Earthworms promote greater richness and abundance in the emergence of plant species across a grassland-forest ecotone

Aims Chalk grasslands are subject to vegetation dynamics that range from species-rich open grasslands to tall and encroached grasslands, and woods and forests. In grasslands, earthworms impact plant communities and ecosystem functioning through the modification of soil physical, chemical and microbiological properties, but also through their selective ingestion and vertical transportation of seeds from the soil seed bank. Laboratory experiments showed that seed–earthworm interactions are species specific, but little is known on the impact of seed–earthworm interactions in the field. The overall aim of this study was to better understand seed–earthworm interactions and their impact on the plant community. First we analyzed the composition of seedlings emerging from casts after earthworm ingestion. Then we compared seedling composition in casts to the plant composition of emerging seedlings from the soil and of the aboveground vegetation along four stages of the secondary succession of chalk grasslands.Methods Four stages of the secondary succession of a chalk grassland—from open sward to woods—were sampled in Upper Normandy, France, in February 2010. Within each successional stage (×3 replicates), we sampled the standing vegetation, soil seed bank at three soil depths (0–2, 2–5 and 5–10cm) and earthworm surface casts along transects. Soil and cast samples were water sieved before samples were spread onto trays and placed into a greenhouse. Emerging seedlings were counted and identified. Effect of successional stage and origin of samples on mean and variability of abundance and species richness of seedlings emerging from casts and soil seed banks were analyzed. Plant compositions were compared between all sample types. We used generalized mixed-effect models and a distance-based redundancy multivariate analysis.Important findings Seedling abundance was always higher in earthworm casts than in the soil seed bank and increased up to 5-fold, 4-fold and 3.5-fold, respectively, in the tall grassland, woods and encroached grassland compared to the soil surface layer. Species richness was also higher in earthworm casts than in the soil seed bank in all successional stages, with a 4-fold increase in the encroached grassland. The plant composition of the standing vegetation was more similar to that of seedlings from casts than to that of seedlings from the soil seed bank. Seedlings diversity emerging from casts in the tall and encroached grasslands tended toward the diversity found in woods. Our results indicate that earthworms may promote the emergence of seedlings. We also suggest that the loss of some plant species in the seed bank and the tall grass vegetation in intermediary successional stages modify the local conditions and prevent the further establishment of early-successional plant species.     

Partitioning of habitable pore space in earthworm burrows

Earthworms affect macro-pore structure of soils. However, some studies suggest that earthworm burrow walls and casts themselves differ greatly in structure from surrounding soils, potentially creating habitat for microbivorours nematodes which accelerate the decomposition and C and N mineralization. In this study aggregates were sampled from the burrow walls of the anecic earthworm Lumbricus terrestris and bulk soil (not altered by earthworms) from mesocosm incubated in the lab for 0, 1, 3, 5 and 16 weeks. Pore volumes and pore sizes were measured in triplicate with Mercury Intrusion Porosimetry (MIP). This method is well suited to establish pore size structure in the context of habitat, because it measures the stepwise intrusion of mercury from the outside of the aggregate into ever smaller pores. The progress of mercury into the aggregate interior thus resembles potential paths of a nematode into accessible habitable pore spaces residing in an aggregate. Total specific pore volume, V(s), varied between 0.13 and 0.18 mL/g and increased from 3 to 16 weeks in both burrow and bulk soil. Differences between total V(s) of bulk and burrow samples were not significant on any sampling date. However, differences were significant for pore size fractions at the scale of nematode body diameter.     

Response of soil nematode community structure and diversity to long-term land use in the black soil region in China

Soil nematodes are sensitive to environmental changes and are used widely as indicators of soil conditions. The community structure and diversity of soil nematodes were studied in different long-term land use regimes in the black soil area in Northeast China. The land use regimes were maintained for 22?years, and included crop land (CL), grass land (GL) and bare land (BL). Soil samples were taken throughout the growing season, and nematodes were extracted and identified. A total of 39 nematode genera with relative abundance over 0.1?% were identified. Heterodera was the dominant genus in CL; Boleodorus was the dominant genus in GL, and Boleodorus, Eucephalobus and Filenchus were the dominant genera in BL. Land use had a significant effect on abundance of all soil nematode tropic groups and ecological indices. Sampling time had an effect on soil nematode abundance, but only on three of the eight nematode ecological indices MI (maturity index of free-living nematode), CI (channel index) and EI (enrichment index). SR (species richness index) was highest in GL where plant species diversity was also high. The CI was the highest in BL among three land uses, which means the soil food web dominated, with fungal decomposition channels in BL. Soil nematode community structure and diversity was shown to be an effective and informative tool for analyzing ecological aspects of land use in black soil regions. The data are inconclusive as to whether the effect of land use on soil nematode parameters is direct, or indirect via inducing changes in soil physicochemical properties.     

施用农用化学品及生防制剂对土壤线虫群落动态影响的研究

1　引　　言在农业生态系统中 ,农业管理 (如化肥、农药等农用化学品的施用等 )对土壤线虫群落组成及多样性产生了显著影响[2～ 4,7] .线虫是农田土壤中的生物因子 ,对植物的生长可以产生直接或间接的影响 .植物寄生线虫对植物的影响主要包括 3种方式 :1)通过在植物的根部取食 ,直接影响植物的营养状况 ,造成植物营养不良 ;2 )通过其口针的穿刺过程 ,传播植物病毒 ,引起植物病毒病 ;3)通过线虫取食植物根部造成的伤口 ,引起与其它植物病原物的复合侵染 .非植物寄生线虫多数是自由生活的线虫 ,在有人为影响的农田中主要是腐生线虫参与土壤中…     

Profiling Nematode Communities in Unmanaged Flowerbed and Agricultural Field Soils in Japan by DNA Barcode Sequencing

Soil nematodes play crucial roles in the soil food web and are a suitable indicator for assessing soil environments and ecosystems. Previous nematode community analyses based on nematode morphology classification have been shown to be useful for assessing various soil environments. Here we have conducted DNA barcode analysis for soil nematode community analyses in Japanese soils. We isolated nematodes from two different environmental soils of an unmanaged flowerbed and an agricultural field using the improved flotation-sieving method. Small subunit (SSU) rDNA fragments were directly amplified from each of 68 (flowerbed samples) and 48 (field samples) isolated nematodes to determine the nucleotide sequence. Sixteen and thirteen operational taxonomic units (OTUs) were obtained by multiple sequence alignment from the flowerbed and agricultural field nematodes, respectively. All 29 SSU rDNA-derived OTUs (rOTUs) were further mapped onto a phylogenetic tree with 107 known nematode species. Interestingly, the two nematode communities examined were clearly distinct from each other in terms of trophic groups: Animal predators and plant feeders were markedly abundant in the flowerbed soils, in contrast, bacterial feeders were dominantly observed in the agricultural field soils. The data from the flowerbed nematodes suggests a possible food web among two different trophic nematode groups and plants (weeds) in the closed soil environment. Finally, DNA sequences derived from the mitochondrial cytochrome oxidase c subunit 1 (COI) gene were determined as a DNA barcode from 43 agricultural field soil nematodes. These nematodes were assigned to 13 rDNA-derived OTUs, but in the COI gene analysis were assigned to 23 COI gene-derived OTUs (cOTUs), indicating that COI gene-based barcoding may provide higher taxonomic resolution than conventional SSU rDNA-barcoding in soil nematode community analysis.     

The Effects of Soil Salinity and Meloidogyne javanica on Tomato

A non-sodic, non-saline sandy loam soil was salinized to anion-cation ratios similar to those naturally occurring in Iraq and California. The interactions of saline soils (conductivities 4, 8, 12 and 16 mmhos/cm) with a moderately salt-tolerant plant (Lycopersicon esculentum ''Marimond'') and a plant parasitic nematode (Meloidogyne javanica) were investigated. Plant parasitic nematodes were shown to be an important modifying influence within the plant environment, either accentuating or ameliorating salinity stress effects.     

Do plant parasitic nematodes have differential effects on the productivity of a fast- and a slow-growing grass species?

We have examined the interaction between plant parasitic nematodes and plant species from different stages of grassland succession. In these grasslands, fertiliser application was stopped in order to restore the former nutrient-poor ecosystems. This management resulted in a reversed succession of high- to low-productivity. Nematodes isolated from a high-productive early-successional field and a low-productive late-successional field were inoculated to sterilised soil planted with seedlings of either Lolium perenne (a fast-growing early-successional species) or Festuca rubra (a slow-growing late-successional species). The experiment was performed at low and high supply rates of nutrients. We hypothesised that at a low nutrient supply rate the growth of L. perenne will be more reduced by nematode herbivory than the growth of F. rubra. Furthermore, we hypothesised that higher numbers of plant parasitic nematodes will develop under L. perenne. We found no support for our first hypothesis, because nematodes did not affect plant growth. Our results suggest that changes in the nutrient availability rather than plant parasitic nematodes affect plant succession in impoverished grasslands. On the other hand, plant species and nutrient supply rate significantly affected the density and composition of the plant parasitic nematode community. In line with our second hypothesis, plant parasitic nematodes reproduced better on the fast-growing L. perenne than on the slow-growing F. rubra. Our results, therefore, suggest that the succession of the plant parasitic nematode community is probably more affected by changes in the plant community than the other way round.     

Molecular diversity of bacterial endosymbionts associated with dagger nematodes of the genus Xiphinema (Nematoda: Longidoridae) reveals a high degree of phylogenetic congruence with their host

Bacterial endosymbionts have been detected in some groups of plant‐parasitic nematodes, but few cases have been reported compared to other groups in the phylum Nematoda, such as animal‐parasitic or free‐living nematodes. This study was performed on a wide variety of plant‐parasitic nematode families and species from different host plants and nematode populations. A total of 124 nematode populations (previously identified morphologically and molecularly) were screened for the presence of potential bacterial endosymbionts using the partial 16S rRNA gene and fluorescence in situ hybridization (FISH) and confocal microscopy. Potential bacterial endosymbionts were only detected in nematode species belonging to the genus Xiphinema and specifically in the X. americanum group. Fifty‐seven partial 16S rRNA sequences were obtained from bacterial endosymbionts in this study. One group of sequences was closely related to the genus ‘Candidatus Xiphinematobacter’ (19 bacterial endosymbiont sequences were associated with seven nematode host species, including two that have already been described and three unknown bacterial endosymbionts). The second bacterial endosymbiont group (38 bacterial endosymbiont sequences associated with six nematode species) was related to the family Burkholderiaceae, which includes fungal and soil–plant bacterial endosymbionts. These endosymbionts were reported for the first time in the phylum Nematoda. Our findings suggest that there is a highly specific symbiotic relationship between nematode host and bacterial endosymbionts. Overall, these results were corroborated by a phylogeny of nematode host and bacterial endosymbionts that suggested that there was a high degree of phylogenetic congruence and long‐term evolutionary persistence between hosts and endosymbionts.     

Molecular and culture-based analyses of prokaryotic communities from an agricultural soil and the burrows and casts of the earthworm Lumbricus rubellus

The microbial populations in no-till agricultural soil and casts of the earthworm Lumbricus rubellus were examined by culturing and molecular methods. Clone libraries of the 16S rRNA genes were prepared from DNA isolated directly from the soil and earthworm casts. Although no single phylum dominated the soil library of 95 clones, the largest numbers of clones were from Acidobacteria (14%), Cytophagales (13%), Chloroflexi (8%), and gamma-Proteobacteria (8%). While the cast clone library of 102 clones was similar to the soil library, the abundances of several taxa were different. Representatives of the Pseudomonas genus as well as the Actinobacteria and Firmicutes increased in number, and one group of unclassified organisms found in the soil library was absent in the cast library. Likewise, soil and cast archaeal 16S rRNA gene libraries were similar, although the abundances of some groups were different. Two hundred and thirty aerobic bacteria were also isolated on general heterotrophic media from casts, burrows, and soil. The cast isolates were both phenotypically and genotypically different from the soil isolates. The cast isolates were more likely to reduce nitrate, grow on acetate and Casamino Acids, and utilize fewer sugars than the soil isolates. On the basis of their ribotypes, the cast isolates were dominated by Aeromonas spp. (28%), which were not found in the soil isolates, and other gamma-Proteobacteria (49%). In contrast, the soil isolates were mostly Actinobacteria (53%), Firmicutes (16%), and gamma-Proteobacteria (19%). Isolates obtained from the sides of earthworm burrows were not different from the soil isolates. Diversity indices for the collections of isolates as well as rRNA gene libraries indicated that the species richness and evenness were decreased in the casts from their levels in the soil. These results were consistent with a model where a large portion of the microbial population in soil passes through the gastrointestinal tract of the earthworm unchanged while representatives of some phyla increase in abundance.     

The role of parasite release in invasion of the USA by European slugs

Several species of European slugs are invasive in the USA, threatening native species and damaging agricultural and horticultural crops. One possible explanation for the success of these invaders is parasite release. To test this hypothesis we collected European slugs in part of their native range (United Kingdom) and in the USA and compared prevalence, distribution and species richness of their nematode parasites. All slugs were dissected and examined for the presence of nematodes. In the UK, nematodes were present at 93% of study sites and 16.4% of all slugs examined were associated with nematodes whereas in the USA the respective figures were 34% of sites and 5.4% of slugs. Nematode species richness was greater in the UK with 12 species being found, seven of which were thought to be truly parasitic as opposed to being phoretic or necromenic. Nine species of nematode were found in the USA, four of which were truly parasitic. Four of the ten European slug species examined in the USA, were entirely free of truly parasitic nematodes whereas all were infected by nematodes in some sites in the UK. There was a significant difference in the prevalence of truly parasitic nematodes in five of these species when comparing their home versus invasive range. A significant difference in parasite prevalence was observed when comparing native and introduced slug species in the USA, however, this was not significant in the UK. Our data support a role for parasite release during the invasion of the USA by European slugs.     

Delivery of macromolecules to plant parasitic nematodes using a tobacco rattle virus vector

Plant parasitic nematodes cause significant damage to crops on a worldwide scale. These nematodes are often soil dwelling but rely on plants for food and to sustain them during reproduction. Complex interactions occur between plants and nematodes during the nematode life cycle with plant roots developing specialized feeding structures through which nematodes withdraw nutrients. Here we describe a novel method for delivering macromolecules to feeding nematodes using a virus-based vector [tobacco rattle virus (TRV)]. We show that the parasitic nematode Heterodera schachtii will ingest fluorescent proteins transiently expressed in plant roots infected with a TRV construct carrying the appropriate protein sequence. A prerequisite for this delivery is the presence of replicating virus in root tips prior to the formation of nematode-induced syncytia. We show also that TRV vectors expressing nematode gene sequences can be used to induce RNAi in the feeding nematodes.     

Migratory plant-parasitic nematodes associated with cereals in Scotland

Ninety-eight soil samples were collected from cereal farms on mainland Scotland and examined for migratory plant-parasitic and predatory nematodes. Soil analysis was also carried out on each sample. Of the 25 parasitic nematode species identified Longidorus elongatus was the most frequently found and while populations were generally low, populations great enough to cause damage were detected. Populations of other plant-parasites were usually low. Individual species were unlikely to cause economic losses but their combined effect could possibly reduce yields. Analysis of the relationships between nematode species suggested that neither competition nor predation were important factors limiting population size. With few exceptions, none of the soil factors measured influenced the occurrence or abundance of the nematodes identified.     

藏东南急尖长苞冷杉林林隙土壤线虫群落特征

为了解西藏东南部急尖长苞冷杉林林隙土壤线虫群落特征,对林隙、非林隙土壤0~30 cm范围内不同深度土层的线虫群落进行调查,并用线虫个体密度、生物多样性指数和营养类群指数等特征值分析了土壤线虫群落的结构及多样性特点.结果表明： 采用浅盘法分离得到土壤线虫26801 条,隶属于2纲5目40科64属;线虫个体密度平均为3552 条·100 g-1干土,表聚性极强.垫咽属、丝垫刃属为林隙土壤线虫优势属;食细菌性线虫为主要营养类群.土壤有机质的分解兼有真菌分解和细菌分解两种途径.线虫的生物多样性及丰富度与林隙面积有关.土壤线虫群落特征表明,林隙具有异于郁闭林分和林间空地的特性,在环境指示方面具有应用潜力.     

Granite Rock Outcrops: An Extreme Environment for Soil Nematodes?

We studied soil nematode communities from the surface of granite flatrock outcrops in the eastern Piedmont region of the United States. The thin soils that develop here experience high light intensity and extreme fluctuations in temperature and moisture and host unique plant communities. We collected soils from outcrop microsites in Virginia (VA) and North Carolina (NC) in various stages of succession (Primitive, Minimal, and Mature) and compared soil properties and nematode communities to those of adjacent forest soils. Nematodes were present in most outcrop soils, with densities comparable to forest soils (P > 0.05). Nematode communities in Mature and Minimal soils had lower species richness than forest soils (P < 0.05) and contained more bacterial-feeders and fewer fungal-feeders (P < 0.05). Primitive soils contained either no nematodes (NC) or only a single species (Mesodorylaimus sp., VA). Nematode communities were similar between Mature and Minimal soils, according to trophic group representation, MI, PPI, EI, SI, and CI (P > 0.05). Forest soils had a higher PPI value (P < 0.05), but otherwise community indices were similar to outcrop soils (P > 0.05). Outcrop nematode communities failed to group together in a Bray-Curtis cluster analysis, indicating higher variability in community structure than the Forest soils, which did cluster together. A high proportion of the nematodes were extracted from outcrop soils in coiled form (33-89%), indicating that they used anhydrobiosis to persist in this unique environment.     

江苏省不同种植年限的西/甜瓜农田土壤线虫群落特征

在江苏省主要西/甜瓜产区,采集不同种植年限的西/甜瓜农田土壤,采用线虫形态学的鉴定方法,分析了不同种植年限西/甜瓜农田土壤线虫的数量、群落结构及相关生态指数的变化。结果表明:所有调查样点中,共发现土壤线虫54属,平均682条·100 g^-1干土,其中原杆属线虫、拟丽突属线虫和矮化属线虫在不同种植年限的西/甜瓜农田土壤中均为优势属;随着种植年限的增加,土壤线虫的总数量虽无显著变化,但其中食细菌类线虫和杂食-捕食类线虫数量显著减少,而植物寄生类线虫数量显著增加;植物寄生类线虫中的矮化属、根结属线虫数量不断增加;随着种植年限的增加,土壤线虫的多样性指数(H)、瓦斯乐斯卡指数(WI)、自由生活线虫成熟度指数(MI)显著降低,线虫通道指数(NCR)、植物寄生线虫成熟度指数(PPI)、PPI/MI显著升高,其他指数无显著变化;相关性分析结果显示,土壤线虫数量与土壤pH、速效磷、有效锌和有效硼均呈显著正相关;土壤p H、有机质、全氮、速效钾、有效硼和有效锌对土壤优势属线虫的影响较大;连续种植西/甜瓜会增加土壤植物寄生类线虫数量,进而可能造成西/甜瓜连作障碍的发生;连续种植6～1...     

新疆长期棉花连作对土壤理化性状与线虫群落的影响

土壤线虫群落特征是评价和指示土壤生态系统健康状况的重要依据。本研究选取不同连作年限(5、10、15、20和25年)的棉田为样地,采用高通量测序技术,探究土壤性状和线虫群落对棉田长期连作的响应。结果表明: 棉田连作10～15年后,土壤pH、电导率显著升高,有机碳、全氮、有效磷、有效钾、硝态氮含量和土壤微生物生物量碳(MBC)显著降低。在连作棉田中共鉴定出土壤线虫3纲7目18科25属,其中螺旋属在不同连作年限的棉田土壤中均为优势属;土壤植物寄生类线虫在不同连作年限中均为优势营养类群,呈现先降低后增加的趋势,连作25年较其他连作年限植物寄生类线虫增加9.1%～208.6%,其中螺旋属线虫增加了392%。随着连作年限的增加,矮化属、茎属、Discopersicus、中环属和中轮属等植物寄生类线虫被检出。连作15年的棉田土壤中,土壤线虫丰富度指数和自由生活线虫成熟度指数显著降低,植物寄生线虫成熟度指数/自由生活线虫成熟度指数显著升高,Shannon多样性指数和瓦斯乐斯卡指数最低;有效磷和MBC是影响土壤线虫群落变化的主要环境因子。这说明棉田连作10～15年会发生土壤养分失衡,土壤线虫多样性降低,土壤食物网稳定性变差,棉花致病类植物寄生线虫增加,产生连作障碍。     

Quantification of the slug parasitic nematode Phasmarhabditis hermaphrodita from soil samples using real time qPCR

Phasmarhabditis hermaphrodita is a nematode parasite that infects and kills several species of slugs. The nematode is produced commercially as a biological control agent for slug pests of agriculture and horticulture. Given the difficulties of distinguishing this species from other nematode species in soil samples, very little is known about its natural ecology or its behaviour and persistence following application for biological control. Here we describe a method to quantify P. hermaphrodita in soil samples based on real time PCR. We designed primers and a dual labelled fluorescent probe that can be used to quantify numbers of P. hermaphrodita and which is capable of distinguishing this species from the morphologically identical Phasmarhabditis neopapillosa. We compared different methods whereby the entire nematode community is extracted prior to DNA extraction, and three methods to extract DNA directly from soil samples. Both nematode extraction and DNA extraction from large (10 g) samples of soil gave reliable estimates of nematode numbers, but methods which extracted DNA from small (1 g or less) soil samples substantially underestimated numbers. However, direct extraction of DNA from soils may overestimate numbers of live nematodes as DNA from dead nematodes was found to persist in soil for at least 6 days. The technique could be modified for detection and quantification of all soil borne parasitic nematodes.     

Above- and belowground insect herbivores differentially affect soil nematode communities in species-rich plant communities

Interactions between above‐ and belowground invertebrate herbivores alter plant diversity, however, little is known on how these effects may influence higher trophic level organisms belowground. Here we explore whether above‐ and belowground invertebrate herbivores which alter plant community diversity and biomass, in turn affect soil nematode communities. We test the hypotheses that insect herbivores 1) alter soil nematode diversity, 2) stimulate bacterial‐feeding and 3) reduce plant‐feeding nematode abundances. In a full factorial outdoor mesocosm experiment we introduced grasshoppers (aboveground herbivores), wireworms (belowground herbivores) and a diverse soil nematode community to species‐rich model plant communities. After two years, insect herbivore effects on nematode diversity and on abundance of herbivorous, bacterivorous, fungivorous and omni‐carnivorous nematodes were evaluated in relation to plant community composition. Wireworms did not affect nematode diversity despite enhanced plant diversity, while grasshoppers, which did not affect plant diversity, reduced nematode diversity. Although grasshoppers and wireworms caused contrasting shifts in plant species dominance, they did not affect abundances of decomposer nematodes at any trophic level. Primary consumer nematodes were, however, strongly promoted by wireworms, while community root biomass was not altered by the insect herbivores. Overall, interaction effects of wireworms and grasshoppers on the soil nematodes were not observed, and we found no support for bottom‐up control of the nematodes. However, our results show that above‐ and belowground insect herbivores may facilitate root‐feeding rather than decomposer nematodes and that this facilitation appears to be driven by shifts in plant species composition. Moreover, the addition of nematodes strongly suppressed shoot biomass of several forb species and reduced grasshopper abundance. Thus, our results suggest that nematode feedback effects on plant community composition, due to plant and herbivore parasitism, may strongly depend on the presence of insect herbivores.     

Effects of native and exotic range‐expanding plant species on taxonomic and functional composition of nematodes in the soil food web

Due to climate warming, many plant species shift ranges towards higher latitudes. Plants can disperse faster than most soil biota, however, little is known about how range‐expanding plants in the new range will establish interactions with the resident soil food web. In this paper we examine how the soil nematode community from the new range responds to range‐expanding plant species compared to related natives. We focused on nematodes, because they are important components in various trophic levels of the soil food web, some feeding on plant roots, others on microbes or on invertebrates. We expected that range expanding plant species have fewer root‐feeding nematodes, as predicted by enemy release hypothesis. We therefore expected that range expanders affect the taxonomic and functional composition of the nematode community, but that these effects would diminish with increasing trophic position of nematodes in the soil food web. We exposed six range expanders (including three intercontinental exotics) and nine related native plant species to soil from the invaded range and show that range expanders on average had fewer root‐feeding nematodes per unit root biomass than related natives. The range expanders showed resistance against rather than tolerance for root‐feeding nematodes from the new range. On the other hand, the overall taxonomic and functional nematode community composition was influenced by plant species rather than by plant origin. The plant identity effects declined with trophic position of nematodes in the soil food web, as plant feeders were influenced more than other feeding guilds. We conclude that range‐expanding plant species can have fewer root‐feeding nematodes per unit root biomass than related natives, but that the taxonomic and functional nematode community composition is determined more by plant identity than by plant origin. Plant species identity effects decreased with trophic position of nematodes in the soil food web.