菌剂对苜蓿青贮发酵品质及微生物群落的影响

doi:10.13560/j.cnki.biotech.bull.1985.2021-0831

摘要/Abstract

摘要：

为了开发适合苜蓿青贮的菌剂产品,研究青贮微生物群落与苜蓿青贮品质的关系。从苜蓿绿汁发酵液中分离筛选产酸能力强的乳酸菌,采用16S rDNA基因序列同源性分析鉴定并复合制备青贮细菌GSSW。青贮样品分为4组,添加苜蓿绿汁发酵液（aFGJ组）,添加宜生贮宝菌剂（YB组）,添加GSSW菌剂（GSSW组）及未添加菌剂作为对照（CK组）,采用高通量测序技术分析不同处理组青贮样品微生物群落多样性并检测理化发酵指标。结果表明：从苜蓿绿汁发酵液中分离得到高产酸乳酸菌株MXLZ-1、MXLZ-2、MXLZ-4经鉴定为Pediococcus parvulus、Lactobacillus plantarum、Pediococcus pentosaceus。与CK组相比,添加青贮菌剂的3组干物质和粗蛋白显著增加（P<0.05）;中性洗涤纤维、酸性洗涤纤维和木质素含量显著降低（P<0.05）;pH及氨氮/总氮显著降低（P<0.05）,乳酸含量显著升高（P<0.05）。4组青贮样品相对丰度较高的属均为Lactobacillis及Pediococcus;而添加菌剂的3组中Lactobacillis丰度高于Pediococcus,CK组相反;4组青贮样品L. plantarum,P. pentosaceus,L. brevis及Enterococcus mundti差异显著,添加3种菌剂均可提高L. plantarum丰度,降低P. pentosaceus及E. mundtii丰度。添加青贮菌剂均能增加有益菌的数量,减少有害菌数量,改善苜蓿青贮发酵品质;L. plantarum和P. pentosaceus的含量及比值对青贮优良发酵品质和组分变化起着重要作用;GSSW菌剂可运用于苜蓿青贮。

关键词: 绿汁发酵液, 苜蓿青贮, 高通量测序, 微生物群落, 发酵品质

Abstract:

In order to develop microbial inoculants suitable for alfalfa silage,the relationship between the microbial community of silage and the quality of alfalfa silage was investigated. The experiment was conducted to isolate and screen lactic acid strains with high acid production capacity from the alfalfa fermented green juice. The 16S rDNA gene sequence homology analysis was used to identify and then to prepare silage bacteria GSSW in compound. Silage samples were divided into 4 groups,adding with fermented green juice（aFGJ）,sila-max silage（YB）,GSSW bacteria,and no addition as control（CK）. A high throughput sequencing technique was used to analyze the microbial community diversity and detect physicochemical fermentation indexes. The results showed that MXLZ-1,MXLZ-2 and MXLZ-4 isolated from the fermented green juice were identified as Pediococcus pentosaceus,Lactobacillus plantarum and Pediococcus parvulus. Compared with CK group,dry matter and crude protein in the other 3 groups significantly increased（P<0.05）;neutral detergent fiber,acid detergent fiber and lignin significantly decreased（P<0.05）;pH and ammonia nitrogen/total nitrogen significantly decreased（P<0.05）,lactic acid significantly increased（P<0.05）. Lactobacillis and Pediococcus were the genera with high relative abundance of silage samples in the 4 groups. The abundance of Lactobacillis was higher than that of Pediococcus group with microbial inoculants,while that of CK was the opposite. Lactobacillus plantarum,Pediococcus pentosaceus,Lactobacillus brevis,and Enterococcus mundti of silage samples in the 4 groups were significantly different. Adding three kinds of microbial inoculants increased the abundance of L. plantarum and decreased the abundance of P. pentosaceus and E. mundtii. Adding silage inoculants increased the number of beneficial bacteria,reduced the number of harmful bacteria,and improved the fermentation quality of alfalfa silage. The GSSW can be used in alfalfa silage.

Key words: fermented green juice, alfalfa silage, high-throughput sequencing, microbial community, fermentation quality

毛婷, 牛永艳, 郑群, 杨涛, 穆永松, 祝英, 季彬, 王治业. 菌剂对苜蓿青贮发酵品质及微生物群落的影响[J]. 生物技术通报, 2021, 37(9): 86-94.

MAO Ting, NIU Yong-yan, ZHENG Qun, YANG Tao, MU Yong-song, ZHU Ying, JI Bin, WANG Zhi-ye. Effects of Microbial Inoculants on the Fermentation Quality and Microbial Community Diversity of Alfalfa Silage[J]. Biotechnology Bulletin, 2021, 37(9): 86-94.

图/表 7

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项目 Index	数值 Numeric value
干物质 Dry matter/（g·kg^-1 FM）	382±0.42
pH	6.43±0.25
酸性洗涤纤维Acid detergent fiber/（g·kg^-1 DM）	345±1.20
中性洗涤纤维Neutral detergent fiber/（g·kg^-1 DM）	452±0.35
粗蛋白Crude protein/（g·kg^-1 DM）	206±0.32
可溶性碳水化合物 Water soluble carbohydrate/（g·kg^-1 DM）	68.3±0.25

项目 Index	数值 Numeric value
干物质 Dry matter/（g·kg^-1 FM）	382±0.42
pH	6.43±0.25
酸性洗涤纤维Acid detergent fiber/（g·kg^-1 DM）	345±1.20
中性洗涤纤维Neutral detergent fiber/（g·kg^-1 DM）	452±0.35
粗蛋白Crude protein/（g·kg^-1 DM）	206±0.32
可溶性碳水化合物 Water soluble carbohydrate/（g·kg^-1 DM）	68.3±0.25

时间 Time/d	添加剂 Silage additive	干物质 DM/（g·kg^-1 FM）	营养成分/（g·kg^-1 DM）
时间 Time/d	添加剂 Silage additive	干物质 DM/（g·kg^-1 FM）	粗蛋白 CP	可溶性碳水化和物 WSC	中性洗涤纤维NDF	酸性洗涤纤维ADF	木质素 Lignin
30	YB	324±0.40^e	253±0.31^e	61.5±0.03^cd	333±0.26^c	286±0.30^b	86±0.52^ab
	aFGJ	323±0.26^e	225±0.25^c	59.7±0.02^ab	359±0.79^e	304±0.15^f	82±0.10^ab
	GSSW	305±0.58^ab	274±0.06^h	62.8±0.00^d	356±0.30^e	291±0.35^cd	87±0.06^b
	CK	304±0.21^ab	216.±0.29^b	60.4±0.05^bc	404±0.44^h	300±0.21^e	86±0.13^ab
50	YB	315±0.12^d	246±0.23^f	58.9±0.06^ab	325±0.17^b	263±0.41^a	83±0.05^ab
	aFGJ	311±0.21^cd	215±0.10^ab	58.6±0.03^a	330±0.06^bc	294±0.12^d	82±0.11^a
	GSSW	304±0.25ab	269±0.25^j	60.7±0.02^bc	343±0.15^d	287±0.16^bc	86±0.15^ab
	CK	299±0.12^a	213±0.29^ab	60.1±0.03^bcd	397±0.25^j	292±0.25^cd	87±0.13^ab
70	YB	317±0.20^d	237±0.25^d	62.5±0.01^d	323±0.32^b	260±0.27^a	83±0.14^ab
	aFGJ	315±0.35^d	210±0.12^a	60.6±0.03^bc	333±0.15^c	294±0.31^d	82±0.22^a
	GSSW	303±0.29^ab	245±0.25^f	67.8±0.21^f	314±0.21^a	284±0.09^b	83±0.16^ab
	CK	296±0.21^a	208±0.21^a	59.9±0.02^bcd	387±0.31^f	285±0.03^b	86±0.11^ab

时间 Time/d	添加剂 Silage additive	干物质 DM/（g·kg^-1 FM）	营养成分/（g·kg^-1 DM）
时间 Time/d	添加剂 Silage additive	干物质 DM/（g·kg^-1 FM）	粗蛋白 CP	可溶性碳水化和物 WSC	中性洗涤纤维NDF	酸性洗涤纤维ADF	木质素 Lignin
30	YB	324±0.40^e	253±0.31^e	61.5±0.03^cd	333±0.26^c	286±0.30^b	86±0.52^ab
	aFGJ	323±0.26^e	225±0.25^c	59.7±0.02^ab	359±0.79^e	304±0.15^f	82±0.10^ab
	GSSW	305±0.58^ab	274±0.06^h	62.8±0.00^d	356±0.30^e	291±0.35^cd	87±0.06^b
	CK	304±0.21^ab	216.±0.29^b	60.4±0.05^bc	404±0.44^h	300±0.21^e	86±0.13^ab
50	YB	315±0.12^d	246±0.23^f	58.9±0.06^ab	325±0.17^b	263±0.41^a	83±0.05^ab
	aFGJ	311±0.21^cd	215±0.10^ab	58.6±0.03^a	330±0.06^bc	294±0.12^d	82±0.11^a
	GSSW	304±0.25ab	269±0.25^j	60.7±0.02^bc	343±0.15^d	287±0.16^bc	86±0.15^ab
	CK	299±0.12^a	213±0.29^ab	60.1±0.03^bcd	397±0.25^j	292±0.25^cd	87±0.13^ab
70	YB	317±0.20^d	237±0.25^d	62.5±0.01^d	323±0.32^b	260±0.27^a	83±0.14^ab
	aFGJ	315±0.35^d	210±0.12^a	60.6±0.03^bc	333±0.15^c	294±0.31^d	82±0.22^a
	GSSW	303±0.29^ab	245±0.25^f	67.8±0.21^f	314±0.21^a	284±0.09^b	83±0.16^ab
	CK	296±0.21^a	208±0.21^a	59.9±0.02^bcd	387±0.31^f	285±0.03^b	86±0.11^ab

时间Time /d	添加剂 Silage additive	pH	氨氮/总氮 Ammonia nitrogen/Total nitrogen	挥发性脂肪酸 Volatile fatty acid/（g·kg^-1DM）
时间Time /d	添加剂 Silage additive	pH	氨氮/总氮 Ammonia nitrogen/Total nitrogen	乳酸 Lactic acid	乙酸 Acetic acid	丁酸 Butyric acid
30	YB	5.02±0.02^ef	60.5±0.03^a	42.3±0.01^e	12.4±0.04^c
	aFGJ	5.13±0.01^fg	63.1±0.03^c	32.9±0.06^a	11.1±0.01^b	0.1±0.01^a
	GSSW	5.08±0.07^e	59.7±0.05^a	43.2±0.04^f	11.2±0.02^b	--
	CK	5.57±0.07ⁱ	69.5±0.07^f	33.4±0.05^a	10.5±0.04^a	0.9±0.12^d
50	YB	4.56±0.02^c	60.3±0.11^a	44.6±0.02^h	18.9±0.04^d	--
	aFGJ	4.82±0.01^d	63.3±0.01^c	40.2±0.02^d	12.5±0.02^c	0.6±0.05^c
	GSSW	4.50±0.02^b	61.9±0.06^b	59.8±0.06^g	11.2±0.03^b	--
	CK	5.21±0.02^h	72.5±0.12^g	33.2±0.06^a	10.2±0.02^a	1.2±0.04^f
70	YB	4.25±0.01^a	67.2±0.05^e	44.5±0.03^h	20.2±0.03^e	--
	aFGJ	4.80±0.02^d	65.1±0.01^d	38.7±0.06^c	13.9±0.05^f	0.6±0.02^c
	GSSW	4.20±0.01^a	64.5±0.03^d	52.3±0.03^k	13.2±0.03^g	--
	CK	5.17±0.05^gh	78.5±0.12^h	35.5±0.04^b	11.2±0.01^b	2.0±0.01^ab