2.1 Study site and soil sampling
Soil was collected from experimental rice fields located in Changsha,
Hunan, China (113°19′ 52″ E, 28° 33′ 04″ N). The site is characterized
by a subtropical climate, with a mean annual temperature of 16.7 °C and
mean annual precipitation of 1457 mm. Moist field soils were collected
from the plow layer (0–20 cm) using a stainless-steel drill (diameter:
5 cm) in September, which is after the growing season in summer. The
soil is derived from Quaternary Red Clay and classified as Haplic
Acrisol according to the IUSS Working Group WRB (WRB, 2015). The soil
received urea (80 kg ha-1 yr-1)
along with biological fertilizer (livestock manure) and rice plant straw
residues (6000 kg ha-1 yr-1) within
a rice–rape rotation. Five soil cores were collected and thoroughly
mixed; they were immediately placed in a gas-permeable plastic bag, and
stored at 4 °C until analysis. A subset of soil samples was air-dried
and passed through a 2-mm sieve to remove fine roots and other plant
residues. The fresh soil samples were used to measure basic chemical
properties such as SOC (11.27 g kg−1) and total N (TN)
(1.3 g kg−1) using the potassium
dichromate-concentrated sulfuric acid-phenanthroline titration method
(Lu, 1999). Dissolved organic C (DOC) (69.84 mg kg−1)
was analyzed using a Shimadzu TOC-VWP analyzer (TOC-VWP, Shimadzu,
Japan). Soil pH (4.98) was measured using a Mettler Toledo 320 pH meter
at a soil-to-water ratio (w:v) of 1:2.5.