Structural changes in ‘Newhall' navel oranges infected with Candiatus Liberibacter Asiaticus

Abstract:【Objective】The objective of this study is to determine the effects of an infection of‘Newhall'navel oranges by Citrus Huanglongbing on the morphology of microtissues and physiological and biochemical activities in order to provide insights on how the pathogen causes damage to the host.The characteristics and mechanism of the damage to the host that is infected would be further analyzed, which could provide some useful information for the prevention and control research of Citrus Huanglongbing.【Methods】Infected leaves and fruit with typical symptoms of Citrus Huanglongbing were collected from an experimental orchard, the apparent healthy leaves and fruit were used as the control.After being wiped clean, the blade tissues 1 cm ×1 cm from both sides of the central leaf veins of the leaves were quickly cut. The pedicel was cut into 1-2 mm thickness from the sheet and put on a platform with conductive adhesive for spraying. Then, the FEI-450 type scanning electron microscope was used to observe and photograph the stomata and pedicel cell the morphology map. After being embedded in paraffin, the leaves and the pedicel were cut into a thickness of 6 μ m sized slices by using a Leica RM2245 slicing machine and the anatomical structure was observed by a Leica DMIL-LED fluorescence inverted microscope after being stained with saffron. The thickness of the phloem, xylem and medullary part, palisade tissue and spongy tissue of the leaves were measured by using the LAS V4.0 and Image-Pro Plus 6.0 software. Contents of the photosynthetic pigments and malondialdehyde (MDA) were measured, and the activities of peroxidase (POD) , catalase (CAT) , and superoxide dismutase (SOD) were recorded. The experiment was repeated three times, and results were analyzed using the SPSS12.0 software.【Results】Observations with a scanning electron microscopy showed that the surrounding tissues of the stomata of the healthy leaves were explanated and smooth, while the sto-mata of the infected leaves were cavated and atrophic, and the surrounding tissues were crinkled, rough and uneven. Moreover, the phloem, xylem and pulp in the diseased leaves became significantly thinner than that of the healthy ones, in which the thickness of the phloem in the diseased leaves was 72.3% of that in the healthy tissue and the thickness of the xylem and pulp in the diseased leaves was 81.2% of that in the healthy tissue. There was no significant change in the thickness of the palisade tissue and the sponge tissue. The cells of the palisade tissue in healthy leaves were long striped, regular and closely arranged, and the intercellular space was small, however, the cells of the palisade tissue in the diseased leaves were elliptical, loosely arranged and had large spaces between the cells. The cells of the healthy pedicel were smooth and their sizes were relatively similar, but the cells infected by Citrus Huanglongbing were uneven, folded, became distorted, swollen, and loose, leaving large intercellular spaces among the cells. The vascular bundles of the infected pedicel were blocked. The content of the photosynthetic pigments in the infected leaves decreased compared to that of the healthy leaves. The contents of the chlorophyll a, chlorophyll b and carotenoid were 19.87%, 22.13% and 47.71% of that in the healthy leaves, respectively. In contrast, the content of the malondialdehyde in the diseased leaves increased significantly, by 120.39%, compared to that of the healthy leaves. The enzymatic activity of POD, CAT and SOD in the leaves of the infected plants were significantly higher than those of the healthy plants, among which the activity of POD increased by 8.43 times.【Conclusion】Our results showed that the infection of leaves and fruit by Citrus Huanglongbing could cause damages to various tissues in the host, which decreased the content of the photosynthetic pigments while significantly increasing the enzymatic activities, which led to more cells being damaged and also weakened their ability to resist the invasion and expansion of pathogenic bacteria.