[Effects of 40 Years of Fertilizer Application on the Characteristics of Soil Enzymatic Stoichiometry in Black Soil].

Microorganisms produce extracellular enzymes to meet elemental requirements and cope with stoichiometric imbalances of resources. To gain insights into the cycling of C, N, and P, the activities of the C∶N∶P acquisition enzymes have been extensively investigated. To detect the effects of long-term fertilization practices on soil nutrient balance and characteristics of soil enzymatic stoichiometry in black soil, four different fertilization treatments were selected： no fertilization （CK）, nitrogen fertilizer （N）, phosphorus fertilizer （P）, and combination of nitrogen and phosphorus fertilizers （NP）. Soil samples were collected in both April 2021 and April 2022 to determine soil enzyme activities and their stoichiometric characteristics. The results showed that soil acid phosphatase and ß-D-glucosidase activities were significantly higher in the N and NP treatments than in CK by 68%-158% and 26%-222%, respectively. Soil ß-N-acetylaminoglucosidase activities were significantly higher in the P and NP treatments, with the highest around 75.48 nmol·ï¼ˆg·h）-1 and 106.81 nmol·ï¼ˆg·h）-1, respectively. Two-way ANOVA analysis showed that N and P inputs had a great impact on soil enzyme activities. Redundancy analysis showed that the main factors controlling enzyme activities were soil pH, microbial biomass phosphorus, and soil available P content. It was found that N inputs significantly increased enzyme vector length, which was ranged from 1.32 to 1.52, and the enzyme vector angles were all larger than 45°, suggesting C and P co-limited in the black soils. These findings suggest that 40 years of fertilization have had a great impact on soil enzymes and the related resource use strategy, which provides great implications for assessing soil nutrients balance and soil sustainability.

Responses of root-associated fungal community structure of mycorrhizal plants to nitrogen and/or phosphorus addition in a subtropical forest.

Root-associated fungi play a vital role in maintaining nutrient absorption and health of host plants. To compare the responses of root-associated fungal community structures to nitrogen (N) and/or phosphorus (P) additions across differential mycorrhizal types, we collected roots of nine plant species belonging to three mycorrhizal types (arbuscular mycorrhiza, ectomycorrhiza, and ericoid mycorrhiza) under control and N and/or P addition treatments from a subtropical forest, and detected the diversity and community composition of fungi inhabiting roots through the high-throughput sequencing technique. The results showed that root-associated fungal communities of all nine plant species were mainly composed of Basidiomycota and Ascomycota. The relative abundance of Ascomycota and Basidiomycota was significantly lower and higher under the P addition than that under control, respectively. The relative abundance of Ascomycota of ericoid mycorrhizal trees was significantly higher than those of arbuscular mycorrhizal and ectomycorrhizal trees, while the relative abundance of Basidiomycota was significantly lower than the other two mycorrhizal types. Compared with the control, P addition significantly reduced the α-diversity and changed community composition of root-associated fungi across different mycorrhizal plant types, while no effect of N addition or mycorrhizal type was observed. Compared with the control and N addition treatments, NP addition caused root-associated fungal communities of all plants becoming integrally divergent. In addition, the fungal communities of ectomycorrhizal mycorrhizal trees became apparently convergent in comparison with those of arbuscular and ericoid mycorrhizal trees under the NP addition. Collectively, our results highlighted that P was a critical factor influencing community structures of tree root-associated fungi in subtropical forest soils. This study would enhance our understanding of the responses and maintenance mechanisms of plant root-associated fungal diversity under global environmental changes in the subtropical region.

Response of gene abundance of ammonia-oxidizing microorganisms and denitrifying microorganisms to nitrogen and phosphorus addition in subtropical forest.

We conducted a nitrogen (N) and phosphorus (P) addition experiment in Qianjiangyuan National Park in 2015, to investigate the response of ammonia-oxidizing microorganisms and denitrifying microorganisms. There were four treatments, including N addition (N), P addition (P), NP, and control (CK). Soil samples were collected in April (wet season) and November (dry season) of 2021. The abundance of amoA gene of ammonia-oxidizing microorganisms (i.e., ammonia-oxidizing archaea, AOA; ammonia-oxidizing bacteria, AOB; comammox) and denitrifying microbial genes (i.e., nirS, nirK, and nosZ) were determined using quantitative PCR approach. The results showed that soil pH was significantly decreased by long-term N addition, while soil ammonium and nitrate contents were significantly increased. Soil available P and total P contents were significantly increased with the long-term P addition. The addition of N (N and NP treatments) significantly increased the abundance of AOB-amoA gene in both seasons, and reached the highest in the N treatment around 8.30×107 copies·g-1 dry soil. The abundance of AOA-amoA gene was significantly higher in the NP treatment than that in CK, with the highest value around 1.17×109 copies·g-1 dry soil. There was no significant difference in N-related gene abundances between two seasons except for the abundance of comammox-amoA. Nitrogen addition exerted significant effect on the abundance of AOB-amoA, nirK and nosZ genes, especially in wet season. Phosphorus addition exerted significant effect on the abundance of AOA-amoA and AOB-amoA genes in both seasons, but did not affect denitrifying gene abundances. Soil pH, ammonium, nitrate, available P, and soil water contents were the main factors affecting the abundance of soil N-related functional genes. In summary, the response of soil ammonia-oxidizing microorganisms and denitrifying microorganisms was more sensitive to N addition than to P addition. These findings shed new light for evaluating soil nutrient availability as well as their response mechanism to global change in subtropical forests.

Imaging asparaginyl endopeptidase (AEP) in the live brain as a biomarker for Alzheimer's disease.

BACKGROUND: Discovery of early-stage biomarkers is a long-sought goal of Alzheimer's disease (AD) diagnosis. Age is the greatest risk factor for most AD and accumulating evidence suggests that age-dependent elevation of asparaginyl endopeptidase (AEP) in the brain may represent a new biological marker for predicting AD. However, this speculation remains to be explored with an appropriate assay method because mammalian AEP exists in many organs and the level of AEP in body fluid isn't proportional to its concentration in brain parenchyma. To this end, we here modified gold nanoparticle (AuNPs) into an AEP-responsive imaging probe and choose transgenic APPswe/PS1dE9 (APP/PS1) mice as an animal model of AD. Our aim is to determine whether imaging of brain AEP can be used to predict AD pathology. RESULTS: This AEP-responsive imaging probe AuNPs-Cy5.5-A&C consisted of two particles, AuNPs-Cy5.5-AK and AuNPs-Cy5.5-CABT, which were respectively modified with Ala-Ala-Asn-Cys-Lys (AK) and 2-cyano-6-aminobenzothiazole (CABT). We showed that AuNPs-Cy5.5-A&C could be selectively activated by AEP to aggregate and emit strong fluorescence. Moreover, AuNPs-Cy5.5-A&C displayed a general applicability in various cell lines and its florescence intensity correlated well with AEP activity in these cells. In the brain of APP/PS1 transgenic mice , AEP activity was increased at an early disease stage of AD that precedes formation of senile plaques and cognitive impairment. Pharmacological inhibition of AEP with δ-secretase inhibitor 11 (10 mg kg-1, p.o.) reduced production of ß-amyloid (Aß) and ameliorated memory loss. Therefore, elevation of AEP is an early sign of AD onset. Finally, we showed that live animal imaging with this AEP-responsive probe could monitor the up-regulated AEP in the brain of APP/PS1 mice. CONCLUSIONS: The current work provided a proof of concept that assessment of brain AEP activity by in vivo imaging assay is a potential biomarker for early diagnosis of AD.

Pharmacological inhibition of asparaginyl endopeptidase by δ-secretase inhibitor 11 mitigates Alzheimer's disease-related pathologies in a senescence-accelerated mouse model.

BACKGROUND: Currently, there is no cure for Alzheimer's disease (AD). Therapeutics that can modify the early stage of AD are urgently needed. Recent studies have shown that the pathogenesis of AD is closely regulated by an endo/lysosomal asparaginyl endopeptidase (AEP). Inhibition of AEP has been reported to prevent neural degeneration in transgenic mouse models of AD. However, more than 90% of AD cases are age-related sporadic AD rather than hereditary AD. The therapeutic efficacy of AEP inhibition in ageing-associated sporadic AD remains unknown. METHODS: The senescence-accelerated mouse prone 8 (SAMP8) was chosen as an approximate model of sporadic AD and treated with a selective AEP inhibitor,: δ-secretase inhibitor 11. Activation of AEP was determined by enzymatic activity assay. Concentration of soluble amyloid ß (Aß) in the brain was determined by ELISA. Morris water maze test was performed to assess the learning and memory-related cognitive ability. Pathological changes in the brain were explored by morphological and western blot analyses. RESULTS: The enzymatic activity of AEP in the SAMP8 mouse brain was significantly higher than that in the age-matched SAMR1 mice. The half maximal inhibitory concentration (IC50) for δ-secretase inhibitor 11 to inhibit AEP in vitro is was around 150 nM. Chronic treatment with δ-secretase inhibitor 11 markedly decreased the brain AEP activity, reduced the generation of Aß1-40/42 and ameliorated memory loss. The inhibition of AEP with this reagent not only reduced the AEP-cleaved tau fragments and tau hyperphosphorylation, but also attenuated neuroinflammation in the form of microglial activation. Moreover, treatment with δ-secretase inhibitor 11 prevented the synaptic loss and alleviated dendritic disruption in SAMP8 mouse brain. CONCLUSIONS: Pharmacological inhibition of AEP can intervene and prevent AD-like pathological progress in the model of sporadic AD. The up-regulated AEP in the brain could be a promising target for early treatment of AD. The δ-secretase inhibitor 11 can be used as a lead compound for translational development of AD treatment.