Sheep Manure Nitrogen Substitution in Acidic Tea Plantation Soil: Gas Emissions and Nitrogen-Cycling Gene Responses During a 30-Day Incubation

Recycling manure nitrogen (N) can reduce mineral fertilizer inputs, but its effects on soil emissions require evaluation. A 30-day, plant-free incubation compared urea alone (N1), 15% (N2) and 30% (N3) substitution with matured sheep manure, and an unfertilized control (N0). Acidic tea plantation soil (pH 4.24; organic matter 49.4 g kg−1) and manure (pH 7.6) were incubated at 80% water-filled pore space. Cumulative molecular N2O over days 1–30 was 894.1 ± 15.7, 911.5 ± 18.4, 960.4 ± 6.0 and 900.6 ± 13.4 ng g−1 dry soil in N0–N3 (mean ± SE; n = 3). Differences from N1 were 48.9 ng g−1 for N2 (unadjusted 95% confidence interval, 2.8 to 95.1) and −10.9 ng g−1 for N3 (−57.0 to 35.3); neither was significant after Holm adjustment. Cumulative CO2 was higher in N3 (1478.0 μg g−1) than in N1 (950.6 μg g−1). Both substitution treatments increased DNA-normalized AOB amoA and nirS on day 30, after N2O fluxes had declined. AOA amoA and nirK lacked significant treatment × day interactions, and nosZ had no significant planned day-specific contrast. Exploratory SEM and Mantel analyses did not establish microbial drivers: no N2O predictor remained significant after categorical-day adjustment, and no Mantel association survived false-discovery-rate correction. The 30% substitution warrants field evaluation, but laboratory nonsignificance does not establish emission equivalence or agronomic benefit.