Metagenomics of soil prokaryotes inhabiting subtropical maize-rice rotation agroecosystem maintained under long-term fertilizer regimes

Long-term ecological research (LTER; 1996-2019) on crop rotation was carried out at Wufeng research field (No. 35) affiliated to Taiwan Agricultural Research Institute (TARI), Taiwan. Maize (Zea mays L. cv. TN1; upland or non-flooded) and rice (Oryza sativa L. ssp. japonica cv. TN67; lowland or flooded) were rotated annually. Long-term treatments included no fertilizer (CK, blank), control full-strength chemical fertilizer (CF, N-P2O5-K2O; 140-80-80 and 120-60-60 kg ha-1 for maize and rice, respectively), swine manure compost (CP), and inorganic nitrogen-supplemented CP (CPN1 and CPN2), green manure (GMN1) and peat (PN1). Sesbania roxburghii Merr. and Trifolium alexandrinum L. were used as green manures for maize and rice, respectively. Experiments were performed at randomized complete block design; quadruplicate plots for each treatment were used. Surface soils (~10 cm depth) proximal to plants were sampled (using a stainless-steel soil sampling tube of core diameter 2 cm) from maize and rice fields during tillering and flowering stages of two consecutive years of crop rotation (22nd-23rd rotation; 2017-2019). Soils collected from five different locations of a plot were mixed to represent a replicate. Four such replicates of the same treatment were pooled to form a representative sample for DNA extraction. Soil prokaryotes were profiled through metagenomics targeted to the hypervariable V3-V4 region of the 16S rRNA gene.