*3.2. Pathogenicity Tests*

Koch's postulates confirmed that "PUE 4.0" strain showed typical symptoms of charcoal rot 20 days after inoculation. In addition, microsclerotia were observed in the vascular system (Xylem) which caused an upward wilt of stem, as shown in Figure 2b. No symptoms were observed in the control group.

**Figure 2.** Pathogenicity tests with "PUE 4.0" strain: (**a**) plant of *Arachis hypogea* with charcoal rot and death of foliage at 20 days after inoculation; (**b**) cross-section of peanut root showing rot and the presence of microsclerotia; (**c**) control group without symptoms.

> Using PCR of repetitive sequence, it was possible to confirm the identity of the reisolation (PUE 4.1) of the original strain. This sequence was deposited in the same database, with the accession number MW585379.

#### *3.3. Percentage of Growth Inhibition In Vitro*

Areas of interaction were observed between *T. harzianum* (T-H3), *T. asperellum* (T-AS1), *T. hamatum* (T-A12), *T. koningiopsis* (T-K11), and the native strain of *T. harzianum* (T-Ah) against *M. phaseolina* (MW585378), where parasitism greater than 50% was obtained at 240 h.

Development rate and the growth speed had significant differences (*p* ≤ 0.05), where *T. koningiopsis* (T-K11) obtained the highest value (Table 2) with 2.18 ± 0.035 mm/hour and 2.23 ± 0.013 cm/d−1, respectively. *M. phaseolina* showed the lowest growth speed (1.67 ± 0.054 cm/d<sup>−</sup>1).

**Table 2.** Rate of development, growth speed, and percentage of inhibition of radial growth and antagonism classification on Bell Scale [34].


\* Media followed by the same letter do not present significant statutory differences (*p* ≤ 0.05) according to Tukey's test. Means followed by the same letter (a, b and c) are not significantly different for *p* ≤ 0.05 according to Tukey test.

The percentage of inhibition of radial growth (PIRG) presents significant differences (*p* = 0.0001). The double confrontation between the different *Trichoderma* species showed an inhibition greater than 50% from the tenth day (Table 2). However, the highest percentage of inhibition was obtained with *T. koningiopsis* (T-K11) obtaining 71.11%. Similarly, *T. harzianum* (T-H3) presented the second-best inhibition with 63.55%; both antagonistic strains presented a class I classification (Figure 3) according to the scale established by Bell et al. [34].
