In-depth insight into the short-term effect of floor management practice on young apple trees development and soil microbial biodiversity and activity

PLOS One | https://doi.org/10.1371/journal.pone.0329979 August 7, 2025 8 / 15 increase of the bacterial metabolic potential towards polymers, compared to control. In the summer, increased metabolism of polymers was induced by Mix2 and of phenolic compounds by both living mulches, compared to the control. Interest- ingly, the most significant changes in the metabolic potential of the soil microbiomes were observed in autumn: for some C-sources the effect of the two living mulches mixtures was similar (e.g., reduced metabolic potential towards amino acids and carboxylic acids), but for others divergent (increased metabolism of carbohydrates, phenolic compounds and amines by Mix2 but decreased by Mix1) (S2 Table and Fig 3). Deepening the analysis of the impact of the two living mulch mixtures on the bacterial community C-source metabolism to individual compounds (Fig 4) allowed to distinguish a group of nine compounds that were hardly metabolized by micro- organisms in spring and autumn belonging to four different classes of C-sources (carbohydrates, amino acids, carboxylic acids, polymers), but 4 out of 9 were among the carbohydrates. One compound (2-hydroxybenzoic acid) was barely used by microorganisms. On the other hand, several amino acids (e.g., L-arginine, L asparagine, etc.) and few organic acids (e.g., L-Malic acid) or polymers compounds (e.g., Tween) were instead commonly utilised by all bacterial communities of the three soil environments throughout the whole vegetative season (Fig 4).

The detailed analysis of differences in bacterial activity induced by the two living mulch mixtures within each sampling point showed that Mix2 induced a higher modification of the C-sources use by the bacterial microbiome compared to the control (23 out of 30) than Mix1 (14 out of 30), and the C-sources profiles were very contrasting (Fig 5, details in S3 Table). However, any C-source resulted to be differentially metabolised by both mixtures and control across the whole vegetative season. Autumn was the period with the highest number of compounds with statistically significant changes (16 out 31 analysed C-sources), while in spring and summer a similar number of compounds resulted with changed activity (6 or 8 out of 31, respectively), though they were not the same C-sources, except for 4-hydroxy benzoic acid. On the other hand, a broader spectrum of compounds with significant changed activity was shared by the spring and summer samples with those of autumn (3 or 4 compounds respectively). Intriguingly, all carbohydrates C-sources (n = 10) showed significant differences in the bacterial microbiome metabolic potential for at least one sampling point. Fig 3. Comparison of the impact of two kinds of living mulch mixtures on bacterial activity (AWCD) towards certain C-sources classes during the vegetative season. Different letters show statistically significant differences between treatments within each C-source class and sampling point for p ≤ 0.05. https://doi.org/10.1371/journal.pone.0329979.g003