecular biology studies around the world. Although the LmjF genome sequence was done following a shotgun strategy and using classical Sanger sequencing, the results were excellent and this genome assembly served as the reference for subsequent genome assemblies in other Leishmania species. Here, we present a new assembly for the genome of this strain (named LMJFC for clarity), generated by the combination of two high throughput sequencing platforms, Illumina short-read sequencing and PacBio Single Molecular Real-Time (SMRT) sequencing, which provides long-read sequences. Apart from resolving uncertain nucleotide positions, several genomic regions have been reorganized and a more precise composition of tandemly repeated gene loci was attained. Additionally, the genome annotation has been improved by adding 542 genes and more accurate coding-sequences defined for around two hundred genes, based on the transcriptome delimitation also carried out in this work. As a result, we are providing gene models (including untranslated regions and introns) for 11,238 genes. Genomic information ultimately determines the biology of every organism; therefore, our understanding of molecular mechanisms will depend on the availability of precise genome sequences and accurate gene annotations. In this regards, this work is providing an improved genome sequence and updated transcriptome annotations for the reference L. major Friedlin strain.

Antimalarial drug resistance has become a real public health problem despite WHO measures. New sequencing technologies make it possible to investigate genomic variations associated with resistant phenotypes at the genome-wide scale. Based on the use of hemisynthetic nanopores, the PromethION technology from Oxford Nanopore Technologies can produce long-read sequences, in contrast to previous short-read technologies used as gold-standard to sequence Plasmodium. Two clones of P.falciparum (Pf3D7 and PfW2) were sequenced in long-read using the PromethION sequencer from Oxford Nanopore Technologies without genomic amplification. This made it possible to create a processing analysis pipeline for human Plasmodium with ONT Fastq only. De novo assembly revealed N50 lengths of 18,488 kb and 17,502 kb for the Pf3D7 and PfW2, respectively. The genome size was estimated at 23,235,407 base pairs for the Pf3D7 clone and 21,712,038 base pairs for the PfW2 clone. The average genome coverage depth was estimated at 787X and 653X for the Pf3D7 and PfW2 clones, respectively. This study proposes an assembly processing pipeline for the human Plasmodium genome using software’s adapted to large ONT data and the high AT percentage of Plasmodium. This search provides all the parameters, which have been optimized, for use with the software selected in the pipeline.

Advances in DNA sequencing technology are facilitating genomic analyses of unprecedented scope and scale, widening the gap between our abilities to generate and fully exploit biological sequence data. Comparable analytical challenges are encountered in other data-intensive fields involving sequential data, such as signal processing, in which dimensionality reduction (i.e., compression) methods are routinely used to lessen the computational burden of analyses. In this work we explore the application of dimensionality reduction methods to numerically represent high-throughput sequence data for three important biological applications of virus sequence data: reference-based mapping, short sequence classification and de novo assembly. Despite using highly compressed sequence transformations to accelerate the processes, our sequence processing approach yielded comparable accuracy to existing approaches, and are ideally suited for sequences originating from highly diverse virus populations. We demonstrate the application of our methodology to both synthetic and real viral pathogen sequence data. Our results show that the use of highly compressed sequence approximations can provide accurate results and that useful analytical performance can be retained and even enhanced through appropriate dimensionality reduction of sequence data.

Background: An outbreak of Mpox disease caused by a mutant clade 1b virus (MPXV) in Kamituga area, South Kivu Province, DRC has rapidly grown to involve neighboring countries in this region. This now has the authorities worried, due to the limited supply of Mpox vaccines with emergence use licensure (EUL) available. As an emergency local response to this challenge, we set out to identify epitopes of clade 1b MPXV surface proteins as targets for the rapid research and development (R & D) of Mpox medical countermeasures. Methods: The H3L (OPG108–heparin binding protein) and A17L (OPG143–myristylated protein) were selected by virtue of their surface localization, stability and known role in MPXV host cell fusion entry. Within the immune epitope database analysis resource (IEDB-AR), surface accessibility, antigenicity and 3-D localisation (DiscoTope) we examined to identify > 9-nomer epitopes Results: Three (3) and six(6) candidate >9-nomer epitopes were derived for H3L and A17L surface fusion protein. Across all parameters, the two best identified epitopes were as follows:—H3L (28-HINDQKFDDVKDNEVMQEKR-47 and 236-YVEHDPRLVAEH-247) and A17L (104-QNEVTPEYIKDLKYATDYIA-123;) 270-KKDYLLGDSDSVAKCCSKTNTKHCPKIFNNNYKTEHC-290). Conclusions: These epitopes— alone, multiplexed (say as a nano-spiked virus-like particles, VLPs) or fused as a chimeric recombinants form immunogens for the rapid production of protein sub-unit vaccines. Inversely, derivative monoclonal antibodies (mAbs) from the same are precursors for therapeutic cocktails and or antigen, rapid diagnostic tests (RDT).

To understand the molecular mechanism associated with growth variability in bivalves, the Solexa/Illumina technology was employed to analyze the transcriptomic profiles of extreme growth rate differences (fast- VS. slow-growing individuals) in one full-sib family of the ark shell Scapharca subcrenata. De novo assembly of S. subcrenata transcriptome yielded 276,082,016 raw reads, which were assembled into 98,502 unique transcripts by Trinity strategy. A total of 6,357 differentially expressed genes (DEGs) were obtained between fast- and slow-growing individuals, with 580 up-regulated expression and 5777 down-regulated expression. Functional annotation revealed that the largest proportion of DEGs were classified to the large or small subunit ribosomal protein, all of which showed significantly lower expression levels in fast-growing group than those in slow-growing group. GO enrichment analysis identified the maximum of DEGs to biological process, followed by molecular function and cellular component. Most of the top enriched KEGG pathways were related to energy metabolism, protein synthesis and degradation. These findings reveal the link between gene expression and contrasting phenotypes in ark shells, which support that fast-growing individuals may be resulted from decreased energy requirements for metabolism maintenance, accompanying with greater efficiency of protein synthesis and degradation in bivalves.

The advent of high throughput sequencing has enabled researchers to systematically evaluate the genetic variations in cancer, resulting in identifying many cancer-associated genes. Although cancers in the same tissue are widely categorized in the same group, they demonstrate many differences concerning their mutational profiles. Hence there is no “silver bullet” for the treatment of a cancer type. This reveals the importance of developing a pipeline to identify cancer-associated genes accurately and re-classify patients with similar mutational profiles. Classification of cancer patients with similar mutational profiles may help discover subtypes of cancer patients who might benefit from specific treatment types. In this study, we propose a new machine learning pipeline to identify protein-coding genes mutated in a significant portion of samples to identify cancer subtypes. We applied our pipeline to 12270 samples collected from the International Cancer Genome Consortium (ICGC), covering 19 cancer types. Here we identified 17 different cancer subtypes. Comprehensive phenotypic and genotypic analysis indicates distinguishable properties, including unique cancer-related signaling pathways, in which, for most of them, targeted treatment options are currently available. This new subtyping approach offers a novel opportunity for cancer drug development based on the mutational profile of patients. We also comprehensive study the causes of mutations among samples in each subtype by mining the mutational signatures, which provides important insight into their active molecular mechanisms. Some of the pathways we identified in most subtypes, including the cell cycle and the Axon guidance pathways, are frequently observed in cancer disease. Interestingly, we also identified several mutated genes and different rates of mutation in multiple cancer subtypes. In addition, our study on “gene-motif” suggests the importance of considering both the context of the mutations and mutational processes in identifying cancer-associated genes. The source codes for our proposed clustering pipeline and analysis are publicly available at: https://github.com/bcb-sut/Pan-Cancer.

Poplar and willow species in the Salicaceae are dioecious, yet have been shown to use different sex determination systems located on different chromosomes. Willows in the section Vetrix are interesting for comparative studies of sex determination systems, yet genomic resources for these species are still quite limited. Only a few annotated reference genome assemblies are available, despite many species in use in breeding programs. Here we present de novo assemblies and annotations of 11 shrub willow genomes from six species. Copy number variation of candidate sex determination genes within each genome was characterized and revealed remarkable differences in putative master regulator gene duplication and deletion. We also analyzed copy number and expression of candidate genes involved in floral secondary metabolism, and identified substantial variation across genotypes, which can be used for parental selection in breeding programs. Lastly, we report on a genotype that produces only female descendants and identified gene presence/absence variation in the mitochondrial genome that may be responsible for this unusual inheritance.

The catalytic parameters describing enzyme catalysis are discussed in the context of the design of de novo enzymes. It is shown that high values of the catalytic efficiency may in some cases reflect tight substrate binding and lead to an immediate levelling-off of rates to reach a low plateau, an undesirable scenario for many industrial applications. It is argued that increasing the catalytic rate constant should be a main target in de novo enzyme engineering.

The Human Herpesviruses persist in the form of a latent infection in specialized cell types. During latency, the herpesvirus genomes associate with cellular histone proteins and the viral lytic genes assemble into transcriptionally repressive heterochromatin. Although there is divergence in the nature of heterochromatin on latent herpesvirus genomes, in general the genomes assemble into forms of heterochromatin that can convert to euchromatin to permit gene expression and therefore reactivation. This reversible form of heterochromatin is known as facultative heterochromatin and is most commonly characterized by polycomb silencing. Polycomb silencing is prevalent on the cellular genome and plays a role in developmentally regulated and imprinted genes, as well as X chromosome inactivation. As herpesviruses initially enter the cell in an un-chromatinized state, they provide an optimal system to study how de novo facultative heterochromatin is targeted to regions of DNA and how it contributes to silencing. Here, we describe how polycomb-mediated silencing potentially assembles onto herpesvirus genomes, synergizing what is known about herpesvirus latency with facultative heterochromatin targeting to the cellular genome. A greater understanding of polycomb silencing of herpesviruses will inform on the mechanism of persistence and reactivation of these pathogenic human viruses and provide clues regarding how de novo facultative heterochromatin forms on the cellular genome.

Autism spectrum disorder (ASD) is a common condition with lifelong implications. The last decade has seen dramatic improvements in DNA sequencing and related bioinformatics and databases. We analyzed the raw DNA sequencing files on the Variantyx® bioinformatics platform for the last 50 ASD patients evaluated with trio whole genome sequencing (trio-WGS). “Qualified” variants were defined as coding, rare, and evolutionarily conserved. Primary Diagnostic Variants (PDV) additionally were in genes directly linked to ASD and matched clinical correlation. A PDV was identified in 34/50 (68%) of cases, including 25 (50%) cases with heterozygous de novo and 10 (20%) with inherited variants. De novo variants in genes directly associated with ASD were far more likely to be Qualifying than non-Qualifying versus a control group of genes (P=0.0002), validating that most are indeed disease related. Sequence reanalysis increased diagnostic yield from 28% to 68%, mostly by inclusion of de novo PDVs in genes not yet reported as ASD associated. Thirty-three subjects (66%) had treatment recommendation(s) based on DNA analyses. Our results demonstrate high yield of trio-WGS for revealing molecular diagnoses in ASD that is greatly enhanced by re-analyzing DNA sequencing files. In contrast to previous reports, de novo variants dominate the findings, mostly representing novel conditions. This has implications to the cause and rising prevalence of autism.

The venom proteome of Temple Pit Viper (Tropidolaemus wagleri) is unique among pit vipers, characterized by a high abundance of a neurotoxic peptide, waglerin. To further explore the genetic diversity of its toxins, the present study de novo assembled the venom gland transcriptome of T. wagleri from west Malaysia. Among the 15 toxin gene families discovered, gene annotation and expression analysis reveal the dominating trend of bradykinin-potentiating peptide/angiotensin-converting enzyme inhibitor-C-type natriuretic peptide (BPP/ACEI-CNP, 76.19% of all-toxin transcription) in the transcriptome, followed by P-III snake venom metalloproteases (13.91%) and other toxins. The transcript TwBNP01 of BPP/ACEI-CNP represents a large precursor gene (209 amino acid residues) containing the coding region for waglerin (24 residues). TwBNP01 shows substantial sequence variations from the corresponding genes of its sister species, T. subannulatus of Northern Philippines, and other viperid species which diversely code for proline-rich small peptides such as bradykinin-potentiating peptides (BPPs). The waglerin/waglerin-like peptides, BPPs and azemiopsin are proline-rich, evolving de novo from multiple hyper-mutatable propeptide regions within the orthologous BPP/ACEI-CNP genes. Neofunctionalization of the peptides results in phylogenetic constraints consistent with a phenotypic dichotomy, where Tropidolaemus spp. and Azemiops feae convergently evolve a neurotoxic trait while vasoactive BPPs evolve only in other species.

The New Zealand brushtail possum (Trichosurus vulpecula), introduced from Australia in the 1850s, is an invasive species that is widespread throughout New Zealand and forms the largest self-sustained reservoir of bovine tuberculosis (Mycobacterium bovis) in the wild. Conservation and agricultural authorities regularly apply a series of population control measures to suppress brushtail possum populations. The evolutionary consequence of more than half a century of intensive population control operations on the species genomic diversity and population structure is hindered by a paucity of available genomic resources for the species. This study is the first to characterise the functional content and diversity of brushtail possum liver and brain cerebral cortex transcriptomes. Raw sequences from hepatic cells and cerebral cortex were assembled into 58,001 and 64,735 transcripts in hepatic cells and cerebral cortex, respectively. Functional annotation and polymorphism assignment of the assembled transcripts demonstrated a considerable level of variation in the core metabolic pathways that represent potential targets for selection pressure exerted by chemical toxicants. This study suggests that the brushtail possum population in New Zealand harbours considerable levels of variation in metabolic pathways that could potentially promote the development of tolerance against chemical toxicants.

Autism spectrum disorder (ASD) is a common condition with lifelong implications and a strong hereditary component suggesting genetic underpinnings. The last decade has seen dramatic improvements in DNA sequencing and related bioinformatics and databases. We analyzed the raw DNA sequencing files on the Variantyx® bioinformatics platform for the last 50 ASD patients evaluated with trio whole genome sequencing (trio-WGS). “Qualified” variants were defined as coding, rare, and evolutionarily conserved. Primary Diagnostic Variants (PDV) additionally were in genes directly linked to ASD and matched clinical correlation. A PDV was identified in 34/50 (68%) of cases, including 25 (50%) cases with heterozygous de novo and 10 (20%) with inherited variants. De novo variants in genes directly associated with ASD were far more likely to be Qualifying than non-Qualifying versus a control group of genes not associated with ASD (P = 0.0002, odds ratio 29), validating that most are indeed disease related. Only 14/34 (41%) of PDV cases had the variant listed on the laboratory report, and reanalysis increased diagnostic yield from 28% to 68%. Variants that we assigned as PDVs yet not on the report were predominately de novo in genes not yet reported as ASD associated. Many subjects both with and without a PDV had inherited Qualifying variants in known ASD-associated genes, suggesting polygenic inheritance. Thirty-three subjects (66%) had treatment recommendation(s) based on DNA analyses. Our results demonstrate high yield of trio-WGS for revealing molecular diagnoses in ASD that is greatly enhanced by re-analyzing DNA sequencing files. In contrast to previous reports, de novo variants dominate the findings, mostly representing novel conditions. This has implications to the cause and rising prevalence of autism.

In this paper we propose the generation of synthetic small and more sophisticated molecule structures that optimize the binding affinity to a target (ASYNT-GAN). To achieve this we leverage on three important achievements in A.I.: Attention, Deep Learning on Graphs and Generative Adversarial Networks. Similar to text generation based on parts of text we are able to generate a molecule architecture based on an existing target. By adopting this approach, we propose a novel way of searching for existing compounds that are suitable candidates. Similar to question and answer Natural Language solutions we are able to find drugs with highest relevance to a target. We are able to identify substructures of the molecular structure that are the most suitable for binding. In addition, we are proposing a novel way of generating the molecule in 3D space in such a way that the binding is optimized. We show that we are able to generate compound structures and protein structures that are optimised for binding to a target.

Long-read single-molecule sequencing has revolutionized de novo genome assembly and enabled the automated reconstruction of reference-quality genomes. It also has been widely used to study structural variants, phase haplotypes and more. Here, we introduce the assembler— SMARTdenovo, which is an SMS assembler that follows the overlap-layout-consensus (OLC) paradigm. SMARTdenovo (RRID: SCR_017622) was designed to be a fast assembler that did not require highly accurate raw reads for error correction, unlike other, contemporaneous SMS assemblers. It has performed well for evaluating congeneric assemblers and has been successful for a variety of assembly projects. It is compatible with Canu for assembling high-quality genomes, and several of the assembly strategies in this program have been incorporated into subsequent popular assemblers. The assembler has been in use since 2015, and here we provide information on the development of SMARTdenovo and how to implement its algorithms into current projects.

The recent covid crisis has proven important lessons for academia and industry regarding digital reorganization. Among fascinating lessons from these times is the huge potential of data analytics and artificial intelligence. The crisis exponentially accelerated the adoption of analytics and artificial intelligence, and this momentum is predicted to continue into the 2020s and over. Moreover, drug development is a costly and time-consuming business, and only a minority of approved drugs return the revenue that exceeds the research and development costs. As a result, there is a huge drive to make drug discovery cheaper and faster. With modern algorithms and hardware, it is not too surprising that the new technologies of artificial intelligence and other computational simulation tools can help drug developers. In only two years of covid research, many novel molecules have been designed/identified using artificial intelligence methods with astonishing results in terms of time and effectiveness. This paper will review the most significant research on artificial intelligence in the de novo drug design for COVID-19 pharmaceutical research.

The real-world outcomes of patients with metastatic prostate cancer (mPCa) are largely unexplored. We investigated the improvements in overall survival (OS) and cancer-specific survival (CSS) in patients with de novo mPCa in latest years. The USA SEER Research Data (2000-2017) were analyzed using the SEER*Stat software. The Kaplan-Meier method and Cox regression were used. Patients with de novo mPCa were allocated to 3 cohorts based on year of diagnosis: A (2000-2003), B (2004-2010), C (2011-2014). Maximum follow-up was fixed to 5 years. Overall, 26434 patients were included. Age, race and metastatic stage significantly affected OS and CSS. After adjustment for age and race, patients in cohort C showed 9% reduced risk of death (HR:0.91 [95% CI, 0.87-0.95], p<0.001) and 8% reduced risk of cancer-specific death (HR:0.91 [95% CI, 0.87-0.95], p<0.001) compared to those in cohort A. After adjustment for age, race and metastatic stage, patients in cohort C showed an improvement in OS and CSS compared to cohort B (HR:0.94 [95% CI, 0.91-0.97], p=0.001 and HR:0.89 [95% CI, 0.85-0.92], p<0.001). Patients with M1c disease had a more pronounced improvement in OS and CSS compared with the other stages. No differences were found between cohort B and C. In conclusion, the prognosis of de novo mPCa remains poor with a median OS of 30 months and a median CSS of 35 months. Limited OS and CSS improvements were observed in latest years.

Objectives: To investigate the etiology of amyotrophic lateral sclerosis (ALS) in an adult female patient. Methods: We conducted trio genome sequencing on a 35-year-old woman with progressive weakness in her left upper limb, as well as on her parents. Prior to sequencing, we performed a comprehensive neurological work-up on the patient, including neurological exam, electrophysiology, biomarker assessment, and brain and spinal cord MRI. Results : The propositus experienced weakness and atrophy in her left hand 6 months before the evaluation. This was associated with brisk reflexes and Hoffman sign in the same arm. Electroneuromyography revealed lower motor neuron involvement in three body regions. Neurofilament light chains were elevated in her cerebrospinal fluid. Brain imaging showed asymmetrical T2 hyperintensity of the corticospinal tracts and T2 linear hypointensity of the precentral gyri. Genome trio sequencing identified a likely pathogenic de novo variant in the propositus’s KIF1A gene (NM_001244008.2): c.574A>G, p.(Ile192Val). Discussion: Pathogenic variants in the KIF1A gene have been associated with a wide range of neurological manifestations called KIF1A-associated neurological diseases (KAND). The present report describe a likely pathogenic de novo variant in KIF1A associated with ALS, expanding the phenotypic spectrum of KAND and our understanding of the pathophysiology of ALS.

Abstract: Obstructive sleep apnea (OSA) is well known to often improve with non-supine positioning as opposed to su¬pine positioning. Emerging research supports a role for sleep position management in patients with central sleep apnea (CSA) as well. We report a case of de novo Complex Sleep apnea Syndrome (CompSAS) in a 78-year-old female, who presented after a car accident due to unclear syncope. Diagnostic polysomnography (PSG) showed a moderate OSA. A CompSAS developed under Automatic positive airway pressure (APAP), while download data of 4 years showed a good adherence. No significant benefit was reported under Adaptive Servo Ventilation (ASV) and BiPAP-ST, while a reduction of CSA in non-supine position was noticed. Oxygen and sleep positional therapy (SPT) were considered resulting in a significant improvement of CSA and sleep quality. Further research on the prevalence of positional CSA is needed.

An electrochemical bioassay based on the rotating droplet electrochemistry by using electron transfer mediator was developed for the evaluation of a wide variety of pollutants such as antibiotics, heavy metals and pesticides in the water environment. Ferricyanide was used as electron transfer mediator for obtaining the catalytic response of Escherichia coli. The electrochemical response of E.coli was measured via hydrodynamic chronoamperometry in a microdroplet on a screen-printed carbon electrode (SPCE). The constructed electrode system successfully evaluate the catalytic response of E.coli solution in presence of ferricyanide. The assay for antibiotic toxicity on E.coli was carried out. The EC50 for ampicillin, sulfamonomethoxine, chlorotetracycline, tetracycline and oxytetracycline valuated by the pre-incubation method were 0.26, 0.77, 5.25, 18.5, 19.0 µM, respectively. The toxicity order was ampicillin > sulfamonomethoxine > chlorotetracycline > tetracycline>oxytetracycline. The proposed method can be used to evaluate the antibiotic toxicities in different real sample, such as pond water, powder and raw milk. Recoveries were found in the range 90 and 99%. The developed methods do not require additional incubation time to evaluate toxicity.

An optical fiber scale sensor based on the detection principle of surface plasmon resonance (SPR) was developed for the rapid, high-sensitivity real-time evaluation of scale precipitation from geothermal fluids. The optical fiber SPR scale sensor was fabricated by depositing a gold thin film on the surface of an optical fiber with an exposed core. The optimal gold film thickness of the sensor was determined to be 30 nm, which achieved a refractive index sensitivity of 2140 nm per refractive index unit. A field test was conducted using geothermal brine from the Obama Binary Geothermal Power Plant in Unzen, Nagasaki Prefecture. A conventional optical fiber scale sensor and the SPR sensor were simultaneously assessed using raw and pH-adjusted brines. For the SPR sensor, a peak shift of 0.27 nm/min was observed at a response time of 1 min, whereas no change in transmittance was observed for the conventional sensor until 180 min. After the experiments, scanning electron microscopy–energy-dispersive spectroscopy analysis was conducted on the sensors, and findings showed that the deposition of Mg–SiO2 scale did not significantly differ between the two sensors. The developed SPR sensor achieved faster scale precipitation detection (tens of minutes to hours) than the conventional sensor.

In Southeast Asia, the Malayan Pit Viper (Calloselasma rhodostoma) is a venomous snake species of medical importance and bioprospecting potential. To unveil the diversity of its toxin genes, this study assembled and analyzed the de novo venom gland transcriptome of C. rhodostoma from Malaysia. The expression of toxin genes dominates the gland transcriptome by 53.78% of total transcript abundance (based on overall FPKM), in which 92 non-redundant transcripts belonging to 16 toxin families were identified. Snake venom metalloproteinase (SVMP, PI>PII>PIII) is the most dominant family (37.84% of all toxin FPKM), followed by phospholipase A2 (29.02%), bradykinin/angiotensinogen-converting enzyme inhibitor-C-type natriuretic peptide (16.30%), C-type lectin (CTL, 10.01%), snake venom serine protease (SVSP, 2.81%), L-amino acid oxidase (2.25%) and others (1.78%). The expressions of SVMP, CTL and SVSP correlate with hemorrhagic, anti-platelet and coagulopathic effects in envenoming. The SVMP metalloproteinase domains encode hemorrhagins (kistomin and rhodostoxin), while disintegrin (rhodostomin from P-II) is platelet-inhibitory. CTL gene homologues uncovered include rhodocytin (platelet aggregators) and rhodocetin (platelet inhibitor), which contribute to thrombocytopenia and platelet dysfunction. The major SVSP is a thrombin-like enzyme (ancrod homolog) responsible for defibrination in consumptive coagulopathy. The findings provide insight into the venom complexity of C. rhodostoma and the pathophysiology of envenoming.

De novo metastatic hormone-sensitive PC (mHSPC) accounts for 5-10% of all prostate cancer (PC) diagnoses but it is responsible for nearly 50% of PC-related deaths. Since 2015, the prognosis of mHSPC has slightly improved thanks to the introduction of new hormonal agents and chemotherapy combined with androgen deprivation therapy from the first-line setting. This review describes the current therapeutic opportunities in de novo mHSPC, focusing on potential molecular biomarkers identified in the main clinical trials that have changed the standard of care, the genomic features of de novo mHSPC, and the principal ongoing trials that are investigating new therapeutic approaches and the efficacy of a biomarker-guided treatment in this setting. The road towards a personalized treatment for de novo mHSPC is still long considering that the randomized clinical trials, which have furnished the basis of the current therapeutic options, stratified patients according to clinical criteria that not necessarily reflect the biological rationale of the chosen therapy. The role of transcriptomic profiling of mHSPC as predictive biomarker requires further validation, as well as it remains to ascertain how the genomic alterations detected in mHSPC, that are considered predictive in the castration-resistant disease, can be exploited in the mHSPC setting.

Snakes are a vital component of wildlife resources and are widely distributed across the globe. Bungarus multicinctus, a highly venomous snake, is found in central and southern China. B. multicinctus is a highly venomous snake and is distributed in central and southern China. Snakes are an ancient group of reptiles, and their genome resources can provide important clues for understanding the evolutionary history of reptiles. Meanwhile, genomic resources play a crucial role in comprehending the evolution of species. So far, the genomic resources of snakes are a rarity. In 2021, a snake sample was collected from Beiliu Longgukeng, Guangxi, which was identified as B. multicinctus through morphological identification. In this study, we present a highly contiguous genome of B. multicinctus with a size of 1.51 Gb. The genome contains a repeat content of 40.15%, with a total length exceeding 620 Mb. Additionally, we annotated a total of 24,869 functional genes. This research is of great significance for comprehending the evolution of B. multicinctus and provides a genomic basis for the genes involved in venom gland function.

Enterovirus A71 (EV-A71) infection is a major cause of hand, foot and mouth disease (HFMD) which may be occasionally associated with severe neurological complications. There are currently a lack of treatment options for EV-A71 infection. The Raf-MEK-ERK signaling pathway, in addition to its critical importance in the regulation of cell growth, differentiation and survival, has been shown to be essential for virus replication. In this study, we investigated the anti-EV-A71 activity of vemurafenib, a clinically approved B-Raf inhibitor used in the treatment of late stage melanoma. Vemurafenib exhibits potent anti-EV-A71 effect in cytopathic effect inhibition and viral load reduction assays, with half maximal effective concentration (EC50) at nanomolar concentration. Mechanistically, vemurafenib interrupts both EV-A71 genome replication and assembly. These findings expand the list of potential antiviral candidates of anti-EV-A71 therapeutics.

Palmitic acid metabolism involves delta-9 and delta-6 desaturase enzymes forming palmitoleic acid (9cis-16:1; n-7 series) and sapienic acid (6cis-16:1; n-10 series), respectively. The corresponding biological consequences and lipidomic research on these positional MUFA isomers are under development. Furthermore, sapienic acid can bring to the de novo synthesis of the n-10 polyunsaturated fatty acid (PUFA) sebaleic acid (5cis,8cis-18:2), but such transformations in cancer cells are not known. The model of Caco-2 cell line was used to monitor sapienic acid supplementation (150 and 300 μM) and evidence the formation of n-10 fatty acids as well as their incorporation at levels of membrane phospholipids and triglycerides. Comparison with palmitoleic and palmitic acids evidenced that lipid remodeling was influenced by the type of fatty acid and positional isomer, with increase of 8cis-18:1, n-10 PUFA and decrease of saturated fats in case of sapienic acid. Cholesteryl esters were formed only in case of sapienic acid. EC₅₀ of sapienic acid (232.3 μM at 96 hrs) was the highest found among the tested fatty acids, thus influencing cell viability that was only reduced at 25% at 300 μM, whereas palmitoleic acid induced cell death. Two-photon fluorescent microscopy with Laurdan as a fluorescent dye provided information on membrane fluidity, highlighting that sapienic acid increases the distribution of fluid regions, probably connected with the formation of 8cis-18:1 and the n-10 PUFA in cell lipidome. Our results bring evidence for MUFA positional isomers and de novo PUFA synthesis for developing lipidomic analysis and cancer research.

Smith-Magenis syndrome is a complex neurobehavioral genetic disorder. The spectrum of SMS phenotype is broad but commonly includes craniofacial, neurobehavioral, and otolaryngologic features. While the etiology of SMS is commonly attributed to one copy interstitial deletion in the 17p11.2 region (90-95% of cases), variants identified by sequence analysis in RAI1 have also been reported in 5-10% of cases. The phenotypic spectrum of patients with RAI1 sequence alterations is not clear. In this study, we report a 9-year-old male with global cognitive and psychomotor developmental delay, musculoskeletal and cardiovascular abnormalities, and dysmorphic craniofacial features. Whole genome quad analysis of the proband, unaffected parent, and unaffected brother identified a novel de novo RAI1:c.2736delC variant. This is the first report of this variant in the literature. This report also highlights the details of the genome analysis and this patient's phenotypic spectrum.

Sphingosine-1-phosphate (S1P) synthesized by sphingosine kinase (SPHK) is a signaling molecule, involved in cell proliferation, growth, differentiation, and survival. Indeed, a sharp increase of S1P was linked to the pathological outcome with inflammation, cancer metastasis or angiogenesis etc. In this regard, the SPHK/S1P axis regulation has been a specific issue in anticancer strategy to turn accumulated sphingosine (SPN) into cytotoxic ceramides (Cers). For these purposes, there have been numerous chemicals synthesized for SPHK inhibition. In this study, we investigated the comparative efficiency of dansylated PF-543 (DPF-543) on the Cers synthesis along with PF-543. DPF-543 deserved attention in strong cytotoxicity, due to the cytotoxic Cers accumulation by ceramide synthase (CerSs). DPF-543 exhibited dual actions on Cers synthesis by enhance the serine palmitoyltransferase (SPT) activity, and by inhibiting SPHKs which eventually induced an unusual environment of the high amount of 3-ketosphinganine and sphinganine (SPA). SPA in turn was consumed to synthesize Cers via de novo pathway. Interestingly, PF-543 increased only the SPN level, but not for SPA. In addition, DPF-543 mildly activates acid sphingomyelinase (aSMase) that contributes a partial increase on Cers. Collectively, a dansyl-modified DPF-543 relatively en-hanced Cers accumulation via de novo pathway which was not observed in PF-543. Our results demonstrated that the structural modification on SPHK inhibitors is still an attractive anticancer strategy by regulating sphingolipid metabolism.

Alkaloids are the most diversified nitrogen-containing secondary metabolites having antioxi-dant, and antimicrobial properties and are extensively used in pharmaceuticals to treat different types of cancer. Nicotiana serves as a reservoir of anticancer alkaloids and is also used as a model plant for the de novo synthesis of various anti-cancer molecules through genetic engineering. Up to 4% of the total dry weight of Nicotiana was found to be composed of alkaloids where nicotine, nornicotine, anatabine, and anabasine are reported as the dominant alkaloids. And, among the alkaloids present in Nicotiana, β-carboline (Harmane and Norharmane), Kynurenines are found to show anti-tumor effects, especially in the case of colon and breast cancer. Creating new or shunting of existing biosynthesis pathways in different species of Nicotiana resulted in de novo or increased synthesis of different anti-tumor molecules or their derivatives or precursors includ-ing Taxadiane (~22.5 µg/g), Artemisinin (~120 μg/g), Parthenolide (~2.05 ng/g), Costunolide (~60 ng/g), Etoposide (~1 mg/g), Crocin (~400 µg/g), Catharanthine (~60 ng/g), Tabersonine (~10 ng/g), Strictosidine (~0.23 mg/g), etc. Enrich precursor pool especially Dimethylallyl Diphos-phate (DMAPP) and down-regulating of other bi-product pathways, compartmentalization or metabolic shunting, or organelle-specific reconstitution of the precursor pool might trigger the enhanced accumulation of the targeted anticancer alkaloid in Nicotiana.

With the advancement of long-read sequencing technologies and their more widespread use for bacterial genomics, several methods for generating genome assemblies from error-prone long reads have been developed. These are complemented by various tools for assembly polishing using either long reads, short reads, or reference genomes. End users are therefore left with a plethora of possible combinations of programs for obtaining a final trusted assembly. Hence, there is also the need for measuring completeness and accuracy of such assemblies, for which, again, several evaluation methods implemented in various programs are available. In order to automatically run all these programs, I developed two workflows for the workflow management system Snakemake for bacterial genome assembly and evaluation of assemblies, which provide end users with an easy-to-run method for both tasks. The workflows are available as open source software under the MIT license at https://github.com/pmenzel/ont-assembly-snake and https://github.com/pmenzel/score-assemblies.

Cercospora beticola Sacc. is an ascomycete pathogen that causes Cercospora leaf spot in sugar beets (Beta vulgaris L.) and other related crops. The disease symptoms are characterized by circular lesions with an ash-gray centre and dark brown to purple margins on the leaves. It can lead to significant yield losses if not effectively managed. This study aims to characterize and assess the impact of bacterial biological control agents against C. beticola. Following a dual-culture screening, 18 bacteria exhibiting over 50% inhibition were selected, with six of them demonstrating more than 80% control. Identification of the bacteria was carried out through sequencing of the16S rDNA, revealing potentially 12 species belonging to six genera including Bacillus which was represented by four species. Additionally, the bacteria underwent in-depth characterization of biochemical and molecular properties, as well as plant-growth promotion. PCR detection of genes responsible for producing antifungal metabolites revealed that 83%, 78%, 89%, and 56% of the selected bacteria possessed the bacillomycin, iturin, fengycin, and surfactin encoding genes, respectively. Infrared spectroscopy analysis confirmed the presence of lipopeptide structure in the bacteria supernatant filtrate. Subsequently, the bacteria were assessed for their effect on sugar beet plants in controlled conditions. The bacteria exhibited notable capabilities, promoting growth in both roots and shoots, resulting in significant increases in root length, weight, and shoot length. The field experiment with four bacterial candidates demonstrated good performance against C. beticola compared to difenoconazole fungicide. These bacteria played a significant role in disease control, achieving a maximum efficacy of 77.42%, slightly below the 88.51% efficacy attained by difenoconazole. Additional field trials are necessary to verify the protective and growth-promoting effects of these candidates, whether applied individually, combined in consortia, or integrated with chemical inputs in sugar beet crop production.

Abstract: Patient blood samples are invaluable in clinical omics databases, yet current methodologies often fail to fully uncover the molecular mechanisms driving patient pathology. While Genome-scale Metabolic Models (GEMs) show promise in systems medicine by integrating various omics data, having only exometabolomic data remains a limiting factor. To address this gap, we introduce a comprehensive pipeline integrating GEMs with patient plasma metabolome. This pipeline constructs case-specific GEMs using literature-based and patient-specific metabolomic data. Novel computational methods, including adaptive sampling and an in-house developed algorithm for the rational exploration of the sampled space of solutions, enhance integration accuracy while improving computational performance. Model characterization involves task analysis in combination with clustering methods to identify critical cellular functions. The new pipeline was applied to a cohort of trauma patients to investigates shock-induced endotheliopathy using patient plasma metabolome data. By analyzing endothelial cell metabolism comprehensively, the pipeline identified critical substrates and contributed to the development of targeted therapeutic strategies. Our study demonstrates the efficacy of integrating patient plasma metabolome data into computational models to analyze endothelial cell metabolism in disease contexts. This approach offers a deeper understanding of metabolic dysregulations and provides insights into diseases with metabolic components and potential treatments.

The acquired immunodeficiency syndrome (AIDS) caused by the human immunodeficiency virus (HIV) continues to be a public health problem. In 2020, 680,000 people died from HIV-related causes, and 1.5 million people were infected. Antiretrovirals are only a way to control HIV infection but not to cure AIDS. As such, effective treatment must be developed to control AIDS. Developing a drug is not an easy task, and there is an enormous amount of work and economic resources invested. For this reason, it is highly convenient to employ computer-aided drug design methods, which can help generate and identify novel molecules. Using the de novo design, new novel molecules can be developed using fragments as building blocks. In this work, we develop a virtual-focused compound library of HIV-1 viral protease inhibitors from natural product fragments. Natural products are characterized by a large diversity of functional groups, many sp3 atoms, and chiral centers. Pseudo-natural products are a combination of natural products fragments that keep the desired structural characteristics from different natural products. An interactive version of chemical space visualization of virtual compounds focused on HIV-1 viral protease inhibitors from natural product fragments is freely available at https://figshare.com/s/ceb58d58e8f5585ce67e.

The purpose of this study was to determine the change in overall survival (OS) for patients with de novo metastatic breast cancer (dnMBC) over time. We conducted a retrospective cohort study with 1981 patients with dnMBC diagnosed between January 1995 and December 2017 at The University of Texas MD Anderson Cancer Center. OS was measured from the date of diagnosis of dnMBC. OS was compared between patients diagnosed during different time periods: 5-year periods and periods defined according to when key agents were approved for clinical use. The median OS was 3.4 years. The 5- and 10-year OS rates improved over time across both types of time periods. A subgroup analysis showed that OS improved significantly over time for the estrogen-receptor-positive/HER2-positive (ER+/HER2+) subtype, and exhibited a tendency toward improvement over time for the ER-negative (ER-)/HER2+ subtype. Median OS was significantly longer in patients with non-inflammatory breast cancer (P = .02) and in patients with ER+ disease, progesterone-receptor-positive disease, HER2+ disease, lower nuclear grade, locoregional therapy, and metastasis to a single organ (all P <.0001). These findings showed that OS at 5 and 10 years after diagnosis in patients with dnMBC improved over time. The significant improvements in OS over time for the ER+/HER2+ subtype and the tendency toward improvement for ER-/HER2+ subtype suggest the contribution of HER2-targeted therapy to survival.

Single cell RNA-seq (scRNA-seq) profiles conceal temporal and spatial tissue developmental information. De novo reconstruction of single cell temporal trajectory has been fairly addressed, but reverse engineering single cell 3D spatial tissue localization is hitherto landmark based, and de novo spatial reconstruction is a compelling computational open problem. Here we show that a new algorithm - named D-CE - for coalescent embedding of single cell transcriptomic networks can address this open problem. We rely merely on the spatial information encoded in the expression patterns of developmental signal transcription factor (DST) genes, and we find that D-CE of cell-cell association DST-transcriptomic networks reliably reconstructs the Geo-seq or single cell samples’ 3D spatial tissue distribution. Comparison to the novoSpaRC and CSOmap (recent and only available de novo 3D spatial reconstruction methods) on 16 datasets and 681 reconstructions, reveals a significantly distinctive superior performance of D-CE.

In China, 65 types of venomous snakes exist, with the Chinese Cobra Naja atra being prominent and a major cause of snakebites in humans. Furthermore, N. atra is a protected animal in some areas, as it has been listed as vulnerable by the International Union for Conservation of Nature. Recently, due to the medical value of snake venoms, venomics has experienced growing research interest. In particular, genomic resources are crucial for understanding the molecular mechanisms of venom production. Here, we report a highly continuous genome assembly of N. atra, based on a snake sample from Huangshan, Anhui, China. The size of this genome is 1.67 Gb, while its repeat content constitutes 37.8% of the genome. A total of 26,432 functional genes were annotated. This data provides an essential resource for studying venom production in N. atra. It may also provide guidance for the protection of this species.

The Oriental ratsnake Ptyas mucosa is a common non-venomous snake of the colubrid family, with a wide geographic range spanning much of South and Southeast Asia. P. mucosa is widely cultivated dut to it used in traditional medicine, scientific research, and handicrafts. Therefore, genome resources could play an important role in the efficacy of traditional medicine and the analysis of the living environment of the species. We collected a snake sample in Hezhou, Guangxi, China, which was identified as P. mucosa by morphological identification. Here we present a highly continuous P. mucosa genome with a genome size of 1.74Gb. The scaffold N50 length is 9.57Mb and the maximal length of scaffold is 78.3Mb, the P. mucosa genome has a CG content of 37.9% and the integrity of the gene reached 86.6%.Assembled using long-reads, the total length of the repeat sequence in the genome reached 735 Mb, and its repeat content was as high as 42.19%. A total of 24,869 functional genes were annotated. This study will assist in the understanding of the P. mucosa, and also provide a basis for medicinal research.

In 1984, Susumu Ohno hypothesized that the nylon-degrading enzyme NylB arose de novo via a frameshift mutation within a hypothetical precursor protein (PR.C). However, Ohno never tested his hypothesis or provided supporting biological evidence. For decades, Ohno’s famous frame-shift hypothesis has been uncritically accepted as the correct explanation for the origin of NylB and has been used to illustrate how simple it is for a totally new enzyme to arise spontaneously. In this paper we test Ohno’s hypothesis in light of data not available in 1984. We searched multiple protein databases and found that the NylB protein is widely occurring, has thousands of homologs, and is found in diverse organisms and diverse habitats. Conserved domain searches showed that the NylB sequence is homologous to beta lactamases - a family of highly conserved enzymes. However, our searches showed that there is no evidence for the existence of Ohno’s hypothetical PR.C protein, nor any credible homolog. Our results effectively falsify Ohno's frameshift hypothesis. We extended this analysis to other nylonases and found all the nylonases we examined had large numbers of homologs throughout the biosphere. This falsifies the long-held assumption that all nylonases evolved after the invention of nylon in 1935.

Garcinia indica (Thouars) Choisy (commonly known as Kokum) is an important medicinal plant in Ayurvedic medicine and belonging to the Clusiaceae family. Kokum is a highly traded medicinal plant species of India, dried rind of fruits and butter from seeds are traded in the markets. This study aimed to assemble the complete chloroplast (cp) genome of G. indica and compare it with the previously published cp genome within the Garcinia taxa to identify the potential species-specific barcode marker for Kokum authentication. The assembled cp genome, with a size of 1,56,891 bp, exhibits a typical quadripartite structure. The large single-copy (LSC) region spans 85,580 bp, and the small single-copy (SSC) region spans 17,181 bp, together comprising 64.5% of the genome. The pair of inverted repeats (IRA and IRB) are each 27,065 bp, covering the remaining 35.5% of the genome. A total of 126 unique genes were identified, including 86 protein-coding genes, 32 tRNA genes, and 8 rRNA genes. Phylogenetic analysis using complete cp genomes from 12 species in the Clusiaceae family indicated that 10 Garcinia species form a sister clade. Notably, the ycf1 gene in the LSC region was more divergent within the Garcinia species. This study presents the first report on the chloroplast genome of G. indica. This chloroplast genome resource provides a basis for identifying new DNA barcode marker as well as species-specific marker for herbal drug authentication of Kokum, and species delineation within the Garcinia genus.

The humpback puffer, Tetraodon palembangensis, also known as Pao palembangensis, is a species of poisonous freshwater pufferfish mainly distributed in Southeast Asia (Thailand, Laos, Malaysia and Indonesia). Despite interesting biological features, such as its very inactive nature, tetrodotoxin production and body expansion mechanisms, molecular research on the humpback puffer is still rare because of the lack of a high-quality reference genome. Here, we reported a first chromosome-level genome assembly of an adult humpback puffer, of which the genome size is 362 Mb with ~1.78 Mb contig N50 and ~15.8 Mb scaffold N50s. Based on the genome, ~61.5Mb (18.11%) repeat sequences were also identified, and totally 19,925 genes were annotated, 99.20% of which could be predicted with function using protein-coding function databases. Finally, a phylogenetic tree was constructed with single-copy gene families from ten teleost fishes. The humpback puffer genome will be a valuable genomic resource to illustrate possible mechanisms of tetrodotoxin synthesis and tolerance, providing clues for future detailed studies of biological toxins.

Concatemeric viral DNA is packaged into Bacteriophage P22 procapsids via a headful packaging mechanism mediated by a molecular machine consisting of small (gp3) and large (gp2) terminase subunits. Although a negative stain reconstruction exists for the terminase holoenzyme, it is not clear how this complex binds dodecameric portal protein located at a 5-fold mismatch vertex. Herein, we describe new assemblies for the holoenzyme. Both native mass spectrometry and transmission electron microscopy reveal that the P22 terminase complex adopts three main assemblies, which include a nonameric S-terminase bound to two L-terminase 1(gp3)9:2(gp2), two nonameric S-terminase bound to five L-terminase 2(gp3)9:5(gp2), and three nonameric S-terminase bound to seven L-terminase 3(gp3)9:7(gp2). Native agarose gel electrophoresis shows that the terminase complex interacts with procapsid with mild crosslinking. These results herein illustrate the P22 terminase complex can adopt a variety of confirmations and assembly states.

Brown-Spotted Pit viper (Protobothrops mucrosquamatus), also known as the Chinese habu, is a widespread and highly venomous snake distributed from from northeastern India to eastern China. Genomics research can help provide much insight in understanding venom components and natural selection in vipers. Here, we collected, sequenced and assembled the genome of a male P. mucrosquamatus individual from China, producing a highly continuous reference genome, with the length of 1.53 Gb and 41.18% repeat element content. From this 24,799 genes were identified, and 97.97% genes could be annotated. Nuclear genome single-copy genes phylogenetic tree including 6 species verified the validity of our genome assembly and annotation process. This research will contribute to further study on Protobothrops biology and the genetic basis of the snake venom.

Closely related to the model plant Arabidopsis thaliana, the genus Boechera is known to contain both sexual and apomictic species or accessions. Boechera retrofracta is a diploid sexually reproducing species and is thought to be an ancestral parent species of the apomictic species Boechera divaricarpa. Here we report the de novo assembly of the B. retrofracta genome using short Illumina and Roche reads from 1 paired-end and 3 mate pair libraries. The distribution of 23-mers from the paired end library has indicated a low level of heterozygosity and the presence of detectable duplications and triplications. The genome size was estimated to be equal 227 Mb. N50 of the assembled scaffolds was 2.3 Mb. 27048 protein-coding genes were predicted using a hybrid approach that combines homology-based and de novo methods. Also repeats, tRNA and rRNA genes were annotated. Finally, genes of B. retrofracta and 6 other Brassicaceae species were used for phylogenetic tree reconstruction. Also, a detailed analysis of evolution of the APOLLO apomixis-associated locus was performed. An assembled genome of B. retrofracta will help in the challenging assembly of the highly heterozygous genomes of hybrid apomictic species such as B. divaricarpa.

Recent advancements in artificial intelligence (AI) and deep learning have revolutionized the field of protein engineering, particularly in the area of de novo protein design. This review article explores the impact of AI-driven approaches on protein design, with a specific focus on the role of structure prediction neural networks, such as AlphaFold and RoseTTAFold. The article discusses the paradigm shift brought about by these networks, which have enabled the design of proteins with unique structures and functions that are not found in nature. The review covers various aspects of AI-driven protein design, including the use of protein language models to harness evolutionary information, the development of de novo protein design workflows utilizing deep learning, and the application of generative models like RFdiffusion and RoseTTAFold All-Atom. The article also highlights the successes and applications of AI-driven protein design across diverse domains, such as enzyme engineering, antibody design, and vaccine development. Additionally, the review identifies current challenges and future directions in the field, emphasizing the need to address limitations in modeling conformational dynamics and designing proteins for in vivo functionality. The article concludes by underscoring the potential of AI-driven protein design to transform various aspects of science and technology, while also acknowledging the importance of interdisciplinary collaboration and the development of robust pipelines for the validation and optimization of designed proteins. Overall, this comprehensive review serves as a valuable resource for researchers and practitioners interested in understanding the current state and future prospects of AI-driven protein design.

Induction heating is a fast, reproducible, and efficient heating method used in various manufacturing processes. However, there is no established additive manufacturing (AM) process based on induction heating using wire as feedstock. This study investigates a novel approach to AM based on inductive heating, where a steel wire is melted and droplets are detached periodically using a two-winding induction coil. The process parameters and energy input into the droplets are characterized. The induction generator exhibits sluggish response to the excitation voltage, resulting in a lag in the coil current. The process is captured using a high-speed camera, revealing regular droplet formation and uniform size and shape when operated within an appropriate process window. Larger drops and increased spatter formation occur outside this window. The proposed method allows for the production of droplets with almost spherical shapes. Further analysis and characterization of droplet formation and energy input provide insights into process optimization and overall efficiency.

Trimeresurus albolabris, also known as the white-lipped pit viper or white-lipped tree viper, is a highly venomous snake distributed across Southeast Asia and the cause of many snakebite cases. In this study, we report the first whole genome assembly of T. albolabris obtained with next-generation sequencing from a specimen collected in Mengzi, Yunnan, China. After genome sequencing and assembly, the genome of this male T. albolabris individual was 1.51 Gb in length and included 38.42% repeat-element content. Using this genome, 21,695 genes were identified, and 99.17% of genes could be annotated using gene functional databases. Our genome assembly and annotation process was validated using a phylogenetic tree, which included six species and focused on single-copy genes of nuclear genomes. This research will contribute to future studies on Trimeresurus biology and the genetic basis of snake venom.

Resistance evolution during exposure to non-lethal levels of antibiotics is influenced by various stress responses of bacteria which are known to affect growth rate. Here, we aim to disentangle how the interplay between resistance development and associated fitness costs is affected by stress responses. We performed de novo resistance evolution of wild-type strains and single-gene knockout strains in stress response pathways using four different antibiotics. Throughout resistance development, the increase in minimum inhibitory concentration (MIC) is accompanied by a gradual decrease in growth rate, most pronounced with amoxicillin or kanamycin. By measuring biomass yield on glucose and whole-genome sequences at intermediate and final timepoints, we identified two patterns of how the stress responses affect the correlation between MIC and growth rate. First, single-gene knockout E. coli strains associated with reactive oxygen species (ROS) acquire resistance faster and mutations related to antibiotic permeability and pumping out occur earlier. This increases the metabolic burden of resistant bacteria. Second, the ΔrelA knockout strain which has reduced (p)ppGpp synthesis, is restricted in its stringent response, leading to diminished growth rates. The ROS-related mutagenesis and the stringent response are vulnerabilities within the fitness of resistant strains that can possibly be targeted to prevent development of resistance.

Mutations, especially those at the protein-protein interaction (PPI) interface, have been associated with various diseases. Meanwhile, though de novo mutations (DNMs) have been proven important in neuropsychiatric disorders, such as developmental delay (DD), the relationship between PPI interface DMNs and DD has not been well studied. Here we curated developmental delay DNM datasets from the PsyMuKB database and showed that DD patients showed a higher rate and deleteriousness in DNM missense on the PPI interface than sibling control. Next, we identified 302 DD-related PsychiPPIs, defined as PPI harboring a statistically significant number of DNM missenses at their interface, and 42 DD candidate genes from PsychiPPI. We then observed that PsychiPPIs preferentially affected hub proteins in the human protein interactome network. When analyzing DD candidate genes using gene ontology and gene spatio-expression, we found that PsychiPPI genes carrying PPI interface mutations, such as FGFR3 and ALOX5, were enriched in development-related pathways and the development of the neocortex, and cerebellar cortex, suggesting their potential involvement in the etiology of DD. Our results demonstrated that DD patients carried an excess burden of PPI-truncating DNM, which could be used to efficiently search for disease-related genes and mutations in large-scale sequencing studies. In conclusion, our comprehensive study indicated the significant role of PPI interface DNMs in developmental delay pathogenicity.

Background. The molecular mechanisms underlying de novo metastasis of luminal breast cancer (dnMBC) remain largely unknown. Materials & Methods. Newly diagnosed dnMBC patients (grade 2/3, ER+, PR+/-, HER2-), with available core needle biopsy (CNB), collected from the primary tumor, were selected from our clinical-pathological database. Tumors from dnMBC patients were 1:1 pairwise matched (n=32) to tumors from newly diagnosed patients who had no distant metastases at baseline (eBC group). RNA was extracted from 5 x 10µm sections of FFPE CNBs. RNA sequencing was performed using the Illumina platform. Differentially expressed genes (DEG)s were assessed using EdgeR, deconvolution was performed using CIBERSORTx to assess immune cell fractions. Paired Wilcoxon test was used to compare dnMBC and eBC groups, and corrected for false discovery rate (FDR). Results. Many regulatory DEGs were significantly downregulated in dnMBC compared to eBC. Also, immune-related and hypoxia-related signatures were significantly upregulated. Paired Wilcoxon analysis showed that CCL17 and neutrophils fraction were significantly upregulated, whereas the memory B-cell fraction was significantly downregulated in the dnMBC group. Conclusion. Primary luminal tumors of dnMBC patients display significant transcriptomic and immunological differences compared to comparable tumors from eBC patients.

The ability to directly look into genome sequences has opened great opportunities in plant breeding. Yet, the assembly of full-length chromosomes remains one of the most difficult problems in modern genomics. Genetic maps are commonly used in de novo genome assembly and are constructed on the basis of a statistical analysis of the number of recombination rather than physical distance in base pairs. This may affect the accuracy of the ordering and orientation of scaffolds within the chromosome, especially in the region of recombination suppression. Here we report the use of Tyr-FISH for validation of genetic and pseudochromosome maps. For probe design we developed a pipeline that is based on selection of a unique sequence with minimal potential fluorescent background arising from non-specific in situ hybridization. In total of 24 unique genes were located on physical chromosomes 2 and 6. The order of markers has been corrected by integration genetic and cytogenetic maps. Tyr-FISH mapping showed that the order of 23.1% (chromosome 2) and 27.3% (chromosome 6) of the tested genes differed between physical chromosomes and pseudochromosomes. Also the position of the mlh1 gene, which was not on the genetic map, was defined on physical chromosome 2. The results suggest the possibility of Tyr-FISH mapping of any gene that may not be neither on the genetic map nor in the assembly. Hence, Tyr-FISH provides valuable information for the improvement of the genome assembly.

The genus Rhodotorula includes basidiomycetous oleaginous yeast species. R. babjevae can produce compounds of biotechnological interest such as lipids, carotenoids and biosurfactants from low value substrates such as lignocellulose hydrolysate. High-quality genome assemblies are needed to develop genetic tools and to understand fungal evolution and genetics. Here, we combined short- and long-read sequencing to resolve the genomes of two R. babjevae strains, CBS 7808 (type strain) and DBVPG 8058 at chromosomal level. Both genomes have a size of 21 Mbp and a GC content of 68.2%. Allele frequency analysis indicated tetraploidy in both strains. They harbor 21 putative chromosomes with sizes ranging from 0.4 to 2.4 Mb. In both assemblies, the mitochondrial genome was recovered in a single contig, which shared 97% pairwise identity. The pairwise identity between the majority of chromosomes ranges from 82% to 87%. We found indications for strain-specific extrachromosomal endogenous DNA. 7,591 protein-coding genes and 7,607 associated transcripts were annotated in CBS 7808 and 7,481 protein-coding genes and 7,516 associated transcripts in DBVPG 8058. CBS 7808 has accumulated a higher number of tandem duplications than DBVPG 8058. We identified large translocation events between putative chromosomes and a high genetic divergence between the two strains.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder, with a highly variable expression of phenotypes (restricted interest or activity and repetitive behavior in communication and social interactions), genes (mutation), markers (alteration of transcription) and pathways. Loss of function of the CC2D1A gene appears to primarily affect the brain, leading to a range of behavioral problems in humans. In our study published in 2020; we found that the expressions of miR-19a-3p, miR-361-5p, miR-150-5p, miR-3613-3p, miR-126-3p and miR-499a-5p were downregulated in the serum samples of autistic patients and their families. Here, acquired non-Mendelian hereditary character in a genetically defined mouse model of autism (Cc2d1a +/-) correlates with the transcriptional alteration of five miRNAs. We seek to test the hypothesis that miRNA levels are varying by changes in RNA/DNA structure during development, thereby creating transcription alteration. Behavioral tests were conducted on the offspring of Cc2d1a +/- mutant and control mice, such as novel object, social interaction, marble burying and tail suspension behavior. Two RNA fractions were isolated from mouse hippocampal tissues and sperm cells by standard TRIzol extraction: total RNA and fraction of RNA bound to DNA in the form of a DNA/RNA hybrid (R-loop). The expression levels of miR-19a-3p, miR-361-5p, miR-150-5p, miR-126-3p and miR-499a-5p were investigated by quantitative real-time RT-PCR. We report differences in the distribution of five miRNAs in the hippocampus between male and female mice in RNA fractions, particularly in colonies of Cc2d1a +/- mice. Furthermore, the number of miRNAs engaged in the DNA/RNA hybrid fraction is generally higher in the mutant pedigree than in the control group. On the other hand, in sperm both fractions are at lower levels than in controls. R-loops contribute to the physiology and pathology of organisms including human disease. Here we report a variation in five miRNA levels between gender and tissue. Our results suggest that the transcription levels of these five miRNAs are directly regulated by their RNA.

Enzymes reliant on pyridoxal 5'-phosphate (PLP), the metabolically active form of vitamin B6, hold significant importance in both biology and medicine. They facilitate various biochemical reactions, particularly in amino acid and neurotransmitter metabolisms. Vitamin B6 is absorbed by organisms in its non-phosphorylated form and phosphorylated within cells by pyridoxal kinase (PLK) and pyridox-(am)-ine 5'-phosphate oxidase (PNPOx). The flavin mononucleotide dependent PNPOx enzyme converts pyridoxine 5'-phosphate and pyridoxamine 5'-phosphate into PLP. PNPOx is vital for both biosynthesis and salvage pathways in organisms producing B6 vitamers. However, for those depending on vitamin B6 as nutrient, PNPOx participates only in the salvage pathway. Transferring PLP produced by PNPOx to client apo-enzymes is indispensable for their catalytic function, proper folding and targeting to specific organelles. PNPOx activity deficiencies due to inborn errors lead to severe neurological pathologies, particularly neonatal epileptic encephalopathy. PNPOx enzymes from different organisms maintain PLP homeostasis through highly regulated mechanisms, including structural alterations throughout the catalytic cycle and allosteric PLP binding influencing substrate transformation at the active site. Despite shared features, species-specific regulations are evident among PNPOx enzymes.

“De-[Constructing] Growth” is offered as a deeper and more useful conceptualization that avoids the negative connotations of, and resistance to, “degrowth” by decoupling profit from unsustainable consumption, production, and inequality.

In this work we looked for a stable and accurate procedure to perform feature selection in datasets with a much higher number of predictors than individuals, as in Genome-Wide Association Studies. Due to the instability in feature selection when many potential predictors are measured, a variable selection procedure is proposed that combines several replications of shrinkage regression models. A weighted formulation is used to define the final predictors. The procedure is applied to investigate Single Nucleotide Polymorphisms (SNPs) predictors associated to Alzheimer’s disease in the Alzheimer’s disease Neuroimaging Initiative (ADNI) dataset. Two data scenarios are investigated: one considering only the set of SNPs and another with the covariates age, sex, educational level and APOE4 genotype. The SNP rs2075650 and the APOE4 genotype are given as risk factors for AD, which is in line with the literature, and other four new SNPs are pointed, opening new hypotheses for in vivo analyses. These experiments demonstrate the potential of the new method for stable feature selection.

The king ratsnake (Elaphe carinata) of the genus Elaphe is a common large non-venomous snake that is widely distributed in Southeast and East Asia, and is an economically important farmed snake species. As a non-venomous snake, the king snake that is predatory on venomous snakes such as cobras and pit vipers. The immune mechanisms of which has been unclear. Despite their economic and research importance, genomic resources which will benefit studies in toxicology, phylogeography and immunogenetics are lacking. In this study, we use single-tube long fragment read (stLFR) sequencing to display the first complete genome of a King ratsnake from Huangshan City, Anhui province in China. The genome size is 1.56GB with a scaffold N50 of 6.53M, the total length of the genome is approximately 621Mb, and the repeat content is 38.90%. Additionally, we predicted 22,339 protein-coding genes, of which 22,065 had functional annotations. Our genome is a potentially useful addition to those currently available for snakes.

Idiopathic granulomatous mastitis (IGM) is a rare condition characterised by chronic inflammation and granuloma formation in the breast. The aetiology of IGM is unclear. By focusing on the protein-coding regions of the genome, where most disease-related mutations often occur, whole-exome sequencing (WES) is a powerful approach for investigating rare and complex conditions, like IGM. We report WES results on paired blood and tissue samples from eight IGM patients. Samples were processed using standard genomic protocols. Somatic variants were called with two analytical pipelines: nf-core/sarek with Strelka2 and GATK4 with Mutect2. Our WES study of eight patients did not find evidence supporting a clear genetic component. The discrepancies between variant calling algorithms, along with the considerable genetic heterogeneity observed amongst the eight IGM cases, indicate that common genetic drivers are not readily identifiable. With only three genes, CHIT1, CEP170, and CTR9, recurrently altered in multiple cases, the genetic basis of IGM remains uncertain. The absence of validation for somatic variants by Sanger sequencing raises further questions about the role of genetic mutations in the disease. These observations suggest that IGM may be influenced by non-genetic factors, such as environmental triggers or immune system dysregulation, rather than being primarily a genetic disorder.

The study of the currently known >3,000 species of snakes can provide valuable insights into the evolution of their genomes. Deinagkistrodon acutus, also known as Sharp-nosed Pit Viper, one hundred-pacer viper or five-pacer viper, is a venomous snake with significant economic, medicinal and scientific importance. Widely distributed in southeastern China and South-East Asia, D. acutus has been primarily studied for its venom. Here, we employed next-generation sequencing to assemble and annotate a highly continuous genome of D. acutus. The genome size is 1.46 Gb; its scaffold N50 length is 6.21 Mb, the repeat content is 42.81%, and 24,402 functional genes were annotated. This study helps to further understand and utilize D. acutus and its venom at the genetic level.

Clupeiformes, such as sardines and herrings, represent an important share of worldwide fisheries. Among those, the European sardine (Sardina pilchardus, Walbaum 1792) exhibits significant commercial relevance. While the last decade showed a steady and sharp decline in capture levels, recent advances in culture husbandry represent promising research avenues. Yet, the complete absence of genomic resources from sardine imposes a severe bottleneck to understand its physiological and ecological requirements. We generated 69 Gbp of paired-end reads using Illumina HiSeq X Ten and assembled a draft genome assembly with an N50 scaffold length of 25579 bp and BUSCO completeness of 82.1% (Actinopterygii). The estimated size of the genome ranges between 655 and 850 Mb. Additionally, we generated a relatively high-level liver transcriptome. To deliver a proof of principle of the value of this dataset, we established the presence and function of enzymes (elovl2, elovl5 and fads2) that have pivotal roles in the biosynthesis of long chain polyunsaturated fatty acids, essential nutrients particularly abundant in oily fish such as sardines. Our study provides the first omics dataset from a valuable economic marine teleost species, the European sardine, an essential resource for their effective conservation, management and sustainable exploitation.

Prunus tenella is a rare and precious relict plant in China. It is an important genetic resource for almond improvement and an indispensable material in ecological protection and landscaping. However, the research of molecular breeding and genetic evolution has been severely restricted, due to the lack of genome information. In this investigation, we created a chromosome-level genomic pattern of P. tenella, 231Mb in length with a contig N50 of 18.1 Mb by Hi-C techniques and high-accuracy PacBio HiFi sequencing. The present assembly predicted 32088 protein-coding genes, and an examination of the genome assembly indicated that 94.7% among all assembled transcripts were alignable to the genome assembly; most (97.24%) were functionally annotated. By phylogenomic genome comparison, we found that P. tenella is an ancient group that diverged approximately 13.4 million years ago (Mya) from 13 additional closely related species and about 6.5 Mya from the cultivated almond. Collinearity analysis revealed that P. tenella is highly syntenic and has high sequence conservation with almond and peach. However, this species also exhibit many presence/absence variants. Moreover, a large inversion at the 7,588 kb position of chromosome 5 was observed, which may have a significant association with phenotypic traits. Lastly, population genetic structure analysis in eight different populations indicated a high genetic differentiation among the natural distribution of P. tenella. This high-quality genome assembly provides critical clues and comprehensive information for the systematic evolution, genetic characteristics, and functional gene research of P. tenella. Moreover, it provides a valuable genomic resource for in-depth study in protecting, developing, and utilizing P. tenella germplasm resources.

Background: Clinical cases reports with snake accidents show that venom bite induces increased oxidative stress including several markers of lipid peroxidation and other oxidative stress marker in plasma. Methods: The main findings of this work were performed with BthTx-II on paw edema of animals treated with the toxin and biochemical measurement of COX-2, PGE2, MDA and the effects of peroxiredoxin inhibitors on edema and myotoxicity were also evaluated. Results: The results show that edema and myotoxocity induced by PLA2 (BthTx-II) induces a strong mobilization of arachidonic acid and an increase in cellular oxidative stress as measured by increased malondialdehydo (MDA) concentration and protein carbonylation. Thus, these findings establish the strong link between oxidative stress, arachidonic acid mobilization and that these events may explain the presence of oxidative stress markers in snake-bitten patients. Experiments performed with animals previously treated with commercially purchased inhibitors showed enzymes such as thioredoxin (TXN), thioredoxin reductase (TXNRD) and other glutathione (GSH)-related antioxidant defenses could play an essential role controlling and defining the end of edema on the late phase of PLA2 BthTx-II-induced process. Conclusion. This study showed that thioate-dependent antioxidant enzymes play an important role in resolving the edema induced by BthTx-II.

This paper, based on the "Unified Theory of Forces," rigorously proves the formula of de Broglie's wave function. This helps us better understand the following important issues: 1) Theoretically prove and understand the essence of de Broglie's wave function; 2) Prove that when the particle's momentum P is determined, the greater the mass of the particle, the more concentrated its wave function distribution, and the weaker the interference effect; 3) Correct the spatial distribution of the wave function of free particles, proving that the wave function of free particles is strictly constrained within a finite spatial range, which is consistent with the facts we observe.

Summary Genome assembly projects have grown exponentially due to breakthroughs in sequencing technologies and assembly algorithms. Evaluating the quality of genome assemblies is critical to ensure the reliability of downstream analysis and interpretation. To fulfil this task, we have developed the AssemblyQC pipeline that performs file-format validation, contaminant checking, contiguity measurement, gene- and repeat-space completeness quantification, telomere inspection, taxonomic assignment, synteny alignment, scaffold examination through Hi-C contact-map visualisation, and assessments of completeness, consensus quality and phasing through K-mer analysis. It produces a comprehensive HTML report with method descriptions, tables, and visualisations. Availability and Implementation The pipeline uses NextFlow for workflow orchestration and adheres to the best-practice established by the nf-core community. This pipeline offers a reproducible, scalable, and portable method to assess the quality of genome assemblies – the code is available online.GitHub: https://github.com/Plant-Food-Research-Open/assemblyqc Supplementary information Pipeline usage documentation, parameter descriptions and example outputs are available on GitHub: https://github.com/Plant-Food-Research-Open/assemblyqc/tree/main/docs. A preview report is also hosted online: https://plant-food-research-open.github.io/assemblyqc

The Peruvian creole cattle (PCC) is a neglected breed, and is an essential livestock resource in the Andean region of Peru. To develop a modern breeding program and conservation strategies for the PCC, a better understanding of the genetics of this breed is needed. We sequenced the whole genome of the PCC using a paired-end 150 strategy on the Illumina HiSeq 2500 platform, obtaining 320 GB of sequencing data. The obtained genome size of the PCC was 2.77 Gb with a contig N50 of 108Mb and 92.59% complete BUSCOs. Also, we identified 40.22% of repetitive DNA of the genome assembly, of which retroelements occupy 32.39% of the total genome. A total of 19,803 protein-coding genes were annotated in the PCC genome. We downloaded proteomes and genomes of the Bovinae subfamily, and conducted a comparative analysis with our draft genome. Phylogenomic analysis showed that PCC is related to Bos indicus. Also, we identified 7,746 family genes shared among the Bovinae subfamily. This first PCC genome is expected to contribute to a better understanding of its genetics to adapt to the tough conditions of the Andean ecosystem, and evolution.

The strong competition in the automotive industry has required manufacturers to implement lean production, both with methods and techniques specific to Industry 4.0. At the same time, universities must provide graduates with specific skills for applying these new production methods and techniques. In this context, a Lean Learning Factory was developed in the Pitesti University Center, which allows students to learn, experiment, and research, in an environment similar to that of enterprises, the new Lean Manufacturing methods and techniques, and Industry 4.0. The research presented in this study had as its objective identifying the minimum number of repetitions necessary to train operators who serve two differently organized workstations, one classic, the other including digital techniques, but on which the same assembly operation is performed. Several indicators were considered in the analysis, such as the number of errors, the number of stops, the effective duration of the work cycle, and the percentage ratio between the standard duration of cyclical activities and the effective duration of the work cycle. The evolution of these indicators was mathematically modelled by regression functions, using the least squares method. The obtained results also highlight the usefulness of applying the DOJO method as a Lean manufacturing-specific learning technique and the efficiency of implementing digital techniques in work organization.

In the framework of the FireStorm project, four portable weather stations were installed in the Lousã/Estrela mountain range. Given that the Portuguese Institute for Sea and Atmosphere’s sur-face network has two weather stations installed in this region, the new data allows an improved monitoring of the vertical variability of near-surface variables in this mountainous region. As most of the wildfires in mainland Portugal affect areas with complex terrain with elevations below 1200 m and major fires continue to burn overnight, it is also relevant to monitor the vertical changes of meteorological variables in the nighttime period, as these may exhibit large variability. This study provides the first assessment of the available data, with focus on the summer seasons of 2021 and 2022. The results highlight the large variability that was observed in the region and suggest that the risk of extreme fire behaviour in the nighttime period may be underestimated.

RaTG13 (a bat derived SARS-like CoV) is the closest relative sequence of SARS-CoV-2 reported till date. The sample from which RaTG13 was sequenced was a bat fecal swab collected in 2013 from Tongguan, Mojiang, Yunnan province, China. The Illumina based sequence of RaTG13, MN996532.1, was deposited on 27^th Jan 2020 and the raw data (Illumina), https://www.ncbi.nlm.nih.gov/sra/SRX7724752[accn]. There are discrepancies in dates about when the metagenome sequencing of RaTG13 sample was done (2018 or 2020), both stated by the same corresponding author. Comparison of the RNA Seq data of RaTG13 fecal swab to the corresponding data from the bat fecal swabs deposited by the same working group using the same methods indicated that the RaTG13 raw data seemed to be different in various aspects. The fecal swab sample showed abnormally less read of bacterial reads in the swab was exceptionally low, i.e. 0.7%, compared to the 20-90% abundance in other fecal swabs from bats processed by similar methods. Also, another raw data in the form of amplicon sequences was deposited in May 2020; however, the dates mentioned on the files of the sequenced amplicons were older (2017, 2018). The genome assembly of RaTG13 could not be done de-novo and the average coverage of the genome ~8%. Also, literature indicates that RaTG13 RBD cannot bind to Rhinolophus ACE-2 receptors. Collectively, the anomalies in the raw data of RaTG13 and other issues pose an important question about the overall authenticity of the RaTG13 genome sequence.

This comprehensive overview of the historical milestones in cell culture underscores key breakthroughs that have shaped the field over time. It begins with Wilhelm Roux's seminal experiments in the 1880s, followed by the pioneering efforts of Ross Granville Harrison, who initiated groundbreaking experiments that fundamentally shaped the landscape of cell culture in the early 20th century. Carrel's influential contributions, notably the immortalization of chicken heart cells, mark a significant advancement in cell culture techniques. Holtfreter, Moscona, and Leighton introduced innovations such as 3D cell culture, laying the groundwork for further exploration. The advent of induced pluripotent stem cells by Takahashi and Yamanaka in 2006 was revolutionary, enabling the reprogramming of differentiated cells into a pluripotent state. Since then, recent innovations include spheroids, organoids, and organ-on-a-chip technologies, aiming to mimic the structure and function of tissues and organs in vitro, pushing the boundaries of biological modeling and disease understanding. In this review, we overview the history of cell culture shedding light on the main discoveries, pitfalls and hurdles that were overcome during the transition from 2D to 3D cell culture techniques. Finally, we discussed the future directions for cell culture research that may accelerate the development of more effective and personalized treatments.

Brazil chicken production is around 13 million tons and about a third is exported to over 150 countries, placing Brazil as the world largest chicken meat producer, and therefore it is crucial to follow the legislation of all importer markets. This study conducted a survey by chance in 45 meat industries able to export. Therefore, 2580 chicken meat samples were collected and submitted to 11 analyte extraction and chromatographic verification of compliance in an accredited laboratory. Ten chemical residues (amoxicillin, bacitracin, colistin, dinitolmide + zoalene, spectinomycin, roxarsone, tiamulin, tylosin, trenbolone acetate and virginiamycin) were investigated in chicken meat and one (halofuginone hydrobromide) in chicken liver. The results showed that no compound exceeded the maximum residue limits established by seven legislations. All residue concentrations found were below the method quantification limit, thereby confirming the capability of Brazilian chicken meat industries in complying to foreign markets.

Background/Objectives: Direct oral anticoagulants (DOACs) are first-line medications for stroke prevention in non-valvular atrial fibrillation (AF). However, variability in drug response poses risks of hemorrhagic or thromboembolic events. The role of genetics in determining DOAC safety is gaining attention, but evidence linking specific genetic polymorphisms to bleeding risk is lacking. Therefore, the aim of our study was to conduct an Exome-Wide Association Study with 198 non-valvular AF patients treated with rivaroxaban or apixaban, including 97 with bleeding complications and 99 without. Methods: DOAC plasma concentrations, 6-β-hydroxycortisol and cortisol levels in urine were measured for CYP3A4 phenotyping. Sequencing was performed on the DNBSEQ G-400 platform. Single nucleotide variants (SNV) associations with bleeding risk were assessed using logistic regression with additive, dominant, and recessive genetic models. Results: No SNVs reached Bonferroni-corrected significance in any genetic model. Polygenic risk scores (PRS) showed weak predictive ability for bleeding with apixaban. For rivaroxaban, regression indicated that ln Css min/D + 1 index increased with PRS, age, and 6-β-hydroxycortisol/cortisol ratio, but decreased with higher 6-β-hydroxycortisol and coronary heart disease presence. Conclusions: The findings revealed that the residual equilibrium concentration of anticoagulants, including dose-adjusted, cannot alone predict bleeding risk in non-valvular AF patients. No new genetic variants associated with bleeding risk were identified.

Assembly theory provides a promising framework to explain the complexity of systems such as molecular structures and the origins of life, with broad applicability across various disciplines. In this study, we explore and consolidate different aspects of assembly theory by introducing a simplified toy model to simulate the autocatalytic formation of complex structures. This model abstracts the molecular formation process, focusing on the probabilistic control of catalysis rather than the intricate interactions found in organic chemistry. We establish a connection between probabilistic catalysis events and key principles of assembly theory, particularly the probability of a possible construction path in the formation of a complex object, and examine how the assembly of complex objects is impacted by the presence of autocatalysis. Our findings suggest that autocatalysis tends to favor longer consecutive construction sequences in environments with low catalysis probability, while this bias diminishes in environments with higher catalysis probabilities, highlighting the significant influence of environmental factors on the assembly of complex structures.

The final assembly of appliances and automotive industries is a rigorous process, that has limited capabilities of full traceability associated with: (1) the parts installed, (2) their fabrication processes, and (3) the assembly work. This is also the case for each of its sub-assemblies The many parts and sub-assemblies that compose the final assembly, make full traceability a challenging fit, that is almost unsurmountable. Such full traceability along the entire supply-chain is not existent today and must be based on documentation of most assembled parts, assembly tasks, and inspection tasks that compose the full assembled product. In addition, security measures are needed to prevent hostile hacking and unauthorized approach to the assembly documentation throughout the entire supply chain. The related documentation and repeated verifications require considerable effort and have many chances for human errors. So, automating these processes has a great value. This article expounds a framework that harnesses block-chain and smart-contract technology to offer traceable and protected documentation of the assembly process. We expand the concept of a Bill-Of-Assembly (BOA) to incorporate data from the bill of materials (BOM), the associated assembly activities, the associated activities’ specification parameters and materials, and the associated assembly resources (machines and/or operators). The paper defines the operation of the BOA with blockchain and smart-contract technology, for attaining full traceability, safety, and security, for the entire assembled product. Future research could extend the proposed approach to facilitate the usage of the BOA data structure in constructing a digital twin of the entire simulated system.

Background and Objectives: ACTION-France (Awareness, Care and Treatment In Obesity MaNagement in France) aims to identify the perceptions, attitudes, behaviors, and potential barriers to effective obesity management in France and guide collaborative actions. Methods: ACTION-France is a cross-sectional survey of people with obesity (PwO) and healthcare professionals (HCPs), in France. The PwO and HCP questionnaires ran from 27/09/2022 to 01/02/2023 and from 19/12/2022 to 31/03/2023, respectively. Results: The study, encompassing 1226 PwO and 166 HCPs, reveals a shared recognition of obesity as a chronic condition. However, despite being requested by most PwO, weight-related discussions are surprisingly infrequent, leading to delayed diagnosis and care. PwO and HCPs held different views as to why: HCPs often attributed it to PwO’s lack of motivation or disinterest, whereas PwO avoided them because they felt weight managing was their own responsibility and were uncomfortable discussing it. When weight was discussed, primarily with general practitioners (GPs), discussions mostly focused on physical activity and dietary. However, results identified a strong psychosocial impact of obesity: 42% of respondents reported anxiety-depressive symptoms, and many more hesitated to engage in certain social activities because of their weight. Psychotherapy was only discussed by 55% of HCPs. Pharmaceutical options were also rarely discussed (19.5% of HCPs), though 56.1% of PwO reported they would want to. Conclusions: HCPs’ and PwO’s perceptions differed significantly and need to converge through enhanced communication. A holistic approach, integrating comprehensive training for GPs and psychological comorbidities, would help to bridge perceptual gaps effectively and foster more empathetic and effective patient care.

Lasiodiplodia theobromae, as one of the causing agents associated with Chinese hickory trunk cankers, has caused huge economic losses to the Chinese hickory industry due to its extremely strong pathogenicity. Though the biological characteristics of this pathogen and the occurrence pattern of this disease have been reported, few studies have focused on investigating the mechanisms responsible for L. theobromae survival strategies and pathogenicity. The high-quality genome data and the efficient transformation system are the basis for researching above mechanism events. In this study we sequenced and assembled L. theobromae strain LTTK16-3, and established the first protoplasmic preparation method and polyethylene glycol (PEG) -mediated genetic transformation system for L. theobromae. These genetic information and transformation methods established the foundation for the future mechanisms study of L. theobromae and set up the possibility of targeted molecular improvements for Chinese hickory canker control.

Cities are complex systems requiring urban design models that balance order and disorder. Collective creativity initiatives engage citizens in these processes, empowering bottom-up approaches that prioritize people and social well-being within urban development. This paper investigates an 'Urban Laboratory' as a case study, examining the potential of collective creativity to address urban complexity. The successful and ongoing project 'El Campo de Cebada' in Madrid, Spain, demonstrates how a community transformed a vacant lot into a vibrant social hub. The phases of this study include case selection, data collection, data analysis, and presentation of the results. This study identifies key enabling factors, including agents, management, social dynamics, infrastructure, and actions. These insights offer a methodological framework for designing future collaborative, resilient, and inclusive urban spaces, addressing the complex needs of communities within our cities.

Any homeostatic protometabolism would have required orchestration of disparate biochemical pathways into integrated circuits. Extraordinarily specific moieties were also required. Assembly Theory conflated with its cousins—Complexity Theory, Chaos theory, Quantum Mechanics, Irreversible Nonequilibrium Thermodynamics and Molecular Evolution theory— collectively have great naturalistic appeal in hopes of their providing the needed exquisite steering and controls. They collectively offer the best hope of circumventing the need for active selection required to formally orchestrate bona fide formal organization (as opposed to the mere self-ordering of chaos theory) [1]. This paper focuses specifically on Assembly Theory’s contribution to life-origin models.

Nyssa yunnanensis is a deciduous tree species in the family Nyssaceae within the order Cornales. As only eight individual trees and two populations have been recorded in China’s Yunnan province, this species has been listed among China’s national Class I protection species since 1999 and also among 120 PSESP (Plant Species with Extremely Small Populations) in the Implementation Plan of Rescuing and Conserving China’s Plant Species with Extremely Small Populations(PSESP) (2011-2-15). Here, we present the draft genome assembly of N. yunnanensis. Using 10X Genomics linked-reads sequencing data, we carried out the de novo assembly and annotation analysis. The N. yunnanensis genome assembly is 1475 Mb in length, containing 288,519 scaffolds with a scaffold N50 length of 985.59 kb. Within the assembled genome, 799.51 Mb was identified as repetitive elements, accounting for 54.24% of the sequenced genome, and a total of 39,803 protein-coding genes were predicted. With the genomic characteristics of N. yunnanensis available, our study might facilitate future conservation biology studies to help protect this extremely threatened tree species.

Oligopeptide transporters (OPTs) are a group of membrane localized proteins that have a broad range of substrate transport capabilities and contribute to numerous biological processes. However, limited information has been reported on OPTs in higher plants. In this study, a comprehensive analysis of the OPT gene family in Medicago truncatula was performed. A total of 26 OPT genes (MtOPT01-MtOPT26) have been identified in the Medicago truncatula genome. Phylogenetic analyses indicated that MtOPTs consisted of two distinct subgroups, 12 MtOPTs belonged to the peptide transport subgroup (PT-OPT) based on their predicted amino acid sequences containing the two highly conserved motifs (NPG and KIPPR) and 14 MtOPTs belonged to yellow stripe subgroup (YS-OPT). The MtOPTs distributed on each of 8 chromosomes in Medicago truncatula. Sequence analysis verified that MtOPTs significant similar to those in other plants. The copy number of MtOPTs was low and the multiply of MtOPTs was simple relatively. Gene structure analysis showed that most of the MtOPTs have various numbers of introns. The multiple of MtOPTs might play different biological roles which were supported by the fact that MtOPTs have a distinct tissue-specific expression pattern. The data obtained in this study will help to better understand the complexity of the MtOPTs gene family and provide new evidence for the function and evolution of the OPT gene family in higher plants.

The multi-stage, multi-level assembly job shop scheduling problem (MsMlAJSP) is commonly encountered in the manufacturing of complex customized products. Ensuring production efficiency while effectively improving energy utilization is a key focus in the industry. For the energy-efficient MsMlAJSP (EEMsMlAJSP), an improved imperialist competitive algorithm based on Q-learning (IICA-QL) is proposed to minimize the maximum completion time and total energy consumption. In IICA-QL, a decoding strategy with energy-efficient triggers based on problem characteristics is designed to ensure solution quality while effectively enhancing search efficiency. Additionally, an assimilation operation with operator parameter self-adaptation based on Q-learning is devised to overcome the challenge of balancing exploration and exploitation with fixed parameters, thus the convergence and diversity of the algorithmic search is enhanced. Finally, the effectiveness of the energy-efficient strategy decoding trigger mechanism and the operator parameter self-adaptation operation based on Q-learning is demonstrated through experimental results, and the effectiveness of IICA-QL for solving EEMsMlAJSP is verified by comparing with other algorithms.

A study of cryogenic drilling in sandwich composites has been carried out. Materials used have been carbon fiber reinforced polymer sandwich sheets with inner foamed polyvinyl chloride core, composites with aerospace, marine, construction and automotive applications, including protec-tion structure of Polar engineering equipment. Experimental tests have been performed in lam-inates with 12 mm and 6 mm of thickness, drilling with liquid nitrogen as refrigerant to reach temperatures below -120ºC under different cutting conditions, in particular, drill rotation speeds between 2000 rpm and 6000 rpm and feed rates between 200 mm/min and 600 mm/min. Variables as thrust forces and circularity error have been measured, due to the importance of its effect on the hole quality, as an operation prior to a possible assembly. A design of experiments has been allowed determining relationships between variables, identifying the influence of cutting condi-tions on the thrust force and the circularity at inlet and outlet holes.

The utilization of electroporation for delivering the CRISPR/Cas9 system components has enabled efficient gene editing in mammalian zygotes, facilitating the development of genome-edited animals. In our study，This research focused on targeting the ACTG1 and MSTN genes in sheep, revealing a threshold phenomenon in electroporation with a voltage tolerance in sheep in vitro fertilization (IVF) zygotes. Various poring voltages near 40V and pulse durations were examined for electroporating sheep zygotes. The study concluded that stronger electric fields required shorter pulse durations to achieve the optimal conditions for high gene mutation rates and reasonable blastocyst development. The investigation also assessed the quality of Cas9/sgRNA ribonucleoprotein complexes (Cas9 RNPs) and their influence on genome editing efficiency in sheep early embryos. It was highlighted that pre-complexation of Cas9 proteins with single-guide RNA (sgRNA) before electroporation was essential for achieving a high mutation rate. The use of suitable electroporation parameters for sheep IVF zygotes led to significantly high mutation rates and heterozygote ratios. By delivering Cas9 RNPs and single-stranded oligodeoxynucleotides (ssODNs) to zygotes through electroporation, targeting the MSTN (Myostatin) gene, a knock-in efficiency of 26% was achieved. The successful generation of MSTN-modified lambs was demonstrated by delivering Cas9 RNPs into IVF zygotes via electroporation.

An explanation of the origin of dark matter is suggested in this work. The argument is based on symmetry considerations about the concept of mass. In the Wigner's view, the rest mass and the spin of a free elementary particle in flat space-time are the two invariants that characterize the associated unitary irreducible representation of the Poincar\'e group. The Poincar\'e group has two and only two deformations with maximal symmetry. They describe respectively the de Sitter (dS) and Anti de Sitter (AdS) kinematic symmetries. Analogously to their shared flat space-time limit, two invariants, spin and energy scale for de Sitter and rest energy for Anti de Sitter, characterize the unitary irreducible representation associated with dS and AdS elementary systems. While the dS energy scale is a simple deformation of the Poincaré rest energy and so has a purely mass nature, AdS rest energy is the sum of a purely mass component and a kind of zero-point energy derived from the curvature. An analysis based on recent estimates on the chemical freeze-out temperature marking in Early Universe the phase transition quark-gluon plasma epoch to the hadron epoch supports the guess that dark matter energy might originate from an effective AdS curvature energy.

Thorough and precise gene structure annotations are essential for maximizing the benefits of genomic data and unveiling valuable genetic insights. The cucumber genome was first released in 2009 and updated in 2019. To increase the accuracy of predicted gene models, 64 published RNA-seq data and 9 new strand-specific RNA-seq data from multiple tissues were used for manual comparison with the gene models. The updated annotation file (version 2.0) contains increased 24,145 predicted genes from previous version (24317 genes) with a higher BUSCO value of 96.9%. 6231 and 1490 transcripts were adjusted and newly added, respectively, accounting for 31.99% of the overall gene tally. These newly added and adjust genes were renamed (CsaV3.1_XGXXXXX), while genes remaining unaltered preserve their original des ignations. A random selection of 21 modified/added genes were validated using RT-PCR analyses. Additionally, tissue-specific patterns of gene expression were examined using newly obtained transcriptome data with the revised gene prediction model. This improved annotation of cucumber genome will provide essential and accurate resources for studies in cucumber.

Background: Alfalfa, the most economically important forage legume worldwide, features modest genetic progress due to long selection cycles and large extent of non-additive genetic variance associated with its autotetraploid genome. Methods: To improve the efficiency of genomic selection in alfalfa, we explored the effects of genome parametrization (as autotetraploid and diploid dosage, plus allele ratios) and SNP marker subsetting (all available SNPs; only genic regions; only non-genic regions) on genomic regressions, together with various levels of filtering on reading depth and missing rates. We used Genotyping by Sequencing-generated data and focused on traits of different genetic complexity, i.e., dry biomass yield in moisture-favorable (FE) and drought stress (SE) environments, leaf size, and onset of flowering, assessed on 143 genotyped plants from a genetically-broad European reference population and their phenotyped half-sib progenies. Results: On average, allele ratio improved the predictive ability compared with other genome parametrizations (+7.9% vs. tetraploid dosage, +12.6% vs. diploid dosage), SNP subsetting offered an advantage compared with any tested subsettings (+3.7% vs. genic regions, +7.6% vs. non-genic regions). However, when focusing on specific traits, different combinations of genome parametrization and subsetting reached better performances. We also released Legpipe2, a SNP calling pipeline tailored for reduced representation (GBS, RAD) medium-sized genotyping experiments.

This study aimed to determine shifts in microbial populations regarding richness and diversity from the daily use of a popular over-the-counter nasal spray. In addition, the finding of nasal commensal bacterial species that overlap with the oral microbiome may prove to be potential probiotics for the “gateway microbiomes.” Nasal swab samples were obtained before and after using the most popular over-the-counter (OTC) nasal spray in 10 participants aged 18–48. All participants were healthy volunteers with no significant medical histories. The participants were randomly assigned a number by randomizing software and consisted of five men and five women. The sampling consisted of placing a nasal swab atraumatically into the nasal cavity. The samples were preserved and sent to Northwestern University Sequencing Center for whole-genome deep sequencing. After 21 days of OTC nasal spray use twice daily, the participants returned for further nasal microbiome sampling. The microbial analysis included all bacteria, archaea, viruses, molds, and yeasts via deep sequencing for species analysis. The Northwestern University Sequencing Center utilized artificial intelligence analysis to determine shifts in species and strains following nasal spray use that resulted in changes in diversity and richness.

Callus induction (CI) is the critical trait for the transformation of desirable genes in plants. A genome-wide association study (GWAS) analysis was conducted on the rice germplasm of 110 diverse indica rice accessions. Three tissue culture media, including B5, MS, and N6, were used for CI in those rice panels. Seven quantitative trait loci (QTLs) on chromosomes 2, 6, 7, and 11 were found to confer a CI percentage in these three media. For the B5 medium, one QTL (qCI-B5-Chr6) was identified on the rice chromosome 6. For the MS medium, two QTLs were identified on the rice chromosomes 2 and 6 (qCI-MS-Chr2 and qCI-MS-Chr6, respectively). For the N6 medium, four QTLs were identified on the rice chromosomes 6, 7, and 11 (qCI-N6-Chr6.1, qCI-N6-Chr6.2, qCI-N6-Chr7, and qCI-N6-Chr11, respectively). Fifty-five genes were identified within the haplotype blocks corresponding to these QTLs. Thirty-one genes showed haplotypes associated with different CI in those media. The qCI-B5-Chr6 was located in the same region as the qCI-N6-Chr6.2. The caleosin-related family protein was identified in this region. Analysis of the gene-based haplotype reveals the association of this gene with different CI in both B5 and N6 media, suggesting that the gene may play a critical role in the CI mechanism. Moreover, several genes, including the beta-tubulin, the zinc finger proteins, the RNP-1 domain-containing protein, and the Lysophosphatidic acid acyltransferase, were associated with different CI in the N6 media. Results from this study provide insight into the potential QTLs and candidate genes for callus induction in rice that will fulfill the understanding of the physiological and biochemical processes involved in callus formation, which is an essential tool in the molecular breeding of rice.

As a form of protection, a nature park is often created to protect and valorise natural and cultural heritage in peripheral rural areas. However, in terms of multi-functionality, new nature parks are incorporated into traditional productive activities, such as recreational and tourist activities, which sometimes compromise sustainability. The research objective is to study the relationship between tourism and sustainability in the Nature Parks of “Sierra de Aracena y Picos de Aroche”, “Sierra Norte de Sevilla” and “Sierra de Hornachuelos” that makes up the Dehesas de Sierra Morena Biosphere Reserve in Andalusia, Spain. Therefore, selective interviews have been carried out with the stakeholders to establish their perception of sustainable tourism and the presence of dominant discourses. The main conclusions indicate (1) the presence of different dominant discourses on sustainability, namely the conservationist and mercantilist ones, with the prevalence of the economic dimension; (2) poor awareness and adaptation to the context of global change; and (3) the presence of competitive relationships that generate difficulties for the governance of sustainable tourism.

We discuss the consequences of the unique symmetry of the de Sitter spacetime. This symmetry leads to the specific thermodynamic properties of the de Sitter vacuum, which produces the thermal bath for matter. The de Sitter spacetime is invariant under the modified translations, ${\bf r}\rightarrow {\bf r} -e^{Ht}{\bf a}$, where $H$ is the Hubble parameter. For $H\rightarrow 0$, this symmetry corresponds to the conventional invariance of Minkowski spacetime under translations ${\bf r}\rightarrow {\bf r} -{\bf a}$. Due to this symmetry, all the comoving observers at any point of the de Sitter space perceive the de Sitter environment as the thermal bath with temperature $T=H/\pi$, which is twice larger than the Gibbons-Hawking temperature of the cosmological horizon. This temperature does not violate the de Sitter symmetry, and thus does not require the preferred reference frame, as distinct from thermal state of matter, which violates the de Sitter symmetry. This leads to the heat exchange between gravity and matter, and to instability of de Sitter state towards the creation of matter, its further heating, and finally to the decay of the de Sitter state. The temperature $T=H/\pi$ determines different processes in the de Sitter environment, which are not possible in the Minkowski vacuum, such as the process of ionization of an atom in the de Sitter environment. This temperature also determines the local entropy of the de Sitter vacuum state, and this allows us to calculate the total entropy inside the cosmological horizon. The result reproduces the Gibbons-Hawking area law, which is related to the cosmological horizon, $S_{\rm hor}=4\pi KA$, where $K=1/16\pi G$. This supports the holographic properties of the cosmological event horizon. We extend the consideration of the local thermodynamics of the de Sitter state using the $f({\cal R})$ gravity. In this thermodynamics, the Ricci scalar curvature ${\cal R}$ and the effective gravitational coupling, $K$, are thermodynamically conjugate variables. The holographic connection between the bulk entropy of the Hubble volume, and the surface entropy of the cosmological horizon remains the same, but with the gravitational coupling $K=df/d{\cal R}$. Such connection takes place only in the $3+1$ spacetime, where there is the special symmetry due to which the variables $K$ and ${\cal R}$ have the same dimensionality. We also consider the lessons from the de Sitter symmetry for the thermodynamics of black and white holes.

Using assembly theory, we investigate the assembly pathways of binary strings of length N formed by joining bits present in the assembly pool and the strings that entered the pool as a result of previous joining operations. We show that the string assembly index is bounded from below by the shortest addition chain for N, and we conjecture about the form of the upper bound. We define the degree of causation for the minimum assembly index and show that for certain N it features regularities that can be used to determine a shortest addition chain. We show that a string with the smallest assembly index for N can be assembled by a binary program of length equal to this index if the length of this string is expressible as a product of Fibonacci numbers. We conjecture that there is no binary program that has a length shorter than the length of the string having the largest assembly index for N that could assemble this string. The results confirm that four Planck areas provide a minimum information capacity that corresponds to a minimum thermodynamic (Bekenstein-Hawking) entropy. Knowing that the problem of determining the assembly index is at least NP-complete, we conjecture that this problem is NP-complete, while the problem of creating the string so that it would have a predetermined largest assembly index is NP-hard. The proof of this conjecture would imply P ≠ NP, since every computable problem and every computable solution can be encoded as a finite binary string.

Scientists can use the Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) protein 9 system, which is a powerful and flexible tool for changing the genome, to change only certain parts of the genome. This has helped researchers figure out how target genes work in biology and disease. CRISPR/Cas9 is made up of a sequence-specific CRISPR RNA (crRNA) set and a generic Cas9 nuclease. The crRNA set tells Cas9 where to cut DNA and cause double-strand breaks. In the next step of cellular DNA repair, insertions, deletions, and swaps are made at specific spots. For CRISPR/Cas9-mediated DNA cleavage to be specific, the target sequences must match the crRNA, and a protospacer nearby motif must be placed after the target sequences. In this article, we talk about how CRISPR/Cas9 works at the molecular level, what it can be used for, what it can't do, and what its future clinical treatment promise might be.

The vast array of omics data in microbiology presents significant opportunities for studying bacterial pathogenesis and creating computational tools for predicting pathogenic potential. However, the field lacks a comprehensive, curated resource that catalogs bacterial strains and their ability to cause human infections. Current methods for identifying pathogenicity determinants often introduce biases and miss critical aspects of bacterial pathogenesis.In response to this gap, we introduce BacSPaD (Bacterial Strains’ Pathogenicity Database), a thoroughly curated database focusing on pathogenicity annotations for a wide range of high-quality, complete bacterial genomes. Our rule-based annotation workflow combines metadata from trusted sources with automated keyword matching, extensive manual curation, and detailed literature review. Our analysis classified 5,502 genomes as pathogenic to humans (HP) and 490 as non-pathogenic to humans (NHP), encompassing 532 species, 193 genera, and 96 families. Statistical analysis demonstrated a significant but moderate correlation between virulence factors and HP classification, highlighting the complexity of bacterial pathogenicity and the need for ongoing research. This resource is poised to enhance our understanding of bacterial pathogenicity mechanisms and aid in the development of predictive models. To improve accessibility and provide key visualization statistics, we developed a user-friendly web interface, accessible at https://bacspad.altrabio.com.

Hybrid electric vehicles (HEVs) are perceived as a first step towards the future of sustainable transport. Of course, battery electric vehicles (EVs) are currently ideal for what is wanted to stand for transport in the future. The lack of infrastructure for these vehicles makes many potential users choose hybrid vehicles in these conditions. This paper analyzes the influence of the engine and electric motor of hybrid vehicles on their performances. Three engines with slightly different power and close characteristics were considered for configured models. Also, two electric motors with very different power but having a very close maximum continuous torque have been used in models. One is an induction motor, and the other is a permanent magnet synchronous motor. ADVISOR software has been used for vehicles configuration and simulation. Series and parallel hybrid vehicles have been considered. The main dynamic performances and the fuel consumption in two driving cycles were compared for the configured models. Three conventional models with the same engines used in HEVs have also been simulated for reference. The results highlight that in the case of HEVs, the choice of combination engine/electric motor is crucial in obtaining the best report between dynamic performances and low fuel consumption, and implicitly a low negative impact on the environment.

Spontaneous tumours in higher plants can develop depending on plant genotype without any pathogen invasion. Spontaneous tumour formation on the taproots is consistently observed in certain inbred lines of radish (Raphanus sativus var. radicula Pers.). In this work, we have sequenced the genomes of two closely related radish inbred lines that differ in their ability to spontaneously form tumours. We have identified numerous single nucleotide variants (amino acid substitutions, insertions or deletions) that are likely to be associated with the spontaneous tumour formation. Furthermore, we performed the identification of CLE and WOX genes the genomic sequences of the radish inbred lines and, as a result, identified two unique radish CLE genes which probably encode proteins with multiple CLE domains. The results obtained provide a basis for investigating the mechanisms of plant tumour formation and also for future genetic and genomic studies of radish.

The flowering period of rice significantly impacts variety adaptability and yield formation. Properly shortening the reproductive period of rice varieties can expand their ecological range without significant yield reduction. Targeted genome editing, like CRISPR/Cas9, is an ideal tool to fine-tune rice growth stages and boost yield synergistically. In this study, we developed a CRISPR/Cas9-mediated multiplex genome editing vector containing five genes related to three traits: Hd2, Ghd7, and DTH8 (flowering stage genes), along with the recessive rice blast resistance gene Pi21 and the aromatic gene BADH2. This vector was introduced into the high-quality Geng rice variety in Zhejiang province, Jiahe212 (JH212), resulting in 34 T0 plants with various effective mutations. Among the 17 mutant T1 lines, several displayed diverse flowering dates, but most exhibited undesirable agronomic traits. Notably, three homozygous mutant lines (JH-C15, JH-C18, and JH-C31) showed slightly earlier flowering dates without significant differences in yield-related traits compared to JH212. Through special Hyg and Cas marker selection of T2 plants, we identified 7, 6, and 2 fragrant glutinous plants devoid of transgenic components, respectively. These single plants will serve as sib lines of JH212 and potential resources for breeding applications, including maintenance lines for Xian-Geng interspecific three-line hybrid rice. In summary, our research offers practical insights and potential resources for targeted improvement of Geng rice varieties, and would also promote the Geng rice varieties expending from the southward to the northward.

Genetic engineering has become an essential element in developing climate-resilient crops and environmental sustainable solutions to respond to the increasing need for global food security. Genome editing using CRISPR/Cas [Clustered regulatory interspaced short palindromic repeat (CRISPR)-associated protein (Cas)] technology is being applied to a variety of organisms, including plants. This technique has become popular because of its high specificity, effectiveness, and low production cost. Therefore, this technology has the potential to revolutionize agriculture and contribute to global food security. Over the past few years, increasing efforts have been seen in its application in developing higher-yielding, nutrition rich, disease resistant, stress-tolerant “crops” fruits and vegetables. Cas proteins such as Cas9, Cas12, Cas13, and Cas14, among others, have distinct architectures and have been used to create new genetic tools that improve features that are important for agriculture. The versatility of Cas has accelerated genomic analysis and facilitated the use of CRISPR/Cas to manipulate and alter nucleic acid sequences in cells of different organisms. This review provides the evolution of CRISPR technology exploring its mechanisms and contrasting it with traditional breeding and transgenic approaches to improve different stress tolerance. We have also discussed the CRISPR/Cas system and explored three Cas proteins that are currently known to exist: Cas12, Cas13, and Cas14 and their potential to generate foreign DNA free or non-transgenic crops that could be easily regulated for commercialization in most countries.

The snap fit is a common mechanical mechanism. We have studied the spherical snap fit carefully for its physical asymmetry which is easy to assemble but difficult to disassemble, and obtained the general law.Because of the complexity of spherical structure, it is difficult to get a theoretical formula to describe its physical asymmetry.In this paper, through the method of combining theory, simulation and experiment, and based on the general rule of cylindrical snap fit obtained in the early stage, pushing assembly and pulling disassembly of spherical snap fit are studied,we not only get the theoretical formula of spherical snap fit, but also further verify the correctness of previous research conclusions on cylindrical snap fit.This research provides strong theoretical support and basis for the optimization design and further research of snap fit.

Snap fit is a common mechanical mechanism. It uses the physical asymmetry that is easy to assemble but difficult to disassemble to provide a simple and fast link between objects. The ingenious combination of geometric shape, bending elasticity and friction of the snap fit is the mechanism behind the easy to assemble but difficult to disassemble disassemble. Yoshida and Wada (2020) has done a groundbreaking work in the analysis of the elastic snap fit. During our study of their paper, while we really enjoyed their research, unfortunately we detected several questioning formulations. After careful checking, we found that those formulations are not typographical, therefore it is necessary to make corrections. This paper reformulates the linear elasticity of a cylindrical snap fit, obtains an exact solution and proposes an accurate relation between the opening angle and bending tangent angle. Under the first order approximation, our formulations can reduced to the results of Yoshida and Wada and hence confirms the scientific correctness of Yoshida and Wada's work. Furthermore, this paper also derives a correct vertical displacement expression, and propose a new way of disassembly by bending for the first time and formulate a scaling law by data fitting. All formulations are validated by finite element simulation and experiment. The research here is helpful to the design of elastic snap fit or adjustable mechanical mechanism and metamaterial cell.

This paper focuses on the role of the most common mineralogical techniques applied to the identification of the different statuary qualities in white, grey and black Göktepe marble. For this purpose, a Roman case of sculpture marble collection from the rural villa of Valdetorres de Jarama (central Iberia), of the 4th century AD, is presented. The mythological statuary, combining white, grey and black marbles, are one of the most outstanding marble collections in Aphrodisian style found in Hispania. The analytical results (Petrography, Cathodoluminescence, C and O isotopes, and Sr and Mn concentration) support the identification of two varieties of “black” Göktepe, traditionally named as bigio morato and bigo antico, as well as the best statuary quality on white Göktepe. In addition, the analytical identification of other Asiatic marbles in Valdetorres collection, a white coarse-grained originally from the quarries of Aphrodisias city and one small piece identified as Carian red from Iasos, corroborates the already suggested strong connection existing between artists and the stone material they chose for their works. Finally, it is noteworthy the identification carried out on the marble of the bases that served as seats for the sculptures, as it is a white marble of lower quality whose analytical characteristics are consistent with the Microasiatic marble of Denizli. The use of these exotic and exceptional raw materials confirms the taste for luxury and decorative richness in Late Antique Hispanic rural villae and contributes to a better understanding of the distribution of Aphrodisian productions in the Western Roman provinces.