THERMOTHERAPY AS A MICROBIAL CONTAMINANT-REDUCING AGENT IN MICROPROPAGATION OF BAMBOO

ABSTRACT The aim of the research was to evaluate the effectiveness of thermotherapy by humid heat of nodal segments to reduce the incidence of microbial contaminants in the introduction phase of the in vitro propagation of Bambusa vulgaris. Two independent experiments were carried out in a completely randomized design using a 4 × 2 factorial arrangement, four levels of temperature (25, 30, 40 and 50 °C) versus two treatment time intervals (5 or 10 min). Each treatment was composed of nine replications, corresponding to one single-nodal segment obtained from primary or secondary branches, in sucrose-free Murashige and Skoog medium with half the concentration of salts, supplemented with 6.5 g L-1 agar, 50 mg L-1 citric acid, 50 mg L-1 ascorbic acid, 200 mg L-1 chloramphenicol and 2 mg L-1 N6-benzylaminopurine. Isolation of prevalent mold fungi per repetition was conducted, and identification at genus level obtained from microcultures. The treatments corresponding to 50 °C for 5 or 10 min reduced significantly the fungal contamination that occurred at 11% and 0%, respectively, in nodal segments from primary branches, and 0% and 11%, respectively, in those from secondary ones. Fungi of genera Alternaria, Bipolaris and Curvularia were isolated from both types of segment, and Cladosporium from secondary branches only. The lowest percentages of mycelial growth detected in nodal segments submitted to both binomials demonstrated the effectiveness of the technique to reduce fungal contamination despite negative interference on sprouting.

摘要 本研究旨在评估采用带节茎段湿热热疗，降低寻常竹（Bambusa vulgaris）离体繁殖启动阶段微生物污染发生率的效果。本研究采用4×2因子设计的完全随机试验方案，开展了两组独立实验，设置4个温度梯度（25、30、40和50℃）与2个处理时长（5min或10min）。每个处理设置9次重复，外植体为取自主枝或侧枝的单节茎段，接种于盐浓度减半的无蔗糖MS培养基（Murashige and Skoog medium）中，该培养基添加了6.5 g·L⁻¹琼脂、50 mg·L⁻¹柠檬酸、50 mg·L⁻¹抗坏血酸、200 mg·L⁻¹氯霉素以及2 mg·L⁻¹ 6-苄氨基腺嘌呤（N6-benzylaminopurine）。对每个重复的优势霉菌进行分离，并通过微培养完成属水平鉴定。采用50℃分别处理5min与10min的实验组，可显著降低真菌污染：取自主枝的带节茎段污染率分别降至11%与0%，取自侧枝的带节茎段污染率则分别降至0%与11%。从两类带节茎段中均分离出了链格孢属（Alternaria）、平脐蠕孢属（Bipolaris）以及弯孢霉属（Curvularia）真菌，而枝孢属（Cladosporium）真菌仅从侧枝来源的茎段中分离得到。经两类温度-时长组合处理的带节茎段中，菌丝生长率均为最低，这表明尽管该技术会对萌芽产生负面干扰，但其降低真菌污染的效果显著。