Pathogen Phytophthora Infestans Research Articles

Study of resistance to Phytophtora infestans of potato breeding material using DNA markers

Potato late blight, caused by the oomycete Phytophthora infestans, is one of the most harmful diseases of the crop. Nightshade crops suffer the most from it, with potatoes (Solanum tuberosum) taking the first place. Late blight causes serious damage to potatoes by sapping out nutrients from the root tissue. The development of molecular marker technology has had a great impact on plant breeding. Various types of molecular markers have been developed, and sequencing technologies have contributed to the improvement of agricultural crops. Late blight resistance based on the R resistance gene is the most effective method for suppressing infection by the pathogen Phytophthora infestans. Using SCAR markers of R genes, it is possible to identify potential donors of resistance genes, facilitate the selection of resistant plants in segregating populations, and determine the preservation of resistance genes after multiple crossings and during long-term cultivation of potato varieties.

Potato β-aminobutyric acid receptor IBI1 manipulates VOZ1 and VOZ2 transcription factor activity to promote disease resistance.

Upon infection with non-pathogenic microorganisms or treatment with natural or synthetic compounds, plants exhibit a more rapid and potent response to both biotic and abiotic stresses. However, the molecular mechanisms behind this phenomenon, known as defense priming, are poorly understood. β-aminobutyric acid (BABA) is an endogenous stress metabolite that enhances plant tolerance to various abiotic stresses and primes plant defense responses, providing the ability to resist a variety of pathogens (broad-spectrum resistance). In this study, we identified an aspartyl-tRNA synthetase (AspRS), StIBI1 (named after Arabidopsis IMPAIRED IN BABA-INDUCED IMMUNITY 1; IBI1), as a BABA receptor in Solanum tuberosum. We elucidated the regulatory mechanisms by which StIBI1 interacts with two NAC (NAM, ATAF1, 2, and CUC2) transcription factors (TFs), StVOZ1 and StVOZ2 (VASCULAR PLANT ONE ZINC FINGER, VOZ), to activate BABA-induced resistance (BABA-IR). StVOZ1 represses, whereas StVOZ2 promotes, immunity to the late blight pathogen Phytophthora infestans. Interestingly, BABA and StIBI1 influence StVOZ1- and StVOZ2-mediated immunity. StIBI1 interacts with StVOZ1 and StVOZ2 in the cytoplasm, reducing the nuclear accumulation of StVOZ1 and promoting the nuclear accumulation of StVOZ2. Our findings indicate that StVOZ1 and StVOZ2 finely regulate potato resistance to late blight through distinct signaling pathways. In summary, our study provides insights into the interaction between the potato BABA receptor StIBI1 and the TFs StVOZ1 and StVOZ2, which affects StVOZ1 and StVOZ2stability and nuclear accumulation to regulate late blight resistance during BABA-IR. This research advances our understanding of the primary mechanisms of BABA-IR in potato and contributes to a theoretical basis for the prevention and control of potato late blight using BABA-IR.

Transcriptional profiling during infection of potato NLRs and Phytophthora infestans effectors using cDNA enrichment sequencing

An accurate assessment of host and pathogen gene expression during infection is critical for understanding the molecular aspects of host-pathogen interactions. Often, pathogen-derived transcripts are difficult to ascertain at early infection stages owing to the unfavourable transcript representation compared to the host genes. In this study, we compare two sequencing techniques, RNAseq and enrichment sequencing (RenSeq and PenSeq) of cDNA, to investigate gene expression patterns in the doubled monoploid potato (DM) infected with the late blight pathogen Phytophthora infestans. Our results reveal distinct advantages of cDNA RenSeq and PenSeq over traditional RNAseq in terms of target gene representation and transcriptional quantification at early infection stages. Throughout the infection time course, cDNA enrichment sequencing enables transcriptomic analyses for more targeted host and pathogen genes. For highly expressed genes that were sampled in parallel by both cDNA enrichment and RNAseq, a high level of concordance in expression profiles is observed, indicative of at least semi-quantitative gene expression representation following enrichment.

OBTAINING TRANSGENIC POTATO PLANTS FOR COUNTERING PHYTOPHTHORA INFESTANS VIA “HIGS” METHOD IMPLEMENTATION

Oomycetes are a group of organisms that include some of the most dangerous plant pathogens capable of causing epiphytoties that lead to reduced crop yields and famine in some countries. The only available method of combating them – repeated application of fungicides in fields and careful selection of seed material – is ineffective for most of these pathogens. This paper proposes to use the HIGS (host-induced gene silencing) approach so that genetically modified plants, when infected with late blight, would cause the pathogen to silence its effector protein genes, which would prevent the pathogen from growing and reproducing in plant. To implement the HIGS method against the pathogen Phytophthora infestans, we developed two genetically engineered constructs, one of which uses the P. infestans RXLR-effector gene AVR3a-b as the RNA interference target, and the other uses a region of the PITG_03155 effector gene, which is conserved for both P. infestans and P. cactorum. After Agrobacterium-mediated genetic transformation of potato plants, we obtained four transgenic plants carrying a construct based on the AVR3a-b gene (two each of the “Milena” and “Gala” varieties) and one of “Milena” variety carrying a construct based on the PITG_03155 gene. These plants will be micro clonally propagated and tested for the presence of interfering RNA to the effector genes of P. infestans, after which the plant lines are planned to be planted under natural pathogenic background conditions and a comparative analysis with control plants for disease incidence and resistance will be conducted.

Subfunctionalization of NRC3 altered the genetic structure of the Nicotiana NRC network.

Nucleotide-binding domain and leucine-rich repeat (NLR) proteins play crucial roles in immunity against pathogens in both animals and plants. In solanaceous plants, activation of several sensor NLRs triggers their helper NLRs, known as NLR-required for cell death (NRC), to form resistosome complexes to initiate immune responses. While the sensor NLRs and downstream NRC helpers display diverse genetic compatibility, molecular evolutionary events leading to the complex network architecture remained elusive. Here, we showed that solanaceous NRC3 variants underwent subfunctionalization after the divergence of Solanum and Nicotiana, altering the genetic architecture of the NRC network in Nicotiana. Natural solanaceous NRC3 variants form three allelic groups displaying distinct compatibilities with the sensor NLR Rpi-blb2. Ancestral sequence reconstruction and analyses of natural and chimeric variants identified six key amino acids involved in sensor-helper compatibility. These residues are positioned on multiple surfaces of the resting NRC3 homodimer, collectively contributing to their compatibility with Rpi-blb2. Upon activation, Rpi-blb2-compatible NRC3 variants form membrane-associated punctate and high molecular weight complexes, and confer resistance to the late blight pathogen Phytophthora infestans. Our findings revealed how mutations in NRC alleles lead to subfunctionalization, altering sensor-helper compatibility and contributing to the increased complexity of the NRC network.

Biocontrol Mechanisms of Three Plant Essential Oils Against Phytophthora infestans Causing Potato Late Blight.

Late blight, caused by the notorious pathogen Phytophthora infestans, poses a significant threat to potato (Solanum tuberosum) crops worldwide, impacting their quality as well as yield. Here, we aimed to investigate the potential use of cinnamaldehyde, carvacrol, and eugenol as control agents against P. infestans and to elucidate their underlying mechanisms of action. To determine the pathogen-inhibiting concentrations of these three plant essential oils (PEOs), a comprehensive evaluation of their effects using gradient dilution, mycelial growth rate, and spore germination methods was carried out. Cinnamaldehyde, carvacrol, and eugenol were capable of significantly inhibiting P. infestans by hindering its mycelial radial growth, zoospore release, and sporangium germination; the median effective inhibitory concentration of the three PEOs was 23.87, 8.66, and 89.65 μl/liter, respectively. Scanning electron microscopy revealed that PEOs caused the irreversible deformation of P. infestans, resulting in hyphal shrinkage, distortion, and breakage. Moreover, propidium iodide staining and extracellular conductivity measurements demonstrated that all three PEOs significantly impaired the integrity and permeability of the pathogen's cell membrane in a time- and dose-dependent manner. In vivo experiments confirmed the dose-dependent efficacy of PEOs in reducing the lesion diameter of potato late blight. Altogether, these findings provide valuable insight into the antifungal mechanisms of PEOs vis-à-vis late blight-causing P. infestans. By utilizing the inherent capabilities of these natural compounds, we could effectively limit the harmful impacts of late blight on potato crops, thereby enhancing agricultural practices and ensuring the resilience of global potato food production.

Characterization of a methyltransferase for iterative N-methylation at the leucinostatin termini in Purpureocillium lilacinum

N-methyltransferase (NMT)-catalyzed methylation at the termini of nonribosomal peptides (NRPs) has rarely been reported. Here, we discover a fungal NMT LcsG for the iterative terminal N-methylation of a family of NRPs, leucinostatins. Gene deletion results suggest that LcsG is essential for leucinostatins methylation. Results from in vitro assays and HRESI-MS-MS analysis reveal the methylation sites as NH2, NHCH3 and N(CH3)2 in the C-terminus of various leucinostatins. LcsG catalysis yields new lipopeptides, some of which demonstrate effective antibiotic properties against the human pathogen Cryptococcus neoformans and the plant pathogen Phytophthora infestans. Multiple sequence alignments and site-directed mutagenesis of LcsG indicate the presence of a highly conserved SAM-binding pocket, along with two possible active site residues (D368 and D395). Molecular dynamics simulations show that the targeted N can dock between these two residues. Thus, this study suggests a method for increasing the variety of natural bioactivity of NPRs and a possible catalytic mechanism underlying the N-methylation of NRPs.

Test of Specificity in Signalling between Potato Plants in Response to Infection by Fusarium Solani and Phytophthora Infestans.

Plant-plant signalling via volatile organic compounds (VOCs) in response to insect herbivory has been widely studied, but its occurrence and specificity in response to pathogen attack has received much less attention. To fill this gap, we carried out a greenhouse experiment using two fungal pathogens (Fusarium solani and Phytophthora infestans) to test for specificity in VOC induction and signalling between potato plants (Solanum tuberosum). We paired potato plants in plastic cages, one acting as VOC emitter and the other as receiver, and subjected emitters to one of the following treatments: no infection (control), infected by F. solani, or infected by P. infestans. We measured total emission and composition of VOCs released by emitter plants to test for pathogen-specificity in VOC induction, and then conducted a pathogen infection bioassay to assess resistance levels on receiver plants by subjecting half of the receivers of each emitter treatment to F. solani infection and the other half to P. infestans infection. This allowed us to test for specificity in plant VOC signalling by comparing its effects on conspecific and heterospecific sequential infections. Results showed that infection by neither F. solani or P. infestans produced quantitative (total emissions) or qualitative (compositional) changes in VOC emissions. Mirroring these patterns, emitter infection treatment (control vs. pathogen infection) did not produce a significant change in pathogen infection levels on receiver plants in any case (i.e., either for conspecific or heterospecific sequential infections), indicating a lack of signalling effects which precluded pathogen-based specificity in signalling. We discuss possible mechanisms for lack of pathogen effects on VOC emissions and call for future work testing for pathogen specificity in plant-plant signalling and its implications for plant-pathogen interactions under ecologically relevant scenarios involving infections by multiple pathogens.

Two structurally different oomycete lipophilic microbe-associated molecular patterns induce distinctive plant immune responses.

Plants recognize a variety of external signals and induce appropriate mechanisms to increase their tolerance to biotic and abiotic stresses. Precise recognition of attacking pathogens and induction of effective resistance mechanisms are critical functions for plant survival. Some molecular patterns unique to a certain group of microbes, microbe-associated molecular patterns (MAMPs), are sensed by plant cells as nonself molecules via pattern recognition receptors. While MAMPs of bacterial and fungal origin have been identified, reports on oomycete MAMPs are relatively limited. This study aimed to identify MAMPs from an oomycete pathogen Phytophthora infestans, the causal agent of potato late blight. Using reactive oxygen species (ROS) production and phytoalexin production in potato (Solanum tuberosum) as markers, two structurally different groups of elicitors, namely ceramides and diacylglycerols, were identified. P. infestans ceramides (Pi-Cer A, B, and D) induced ROS production, while diacylglycerol (Pi-DAG A and B), containing eicosapentaenoic acid (EPA) as a substructure, induced phytoalexins production in potato. The molecular patterns in Pi-Cers and Pi-DAGs essential for defense induction were identified as 9-methyl-4,8-sphingadienine (9Me-Spd) and 5,8,11,14-tetraene-type fatty acid (5,8,11,14-TEFA), respectively. These structures are not found in plants, but in oomycetes and fungi, indicating that they are microbe molecular patterns recognized by plants. When Arabidopsis (Arabidopsis thaliana) was treated with Pi-Cer D and EPA, partially overlapping but different sets of genes were induced. Furthermore, expression of some genes is upregulated only after the simultaneous treatment with Pi-Cer D and EPA, indicating that plants combine the signals from simultaneously recognized MAMPs to adapt their defense response to pathogens.

Understanding the Temporal Dynamics of Invasive Late Blight Populations in India for Improved Management Practices.

The microbial oomycete pathogen Phytophthora infestans causes severe epidemics of potato late blight in crops globally. Disease management benefits from an understanding of the diversity of pathogen populations. In this study, we explore the dynamics of P. infestans populations in the late blight-potato agro-ecosystem across the Indian subcontinent. Investigations of the macroecological observations at the field level and microbial ecological principles provided insights into future pathogen behavior. We use a comprehensive simple sequence repeat allele dataset to demonstrate that an invasive clonal lineage called EU_13_A2 has dominated populations over 14 years across India, Bangladesh, and Pakistan. Increasing levels of subclonal variation were tracked over time and space, and, for the first time, populations in Asia were also compared with the source populations from Europe. Within India, a regional pathogen population structure was observed with evidence for local migration, cross-border movement between surrounding countries, and introductions via imports. There was also evidence of genetic drift and between-season transmission of more strongly pathogenic subclones with a complete displacement of some subclonal types. The limited introduction of novel genotypes and the use of resistant potato cultivars could contribute to the dominance of the 13_A2 lineage. The insights will contribute to the management of the pathogen in these key global potato production regions.

CRISPR/Cas9 genome editing of potato StDMR6-1 results in plants less affected by different stress conditions.

Potato is the third most important food crop, but cultivation is challenged by numerous diseases and adverse abiotic conditions. To combat diseases, frequent fungicide application is common. Knocking out susceptibility genes by genome editing could be a durable option to increase resistance. DMR6 has been described as a susceptibility gene in several crops, based on data that indicates increased resistance upon interruption of the gene function. In potato, Stdmr6-1 mutants have been described to have increased resistance against the late blight pathogen Phytophthora infestans in controlled conditions. Here, we present field evaluations of CRISPR/Cas9 mutants, in a location with a complex population of P. infestans, during four consecutive years that indicate increased resistance to late blight without any trade-off in terms of yield penalty or tuber quality. Furthermore, studies of potato tubers from the field trials indicated increased resistance to common scab, and the mutant lines exhibit increased resistance to early blight pathogen Alternaria solani in controlled conditions. Early blight and common scab are problematic targets in potato resistance breeding, as resistance genes are very scarce. The described broad-spectrum resistance of Stdmr6-1 mutants may further extend to some abiotic stress conditions. In controlled experiments of either drought simulation or salinity, Stdmr6-1 mutant plants are less affected than the background cultivar. Together, these results demonstrate the prospect of the Stdmr6-1 mutants as a useful tool in future sustainable potato cultivation without any apparent trade-offs.

Botrytis cinerea detoxifies the sesquiterpenoid phytoalexin rishitin through multiple metabolizing pathways

Botrytis cinerea is a necrotrophic pathogen that infects across a broad range of plant hosts, including high-impact crop species. Its generalist necrotrophic behavior stems from its ability to detoxify structurally diverse phytoalexins. The current study aims to provide evidence of the ability of B. cinerea to tolerate the sesquiterpenoid phytoalexin rishitin, which is produced by potato and tomato. While the growth of potato pathogens Phytophthora infestans (late blight) and Alternaria solani (early blight) was severely inhibited by rishitin, B. cinerea was tolerant to rishitin. After incubation of rishitin with the mycelia of B. cinerea, it was metabolized to at least six oxidized forms. Structural analysis of these purified rishitin metabolites revealed a variety of oxidative metabolism including hydroxylation at C7 or C12, ketone formation at C5, and dihydroxylation at the 10,11-olefin. Six rishitin metabolites showed reduced toxicity to P. infestans and A. solani, indicating that B. cinerea has at least 5 distinct enzymatic reactions to detoxify rishitin. Four host-specialized phytopathogenic Botrytis species, namely B. elliptica, B. allii, B. squamosa, and B. tulipae also had at least a partial ability to metabolize rishitin as B. cinerea, but their metabolic capacity was significantly weaker than that of B. cinerea. These results suggest that the ability of B. cinerea to rapidly metabolize rishitin through multiple detoxification mechanisms could be critical for its pathogenicity in potato and tomato.

A RabGAP negatively regulates plant autophagy and immune trafficking

Plants rely on autophagy and membrane trafficking to tolerate stress, combat infections, and maintain cellular homeostasis. However, the molecular interplay between autophagy and membrane trafficking is poorly understood. Using an AI-assisted approach, we identified Rab3GAP-like (Rab3GAPL) as a key membrane trafficking node that controls plant autophagy negatively. Rab3GAPL suppresses autophagy by binding to ATG8, the core autophagy adaptor, and deactivating Rab8a, a small GTPase essential for autophagosome formation and defense-related secretion. Rab3GAPL reduces autophagic flux in three model plant species, suggesting that its negative regulatory role in autophagy is conserved in land plants. Beyond autophagy regulation, Rab3GAPL modulates focal immunity against the oomycete pathogen Phytophthora infestans by preventing defense-related secretion. Altogether, our results suggest that Rab3GAPL acts as a molecular rheostat to coordinate autophagic flux and defense-related secretion by restraining Rab8a-mediated trafficking. This unprecedented interplay between a RabGAP-Rab pair and ATG8 sheds new light on the intricate membrane transport mechanisms underlying plant autophagy and immunity.

Оценка устойчивости перспективных гибридов цветного картофеля к фитофторозу

Abstract. Potatoes with pigmented tubers contain anthocyanins and other phenolic compounds that demonstrate antioxidant activity and have a beneficial effect on human health. The nutritional value and other consumer qualities of available potato varieties must be combined with resistance to various infections, primarily late blight, which leads to significant yield losses and considerable costs for chemical protection. The purpose of this work was to assess late blight resistance of 46 promising potato hybrids with different pigmentation patterns, both in the field and by laboratory methods, including molecular screening. The objectives of this study included obtaining promising hybrids of colored potatoes characterized by a set of economically importatnt traits, laboratory and field assessment of the resistance of leaves and tubers to the late blight pathogen Phytophthora infestans, as well as molecular screening of late blight resistance gene markers (Rpi genes). The production, cultivation and evaluation of the hybrid material was carried out using standard methods. Field and laboratory resistance of leaves and tubers was assessed on a 9-point scale. Molecular screening was carried out using PCR analysis. The scientific novelty of the work lies in the fact that for the first time a comprehensive assessment of the resistance to late blight of hybrid material of colored potatoes obtained on the territory of Russian Federation was carried out. According to the results obtained, the studied samples demonstrated a high level of resistance of tubers and leaves in field conditions and laboratory tests. However, the results of the study did not reveal a relationship between laboratory or field resistance and the presence of Rpi gene markers, which may be due to the influence of many independent factors that determine horizontal resistance.

Lipid transfer protein StLTPa enhances potato disease resistance against different pathogens by binding and disturbing the integrity of pathogens plasma membrane.

Potato is the third most important food crop worldwide. Potato production suffers from severe diseases caused by multiple detrimental plant pathogens, and broad-spectrum disease resistance genes are rarely identified in potato. Here we identified the potato non-specific lipid transfer protein StLTPa, which enhances species none-specific disease resistance against various pathogens, such as the oomycete pathogen Phytophthora infestans, the fungal pathogens Botrytis cinerea and Verticillium dahliae, and the bacterial pathogens Pectobacterium carotovorum and Ralstonia solanacearum. The StLTPa overexpression potato lines do not show growth penalty. Furthermore, we provide evidence that StLTPa binds to lipids present in the plasma membrane (PM) of the hyphal cells of P. infestans, leading to an increased permeability of the PM. Adding of PI(3,5)P2 and PI(3)P could compete the binding of StLTPa to pathogen PM and reduce the inhibition effect of StLTPa. The lipid-binding activity of StLTPa is essential for its role in pathogen inhibition and promotion of potato disease resistance. We propose that StLTPa enhances potato broad-spectrum disease resistance by binding to, and thereby promoting the permeability of the PM of the cells of various pathogens. Overall, our discovery illustrates that increasing the expression of a single gene in potato enhances potato disease resistance against different pathogens without growth penalty.

Modelling the displacement and coexistence of clonal lineages of Phytophthora infestans through revisiting past outbreaks

AbstractThe continuous changes in the lineage proportions of populations in the clonal plant pathogen Phytophthora infestans on potato and tomato crops have been perplexing to researchers and disease managers. Sudden outbreaks of newly emergent genotypes are often associated with these rapid composition changes. Modelling can predict the persistence and displacement of pathogen genotypes with differential fitness among hosts. Building upon previous models, we combined analytical and simulation methods to model the outcome of interactions between competing lineages on different hosts. Model inputs include pathogenesis parameters, and the outputs are fitness and lineage proportions within each host. Analytical solutions yielding complete displacement, partial coexistence‐displacement and complete coexistence were described. In a retrospective study, the lesion growth rate and sporulation density of P. infestans lineages on potato and tomato from pathogenicity trials were used as inputs. Output lineage frequencies were compared with historical epidemiological situations to check model accuracy. The results showed that pathogenesis traits measured from empirical trials could simulate lineage constituents on potato and tomato and estimate genotypic fitness with reasonable accuracy. The model also showed promise in predicting ongoing lineage displacements in the subsequent year or few years, even when the displaced lineage was still highly prevalent during the time of isolation. However, large uncertainties remain at temporal–spatial scales owing to complex meta‐population dynamics in some regions and adaptation to local environmental factors. This simulation model provides a new tool for forecasting pathogen compositions and can be used to identify potentially problematic genotypes based on pathogen life‐history traits.

Nutritional and anti-nutritional compositional analysis of transgenic potatoes with late blight resistance.

Late blight, caused by the pathogen Phytophthora infestans, is a devastating disease affecting potato production globally, with adverse effects in Africa where limited access to fungicides exacerbates its impact. Outbreaks of late blight lead to reduced yields and substantial economic losses to potato farmers and agricultural systems. The development of resistant potato varieties, tailored to African agroecological conditions, offers a viable solution in mitigating the devastating effects of late blight on potato cultivation. Leading to this study, two consumer-preferred varieties, Victoria and Shangi, with high susceptibility to late blight were targeted for conferring late blight resistance through genetic engineering. This was achieved by inserting R genes from wild relatives of potato displaying resistance to the disease. The intended effect of conferring resistance to the late blight disease has been consistently observed over twenty experimental field trials spanning 8years at three locations in Uganda and Kenya. In this study, we assessed whether the genetic transformation has led to any significant unintended effects on the nutritional and anti-nutritional composition of potato tubers compared to the non-transgenic controls grown under the same agroecological conditions. The compositional assessments were conducted on commercial-size potato tubers harvested from regulatory trials at three locations in Uganda and Kenya. Statistical analysis was conducted using two-way analysis of variance comparing transgenic and non-transgenic samples. Overall, the results showed that the transgenic and non-transgenic samples exhibited similar levels of nutritional and antinutritional components. Variations detected in the levels of the analysed components fell within the expected ranges as documented in existing literature and potato composition databases. Thus, we conclude that there are no biologically significant differences in the nutritional and anti-nutritional composition of transgenic and non-transgenic potato tubers engineered for resistance to late blight.

Anti-Oomycete Effect and Mechanism of Salicylic Acid on Phytophthora infestans.

Pathogenic oomycetes infect a wide variety of organisms, including plants, animals, and humans, and cause massive economic losses in global agriculture, aquaculture, and human health. Salicylic acid (SA), an endogenous phytohormone, is regarded as an inducer of plant immunity. Here, the potato late blight pathogen Phytophthora infestans was used as a model system to uncover the inhibitory mechanisms of SA on pathogenic oomycetes. In this research, SA significantly inhibited the mycelial growth, sporulation, sporangium germination, and virulence of P. infestans. Inhibition was closely related to enhanced autophagy, suppression of translation initiation, and ribosomal biogenesis in P. infestans, as shown by multiomics analysis (transcriptomics, proteomics, and phosphorylated proteomics). Monodansylcadaverine (MDC) staining and Western blotting analysis showed that SA promoted autophagy in P. infestans by probably targeting the TOR signaling pathway. These observations suggest that SA has the potential to control late blight caused by P. infestans.

A Variant of LbCas12a and Elevated Incubation Temperatures Enhance the Rate of Gene Editing in the Oomycete Phytophthora infestans.

CRISPR-Cas editing systems have proved to be powerful tools for functional genomics research, but their effectiveness in many non-model species remains limited. In the potato and tomato pathogen Phytophthora infestans, an editing system was previously developed that expresses the Lachnospiracae bacterium Cas12a endonuclease (LbCas12a) and guide RNA from a DNA vector. However, the method works at low efficiency. Based on a hypothesis that editing is constrained by a mismatch between the optimal temperatures for P. infestans growth and endonuclease catalysis, we tested two strategies that increased the frequency of editing of two target genes by about 10-fold. First, we found that editing was boosted by a mutation in LbCas12a (D156R) that had been reported to expand its catalytic activity over a broader temperature range. Second, we observed that editing was enhanced by transiently incubating transformed tissue at a higher temperature. These modifications should make CRISPR-Cas12a more useful for interrogating gene and protein function in P. infestans and its relatives, especially species that grow optimally at lower temperatures. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Bio-synthesized ZnO NPs using cow urine in vitro & in vivo efficacy against late blight pathogen

Biosynthesis of nanoparticles and their use to harvest agricultural products are a brilliant approach for vegetables and crops with large cultivation areas. Phytophthora, an oomycete, is one of the most serious challenges to tomato crop productivity. In this study, ZnO nanoparticles (ZnO NPs) have been synthesized using cow urine. The X-ray diffraction pattern confirms the hexagonal structure, and symmetry of ZnO NPs have a space group (P63mc). SEM and FTIR identified surface morphology and formation of metal oxide functional groups. A prominent absorption peak executes the electronic transitions from the valence band to the conduction band. In vitro and in vivo efficacy of ZnO NPs, cow urine, and the combination of (CU + ZnONPs) were checked against late blight pathogen (Phytophthora infestans). The role of ROS in ZnO NPs represents by a reduction in the growth inhibition of Phytophthora infestans and the cytotoxic effect of ZnO NPs mediated through ROS.