A novel optical interference technique to measure minute root elongations of Japanese red pine ( Pinus densiflora Seibold & Zucc.) seedlings infected with ectomycorrhizal fungi

Abstract

This study presents a new technique to measure root elongation of Japanese red pine ( Pinus densiflora Seibold & Zucc.) seedlings with very high sensitivity in the order of sub-nanometer by using a novel optical interference method called statistical interferometry. The principle of the statistical interferometry is based on the statistics of a speckle field, which is generated when a rough surface is illuminated by a laser light. The technique facilitates to obtain minute root elongation measurements in the order of sub-seconds. The root elongation behavior of Pinus densiflora seedlings infected with ectomycorrhizal fungi, Pisolithus sp. ( Ps) and Cenococcum geophilum Fr. ( Cg), was investigated in comparison with that of an uninfected control. In the experiments, two points on a root with the separation of 3 mm were illuminated by laser beams and the elongation was measured continuously by analyzing speckle patterns successively taken by a CCD camera. The root elongation rate (RER), measured as the length of root elongation per second per millimeter (mean ± S.D.) for Ps-infected, Cg-infected and uninfected seedlings were 10.85 ± 2.41, 5.54 ± 1.43, and 2.41 ± 1.01 nm s −1 mm −1, respectively. We found that the RERs of seedlings infected with ectomycorrhizal fungi were significantly higher than that of the uninfected seedlings, and the seedlings infected with Ps fungi showed the highest RER. We conducted another experiment to observe two-dimensional root growth, in which the growth measurements were obtained for 4 months. From this experiment, we observed that root growth of ectomycorrhizal fungi infected seedlings were higher than that of the uninfected seedlings. The evaluation of results from these two techniques proves that the proposed statistical interferometry is much faster and very sensitive technique, where the time required for growth monitoring is 10 7 times less than the other. We can conclude that, at the scale of either very short time or relatively long time, the symbiotic relationship between root and ectomycorrhiza has a positive effect steadily on the root elongation.