Search Results (76)

DNA is inherently unstable and is susceptible to damage from both endogenous sources (such as reactive oxygen species) and exogenous factors (including UV, ionizing radiation, and chemicals). The accumulation of DNA damage manifests as genetic mutations, chromosomal instability, and the stalling of DNA replication and transcription processes. Accumulated DNA damage influences apoptosis and cell cycle checkpoints, serving as one of the key triggers for the manifestation of the senescent phenotype. Both aging and cancer are associated with the accumulation of mutations in somatic cells. Disruption of cell cycle control and uncontrolled proliferation are fundamental characteristics of any cancer cell, with the majority of anticancer drugs acting as inhibitors of cyclin-dependent kinases, thereby inducing a transition of cells into a senescent state. Consequently, disturbances in the dynamics and regulation of inflammatory responses, oxidative stress, cell proliferation, DNA damage repair, and epigenetic anomalies, along with the influence of retroviruses and transposons, lead to the accumulation of senescent cells within the human body, characterized by blocked replication and cell cycle, as well as a distinct secretory phenotype. The age-related or disease-associated accumulation of these senescent cells significantly alters the physiology of tissues and the organism as a whole. Many secondary metabolites of higher plants exhibit senolytic and senomorphic activities, although most of them are not fully characterized. In this review, we will explore the principal signaling pathways in mammalian cells that govern the cell cycle and cellular senescence, with a particular emphasis on how their dynamics, expression, and regulation have been modified through the application of senotherapeutic compounds. The second section of the review will identify key target genes for the metabolic engineering, primarily aimed at enhancing the accumulation of plant secondary metabolites with potential therapeutic benefits. Lastly, we will discuss the rationale for utilizing liver cells as a model system to investigate the effects of senolytic compounds on human physiology and health, as well as how senotherapeutic substances can be leveraged to improve gene therapy approaches based on CRISPR/Cas9 and prime-editing technologies. Full article

The growth of aquaculture production in recent years has revealed multiple challenges, including the rise of antimicrobial resistance (AMR) in aquatic animal production, which is currently attracting significant attention from multiple one-health stakeholders. While antibiotics have played a major role in the treatment of bacterial infections for almost a century, a major consequence of their use is the increase in AMR, including the emergence of AMR in aquaculture. The AMR phenomenon creates a situation where antibiotic use in one system (e.g., aquaculture) may impact another system (e.g., terrestrial–human). Non-prudent use of antibiotics in aquaculture and animal farming increases the risk of AMR emergence, since bacteria harboring antibiotic resistance genes can cross between compartments such as wastewater or other effluents to aquatic environments, including intensive aquaculture. Transferable antimicrobial resistance gene (AMG) elements (plasmids, transposons, integrons, etc.) have already been detected in varying degrees from pathogenic bacteria that are often causing infections in farmed fish (Aeromonas, Vibrio, Streptococcus, Pseudomonas, Edwardsiella, etc.). This review of current veterinary approaches for the prevention and control of AMR emergence in aquaculture focuses on the feasibility of alternatives to antimicrobials and supplemental treatment applications during on-farm bacterial disease control and prevention. The use of vaccines, bacteriophages, biosurfactants, probiotics, bacteriocins, and antimicrobial peptides is discussed. Full article

Background: Infections caused by carbapenem-resistant Enterobacterales have steadily multiplied over time, becoming a major threat to healthcare systems due to limited therapeutic options and high case-fatality rates. Case report: We studied a patient who, after being discharged from an ICU, developed salmonellosis caused by an antibiotic-susceptible S. enteritidis. After undergoing treatment with ciprofloxacin, the patient presented an episode of asymptomatic bacteriuria originated by a carbapenem and ciprofloxacin-resistant S. enteritidis. Results: Whole genome sequencing analysis revealed that both Salmonella isolates belonged to the same strain, and that isolate SEn_T2 acquired a plasmid carrying both bla_NDM-1 and qnrA1 genes (pIncCSEn) which was previously present in the patient’s gut in at least one Enterobacter cloacae isolate. Additionally, pIncCSEN was identified as a putatively new sub-lineage of IncC2 plasmids which lacked the first copy of the methyltransferase gene dcm and the rhs gene. The resistance genes bla_NDM-1 and qnrA1 were incorporated into a Tn21-derived transposon that included a complex class 1 integron whose genetic arrangement was: intI1- dfrA12- orfF- aadA2- qacEΔ1-sul1-ISCR1- trpF- ble- bla_NDM-1 (in reverse direction)- ISAba125-ISCR1- qnrA- cmlA1- qacEΔ1-sul1. Conclusions: Antimicrobial persistence and co-selection of antibiotic resistance play an important role in the dissemination of antimicrobial resistance genes; in this regard, a joint effort involving the infection control team, effective antibiotic stewardship, and genomic surveillance could help mitigate the spread of these multidrug resistant microorganisms. Full article

Objectives: Linezolid resistance among Enterococcus species poses a growing clinical and public health concern, especially due to the dissemination of transferable resistance genes, such as optrA. This study aimed to evaluate the prevalence of linezolid resistance and to characterize the molecular epidemiology and genetic contexts of optrA-positive linezolid-resistant Enterococcus (LRE) isolates from clinical and animal sources in South Korea. Methods: A total of 2156 Enterococcus isolates, collected through nationwide surveillance from hospitalized patients and healthy livestock (pigs, cattle, and chickens) between 2017 and 2019, were retrospectively analyzed. Phenotypic susceptibility testing, optrA gene screening, and whole-genome sequencing were performed to investigate genetic environments and phylogenetic relationships. Results: The prevalence of linezolid resistance was 0.2% in clinical isolates, 3.3% in pigs, 4.3% in cattle, and 1.4% in chickens. optrA-positive linezolid-resistant isolates were less frequent, with rates of 0.1%, 1.4%, 0.9%, and 1.0%, respectively. Multilocus sequence typing identified sequence types (STs) 330 and ST476 in E. faecalis from humans, with no shared STs between human and livestock isolates. The optrA gene was located either chromosomally, frequently associated with transposon Tn6674, or on multidrug resistance plasmids. Notably, optrA variants exhibited host-specific distribution patterns. Phylogenetic analysis demonstrated considerable genomic diversity, and Korean ST476 isolates were genetically related to international strains reported from livestock, poultry products, and wild birds, suggesting potential global dissemination. Conclusions: This study provides a comprehensive, nationally representative assessment of linezolid resistance in South Korea. The findings highlight the zoonotic potential and possible international dissemination of optrA-carrying ST476 lineages, underscoring the need for integrated One Health surveillance to monitor and control the spread of transferable resistance genes. Full article

Utilizing high-throughput Illumina sequencing, we examined how small RNA (sRNA) profiles vary in Chinese white poplar (Populus tomentosa) across two pivotal infection stages by the rust fungus Melampsora larici-populina: the biotrophic growth phase (T02; 48 h post infection) and the urediniospore development and dispersal phase (T03; 168 h), both essential for plant colonization and prolonged biotrophic engagement. Far exceeding random expectations, siRNA clusters predominantly arose from transposon regions, with pseudogenes also contributing significantly, and infection-stage-specific variations were notably tied to these transposon-derived siRNAs. As the infection advanced, clusters of 24 nt siRNAs in transposon and intergenic regions exhibited pronounced abundance shifts. An analysis of targets indicated that Populus sRNAs potentially regulate 95% of Melampsora larici-populina genes, with pathogen effector genes showing heightened targeting by sRNAs during the biotrophic and urediniospore phases compared to controls, pointing to selective sRNA-target interactions. In contrast to conserved miRNAs across plant species, Populus-specific miRNAs displayed a markedly greater tendency to target NB-LRR genes. These observations collectively highlight the innovative roles of sRNAs in plant defense, their evolutionary roots, and their dynamic interplay with pathogen coevolution. Full article

Mucor lusitanicus has emerged as a model organism for studying RNAi in early-diverging fungi. This fungus exhibits intricate RNAi pathways that play crucial roles in regulating gene expression, destroying invasive exogenous genetic material, and controlling the movement of transposable elements (TEs) to ensure genome stability. One of the most fascinating RNAi pathways of this fungus is the non-canonical RNAi pathway (NCRIP), which is independent of Dicer and Argonaute proteins and uses the atypical RNase III R3B2 to degrade specific target messenger RNAs (mRNAs), playing an essential role in genome stability and virulence. Despite accumulating data suggesting that this pathway is a degradation mechanism, there has been no conclusive evidence. Here, we conducted a comparative transcriptomic analysis of mRNA and small RNAs regulated by r3b2, identifying 35 direct NCRIP targets. Most of these direct NCRIP targets correspond to TEs, highlighting the significant role of this RNAi pathway in TE control. Detailed functional analysis of the NCRIP targets confirmed the crucial role of r3b2 in regulating gene expression of protein-coding genes and controlling TEs other than centromeric GremLINE1 transposons, emphasizing the important role of r3b2 in genome stability. Interestingly, the RNAs of the NCRIP targets harbor a unique motif consisting of CAG repeats which are known to form hairpin structures which are targeted by RNA interference. Additionally, the generation of transformants expressing mRNAs containing the luciferase reporter gene along direct NCRIP targets reveals that this RNAi pathway is a true degradation mechanism for specific mRNAs. These results are expected to contribute to the understanding of the regulation of the NCRIP pathway through the analysis of its direct targets identified here. Full article

Alternative splicing of ddx4 (DEAD-box helicase 4), a key germline marker gene, has been reported to generate sex-specific transcripts in zebrafish gonads. The biological functions and regulatory mechanisms of the ddx4 ovary-specific transcript (ddx4-L) during oogenesis remain unclear. In this study, we found that ddx4-L mutants, in which ddx4-L was specifically deleted, had enlarged ovaries but laid fewer eggs, along with having a lower fertilization rate compared to WT controls. RNA-seq analysis was performed to detect the changes in gene expression between WT and ddx4-L mutant ovaries. A total of 524 upregulated and 610 downregulated DEGs were identified. GO and GSEA enrichment analyses showed that genes involved in fertilization and reproduction biological processes were significantly downregulated. More specifically, we observed a remarkable reduction in Sycp1, a core component of synaptonemal complex, in ddx4-L mutant ovaries at both the mRNA and protein levels. In addition, the expressions of transposon elements, as well as the events of alternative splicing, alternative polyadenylation, and RNA editing, were analyzed based on the RNA-seq data. We found that the deletion of ddx4-L resulted in derepression of DNA transposons in zebrafish ovaries, possibly causing genome instability. In conclusion, our work demonstrates that the ovary-specific ddx4 transcript plays important roles in oocyte meiosis and DNA transposon repression, which extends our understanding of the biological functions and regulatory mechanisms of sex-specific alternative splicing in zebrafish oogenesis and reproduction. Full article

Currently, pJP4 is one of the best-known plasmids for the biodegradation of xenobiotics that mediate the degradation of 2,4-dichlorophenoxyacetic acid (2,4-D), which is associated with serious health and environmental risks. Although the sequencing and proposed theory of pJP4 formation occurred almost 20 years ago (2004), pJP4 is still the model object of many studies focused on the biodegradation of 2,4-D. The uniqueness of this plasmid is due to the presence of two evolutionarily distinct gene clusters, tfd_I and tfd_II, controlling the degradation of 2,4-D. Recent advances in plasmid biology, especially those concerning the characterization of new IncP-1 plasmids and the systematization of tfd gene cluster findings, serve as a basis for proposing new insights into the formation of the clusters’ architecture of the canonical plasmid, pJP4, and their distribution among other plasmids. In the present work, a comparative genomic and phylogenetic in silico study of plasmids with tfd_I and tfd_II clusters was carried out. The possible initial distribution patterns of tfd_I clusters among plasmids of different incompatibility groups (non-IncP-1) and tfd_II clusters among IncP-1 plasmids using the IS1071-based composite transposon were revealed. A new theory on the formation of the architecture of the tfd_I and tfd_II clusters of pJP4 through sequential internal rearrangements, recombination, and ISJP4 insertion, is proposed. In addition, small gene clusters resulting from internal rearrangements of pJP4 (tfd_IISA and ORF31/32) served as fingerprints for exploring the distribution of tfd_I and tfd_II clusters. The revealed patterns and formulated theory extend the frontiers of plasmid biology and will be beneficial for understanding the role of plasmids in bacterial adaptation to xenobiotic-contaminated environments. Full article

Chimeric antigen receptor (CAR) T-cell therapy has achieved notable success in treating hematological cancers but faces significant challenges in solid-tumor treatment and overall efficacy. Key limitations include T-cell exhaustion, tumor relapse, immunosuppressive tumor microenvironments (TME), immunogenicity, and antigen heterogeneity. To address these issues, various genetic engineering strategies have been proposed. Approaches such as overexpression of transcription factors or metabolic armoring and dynamic CAR regulation are being explored to improve CAR T-cell function and safety. Other efforts to improve CAR T-cell efficacy in solid tumors include targeting novel antigens or developing alternative strategies to address antigen diversity. Despite the promising preclinical results of these solutions, challenges remain in translating CAR T-cell therapies to the clinic to enable economically viable access to these transformative medicines. The efficiency and scalability of autologous CAR T-cell therapy production are hindered by traditional, manual processes which are costly, time-consuming, and prone to variability and contamination. These high-cost, time-intensive processes have complex quality-control requirements. Recent advancements suggest that smaller, decentralized solutions such as microbioreactors and automated point-of-care systems could improve production efficiency, reduce costs, and shorten manufacturing timelines, especially when coupled with innovative manufacturing methods such as transposons and lipid nanoparticles. Future advancements may include harmonized consumables and AI-enabled technologies, which promise to streamline manufacturing, reduce costs, and enhance production quality. Full article

Although biofilms contribute to bacterial tolerance to desiccation and survival in low-moisture foods, the molecular mechanisms underlying biofilm formation have not been fully understood. This study created a mutant library from Salmonella Enteritidis using mini-Tn10 transposon mutagenesis. The biofilm-forming potential of acquired mutants was assessed before the genomic DNA of the mutants that formed significantly (p ≤ 0.05) less biofilm mass than their wildtype parent strain was extracted for deep DNA sequencing. The gene of each mutant interrupted by mini-Tn10 insertion was identified by aligning obtained sequencing data with the reference Genbank sequences using a BLAST search. Sixty-four mutant colonies were selected, and five mutants that formed the least amount of biofilm mass compared to the wildtype parent strain were selected for sequencing analysis. The results of the BLAST search revealed that the gene interrupted by mini-Tn10 in each mutant is responsible for the biosynthesis of aldehyde dehydrogenase (EutE), cysteine desulfurase (SufS or SufE), a transporter protein, porin OmpL, and a ribbon–helix–helix protein from the CopG family, respectively. Knock-off mutant construction is a possible approach to verify the potential of the identified genes to serve as targets of antimicrobial intervention to control Salmonella colonization on low-moisture foods and in their production environment. Full article

The Maculipennis subgroup of malaria mosquitoes includes both dominant malaria vectors and non-vectors in Eurasia. Understanding the genetic factors, particularly chromosomal inversions, that differentiate Anopheles species can provide valuable insights for vector control strategies. Although autosomal inversions between the species in this subgroup have been characterized based on the chromosomal banding patterns, the number and positions of rearrangements in the X chromosome remain unclear due to the divergent banding patterns. Here, we identified two large X chromosomal inversions, approximately 13 Mb and 10 Mb in size, using fluorescence in situ hybridization. The inversion breakpoint regions were mapped by hybridizing 53 gene markers with polytene chromosomes of An. messeae. The DNA probes were designed based on gene sequences from the annotated An. atroparvus genome. The two nested inversions resulted in five syntenic blocks. Only two small syntenic blocks, which encompass 181 annotated genes in the An. atroparvus genome, changed their position and orientation in the X chromosome. The analysis of the An. atroparvus genome revealed an enrichment of gene ontology terms associated with immune system and mating behavior in the rearranged syntenic blocks. Additionally, the enrichment of DNA transposons was found in sequences homologous to three of the four breakpoint regions. This study demonstrates the successful application of the physical genome mapping approach to identify rearrangements that differentiate species in insects with polytene chromosomes. Full article

Information regarding Klebsiella aerogenes haboring carbapenemase in Japan is limited. A comprehensive nationwide survey was conducted from September 2014 to December 2022, and 67 non-duplicate strains of carbapenem-resistant K. aerogenes were isolated from 57 healthcare facilities in Japan. Through genetic testing and whole-genome sequencing, six strains were found to possess carbapenemases, including imipenemase (IMP)-1, IMP-6, New Delhi metallo-β-lactamase (NDM)-1, and NDM-5. The strain harboring bla_NDM-5 was the novel strain ST709, which belongs to the clonal complex of the predominant ST4 in China. The novel integron containing bla_IMP-1 featured the oxacillinase-101 gene, which is a previously unreported structure, with an IncN₄ plasmid type. However, integrons found in the strains possessing bla_IMP-6, which were the most commonly identified, matched those reported domestically in Klebsiella pneumoniae, suggesting the prevalence of identical integrons. Transposons containing bla_NDM are similar or identical to the transposon structure of K. aerogenes harboring bla_NDM-5 previously reported in Japan, suggesting that the same type of transposon could have been transmitted to K. aerogenes in Japan. This investigation analyzed mobile genetic elements, such as integrons and transposons, to understand the spread of carbapenemases, highlighting the growing challenge of carbapenem-resistant Enterobacterales in Japan and underscoring the critical need for ongoing surveillance to control these pathogens. Full article

Streptococcus pneumoniae, Streptococcus pyogenes (GAS), and Streptococcus agalactiae (GBS) are bacteria that can cause a range of infections, some of them life-threatening. This review examines the spread of antibiotic resistance and its mechanisms against antibiotics for streptococcal infections. Data on high-level penicillin-resistant invasive pneumococci have been found in Brazil (42.8%) and Japan (77%). The resistance is caused by mutations in genes that encode penicillin-binding proteins. Similarly, GAS and GBS strains reported from Asia, the USA, and Africa have undergone similar transformations in PBPs. Resistance to major alternatives of penicillins, macrolides, and lincosamides has become widespread among pneumococci and streptococci, especially in Asia (70–95%). The combination of several emm types with erm(B) is associated with the development of high-level macrolide resistance in GAS. Major mechanisms are ribosomal target modifications encoded by erm genes, ribosomal alterations, and active efflux pumps that regulate antibiotic entry due to mefA/E and msrD genes. Tetracycline resistance for streptococci in different countries varied from 22.4% in the USA to 83.7/100% in China, due to tet genes. Combined tetracycline/macrolide resistance is usually linked with the insertion of ermB into the transposon carrying tetM. New quinolone resistance is increasing by between 11.5 and 47.9% in Asia and Europe. The mechanism of quinolone resistance is based on mutations in gyrA/B, determinants for DNA gyrase, or parC/E encoding topoisomerase IV. The results for antibiotic resistance are alarming, and urgently call for increased monitoring of this problem and precautionary measures for control to prevent the spread of resistant mutant strains. Full article

Pathogenic Aeromonas spp. are the etiological agents of Motile Aeromonas Septicemia (MAS). This study aimed to identify the pathogen of diseased tadpoles (Quasipaa spinosa) and the antibiotic-resistance characteristics of this bacterium. A Gram-negative bacterium, named strain QST31, was isolated from the ascites of diseased tadpoles and was identified as Aeromonas media based on physiological and biochemical tests, as well as molecular identification. Artificial infection experiments showed that strain QST31 was highly virulent to tadpoles, with an LC₅₀ of 2.56 × 10⁷ CFU/mL. The antimicrobial susceptibility of strain QST31 was evaluated using the disk diffusion method, and the results indicated that strain QST31 was resistant to 28 antibacterial agents. In addition, the whole genome of strain QST31 was sequenced, and the presence of antimicrobial resistance genes, integron, and transposon was investigated. Genes involved in adherence, hemolysis, type II secretion system (T2SS), T6SS, iron uptake system, and quorum sensing were identified in the genome of strain QST31. More than 12 antimicrobial resistance genes were predicted in the genome of strain QST31. Interestingly, a novel Tn7709 transposon harboring sul1, aadA16, catB3, bla_OXA-21, aac(6′)-IIa, and tet(A) genes was identified. In conclusion, this is the first report on the isolation and identification of pathogenic A. media with multidrug resistance genes from diseased tadpoles. The results revealed that preventing and controlling aquatic animal diseases caused by multidrug resistance A. media will be a huge challenge in the future. Full article

Cancer therapies developed using bacteria and their components have been around since the 19th century. Compared to traditional cancer treatments, the use of bacteria-derived compounds as cancer therapeutics could offer a higher degree of specificity, with minimal off-target effects. Here, we explored the use of soluble bacteria-derived toxins as a potential squamous cell carcinoma (SCC) therapeutic. We optimized a protocol to generate Staphylococcus aureus biofilm-conditioned media (BCM), where soluble bacterial products enriched in the development of biofilms were isolated from a bacterial culture and applied to SCC cell lines. Bioactive components of S. aureus ATCC 29213 (SA29213) BCM display selective toxicity towards cancerous human skin SCC-12 at low doses, while non-cancerous human keratinocyte HaCaT and fibroblast BJ-5ta are minimally affected. SA29213 BCM treatment causes DNA damage to SCC-12 and initiates Caspase 3-dependent-regulated cell death. The use of the novel SA29213 bursa aurealis transposon mutant library led to the identification of S. aureus alpha hemolysin as the main bioactive compound responsible for the observed SCC-12-specific toxicity. The antibody neutralisation of Hla eradicates the cytotoxicity of SA29213 BCM towards SCC-12. Hla displays high SCC-12-specific toxicity, which is exerted primarily through Hla-ADAM10 interaction, Hla oligomerisation, and pore formation. The high target specificity and potential to cause cell death in a controlled manner highlight SA29213 Hla as a good candidate as an alternative SCC therapeutic. Full article