elocation-id: e3076
The objective of the research was to evaluate the in vitro antifungal activity of two extracts, creosote bush (Larrea tridentata L.) and mustard (Sinapis alba L.), nanoformulated with silicon oxide and zinc oxide nanoparticles (200 to 400 μ globular) on the mycelial growth and sporulation of Fusarium solani, one of the plant pathogens causing vascular wilt and root rot in at least 100 economically important crops. Inhibitory concentrations and the number of conidia were determined using the poisoned media method. Data were analyzed using probit analysis, Anova, and Tukey’s test (p≤ 0.05). The results showed that mustard-only treatments have the most effective ID50 with 920.57 ppm; nevertheless, the mixture of mustard and creosote bush with SO2 presented significant results on sporulation, with a formation of 0.35 and 0.48 million conidia ml-1 for creosote bush 3% SO2 and mustard 5% SO2 compared to the control (7.78).
mycelial growth, nanoparticles, sporulation.
In Mexico, fruit and vegetable production is an important source of economic resources since, in addition to being a profitable agricultural activity, it requires labor for the traceability of the product as these are the most exported. They are also important for their nutritional value since they contain vitamins, fibers, minerals, and antioxidants that complement the daily diet of people (Cedillo-Portugal, 2017). For export, phytosanitary requirements established by each of the countries with which Mexico has a trade agreement must be complied with, which means that they have to be free of pathogens that cause different diseases in these crops; however, the most important diseases that cause the most production losses are fungi, which affect the quality of the product and lead to entry rejection for the exporting country (SENASICA, 2020).
The list of plant diseases presented by the American Phytopathological Society reports that more than 81 of the 101 economically important plants have at least one disease caused by Fusarium spp. (Leslie and Summerell, 2006). Within the genus Fusarium is the species F. solani, one of the phytopathogens that cause vascular wilt and root rot in at least 100 agricultural crops (Šišić, 2018).
For its control, the intensive application of fungicides is mainly used, which generates health risks and damage to the environment; in addition, it is unfavorable and inadequate to control the disease, so it is important to have alternatives for its control (El-Mohamedy et al., 2014).
Some of the alternatives for controlling fungal diseases are using plant extracts and essential oils. Research has shown that they are safe for the environment and consumers and effective for the control of diseases caused by Fusarium; tests have also been carried out with commercial botanical formulations in mixtures with plant extracts and biological control microorganisms to analyze interactions (Villa-Martínez et al., 2015).
On the other hand, there is a technology with a remarkable impact in different sectors; in agriculture, it has diverse applications, mainly in the production of nanopesticides, the production of macro and micronutrients at the nano level, making them more efficient and sustainable (Lira et al., 2018). This study aimed to evaluate creosote bush and mustard extracts, nanoformulated with silicon oxide (SO2) and zinc oxide (ZnO), against the mycelial growth and spore production of F. solani.
The fungus F. solani was reactivated in potato dextrose agar (PDA) medium after being identified by morphological characteristics according to Lesley and Summer (2006); the strain was obtained from the Toxicology Laboratory of the Department of Parasitology at the Antonio Narro Autonomous Agrarian University (UAAAN), for its acronym in Spanish. The extracts and nanoparticles (200 to 400 μ globular) were acquired from the company Culta SA de CV. Fourteen treatments were obtained, which included pure extracts and nanoformulations with SiO2 and ZnO at 1, 3, and 5% per 100 ml of the extract.
To prepare the concentrations (0, 10, 100, 50, 1 000, 3 000, and 5 000 ppm), the extract at 100% was considered. To carry out the bioassays, the methodology followed was PDA poisoned medium in Petri dishes, leaving 24 h for solidification; a fragment of 0.5 cm of F. solani was placed in each Petri dish, they were incubated at 26 °C, and the measurement of the growth of the mycelium was taken with a vernier at 24 h until the control reached the dimensions of the Petri dish.
Mycelial growth was considered to obtain the percentage of inhibition using the formula: % inhibition= mycelial growth of the control-mycelial growth of the treatment/mycelial growth of the control x 100 (Ezziyyani et al., 2004). Spore counting was performed with a Neubauer chamber based on quantifying spore suspension (Bustillo, 2010).
To obtain the medium inhibitory dose (ID50), a probit analysis was performed with the results of percentage of inhibition; on the other hand, for spore formation, a completely randomized analysis of variance was performed with four repetitions. Each repetition consisted of three dishes per evaluation and a comparison of means test with the Tukey method (p≤ 0.05), using the SAS System version 9.0 statistical program.
The values of the creosote bush and mustard treatments, alone and with the SO2 and ZnO nanoparticles, are shown in Table 1, where it was observed that in the case of the treatments with creosote bush and SO2, the one with 5% showed the lowest value of ID50 with 3 140 ppm, followed by creosote bush 1% SO2 with 3 204 ppm. The highest ID50 values were creosote bush 3% SO2 and creosote bush without nanoparticles, with values of 3 351 and 3 238 ppm, respectively. For the mixture of creosote bush and ZnO, they presented values higher than those with SO2, with creosote bush 1% ZnO (ID50 of 4 739 ppm) being the highest result.
In this regard, the results of Tequida-Meneses et al. (2002), when evaluating the antifungal activity of extracts (creosote bush), showed inhibition of Fusarium spp., up to 100%, while Huang (2011), when evaluating silicon oxide, shows that the severity of the disease by F. oxysporum f. sp. radicis-lycopersici decreased considerably when transplanting tomato plants in nutrient solution added with this element. For mustard treatments, the lowest values occurred in mustard without nanoparticles, with an ID50 of 920.47 ppm; Drakopoulos et al. (2010) showed that mustard helps control diseases caused by Fusarium.
On the other hand, mustard with 3% ZnO had the highest ID50, with 1329 ppm, followed by mustard with SO2 at 1, 3, and 5% (1 281, 1 308, and 1 298 ppm, respectively), while mustard with ZnO at 1% and 5% obtained an ID50 of 1 231 and 1 244 ppm. This coincides with what was reported by Siddiqui et al. (2019) in a study on the inhibitory effect of zinc nanoparticles, where it was found to be the best treatment on F. solani. Regarding sporulation activity, mustard treatments at the highest dose (500 ppm) showed total inhibition (Table 2).
For the extract of creosote bush with 3% SO2 at the dose of 5 000 ppm, it presented the lowest number of spores, 0.35 million conidia ml-1, followed by 5% SO2 at the dose of 3 000 ppm (0.48); the highest values were shown by creosote bush with 3% ZnO at the dose of 5 000 ppm, with 45.40 million ml-1, followed by 1% ZnO at the dose of 3 000 ppm (35.25), surpassing the control, which presented the highest value with 7.8 million conidia ml-1.
For mustard extract, the concentration of 5 000 ppm, either alone or in mixture with nanoparticles, inhibited the production of spores by 100%; other results with low values were shown by mustard alone at the dose of 3 000 ppm, with 0.37 million ml-1, followed by mustard plus 5% SO2 at the dose of 3 000 ppm with 0.48, and mustard with 3% SO2 at the dose of 3 000 ppm with 1.49.
These results are consistent with those reported by Singh et al. (2017), who tested 12 botanicals against Fusarium oxysporum, where mustard inhibited 93.75% of the fungus. Extracts of creosote bush have been used as an ecological strategy and have been shown as an alternative for integrated management for Fusarium (Peñuelas-Rubio) and these, being enriched with nanoparticles, increase their effectiveness for the control of F. solani.
The treatments win ZnO showed the highest values; with creosote bush, 70% of the results are higher than the control, where the lowest spore production was presented by creosote bush with 3% Zn at a concentration of 1 000 ppm, with 15.41 million conidia ml-1, while in those of mustard, the highest number of spores was observed at the concentration of 1 000 ppm, after the control, with 15.38 million conidia ml-1; Labiadh et al. (2016) reported that ZnO nanoparticles have a antibacterial effect better than an antifungal effect.
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