Aspergillus flavus is a common filamentous fungus widely present in the soil, air, and in crops. This facultative pathogen of both animals and plants produces aflatoxins, a group of mycotoxins with strong teratogenic and carcinogenic properties. Peanuts are highly susceptible to aflatoxin contamination and consumption of contaminated peanuts poses serious threats to the health of humans and domestic animals. Currently, the competitive displacement of aflatoxin-producers from agricultural environments by atoxigenic A. flavus is the most effective method of preventing crop aflatoxin contamination. In the current study, 47 isolates of A. flavus collected from peanut samples originating in Shandong Province were characterized with molecular methods and for aflatoxin-producing ability in laboratory studies. Isolates PA04 and PA10 were found to be atoxigenic members of the L strains morphotype. When co-inoculated with A. flavus NRRL3357 at ratios of 1:10, 1:1, and 10:1 (PA04/PA10: NRRL3357), both atoxigenic strains were able to reduce aflatoxin B1 (AFB1) levels, on both culture media and peanut kernels, by up to 90%. The extent to which atoxigenic strains reduced contamination was correlated with the inoculation ratio. Abilities to compete of PA04 and PA10 were also independently verified against local aflatoxin-producer PA37. The results suggest that the two identified atoxigenic strains are good candidates for active ingredients of biocontrol products for the prevention of aflatoxin contamination of peanuts in Shandong Province.
Microtubule is a well-known structural protein participating in cell division, motility and vesicle traffic. In this study, we found that β2 -tubulin, one of the microtubule components, plays an important role in regulating secondary metabolite deoxynivalenol (DON) biosynthesis in Fusarium graminearum by interacting with isocitrate dehydrogenase subunit 3 (IDH3). We found IDH3 negatively regulate DON biosynthesis by reducing acetyl-CoA accumulation in F. graminearum and DON biosynthesis was stimulated by exogenous acetyl-CoA. In addition, the expression of IDH3 significantly decreased in the carbendazim-resistant mutant nt167 (Fgβ2F167Y ). Furthermore, we found that carbendazim-resistance associated β2 -tubulin substitutions reducing the interaction intensity between β2 -tubulin and IDH3. Interestingly, we demonstrated that β2 -tubulin inhibitor carbendazim can disrupt the interaction between β2 -tubulin and IDH3. The decreased interaction intensity between β2 -tubulin and IDH3 resulted in the decreased expression of IDH3, which can cause the accumulation of acetyl-CoA, precursor of DON biosynthesis in F. graminearum. Thus, we revealed that carbendazim-resistance associated β2 -tubulin substitutions or carbendazim treatment increases DON biosynthesis by reducing the interaction between β2 -tubulin and IDH3 in F. graminearum. Taken together, the novel findings give the new perspectives of β2 -tubulin in regulating secondary metabolism in phytopathogenic fungi.
Abstract Microtubule is a well-known structural protein participating in cell division, motility and vesicle traffic. In this study, we found that β 2 -tubulin, one of the microtubule components, plays an important role in regulating secondary metabolite deoxynivalenol (DON) biosynthesis in Fusarium graminearum by interacting with isocitrate dehydrogenase subunit 3 (IDH3). We found IDH3 negatively regulate DON biosynthesis by reducing acetyl-CoA accumulation in F. graminearum and DON biosynthesis was stimulated by exogenous acetyl-CoA. In addition, the expression of IDH3 significantly decreased in the carbendazim-resistant mutant nt167 (Fgβ F167Y ). Furthermore, we found that carbendazim-resistance associated β 2 -tubulin substitutions reducing the interaction intensity between β 2 -tubulin and IDH3. Interestingly, we demonstrated that β 2 -tubulin inhibitor carbendazim can disrupt the interaction between β 2 -tubulin and IDH3. The decreased interaction intensity between β 2 -tubulin and IDH3 resulted in the decreased expression of IDH3, which can cause the accumulation of acetyl-CoA, precursor of DON biosynthesis in F. graminearum . Thus, we revealed that carbendazim-resistance associated β 2 -tubulin substitutions or carbendazim treatment increases DON biosynthesis by reducing the interaction between β 2 -tubulin and IDH3 in F. graminearum . Taken together, the novel findings give the new perspectives of β 2 -tubulin in regulating secondary metabolism in phytopathogenic fungi. Author Summary The deoxynivalenol (DON) biosynthesis is increased in carbendazim-resistant strains in Fusarium graminearum . To date, the molecular mechanism between the carbendazim-resistant substitution and the increased DON production remained elusive. Here we found that acetyl-CoA-associated enzyme IDH3 negatively regulates acetyl-CoA and DON biosynthesis. Moreover, β 2 tubulin interacted with IDH3 physically and increase its expression. We further found that carbendazim-resistant substitution in β 2 tubulin reducing the interaction between β 2 tubulin and IDH3, which resulted in the decreased expression of IDH3. In addition, we demonstrated that carbendazim disrupting the binding between β 2 tubulin and IDH3, which also decreases the expression of IDH3. Taken together, our results give a newly insights into the mechanism of β 2 tubulin and its carbendazim-resistant substitution in regulating DON biosynthesis.
Cytisine is a naturally occurring bioactive compound, an alkaloid mainly isolated from legume plants. In recent years, various biological activities of cytisine have been explored, showing certain effects in smoking cessation, reducing drinking behavior, anti-tumor, cardiovascular protection, blood sugar regulation, neuroprotection, osteoporosis prevention and treatment, etc. At the same time, cytisine has the advantages of high efficiency, safety, and low cost, has broad development prospects, and is a drug of great application value. However, a summary of cytisine's biological activities is currently lacking. Therefore, this paper summarizes the pharmacological action, mechanism, and pharmacokinetics of cytisine by referring to numerous databases, and analyzes the new and core targets of cytisine with the help of computer simulation technology, to provide reference for doctors.
Abstract In Fusarium graminearum , a trichothecene biosynthetic complex known as the toxisome forms ovoid and spherical structures in the remodelled endoplasmic reticulum (ER) under mycotoxin‐inducing conditions. Previous studies also demonstrated that disruption of actin and tubulin results in a significant decrease in deoxynivalenol (DON) biosynthesis in F. graminearum . However, the functional association between the toxisome and microtubule components has not been clearly defined. In this study we tested the hypothesis that the microtubule network provides key support for toxisome assembly and thus facilitates DON biosynthesis. Through fluorescent live cell imaging, knockout mutant generation, and protein–protein interaction assays, we determined that two of the four F. graminearum tubulins, α 1 and β 2 tubulins, are indispensable for DON production. We also showed that these two tubulins are directly associated. When the α 1 –β 2 tubulin heterodimer is disrupted, the metabolic activity of the toxisome is significantly suppressed, which leads to significant DON biosynthesis impairment. Similar phenotypic outcomes were shown when F. graminearum wild type was treated with carbendazim, a fungicide that binds to microtubules and disrupts spindle formation. Based on our results, we propose a model where α 1 –β 2 tubulin heterodimer serves as the scaffold for functional toxisome assembly in F. graminearum .
Sansevieria trifasciata originates from tropical West Africa. It is widely planted as a potted ornamental in China for improving indoor air quality (1). In February 2011, leaves of S. trifasciata plants in an ornamental market of Anle, Luoyang City, China, were observed with sunken brown lesions up to 20 mm in diameter, and with black pycnidia present in the lesions. One hundred potted plants were examined, with disease incidence at 20%. The symptomatic leaves affected the ornamental value of the plants. A section of leaf tissue from the periphery of two lesions from a plant was cut into 1 cm2 pieces, soaked in 70% ethanol for 30 s, sterilized with 0.1% HgCl2 for 2 min, then washed five times in sterilized distilled water. The pieces were incubated at 28°C on potato dextrose agar (PDA). Colonies of two isolates were brown with submerged hyphae, and aerial mycelium was rare. Abundant and scattered pycnidia were reniform, dark brown, and 200 to 350 × 100 to 250 μm. There were two types of setae on the pycnidia: 1) dark brown setae with inward curved tops, and 2) straight, brown setae. Conidia were hyaline, unicellular, cylindrical, and 3.75 to 6.25 × 1.25 to 2.50 μm. Morphological characteristics suggested the two fungal isolates were a Chaetomella sp. To confirm pathogenicity, six mature leaves of a potted S. trifasciata plant were wounded with a sterile pin after wiping each leaf surface with 70% ethanol and washing each leaf with sterilized distilled water three times. A 0.5 cm mycelial disk cut from the margin of a 5-day-old colony on a PDA plate was placed on each pin-wounded leaf, ensuring that the mycelium was in contact with the wound. Non-colonized PDA discs were placed on pin-wounded leaves as the control treatment. Each of two fungal isolates was inoculated on two leaves, and the control treatment was done similarly on two leaves. The inoculated plant was placed in a growth chamber at 28°C with 80% relative humidity. After 7 days, inoculated leaves produced brown lesions with black pycnidia, but no symptoms developed on the control leaves. A Chaetomella sp. was reisolated from the lesions of inoculated leaves, but not from the control leaves. An additional two potted plants were inoculated using the same methods as replications of the experiment, with identical results. To confirm the fungal identification, the internal transcribed spacer (ITS) region of rDNA of the two isolates was amplified using primers ITS1 and ITS4 (2) and sequenced. The sequences were identical (GenBank Accession No. KC515097) and exhibited 99% nucleotide identity to the ITS sequence of an isolate of Chaetomella sp. in GenBank (AJ301961). To our knowledge, this is the first report of a leaf spot of S. trifasciata caused by Chaetomella sp. in China as well as anywhere in the world. References: (1) X. Z. Guo et al. Subtropical Crops Commun. Zhejiang 27:9, 2005. (2) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, CA, 1990.
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