ULTRASTRUCTURAL EFFECTS OF AAL-TOXIN T-A FROM THE FUNGUS ALTERNARIA-ALTERNATA ON BLACK NIGHTSHADE (SOLANUM-NIGRUM L.) LEAF DISCS AND CORRELATION WITH BIOCHEMICAL MEASURES OF TOXICITY

Ultrastructural effects of AAL-toxin T-A from Alternaria alternata on black nightshade (Solanum nigrum L.) leaf discs and correlation with b iochemical measures of toxicity. In black nightshade (Solanum nigrum L .) leaf discs floating in solutions of AAL-toxin T-A (0.01 - 200 mu M) under continuous light of 25 degrees C, electrolyte leakage, chloroph yll loss, autolysis, and photobleaching were observed within 24 h. Ele ctrolyte leakage, measured by the conductivity increase in the culture medium, began after 12 h with 200 mu M AAL-toxin T-A, but was observe d after 24 h with 0.01 to 50 mu M AAL-toxin T-A, when it ranged from 2 5% to 63% of total releasable electrolytes, respectively. After 48 h i ncubation, leakage ranged from 39% to 79% of total for 0.01 to 200 mu M AAL-toxin T-A, respectively, while chlorophyll loss ranged from 5% t o 32% of total, respectively. Ultrastructural examination of black nig htshade leaf discs floating in 10 mu M AAL-toxin T-A under continuous light at 25 degrees C revealed cytological damage beginning at 30 h, c onsistent with the time electrolyte leakage and chlorophyll reduction were observed. After 30 h incubation chloroplast starch grains were en larged in control leaf discs, but not in AAL-toxin T-A-treated discs, and the thylakoids of treated tissue contained structural abnormalitie s. After 36-48 incubation with 10 mu M AAL-toxin T-A, all tissues were destroyed with only cell walls, starch grains, and thylakoid fragment s remaining. Toxicity was light-dependent, because leaf discs incubate d with AAL-toxin T-A in darkness for up to 72 h showed little phytotox ic damage. Within 6 h of exposure to greater than or equal to 0.5 mu M toxin, phytosphingosine and sphinganine in black nightshade leaf disc s increased markedly, and continued to increase up to 24 h exposure. T hus, physiological and ultrastructural changes occurred in parallel wi th disruption of sphingolipid synthesis, consistent with the hypothesi s that AAL-toxin T-A causes phytotoxicity by interrupting sphingolipid biosynthesis, thereby damaging cellular membranes. (C) 1998 Elsevier Science Ltd. All lights reserved.