Scientia Agricultura Sinica ›› 2023, Vol. 56 ›› Issue (24): 4854-4865.doi: 10.3864/j.issn.0578-1752.2023.24.005

• PLANT PROTECTION • Previous Articles     Next Articles

The Effect of 3-Oxyacyl ACP Reductase Gene FgOAR1 on the Growth, Development and Pathogenicity of Fusarium graminearum

GONG AnDong(), LEI YinYu, WU NanNan, LIU JingRong, SONG MengGe, ZHANG YiMei(), YANG Guang, YANG Peng   

  1. College of Life and Science, Xinyang Normal University, Xinyang 464000, Henan
  • Received:2023-07-28 Accepted:2023-09-29 Online:2023-12-21 Published:2023-12-21
  • Contact: ZHANG YiMei

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Abstract:

【Background】Fusarium graminearum is the main pathogenic fungus which can infect wheat and result in Fusarium head blight. The infection of F. graminearum leads to huge reduction in crop quality and yield, and produces different types of mycotoxins that endanger grain production and human health. 3-oxoacyl ACP reductase (OAR1), catalyzing carboxyacyl carrier protein to oxyacyl carrier protein, which plays an important role in the fatty acid biosynthesis of F. graminearum.【Objective】To reveal the biological function of FgOAR1 in F. graminearum, the FgOAR1 deletion mutants were constructed. The phenotype, cell structure, conidia concentration, sexual reproduction and pathogenicity of mutant were analyzed and compared with wild-type strain PH-1. The results will uncover the function of FgOAR1 in the growth and pathogenic process of F. graminearum, and provide scientific evidences for the identification of novel antifungal target and control of Fusarium head blight.【Method】Wild-type strain F. graminearum PH-1 was used as material in this study. Split-Marker gene knockout technology was conducted to construct the ΔFgOAR1 mutants. The ΔFgOAR1-C reverting strain was obtained by the Gap-repair method. All strains were individually inoculated in common media (PDA, CM, YPG), and the stress selection medium containing Congo-Red, SDS, NaCl and H2O2, respectively. The mycelium phenotypes and fatty acid content were recorded. The strains were inoculated in CMC medium to analyze the conidia concentration. The carrot culture-medium was used to analyze the sexual reproduction of each strain. All strains were individually inoculated to the wheat coleoptile and spikelet to analyze the pathogenicity and mycotoxin content, respectively. The disease incidence of mutant strains was calculated and compared with PH-1 strain.【Result】Compared with the wild-type strain PH-1, the growth of ΔFgOAR1 mutant showed no difference in PDA, YEG and CM media. Whereas, under the pressure of 0.7 mol·L-1 NaCl, 0.03% H2O2, 0.01% SDS and 300 μg·mL-1 Congo-Red, the growth of ΔFgOAR1 mutant was significantly reduced compared with PH-1. The evidence demonstrated that FgOAR1 is related to the cell membrane and cell wall formation in F. graminearum. In CMC medium, the conidia concentration of ΔFgOAR1 mutant (8.1×105 conidia/mL) was less than PH-1 (1.26×106 conidia/mL) with significant difference; The pathogenicity analysis further demonstrated that the disease incidence of ΔFgOAR1 mutant was significantly lower than PH-1 in wheat coleoptile and spikelet inoculation tests.【Conclusion】FgOAR1 participates in the fatty acid biosynthesis, and plays an important role in cell membrane formation of F. graminearum. The deletion of FgOAR1 resulted in the reduced resistance to pressures, decreased conidia production and pathogenicity. Therefore, FgOAR1 is important for the growth, development and pathogenicity process of F. graminearum.

Key words: Fusarium graminearum, Fusarium head blight, gene knockout, FgOAR1, pathogenicity

Table 1

The primer sequences used in this study"

引物名称<BOLD>P</BOLD>rimer name 引物序列<BOLD>P</BOLD>rimer sequence (5′→3′)
FgOAR1_1F TCTCGATACCTCCTACCTACCT
FgOAR1_2R TTGACCTCCACTAGCTCCAGCCAAGCCTCAACACATGCTCTACACCCT
FgOAR1_3F GAATAGAGTAGATGCCGACCGCGGGTTCCAGGGTCTCAAAGCAAC
FgOAR1_4R ATCCCAGAAAATCCGCAA
HYG/F GGCTTGGCTGGAGCTAGTGGAGGTCAA
HY/R GTATTGACCGATTCCTTGCGGTCCGAA
YG/F GATGTAGGAGGGCGTGGATATGTCCT
HYG/R AACCCGCGGTCGGCATCTACTCTATTC
FgOAR1_5F ATCGGAGTTGAGGTGACAG
FgOAR1_6R GAAGAGGGGAGGAAAGGTT
FgOAR1_7F GGTCAATAGAGCAGGTCGC
FgOAR1_8R CAGGCGGTGGTAACTTCA
h850 TTCCTCCCTTTATTTCAGATTCAA
h852 ATGTTGGCGACCTCGTATTGG
h855R GTCGATGCGACGCAATCGT
h856F GCTGATCTGACCAGTTGC
FgOAR1_hbF CGACTCACTATAGGGCGAATTGGGTACTCAAATTGGTGACAGCGGCGTTAGGACA
FgOAR1_hbR CACCACCCCGGTGAACAGCTCCTCGCCCTTGCTCACGTACATCTCCAGTCCACCACT
insert_F TAACGCCAGGGTTTTCCCAGTCA
insert_R CGTGCTGCTTCATGTGGTCGG

Fig. 1

Multiple alignment of amino acid sequence of OAR1 in different fungi"

Fig. 2

PCR detection of FgOAR1 deletion mutant and complementary mutant"

Fig. 3

The colony morphology of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Table 2

The growth rate determination of three strains"

菌株
Strain		 生长速率Growth rate (cm·d-1)
PDA 5×YEG CM
PH-1 1.20±0.05 0.83±0.10 0.85±0.10
ΔFgOAR1 1.19±0.01 0.87±0.02 0.84±0.11
ΔFgOAR1-C 1.23±0.09 0.85±0.12 0.85±0.10

Table 3

The growth rate of PH-1、ΔFgOAR1 and ΔFgOAR1-C strains under different stress cultural conditions"

菌株
Strain		 生长速度Growth rate (cm·d-1)
CM Congo-Red NaCl SDS H2O2
PH-1 1.20±0.05 1.15±0.10 0.98±0.11 0.54±0.15 0.75±0.11
ΔFgOAR1 1.10±0.01 0.81±0.02* 0.62±0.17* 0.37±0.09* 0.60±0.09*
ΔFgOAR1-C 1.23±0.09 1.16±0.12 0.88±0.11 0.57±0.19 0.73±0.10

Fig. 4

The colony morphology of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains under different stress cultural conditions"

Fig. 5

Fatty acid content determination of PH-1 and ΔFgOAR1 strains"

Fig. 6

Conidia concentration determination and germination observation of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Fig. 7

Sexual reproduction of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Fig. 8

Pathogenicity of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Fig. 9

The expression of FgOAR1 under different growth conditions"

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