Superoxide radical induces sclerotial differentiation in filamentous phytopathogenic fungi: a superoxide dismutase mimetics study

Abstract

This study shows that the superoxide radical (O 2˙" ), a direct indicator of oxidative stress, is involved in the differentiation of the phytopathogenic filamentous fungi Rhizoctonia solani, Sclerotinia sclerotiorum, Sclerotium rolfsii and Sclerotinia minor, shown by using superoxide dismutase (SOD) mimetics to decrease their sclerotial differentiation. The production rate of O 2˙" and SOD levels in these fungi, as expected, were significantly lowered by the SOD mimetics, with concomitant decrease of the indirect indicator of oxidative stress, lipid peroxidation. INTRODUCTIONSclerotial differentiation is a primitive form of differentiation exhibited by certain phytopathogenic filamentous fungi. It is expressed by four main types of sclerotia (compact bodies of aggregated hyphae), which are loose, terminal, lateral-chained and lateral-simple. These types are represented mainly, but not solely, by Rhizoctonia solani, Sclerotinia sclerotiorum, Sclerotium rolfsii and Sclerotinia minor, respectively (Chet et al., 1967(Chet et al., , 1969Chet & Henis, 1975;Le Tourneau, 1979;Willetts, 1971Willetts, , 1978Willetts, , 1997Willetts & Wong, 1980). These fungi are of great agricultural interest because they cause stem-rot diseases in a wide variety of economically important agricultural crops. Therefore, elucidating the mechanisms of sclerotium biogenesis of these fungi may lead to the development of non-toxic ways to combat plant diseases, which could be used as alternatives to the traditional toxic fungicides. Moreover, understanding sclerotial differentiation is very important because it will help to understand more complex forms of differentiation in other organisms as well as whether their mechanisms share common biochemical pathways.It has been proposed that sclerotial differentiation is related to high oxidative stress (Georgiou, 1997), based on the finding that sclerotium biogenesis in S. rolfsii was accompanied by the accumulation of high levels of lipid peroxidation products. This finding has been supported by other studies that developed into a theory of oxidative stress as an inducer of sclerotial metamorphosis (Georgiou et al., 2006). Specifically, hydroxyl radical scavengers, vitamin C and b-carotene (both also acting as endogenous fungal antioxidants), decreased sclerotial differentiation in S. rolfsii, S. minor, S. sclerotiorum and R. solani. In more recent studies, we demonstrated a strong relationship between the thiol redox state of these fungi and their sclerotial differentiation (Patsoukis & Georgiou, 2007a, b, 2008a. Specifically, in R. solani, S. rolfsii and S. minor, there was a decrease in the total amount of reduced thiols during transformation of the undifferentiated mycelium into sclerotium, which, for S. sclerotiorum was accompanied by an increase in the total oxidized components. Moreover, the antioxidant thiol redox state modulator N-acetylcysteine inhibited sclerotial differentiation in S. rolfsii, R. solani and S. sclerotiorum.Until now, our studies have associated only indirect indicators ...

Functional Analysis of the Trichoderma harzianum nox1 Gene, Encoding an NADPH Oxidase, Relates Production of Reactive Oxygen Species to Specific Biocontrol Activity against Pythium ultimum

“…We observed motifs for binding the regulators of oxygen metabolism, sexual development, and defense PR proteins in the nox1 promoter region. These data are in agreement with the role of Nox proteins as critical enzymes for sexual development (2,24,30) and sclerotial differentiation (39) in filamentous fungi, plant infection processes (10), and antimicrobial activity due to their involvement in ROS production (15,51).…”

“…It is well documented that ROS can induce sclerotial metamorphosis in R. solani (39), but this pathogen can also repress ROS formation through the production of the scavenger ascorbic acid in certain concentration gradients and proportions in response to oxidative stress caused by ROS during mycelial and sclerotial differentiation (13). On the other hand, B. cinerea seems to be better adapted to oxidative stress than other phytopathogens since it has been observed that ROS function generated by Nicotiana benthamiana has a positive role in expansion of disease lesions during B. cinerea-plant interaction (3).…”

Role of Oxidative Stress in Sclerotial Differentiation and Aflatoxin B1 Biosynthesis in Aspergillus flavus

“…Based on previous studies in sclerotiogenic fungi, high levels of H 2 O 2 inhibit SD because H 2 O 2 promotes intense cell proliferation throughout the whole mycelial colony. Such extensive cell proliferation surpasses the localized cell proliferation that is required for the generation of sclerotial initials (that are developed to mature sclerotia) (4,36,(46)(47)(48). Therefore, our data suggest that the increase in the SOD activity, combined with the decrease in CAT activity in the mutant strain compared to the control and the consequent increase in the concentration of H 2 O 2 , could contribute to the prevention of SD in the ⌬veA mutant strain.…”

“…Enzyme activities were expressed per protein, the concentration of which was determined as previously described (28). The SOD specific activity was assayed as previously reported (36) and expressed as SOD units per mg of protein (using a standard curve made with pure bovine erythrocyte SOD). The CAT specific activity was measured with a Clark-type O 2 electrode (controlled by Oxygraph Plus software [Hansatech Instruments Co., England] provided by Hansatech) by monitoring the decomposition of H 2 O 2 to O 2 (in a 1-ml assay solution containing 0.3 mM H 2 O 2 in 10 m⌴ phosphate buffer [pH 7.0] with or without a 1 mM concentration of the CAT inhibitor ATR).…”

Reproduction without sex: conidiation in the filamentous fungus Trichoderma

“…Oxidative stress has been suggested to be involved in the differentiation of several fungi, including Aspergillus nidulans, N. crassa, Rhizoctonia solani, Sclerotinia sclerotiorum and Sclerotinia minor, and in Sclerotium rolfsii, sclerotial biogenesis in response to mycelial injury occurs as a result of oxidative stress (Aguirre, et al, 2005;Georgiou et al, 2006;Papapostolou & Georgiou, 2010).…”