Complete Sequence Research Articles

All-Atom Protein Sequence Design Based on Geometric Deep Learning.

Designing sequences for specific protein backbones is a keystep in creating new functional proteins. Here, we introduceGeoSeqBuilder, a deep learning framework that integratesprotein sequence generation with side chain conformationprediction to produce the complete all-atom structures fordesigned sequences. GeoSeqBuilder uses spatial geometricfeatures from protein backbones and explicitly includesthree-body interactions of neighboring residues. GeoSeqBuilderachieves native residue type recovery rate of 51.6%,comparable to ProteinMPNN and otherleadingmethods,while accurately predicting side chain conformations. Wefirst used GeoSeqBuilder to design sequences for thioredoxinand a hallucinated three-helical bundle protein. All the15 tested sequences expressed as soluble monomeric proteinswith high thermal stability, and the 2 high-resolutioncrystal structures solved closely match the designed models.The generated protein sequences exhibit low similarity(minimum 23%) to the original sequences, with significantlyaltered hydrophobic cores. We further redesigned the hydrophobiccore of glutathione peroxidase 4, and 3 of the 5designs showed improved enzyme activity. Although furthertesting is needed, the high experimental success ratein our testing demonstrates that GeoSeqBuilder is a powerfultool for designing novel sequences for predefined proteinstructures with atomic details. GeoSeqBuilder is availableat https://github.com/PKUliujl/GeoSeqBuilder.

Multimodal Convolutional Neural Network-Based Algorithm for Real-Time Detection and Differentiation of Malignant and Inflammatory Biliary Strictures in Cholangioscopy: a Proof of Concept Study with Video

Deep learning algorithms gained attention for detection (CADe) of biliary tract cancer (BTC) in digital single-operator cholangioscopy (dSOC). We developed a multimodal convolutional neural network (CNN) for detection (CADe) characterization and discriminating (CADx) between malignant, inflammatory and normal biliary tissue in raw dSOC videos. In addition, clinical metadata was included in the CNN algorithm to overcome limitations of image-only models. Based on dSOC videos and images of 111 patients (total of 15,158 still frames), we developed and validated a real-time CNN-based algorithm for CADe and CADx. We established an image-only model and metadata injection approach. In addition, we validated frame-wise and case-based predictions on complete dSOC video sequences. Model embeddings were visualized and class-activation maps highlighted relevant image regions. The concatenation-based CADx approach achieved a per-frame AUC of 0.871, sensitivity of 0.809 (95% CI: [0.784-0.832]), specificity of 0.773 [0.761-0.785], PPV of 0.450 [0.423-0.467], and NPV of 0.946 [0.940-0.954] with respect to malignancy on 5,715 test frames from complete videos of 20 patients. For case-based diagnosis using average prediction scores, six out of eight malignant cases and all twelve benign cases were identified correctly. Our algorithm distinguishes malignant and inflammatory bile duct lesions in dSOC videos, indicating the potential of CNN-based diagnostic support systems for both, CADe and CADx. The integration of non-image data can improve CNN based support systems, targeting current challenges in the assessment of biliary strictures.

A multi-center study on genetic variations in the fusion protein of respiratory syncytial virus from children with Acute Lower Respiratory Tract Infections in China during 2017-2021

Respiratory syncytial virus (RSV) is a significant cause of acute lower respiratory tract infection (ALRTI) in children under five years of age. Between 2017 and 2021, 396 complete sequences of the RSV F gene were obtained from 500 RSV-positive throat swabs collected from ten hospitals across nine provinces in China. In addition, 151 sequences from China were sourced from GenBank and GISAID, making a total of 549 RSV F gene sequences subjected to analysis. Phylogenetic and genetic diversity analyses revealed that the RSV F genes circulating in China from 2017 to 2021 have remained relatively conserved, although some amino acids (AAs) have undergone changes. AA mutations with frequencies ≥ 10% were identified at six sites and the p27 region: V384I (site I), N276S (site II), R213S (site Ø), and K124N (p27) for RSV A; F45L (site I), M152I/L172Q/S173L/I185V/K191R (site V), and R202Q/I206M/Q209R (site Ø) for RSV B. Comparing mutational frequencies in RSV-F before and after 2020 revealed minor changes for RSV A, while the K191R, I206M, and Q209R frequencies increased by over 10% in RSV B. Notably, the nirsevimab-resistant mutation, S211N in RSV B, increased in frequency from 0% to 1.15%. Both representative strains aligned with the predicted RSV-F structures of their respective prototypes exhibited similar conformations, with low root-mean-square deviation values. These results could provide foundational data from China for the development of RSV mAbs and vaccines.

Genomic insights into epidemic plasmids carrying bla CTX-M and mcr-1 genes in Escherichia coli from Lebanese broiler production.

In a previous nationwide survey in the Lebanese broiler production, multidrug-resistant CTX-M-producing E. coli were found to carry the mobile colistin resistance gene mcr-1. To investigate the mobile genetic supports responsible for the spread of these resistance genes among E. coli in healthy broilers in Lebanon. Thirty-three bla CTX-M and mcr-1 positive E. coli of various sequence types from 17 broilers farms were subjected to conjugation assays. Long-read sequencing (Oxford Nanopore Technologies) and hybrid assembly were performed to determine complete plasmid sequences and their phylogenetic diversity. Twenty-nine conjugative IncFII plasmids harboured the extended-spectrum β-lactamase genes bla CTX-M-3 (n = 25) or bla CTX-M-55 (n = 4). Highly related IncF2:A-:B-/bla CTX-M-3 plasmids differing only through IS-mediated genetic rearrangements in antibiotic resistance gene clusters were found in genetically diverse E. coli strains isolated from distant farms. The mobile colistin resistance genes mcr-1.1 and mcr-1.26 were carried by IncX4 and IncI2 plasmids. Worryingly, in one isolate, the ISEcp1-bla CTX-M-55 transposable unit was found integrated in a mcr-1.26-carrying IncX4 plasmid. Beside expanded cephalosporins and colistin resistances, all E. coli isolates were multidrug-resistant with different additional resistances against aminoglycosides, (fluoro)quinolones, fosfomycin, phenicols, sulphonamides, tetracycline and trimethoprim. Closely related blaCTX-M-3/55-borne IncF2:A-:B- plasmids harbouring variable MDR regions and mcr-1 carrying IncX4 plasmids are widely disseminated in the E. coli population of healthy broilers in Lebanon. Further surveillance programmes of antimicrobial resistance and interventions to reduce the abusive use of medically important antibiotics are necessary to limit the spread of resistances in food-producing animals in Lebanon.

Detection and molecular characterization of a novel mitovirus associated with Passiflora edulis Sims.

Mitoviruses are cryptic capsidless viruses belonging to the family Mitoviridae that replicate and are maintained in the mitochondria of fungi. Complete mitovirus-like sequences were recently assembled from plant transcriptome data and plant leaf tissue samples. Passion fruit (Passiflora spp.) is an economically important crop for numerous tropical and subtropical countries worldwide, and many virus-induced diseases impact its production. From a large-scale genomic study targeting viruses infecting Passiflora spp. in Brazil, we detected a de novo-assembled contig with similarity to other plant-associated mitoviruses. The contig is ∼2.6kb long, with a single open reading frame (ORF) encoding an RNA-dependent RNA polymerase (RdRP). This contig has been named "passion fruit mitovirus-like 1" (PfMv1). An alignment of the predicted amino acid sequence of the RdRP of PfMv1 and those of other plant-associated mitoviruses revealed the presence of the six conserved motifs of mitovirus RdRPs. PfMv1 has 79% coverage and 50.14% identity to Humulus lupulus mitovirus 1. Phylogenetic analysis showed that PfMV1 clustered with other plant-associated mitoviruses in the genus Duamitovirus. Using RT-PCR, we detected a PfMv1-derived fragment, but no corresponding DNA was identified, thus excluding the possibility that this is an endogenized viral-like sequence. This is the first evidence of a replicating mitovirus associated with Passiflora edulis, and it should be classified as a member of a new species, for which we propose the name "Duamitovirus passiflorae".

Epidemiology and molecular evolution of GI.1 sapovirus in the recent era.

Sapovirus (SaV) infection is increasing worldwide. Herein, we provided evidence of a significant increase in SaV infection in Japan during 2010-2022, primarily due to the considerable (p = 0.0003) rise of the GI.1 genotype. Furthermore, we found that all major and minor SaV outbreaks in Japan, including the largest SaV outbreak in 2021-2022, were caused by the GI.1 genotype. Therefore, to get insight into the underlying molecular mechanism behind this rising trend of the SaV GI.1 type, we selected 15 SaV GI.1 outbreak strains for complete genome analysis through next-generation sequencing. Phylogenetically, our strains remained clustered in different branches in lineages I and II among the GI.1 genotype. We showed all amino acid (aa) substitutions in different open reading frames (ORFs) in these strains. Importantly, we have demonstrated that the strains involved in the largest SaV outbreak in Japan in 2021-2022 belonged to lineage II and possessed the third ORF. We have identified some unique aa mutations in these major outbreak strains in the NS1 and NS6-NS7 regions that are thought to be associated with viral pathogenicity, cell tropism, and epidemiological competence. Thus, in addition to enriching the database of SaV's complete sequences, this study provides insights into its important mutations.

DANTE and DANTE_LTR: lineage-centric annotation pipelines for long terminal repeat retrotransposons in plant genomes.

Long terminal repeat (LTR) retrotransposons constitute a predominant class of repetitive DNA elements in most plant genomes. With the increasing number of sequenced plant genomes, there is an ongoing demand for computational tools facilitating efficient annotation and classification of LTR retrotransposons in plant genome assemblies. Herein, we introduce DANTE, a computational pipeline for Domain-based ANnotation of Transposable Elements, designed for sensitive detection of these elements via their conserved protein domain sequences. The identified protein domains are subsequently inputted into the DANTE_LTR pipeline to annotate complete element sequences by detecting their structural features, such as LTRs, in adjacent genomic regions. Leveraging domain sequences allows for precise classification of elements into phylogenetic lineages, offering a more granular annotation compared with coarser conventional superfamily-based classification methods. The efficiency and accuracy of this approach were evidenced via annotation of LTR retrotransposons in 93 plant genomes. Results were benchmarked against several established pipelines, showing that DANTE_LTR is capable of identifying significantly more intact LTR retrotransposons. DANTE and DANTE_LTR are provided as user-friendly Galaxy tools accessible via a public server (https://repeatexplorer-elixir.cerit-sc.cz), installable on local Galaxy instances from the Galaxy tool shed or executable from the command line.

PQEB1: a hospital outbreak plasmid lineage carrying bla KPC-2.

While conducting genomic surveillance of carbapenemase-producing Enterobacteriaceae (CPE) from patient colonisation and clinical infections at Birmingham's Queen Elizabeth Hospital (QE), we identified an N-type plasmid lineage, pQEB1, carrying several antibiotic resistance genes, including the carbapenemase gene bla KPC-2. The pQEB1 lineage is concerning due to its conferral of multidrug resistance, its host range and apparent transmissibility, and its potential for acquiring further resistance genes. Representatives of pQEB1 were found in three sequence types (STs) of Citrobacter freundii, two STs of Enterobacter cloacae, and three species of Klebsiella. Hosts of pQEB1 were isolated from 11 different patients who stayed in various wards throughout the hospital complex over a 13 month period from January 2023 to February 2024. At present, the only representatives of the pQEB1 lineage in GenBank were carried by an Enterobacter hormaechei isolated from a blood sample at the QE in 2016 and a Klebsiella pneumoniae isolated from a urine sample at University Hospitals Coventry and Warwickshire (UHCW) in May 2023. The UHCW patient had been treated at the QE. Long-read whole-genome sequencing was performed on Oxford Nanopore R10.4.1 flow cells, facilitating comparison of complete plasmid sequences. We identified structural variants of pQEB1 and defined the molecular events responsible for them. These have included IS26-mediated inversions and acquisitions of multiple insertion sequences and transposons, including carriers of mercury or arsenic resistance genes. We found that a particular inversion variant of pQEB1 was strongly associated with the QE Liver speciality after appearing in November 2023, but was found in different specialities and wards in January/February 2024. That variant has so far been seen in five different bacterial hosts from six patients, consistent with recent and ongoing inter-host and inter-patient transmission of pQEB1 in this hospital setting.

Multiomics analyses reveal high yield-related genes in the hypothalamic-pituitary-ovarian/liver axis of chicken

Egg production, regulated by multiple tissues, is among the most important economic traits in poultry. However, current research only focuses on the hypothalamic-pituitary-ovarian axis, ignoring the most important organ for substance metabolism in the body, the liver. Eggs are rich in lipids, proteins, and other nutrients, which are biosynthesized in the liver. Therefore, here the liver was included in the study of the hypothalamic-pituitary axis. This study used hypothalamus (HH_vs_LH), pituitary (HP_vs_LP), liver (HL_vs_LL), and ovary (HO_vs_LO) tissue samples from high- and low-laying Chengkou mountain chickens (CMCs) for epihistological, transcriptome and metabolomic analyses aimed at improving the reproductive performance of CMC. The results showed that the liver of the high-laying group was yellowish, the cell boundary was clear, and the lipid droplets were evenly distributed. The ovaries of the high-laying group had a complete sequence of hierarchical follicles, which were rich in yolk. In contrast, the ovaries of the low-laying group were atrophic, except for a few small yellow follicles, and numerous primordial follicles that remained. The transcriptome sequences yielded 167.11 Gb of clean data, containing 28,715 genes. Furthermore, 285, 822, 787, and 1,183 differentially expressed genes (DEGs) were identified in HH_vs_LH, HP_vs_LP, HL_vs_LL and HO_vs_LO and the DEGs significantly enriched 77, 163, 170, 171 pathways, respectively. Metabolome sequencing yielded 21,808 peaks containing 4,006 metabolites. The differential metabolite analysis yielded 343 and 682 significantly different metabolites (SDMs) that significantly enriched 136 and 87 pathways in the liver and ovaries, respectively. A combined analysis of the transcriptome and metabolome of the liver and ovaries identified “CYP51A1-4α-carboxy-stigmasta7, 24(24(1))-dien-3β-ol” and “ACSS1B-estrone 3-sulfate” and other multiple gene-metabolite pairs. The DEGs in the hypothalamus and pituitary mainly enriched signaling transduction. In contrast, the DEGs and SDMs in the liver and ovaries mainly enriched the substance metabolism pathways: “gap junction”, “extracellular matrix (ECM)-receptor interaction”, “Steroid biosynthesis”, and “Steroid hormone biosynthesis”. These results suggest that the hypothalamic-pituitary axis may affect egg production mainly by regulating lipid metabolism in the liver and ovaries.

Meta-transcriptomic analysis reveals the geographical expansion of known sugarbeet-infecting viruses and the occurrence of a novel virus in sugarbeet in the United States.

In this study, meta-transcriptome sequencing was conducted on a total of 18 sugarbeet (Beta vulgaris L. subsp. vulgaris) sample libraries to profile the virome of field-grown sugarbeet to identify the occurrence and distribution of known and potentially new viruses from five different states in the United States. Sugarbeet roots with symptoms resembling rhizomania caused by beet necrotic yellow vein virus (BNYVV), or leaves exhibiting leaf-curling, yellowing to browning, or green mosaic were collected from the sugarbeet growing areas of California, Colorado, Idaho, Minnesota, and North Dakota. In silico analysis of de novo assembled contigs revealed the presence of nearly full-length genomes of BNYVV, beet soil-borne virus (BSBV), and beet soil-borne mosaic virus (BSBMV), which represent known sugarbeet-infecting viruses. Among those, BNYVV was widespread across the locations, whereas BSBV was prevalent in Minnesota and Idaho, and BSBMV was only detected in Minnesota. In addition, two recently reported Beta vulgaris satellite virus isoforms (BvSatV-1A and BvSatV-1B) were detected in new locations, indicating the geographical expansion of this known virus. Besides these known sugarbeet-infecting viruses, the bioinformatic analysis identified the widespread occurrence of a new uncharacterized Erysiphe necator-associated abispo virus (En_abispoV), a fungus-related virus that was identified in all 14 libraries. En_abispoV contains two RNA components, and nearly complete sequences of both RNA1 and RNA2 were obtained from RNASeq and were further confirmed by primer-walking RT-PCR and Sanger sequencing. Phylogenetic comparison of En_abispoV isolates obtained in this study showed varying levels of genetic diversity within RNA1 and RNA2 compared to previously reported isolates. The undertaken meta-transcriptomic approach revealed the widespread nature of coexisting viruses associated with field-grown sugarbeet exhibiting virus disease-like symptoms in the United States.

First insight into the whole genome sequence variations in clarithromycin resistant Helicobacter pylori clinical isolates in Russia

Clarithromycin (CLR) is currently a key antibiotic for Helicobacter pylori infection treatment, however, the data on CLR resistance patterns in Russia are missing. Here, we applied WGS-based approach to H. pylori clinical isolates from Russia to comprehensively investigate sequence variation, identify putative markers of CLR resistance and correlate them with phenotypic susceptibility testing. The phenotypic susceptibility of 44 H. pylori isolates (2014–2022) to CLR was determined by disc diffusion method: 23 isolates were CLR-resistant and 21-CLR-susceptible. All isolates were subjected to WGS and submitted to GenBank. Based on complete sequence analysis, we showed that among all sequence variants, the combination of mutations A2146G/A2147G in the 23S rRNA gene is the most reliable for prediction of phenotypic susceptibility. For the first time, the average number of mutations in 106 virulence-associated genes between resistant and susceptible groups were compared. Moreover, this study presents the first WGS insight into genetic diversity of H. pylori in Russia with a particular focus on the molecular basis of drug resistance: the novel mutations were described as potential markers for the resistance development. Of these, the most prominent was a frameshift deletion (252:CGGGT) in HP0820 coding region, which is a good candidate for further investigation.

EDNAqua-Plan—Standardisation Overview for eDNA Sequencing of Aquatic Organisms and the Downstream Data Ecosystem

The eDNAqua-Plan project stands as a beacon of innovation in the biomonitoring of marine and freshwater ecosystems, propelled by the urgent need to integrate DNA-based approaches in aquatic bioassessment and monitoring frameworks. The broad utilisation of cutting-edge environmental DNA (eDNA) and DNA barcoding methodologies is dependent on complete, reliable, and accessible reference DNA sequence data (Rimet et al. 2021). Complete and interoperable metadata is crucial to allow a broad reuse of (e)DNA data and analysis outputs, and for a broader uptake of results by end users. The eDNAqua-Plan project aims to address key limitations to the routine implementation of eDNA-based monitoring methods in Europe by developing plans for federated DNA barcode reference libraries and eDNA data repositories to support DNA-based environmental monitoring. This will ensure a sustainable and reliable infrastructure to underpin its broad use, thereby paving the way for more effective conservation and management strategies. The project is working towards creating a comprehensive overview of standardisation efforts and data workflows, through collaborations with other projects, initiatives and infrastructures for aquatic monitoring across the European Union (EU) and associated countries. We are analysing existing archives (e.g., International Nucleotide Sequence Database Collaboration (INSDC), Barcode of Life Data System (BOLD), Global Biodiversity Information Facility (GBIF), Ocean Biodiversity Information System (OBIS)), portals, and papers to determine current and best practices through the use of questionnaires, manual evaluation of repositories, and machine learning methods (LLMs). This includes an overview of the usage of existing metadata and data standards (e.g., Minimum Information about any (X) Sequence Specifications from the Genomics Standards Consortium (GSC), Darwin Core standard). The results are being integrated by a team of experts in marine and freshwater biomonitoring. With a diverse consortium comprising 18 partner institutions from 11 countries and one international institute, eDNAqua-Plan brings together experts in marine and freshwater monitoring, eDNA analysis, and data science. The collective effort by this consortium will lay the groundwork for the creation of a digital ecosystem of eDNA repositories and an integrated reference library of marine and freshwater species, adhering to FAIR (Findable, Accessible, Interoperable, and Reusable) principles.

Pollen-Food Allergy Syndrome: From Food Avoidance to Deciphering the Potential Cross-Reactivity between Pru p 3 and Ole e 7.

Cross-reactivity between nonspecific lipid transfer proteins could cause anaphylaxis, further influencing food avoidance and nutrient deficiencies. The one affecting olive pollen (Ole e 7) and peach (Pru p 3) may underlie a variety of pollen-food syndromes, though a deep molecular analysis is necessary. Three Ole e 7-monosensitised patients (MON_OLE), three Pru p 3-monosensitised patients (MON_PRU) and three bisensitised patients (BI) were selected. For epitope mapping, both digested proteins were incubated with patient sera, and the captured IgE-bound peptides were characterised by LC-MS. The analysis revealed two Ole e 7 epitopes and the three Pru p 3 epitopes previously described. Interestingly, the "KSALALVGNKV" Ole e 7 peptide was recognised by MON_OLE, BI and MON_PRU patients. Conversely, all patients recognised the "ISASTNCATVK" Pru p 3 peptide. Although complete sequence alignment between both proteins revealed 32.6% identity, local alignment considering seven residue fragments showed 50 and 57% identity when comparing "ISASTNCATVK" with Ole e 7 and "KSALALVGNKV" with Pru p 3. This study mapped sIgE-Ole e 7-binding epitopes, paving the way for more precise diagnostic tools. Assuming non-significant sequence similarity, structural homology and shared key residues may underlie the potential cross-reactivity between Ole e 7 and Pru p 3 nsLTPs.

Amugulang virus, a novel hantavirus harboured by small rodents in Hulunbuir, China

The Hulunbuir region, known for its diverse terrain and rich wildlife, is a hotspot for various natural epidemic diseases. Between 2021 and 2023, we collected 885 wild rodent samples from this area, representing three families, seven genera, and eleven species. Metagenomic analysis identified three complete nucleic acid sequences from the S, M, and L segments of the Hantaviridae family, which were closely related to the Khabarovsk virus. The nucleotide coding sequences for S, M, and L (1392 nt, 3465 nt, and 6491 nt, respectively) exhibited similarities of 82.34%, 81.68%, and 81.94% to known sequences, respectively, while protein-level analysis indicated higher similarities of 94.92%, 94.41%, and 95.87%, respectively. Phylogenetic analysis placed these sequences within the same clade as the Khabarovsk, Puumala, Muju, Hokkaido, Topografov, and Tatenalense viruses, all of which are known to cause febrile diseases in humans. Immunofluorescence detection of nucleic acid-positive rodent kidney samples using sera from patients with hemorrhagic fever and renal syndrome confirmed the presence of viral particles. Based on these findings, we propose that this virus represents a new member of the Hantaviridae family, tentatively named the Amugulang virus, after its primary distribution area.

Deciphering the Plastome and Molecular Identities of Six Medicinal "Doukou" Species.

The genus Amomum includes over 111 species, 6 of which are widely utilized as medicinal plants and have already undergone taxonomic revision. Due to their morphological similarities, the presence of counterfeit and substandard products remains a challenge. Accurate plant identification is, therefore, essential to address these issues. This study utilized 11 newly sequenced samples and extensive NCBI data to perform molecular identification of the six medicinal "Doukou" species. The plastomes of these species exhibited a typical quadripartite structure with a conserved gene content. However, independent variation shifts of the SC/IR boundaries existed between and within species. The comprehensive set of genetic sequences, including ITS, ITS1, ITS2, complete plastomes, matK, rbcL, psbA-trnH, and ycf1, showed varying discrimination of the six "Doukou" species based on both distance and phylogenetic tree methods. Among these, the ITS, ITS1, and complete plastome sequences demonstrated the highest identification success rate (3/6), followed by ycf1 (2/6), and then ITS2, matK, and psbA-trnH (1/6). In contrast, rbcL failed to identify any species. This research established a basis for a reliable molecular identification method for medicinal "Doukou" plants to protect wild plant resources, promote the sustainable use of medicinal plants, and restrict the exploitation of these resources.

The complete mitochondrial genome and microsatellite discovery of the Chilean kelp crab Taliepus dentatus (Brachuyra, Majoidea)

Abstract The complete sequence of the mitochondrial genome and fifteen microsatellite loci for the Chilean kelp crab Taliepus dentatus are reported for the first time. The DNA assembly obtained from an Illumina sequencing revealed a complete mitochondrial genome of 15 603 bp in length. The gene composition and arrangement were similar to that reported for most decapod crabs, containing 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes and a non-coding region. A phylogeny performed with 10 species obtained from GenBank placed T. dentatus close to individuals of the Family Epialtidae. From the 6397 repeated sequences encountered in the data, 15 microsatellite loci primers were designed and five tested for variability in specimens obtained from two sampling sites separated by 300 km of distance. The mean number of alleles per locus was nine (5–15 alleles per locus) and the observed heterozygosities ranged from 0.34 to 0.8. Departure from Hardy–Weinberg equilibrium was observed in two out of the five microsatellites, while none of the loci exhibited significant linkage disequilibrium. This genetic resource will be useful for phylogeny, population genetics and parentage analysis of the commercial species T. dentatus.

Comparative study on the transcriptional activities of grass carp (Ctenopharyngodon idellus) peroxisome proliferator-activated receptors induced by different fatty acids

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that are part of the nuclear hormone receptor family, playing a crucial role in gene expression regulation. They serve as a connection between lipid metabolism disorders and innate immunity by being activated by fatty acids and their derivatives, facilitating signal transduction between the cell surface and nucleus. However, the specific transcriptional effects of different fatty acids (FAs) in fish are not yet fully understood. In our research, we identified and characterized PPARs in grass carp (Ctenopharyngodon idellus). The complete coding sequences of pparαa, pparαb, pparγ, pparδa, and pparδb were 1443 bp, 1404 bp, 1569 bp, 1551 bp, and 1560 bp in length, respectively. Pparα showed the highest expression in the liver, pparγ was mainly expressed in abdominal adipose tissue, and pparδ exhibited increased expression in the heart compared to other tissues. Gene localization analysis revealed that only pparδa was present in both the nucleus and cytoplasm, while the other four genes were exclusively located in the nucleus. Furthermore, our study explored the influence of various fatty acids (docosahexaenoic acid, palmitic acid, lauric acid and oleic acid at concentrations of 0, 50, 100, and 200 μM) on the transcriptional activities of different PPARs, demonstrating the diverse effects of fatty acid ligands on PPAR transcriptional activity. These results have significant implications for understanding the regulation of PPARs transcriptional activity.

A Standardized Pipeline for Assembly and Annotation of African Swine Fever Virus Genome.

Obtaining a complete good-quality sequence and annotation for the long double-stranded DNA genome of the African swine fever virus (ASFV) from next-generation sequencing (NGS) technology has proven difficult, despite the increasing availability of reference genome sequences and the increasing affordability of NGS. A gap analysis conducted by the global African swine fever research alliance (GARA) partners identified that a standardized, automatic pipeline for NGS analysis was urgently needed, particularly for new outbreak strains. Whilst there are several diagnostic and research labs worldwide that collect isolates of the ASFV from outbreaks, many do not have the capability to analyze, annotate, and format NGS data from outbreaks for submission to NCBI, and some publicly available ASFV genomes have missing or incorrect annotations. We developed an automated, standardized pipeline for the analysis of NGS reads that directly provides users with assemblies and annotations formatted for their submission to NCBI. This pipeline is freely available on GitHub and has been tested through the GARA partners by examining two previously sequenced ASFV genomes; this study also aimed to assess the accuracy and limitations of two strategies present within the pipeline: reference-based (Illumina reads) and de novo assembly (Illumina and Nanopore reads) strategies.

Genomic Analysis of a Novel Torradovirus "Rehmannia Torradovirus Virus": Two Distinct Variants Infecting Rehmannia glutinosa.

Rehmannia glutinosa, a crucial medicinal plant native to China, is extensively cultivated across East Asia. We used high-throughput sequencing to identify viruses infecting R. glutinosa with mosaic, leaf yellowing, and necrotic symptoms. A novel Torradovirus, which we tentatively named "Rehmannia torradovirus virus" (ReTV), was identified. The complete sequences were obtained through reverse-transcription polymerase chain reaction (RT-PCR), 5' and 3' rapid amplification of cDNA ends, and Sanger sequencing. The amino acid sequence alignment between the ReTV-52 isolate and known Torradovirus species in the Pro-Pol and coat protein regions were 51.3-73.3% and 37.1-68.1%, respectively. Meanwhile, the amino acid sequence alignment between the ReTV-8 isolate and known Torradovirus species in the Pro-Pol and coat protein regions were 52.7-72.8% and 36.8-67.5%, respectively. The sequence analysis classified ten ReTV strains into two variants. The ReTV-52 genome has two RNA segments of 6939 and 4569 nucleotides, while that of ReTV-8 consists of two RNA segments containing 6889 and 4662 nucleotides. Sequence comparisons and phylogenetic analysis showed ReTV strains clustered within the Torradovirus, exhibiting the closet relation to the squash chlorotic leaf spot virus. The RT-PCR results showed a 100% ReTV detection rate in all 60 R. glutinosa samples. Therefore, ReTV should be classified as a novel Torradovirus species. ReTV is potentially dangerous to R. glutinosa, and necessitating monitoring this virus in the field.

Southern African Origin of HTLV-1 in Romania.

In Europe, most HTLV-1-infected individuals originate from highly endemic regions such as West Indies, sub-Saharan Africa, and South America. The only genuine endemic region for HTLV-1 in Europe is Romania where ATL series have been reported among Romanian patients. Our objective is to better understand the origin of this endemic focus based on a study of the genetic diversity of HTLV-1 in Romanians. DNA was obtained from PBMCs/buffy coats of 11 unrelated HTLV-1-infected individuals of Romanian origin. They include 9 ATL cases and 2 asymptomatic carriers. LTR sequences were obtained for all specimens. Complete genomic HTLV-1 sequences were obtained using four PCR series on 10 specimens. Phylogenetic trees were generated from multiple alignments using HTLV-1 prototypic sequences and the new generated sequences. Most of the complete LTR sequences (756-bp) showed low nucleotide diversity, ranging from 0% to 0.8% difference, and were closely related (less than 0.8% divergence) to the only previously characterized Romanian strain, RKI2. One strain, ROU7, diverged slightly (1.5% on average) from the others. Phylogenetic analyses both on partial LTR and the complete genome demonstrate that the 11 sequences belong to the HTLV-1a cosmopolitan genotype and 10 of them belong to the previously denominated a-TC Mozambique-Southern Africa A subgroup. In this study, we demonstrated that the HTLV-1 present in Romania most probably originated in Southern Africa. As most Romanian HTLV-1 strains are very closely related, we can assume that HTLV-1 has been introduced into the Romanian population recently. Further studies are ongoing to decipher the routes of arrival and dissemination of these HTLV-1 strains, and to date the emergence of this endemic focus in Central Europe.