Isolation of Bioactive Compounds, Antibacterial Activity, and Action Mechanism of Spore Powder From Aspergillus niger xj

Abstract

Aspergillus fungi can produce a wide range of secondary metabolites, and they have represented a potential resource of novel bioactive compounds. Bacterial plant diseases have a serious impact on the sustainable development of agriculture worldwide, so it is necessary to use natural antibacterial compounds in microorganisms to control plant pathogens. This study was conducted to investigate the bioactive compounds of Aspergillus niger xj, three plant pathogens (Agrobacterium tumefaciens T-37, Erwinia carotovora EC-1, and Ralstonia solanacearum RS-2) were used as indicator bacteria, according to the biological activity tracking, five compounds were isolated from A. niger xj spore powder, and characterization of compounds was done by NMR (¹H-NMR and ¹³C-NMR) and EI-MS and was identified as ergosterol (1), β-sitosterol (2), 5-pentadecylresorcinol (3), 5-hydroxymethyl-2-furancarboxylic acid (4), and succinimide (5). Compounds 3 and 5 were isolated from A. niger xj for the first time. The minimum inhibitory concentration (MIC) of five compounds against three plant pathogens was evaluated, the results showed that compound 4 exhibited the strongest antibacterial activity against tested bacteria, and RS-2 was the most sensitive to compound 4, showing the lowest MIC of 15.56 μg/ml. We concluded that the mechanism of action of the compound 4 against RS-2 might be described as compound 4 acting on bacterial protein synthesis and intracellular metabolism according to the results of the scanning electron microscopy observation, permeability of cell membrane and SDS-PAGE. These results indicated that compound 4 has good potential to be as a biocontrol agent. In conclusion, the results from this study demonstrated that the compounds with antibacterial activity are of great significance of the prevention and control of plant phytopathogenic bacteria, and they may be applicable to exploring alternative approaches to integrated control of phytopathogens.

Keywords: Aspergillus niger xj; antibacterial activity; antibacterial mechanism; bioactive compounds; isolation and identification; minimum inhibitory concentration.

FIGURE 1

The antibacterial activity guided fractionation of petroleum ether extract.

FIGURE 2

Antibacterial activities of the organic extract fractions of the spore powder against the indicator bacteria. Different lowercase letters indicate significant differences (p < 0.05). T-37, EC-1, and RS-2 represent Agrobacterium tumefaciens T -37, Erwinia carotovora EC-1, and Ralstonia solanacearum RS-2, respectively. DMSO represents dimethyl sulfoxide. CHL represents chloramphenicol.

FIGURE 3

Antibacterial activities of the subfractions A1–A11 of the spore powder against the indicator bacteria. Different lowercase letters indicate significant differences (p < 0.05).

FIGURE 4

The chemical structures of compounds isolated from A. niger xj.

FIGURE 5

The EC50 value of compound 4 against RS-2.

FIGURE 6

SEM of RS-2 (A) untreated cells and (B) after treatment with compound 4 at EC50 value for 6 h.

FIGURE 7

Effect of compound 4 on cell membrane permeability of RS-2.

FIGURE 8

Effect of compound 4 on protein synthesis in RS-2 cells. (A) SDS-PAGE of RS-2 cells treated with compound 4. M: Maker; 1, 4, 7, and 10 were the control treatments of 0, 4, 8, and 12 h, respectively. 2, 5, 8, and 11 were treated with compound 4 at 0, 4, 8, and 12 h, respectively. 3, 6, 9, and 12 were treated with chloramphenicol for 0, 4, 8, and 12 h, respectively. (B) The grayscale value map of the effect of compound 4 on protein synthesis of RS-2. Different lowercase letters indicate significant differences among treatments at the same exposure time (p < 0.05).