Search Results (1,529)

Transposable elements (TEs) are major components of plant genomes and play crucial roles in adaptive genome evolution and stress tolerance. Under abiotic stress, activated TEs can generate abundant genetic variation and regulate the expression of stress-responsive genes. As a pioneer species in desert and saline–alkali environments, Tamarix chinensis L. has been little studied with respect to the abundance and evolutionary relationships of its LTR retrotransposons, particularly their activation patterns under salt and alkali stresses. This study aimed to investigate the characteristics of the reverse transcriptase (RT) domain of LTR retrotransposons in T. chinensis and to determine their patterns of activation in response to salt and alkali stresses. A total of 629 Ty1-copia and 607 Ty3-gypsy RT nucleotide sequences, which displayed high AT/GC ratios and evidence of stop codon insertions, were identified in T. chinensis by amplicon sequencing. Among these, 211 Ty1-copia and 117 Ty3-gypsy RT sequences with potential transpositional activity each contained distinct domains, suggesting a high degree of conservation. Phylogenetic analysis revealed that the RT sequences of T. chinensis are closely related to those of mangrove, wild potato, and Ipomoea, and may have undergone horizontal transfer. Expression analysis showed that 634 and 181 RT sequences were activated under salt and alkali stresses, respectively, with the majority belonging to salt-induced Ty1-copia families. Compared with the control group, under salt and alkali stresses, the cTy1-copia elements (Ty1-copia with amplificated from cDNA of T. chinensis, the same below) with dominant abundance were mainly concentrated in the Angela subfamily, while the cTy3-gypsy elements induced by alkali stress were primarily distributed in the Tekay and Reina subfamilies. Furthermore, four cTy1-copia and five cTy3-gypsy were identified as candidate key LTR retrotransposons responsive to salt and alkali stresses. Overall, this study provides new insights into the epigenetic mechanisms underlying the adaptation of T. chinensis to saline and alkali stresses and offers a theoretical basis for its potential applications in saline–alkali land reclamation. Full article

SARS-CoV-2 persists worldwide, driving the demand for effective antivirals that inhibit replication and limit the emergence of resistant variants. Lycorine, a non-nucleoside inhibitor of SARS-CoV-2 RNA-dependent RNA polymerase, exhibits antiviral activity without direct mutagenic effects. Here, we examine the occurrence of single-nucleotide variants (SNVs) and insertions/deletions (indels) in SARS-CoV-2 B.1.499 strain during serial passages in Vero cells, comparing lycorine-treated cultures (2.5 and 5 µg/mL) with untreated controls. Whole-genome sequencing was used to assess mutation patterns and frequencies. Lycorine-treated passages displayed greater variant diversity than controls, with fixed mutations mainly affecting non-structural proteins (Nsp3-F1375A, Nsp5-L50F, and Nsp14-G265D) and the envelope protein (E-S6L). A 15-nucleotide deletion in the spike gene (QTQTN motif) occurred in both groups but became fixed only in untreated passages, suggesting negative selection under lycorine pressure. Notably, the L50F mutation in Nsp5, previously linked to nirmatrelvir resistance, was found exclusively in lycorine-treated passages. Additionally, a 1-nucleotide deletion in the accessory gene ORF8, detected only under lycorine treatment, resulted in a frameshift mutation that added four amino acids, potentially altering the protein’s function. Overall, lycorine induces a distinct mutation profile, favoring replication-related variants while suppressing deleterious deletions. These findings suggest potential mechanisms of cross-resistance and highlight the importance of monitoring resistance during clinical use. Full article

Objectives: To explore the genetic basis of phage resistance in sequentially generated capsular mutants of phage-resistant Klebsiella pneumoniae using an established phage library. Methods: Sequential induction strategies were employed to obtain phage-resistant K. pneumoniae capsular mutants by exposing ST11-K64 K. pneumoniae Kp2325 to different single phages. Whole genome sequencing and bioinformatic analysis were used to elucidate the capsular-related genetic changes in phage-resistant mutants. Phenotypic changes were assessed through gene complementation, growth assays, phage cleavage spectrum analysis, TEM for phage morphology, CPS analysis, biofilm formation, and virulence assays. Results: Three sequentially generated phage-resistant K. pneumoniae capsular mutants were obtained, designated R1, R2 and R3. The narrowing of the phage cleavage spectrum and the evolutionary trade-offs of biological phenotypes were observed. Key genetic changes included: (1) ISKpn26 insertion disrupting wcaJ in R1; (2) combined wcaJ insertion and 9-bp deletion in waaH in R2; and (3) CPS gene cluster deletion in R3 were identified as key mechanisms of phage resistance in K. pneumoniae mutants R1, R2 and R3, respectively. Conclusions: Sequential exposure to different single phages led to rapid evolution of phage resistance in K. pneumoniae via genetic mutations that disrupt capsular synthesis. These findings highlight the critical role of bacterial capsule in phage–host interactions and emphasize the need to use phage cocktails targeting different types of receptors to counteract the evolution of bacterial defense mechanisms in phage therapy. Full article

Freshwater mussels in the genera Fusconaia, Pleurobema, and Pleuronaia are similar in their external shell morphology, which has made the identification and classification of species within these genera difficult and led to many taxonomic revisions. Large samples (N = 464) of select mussel species in these genera were collected from 2012 through 2014, primarily in the upper Tennessee River basin of Tennessee and Virginia, USA. Mitochondrial ND1 and nuclear ITS1 DNA sequences were analyzed to assess phylogenetic relationships among taxa. Ten species were verified as phylogenetically distinct at ND1, two of which were cryptic and previously unrecognized species. Described herein as Pleurobema parmaleei and Pleuronaia estabrookianus, each species clade was diverged at this gene region by ~3.0% from the respective closest congener. The nuclear ITS1 gene region’s nucleotide-site insertion/deletion (indel) patterns were analyzed as single mutational events rather than as fifth character states or missing data. Most species, including these two, were phylogenetically distinct at the ITS1 region when incorporating indels into analyses, but some estimated interspecific pairwise distances were lower than corresponding intraspecific estimates. Among morphological traits assessed for each species, differences in foot color and gravidity characteristics illustrated differences between phylogenetically recognized species and their closest congeners. Due to the limited known geographical distributions of these two cryptic species, each may require protection under the U.S. Endangered Species Act. While this study collected large sample sizes for each species, many streams in the basin remain unsampled and could potentially contain populations of these species or additional cryptic species. Full article

Rational design of capripoxvirus-based vaccine vectors can be achieved by knockout of immunomodulatory genes. In this study, the effect of knockout of the immunomodulatory genes LSDV005, LSDV008 and LSDV066 on the replication of Lumpy skin disease virus in cell cultures and the immune response to an integrated foreign antigen were assessed. The knockout of genes was performed by homologous recombination under conditions of temporary dominant selection. It was found that single knockout of the LSDV005 gene and the LSDV008 gene did not affect the replicative activity of recombinant viruses in vitro (Atyrau-5 and Atyrau-B). Both single knockout of the LSDV066 gene and in combination with knockout of LSDV005 or LSDV008 led to a decrease in the replicative activity of recombinant LSDVs. The recombinant Atyrau-5J(IL18) with LSDV005 gene knockout induced production of antibodies to the integrated antigen in mice. Prime-boost vaccination with all studied recombinants increased the level of interferon-γ. In addition, during immunization with the recombinant Atyrau-5J(IL18) secretion of interleukin-2 was significantly increased. The study of the functions of immunomodulatory genes and their effect on the expression of inserted sequences of foreign antigens is promising for the creation of highly effective polyvalent vector vaccines for animals. Full article

Background: Digital polymerase chain reaction (dPCR) is a highly sensitive molecular method that allows rapid detection of bacterial DNA and resistance genes, requiring only a small blood volume. Although not a new technology, its application in pediatric patients with suspected catheter-related bloodstream infection (CRBSI) remains limited. Case presentation: A 16-year-old female, diagnosed with recurrent acute myelogenous leukemia, received re-induction chemotherapy through a peripherally inserted central venous catheter (PICC). The patient developed a fever, and the blood culture (BC) drawn from the PICC was positive for methicillin-resistant S. epidermidis, leading to suspicion of CRBSI. Several antibiotics were used, and the PICC was replaced. Eventually, the fever subsided, and the BC was negative after PICC removal. The levels of S. epidermidis-specific DNA sequences and mecA genes were correlated with the results of the BC and clinical course. Turnaround time was significantly shorter in dPCR (3.5 h) than in the BC (14–21 h); dPCR was performed using only 400 µL of blood. Conclusions: This case highlights the potential of dPCR as a complementary tool to conventional BCs in the management of pediatric CRBSI. dPCR may support rapid decision-making and monitoring of the treatment response, particularly when sample volumes are limited. Full article

Background/Objectives: Short inverted repeats (SIRs) are abundant DNA motifs capable of forming secondary structures, such as hairpins and cruciforms, that can induce genome instability. However, their mutational consequences in cancer, particularly in osteosarcoma (OS), remain largely unexplored. Methods: In this study, we systematically identified over 5.2 million SIRs in the human genome and analyzed their mutational patterns across six common cancer types. Results: We found that increased small insertion and deletion (INDEL) density within SIR spacer regions represents a consistent feature across cancers, whereas elevated single nucleotide variant (SNV) and structural breakpoint density is cancer-type specific. Integrating whole-genome sequencing data from 13 OS patients, we found that both SNVs and INDELs are significantly enriched within SIR spacer regions in OS. Notably, genomic regions with higher SIR density tend to accumulate more somatic mutations, suggesting a link between SIR abundance and local genome instability. SIR-associated mutations frequently occur in oncogenes and tumor suppressor genes, including TP53, NFATC2, MECOM, LRP1B, RB1, CNTNAP2, and PTPRD, as well as in long non-coding RNAs. Mutational signature analysis further suggests that defective DNA mismatch repair and homologous recombination may act in concert with SIR-induced DNA structural instability to drive OS development. Conclusions: Our findings highlight SIRs as mutational hotspots and potential drivers of osteosarcoma pathogenesis. Full article

Background: Telomeres are nucleoprotein complexes that maintain chromosome integrity in eukaryotes. In insects, the canonical telomeric repeat (TTAGG)n is considered ancestral, though alternative motifs exist across various orders. Neuroptera, comprising about 5800 species, remains understudied regarding telomeric sequences, with data available for only seven species across three families. Previous studies reported the absence of (TTAGG)n in Chrysopidae species, contrasting with its presence in other Neuroptera families. This study aimed to identify and characterize telomeric motifs in Chrysopidae using chromosome-level genome assemblies and search for retrotransposon insertions. Methods: We analyzed chromosome-level genome assemblies from four Chrysopidae species: three Chrysopinae—Chrysoperla carnea (Stephens, 1836), Chrysopa pallens (Rambur, 1838), and Nineta flava (Scopoli, 1763); and one Nothochrysinae—Nothochrysa capitata (Fabricius, 1793). Terminal sequences of chromosome pseudomolecules were examined using Geneious Prime^®, applying five specific criteria for optimal telomeric sequence identification. We searched for SART and TRAS retrotransposons using the graphical sequence panel in GenBank. Results: We identified (TTGGG)n as the telomeric motif in N. flava, representing the first report of this pentanucleotide repeat in telomeres of Neuroptera. Arrays ranged from 228 to 8005 bp across seven terminal locations in five chromosome pseudomolecules. In N. capitata, we detected (TTAGG)n arrays (2316–3808 bp) at four terminal locations. No telomeric motifs meeting all criteria were found in C. carnea and C. pallens. No SART/TRAS retrotransposons were detected in any species. Conclusions: This study reveals previously unknown telomeric diversity within Chrysopidae, with both canonical (TTAGG)n and novel (TTGGG)n motifs present. The discovery of (TTGGG)n in Neuroptera expands known telomeric sequence diversity in this order. Full article

Background/Objectives: Actinobacillus pleuropneumoniae is an important respiratory tract pathogen of swine worldwide. Insertion sequences (ISs) play a major role in the transfer of antimicrobial resistance (AMR) among various porcine respiratory tract pathogens. In this study, three A. pleuropneumoniae genomes were investigated for the presence of a novel IS. Methods: Analysis of the draft genomes of three A. pleuropneumoniae serovar 8 isolates (AP_1, AP_120, AP_123) suggested the presence of a novel IS. A closed whole-genome sequence was generated for strain AP_123 by hybrid assembly of Oxford Nanopore MinION long-reads and Illumina MiSeq short-reads, followed by sequence analysis using standard online tools. Transfer was tested by natural transformation. Antimicrobial susceptibility testing was conducted by broth microdilution following Clinical and Laboratory Standards Institute standards. Results: A novel IS, designated ISApl4, was detected in all three genomes. ISApl4 is 712 bp in size and has a transposase gene (tnp) of 654 bp. Moreover, it has perfect terminal 14-bp inverted repeats and produces 8-bp direct repeats at its integration site. This IS was found in 39 copies in the AP_123 genome, two of which formed the 5,765-bp pseudo-compound transposon Tn7560. This transposon carries four AMR genes: sul2 (sulfonamide resistance), strA-strB (streptomycin resistance), and tet(Y) (tetracycline resistance). RT-PCR confirmed tnp gene expression and horizontal transfer of Tn7560 into A. pleuropneumoniae MIDG2331. Conclusions: This study identified the novel ISApl4 in porcine A. pleuropneumoniae and its association with the novel pseudo-compound transposon Tn7560, which proved to be an active element capable of disseminating multidrug resistance amongst A. pleuropneumoniae. Full article

Extended-spectrum β-lactamases (ESBLs) and plasmid-mediated AmpC (pAmpC) β-lactamases represent a threat for public health. Their dissemination is often mediated by mobile genetic elements (MGEs), but plasmid identification and characterization could be hindered by sequencing limitations. Hybrid assembly may overcome these barriers. Eight ESBL/pAmpC-producing E. coli isolates from broilers were sequenced using Illumina (short-read) and Oxford Nanopore MinION (long-read). Assemblies were generated individually and using a hybrid approach. Plasmids were typed, annotated, and screened for antimicrobial resistance genes (ARGs), MGEs, and virulence factors. Short-read assemblies were highly fragmented, while long reads improved contiguity but showed typing errors. Hybrid assemblies produced the most accurate and complete plasmids, including more circularized plasmids. Long and hybrid assemblies detected IS26 associated with ESBL genes and additional virulence genes not identified by short reads. ARG profiles were consistent across methods, but structural resolution and contextualization of resistance loci were superior in hybrid assembly. Hybrid assembly integrates the strengths of short- and long-read sequencing, enabling accurate plasmid reconstruction and improved detection of resistance-associated MGEs. This approach may enhance genomic surveillance of ESBL/pAmpC plasmids and support strategies to mitigate antimicrobial resistance. Full article

Background/Objectives: The global dissemination of tet(X) variants critically threatens tigecycline efficacy as a last-resort antibiotic. The aim of this study was to characterize a tet(X6)-carrying integrative and conjugative element (ICE) in a multidrug-resistant Chryseobacterium lecithinasegens strain, SHZ29, isolated from Shanghai, China. Methods: Minimum inhibitory concentrations (MICs) were determined by broth microdilution for SHZ29. Whole genomic sequencing and bioinformatic analysis were performed to depict the structure of the novel tet(X6)-carrying ICE. Inverse PCR and conjugation experiments were conducted to investigate the transfer ability of the ICE. Results: We depicted a novel tet(X6)-carrying ICE, named ICECleSHZ29, which is 74,906 bp in size and inserted into the 3′ end of tRNA-Met-CAT gene of the C. lecithinasegens strain SHZ29, with 17 bp direct repeats (5′-tcccgtcttcgctacaa-3′). This ICE possesses a 38 kb conserved backbone and four variable regions (VR1-4), with VR3 aggregating multiple resistance genes, including tet(X6), tet(X2), erm(F), ere(D), floR, catB, sul2, ant(6)-I and bla_OXA-1327. NCBI database searching identified 13 additional ICEs sharing a similar backbone to ICECleSHZ29. These ICECleSHZ29-like ICEs could be classified into two types based on their distinct insertion sites: Type I, inserted at the tRNA-Met-CAT gene; and Type II, inserted at the tRNA-Glu-TTC gene. Phylogenetic analysis indicated that differences in integrases may result in differences in the insertion site among these ICEs. A circular intermediate form of ICECleSHZ29 was detected by inverse PCR. However, the conjugation experiments using Escherichia coli EC600 as recipients failed. Conclusions: To our knowledge, this study provides the first report of tet(X6) in C. lecithinasegens and characterizes its carrier, a novel ICE: ICECleSHZ29. Full article

Damping-off disease hinders rice seedling growth and reduces yield. Current control methods, such as seed or soil sterilization, rely on chemicals that cause environmental pollution and promote pathogen resistance. As a sustainable alternative, we targeted the damping-off resistance-related gene OsDGTq1 using CRISPR/Cas9. Field experiments first verified OsDGTq1’s significance in resistance. The CRISPR/Cas9 system, delivered via Agrobacterium-mediated transformation, was used to edit OsDGTq1 in rice cultivar Ilmi. Lesions from major damping-off pathogens, Rhizoctonia solani and Pythium graminicola, were observed on G₀ plants. All 37 regenerated plants contained T-DNA insertions. Among them, edits generated by sgRNA1-1, sgRNA1-2, and sgRNA1-3 resulted in the insertion of two thymine bases as target mutations. Edited lines were assigned names and evaluated for agronomic traits, seed-setting rates, and pathogen responses. Several lines with edited target genes showed distinct disease responses and altered gene expression compared to Ilmi, likely due to CRISPR/Cas9-induced sequence changes. Further studies in subsequent generations are needed to confirm the stability of these edits and their association with resistance. These results confirm that genome editing of OsDGTq1 alters resistance to damping-off. The approach demonstrates that gene-editing technology can accelerate rice breeding, offering an environmentally friendly strategy to develop resistant varieties. Such varieties can reduce chemical inputs, prevent pollution, and minimize seedling loss, ultimately enhancing food self-sufficiency and stabilizing rice supply. Full article

Polyproline residues are well known to induce ribosomal stalling during translation. Our previous work demonstrated that inserting a short translation-enhancing peptide, Ser-Lys-Ile-Lys (SKIK), immediately upstream of such difficult-to-translate sequences can significantly alleviate ribosomal stalling in Escherichia coli. In this study, we provide a quantitative evaluation of its translational effect by kinetically analyzing the influence of the SKIK peptide on polyproline motifs using a reconstituted E. coli in vitro translation system. Translation rates estimated under reasonable assumptions fitted well to a Hill equation within a Michaelis–Menten-like kinetic framework. We further revealed that repetition of the SKIK tag did not provide any positive effect on translation. Moreover, introduction of the SKIK tag increased the production of polyproline-containing proteins, including human interleukin 11, human G protein signaling modulator 3, and DUF58 domain–containing protein from Streptomyces sp. in E. coli cell-free protein synthesis. These findings not only provide new insight into the fundamental regulation of translation by nascent peptides but also demonstrate the potential of the SKIK peptide as a practical tool for synthetic biology, offering a strategy to improve the production of difficult-to-express proteins. Full article

The global rise of antimicrobial resistance (AMR), exacerbated by the COVID-19 pandemic, has led to a surge in infections caused by multidrug-resistant (MDR) bacteria. A key driver of this phenomenon is co-selection, where exposure to one antimicrobial promotes resistance to others via horizontal gene transfer (HGT) mediated by mobile genetic elements (MGEs). Carbapenem-resistant Enterobacteriaceae, known for their genomic plasticity, are particularly worrisome; yet genomic data from Latin America—especially Ecuador—remain scarce. This study investigated four carbapenem-resistant clinical isolates (two Klebsiella pneumoniae ST1440 and two Serratia marcescens) from tracheal aspirates of three ICU patients during a COVID-19 outbreak at Hospital IESS Quito Sur, Ecuador. Phenotypic profiling and whole-genome sequencing were performed, followed by bioinformatic reconstruction of plasmid content. Nineteen plasmids were identified, carrying 70 resistance-related genes, including antimicrobial resistance genes (ARGs), metal resistance genes (MRGs), integrons, transposons, and insertion sequences. Hierarchical clustering revealed six distinct gene clusters, with several co-localizing ARGs and genes for resistance to disinfectants and heavy metals—suggesting strong co-selective pressure. Conjugative plasmids harboring high-risk elements such as blaKPC-2, qacE, and Tn4401 were found in multiple isolates, indicating potential interspecies dissemination. These findings emphasize the importance of plasmid-mediated resistance during the pandemic and highlight the urgent need to enhance genomic surveillance and infection control, particularly in resource-limited healthcare settings. Full article

Lesion mimic mutants (LMMs) represent valuable biological tools for investigating plant defense mechanisms and cell death. Although multiple genes triggering lesion mimic formation have been identified, the connection between the lesion mimic phenotype and primary nutrient biosynthesis remains poorly understood. In our study, we characterized a novel rice LMM, lmm9, which exhibited persistent reddish-brown necrotic lesions from seedling stage to maturity, coupled with compromised agronomic traits and increased mortality rates. Map-based cloning and whole-genome sequencing identified a causal insertion in the promoter of Os03g03450/OsPAT1, the sole homolog of Arabidopsis PAT1 in rice, resulting in reduced gene expression. Genetic complementation and RNAi assays confirmed that downregulation of OsPAT1 led to lesion mimic formation in lmm9. OsPAT1 could translate into two variants—the predominant OsPAT1.1 and the C-terminal variant OsPAT1.2. Structural modeling demonstrated high conservation between OsPAT1 and yeast TRP4, and OsPAT1.1 combining the plastid signal sequence of Arabidopsis PAT1 successfully complemented the trp4 mutant in yeast. Notably, OsPAT1.1 and OsPAT1.2 showed different localization patterns, with OsPAT1.1 targeted to mitochondria and OsPAT1.2 localized to chloroplasts. Transcription analysis showed significant upregulation of tryptophan biosynthesis pathway genes in lmm9, consequently increasing the relative abundance of tryptophan and associated metabolites. Our findings provided further evidence that mutations in tryptophan biosynthetic genes can induce lesion mimic phenotypes in rice and would enhance the understanding of metabolic homeostasis in plant stress responses and cell death regulation. Full article