**4. Discussion**

Our results support the hypothesis that natural birch regeneration has a positive effect on the soil organic carbon accumulation. The tendency to increase carbon stocks was observed in the studied chronosequence of the birch tree stands. Several studies have shown that the decomposition of soil organic matter exceeded the input of organic matter from the trees in the initial following afforestation [26,27]. In the younger stands, the soil organic carbon accumulation was the greatest in the surface layer. In the 5-cm layer, accumulation accounted for about 30% of the total accumulation in the 50-cm deep soil column. Conversely, carbon accumulation was considerably lower than in the deeper soil layers in the 5-cm layer of the soils from the older stands. In the group I soils, accumulation in the deeper layers constituted 40% of the total stock of carbon, while it accounted for 60% in group IV. This increase in C accumulation in the deeper layers of the soils is associated not only with the processes of transporting dissolved organic compounds downwards, but also with an effect of supplying organic debris from the root systems. This is confirmed by the increase in root biomass in particular groups of stands. Subsoil soil organic carbon (SOC) storage may be promoted by the translocation of OM into deeper soil layers as DOC with the percolating water and due to bioturbation by soil animals [28]. Kotroczó et al. [29] found that plants cause greater changes in soil properties through their roots and secretions than via litter. In this sense, aboveground OM only probably has limited effects on SOM levels when compared to belowground OM [28]. Roots are a key component of the belowground part of the forest ecosystem, constituting the basic source of SOM that significantly affects soil microbiological activity [30,31]. Over time, soil organic matter input increases with the productivity of the forest stands, and the soils switch from being a C source to a C sink [32]. According to Laganiere et al. [33], the positive impact of afforestation on soil organic carbon stock is more pronounced in the cropland soils than in pastures or natural grasslands. Afforestation usually results in the establishment of higher plant biomass, and trees modify the quality and quantity of litter inputs and microclimatic conditions, such as moisture and temperature. Deng and Shangguang [34] highlight the importance of previous land use, tree species, soil depth, and forest age in determining soil C and N changes in a range of environments

and land use transitions. In our study, birch stands, through aboveground and belowground biomass accumulation, had a positive e ffect on the quality of SOM, as expressed by the C/N ratio, which is an indicator of the extent of plant nitrogen being made available to plant residues. Li et al. [35] state that land use changes from agricultural areas to forest alter the ratios between soil C, N, and P. Springob and Krichmann [36] found that a soil C/N ratio of >20 could limit SOM mineralisation. According to Cools et al. [37], tree species are the main factor in explaining the variability of the C/N ratio. The content of better decomposed soil organic matter increases with stand age. At the same time, soil acidity and nutrient uptake increase with tree growth. Riqueiro-Rodríguez [38] note that the Pinus radiate more drastically decreases the soil pH than *Betula alba*. In another study, the acidifying a ffect of afforestation on mineral soil has been confirmed by a significant decrease in soil pH in the 0–5-cm layer and by a slightly weaker decrease in the 5–15-cm layer [39].

The results of the cluster analysis confirmed the distinctness in terms of C content and enzymatic activity of the soils of younger stands when compared to the soils of older stands. The soil parameters pH and soil organic carbon are important factors that shape dehydrogenase activity [15,40]. The highest pH, with the highest alkaline cation content, resulted in the highest dehydrogenase activity in soils of the younger stands (Groups I and II), reflecting the previous agricultural use of the soils and the associated intensive fertilisation and liming. According to Rousk et al. [41], pH is the main determinant of the structure of soil microbial populations. Soil pH directly determines plant growth, nutrient absorption, and the intensity of biological and chemical processes in the soil. In this work, dehydrogenase activity was positively associated with exchangeable Ca, K, and Mg contents, with a higher content of basic ions leading to an increase in pH, which results in the stimulation of soil microorganisms. Soil pH may be the major factor controlling the biomass and composition of microbial communities and their maintenance demand [42]. When assessing the properties of soils that were subjected to long-term agricultural use, several authors have considered the high plant-nutrient content as evidence of systematic fertilisation [43]. For example, Ren et al. [44] have noted that catalase, saccharase, urease, and alkaline phosphatase were significantly increased by land-use conversion from farmland to forest. According to this, significant correlations between soil enzyme activities and soil properties indicate that the soil enzyme activities are closely related to soil nutrients dynamics [18]. Dehydrogenase activity di ffered among the soils of the studied birch stands. The activity of this enzyme reflects that changes in the soil that are associated with the growth of the birch stands. According to previous studies, enzymatic activity is strongly stimulated by SOM [15,18], and processes that are related to organic matter transformations are carried out with the participation of soil microorganisms and their enzymes [45]. In our study, no direct relationship between dehydrogenase activity and carbon accumulation was found. Dehydrogenase activity was high in the soil of the younges<sup>t</sup> stands (first age class), and subsequently considerably decreased in class II. Our results indicate a trend to increased dehydrogenase activity in the soils of the oldest stands (IV group of stands). The highest pH, with the highest alkaline cation content, resulted in the highest dehydrogenase activity in soils of the younger stands, reflecting the previous agricultural use of the soils and the associated intensive fertilization and liming. The e ffects of fertilization disappear in the following years of tree stand growth. Forest stands grow and provide more litter fall to the soil, which stimulates the dehydrogenase activity. With age, greater amounts of carbon were accumulated in the surface soil layers. With increased litter input and in the absence of soil cultivation, conversion from cropland to forest could result in increased SOM stocks [46]. Similarly, Kara et al. [47] and Kang et al. [48] sugges<sup>t</sup> that long-term a fforestation could significantly enhance SOM contents, accumulate microbial biomass, and improve potential enzyme activities.
