Endophytic colonization of five Trichoderma species and their effects on growth of a Eucalyptus hybrid

Abstract

The study aimed to evaluate the effectiveness of endophytic colonization via leaf and root inoculation of five Trichoderma species in a Eucalyptus hybrid, as well as the effects of inoculation on plant growth. The experimental design was completely randomized in a 6 × 2 factorial scheme. Plant growth was evaluated during the experimental period at three different times: 20 days after inoculation (d.a.i), 40 d.a.i., and 60 d.a.i. A statistical difference was observed between the inoculation methods during each period and between the Trichoderma species. Plants inoculated with T. asperellum showed the greatest growth among the treatments. Root-inoculated plants produced the greatest growth response. This showed that the presence of Trichoderma in the roots assisted in nutrient assimilation, promoted greater plant growth, when compared with leaf-inoculated plants. Evaluation of the effectiveness of endophytic colonization was performed at each sampling period by collecting leaf samples, and at 60 d.a.i., by collecting leaf, stem, and root samples. T. longibrachiatum and T. harzianum were isolated from leaves at 20 d.a.i., with an increase in the number of colonized plants throughout the evaluation of leaf-inoculated plants. In root-inoculated plants, treatment with T. longibrachiatum, T. harzianum, and T. asperellum presented the highest endophytic colonization in the stem and root samples (at 60 d.a.i.).

Keywords: Endophytic colonization; Eucalyptus spp.; Symbiotic relationships; Trichoderma spp.; Vegetative growth.

Fig. 1

Height of Eucalyptus plants (cm) endophytically colonized by Trichoderma isolates at 20 (a), 40 (b), and 60 (c) d.a.i. when leaf-inoculated, and 20 (d), 40 (e), and 60 (f) d.a.i when root-inoculated. Uppercase letters represent interactions between days after inoculation (p < 0.05), lowercase letters represent Trichoderma species interactions (p < 0.05), and * represents interactions between inoculation methods (p < 0.05)

Fig. 2

Stem diameter of Eucalyptus plants (mm) endophytically colonized by Trichoderma isolates at 20 (a), 40 (b), and 60 (c) d.a.i. when leaf-inoculated, and 20 (d), 40 (e), and 60 (f) d.a.i when root-inoculated. Uppercase letters represent interactions between days after inoculation (p < 0.05), lowercase letters represent Trichoderma species interactions (p < 0.05), and * represents interactions between inoculation methods (p < 0.05)

Fig. 3

Number of leaves on Eucalyptus plants endophytically colonized by Trichoderma isolates at 20 (a), 40 (b), and 60 (c) d.a.i. when leaf-inoculated, and 20 (d), 40 (e), and 60 (f) d.a.i when root-inoculated. Uppercase letters represent interactions between days after inoculation (p < 0.05), lowercase letters represent Trichoderma species interactions (p < 0.05), and * represents interactions between inoculation methods (p < 0.05)

Fig. 4

Number of branches on Eucalyptus plants endophytically colonized by Trichoderma isolates at 20 (a), 40 (b), and 60 (c) d.a.i. when leaf-inoculated, and 20 (d), 40 (e), and 60 (f) d.a.i when root-inoculated. Uppercase letters represent interactions between days after inoculation (p < 0.05), lowercase letters represent Trichoderma species interactions (p < 0.05), and * represents interactions between inoculation methods (p < 0.05)

Fig. 5

Total fresh biomass (g) of Eucalyptus plants endophytically colonized by Trichoderma isolates at 60 d.a.i., leaf-inoculates (a) and, root-inoculated (b). The black bars represent the mean fresh mass of roots, and the white bar represents the mean fresh mass of shoots (the sum of the mass of stems, branches, and leaves). The bars when stacked (black bar + white bar) corresponds to the average of total fresh biomass. Uppercase letters represent significant interactions between inoculation methods (p < 0.05), and lowercase letters represent significant interactions between Trichoderma species (p < 0.05)

Fig. 6

Total dry biomass (g) of Eucalyptus plants endophytically colonized by Trichoderma isolates at 60 d.a.i., leaf-inoculated (a) and, root-inoculated (b). The black bars represent the mean dry mass of roots, and the white bar represents the mean dry mass of shoots (the sum of the mass of stems, branches, and leaves). The bars when stacked (black bar + white bar) corresponds to the average of total dry biomass. Uppercase letters represent significant interactions between inoculation methods (p < 0.05), and lowercase letters represent significant interactions between Trichoderma species (p < 0.05)

Fig. 7

Sections of Eucalyptus plants 60 d.a.i. with presence of Trichoderma species. a Trichoderma harzianum present in leaf and stem sections of plants treated with leaf inoculation method. b T. longibrachiatum present in stem and root sections of plants treated with root inoculation method. c T. asperellium present in root sections of plants treated with root inoculation method

Fig. 8

Sections of Eucalyptus plants 60 d.a.i. with presence of Trichoderma species. a Visualization of the morphological structure of Trichoderma harzianum grown in Petri dishes isolated from parts of leaves and branches. b Conidia and conidiophores of T. harzianum isolated from leaves visualized in slides under optical microscope (×400 magnification)