Study On Pathophysiology Of Pinus Thunbergii And Pinus Massoniana Naturally Infected By Pine Wilt Nematode

Pine wilt disease is a fatal one caused by infection of the pine wilt nematode Bursaphelenchus xylophilus. The disease was first identified in Japan, where an outbreak occurred, and has been since spread to many countries including USA, Canada. Mexico. Portugal, South Korea and China. The pathogenesis of pine wilt is not clear, because it is affected by multiple factors including the plant hosts, longhorned beetles, nematodes and the microbes associated with them, as well as abiotic environmental factors. Pine wilt has posed enormous pressure on the ecosystem and caused huge economic loss in the disease epidemic countries. Since the beginning of the1980s’, pine wilt has emerged and became widespread in China. It has been continuously threatening and destroying the pine ecosystem, and is one of the most dangerous biological disasters in the forest in China.In this study, from a pine wilt epidemic area, we selected naturally infected Japanese black pine s (Pinus thunbergii) and Masson pines (P. massoniana) at different disease stages, isolated the nematode pathogen. and studied the host changes in photosynthesis and other physiological characteristics including color changes of needles and resinosis. The results are listed as follows:1. Under saturating light intensity, both of the two pine species had the highest net photosynthetic rate (Pn) at the onset of the disease (higher than those of healthy pines). and the pines showed certain extent of defense responses. With disease progression, the Pn of the two species decreased to negative values, indicating that nematode infection interfered with the photosynthesis of Japanese black pines and Masson pines. In addition, analysis of chlorophyll fluorescence indicated that the decreases in photosynthetic rates were due to the decrease in the efficiency of photochemical reactions (ΦPSII) in photosystem Ⅱ, which was caused by the declines in the efficiency of light harvesting (Fv’/Fm’) and the number of active PSII centers (qP).2. During the onset of disease, the chlorophyll and carotenoid contents, as well as the water content in the needles of the two pine species slightly increased, and then dropped, which caused wilt and decrease in photosynthetic rate. In the diseased pines, the activity of catalase (CAT) reduced, and the activities of peroxidase (POD) and superoxide dismutase (SOD) also deceased eventually after slight increases at the beginning of the disease development. These changes indicated that nematode infection destroyed the antioxidant enzyme systems of Japanese black pines and Masson pines. Although the host plants had compensation mechanisms in response to stress in the early stage of the disease, the self-protection function was diminished and lost ultimately with the substantial proliferation of the pine wilt nematodes.3. We have also analyzed the contents of reducing sugar and mineral nutrients in the diseased pine conifers. With disease progression, the contents of reducing sugar and nitrogen element decreased in both Japanese black pine and Masson pine. The changes in other minerals were inconsistent between the two types of pines. These results indicated that the structural damage caused by nematode infection significantly compromised metabolism, and affected the absorption, movement and utilization of different nutrients in the host plants.4. We have measured the reflective spectrum of the Japanese black pines and Masson pines infected by pine wilt nematodes using an ASD field spectrometer, and analyzed the spectral characteristic parameters and the changes in chlorophyll content. To better understand the relationship between the disease severity and reflective spectrum, different wavelengths were used for the measurement of pines that were infected for various time periods. Reflective spectrum curves obtained with wavelength in the mid-infrared range were useful in indicating the onset of pine wilt disease. Specifically, the changes in parameters such as position of red edge, the height of green peak, the red valley reflectivity, the slope of red edge, and water stress band reflectivity were consistent with the disease symptoms of the two types of pines. The chlorophyll contents in the two pine species decreased with disease progression, and were significantly and linearly correlated with the spectral characteristic parameters.5. Histopathological analyses showed typical lesions on the branches of Japanese black pine and Masson pine throughout the disease progression. During the early stages of disease development. the cell walls of cortical parenchyma cells on branches of Masson pines started to lignify. which was accompanied by cell ruptures and fusions to form cavities. During the middle stages of disease development, the cell walls of cortical parenchyma cells, periderm phelloderm cells, and phloem parenchyma cells also lignified. There was an increase in cell contents, and lignification was also observed in cambium cells. The resin canal deformed due to the rupture of surrounding cells. At the late stages of disease development, the majority of tissues outside the xylem were completely lignified and a large number of cells had ruptured. Japanese black pine and Masson pine had similar disease symptoms. Compared to Masson pine, the disease onset was delayed in Japanese black pine, and the disease was less severe. This might be due to the difference in resistance of the two different pine species to pine wilt disease. There was also difference in histopathology in sections obtained from upper, middle and lower canopies of the host plants at the same disease stage, which might be related to the differences in infection location, as well as the number and distribution of pine wood nematodes in the plants.