## 3 Results and Discussion
### Biological Characterisation of Biosolids
However, this increase in URS activity in agricultural soils with the addition of biosolids in the short-term applications could also be attributed to the fact that biosolids provide both labile organic matter and microorganisms with active enzymatic machinery ready for N transformation. Hence, the importance of evaluating URS activity in biosolids before application to soil becomes clear, especially since biosolids have been presented as an organic amendment with an important content of nitrogen compounds, such as amides, peptides and proteins, as observed in studies and analysis of FTIR spectra (Figures 2 and 3). Determining this enzymatic activity over time and in biosolids can be a usable indicator of quality and functionality to ensure the biotransformation of nitrogen-rich organic matter in soils prior to application. Other biological parameters related to soil quality and functionality that have been used in assessing the effect of organic amendments such as biosolids on agricultural soils are the metabolic quotient (qCO2) and specific hydrolytic activity (qFDA) [47; 66; 130]. The qCO2 expresses the amount of CO2 produced per unit of biomass and time. This indicator has been used as an indicator of the physiological status of microorganisms in biosolids [66; 131]. The qCO2 has also been used to detect alterations or stress of MB in the soil or in organic amendments due to disturbances or environmental changes [66; 67; 63]. The qCO2 obtained in the biosolids analysed in this study for each time of year were 157.95, 124.31, 115.42, and 270.19 mg C-CO2 g-1 dry biosolids d-1 for spring, summer, autumn and winter, respectively. Winter qCO2 was significantly different (F = 41.951, \(p\) = 0.000) from the rest of the year. Therefore, it can be attributed that during the winter the biosolids presented an alteration that allowed them to increase their respiration rate (Table 3). This increase could be possibly due to a lower MB content or climatic conditions, as already mentioned. The lower MB content or climatic conditions can propitiate a structural change of the microbial community [113]. Although there are no reported data in the literature on qCO\({}_{2}\) for biosolids per se, [PERSON] et al. [66] established that qCO\({}_{2}\) in soils amended with biosolids presented a greater impact as the application rate of biosolids to the soil increased, thus confirming that qCO\({}_{2}\) can emphasise the effects of biosolids on soil properties. [PERSON] et al. [131] in a field study on the application of biosolids to agricultural soils in the long term (\(>\)20 years) found that qCO\({}_{2}\) was lower in soil with application of biosolids than control soil without biosolids, thus concluding that there was an improvement in the microbial metabolism of C as a result of stress relief in soil microorganisms, due also to an increase in C sequestration in soils with biosolids. Thus, they expanded the concept of qCO\({}_{2}\) to assess microbial stress relief and restoration of microbial physiology in agricultural soils with biosolids amendment. [PERSON] et al. [72] evaluated the application of 2 types of biosolids: aerobic thermophilic (AT) and anaerobic mesophilic (AM) to an agricultural soil to establish if there are differences of these biosolids according to the stabilisation processes with which they were obtained and found that qCO\({}_{2}\) was higher in the soil with biosolids AM both with respect to the control soil (without amendment), as well as in the soil with biosolids AT. They concluded that AT biosolids produced minor changes in soil microbiota and improved soil microbial activity. Therefore, it is important to monitor these parameters in biosolids before their application to agricultural soils, to expect or not possible alterations to the soil microbiota and therefore to establish an adequate management of biosolids. The qFDA is an index established by [PERSON] et al. [30] and was based on the measurement of the global hydrolytic capacity of a specific microbial biomass.