Enhanced stress resilience in potato by deletion of Parakletos.

Continued climate change impose multiple stressors on crops, including pathogens, salt, and drought, severely impacting agricultural productivity. Innovative solutions are necessary to develop resilient crops. Here, using quantitative potato proteomics, we identify Parakletos, a thylakoid protein that contributes to disease susceptibility. We show that knockout or silencing of Parakletos enhances resistance to oomycete, fungi, bacteria, salt, and drought, whereas its overexpression reduces resistance. In response to biotic stimuli, Parakletos-overexpressing plants exhibit reduced amplitude of reactive oxygen species and Ca2+ signalling, and silencing Parakletos does the opposite. Parakletos homologues have been identified in all major crops. Consecutive years of field trials demonstrate that Parakletos deletion enhances resistance to Phytophthora infestans and increases yield. These findings demark a susceptibility gene, which can be exploited to enhance crop resilience towards abiotic and biotic stresses in a low-input agriculture.

A new bacterial consortia for management of Fusarium head blight in wheat.

Fusarium head blight (FHB) is a significantly important disease in cereals primarily caused by Fusarium species. FHB control is largely executed through chemical strategies, which are costlier to sustainable wheat production, resulting in leaning towards sustainable sources such as resistance breeding and biological control methods for FHB. The present investigation was aimed at evaluating newly identified bacterial consortium (BCM) as biocontrol agents for FHB and understanding the morpho-physiological traits associated with the disease resistance of spring wheat. Preliminary evaluation through antagonistic plate assay and in vivo assessment indicated that BCM effectively inhibited Fusarium growth in spring wheat, reducing area under disease progress curve (AUDPC) and deoxynivalenol (DON), potentially causing type II and V resistance, and improving single spike yield (SSPY). Endurance to FHB infection with the application of BCM is associated with better sustenance of spike photosynthetic performance by improving the light energy harvesting and its utilization. Correlation and path-coefficient analysis indicated that maximum quantum yield (QY_max) is directly influencing the improvement of SSPY and reduction of grain DON accumulation, which is corroborated by principal component analysis. The chlorophyll fluorescence traits identified in the present investigation might be applied as a phenotyping tool for the large-scale identification of wheat sensitivity to FHB.

Genomic selection in plant breeding: Key factors shaping two decades of progress.

Genomic selection, the application of genomic prediction (GP) models to select candidate individuals, has significantly advanced in the past two decades, effectively accelerating genetic gains in plant breeding. This article provides a holistic overview of key factors that have influenced GP in plant breeding during this period. We delved into the pivotal roles of training population size and genetic diversity, and their relationship with the breeding population, in determining GP accuracy. Special emphasis was placed on optimizing training population size. We explored its benefits and the associated diminishing returns beyond an optimum size. This was done while considering the balance between resource allocation and maximizing prediction accuracy through current optimization algorithms. The density and distribution of single-nucleotide polymorphisms, level of linkage disequilibrium, genetic complexity, trait heritability, statistical machine-learning methods, and non-additive effects are the other vital factors. Using wheat, maize, and potato as examples, we summarize the effect of these factors on the accuracy of GP for various traits. The search for high accuracy in GP-theoretically reaching one when using the Pearson's correlation as a metric-is an active research area as yet far from optimal for various traits. We hypothesize that with ultra-high sizes of genotypic and phenotypic datasets, effective training population optimization methods and support from other omics approaches (transcriptomics, metabolomics and proteomics) coupled with deep-learning algorithms could overcome the boundaries of current limitations to achieve the highest possible prediction accuracy, making genomic selection an effective tool in plant breeding.

A thermostable organic solvent-tolerant lipase from Brevibacillus sp.: production and integrated downstream processing using an alcohol-salt-based aqueous two-phase system.

Lipases are used for the synthesis of different compounds in the chemical, pharmaceutical, and food industries. Most of the reactions are carried out in non-aqueous media and often at elevated temperature, requiring the use of organic solvent-tolerant thermostable lipases. However, most known lipases are not stable in the presence of organic solvents and at elevated temperature. In this study, an organic solvent-tolerant thermostable lipase was obtained from Brevibacillus sp. SHI-160, a moderate thermophile isolated from a hot spring in the East African Rift Valley. The enzyme was optimally active at 65°C and retained over 90% of its activity after 1 h of incubation at 70°C. High lipase activity was measured in the pH range of 6.5 to 9.0 with an optimum pH of 8.5. The enzyme was stable in the presence of both polar and non-polar organic solvents. The stability of the enzyme in the presence of polar organic solvents allowed the development of an efficient downstream processing using an alcohol-salt-based aqueous two-phase system (ATPS). Thus, in the presence of 2% salt, over 98% of the enzyme partitioned to the alcohol phase. The ATPS-recovered enzyme was directly immobilized on a solid support through adsorption and successfully used to catalyze a transesterification reaction between paranitrophenyl palmitate and short-chain alcohols in non-aqueous media. This shows the potential of lipase SHI-160 to catalyze reactions in non-aqueous media for the synthesis of valuable compounds. The integrated approach developed for enzyme production and cheap and efficient downstream processing using ATPS could allow a significant reduction in enzyme production costs. The results also show the potential of extreme environments in the East African Rift Valley as sources of valuable microbial genetic resources for the isolation of novel lipases and other industrially important enzymes.

Unlocking the Potential of the CA2, CA7, and ITM2C Gene Signatures for the Early Detection of Colorectal Cancer: A Comprehensive Analysis of RNA-Seq Data by Utilizing Machine Learning Algorithms.

Colorectal cancer affects the colon or rectum and is a common global health issue, with 1.1 million new cases occurring yearly. The study aimed to identify gene signatures for the early detection of CRC using machine learning (ML) algorithms utilizing gene expression data. The TCGA-CRC and GSE50760 datasets were pre-processed and subjected to feature selection using the LASSO method in combination with five ML algorithms: Adaboost, Random Forest (RF), Logistic Regression (LR), Gaussian Naive Bayes (GNB), and Support Vector Machine (SVM). The important features were further analyzed for gene expression, correlation, and survival analyses. Validation of the external dataset GSE142279 was also performed. The RF model had the best classification accuracy for both datasets. A feature selection process resulted in the identification of 12 candidate genes, which were subsequently reduced to 3 (CA2, CA7, and ITM2C) through gene expression and correlation analyses. These three genes achieved 100% accuracy in an external dataset. The AUC values for these genes were 99.24%, 100%, and 99.5%, respectively. The survival analysis showed a significant logrank p-value of 0.044 for the final gene signatures. The analysis of tumor immunocyte infiltration showed a weak correlation with the expression of the gene signatures. CA2, CA7, and ITM2C can serve as gene signatures for the early detection of CRC and may provide valuable information for prognostic and therapeutic decision making. Further research is needed to fully understand the potential of these genes in the context of CRC.

Comparative transcriptome profiling provides insights into the growth promotion activity of Pseudomonas fluorescens strain SLU99 in tomato and potato plants.

The use of biocontrol agents with plant growth-promoting activity has emerged as an approach to support sustainable agriculture. During our field evaluation of potato plants treated with biocontrol rhizobacteria, four bacteria were associated with increased plant height. Using two important solanaceous crop plants, tomato and potato, we carried out a comparative analysis of the growth-promoting activity of the four bacterial strains: Pseudomonas fluorescens SLU99, Serratia plymuthica S412, S. rubidaea AV10, and S. rubidaea EV23. Greenhouse and in vitro experiments showed that P. fluorescens SLU99 promoted plant height, biomass accumulation, and yield of potato and tomato plants, while EV23 promoted growth in potato but not in tomato plants. SLU99 induced the expression of plant hormone-related genes in potato and tomato, especially those involved in maintaining homeostasis of auxin, cytokinin, gibberellic acid and ethylene. Our results reveal potential mechanisms underlying the growth promotion and biocontrol effects of these rhizobacteria and suggest which strains may be best deployed for sustainably improving crop yield.

Potential role of the regulatory miR1119-MYC2 module in wheat (Triticum aestivum L.) drought tolerance.

MicroRNA (miRNA)-target gene modules are essential components of plants' abiotic stress signalling pathways Little is known about the drought-responsive miRNA-target modules in wheat, but systems biology approaches have enabled the prediction of these regulatory modules and systematic study of their roles in responses to abiotic stresses. Using such an approach, we sought miRNA-target module(s) that may be differentially expressed under drought and non-stressed conditions by mining Expressed Sequence Tag (EST) libraries of wheat roots and identified a strong candidate (miR1119-MYC2). We then assessed molecular and physiochemical differences between two wheat genotypes with contrasting drought tolerance in a controlled drought experiment and assessed possible relationships between their tolerance and evaluated traits. We found that the miR1119-MYC2 module significantly responds to drought stress in wheat roots. It is differentially expressed between the contrasting wheat genotypes and under drought versus non-stressed conditions. We also found significant associations between the module's expression profiles and ABA hormone content, water relations, photosynthetic activities, H2O2 levels, plasma membrane damage, and antioxidant enzyme activities in wheat. Collectively, our results suggest that a regulatory module consisting of miR1119 and MYC2 may play an important role in wheat's drought tolerance.

Metabolic Processes and Biological Macromolecules Defined the Positive Effects of Protein-Rich Biostimulants on Sugar Beet Plant Development.

Protein-based biostimulants (PBBs) have a positive effect on plant development, although the biological background for this effect is not well understood. Here, hydrolyzed wheat gluten (HWG) and potato protein film (PF) in two levels (1 and 2 g/kg soil) and in two different soils (low and high nutrient; LNC and HNC) were used as PBBs. The effect of these PBBs on agronomic traits, sugars, protein, and peptides, as well as metabolic processes, were evaluated on sugar beet in comparison with no treatment (control) and treatment with nutrient solution (NS). The results showed a significant growth enhancement of the plants using HWG and PF across the two soils. Sucrose and total sugar content in the roots were high in NS-treated plants and correlated to root growth in HNC soil. Traits related to protein composition, including nitrogen, peptide, and RuBisCO contents, were enhanced in PBB-treated plants (mostly for HWG and PF at 2 g/kg soil) by 100% and >250% in HNC and LNC, respectively, compared to control. The transcriptomic analysis revealed that genes associated with ribosomes and photosynthesis were upregulated in the leaf samples of plants treated with either HWG or PP compared to the control. Furthermore, genes associated with the biosynthesis of secondary metabolites were largely down-regulated in root samples of HWG or PF-treated plants. Thus, the PBBs enhanced protein-related traits in the plants through a higher transcription rate of genes related to protein- and photosynthesis, which resulted in increased plant growth, especially when added in certain amounts (2 g/kg soil). However, sucrose accumulation in the roots of sugar beet seemed to be related to the easy availability of nitrogen.

Inbreeding Effects on the Performance and Genomic Prediction for Polysomic Tetraploid Potato Offspring Grown at High Nordic Latitudes.

Inbreeding depression (ID) is caused by increased homozygosity in the offspring after selfing. Although the self-compatible, highly heterozygous, tetrasomic polyploid potato (Solanum tuberosum L.) suffers from ID, some argue that the potential genetic gains from using inbred lines in a sexual propagation system of potato are too large to be ignored. The aim of this research was to assess the effects of inbreeding on potato offspring performance under a high latitude and the accuracy of the genomic prediction of breeding values (GEBVs) for further use in selection. Four inbred (S1) and two hybrid (F1) offspring and their parents (S0) were used in the experiment, with a field layout of an augmented design with the four S0 replicated in nine incomplete blocks comprising 100, four-plant plots at Umeå (63°49'30″ N 20°15'50″ E), Sweden. S0 was significantly (p < 0.01) better than both S1 and F1 offspring for tuber weight (total and according to five grading sizes), tuber shape and size uniformity, tuber eye depth and reducing sugars in the tuber flesh, while F1 was significantly (p < 0.01) better than S1 for all tuber weight and uniformity traits. Some F1 hybrid offspring (15-19%) had better total tuber yield than the best-performing parent. The GEBV accuracy ranged from -0.3928 to 0.4436. Overall, tuber shape uniformity had the highest GEBV accuracy, while tuber weight traits exhibited the lowest accuracy. The F1 full sib's GEBV accuracy was higher, on average, than that of S1. Genomic prediction may facilitate eliminating undesired inbred or hybrid offspring for further use in the genetic betterment of potato.

RNA silencing proteins and small RNAs in oomycete plant pathogens and biocontrol agents.

Introduction: Oomycetes cause several damaging diseases of plants and animals, and some species also act as biocontrol agents on insects, fungi, and other oomycetes. RNA silencing is increasingly being shown to play a role in the pathogenicity of Phytophthora species, either through trans-boundary movement of small RNAs (sRNAs) or through expression regulation of infection promoting effectors. Methods: To gain a wider understanding of RNA silencing in oomycete species with more diverse hosts, we mined genome assemblies for Dicer-like (DCL), Argonaute (AGO), and RNA dependent RNA polymerase (RDRP) proteins from Phytophthora plurivora, Ph. cactorum, Ph. colocasiae, Pythium oligandrum, Py. periplocum, and Lagenidium giganteum. Moreover, we sequenced small RNAs from the mycelium stage in each of these species. Results and discussion: Each of the species possessed a single DCL protein, but they differed in the number and sequence of AGOs and RDRPs. SRNAs of 21nt, 25nt, and 26nt were prevalent in all oomycetes analyzed, but the relative abundance and 5' base preference of these classes differed markedly between genera. Most sRNAs mapped to transposons and other repeats, signifying that the major role for RNA silencing in oomycetes is to limit the expansion of these elements. We also found that sRNAs may act to regulate the expression of duplicated genes. Other sRNAs mapped to several gene families, and this number was higher in Pythium spp., suggesting a role of RNA silencing in regulating gene expression. Genes for most effector classes were the source of sRNAs of variable size, but some gene families showed a preference for specific classes of sRNAs, such as 25/26 nt sRNAs targeting RxLR effector genes in Phytophthora species. Novel miRNA-like RNAs (milRNAs) were discovered in all species, and two were predicted to target transcripts for RxLR effectors in Ph. plurivora and Ph. cactorum, indicating a putative role in regulating infection. Moreover, milRNAs from the biocontrol Pythium species had matches in the predicted transcriptome of Phytophthora infestans and Botrytis cinerea, and L. giganteum milRNAs matched candidate genes in the mosquito Aedes aegypti. This suggests that trans-boundary RNA silencing may have a role in the biocontrol action of these oomycetes.

Biological management of coffee wilt disease (Fusarium xylarioides) using antagonistic Trichoderma isolates.

Coffee wilt disease (CWD) is a serious threat to the food security of small-scale farmers in Ethiopia, causing significant reductions in coffee yield. Currently, there are no effective control measures available against the causative agent of CWD, Fusarium xylarioides. The main objective of this study was therefore to develop, formulate, and evaluate a range of biofungicides against F. xylarioides, derived from Trichoderma species and tested under in vitro, greenhouse, and field conditions. In total, 175 Trichoderma isolates were screened as microbial biocontrol agents against F. xylarioides. The efficacy of two biofungicide formulations, wettable powder and water dispensable granules, were tested on the susceptible Geisha coffee variety in three different agro-ecological zones in southwestern Ethiopia over three years. The greenhouse experiments were set up using a complete block design, while in the field a randomized complete block design was used, with twice yearly applications of biofungicide. The test pathogen spore suspension was applied to the coffee seedlings by soil drenching, and the subsequent incidence and severity of CWD evaluated annually. The mycelial growth inhibition profiles of the Trichoderma isolates against F. xylarioides ranged from 44.5% to 84.8%. In vitro experiments revealed that T. asperelloides AU71, T. asperellum AU131 and T. longibrachiatum AU158 reduced the mycelial growth of F. xylarioides by over 80%. The greenhouse study indicated that wettable powder (WP) of T. asperellum AU131 had the highest biocontrol efficacy (84.3%), followed by T. longibrachiatum AU158 (77.9%) and T. asperelloides AU71 (71.2%); they also had a significant positive impact on plant growth. The pathogen-treated control plants had a disease severity index of 100% across all the field experiments, and of 76.7% in the greenhouse experiments. In comparison to untreated controls, the annual and cumulative disease incidence over the three years of the study period varied from 46.2 to 90%, 51.6 to 84.5%, and 58.2 to 91%, at the Teppi, Gera and Jimma field experimental locations. Overall, the greenhouse and field experiments and in vitro assays support the biocontrol potential of Trichoderma isolates, and T. asperellum AU131 and T. longibrachiatum AU158 in particular are recommended for the management of CWD under field conditions.

Delineation of Genotype X Environment Interaction for Grain Yield in Spring Barley under Untreated and Fungicide-Treated Environments.

Barley (Hordeul vulgare L.) is the fourth most important cereal crop based on production and cultivated area. Biotic stresses, especially fungal diseases in barley, are devastating, incurring high possibilities of absolute yield loss. Identifying superior and stable yielding genotypes is crucial for accompanying the increasing barley demand. However, the identification and recommendation of superior genotypes is challenging due to the interaction between genotype and environment. Hence, the present investigation was aimed at evaluating the grain yield of different sets of spring barley genotypes when undergoing one of two treatments (no treatment and fungicide treatment) laid out in an alpha lattice design in six to seven locations for five years, through additive main effects and multiplicative interaction (AMMI), GGE biplot (genotype + genotype X environment), and stability analysis. The combined analysis of variance indicated that the environment was the main factor that contributed to the variation in grain yield, followed by genotype X environment interaction (GEI) effects and genotypic effects. Ten mega environments (MEs) with five MEs from each of the treatments harboured well-adapted, stable yielding genotypes. Exploiting the stable yielding genotypes with discreet use of the representative and discriminative environments identified in the present study could aid in breeding for the improvement of grain yield in spring barley genotypes.

Antagonistic and plant growth promotion of rhizobacteria against Phytophthora colocasiae in taro.

Taro leaf blight caused by Phytophthora colocasiae adversely affects the growth and yield of taro. The management of this disease depends heavily on synthetic fungicides. These compounds, however, pose potential hazards to human health and the environment. The present study aimed to investigate an alternative approach for plant growth promotion and disease control by evaluating seven different bacterial strains (viz., Serratia plymuthica, S412; S. plymuthica, S414; S. plymuthica, AS13; S. proteamaculans, S4; S. rubidaea, EV23; S. rubidaea, AV10; Pseudomonas fluorescens, SLU-99) and their different combinations as consortia against P. colocasiae. Antagonistic tests were performed in in vitro plate assays and the effective strains were selected for detached leaf assays and greenhouse trials. Plant growth-promoting and disease prevention traits of selected bacterial strains were also investigated in vitro. Our results indicated that some of these strains used singly (AV10, AS13, S4, and S414) and in combinations (S4+S414, AS13+AV10) reduced the growth of P. colocasiae (30-50%) in vitro and showed disease reduction ability when used singly or in combinations as consortia in greenhouse trials (88.75-99.37%). The disease-suppressing ability of these strains may be related to the production of enzymes such as chitinase, protease, cellulase, and amylase. Furthermore, all strains tested possessed plant growth-promoting traits such as indole-3-acetic acid production, siderophore formation, and phosphate solubilization. Overall, the present study revealed that bacterial strains significantly suppressed P. colocasiae disease development using in vitro, detached leaf, and greenhouse assays. Therefore, these bacterial strains can be used as an alternative strategy to minimize the use of synthetic fungicides and fertilizers to control taro blight and improve sustainable taro production.

Finger millet RNA-seq reveals differential gene expression associated with tolerance to aluminum toxicity and provides novel genomic resources.

Eleusine coracana, finger millet, is a multipurpose crop cultivated in arid and semi-arid regions of Africa and Asia. RNA sequencing (RNA-seq) was used in this study to obtain valuable genomic resources and identify genes differentially expressed between Al-tolerant and Al-susceptible genotypes. Two groups of finger millet genotypes were used: Al-tolerant (215836, 215845, and 229722) and Al-susceptible (212462, 215804 and 238323). The analysis of the RNA-seq data resulted in 198,546 unigenes, 56.5% of which were annotated with significant hits in one or more of the following six databases: NR (48.8%), GO (29.7%), KEGG (45%), PlantTFDB (19.0%), Uniprot (49.2%), and NT (46.2%). It is noteworthy that only 220 unigenes in the NR database had significant hits against finger millet sequences suggesting that finger millet's genomic resources are scarce. The gene expression analysis revealed that 322 genes were significantly differentially expressed between the Al-tolerant and Al-susceptible genotypes, of which 40.7% were upregulated while 59.3% were downregulated in Al-tolerant genotypes. Among the significant DEGs, 54.7% were annotated in the GO database with the top hits being ATP binding (GO:0005524) and DNA binding (GO:0003677) in the molecular function, DNA integration (GO:0015074) and cell redox homeostasis in the biological process, as well as cellular anatomical entity and intracellular component in the cellular component GO classes. Several of the annotated DEGs were significantly enriched for their corresponding GO terms. The KEGG pathway analysis resulted in 60 DEGs that were annotated with different pathway classes, of which carbohydrate metabolism and signal transduction were the most prominent. The homologs of a number of significant DEGs have been previously reported as being associated with Al or other abiotic stress responses in various crops, including carboxypeptidase SOL1, HMA3, AP2, bZIP, C3H, and WRKY TF genes. A more detailed investigation of these and other DEGs will enable genomic-led breeding for Al tolerance in finger millet. RNA-seq data analysis also yielded 119,073 SNP markers, the majority of which had PIC values above 0.3, indicating that they are highly informative. Additionally, 3,553 single-copy SSR markers were identified, of which trinucleotide SSRs were the most prevalent. These genomic resources contribute substantially to the enrichment of genomic databases for finger millet, and facilitate future research on this crop.

Antifungal compounds, GC-MS analysis and toxicity assessment of methanolic extracts of Trichoderma species in an animal model.

Fungi of the genus Trichoderma have been marketed for the management of diseases of crops. However, some Trichoderma species may produce toxic secondary metabolites and it should receive due attention to ensure human safety. In this study, we investigated the in vitro antagonistic potential of T. asperellum AU131 and T. longibrachiatum AU158 as microbial biocontrol agents (MBCAs) against Fusarium xylarioides and the associated antagonistic mechanism with bioactive substances. Swiss albino mice were used to evaluate the in vivo toxicity and pathogenicity of T. asperellum AU131 and T. longibrachiatum AU158 methanolic extracts and spore suspensions, respectively, in a preliminary safety assessment for use as biofungicides. Gas Chromatography-Mass Spectrometry (GC-MS) was used to profile volatile organic metabolites (VOCs) present in the methanolic extracts. The agar diffusion assay of the methanolic extracts from both T. asperellum AU131 and T. longibrachiatum AU158 were effective at a concentration of 200 µg/mL (1×107 spores/mL), causing 62.5%, and 74.3% inhibition, respectively. A GC-MS analysis of methanolic extracts from both bioagents identified 23 VOCs which classified as alcohols, acids, sesquiterpenes, ketones and aromatic compounds. The oral administration of methanolic extracts and spore suspensions of each Trichoderma species to female Swiss albino mice over 14 days did not show any significant signs of toxicity, mortality or changes to body weight. It can be concluded that the tested spore suspensions and methanolic extracts were not pathogenic or toxic, respectively, when administered to Swiss albino mice at various doses.

A Quantitative Luminol-Based Assay for ROS Burst Detection in Potato Leaves in Response to Biotic Stimuli.

Reactive oxygen species (ROS) are important signaling agents in plants and animals. They are involved in diverse processes, including activation of immune responses to pathogen infection. Biphasic detection of ROS in response to pathogen perception is becoming more popular even in important crops like potato as means of screening different germ plasms and mutants generated by for example CRISPR-Cas9 as well as identifying signaling pathways. Here we describe a detailed protocol for quantifying ROS bursts induced in potato leaf discs in response to a bacterial elicitor and Phytophthora infestans.

A Detached Leaf Assay for Rapidly Screening Plant Pathogen-Biological Control Agents.

The detached leaf assay (DLA) is a nondestructive method for evaluating interactions between plants and disease-causing agents that allows quick characterization of potential pathogens' infectivity and plants' resistance to them. Here we show its utility for also assessing potential biological control agents (BCAs), by demonstrating its applicability for screening potential BCAs for the strawberry (Fragaria × ananassa) pathogen Phytophtora cactorum.