Periostin Is a Biomarker for Anterior Shoulder Instability: Proteomic Analysis of Synovial Fluid.

BACKGROUND: The incremental biological changes in the synovial microenvironment of the shoulder in acute and chronic instability that may contribute to joint degeneration are poorly understood. Proteomic analysis of synovial fluid in patients with shoulder instability may improve our understanding of proteins that are shed into shoulder synovial fluid after an injury. HYPOTHESIS: Injury-specific factors such as the direction of instability and the severity of glenoid and humeral bone loss are associated with the proteome of synovial fluid in patients with shoulder instability. STUDY DESIGN: Descriptive laboratory study. METHODS: Synovial fluid lavage samples were compared between patients with anterior (n = 12) and posterior (n = 8) instability and those without instability (n = 5). Synovial proteins were identified with liquid chromatography-tandem mass spectrometry. Orthogonal validation of protein targets found to be significant on tandem mass spectrometry was performed in a separate set of prospective patients with Western blotting. Data were processed and analyzed, and P values were adjusted with the Benjamini-Hochberg method for multiple comparisons. RESULTS: A total of 25 patients were included. Tandem mass spectrometry identified 720 protein groups in synovial fluid of patients with shoulder instability. There were 4 synovial proteins that were significantly expressed in patients with anterior instability relative to posterior instability: periostin (POSTN) (adjusted P value = .03; log fold change [logFc] = 4.7), transforming growth factor beta-induced protein ig-h3 (adjusted P value = .05; logFc = 1.7), collagen type VI alpha-3 chain (adjusted P value = .04; logFc = 2.6), and coagulation factor V (adjusted P value = .04; logFc = -3.3). Among these targets, POSTN showed a moderate correlation with the Hill-Sachs lesion size (r = 0.7). Prospective validation with Western blotting confirmed a significantly higher level of POSTN in synovial fluid of patients with anterior instability (P = .00025; logFc = 5.1). CONCLUSION: Proteomic analysis enriched our understanding of proteins that were secreted into shoulder synovial fluid of patients with shoulder instability. The identification of POSTN, a proinflammatory catabolic protein involved with tissue remodeling and repair, as a significant target in anterior shoulder instability is a novel finding. Therefore, further study is warranted to determine the role that POSTN may play in the progression of bone loss and posttraumatic osteoarthritis. CLINICAL RELEVANCE: Proteomic analysis of synovial fluid in patients with shoulder instability improved our understanding of this abnormality after an injury.

The association between haemosporidian infection and non-breeding moult location in great reed warblers revisited by combining feather stable isotope profiles and geolocator data.

Stable isotope analysis provides valuable insights into the ecology of long-distance migratory birds during periods spent away from a specific study site. In a previous study, Swedish great reed warblers (Acrocephalus arundinaceus) infected with haemosporidian parasites differed in feather isotope ratios compared to non-infected birds, suggesting that infected and non-infected birds spent the non-breeding season in different locations or habitats. Here, we use a novel dataset comprising geolocator data, isotopes, and haemosporidian infection status of 92 individuals from four Eurasian populations to investigate whether parasite transmission varies with geography or habitats. We found that the probability of harbouring Plasmodium and Leucocytozoon parasites was higher in birds moulting in the eastern region of the non-breeding grounds. However, no geographic pattern occurred for Haemoproteus infections or overall infection status. In contrast to the previous study, we did not find any relationship between feather isotope ratios and overall haemosporidian infection for the entire current dataset. Plasmodium-infected birds had lower feather δ15N values indicating that they occupied more mesic habitats. Leucocytozoon-infected birds had higher feather δ34S values suggesting more coastal sites or wetlands with anoxic sulphate reduction. As the composition and prevalence of haemosporidian parasites differed between the old and the current dataset, we suggest that the differences might be a consequence of temporal dynamics of haemosporidian parasites. Our results emphasize the importance of replicating studies conducted on a single population over a restricted time period, as the patterns can become more complex for data from wider geographical areas and different time periods.

Animal tracing with sulfur isotopes: Spatial segregation and climate variability in Africa likely contribute to population trends of a migratory songbird.

Climatic conditions affect animals but range-wide impacts at the population level remain largely unknown, especially in migratory species. However, studying climate-population relationships is still challenging in small migrants due to a lack of efficient and cost-effective geographic tracking method. Spatial distribution patterns of environmental stable isotopes (so called 'isoscapes') generally overcome these limitations but none of the currently available isoscapes provide a substantial longitudinal gradient in species-rich sub-Saharan Africa. In this region, sulphur (δ34 S) has not been sufficiently explored on a larger scale. We developed a δ34 S isoscape to trace animal origins in sub-Saharan Africa by coupling known-origin samples from tracked migratory birds with continental remotely sensed environmental data building on environment-δ34 S relationships using a flexible machine learning technique. Furthermore, we link population-specific nonbreeding grounds with interannual climatic variation that might translate to breeding population trends. The predicted δ34 S isotopic map featured east-west and coast-to-inland isotopic gradients and was applied to predict nonbreeding grounds of three breeding populations of Eurasian Reed Warblers Acrocephalus scirpaceus with two distinct migratory phenotypes. Breeding populations as well as migratory phenotypes exhibited large-scale segregation within the African nonbreeding range. These regions also differed substantially in the interannual climatic variation, with higher interannual variability in the eastern part of the range during 2001-2012. Over the same period, the eastern European breeding population seemed to have experienced a more steep decline in population size. The link between migratory patterns and large-scale climatic variability appears important to better understand population trajectories in many declining migratory animals. We believe animal tracing using sulphur isotopes will facilitate these efforts and offers manifold ecological and forensic applications in the biodiversity hotspot of sub-Saharan Africa.

Toxicity and Starvation Induce Major Trophic Isotope Variation in Daphnia Individuals: A Diet Switch Experiment Using Eight Phytoplankton Species of Differing Nutritional Quality.

Stable isotope values can express resource usage by organisms, but their precise interpretation is predicated using a controlled experiment-based validation process. Here, we develop a stable isotope tracking approach towards exploring resource shifts in a key primary consumer species Daphnia magna. We used a diet switch experiment and model fitting to quantify the stable carbon (δ13C) and nitrogen (δ15N) isotope turnover rates and discrimination factors for eight dietary sources of the plankton species that differ in their cellular organization (unicellular or filamentous), pigment and nutrient compositions (sterols and polyunsaturated fatty acids), and secondary metabolite production rates. We also conduct a starvation experiment. We evaluate nine tissue turnover models using Akaike's information criterion and estimate the repetitive trophic discrimination factors. Using the parameter estimates, we calculate the hourly stable isotope turnover rates. We report an exceedingly faster turnover value following dietary switching (72 to 96 h) and a measurable variation in trophic discrimination factors. The results show that toxic stress and the dietary quantity and quality induce trophic isotope variation in Daphnia individuals. This study provides insight into the physiological processes that underpin stable isotope patterns. We explicitly test multiple alternative dietary sources and fasting and discuss the parameters that are fundamental for field- and laboratory-based stable isotope studies.

Longitudinal Proteomic Analysis of Plasma across Healthy Pregnancies Reveals Indicators of Gestational Age.

Longitudinal changes in the blood proteome during gestation relate to fetal development and maternal homeostasis. Charting the maternal blood proteome in normal pregnancies is critical for establishing a baseline reference when assessing complications and disease. Using mass spectrometry-based shotgun proteomics, we surveyed the maternal plasma proteome across uncomplicated pregnancies. Results indicate a significant rise in proteins that govern placentation and are vital to the development and health of the fetus. Importantly, we uncovered proteome signatures that strongly correlated with gestational age. Fold increases and correlations between the plasma concentrations of ADAM12 (ρ = 0.973), PSG1 (ρ = 0.936), and/or CSH1/2 (ρ = 0.928) with gestational age were validated with ELISA. Proteomic and validation analyses demonstrate that the maternal plasma concentration of ADAM12, either independently or in combination with either PSG1 or CSH1/2, correlates with gestational age within ±8 days throughout pregnancy. These findings suggest that the gestational age in healthy pregnancies may be determined by referencing the concentration of select plasma proteins.

Mitochondrial phosphoenolpyruvate carboxylase contributes to carbon fixation in the diatom Phaeodactylum tricornutum at low inorganic carbon concentrations.

Photosynthetic carbon fixation is often limited by CO2 availability, which led to the evolution of CO2 concentrating mechanisms (CCMs). Some diatoms possess CCMs that employ biochemical fixation of bicarbonate, similar to C4 plants, but whether biochemical CCMs are commonly found in diatoms is a subject of debate. In the diatom Phaeodactylum tricornutum, phosphoenolpyruvate carboxylase (PEPC) is present in two isoforms, PEPC1 in the plastids and PEPC2 in the mitochondria. We used real-time quantitative polymerase chain reaction, Western blots, and enzymatic assays to examine PEPC expression and PEPC activity, under low and high concentrations of dissolved inorganic carbon (DIC). We generated and analyzed individual knockout cell lines of PEPC1 and PEPC2, as well as a PEPC1/2 double-knockout strain. While we could not detect an altered phenotype in the PEPC1 knockout strains at ambient, low or high DIC concentrations, PEPC2 and the double-knockout strains grown under ambient air or lower DIC availability conditions showed reduced growth and photosynthetic affinity for DIC while behaving similarly to wild-type (WT) cells at high DIC concentrations. These mutants furthermore exhibited significantly lower 13 C/12 C ratios compared to the WT. Our data imply that in P. tricornutum at least parts of the CCM rely on biochemical bicarbonate fixation catalyzed by the mitochondrial PEPC2.

Rapid adaptation through genomic and epigenomic responses following translocations in an endangered salmonid.

Identifying the molecular mechanisms facilitating adaptation to new environments is a key question in evolutionary biology, especially in the face of current rapid and human-induced changes. Translocations have become an important tool for species conservation, but the attendant small population sizes and new ecological pressures might affect phenotypic and genotypic variation and trajectories dramatically and in unknown ways. In Scotland, the European whitefish (Coregonus lavaretus) is native to only two lakes and vulnerable to extirpation. Six new refuge populations were established over the last 30 years as a conservation measure. In this study, we examined whether there is a predictable ecological and evolutionary response of these fishes to translocation. We found eco-morphological differences, as functional traits relating to body shape differed between source and refuge populations. Dual isotopic analyses suggested some ecological release, with the diets in refuge populations being more diverse than in source populations. Analyses of up to 9117 genome-mapped SNPs showed that refuge populations had reduced genetic diversity and elevated inbreeding and relatedness relative to source populations, though genomic differentiation was low (F ST = 0.002-0.030). We identified 14 genomic SNPs that showed shared signals of a selective response to translocations, including some located near or within genes involved in the immune system, nervous system and hepatic functions. Analysis of up to 120,897 epigenomic loci identified a component of consistent differential methylation between source and refuge populations. We found that epigenomic variation and genomic variation were associated with morphological variation, but we were not able to infer an effect of population age because the patterns were also linked with the methodology of the translocations. These results show that conservation-driven translocations affect evolutionary potential by impacting eco-morphological, genomic and epigenomic components of diversity, shedding light on acclimation and adaptation process in these contexts.

Parallel and non-parallel changes of the gut microbiota during trophic diversification in repeated young adaptive radiations of sympatric cichlid fish.

BACKGROUND: Recent increases in understanding the ecological and evolutionary roles of microbial communities have underscored the importance of their hosts' biology. Yet, little is known about gut microbiota dynamics during the early stages of ecological diversification and speciation. We sequenced the V4 region of the 16s rRNA gene to study the gut microbiota of Nicaraguan Midas cichlid fish (Amphilophus cf. citrinellus). Specifically, we tested the hypothesis that parallel divergence in trophic ecology in extremely young adaptive radiations from two crater lakes is associated with parallel changes of their gut microbiota. RESULTS: Bacterial communities of fish guts and lake water were highly distinct, indicating that the gut microbiota is shaped by host-specific factors. Among individuals of the same crater lake, differentiation in trophic ecology was weakly associated with gut microbiota differentiation, suggesting that diet, to some extent, affects the gut microbiota. However, differences in trophic ecology were much more pronounced across than within species whereas similar patterns were not observed for taxonomic and functional differences of the gut microbiota. Across the two crater lakes, we could not detect conclusive evidence for parallel changes of the gut microbiota associated with trophic ecology. CONCLUSIONS: A lack of clearly differentiated niches during the early stages of ecological diversification might result in non-parallel changes of gut microbial communities, as observed in our study system as well as in other recently diverged fish species. Video Abstract.

Foraging niche shift maintains breeding parameters of a colonial waterbird during range expansion.

Relating the effects of foraging niche variation to reproductive dynamics is critical to understand species response to environmental change. We examined foraging niche variations of the slender-billed gull (Chroicocephalus genei), a nomadic colonial waterbird species during its range expansion along the French Mediterranean coast over a 16-year period (1998-2013). We investigated whether range expansion was associated with a change in chick diet, breeding success, and chicks body condition. We also examined whether breeding success and chicks body condition were explained by diet and colonial characteristics (number of pairs, laying phenology, habitat, and locality). Diet was characterized using dual-stable isotopic proxies (δ 13C and δ 15N) of feather keratin from 331 individuals subsampled from a total of 4,154 chicks ringed and measured at 18 different colonies. δ 13C decreased and δ 15N increased significantly during range expansion suggesting that chicks were fed from preys of increasing trophic level found in the less salty habitat colonized by the end of the study period. Niche shift occurred without significant change of niche width which did not vary among periods, habitats, or localities either. Breeding success and chick body condition showed no consistent trends over years. Breeding success tended to increase with decreasing δ 13C at the colony level while there was no relationship between stable isotope signatures and chick body condition. Overall, our results suggest that even if range expansion is associated with foraging niche shift toward the colonization of less salty and more brackish habitats, the shift had marginal effect on the breeding parameters of the Slender-billed gull. Niche width appears as an asset of this species, which likely explains its ability to rapidly colonize new locations.

The human tubal lavage proteome reveals biological processes that may govern the pathology of hydrosalpinx.

Hydrosalpinx, the blockage of fallopian tubes, can result from pelvic inflammatory disease. Hydrosalpinx is a cause of infertility and negatively impacts in vitro fertilization. To better understand the pathobiology of hydrosalpinx, we compared the proteome of lavages from disease vs. healthy fallopian tubes. Results indicate a disruption of redox homeostasis and activation of the complement system, immune cell infiltration, and phagocytosis; pathways that may drive tubal injury. To our surprise among the most prominent proteins with hydrosalpinx was mesothelin (MSLN), which until now has only been associated with epithelial malignancies. Analogous to mesothelioma and ovarian carcinoma, a significant increase of MSLN was detected in plasma from patients with hydrosalpinx. This finding suggests MSLN may provide clinical diagnosis in lieu of the current approaches that require invasive imaging. Importantly, these findings implicate MSLN in a benign disease, indicating that the activation and role of MSLN is not restricted to cancer.

Rapid niche expansion by selection on functional genomic variation after ecosystem recovery.

It is well recognized that environmental degradation caused by human activities can result in dramatic losses of species and diversity. However, comparatively little is known about the ability of biodiversity to re-emerge following ecosystem recovery. Here, we show that a European whitefish subspecies, the gangfisch Coregonus lavaretus macrophthalmus, rapidly increased its ecologically functional diversity following the restoration of Lake Constance after anthropogenic eutrophication. In fewer than ten generations, gangfisch evolved a greater range of gill raker numbers (GRNs) to utilize a broader ecological niche. A sparse genetic architecture underlies this variation in GRN. Several co-expressed gene modules and genes showing signals of positive selection were associated with GRN and body shape. These were enriched for biological pathways related to trophic niche expansion in fishes. Our findings demonstrate the potential of functional diversity to expand following habitat restoration, given a fortuitous combination of genetic architecture, genetic diversity and selection.

Dietary nutrient allocation to somatic tissue synthesis in emerging subimago freshwater mayfly Ephemera danica.

BACKGROUND: The relative importance of nutrients derived from different sources for tissue synthesis is crucial for predicting a species responds to changes in food availability. The ecological and physiological strategies that govern the incorporation and routing of nutrients for reproduction are often well understood. However, the role and adaptive value of both species and individual variation during early life-stage remain elusive. In freshwater systems, dietary nutrient allocation to somatic tissue should be favoured when dietary source peaks and resource limitation may hinder flexible resource allocation. We used carbon and nitrogen stable isotopes (δ13C and δ15N) to examine metabolic nutrient routing and resource allocation from four dietary sources used to biosynthesize three somatic tissues of emerging subimago Ephemera danica. Aquatic emerging insects, such as the mayfly E. danica, are well suited for such studies. This is because, while burrowing nymph phase is a detritivores feeders with several early life-stages of metamorphosis, adult insects do not feed during this period but do utilize energy. RESULTS: Constructed models to predict percent proportional contribution of source to tissue showed that terrestrial detritus was the dominant nutrient source for abdomen, head and wing with mean values of 57%, 65% and 73%, respectively. There was evidence for differential resource allocation, as insect partitioned periphyton and sediment (but also seston) elements for tissue synthesis. Utilizing individual-specimen based relationship in isotope value; we derived tissue specific isotopic niche estimates, for the different tissue-source combinations. CONCLUSIONS: Results indicate that tissue selection is crucial for isotopic ecological measurements in arthropods. Mayfly has long been used as bio-indicator of freshwater ecosystems and their larvae show rapid response to environmental changes. In light of the recent evidence of drastic reduction in flying insect mass in Germany, developing a system using isotopic tools to trace nutrient flow in this important taxon will assist conservation and management efforts.

Seasonality at the equator: isotope signatures and hormonal correlates of molt phenology in a non-migratory Amazonian songbird.

BACKGROUND: Birds, across their annual cycle, progress through sequences of life-history stages such as reproduction and molt. The mechanisms that control annual avian itineraries involve endocrine responses triggered by seasonal environmental factors, including changes in resource availability and/or photoperiod. However, at equatorial latitudes birds are exposed to different degrees of seasonality, and the mechanisms underlying phenology of birds near the equator remain less explored. We studied the silver-beaked tanager, an endemic Amazonian songbird, from an equatorial lowland population. Remarkably, in this species, song behavior has been shown to be seasonally aligned to minimal changes in day length near the equator. Here, we aimed to further explore the phenology of silver-beaked tanagers by assessing shifts of food sources utilization as potential ultimate factors. We measured triple isotopic tracers of carbon (δ13C), nitrogen (δ15N) and sulphur (δ34S) in blood and feathers of birds throughout a whole year. In addition, we assessed the degree of seasonality in the molting activity, in relation to circulating levels of corticosterone, as well as to testosterone as a proxy of the reproductive condition of males. RESULTS: There was important seasonal variation of δ34S values in relation to rainfall patterns and changes in estuarine water composition. Despite the seasonal rainfall, we found no substantial variation in the foraging ecology of birds over seasons. This was accompanied by uniform levels of corticosterone throughout the year, probably associated with the absence of drastic seasonal resource shortages. Even so, silver-beaked tanagers showed a marked seasonal molting schedule, which was related to variation in the circulating levels of both corticosterone and testosterone. CONCLUSIONS: These findings suggest that foraging niche is not life history stage-dependent in silver-beaked tanagers, and highlight rainfall as an important environmental cue for bird phenology. Our stable isotope results encourage further studies addressing the influence of estuarine water dynamics on bird timing. In addition, the results suggest a primary role of steroid hormones in regulating seasonal life history stages under the absence of a marked photoperiod. Contrary to what might be expected for a tropical songbird, our physiological data in silver-beaked tanagers do not support reproduction-molt overlapping.

Dual-tracer-based isotope turnover rates in a highly invasive mysid Limnomysis benedeni from Lake Constance.

Understanding the ecological patterns of invasive species and their habitats require an understanding of the species' foraging ecology. Stable carbon (δ13C) and nitrogen (δ15N) isotope values provide useful information into the study of animal ecology and evolution, since the isotope ratios of consumers reflect consumer's dietary patterns. Nevertheless, the lack of species- and element-specific laboratory-derived turnover rates could limit their application. Using a laboratory-based dual stable isotope tracer approach (Na15 NO 3 and NaH13 CO 3), we evaluated the δ15N and δ13C isotope turnover rates in full-grown adult invasive Limnomysis benedeni from Lake Constance. We provide δ15N and δ13C turnover rates based on nonlinear least-squares regression and posterior linear regression models. Model precisions and fit were evaluated using Akaike's information criterion. Within a couple of days, the δ15N and δ13C of mysids began to change. Nevertheless, after about 14 days, L. benedeni did not reach equilibrium with their new isotope values. Since the experiment was conducted on adult subjects, it is evident that turnover was mainly influenced by metabolism (in contrast to growth). Unlike traditional dietary shifts, our laboratory-based dual stable isotope tracer approach does not shift the experimental organisms into a new diet and avoids dietary effects on isotope values. Results confirm the application of isotopic tracers to label mysid subpopulations and could be used to reflect assimilation and turnover from the labeled dietary sources. Field-based stable isotope studies often use isotopic mixing models commonly assuming diet-tissue steady state. Unfortunately, in cases where the isotopic composition of the animal is not in equilibrium with its diet, this can lead to highly misleading conclusions. Thus, our laboratory-based isotopic incorporation rates assist interpretation of the isotopic values from the field and provide a foundation for future research into using isotopic tracers to investigate invasion ecology.

The Effect of Parasite Infection on Stable Isotope Turnover Rates of δ15N, δ13C and δ34S in Multiple Tissues of Eurasian Perch Perca fluviatilis.

Stable isotope analysis of commercially and ecologically important fish can improve understanding of life-history and trophic ecology. However, accurate interpretation of stable isotope values requires knowledge of tissue-specific isotopic turnover that will help to describe differences in the isotopic composition of tissues and diet. We performed a diet-switch experiment using captive-reared parasite-free Eurasian perch (Perca fluviatilis) and wild caught specimens of the same species, infected with the pike tapeworm Triaenophorus nodulosus living in host liver tissue. We hypothesize that metabolic processes related to infection status play a major role in isotopic turnover and examined the influence of parasite infection on isotopic turn-over rate of carbon (δ13C), nitrogen (δ15N) and sulphur (δ34S) in liver, blood and muscle. The δ15N and δ13C turnovers were fastest in liver tissues, followed by blood and muscle. In infected fish, liver and blood δ15N and δ13C turnover rates were similar. However, in infected fish, liver and blood δ13C turnover was faster than that of δ15N. Moreover, in infected subjects, liver δ15N and δ13C turnover rates were three to five times faster than in livers of uninfected subjects (isotopic half-life of ca.3-4 days compared to 16 and 10 days, respectively). Blood δ34S turnover rate were about twice faster in non-infected individuals implying that parasite infection could retard the turnover rate of δ34S and sulphur containing amino acids. Slower turnover rate of essential amino acid could probably decrease individual immune function. These indicate potential hidden costs of chronic and persistent infections that may have accumulated adverse effects and might eventually impair life-history fitness. For the first time, we were able to shift the isotope values of parasites encapsulated in the liver by changing the dietary source of the host. We also report variability in isotopic turnover rates between tissues, elements and between infected and parasite-free individuals. These results contribute to our understanding of data obtained from field and commercial hatcheries; and strongly improve the applicability of the stable isotope method in understanding life-history and trophic ecology of fish populations.

Seasonal rainfall at long-term migratory staging sites is associated with altered carry-over effects in a Palearctic-African migratory bird.

BACKGROUND: An understanding of year-round habitat use is essential for determining how carry-over effects shape population dynamics in long-distance migratory songbirds. The recent discovery of long-term migratory staging sites in many species, prior to arrival at final wintering sites, adds complexity to efforts to decipher non-breeding habitat use and connections between sites. We investigated whether habitat conditions during migratory staging carry over to influence great reed warbler (Acrocephalus arundinaceus) body condition at final wintering sites in Zambia. We asked whether the presence/absence and strength of such carry-over effects were modified by contrasting rainfall conditions during 2 years. RESULTS: First, we found that individuals staging in a dry year had higher corticosterone (CORTf) and stable nitrogen isotope values (suggesting higher aridity) than birds staging in a wet year, indicating that regional weather affected staging conditions. Second, we found that carry-over effects from staging habitat conditions (measured via carbon and nitrogen isotopes) to final winter site body condition (measured via scaled mass index and ß-hydroxybutyrate) were only present in a dry year, suggesting that environmental factors have consequences for the strength of carry-over effects. Our results also suggest that wet conditions at final winter sites may buffer the effects of poor staging conditions, at least in the short term, since individuals that staged in a dry year had higher scaled mass indices in Zambia than individuals that staged in a wet year. CONCLUSIONS: This study provides a first insight into the connections between long-term migratory staging sites and final wintering sites, and suggests that local environmental factors can modify the strength of carry-over effects for long-distance migratory birds.

Induced responses to grazing by an insect herbivore (Acentria ephemerella) in an immature macrophyte (Myriophyllum spicatum): an isotopic study.

While the mechanisms by which adult terrestrial plants deploy constitutive and induced responses to grazing pressure are well known, the means by which young aquatic plants defend themselves from herbivory are little studied. This study addresses nitrogen transport in the aquatic angiosperm Myriophyllum spicatum in response to herbivore exposure. Nitrogen tracers were used to monitor nitrogen uptake and reallocation in young plants in response to grazing by the generalist insect herbivore Acentria ephemerella. Total nitrogen content (N%) and patterns of nitrogen uptake and allocation (δ(15)N) were assessed in various plant tissues after 24 and 48 h. Following 24 h exposure to herbivore damage (Experiment 1), nitrogen content of plant apices was significantly elevated. This rapid early reaction may be an adaptation allowing the grazer to be sated as fast as possible, or indicate the accumulation of nitrogenous defense chemicals. After 48 h (Experiment 2), plants' tips showed depletion in nitrogen levels of ca. 60‰ in stem sections vulnerable to grazing. In addition, nitrogen uptake by grazed and grazing-prone upper plant parts was reduced and nutrient allocation into the relatively secure lower parts increased. The results point to three conclusions: (1) exposure to an insect herbivore induces a similar response in immature M. spicatum as previously observed in mature terrestrial species, namely a rapid (within 48 h) reduction in the nutritional value (N%) of vulnerable tissues, (2) high grazing intensity (100% of growing tips affected) did not limit the ability of young plants to induce resistance; and (3) young plants exposed to herbivory exhibit different patterns of nutrient allocation in vulnerable and secure tissues. These results provide evidence of induced defense and resource reallocation in immature aquatic macrophytes which is in line with the responses shown for mature aquatic macrophytes and terrestrial plants.

Compound-specific δ(13)C analyses reveal sterol metabolic constraints in an aquatic invertebrate.

RATIONALE: Dietary sterol deficiencies can have severe life history consequences for consumers. Compound-specific stable isotope analysis (CSIA) was applied to the exploration of the sterol metabolic constraints and bioconversion capacities of the amphipod Gammarus roeselii. Evaluating structural sterol requirements has great potential to improve our understanding of the ecological relevance of sterols as limiting nutrients. METHODS: Juvenile G. roeselii were reared on food mixtures consisting of different ratios of the two algae Scenedesmus obliquus (cultivated with (13)C-labeled NaHCO3) and Nannochloropsis limnetica (unlabeled), which have been shown previously to differ in food quality. We measured the sterol content and composition using a gas chromatograph equipped with a flame ionization detector and the δ(13)C values of sterols using compound-specific isotope ratio mass spectrometry to examine potential sterol-mediated nutritional constraints of G. roeselii. RESULTS: In the food mixtures, δ(13)C values of cholesterol, synthesized by N. limnetica, were -25‰ and those of the Δ(7)-phytosterols, chondrillasterol and fungisterol, synthesized by S. obliquus, were 7 and 18‰, respectively. Although the cholesterol concentrations in G. roeselii decreased with increasing proportion of dietary S. obliquus, the δ(13)C values remained constant at -25‰. Lathosterol, which appeared in G. roeselii at high dietary proportions of S. obliquus, had a δ(13)C value of 35‰. CONCLUSIONS: We provide evidence that the the Δ(7)-phytosterols present in S. obliquus cannot be metabolized to cholesterol in G. roeselii, resulting in the accumulation of lathosterol in the animals and potentially in sterol-limited growth. These findings emphasize the advantage of CSIA in revealing the physiological mechanisms associated with nutritional constraints.

Isotopic evidence for dietary niche overlap between barking deer and four-horned antelope in Nepal.

BACKGROUND: Morphologically similar sympatric species may have a high degree of niche overlap. Barking deer Muntiacus vaginalis and four-horned antelope Tetracerus quadricornis are solitary ungulates of the Indian sub-continent. Limited information is available regarding their trophic ecology, particularly of the endemic four-horned antelope. We present stable carbon (δ(13)C), nitrogen (δ(15)N), and sulphur (δ(34)S) isotopic values, and nitrogen content (%N) of faeces from barking deer and four-horned antelope living in lowland Nepal to assess trophic niche differentiation of these herbivores along the browser-grazer continuum. We also describe trophic differences between those two species in ecological niches and seasonal effects on their diets. RESULTS: We found that the barking deer and four-horned antelope consumed C3 plant sources exclusively. The niche partitioning in their diet was reflected by δ(34)S values. Some seasonal effects observed were: δ(13)C and δ(15)N were significantly lower in the dry season diet of four-horned antelope than that of barking deer, while δ(34)S values were significantly higher in the winter diet; monsoon diet was similar for both species. Faecal N levels for barking deer and four-horned antelope were similar throughout all the seasons, indicating that both species adapted their feeding behaviour so as to maximize protein intake, in accordance with season and environment. CONCLUSIONS: Barking deer and four-horned antelope both are browsers; their dietary sources overlapped during monsoon but differed during the dry season. Conservation actions focused on resource management during the dry season to reduce food scarcity and competition over limited resources is likely to be the most effective.