3株酵母菌的分离鉴定及发酵液除草潜力的研究

施生姣; 宋维敏; 程亮; 魏有海

doi:10.19303/j.issn.1008-0384.2022.008.013

3株酵母菌的分离鉴定及发酵液除草潜力的研究

doi: 10.19303/j.issn.1008-0384.2022.008.013

施生姣^{1, 2, 3, 4, 5,},
宋维敏^{1, 2, 3, 4, 5},
程亮^{2, 3, 4, 5},
魏有海^{2, 3, 4, 5, ,}

1.
青海大学, 青海　西宁　810016
2.
青海省农林科学院, 青海　西宁　810016
3.
青海省农业有害生物综合治理重点实验室, 青海　西宁　810016
4.
农业农村部西宁作物有害生物科学观测实验站, 青海　西宁　810016
5.
青藏高原生物技术教育部重点实验室, 青海　西宁　810016

基金项目: 现代农业技术体系建设专项（CARS-07-C-3、CARS-08-C-3）；科技部第二次青藏高原综合科学考察研究项目(2019QZKK0303)；青海省农业有害生物综合治理重点实验室（2022-ZJ-Y10）

详细信息

作者简介:
施生姣（1997−），女，硕士研究生，研究方向：农田杂草综合防治（E-mail：2964307528@qq.com）

通讯作者:
魏有海（1972−），男，硕士，研究员，研究方向：农田杂草综合治理（E-mail：youhaiweiqh@163.com）

中图分类号: S 476.1
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出版历程
- 收稿日期: 2022-03-20
- 修回日期: 2022-06-20
- 网络出版日期: 2022-10-05
- 刊出日期: 2022-08-28

Identification and Herbicidal Efficacies of Three Yeast Strains

SHI Shengjiao^{1, 2, 3, 4, 5
,},
SONG Weimin^{1, 2, 3, 4, 5},
CHENG Liang^{2, 3, 4, 5},
WEI Youhai^{2, 3, 4, 5
, ,}

1.
Qinghai University, Xining, Qinghai　810016, China
2.
Qinghai Academy of Agriculture and Forestry Sciences, Xining, Qinghai　810016, China
3.
Key Laboratory of Agricultural Integrated Pest Management of Qinghai Province, Xining, Qinghai　810016, China
4.
Scientific Observing and Experimental Station of Crop Pest in Xining, Ministry of Agriculture and Rural Affairs, Xining, Qinghai　810016, China
5.
Key Laboratory of Qinghai-Tibetan Plateau Biotechnology, Ministry of Education, Xining, Qinghai　810016, China

摘要

摘要: 目的探究分离菌株6-3、12-6和16-8对青海农田优势杂草猪殃殃、密花香薷、藜的除草活性以及对作物的安全性，为微生物除草剂的开发提供理论依据。方法采用温室盆栽法进行菌株的除草活性及作物的安全性测定；通过形态学观察结合ITS及26S rDNA基因序列对菌株进行分子生物学鉴定。结果接种菌株16-8、12-6和6-3发酵液7 d后对猪殃殃的伤害率为90.17%、87.50%和0%，鲜重抑制率为88.63%、75.80%和9.41%；对密花香薷的伤害率为66.67%、75.00%和36.67%，鲜重抑制率为56.90%、62.72%和46.29%；对藜的伤害率为95.00%、37.50%和25.00%，鲜重抑制率为78.77%、43.25%和43.46%。作物安全性试验结果表明，3株生防菌发酵液对供试的燕麦、蚕豆、青稞、小麦、油菜、豌豆和马铃薯等7种作物表现安全。经鉴定16-8菌株为库德里阿兹威毕赤酵母菌，12-6为马克斯克鲁维酵母菌、6-3为季也蒙迈耶氏酵母菌。结论 16-8菌株和12-6菌株对不同优势杂草表现出较好的抑制效果，有开发为微生物除草剂的潜力，可开展杀草谱评价和微生物除草剂研发相关研究。
- 杂草 /
- 生防菌株 /
- 菌种鉴定 /
- 除草活性 /
- 作物安全性
Abstract: Objective Three yeast strains with herbicidal activity were isolated for the determinations of their efficacy on weeds and application safety on crops. Method In pot culture, the herbicidal efficacies of Strains 6-3, 12-6, and 16-8 on Galium spurium, Elsholtzia densa, and Chenopodium album, the weeds of major concern in Qinghai Province, were examined. A test on the application safety on crops was conducted in a greenhouse. In addition to morphological characteristics, ITS and 26S rDNA gene sequencing were performed to biologically identify the 3 strains. Results Seven days after treatment with the yeast fermentation broths of 16-8, 12-6, and 6-3, the G. spurium plants sustained the injury rates of 90.17%, 87.50%, and 0%, respectively, with the fresh weight losses of 88.63%, 75.80%, and 9.41%, respectively; E. densa showed the rates of 66.67%, 75.00%, and 36.67%, respectively, with the weight losses of 56.90%, 62.72%, and 46.29%, respectively; and C. album displayed the injury rates of 95.00%, 37.50%, and 25.00%, respectively, with the weight losses of 78.77%, 43.25%, and 43.46%, respectively. The treatments on oat, broad beans, barley, wheat, rapeseeds, peas, and potatoes were shown to be safe for food consumption and crop cultivation. Strain 16-8 was confirmed to be Pichia kudriavzevii, 12-6 to be Kluyveromyces marxianus, and 6-3 to be Meyerozyma guilliermondii. Conclusion Of the 3 yeast strains, 16-8 and 12-6 were strong in inhibiting the growth of the weeds of major concern in the province and could be developed as natural herbicides. The results might also lead to a broadened study on microbe-origin herbicides and related research.
- Weeds /
- biocontrol strains /
- strains identification /
- herbicidal activity /
- crop safety

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图 1 3株酵母菌发酵液对多种杂草幼苗生长的抑制作用

A：猪殃殃；B：密花香薷；C：藜

Figure 1. Inhibition effect of yeast fermentation broths on growth of weed seedlings

A：G. aparine；B：E. densa；C：C. album.

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图 2 除草剂和16-8菌株混用后7 d的除草效果

D：氯氟吡氧乙酸推荐剂量；E：氯氟吡氧乙酸50%推荐剂量；F：氯氟吡氧乙酸25%推荐剂量；G：氯氟吡氧乙酸推荐剂量+16-8菌株发酵液；H：氯氟吡氧乙酸50%推荐剂量+16-8菌株发酵液；I：氯氟吡氧乙酸25%推荐剂量+16-8菌株发酵液J:16-8菌株发酵液；K：清水对照。

Figure 2. Efficacy of herbicide, 16-8 or herbicide+16-8 on weeds 7 d after treatment

D: fluroxypyr at recommended dose; E: fluroxypyr at half recommended dose; F: fluroxypyr at 25% of recommended dose; G: recommended dose+16-8 fermentation broth; H: 50% of recommended fluroxypyr dose+16-8 fermentation broth; I: 25% of recommended fluroxypyr dose+16-8 fermentation broth J: 16-8 fermentation broth alone; K: water control.

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图 3 分离菌株菌落形态及其孢子显微观察

L：16-8菌落形态（左，正面；右，反面）；M：12-6菌落形态（左，正面；右，反面）；N：6-3菌落形态（左，正面；右，反面）；O：16-8孢子形态；P：12-6孢子形态；Q：6-3孢子形态。

Figure 3. Colony morphology and microscopic images of 6-3, 12-6, and 16-8 spores

L：front-view of 16-8 colony (left); back-view of 16-8 colony (right); M：front-view of 12-6 colony (left); back-view of 12-6 colony (right); N: front-view of 6-3 colony (left); back-view of 6-3 colony (right); O: morphology of 16-8 spores; P: morphology of 12-6 spores; Q: morphology of 6-3 spores.

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图 4 3个菌株（16-8、12-6和6-3）基于ITS及26S rDNA基因构建的邻接法系统发育树

Figure 4. Phylogenic NJ trees based on ITS rDNA and 26S rDNA sequences of Strains 16-8, 12-6, and 6-3

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表 1 3株酵母菌菌株发酵液对杂草幼苗生长的抑制作用

Table 1. Inhibition effect of yeast fermentation broths on growth of weed seedlings

菌株 strain	猪殃殃 G. aparine.			密花香薷 E. densa			藜 C. album
菌株 strain	伤害率 Injury rate /%	鲜重 Fresh weight /g	鲜重抑制率 Fresh weight inhibition rate /%	伤害率 Injury rate/%	鲜重 Fresh weight/g	鲜重抑制率 Fresh weight inhibition rate/%	伤害率 Injury rate%	鲜重 Fresh weight/g	鲜重抑制率 Fresh weight inhibition rate/%
16-8	90.17	1.5±0.22	88.63 a	66.67	9.7±0.15	56.90 b	95.00	1.6±0.21	78.77 a
12-6	87.50	3.2±0.12	75.80 b	75.00	8.4±0.24	62.72 a	43.33	4.1±0.11	43.25 b
6-3	0	11.9±0.14	9.41 c	36.67	12.1±0.31	46.29 c	41.67	4.1±0.12	43.46 b
CK	0	13.1±0.23	0 d	0	22.6±0.15	0 d	0	7.3±0.15	0 c
同列数据后不同小写字母表示差异达0.05显著水平（P＜0.05），表3~4同。 Data with different lowercase letters on same column indicate significant different at P＜0.05, Same for table3-4.

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表 2 3株酵母菌株发酵液对不同作物的安全性比较

Table 2. Effect of yeast fermentation broths on crop safety

菌株 strain	燕麦 Avena sativa	蚕豆 Vicia faba.	豌豆 Pisum sativum	小麦 Triticum aestivum	青稞 Hordeum vulgare	油菜 Brassica napus	马铃薯 Solanum tuberosum
16-8	NS	LS	LS	NS	NS	NS	NS
6-3	NS	NS	NS	NS	NS	NS	NS
12-6	NS	NS	NS	NS	NS	NS	NS
CK	NS	NS	NS	NS	NS	NS	NS
NS表示无症状；LS表示有轻微反应；MS表示中等感病；SS表示严重感病。 NS: asymptomatic; LS: mild reaction; MS: moderately susceptible; SS: severely susceptible.

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表 3 3株酵母菌株发酵液对不同作物抑制作用

Table 3. Inhibition effect of yeast fermentation broths on crops （单位：%）

菌株 strain	燕麦 A. sativa		蚕豆 V. faba		豌豆 P. sativum		小麦 T. aestivum
菌株 strain	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate
6-3	0.25±1.35 b	1.08±0.46 b	12.73±0.88a	14.73±2.43 a	10.69±0.13 a	22.61±0.43 a	0.17±0.31 c	3.20±0.17 a
12-6	0.53±1.13 a	3.71±0.21 a	0.03±0.39c	2.34±0.37 c	0.05±0.29 b	0.44±0.40 b	1.26±0.57 a	0.58±0.31c
16-8	0.02±0.55 c	0.04±0.16 c	1.54±0.67b	5.71±1.66 b	0.16±0.67b	9.46±0.71c	1.16±0.17 ab	2.15±0.25 b

菌株 strain	青稞 H. vulgare		油菜 B. napus		马铃薯 S. tuberosum
菌株 strain	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate	株高抑制率 Inhibition rate of plant height	鲜重抑制率 Fresh weight inhibition rate
6-3	0.12±0.52 a	3.51±0.37 a	8.26±0.07 a	0.01±0.01 a	0.20±0.55c	5.66±5.53 a
12-6	0.01±0.17b	1.56±0.35 b	0.13±0.15 c	0.00±0.06 a	0.49±0.14 b	1.58±13.57 c
16-8	0.07±0.41 c	0.21±0.39 c	2.62±0.33 b	0.01±0.04 a	0.68±0.33 a	2.74±6.76 b

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表 4 氯氟吡氧乙酸与16-8菌株发酵液混用对猪殃殃的防效

Table 4. Efficacy of 16-8 fermentation broth per se or in combination with fluroxypyr on controlling G. aparine

处理 Treatment	剂量Dosage/ (mL·hm⁻²+mL·m^-2)	处理后7 d After treatment 7 d		处理后14 d After treatment 14 d
处理 Treatment	剂量Dosage/ (mL·hm⁻²+mL·m^-2)	鲜重 Fresh weight/g	鲜重抑制率 Fresh weight inhibition rate/%	鲜重 Fresh weight/g	鲜重抑制率 Fresh weight inhibition rate/%
氯氟吡氧乙酸乳油（推荐剂量） Fluroxypyr EC at the recommended dose	750+0	2.45±0.13	62.60 b	1.57±0.04	79.42 b
氯氟吡氧乙酸乳油（50%推荐剂量） Fluroxypyr EC at 50 % of the recommended dose	375+0	3.02±0.37	53.89 bc	2.27±0.19	70.25 c
氯氟吡氧乙酸（25%推荐剂量） Fluroxypyr EC at 25 % of the recommended dose	187.5+0	3.55±0.26	45.80 c	2.84±0.12	62.78 d
氯氟吡氧乙酸乳油（推荐剂量）+16-8发酵液 Fluroxypyr EC at the recommended dose +16-8 strain fermentation broth	750+250	0.78±0.10	88.09 a	0.51±0.07	93.32 a
氯氟吡氧乙酸+16-8发酵液 Fluroxypyr EC at 50 % of the recommended dose +16-8 strain fermentation broth	375+250	0.94±0.01	85.65 a	0.64±0.03	91.61 a
氯氟吡氧乙酸+16-8发酵液 Fluroxypyr EC at 25 % of the recommended dose +16-8 strain fermentation broth	187.5+250	1.11±0.25	83.05 a	0.87±0.20	88.60 a
16-8发酵液16-8 strain fermentation broth	0+250	0.97±0.13	85.19 a	0.79±0.11	89.65 a
清水对照 Water control	0+0	6.55±0.39	0 d	7.63±0.25	0 e

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