Land use is known to be one of the major factors driving soil microbial and physico‐chemical properties. Understanding its long‐term effect remains a major challenge in assessing current soil functioning. Here, soil microbial and physico‐chemical properties of recent and very recent forests (forests developed in 1958 vs. after 1958) were compared with those of ancient forests (present in 1860 and 1958) to assess the effect of Land Use Legacy (LUL). LUL effects were further analysed depending on contrasting (i) climate conditions (sub‐humid vs. humid Mediterranean climates) and (ii) seasons (winter and summer), to examine whether LUL modified microbial responses to different spatio‐temporal climate conditions. Microbial indicators (lignocellulolytic activities, basal respiration, and microbial biomass) and physico‐chemical properties (C and N contents, mineralogical analyses, pH and conductivity) were assessed. A strong effect of past agricultural practice (terrace cultivation) was observed in soils from very recent forests: reduced microbial biomass and activities as well as number of Quercus pubescens stems together with increased phosphorous content and pH. Interestingly, LUL effect did not affect microbial and physico‐chemical responses to seasonal contrasts (winter vs. summer). Microbial response to LUL was not influenced by climate while climate modified LUL effects on some physico‐chemical properties (CaCO3, Corg, and K content). Moreover, soil recovered “pristine” physico‐chemical and microbial functional properties after at least 60 years of reforestation.

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Second-generation ethanol process uses lignocellulose, but a pre-treatment is required to degrade lignin before saccharification. Biological pre-treatment, using phenoloxidases, represents a cheap and ecofriendly option. Mould contamination can be overcome by heat pre-conditioning of substrates. Pasteurisation can be a mild and sustainable option. A 5-level response-surface experimental design was performed to test the effect of different time and temperatures on lignocellulolytic activities and substrate dephenolisation. Substrate microbial communities were characterised via catabolic profiles to determine functional diversity changes after pasteurisation. Temperatures ranging from 70 to 75 C and time from 5 to 10 h led to the highest laccase activities and 30% of substrate dephenolisation was achieved while avoiding mould contamination. The per cent of O-Alkyl (polysaccharides) did not vary significantly, meaning that polysaccharides were not extensively consumed. Microbial communities were less catabolically diversified (H' ¼ 1.2) but more active (AWCD ¼ 0.9) and could degrade polysaccharides.