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Aspergillus flavus grown in peptone as the carbon source exhibits spore density- and peptone concentration-dependent aflatoxin biosynthesis.

Yan S, Liang Y, Zhang J, Liu CM - BMC Microbiol. (2012)

Bottom Line: Metabolomic studies revealed that, in addition to inhibited AF biosynthesis, mycelia grown in peptone media with a high initial spore density showed suppressed fatty acid biosynthesis, reduced tricarboxylic acid (TCA) cycle intermediates, and increased pentose phosphate pathway products.Additions of TCA cycle intermediates had no effect on AF biosynthesis, suggesting the inhibited AF biosynthesis was not caused by depleted TCA cycle intermediates.This switching ability may offer Aspergillus species a competition advantage in natural ecosystems, producing AFs only when self-population is low and food is scarce.

View Article: PubMed Central - HTML - PubMed

Affiliation: Practaculture College, Gansu Agricultural University, Lanzhou 730070, China.

ABSTRACT

Background: Aflatoxins (AFs) are highly carcinogenic compounds produced by Aspergillus species in seeds with high lipid and protein contents. It has been known for over 30 years that peptone is not conducive for AF productions, although reasons for this remain unknown.

Results: In this study, we showed that when Aspergillus flavus was grown in peptone-containing media, higher initial spore densities inhibited AF biosynthesis, but promoted mycelial growth; while in glucose-containing media, more AFs were produced when initial spore densities were increased. This phenomenon was also observed in other AF-producing strains including A. parasiticus and A. nomius. Higher peptone concentrations led to inhibited AF production, even in culture with a low spore density. High peptone concentrations did however promote mycelial growth. Spent medium experiments showed that the inhibited AF production in peptone media was regulated in a cell-autonomous manner. mRNA expression analyses showed that both regulatory and AF biosynthesis genes were repressed in mycelia cultured with high initial spore densities. Metabolomic studies revealed that, in addition to inhibited AF biosynthesis, mycelia grown in peptone media with a high initial spore density showed suppressed fatty acid biosynthesis, reduced tricarboxylic acid (TCA) cycle intermediates, and increased pentose phosphate pathway products. Additions of TCA cycle intermediates had no effect on AF biosynthesis, suggesting the inhibited AF biosynthesis was not caused by depleted TCA cycle intermediates.

Conclusions: We here demonstrate that Aspergillus species grown in media with peptone as the sole carbon source are able to sense their own population densities and peptone concentrations to switch between rapid growth and AF production. This switching ability may offer Aspergillus species a competition advantage in natural ecosystems, producing AFs only when self-population is low and food is scarce.

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Metabolites with different contents in cultures initiated with high or low spore densities. (A) A PLS scores plot, performed using SIMCA-P V11.0, for metabolites extracted from mycelia cultured for 2, 3, 4 and 5 days in PMS media with initial spore densities of 104 (black) and 106 (gray) spores/ml, with 3 replicates in each treatment. (B) Scatter loading plots obtained from PLS analyses of the entire GC-Tof-MS dataset. (C and D) Total ion chromatographies of metabolites extracted from mycelia of A. flavus grown in PMS media for 3 days with the initial spore densities of 104 (C) and 106 spores/ml (D). Metabolites with significant differences in quantity between (C) and (D) are labeled.
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Figure 6: Metabolites with different contents in cultures initiated with high or low spore densities. (A) A PLS scores plot, performed using SIMCA-P V11.0, for metabolites extracted from mycelia cultured for 2, 3, 4 and 5 days in PMS media with initial spore densities of 104 (black) and 106 (gray) spores/ml, with 3 replicates in each treatment. (B) Scatter loading plots obtained from PLS analyses of the entire GC-Tof-MS dataset. (C and D) Total ion chromatographies of metabolites extracted from mycelia of A. flavus grown in PMS media for 3 days with the initial spore densities of 104 (C) and 106 spores/ml (D). Metabolites with significant differences in quantity between (C) and (D) are labeled.

Mentions: To determine metabolic differences in A. flavus grown in PMS media with high or low initial spore densities, metabolites in mycelia cultured for 2, 3, 4 and 5 days were analyzed by gas chromatography time-of-flight mass spectrometry (GC-Tof-MS) using methods described previously [49,50]. Multi-variate analyses showed that mycelia inoculated with 104 spores/ml clustered separately from mycelia inoculated with 106 spores/ml, suggesting evident metabolic differences between these two cultures (Figure 6A & B). Striking differences in levels were observed in 24 metabolites on the 3rd day (Figure 6C & D, and Table 1). In PMS cultures initiated with 106 spores/ml, a condition without AF production, the level of three TCA cycle intermediates, namely malic acid, fumaric acid and succinic acid, accumulated significantly less than those in cultures initiated with 104 spores/ml This suggests that the TCA cycle was more active in the high density culture. Similarly, levels of four fatty acids, palmitic acid, stearic acid, oleic acid and linoleic acid, were reduced in cultures initiated with the high spore density (Table 1), indicating that fatty acid biosynthesis was generally inhibited in the high density culture. In contrast, many sugar metabolites including ribitol, glucopyranoside, gluconolactone-6-P, glycerol, butanediamine, ethylamine and galactose, were accumulated more in the high density cultures (Table 1), suggesting that the PP pathway was active. In addition, nucleotides and compounds involved in amino acid metabolism were less abundant in cultures initiated with the high spore density (Table 1), which may be the consequence of the rapid mycelial growth.


Aspergillus flavus grown in peptone as the carbon source exhibits spore density- and peptone concentration-dependent aflatoxin biosynthesis.

Yan S, Liang Y, Zhang J, Liu CM - BMC Microbiol. (2012)

Metabolites with different contents in cultures initiated with high or low spore densities. (A) A PLS scores plot, performed using SIMCA-P V11.0, for metabolites extracted from mycelia cultured for 2, 3, 4 and 5 days in PMS media with initial spore densities of 104 (black) and 106 (gray) spores/ml, with 3 replicates in each treatment. (B) Scatter loading plots obtained from PLS analyses of the entire GC-Tof-MS dataset. (C and D) Total ion chromatographies of metabolites extracted from mycelia of A. flavus grown in PMS media for 3 days with the initial spore densities of 104 (C) and 106 spores/ml (D). Metabolites with significant differences in quantity between (C) and (D) are labeled.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3412747&req=5

Figure 6: Metabolites with different contents in cultures initiated with high or low spore densities. (A) A PLS scores plot, performed using SIMCA-P V11.0, for metabolites extracted from mycelia cultured for 2, 3, 4 and 5 days in PMS media with initial spore densities of 104 (black) and 106 (gray) spores/ml, with 3 replicates in each treatment. (B) Scatter loading plots obtained from PLS analyses of the entire GC-Tof-MS dataset. (C and D) Total ion chromatographies of metabolites extracted from mycelia of A. flavus grown in PMS media for 3 days with the initial spore densities of 104 (C) and 106 spores/ml (D). Metabolites with significant differences in quantity between (C) and (D) are labeled.
Mentions: To determine metabolic differences in A. flavus grown in PMS media with high or low initial spore densities, metabolites in mycelia cultured for 2, 3, 4 and 5 days were analyzed by gas chromatography time-of-flight mass spectrometry (GC-Tof-MS) using methods described previously [49,50]. Multi-variate analyses showed that mycelia inoculated with 104 spores/ml clustered separately from mycelia inoculated with 106 spores/ml, suggesting evident metabolic differences between these two cultures (Figure 6A & B). Striking differences in levels were observed in 24 metabolites on the 3rd day (Figure 6C & D, and Table 1). In PMS cultures initiated with 106 spores/ml, a condition without AF production, the level of three TCA cycle intermediates, namely malic acid, fumaric acid and succinic acid, accumulated significantly less than those in cultures initiated with 104 spores/ml This suggests that the TCA cycle was more active in the high density culture. Similarly, levels of four fatty acids, palmitic acid, stearic acid, oleic acid and linoleic acid, were reduced in cultures initiated with the high spore density (Table 1), indicating that fatty acid biosynthesis was generally inhibited in the high density culture. In contrast, many sugar metabolites including ribitol, glucopyranoside, gluconolactone-6-P, glycerol, butanediamine, ethylamine and galactose, were accumulated more in the high density cultures (Table 1), suggesting that the PP pathway was active. In addition, nucleotides and compounds involved in amino acid metabolism were less abundant in cultures initiated with the high spore density (Table 1), which may be the consequence of the rapid mycelial growth.

Bottom Line: Metabolomic studies revealed that, in addition to inhibited AF biosynthesis, mycelia grown in peptone media with a high initial spore density showed suppressed fatty acid biosynthesis, reduced tricarboxylic acid (TCA) cycle intermediates, and increased pentose phosphate pathway products.Additions of TCA cycle intermediates had no effect on AF biosynthesis, suggesting the inhibited AF biosynthesis was not caused by depleted TCA cycle intermediates.This switching ability may offer Aspergillus species a competition advantage in natural ecosystems, producing AFs only when self-population is low and food is scarce.

View Article: PubMed Central - HTML - PubMed

Affiliation: Practaculture College, Gansu Agricultural University, Lanzhou 730070, China.

ABSTRACT

Background: Aflatoxins (AFs) are highly carcinogenic compounds produced by Aspergillus species in seeds with high lipid and protein contents. It has been known for over 30 years that peptone is not conducive for AF productions, although reasons for this remain unknown.

Results: In this study, we showed that when Aspergillus flavus was grown in peptone-containing media, higher initial spore densities inhibited AF biosynthesis, but promoted mycelial growth; while in glucose-containing media, more AFs were produced when initial spore densities were increased. This phenomenon was also observed in other AF-producing strains including A. parasiticus and A. nomius. Higher peptone concentrations led to inhibited AF production, even in culture with a low spore density. High peptone concentrations did however promote mycelial growth. Spent medium experiments showed that the inhibited AF production in peptone media was regulated in a cell-autonomous manner. mRNA expression analyses showed that both regulatory and AF biosynthesis genes were repressed in mycelia cultured with high initial spore densities. Metabolomic studies revealed that, in addition to inhibited AF biosynthesis, mycelia grown in peptone media with a high initial spore density showed suppressed fatty acid biosynthesis, reduced tricarboxylic acid (TCA) cycle intermediates, and increased pentose phosphate pathway products. Additions of TCA cycle intermediates had no effect on AF biosynthesis, suggesting the inhibited AF biosynthesis was not caused by depleted TCA cycle intermediates.

Conclusions: We here demonstrate that Aspergillus species grown in media with peptone as the sole carbon source are able to sense their own population densities and peptone concentrations to switch between rapid growth and AF production. This switching ability may offer Aspergillus species a competition advantage in natural ecosystems, producing AFs only when self-population is low and food is scarce.

Show MeSH