霉变饲用玉米真菌的分离、鉴定与乳酸菌素对其的防霉抑菌效果

doi:10.3969/j.issn.1004-1524.2021.07.14

浙江农业学报 ›› 2021, Vol. 33 ›› Issue (7): 1283-1291.DOI: 10.3969/j.issn.1004-1524.2021.07.14

霉变饲用玉米真菌的分离、鉴定与乳酸菌素对其的防霉抑菌效果

江宇航¹^,²(), 辛维岗¹^,², 张棋麟¹^,², 邓先余¹^,², 王峰¹^,², 林连兵¹^,²^,^*()

1.昆明理工大学生命科学与技术学院,云南昆明 650500
2.云南省高校饲用抗生素替代技术工程研究中心,云南昆明 650500

收稿日期:2020-10-12 出版日期:2021-07-25 发布日期:2021-08-06
通讯作者: 林连兵
作者简介:*林连兵,E-mail: linlb@kmust.edu.cn
江宇航(1996—),男,四川遂宁人,硕士研究生,研究方向为微生物资源开发与利用。E-mail: 1584727524@qq.com
基金资助:
国家自然科学基金(31760042)

Isolation and identification of fungi from mildewed feed corn and study on anti-mildew and antifungal effects of lactobacillin

JIANG Yuhang¹^,²(), XIN Weigang¹^,², ZHANG Qili¹^,², DENG Xianyu¹^,², WANG Feng¹^,², LIN Lianbing¹^,²^,^*()

1. Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming 650500, China
2. Engineering Research Center for Replacement Technology of Feed Antibiotics of Yunnan College, Kunming 650500, China

Received:2020-10-12 Online:2021-07-25 Published:2021-08-06
Contact: LIN Lianbing

摘要/Abstract

摘要：

为明确霉变饲用玉米中真菌的组成,探讨乳酸菌素对霉变真菌的抑菌效果和延缓饲用玉米霉变的作用。本研究采用传统真菌培养方法,对霉变饲用玉米中的霉变真菌进行分离,通过ITS扩增子测序确定霉变饲用玉米中可培养真菌的分类学地位;利用牛津杯法评估乳酸菌素对霉变真菌的抑制效果,并设置不同添加比例(10%、20%、30%)的乳酸菌素粗提液,探究乳酸菌素对饲用玉米发生霉变的周期与霉变程度(以结块、霉斑、霉味、霉菌数等参数作为观测指标)的延缓效果。结果发现:霉变饲用玉米中真菌种类多样性高,筛选得到了隶属曲霉属(Aspergillus)、青霉属(Penicillium)、毛霉属(Mucor)、镰刀菌属(Fusarium)和地霉属(Geotrichum)等三门5属的霉变真菌。乳酸菌素对霉变饲用玉米中各种霉菌均有良好的抑制效果,尤其是对青霉属中的皮壳青霉(Penicillium crustosum)抑菌活性最佳,抑菌圈直径可达(16.54±0.23)mm,对地霉属中的白地霉(Geotrichum candidum)抑菌活性最差,抑菌圈直径仅为(9.26±0.16)mm,但均显著(P<0.05)高于对照组。添加不同添加比例的乳酸菌素粗提液至饲用玉米中,在10 d内均无明显霉变现象出现,在30 d时各处理组与对照组之间出现了明显的霉变程度差异。此外,随着乳酸菌素粗提液添加比例的增加,饲用玉米霉变程度逐渐降低,添加比例30%的处理组中,真菌菌落仅有(9.28±0.59) × 10⁷ CFU·g^-1。总而言之,霉变饲用玉米中真菌种类丰富,存在多种产毒素霉菌;利用乳酸菌素可有效抑制霉变真菌,减缓饲用玉米发生霉变的周期与霉变程度。

关键词: 饲用玉米, 霉变真菌, 菌种鉴定, 乳酸菌素, 抑菌效果

Abstract:

To clarify composition of fungi species in mildewed feed corn and explore antifungal effect of lactobacillin on mildewed fungi and the effect of delaying feed corn mildewing, in this study, mildew fungi were isolated from mildewed feeding corn by using traditional fungi culture method, and the taxonomic status of culturable fungi in mildewed feed corn was determined by sequencing of ITS amplicons. The inhibitory effect of lactobacillin on mildew fungi was evaluated using the Oxford Cup method, and crude lactobacillin with different proportions (10%, 20%, 30%) were set to explore mildew cycle and mildew degree of lactobacillin on delayed feeding corn mildew (caking, mildew spot, mildew taste, and mildew number were taken as observation indices). The results showed that diversity of fungi species in mildewed feeding corn was high, and five genera of fungus (Aspergillus, Penicillium, Mucor, Fusarium and Geotrichum) belonging to three phyla were screened. Lactobacillin had a good inhibitory effects on mildew fungi of various genera in mildewed feed corn, it had the best antifungal activity against Penicillium crustosum, the diameter of the inhibition circle could reach (16.54 ± 0.23) mm. The antifungal activity against Geotrichum candidum was the worst, only with the diameter of inhibition zone of (9.26±0.16) mm, but their diameter of inhibition zones were significantly (P <0.05) higher than that of the control group. Different proportions of lactobacillin crude extract were added to feeding corn, there was no obvious mildew phenomen within 10 days, and there was obvious difference in the degree of mildew between the treatment groups and the control group on the 30th day. In addition, with the increasing of proportion of added lactobacillin, the mildewing degree of feed corn gradually decreased, and the fungi colonies were only (9.28±0.59) × 10⁷ CFU·g^-1 in the treatment group with 30% addition proportion. In conclusion, there were rich species of fungi in moldy feed corn, and there were a variety of toxin-producing molds; Lactobacillin could effectively inhibit mildew fungi, slow down the cycle and degree of mildew in feed corn.

Key words: feed corn, mildew fungi, strain identification, lactobacillin, antibacterial activity

中图分类号:

S513

江宇航, 辛维岗, 张棋麟, 邓先余, 王峰, 林连兵. 霉变饲用玉米真菌的分离、鉴定与乳酸菌素对其的防霉抑菌效果[J]. 浙江农业学报, 2021, 33(7): 1283-1291.

JIANG Yuhang, XIN Weigang, ZHANG Qili, DENG Xianyu, WANG Feng, LIN Lianbing. Isolation and identification of fungi from mildewed feed corn and study on anti-mildew and antifungal effects of lactobacillin[J]. Acta Agriculturae Zhejiangensis, 2021, 33(7): 1283-1291.

图/表 5

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门 Phylum	菌株 Strain	分子鉴定 Molecular identification	序列一致性 Sequence identity/%	GenBank 登录号 GenBank accession No.
子囊菌门Ascomycota	MR-1	黄曲霉Aspergillus flavus	100	MG818938
半知菌亚门Deuteromycotina	MR-4	米曲霉 Aspergillus oryzae	100	MK748312
	MR-6	黑曲霉 Aspergillus niger	100	MN509035
	MR-7	皮壳青霉 Penicillium crustosum	99	MG975627
	MR-10	拟轮枝镰孢菌 Fusarium verticillioides	100	MN429250
	MR-12	白地霉 Geotrichum candidum	99	KT828737
接合菌亚门Zygomycotina	MR-8	卷枝毛霉 Mucor circinelloides	99	MK559692

门 Phylum	菌株 Strain	分子鉴定 Molecular identification	序列一致性 Sequence identity/%	GenBank 登录号 GenBank accession No.
子囊菌门Ascomycota	MR-1	黄曲霉Aspergillus flavus	100	MG818938
半知菌亚门Deuteromycotina	MR-4	米曲霉 Aspergillus oryzae	100	MK748312
	MR-6	黑曲霉 Aspergillus niger	100	MN509035
	MR-7	皮壳青霉 Penicillium crustosum	99	MG975627
	MR-10	拟轮枝镰孢菌 Fusarium verticillioides	100	MN429250
	MR-12	白地霉 Geotrichum candidum	99	KT828737
接合菌亚门Zygomycotina	MR-8	卷枝毛霉 Mucor circinelloides	99	MK559692

菌株 Strains	抑菌圈直径 Inhibition zone diameter	对照组 Control group
MR-1	11.36±0.06^*	0.30±0.03
MR-4	12.24±0.18^*	0.16±0.06
MR-6	15.65±0.22^*	0.12±0.04
MR-7	16.54±0.23^*	0.31±0.10
MR-8	13.36±0.12^*	0.52±0.06
MR-10	14.45±0.13^*	0.23±0.01
MR-12	9.26±0.16^*	0.41±0.02

菌株 Strains	抑菌圈直径 Inhibition zone diameter	对照组 Control group
MR-1	11.36±0.06^*	0.30±0.03
MR-4	12.24±0.18^*	0.16±0.06
MR-6	15.65±0.22^*	0.12±0.04
MR-7	16.54±0.23^*	0.31±0.10
MR-8	13.36±0.12^*	0.52±0.06
MR-10	14.45±0.13^*	0.23±0.01
MR-12	9.26±0.16^*	0.41±0.02

t/d	处理组Treatment group			对照组Control group
t/d	10%	20%	30%	CA10%	CA20%		CA30%
5	—,—,—	—,—,—	—,—,—	—,—,—		—,—,+		—,*,+
10	—,—,—	—,—,—	—,—,—	—,*,+		〇,*,++		〇〇,**,++
15	〇,*,—	—,*,+	—,*,—	〇,*,+		〇〇,**,++		〇〇〇,***,+++
20	〇,*,++	〇,*,++	〇,*,+	〇,**,++		〇〇〇,***,+++		〇〇〇〇,****,+++
25	〇〇,*,++	〇,**,++	〇,**,+	〇〇,***,++		〇〇〇〇,****,+++		〇〇〇〇〇,*****,++++
30	〇〇,**,++	〇,**,++	〇,**,+	〇〇〇,***,+++		〇〇〇〇〇,*****,++++		〇〇〇〇〇,*****,++++

霉变饲用玉米真菌的分离、鉴定与乳酸菌素对其的防霉抑菌效果

Isolation and identification of fungi from mildewed feed corn and study on anti-mildew and antifungal effects of lactobacillin

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