Abstract
Nematodes are abundant metazoans that play crucial roles in nutrient recycle in the pedosphere. Although high-throughput amplicon sequencing is a powerful tool for the taxonomic profiling of soil nematodes, polymerase chain reaction (PCR) primers for amplification of the 18S ribosomal RNA (SSU) gene and preparation of template DNAs have not been sufficiently evaluated. We investigated nematode community structure in copse soil using four nematode-specific (regions 1–4) and two universal (regions U1 and U2) primer sets for the SSU gene regions with two DNAs prepared from copse-derived mixed nematodes and whole soil. The major nematode-derived sequence variants (SVs) identified in each region was detected in both template DNAs. Order level taxonomy and feeding type of identified nematode-derived SVs were distantly related between the two DNA preparations, and the region U2 was closely related to region 4 in the non-metric multidimensional scaling (NMDS) based on Bray-Curtis dissimilarity. Thus, the universal primers for region U2 could be used to analyze soil nematode communities. We further applied this method to analyze the nematodes living in two sampling sites of a sweet potato-cultivated field, where the plants were differently growing. The structure of nematode-derived SVs from the two sites was distantly related in the principal coordinate analysis (PCoA) with weighted unifrac distances, suggesting their distinct soil environments. The resultant ecophysiological status of the nematode communities in the copse and field on the basis of feeding behavior and maturity indices was fairly consistent with those of the copse- and the cultivated house garden-derived nematodes in prior studies. These findings will be useful for the DNA metabarcoding of soil eukaryotes, including nematodes, using soil DNAs.
Highlights
soil DNA (Soil) ecosystem is composed of microbiota and soil [1]
Hundreds of sequence variants (SVs) were identified by Illumina MiSeqassisted amplicon sequencing (Table 2): the largest (883) and smallest (338) numbers of SVs from the nematode and soil DNAs were found in regions 1 and 3, respectively
This study investigated nematode-derived SVs identified from complex nematode genomic DNAs and whole soil DNAs prepared from copse soils by using high-throughput amplicon sequencing with four nematode-specific and two universal primer sets for the 18S ribosomal RNA (SSU) gene
Summary
Soil ecosystem is composed of microbiota and soil [1]. Soil biota is influenced by soil environments because soils are heterologous with different chemical and physical properties, including pH, nutrients, and water contents. Nematodes are abundant animals with various species and feeding habitats and are widely distributed in freshwater, marine, and terrestrial environments [4,5,6,7] They play a crucial role in soil nutrient recycling [8, 9] and occupy the ecological position in the pedosphere comparable to that of planktons in the hydrosphere. Nematode taxonomic compositions are influenced by ecosystem type and various factors, such as food availability and abundance, physical, and chemical parameters (e.g., pH, temperature) [8, 9], soil properties [9, 10], latitude [11, 12], and agricultural conditions (e.g., tillage, cultivated plants, fertilizers) [13,14,15,16,17,18,19,20,21,22]. The taxonomic composition and abundance of nematodes have been used as an indicator of biological, environmental, and toxicological conditions in soils [4, 23, 24]
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