The decline and mortality of oak trees represent a significant global concern, threatening essential ecosystem services. Increased rates of oak decline and mortality are observed in temperate regions, attributed to a complex interplay of biotic and abiotic stressors, including temperature changes, pollution, invasive pests, and pathogens. The intricate interactions between these factors make identifying and addressing the underlying soil stressors challenging for effective oak health management. In the UK and Europe, increased nitrogen (N) deposition and acidifying compounds are implicated in disrupting nutrient cycles and diminishing the stress tolerance of oak trees. Elevated soil N can stimulate vegetative growth, potentially increasing vulnerability to insect pests and pathogens, while N deficiency can result in weaker trees susceptible to attacks. The interplay between soil N availability and tree health is further complicated by factors like plant pathogens, insect pests, plant physiology, root microclimate, and the soil microbiome. Soil microorganisms play a crucial role in N cycling and regulate N availability for plants. Nitrification, the oxidation of ammonium (NH₄⁺) to nitrite (NO₂⁻) and then nitrate (NO₃⁻), is a key process driven by AOB and AOA. Denitrification, the reduction of nitrate to gaseous nitrogen, is another significant process. While AOA and AOB coexist, they exhibit different responses to soil environmental factors, suggesting niche differentiation. AOA generally dominate in acidic soils and favor low ammonium environments, unlike AOB. Plant activity can also modulate AOA and AOB community abundance, with higher AOB abundance often observed under plant canopies. Understanding the mechanisms controlling the abundance and diversity of soil microbes involved in N transformations is crucial for determining if N-cycling destabilization is linked to oak stress and decline.

The literature review extensively covers previous research on oak decline, highlighting the various biotic and abiotic factors contributing to this phenomenon. Studies on the impact of nitrogen deposition and soil acidification on tree health are reviewed, emphasizing the complex interactions between nitrogen availability, vegetative growth, and susceptibility to pests and pathogens. The role of soil microorganisms in nitrogen cycling is thoroughly discussed, including the contrasting roles of AOB and AOA in nitrification, their responses to environmental factors, and the influence of plant communities on their abundance. The literature also supports the hypothesis that changes in microbial utilization of organic carbon and nitrogen can alter overall soil microbial community structure and impact nitrification processes. Overall, the review establishes a strong foundation for the current study by highlighting the gaps in knowledge regarding the links between soil N cycling, microbial communities, and oak tree health.

This study investigated the abundance and diversity of microbial communities driving nitrogen transformations in soil associated with symptomatic (declining) and asymptomatic (*Quercus robur* and *Q. petraea*) oak trees across seven woodlands in the UK. Ten symptomatic and ten asymptomatic trees were selected at each site based on crown condition and the presence of stem lesions indicative of acute oak decline or root decay fungi. Soil samples (0-20 cm) were collected from three cores per tree. Soil chemical analyses determined moisture content, total C and N concentrations, pH, available nitrate and ammonium, and nitrification potential. DNA was extracted from soil samples, and the abundance of key genes (amoA for AOB and AOA, nirS, nirK, and nosZ for denitrification) was quantified using qPCR. Amplicon sequencing was performed for 16S rRNA genes (bacteria and archaea) and the functional genes to assess microbial community composition. Piecewise structural equation modelling (Piecewise SEM) analyzed the relationships between soil abiotic variables, N-cycle microbial gene abundances, and tree health. A quasibinomial generalized linear model (glm) assessed the AOB:AOA ratio associated with tree health. Non-metric multidimensional scaling (NMDS) visualized compositional differences in microbial communities. Phylogenetic analysis was conducted using the 50 most abundant OTUs from each gene.

Soil chemical characteristics varied significantly across and within the seven sites, with pH ranging from 3.6 to 8.3 and NH₄⁺ concentrations exhibiting high variability. AOA *amoA* gene abundance ranged from 2.6 x 10¹ to 1.4 x 10⁵ copies g⁻¹ dry soil, while AOB *amoA* abundance ranged from 3.1 x 10¹ to 4.1 x 10⁵ copies g⁻¹ dry soil. The abundance of *nirS*, *nirK*, and *nosZ* genes also varied significantly across sites. Piecewise SEM revealed a positive association between asymptomatic trees and higher AOB *amoA* gene abundance, driven by soil pH. Higher pH soils were associated with increased AOB abundance. No direct relationship was found between AOA *amoA* abundance and tree health, but AOA abundance was linked to lower soil NH₄⁺ concentrations. Denitrification gene abundance was primarily driven by the soil C:N ratio, with positive correlations between different denitrification genes and also positive correlations between AOB abundance and denitrification genes. Soil parameters showed complex interrelationships, with soil pH influenced by the C:N ratio, and moisture content influencing several parameters, including NO₂⁺+NO₃⁻, NH₄⁺, total carbon, and nitrification potential.

The findings indicate an indirect effect of soil pH on oak health, mediated by AOB abundance. Higher soil pH promotes a more abundant AOB community, which is associated with healthier oak trees. This suggests that soil acidification negatively impacts AOB, potentially disrupting N cycling and contributing to oak decline. The lack of a direct relationship between AOA abundance and tree health, coupled with its association with NH₄⁺ concentrations, further emphasizes the specific roles of AOB and AOA in the soil N cycle. The strong influence of the C:N ratio on denitrification gene abundance points to the importance of organic matter dynamics in shaping the microbial community and N transformations. The complex interrelationships among soil parameters and microbial communities highlight the integrated nature of the soil ecosystem and its impact on tree health. The results suggest that improving soil conditions through strategies that balance C:N ratios may enhance AOB abundance and improve N cycling, which could indirectly alleviate stress on declining oak trees.

This study demonstrates a significant indirect effect of soil pH on oak decline, mediated by AOB abundance and soil nitrogen cycling. Ameliorating soil acidification through C:N ratio management could positively influence AOB populations, potentially enhancing nitrogen availability and improving oak tree health. Further research should focus on manipulative field experiments to test the causal links identified here and to investigate the long-term effects of soil management practices on oak health and soil microbial communities.

The study's cross-sectional design limits the ability to establish definitive causal relationships. While the piecewise SEM helps elucidate potential pathways, it doesn't replace manipulative experiments. The focus on specific genes might not fully capture the complexity of microbial community interactions influencing N cycling. The number of sampling sites and trees per site could potentially impact the generalizability of the findings. Further research is needed to confirm these relationships and investigate the underlying mechanisms.