Pathological characterization and management of Lasiodiplodia theobromae, a hemi-biotroph with an interkingdom host range.

Heart rot disease, caused by Lasiodiplodia theobromae, is destructive for date palms and other woody plants. The disease was reported in several oasis in Egypt, and the pathogen was found in association with infected trees suffering die-back and rachis blight. Seven phylogenetically distinct fungal isolates were selected, and their pathogenicity was confirmed on date palms. The isolates exhibited variable degrees of virulence on inoculated leaves, which confirms the variation. We examined the antifungal effect of microbial bioagents and plant extracts on heart rot disease. The isolates of Trichoderma spp. gave moderate reduction of the pathogen's linear growth (40-60%), while their exudates were ultimately ineffective. Bacillus spp. isolates, except for B. megaterium, were more effective against spore germination as they gave 80-90% reduction on average. Among the examined plant extracts garlic sap gave 98.67% reduction of linear growth followed by artemisia (15.5%) and camphor (24.8%). The extraction methods greatly influenced the antifungal efficiency of each extract as exposure to organic solvents significantly decreased the efficiency of all extracts, while hot water extraction negatively affected garlic sap only. Successful bioagents and plant extracts were further assayed for the suppression of heart rot disease on date palms. Both T. album and T. harzianum gave comparable degrees of suppression as by commercial fungicides. In addition, treatment before or during pathogen inoculation was the most effective as it significantly enhanced the expression of defense-related enzymes. Our findings suggest bio-pesticides possessing a dual role in disease suppression and defense boosters for date palms suffering heart rot disease.

bZIP60 and Bax inhibitor 1 contribute IRE1-dependent and independent roles to potexvirus infection.

IRE1, BI-1, and bZIP60 monitor compatible plant-potexvirus interactions though recognition of the viral TGB3 protein. This study was undertaken to elucidate the roles of three IRE1 isoforms, the bZIP60U and bZIP60S, and BI-1 roles in genetic reprogramming of cells during potexvirus infection. Experiments were performed using Arabidopsis thaliana knockout lines and Plantago asiatica mosaic virus infectious clone tagged with the green fluorescent protein gene (PlAMV-GFP). There were more PlAMV-GFP infection foci in ire1a/b, ire1c, bzip60, and bi-1 knockout than wild-type (WT) plants. Cell-to-cell movement and systemic RNA levels were greater bzip60 and bi-1 than in WT plants. Overall, these data indicate an increased susceptibility to virus infection. Transgenic overexpression of AtIRE1b or StbZIP60 in ire1a/b or bzip60 mutant background reduced virus infection foci, while StbZIP60 expression influences virus movement. Transgenic overexpression of StbZIP60 also confers endoplasmic reticulum (ER) stress resistance following tunicamycin treatment. We also show bZIP60U and TGB3 interact at the ER. This is the first demonstration of a potato bZIP transcription factor complementing genetic defects in Arabidopsis. Evidence indicates that the three IRE1 isoforms regulate the initial stages of virus replication and gene expression, while bZIP60 and BI-1 contribute separately to virus cell-to-cell and systemic movement.

Non-proteinogenic amino acids mitigate oxidative stress and enhance the resistance of common bean plants against Sclerotinia sclerotiorum.

White mold, caused by the necrotrophic fungus Sclerotinia sclerotiorum, is a challenging disease to common bean cultivation worldwide. In the current study, two non-proteinogenic amino acids (NPAAs), γ-aminobutyric acid (GABA) and ß-alanine, were suggested as innovative environmentally acceptable alternatives for more sustainable management of white mold disease. In vitro, GABA and ß-alanine individually demonstrated potent dose-dependent fungistatic activity and effectively impeded the radial growth and development of S. sclerotiorum mycelium. Moreover, the application of GABA or ß-alanine as a seed treatment followed by three root drench applications efficiently decreased the disease severity, stimulated plant growth, and boosted the content of photosynthetic pigments of treated S. sclerotiorum-infected plants. Furthermore, although higher levels of hydrogen peroxide (H2O2), superoxide anion (O2 •-), and malondialdehyde (MDA) indicated that S. sclerotiorum infection had markedly triggered oxidative stress in infected bean plants, the exogenous application of both NPAAs significantly reduced the levels of the three studied oxidative stress indicators. Additionally, the application of GABA and ß-alanine increased the levels of both non-enzymatic (total soluble phenolics and flavonoids), as well as enzymatic (catalase [CAT], peroxidases [POX], and polyphenol oxidase [PPO]) antioxidants in the leaves of S. sclerotiorum-infected plants and improved their scavenging activity and antioxidant efficiency. Applications of GABA and ß-alanine also raised the proline and total amino acid content of infected bean plants. Lastly, the application of both NPAAs upregulated the three antioxidant-related genes PvCAT1, PvCuZnSOD1, and PvGR. Collectively, the fungistatic activity of NPAAs, coupled with their ability to alleviate oxidative stress, enhance antioxidant defenses, and stimulate plant growth, establishes them as promising eco-friendly alternatives for white mold disease management for sustainable bean production.

Gamma-rays induced mutations increase soybean oil and protein contents.

Mutation breeding is one of the effective techniques used for improving desired traits such as yield quality and quantity in economic crops. The present study aims to develop oil and protein contents in addition to high yield attributes in soybean using gamma rays as a mutagen. Seeds of the soybean genotypes Giza 21, Giza 22, Giza 82, Giza 83 and 117 were treated with gamma rays doses 50, 100, 200 and 300 Gy. Plants were then scored based on morphological parameters correlated with yield quantity including plant height, seed weight and valuable protein and oil contents. Mutant lines exhibiting the highest yield attributes were selected and used as parents for M2 generation. The M2 progeny was further assessed based on their ability to maintain their yield attributes. Twenty mutant lines were selected and used as M3 lines. The yield parameters inferred a positive effect of gamma irradiation on the collected M3 mutant lines compared to their parental genotypes. 100 Gy of gamma rays gave the highest effect on the number of pods, branches and seeds per plant in addition to protein content, while 200 Gy was more effective in increasing plant height, number of pods per plant, and oil content. Six mutant lines scored the highest yield parameters. Further assessment inferred an inverse relationship between oil and protein content in most of the tested cultivars with high agronomic features. However, four mutant lines recorded high content of oil and protein besides their high seed yield as well, which elect them as potential candidates for large-scale evaluation. The correlation among examined parameters was further confirmed via principal component analysis (PCA), which inferred a positive correlation between the number of pods, branches, seeds, and seed weight. Conversely, oil and protein content were inversely correlated in most of yielded mutant lines. Together, those findings introduce novel soybean lines with favorable agronomic traits for the market. In addition, our research sheds light on the value of using gamma rays treatment in enhancing genetic variability in soybean and improving oil, protein contents and seed yield.

Association of High and Low Molecular Weight Glutenin Subunits with Gluten Strength in Tetraploid Durum Wheat (Triticum turgidum spp. Durum L.).

The gluten strength and the composition of high- and low-molecular-weight glutenin subunits (HMWGSs and LMWGSs) of fifty-one durum wheat genotypes were evaluated using sodium dodecyl sulfate (SDS) sedimentation testing and SDS polyacrylamide gel electrophoresis (SDS-PAGE). This study examined the allelic variability and the composition of HMWGSs and LMWGSs in T. durum wheat genotypes. SDS-PAGE was proven to be a successful method for identifying HMWGS and LMWGS alleles and their importance in determining the dough quality. The evaluated durum wheat genotypes with HMWGS alleles 7+8, 7+9, 13+16, and 17+18 were highly correlated with improved dough strength. The genotypes containing the LMW-2 allele displayed stronger gluten than those with the LMW-1 allele. The comparative in silico analysis indicated that Glu-A1, Glu-B1, and Glu-B3 possessed a typical primary structure. The study also revealed that the lower content of glutamine, proline, glycine, and tyrosineand the higher content of serine and valine in the Glu-A1 and Glu-B1 glutenin subunits, and the higher cysteine residues in Glu-B1 and lower arginine, isoleucine, and leucine in the Glu-B3 glutenin, are associated with the suitability of durum wheat for pasta making and the suitability of bread wheat with good bread-making quality. The phylogeny analysis reported that both Glu-B1 and Glu-B3 had a closer evolutionary relationship in bread and durum wheat, while the Glu-A1 was highly distinct. The results of the current research may help breeders to manage the quality of durum wheat genotypes by exploiting the allelic variation in glutenin. Computational analysis showed the presence of higher proportions of glutamine, glycine, proline, serine, and tyrosine than the other residues in both HMWGSs and LMWGSs. Thus, durum wheat genotype selection according to the presence of a few protein components effectively distinguishes the strongest from the weakest types of gluten.

New Insights into Hop Latent Viroid Detection, Infectivity, Host Range, and Transmission.

Hop latent viroid (HLVd), a subviral pathogen from the family Pospiviroidae, is a major threat to the global cannabis industry and is the causative agent for "dudding disease". Infected plants can often be asymptomatic for a period of growth and then develop symptoms such as malformed and yellowing leaves, as well as stunted growth. During flowering, HLVd-infected plants show reduced levels of valuable metabolites. This study was undertaken to expand our basic knowledge of HLVd infectivity, transmission, and host range. HLVd-specific primers were used for RT-PCR detection in plant samples and were able to detect HLVd in as little as 5 picograms of total RNA. A survey of hemp samples obtained from a diseased production system proved sole infection of HLVd (72%) with no coexistence of hop stunt viroid. HLVd was infectious through successive passage assays using a crude sap or total RNA extract derived from infected hemp. HLVd was also highly transmissible through hemp seeds at rates of 58 to 80%. Host range assays revealed new hosts for HLVd: tomato, cucumber, chrysanthemum, Nicotiana benthamiana, and Arabidopsis thaliana (Col-0). Sequence analysis of 77 isolates revealed only 3 parsimony-informative sites, while 10 sites were detected among all HLVd isolates available in the GenBank. The phylogenetic relationship among HLVd isolates allowed for inferring two major clades based on the genetic distance. Our findings facilitate further studies on host-viroid interaction and viroid management.

Exploring the Host Range of Rose rosette Virus among Herbaceous Annual Plants.

To study the host range of Rose rosette virus (RRV), we employed crude sap inoculum extracted from RRV-infected roses and the RRV infectious clone. We inoculated plants from the families Solanaceae, Cucurbitaceae, Leguminosae, Malvaceae, Amaranthaceae, and Brassicaceae. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect RRV in the inoculated plants throughout their growth stages. Interestingly, RRV was detected in the newly developed leaves of tomato, pepper, tobacco, cucumber, squash, zucchini, pumpkin, pea, peanut, soybean, spinach, okra, and Chenopodium spp. The speed of upward advancement of RRV within infected plants was variable between plants as it took two to three weeks for some plant species and up to five weeks in other plant species to emerge in the newest leaves. No severe symptoms were detected on most of the inoculated plants. Chenopodium spp., spinach, cucumber and Nicotiana rustica exhibited either chlorotic or necrotic lesions with variable shapes and patterns on the systemically infected leaves. Double membrane-bound particles of 80-120 nm in diameter were detected by transmission electron microscopy in the infected tissues of cucumber, pepper, and N. benthamiana plants. This finding infers the validity of mechanical inoculation for RRV on a wide range of plants that would serve as potential natural reservoirs.

Phylogenetic comparative analysis: Chemical and biological features of caseins (alpha-S-1, alpha-S-2, beta- and kappa-) in domestic dairy animals.

Caseins determine the physicochemical, physiological, and biological characteristics of milk. Four caseins-alpha-S-1, alpha-S-2, beta, and kappa-were analyzed phylogenetically and in silico and characterized regarding chemical, antimicrobial, and antioxidant features in five dairy animals: Arabian camels, sheep, goats, cattle, and water buffalos. The sequence of full-length amino acids of the four caseins for the five species was retracted from the NCBI GenBank database. Multiple sequence alignment is used to examine further the candidate sequences for phylogenetic analysis using Clustal X and NJ-Plot tools. The results revealed that sheep and goats possess strong similarities (98.06%) because of their common ancestor. The same was observed with cattle and water buffalos (96.25%). The Arabian camel was located in a single subclade due to low similarity in casein residues and compositions with other dairy animals. Protein modeling showed that alpha-S1- and alpha-S2-caseins possess the highest number of phosphoserine residues. The in silico computed chemical properties showed that ß-casein recorded highest hydrophobicity index and lowest basic amino acid content, while α-S2-casein showed the opposite. The computed biological parameters revealed that α-S2-casein presented the highest bactericidal stretches. Only Arabian camel ß-casein and k-casein showed one bactericidal stretches. The analysis also revealed that ß-casein, particularly in Arabian camels, possesses the highest antioxidant activity index. These results support the importance of the bioinformatics resources to determine milk casein micelles' chemical and biological activities.

Polyphasic Characterization of Four Aspergillus Species as Potential Biocontrol Agents for White Mold Disease of Bean.

The genus Aspergillus comprises several species that play pivotal roles in agriculture. Herein, we morphologically and physiologically characterized four genetically distinct Aspergillus spp., namely A. japonicus, A. niger, A. flavus, and A. pseudoelegans, and examined their ability to suppress the white mold disease of bean caused by Sclerotinia sclerotiorum in vitro and under greenhouse conditions. Seriation type of Aspergillus spp. correlates with conidiospores discharge as detected on the Petri glass lid. Members of Nigri section cover their conidial heads with hard shells after prolonged incubation. In addition, sporulation of the tested Aspergillus isolates is temperature sensitive as it becomes inhibited at low temperatures and the colonies become white. Examined Aspergillus spp. were neither infectious to legumes nor aflatoxigenic as confirmed by HPLC except for A. flavus and A. pseudoelegans which, secreted 5 and 1 ppm of aflatoxin B1, respectively. Co-inoculations of Sclerotinia's mycelium or sclerotia with a spore suspension of Aspergillus spp. inhibited their germination on PDA at 18 °C and 28 °C, and halted disease onset on detached common bean and soybean leaves. Similarly, plants treated with A. japonicus and A. niger showed the highest survival rates compared to untreated plants. In conclusion, black Aspergillus spp. are efficient biocides and safe alternatives for the management of plant diseases, particularly in organic farms.

Emergence, evolution, and vaccine production approaches of SARS-CoV-2 virus: Benefits of getting vaccinated and common questions.

The emergence of coronavirus disease 2019 (COVID-19) pandemic in Wuhan city, China at the end of 2019 made it urgent to identify the origin of the causal pathogen and its molecular evolution, to appropriately design an effective vaccine. This study analyzes the evolutionary background of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 or SARS-2) in accordance with its close relative SARS-CoV (SARS-1), which was emerged in 2002. A comparative genomic and proteomic study was conducted on SARS-2, SARS-1, and Middle East respiratory syndrome coronavirus (MERS), which was emerged in 2012. In silico analysis inferred the genetic variability among the tested viruses. The SARS-1 genome harbored 11 genes encoding 12 proteins, while SARS-2 genome contained only 10 genes encoding for 10 proteins. MERS genome contained 11 genes encoding 11 proteins. The analysis also revealed a slight variation in the whole genome size of SARS-2 comparing to its siblings resulting from sequential insertions and deletions (indels) throughout the viral genome particularly ORF1AB, spike, ORF10 and ORF8. The effective indels were observed in the gene encoding the spike protein that is responsible for viral attachment to the angiotensin-converting enzyme 2 (ACE2) cell receptor and initiating infection. These indels are responsible for the newly emerging COVID-19 variants αCoV, ßCoV, γCoV and δCoV. Nowadays, few effective COVID-19 vaccines developed based on spike (S) glycoprotein were approved and become available worldwide. Currently available vaccines can relatively prevent the spread of COVID-19 and suppress the disease. The traditional (killed or attenuated virus vaccine and antibody-based vaccine) and innovated vaccine production technologies (RNA- and DNA-based vaccines and viral vectors) are summarized in this review. We finally highlight the most common questions related to COVID-19 disease and the benefits of getting vaccinated.

Silicon Dioxide Nanoparticles Induce Innate Immune Responses and Activate Antioxidant Machinery in Wheat Against Rhizoctonia solani.

The phytopathogenic basidiomycetous fungus, Rhizoctonia solani, has a wide range of host plants including members of the family Poaceae, causing damping-off and root rot diseases. In this study, we biosynthesized spherical-shaped silicon dioxide nanoparticles (SiO2 NPs; sized between 9.92 and 19.8 nm) using saffron extract and introduced them as a potential alternative therapeutic solution to protect wheat seedlings against R. solani. SiO2 NPs showed strong dose-dependent fungistatic activity on R. solani, and significantly reduced mycelial radial growth (up to 100% growth reduction), mycelium fresh and dry weight, and pre-, post-emergence damping-off, and root rot severities. Moreover, the impact of SiO2 NPs on the growth of wheat seedlings and their potential mechanism (s) for disease suppression was deciphered. SiO2 NPs application also improved the germination, vegetative growth, and vigor indexes of infected wheat seedlings which indicates no phytotoxicity on treated wheat seedlings. Moreover, SiO2 NPs enhanced the content of the photosynthetic pigments (chlorophylls and carotenoids), induced the accumulation of defense-related compounds (particularly salicylic acid), and alleviated the oxidative stress via stimulation of both enzymatic (POD, SOD, APX, CAT, and PPO) and non-enzymatic (phenolics and flavonoids) antioxidant defense machinery. Collectively, our findings demonstrated the potential therapeutic role of SiO2 NPs against R. solani infection via the simultaneous activation of a multilayered defense system to suppress the pathogen, neutralize the destructive effect of ROS, lipid peroxidation, and methylglyoxal, and maintain their homeostasis within R. solani-infected plants.

Soybean ß-conglycinin and catfish cutaneous mucous p22 glycoproteins deteriorate sporangial cell walls of Pseudoperonospora cubensis and suppress cucumber downy mildew.

BACKGROUND: Cucumber plants suffer from a serious threatening disease, downy mildew, throughout the growing seasons irrespective of the weather temperature. The causal agent, Pseudoperonospora cubensis, tends to evolve rapidly upon sequential applications of chemical fungicides and generate new progeny possessing tolerance to such fungicides. Glycoproteins represent an environmentally safe alternative for chemically synthetized fungicides and do not trigger fungicide resistance. We studied the antifungal activity of four glycoproteins namely soybean ß-conglycinin, chickpea vicilin, duck egg ovomucin and catfish p22 against P. cubensis. Ten commercial fungicides of different chemical groups were used as positive controls of glycoprotein treatments. RESULTS: The results revealed that soybean ß-conglycinin and catfish p22 glycoproteins possess significant antifungal activity against P. cubensis. The amount of disease suppression caused by ß-conglycinin and p22 was comparable to the highly efficient chemical fungicides containing copper oxychloride, cymoxanil and fosetyl Al as active ingredients. Vicilin and ovomucin were less efficient biocides as they gave moderate suppression of disease severity. However, all tested glycoproteins provided full protection for the newly emerged cucumber leaves. Microscopic examination of glycoprotein-treated leaves inferred the ability of catfish p22 and soybean ß-conglycinin to disrupt the integrity of sporangial cell walls of P. cubensis rendering them non-viable compared to untreated ones. Expression levels of total phenolic compounds and the antioxidant enzymes catalase, superoxide dismutase and peroxidase were elevated upon glycoproteins application, which infers their involvement in disease suppression. CONCLUSION: This report emphasizes the direct and indirect roles of glycoproteins in safe management of cucumber downy mildew disease. © 2021 Society of Chemical Industry.

Aspergillus spp. eliminate Sclerotinia sclerotiorum by imbalancing the ambient oxalic acid concentration and parasitizing its sclerotia.

Sclerotinia sclerotiorum, a pathogen of more than 600 host plants, secretes oxalic acid to regulate the ambient acidity and provide conducive environment for pathogenicity and reproduction. Few Aspergillus spp. were previously proposed as potential biocontrol agents for S. sclerotiorum as they deteriorate sclerotia and prevent pathogen's overwintering and initial infections. We studied the nature of physical and biochemical interactions between Aspergillus and Sclerotinia. Aspergillus species inhibited sclerotial germination as they colonized its rind layer. However, Aspergillus-infested sclerotia remain solid and viable for vegetative and carpogenic germination, indicating that Aspergillus infestation is superficial. Aspergillus spp. of section Nigri (Aspergillus japonicus and Aspergillus niger) were also capable of suppressing sclerotial formation by S. sclerotiorum on agar plates. Their culture filtrate contained high levels of oxalic, citric and glutaric acids comparing to the other Aspergillus spp. tested. Exogenous supplementation of oxalic acid altered growth and reproduction of S. sclerotiorum at low concentrations. Inhibitory concentrations of oxalic acid displayed lower pH values comparing to their parallel concentrations of other organic acids. Thus, S. sclerotiorum growth and reproduction are sensitive to the ambient oxalic acid fluctuations and the environmental acidity. Together, Aspergillus species parasitize colonies of Sclerotinia and prevent sclerotial formation through their acidic secretions.

A Long Non-Coding RNA of Citrus tristeza virus: Role in the Virus Interplay with the Host Immunity.

During infection, Citrus tristeza virus (CTV) produces a non-coding subgenomic RNA referred to as low-molecular-weight tristeza 1 (LMT1), which for a long time has been considered as a by-product of the complex CTV replication machinery. In this study, we investigated the role of LMT1 in the virus infection cycle using a CTV variant that does not produce LMT1 (CTV-LMT1d). We showed that lack of LMT1 did not halt virus ability to replicate or form proper virions. However, the mutant virus demonstrated significantly reduced invasiveness and systemic spread in Nicotiana benthamiana as well as an inability to establish infection in citrus. Introduction of CTV-LMT1d into the herbaceous host resulted in elevation of the levels of salicylic acid (SA) and SA-responsive pathogenesis-related genes beyond those upon inoculation with wild-type (WT) virus (CTV-WT). Further analysis showed that the LMT1 RNA produced by CTV-WT or via ectopic expression in the N. benthamiana leaves suppressed SA accumulation and up-regulated an alternative oxidase gene, which appeared to mitigate the accumulation of reactive oxygen species. To the best of our knowledge, this is the first report of a plant viral long non-coding RNA being involved in counter-acting host response by subverting the SA-mediated plant defense.

Functional diversification upon leader protease domain duplication in the Citrus tristeza virus genome: Role of RNA sequences and the encoded proteins.

Viruses from the family Closteroviridae show an example of intra-genome duplications of more than one gene. In addition to the hallmark coat protein gene duplication, several members possess a tandem duplication of papain-like leader proteases. In this study, we demonstrate that domains encoding the L1 and L2 proteases in the Citrus tristeza virus genome underwent a significant functional divergence at the RNA and protein levels. We show that the L1 protease is crucial for viral accumulation and establishment of initial infection, whereas its coding region is vital for virus transport. On the other hand, the second protease is indispensable for virus infection of its natural citrus host, suggesting that L2 has evolved an important adaptive function that mediates virus interaction with the woody host.

A 5'-proximal region of the Citrus tristeza virus genome encoding two leader proteases is involved in virus superinfection exclusion.

Superinfection exclusion (SIE), a phenomenon in which a primary virus infection prevents a secondary infection with the same or closely related virus, has been observed with various viruses. Earlier we demonstrated that SIE by Citrus tristeza virus (CTV) requires viral p33 protein. In this work we show that p33 alone is not sufficient for virus exclusion. To define the additional viral components that are involved in this phenomenon, we engineered a hybrid virus in which a 5'-proximal region in the genome of the T36 isolate containing coding sequences for the two leader proteases L1 and L2 has been substituted with a corresponding region from the genome of a heterologous T68-1 isolate. Sequential inoculation of plants pre-infected with the CTV L1L2T68 hybrid with T36 CTV resulted in superinfection with the challenge virus, which indicated that the substitution of the L1-L2 coding region affected SIE ability of the virus.