Optimization of the tetrazolium test for neotropical orchids: influence of preconditioning and temperature

Abstract

Orchids face significant conservation challenges due to habitat loss and illegal collection, making ex situ conservation strategies essential. This study aimed to optimize the tetrazolium test for four Orchidaceae species (Cattleya forbesii, C. wittigiana, Encyclia argentinensis, and Ionopsis utricularioides). The experiment followed a completely randomized design, in a 3 × 3 factorial arrangement (six replications), evaluating preconditioning (distilled water for 24 h, 10 % sucrose for 24 h, and control) and incubation temperatures (water bath at 40 ± 0.5 ºC for 24 h, oven at 40 ± 2 ºC for 24 h, and room temperature at 25 ± 2 ºC for 24 h). Estimated viability was validated by in vitro germination. At room temperature, the distilled water and sucrose yielded comparable results for C. forbesii, C. wittigiana, and I. utricularioides; whereas, for E. argentinensis, the sucrose resulted in the highest estimate. The tetrazolium test overestimated germination across all taxa, suggesting it detects metabolic activity rather than the complex sequence of events required for germination. The optimization was species-specific; however, the combination of distilled water and room temperature is recommended as a practical protocol for the studied species, with sucrose as an alternative when optimizing the test for E. argentinensis.

KEYWORDS: Ex situ conservation, germplasm, Orchidaceae, seed viability.