Use of antimicrobial agents and ultrasound to control endogenous contaminants in the micropropagation of a hybrid clone of Eucalyptus urophylla x E. globulus

Abstract

Micropropagation is a technique widely applied aiming at the large-scale multiplication of selected individuals. However,
challenges such as endogenous microbial contamination can lead to substantial losses of plant materials. Endophytic
contamination consists of resident microorganisms that actively participate in the plant’s metabolism, making them difficult to control. In this context, the use of antimicrobial agents in plant tissue culture has increased as an alternative approach to optimize clonal cleaning. Additionally, the application of ultrasound (sonication) may enhance the penetration and effectiveness of these agents within plant tissues. Therefore, the present study aimed to evaluate the effects and efficacy of disinfectant agents combined with ultrasound in controlling endophytic microbiota during the micropropagation of a hybrid clone of Eucalyptus urophylla × Eucalyptus globulus. Explants were inoculated into JADS culture medium supplemented with two β-lactam antibiotics (Timentin® and Cefotaxime®) and two biocides [PPM® (Plant Preservative Mixture) and FZ (Fungozur B7DM)], combined with three ultrasound exposure times (0, 9, and 18 s), for a total of 15 treatments, including the control. At 15 and 30 days of in vitro culture, survival rate (%), contaminated area (mm²), rooting rate (%), height, shoot length (mm), and callus formation (%) were evaluated. The use of FZ or PPM® resulted in lower contamination rates; however, FZ promoted taller and more developed plantlets. The use of ultrasound sonication increased the level of contamination and did not significantly affect explant development. For clonal cleaning of the Eucalyptus hybrid, the use of FZ without sonication is recommended, regardless of the duration of in vitro culture.

Keywords: Disinfestation; Clonal Cleansing; Endophytic; Biocide; Sonication.