Net microbial amino sugar accumulation process in soil as influenced by different plant material inputs

Identifying the impact of plant material inputs on soil amino sugar synthesis may advance our knowledge of microbial transformation processes in soils. In a 12-week laboratory microcosm incubation, 1, 2, 4, and 6% (w/w) soybean leaf or maize stalk were initially added to soil, respectively, whereas soil without plant addition was used as a control. The results showed that adding organic materials to the soil led to a net accumulation of amino sugars, because of greater microbial synthesis. The ratios of glucosamine to galactosamine and of glucosamine to muramic acid, two indicators differentiating the relative contribution to soil organic matter of fungi and bacteria, showed substantial variance across the gradient of substrate addition. Our results suggest that the amount of nutrients in a given substrate is the primary attribute determining microbial net accumulation of soil amino sugars, especially in the relatively short term, whereas the composition of nutrients might be more important in the relatively long term when nutrients are not sufficient. The use of the two ratios (glucosamine to galactosamine and glucosamine to muramic acid) reflects different dynamics of galactosamine and muramic acid during the decomposition of organic substrates in soils. Muramic acid, compared with galactosamine, is more likely to accumulate in the soil active organic fraction under abundant nutrient conditions, whereas it would be decomposed along with active organic matter when the nutrients are scarce and remain in minor quantities in the clay fraction without being attacked by microbes.     

Determination of 15N abundance of amino acids in soil hydrolysates

¹⁵N abundance of amino acids in soil hydrolysates was determined by emission spectroscopic method.     

尿素向氨基糖的转化以及对土壤氨基糖库动态的影响

采用13CO(NH2)2为底物进行黑土培养实验,利用气相色谱/质谱技术测定土壤中三种氨基糖含量以及同位素富集比例,根据其微生物标识物作用探讨土壤中不同微生物群落对于尿素碳的同化利用特征及黑土氨基糖库对于尿素添加的响应。研究结果表明,尿素碳可以被土壤微生物同化利用,但是可利用性显著低于葡萄糖。氨基葡萄糖中13C富集比例显著高于胞壁酸,表明真菌对尿素碳的同化能力高于细菌。尿素添加使土壤有机碳含量有所下降,同时土壤氨基糖总量及其与有机碳的相对比例也显著降低,说明在碳源严重受限条件下,氨基糖可被优先分解利用以补充碳源供给。胞壁酸含量虽低,但其调节并平衡碳氮元素供给与需求的能力较强;氨基葡萄糖稳定性高于胞壁酸,但在碳源缺乏时也可部分分解。土壤氨基糖的动态与土壤碳氮的可利用性及其耦合作用密切相关,在平衡土壤碳氮需求方面具有一定的调节作用。     

Free amino sugar reactions in soil in relation to soil carbon and nitrogen cycling

Amino sugars represent a major constituent of microbial cell walls and hydrolyzed soil organic matter. Despite their potential importance in soil nitrogen cycling, comparatively little is known about their dynamics in soil. The aim of this study was therefore to quantify the behaviour of glucosamine in two contrasting grassland soil profiles. Our results show that both free amino sugars and amino acids represented only a small proportion of dissolved organic N and C pool in soil. Based upon our findings we hypothesize that the low concentrations of free amino sugars found in soils is due to rapid removal from the soil solution rather than slow rates of production. Further, we showed that glucosamine removal from solution was a predominantly biotic process and that its half-life in soil solution ranged from 1 to 3 h. The rates of turnover were similar to those of glucose at low substrate concentrations, however, at higher glucosamine concentrations its microbial use was much less than for glucose. We hypothesized that this was due to the lack of expression of a low affinity transport systems in the microbial community. Glucosamine was only weakly sorbed to the soil's solid phase (K_d=6.4±1.0) and our results suggest that this did not limit its bioavailability in soil. Here we showed that glucosamine addition to soil resulted in rapid N mineralization and subsequent NO₃⁻ production. In contrast to some previous reports, our results suggest that free amino sugars turn over rapidly in soil and provide a suitable substrate for both microbial respiration and new biomass formation.     

免耕秸秆还田和传统耕作方式下东北黑土氨基糖态碳的积累特征

以吉林德惠市中层黑土进行7年田间定位试验的小区土壤为研究对象,对免耕(NT)和传统耕作下(CT)耕层(0～20 cm)氨基糖态碳含量的变化特征进行了分析。结果表明,与传统耕作相比,实施免耕7年后整个耕层土壤中氨基糖态碳含量显著增加(p<0.05),以表层(0～5 cm)增加幅度最大,高达94.7%。说明在研究地区,免耕措施有利于微生物代谢物如细胞壁物质等作为潜在的碳源逐渐积累在土壤中。免耕土壤中不同微生物来源氨基糖态碳的含量均较传统耕作有显著增加,但是变化特征有所不同,其中免耕条件下真菌来源的氨基葡萄糖的积累量较传统耕作高出1倍多,而且氨基葡萄糖与细菌来源的胞壁酸的比值(6.9～7.3)显著高于传统耕作(4.7～5.4),暗示实施免耕秸秆还田7年后土壤中真菌已逐渐转为优势群体,而真菌占优势的农田生态系统具有更大的固碳潜力。     

Rapid and sensitive quantification of amino acids in soil extracts by capillary electrophoresis with laser-induced fluorescence

A growing body of research is arguing that amino acids are key components of the soil nitrogen cycle. For example, we now know that many plants can take up intact amino acids, even in competition with soil microbes. Our growing recognition of the importance of amino acids is not matched by knowledge of the amounts and type of amino acids in the soil, certainly not in comparison with our encyclopaedic knowledge of inorganic N. The primary reason that less is known about amino acids than inorganic N is that measuring the amounts of individual amino acids with conventional chromatographic techniques is slow and typically requires extensive sample clean-up if KCl extracts are analysed. The aim of this study was to develop capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) as a more rapid alternative for measuring individual amino acids in KCl extracts of soil. The CE-LIF method separated 17 common amino acids within 12 min, with detection limits between 7 and 250 nM. One molar KCl extracts could be analysed without any sample clean-up or de-salting, and spike and recovery tests indicated that the complex matrix of soil extracts did not affect quantification. Further evidence of the suitability and robustness of the method came from the repeated analysis (n=5) of the same soil KCl extract. The relative standard deviation of migration times for replicate analyses were <0.2% while relative standard deviations for peak areas were <5%. To demonstrate application of the CE-LIF method to real world problems it was used to analyse amino acids in 1 M KCl extracts from a sub-alpine grassland and a Eucalyptus regnans forest. The most abundant amino acids were Ala, Gly and Arg. Other amino acids present at smaller concentrations or in a minority of samples were Asn, Cit, GABA, Glu, His, Phe, Leu, and Lys. The proposed CE-LIF method offers significant advantages over chromatographic methods via its rapidity, reproducibility and, most importantly, its ability to analyse crude KCl extracts.     

Microbial response time to sugar and amino acid additions to soil

Soil microbial respiration is derived predominantly from the turnover of carbohydrates and proteins in soil. In most agricultural ecosystems, these C compounds enter soil mainly from rhizodeposition (root exudation and turnover). Our aim was to determine how long it takes for the microbial population to reach their maximum mineralization potential after the addition of low-molecular-weight (MW) rhizodeposits to the soil. We added sugar in the form of glucose and amino acids in the form of glycine to an arable, grazed grassland, Eucalyptus forest and boreal forest soil and monitored CO₂ efflux over a 6-h period. Artificial rainwater amended (zero C addition) or unamended soils were used as controls. The Michaelis-Menten substrate utilization profiles showed vastly different patterns of microbial mineralization capacity and substrate affinity between the soils. However, in all soils we showed that activation of the soil microbial community to C addition occurred almost instantaneously (?60 s) with the average time taken to reach half maximal CO₂ production being 14±8 min for glucose and 10±8 min for glycine. After reaching their maximal mineralization potential, the rate of CO₂ evolution remained constant for the remainder of the experiment. Our results showed that while substrate uptake and mineralization within the soil microbial biomass was activated quickly, subsequent adaptation and upregulation of its C processing capacity did not occur at least in the short term. The fast rate of microbial activation and substrate use we partially attribute to the large degree of functional redundancy that exists within the soil microbial community for processing rhizodeposits.     

Temporal responses of soil microorganisms to substrate addition as indicated by amino sugar differentiation

Amino sugars are one of the important microbial residue biomarkers which are associated with soil organic matter cycling. However, little is known about their transformation kinetics in response to available substrates because living biomass only contributes a negligible portion to the total mass of amino sugars. By using ¹⁵N tracing technique, the newly synthesized (labeled) amino sugars can be differentiated from the native portions in soil matrix, making it possible to evaluate, in quantitative manner, the transformation pattern of amino sugars and to interpret the past and ongoing changes of microbial communities during the assimilation of extraneous ¹⁵N. In this study, laboratory incubations of soil samples were conducted by using ¹⁵NH₄⁺ as nitrogen source with or without glucose addition. Both the ¹⁵N enrichment (expressed as atom percentage excess, APE) and the contents of amino sugars were determined by an isotope-based gas chromatography-mass spectrometry. The significant ¹⁵N incorporation into amino sugars was only observed in glucose plus ¹⁵NH₄⁺ amendment with the APE arranged as: muramic acid (MurN) > glucosamine (GlcN) > galactosamine (GalN). The dynamics of ¹⁵N enrichment in bacterial-derived MurN and fungal-derived GlcN were fitted to the hyperbolic equations and indicative for the temporal responses of different soil microorganisms. The APE plateau of MurN and fungal-derived GlcN represented the maximal extent of bacterial and fungal populations, respectively, becoming active in response to the available substrates. The different dynamics of the ¹⁵N enrichment between MurN and GlcN indicated that bacteria reacted faster than fungi to assimilate the labile substrates initially, but fungus growth was dominant afterward, leading to integrated microbial community structure over time. Furthermore, the dynamics of labeled and unlabeled portions of amino sugars were compound-specific and substrate-dependent, suggesting their different stability in soil. GlcN tended to accumulate in soil while MurN was more likely degraded as a carbon source when nitrogen supply was excessive.     

The automated determination of glucosamine,galactosamine, muramic acid,and mannosamine in soil and root hydrolysates by HPLC

Amino sugars make a significant contribution to soil organic N and are mainly of microbial origin. The most important amino sugars in soil are glucosamine, galactosamine, muramic acid, and mannosamine. A method was developed for the simultaneous determination of these four amino sugars by high‐performance reverse‐phase liquid chromatography in standard solutions, soil and root hydrolysates. Pre‐column derivatization with o‐phthaldialdehyde (OPA) was used in an automated sample injector with thermostatic regulation of the reagent at 4 °C. The separation of the four amino sugars was fully satisfactory and was not disturbed by other fluorescent components in the soil and root hydrolysates.     

Shifts in amino sugar and ergosterol contents after addition of sucrose and cellulose to soil

An incubation experiment with organic soil amendments was carried out with the aim to determine whether formation and use of microbial tissue (biomass and residues) could be monitored by measuring glucosamine and muramic acid. Living fungal tissue was additionally determined by the cell-membrane component ergosterol. The organic amendments were fibrous maize cellulose and sugarcane sucrose adjusted to the same C/N ratio of 15. In a subsequent step, spherical cellulose was added without N to determine whether the microbial residues formed initially were preferentially decomposed. In the non-amended control treatment, ergosterol remained constant at 0.44 μg g⁻¹ soil throughout the 67-day incubation. It increased to a highest value of 1.9 μg g⁻¹ soil at day 5 in the sucrose treatment and to 5.0 μg g⁻¹ soil at day 33 in the fibrous cellulose treatment. Then, the ergosterol content declined again. The addition of spherical cellulose had no further significant effects on the ergosterol content in these two treatments. The non-amended control treatment contained 48 μg muramic acid and 650 μg glucosamine g⁻¹ soil at day 5. During incubation, these contents decreased by 17% and 19%, respectively. A 33% increase in muramic acid and an 8% increase in glucosamine were observed after adding sucrose. Consequently, the ratio of fungal C to bacterial C based on bacterial muramic acid and fungal glucosamine was lowered in comparison with the other two treatments. No effect on the two amino sugars was observed after adding cellulose initially or subsequently during the second incubation period. This indicates that the differences in quality between sucrose and cellulose had a strong impact on the formation of microbial residues. However, the amino sugars did not indicate a preferential decomposition of microbial residues as N sources.     

Influence of crops, crop residues and manure on amino acid and amino sugar fractions of organic nitrogen in soil

A field experiment was carried out to study the effect of: (1) the cultivation of a cereal (pearl millet) and two legumes (mung bean and clusterbean), and (2) incorporation of crop residues and manure in soil, on depletion or enrichment of pools of amino acid-N and amino sugar-N in soil. Both legumes enriched amino acid and amino sugar fractions but pearl millet cultivation depleted them. Enrichment of these fractions was partly attributed to the conversion of N biologically fixed during legume cultivation to these fractions and partly to the conversion of unknown and non-hydrolysable N fractions to amino acid-N and amino sugar-N. Mineralization of amino acid-N and amino sugar-N fractions along with their conversion to unknown and non-hydrolysable fractions were the plausible reasons for the decline in their concentrations after pearl millet cultivation. Application of residues or manure reversed this effect of pearl millet.     

A novel in situ water perfusion and extraction method for soil amino acid quantification

Classic methods of soil amino acid (SAA) determination involve field collection of soils and disturbance during storage, sieving, mixing and extraction in the laboratory. We describe a novel inexpensive method for extraction of SAA's in situ with minimal soil disturbance. In a comparison between methods of SAA extraction from a semiarid grassland site, glu-x and arginine were the dominant SAA nitrogen sources detected by the novel method while serine and glycine were the dominant SAA nitrogen sources detected using the classic method. The extraction method employed is likely to be useful for accurate determination of SAA availability to plants and soil microbes.     

Studies on the rate of decomposition of plant residues in soil by following the changes in sugar components

Uniformly ¹⁴C labelled straw and ryegrass leaf were incubated in soil in the proportions 0.5 and 1% w/w, respectively at 12°C for various periods up to 5 years.
Samples were hydrolysed with 2.5 m H₂SO₄ under reflux for 20 min to release non-cellulosic sugars and subsequently with 12 M/0.5 m H₂SO₄ to release glucose from cellulose-like polymers. Total radioactivity was determined in whole soil and hydrolysates and in individual sugars.
Fitting the data to simple exponential equations, total and 2.5 m H₂SO₄ hydrolysate radioactivity were best expressed by three part exponential functions, whereas radioactivity in 12M/0.5M H₂SO₄ hydrolysates and sugars were best expressed by two part functions, except for arabinose from the straw, for which a three-part function was better.
A comparison of the fit was made with that using the single-term exponential expressions devised by Janssen, 1984 and Weibull, 1951 and with a modification of the two part exponential expression by the substitution of one term by the Weibull expression. Apart from the Weibull expression on its own, there was no significant difference between the expressions.
It is suggested that, even for well defined carbohydrate components of the substrate, the decomposability changes with time because of their increasing inaccessibility to microorganisms, rather than that several components are present each with a different constant rate of decomposition.     

Dynamics of soil amino sugar pools during decomposition processes of corn residues as affected by inorganic N addition

Purpose  

Identifying the impact of inorganic-nitrogen (N) availability on soil amino sugar dynamics during corn (Zea mays L.) residue decomposition may advance our knowledge of microbial carbon (C) and N transformations and the factors controlling these processes in soils. Amino sugars are routinely used as microbial biomarkers to investigate C and N sequestration in microbial residues, and they are also involved in microbial-mediated soil organic matter (SOM) turnover. We conducted a 38-week incubation study using a Mollisol which was amended with corn residues and four levels of inorganic N (i.e., 0, 60.3, 167.2, and 701.9 mg N kg⁻¹ soil). The objective of this study was to examine the effects of inorganic-N availability on fungal and bacterial formation and stabilization of heterogeneous amino sugars during the corn residue decomposition in soil.     

氨基酸菌肥施用量对茶树生长及土壤肥力的影响

为探究不同氨基酸菌肥施用量对茶树生长和土壤肥力含量的影响,以鸿雁12号茶树为研究对象,采用田间微区试验的方法,设置了不施用氨基酸菌肥作对照组(CK)和氨基酸菌肥施用量分别为75 kg·hm^-2(T1)、150 kg·hm^-2(T2)、225 kg·hm^-2(T3)和300 kg·hm^-2(T4)的处理,研究不同氨基酸菌肥施用量对茶树生长、茶叶品质及土壤肥力的影响。结果表明：氨基酸菌肥施用后能显著增加茶叶芽头密度及茶叶产量,提高茶叶中氨基酸含量和土壤中碱解氮含量。试验6个月后与CK相比,T4组的芽头密度显著增加了63.4%,茶叶产量显著增加了71.5%;T1～T4处理茶叶中氨基酸含量分别增加了9.73%、6.41%、7.57%和13.2%,土壤中碱解氮含量分别增加了25.0%、36.0%、46.0%和37.6%。综上,氨基酸菌肥的施用能提高茶叶产量及品质,改善茶园土壤环境质量,为茶园生产的合理施肥及优质高效生产提供数据参考。     

Effect of plant materials on microbial transformation of amino sugars in three soil microcosms

Amino sugars, being predominantly of microbial origin, can help elucidate the role of microbes in carbon and nitrogen cycling in soils. However, little is known about the microbial degradation and synthesis of soil amino sugars as affected by plant-derived organic materials. We conducted a 30-week microcosm study using three soils amended with soybean leaf or maize stalk to investigate changes in the amounts and patterns of amino sugars over time. The total soil amino sugar content initially increased during the incubation, but later decreased. Amino sugar content of soil amended with maize stalk peaked at an earlier time than it did for soybean leaf, suggesting nutrient quantity and substrate composition influence microbial transformation. Temporal dynamics of the proportion of total soil amino sugar to organic matter after plant material addition conformed to parabolic models (r > 0.8; p < 0.01), which tended to converge over time. The models predicted that the proportions would ultimately approach the initial values as determined before amendment. These findings suggest that soil organic matter has the ability to maintain a baseline steady-state level of amino sugars, and support the interpretation of soil amino sugar reservoir as two components: the Stable Pool (SP) and the Transition Pool (TP).     

Complete amino acid analysis in hydrolysates of foods and feces by liquid chromatography of precolumn phenylisothiocyanate derivatives

The amino acid analysis method using precolumn phenylisothiocyanate (PITC) derivatization and liquid chromatography was modified for accurate determination of methionine (as methionine sulfone), cysteine/cystine (as cysteic acid), and all other amino acids, except tryptophan, in hydrolyzed samples of foods and feces. A simple liquid chromatographic method (requiring no derivatization) for the determination of tryptophan in alkaline hydrolysates of foods and feces was also developed. Separation of all amino acids by liquid chromatography was completed in 12 min compared with 60-90 min by ion-exchange chromatography. Variation expressed as coefficients of variation (CV) for the determination of most amino acids in the food and feces samples was not more than 4%, which compared favorably with the reproducibility of ion-exchange methods. Data for amino acids and recoveries of amino acid nitrogen obtained by liquid chromatographic methods were also similar to those obtained by conventional ion-exchange procedures.     

Characterization of the soil aeration status by plant and soil analyses

In differently aerated sites of two catenae under forest (Luvisols) and meadow (Gleysols) in Poland it was investigated if aeration in the period 1980–1983 could be characterized by the indicator I₀ = N (in plants : soils): Mn (in plants : soils). In the Luvisol catena I₀ was significantly correlated with ODR and in the Gleysol catena with ODR and Eh, but in neither of them with air volume. In both catenae “active” and “total” manganese fractions were a suitable basis for I₀ calculation. In the Gleysol catena the investigation of top- (0–20) and subsoil (20–50 cm) appeared to be necessary when only the “active” Mn is considered.     

植物对氨基酸的吸收利用及氨基酸在农业中的应用

植物对氨基酸的吸收、转运、代谢以及氨基酸在肥料和农药上的应用国内外已有报道。已有研究证明,植物可直接吸收土壤中的氨基酸分子,其吸收后的转运、分配、代谢因氨基酸种类而异,产生的生理效应也不相同;氨基酸农药易被日光分解或被自然界微生物降解,在土壤中、植物体内不留残毒,其降解产物还可作为农作物的营养物质,提高农作物的品质和产量,施用这类农药,人畜安全,没有公害;氨基酸肥具有促进植株生长发育、增强抗逆性、改善土壤状况和提高作物产量的作用。