Season, thermotherapy and surface sterilization play important roles in microbial contamination of hazelnut in vitro cultures

Abstract

The initiation of cultivars for hazelnut micropropagation from mature field-grown plants is challenging due to high contamination levels and low regeneration rates. This study aimed to develop an efficient protocol for establishing in vitro cultures, the crucial first step in hazelnut micropropagation. Three independent experiments were conducted to achieve this goal. In the first experiment, plant materials were collected directly from the field at different times of the year to initiate cultures, with July to August identified as the most suitable period. In the second experiment, one-year-old potted suckers underwent a sanitation program in the greenhouse and were subjected to thermotherapy in the growth chamber before being utilized as explant sources. Nodal explants obtained from these plants were then used to initiate cultures. In the third experiment, various doses and durations of sterilization agents were applied to optimize surface sterilization. The combination of 1% silver nitrate (AgNO3), 0.1% mercury chloride (HgCl2), and 3% hydrogen peroxide (H2O2) after treatments with 70% ethanol and 2% sodium hypochlorite (NaOCl) yielded the best results, with the highest percentage of viable shoots. Overall, thermotherapy treatment to the mother plant was the most effective method for initiating cultures from plants in highly contaminated environments. While cultures initiated with explants from field-grown plants exhibited 53.7% visually clean shoots, this rate increased to 97.4% in plants subjected to thermotherapy. This study, for the first time, evaluated the effectiveness of applying thermotherapy to mother plants to reduce the incidence of microbial contaminants in the establishment stage of hazelnut in vitro propagation.