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1.
Measurement of stress hormone metabolites in fecal samples has become a common method to assess physiological stress in wildlife populations. Glucocorticoid metabolite (GCM) measurements can be collected noninvasively, and studies relating this stress metric to anthropogenic disturbance are increasing. However, environmental characteristics (e.g., temperature) can alter measured GCM concentration when fecal samples cannot be collected immediately after defecation. This effect can confound efforts to separate environmental factors causing predeposition physiological stress in an individual from those acting on a fecal sample postdeposition. We used fecal samples from American pikas (Ochotona princeps) to examine the influence of environmental conditions on GCM concentration by (1) comparing GCM concentration measured in freshly collected control samples to those placed in natural habitats for timed exposure, and (2) relating GCM concentration in samples collected noninvasively throughout the western United States to local environmental characteristics measured before and after deposition. Our timed‐exposure trials clarified the spatial scale at which exposure to environmental factors postdeposition influences GCM concentration in pika feces. Also, fecal samples collected from occupied pika habitats throughout the species' range revealed significant relationships between GCM and metrics of climate during the postdeposition period (maximum temperature, minimum temperature, and precipitation during the month of sample collection). Conversely, we found no such relationships between GCM and metrics of climate during the predeposition period (prior to the month of sample collection). Together, these results indicate that noninvasive measurement of physiological stress in pikas across the western US may be confounded by climatic conditions in the postdeposition environment when samples cannot be collected immediately after defecation. Our results reiterate the importance of considering postdeposition environmental influences on this stress metric, especially in multiregional comparisons. However, measurements of fecal GCM concentration should prove useful for population monitoring within an eco‐region or when postdeposition exposure can be minimized.  相似文献   

2.
Data on the gut microbiota (GM) of wild animals are key to studies on evolutionary biology (host–GM interactions under natural selection), ecology and conservation biology (GM as a fitness component closely connected to the environment). Wildlife GM sampling often requires non-invasive techniques or sampling from dead animals. In a controlled experiment profiling microbial 16S rRNA in 52 house mice (Mus musculus) from eight families and four genetic backgrounds, we studied the effects of live- and snap-trapping on small mammal GM and evaluated the suitability of microbiota from non-fresh faeces as a proxy for caecal GM. We compared CM from individuals sampled 16–18 h after death with those in live traps and caged controls, and caecal and faecal GM collected from mice in live-traps. Sampling delay did not affect GM composition, validating data from fresh cadavers or snap-trapped animals. Animals trapped overnight displayed a slight but significant difference in GM composition to the caged controls, though the change only had negligible effect on GM diversity, composition and inter-individual divergence. Hence, the trapping process appears not to bias GM profiling. Despite their significant difference, caecal and faecal microbiota were correlated in composition and, to a lesser extent, diversity. Both showed congruent patterns of inter-individual divergence following the natural structure of the dataset. Thus, the faecal microbiome represents a good non-invasive proxy of the caecal microbiome, making it suitable for detecting biologically relevant patterns. However, care should be taken when analysing mixed datasets containing both faecal and caecal samples.Subject terms: Evolutionary genetics, Evolutionary ecology, Metagenomics  相似文献   

3.
In Japan, the three main crane species are the endangered red-crowned crane (Grus japonensis) inhabiting Hokkaido, the northernmost island of Japan; the vulnerable hooded crane (Grus monacha); and the vulnerable white-naped crane (Grus vipio). Both the hooded and white-naped cranes migrate in winter to Izumi in Kyushu, the southern island of Japan. In this study, we investigated the cranes and their coccidian parasites, through a targeted molecular approach using faecal DNA to develop a noninvasive method for infectious disease research. To determine the origin of noninvasively collected faecal samples, host species were identified by sequencing a region of approximately 470 bp of the mitochondrial 16S ribosomal RNA gene in the faecal DNA. Furthermore, to avoid sample redundancy, individual determination was performed by fragment analysis using microsatellite and sex-linked markers. For microsatellite genotyping, previously reported markers and markers isolated in this study were examined, and seven loci for red-crowned cranes, eight for hooded cranes and six for white-naped cranes displayed polymorphisms. A low error rate was demonstrated by comparing microsatellite data generated from faecal DNA samples with that generated from feather DNA samples, indicating a high reliability. Polymerase chain reaction-based capillary electrophoresis (PCR-CE), employing genetic markers in the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA, was employed to detect crane coccidia. The sensitivity of detection of PCR-CE using faecal DNA was inferior to that with traditional microscopy; however, our results suggest that PCR-CE can depict crane coccidia diversity with higher resolution and it is a useful tool to characterize community composition of coccidia in detail.  相似文献   

4.
Noninvasive sampling methods for studying intestinal microbiomes are widely applied in studies of endangered species and in those conducting temporal monitoring during manipulative experiments. Although existing studies show that noninvasive sampling methods among different taxa vary in their accuracy, no studies have yet been published comparing nonlethal sampling methods in adult amphibians. In this study, we compare microbiomes from two noninvasive sample types (faeces and cloacal swabs) to that of the large intestine in adult cane toads, Rhinella marina. We use 16S rRNA gene sequencing to investigate how microbial communities change along the digestive tract and which nonlethal sampling method better represents large intestinal microbiota. We found that cane toads' intestinal microbiota was dominated by Bacteroidetes, Proteobacteria and Firmicutes and, interestingly, we also saw a high proportion of Fusobacteria, which has previously been associated with marine species and changes in frog immunity. The large and small intestine of cane toads had a similar microbial composition, but the large intestine showed higher diversity. Our results indicate that cloacal swabs were more similar to large intestine samples than were faecal samples, and small intestine samples were significantly different from both nonlethal sample types. Our study provides valuable information for future investigations of the cane toad gut microbiome and validates the use of cloacal swabs as a nonlethal method to study changes in the large intestine microbiome. These data provide insights for future studies requiring nonlethal sampling of amphibian gut microbiota.  相似文献   

5.
Due to its conspicuous manifestations and its capacity to shape the configuration and dynamics of wild populations, territorial behavior has long intrigued ecologists. Territoriality and other animal interactions in situ have traditionally been studied via direct observations and telemetry. Here, we explore whether noninvasive genetic sampling, which is increasingly supplementing traditional field methods in ecological research, can reveal territorial behavior in an elusive carnivore, the wolverine (Gulo gulo). Using the locations of genotyped wolverine scat samples collected annually over a period of 12 years in central Norway, we test three predictions: (1) male home ranges constructed from noninvasive genetic sampling data are larger than those of females, (2) individuals avoid areas used by other conspecifics of the same sex (intrasexual territoriality), and (3) avoidance of same‐sex territories diminishes or disappears after the territory owner's death. Each of these predictions is substantiated by our results: sex‐specific differences in home range size and intrasexual territoriality in wolverine are patently reflected in the spatial and temporal configuration of noninvasively collected genetic samples. Our study confirms that wildlife monitoring programs can utilize the spatial information in noninvasive genetic sampling data to detect and quantify home ranges and social organization.  相似文献   

6.
The koala, an Australian icon, has been added to the threatened species list. Rationale for the listing includes proposed declines in population size, threats to populations (e.g. disease) and loss and fragmentation of habitat. There is now an urgent need to obtain accurate data to assess the status of koala populations in Australia, to ensure the long‐term viability of this species. Advances in genetic techniques have enabled DNA analysis to study and inform the management of wild populations; however, sampling of individual koalas is difficult in tall, often remote, eucalypt forest. The collection of faecal pellets (scats) from the forest floor presents an opportunistic sampling strategy, where DNA can be collected without capturing or even sighting an individual. Obtaining DNA via noninvasive sampling can be used to rapidly sample a large proportion of a population; however, DNA from noninvasively collected samples is often degraded. Factors influencing DNA quality and quantity include environmental exposure, diet and methods of sample collection, storage and DNA isolation. Reduced DNA quality and quantity can introduce genotyping errors and provide inaccurate DNA profiles, reducing confidence in the ability of such data to inform management/conservation strategies. Here, we present a protocol that produces a reliable individual koala genotype from a single faecal pellet and highlight the importance of optimizing DNA isolation and analysis for the species of interest. This method could readily be adapted for genetic studies of mammals other than koalas, particularly those whose diet contains high proportions of volatile materials that are likely to induce DNA damage.  相似文献   

7.
Population‐scale molecular studies of endangered and cryptic species are often limited by access to high‐quality samples. The use of noninvasively collected samples or museum‐preserved specimens reduces the pressure on modern populations by removing the need to capture and handle live animals. However, endogenous DNA content in such samples is low, making shotgun sequencing a financially prohibitive approach. Here, we apply a target enrichment method to retrieve mitochondrial genomes from 65 museum specimens and 56 noninvasively collected faecal samples of two endangered great ape species, Grauer's gorilla and the eastern chimpanzee. We show that the applied method is suitable for a wide range of sample types that differ in endogenous DNA content, increasing the proportion of target reads to over 300‐fold. By systematically evaluating biases introduced during target enrichment of pooled museum samples, we show that capture is less efficient for fragments shorter or longer than the baits, that the proportion of human contaminating reads increases postcapture although capture efficiency is lower for human compared to gorilla fragments with a gorilla‐generated bait, and that the rate of jumping PCR is considerable, but can be controlled for with a double‐barcoding approach. We succeed in capturing complete mitochondrial genomes from faecal samples, but observe reduced capture efficiency as sequence divergence increases between the bait and target species. As previously shown for museum specimens, we demonstrate here that mitochondrial genome capture from field‐collected faecal samples is a robust and reliable approach for population‐wide studies of nonmodel organisms.  相似文献   

8.
黑麂粪便DNA提取及其PCR检测   总被引:6,自引:1,他引:5  
采集了黑麂(Muntiacus crinifrons)的新鲜粪便以及在野外自然条件下保存较长时间的粪便样品,晾干后带回实验室,提取其DNA;同时提取黑麂肌肉、皮张样品的DNA,用以对比粪便样品的提取效果。电泳检测结果显示,此方法使用实验室中常用的分子生物学试剂,可以从黑麂粪便样品中抽提到高质量的粪便DNA并克服分子粪便学研究中常见的PCR反应抑制物的影响。为其它濒危鹿科动物的非损伤性取样提供了的新途径,为其遗传结构、遗传多样性现状等研究提供了更加广阔的取材空间。  相似文献   

9.
ABSTRACT Noninvasive scat sampling methods can generate large samples sizes, collected over vast landscapes, ideal for addressing wildlife conservation and management questions. However, the cost of genotyping scat samples limits the accessibility of these techniques. We describe detection-dog methods for matching large numbers of scat samples to the individual, reducing or eliminating the need for sample genotyping. Three dogs correctly matched 25 out of 28 samples from 6 captive maned wolves (Chrysocyon brachyurus) of known identity. Sample scent-matching can increase overall accessibility and breadth of applications of noninvasive scat-collection methods to important landscape scale problems in wildlife sciences.  相似文献   

10.
Gastrointestinal disturbances are a side-effect frequently associated with haematological malignancies due to the intensive cytotoxic treatment given in connection with bone marrow transplantation (BMT). However, intestinal microbiota changes during chemotherapy remain poorly described, probably due to the use of culture-based and low-resolution molecular methods in previous studies. The objective of our study was to apply a next generation DNA sequencing technology to analyse chemotherapy-induced changes in faecal microbiota. We included eight patients with non-Hodgkin’s lymphoma undergoing one course of BMT conditioning chemotherapy. We collected a prechemotherapy faecal sample, the day before chemotherapy was initiated, and a postchemotherapy sample, collected 1 week after the initiation of chemotherapy. Total DNA was extracted from faecal samples, denaturing high-performance liquid chromatography based on amplification of the V6 to V8 region of the 16S ribosomal RNA (rRNA) gene, and 454-pyrosequencing of the 16 S rRNA gene, using PCR primers targeting the V5 and V6 hypervariable 16S rRNA gene regions were performed. Raw sequence data were screened, trimmed, and filtered using the QIIME pipeline. We observed a steep reduction in alpha diversity and significant differences in the composition of the intestinal microbiota in response to chemotherapy. Chemotherapy was associated with a drastic drop in Faecalibacterium and accompanied by an increase of Escherichia. The chemotherapy-induced shift in the intestinal microbiota could induce severe side effects in immunocompromised cancer patients. Our study is a first step in identifying patients at risk for gastrointestinal disturbances and to promote strategies to prevent this drastic shift in intestinal microbiota.  相似文献   

11.
Target‐capture approach has improved over the past years, proving to be very efficient tool for selectively sequencing genetic regions of interest. These methods have also allowed the use of noninvasive samples such as faeces (characterized by their low quantity and quality of endogenous DNA) to be used in conservation genomic, evolution and population genetic studies. Here we aim to test different protocols and strategies for exome capture using the Roche SeqCap EZ Developer kit (57.5 Mb). First, we captured a complex pool of DNA libraries. Second, we assessed the influence of using more than one faecal sample, extract and/or library from the same individual, to evaluate its effect on the molecular complexity of the experiment. We validated our experiments with 18 chimpanzee faecal samples collected from two field sites as a part of the Pan African Programme: The Cultured Chimpanzee. Those two field sites are in Kibale National Park, Uganda (N = 9) and Loango National Park, Gabon (N = 9). We demonstrate that at least 16 libraries can be pooled, target enriched through hybridization, and sequenced allowing for the genotyping of 951,949 exome markers for population genetic analyses. Further, we observe that molecule richness, and thus, data acquisition, increase when using multiple libraries from the same extract or multiple extracts from the same sample. Finally, repeated captures significantly decrease the proportion of off‐target reads from 34.15% after one capture round to 7.83% after two capture rounds, supporting our conclusion that two rounds of target enrichment are advisable when using complex faecal samples.  相似文献   

12.
The gut microbiomes of birds and other animals are increasingly being studied in ecological and evolutionary contexts. Numerous studies on birds and reptiles have made inferences about gut microbiota using cloacal sampling; however, it is not known whether the bacterial community of the cloaca provides an accurate representation of the gut microbiome. We examined the accuracy with which cloacal swabs and faecal samples measure the microbiota in three different parts of the gastrointestinal tract (ileum, caecum, and colon) using a case study on juvenile ostriches, Struthio camelus, and high‐throughput 16S rRNA sequencing. We found that faeces were significantly better than cloacal swabs in representing the bacterial community of the colon. Cloacal samples had a higher abundance of Gammaproteobacteria and fewer Clostridia relative to the gut and faecal samples. However, both faecal and cloacal samples were poor representatives of the microbial communities in the caecum and ileum. Furthermore, the accuracy of each sampling method in measuring the abundance of different bacterial taxa was highly variable: Bacteroidetes was the most highly correlated phylum between all three gut sections and both methods, whereas Actinobacteria, for example, was only strongly correlated between faecal and colon samples. Based on our results, we recommend sampling faeces, whenever possible, as this sample type provides the most accurate assessment of the colon microbiome. The fact that neither sampling technique accurately portrayed the bacterial community of the ileum nor the caecum illustrates the difficulty in noninvasively monitoring gut bacteria located further up in the gastrointestinal tract. These results have important implications for the interpretation of avian gut microbiome studies.  相似文献   

13.
Microorganisms within the gastrointestinal tract significantly influence metabolic processes within their mammalian host, and recently several groups have sought to characterise the gastrointestinal microbiota of individuals affected by metabolic disease. Differences in the composition of the gastrointestinal microbiota have been reported in mouse models of type 2 diabetes mellitus, as well as in human patients. Diabetes mellitus in cats has many similarities to type 2 diabetes in humans. No studies of the gastrointestinal microbiota of diabetic cats have been previously published. The objectives of this study were to compare the composition of the faecal microbiota of diabetic and non-diabetic cats, and secondarily to determine if host signalment and dietary factors influence the composition of the faecal microbiota in cats. Faecal samples were collected from insulin-treated diabetic and non-diabetic cats, and Illumina sequencing of the 16S rRNA gene and quantitative PCR were performed on each sample. ANOSIM based on the unweighted UniFrac distance metric identified no difference in the composition of the faecal microbiota between diabetic and non-diabetic cats, and no significant differences in the proportions of dominant bacteria by phylum, class, order, family or genus as determined by 16S rRNA gene sequencing were identified between diabetic and non-diabetic cats. qPCR identified a decrease in Faecalibacterium spp. in cats aged over ten years. Cat breed or gender, dietary carbohydrate, protein or fat content, and dietary formulation (wet versus dry food) did not affect the composition of the faecal microbiota. In conclusion, the composition of the faecal microbiota was not altered by the presence of diabetes mellitus in cats. Additional studies that compare the functional products of the microbiota in diabetic and non-diabetic cats are warranted to further investigate the potential impact of the gastrointestinal microbiota on metabolic diseases such as diabetes mellitus in cats.  相似文献   

14.
Vervet monkeys, Chlorocebus pygerythrus, thrive in urban areas of KwaZulu-Natal, South Africa, and present a suitable model to assess parasitic load as a measure of anthropogenic disturbance, such as urbanization. We collected vervet monkey faecal samples from four study sites representing a gradient of land use and urbanization. We assessed faecal parasites using the faecal flotation method calculating eggs per gram and parasite richness. Overall, the more urban vervet monkey populations had a significantly higher parasite richness and abundance. Our study shows the applicability of using parasite load to measure the effect of urbanization on wildlife.  相似文献   

15.
Fecal DNA collected noninvasively can provide valuable information about genetic and ecological characteristics. This approach has rarely been used for equids, despite the need for conservation of endangered species and management of abundant feral populations. We examined factors affecting the efficacy of using equid fecal samples for conservation genetics. First, we evaluated two fecal collection methods (paper bag vs. ethanol). Then, we investigated how time since deposition and month of collection impacted microsatellite amplification success and genotyping errors. Between May and November 2014, we collected feral horse fecal samples of known age each month in a feral horse Herd Management Area in western Colorado and documented deterioration in the field with photographs. Samples collected and dried in paper bags had significantly higher amplification rates than those collected and stored in ethanol. There was little difference in the number of loci that amplified per sample between fresh fecal piles and those that had been exposed to the environment for up to 2 months (in samples collected in paper bags). After 2 months of exposure, amplification success declined. When comparing fresh (0–2 months) and old (3–6 months) fecal piles, samples from fresh piles had more matching genotypes across samples, better amplification success and less allelic dropout. Samples defecated during the summer and collected within 2 months of deposition had highest number of genotypes matching among samples, and lowest rates of amplification failure and allelic dropout. Due to the digestive system and amount of fecal material produced by equids, as well as their occurrence in arid ecosystems, we suggest that they are particularly good candidates for noninvasive sampling using fecal DNA.  相似文献   

16.
Knowledge of population demographics is important for species management but can be challenging in low‐density, wide‐ranging species. Population monitoring of the endangered Sonoran pronghorn (Antilocapra americana sonoriensis) is critical for assessing the success of recovery efforts, and noninvasive DNA sampling (NDS) could be more cost‐effective and less intrusive than traditional methods. We evaluated faecal pellet deposition rates and faecal DNA degradation rates to maximize sampling efficiency for DNA‐based mark–recapture analyses. Deposition data were collected at five watering holes using sampling intervals of 1–7 days and averaged one pellet pile per pronghorn per day. To evaluate nuclear DNA (nDNA) degradation, 20 faecal samples were exposed to local environmental conditions and sampled at eight time points from one to 124 days. Average amplification success rates for six nDNA microsatellite loci were 81% for samples on day one, 63% by day seven, 2% by day 14 and 0% by day 60. We evaluated the efficiency of different sampling intervals (1–10 days) by estimating the number of successful samples, success rate of individual identification and laboratory costs per successful sample. Cost per successful sample increased and success and efficiency declined as the sampling interval increased. Results indicate NDS of faecal pellets is a feasible method for individual identification, population estimation and demographic monitoring of Sonoran pronghorn. We recommend collecting samples >7 days old and estimate that a sampling interval of 4–7 days in summer conditions (i.e. extreme heat and exposure to UV light) will achieve desired sample sizes for mark–recapture analysis while also maximizing efficiency.  相似文献   

17.
Large‐scale genomic studies of wild animal populations are often limited by access to high‐quality DNA. Although noninvasive samples, such as faeces, can be readily collected, DNA from the sample producers is usually present in low quantities, fragmented, and contaminated by microorganism and dietary DNAs. Hybridization capture can help to overcome these impediments by increasing the proportion of subject DNA prior to high‐throughput sequencing. Here we evaluate a key design variable for hybridization capture, the number of rounds of capture, by testing whether one or two rounds are most appropriate, given varying sample quality (as measured by the ratios of subject to total DNA). We used a set of 1,780 quality‐assessed wild chimpanzee (Pan troglodytes schweinfurthii) faecal samples and chose 110 samples of varying quality for exome capture and sequencing. We used multiple regression to assess the effects of the ratio of subject to total DNA (sample quality), rounds of capture and sequencing effort on the number of unique exome reads sequenced. We not only show that one round of capture is preferable when the proportion of subject DNA in a sample is above ~2%–3%, but also explore various types of bias introduced by capture, and develop a model that predicts the sequencing effort necessary for a desired data yield from samples of a given quality. Thus, our results provide a useful guide and pave a methodological way forward for researchers wishing to plan similar hybridization capture studies.  相似文献   

18.
We studied the diets of four sympatric carnivores in the flooding savannas of western Venezuela by analysing predator DNA and prey remains in faeces. DNA was isolated and a portion of the cytochrome b gene of the mitochondrial genome amplified and sequenced from 20 of 34 scats. Species were diagnosed by comparing the resulting sequences to reference sequences generated from the blood of puma (Puma concolor), jaguar (Panthera onca), ocelot (Leopardus pardalus) and crab-eating fox (Cerdocyon thous). Scat size has previously been used to identify predators, but DNA data show that puma and jaguar scats overlap in size, as do those of puma, ocelot and fox. Prey-content analysis suggests minimal prey partitioning between pumas and jaguars. In field testing this technique for large carnivores, two potential limitations emerged: locating intact faecal samples and recovering DNA sequences from samples obtained in the wet season. Nonetheless, this study illustrates the tremendous potential of DNA faecal studies. The presence of domestic dog (Canis familiaris) in one puma scat and of wild pig (Sus scrofa), set as bait, in one jaguar sample exemplifies the forensic possibilities of this noninvasive analysis. In addition to defining the dietary habits of similar size sympatric mammals, DNA identifications from faeces allow wildlife managers to detect the presence of endangered taxa and manage prey for their conservation.  相似文献   

19.
Animal vectors are essential for the movement of invertebrate resting eggs between water bodies. However, differences in habitat preferences and feeding behaviour between bird species may result in variations in the dispersal of invertebrates via these birds, even if the different bird species live in the same lake. To test such effects, faecal samples from Anas platyrhynchos (collected in autumn and spring) and Tadorna tadorna (collected in spring) were cultured in water at 20° C and 12 L: 12 D conditions in order to quantify the resting eggs which could be internally transported by these birds. One half of each faecal sample was initially cultured at a conductivity of 0.6 mS cm−1 and the other half at 6 mS cm−1. 1054 invertebrates hatched from a total of 60 faecal samples, including cladocerans, copepods, ostracods, rotifers and ciliates, with a wide variability among faeces. Autumn yielded a low proportion of samples with hatchlings (12.5%) compared to spring (90%). Significant differences were observed between birds, but not between conductivity treatments. Thus, our results imply different hatching dynamics affected by disperser and season, but most species transported as resting eggs by birds seem to have a wide tolerance to hatch under variable salinity conditions. These differences may largery influence the metacommunity dynamics of lake networks, and could be a key factor to consider in wetland conservation planning.  相似文献   

20.
非损伤性取样被广泛应用在动物保护遗传学、分子生态学和分子进化等研究领域.随着基因组测序技术的发展和基因组学时代的到来,如何从非损伤性取样样品中获取能够用于进行基因组测序的高质量DNA是研究者面临的难题.本文总结和比较了非损伤性取样中最常用的粪便样品和考古材料或博物馆标本两类样品中富集宿主DNA的方法及应用,以期为非损伤...  相似文献   

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