Development and use of nanoparticles from Lantana plant (Lantana camara) and Toad plant (Tabernaemontana elegans) in the management of Southern root-knot nematode (Meloidogyne incognita).

Abstract

Recent research on innovative nematicides has gained significant interest in addressing global crop losses caused by plant-parasitic nematodes. While chemical nematicides remain the primary control method, they pose risks to human health and the environment due to their toxicity, pollution potential, and residual effects. As a result, there is a growing demand for safer, more sustainable alternatives. Nanotechnology offers a promising solution by enhancing the effectiveness and targeted delivery of agrochemicals. Plant-based nanoparticle synthesis presents an eco-friendly approach by utilising natural bioactive compounds as reducing and stabilizing agents. The present study was carried out to explore the synthesis of green nanoparticles (NPs) derived from Lantana camara (NPlc) and Tabernaemontana elegans (NPte) using Aloe vera as reducing and stabilising agent (A. vera) and the sustainable management of plant-parasitic nematodes. This was achieved through four objectives: (1) to develop nanoparticles from nematicidal plants (T. elegans and L. camara) and determine their phytochemical composition, (2) to evaluate the suitability of A. vera gel as a reducing and stabilizing agent in the formulation of nanoparticles from T. elegans and L. camara, (3) to examine the effects of the prepared nanoparticles on hatch and mortality of M. incognita second-stage juveniles (J2) under in vitro conditions and (4) to explore the potential of the prepared nanoparticles in mitigating the impact of M. incognita and enhancing plant growth variables under greenhouse conditions. To achieve Objective 1 and 2, nanoparticles were synthesised by mixing four rations (25:75, 50:50,75:25, and 0:100 v/v) of A. vera gel with each plant extract. The experiment was a randomised complete design with five replications. The mixture was stirred using a magnetic stirrer for 12 h at room temperature. Nanoparticles and their mother plant extracts were screened for glycosides, flavonoids, alkaloids, tannins, phenols, saponins, reducing sugars and terpenoids phytochemicals using previously described qualitative methods, with energy-dispersive X-ray spectroscopy used to determine the chemical purity and elemental composition. Physical characterisation of the synthesised nanoparticles was done using UV-Vis spectroscopy, transmission electron microscopy (TEM), scanning electron microscopy (SEM) and selected area electron diffraction (SAED) patterns. Plant-based nanoparticles were successfully synthesised from T. elegans and L. camara plant extracts with Aloe vera as reducing and stabilising agent. Generally, all pure plant extracts exhibited a high abundance of phytochemicals measured, with synthesised nanoparticles having an improved levels of phytochemicals when compared to the pure forms. Adding A. vera increased the levels of phytochemicals such as saponins, terpenoids and reducing sugars on both L. camara and T. elegans nanoparticles. Synthesised nanoparticle concentrations of 50:50 and 25:75 (extract : A. vera) had more phytochemicals at a higher levels. Energy-dispersive X-ray spectroscopy (EDS) spectrum also indicated presence of key elements such as potassium (K), phosphorus (P), magnesium (Mg), chlorine (Cl), and copper (Cu), in the synthesised nanoparticles. The UV-Vis spectra peak intensity and wavelength observed in the two plant extracts, L. camara and T. elegans nanoparticles in the presence of A. vera extracts were slightly shifted and broader, suggesting an interaction between the biomolecules from both extracts during the nanoparticle formation process. New peaks in the nanoparticle spectrum indicate the formation of distinct molecular species or structural changes due to nanoparticle synthesis. Electron microscopy provided detailed structural and morphological properties, which were a variety of shapes such as cubic, triangular, platelet, and irregular forms. The nanoparticle sizes varied from 5 to 21 nm, far smaller than the pure extract sizes that were greater than 450 nm. The Selected Area Electron Diffraction (SAED) patterns of nanoparticles where generally bright with distinct spots for 50:50 and 25:75 ratio concentrations, indicative of well-developed polycrystalline structures. In Objective 3, the M. incognita J2 hatch inhibition test was conducted under in-vitro conditions. Mortality J2 hatch rates differed significantly (P ≤ 0.05) across these time intervals, with the highest rates recorded at 72 h and the lowest at 24 h. A consistent trend of increasing M. incognita juvenile hatch over time was observed. In addition, NPlc and NPte at the 50:50 (extract: A. vera) concentration demonstrated the lowest M. incognita juvenile hatch rates, followed by 25:75 (extract: A. vera). The mortality assay was conducted under in-vitro conditions as in the juvenile hatch inhibition, except that 100 M. incognita J2 were added into the Petri dish. Meloidogyne incognita J2 nematodes were considered dead if they were not mobile after transferring them into distilled water for 30 s even if they were probed with needles. The percentage of juvenile mortality was calculated. The highest mortality rate was recorded at 72 hours, while the lowest occurred at 24 hours, indicating a clear trend of increased juvenile mortality with extended exposure durations. The highest mortality rates were recorded at the 50:50 and 25:75 concentrations (extract: A. vera) of both NPlc and NPte, with no significant differences between these two concentrations. In Objective 4, plastic pots of 30-cm-diameter, were filled with a soil mixture of steam pasteurized loamy and sandy soil at a ratio of 1:3 (v/v). Each pot was transplanted with one "Star 9009" tomato seedling. Two weeks after transplanting, seedlings were inoculated with 5000 M. incognita eggs and J2 using a 20 mL plastic syringe. One-week post-inoculation, nanoparticle treatments were applied to the pots. The experiment followed a 2 x 6 x 3 factorial arrangement in a randomized complete block design (RCBD) with five replications. The first factor consisted of two plant extracts (L. camara and T. elegans). The second factor consisted of five previously developed nanoparticle formulations; a positive control (Nemacur® 10GR at 5 g/plant); 100 % A. vera; and a negative control (nematode-inoculated plants without treatment) were also included. The third factor comprised of three nanoparticle application rates of 5 ml, 10 ml, and 15 ml. At 56 days, plant and nematodes variables were measured. Nematode management results highlighted a trend where increasing concentrations of A. vera in nanoparticle formation contributed to reductions in the number of M. incognita nematode J2 within the roots. The 50:50 and 25:75 nanoparticle concentration were particularly effective in this regard, achieving the greatest reductions compared to other treatments. The application of nanoparticles directly influenced key plant growth parameters. This study represents the first documented use of A. vera gel in the green synthesis of nanoparticles from T. elegans and L. camara. The findings of this study conclusively highlight the capacity of green-synthesised nanoparticles to not only mitigate nematode infestations but also to enhance overall crop productivity, making them a valuable tool for sustainable agriculture. The study also paves the way for future research into optimising nanoparticle synthesis, conducting large-scale field trials, and evaluating the long-term environmental and economic impacts of their application.