Phosphorus (P) is a critical nutrient for plant growth, yet its uptake is often hindered by soil factors like clay minerals and metal oxides such as aluminum (Al), iron (Fe), and calcium (Ca), which bind P and limit its availability. Phosphate-solubilizing bacteria (PSB) have the unique ability to convert insoluble P into a soluble form, thereby fostering plant growth. This study aimed to assess the efficacy of inoculation of Bacillus megaterium B119 (rhizospheric) and B. subtilis B2084 (endophytic) via seed treatment in enhancing maize yield, grain P content, and enzyme activities across two distinct soil types in field conditions. Additionally, we investigated various mechanisms contributing to plant growth promotion, compatibility with commercial inoculants, and the maize root adhesion profile of these strains. During five crop seasons in two experimental areas in Brazil, Sete Lagoas-MG and Santo Antônio de Goiás-GO, single inoculations with either B119 or B2084 were implemented in three seasons, while a co-inoculation with both strains was applied in two seasons. All treatments received P fertilizer according to plot recommendations, except for control. Both the Bacillus strains exhibited plant growth-promoting properties relevant to P dynamics, including phosphate solubilization and mineralization, production of indole-3-acetic acid (IAA)-like molecules, siderophores, exopolysaccharides (EPS), biofilms, and phosphatases, with no antagonism observed with Azospirillum and Bradyrizhobium. Strain B2084 displayed superior maize root adhesion compared to B119. In field trials, single inoculations with either B119 or B2084 resulted in increased maize grain yield, with relative average productivities of 22 and 16% in Sete Lagoas and 6 and 3% in Santo Antônio de Goiás, respectively. Co-inoculation proved more effective, with an average yield increase of 24% in Sete Lagoas and 11% in Santo Antônio de Goiás compared to the non-inoculated control. Across all seasons, accumulated grain P content correlated with yield, and soil P availability in the rhizosphere increased after co-inoculation in Santo Antônio de Goiás. These findings complement previous research efforts and have led to the validation and registration of the first Brazilian inoculant formulated with Bacillus strains for maize, effectively enhancing and P grain content.

This study explored how different sugarcane vinasses influence the structure and composition of soil bacterial communities in two tropical Oxisols with contrasting textures. In a controlled microcosm experiment with sugarcane seedlings, two concentrations of three vinasse types were applied, and bacterial communities were monitored over 10, 30, and 60 days using T-RFLP and 16S rRNA gene sequencing. Across all treatments, vinasse application led to clear changes in bacterial community structure in both soils, regardless of the time point. Certain bacterial groups, such as Sphingobacteriia, Alphaproteobacteria, and Gammaproteobacteria, became more abundant—likely responding to increased carbon availability, higher pH, and greater soil moisture. At the same time, other groups declined, possibly due to excess nutrients like potassium and sulfur. Notably, these shifts occurred even when standard biochemical indicators suggested no major impact, highlighting the sensitivity of microbial community-level responses. These findings point to the importance of looking beyond traditional soil quality metrics when assessing the environmental effects of organic residue applications. Incorporating microbial indicators can offer a more nuanced understanding of how practices like vinasse reuse affect soil functioning in tropical agroecosystems.

The genus Bacillus includes several species recognized for their agricultural benefits, such as biological control, induction of plant defenses, and promotion of plant growth. This study aims to evaluate the agronomic potential of the phosphate-solubilizing bacterium Bacillus thuringiensis strain CNPMS B116 in maize cultivation. Field trials were conducted over four crop seasons in Sete Lagoas and Santo Antônio de Goiás, with treatments including B. thuringiensis B116, B. megaterium B119, B. subtilis B2084, or no inoculation, combined with triple superphosphate (TSP) or rock phosphate plus TSP (RP + TSP). The evaluated outcome included nutrient content (nitrogen, phosphorus, and potassium), grain yield, and phosphate use efficiency. The compatibility assay of B116 with elite strains (Azospirillum and Bradyrhizobium) was tested using the cross-streak method in vitro. In field trials, nitrogen, phosphorus, and potassium accumulation in maize grains increased, along with productivity across all fertilization treatments. At Sete Lagoas, maize yield increased by 30

The increasing demand for sustainable food production has driven a surge in the use and commercialization of biological inputs, including biofertilizers. In this context, biofertilizers offer potential benefits for nutrient use efficiency, crop yield and sustainability. However, inconsistent definition of the term “biofertilizer” and regulations, particularly in the USA, hinder market growth and consumer confidence. While the European Union, and countries like Brazil, India, and China have made progress in this area, the USA market, projected to exceed 1 billion by 2029, lacks clear guidelines for biofertilizer production and sale. The USA market is dominated by Rhizobium genus, Mycorrhizae fungi, and Azospirillum species and based products targeting various crops. Although there is a growing and promising market for the use of biofertilizers, there are still many challenges to overcome, and to fully realize the potential of biofertilizers, future research should focus on modes of action, specific claims, and robust regulations that must be established. • The term “biofertilizer” lacks a universally accepted definition • It is necessary establishing a national regulation for biofertilizers in the USA • The biofertilizer market is growing fast and the biggest one is in America

Bacterial inoculants have been used in agriculture to improve plant performance. However, laboratory and field requirements must be completed before a candidate can be employed as an inoculant. Therefore, this study aimed to evaluate the parameters for inoculant formulation and the potential of Bacillus subtilis (B70) and B. pumilus (B32) to improve phosphorus availability in maize (Zea mays L.) crops. In vitro experiments assessed the bacterial ability to solubilize and mineralize phosphate, their adherence to roots, and shelf life in cassava starch (CS), carboxymethyl cellulose (CMC), peat, and activated charcoal (AC) stored at 4 °C and room temperature for 6 months. A field experiment evaluated the effectiveness of strains to increase the P availability to plants growing with rock phosphate (RP) and a mixture of RP and triple superphosphate (TS) and their contribution to improving maize yield and P accumulation in grains. The B70 was outstanding in solubilizing RP and phytate mineralization and more stable in carriers and storage conditions than B32. However, root adherence was more noticeable in B32. Among carriers, AC was the most effective for preserving viable cell counts, closely similar to those of the initial inoculum of both strains. Maize productivity using the mixture RPTS was similar for B70 and B32. The best combination was B70 with RP, which improved the maize yield (6532 kg ha−1) and P accumulation in grains (15.95 kg ha−1). Our results indicated that the inoculant formulation with AC carrier and B70 is a feasible strategy for improving phosphorus mobilization in the soil and maize productivity.

Sugarcane vinasse is a liquid waste derived from ethanol production. In Brazil, large amounts of this waste are applied to soil as fertigation. In general, this management seems to be beneficial for soil fertility and for some biological parameters, though the published information about the effects of sugarcane vinasse on the soil biota is controversial and the results may vary according to the vinasses’ composition. In this study, we assessed the effects of different sugarcane vinasses on microbial growth and activity indicators to verify their influence on soil microbial metabolism. For this purpose, we used two vinasses from different distillery plants (VA and VB) and a vinasse from a laboratory production (VC). Increasing concentrations of these vinasses were amended on two tropical Oxisols, with 33.6% (RL) and 17.6% (RYL) of clay, in a microcosm experiment with sugarcane plants. Ten, 30 and 60 days after application, we assessed the effects of the vinasses on soil microbial biomass carbon (MBC), basal soil respiration (C–CO2), the metabolic quotient (qCO2) and dehydrogenase activity of the soils. We found an increase in MBC, C–CO2, qCO2, as well as in dehydrogenase activity with increasing vinasse concentrations in both soils, when compared to the control. These changes were attributed mainly to the addition of carbon sources (C) of the vinasses to the soils, which improves the general biological activity.

A batata (Solanum tuberosum) apresenta grande relevância alimentar e economica, devido ao fato de ser muito consumida pelos seres humanos. Esta cultura apresenta grande exigencia em todos os tipos de tratos culturais, principalmente sobre o manejo do solo para sua implantacao. Este trabalho tem por objetivo realizar um levantamento sobre o manejo de solo e adubacao da cultura da batata na regiao de Guarapuava, PR. Foram acompanhadas tres propriedades na regiao de Guarapuava, Parana, localizadas nos municipios de Guarapuava, Candoi e Foz do Jordao, foi realizado o levantamento das caracteristicas quimicas dos solos por meio da analise de solo e tambem os seguintes procedimentos de manejo de solo: escarificacao, gradagem aradora e uma escarificacao com enxada rotativa. A cultura da batata necessita de um intenso manejo de solo, para que esta nao encontre nenhum obstaculo que possa impedir o seu desenvolvimento vegetativo e de seus tuberculos, observou-se tambem que a quantidade de nutrientes aplicados e bastante elevada e nem sempre apresenta criterios tecnicos para escolha da dose de fertilizantes. A batata apresenta grande demanda de nutrientes em todo o ciclo e tambem de um preparo intenso de solo, pois o mesmo nao deve apresentar nenhum tipo de barreira, sendo que este manejo deve ser realizado com no minimo 30 dias antes do plantio, para que o solo ja esteja apto ao plantio.

Arbuscular mycorrhizal fungi (AMF) are very important to plant nutrition, mostly in terms of acquisition of P and micronutrients. While Acacia mangium is closely associated with AMF throughout the whole cycle, Eucalyptus grandis presents this symbiosis primarily at the seedling stage. The aim of this study was to evaluate the dynamics of AMF in these two tree species in both pure and mixed plantations during the first 20 months after planting. We evaluated the abundance, richness and diversity of AMF spores, the rate of AMF mycorrhizal root colonization, enzymatic activity and soil and litter C, N and P. There was an increase in AMF root colonization of E. grandis when intercropped with A. mangium as well as an increase in the activity of acid and alkaline phosphatase in the presence of leguminous trees. AMF colonization and phosphatase activities were both involved in improvements in P cycling and P nutrition in soil. In addition, P cycling was favored in the intercropped plantation, which showed negative correlation with litter C/N and C/P ratios and positive correlation with soil acid phosphatase activity and soil N and P concentrations. Intercropping A. mangium and E. grandis maximized AMF root colonization of E. grandis and phosphatase activity in the soil, both of which accelerate P cycling and forest performance.

Our knowledge of the rhizosphere bacterial communities in deep soils and the role of Eucalyptus and Acacia on the structure of these communities remains very limited. In this study, we targeted the bacterial community along a depth profile (0 to 800 cm) and compared community structure in monospecific or mixed plantations of Acacia mangium and Eucalyptus grandis. We applied quantitative PCR (qPCR) and sequence the V6 region of the 16S rRNA gene to characterize composition of bacterial communities. We identified a decrease in bacterial abundance with soil depth, and differences in community patterns between monospecific and mixed cultivations. Sequence analysis indicated a prevalent effect of soil depth on bacterial communities in the mixed plant cultivation system, and a remarkable differentiation of bacterial communities in areas solely cultivated with Eucalyptus. The groups most influenced by soil depth were Proteobacteria and Acidobacteria (more frequent in samples between 0 and 300 cm). The predominant bacterial groups differentially displayed in the monospecific stands of Eucalyptus were Firmicutes and Proteobacteria. Our results suggest that the addition of an N2-fixing tree in a monospecific cultivation system modulates bacterial community composition even at a great depth. We conclude that co-cultivation systems may represent a key strategy to improve soil resources and to establish more sustainable cultivation of Eucalyptus in Brazil.

Sodium salicylate (NaS) and aspirin (ASA) are known to have a variety of effects on microorganisms, such as fungus (C. albicans and C. neoformans), moreover, it have effects in leukocyte adhesion and migration in vitro. In this report, we investigated the effect of ASA and NaS in neutrophil migration and cytokine production in C. albicans-induced peritonitis murine model. For this, mice were treated intraperitoneally (i.p) or orally (po) with NaS or ASA; after they were stimulated i.p. with C. albicans, the cellular migration was evaluated 24 h after stimulation. NaS, in mice treated i.p., unlike ASA, was able to inhibit the neutrophil migration and proinflammatory cytokine production induced by C. albicans, such as TNF-α, IL-1, IFN-γ, IL-12, and IL-10, but did not alter the IL-4 levels in these animals. However, the po treatment with same the dose of NaS or ASA did not affect the influx of this cell for inflammatory site. These results suggest that the NaS inhibits cellular migration and proinflammatory cytokine by different anti-inflammatory mechanism compared to ASA.

Pasture degradation is a concern, especially in susceptible sandy soils for which strategies to recover them must be developed. Microbiological and biochemical soil health indicators are useful in the guindace of soil management practices and sustainable soil use. We assessed the success of three Panicum maximum Jacq. cultivars in the reclamation of a pasture in a sandy Typic Acrudox in the northwest of the state of Parana, Brazil, based on soil health indicators. On a formerly degraded pasture with Urochloa brizantha (Hochst. ex A. Rich.) R.D. Webster, a trial with three P. maximum (cv. Massai, Tanzania, or Mombaca) was conducted. Lime and phosphate were applied at set-up, and mineral N and K as topdressing. A remnant of degraded pasture adjacent to the trial was used as control. Twenty-three chemical, physical, microbiological and biochemical attributes were assessed for the 0-10 cm topsoil. The procedures for reclamation improved most of the indicators of soil health in relation to the degraded pasture, such as soil P, mineral N, microbial biomass C, ammonification rate, dehydrogenase activity and acid phosphatase. CO2 evolution decreased, whereas microbial biomass C increased in the pasture under reclamation, resulting in a lower metabolic quotient (qCO(2)) that points to a decrease in metabolic stress of the microbial community. The reclamation of the pasture with P. maximum, especially cv. Mombaca, were evidenced by improvements in the microbiological and biochemical soil health indicators, showing a recovery of processes related to C, N and P cycling in the soil.

•Mulching is pivotal to improve sustainability in tropical and subtropical soils.•No-tillage increased both total and microbial C and N in soil.•No-tillage associated to oat as mulching crop in winter increased P in soil.•N-cycling attributes were more effective to separate soil use and management.•Glutaminase activity was very sensitive indicator of soil management.

Forest recovery, especially of riparian forests, has become a key factor in the environmental planning of agriculture, to protect water resources and the biota. However, traditional agricultural crops have not only affected nutrient cycling by the loss of organic matter and of microbial biomass and activity, but also increased soil compaction, which can later affect reforestation. The aim of this study was to identify the soil (physical, chemical and microbiological) properties that most influence the litter moisture contents of C, N, and P in areas of recovering riparian forests (20, 10 and 5 years old), in comparison with a native site (NT). The analyses ANOVA, Duncan's test and Pearson's correlation were used to identify the properties influenced by time of recovery and to identify the relationship between them. Multivariate analysis and partition of inertia (pRDA) were used to identify the most important soil properties to explain variations in litter C, N, P, and moisture content. The highest litter C: N ratios (26.8 and 29.9, respectively) were found at the recovery sites after 20 and 5 years. Among the microbiological properties, basal respiration (CO2-C), microbial biomass carbon (MBC) and dehydrogenase and urease activities increased with the age of the recovery sites. Soil total C and N, NO3-N, microbial biomass nitrogen (MBN) and dehydrogenase and urease activities showed a positive correlation with litter C and N content, while microporosity shower a negative correlation. According to the pRDA analysis, NBM and soil C were the most important variables to explain litter C and N content. Besides, these variables together explained more than half of the total variation or 28.2 %. The improvement of the soil physical conditions and the addition of organic matter with higher N content could be a possibility of improving the soil microbiological and chemical conditions and accelerate the recovery process.

Carbon plays a key role in determining soil health, which is defined as the soil's capacity to maintain environmental functions and biological productivity. In this study, C cycling was evaluated in soils along a gradient of land use, from native forest (NF), reforested sites (secondary forest - SF, Araucaria angustifolia - AR, Pinus elliottii - PI), clear-cut P. elliottii stands (CT) to farmland (AG). NF, AR, and SF sites had lower litter C:N ratios than PI, CT, and AG sites. Soils under forests had more organic C than CT and AG soils, whereas soils with native species had more microbial biomass C than PI, CT and AG soils. Both metabolic quotient (qCO(2)) and dehydrogenase activity increased with land use. Multivariate analysis revealed that soils of AR and SF were similar to NF and differed from CT and AG, which had higher qCO(2) and dehydrogenase activity, suggesting microbial stress. Litter C:N ratios and soil microbial biomass C, moisture, CO2 evolution, and cellulase activity discriminated most effectively between land uses. Reforestation with native species restored soil properties to levels similar to those in NF, being more sustainable, whereas reforestation with the exotic P. elliottii made soils more similar to AG soils. (C) 2013 Elsevier Masson SAS. All rights reserved.

Soil microorganisms and microbial processes are influenced by the quality and quantity of plant waste entering the soil, by its seasonal and spatial distribution, by the ratio of above- to below-ground inputs, and by changes in nutrient inputs. Soil management strategies sometimes promote mixed-species plantations to mitigate the loss of soil nutrients and improve biogeochemical cycling. The objective of this study was to explore changes in microbiological and chemical attributes of soils and litter in the early stages of the second rotation of mixed and pure plantations of Eucalyptus grandis and Acacia mangium, and to look for correlations between attributes. Soil samples at 0-10 cm depth were collected two, seven, 14, and 20 months after planting in the following treatments: monocultures of A. mangium and E. grandis, a monoculture of E. grandis with N-fertilizer, and an intercropped plantation with E. grandis and A. mangium. Microbial soil attributes varied dramatically between treatments 20 months after planting. Total C, N and P contents in litter showed the strongest correlations with microbial biomass C and N (C-mic and N-mic), microbial respiration, and dehydrogenase activity in all sampling periods. Lower C/N and C/P ratios in litter and lower C/N and C-mic/tC ratios in soils after 20 months in the intercropped plantation illustrated the system's capacity for supplying inputs of high-quality organic matter rich in N and P, but this did not result in higher contents of these elements or greater microbial activity in soils. An implication of this finding is that, at least in the initial growth phase of these plantations, chemical attributes of the litter and variation in those attributes govern microbial processes and, consequently, are mostly responsible for plant development. Canonical discriminant analysis revealed changes in the microbiological and chemical attributes of soil in the intercropped plantation due to the plants growth and the leaf litter accumulation. Twenty months after planting, the different plantations could be discriminated by differences in litter chemistry (C, N, and P), total soil C, N-mic, and dehydrogenase activity, which were very similar in intercropped plantations and E. grandis with N-fertilizer. These results from the early stages of plantation development are important for understanding the dynamics of soil attributes in these systems, and especially in intercropped plantations. In intercropped areas the cumulative effect of microbial attributes reflects a more sustainable system. Long-term studies are needed to identify patterns that develop after 20 months, during the growth period of these plantations. (C) 2012 Elsevier B.V. All rights reserved.

A recuperação de áreas de floresta, principalmente de matas ciliares, tornou-se fator-chave para a adequação ambiental da agricultura, com o propósito de proteger os recursos hídricos e a biota. Entretanto, o cultivo agrícola tradicional, além de ter alterado a ciclagem de nutrientes com a perda de matéria orgânica, biomassa e atividade microbiana, causou também a compactação do solo, o que influencia a posterior implantação de reflorestamentos. O objetivo deste trabalho foi identificar quais atributos do solo (físicos, químicos e microbiológicos) mais influenciam o teor de C, N, P e umidade da serapilheira em áreas de mata ciliar com diferentes idades de recuperação (20, 10 e cinco anos), em comparação com uma área nativa (NT). A partir da ANOVA e dos testes de médias (Duncan) e correlação (Pearson), foram verificados quais atributos acompanham o tempo de recuperação e qual a relação entre eles. A análise multivariada de partição da inércia a partir da análise de redundância (pRDA) foi feita a fim de identificar quais atributos do solo foram mais importantes para discriminar entre as áreas, com base nos teores de C, N, P e umidade da serapilheira encontrados nessas áreas. Maior relação C/N da serapilheira foi encontrada nas áreas de 20 (26,8) e de cinco (29,9) anos. Entre os atributos microbiológicos avaliados, observou-se que a respiração basal do solo (C-CO2), o carbono da biomassa microbiana (CBM) e a atividade das enzimas desidrogenase e urease foram diretamente proporcionais ao aumento da idade das áreas. O C e N totais do solo, o N-NO3, o nitrogênio da biomassa microbiana (NBM), a atividade das enzimas desidrogenase e urease, bem como a macroporosidade correlacionaram-se positivamente e a microporosidade, negativamente, com o C e N da serapilheira. A partir da pRDA, o NBM e o carbono total do solo foram os atributos mais importantes para explicar o teor de N e C da serapilheira; a explicação compartilhada foi mais da metade de toda a explicação observada ou 28,2 %. A melhoria das condições físicas do solo e a adição de matéria orgânica que resultam na formação de uma serapilheira mais rica em N pode ser uma forma de melhorar as condições microbiológicas e químicas do solo e de acelerar o processo de recuperação.

Studies on microbial activity and biomass in forestry plantations often overlook the role of litter, typically focusing instead on soil nutrient contents to explain plant and microorganism development. However, since the litter is a significant source of recycled nutrients that affect nutrient dynamics in the soil, litter composition may be more strongly correlated with forest growth and development than soil nutrient contents. This study aimed to test this hypothesis by examining correlations between soil C, N, and P; litter C, N, P, lignin content, and polyphenol content; and microbial biomass and activity in pure and mixed second-rotation plantations of Eucalyptus grandis and Acacia mangium before and after senescent leaf drop. The numbers of cultivable fungi and bacteria were also estimated. All properties were correlated with litter C, N, P, lignin and polyphenols, and with soil C and N. We found higher microbial activity (CO2 evolution) in litter than in soil. In the E. grandis monoculture before senescent leaf drop, microbial biomass C was 46 % higher in litter than in soil. After leaf drop, this difference decreased to 16 %. In A. mangium plantations, however, microbial biomass C was lower in litter than in soil both before and after leaf drop. Microbial biomass N of litter was approximately 94 % greater than that of the soil in summer and winter in all plantations. The number of cultivable fungi and bacteria increased after leaf drop, especially so in the litter. Fungi were also more abundant in the E. grandis litter. In general, the A. mangium monoculture was associated with higher levels of litter lignin and N, especially after leaf drop. In contrast, the polyphenol and C levels in E. grandis monoculture litter were higher after leaf drop. These properties were negatively correlated with total soil C and N. Litter in the mixed stands had lower C:N and C:P ratios and higher N, P, and C levels in the microbial biomass. This suggests more effective nutrient cycling in mixed plantations in the long term, greater stimulation of microbial activity in litter and soil, and a more sustainable system in general.

Soil Health refers to the ecological equilibrium and the functionality of a soil and its capacity to maintain a well balanced ecosystem with high biodiversity above and below surface, and productivity. To understand and use soil health as a tool for sustainability, physical, chemical, and biological properties must be employed to verify which respond to the soil use and management within a desired timescale. Attributes with a rapid response to natural or anthropogenic actions are considered good indicators of soil health. Among the physical indicators, soil texture, aggregation, moisture, porosity, and bulk density have been used, while among chemical indicators total C and N, mineral nutrients, organic matter, cation exchange capacity, among others are well established. However, most of them generally have a slow response, when compared to the biological ones, such as microbial biomass C and N, biodiversity, soil enzymes, soil respiration, etc., in addition to macro and mesofauna. Thus, a systemic approach based on different kinds of indicators (physical, chemical and biological) in assessing soil health would be safer than using only one kind of attribute. Many human activities have caused desertification, loss of biodiversity, disruption of aggregates, loss of organic matter and nutrients, among others. Today, it is imperious to maintain soil health and productivity with increasing emphasis on reforestation and recuperation of degraded areas through the use of organic amendments, reintroduction of plants, soil fauna and microorganisms. This review focused on an integrative view on indicators of soil health to be used as tools for prediction of sustainability in production systems.

Araucaria angustifolia (Bert.) O. Kuntze is the main component of the Mixed Ombrophilous forest and, in the State of São Paulo, it is associated with a high diversity of soil organisms, essential for the maintenance of soil quality, making the conservation of this ecosystem a major and pressing challenge. The objective of this study was to identify the physical and chemical properties that are most closely correlated with dehydrogenase enzyme activity, basal respiration and microbial biomass under native (NF) and replanted (RF) Araucaria angustifolia forests in three regions of the state of São Paulo, in winter and summer. The main differentiating factors between the areas were also determined. Each forest was represented by three true replications; at each site, from around the araucaria trees, 15 soil samples (0-20 cm) were collected to evaluate the soil physical, chemical and microbiological properties. At the same points, forest litter was sampled to assess mass and chemical properties. The following microbiological properties were evaluated: microbial biomass carbon (MBC), basal respiration (CO2-C), metabolic quotient (Q: CO2), dehydrogenase enzyme activity (DHA) as well as the physical properties (moisture, bulk density, macroporosity and total porosity), soil chemical properties [pH, organic carbon (org-C), P, Ca, K, Mg, Al, H+Al], litter dry mass, and C, N and S contents. The data were subjected to analysis of variance (TWO-WAY: ANOVA). A Canonical Discriminant Analysis (CDA) and a Canonical Correlation Analysis (CCA) were also performed. In the soil under NF, the values of K, P, soil macroporosity, and litter dry mass were higher and Q: CO2 and DHA lower, regardless of the sampling period, and DHA was lower in winter. In the RF areas, the levels of moisture, porosity and Q: CO2 were higher in both sampling periods, and DHA was higher in winter. The MBC was only higher under NF in the summer, while the litter contents of C, N and S were greater in winter. In winter, CCA showed a high correlation of DHA with CO2-C, pH and H+Al, while in the summer org-C, moisture, Mg, pH and litter C were more associated with DHA and CO2-C. The CDA indicated H+Al, available P, total porosity, litter S content, and soil moisture as the most discriminating variables between NF and RF, but moisture was the most relevant, in both seasons and CO2-C only in winter. The combined analysis of CCA and CDA showed that the contribution of the microbiological variables to a differentiation of the areas was small at both samplings, which may indicate that the period after reforestation was long enough to allow an almost complete recovery of the microbial activity.