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The death and structural alterations in the secondary xylem of Gmelina arborea trees in response to fungal invasion were investigated by light and electron microscopy methods. In vivo experiments indicate that Alternaria alternata is an aggressive pathogen which invades the deeper part of the wood in the branches––up to 4 cm from the site of infection––within 10 days of inoculation. The mycelial invasion occurred through pits and boreholes formed by the fungus in the vessels, fibers, and axial and ray parenchyma cells. Scanning electron microscopy also revealed the colonization of fungus through pit and vessel elements. In vitro studies showed that ray cells and fibers were more prone to cell wall damage in contrast to vessels during the initial stage of infection. The etiology of infection found in the infected samples collected from the living trees and samples of in vivo and in vitro experiments showed similar patterns of infection and confirmed that A. alternata is responsible for the mortality and decay in Gmelina trees. It causes a typical soft-rot pattern with cell wall alterations such as cavity formation by “L” bending. The separation of fibers at the compound middle lamella region indicated that the fungus is capable of degrading both lignin-rich regions of cell walls. The boreholes in the fiber wall also showed distinct variations in size and shape. The penetration of fungal hyphae through the middle lamella and removal of lignin from the cell walls was also confirmed with transmission electron microscopy studies.