Production of extracellular amylase contributes to the colonization of Bacillus cereus 0–9 in wheat roots View Full Text


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Article Info

DATE

2022-08-22

AUTHORS

Qiubin Huang, Huiping Liu, Juanmei Zhang, Shaowei Wang, Fengying Liu, Chengdie Li, Gang Wang

ABSTRACT

BackgroundBacteria usually secrete a variety of extracellular enzymes to degrade extracellular macromolecules to meet their nutritional needs and enhance their environmental adaptability. Bacillus cereus 0–9, a biocontrol bacterial strain isolated from wheat roots, has three genes annotated as encoding amylases in the genome, but their functions are unknown, and whether they are involved in the colonization process of the bacterium remains to be further studied.MethodsMutant gene strains and fluorescently tagged strains were constructed by homologous recombination, and amylase protein was expressed in the prokaryotic Escherichia coli BL21(DE3) expression system. The iodine staining method was used to measure the activity of amylase proteins. We further observed the colonization abilities of the test strains in wheat roots through frozen section technology.ResultsThe results showed that there were three amylase-encoding genes, amyC, amyP and amyS, in the B. cereus 0–9 genome. Among the three amylase encoding genes, only amyS produced extracellular amylase whose secretion was related to signal peptide at position 1–27. The AmyS protein encoded by the amyS gene is an α-amylase. The growth of Rhizoctonia cerealis was inhibited 84.7% by B. cereus 0–9, but the biocontrol ability of the ΔamyS strain decreased to 43.8% and that of ΔamyS/amyS was restored when the amyS gene was complemented. Furthermore, the biocontrol ability of the ΔamySec strain was decreased to 46.8%, almost the same as that of the ΔamyS mutant. Due to the deletion of the amyS gene, the colonization capacities of ΔamyS (RFP) and ΔamySec (RFP) in wheat roots decreased, while that of ΔamyS/amyS (RFP) was restored after the amyS gene was complemented, indicating that the amyS gene influences the colonization of B. cereus 0–9 in wheat roots. In addition, the colonization and biocontrol abilities of the mutant were restored after the addition of sugars, such as glucose and maltose.ConclusionsB. cereus 0–9 encodes three genes annotated as amylases, amyC, amyP and amyS. Only the deletion of the amyS gene with a signal peptide did not produce extracellular amylase. The AmyS protein encoded by the amyS gene is an α-amylase. Our results indicated that the amyS gene is closely related to the colonization abilities of B. cereus 0–9 in wheat roots and the biocontrol abilities of B. cereus 0–9 to fight against R. cerealis. The extracellular amylase produced by B. cereus 0–9 can hydrolyze starch and use glucose, maltose and other nutrients to meet the needs of bacterial growth. Therefore, it is very possible that the secretion and hydrolytic activities of extracellular amylase can promote the colonization of B. cereus 0–9 in wheat roots and play important roles in the prevention and control of plant diseases. Our results contribute to exploring the mechanisms of microbial colonization in plant roots. More... »

PAGES

205

References to SciGraph publications

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        "description": "BackgroundBacteria usually secrete a variety of extracellular enzymes to degrade extracellular macromolecules to meet their nutritional needs and enhance their environmental adaptability. Bacillus cereus 0\u20139, a biocontrol bacterial strain isolated from wheat roots, has three genes annotated as encoding amylases in the genome, but their functions are unknown, and whether they are involved in the colonization process of the bacterium remains to be further studied.MethodsMutant gene strains and fluorescently tagged strains were constructed by homologous recombination, and amylase protein was expressed in the prokaryotic Escherichia coli BL21(DE3) expression system. The iodine staining method was used to measure the activity of amylase proteins. We further observed the colonization abilities of the test strains in wheat roots through frozen section technology.ResultsThe results showed that there were three amylase-encoding genes, amyC, amyP and amyS, in the B. cereus 0\u20139 genome. Among the three amylase encoding genes, only amyS produced extracellular amylase whose secretion was related to signal peptide at position\u00a01\u201327. The AmyS protein encoded by the amyS gene is an \u03b1-amylase. The growth of Rhizoctonia cerealis was inhibited 84.7% by B. cereus 0\u20139, but the biocontrol ability of the \u0394amyS strain decreased to 43.8% and that of \u0394amyS/amyS was restored when the amyS gene was complemented. Furthermore, the biocontrol ability of the \u0394amySec strain was decreased to 46.8%, almost the same as that of the \u0394amyS mutant. Due to the deletion of the amyS gene, the colonization capacities of \u0394amyS (RFP) and \u0394amySec (RFP) in wheat roots decreased, while that of \u0394amyS/amyS (RFP) was restored after the amyS gene was complemented, indicating that the amyS gene influences the colonization of B. cereus 0\u20139 in wheat roots. In addition, the colonization and biocontrol abilities of the mutant were restored after the addition of sugars, such as glucose and maltose.ConclusionsB. cereus 0\u20139 encodes three genes annotated as amylases, amyC, amyP and amyS. Only the deletion of the amyS gene with a signal peptide did not produce extracellular amylase. The AmyS protein encoded by the amyS gene is an \u03b1-amylase. Our results indicated that the amyS gene is closely related to the colonization abilities of B. cereus 0\u20139 in wheat roots and the biocontrol abilities of B. cereus 0\u20139 to fight against R. cerealis. The extracellular amylase produced by B. cereus 0\u20139 can hydrolyze starch and use glucose, maltose and other nutrients to meet the needs of bacterial growth. Therefore, it is very possible that the secretion and hydrolytic activities of extracellular amylase can promote the colonization of B. cereus 0\u20139 in wheat roots and play important roles in the prevention and control of plant diseases. Our results contribute to exploring the mechanisms of microbial colonization in plant roots.", 
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    29 schema:description BackgroundBacteria usually secrete a variety of extracellular enzymes to degrade extracellular macromolecules to meet their nutritional needs and enhance their environmental adaptability. Bacillus cereus 0–9, a biocontrol bacterial strain isolated from wheat roots, has three genes annotated as encoding amylases in the genome, but their functions are unknown, and whether they are involved in the colonization process of the bacterium remains to be further studied.MethodsMutant gene strains and fluorescently tagged strains were constructed by homologous recombination, and amylase protein was expressed in the prokaryotic Escherichia coli BL21(DE3) expression system. The iodine staining method was used to measure the activity of amylase proteins. We further observed the colonization abilities of the test strains in wheat roots through frozen section technology.ResultsThe results showed that there were three amylase-encoding genes, amyC, amyP and amyS, in the B. cereus 0–9 genome. Among the three amylase encoding genes, only amyS produced extracellular amylase whose secretion was related to signal peptide at position 1–27. The AmyS protein encoded by the amyS gene is an α-amylase. The growth of Rhizoctonia cerealis was inhibited 84.7% by B. cereus 0–9, but the biocontrol ability of the ΔamyS strain decreased to 43.8% and that of ΔamyS/amyS was restored when the amyS gene was complemented. Furthermore, the biocontrol ability of the ΔamySec strain was decreased to 46.8%, almost the same as that of the ΔamyS mutant. Due to the deletion of the amyS gene, the colonization capacities of ΔamyS (RFP) and ΔamySec (RFP) in wheat roots decreased, while that of ΔamyS/amyS (RFP) was restored after the amyS gene was complemented, indicating that the amyS gene influences the colonization of B. cereus 0–9 in wheat roots. In addition, the colonization and biocontrol abilities of the mutant were restored after the addition of sugars, such as glucose and maltose.ConclusionsB. cereus 0–9 encodes three genes annotated as amylases, amyC, amyP and amyS. Only the deletion of the amyS gene with a signal peptide did not produce extracellular amylase. The AmyS protein encoded by the amyS gene is an α-amylase. Our results indicated that the amyS gene is closely related to the colonization abilities of B. cereus 0–9 in wheat roots and the biocontrol abilities of B. cereus 0–9 to fight against R. cerealis. The extracellular amylase produced by B. cereus 0–9 can hydrolyze starch and use glucose, maltose and other nutrients to meet the needs of bacterial growth. Therefore, it is very possible that the secretion and hydrolytic activities of extracellular amylase can promote the colonization of B. cereus 0–9 in wheat roots and play important roles in the prevention and control of plant diseases. Our results contribute to exploring the mechanisms of microbial colonization in plant roots.
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    38 Escherichia coli
    39 R. cerealis
    40 ResultsThe results
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    43 activity
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    45 addition
    46 addition of sugar
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    50 bacterial growth
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    79 iodine staining method
    80 macromolecules
    81 maltose
    82 mechanism
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    84 microbial colonization
    85 mutants
    86 need
    87 nutrients
    88 nutritional needs
    89 peptides
    90 plant diseases
    91 plant roots
    92 position
    93 prevention
    94 process
    95 production
    96 protein
    97 recombination
    98 results
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    100 roots
    101 secretion
    102 signal peptide
    103 staining method
    104 starch
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