Extracellular Enzyme Activities Research Articles

Soil microbial activity profiles associated with organic compost fertilizers in an urban garden

AbstractBiological soil health is recognized as an important component of sustainable agriculture due to microbial biomineralization of nutrients. However, soil health can be difficult to assess consistently across urban agricultural systems due to diverse land use histories, soil heterogeneity, and lack of mechanistic links to agricultural management practices (e.g., recycled compost addition) and crop outcomes. In this study, we characterized soil microbial activity profiles in an urban agriculture system in Minnesota, USA, including microbial abundance, soil respiration, extracellular enzyme activity, and crop yield. Garden plots were fertilized with recycled organic compost (either manure or municipal) at high or low rates (ranging from 2.6 to 39 tons ha−1) targeted to crop N and P demands. Control plots received inorganic fertilizer or no fertilizer. We found that a high application rate of manure compost supported 6–10x higher basal respiration than municipal compost or inorganic fertilizer. Enzyme activity data demonstrated that soil microbial communities exhibited unique profiles of biochemical function that varied among fertilizers of different compositions. Microbial biochemical function predicted 50% of the variability in bell pepper (Capsicum annuum) yield, while soil microbial community size alone was a poor predictor of yield. Yield was highest in plots fertilized with municipal compost, outperforming inorganic fertilizer by threefold. High‐yield plots exhibited higher ratios of N to P enzyme activity compared to those with lower yield. Our findings demonstrate that “more is better” may not necessarily be true regarding soil microorganisms in biological soil health, and that measures of soil microbial biochemical function may be more important.

Study on the Potential Probiotics Isolated from Marine Aquaculture System and Evaluation for Aquaculture Application

In recent years, more attention has been paid to the application of host‐associated microorganisms as an appropriate source of probiotics in aquaculture. These probiotics promoted effective colonization and application and provided better benefits than other sources. A total of 32 isolates were collected from the water, shrimp, and sediment of the cultured pond of Litopenaeus vannamei. Probiotic effects were comprehensively evaluated by extracellular enzyme activity, virulence genes detection, bacteriostatic activity, drug susceptibility test, tolerance to adverse environment, and challenge experiment. Six of these isolates exhibited bacteriostatic activity against pathogens, such as Vibrio cholerae, V. owensii, V. fluvialis, V. campbellii, V. rotiferianus, Photobacterium damselae, V. vulnificus, and V. parahaemolyticus. Using 16S rRNA gene sequencing analysis, isolates were identified as Bacillus subtilis (P8, P29, and P30), B. velezensis (P12), Prestia flexa (P21), and Cytobacillus firmus (P23). They were tolerant to high temperature, low acid (pH 2.0), bile salt (2%), and the simulated gastrointestinal environment, secreting phospholipase, amylase, protease, gelatinase, and lipase. They were sensitive to only two or fewer drugs among 27 antibacterial drugs and did not carry any of the tested virulence genes. Biosafety in vivo evaluations showed that they were safe for Ctenopharyngodon idella (P12 ＞ P23 ＝ P29 ＞ P8 ＞ P30 ＝ P21). The results suggest that P8, P12, P23, P21, P29, and P30 have high probiotic potential and can be used as good probiotics in aquaculture and the development of ecological preparations.

Effects of thinning on accumulation of soil microbial residue carbon of Picea asperata plantations in sub-alpine region of western Sichuan, China

Microbial residues are an important component of soil organic carbon (SOC). It is unclear how long-term thinning affects the accumulation characteristics of microbial residue carbon (C). We analyzed the differences in soil physicochemical properties, microbial communities, extracellular enzyme activities, and microbial residue C in topsoil (0-10 cm) and subsoil (20-30 cm) in Picea asperata plantation of non-thinned (control, 4950 trees·hm-2) and thinned for 14 years (1160 trees·hm-2) stands, aiming to reveal the regulatory mechanism of thinning on microbial residue C accumulation. The results showed that thinning significantly increased SOC content, total nitrogen content, available phosphorus content, the proportion of particulate organic C, soil water content, C-cycle hydrolase, and acid phosphatase activities, but significantly reduced the proportion of mineral-associated organic C. Thinning significantly affected the content of fungal and microbial residue C, and the contribution of microbial residue C to SOC, and these effects were independent of soil layer. The content of fungal and microbial residue C was 25.0% and 24.5% higher under thinning treatments. However, thinning significantly decreased the contribution of microbial residue C to SOC by 12.3%, indicating an increase in the proportion of plant-derived C in SOC. Stepwise regression analysis showed that total nitrogen and soil water content were key factors influencing fungal and micro-bial residue C accumulation. In summary, thinning promoted microbial residue C sequestration by altering soil pro-perties and changed the composition of SOC sources.

Thermophilic bacteria of the Verhne-Vilyuchinskie and Sredne-Paratunskie hot springs (Kamchatka, Far East)

The number of physiological groups, biodiversity, enzymatic activity and some physiological and biochemical properties of microorganisms isolated from thermal waters and microbial mats of Kamchatka were studied. A low content of microorganisms with a predominance of denitrifying (Verhne-Vilyuchinsky) and nitrifying bacteria (Sredne-Paratunsky springs) was shown. Gram-positive spore- forming rods of the genus Bacillus predominated among the isolated bacteria. The isolated microorganisms had extracellular enzymatic activity and were capable of growing in wide ranges of temperatures, pH and NaCl concentration. Strains of thermophilic bacteria are of interest for biotechnology purposes.

Biological evaluation of mangrove endophytic fungi Aspergillus terreus derived from Sonneratia alba

Mangroves represent highly diverse marine ecosystems, encompassing various vegetation such as shrubs, plants, trees, palms, and other plant forms that have adapted to thrive in both freshwater and saline environments. Within these mangrove ecosystems, secondary metabolites are present, contributing to various pharmacological functions and holding significant ecological importance. Endophytic fungi constitute a substantial and quantifiable portion of fungal biodiversity and are acknowledged for their impact on the composition and diversity of plant communities. This study aims to investigate the extracellular proteins and enzyme activity exhibited by endophytic fungi discovered in the leaves of Sonneratia alba mangroves collected in Buton, Southeast Sulawesi, Indonesia. The primary objective is to evaluate the compatibility of marine fungi with antibacterial and antioxidant activity. The research involves analyzing the biological activities of marine fungi containing cellulase enzyme through methods such as DNS for reducing sugar, protein concentration determination, antioxidant analysis, and total phenol content assessment. Sample A (shaker) has highest enzyme activity at Day 4 and Sample B has highest enzyme activity at Day 6 (static condition). The protein concentration in the purified cellulase from Aspergillus terreus surpassed that of crude enzyme extracts.

Land management of formerly subtropical Atlantic Forest reduces soil carbon stocks and alters microbial community structure and function

Soil carbon (C) sequestration has been proposed as one means to mitigate the effects of climate change and to guarantee food safety. However, forest conversion to managed land uses can deplete soil C stocks and affect soil microbial communities, potentially impacting a wide range of ecosystem services. Our objective was to investigate the effect of forest conversion to intensive land uses on soil C dynamics and soil microbial community structure and function in the subtropical portion of the Atlantic Forest biome. We evaluated four land uses - forest fragment, grassland, and agriculture with five or fifty years of cultivation. We analyzed soil C pools, edaphic variables, and soil bacterial/archaeal (16S) and fungal (ITS) communities. Forest conversion to managed land uses reduced soil C stocks and altered soil microbial community structure, with implications for soil C dynamics. Bacterial/archaeal and fungal community composition were strongly correlated with pH, base saturation, and calcium (Ca) and magnesium (Mg) content. However, soil C pools were not associated with potential extracellular enzyme activity or microbial diversity. Finally, the fungal species Mortierella minutissima ASV_1 and several bacterial groups were positively correlated with soil C stocks, potentially representing indicators of soil C storage. Our study indicates that land use change affects the interplay between soil microbial communities and soil C pools, with consequences to soil C storage. Our findings may support the development of soil C enhancement programs within the Atlantic Forest biome.

Evaluation of extracellular alkaline proteases from Bacillus for environmentally friendly detergent additives

Growing demand for environmentally friendly and sustainable enzyme solutions drives markets for alkaline proteases across various industries. Alkaline proteases produced by different Bacillus strains exhibit unique properties and versatility. Production of alkaline proteases from Bacillus amyloliquefaciens TBRC 2902, B. siamensis TBRC 1180, B. subtilis TBRC 6663, and B. velezensis TBRC 7773 was optimized by adjusting production media, considering carbon and nitrogen sources to enhance extracellular enzyme activities. Optimized extracellular protease activity was achieved using tapioca starch for TBRC 2902 and TBRC 1180, and soluble starch for TBRC 7773, while both carbon sources were optimal for TBRC 6663. Skim milk was an equally effective nitrogen source for TBRC 2902, TBRC 1180, and TBRC 7773, whereas soytone was as effective as yeast extract for TBRC 6663. Inorganic nitrogen sources, such as diammonium hydrogen phosphate for TBRC 2902 and potassium nitrate for TBRC 1180, enhanced alkaline protease activity by 10-20%. The protease from B. siamensis TBRC 1180 exhibited optimal pH and temperature values of 9.0 and 60°C, respectively, while the others had optimal values of 8.0 and 50°C. All enzymes tolerated non-ionic surfactants, retaining over 40% activity after 24 hours exposure to Triton X-100, Tween-20, and Tween-80, indicating their potential as detergent additives.

Responses of soil microbial metabolic limitations to erosion under contrasting soil types and management practices

AbstractSoil microbial metabolism plays an important role in supporting soil services and functionalities but is usually restricted by the availability of soil nutrients. Although soil microbial metabolism limitation (SMML) has been widely addressed with regard to various driving factors, its response to erosion has not been examined but is urgently needed given the important influence of erosion on global environments. To understand how erosion affects SMML and whether erosion's effect varies with tillage practices and soil types, we conducted a simulated erosion experiment to compare eroded and noneroded plots with contrasting soil types (Mollisol vs. loess soils) and managements (conventional tillage versus no‐tillage in plots with crops versus without crops). We measured soil extracellular enzymatic activities and quantified SMML with extracellular enzymatic stoichiometry models. In both soils, the microbes were limited by carbon (C) and phosphorus (P), but not by nitrogen, and microbial C and P limitations were greater in Mollisol than loess soils. For the Mollisol, erosion significantly increased the relative C limitation, whereas the presence of crops eliminated this increase. For the loess soils, erosion enhanced the relative C limitation regardless of crop presence and tillage management. Erosion consistently increased microbial P limitation in plots with crops, with greater effects under no‐tillage (+2.9%) compared to conventional tillage (+0.7%) conditions regardless of soil type (p < 0.05). In plots without crops, microbial P limitation was increased by erosion under no‐tillage (+2.1%) but was decreased under conventional tillage (−5.6%) (p < 0.05). Additionally, conventional tillage exacerbated microbial C limitation but alleviated microbial P limitation. These results indicated that erosion shifted microbial C and P limitations depending on soil type, tillage practice and the presence of plants, highlighting the importance of considering all these factors when assessing soil microbial metabolic limitation at various spatial scales.

ISOLATION, IDENTIFICATION, AND CHARACTERIZATION OF MICROORGANISMS FROM INDUSTRIAL DAIRY WASTE AND EVALUATION OF ITS BIODEGRADABILITY

Microscopic organisms, commonly referred to as microbes, are organisms that are not visible to the naked eye. While some of these microorganisms can lead to diseases, numerous beneficial ones play essential roles in various domains. These advantageous microorganisms are pivotal in ecological systems, contributing to processes like biodegradation and bioremediation. They also find significant applications in diverse industries such as agriculture, food production, and the development of probiotics. This research aims to investigate the potential discovery of various biodegradable microorganisms capable of breaking down organic waste and similar materials. The focus is on finding effective ways to manage these materials in the context of global warming and related environmental concerns. The isolation of microorganisms involved the collection of samples from dairy industry effluents from dairy farms of Dehradun, Uttarakhand. Six isolates were screened following sample collection and examination. To distinguish the gathered microorganisms, various analyses were conducted. These included testing the isolates on nutrient agar at room temperature for 24 hours, with observations made on characteristics such as size, shape, color, margin, height, opacity, and stability. Morphological features like gram nature and motility were also examined. Subsequent to physical-biochemical characterization, tests for catalase, oxidase, and indole citrate were conducted. Biochemical tests were carried out to assess extracellular enzymatic activities, including caseinase and starch hydrolysis. Additionally, carbohydrate utilization tests were performed for lactose, maltose, inositol, xylose, among others. Out of the 06 isolates, two exhibited protease production and were selected for further investigation. Furthermore, two isolates demonstrated effective antibacterial activity and susceptibility to different antibiotics.

Screening and Molecular Identification of Potential Lignolytic White Rot Fungi Isolated from Western Ghats

ABSTRACT: The mounting concerns over the usage of enormous amount of wood and perilous chemicals in paper industry have led the researchers to develop ecofriendly green technologies for pulp and paper production. Lignocellulosic agro-waste can be an excellent alternative to the wood in papermaking. White rot fungi with ligninolytic properties can be effectively used for pretreatment of agro-wastes for pulp making. This study isolates and identifies previously unexplored strains of white rot fungi from Western Ghats of Kerala, India which can be used for developing biological systems for lignocellulosic treatment in pulp and paper production. Ten isolates were identified using molecular techniques and screened for their extracellular enzyme activities, including laccase, Manganese peroxidase and Lignin peroxidase. Among the 10 isolates, Trametes versicolor (Laccase activity 31.79 U/ml, Manganese peroxidase (MnP) 42.336 U/ml and Lignin peroxidase (LiP) - 50.65U/ml, Favolus teniculus (Laccase - 41.54 U/ml, MnP - 44.07 U/ml &LiP - 30.54), Coriolopsis byrsina (Laccase - 42.56 U/ml, MnP - 43.54 U/ml and LiP - 25.14 U/ml) Lenzitus betulina (Laccase - 37.15 U/ml, MnP - 38.97 U/ml and LiP - 30.43 U/ml) exhibited highly promising lignolytic enzymatic system. The study’s findings may provide a better eco-friendly substitute for conventional chemical treatments in various industrial applications.

Isolation, Characterization, and Selection of Bacterial Endophytes from Soybean (Glycine max) Nodules and Roots in Some Soils of Cameroon for Promoting Growth of Forage Legume Plants

Aim: The present study aimed to isolate, characterize and select the most effective bacterial endophytes to enhance soybean growth, biomass, and yield. Methodology: Seven soil samples were collected from the rhizosphere of forage legumes in fields across three agroecological zones of Cameroon. Bacterial endophytes were isolated from soybean roots and nodules and cultured on nutrient agar. The isolates were screened for the tolerance tests, extracellular enzymatic activity, P-solubilization activity, and IAA production. The best isolates were selected using a two-factor block factorial design. Each treatment was replicated six times. Results: A total of 85 bacterial endophytes were isolated. Characterization results of 22 preselected isolates revealed an optimal growth temperature of 37°C and a pH range between 6 and 7. Furthermore, the 22 isolates solubilized inorganic phosphate, 7 produced IAA, and 8 exhibited amylase activity. PCR analysis of the nifH and nodC genes showed that the isolates possessed the nifH gene as a nitrogen fixation marker and the nodC gene as a nodulation marker. The findings show that, out of the 22 bacterial endophyte isolates, NTT1 and BOSH9 were the most effective in increasing plant height by 26.74% and 31.78%, respectively. Additionally, they resulted in an increase in biomass of 94.24% to 120.48% and an increase of 71.59% to 76.70% in grain yield compared to control treatment. Conclusion: The selected isolates significantly enhance plant growth, increase biomass, and improve soybean grain yield. However, their potential use as biofertilizers in agriculture will require further investigation under real field conditions.

Organic nutrient availability and extracellular enzyme activities influence harmful algal bloom proliferation in a coastal aquaculture area

Small-scale but highly destructive harmful algal blooms (HABs) are increasingly observed in coastal aquaculture areas. However, the impact of elevated organic nutrients generated during mariculture production and their bioavailability for HABs are not yet fully understood. This study investigated the population dynamics of HAB-causing species and the hydrolytic dynamics of different extracellular enzyme activities (EEAs) along with variations in environmental inorganic and organic nutrients during a dinoflagellate bloom of Prorocentrum shikokuense adjacent to the coastal aquaculture area in Sansha Town, located in the East China Sea. The results indicated that both inorganic and organic nutrients were enriched in the study area, with organic nutrients comprising more than half of the total nutrient content. The presence of enriched organic nutrients and a high N:P ratio significantly contributed to bloom occurrence. Organic nutrient concentrations played a predominant role in controlling both the density and duration of the P. shikokuense bloom. The significant increase in EEAs during the bloom stages, primarily contributed by P. shikokuense, emphasized the crucial hydrolytic role of extracellular enzymes in nutrient recycling and supply for the bloom. The enzymatic kinetic parameters indicated enhanced efficiency in the recycling of organic nutrients as the bloom progressed. This study highlights the significant role of organic nutrients facilitated by EEAs in determining the occurrence, duration, and magnitude of HABs in coastal aquaculture areas.

Heterogeneity in Neutrophil Extracellular Traps from Healthy Human Subjects.

Neutrophil extracellular traps (NETs), a key component of early defense against microbial infection, are also associated with tissue injury. NET composition has been reported to vary with some disease states, but the composition and variability of NETs across many healthy subjects provide a critical comparison that has not been well investigated. We evaluated NETs from twelve healthy subjects of varying ages isolated from multiple blood draws over a three-and-one-half-year period to delineate the variability in extracellular DNA, protein, enzymatic activities, and susceptibility to protease inhibitors. We calculated correlations for NET constituents and loss of human bronchial epithelial barrier integrity, measured by transepithelial electrical resistance, after NET exposure. We found that although there was some variability within the same subject over time, the mean NET total DNA, dsDNA, protein, LDH, neutrophil elastase (NE), and proteinase 3 (PR3) in isolated NETs were consistent across subjects. NET serine protease activity varied considerably within the same donor from day to day. The mean NET cathepsin G and MPO were significantly different across donors. IL-8 > IL-1RA > G-CSF were the most abundant cytokines in NETs. There was no significant difference in the mean concentration or variability of IL-8, IL-1RA, G-CSF, IL-1α, IL-1β, or TNF-α in different subjects' NETs. NET DNA concentration was correlated with increased NET neutrophil elastase activity and higher NET IL-1RA concentrations. The mean reduction in protease activity by protease inhibitors was significantly different across donors. NET DNA concentration correlated best with reductions in the barrier integrity of human bronchial epithelia. Defining NET concentration by DNA content correlates with other NET components and reductions in NET-driven epithelial barrier dysfunction, suggesting DNA is a reasonable surrogate measurement for these complex structures in healthy subjects.

Microbial carbon and nitrogen limitation in Larix gmelinii forests along an altitudinal gradient: Evidence from ecoenzymatic stoichiometry and vector analysis

Mountain ecosystems have obvious variations in environmental factors at small spatial scales and thus provide a natural platform for investigating climate change. To investigate the characteristics of extracellular enzymes and microbial metabolic limitations in the hostile environmental conditions of cold temperate mountain ecosystems, soil physicochemical properties and enzyme activities were measured in the 0–10 cm and 10–20 cm soil layers of Larix gmelinii forests at elevations of 830 m, 950 m, 1100 m, and 1300 m. Our results showed that altitude and soil depth had a significant effect on 1,4-β-glucosidase, 1,4-β-N-acetyl-glucosaminidase, leucine aminopeptidase, and acid phosphatase activities. Extracellular enzyme activities decreased with increasing altitude and soil depth. The stoichiometric ratio of soil C-, N-, and P-acquiring enzymes after logarithmic transformation was 1:1.27:0.72 across all altitudes and soil depths. The metabolism of the microorganisms was mainly limited by C and N, and a high C limitation was observed at higher-altitude sites. With increased soil depth, the N limitation gradually weakened. Moreover, soil bulk density and pH were the main factors affecting extracellular enzyme activities and microbial nutrient constraints. Our study revealed that the carbon and nitrogen limitation of soil microbes was induced by the variation in soil factors in a cold temperate mountain forest ecosystem. This finding provides important insights into the mechanism of microbial metabolic limitation in the face of climate change.

Variations in crop yield caused by different ratios of organic substitution are closely related to microbial ecological clusters in a fluvo-aquic soil

ContextPartial substitution of chemical fertilizer with organic fertilizer (PSCFOF) can improve soil quality and reduce the overreliance of crop yield on chemical fertilizers. However, the relationship between alterations in crop yield and soil microbial community in response to various substitution ratios is not comprehensively understood. ObjectivesThis study aimed to investigate the impact of PSCFOF on soil nutrient supply, enzyme activity, bacterial and fungal community, and crop yield. MethodsAn 11-year (2009 – 2021) field experiment was carried out in a winter wheat–summer maize double-cropping system with four fertilization regimes (no fertilizer, NF; pure chemical fertilizer, NPK; 30% organic substitution, NPKLM; and 50% organic substitution, NPKHM). Co-occurrence network analysis was applied to examine the importance of ecological clusters and keystone species in controlling microbial activities and crop yields. ResultsThere were no significant differences in soil nutrient contents and annual crop yield between the NPK and NPKLM treatments. However, the NPKHM treatment resulted in significant reductions in crop yield by 9.49%− 16.78%, despite increasing soil organic matter (SOM), total nitrogen (TN), available nitrogen (AN), and available phosphorus (AP) by 9.05%, 7.26%, 11.67%, and 26.62%, respectively. This contrasting effect was primarily attributed to the influence of a specific ecological cluster (module 2), rather than the entire microbial community. Long-term 50% organic substitution would suppress keystone species within module 2, such as Aggregatilinea, Luteimonas, Usitatibacter and Microvirga. These species were specifically correlated with soil carbon enzyme activities, including β-glucosidase, invertase, and dehydrogenase, and contributed significantly and positively to crop yield. ConclusionsOrganic substitution at different ratios altered soil nutrient content, enzyme activities and microbial community in different intensities and directions. Long-term 30% organic substitution could preserve soil fertility and microbial community, leading to stable crop yield; whereas 50% organic substitution could inhibit keystone species, reduce extracellular enzyme activity and ultimately lead to a decrease in crop yield. ImplicationsOur results highlight the importance of ecological clusters within soil microbial co-occurrence network in regulating crop yield, presenting a new perspective on the variations in crop yield resulting from varying ratios of organic substitution.

Grazing exclusion alters denitrification N2O/(N2O + N2) ratio in alpine meadow of Qinghai–Tibet Plateau

Grazing exclusion has been implemented worldwide as a nature-based solution for restoring degraded grassland ecosystems that arise from overgrazing. However, the effect of grazing exclusion on soil nitrogen cycle processes, subsequent greenhouse gas emissions and underlying mechanisms remain unclear. Here, we investigated the effect of four-year grazing exclusion on plant communities, soil properties, and soil nitrogen cycle-related functional gene abundance in an alpine meadow on the Qinghai–Tibet Plateau. Using an automated continuous-flow incubation system, we performed an incubation experiment and measured soil-borne N2O, N2, and CO2 fluxes to three successive “hot moment” events (precipitation, N deposition, and oxic-to-anoxic transition) between grazing-excluded and grazing soil. Higher soil N contents (total nitrogen, NH4+, NO3−) and extracellular enzyme activities (β-1,4-glucosidase, β-1,4-N-acetyl-glucosaminidase, cellobiohydrolase) are observed under grazing exclusion. The aboveground and litter biomass of plant community was significantly increased by grazing exclusion, but grazing exclusion decreased the average number of plant species and microbial diversity. The N2O + N2 fluxes observed under grazing exclusion were higher than those observed under free grazing. The N2 emissions and N2O/(N2O + N2) ratios observed under grazing exclusion were higher than those observed under free grazing in oxic conditions. Instead, higher N2O fluxes and lower denitrification functional gene abundances (nirS, nirK, nosZ, and nirK + nirS) under anoxia were found under grazing exclusion than under free grazing. The N2O site-preference value indicates that under grazing exclusion, bacterial denitrification contributes more to higher N2O production compared with under free grazing (81.6 % vs. 59.9 %). We conclude that grazing exclusion could improve soil fertility and plant biomass, nevertheless it may lower plant and microbial diversity and increase potential N2O emission risk via the alteration of the denitrification end-product ratio. This indicates that not all grassland management options result in a mutually beneficial situation among wider environmental goals such as greenhouse gas mitigation, biodiversity, and social welfare.

Altered intra-annual precipitation patterns affect the N-limitation status of soil microorganisms in a semiarid alpine grassland

The ongoing intensification of the hydrological cycle due to global climate change alters intra-annual precipitation variability. Changes in precipitation patterns lead to disparities in soil moisture; however, the responses of soil extracellular enzyme activities (EEAs) and microbial metabolism limitations to them is unclear. This study conducted an in situ field experiment in the alpine grasslands of the Kunlun Mountains to simulate the same amount of precipitation but change the distribution time of precipitation within the plant growing season. We examined the effects of variability in intra-annual precipitation patterns on the stoichiometry of topsoil (0–5 and 5–20 cm) properties, microbial biomass, and EEAs. Results showed that altered precipitation patterns significantly increased soil moisture’s disparity index (D) by 57 %–89 %. The activity of nitrogen-acquiring enzymes (L-leucine aminopeptidase) was increased by 20.82–32.08 % (P < 0.05) with increased variability of precipitation, while phosphorus-acquiring enzymes (Acid phosphatase) and carbon-acquiring enzymes (β-glucosidase) both decrease by 9.25–23.35 % and by 17.86–33.04 %, respectively (P < 0.05). We determined that soil microorganisms in the study region were metabolism-limited by nitrogen (vector angles < 45◦), and increased precipitation variability exacerbated nitrogen limitation (decrescent vector angles) and alleviated carbon limitation (shortened vector length). In addition, soil EEAs can be both interactively influenced by biotic and abiotic factors, with abiotic factors (i.e., D, soil organic carbon content, and soil total nitrogen content) having a stronger effect. Our study provides an effective strategy for regulating soil EEAs, microbial biomass, and microbial metabolism limitation and contributes to developing nutrient management and restoration strategies in semiarid alpine grassland ecosystems.

Isolation and Identification of Bacillus subtilis LY-1 and Its Antifungal and Growth-Promoting Effects.

Peanut root rot, caused by Fusarium spp., is a devastating fungal disease. As part of a program to obtain a biocontrol agent to control peanut root rot in the field, a bacterial strain LY-1 capable of inhibiting the growth of the fungus in vitro was isolated from rhizosphere soil samples collected from wild mint by agar disk dilution and dual-culture assay. Strain LY-1 was identified as Bacillus subtilis based on morphological characteristics, 16S rDNA, and gyrA sequence analyses. The bacterial suspension and cell-free culture filtrate of LY-1 could significantly inhibit the growth of Fusarium oxysporum, Fusarium proliferatum and Fusarium solani, but volatile organic compounds from the cultures had only a weak effect on mycelial growth. The percentage inhibition of 20% concentration of the cell-free culture filtrate of LY-1 on conidium production of each of the three Fusarium species was greater than 72.38%, and the percentage inhibition by the culture filtration on the germination of conidia of the three species was at least 62.37%. The production of extracellular enzyme activity by LY-1 was studied in functional assays, showing protease, cellulase, amylase, chitinase, and β-1,3-glucanase activity, while LY-1 contained a gene encoding iturin, an antifungal lipopeptide. In addition, under pot culture in a greenhouse, culture filtrate of LY-1 significantly promoted the growth of peanut, increasing the fresh and dry mass of the plant by 30.77% and 27.27%, respectively, in comparison with the no-filtrate control. The culture filtrate of LY-1 increased the resistance of peanut plants to F. oxysporum, with the biocontrol efficiency reaching 44.71%. In conclusion, B. subtilis LY-1, a plant-growth-promoting rhizobacterium, was able to protect peanuts from Fusarium spp. infection.

The effect of intercropping leguminous green manure on theanine accumulation in the tea plant: A metagenomic analysis.

Intercropping is a widely recognised technique that contributes to agricultural sustainability. While intercropping leguminous green manure offers advantages for soil health and tea plants growth, the impact on the accumulation of theanine and soil nitrogen cycle are largely unknown. The levels of theanine, epigallocatechin gallateand soluble sugar in tea leaves increased by 52.87% and 40.98%, 22.80% and 6.17%, 22.22% and 29.04% in intercropping with soybean-Chinese milk vetch rotation and soybean alone, respectively. Additionally, intercropping significantly increased soil amino acidnitrogen content, enhanced extracellular enzyme activities, particularly β-glucosidase and N-acetyl-glucosaminidase, as well as soil multifunctionality. Metagenomics analysis revealed that intercropping positively influenced the relative abundances of several potentially beneficial microorganisms, including Burkholderia, Mycolicibacteriumand Paraburkholderia. Intercropping resulted in lower expression levels of nitrification genes, reducing soil mineral nitrogen loss and N2 O emissions. The expression of nrfA/H significantly increased in intercropping with soybean-Chinese milk vetch rotation. Structural equation model analysis demonstrated that the accumulation of theanine in tea leaves was directly influenced by the number of intercropping leguminous green manure species, soil ammonium nitrogen and amino acid nitrogen. In summary, the intercropping strategy, particularly intercropping with soybean-Chinese milk vetch rotation, could be a novel way for theanine accumulation.

Soil extracellular enzyme activity and microbial resource limitation exhibited close relationships with groundwater table decline in desert wetlands

In groundwater-fed desert wetlands, groundwater table (GWT) decline leads to the loss of carbon and nutrients, followed by the expansion of desertification. How the GWT decline affects soil nutrient cycling and microbial resource limitation is still unclear. Here, we sampled soils along a steep GWT decline gradient in desert wetlands and surroundings traversing marshland (ML), sandy grassland (SG) and bare sandy land (SL) in the southern margin of Mu Us Sandy Land, China, then determined the extracellular enzyme activity (EEA, including carbon (C)-, nitrogen (N)- and phosphorus (P)-acquiring enzymes) and related soil environmental variables, and quantified the relative value of microbial metabolic limitation by vector analysis. Results showed that GWT decline eventually inhibited the activities of soil C-, N- and P-acquiring enzymes, which might be unfavourable for nutrient cycling. The relationship between soil microbial C limitation and GWT showed a downwards unimodal trend, despite the largest SOC value observed in ML. However, microbial P limitation was relieved by a slight GWT decline, and the continuous decline in GWT eventually shifted microorganisms from P to N limitation. We further provide evidence that DOC and C:P were the key factors explaining the variations in microbial C and nutrient limitations along the decline in GWT, respectively. Altogether, these findings suggest that soil EEA and microbial resource limitation exhibit close relationships with GWT decline in desert wetlands. Moreover, they highlight that alleviating microbial N limitation may be the primary solution for vegetation restoration and carbon sequestration in sandy land around wetlands.