耐盐深绿木霉TW320和拟康宁木霉TW1876生长和产孢条件优化

doi:10.3976/j.issn.1002-4026.2022.06.009

摘要/Abstract

摘要：

为优化两株耐盐木霉-深绿木霉(Trichoderma atroviride)TW320和拟康宁木霉(Trichoderma koningiopsis)TW1876的生长和产孢条件,研究了不同pH、不同培养基,不同碳源、氮源和不同碳氮比对两株木霉菌的菌落生长和孢子产量的影响。结果显示:耐盐深绿木霉(T. atroviride)TW320在 pH =4~11 的马铃薯葡萄糖琼脂 (potato dextrose agar, PDA)培养基上均可生长和产孢,且菌丝生长和产孢的最佳pH为5,在大豆玉米粉培养基上生长速度较快,产孢量最多,该菌株菌丝生长和产孢的最佳碳源为木糖,最佳氮源是酵母膏,当碳氮比为12:1时,最有利于TW320产孢;拟康宁木霉(T. koningiopsis)TW1876菌丝生长和产孢的最佳pH为11,最优培养基为PDA培养基,以乳糖作为最佳碳源,甘氨酸作为最佳氮源,且碳氮比为9:1时,TW1876产孢量最多。

关键词: 耐盐木霉, 菌落直径, 产孢量, 碳氮比

Abstract:

The effects of different pH, medium, carbon source, nitrogen source, and C/N ratios on the colony growth and spore yield of two salt-tolerant Trichoderma strains, Trichoderma atroviride TW320 and Trichoderma. koningiopsis TW1876, were studied in order to optimize the growth and spore production conditions of the two strains. The results showed that the salt-tolerant T. atroviride TW320 could grow and produce spores on potato dextrose agar (PDA) medium with a pH of 4 to 11, and the optimum pH for mycelial growth and spore production was 5. On soybean corn flour medium, TW320 grew the fastest and produced the most spores. The best carbon source for TW320 mycelial growth and sporulation was xyaccharide, respectively, and the best nitrogen source for TW320 was yeast extract. When the C/N ratio is 12:1, TW320 produces the most spores. The optimum pH for mycelial growth and sporulation of T. koningiopsis TW1876 was 11, and the optimal medium was PDA. TW1876 has the highest sporulation capacity, with lactose being the best carbon source, glycine being the best nitrogen source, and the C/N ratio being 9:1.

Key words: salt-tolerant Trichoderma, colony diameter, Spore production, C/N ratio

中图分类号:

Q939.9

胡广艳, 赵忠娟, 杨合同. 耐盐深绿木霉TW320和拟康宁木霉TW1876生长和产孢条件优化[J]. 山东科学, 2022, 35(6): 65-73.

HU Guang-yan, ZHAO Zhong-juan, YANG He-tong. Optimization of growth and spore-producing conditions of salt-tolerant Trichoderma atroviride TW320 and Trichoderma koningiopsis TW1876[J]. Shandong Science, 2022, 35(6): 65-73.

图/表 11

表1

各培养基成分"

培养基	培养基配方	灭菌方式
PDA	土豆200 g/L,葡萄糖20 g/L,琼脂15 g/L	115 ℃高压,30 min
Martin	葡萄糖10 g/L,蛋白胨5 g/L,KH₂P $O 4$ 1.0 g/L,MgSO₄·7H₂O 0.5 g/L,琼脂15 g/L	115 ℃高压,30 min
CDA	NaNO₄ 3.0 g/L,K₂HP $O 4$ 1.0 g/L,MgSO₄·7H₂O 0.5 g/L, KCl 0.5 g/L,FeS $O 4$ 0.01 g/L,蔗糖30 g/L,琼脂20 g/L	115 ℃高压,30 min
SNA	KH₂PO₄ 1.0 g/L,KNO₃1.0 g/L, MgSO₄·7H₂O 0.5 g/L, KCl 0.5 g/L,葡萄糖0.2 g/L,蔗糖0.2 g/L,琼脂20 g/L	115 ℃高压,30 min
CMD	玉米粉30 g/L,葡萄糖20 g/L,琼脂20 g/L	115 ℃高压,30 min
大豆玉米粉	大豆粉4.65 g/L,玉米粉12.75 g/L,蔗糖10 g/L,NH₄NO₃ 2.0 g/L, MgSO₄·7H₂O 1.0 g/L, KH₂PO₄ 3.32 g/L,琼脂20 g/L	115 ℃高压,30 min
MPA	Ca₃(PO₄)₂ 2.5 g/L,葡萄糖13 g/L, (NH₄)₂SO₄ 0.5 g/L, NaCl 0.2 g/L, MgSO₄·7H₂O 0.1 g/L, KCl 0.2 g/L,酵母粉0.5 g/L, MnSO₄ 0.000 2 g/L, FeSO₄·7H₂O 0.000 2 g/L,琼脂15 g/L	115 ℃高压,30 min
OA	燕麦片20 g/L,琼脂12.5 g/L	121 ℃高压,90 min
MEA	麦芽膏粉130 g/L,琼脂15 g/L	115 ℃高压,30 min

表1

图1

表2

图2

表3

图3

表4

图4

表5

图5

表6

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pH	TW320		TW1876
pH	菌落直径/cm	产孢量/(cfu·mL^-1)	菌落直径/cm	产孢量/(cfu·mL^-1)
4	8±0.07a	(3.25±1.69×10⁷bc	7.02±0.11a	(0.64±0.26)×10⁸cd
5	8±0.14a	(4.94±3.42)×10⁷ab	7.04±0.11a	(0.49±0.26)×10⁸d
6	7.95±0.06a	(4.12±1.6)×10⁷bc	7.12±0.26a	(0.47±0.29)×10⁸d
7	7.66±0.11b	(3.41±1.93)×10⁷bc	6.76±0.21b	(0.75±0.35)×10⁸cd
8	7.38±0.13c	(3.95±2.51)×10⁷bc	6.66±0.11b	(1.41±0.19)×10⁸ab
9	6.82±0.2e	(2.07±0.75)×10⁷c	6.6±0.07b	(1.03±0.28)×10⁸bc
10	7.06±0.05d	(2.39±2.35)×10⁷c	6.66±0.05b	(0.77±0.47)×10⁸cd
11	7.02±0.16d	(3.06±1.67)×10⁷bc	6.62±0.11b	(1.76±0.88)×10⁸a

培养基	TW320		TW1876
培养基	菌落直径/cm	产孢量/(cfu·mL^-1)	菌落直径/cm	产孢量/(cfu·mL^-1)
SNA	2.74±0.11g	(0.05±0.01)×10⁷b	4.22±0.13d	(0.5±0.26)×10⁷b
PDA	7.52±0.28a	(0.57±0.5)×10⁷b	6.74±0.24a	(9.68±3.64)×10⁷a
Martin	5.8±0.12d	(0.32±0.2)×10⁷b	5.52±0.13c	(0.13±0.06)×10⁷b
CMD	6.38±0.2c	(0.13±0.08)×10⁷b	6.26±0.11b	(7.97±5.24)×10⁷a
大豆玉米粉	6.92±0.11b	(3.13±2.71)×10⁷a	6.38±0.08b	(0.07±0.01)×10⁷b
MPA	4.68±0.19e	(0.42±0.24)×10⁷b	5.32±0.19c	(0.63±0.42)×10⁷b
CDA	1.68±0.3h	(0.07±0.04)×10⁷b	3±0.16f	(0.25±0.16)×10⁷b
OA	4.78±0.19e	(0.67±0.64)×10⁷b	5.34±0.09c	(7.02±2.49)×10⁷a
MEA	3.26±0.17f	(0.2±0.11b)×10⁷	3.44±0.27e	(0.09±0.07)×10⁷b

碳源	TW320		TW1876
碳源	菌落直径/cm	产孢量/(cfu·mL^-1)	菌落直径/cm	产孢量/(cfu·mL^-1)
甘露醇	2.02±0.68c	(0.65±0.3)×10⁶b	2.46±0.4cd	(0.98±0.16)×10⁶c
木糖	2.76±0.19b	(2.93±1.59)×10⁶a	3.12±0.13b	(2.95±0.93)×10⁶bc
蔗糖	2.56±0.13b	(0.58±0.2)×10⁶b	2.68±0.15c	(2.6±1.01)×10⁶bc
乳糖	2.74±0.27b	(1.02±0.94)×10⁶b	3.12±0.19b	(6.75±2.21)×10⁶a
葡萄糖	2.7±0.44b	(1.02±0.49)×10⁶b	3.18±0.22b	(2.47±0.6)×10⁶bc
淀粉	2.94±0.26b	(0.22±0.18)×10⁶b	3.68±0.27a	(4.05±1.26)×10⁶abc
麦芽糖	3.5±0.65a	(1.22±1.29)×10⁶b	3.42±0.15ab	(5.45±4.46)×10⁶ab
无碳源	1.62±0.08c	(0.52±0.45)×10⁶b	2.18±0.13d	(0.47±0.25)×10⁶c

氮源	TW320		TW1876
氮源	菌落直径/cm	产孢量/(CFU·mL^-1)	菌落直径/cm	产孢量/(CFU·mL^-1)
磷酸氢二铵	2.06±0.18de	(0.93±0.1)×10⁶bc	1.8±0.12d	(0.58±0.29)×10⁶cd
蛋白胨	4.54±0.19b	(1.48±0.15)×10⁶bc	3.74±0.21b	(1.8±1.24)×10⁶bcd
酵母膏	5.54±0.19a	(5.85±2.98)×10⁶a	4.86±0.21a	(1.03±0.28)×10⁶cd
尿素	1.32±0.16f	(0.28±0.1)×10⁶c	1.56±0.25d	(0.83±0.93)×10⁶cd
硫酸铵	3.5±0.63c	(3.12±2.59)×10⁶b	3.78±0.22b	(2.52±0.94)×10⁶abc
硝酸铵	3.14±0.26c	(1.37±0.55)×10⁶bc	3.62±0.19b	(3.17±1.65)×10⁶ab
甘氨酸	3.42±0.44c	(1.08±0.47)×10⁶bc	3.66±0.21b	(3.83±2.06)×10⁶a
硝酸钠	1.74±0.25ef	(0.5±0.1)×10⁶c	2.52±0.13c	(0.42±0.3)×10⁶d
牛肉膏	4.78±0.26b	(2.33±0.93)×10⁶bc	3.53±0.22b	(1.08±0.4)×10⁶cd
无氮源	2.34±0.72d	(0.08±0.03)×10⁶c	2.7±0.14c	(0.3±0.13)×10⁶d

碳氮比	TW320		TW1876
碳氮比	菌落直径/cm	产孢量/(cfu·mL^-1)	菌落直径/cm	产孢量/(cfu·mL^-1)
3:1	4.5±0.14c	(0.66±0.31)×10⁷b	3.12±0.04a	(6.88±3.69)×10⁶ab
6:1	5.48±0.13b	(0.98±0.52)×10⁷b	2.94±0.05b	(3.07±1.24)×10⁶c
9:1	6.28±0.18a	(0.49±0.08)×10⁷b	3.14±0.11a	(7.41±2.72)×10⁶a
12:1	6.42±0.13a	(2.18±2.53)×10⁷a	3.02±0.04b	(4.67±2.49)×10⁶bc