Complexity of Brassica oleracea–Alternaria brassicicola Susceptible Interaction Reveals Downregulation of Photosynthesis at Ultrastructural, Transcriptional, and Physiological Levels

Abstract

Black spot disease, caused by Alternaria brassicicola in Brassica species, is one of the most devastating diseases all over the world, especially since there is no known fully resistant Brassica cultivar. In this study, the visualization of black spot disease development on Brassica oleracea var. capitata f. alba (white cabbage) leaves and subsequent ultrastructural, molecular and physiological investigations were conducted. Inter- and intracellular hyphae growth within leaf tissues led to the loss of host cell integrity and various levels of organelle disintegration. Severe symptoms of chloroplast damage included the degeneration of chloroplast envelope and grana, and the loss of electron denseness by stroma at the advanced stage of infection. Transcriptional profiling of infected leaves revealed that photosynthesis was the most negatively regulated biological process. However, in infected leaves, chlorophyll and carotenoid content did not decrease until 48 hpi, and several chlorophyll a fluorescence parameters, such as photosystem II quantum yield (Fv/Fm), non-photochemical quenching (NPQ), or plant vitality parameter (Rdf) decreased significantly at 24 and 48 hpi compared to control leaves. Our results indicate that the initial stages of interaction between B. oleracea and A. brassicicola are not uniform within an inoculation site and show a complexity of host responses and fungal attempts to overcome host cell defense mechanisms. The downregulation of photosynthesis at the early stage of this susceptible interaction suggests that it may be a part of a host defense strategy, or, alternatively, that chloroplasts are targets for the unknown virulence factor(s) of A. brassicicola. However, the observed decrease of photosynthetic efficiency at the later stages of infection is a result of the fungus-induced necrotic lesion expansion.

The Effect of Photoperiod on Necrosis Development, Photosynthetic Efficiency and ‘Green Islands’ Formation in Brassica juncea Infected with Alternaria brassicicola

“…At the initial stages of B. oleracea infection, A. brassicicola induces gradual changes in the chloroplast ultrastructure, depending on the distance from the infecting hyphae. Most frequently, chloroplasts changed their shape from lenticular to round, disintegrate their envelope and stroma, as well as observe the disappearance of grana and damaged thylakoids [35]. Similarly, damaged chloroplasts with large starch grains or their hydrolyzed remains have been previously described at infection sites of B. juncea leaves infected with A. brassicicola [19], as well as in this study (Figure 8).…”

“…Reduction of the chl a:b ratio was mostly related to a significant decrease in chl a content. A similar trend of a decreased chlorophyll content has been described during progressive infection of B. oleracea with A. brassicicola [35] and B. juncea infected with A. brassicae [48]. Chlorophyll content decreased in tolerant and susceptible B. napus and B. rapa cultivars to A. brassicicola infection with much stronger degradation in susceptible cultivars [49].…”

“…A 12 h or longer light period per day caused a significant decrease in the maximum quantum yield of PSII after fungal infection, depending on the time after inoculation and leaf position (age of a leaf). A decrease in the F v /F m value caused by A. brassicicola has been previously observed in other species of the Brassicaceae family, e.g., B. oleracea [35], as well as in Arabidopsis thaliana [55]. The observed decrease in photochemical activity in the following hours after inoculation was probably the result of progressive degradation of chloroplasts and disturbance of the thylakoid system, as well as the development of necrotic lesions, which reduced the assimilation area of the leaf.…”

“…In general, necrotrophic fungi-induced lesions of foliar tissues often develop preferentially on older leaves of susceptible cultivars, whereas younger leaves show some level of tolerance/resistance to a fungus. Such a phenomenon is also characteristic for Brassica species (B. oleracea and B. rapa) infected with various pathogens, such as the oomycete Hyaloperonospora parasitica or a typical necrotroph such as A. brassicicola [33][34][35]. Moreover, a previous study revealed that necrotic lesions on B. juncea developed in a leaf position-dependent manner during A. brassicicola infection-the older the leaf, the larger the necrosis [19].…”

“…In many cases, damage to the photosynthetic apparatus caused by A. brassicicola or other Alternaria species has been correlated with an early increase in heat dissipation to protect PSII from further damage caused by excess excitation energy [54,60]. On the other hand, the PSII response to the stress caused by infection by necrotrophic fungi is also often associated with a rapid decrease in NPQ, both in the necrotic region and in adjacent areas of the leaf blade [35,61]. This is due to insufficient mobilization of the photoprotective mechanisms around the sites of inoculation [62].…”