Research in Plant Metabolites

Research in Plant Metabolites

In vitro antifungal activity of plant-derived metabolites against Curvularia tsudae isolated from Lawsonia roots

Document Type : Original Article

Authors
Mahsa Moshayedi 1
Hadi Rahanandeh 2
Arash Sharifi Eshkalak 1
1 Member of the Agricultural and Natural Resources Engineering Organization of Gilan Province, Rasht, Iran
2 Department of Plant protection, Ra.C., Islamic Azad University, Rasht, Iran
10.22034/jrpsm.2026.570645.1105
Abstract
Background and Objectives:Ornamental coniferous plants play an essential role in urban green spaces, parks, and landscape design due to their aesthetic value, ecological functions, and adaptability to diverse environmental conditions. Among these plants, Lawson cypress (Chamaecyparis lawsoniana) is widely cultivated because of its desirable growth form, foliage color, and ornamental appeal. However, root decline and root-associated disorders represent a major constraint to the sustainability and visual quality of these plants. Such disorders are frequently associated with soil-borne and root-associated fungi, particularly dematiaceous fungi, which may persist as saprophytes or endophytes and become opportunistic pathogens under stress conditions.
Species belonging to the genus Curvularia have increasingly been reported from the rhizosphere and root tissues of ornamental and woody plants and are often linked to symptoms such as reduced vigor, chlorosis, and progressive decline. Climate change–related stresses, including fluctuations in temperature and soil moisture, can further disturb the balance between host plants and associated microorganisms, facilitating the pathogenic behavior of otherwise latent fungi. Management of root-associated fungal diseases traditionally relies on chemical fungicides. Although effective in the short term, repeated fungicide applications raise concerns regarding the development of resistant fungal populations, environmental contamination, and potential risks to human health. Consequently, there is growing interest in environmentally friendly alternatives, particularly plant-derived metabolites and essential oils, which are rich in bioactive compounds with documented antifungal properties. The present study aimed to isolate and identify a fungal species associated with root decline symptoms in C. lawsoniana and to evaluate the in vitro antifungal activity of selected plant essential oils against the identified isolate. In addition, the efficacy of these natural products was compared with commonly used chemical fungicides to assess their potential as sustainable disease management options.
Materials and Methods:Root samples were collected from Lawson cypress plants exhibiting decline and root rot symptoms in Guilan Province, northern Iran. After surface sterilization, root segments were plated on potato dextrose agar (PDA), and the dominant fungal isolate was purified through successive subculturing. Morphological identification was conducted based on colony characteristics and microscopic features of conidiophores and conidia. Molecular identification was performed by amplification and sequencing of the internal transcribed spacer (ITS) region of rDNA using universal primers ITS5 and ITS4. The obtained sequence was compared with reference sequences in the GenBank database using BLAST analysis. Phylogenetic relationships were inferred using the Neighbor-Joining method, and the evolutionary stability of branches was assessed by bootstrap analysis. Twelve plant essential oils, including thyme (Thymus vulgaris), ajwain (Trachyspermum ammi), kakuti (Ziziphora clinopodioides), lemongrass (Cymbopogon citratus), rosemary (Rosmarinus officinalis), peppermint (Mentha piperita), lavender (Lavandula angustifolia), fennel (Foeniculum vulgare), myrtle (Myrtus communis), eucalyptus (Eucalyptus globulus), galbanum (Ferula gummosa), and Persian hogweed (Heracleum persicum), were evaluated for antifungal activity. Antifungal assays were conducted using three complementary in vitro methods: disk diffusion, poisoned food (medium incorporation), and spore germination tests at different concentrations. Five chemical fungicides—mancozeb, thiophanate-methyl, copper oxychloride, iprodione + carbendazim (Rovral-TS), and Bordeaux mixture—were tested at 2000 mg L⁻¹ as chemical controls. All experiments were arranged in a completely randomized factorial design with three replications. Data were analyzed using analysis of variance (ANOVA), and mean comparisons were performed using Duncan’s multiple range test at P ≤ 0.05.
Results:Based on morphological traits and ITS rDNA sequence analysis, the fungal isolate obtained from Lawson cypress roots was identified as Curvularia tsudae and deposited in GenBank under accession number PX096193. Phylogenetic analysis confirmed that the isolate clustered within the Curvularia tsudae species complex, consistent with previous reports indicating limited species-level resolution of the ITS region within this genus. The disk diffusion assay revealed significant differences among essential oils in their ability to inhibit mycelial growth. Thyme essential oil exhibited the highest inhibitory effect, while several other oils showed limited or no significant activity compared with the control.
In the poisoned food assay, a stronger antifungal response was observed. Thyme, ajwain, and lemongrass essential oils completely inhibited mycelial growth of C. tsudae at higher concentrations, indicating pronounced antifungal efficacy. Other essential oils displayed moderate to low inhibitory effects, with inhibition generally increasing as concentration increased. Spore germination assays demonstrated that lemongrass and ajwain essential oils were the most effective in suppressing spore germination, whereas thyme essential oil showed a moderate inhibitory effect. In contrast, all tested chemical fungicides completely inhibited spore germination, confirming their high efficacy under laboratory conditions. Evaluation of fungicidal versus fungistatic activity revealed that thyme, ajwain, and lemongrass essential oils exerted true fungicidal effects, as no fungal regrowth occurred after transfer to fresh medium. Kakuti essential oil, however, exhibited a fungistatic effect, allowing fungal regrowth after removal of the treatment. Chemical fungicides varied in their mode of action, with some acting fungicidally and others primarily fungistatically.
Conclusion:This study demonstrated that Curvularia tsudae is associated with root decline symptoms in Lawson cypress and highlighted the considerable in vitro antifungal potential of selected plant-derived essential oils against this fungus. In particular, thyme, ajwain, and lemongrass essential oils showed strong inhibitory effects on mycelial growth and spore germination and exhibited fungicidal activity comparable, though not identical, to that of chemical fungicides. Although chemical fungicides achieved complete inhibition of spore germination, the observed efficacy of certain essential oils underscores their potential as environmentally friendly alternatives or complementary tools in integrated disease management strategies. These findings support further investigations under greenhouse and field conditions to evaluate the practical applicability, formulation stability, and phytotoxicity of these natural products. Overall, plant essential oils represent promising candidates for reducing reliance on synthetic fungicides and promoting sustainable management of root-associated fungal diseases in ornamental plants.
Keywords
Essential oils
Mycelial growth inhibition
Spore germination
Biological control
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