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Zucchini yellow mosaic virus (ZYMV) poses a significant threat to squash crops, causing severe symptoms and substantial yield losses. This study investigates the potential of Streptomyces fungicidicus as a biocontrol agent for managing ZYMV in squash plants by inducing systemic resistance. Approximately 95% of field-collected squash samples were positive for ZYMV, exhibiting chlorotic mottling, vein banding, and leaf distortion. The virus was isolated, purified, and confirmed through RT-PCR (Accession no. PV131044) and transmission electron microscopy (TEM), which displayed flexuous, filamentous particles typical of ZYMV. Among the isolated and tested Streptomyces spp., Streptomyces fungicidicus SF1-RSI2 (Accession no. PV489988) exhibited promising antiviral activity. GC–MS analysis of the SF1-RSI2 culture filtrate identified 35 bioactive compounds, with (–)-spathulenol being the most abundant (13.1%), followed by 9-octadecenoic acid methyl ester (9.24%) and triacetin (8.88%), suggesting a complex mixture of metabolites potentially contributing to the observed antiviral effects. Under greenhouse conditions, the foliar application of SF1-RSI2 culture filtrate, either as a pre-ZYMV-inoculation (protective) or post-ZYMV-inoculation (curative) treatment, enhanced plant growth, delayed symptom onset by up to 3 days, and reduced viral accumulation by up to 49.7% at 5 days post-inoculation (dpi) compared to untreated infected plants. The SF1-RSI2 applications significantly mitigated the detrimental effects of ZYMV on plant growth, chlorophyll content, and oxidative stress markers, such as H₂O₂ and malondialdehyde (MDA), while also notably increasing peroxidase enzyme activity and total phenolic content, indicating enhanced systemic resistance. Additionally, it influenced the transcriptional levels of defense-related genes, with protective treatment resulting in the highest expression levels of Cinnamate-4-hydroxylase ( C4H ), Cinnamate-3-hydroxylase ( C3H ), and Chalcone synthase ( CHS ), which are essential for plant defense mechanisms. HPLC analysis revealed a substantial increase in polyphenolic compounds, particularly chlorogenic acid, in plants treated with SF1-RSI2, highlighting its role in strengthening plant defenses. The findings suggest that S. fungicidicus promotes plant growth and enhances defense mechanisms, presenting a viable biocontrol strategy for managing viral infections in squash.

Identifying a viable substitute for the limited array of current antifungal agents stands as a crucial objective in modern agriculture. Consequently, extensive worldwide research has been undertaken to unveil eco-friendly and effective agents capable of controlling pathogens resistant to the presently employed fungicides. This study explores the efficacy of Trichoderma isolates in combating tomato leaf spot disease, primarily caused by Alternaria alternata . The identified pathogen, A. alternata Alt3, was isolated and confirmed through the ITS region (OQ888806). Six Trichoderma isolates were assessed for their ability to inhibit Alt3 hyphal growth using dual culture, ethyl acetate extract, and volatile organic compounds (VOCs) techniques. The most promising biocontrol isolate was identified as T. afroharzianum isolate TRI07 based on three markers: ITS region (OQ820171), translation elongation factor alpha 1 gene (OR125580), and RNA polymerase II subunit gene (OR125581). The ethyl acetate extract of TRI07 isolate was subjected to GC–MS analysis, revealing spathulenol, triacetin, and aspartame as the main compounds, with percentages of 28.90, 14.03, and 12.97%, respectively. Analysis of TRI07-VOCs by solid-phase microextraction technique indicated that the most abundant compounds included ethanol, hydroperoxide, 1-methylhexyl, and 1-octen-3-one. When TRI07 interacted with Alt3, 34 compounds were identified, with major components including 1-octen-3-one, ethanol, and hexanedioic acid, bis(2-ethylhexyl) ester. In greenhouse experiment, the treatment of TRI07 48 h before inoculation with A. alternata (A3 treatment) resulted in a reduction in disease severity (16.66%) and incidence (44.44%). Furthermore, A3 treatment led to improved tomato growth performance parameters and increased chlorophyll content. After 21 days post-inoculation, A3 treatment was associated with increased production of antioxidant enzymes (CAT, POD, SOD, and PPO), while infected tomato plants exhibited elevated levels of oxidative stress markers MDA and H 2 O 2 . HPLC analysis of tomato leaf extracts from A3 treatment revealed higher levels of phenolic acids such as gallic, chlorogenic, caffeic, syringic, and coumaric acids, as well as flavonoid compounds including catechin, rutin, and vanillin. The novelty lies in bridging the gap between strain-specific attributes and practical application, enhancing the understanding of TRI07’s potential for integrated pest management. This study concludes that TRI07 isolate presents potential natural compounds with biological activity, effectively controlling tomato leaf spot disease and promoting tomato plant growth. The findings have practical implications for agriculture, suggesting a sustainable biocontrol strategy that can enhance crop resilience and contribute to integrated pest management practices.

One of the tomato’s acutely devastating diseases is Alternaria leaf spot, lowering worldwide tomato production. In this study, one fungal isolate was isolated from tomatoes and was assigned to Alternaria alternata TAA-05 upon morphological and molecular analysis of the ITS region and 18SrRNA, endoPG , Alt a1, and gapdh genes. Also, Urtica dioica and Dodonaea viscosa methanol leaf extracts (MLEs) were utilized as antifungal agents in vitro and compared to Ridomil, a reference chemical fungicide. The in vitro antifungal activity results revealed that Ridomil (2000 µg/mL) showed the highest fungal growth inhibition (FGI) against A. alternata (96.29%). Moderate activity was found against A. alternata by D. viscosa and U. dioica MLEs (2000 µg/mL), with an FGI value of 56.67 and 54.81%, respectively. The abundance of flavonoid and phenolic components were identified by HPLC analysis in the two plant extracts. The flavonoid compounds, including hesperidin, quercetin, and rutin were identified using HPLC in D. viscosa MLE with concentrations of 11.56, 10.04, and 5.14 µg/mL of extract and in U. dioica MLE with concentrations of 12.45, 9.21, and 5.23 µg/mL, respectively. α-Tocopherol and syringic acid, were also identified in D. viscosa MLE with concentrations of 26.13 and 13.69 µg/mL, and in U. dioica MLE, with values of 21.12 and 18.33 µg/mL, respectively. Finally, the bioactivity of plant extracts suggests that they play a crucial role as antifungal agents against A. alternata . Some phenolic chemicals, including coumaric acid, caffeic acid, ferulic acid, and α -tocopherol, have shown that they may be utilized as environmentally friendly fungicidal compounds.

The current investigation identified and characterized Alternaria alternata as the causal agent of tomato leaf spot through morphological traits and multilocus phylogenetic analysis [internal transcribed spacer (ITS), translation elongation factor 1-alpha ( tef1 -α), and the RNA polymerase II second-largest subunit ( RPB2 )], thereby confirming its classification within the A. alternata complex. Four Trichoderma isolates (Ham34, Ham35, Ham36, and Ham37) were recovered from loamy rhizosphere soil associated with healthy tomato plants. Among them, Ham34 ( Trichoderma hamatum ) showed the most significant antagonistic activity in dual-culture assays. Ethyl acetate extracts of all isolates exhibited concentration-dependent antifungal effects, with Ham34 showing the most potent inhibition (57.8% at 2000 µg/mL). Gas chromatography-mass spectrometry (GC-MS) profiling of the Ham34 extract identified bioactive metabolites, including (-)-spathulenol (28.2%) and glycerol 1,2-diacetate (21.4%). Among the four tested plant extracts, Rumex dentatus (RD) showed superior activity, achieving 68.5% inhibition at 2000 µg/mL, followed by moderate inhibition from Cichorium intybus , Conium maculatum , and Capsicum annuum . High-performance liquid chromatography (HPLC) profiling of Rumex dentatus extract revealed high levels of phenolic and flavonoid compounds, with a total concentration of 108.63 µg/mL. The primary identified constituents were rutin (33.1 µg/mL), gallic acid (22.7 µg/mL), and chlorogenic acid (17.9 µg/mL). Under greenhouse conditions, tomato plants ( Solanum lycopersicum cv. Super Strain B) were treated with Ham34 ethyl acetate extract, RD extract, and their combination (Ham34 + RD). The combined treatment significantly reduced disease incidence (11.1%) and severity (16.7%), outperforming Ridomil Gold® in disease suppression. Growth parameters and chlorophyll content (SPAD) were significantly enhanced in the combined treatment, with increased shoot and root biomass and length. Biochemical analyses revealed that the combined Ham34 + RD treatment enhanced antioxidant enzyme activities, including catalase (CAT) and peroxidase (POD), and increased total soluble protein content to 342.8 µg/mL compared to 331.1 µg/mL in the untreated control. Moreover, the treatment elevated total phenolic content to 256 mg GAE/g, indicating improved stress mitigation and enhanced plant vigor. This eco-friendly approach provides a low-cost, chemically stable, and safe alternative to synthetic fungicides, highlighting the synergistic potential of integrating R. dentatus and T. hamatum for sustainable management of A. alternata -induced tomato leaf spot.