Field sampling and soil analysis
Soils were collected from each site from late July to early August,
which is within the plant growing season. At each site, at least one
typical forest was selected. In each forest with 3-5 plots, the upper 10
cm of mineral soil was sampled from 8–10 locations within a plot
(100–150 m2), coarsely sieved to 5.6 mm to minimize
disturbance, and gently combined to produce a composite sample. All soil
samples were transported with a low-temperature incubator with ice
(< 5 °C) to laboratory, where they were sieved to 2 mm, and
then homogenized. One part was stored at 4 °C for incubation and
microbial measurement, and the rest was air-dried for chemical
determination.
In this experiment, the initial SOC and the total N contents were
determined using an elemental analyzer. Labile organic C (LOC) which is
oxidized by 333 mM potassium permanganate was measured according to the
method described by Blair et al. (1995). Soil pH was measured with a pH
meter from a soil slurry with 1:2.5 ratio by volume of soil to distilled
water. The soil texture (sand, silt, and clay contents), mineral N,
total and available P, and exchangeable cations (K+,
Na+, Ca2+, Mg2+)
were determined according to the common methods described by Lu (2000).
Microbial biomass and community composition was determined through
phospholipid fatty acid (PLFA) analysis described by Bardgett et al.
(1996). Phospholipids were extracted from 1.5 g of soil fresh weight and
analyzed using an Agilent 6890 Gas Chromatograph. Gram-positive bacteria
were identified by the terminal and mid-chain branched fatty acids
(i15:0, a15:0, i16:0, i17:0, a17:0), and cyclopropyl saturated and
monosaturated fatty acids (16:1ω7c, cy-17:0, 18:1ω7c, 8cy-19:0) were
considered indicative of gram-negative bacteria (Rinnan & Bååth, 2009).
Fungi were the sum of 18:1ω9c, 18:1ω9t, and 18:2ω9,12c, and
actinomycetes were the sum of 10Me16:0, 10Me17:0 and 10Me18:0
(Moore-Kucera & Dick, 2008). The total PLFA concentration was
calculated from the identified PLFAs (15:0, 14:0, 16:1, 16:1ω5, 16:0,
17:1ω8, 7Me-17:0, br17:0, br18:0, 18:1ω5, 18:0, 19:1; and those listed
above). The ratios of fungal to bacterial (F:B) PLFA and gram-positive
to gram-negative (GP:GN) PLFA were taken to represent the relative
abundance metrics of these groups.