宁南山区不同植被恢复方式下土壤线虫群落特征:形态学鉴定与高通量测序法比较

doi:10.17520/biods.2021124

生物多样性 ›› 2021, Vol. 29 ›› Issue (11): 1513-1529. DOI: 10.17520/biods.2021124

所属专题：土壤生物与土壤健康

• 研究报告: 动物多样性 • 上一篇下一篇

宁南山区不同植被恢复方式下土壤线虫群落特征:形态学鉴定与高通量测序法比较

王楠¹^,², 黄菁华¹^,³^,^*(), 霍娜¹^,², 杨盼盼³^,⁴, 张欣玥¹^,⁵, 赵世伟¹^,²^,³

1.西北农林科技大学黄土高原土壤侵蚀与旱地农业国家重点实验室, 陕西杨凌 712100
2.西北农林科技大学资源环境学院, 陕西杨凌 712100
3.中国科学院水利部水土保持研究所, 陕西杨凌 712100
4.中国科学院大学, 北京 100049
5.西北农林科技大学林学院, 陕西杨凌 712100

收稿日期:2021-04-02 接受日期:2021-06-03 出版日期:2021-11-20 发布日期:2021-08-12
通讯作者: 黄菁华
作者简介:*E-mail: jhhuang@nwafu.edu.cn
基金资助:
国家自然科学基金(31500449);陕西省自然科学基础研究计划(2020JQ-435);陕西省引进博士经费(A279021836)

Characteristics of soil nematode community under different vegetation restoration approaches in the mountainous region of southern Ningxia: A comparative study based on morphological identification and high-throughput sequencing methods

Nan Wang¹^,², Jinghua Huang¹^,³^,^*(), Na Huo¹^,², Panpan Yang³^,⁴, Xinyue Zhang¹^,⁵, Shiwei Zhao¹^,²^,³

1 State Key Laboratory of Soil Erosion and Dryland Farming on Loess Plateau, Northwest A&F University, Yangling, Shaanxi 712100
2 College of Natural Resources and Environment, Northwest A&F University, Yangling, Shaanxi 712100
3 Institute of Soil and Water Conservation, Chinese Academy of Sciences & Ministry of Water Resources, Yangling, Shaanxi 712100
4 University of Chinese Academy of Sciences, Beijing 100049
5 College of Forestry, Northwest A&F University, Yangling, Shaanxi 712100

Received:2021-04-02 Accepted:2021-06-03 Online:2021-11-20 Published:2021-08-12
Contact: Jinghua Huang

摘要/Abstract

摘要：

线虫是土壤食物网的重要组分, 也是土壤健康与生态系统恢复的重要指示生物, 因此准确测定线虫群落特征是发挥其生态指示作用的基础。传统线虫学研究多采用形态学鉴定方法, 但高通量测序技术近年来也逐渐受到重视。然而, 关于这两种方法的对比研究目前仍比较缺乏。本研究同时采用形态学鉴定和高通量测序法, 在黄土高原宁夏南部山区, 对不同植被恢复方式下(农田、自然恢复草地、柠条(Caragana korshinskii)人工林地和苜蓿人工草地)土壤线虫的数量、群落格局和生态指数进行了测定和比较。结果表明: (1)高通量测序仅能提供线虫类群的相对多度, 而形态学鉴定法能够测定土壤线虫的绝对多度, 后者测定结果表明3种植被恢复样地, 特别是自然恢复草地和柠条人工林地, 较农田具有更高的土壤线虫多度; (2)高通量测序法检获的线虫类群数(3纲4目26科42属)高于形态学鉴定法的测定结果(2纲3目18科27属), 但两种方法仅检获15个共有线虫属, 前者检测到的植物寄生线虫属数(22属)较后者(7属)显著增加, 而食细菌线虫和杂食-捕食线虫则相反; (3)在两种方法下, 相比农田, 3种植被恢复样地尤其是自然恢复草地和柠条人工林地, 其食微线虫的相对多度均显著下降, 而植物寄生线虫和杂食-捕食线虫的相对多度大幅上升, 这也导致线虫成熟度指数(MI)和植物寄生线虫指数(PPI)的提高及瓦斯乐卡指数(WI)的显著下降; (4)相比形态学鉴定法, 高通量测序法能检测到更丰富多样的植物寄生线虫, 在该方法下土壤线虫群落的组成、结构和生态指数在植被恢复样地与农田之间的差异也更为显著。综上所述, 采用形态学鉴定和高通量测序法测定的不同植被恢复方式下的线虫群落特征具有显著差异。

关键词: 形态学鉴定, 高通量测序, 土壤线虫, 群落结构, 植被恢复

Abstract

Aims Nematodes are considered as an important part of the soil food web, and their community characteristics are an effective indicator of soil health and ecosystem restoration. The accurate measurement of soil nematode communities is necessary to better understand their ecological function. Historically, comparisons in nematode morphology has been used to understand nematode community characteristics. In recent years, the use of high-throughput sequencing (HTS) methods has become more popular. However, relatively little is known about how these two methods compare when analyzing soil nematode communities.
Methods Here, we used morphological identification and HTS methods to simultaneously analyze soil nematode abundance, community composition and structure, and ecological indexes, under different vegetation restoration approaches (cropland, naturally restored grassland, Caragana korshinskiiplantation and Medicago sativaartificial grassland) in the mountain area of southern Ningxia in the Loess Plateau region.
Result We found that morphological identification is a more accurate method to determine the absolute abundance of soil nematodes, while HTS can only obtain relative abundance data. In our study, the morphological method showed higher abundance of soil nematodes in the sites under vegetation restoration, especially in the naturally restored grassland and C. korshinskiiplantation. The HTS method, on the other hand, detected more nematode genera (42 genera belonging to 3 classes, 4 orders, and 26 families) than morphological method (27 genera belonging to 2 classes, 3 orders, and 18 families). However, only 15 genera were simultaneously identified with both methods, because the HTS method detected more plant-parasitic nematode genera (22) but fewer genera of bacterial-feeding nematodes and omnivores-predators than the morphological method. One major result indicated by both methods showed that the relative abundance of microbial-feeding nematodes greatly decreased, while those of plant parasites and omnivores-predators substantially increased, in all the sites under vegetation restoration when compared with nematodes in the farmland. This was especially the case in the naturally restored grassland and C. korshinskiiplantation, accompanied with the increases of maturity index (MI) and plant-parasitic index (PPI) and a decrease in the Wasilewska index (WI). When compared with the morphological method, the HTS method could detect more abundant and diverse plant parasites. Therefore, more significant differences were found in the composition, structure and ecological indexes of soil nematode communities when the HTS method was applied.
Conclusion Overall, the characteristics of soil nematode communities and their response patterns to vegetation restoration highly depended on the applied methodology, which greatly influences the understanding and evaluation of how vegetation restoration impacts the soil ecosystem.

Key words: morphological identification, high-throughput sequencing, soil nematodes, community structure, vegetation restoration

王楠, 黄菁华, 霍娜, 杨盼盼, 张欣玥, 赵世伟 (2021) 宁南山区不同植被恢复方式下土壤线虫群落特征:形态学鉴定与高通量测序法比较. 生物多样性, 29, 1513-1529. DOI: 10.17520/biods.2021124.

Nan Wang, Jinghua Huang, Na Huo, Panpan Yang, Xinyue Zhang, Shiwei Zhao (2021) Characteristics of soil nematode community under different vegetation restoration approaches in the mountainous region of southern Ningxia: A comparative study based on morphological identification and high-throughput sequencing methods. Biodiversity Science, 29, 1513-1529. DOI: 10.17520/biods.2021124.

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https://www.biodiversity-science.net/CN/Y2021/V29/I11/1513

图/表 9

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样地 Sampling site	植被特征 Vegetation characteristics	恢复年限 Restoration year	经度 Longitude	纬度 Latitude	海拔 Altitude (m)	坡向 Aspect	坡度 Slope degree (°)
农田 Cropland (CL)	玉米 Corn	>20	106°27′54′′ E	36°0′42′′ N	1,530	西偏北14° W14°N	7
自然恢复草地 Natural grassland (NR)	长芒草 Stipa bungeana 百里香 Thymus mongolicus 冰草 Agropyron cristatum	24	106°28′12′′ E	36°0′35′′ N	1,673	西偏北12° W12°N	24
柠条人工林地 Caragana korshinskii plantation (CK)	柠条 Caragana korshinskii 长芒草 Stipa bungeana 冰草 Agropyron cristatum	24	106°28′12′′ E	36°0′40′′ N	1,670	西偏北22° W22°N	23
苜蓿人工草地 Medicago sativa grassland (MS)	紫花苜蓿 Medicago sativa 长芒草 Stipa bungeana	12	106°28′02′′ E	36°0′46′′ N	1,591	西偏北16° W16°N	15

样地 Sampling site	植被特征 Vegetation characteristics	恢复年限 Restoration year	经度 Longitude	纬度 Latitude	海拔 Altitude (m)	坡向 Aspect	坡度 Slope degree (°)
农田 Cropland (CL)	玉米 Corn	>20	106°27′54′′ E	36°0′42′′ N	1,530	西偏北14° W14°N	7
自然恢复草地 Natural grassland (NR)	长芒草 Stipa bungeana 百里香 Thymus mongolicus 冰草 Agropyron cristatum	24	106°28′12′′ E	36°0′35′′ N	1,673	西偏北12° W12°N	24
柠条人工林地 Caragana korshinskii plantation (CK)	柠条 Caragana korshinskii 长芒草 Stipa bungeana 冰草 Agropyron cristatum	24	106°28′12′′ E	36°0′40′′ N	1,670	西偏北22° W22°N	23
苜蓿人工草地 Medicago sativa grassland (MS)	紫花苜蓿 Medicago sativa 长芒草 Stipa bungeana	12	106°28′02′′ E	36°0′46′′ N	1,591	西偏北16° W16°N	15

样地 Sampling site	pH	土壤含水量 SM (%)	有机碳 SOC (g/kg)	全氮 TN (g/kg)	速效磷 Olsen-P (mg/kg)	铵态氮 NH₄⁺-N (mg/kg)	硝态氮 NO₃^--N (mg/kg)
农田 Cropland (CL)	8.24^a	6.40^a	7.34^b	0.80^c	45.58^a	0.73^ab	1.76^a
自然恢复草地 Natural grassland (NR)	8.02^b	5.55^a	15.90^a	1.42^a	5.20^b	1.46^a	2.70^a
柠条人工林地 Caragana korshinskii plantation (CK)	8.00^b	6.02^a	17.47^a	1.32^ab	5.73^b	0.62^ab	2.25^a
苜蓿人工草地 Medicago sativa grassland (MS)	8.26^a	6.37^a	8.11^b	0.97^bc	7.40^b	0.40^b	1.20^a

样地 Sampling site	pH	土壤含水量 SM (%)	有机碳 SOC (g/kg)	全氮 TN (g/kg)	速效磷 Olsen-P (mg/kg)	铵态氮 NH₄⁺-N (mg/kg)	硝态氮 NO₃^--N (mg/kg)
农田 Cropland (CL)	8.24^a	6.40^a	7.34^b	0.80^c	45.58^a	0.73^ab	1.76^a
自然恢复草地 Natural grassland (NR)	8.02^b	5.55^a	15.90^a	1.42^a	5.20^b	1.46^a	2.70^a
柠条人工林地 Caragana korshinskii plantation (CK)	8.00^b	6.02^a	17.47^a	1.32^ab	5.73^b	0.62^ab	2.25^a
苜蓿人工草地 Medicago sativa grassland (MS)	8.26^a	6.37^a	8.11^b	0.97^bc	7.40^b	0.40^b	1.20^a

样地 Sampling site	序列数 Sequence number	OTU数目 Number of OTUs	不能识别OTU比例 Proportion of unidentified OTUs (%)	覆盖率 Coverage (%)
农田 Cropland (CL)	447^ab	26	59.82	98.23^a
自然恢复草地 Natural grassland (NR)	633^ab	38	59.12	98.44^a
柠条人工林地 Caragana korshinskii plantation (CK)	938^a	48	57.53	98.24^a
苜蓿人工草地 Medicago sativa grassland (MS)	151^b	42	57.40	98.02^a

宁南山区不同植被恢复方式下土壤线虫群落特征:形态学鉴定与高通量测序法比较

Characteristics of soil nematode community under different vegetation restoration approaches in the mountainous region of southern Ningxia: A comparative study based on morphological identification and high-throughput sequencing methods

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线虫属 Nematode genus	营养类群 Trophic group	c-p值 c-p value	相对多度 Relative abundance (%)
			农田 (CL)		自然恢复草地 (NR)		柠条人工林地 (CK)		苜蓿人工草地 (MS)
			I	II	I	II	I	II	I	II
头叶属 Cephalobus	BF	2	19.5	22.0	0.6	2.7	7.6	5.4	22.6	5.8
拟丽突属 Acrobeloides	BF	2	0.3	11.4	-	1.9	2.5	4.2	0.3	5.0
管咽属 Aulolaimus	BF	3	-	-	-	-	-	-	-	0.8
中杆属 Mesorhabditis	BF	1	-	5.1	-	-	-	-	-	-
三等齿属 Pelodera	BF	1	-	4.6	-	-	-	-	-	-
无咽属 Alaimus	BF	4	-	1.6	-	-	-	-	-	-
丽突属 Acrobeles	BF	2	1.7	-	11.5	-	8.3	-	6.1	-
鹿角唇属 Cervidellus	BF	2	2.4	-	4.1	-	6.0	-	0.6	-
板唇属 Chiloplacus	BF	2	-	-	1.9	-	0.3	-	-	-
真头叶属 Eucephalobus	BF	2	5.6	-	3.5	-	2.2	-	2.9	-
线虫属 Nematode genus	营养类群 Trophic group	c-p值 c-p value	相对多度 Relative abundance (%)
			农田 (CL)		自然恢复草地 (NR)		柠条人工林地 (CK)		苜蓿人工草地 (MS)
			I	II	I	II	I	II	I	II
小杆属 Rhabditis	BF	1	6.0	-	-	-	0.3	-	-	-
茎属 Ditylenchus	PP	2	13.5	11.4	4.5	0.1	7.0	0.2	16.8	2.1
垫刃属 Tylenchus	PP	2	0.3	-	6.0	-	2.2	<0.1	1.3	-
螺旋属 Helicotylenchus	PP	3	1.2	-	1.9	-	2.2	0.1	-	2.5
短体属 Pratylenchus	PP	3	1.9	-	1.6	-	0.3	-	1.0	1.2
矮化属 Tylenchorhynchus	PP	3	3.9	-	21.1	-	16.2	0.4	4.5	0.4
剑属 Xiphinema	PP	5	0.3	-	0.3	1.2	1.3	2.0	-	0.7
盘旋属 Rotylenchus	PP	3	3.4	-	4.4	<0.1	4.8	-	18.9	-
针属 Paratylenchus	PP	2	-	<0.1	-	61.5	-	39.9	-	34.4
细小线虫属 Gracilacus	PP	2	-	-	-	0.2	-	0.1	-	0.2
根结线虫属 Meloidogyne	PP	3	-	-	-	-	-	0.4	-	-
细纹垫刃属 Lelenchus	PP	2	-	0.3	-	-	-	-	-	0.8
新平滑垫刃属 Neopsilenchus	PP	2	-	-	-	-	-	0.2	-	0.2
伞滑刃属 Bursaphelenchus	PP	2	-	-	-	-	-	0.2	-	0.7
米卡垫刃属 Miculenchus	PP	2	-	-	-	-	-	0.2	-	-
轮线虫属 Criconemoides	PP	3	-	-	-	10.2	-	13.6	-	4.5
大默林属 Amplimerlinius	PP	3	-	-	-	11.0	-	0.4	-	0.4
裸矛属 Psilenchus	PP	2	-	-	-	-	-	-	-	1.2
突腔唇属 Ecphyadophora	PP	2	-	-	-	0.1	-	-	-	-
叉针属 Boleodorus	PP	2	-	0.6	-	0.1	-	0.1	-	0.4
剑尾垫刃属 Malenchus	PP	2	-	0.8	-	-	-	-	-	-
巴兹尔属 Basiria	PP	2	-	-	-	-	-	-	-	1.1
牙咽属 Dorylaimellus	PP	5	-	-	-	-	-	-	-	0.2
垫咽属 Tylencholaimus	FF	4	4.1	-	9.6	0.2	6.3	0.5	12.7	0.4
滑刃属 Aphelenchoides	FF	2	0.6	-	1.0	0.2	-	0.4	-	0.8
真滑刃属 Aphelenchus	FF	2	25.4	8.8	6.1	0.2	6.0	3.3	3.2	9.5
艾普鲁斯属 Aprutides	FF	2	-	-	-	0.1	-	-	-	-
瘤咽属 Tylencholaimellus	FF	4	-	28.0	-	0.8	-	1.4	-	11.2
丝尾垫刃属 Filenchus	FF	2	-	1.8	-	-	-	-	-	-
孔咽属 Aporcelaimellus	OP	5	2.0	0.1	1.6	0.7	3.8	1.1	2.9	4.3
前矛线属 Prodorylaimus	OP	5	-	-	0.3	0.2	-	0.1	-	-
Allodorylaimus	OP	4	-	3.4	-	0.8	-	0.8	-	1.5
中矛线属 Mesodorylaimus	OP	5	-	-	-	3.3	-	9.8	-	-
独壁齿属 Campydora	OP	4	-	-	-	4.1	-	14.4	-	8.3
真矛线属 Eudorylaimus	OP	4	-	-	-	-	1.9	-	1.0	-
高知属 Kochinema	OP	4	-	-	1.9	-	1.3	-	0.3	-
微矛线属 Microdorylaimus	OP	4	-	-	-	-	0.6	-	-	-
缢咽属 Paraxonchium	OP	5	1.0	-	-	-	0.3	-	-	-
索努斯属 Thonus	OP	4	4.7	-	3.5	-	7.9	-	2.2	-
盘咽属 Discolaimus	OP	5	-	-	0.3	0.4	0.3	0.8	0.3	1.6
Paractinolaimus	OP	5	-	<0.1	-	-	-	-	-	-
狭咽属 Discolaimium	OP	5	1.9	-	7.6	0.4	5.4	0.8	2.2	1.6
Paravulvus	OP	5	0.3	-	6.7	-	4.8	-	0.3	-
属数 Genera number			21	16	22	23	25	27	19	28

线虫属 Nematode genus	营养类群 Trophic group	r	P
头叶属 Cephalobus	BF	0.455	0.14
拟丽突属 Acrobeloides	BF	-0.007	0.98
茎属 Ditylenchus	PP	0.061	0.85
垫刃属 Tylenchus	PP	-0.168	0.60
螺旋属 Helicotylenchus	PP	-0.305	0.34
短体属 Pratylenchus	PP	-0.080	0.80
盘旋属 Rotylenchus	PP	-0.183	0.57
矮化属 Tylenchorhynchus	PP	-0.062	0.85
剑属 Xiphinema	PP	0.202	0.53
滑刃属 Aphelenchoides	FF	-0.196	0.54
真滑刃属 Aphelenchus	FF	0.266	0.40
垫咽属 Tylencholaimus	FF	-0.144	0.66
孔咽属 Aporcelaimellus	OP	0.020	0.95
前矛线属 Prodorylaimus	OP	0.291	0.36
盘咽属 Discolaimus	OP	-0.292	0.36

	H'		λ	J'	SR	MI	PPI	WI	NCR
形态学鉴定法 Morphological identification method
农田 Cropland (CL)	2.24^bc	0.15^ab		0.81^bc	2.97^a	2.38^c	2.43^a	2.90^a	0.54^a
自然恢复草地 Natural grassland (NR)	2.48^ab	0.12^b		0.86^ab	2.53^a	3.10^a	2.75^a	0.98^a	0.57^a
柠条人工林地 Caragana korshinskii plantation (CK)	2.68^a	0.09^b		0.90^a	2.81^a	3.07^a	2.80^a	1.33^a	0.66^a
苜蓿人工草地 Medicago sativa grassland (MS)	2.05^c	0.19^a		0.76^c	2.20^a	2.73^b	2.51^a	1.53^a	0.69^a
高通量测序 High-throughput sequencing
农田 Cropland (CL)	1.54^a		0.28^a	0.60^a	2.21^b	2.76^a	2.00^a	4.58^a	0.49^a
自然恢复草地 Natural grassland (NR)	0.95^a		0.46^a	0.30^b	3.67^ab	3.71^a	2.29^a	0.08^b	0.67^a
柠条人工林地 Caragana korshinskii plantation (CK)	1.45^a		0.30^a	0.42^ab	5.04^a	3.55^a	2.44^a	0.31^b	0.63^a
苜蓿人工草地 Medicago sativa grassland (MS)	1.79^a		0.24^a	0.56^ab	4.62^a	3.39^a	2.27^a	0.90^b	0.34^a

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