Effect of carbon addition and predation on acetate-assimilating bacterial cells in groundwater

Groundwater microbial community dynamics are poorly understood due to the challenges associated with accessing subsurface environments. In particular, microbial interactions and their impact on the subsurface carbon cycle remain unclear. In the present project, stable isotope probing with uniformly labeled [¹³C]-acetate was used to identify metabolically active and inactive bacterial populations based on their ability to assimilate acetate and/or its metabolites. Furthermore, we assessed whether substrate availability (bottom–up control) or grazing mortality (top–down control) played a greater role in shaping bacterial community composition by separately manipulating the organic carbon supply and the protozoan grazer population. A community fingerprinting technique, terminal restriction fragment length polymorphism, revealed that the bacterial community was not affected by changes in acetate availability but was significantly altered by the removal of protozoan grazers. In silico identification of terminal restriction fragments and 16S rRNA gene sequences from clone libraries revealed a bacterial community dominated by Proteobacteria, Firmicutes , and Bacteroidetes . Elucidation of the factors that structure the bacterial community will improve our understanding of the bacterial role in the carbon cycle of this important subterranean environment.     

Microbial Community Structures in Terrestrial Subsurface Sediments from the Southern Kanto Plain,Japan

Abstract

Microbial community structure reflects the surrounding natural environment and changes to that environment. Although the subsurface at 5–100?m depth is important for human activities and there are potential risks of environmental pollution in this region, there have been only a few reports of subsurface microbial community structures in terrestrial areas. We investigated the diversity and community compositions of Bacteria and Archaea in boring cores collected from various depths at three different sites in the southern Kanto Plain, Japan. The results of 16S rRNA gene amplicon sequencing using MiSeq showed that the microbial community composition varied with the geological unit. Proteobacteria (Alphaproteobacteria and Gammaproteobacteria) were dominant members within sediments accumulated during the Pleistocene in the Musashino Upland. In contrast, Acidobacteria and Chloroflexi characteristically appeared in the Holocene layers of the Arakawa Lowland. These data suggest that the subsurface microbial composition is controlled by the geological features of the sediments.     

Changes in Prokaryote and Eukaryote Assemblages Along a Gradient of Hydrocarbon Contamination in Groundwater

Groundwater biota are particularly sensitive to environmental perturbations such as groundwater contamination. The diversity of prokaryotic and eukaryotic biota has been examined along a gradient of chlorinated hydrocarbon (CHC) contamination in the Botany Sands, an urban coastal sand-bed aquifer (Sydney, Australia). Molecular techniques were used to analyze the richness and composition of prokaryote and eukaryote assemblages using 16S and 18S rDNA, respectively. Taxon richness did not change significantly along the gradient for either prokaryotes or eukaryotes; however, significant shifts in assemblage composition were evident for both groups. Assemblage changes were most strongly correlated with concentrations of the major CHC, cis-1,2-dichloroethene, but the concentrations of a number of the contaminants were also correlated, making it difficult to infer if effects were due to any particular contaminant. The presence of cis-1,2-dichloroethene and other secondary ethenes suggests in situ breakdown of the primary CHCs via natural attenuation. The current focus of management of the Botany aquifer is to stop the contaminant plume reaching the adjoining estuary. This approach is clearly justified given the changes evident in the microbial assemblages in the groundwater, which are a likely consequence of the contamination.     

Relating Microbial Community Structure and Geochemistry in Deep Regolith Developed on Volcaniclastic Rock in the Luquillo Mountains,Puerto Rico

Fe oxidation is often the first chemical reaction that initiates weathering and disaggregation of intact bedrock into regolith. Here we explore the use of pyrosequencing tools to test for evidence that bacteria participate in these reactions in deep regolith. We analyze regolith developed on volcaniclastic rocks of the Fajardo formation in a ridgetop within the rainforest of the Luquillo Mountains of Puerto Rico. In the 9-m-deep regolith profile, the primary minerals chlorite, feldspar, and pyroxene are detected near 8.3 m but weather to kaolinite and Fe oxides found at shallower depths. Over the regolith profile, both total and heterotrophic bacterial cell counts generally increase from the bedrock to the surface. Like other soil microbial studies, the dominant phyla detected are Proteobacteria, Acidobacteria, Planctomycetes, and Actinobacteria. Proteobacteria (α, β, γ and δ) were the most abundant at depth (6.8–9 m, 41–44%), while Acidobacteria were the most abundant at the surface (1.4–4.4 m, 37–43%). Despite the fact that Acidobacteria dominated surficial communities while Proteobacteria dominated near bedrock, the near-surface and near-bedrock communities were not statistically different in structure but were statistically different from mid-depth communities. Approximately 21% of all sequences analyzed did not match known sequences: the highest fraction of unmatched sequences was greatest at mid-depth (45% at 4.4 m). At the regolith-bedrock interface where weathering begins, several lines of evidence are consistent with biotic Fe oxidation. At that interface, iron-related bacterial activity tests and culturing indicate the presence of iron-related bacteria, and phylogenetic analyses identified sub-phyla containing known iron-oxidizing microorganisms. Cell densities of iron-oxidizers in the deep saprolite were estimated to be on the order of 10⁵ cells g^?1. Overall Fe loss was also observed at the regolith-bedrock interface, consistent with bacterial production of organic acids and leaching of Fe-organic complexes. Fe-organic species were also detected to be enriched near the bedrock-regolith interface. In this and other deep weathering profiles, chemolithoautotrophic bacteria that use Fe for energy and nitrate or oxygen as an electron acceptor may play an important role in initiating disaggregation of bedrock.     

Deep Subsurface Microbial Biomass and Community Structure in Witwatersrand Basin Mines

The extreme environments of South Africa mines were investigated to determine microbial community structure and biomass in the deep subsurface. These community parameters were determined using phospholipid fatty acid (PLFA) technique. Air, water and rock samples were collected from several levels and shafts in eight different mines. Biomass estimates ranged over nine orders of magnitude. Biofilm samples exhibited the highest biomass with quantities ranging from 10 ³ to 10 ⁷ pmol PLFA g ^?1 . Rock samples had biomass ranging from 10 ³ to 10 ⁶ pmol PLFA g ^?1 . Mine service waters and rock fracture waters had biomass estimates ranging from 10 ⁰ to 10 ⁶ pmol PLFA L ^?1 . Air samples biomass values ranged from 10 ^?2 to 10 ⁰ pmol PLFA L ^?1 . The biomass estimates were similar to those estimates for other deep subsurface sites. Redundancy analysis of the PLFA profiles distinguished between the sample types, where signature lipid biomarkers for aerobic and anaerobic prokaryotes, sulfate-and metal-reducing bacteria were associated with biofilms. Rock samples were enriched in 18:1 ω 9 c , 18:2 ω 6, br17:1s and br18:1s, which are indicative of microeukaryotes and metal- reducing bacteria. Air samples were enriched with 22:0, 17:1, 18:1, and a polyunsaturated fatty acid. Service waters had monounsaturated fatty acids. Fracture waters contained i17:0 and 10Me18:0 which indicated gram-positive and other anaerobic bacteria. When the fracture and service water sample PLFA responses to changes in environmental parameters of temperature, pH, and anion concentrations were analyzed, service waters correlated with higher nitrate and sulfate concentrations and the PLFAs 18:1 ω 7 c and 16:1 ω 7 c . Dreifontein shaft 5 samples correlated with chloride concentrations and terminally branched saturated fatty acids and branched monounsaturated fatty acids. Kloof, Tau Tona, and Merriespruit fracture waters aligned with temperature and pH vectors and 18:0, 20:0 and 22:6 ω 3. The redundancy analysis provided a robust method to understand the PLFA responses to changes in environmental parameters.     

Novel approach using substrate-mediated radiolabelling of RNA to link metabolic function with the structure of microbial communities

A novel concept was developed applying radioisotope-labelled substrate incorporation into the biomass. The resulting radiolabelled RNA was used both as an indicator of activity and as a template for gaining structural and functional information about a substrate-utilizing microbial community. Sequences of PCR products are separated via cloning or using molecular fingerprinting techniques. Nucleic acids from predominant clones or the whole molecular fingerprinting pattern are transferred to a membrane and hybridized with the radiolabelled sample RNA. Scanning of the hybridized blots for radioactivity indicates the members involved in the utilization of the substrate. This novel 'random walk' approach using radioisotope probing was evaluated in a model community experiment.     

Geochemistry and Microbial Populations in Sediments of the Northern Baffin Bay,Arctic

The Northern Baffin Bay between Greenland and Canada is a remote Arctic area restricted in primary production by seasonal ice cover, with presumably low sedimentation rates, carbon content and microbial activities in its sediments. Our aim was to study the so far unknown subseafloor geochemistry and microbial populations driving seafloor ecosystems. Shelf sediments had the highest organic carbon content, numbers of Bacteria and Archaea, and microcosms inoculated from Shelf sediments showed highest sulfate reduction and methane production rates. Sediments in the central deep area and on the southern slope contained less organic carbon and overall lower microbial numbers. Similar 16S rRNA gene copy numbers of Archaea and Bacteria were found for the majority of the sites investigated. Sulfate in pore water correlated with dsrA copy numbers of sulfate-reducing prokaryotes and differed between sites. No methane was found as free gas in the sediments, and mcrA copy numbers of methanogenic Archaea were low. Methanogenic and sulfate-reducing cultures were enriched on a variety of substrates including hydrocarbons. In summary, the Greenlandic shelf sediments contain vital microbial communities adapted to their specific environmental conditions.     

中美医学院病原生物学教学比较和借鉴

从课程设置、教学方法、教材建设和教学手段等方面对中国和美国高校病原生物学教学进行了比较和探讨.     

中美医学院病原生物学教学比较和借鉴

从课程设置、教学方法、教材建设和教学手段等方面对中国和美国高校病原生物学教学进行了比较和探讨。     

Geomicrobial Processes and Biodiversity in the Deep Terrestrial Subsurface

The concept of a deep microbial biosphere has advanced over the past several decades from a hypothesis viewed with considerable skepticism to being widely accepted. Phylogenetically diverse prokaryotes have been cultured from or detected via characterization of directly-extracted nucleic acids from a wide range of deep terrestrial environments. Recent advances have linked the metabolic potential of these microorganisms, determined directly or inferred from phylogeny, to biogeochemical reactions determined via geochemical measurements and modeling. Buried organic matter or kerogen is an important source of energy for sustaining anaerobic heterotrophic microbial communities in deep sediments and sedimentary rock although rates of respiration are among the slowest rates measured on the planet. In contrast, Subsurface Lithoautotrophic Microbial Ecosystems based on H ₂ as the primary energy source appear to dominate in many crystalline rock environments. These photosynthesis-independent ecosystems remain an enigma due to the difficulty in accessing and characterizing appropriate samples. Deep mines and dedicated rock laboratories, however, may offer unprecedented opportunities for investigating subsurface microbial communities and their interactions with the geosphere.     

Vertical and Horizontal Distribution of Bacterial Communities in Alluvial Groundwater of the Nakdong River Bank

As a collaborative effort to characterize a pilot test site for managed aquifer recharge (MAR), vertical and horizontal distributions of microbial communities in the river bank subsurface were investigated to assess the ecological effects after the operation of the MAR using the river water adjacent to the site. Along with a geochemical analysis, barcoded pyrosequencing was performed using the genomic DNAs extracted from the subsurface groundwater/sediment samples retrieved from three multilevel wells among the installed cluster of 14 boreholes. A total of 9 samples from 3 depths (10, 15–25, and 33 m below the ground surface) of each borehole showed higher bacterial abundance and diversity in the shallow (10 m) depths than in the deep (33 m) groundwater. In addition, there was a slight separation of the microbial communities between the depths based on the nonmetric multidimensional scaling analysis of the Yue and Clayton distance and the distance-weighted UniFrac analysis. The phylum Proteobacteria was dominant in all the samples at the sequence abundance of 64.0–97.8% with the total operational taxonomic units of 3375 at the species level, while among the total 288 genera, the genus Pseudomonas and an unclassified genus from Betaproteobacteria were the most abundant across the samples. The community separation between the shallow and the deep groundwater seemed to be correlated with depth differences, supported by differences in the dissolved oxygen (DO) concentration and oxidation-reduction potential (ORP). In the study site, unusually high values of electrical conductivity (EC) were found in the deep groundwater, but those values were unlikely to contribute to the community separation between the shallow and deep groundwater, unlike the DO and ORP values, which were found to influence the community differences.     

Identification of Metabolically Active Bacteria in the Gut of the Generalist Spodoptera littoralis via DNA Stable Isotope Probing Using 13C-Glucose

Guts of most insects are inhabited by complex communities of symbiotic nonpathogenic bacteria. Within such microbial communities it is possible to identify commensal or mutualistic bacteria species. The latter ones, have been observed to serve multiple functions to the insect, i.e. helping in insect reproduction¹, boosting the immune response², pheromone production³, as well as nutrition, including the synthesis of essential amino acids^4, among others.    Due to the importance of these associations, many efforts have been made to characterize the communities down to the individual members. However, most of these efforts were either based on cultivation methods or relied on the generation of 16S rRNA gene fragments which were sequenced for final identification. Unfortunately, these approaches only identified the bacterial species present in the gut and provided no information on the metabolic activity of the microorganisms.To characterize the metabolically active bacterial species in the gut of an insect, we used stable isotope probing (SIP) in vivo employing ¹³C-glucose as a universal substrate. This is a promising culture-free technique that allows the linkage of microbial phylogenies to their particular metabolic activity. This is possible by tracking stable, isotope labeled atoms from substrates into microbial biomarkers, such as DNA and RNA⁵. The incorporation of ¹³C isotopes into DNA increases the density of the labeled DNA compared to the unlabeled (¹²C) one. In the end, the ¹³C-labeled DNA or RNA is separated by density-gradient ultracentrifugation from the ¹²C-unlabeled similar one⁶. Subsequent molecular analysis of the separated nucleic acid isotopomers provides the connection between metabolic activity and identity of the species.Here, we present the protocol used to characterize the metabolically active bacteria in the gut of a generalist insect (our model system), Spodoptera littoralis (Lepidoptera, Noctuidae). The phylogenetic analysis of the DNA was done using pyrosequencing, which allowed high resolution and precision in the identification of insect gut bacterial community. As main substrate, ¹³C-labeled glucose was used in the experiments. The substrate was fed to the insects using an artificial diet.     

Sr Isotope Tracing of Differences in the Effects of Microorganisms on Weathering of Calcite and Apatite

To study differences in the effects of microorganisms on weathering of calcite and apatite, one strain of Aspergillus niger (A. niger) and one strain of Penicillium glaucum (P. glaucum), which respectively contain the mixture of calcite and apatite were cultivated for 24 days in the sucrose-potato culture medium, supernatant was taken every three days from the culture medium, followed by the determination of Ca²⁺ and Sr²⁺ contents and Sr isotopic ratios. The results of measurement showed that the Sr isotope ratios in the supernatant from the culture medium are intermediate between those of the end-member constituents calcite and apatite (0.70721-0.70861). Results of isotope mixing equations to calculation showed that in the first 15 days A. niger played a dominant role in weathering of calcite in the apatite/calcite mixture. The contribution rate of apatite for Ca²⁺ in the solution increased from 39.0% on the 18th day to 61.6% on the 24th day; P. glaucum played a key role in weathering of apatite in the first 3 days. Ions dissolved from apatite account for 73.9% of the total. It is known from the results of Sr isotope tracing that in the prior period of fungus cultivation A. niger plays a key role in weathering of calcite while P. glaucum plays a key role in weathering of apatite. The ability of P. glaucum to weather calcite tends to intensify progressively over time. Therefore, Sr isotope tracing can be used to accurately recognize differences in the effects of microorganisms on weathering of minerals.     

The phylogenetic composition and structure of soil microbial communities shifts in response to elevated carbon dioxide

One of the major factors associated with global change is the ever-increasing concentration of atmospheric CO₂. Although the stimulating effects of elevated CO₂ (eCO₂) on plant growth and primary productivity have been established, its impacts on the diversity and function of soil microbial communities are poorly understood. In this study, phylogenetic microarrays (PhyloChip) were used to comprehensively survey the richness, composition and structure of soil microbial communities in a grassland experiment subjected to two CO₂ conditions (ambient, 368 p.p.m., versus elevated, 560 p.p.m.) for 10 years. The richness based on the detected number of operational taxonomic units (OTUs) significantly decreased under eCO₂. PhyloChip detected 2269 OTUs derived from 45 phyla (including two from Archaea), 55 classes, 99 orders, 164 families and 190 subfamilies. Also, the signal intensity of five phyla (Crenarchaeota, Chloroflexi, OP10, OP9/JS1, Verrucomicrobia) significantly decreased at eCO₂, and such significant effects of eCO₂ on microbial composition were also observed at the class or lower taxonomic levels for most abundant phyla, such as Proteobacteria, Firmicutes, Actinobacteria, Bacteroidetes and Acidobacteria, suggesting a shift in microbial community composition at eCO₂. Additionally, statistical analyses showed that the overall taxonomic structure of soil microbial communities was altered at eCO₂. Mantel tests indicated that such changes in species richness, composition and structure of soil microbial communities were closely correlated with soil and plant properties. This study provides insights into our understanding of shifts in the richness, composition and structure of soil microbial communities under eCO₂ and environmental factors shaping the microbial community structure.     

Differences in the soil microbial community and carbon‐use efficiency following development of Vochysia guatemalensis tree plantations in unproductive pastures in Costa Rica

This study shows that Vochysia guatemalensis tree plantations were associated with enhanced soil biotic and abiotic characteristics in previously cleared forests in the northern zone of Costa Rica, suggesting the possible use of this practice as a restoration strategy for local land owners. Soil samples from a primary forest, secondary forest, and a 13‐year‐old plantation of V. guatemalensis had greater relative abundances of DNA sequences of microbial genera critical for carbon‐use (C‐use) efficiency (i.e. the saprobe, complex C and wood rot/lignin decomposer fungi, and bacterial lignin and other complex C degraders), and greater levels of total organic carbon, C‐biomass, and microbial quotients as indicators of enhanced C‐use efficiency, than found in soils of adjacent 5‐year‐old V. guatemalensis plantations and abandoned non‐productive pasture/grasslands (GRs). The major research conclusions were that (1) conversion of forested land into abandoned pasture/GRs decreased the C‐use efficiency in the soils and the microbial groups associated with C‐use efficiency; (2) soils in plantations of V. guatemalensis were associated with increased abundances of the DNA of these same microbial groups and enhanced C‐use efficiency; (3) DNA‐based taxonomic analysis of microbes and analysis of the microbial quotient values can be used to monitor soil ecosystems for assessment of the efficacy of restoration activities. Thus, planting V. guatemalensis on damaged lands in the Maquenque National Wildlife Refuge should be encouraged to provide a sustainable forestry crop that can be harvested rotationally, while improving soil ecosystem health and reducing the pressure to harvest other forest sites.     

The response of soil microorganisms and roots to elevated CO2 and temperature in a terrestrial model ecosystem

We investigate the response of soil microorganisms to atmospheric CO2 and temperature change within model terrestrial ecosystems in the Ecotron. The model communities consisted of four plant species (Cardamine hirsuta, Poa annua, Senecio vulgaris, Spergula arvensis), four herbivorous insect species (two aphids, a leaf-miner, and a whitefly) and their parasitoids, snails, earthworms, woodlice, soil-dwelling Collembola (springtails), nematodes and soil microorganisms (bacteria, fungi, mycorrhizae and Protista). In two successive experiments, the effects of elevated temperature (ambient plus 2 °C) at both ambient and elevated CO2 conditions (ambient plus 200 ppm) were investigated. A 40:60 sand:Surrey loam mixture with relatively low nutrient levels was used. Each experiment ran for 9 months and soil microbial biomass (Cmic and Nmic), soil microbial community (fungal and bacterial phospholipid fatty acids), basal respiration, and enzymes involved in the carbon cycling (xylanase, trehalase) were measured at depths of 0–2, 0–10 and 10–20 cm. In addition, root biomass and tissue C:N ratio were determined to provide information on the amount and quality of substrates for microbial growth.Elevated temperature under both ambient and elevated CO2 did not show consistent treatment effects. Elevation of air temperature at ambient CO2 induced an increase in Cmic of the 0–10 cm layer, while at elevated CO2 total phospholipid fatty acids (PLFA) increased after the third generation. The metabolic quotient qCO2 decreased at elevated temperature in the ambient CO2 run. Xylanase and trehalase showed no changes in both runs. Root biomass and C:N ratio were not influenced by elevated temperature in ambient CO2. In elevated CO2, however, elevated temperature reduced root biomass in the 0–10 cm and 30–40 cm layers and increased N content of roots in the deeper layers. The different response of root biomass and C:N ratio to elevated temperature may be caused by differences in the dynamics of root decomposition and/or in allocation patterns to coarse or fine roots (i.e. storage vs. resource capture functions). Overall, our data suggests that in soils of low nutrient availability, the effects of climate change on the soil microbial community and processes are likely to be minimal and largely unpredicatable.     

黄土高原石油污染土壤微生物群落结构及其代谢特征

针对污染胁迫下土壤微生物群落变化和代谢变异等问题,基于平板稀释法和Biolog微平板分析方法,研究了陕北黄土高原石油污染土壤微生物群落结构、代谢特征及其功能多样性。结果表明,不同类群的土壤微生物对石油污染胁迫的响应不同,污染土壤细菌和真菌数量高出清洁土壤1个数量级,而污染土壤的放线菌数量极显著减少(P0.01);污染土壤和清洁土壤微生物对糖类和多聚物类碳源较易利用,污染土壤微生物总体上代谢碳源的种类和活性均低于清洁土壤。微生物群落主成分分析(PCA)表明,石油污染土壤和清洁土壤的微生物群落存在显著差异(P0.01),起分异作用的碳源主要为糖类,其次是羧酸类和氨基酸类;随着土壤石油含量增加,典型变量值变异(离散)增大,土壤微生物群落结构稳定性降低。微生物群落多样性分析表明,Shannon丰富度指数(H)、McIntosh均一度指数(U)和Simpson优势度指数(1/D)均达到极显著差异(P0.01),污染土壤微生物群落H和U低于清洁土壤,但是一定浓度的石油污染可以刺激土壤微生物群落中优势种群的生长,1/D增高。研究结果为陕北黄土高原石油污染区土壤微生物修复提供理论基础。     

用ERIC-PCR结合分子杂交监测焦化废水处理系统(A2/O)中微生物群落结构的变化

建立一种不依赖纯培养 ,可以在废水处理工业现场使用的监测微生物群落结构变化的分子技术。以处理焦化工业废水(A2 /O生物膜工艺 )不同构筑物中的悬浮污泥的微生物群落为研究对象 ,每周采样 1次 ,连续 4周。获得悬浮污泥总 DNA的ERIC- PCR指纹图谱 ,结合分子杂交进一步区分相同条带间的不同序列信息。结果表明 ,在缺氧池 (A2池 )和好氧池 (O池 )之间 ,各个采样点的 ERIC- PCR图谱差异不大 ,悬浮污泥在各构筑物之间交流充分 ;同一采样点的图谱在不同采样时期具有明显差异 ,显示了在此期间微生物群落的连续动态变化过程。通过对生物膜系统中悬浮污泥的微生物群落结构的指纹图谱分析 ,可开发出对该系统微生物群落结构动态变化进行检测的技术     

Using the Stable Carbon and Nitrogen Isotope Compositions of Vervet Monkeys (Chlorocebus pygerythrus) to Examine Questions in Ethnoprimatology

This study seeks to understand how humans impact the dietary patterns of eight free-ranging vervet monkey (Chlorocebus pygerythrus) groups in South Africa using stable isotope analysis. Vervets are omnivores that exploit a wide range of habitats including those that have been anthropogenically-disturbed. As humans encroach upon nonhuman primate landscapes, human-nonhuman primate interconnections become increasingly common, which has led to the rise of the field of ethnoprimatology. To date, many ethnoprimatological studies have examined human-nonhuman primate associations largely in qualitative terms. By using stable carbon (δ¹³C) and nitrogen (δ¹⁵N) isotope analysis, we use quantitative data to understand the degree to which humans impact vervet monkey dietary patterns. Based on initial behavioral observations we placed the eight groups into three categories of anthropogenic disturbance (low, mid, and high). Using δ¹³C and δ¹⁵N values we estimated the degree to which each group and each anthropogenically-disturbed category was consuming C₄ plants (primarily sugar cane, corn, or processed foods incorporating these crops). δ¹³C values were significantly different between groups and categories of anthropogenic-disturbance. δ¹⁵N values were significantly different at the group level. The two vervet groups with the highest consumption of C₄ plants inhabited small nature reserves, appeared to interact with humans only sporadically, and were initially placed in the mid level of anthropogenic-disturbance. However, further behavioral observations revealed that the high δ¹³C values exhibited by these groups were linked to previously unseen raiding of C₄ crops. By revealing these cryptic feeding patterns, this study illustrates the utility of stable isotopes analysis for some ethnoprimatological questions.