Persistence and functional impact of a microbial inoculant on native microbial community structure,nutrient digestion and fermentation characteristics in a rumen model

Small sub-unit (SSU) rRNA-targeted oligonucleotide probes were used to monitor the persistence of a genetically engineered bacterium inoculated in model rumens. Eight dual flow continuous culture fermenters were operated with either standard artificial saliva buffer or buffer with chondroitin sulfate (0.5 g/l) added. After 168 h of operation, fermenters were inoculated with Bacteroides thetaiotaomicron BTX (BTX), at approximately 1% of total bacteria. B. thetaiotaomicron was quantified using a species-specific probe and shown to persist in fermenters 144 h after inoculation (relative abundance 0.48% and 1.42% of total SSU rRNA with standard and chondroitin sulfate buffers, respectively). No B. thetaiotaomicron SSU rRNA was detected in fermenter samples prior to inoculation with strain BTX. Relative abundances of Bacteria, Eucarya and Archaea were not affected by either inoculation or buffer type. Fiber digestion, in particular the hemicellulose fraction, increased after strain BTX addition. Chondroitin sulfate addition to the buffer increased bacterial nitrogen flow in fermenters, but did not alter fiber digestion. Neither inoculum nor buffer type altered total short chain fatty acid (VFA) concentrations but proportions of individual VFA differed. In model rumens, B. thetaiotaomicron BTX increased fiber digestion when added to mixed ruminal microbes, independent of chondroitin sulfate addition; but further study is needed to determine effects on other fiber-digesting bacteria.     

Ecoenzymatic stoichiometry of microbial nutrient acquisition in tropical soils

The relative activities of soil enzymes involved in mineralizing organic carbon (C), nitrogen (N), and phosphorus (P) reveal stoichiometric and energetic constraints on microbial biomass growth. Although tropical forests and grasslands are a major component of the global C cycle, the effects of soil nutrient availability on microbial activity and C dynamics in these ecosystems are poorly understood. To explore potential microbial nutrient limitation in relation to enzyme allocation in low latitude ecosystems, we performed a meta-analysis of acid/alkaline phosphatase (AP), β-1,4-glucosidase (BG), and β-1,4-N-acetyl-glucosaminidase (NAG) activities in tropical soils. We found that BG:AP and NAG:AP ratios in tropical soils are significantly lower than those of temperate ecosystems overall. The lowest BG:AP and NAG:AP ratios were associated with old or acid soils, consistent with greater biological phosphorus demand relative to P availability. Additionally, correlations between enzyme activities and mean annual temperature and precipitation suggest some climatic regulation of microbial enzyme allocation in tropical soils. We used the results of our analysis in conjunction with previously published data on soil and biomass C:N:P stoichiometry to parameterize a biogeochemical equilibrium model that relates microbial growth efficiency to extracellular enzyme activity. The model predicts low microbial growth efficiencies in P-limited soils, indicating that P availability may influence C cycling in the highly weathered soils that underlie many tropical ecosystems. Therefore, we suggest that P availability be included in models that simulate microbial enzyme allocation, biomass growth, and C mineralization.     

Effects of a surface wildfire on soil nutrient and microbial functional diversity in a shrubbery

The aim of the study was to determine effects of a wildfire on soil nutrients and soil microbial functional diversity in short-term time scales. Burned and unburned control soil samples were collected 1 day, and 2, 4, 8, 10, 12 and 15 months after a shrubbery fire in Yumin county of Xinjiang, Northwest China. Nutrients of soil in each sampling time were detected and soil microbial functional diversity was measured by Biolog Eco plates. Results of the study showed that soil nutrients were significantly affected by fire. Soil pH increased immediately after the wildfire and was higher than that of unburned soil during 15 months post fire. Soil organic matter and total N significantly decreased immediately after the fire and was even lower than control soil at the 15th month post fire. Soil available P level increased sharply during the 4th month after the fire, and later reached to the maximum value with eight times higher than that of unburned soil. Soil available N and available K were more than the control site in 2 months after the fire, then decreased, but available N began to increase, when vegetations restored 1 year after the fire. Soil microbial activity and functional diversity recovered gradually after fire. The average well color development (AWCD) and functional diversity indices (Shannon index, Simpson index, and McIntosh index) decreased significantly 1 day after the fire, but then increased and were similar to that of undisturbed soil 15 months after the fire, when plant started to regenerate in burned area. The changes in soil nutrients after the fire affected soil microbial activity and functional diversity. Correlation analysis revealed that AWCD was negatively correlated with soil pH and positively correlated with soil total N and available N, Shannon and Simpson index had positive significantly correlation with soil total N and McIntosh index had positive significantly correlation with available N. Result of principal component analysis based on the data of carbons metabolism showed that microbial catabolic profiles of burned soils of each sampling time after the wildfire were different and all were distinct from those of unburned soils, which might suggest that microbial community structure of fire-impacted area changed dynamically on monthly scale and was distinct from that of the control site in 15 months after fire, although microbial activity or richness showed similar to pre-fire level at the 15th month post-fire.     

Soil organic C and nutrient contents under trees with different functional characteristics in seasonally dry tropical silvopastures

Aims

The selection of tree characteristics is critical for the outcome of the tree effects on soil fertility in silvopastoral pastures. This study aims to quantify the effects of trees on soil nutrient and C stocks, as well as assessing differences on the effects between legume (Albizia saman; Enterolobium cyclocarpum) and non-legume tree species (Tabebuia rosea; Guazuma ulmifolia).

Methods

In Central Nicaragua, soil was sampled (0–10 cm deep) in paired plots, under both a canopy and in open grassland, in 12 sites per tree species and analysed for organic C, total N stocks, available P and extractable K⁺, Ca²⁺ and Mg²⁺. To assess the effects of herbaceous composition and cattle to soil proprieties, we recorded the cover of plant groups and assessed the mass of dung in each plot.

Results

Soil organic C and N, available P and extractable K⁺ and Ca²⁺ were higher under the tree canopy than under paired open grassland. The basal area of trees was positively related with the canopy effect on soil variables, thus suggesting that the age or sizes of the trees are relevant factors associated with the content of soil C and nutrients. No specific effects related to the legume species group were detected.

Conclusions

Our results indicate that in fertile seasonally dry subtropical pastures, scattered trees have an overall effect on soil fertility, and that the magnitude of the effect depends more on the tree characteristics (i.e. basal area, crown area) than on whether the species is a legume or not.     

Carbon and nutrient limitation of soil microorganisms and microbial grazers in a tropical montane rain forest

We investigated the role of carbon, nitrogen and phosphorus as limiting factors of microorganisms and microbial grazers (testate amoebae) in a montane tropical rain forest in southern Ecuador. Carbon (as glucose), nitrogen (as NH₄NO₃) and phosphorus (as NaH₂PO₄) were added separately and in combination bimonthly to experimental plots for 20 months. By adding glucose and nutrients we expected to increase the growth of microorganisms as the major food resource of testate amoebae. The response of microorganisms to experimental treatments was determined by analysing microbial biomass (SIR), fungal biomass and microbial community composition as measured by phospholipid fatty acids (PLFAs). We hypothesized that the response of testate amoebae is closely linked to that of microorganisms. Carbon addition strongly increased ergosterol concentration and, less pronounced, the amount of linoleic acid as fungal biomarker, suggesting that saprotrophic fungi are limited by carbon. Microbial biomass and ergosterol concentrations reached a maximum in the combined treatment with C, N and P indicating that both N and P also were in short supply. In contrast to saprotrophic fungi and microorganisms in total, testate amoebae suffered from the addition of C and reached maximum density by the addition of N. The results indicate that saprotrophic fungi in tropical montane rain forests are mainly limited by carbon whereas gram positive and negative bacteria benefit from increased availability of P. Testate amoebae suffered from increased dominance of saprotrophic fungi in glucose treatments but benefited from increased supply of N. The results show that testate amoebae of tropical montane rain forests are controlled by bottom–up forces relying on specific food resources rather than the amount of bacterial biomass with saprotrophic fungi functioning as major antagonists. Compared to temperate systems microbial food webs in tropical forests therefore may be much more complex than previously assumed with trophic links being rather specific and antagonistic interactions overriding trophic interactions.     

热带森林植物功能群及其动态研究进展

热带森林极高的物种多样性和结构复杂性给生态学研究带来了很多挑战。植物功能群是对特定环境响应相似或对主要的生态过程具有相似作用的物种组合。应用植物功能群的方法是有效减少热带森林群落复杂性,并揭示其格局和过程的良好途径。在介绍植物功能群概念和划分途径的基础上,分析了热带森林植物功能群的时空动态规律。一般来讲,划分植物功能群通常有3种途径,并可通过5个步骤来完成。热带森林植物功能群的空间分布常受景观格局的制约,而环境异质性往往是影响不同植物功能群组配比例变化的直接原因。不同类型的植物功能群随演替过程发生显著的替代,而干扰体系和全球气候变化对功能群的动态过程具有重要的驱动作用。以功能群为基础的动态模型在模拟热带林群落动态和预测植被潜在分布等方面具有广阔的发展前景。探索有效的植物功能分类方法、构建完善的植物功能性状数据库、开发更为精确的功能群动态模型以及加强基于景观水平的植物功能群动态机制的认识等是未来热带森林植物功能群及其动态研究的重要方向。     

Priming and microbial nutrient limitation in lowland tropical forest soils of contrasting fertility

Priming is an increase in soil organic carbon decomposition following input of labile organic carbon. In temperate soils where biological activity is limited commonly by nitrogen availability, priming is expected to occur through microbial acquisition of nitrogen from organic matter or stimulated activity of recalcitrant-carbon degrading microorganisms. However, these priming mechanisms have not yet been assessed in strongly weathered tropical forest soils where biological activity is often limited by the availability of phosphorus. We examined whether microbial nutrient limitation or community dynamics drive priming in three lowland tropical forest soils of contrasting fertility (‘low’, ‘mid’ and ‘high’) by applying C₄-sucrose (alone or in combination with nutrients; nitrogen, phosphorus and potassium) and measuring (1) the δ¹³C-signatures in respired CO₂ and in phospholipid fatty acid (PLFA) biomarkers, and (2) the activities of enzymes involved in nitrogen (N-acetyl β-glucosaminidase), phosphorus (phosphomonoesterase) and carbon (β-glucosidase, cellobiohydrolase, xylanase, phenol oxidase) acquisition from organic compounds. Priming was constrained in part by nutrient availability, because priming was greater when sucrose was added alone compared to when added with nutrients. However, the greatest priming with sucrose addition alone was detected in the medium fertility soil. Priming occurred in parallel with stimulated activity of phosphomonoesterase and phenol oxidase (but not N-acetyl β-glucosaminidase); when sucrose was added with nutrients there were lower activities of phosphomonoesterase and phenol oxidase. There was no evidence according to PLFA δ¹³C-incorporation that priming was caused by specific groups of recalcitrant-carbon degrading microorganisms. We conclude that priming occurred in the intermediate fertility soil following microbial mineralization of organic nutrients (phosphorus in particular) and suggest that priming was constrained in the high fertility soil by high nutrient availability and in the low fertility soil by the low concentration of soil organic matter amenable to priming. This first study of priming mechanisms in tropical forest soils indicates that input of labile carbon can result in priming by microbial mineralization of organic nutrients, which has important implications for understanding the fate of organic carbon in tropical forest soils.     

Moderate changes in nutrient input alter tropical microbial and protist communities and belowground linkages

We investigated the response of soil microbial communities in tropical ecosystems to increased nutrient deposition, such as predicted by anthropogenic change scenarios. Moderate amounts of nitrogen and phosphorus and their combination were added along an altitudinal transect. We expected microorganisms and microbial grazers (testate amoebae) to significantly respond to nutrient additions with the effect increasing with increasing altitude and with duration of nutrient additions. Further, we expected nutrients to alter grazer–prey interrelationships. Indeed, nutrient additions strongly altered microbial biomass (MB) and community structure as well as the community structure of testate amoebae. The response of microorganisms varied with both altitude and duration of nutrient addition. The results indicate that microorganisms are generally limited by N, but saprotrophic fungi also by P. Also, arbuscular mycorrhizal fungi benefited from N and/or P addition. Parallel to MB, testate amoebae benefited from the addition of N but were detrimentally affected by P, with the addition of P negating the positive effect of N. Our data suggests that testate amoeba communities are predominantly structured by abiotic factors and by antagonistic interactions with other microorganisms, in particular mycorrhizal fungi, rather than by the availability of prey. Overall, the results suggest that the decomposer system of tropical montane rainforests significantly responds to even moderate changes in nutrient inputs with the potential to cause major ramifications of the whole ecosystem including litter decomposition and plant growth.     

Plant litter quality regulates soil eco-enzymatic stoichiometry and microbial nutrient limitation in a citrus orchard

Plant and Soil - The impacts associated with litter decomposition in intensive cropping on eco-enzymatic stoichiometry and microbial nutrient limitation are still under debate. To evaluate the...     

Silicon controls microbial decay and nutrient release of grass litter during aquatic decomposition

The decomposition rate of plant litter is important for the carbon cycle. Element stoichiometry and hardly degradable carbon compounds are main factors controlling the decomposition rate of plant litter. Recent research has linked these factors to silicon availability during plant growth, but no research focused on the effect of silicon on litter decomposition. We therefore conducted a batch experiment to assess the effect of silicon availability to plants on litter degradation, nutrient release and multi elemental stoichiometry. Experiments were conducted in the presence or absence of invertebrate shredders (Gammarus pulex). We show that nutrient content (affected by silicon availability during plant growth) has a strong impact on nutrient turnover, while DOC, N, and Mn were mainly controlled by invertebrate feeding. The carbon turnover during microbial litter decay was strongly influenced by the silicon availability during plant growth, with quicker potential C turnover of litter with higher silicon content. In both Si-rich and Si-poor litter, feeding by invertebrate shredders positively impacted turnover rates, but effects were less pronounced in Si-rich litter. It can be concluded that silicon availability in wetlands dominated by reed plays an important role in carbon sequestration, nutrient cycling, and remobilization during aquatic litter decay.     

Broussonetia papyrifera controls nutrient return to soil to facilitate its invasion in a tropical forest of Ghana

Aims Non-native invasive plants can alter soil chemistry through litter production and decomposition to facilitate their invasion. However, the important roles of these underlying processes in plant invasion remain poorly understood, particularly in tropical forest ecosystems. Here, we compared litter production, quality and decomposition of two invasive species (Broussonetia papyriferaandCedrela odorata) and two co-occurring native species (CeltismildbraediiandFuntumia elastica), and soil properties under them to elucidate their roles in the invasion of a tropical forest in Ghana.     

Plant functional group richness-affected microbial community structure and function in a full-scale constructed wetland

We examined the linkages between plant functional group richness and microbial community structure and functions in a full-scale vertical flow constructed wetland (VFCW), where five plant functional group richness levels (including zero (control), one, two, three and four functional groups) were applied. Most diagnostic phospholipid fatty acid (PLFA) abundances and enzyme activities were higher in planted treatments than in the control (p < 0.05). Among the diagnostic PLFAs determined, only the fungal PLFA (18:2ω6,9) abundance was related to plant functional group richness level (p < 0.05). For the enzyme activities determined, dehydrogenase, invertase, urease and acid phosphatase activities responded positively to plant functional group richness (p < 0.05). Redundancy analysis (RDA) identified that the Gram-positive and Gram-negative bacteria, actinomycetes and fungi abundances affected enzyme activities. Principal component analyses (PCAs) revealed that the enzyme profiles had greater resolving power in distinguishing plant functional group richness than the PLFA profiles. We conclude that plant functional group richness was closely linked to microbial community structure and functions in the full-scale VFCW we studied.     

Groundwater controls on nutrient cycling in a Mojave desert stream

1. Groundwater fluxes of nitrogen and dissolved organic carbon (DOC) were investigated in Grape Vine Canyon Stream in the Mojave Desert focusing on the rate of inputs and the fate of groundwater-derived nutrients in the stream. Discharge rates from different ground waters were measured using an end-member mixing model coupled with injections of a conservative solute tracer into the stream channel.
2. In surface water, nitrate concentration averaged 1.13 mg N L^–1 and DOC concentration averaged 1.82 mg C L^–1.
3. Groundwater discharge into Grape Vine Canyon Stream was derived from three sources. Nitrate concentration varied among the three groundwater sources with mean concentrations of 0.56, 0.94 and 0.08 mg N L^–1. DOC, in contrast, did not vary among ground water sources, with an overall average concentration of 2.96 mg C L^–1.
4. In the surface stream, nitrate concentration was two-fold greater than the concentration predicted from groundwater input, indicating that in-stream processes generated nitrate. Stream DOC concentration was lower than predicted based upon groundwater input rate. The production of nitrate and loss of DOC suggest that DOC is lost through mineralisation of dissolved organic matter, possibly resulting in the mineralisation of dissolved organic nitrogen to ammonium and subsequent transformation to nitrate via nitrification. In further support of this hypothesised linkage, DOC loss explained 80–89% of the variance in nitrate production in Grape Vine Canyon Stream.     

Plant biomass and nutrient levels of a tropical macrophyte (Cyperus papyrus L.) receiving domestic wastewater

The aboveground biomass and nutrient content ofCyperus papyrus were determined in a small tropical swamp receiving domestic wastewater. It was found that the biomass (4,955 g.m^–2 dry weight) was the highest ever reported for papyrus. The levels of both nitrogen and phosphorus in the plant organs were very high. The nitrogen concentrations of the various plant organs were 4.8% roots, 8.4% rhizomes, 4.5% scales, 4.8% culms, and 6.2% umbels on dry weight basis. As to phosphorus the concentrations were 0.09% roots, 0.11% rhizomes, 0.09% scales, 0.10% culms, and 0.13% umbels. The high biomass and nutrient contents of the plants may have been caused by the high nutrient levels in the surrounding water. Comparison of the nitrogen to phosphorus ratios in the plants to those in the surrounding water showed that the plants stored very high amounts of nitrogen.     

Plant functional traits with particular reference to tropical deciduous forests: a review

Functional traits (FTs) integrate the ecological and evolutionary history of a species, and can potentially be used to predict its response as well as its influence on ecosystem functioning. Study of inter-specific variation in the FTs of plants aids in classifying species into plant functional types (PFTs) and provides insights into fundamental patterns and trade-offs in plant form and functioning and the effect of changing species composition on ecosystem functions. Specifically, this paper focuses on those FTs that make a species successful in the dry tropical environment. Following a brief overview, we discuss plant FTs that may be particularly relevant to tropical deciduous forests (TDFs). We consider the traits under the following categories: leaf traits, stem and root traits, reproductive traits, and traits particularly relevant to water availability. We compile quantitative information on functional traits of dry tropical forest species. We also discuss trait-based grouping of plants into PFTs. We recognize that there is incomplete knowledge about many FTs and their effects on TDFs and point out the need for further research on PFTs of TDF species, which can enable prediction of the dynamics of these forests in the face of disturbance and global climate change. Correlations between structural and ecophysiological traits and ecosystem functioning should also be established which could make it possible to generate predictions of changes in ecosystem services from changes in functional composition.     

Plant functional assembly is mediated by rainfall and soil conditions in a seasonally dry tropical forest

Understanding how species assembly is influenced by the interplay of climate, local environmental conditions and human-caused disturbances remains a central question in ecology and conservation. Here, we assess how plant species abundance is determined by combinations of functional traits (ecological strategies) and interacting gradients of rainfall, soil conditions (fertility and field capacity) and chronic anthropogenic disturbance in a Caatinga dry tropical forest, Brazil. We tested for trait–environment relationships using multivariate methods (RLQ) accounting for groups of species sharing similar responses to gradients and similar expression of multiple traits (i.e. response groups). Overall, species’ abundances changed predictably in response to rainfall and soil fertility, and were mediated by functional traits, i.e. species with particular trait combinations tended to respond similarly to multifactorial conditions. Briefly, three ecological strategies emerged: species with low wood density and soft (i.e. lower dry matter content), thick leaves converged into a trait syndrome characterizing a drought-avoidance strategy through water storage. They were particularly abundant under extremely low precipitation and relatively high soil field capacity. Under conditions of increasing rainfall and decreasing soil field capacity, species with high wood density were favored, consistent with a drought-tolerance strategy. However, these species fell into two groups relative to leaf-investment: more conservative leaves (low SLA) on relatively fertile soils vs. thinner and softer (i.e. high SLA) leaves on unfertile soils. In seasonally dry tropical forests, low SLA on relatively fertile soils may represent a water conservation strategy. Unexpectedly, no ecological strategy emerged in response to disturbance. The patterns we uncovered help to understand the interplay between precipitation, soil fertility and anthropogenic disturbance in plant species filtering in seasonally dry tropical forests. Moreover, our results underline that impacts of future climate change will depend on how rainfall patterns covary with finer-scale environmental factors such as soil fertility and field capacity.     

Plant functional traits and soil microbial biomass in different vegetation zones on the Loess Plateau

Plant functional traits built the relationships between plant diversity, species composition, and physiology along with the environmental changes, thus influencing soil microbial community. As the sensitivity indicators, soil microbial biomass and plant functional traits responses soil micro-organism and plant characteristics in direct way. Ten plant functional traits of 149 species and soil microbial biomass (carbon, nitrogen, and phosphorus) were analyzed across the different vegetation types (forest, forest-steppe, and steppe) that are divided by environmental gradient (temperature and precipitation), aimed to find the correlations among them. Our results confirmed the greatest values of plant functional traits (except the leaf density and the fine root density) that were distributed in the steppe zone, mainly due to the different mean annual temperature and mean annual precipitation conditions. For different plant growth forms, the plant functional traits were significant differences among the vegetation zones. The advantages of higher rate nutrient cycling, plentiful biomass supplements, and favorite habit conditions lead to the forest-steppe zone with the highest C_mic and N_mic concentrations. The canonical correlation analysis indicated that leaf nitrogen, root nitrogen, and fine root densities were correlated with root exudate and tissue which affected the concentrations of soil organic carbon (SOC) and total nitrogen (N), consequently impacting soil microbial biomass carbon (C_mic) and soil microbial biomass nitrogen (N_mic). Soil is the medium that connects micro-organism and plant root system that influenced leaf nitrogen, root nitrogen, and fine root density of plant functional traits, the concentrations of SOC and total N that plant feedback are consequently influencing C_mic and N_mic.     

Soil and species effects on bark nutrient storage in a premontane tropical forest

Plant and Soil - The phylogenetic variations of fine root traits, which are related to plant growth and development as well as to physiological and ecological processes, are not fully understood....     

Plant functional group responses in an African tropical forest recovering from disturbance

Background: Newly protected, tropical forests recovering from logging and clearance are increasingly important targets for conservation. Recovery is typically evaluated or monitored by using a few easily identified species groups, but this may not be sufficient as unmonitored groups of equivalent ecological importance are unlikely to respond in a similar manner due to physiological or dispersal differences.

Aims: We compared four groups of plants: large trees, shrubs and saplings, herbs and ferns in a forest recovering from disturbance. We quantified the relative importance of disturbance history and local environmental conditions in determining species richness and community composition of these groups and assessed whether the groups could act as surrogate indicators for one another.

Methods: Vegetation was surveyed on a gradient of disturbance intensity and recovery time (20–60 years) in Mabira Forest, Uganda. We looked for correlations between species groups in richness or composition across sites and used constrained ordinations to identify important environmental variables.

Results: Neither species richness nor composition patterns were correlated between groups. All groups were weakly related to disturbance history; trees and shrubs were related to soil, herbs to microhabitat and ferns to microclimate.

Conclusions: No group acted as an indicator for another. The relative influence of disturbance and local environmental effects varied in the manner predicted, with disturbance stronger for trees and environment stronger for ferns.     

Trophic functioning and nutrient flux in a highly productive tropical lagoon

Chiku Lagoon is a highly productive tropical lagoon with high fishery yields. Trophic networks and stoichiometrically linked water-salt-nutrient budgets were constructed to relate the functioning of the food web to nonconservative behavior of nutrients in the lagoon. Network analysis showed that the lagoon is more dependent on phytoplankton than detritus and periphyton to generate food sources for consumers. Nevertheless, detritivory is more important than herbivory in the food web. Transfer efficiency is high at low trophic levels, but declines at higher levels due to the high fishery pressure. Thus, only a small fraction of organic matter (15%) is recycled, and this all through detrital pathways, most of which involve only two compartments. Summation of individual rate measurements for primary production and respiration yielded an estimate of +249 g C m^-2 year^-1, suggesting an autotrophic ecosystem. An alternative biogeochemical approach demonstrated that the lagoon is a large sink for total dissolved nitrogen and phosphorus, and the net system metabolism was calculated to be +144 g C m^-2 year^-1, thus providing a biogeochemical explanation for the high productivity of Chiku Lagoon. Our results suggest that the high fishery yield in Chiku Lagoon can be attributed to high planktonic productivity induced by the high rate of nutrient loading, and the straight-through pathways of the food web.