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 11 / 15 was consociated with white clover or when a more vigorous rootstock was utilized [38]. However, the negative effects of this orchard floor management practice can be mitigated by delaying the introduction of living mulch for several years after the trees were planted [39,40]. The living mulch mixtures induced also a lower level of K in soil and in the apple tree leaves, which have likely affected tree growth and the yield potential [41,42]. However, it should also be considered that a persistent high soil nitrate nitro- gen level could have an effect on the balance between vegetative and reproductive growth [43]. The fluctuations in soil Mg and Ca content did not always correspond to changes observed in the leaves Nevertheless, the changes of the macroelements levels in the leaves, even if statistically significant, still kept their content within the optimal or high content according to Polish standards for apple trees [44]. It is noteworthy that the evaluation of the living mulch mixtures impact considering all nutrients content allowed to discriminate between the two living mulches and the control, pointing to an effect that would comprehensively integrate the physiological role of each nutrient. The introduction of mixtures of flowering plants in orchards (flower strips) is commonly considered a practice aiming at promoting functional agrobiodiversity [45], which generally is carried out in the alleyways or hedgerows of the orchard. In

the present study we wanted to test the possibility of exploiting flower strips also for row management, which could also improve their expected performance on functional biodiversity [7]. Mix2, a living mulch mixture designed as a flower strip, had a limited negative impact on soil and tree nutrients status and tree growth compared to both control and the classical leguminous-grass living mulch (Mix1). Even though the performance of Mix2 on pest control or pollinators was not eval- uated in the present study, the obtained results suggest that such approach could be a sustainable method for tree row management from the agronomical point of view, which could provide a variety of ecosystem services, without impairing the main crop performance. The two living mulch mixtures generally modified soil bacterial activity and biodiversity and consequently the bac- terial microbiome capacity of exploiting various growing substrates and C-sources. Such impact has been observed also in other studies on apple orchards [46,47]. Living mulches based only on white clover or a grass significantly modified various soil enzymatic activities (e.g., invertase, urease or alkaline phosphatase), and increased the soil bacterial carbon metabolic activity and the bacterial community diversity [48]. It is noteworthy that both living mulch mixtures induced significant fluctuations in the potential activity of soil bacterial population towards phenolic com- pounds compared to control. Both mixtures seemed to impact weed diversity (visual observations), and the proportion

between monocotyledons and dicotyledons, similarly to other reports [49–52]. Considering that the concentration of phenolic acids in soil can range from 2.1% to 4.4% for the roots of monocotyledonous plants and from 0.1% to 0.6% for the roots of dicotyledonous plants [53], the different composition of the living mulches in this respect would account for the modifications of the bacterial population and its metabolic activity. Furthermore, the soil microbiome associated with another clover, Trifolium pratense, was found to include microorganisms belonging to the genera Sphingobium and Novosphingobium [54], which have a wide range of metabolic capabilities, including the degradation of phenolic compounds [55,56]. The soil microbiome associated with Mix1 showed a greater capacity to utilise glucose-1-phosphate and D-xylose in various periods of the vegetative season than the control, which may indicate the improved use of lignin or hemicellulo- ses derived from the living mulch plant residues as a carbon source [57]. Interestingly, the Mix2 soil bacterial microbiome showed similar properties only in autumn. The bacteria populations associated with the two living mulch mixtures differed in the activity towards various compounds that have been reported as plant root exudates (e.g., L-asparagine, L-serine, L-arginine, phenylethylamine, D-malic acid or 4-hydroxy benzoic acid, D-xylose, β-methyl-D-glucoside) of some species present in the mixtures, including clover [58–60]. Even though an unambiguous understanding of impact of the tested

living mulches on the soil bacterial communities and their C-sources use requires more thorough research [61], it appears evident that the introduced living mulch mixtures, can significantly modify the activity and biodiversity of the soil microbi- ome in addition to the impact on above-ground biodiversity.