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The increasing prevalence of carbapenem-resistant Enterobacter cloacae complex (CREC) poses great challenges to infection treatment in the clinical setting. In this study, we reported the emergence of carbapenemase in a rare species, Enterobacter chuandaensis, belonging to the Enterobacter cloacae complex (ECC). We elucidated the genetic characteristics of carbapenem-resistant isolate FAHZZU5885, co-harboring blaNDM−1 and blaKPC−2. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and average nucleotide identity (ANI) analysis were used to identify E. chuandaensis. S1 nuclease pulsed-field gel electrophoresis (S1-PFGE) and Southern blotting were used to clarify the number and size of the plasmids in FAHZZU5885. Antimicrobial phenotypes were identified by antimicrobial susceptibility testing (AST), and the characteristics of the strain were examined with whole-genome sequencing (WGS). The conjugation experiment and stability assay were conducted to verify the transferability and stability of the plasmid carrying carbapenemase-encoding genes. E. chuandaensis FAHZZU5885 was isolated from a perianal swab of a patient admitted to the ICU. This strain simultaneously carried blaNDM−1 and two blaKPC−2 genes. FAHZZU5885 was resistant to most of the tested antibiotics except for amikacin, tigecycline, and colistin. Two blaKPC−2 were located separately on two different plasmids, the ~ 120 kb IncFIA-IncFII plasmid and the ~ 80 kb IncR plasmid. Both plasmids shared the conserved sequence klcA-korC-ISkpn6-blaKPC−2-ISkpn27-tnpR-tnpA. The blaNDM−1-bearing plasmid had the potential to transfer and can remain stable after successive passages. In addition, the blaNDM−1 was carried on a ~ 80 kb IncFII plasmid with the conserved sequence ISAba125-blaNDM−1-ble-trpF-dsbD-cutA-groS-groL. In summary, this study marks the first report of the multidrug-resistant E. chuandaensis strain FAHZZU5885 harboring two blaKPC−2-bearing plasmids, indicating the potential for the further dissemination of carbapenemase-encoding genes in novel species. The findings contribute to enhancing our understanding of CREC strains, emphasizing the need for continued and comprehensive surveillance of this species. [ABSTRACT FROM AUTHOR]

Background: The production of β-lactamases is the most prevalent resistance mechanism for β-lactam antibiotics in Gram-negative bacteria. Presently, over 4900 β-lactamases have been discovered, and they are categorized into hundreds of families. In each enzyme family, amino acid substitutions result in subtle changes to enzyme hydrolysis profiles; in contrast, certain conserved sequences retained by all of the family members can serve as important markers for enzyme family identification. Results: The SHV family was chosen as the study object. First, a unique 10-mer peptide was identified as SHV family's epitope by an approach of protein fingerprint analysis. Then, an SHV-specific magnetic epitope-imprinted gel polymer (MEI-GP) was prepared by an epitope surface imprinting technique, and its sorption behavior and recognition mechanism for template epitope and SHV were both elaborated. Finally, the MEI-GP was successfully applied to selectively extract SHV from bacteria, and the extracted SHV was submitted to MALDI-TOF MS for specific determination. By following this strategy, other β-lactamase families can also be specifically detected. According to the molecular weight displayed in mass spectra, the kind of β-lactamase and its associated hydrolysis profile on β-lactams can be easily identified. Based on this, an initial drug option scheme can be quickly formulated for antimicrobial therapy. From protein extraction to medication guidance reporting, the mean time to detection (MTTD) was less than 2 h, which is much faster than conventional phenotype-based methods (at least 16–20 h) and gene-based techniques (usually about 8 h). Conclusions: This enzyme-specific detection strategy combined the specificity of epitope imprinting with the sensitivity of mass spectrometry, enabling β-lactamase to be selectively extracted from bacteria and clearly presented in mass spectra. Compared with other drug resistance detection methods, this technique has good specificity, high sensitivity (≤ 15 mg of bacteria), a short MTTD (less than 2 h), and simple operation, and therefore has a broad application prospect in clinical medicine. [ABSTRACT FROM AUTHOR]

Introduction: In clinical diagnosis of porcine diseases, co-infection with multiple viruses often leads to similar clinical symptoms. Postweaning multisystemic wasting syndrome (PMWS) can be caused by infections with TTSuV or PCV2, while PCV2, PRV, and PPV can cause respiratory and reproductive disorders in pigs. The overlapping clinical and pathological features of these infections necessitate the development of a rapid and specific method for differentiating and detecting these four DNA viruses. Methods: In this study, four pairs of primers and TaqMan probes were designed targeting the conserved sequence of TTSuV, the Rep gene of PCV2, the gE gene of PRV, and the VP2 gene of PPV. After optimizing reaction conditions, including annealing temperature, primer concentration, and probe concentration, a quadruplex real-time PCR method was developed. Results: This method can specifically detect TTSuV1, PCV2, PRV, and PPV simultaneously, with no cross-reactivity with ASFV, CSFV, PRRSV, PEDV, PSV, and TGEV. The minimum detection limit for each virus was 10 copies/ml, and the inter-assay and intra-assay coefficients of variation ranged from 0.33% to 1.43%. Subsequently, 150 clinical samples were tested to evaluate the practical applicability of this method. The positive rates for TTSuV1, PCV2, PRV, and PPV were 8.6% (13/150), 10.67% (16/150), 14% (21/150), and 11.33% (17/150), respectively. Discussion: The results indicate that the established quadruplex real-time PCR method can assist in the accurate and rapid diagnosis of TTSuV1, PCV2, PRV, and PPV in clinical settings, providing robust support for the prevention and control of these infections. [ABSTRACT FROM AUTHOR]

Genome annotation is an important but challenging task. Accurate identification of short interspersed nuclear elements (SINEs) is particularly difficult due to their lack of highly conserved sequences. AnnoSINE is state-of-the-art software for annotating SINEs in plant genomes, but it is computationally inefficient for large genomes. Moreover, its applicability to animals is limited due to the absence of animal pHMMs in its HMM library. Therefore, we propose AnnoSINE_v2, which extends accurate SINE annotation for animal genomes with greatly optimized computational efficiency. Our results show that AnnoSINE_v2's annotation of SINEs has over 20% higher F1-score compared to the existing tools on animal genomes and enables the processing of complicated genomes, like human and zebrafish, which were beyond the capabilities of AnnoSINE_v1. AnnoSINE_v2 is freely available on Conda and GitHub: https://github.com/liaoherui/AnnoSINE_v2. [ABSTRACT FROM AUTHOR]

DNA transposons are diverse in fish genomes and have been described to generate genomic evolutionary novelties. hAT transposable element data are scarce in Teleostei genomes, making it challenging to conduct comparative genomic studies to understand their neutrality or function. This study aimed to perform a genomic and molecular characterization of hAT copies to assess the diversity of these elements and associate changes in these sequences to genomic and karyotypic novelties in Apareiodon sp. The data revealed that hAT TEs are highly abundant in the Apareiodon sp. genome, with few possibly autonomous copies. Highly conserved sequences with likely functional transposases were observed in nine hAT elements. A great diversity of hAT subgroups was observed, especially from Ac, Charlie, Blackjack, Tip100, hAT6, and hAT5, and a similar wave of hAT genomic invasion was identified in the genome for these six groups of hAT sequences. The data also revealed a distinct number of microsatellites within degenerated hAT copies. hAT sites were demonstrated to be dispersed in the Apareiodon sp. chromosomes and not involved in W chromosome-specific region differentiation. In conclusion, the genomic analysis revealed a great diversity of hAT elements, possible autonomous copies, and differentiation of degenerated transposable elements into tandem sequences. [ABSTRACT FROM AUTHOR]

Pyrone-2,4-dicarboxylic acid (PDC) is a valuable polymer precursor that can be derived from the microbial degradation of lignin. The key enzyme in the microbial production of PDC is 4-carboxy-2-hydroxymuconate-6-semialdehyde (CHMS) dehydrogenase, which acts on the substrate CHMS. We present the crystal structure of CHMS dehydrogenase (PmdC from Comamonas testosteroni) bound to the cofactor NADP, shedding light on its three-dimensional architecture, and revealing residues responsible for binding NADP. Using a combination of structural homology, molecular docking, and quantum chemistry calculations, we have predicted the binding site of CHMS. Key histidine residues in a conserved sequence are identified as crucial for binding the hydroxyl group of CHMS and facilitating dehydrogenation with NADP. Mutating these histidine residues results in a loss of enzyme activity, leading to a proposed model for the enzyme's mechanism. These findings are expected to help guide efforts in protein and metabolic engineering to enhance PDC yields in biological routes to polymer feedstock synthesis. [ABSTRACT FROM AUTHOR]

The water‐lily clade represents the second earliest‐diverging branch of angiosperms. Most of its species belong to Nymphaeaceae, of which the "core Nymphaeaceae"—comprising the genera Euryale, Nymphaea and Victoria—is the most diverse clade. Despite previous molecular phylogenetic studies on the core Nymphaeaceae, various aspects of their evolutionary relationships have remained unresolved. The length‐variable introns and intergenic spacers are known to contain most of the sequence variability within the water‐lily plastomes. Despite the challenges with multiple sequence alignment, any new molecular phylogenetic investigation on the core Nymphaeaceae should focus on these noncoding plastome regions. For example, a new plastid phylogenomic study on the core Nymphaeaceae should generate DNA sequence alignments of all plastid introns and intergenic spacers based on the principle of conserved sequence motifs. In this investigation, we revisit the phylogenetic history of the core Nymphaeaceae by employing such an approach. Specifically, we use a plastid phylogenomic analysis strategy in which all coding and noncoding partitions are separated and then undergo software‐driven DNA sequence alignment, followed by a motif‐based alignment inspection and adjustment. This approach allows us to increase the reliability of the character base compared to the default practice of aligning complete plastomes through software algorithms alone. Our approach produces significantly different phylogenetic tree reconstructions for several of the plastome regions under study. The results of these reconstructions underscore that Nymphaea is paraphyletic in its current circumscription, that each of the five subgenera of Nymphaea is monophyletic, and that the subgenus Nymphaea is sister to all other subgenera of Nymphaea. Our results also clarify many evolutionary relationships within the Nymphaea subgenera Brachyceras, Hydrocallis and Nymphaea. In closing, we discuss whether the phylogenetic reconstructions obtained through our motif‐based alignment adjustments are in line with morphological evidence on water‐lily evolution. [ABSTRACT FROM AUTHOR]

This study offers an in-depth analysis of the mitochondrial genome of Selene dorsalis (Gill 1863), a species native to the Eastern Atlantic Ocean. The circular mitochondrial DNA molecule measures 16,541 base pairs and comprises 13 protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes, two ribosomal RNA genes, and a control region (CR). The nucleotide composition exhibits a notable adenine-thymine (AT) bias, accounting for 53.13%, which aligns with other species in the Carangidae family. Most PCGs initiate with the ATG codon, with the exception of Cytochrome C oxidase subunit I, which starts with GTG. Analysis of relative synonymous codon usage reveals that leucine and serine are the most prevalent amino acids in the mitochondrial genome of S. dorsalis and its congeners (S. vomer and S. setapinnis). All tRNAs display the typical cloverleaf structure, though tRNA Serine (S1) lacks a dihydrouracil arm. Pairwise comparisons of synonymous and nonsynonymous substitutions for all PCGs yielded values below '1', indicating strong purifying selection. The CR spans 847 bp, representing 5.12% of the mitochondrial genome, and is characterized by high AT content (62.81%). It is situated between tRNA-Pro (TGG) and tRNA-Phe (GAA). The CR contains conserved sequence blocks, with CSB-1 being the longest at 22 bp and CSB-D the shortest at 18 bp. Phylogenetic analysis, using Bayesian and Maximum-likelihood trees constructed from concatenated PCGs across 72 species, successfully differentiates S. dorsalis from other carangids. This study also explores how ocean currents and gyres might influence lineage diversification and parapatric speciation of Selene species between the Atlantic and Pacific Oceans. These results highlight the importance of the mitochondrial genome in elucidating the structural organization and evolutionary dynamics of S. dorsalis and its relatives within marine ecosystems. [ABSTRACT FROM AUTHOR]

The evolutionary history of emperors, particularly in the Atlantic and Indo-West Pacific Oceans, remains largely unmapped. This study explores the maternal lineage evolution of Lethrinids by examining the complete mitogenome of Lethrinus atlanticus, which is endemic to the Eastern Atlantic Ocean. Utilizing advanced next-generation sequencing, we found that the mitogenome spans 16,789 base pairs and encompasses 37 genes, including 13 protein-coding genes (PCGs), two ribosomal RNAs, 22 transfer RNAs, and an AT-rich control region (CR). Our analysis indicates a preference for AT base pairs in the L. atlanticus mitogenome (53.10%). Most PCGs begin with the ATG codon, except for COI, which starts with GTG. Relative synonymous codon usage reveals high frequencies for alanine, leucine, proline, serine, and threonine. The ratio of nonsynonymous to synonymous substitutions suggests strong negative selection across all PCGs in Lethrinus species. Most transfer RNAs exhibit typical cloverleaf structures, with the exception of tRNA-serine (GCT), which lacks a dihydrouracil stem. Comparative analysis of conserved sequence blocks across the CRs of three Lethrinus species shows notable differences in length and nucleotide composition. Phylogenetic analysis using concatenated PCGs clearly distinguishes all Lethrinus species, including L. atlanticus, and sheds light on the evolutionary relationships among Spariformes species. The estimated divergence time of approximately 20.67 million years between L. atlanticus and its Indo-West Pacific relatives provides insights into their historical separation and colonization during the late Oligocene. The distribution of Lethrinids may be influenced by ocean currents and ecological factors, potentially leading to their speciation across the Eastern Atlantic and Indo-West Pacific. This study enhances our understanding of the genetic diversity and phylogenetic relationships within Lethrinus species. Further exploration of other emperor fish mitogenomes and comprehensive genomic data could provide vital insights into their genetic makeup, evolutionary history, and environmental adaptability in marine ecosystems globally. [ABSTRACT FROM AUTHOR]

The hedgehog signaling pathway plays an important role in early development and growth of most vertebrates. Sonic hedgehog (shh) gene is a critical regulator of embryonic development in many species, including humans. However, it is not clear what roles shh can play in the development of fish. In this paper, shh gene was cloned from Pseudopleuronectes yokohamae. The full‐length complementary DNA (cDNA) of P. yokohamae sonic hedgehog gene (Pyshh) comprises 3194 bp, with a 1317‐bp open reading frame (ORF) that encodes a polypeptide of 438 amino acids with a typical HH‐signal domain and Hint‐N domain. The conserved sequences of the protein among species were predicted by using multiple sequence comparison. The phylogenetic tree construction showed that PySHH is clustered in a branch of Pleuronectidae. To explore the expression of Pyshh gene in various tissues of P. yokohamae, we used real‐time fluorescence quantitative PCR technology to detect it. The results showed that Pyshh gene is widely distributed in various tissues of P. yokohamae juveniles, different tissues of adult males and females, and is particularly expressed in immune organs. The Pyshh gene expression was higher in the muscle and brain of juvenile fish, and higher in bone, gill, and skin of male fish than that of female fish, suggesting that Pyshh might be involved in the formation of immune organs of P. yokohamae. The expression of Pyshh gene significantly upregulated from the gastrula stage to the hatching stage. Western blotting of the expression levels of PySHH during different embryonic development stages revealed that PySHH levels increased gradually during development stages from oosperm stage to hatching stage. These results indicate that Pyshh is highly conserved among species and plays a critical role in the complex process of embryonic development. Its precise regulation is essential for the proper formation of many organs and tissues in the body, and disruptions in its function may have serious consequences for the formation of immune organs in fish. [ABSTRACT FROM AUTHOR]

Xylophilus ampelinus is a xylematic bacterium causing bacterial blight of grapevine, a disease regarded as a potential threat for viticulture in several countries. Currently, PCR detection is pivotal in diagnostic protocols due to the bacterium's infrequent occurrence in the field and the technical advantages of PCR. Recent metagenomic studies have unveiled diversity in its taxonomic domain, unknown when the most widely used assays for the detection of X. ampelinus infections were developed. In particular, PCR assays relying on highly conserved sequence regions, such as those surrounding ribosomal RNA genes, may be substituted with more specific PCR assays. In this study, we first investigated the diversity of detectable grapevine endophytes related to but different from X. ampelinus and delineated the genotaxonomic boundaries of the species in relation to the known (meta)genomes of closely related bacteria. Then, by exploiting the wealth of genomes now available for bacteria classified in the Burkholderiales, we devised several sets of primers targeting only X. ampelinus and its closest relatives. These primers were employed to (i) genotaxonomically circumscribe the grapevine endophyte species related to but distinguishable from X. ampelinus and (ii) develop a robust multiplex PCR assay expected to be specific for the species X. ampelinus based on in vitro and in silico evidence. The adoption of the multiplex PCR assay presented here is expected to reduce the risk of false positives in the diagnosis of bacterial blight of grapevine. [ABSTRACT FROM AUTHOR]

The emergence of porcine deltacoronavirus (PDCoV) presents a significant threat to both human and animal health due to its ability to cause highly contagious enteric diseases. This underscores the crucial need for timely and accurate diagnosis to facilitate effective epidemiological investigation and clinical management. This research aimed to establish a visual detection method based on reverse transcription loop-mediated isothermal amplification (RT-LAMP) for PDCoV testing. In this study, six pairs of primers were designed according to the conserved sequences of PDCoV ORF1a/b genes. The primer sets and parameters that affect LAMP reaction were optimized. The visual RT-LAMP method was developed by incorporating methyl red into the optimized reaction system, it exclusively detected PDCoV without cross-reactivity with other viruses and the detection limits for PDCoV could reach 10 copies/µL. In comparison with RT-PCR for testing 132 clinical samples, the relative specificity and sensitivity of the visual RT-LAMP were found to be 99.2 and 100%, respectively, with a concordance rate of 99.2% and a kappa value of 0.959, indicating that the visual RT-LAMP is a reliable method for the application of PDCoV detection in clinical samples. [ABSTRACT FROM AUTHOR]

The heart is a multicellular system, and the intercellular crosstalk mechanism is very important for the growth and development of the heart and even the organs, tissues, and cells at a distance. As a kind of extracellular vesicle, exosomes are released by different types of cells and can carry specific genetic material, endosomal proteins, cytokines, etc., which are the main material basis for mediating cell crosstalk mechanism. Among them, microRNA carried by cardiac cells-derived exosomes have highly conserved sequences and play a key role in regulating the function of organs, tissues, and cells related to cardiovascular diseases and their complications and comorbidities, which have attracted extensive attention in the medical community in recent years. Following up on the latest research progress at home and abroad, this review systematically summarized the regulatory role of microRNA from cardiac cells-derived exosomes in various cell crosstalk, including not only cardiac cells (including cardiomyocytes, fibroblasts, myofibroblast, cardiac progenitor cells, cardiac microvascular endothelial cells, cardiosphere-derived cells, etc.) but also tumor cells, bone marrow progenitor cells, and other tissue cells, in order to provide a reference for the prevention and treatment of cardiovascular diseases and their complications and comorbidities. [ABSTRACT FROM AUTHOR]

Petrocodon is a small genus in the family Gesneriaceae, which is special for its remarkable floral diversity, and has high ornamental value. In this study, the complete chloroplast genome sequence and genome characteristics of Petrocodon longitubus are first reported. The genome size is 152,958 bp, including a large single-copy region (LSC, 83,901 bp), a small single-copy region (SSC, 18,255 bp), and two inverted repeat sequences (IRs, 25,401 bp, each). The chloroplast genome of P. longitubus was analyzed, revealing a total GC content of 37.47%. A total of 131 genes were de novo assembled, consisting of 87 protein-coding genes, 36 tRNA genes, and 8 rRNA genes. A comparative analysis was conducted between the chloroplast genome of P. longitubus and three other published species of Petrocodon. The chloroplast genome of four Petrocodon species was found to have a double-chain ring structure, with a size ranging from 152,958 to 153,292 bp. Chloroplast genome size had indistinguishable. Four Petrocodon species was ra elatively conserved sequence, with 87 or 88 protein-coding genes, and 8 rRNA were the most conserved, which contains 42 ~ 50 SSR sites, which are mainly mononucleotides and dinucleotides, 4 boundary transition regions, then trinucleotides, pentanucleotides and hexanucleotides have been not detected. The non-preferred codons of the chloroplast genome in the four Petrocodon species are those ending in A, C, G, or T. The chloroplast genomes of these four Petrocodon species are highly similar to each other and to several Primulina species. Phylogenetic trees indicate that P. longitubus and other Petrocodon species were grouped together in a clade, with P. longitubus form a single clade. The results support the scientific naming of P. Longitubusr based on horticultural traits and further clarify the systematic status using molecular information. [ABSTRACT FROM AUTHOR]

Simple Summary: Krüppel-like factors (KLFs) are essential transcription factors found in many eukaryotes. In this study, the focus is on identifying and analyzing the KLF gene family members in chickens using bioinformatic tools, and comparing them with representative classes such as fish, amphibians, birds, and mammals. The analysis includes the structural characterization, evolutionary relationship assessment, and functional predictions. Additionally, in this study, the impact of miRNA-22 is explored, associated with lipid metabolism, on the expression of KLF genes in the liver, heart, and muscle tissues of Qingyuan partridge chickens. The results indicate that the avian KLF gene family is evolutionarily closer to that of mammals, and all chicken KLFs are non-transmembrane proteins. Moreover, the effect of miRNA-22 on KLF expression varies across different tissues. These findings provide a scientific basis for further research into the functions of KLFs in chickens. Krüppel-like factors (KLFs) are a class of fundamental transcription factors that are widely present in various eukaryotes from nematodes to humans, named after their DNA binding domain which is highly homologous to the Krüppel factor in fruit flies. To investigate the composition, organization, and evolutionary trajectory of KLF gene family members in chickens, in our study, we leveraged conserved sequences of KLF genes from representative classes across fish, amphibians, birds, and mammals as foundational sequences. Bioinformatic tools were employed to perform homology alignment on the chicken genome database, ultimately identifying the KLF family members present in chickens. The gene structure, phylogenetic analysis, conserved base sequences, physicochemical properties, collinearity analysis, and protein structure were then analyzed using bioinformatic tools. Additionally, the impact of miRNA-22, related to poultry lipid metabolism, on the expression of the KLF gene family in the liver, heart, and muscle of Qingyuan partridge chickens was explored. The results showed that: (1) compared to fish, the KLF family in birds is more closely related to mammals and amphibians; (2) KLFs within the same subgroups are likely to be derived from a common ancestral gene duplication; (3) KLF3/8/12 in the same subgroup may have some similar or overlapping functions; (4) the motif 4 of KLF5 was most likely lost during evolution; (5) KLF9 may perform a similar function in chickens and pigs; (6) there are collinear relationships between certain KLF genes, indicating that there are related biomolecular functions between these KLF genes; (7) all members of the KLF family in chickens are non-transmembrane proteins; and (8) interference and overexpression of miRNA-22 in Qingyuan partridge chickens can affect the expression levels of KLF genes in liver, heart, and muscle. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Henan Agricultural Sciences is the property of Editorial Board of Journal of Henan Agricultural Sciences and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Anthocyanins, constituents of flavonoid compounds prevalent in plants, possess significant value in both plant development and human nutrition. The regulation of anthocyanin biosynthesis primarily involves the orchestration of MYB, bHLH, and WD40 transcription factors. Consequently, the bHLH family assumes a pivotal role in modulating plant developmental processes. In the present investigation, a transcription factor, denoted as LvbHLH13, was identified as a positive regulator of anthocyanin pigmentation in lily petals. LvbHLH13 is classified within the IIId subgroup of Arabidopsis bHLH proteins. Functional analyses involving the transient expression and gene silencing of LvbHLH13 revealed its capacity to enhance and diminish anthocyanin accumulation, respectively, by modulating the LvMYB5 expression, thereby influencing the downstream structural gene expression. The overexpression of LvbHLH13 resulted in an increase in the expression of the downstream structural genes related to anthocyanin synthesis, whereas silencing of LvbHLH13 correspondingly decreased the expression. Yeast one-hybrid and EMSA assays demonstrated the interaction between LvbHLH13 and the LvMYB5 promoter, leading to the activation of anthocyanin biosynthesis. A further luciferase (LUC) analysis corroborated the stimulatory effect of LvbHLH13 on the LvMYB5 promoter sequence. Consequently, LvbHLH13 assumed a crucial role in lily-petal pigmentation. A yeast two-hybrid analysis revealed that LvbHLH13 diverged from typical bHLH transcription factor behavior as it did not form a complex with MYB to regulate anthocyanin biosynthesis. This discrepancy could be attributed to the deletion of the N-terminal conserved sequence of LvbHLH13. This study provides a new bHLH candidate and bHLH-MYB partner to explore the anthocyanin regulatory network in further research and provides new opportunities for breeding lilies with various anthocyanin contents. These findings lay a theoretical foundation for subsequent investigations into lily flower coloring mechanisms. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Northwest A & F University - Natural Science Edition is the property of Editorial Department of Journal of Northwest A&F University (Natural Science Edition) and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Small open reading frames (smORFs) shorter than 100 codons are widespread and perform essential roles in microorganisms, where they encode proteins active in several cell functions, including signal pathways, stress response, and antibacterial activities. However, the ecology, distribution and role of small proteins in the global microbiome remain unknown. Here, we construct a global microbial smORFs catalog (GMSC) derived from 63,410 publicly available metagenomes across 75 distinct habitats and 87,920 high-quality isolate genomes. GMSC contains 965 million non-redundant smORFs with comprehensive annotations. We find that archaea harbor more smORFs proportionally than bacteria. We moreover provide a tool called GMSC-mapper to identify and annotate small proteins from microbial (meta)genomes. Overall, this publicly-available resource demonstrates the immense and underexplored diversity of small proteins. Here, the authors built a non-redundant catalogue of nearly 1 billion putative small proteins from the global microbiome as a publicly-available resource, and highlight how some highly prevalent and evolutionarily conserved sequences lack functional annotation. [ABSTRACT FROM AUTHOR]

Avian leukemia virus (ALV) is one of the main pathogens of poultry tumor diseases, and has caused significant economic losses to the poultry industry since its discovery. Therefore, establishing a rapid detection method is essential to effectively prevent and control the spread of ALV. In this study, specific CRISPR RNA (crRNA) and recombinase-aided amplification (RAA) primers with T7 promoter were designed based on the relatively conserved sequence of avian leukemia virus. When crRNA recognized the target sequence, Cas13a protein was activated to cut the reporting probes, and then the detection results were read by using lateral flow dipstick (LFD). The RAA-CRISPR/Cas13a-LFD reaction system was constructed. The RAA amplification time, Cas13a protein concentration, crRNA concentration and CRISPR reaction time were optimized to evaluate the specificity, sensitivity and reproducibility of the system. Finally, RAA-CRISPR/Cas13a-LFD method was compared with Polymerase chain reaction (PCR)-Agarose electrophoresis method and qPCR method in the detection of clinical samples, and the reliability of RAA-CRISPR/Cas13a-LFD method was evaluated. The results showed that the RAA-CRISPR/Cas13a-LFD method could effectively amplify the target gene at 37°C for 40 min, and the test results could be determined by LFD visual observation. The method had good specificity and no cross-reaction with Marek's disease virus (MDV), Fowl adenovirus (FAdV), Infectious bursal disease virus (IBDV), Newcastle disease virus (NDV), Infectious laryngotracheitis virus (ILTV), and Infectious bronchitis virus (IBV). The minimum detection limit of the method was 100 copies/μL, and it had good repeatability and stability. The coincidence rate of clinical detection reached 97.69% and 99.23%. In summary, this study established a simple, efficient, accurate and visualized ALV detection method, which can be used for the prevention and rapid clinical diagnosis of avian leukosis (AL). [ABSTRACT FROM AUTHOR]

Phenylalanine ammonia lyase (PAL) is a widely studied enzyme in plant biology due to its role in connecting primary metabolism to secondary phenylpropanoid metabolism, significantly influencing plant growth, development, and stress response. Although PAL genes have been extensively studied in various plant species but their exploration in cucumber has been limited. This study successfully identified 11 CsPAL genes in Cucumis sativus (cucumber). These CsPAL genes were categorized based on their conserved sequences revealing patterns through MEME analysis and multiple sequence alignment. Interestingly, cis-elements related to stress were found in the promoter regions of CsPAL genes, indicating their involvement in responding to abiotic stress. Furthermore, these gene's promoters contained components associated with light, development and hormone responsiveness. This suggests that they may have roles in hormone developmental processes. MicroRNAs were identified as a key regulators for the CsPAL genes, playing a crucial role in modulating their expression. This discovery underscores the complex regulatory network involved in the plant's response to various stress conditions. The influence of these microRNAs further highlights the complicated mechanisms that plants use to manage stress. Gene expression patterns were analyzed using RNA-seq data. The significant upregulation of CsPAL9 during HT3h (heat stress for 3 h) and the heightened upregulation of both CsPAL9 and CsPAL7 under HT6h (heat stress for 6 h) in the transcriptome study suggest a potential role for these genes in cucumber's tolerance to heat stress. This comprehensive investigation aims to enhance our understanding of the PAL gene family's versatility, offering valuable insights for advancements in cucumber genetics. [ABSTRACT FROM AUTHOR]

Nitrogen (N) is essential for sugar beet (Beta vulgaris L.), a highly N-demanding sugar crop. This study investigated the morphological, subcellular, and microRNA-regulated responses of sugar beet roots to low N (LN) stress (0.5 mmol/L N) to better understand the N perception, uptake, and utilization in this species. The results showed that LN led to decreased dry weight of roots, N accumulation, and N dry matter production efficiency, along with damage to cell walls and membranes and a reduction in organelle numbers (particularly mitochondria). Meanwhile, there was an increase in root length (7.2%) and branch numbers (29.2%) and a decrease in root surface area (6.14%) and root volume (6.23%) in sugar beet after 7 d of LN exposure compared to the control (5 mmol/L N). Transcriptomics analysis was confirmed by qRT-PCR for 6 randomly selected microRNAs, and we identified 22 differentially expressed microRNAs (DEMs) in beet root under LN treatment. They were primarily enriched in functions related to binding (1125), ion binding (641), intracellular (437) and intracellular parts (428), and organelles (350) and associated with starch and sucrose metabolism, tyrosine metabolism, pyrimidine metabolism, amino sugar and nucleotide sugar metabolism, and isoquinoline alkaloid biosynthesis, as indicated by the GO and KEGG analyses. Among them, the upregulated miR156a, with conserved sequences, was identified as a key DEM that potentially targets and regulates squamosa promoter-binding-like proteins (SPLs, 104889216 and 104897537) through the microRNA-mRNA network. Overexpression of miR156a (MIR) promoted root growth in transgenic Arabidopsis, increasing the length, surface area, and volume. In contrast, silencing miR156a (STTM) had the opposite effect. Notably, the fresh root weight decreased by 45.6% in STTM lines, while it increased by 27.4% in MIR lines, compared to the wild type (WT). It can be inferred that microRNAs, especially miR156, play crucial roles in sugar beet root's development and acclimation to LN conditions. They likely facilitate active responses to N deficiency through network regulation, enabling beet roots to take up nutrients from the environment and sustain their vital life processes. [ABSTRACT FROM AUTHOR]

Potato aphids are the most destructive sucking insect pests of potato crops worldwide and cause financial losses to potato crops. Accurate identification is most famous for the implementation of an appropriate control strategy. DNA barcoding is the most accurate and reliable technique for the identification of aphid species. Therefore, an experiment was conducted based on DNA barcoding of aphid species. Aphid specimens were collected from different districts of Punjab, Pakistan. Cytochrome C Oxidase I (CO1) gene primers were used for the amplification of conserved sequence mtDNA of the collected specimens. The results revealed that one species of potato aphid identified as Macrosiphum euphorbiae with 97–99% good query coverage and 97–100% similarity with other potato aphid species present on the NCBI database and the other species identified as Myzus persicae with good query coverage from 95 to 100%. In contrast, the similarity index observed from 86 100% related to that of available on the NCBI database. The phylogenetic studies showed that M. euphorbiae shares the same cluster with M. euphorbiae isolate Shimla_A7, M. euphorbia isolate 10BBCHEM-0973, M. euphorbia, and M. euphorbia voucher. On the other hand, M. persicae shares the same cluster with M. persicae voucher APOM13 and M. persicae voucher APOL13. Aphids are becoming dangerous pests. Therefore, this study will help us to develop species-specific M. euphorbiae and M. persicae control measures. [ABSTRACT FROM AUTHOR]

Sorghum (Sorghum bicolor) is the fifth largest cereal crop in the world and is an important food and feed crop. However, the understanding of the chloroplast genome of sorghum is limited, especially for some landrace varieties. In this study, the complete chloroplast (cp) genomes of 10 sorghum varieties from southwestern China were assembled and analysed in depth. The genome sizes ranged between 140.644 and 140.814 bp; the b3 variety had the smallest genome size, and the b6, b7 and b8 varieties had the largest. Highly conserved sequences were found among the 10 cp genomes. A total of 136 genes, including 88 protein-coding genes, 40 tRNAs and 8 rRNAs, were found in each cp genome. Comparative genomic analysis revealed that the large single-copy (LSC) region had more SNPs and indels than the inverted repeat (IR) and small single-copy (SSC) regions. Furthermore, some genes, including matK, matK-trnk, psbM-petN, rps12-rpl20, and ccsA, exhibited greater sequence variation than others. Furthermore, phylogenetic analysis revealed that b3 had a distant genetic relationship with the other 9 varieties. Our findings provide valuable information for a deeper understanding of the genetic structure of sorghum germplasm resources globally and provide an important foundation for subsequent genetic improvement of sorghum. [ABSTRACT FROM AUTHOR]

Phosphatidylethanolamine-binding proteins (PEBPs) play a crucial role in the growth and development of various organisms. Due to the low sequence similarity compared to plants, humans, and animals, the study of pebp genes in fungi has not received significant attention. The redifferentiation of fruiting bodies is exceedingly rare in fungal development. Hitherto, only a few studies have identified the Capebp2 gene as being associated with this phenomenon in Cyclocybe aegerita. Thus, exploring the role of pebp genes in fruiting body development is imperative. In the present study, four Capebp genes (Capebp1, Capebp3, Capebp4, and Capebp5) were cloned from the AC0007 strain of C. aegerita based on genome sequencing and gene prediction. The findings indicate that the pebp family, in C. aegerita, comprises a total of five genes. Moreover, the sequence similarity was low across the five CAPEBP protein sequences in C. aegerita, and only a few conserved sequences, such as HRY and RHF, were identical. Expression analyses revealed that, similarly to Capebp2, the four Capebp genes exhibit significantly higher expression levels in the fruiting bodies than in the mycelium. Furthermore, overexpressed and RNA interference Capebp1 or Capebp5 transformants were analyzed. The results demonstrate that overexpression of Capebp1 or Capebp5 could induce the regeneration of the lamella or fruiting body, whereas the knockdown of Capebp1 or Capebp5 could lead to the accelerated aging of fruiting bodies. These findings highlight a significant role of Capebp genes in the generation of C. aegerita fruiting bodies and provide a foundation for further exploration into their involvement in basidiomycete growth and development. [ABSTRACT FROM AUTHOR]

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Parallel evolution occurs when distinct lineages with similar ancestral states converge on a new phenotype. Parallel evolution has been well documented at the organ, gene pathway, and amino acid sequence level but in theory, it can also occur at individual nucleotides within noncoding regions. To examine the role of parallel evolution in shaping the biology of mammalian complex traits, we used data on single-nucleotide polymorphisms (SNPs) influencing human intraspecific variation to predict trait values in other species for 11 complex traits. We found that the alleles at SNP positions associated with human intraspecific height and red blood cell (RBC) count variation are associated with interspecific variation in the corresponding traits across mammals. These associations hold for deeper branches of mammalian evolution as well as between strains of collaborative cross mice. While variation in RBC count between primates uses both ancient and more recently evolved genomic regions, we found that only primate-specific elements were correlated with primate body size. We show that the SNP positions driving these signals are flanked by conserved sequences, maintain synteny with target genes, and overlap transcription factor binding sites. This work highlights the potential of conserved but tunable regulatory elements to be reused in parallel to facilitate evolutionary adaptation in mammals. [ABSTRACT FROM AUTHOR]

Stability under high‐temperature environments is crucial for amylase to function in starch‐based industries. This study develops a method of modifying protein surfaces by combining computer‐aided tools including FoldX, PoPMuSiC, Discovery Studio, and I‐Mutant 2.0 with conserved sequence analysis. A truncated α‐amylase ∆AmyPTG from Parageobacillus thermoglucosidasius DSMZ 2542 is rationally designed through this method to improve its thermostability. Seven single‐site variants are constructed and five of them displayed enhanced thermostability. Next, three double‐site variants are constructed with one particularly successful double‐site variant N31RT213R, exhibiting a 4.3‐fold longer half‐life at 80 °C. Notably, the specific activity of N31RT213R reaches 10 567.16 U mg−1, higher than ∆AmyPTG (6645.43 U mg−1). When applied to the corn starch liquefaction reaction, the mutant N31RT213R gets a higher yield of product concentration of about 255.70 µg mL−1, compared to 190.72 µg mL−1 for ∆AmyPTG. Intramolecular forces analysis and surface electrostatic charges analysis are conducted to determine possible causes for the improvement. Also, molecular dynamics simulation is used to analyze the flexibility shifts of the entire protein. This innovative rational engineering approach has proven to be a successful strategy for the selection of hot spots for protein thermostability evolution and has the potential to be applied to other enzymes. [ABSTRACT FROM AUTHOR]

Currently, the main α-amylase family GH13 has been divided into 47 subfamilies in CAZy, with new subfamilies regularly emerging. The present in silico study was performed to highlight the groups, represented by the maltogenic amylase from Thermotoga neapolitana and the α-amylase from Haloarcula japonica, which are worth of creating their own new GH13 subfamilies. This enlarges functional annotation and thus allows more precise prediction of the function of putative proteins. Interestingly, those two share certain sequence features, e.g. the highly conserved cysteine in the second conserved sequence region (CSR-II) directly preceding the catalytic nucleophile, or the well-preserved GQ character of the end of CSR-VII. On the other hand, the two groups bear also specific and highly conserved positions that distinguish them not only from each other but also from representatives of remaining GH13 subfamilies established so far. For the T. neapolitana maltogenic amylase group, it is the stretch of residues at the end of CSR-V highly conserved as L-[DN]. The H. japonica α-amylase group can be characterized by a highly conserved [WY]-[GA] sequence at the end of CSR-II. Other specific sequence features include an almost fully conserved aspartic acid located directly preceding the general acid/base in CSR-III or well-preserved glutamic acid in CSR-IV. The assumption that these two groups represent two mutually related, but simultaneously independent GH13 subfamilies has been supported by phylogenetic analysis as well as by comparison of tertiary structures. The main α-amylase family GH13 has thus been expanded by two novel subfamilies GH13_48 and GH13_49. Key points: • In silico analysis of two groups of family GH13 members with characterized representatives • Identification of certain common, but also some specific sequence features in seven CSRs • Creation of two novel subfamilies—GH13_48 and GH13_49 within the CAZy database [ABSTRACT FROM AUTHOR]

Phenylalanine ammonia lyase (PAL) is a pivotal enzyme bridging primary and secondary phenylpropanoid metabolism, influencing plant growth, development, and stress responses. Despite extensive studies on PAL genes across various plant species, their investigation in barley, a critical staple food globally, has been relatively scarce. In this study, we have successfully identified 10 HvPAL genes, designated as HvPAL genes, in Hordeum vulgare (barley). These HvPAL genes were categorized based on their conserved sequences, which revealed patterns through MEME analysis and multiple sequence alignment. Interestingly, we found cis elements related to stress in the promoter regions of HvPAL genes, indicating their involvement in the response to pathogens. Furthermore, these gene promoters contained components associated with light, development, and hormone responsiveness. This suggests that they may play a role in hormonal developmental processes. MicroRNAs were also identified as regulators of the HvPAL genes we identified highlighting their significance in barley. To further investigate these gene expression patterns, we analyzed the RNA-seq data revealed the upregulating of HvPAL 2, HvPAL3, and HvPAL8, and downregulating HvPAL 5, HvPAL 6, and HvPAL9 genes in this study. This study focused on the regulation of PAL genes in response to 23 different races of Puccinia graminis f. sp. tritici in barley. These results suggest ways to improve traits and develop barley varieties that are resistant to pathogens by selectively increasing the expression of certain HvPAL genes that were not previously regulated. This thorough investigation aims to expand our knowledge of the versatility of the PAL gene family, providing insights for advancements in host -pathogen genetics. [ABSTRACT FROM AUTHOR]

Receptor‐G protein promiscuity is frequently observed in class A G protein‐coupled receptors (GPCRs). In particular, GPCRs can couple with G proteins from different families (Gαs, Gαq/11, Gαi/o, and Gα12/13) or the same family subtypes. The molecular basis underlying the selectivity/promiscuity is not fully revealed. We recently reported the structures of kappa opioid receptor (KOR) in complex with the Gi/o family subtypes [Gαi1, GαoA, Gαz, and Gustducin (Gαg)] determined by cryo‐electron microscopy (cryo‐EM). The structural analysis, in combination with pharmacological studies, provides insights into Gi/o subtype selectivity. Given the conserved sequence identity and activation mechanism between different G protein families, the findings within Gi/o subtypes could be likely extended to other families. Understanding the KOR‐Gi/o or GPCR‐G protein selectivity will facilitate the development of more precise therapeutics targeting a specific G protein subtype. [ABSTRACT FROM AUTHOR]

CABYR is a protein that regulates calcium-binding tyrosine phosphorylation located in sperm flagella, which increases significantly during sperm capacitation and is primarily involved in sperm capacitation and motility. This study aimed to analyze the expression regulation pattern, sequence characteristics and potential biological function of CABYR gene in Banna mini-pig inbred line (BMI). The testes of adult BMI boars were used for RNA-seq; the complete coding sequence of CABYR was cloned; the sequence, structural characteristics, interacting proteins, KEGG and GO of CABYR were analyzed; the ceRNA regulatory network of CABYR was built using RNA-seq data. The average expression level and TPM value of CABYR gene obtained by RNA-seq was 11 187.50 and 225.29, respectively. The full-length CDS of CABYR was 1149 bp (GenBank accession number: OK042309) encoding 382 amino acids (GenBank login number: UYO37316). The amino acid sequence alignment analysis of multiple species revealed that the similarity between BMI and other species exceeded 73%, indicating a relatively conserved sequence. The CABYR protein was found to contain the DD_CABYR_SP17 conserved domain, the results of species phylogenetic tree analysis met the clustering criteria. Other analyses, including protein-protein interaction (PPI) networks, KEGG, and GO, revealed that BMI CABYR interacts with 50 proteins involved in a variety of functions, such as sexual reproduction and reproductive processes. CABYR was involved in 7 GOs, including five cellular components, one molecular functions, and one biological processes, according to functional annotation. Six miRNAs were found to regulate the CABYR gene via targeted interactions. Additionally, 8, 6, and 1 lncRNAs were found to interact with ssc-miR-4331-3p, ssc-miR-744, and ssc-miR-491, respectively, in association with CABYR. This study examines the expression of CABYR in the testis of BMI, as well as its molecular structure characteristics and expression regulatory network. These findings provide a foundation for future research on the involvement of the CABYR gene in spermatogenesis processes in BMI pigs, particularly in relation to crucial biological processes such as sperm motility and fibrous sheath development. [ABSTRACT FROM AUTHOR]

Cyclin-dependent kinase (CDK), the pivotal controller of cell cycle progression, is involved in stress resistance and plant growth. Moso bamboo (Phyllostachys edulis) with surprisingly rapid growth depends on continuous cell division, but the CDK gene family information is still lacking. Here, we performed the systematic identification and analysis of the CDK genes in moso bamboo. 40 PeCDK genes were identified and divided into eight subgroups via phylogeny. The same subgroup possessed extremely conserved sequences and similar gene structure. In addition, evolutionary analysis suggested that the PeCDK gene family may have experienced extensive duplication events primarily 6.85–13.71 million years ago (MYA), with segmental duplication dominating. Furthermore, the multiple promoter cis-elements determined in PeCDK genes indicated their potential functions. The expression patterns revealed that most PeCDK genes participated in responses to drought, salt, ABA and SA signals, cell wall synthesis, organ developmental processes and callus growth. The insights gained provide beneficial reference for clarifying the functions of PeCDKs as well as exploring the growth mechanism in bamboo. [ABSTRACT FROM AUTHOR]

The SLC35 (Solute Carrier 35) family members acting as nucleotide sugar transporters are typically localized in the endoplasmic reticulum or Golgi apparatus. It is, therefore, intriguing that some reports document the presence of orphan transporters SLC35F1 and SLC35F6 within the endosomal and lysosomal system. Here, we compared the subcellular distribution of these proteins and found that they are concentrated in separate compartments; i.e., recycling endosomes for SLC35F1 and lysosomes for SLC35F6. Swapping the C-terminal tail of these proteins resulted in a switch of localization, with SLC35F1 being trafficked to lysosomes while SLC35F6 remained in endosomes. This suggested the presence of specific sorting signals in these C-terminal regions. Using site-directed mutagenesis, fluorescence microscopy, and cell surface biotinylation assays, we found that the EQERLL360 signal located in the cytoplasmic tail of human SLC35F6 is involved in its lysosomal sorting (as previously shown for this conserved sequence in mouse SLC35F6), and that SLC35F1 localization in the recycling pathway depends on two YXXΦ-type signals: a Y367KQF sequence facilitates its internalization from the plasma membrane, while a Y392TSL motif prevents its transport to lysosomes, likely by promoting SLC35F1 recycling to the cell surface. Taken together, these results support that some SLC35 members may function at different levels of the endosomal and lysosomal system. [ABSTRACT FROM AUTHOR]

Lycopene β cyclase (LCY-B) catalyzes the conversion of linear lycopene to cyclic β carotene, a crucial component of the photosynthetic machinery and a source of vitamin A for humans and animals. The gene was amplified in fragments using primers designed from conserved sequences of available nucleotide sequences from NCBI and a sequence of 1494 bp was observed upon amplification of the complete gene. The deduced amino acid sequence had a significant overall similarity with sequences of other Capsicum plants. It was observed that C. frutescens occurred in the same cluster as C. annuum and C. baccatum, while LCYB from all the other plants of the Solanaceae family were placed in one cluster signifying the conserved nature of the protein. The tertiary structure obtained from the protein was validated using the Ramachandran plot and ERRANT scores were determined. The protein function analysis gives insight into important parameters of the protein such as the number of cysteine residues, glycosylation sites, phosphorylation sites, etc. [ABSTRACT FROM AUTHOR]

Nucleotide-binding oligomerization domain containing 2 (NOD2), located in the cell cytoplasm, is a pattern recognition receptor belonging to the innate immune receptor family. It mediates the innate immune response by identifying conserved sequences in bacterial peptide glycans and plays an essential role in maintaining immune system homeostasis. Gene mutations of NOD2 lead to the development of autoimmune diseases such as Crohn's disease and Blau syndrome. Recently, NOD2 has been shown to be associated with the pathogenesis of diabetes, cardiac-cerebral diseases, and cancers. However, the function of NOD2 in these non-communicable diseases (CNCDs) is not well summarized in reviews. Our report mainly discusses the primary function and molecular mechanism of NOD2 as well as its potential clinical significance in CNCDs. [ABSTRACT FROM AUTHOR]

To establish a rapid detection method for norovirus GII.2 genotype, this study employed reverse transcription recombinase polymerase amplification (RT-RPA) combined with CRISPR/Cas12a and lateral flow strip (RT-RPA-Cas12a-LFS). Here, the genome of norovirus GII.2 genotype was compared to identify highly conserved sequences, facilitating the design of RT-RPA primers and crRNA specific to the conserved regions of norovirus GII.2. Subsequently, the reaction parameters of RT-RPA were optimized and evaluated using agar-gel electrophoresis and LFS. The results indicate that the conserved sequences of norovirus GII.2 were successfully amplified through RT-RPA at 37°C for 25 minutes. Additionally, CRISPR/Cas12a-mediated cleavage detection was achieved through LFS at 37°C within 10 minutes using the amplification products as templates. Including the isothermal amplification reaction time, the total time is 35 minutes. The established RT-RPA-Cas12a-LFS method demonstrated specific detection of norovirus GII.2, yielding negative results for other viral genomes, and exhibited an excellent detection limit of 10 copies/μl. The RT-RPA-Cas12a-LFS method was further compared with qRT-PCR by analyzing 60 food-contaminated samples. The positive conformity rate was 100%, the negative conformity rate was 95.45%, and the overall conformity rate reached 98.33%. This detection method for norovirus GII.2 genotype is cost-effective, highly sensitive, specific, and easy to operate, offering a promising technical solution for field-based detection of the norovirus GII.2 genotype. [ABSTRACT FROM AUTHOR]

Background: Due to their resistance and difficulty in treatment, biofilm-associated infections are problematic among hospitalized patients globally and account for 60% of all bacterial infections in humans. Antibiofilm peptides have recently emerged as an alternative treatment since they can be effectively designed and exert a different mode of biofilm inhibition and eradication. Methods: A novel antibiofilm peptide, BiF, was designed from the conserved sequence of 18 α-helical antibiofilm peptides by template-assisted technique and its activity was improved by hybridization with a lipid binding motif (KILRR). Novel antibiofilm peptide derivatives were modified by substituting hydrophobic amino acids at positions 5 or 7, and both, with positively charged lysines (L5K, L7K). These peptide derivatives were tested for antibiofilm and antimicrobial activities against biofilm-forming Staphylococcus epidermidis and multiple other microbes using crystal violet and broth microdilution assays, respectively. To assess their impact on mammalian cells, the toxicity of peptides was determined through hemolysis and cytotoxicity assays. The stability of candidate peptide, BiF2_5K7K, was assessed in human serum and its secondary structure in bacterial membrane-like environments was analyzed using circular dichroism. The action of BiF2_5K7K on planktonic S. epidermidis and its effect on biofilm cell viability were assessed via viable counting assays. Its biofilm inhibition mechanism was investigated through confocal laser scanning microscopy and transcription analysis. Additionally, its ability to eradicate mature biofilms was examined using colony counting. Finally, a preliminary evaluation involved coating a catheter with BiF2_5K7K to assess its preventive efficacy against S. epidermidis biofilm formation on the catheter and its surrounding area. Results: BiF2_5K7K, the modified antibiofilm peptide, exhibited dose-dependent antibiofilm activity against S. epidermidis. It inhibited biofilm formation at subinhibitory concentrations by altering S. epidermidis extracellular polysaccharide production and quorum-sensing gene expression. Additionally, it exhibited broad-spectrum antimicrobial activity and no significant hemolysis or toxicity against mammalian cell lines was observed. Its activity is retained when exposed to human serum. In bacterial membrane-like environments, this peptide formed an α-helix amphipathic structure. Within 4 h, a reduction in the number of S. epidermidis colonies was observed, demonstrating the fast action of this peptide. As a preliminary test, a BiF2_5K7K-coated catheter was able to prevent the development of S. epidermidis biofilm both on the catheter surface and in its surrounding area. Conclusions: Due to the safety and effectiveness of BiF2_5K7K, we suggest that this peptide be further developed to combat biofilm infections, particularly those of biofilm-forming S. epidermidis. [ABSTRACT FROM AUTHOR]

Tomato spotted wilt virus (TSWV) and related thrips-borne orthotospoviruses are a threat to food and ornamental crops. Orthotospoviruses have the capacity for rapid genetic change by genome segment reassortment and mutation. Genetic resistance is one of the most effective strategies for managing orthotospoviruses, but there are multiple examples of resistance gene breakdown. Our goal was to develop effective multigenic, broad-spectrum resistance to TSWV and other orthotospoviruses. The most conserved sequences for each open reading frame (ORF) of the TSWV genome were identified, and comparison with other orthotospoviruses revealed sequence conservation within virus clades; some overlapped with domains with well-documented biological functions. We made six hairpin constructs, each of which incorporated sequences matching portions of all five ORFs. Tomato plants expressing the hairpin transgene were challenged with TSWV by thrips and leaf-rub inoculation, and four constructs provided strong protection against TSWV in foliage and fruit. To determine if the hairpin constructs provided protection against other emerging orthotospoviruses, we challenged the plants with tomato chlorotic spot virus and resistance-breaking TSWV and found that the same constructs also provided resistance to these related viruses. Antiviral hairpin constructs are an effective way to protect plants from multiple orthotospoviruses and are an important strategy in the fight against resistance-breaking TSWV and emerging viruses. Targeting of all five viral ORFs is expected to increase the durability of resistance, and combining them with other resistance genes could further extend the utility of this disease control strategy. [ABSTRACT FROM AUTHOR]

Osmolytes are produced by various microorganisms as a defense mechanism to protect cells and macromolecules from damage caused by external stresses in harsh environments. Due to their useful stabilizing properties, these molecules are applied as active ingredients in a wide range of cosmetics and healthcare products. The metabolic pathways and biocatalytic syntheses of glycosidic osmolytes such as 2-O-α-d-glucosyl-d-glycerate often involve the action of a glycoside phosphorylase. Here, we report the discovery of a glucosylglycerate phosphorylase from carbohydrate-active enzyme family GH13 that is also active on sucrose, which contrasts the strict specificity of known glucosylglycerate phosphorylases that can only use α-d-glucose 1-phosphate as glycosyl donor in transglycosylation reactions. The novel enzyme can be distinguished from other phosphorylases from the same family by the presence of an atypical conserved sequence motif at specificity-determining positions in the active site. The promiscuity of the sucrose-active glucosylglycerate phosphorylase can be exploited for the high-yielding and rapid synthesis of 2-O-α-d-glucosyl-d-glycerate from sucrose and d-glycerate. Key points: • A Xylanimonas protaetiae glycoside phosphorylase can use bothd-glycerate and fructose as glucosyl acceptor with high catalytic efficiency • Biocatalytic synthesis of the osmolyte 2-O-α-d-glucosyl-d-glycerate • Positions in the active site of GH13 phosphorylases act as convenient specificity fingerprints [ABSTRACT FROM AUTHOR]

Thermotoga maritima is a hypertherinophilic, anaerobic organism. As a member of the order Thermo - togates, T maritima carry a unique ability as only fermentative bacterium to produce hydrogen as clean energy. We noticed a hypothetical protein TM] 070 [Protein Data Bank, PDB 1NC71. This protein is characterized as a member of DUE domain of unknown function. Interestingly. this sequence length of 138 amino acids matches to only known structure of cognate transducer in Anabaena PCC 7120 [2Ii'7-21I9 etc]. The scaffold of both transducer and TM 1070 protein is tetrameric. We analyzed 114 highly conserved sequence two these two proteins. The detailed structural analysis of all three space groups (P4, (2 and P21. 21,21-large crystal) transducer forms the same tightly packed ten·ainer with C4 symmetry. In the P4 space group, this tetramer is formed by crystallographic symmetry, while in the other two space groups the tetramer syinmetry is non-crystallographic. The same tetramer was observed in the only structural homolog of transducer iii the PDB, protein structure of 1 NC7. This 1 14 residue protein possesses significant sequence identity with transducer. In this project we have focused on motif structural insight using PyMol, AxPyMol and NovaFold A I. Interestingly, we found well-coordinated divalent magnesium present in the core of tetrameric assesmbly of INC7 with polar pocket [Thr amino acid]. It is likely that magnesium is crucial for phosphory] transfer, as found in transducer leading to destabilization of tetramer as part of signaling state. Our detailed motif characterization reveal a novel ligand binding to this DUF tetramer. [ABSTRACT FROM AUTHOR]

Most coronavirus vaccines focus on the spike (S) antigen, but the frequent mutations in S raise concerns about the vaccine efficacy against new variants. Although additional antigens with conserved sequences are have been tested, the extent to which these vaccines can provide immunity against different coronavirus species remains unclear. In this study, we assessed the potential of nucleocapsid (N) as a coronavirus vaccine antigen. Immunization with MERS-CoV N induced robust immune responses, providing significant protection against MERS-CoV. Notably, MERS-CoV N elicited cross-reactive T cell responses to SARS-CoV-2 N and significantly reduced lung inflammation following a SARS-CoV-2 challenge in the transient hACE2 mouse model. However, in K18-hACE transgenic mice, the vaccine showed limited protection. Collectively, our findings suggest that coronavirus N can be an effective vaccine antigen against homologous viruses, but its efficacy may vary across different coronaviruses, highlighting the need for further research on pan-coronavirus vaccines using conserved antigens. • DNA vaccines expressing MERS-CoV Nucleocapsid protein induces MERS-CoV-specific antibody and T cell responses. • MERS-CoV N provide protection against lethal MERS-CoV infection. • MERS-CoV N also induces immune responses that cross-react with the other β-coronaviruses such as SARS-CoV and SARS-CoV-2. • MERS-CoV N-based vaccine offers partial or limited protection against SARS-CoV-2 infection depending on host susceptibility. [ABSTRACT FROM AUTHOR]

Viral spike proteins mutate frequently, but conserved features within these proteins often have functional importance and can inform development of anti-viral therapies which circumvent the effects of viral sequence mutations. Through analysis of large numbers of viral spike protein sequences from several viral families, we found highly (>99%) conserved patterns within their intracellular domains. The patterns generally consist of one or more basic amino acids (arginine or lysine) adjacent to a cysteine, many of which are known to undergo acylation. These patterns were not enriched in cellular proteins in general. Molecular dynamics simulations show direct electrostatic and hydrophobic interactions between these conserved residues in hemagglutinin (HA) from influenza A and B and the phosphoinositide PIP2. Super-resolution microscopy shows nanoscale colocalization of PIP2 and several of the same viral proteins. We propose the hypothesis that these conserved viral spike protein features can interact with phosphoinositides such as PIP2. • Conservation in viral protein sequences often implies functional importance. • We found intracellular domains of viral spike proteins almost always contain a basic residue close to a cysteine or other potentially acylated residue. • Molecular dynamics simulations show direct interaction between these features and PIP2. • Super-resolution microscopy shows quantitative enrichment of PIP2 near viral spike proteins expressed in cells. • We hypothesize these viral spike protein features can interact with phosphoinositides. [ABSTRACT FROM AUTHOR]

[Display omitted] • mtDNA from hares has both short (SR) and long (LR) tandem repeat elements. • SRs in most hares consist of two randomly alternating 10 bp sequence motifs. • LR copy number varies between individuals but has a species-specific median value. • LR number can be altered by mitochondrial transfer in cultured hare cells. The non-coding regions of the mitochondrial DNAs (mtDNAs) of hares, rabbits, and pikas (Lagomorpha) contain short (∼20 bp) and long (130–160 bp) tandem repeats, absent in related mammalian orders. In the presented study, we provide in-depth analysis for mountain hare (Lepus timidus) and brown hare (L. europaeus) mtDNA non-coding regions, together with a species- and population-level analysis of tandem repeat variation. Mountain hare short tandem repeats (SRs) as well as other analyzed hare species consist of two conserved 10 bp motifs, with only brown hares exhibiting a single, more variable motif. Long tandem repeats (LRs) also differ in sequence and copy number between species. Mountain hares have four to seven LRs, median value five, while brown hares exhibit five to nine LRs, median value six. Interestingly, introgressed mountain hare mtDNA in brown hares obtained an intermediate LR length distribution, with median copy number being the same as with conspecific brown hare mtDNA. In contrast, transfer of brown hare mtDNA into cultured mtDNA-less mountain hare cells maintained the original LR number, whereas the reciprocal transfer caused copy number instability, suggesting that cellular environment rather than the nuclear genomic background plays a role in the LR maintenance. Due to their dynamic nature and separation from other known conserved sequence elements on the non-coding region of hare mitochondrial genomes, the tandem repeat elements likely to represent signatures of ancient genetic rearrangements. clarifying the nature and dynamics of these rearrangements may shed light on the possible role of NCR repeated elements in mitochondria and in species evolution. [ABSTRACT FROM AUTHOR]

Facioscapulohumeral muscular dystrophy type 1 (FSHD1), the second most common myopathy affecting adults, is linked to a contraction to less than 11 Repeated Units (RU) of D4Z4 macrosatellite at 4q35 locus and associated with the 4qA161 permissive haplotype. To date, FSHD1 has no cure and pioneer studies showed that the absence of the 4q telomeric region in phenotypically normal cases indicates that haploinsufficiency of the region containing D4Z4 RU does not cause FSHD. In this study, we applied a CRISPR/CasX strategy to ablate the D4Z4 region as a therapeutic strategy for FSHD1 after characterization of an isogenic human induced Pluripotent Stem Cell (hiPSC) line obtained from iStem. We firstly determined the haplotype on the 4q35 alleles of interest of the hiPSC line and highlighted the presence of the permissive 4qA161 haplotype. To discriminate and quantify the D4Z4 RU in the 4q35/10q26 alleles, a double restriction enzyme digestion (EcoRI ± BlnI) coupled to a Southern Blot was performed and validated by Molecular Combing. We confirmed that the hiPSC line contains one 4qA allele, one 4qB allele, and two 10qA alleles with respectively 30, 27, 19 and 12 D4Z4 RU, thus recapitulating the human genetic configuration. We sequenced DNA of 15 human cell samples confirming the identification of conserved sequences and identifying undesired SNPs. Then we selected the CRISPR/CasX, which is compatible in silico with the excision of the 4q region in all of these samples. We designed CasX Single guide RNAs (SgRNAs) and tested them in one FSHD1 fibroblast cell line. Importantly, the SgRNAs induced INDELs in the 4q telomeric region, thus leading to haploinsufficiency. We obtained a CRISPR/CasX SgRNAs maximal efficiency of 27.3%. The validation of the D4Z4 locus ablation will be verified with the Nanopore technology. Our work represents a significant step toward an in vitro gene therapy development for FSHD1. [ABSTRACT FROM AUTHOR]

Synthetic pyrethroids are widely used insecticides which may cause chronic diseases in non-target organisms upon long-term exposure. Microbial degradation offers a reliable method to remove them from the environment. This study focused on Brevibacillus parabrevis BCP-09 and its enzymes for degrading pyrethroids. The predicted deltamethrin-degrading genes phnA and mhpC were used to construct recombinant plasmids. These plasmids, introduced into Escherichia coli BL21(DE3) cells and induced with L-arabinose. The results indicated that the intracellular crude enzyme efficiently degraded deltamethrin by 98.8 %, β-cypermethrin by 94.84 %, and cyfluthrin by 73.52 % within 24 h. The hydrolytic enzyme MhpC possesses a catalytic triad Ser/His/Asp and a typical "Gly-X-Ser-X-Gly" conservative sequence of the esterase family. Co-cultivation of induced E. coli PhnA and E. coli MhpC resulted in degradation rates of 41.44 ± 3.55 % and 60.30 ± 4.55 %, respectively, for deltamethrin after 7 d. This study states that the degrading enzymes from B. parabrevis BCP-09 are an effective method for the degradation of pyrethroids, providing available enzyme resources for food safety and environmental protection. [Display omitted] • Strain BCP-09 intracellular crude enzyme degrades deltamethrin by 98.8 %. • MhpC has a catalytic triad (Ser/His/Asp) and a conserved sequence "Gly-X-Ser-X-Gly". • Induced by inducer, PhnA and MhpC fusion protein was expressed in E. coli BL21(DE3). • After 7 d, E. coli PhnA degraded deltamethrin by 41.44 %, MhpC degraded it by 60.30 %. [ABSTRACT FROM AUTHOR]