Effect on Spore Producing Ability of Biocontrol Bacterium Bacillus subtilis CS16: Calcium Ions and Manganese Ions in Liquid Fermentation Process

doi:10.11924/j.issn.1000-6850.casb2023-0072

Abstract

Abstract:

This study was conducted to optimize the fermentation conditions for spore production of strain Bacillus subtilis CS16 using the methods of single factor and orthogonal experiment. The effects of type and concentration of mineral salts on the spore count and sporulation rate of Bacillus subtilis CS16 were studied, and then fermentation technology was further optimized by orthogonal experiment on the basis of single-factor experiments. The results showed that the addition of CaCl₂ or MnSO₄·H₂O in the medium significantly increased the spore count (P<0.05). Orthogonal experiment results indicated that the highest spore count reached 2.37×10⁹cfu/mL after 48 h of fermentation when 0.8% of CaCl₂ and 0.04% of MnSO₄·H₂O were simultaneously added, which was 8.4 times and 6.5 times of the highest spore count compared with CaCl₂ and MnSO₄·H₂O were added alone, respectively. The spore count increased almost 20 times compared with the control group (CK), and the sporulation rate reached to 65.02%, which increased nearly by 50% compared with the CK group. Simultaneous addition of Ca²⁺ and Mn²⁺ had a synergistic effect on the spore formation of CS16, which could effectively increase the spore count and sporulation rate in CS16 liquid fermentation process.

Key words: Bacillus subtilis CS16, spore count, sporulation rate, mineral salts

LIN Chenqiang, CHEN Longjun, SU Guangqiu, JIA Xianbo, CHEN Jichen. Effect on Spore Producing Ability of Biocontrol Bacterium Bacillus subtilis CS16: Calcium Ions and Manganese Ions in Liquid Fermentation Process[J]. Chinese Agricultural Science Bulletin, 2023, 39(36): 125-131.

Figures/Tables 8

References 26

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水平	因素
水平	A.CaCl₂浓度/%	B.MnSO₄·H₂O浓度/%	C.培养时间/h
1	0.4	0.04	40
2	0.6	0.06	48
3	0.8	0.08	56

水平	因素
水平	A.CaCl₂浓度/%	B.MnSO₄·H₂O浓度/%	C.培养时间/h
1	0.4	0.04	40
2	0.6	0.06	48
3	0.8	0.08	56

处理	培养时间
处理	48 h	52 h	56 h	60 h	64 h	68 h	72 h	76 h	80 h
CK	5.4%	5.3%	5.9%	9.5%	10.8%	11.4%	14.4%	17.0%	18.4%
0.6%CaCl₂	8.3%	13.6%	12.6%	11.9%	16.0%	22.7%	13.9%	20.0%	17.0%

处理	培养时间
处理	48 h	52 h	56 h	60 h	64 h	68 h	72 h	76 h	80 h
CK	5.4%	5.3%	5.9%	9.5%	10.8%	11.4%	14.4%	17.0%	18.4%
0.6%CaCl₂	8.3%	13.6%	12.6%	11.9%	16.0%	22.7%	13.9%	20.0%	17.0%

试验号	A CaCl₂浓度	B MnSO₄·H₂O浓度	C 培养时间	活菌总数/ (cfu/mL)	芽孢数/(cfu/mL)	芽孢率/%
1	1	1	1	4.65×10⁹	9.80×10⁸	21.08
2	2	2	1	6.95×10⁹	1.23×10⁹	17.75
3	3	3	1	3.55×10⁹	8.83×10⁸	24.69
4	3	2	2	3.65×10⁹	2.37×10⁹	65.02
5	2	1	2	4.95×10⁹	1.99×10⁹	40.20
6	1	3	2	2.51×10⁹	9.03×10⁸	36.04
7	1	2	3	3.30×10⁹	2.35×10⁹	71.31
8	2	3	3	2.77×10⁹	1.22×10⁹	43.98
9	3	1	3	3.40×10⁹	1.68×10⁹	49.51
活菌总数	k₁	3.49×10⁹	4.33×10⁹	5.05×10⁹
	k₂	4.89×10⁹	4.63×10⁹	3.70×10⁹
	k₃	3.53×10⁹	2.94×10⁹	3.16×10⁹
	R	1.36×10⁹	1.69×10⁹	1.89×10⁹
	主次性	C>B>A
	最佳组合	A₂B₂C₁
芽孢数	k'₁	1.41×10⁹	1.55×10⁹	1.03×10⁹
	k'₂	1.48×10⁹	1.98×10⁹	1.75×10⁹
	k'₃	1.64×10⁹	1.00×10⁹	1.75×10⁹
	R'	0.23×10⁹	0.98×10⁹	0.72×10⁹
	主次性	B>C>A
	最佳组合	A₃B₂C₂
芽孢率	k"₁	42.81	36.93	21.17
	k"₂	33.98	51.36	47.09
	k"₃	46.41	34.90	54.93
	R"	12.43	16.46	33.76
	主次性	C>B>A
	最佳组合	A₃B₂C₃