A novel metabarcoding strategy for studying nematode communities

Nematodes are most abundant soil metazoa. How, the ecology of this group of organism remains poorly elucidated, partly due to the lack of reliable and validated sequencing strategies. In this study, we tested four primer sets for amplicon sequencing: JB3/JB5, SSU_04F/SSU_22R, Nemf/18Sr2b from earlier studies; and MMSF/MMSR, a newly developed primer set from this study. In order to test these primer sets, we used 22 samples of individual nematode species, 20 mock communities, 20 soil samples, 20 mock communities in soil, and 4 root/rhizosphere soil samples. We successfully amplified the target regions (COI gene; V1-V2, V4-V8 region of 18S rRNA gene) from these 86 DNA samples and sequenced the amplicons on an Illumina MiSeq sequencing platform. We found that the MMSF/MMSR and Nemf/18Sr2b were very efficient in detecting soil nematode compared to JB and SSU primer sets based on annotation of sequence reads at genus and in some cases at species level. Therefore, these primer sets could be used to study nematode communities in agricultural environments.