Seed biopriming with Trichoderma harzianum in soybean subjected to salt stressSoybean (Glycine max [L.] Merril) originating in East Asia, especially in Northeast China, is considered one of the most important crops in the world, mainly as a source of protein and vegetable oil (Silva et al., 2022). Production on Brazilian soil is a global highlight, where it covers approximately 40 million hectares, showing an increase of 3.8% compared to the previous harvest (Rocha et al., 2024).
However, abiotic factors considered stressful such as salinity can impair seed germination and seedling phenotypic responses. It is estimated that 20% of global agricultural land and 33% of irrigated land are affected by high salinity (Ludwig et al., 2023). Occurrence of salinity may be associated with natural salinity of soils, poorly managed fertilizers, inadequate irrigation and fertigation, problems with drainage, among other factors.
Salt stress in addition to increasing water deficit has negative effects on seed germination and vigor of seedlings. This effect is a response to oxidative and ionic stress (Dehnavi et al., 2020; Ren et al., 2020; Ors et al., 2021). The period from sowing to emergence is crucial for defining crop yield potential and plant stand in the area, making the implementation of methods that prevent and mitigate these stress-causing agents essential for the success of agriculture (Ludwig et al., 2023).
Fungi of the genus Trichoderma, such as the harzianum species, have great potential for use in agriculture. This group contributes to the control of pathogens in cultivated plants, acts as growth promoters and inducers of resistance and tolerance to isolated or multiple abiotic stresses (Maruyama et al., 2020; Pani et al., 2021; Woo et al., 2023). According to these authors, this group of organisms stimulates root growth by favoring the synthesis of hormones, in addition to favoring the increase in the activity of antioxidant enzymes that favor osmoregulation, with a mitigating effect for some plants.
The study of the use of these isolates has become increasingly frequent, with the aim of promoting seed germination and plant growth, especially owing to fact that for several cultivars there is insufficient information. We hypothesized that (i) the cultivar we studied is sensitive to salt stress, but (ii) biopriming with T. harzianum alleviates the stressful effect of this condition and (iii) the role of fungi varyng with the dose used. We aimed to evaluate the effect of doses of T. harzianum on seed germination and growth of soybean seedlings subjected to salt stress.
The test was carried out at the Plant Nutrition and Metabolism Laboratory, in the Faculty of Agricultural Sciences, Universidade Federal da Grande Dourados (UFGD), in Dourados municipally, Mato Grosso do Sul state, Brazil. We used seeds from cv. BMX NEXUS 64IX66 RSF I2X, which were previously subjected to biopriming with three doses of T. harzianum: CK (without biopriming), 5, and 10 mL kg seed–1, and rested for 30 minutes before sowing. Subsequently, seeds were subjected to four levels of osmotic potential: 0.0 (control, distilled water), –0.2, –0.4, and –0.6 MPa, induced by sodium chloride (NaCl).
The experimental design used was completely randomized, and treatments were arranged in a 3 × 4 factorial scheme, with four replications of 50 seeds. Trichoderma harzianum isolate used was IB19/17 (2 x 109 viable conidia g formulated product–1).
The germination test was carried out using Germitest® paper rolls moistened with distilled water or the corresponding saline solution, in an amount of 2.5 times the weight of the dry paper. Seeds were kept in B.O.D., at a constant temperature of 25 °C and under a 12-hour photoperiod. At 8 days, the percentage of germination was evaluated considering criteria established by Rules for Seed Analysis (Brasil, 2009). The germination speed index (GSI) was calculated according to Maguire (1962). At the end of test, ten seedlings were randomly evaluated, measuring the hypocotyl and radicle length, using a ruler, results expressed in centimeters.
The data were subjected to normality tests (Shapiro-Wilk) and homoscedasticity of variance (Hartley). All data were subjected to analysis of variance (F test, p ≤ 0.05), and the means of isolated effects and interactions were compared using the Tukey test (p ≤ 0.05) ± standard error for biopriming with T. harzianum and analysis of regression for osmotic potentials (p ≤ 0.05).
Based in our results, we partially accept our initial hypothesis, as the soybean cultivar studied is sensitive to salt stress, since the values of seed germination indicators and seedling growth decreased with linear adjustment as the osmotic potential increased, but the biopriming with T. harzianum did not mitigate the stressful effect. Under higher salinity level, represented here by –0.6 MPa, the germination and GSI values were lower with 10 mL of T. harzianum, while in CK the values were higher in the same condition (Figure 1). We observed the effect of salt stress on seed germination and GSI from the osmotic potential of –0.2 and –0.4 MPa, respectively.
Germination (a) and germination speed index – GSI (b) of soybean seeds subjected to biopriming with doses of Trichoderma harzianum (Th) and different osmotic potentials. Different letters compare the effect of T. harzianum doses in each osmotic potential by Tukey test (p ≤ 0.05) ± standard error. CK: control (without biopriming). * or ** (p ≤ 0.05 and 0.01, respectively).
Salinity promotes adverse effects on seed germination and seedling growth through physiological and biochemical changes, such as osmotic stress, ionic toxicity, and oxidative damage (Ren et al., 2020), reflecting delayed germination and abnormal seedlings. Responses at T. harzianum may varied, and for some species the inoculation with this fungus has contributed to defense induction responses and tolerance to several stresses for some species or cultivars (Sood et al., 2020; Naeimi et al., 2020; Tyśkiewicz et al., 2022), different from what was observed in our study, that this fungi isolate did not contribute to alleviating stressful effect of salinity.
Seedling length values showed the same response trend to germination indicators, with a reduction as osmotic potential increased (Figure 2). We observed a more pronounced reduction in hypocotyl than in radicle, resulting in lower phenotypic responses (Figure 3). Exposure to higher salinity levels promoted a reduction especially in rootlet lengths, in addition to the volume of secondary roots.
Radicle (a) and hypocotyl (b) length of soybean seedlings from seeds subjected to biopriming with doses of Trichoderma harzianum (Th) and different osmotic potentials. Different letters compare the effect of T. harzianum doses in each osmotic potential by Tukey test (p ≤ 0.05) ± standard error. CK: control (without biopriming). * or ** (p ≤ 0.05 and 0.01, respectively).
Phenotypic responses of soybean seedlings from seeds subjected to biopriming with Trichoderma harzianum (Th) and different osmotic potentials. (a) CK (control, without biopriming), (b) 5 mL, and (c) 10 mL of T. harzianum.
Why did biopriming with T. harzianum not alleviate the negative effects of salinity? We believe that the salinity level may have influenced the activity of the fungi! High salt concentration can inhibit the growth and functionality of microorganisms, including the genus Trichoderma, which can reduce their ability to promote germination and seedling growth (Otlewska et al., 2020; Zhang et al., 2021, 2024), and microorganisms can compete for limited resources with seeds, aggravating stress on plants (Dalling et al., 2011; Chepsergon and Moleleki, 2023).
Another important point to be highlighted is that the doses of T. harzianum used in our study for biopriming may have been high for the cultivar. Some Trichoderma strains can produce secondary compounds that can be phytotoxic at certain concentrations (Machado et al., 2012). In addition, under salt stress, seed germination may become more sensitive to these compounds, which could possibly explain the lower results with 10 mL of T. harzianum, especially compared to CK, whether or not they are exposed to salt stress.
In addition, Trichoderma can acidify the environment, which can influence the endogenous concentration of auxin, volatile organic compounds and other bioactive molecules, stimulating or suppressing different plant processes in a specific way (Pelagio-Flores et al., 2017; Nieto-Jacobo et al., 2017). Pelagio-Flores et al. (2017) observed that acidification by Trichoderma induced auxin redistribution within Arabidopsis columella root cap cells, promoting root tip curvature and growth inhibition.
In future perspectives, we suggest that new studies with lower doses than those tested should be carried out in order to verify the mitigation responses of this same cultivar. In addition, studies of antioxidant protection responses should be carried out in order to understand the mechanisms of action of T. harzianum on biochemical and physiological aspects of seeds. In conclusion, salt stress impaired seed germination and seedling length of the evaluated soybean cultivar and biopriming with T. harzianum did not alleviate the stressful effect of this adverse condition.
Acknowledgements
The authors thank CAPES and CNPq, for granting the scholarships, and the FUNDECT, for financial support.
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Publication Dates
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Publication in this collection
13 Jan 2025 -
Date of issue
2024
History
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Received
30 July 2024 -
Accepted
30 Oct 2024
