Search Results (169)

Quinoa (Chenopodium quinoa), a stress-tolerant pseudocereal ideal for studying abiotic stress responses, was used to systematically identify optimal reference genes for qPCR normalization under gradient stresses: low temperatures (LT group: −2 °C to −10 °C), heat (HT group: 39° C to 45 °C), and drought (DR group: 7 to 13 days). Through multi-algorithm evaluation (GeNorm, NormFinder, BestKeeper, the ΔCt method, and RefFinder) of eleven candidates, condition-specific optimal genes were established as ACT16 (Actin), SAL92 (IT4 phosphatase-associated protein), SSU32 (Ssu72-like family protein), and TSB05 (Tryptophan synthase beta-subunit 2) for the LT group; ACT16 and NRP13 (Asparagine-rich protein) for the HT group; and ACT16, SKP27 (S-phase kinase), and NRP13 for the DR group, with ACT16, NRP13, WLIM96 (LIM domain-containing protein), SSU32, SKP27, SAL92, and UBC22 (ubiquitin-conjugating enzyme E2) demonstrating cross-stress stability (global group). DHDPS96 (dihydrodipicolinate synthase) and EF03 (translation elongation factor) showed minimal stability. Validation using stress-responsive markers—COR72 (LT), HSP44 (HT), COR413-PM (LT), and DREB12 (DR)—confirmed reliability; COR72 and COR413-PM exhibited oscillatory cold response patterns, HSP44 peaked at 43 °C before declining, and DREB12 showed progressive drought-induced upregulation. Crucially, normalization with unstable genes (DHDPS96 and EF03) distorted expression profiles. This work provides validated reference standards for quinoa transcriptomics under abiotic stresses. Full article

Plant microbiota contributes to nutrient absorption, and the production of hormones and vitamins, and plays a crucial role in responding to environmental stress. We hypothesized that Vaccinium spp. harbour a unique microbiota that enables them to coexist in extreme environments such as saline, nutrient-poor, and waterlogged conditions. Upon examining Bacillus spp. endophytes isolated from blueberries, cranberries and lingonberries in vitro, we identified B. halotolerans (Bil-LT1_1, Bil-LT1_2) and B. velezensis (Cran-LT1_8, Ling-NOR4_15) strains that inhibit the growth of five pathogenic fungi and five foodborne bacteria. Whole-genome sequencing provided insights into genome organization and plasticity, helping identify mobile elements and genes potentially acquired through horizontal gene transfer. Functional annotation identified genes associated with plant colonization, stress tolerance, biocontrol activity, and plant growth promotion. Comparative genomic analyses revealed key biosynthetic gene clusters (BGCs) responsible for producing antifungal metabolites, including lipopeptides and polyketides. Genes supporting plant nutrition, growth, and environmental adaptation were present also in these strains. Notably, isolated endophytes exhibited particularly high levels of genomic plasticity, likely due to horizontal gene transfer involving gene ontology (GO) pathways related to survival in polymicrobial and foreign environments. Full article

GCN5-containing SAGA complex is evolutionarily conserved across yeast, plants, and humans and acts as a general transcription coactivator in the genome-wide regulation of genes. In Plasmodium falciparum, PfGCN5 forms a divergent complex, and the mis-localization of this complex by deleting the PfGCN5 bromodomain (ΔBrd) causes a plethora of growth defects. To directly test the PfGCN5 function, we performed conditional knockdown (KD) of PfGCN5. Whereas PfGCN5 KD phenotypically recapitulated the ΔBrd growth defects, it caused fewer transcriptional alterations compared to ΔBrd. To decipher the mechanism by which PfGCN5 regulates gene expression, we applied a new chromatin landscape analysis tool, CUT&Tag-seq, to map the chromatin localization of PfGCN5 and its deposited histone mark H3K9ac. Compared to ChIP-seq, CUT&Tag-seq identified substantially more H3K9ac peaks in the promoters of its target genes, with the peak intensity positively correlated with the levels of gene expression. CUT&Tag-seq analysis was remarkably more sensitive in mapping chromatin positions of PfGCN5, which colocalized with H3K9ac. The genes enriched with PfGCN5/H3K9ac signals at their promoters are involved in broad biological processes. Notably, PfGCN5′s positions overlapped with sequence motifs recognized by multiple apetela2 (AP2)-domain-containing transcription factors (AP2 TFs), suggesting that they recruited PfGCN5 to these promoters. Additionally, PfGCN5 was also colocalized with AP2-LT, further validating that AP2-LT is an integral component of the PfGCN5 complex. Collectively, these findings establish PfGCN5 as a master gene regulator in controlling general and parasite-specific cellular processes in this low-branching parasitic protist. Full article

Background/Objectives: Rising antibiotic resistance underscores the urgent need for effective vaccines against shigellosis. Our previous research identified the Shigella flexneri 2a truncated mutant (STM), a wzy gene knock-out strain cultivated in shake-flasks, as a promising broadly protective Shigella vaccine candidate. Expanding on this finding, our current study explores the feasibility of transitioning to a fermentor-grown STM as a vaccine candidate for further clinical development. Methods: The STM and STM-Cj, engineered to express the conserved Campylobacter jejuni N-glycan antigen, were grown in animal-free media, inactivated with formalin, and evaluated for key antigen retention and immunogenicity in mice. Results: The fermentor-grown STM exhibited significantly increased production yields and retained key antigens after inactivation. Immunization with the STM, particularly along with the double-mutant labile toxin (dmLT) adjuvant, induced robust immune responses to the conserved proteins IpaB, IpaC, and PSSP-1. Additionally, it provided protection against homologous and heterologous Shigella challenges in a mouse pulmonary model. The STM-Cj vaccine elicited antibody responses specific to the N-glycan while maintaining protective immune responses against Shigella. These findings underscore the potential of the fermentor-grown STM as a safe and immunogenic vaccine platform for combating shigellosis and possibly other gastrointestinal bacterial infections. Conclusions: Further process development to optimize growth and key antigen expression as well as expanded testing in additional animal models for the assessment of protection against Shigella and Campylobacter are needed to build on these encouraging initial results. Ultimately, clinical trials are essential to evaluate the efficacy and safety of STM-based vaccines in humans. Full article

Euphrates poplar (Populus euphratica) is known as a system model to study the genomic mechanisms underlying the salt resistance of woody species. To characterize how dynamic gene regulatory networks (GRNs) drive the defense response of this species to salt stress, we performed mRNA sequencing of P. euphratica roots under short-term (ST) and long-term (LT) salt stress treatments across multiple time points. Comparisons of these transcriptomes revealed the diverged gene expression patterns between the ST and LT treated samples. Based on the informative, dynamic, omnidirectional, and personalized networks model (idopNetwork), inter- and intra-module networks were constructed across different time points for both the ST and LT groups. Through the analysis of the inter-module network, we identified module 4 as the hub, containing the largest number of genes. Further analysis of the gene network within module 4 revealed that gene XM_011048240.1 had the most prominent interactions with other genes. Under short-term salt stress, gene interactions within the network were predominantly promoted, whereas under long-term stress, these interactions shifted towards inhibition. As for the gene ontology (GO) annotation of differentially expressed genes, the results suggest that P. euphratica may employ distinct response mechanisms during the early and late stages of salt stress. Taking together, these results offer valuable insights into the regulatory mechanism involved in P. euphratica’s stress response, advancing our understanding of complex biological processes. Full article

Background/Objectives: Antibiotic resistance is a serious public health problem threatening the treatment of infectious diseases caused by Escherichia coli, the main source of food contamination and responsible for many infectious diseases with high indices of AR profiles. Our objective was to study the presence of Escherichia coli in foods that are distributed and prepared on the street, characterizing its sensitivity profile and resistance to antibiotic drugs commonly prescribed in this geographical area. Methods: Standard procedures were performed to identify and isolate E. coli colonies from food samples collected during a three-year study. Susceptibility assays were conducted to determine the antibiotic resistance profile, and Colony PCR assays were performed to determine the pathogenic and antibiotic resistance genes. Results: A total of 189 food samples were collected, and 100% of the samples were positive for E. coli, with higher percentages of contamination for vegetables and fruits. ETEC (lt) and UPEC (vat, cnf1, hylA) genes were identified in 100% of the samples and DAEC (afa) in 27%. E. coli exhibited high percentages of resistance against ampicillin and amoxicillin/clavulanic acid (100%) and cephalexin (45%). The most effective antibiotics were tetracycline, TMP-SMX, polymyxin, and quinolones. The AR genes tetA, sul1, catA1, strA, qnrS, and floR were identified among the samples. Conclusions: Food prepared and marketed on the streets seriously threatens human health. Ampicillin and amoxicillin/clavulanic acid should not be used to treat infections caused by the multidrug-resistant ETEC and UPEC identified in this area. To our knowledge, this is the first study that explores the status of AR in this geographical area. Full article

As a common food-borne pathogen, Vibrio parahaemolyticus comes into direct or indirect contact with gastric acid after ingestion. However, the mechanisms by which Vibrio parahaemolyticus passes through the gastric acid barrier, recovers, and causes pathogenicity remain unclear. In this study, static in vitro digestion simulation experiments showed that some strains can pass through the gastric acid barrier by utilizing microacid tolerance mechanisms and altering their survival state. Food digestion simulation experiments showed that food matrices could help bacteria escape gastric acid stress, with significantly different survival rates observed for bacteria in various food matrices after exposure to gastric acid. Interestingly, surviving Vibrio parahaemolyticus showed a significantly shorter growth lag time (LT) during recovery. Transcriptome sequencing (RNA-seq) analyses indicated that the bacteria adapted to gastric acid stress by regulating the two-component system through stress proteins secreted via the ribosomal pathway. Pathogenic Vibrio parahaemolyticus that successfully passes through the gastric acid barrier potentially exhibits enhanced pathogenicity during recovery due to the significant upregulation of virulence genes such as tdh and yscF. This study provides a scientific basis for revealing the tolerance mechanisms of food-borne pathogens represented by Vibrio parahaemolyticus in the human body. Full article

Background/Objectives: Primary cutaneous B-cell lymphomas (PCBCL) are a rare and heterogeneous group of non-Hodgkin lymphomas that are confined to the skin at diagnosis and exhibit a tendency for cutaneous recurrence. The 5th edition of the World Health Organization and the 2022 International Consensus Classification recognize three main subtypes: primary cutaneous follicle center lymphoma (PCFCL), primary cutaneous marginal zone lymphoma/lymphoproliferative disorder (PCMZL/LPD), and primary cutaneous diffuse large B-cell lymphoma, leg type (PCDLBCL,LT). These subtypes differ in clinical behavior, histopathologic features, immunophenotype, and molecular alterations. Diagnosis and management remain challenging for clinicians. This review aims to provide a comprehensive overview of the defining features and current treatment strategies for PCBCL. Methods: This narrative review synthesizes current literature on the clinical, morphologic, immunohistochemical, and molecular characteristics of PCBCL. It also evaluates the diagnostic utility of immunohistochemistry, gene expression profiling, and molecular assays, particularly in distinguishing primary cutaneous disease from secondary cutaneous involvement by systemic lymphomas. Results: PCFCL arises from germinal center B-cells and must be differentiated from nodal follicular lymphoma. PCMZL/LPD is derived from post-germinal center B-cells and is often linked to chronic antigenic stimulation. Both PCFCL and PCMZL/LPD are indolent and associated with favorable outcomes. By contrast, PCDLBCL,LT is an aggressive lymphoma characterized by genetic alterations activating the NF-κB pathway, commonly including mutations to MYD88 and CD79B. Treatment strategies vary by subtype, ranging from localized therapies for indolent lymphomas to systemic chemoimmunotherapy for aggressive PCBCL. Emerging therapies, such as Bruton tyrosine kinase inhibitors and immunoregulatory agents, are being investigated for relapsed/refractory disease. Conclusions: PCBCL encompass distinct clinicopathologic entities with subtype-specific diagnostic and therapeutic considerations. While current management is guided by clinical behavior, significant knowledge gaps remain regarding the molecular mechanisms underlying skin tropism, immune evasion, and disease progression. Future research could focus on improving molecular characterization and developing personalized and immune-based therapies to enhance outcomes. This review consolidates current knowledge and highlights innovations aimed at advancing the diagnosis and treatment of PCBCL in clinical practice. Full article

Hepatoblastoma (HB) is the most common malignant liver tumor in children under five years of age. Although globally rare, it accounts for a large proportion of liver cancer in children and has poor survival rates in high-risk and metastatic cases. This review discusses the molecular mechanisms, diagnostic methods, and therapeutic strategies of HB. Mutations in the CTNNB1 gene and the activation of the Wnt/β-catenin pathway are essential genetic factors. Furthermore, genetic syndromes like Beckwith–Wiedemann syndrome (BWS) and Familial Adenomatous Polyposis (FAP) considerably heighten the risk of associated conditions. Additionally, epigenetic mechanisms, such as DNA methylation and the influence of non-coding RNAs (ncRNAs), are pivotal drivers of tumor development. Diagnostics include serum biomarkers, immunohistochemistry (IHC), and imaging techniques. Standard treatments are chemotherapy, surgical resection, and liver transplantation (LT). Emerging therapies like immunotherapy and targeted treatments offer hope against chemotherapy resistance. Future research will prioritize personalized medicine, novel biomarkers, and molecular-targeted therapies to improve survival outcomes. Full article

Nitric oxide (NO) functions as a signaling molecule in plant adaptation to changing environmental conditions. NO levels were found to increase in plants in response to low temperatures (LTs). However, knowledge of the pathways involved in enhanced NO production under cold stress is still limited. For this reason, we aimed to determine the role of different NO sources in NO generation in cucumber roots exposed to 10 °C for short (1 d) and long (6 d) periods. The short-term treatment of seedlings with LT markedly increased plasma membrane-bound nitrate reductase (PM-NR) activity and induced the expression of three genes encoding NR in cucumber (CsNR1-3). On the other hand, long-term exposure was related to both increased cytoplasmic NR (cNR) activity and induced expression of the CsARC gene, encoding the amidoxime-reducing component (ARC) protein. The decrease in nitrite reductase (NiR) activity and the higher NO₂⁻/NO₃⁻ ratio in the roots of plants exposed to LTs for 1 d suggest that tissue conditions may favor NR-dependent NO production. Regardless of NR stimulation, a significant increase in NOS-like activity was observed in the roots, especially during the long-term treatment of plants with LT. These results indicate that diverse NO-producing routes, both reductive and oxidative, are activated in cucumber tissues at different stages of cold stress. Full article

Enterotoxigenic Escherichia coli (ETEC) produces two types of enterotoxins, LTs and STs, as well as several colonization factors (CFs), including CS21, CS3 fimbriae, and flagellar structures. This study investigated how these structures contribute to ETEC colonization and the immune response in HT-29 and HuTu-80 intestinal cells. ETEC strains with single, double, and triple mutations in the lngA, cstH, and fliC genes were generated and confirmed using PCR and Western blotting. The colonization of HT-29 and HuTu-80 intestinal cells by the ETEC E9034A strain, which was fully sequenced using a hybrid approach involving both Illumina and Oxford Nanopore technologies, was used to generate the mutant and recombinant proteins. The colonization and adherence of E9034A and its mutants were assessed through colony-forming unit (CFU) counts. Cytokine levels were assessed using flow cytometry and analyzed via FlowJo 7.6.1. Quantitative analysis revealed that the absence of the lngA, cstH, and fliC genes significantly (p < 0.01) reduced ETEC adherence to HT-29 and HutU-80 cells. In addition, only ETEC strains expressing the FliC protein induced IL-8 secretion. These findings suggest that LngA, CstH, and FliC in ETEC E9034A enhance adherence to intestinal cells and trigger the release of IL-8. Full article

Walnuts are among the globally significant woody food and oil tree species. At high latitudes, they frequently experience late-frost damage, inducing low-temperature stress, which significantly affects walnut seedlings. The aim of this study was to investigate the physiological and biochemical alterations in walnut seedlings under low-temperature (LT) stress along with its underlying molecular mechanisms. Physiological indices were determined, and the transcriptome and miRNA were sequenced by sampling leaves (0 h, 6 h, 12 h, 24 h, and 48 h) of two-month-old live seedlings of walnuts treated with a low temperature of 4 °C. The results indicated that LT stress induced an increase in electrical conductivity and malondialdehyde content while simultaneously causing a reduction in Fv/Fm. From the transcriptome comparison between the control and treated groups, a total of 12,566 differentially expressed genes (DEGs) were identified, consisting of 6829 up-regulated and 5737 down-regulated genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the DEGs were primarily enriched in polysaccharide metabolic processes, responses to abscisic acid and phenylpropanoid biosynthesis pathways. Furthermore, the miRNA database identified 1052 miRNAs in response to low-temperature stress in walnuts; these miRNAs were found to target 7043 predicted genes. Through the integration and analysis of transcriptome and miRNA data, 244 differential DEGs were identified. Following GO and KEGG enrichment analyses of the differential target genes, we identified that these genes primarily regulate pathways involved in starch and sucrose metabolism, glyoxylate and dicarboxylate metabolism, and glycerophospholipid biosynthesis, as well as phenylalanine, tyrosine, and tryptophan biosynthesis, in walnut leaves under LT stress. Additionally, we conducted an in-depth analysis of the associations between differentially expressed genes (DEGs) and differentially expressed microRNAs (DEMs) within the starch and sucrose metabolism pathway. Real-time fluorescent quantitative PCR (qRT-PCR) validation of the expression patterns of a subset of differential genes confirmed the accuracy of the transcriptome data. This study unveils the potential molecular mechanisms underlying walnut’s response to low-temperature stress, providing valuable genetic resources for future research on the cold tolerance mechanisms of walnut in response to late-frost damage. Full article

The clustered, regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) system is the most widely used gene-editing tool to date. However, its application in the genetic improvement of forestry trees has been largely limited. Here, we first established a highly efficient multi-target editing system in the magnoliid woody plant Liriodendron tulipifera. Using phytoene desaturase gene (PDS) as an example, we systematically compared CRISPR/Cas9 and CRSPR/Cpf1 expression systems for loss-of-function analysis and conducted genetic transformations using transient and stable transformation. Ultimately, our findings indicated that the CRISPR/Cas9 system, when applied to transformation based on single-cell-originated somatic embryogenesis, yielded the highest gene-editing efficiency, with mutation rates of nearly 100%. Furthermore, we obtained a total of 137 regeneration plantlets via somatic embryogenesis, of which 82.48% exhibited an albino phenotype. The Illumina sequencing results of albino seedlings and the callus tissue obtained from dedifferentiation of mutant plants revealed that the mutation at the T1 target site was homozygous. These results indicate that CRISPR/Cas9-based multiplex genome-editing technology can not only accelerate the identification of gene function but also be incorporated into the genetic improvement and breeding of tulip trees, supporting the scale propagation of genome-edited plantlets via somatic embryogenesis. Full article

Low-temperature (LT) stress seriously affects the distribution, seedling survival, and grain yield of maize. At the seedling emergence stage, maize’s coleoptile is one of the most sensitive organs in sensing LT signaling and, in general, it can envelop young leaves to protect them from LT damage. In addition, brassinolides (BRs) have been shown to enhance LT tolerance from various species, but the effects of BRs on coleoptiles in maize seedlings under LT stress are unclear. Therefore, in this study, the pre-cultured coleoptiles of Zheng58 seedlings were treated with or without 2.0 μM 24-epibrassinolide (EBR) at 25 °C and 10 °C environments for five days to analyze their physiological and transcriptomic changes. Physiological analysis showed that a 10°C LT stress increased the content of glucose (0.43 mg g⁻¹ FW), sucrose (0.45 mg g⁻¹ FW), and starch (0.76 mg g⁻¹ FW) of Zheng58 coleoptiles compared to a 25°C environment. After the coleoptiles were exposed to a 2.0 μM EBR application under 10°C temperature for five days, the contents of these three sugars continued to increase, and reached 2.68 mg g⁻¹ FW, 4.64 mg g⁻¹ FW, and 9.27 mg g⁻¹ FW, respectively, indicating that sugar signaling and metabolism played key roles in regulating LT tolerance in the coleoptiles of maize seedlings. Meanwhile, a transcriptome analysis showed that 84 and 15 differentially expressed genes (DEGs) were enriched in the sucrose and starch metabolism and photosynthesis pathways, respectively, and multiple DEGs involved in these pathways were significantly up-regulated under LT stress and EBR stimulation. Further analysis speculated that the four DEGs responsible for sucrose-phosphate synthetase (SPS, i.e., Zm00001d048979, probable sucrose-phosphate synthase 5 and Zm00001d012036, sucrose-phosphate synthase 1), sucrose synthase (SUS, Zm00001d029091, sucrose synthase 2 and Zm00001d029087, sucrose synthase 4) were crucial nodes that could potentially link photosynthesis and other unknown pathways to form the complex interaction networks of maize LT tolerance. In conclusion, our findings provide new insights into the molecular mechanisms of exogenous EBR in enhancing LT tolerance of maize seedlings and identified potential candidate genes to be used for LT tolerance breeding in maize. Full article

Lasiodiplodia theobromae is a pathogenic fungus associated with tropical perennial fruit plants worldwide. In apple trees, L. theobromae causes dieback and canker, a disease that affects the architecture of the wood producing the progressive death of branches and stems, from the tips to the base, invading the vascular tissue, manifesting necrotic lesions in the bark, impeding the flow of nutrients and water. The present work reports the whole genome de novo sequencing (WGS) of L. theobromae strain Bot-2018-LT45 isolated from apple trees with dieback symptoms. Genomic DNA of L. theobromae was sequenced using Illumina paired-end short-read technology (NovaSeq6000) and PacBio SMRTbell^TM (Single Molecule, Real-Time) long-read technology. The genome size was 44.17 Mb. Then, assembly and annotation revealed a total of 12,948 genes of which 11,634 encoded proteins. The genome was assembled into 34 contigs with an N50 (Mb) value of 3.23. This study is the first report of the L. theobromae genome de novo obtained from apple trees with dieback and canker symptoms in the Maule Region, Chile. This genetic information may set the basis for future study of the mechanisms of L. theobromae and establish the possibility of specific molecular improvements for the control of dieback and canker. Full article