Isotopic niche partitioning in two sympatric howler monkey species.

OBJECTIVES: Ecological similarity between species can lead to interspecific trophic competition. However, when ecologically similar species coexist, they may differ in foraging strategies and habitat use, which can lead to niche partitioning. As the body tissues of consumers contain a stable isotope signature that reflects the isotopic composition of their diet, stable isotope analysis is a useful tool to study feeding behavior. We measured the isotopic niche width, which is a proxy for trophic niche width, of mantled (Alouatta palliata) and black (A. pigra) howler monkeys. Specifically, studied populations in allopatry and sympatry to assess whether these species showed niche partitioning. MATERIALS AND METHODS: Between 2008 and 2012, we collected hair samples from 200 subjects (113 black and 87 mantled howler monkeys) and used continuous flow isotope ratio mass spectrometry to estimate δ13 C and δ15 N. We described the isotopic niche width of each species in allopatry and sympatry with the Bayesian estimation of the standard ellipse areas. RESULTS: In allopatry, isotopic niche width and isotopic variation were similar in both species. In sympatry, black howler monkeys had a significantly broader isotopic niche, which was mainly determined by high δ15 N values, and included the majority of mantled howler monkeys' isotopic niche. The isotopic niche of mantled howler monkeys did not differ between sympatry and allopatry. CONCLUSIONS: The coexistence of these ecologically similar species may be linked to trophic niche adjustments by one species, although the particular features of such adjustments (e.g., dietary, spatial, or sensory partitioning) remain to be addressed.

Population control of an overabundant species achieved through consecutive anthropogenic perturbations.

The control of overabundant vertebrates is often problematic. Much work has focused on population-level responses and overabundance due to anthropogenic subsidies. However, far less work has been directed at investigating responses following the removal of subsidies. We investigate the consequences of two consecutive perturbations, the closure of a landfill and an inadvertent poisoning event, on the trophic ecology (δ13C, δ15N, and δ34S), survival, and population size of an overabundant generalist seabird species, the Yellow-legged Gull (Larus michahellis). We expected that the landfill closure would cause a strong dietary shift and the inadvertent poisoning a decrease in gull population size. As a long-lived species, we also anticipated adult survival to be buffered against the decrease in food availability but not against the inadvertent poisoning event. Stable isotope analysis confirmed the dietary shift towards marine resources after the disappearance of the landfill. Although the survival model was inconclusive, it did suggest that the perturbations had a negative effect on survival, which was followed by a recovery back to average values. Food limitation likely triggered dispersal to other populations, while poisoning may have increased mortality; these two processes were likely responsible for the large fall in population size that occurred after the two consecutive perturbations. Life-history theory suggests that perturbations may encourage species to halt existing breeding investment in order to ensure future survival. However, under strong perturbation pulses the resilience threshold might be surpassed and changes in population density can arise. Consecutive perturbations may effectively manage overabundant species.

From recruitment to senescence: food shapes the age-dependent pattern of breeding performance in a long-lived bird.

We used a long-term data set (26 years) from Audouin's Gull (Larus audouinii), a long-lived seabird, to address the relationship between the age-dependent pattern of reproductive performance and environmental conditions during breeding. Although theoretical models predict that the youngest and oldest breeders (due to inexperience and senescence, respectively) will perform less well than intermediate age classes, few empirical data exist regarding how this expected pattern varies with food availability. To assess the influence of age and food availability (corrected by population size of the main consumers to take into account density dependence) on a number of breeding parameters (laying dates, egg volume, clutch size, and hatching success), we modeled mean and variances of these parameters by incorporating heterogeneity into generalized linear models. All parameters varied with age and to different degrees, depending on food availability. As expected, performance improved with increased food supply, and the observed age pattern was quadratic, with poorer breeding performances occurring in extreme ages. For most parameters (except for laying dates, for which age and food did not interact), the pattern changed with food somewhat unexpectedly; the differences in performance between age classes were higher (i.e., the quadratic pattern was more noticeable) when food was more readily available than when food availability was lower. We suggest that, under poor environmental conditions, only high-quality individuals of the younger and older birds bred and that the differences in breeding performance between age classes were smaller. Although variances for egg volume were constant, variances for laying dates were highest for the youngest breeders and tended to decrease with age, either due to the selection of higher-quality individuals or to a greater frequency of birds skipping breeding with age, especially when food was in low supply. Our results show that mean and variances of breeding parameters changed with age, but that this pattern was different for each parameter and also varied according to food availability. It is likely that, other than food, certain additional factors (e.g., sex, cohort effects, density dependence) also influence changes in breeding performance with age, and this may preclude the finding of a common pattern among traits and among studies on different taxa.

Distinguishability and agreement with continuous data.

The analysis of concordance among repeated measures has received a huge amount of attention in the statistical literature leading to a range of different approaches. However, because all the approaches are able to assess the closeness among the readings taken on the same subject, the conclusions about the degree of concordance should be similar regardless the approach applied. Here, two indices to assess the concordance among continuous repeated measures, the intraclass correlation coefficient and the total deviation index, are applied and compared in two case examples. The first example concerns the repeatability of individual nutrient allocation strategy assessed by stable isotope analysis. The second example dealt with the assessment of the concordance of functional magnetic resonance imaging data that shows spatial correlation. The results differ depending upon the approach applied leading to contradictory conclusions about the degree of concordance. The reason behind these results is discussed reaching the conclusion that the total deviation index is just assessing agreement among repeated measurements, whereas the intraclass correlation coefficient assesses the concept of distinguishability among subjects that involves agreement among repeated measurements and spread of subjects at once. Therefore, the best way to select the right approach is to understand the right question behind the research hypothesis.

Assessment of trace elements and stable isotopes of three Ardeid species at Birama Swamp, Cuba.

The Birama Swamp is the second largest wetland in the Caribbean region and it is inhabited by large populations of waterbirds. Here we report, for the first time, the foraging ecology and pollutant levels of three Ardeidae species: Cattle egret (Bubulcus ibis), Snowy egret (Egretta thula), and Tricolored heron (E. tricolor) breeding in this wetland using stable-isotope (δ (15)N and δ (13)C) and trace elements [mercury (Hg), lead (Pb), and selenium (Se)] analysis of chick feathers. Our results showed that individuals from all species occupied similar trophic levels. However, we found significant differences for δ (13)C, with the highest values in cattle egret indicating its use of terrestrial habitats and a generalist and opportunist behavior. No significant differences were found for Pb among species. Yet, Hg levels were greater and similar in tricolored heron and snowy egret than in cattle egret, which was associated with their greater use of aquatic environments. Snowy egret had the lowest values of Se differing significantly with the other two species suggesting a different relative use of prey type. Modeling log-Hg concentration in relation to δ (15)N and δ (13)C showed an independent and significant relationship among species but without interaction with species level indicating that within a particular species, higher Hg levels were associated with higher δ (15)N values. There was no interaction between δ (15)N and δ (13)C in the general linear models for Se and Pb in all species. We found an association between δ (15)N and species in Pb for snowy egret. The foraging habitat use of these species and the low levels of pollutants, which are lower than in other similar habitats in other areas of the world, indicated that there is not risk of negative effects in juvenile birds of the Birama Swamp colony that may impair their survival. Our results can be used as a baseline to achieve management regulations.

Optimum threshold estimation based on cost function in a multistate diagnostic setting.

In the diagnostic area, the usual setting considers two populations: nondiseased and diseased. The use of the standard ROC analysis methodology is well established. Sometimes, however, diagnostic problems inherently include more than two classification states. For example, 'yes, uncertain, no' or 'low, normal, high'. Here we consider a three-normal distribution setting and derive estimators for the optimum thresholds between states based on a cost function. These estimators can be extended for clinical contexts with more than three states. This approach is well known for the two-state setting and its advantage lies in the fact that it accounts for the specific context's properties, such as disease prevalence and classification costs. Here we calculated the variance of the estimators by the use of parametric methods on nonlinear equations and we constructed confidence intervals accounting for possible uncertainty in the threshold estimation. We conducted a simulation study to assess the performance of these estimators and the confidence intervals. Comparisons with the naive threshold estimation method of joining the distributions two-by-two and applying standard ROC techniques proved that the latter method is not reliable for all parameter combinations and should be avoided.

Are the toxic sediments deposited at Flix reservoir affecting the Ebro river biota? Purple heron eggs and nestlings as indicators.

The Flix reservoir, in the low course of the Ebro River, contains thousands of tons of polluted sediments, accumulated from the activities of a chemical factory. An ongoing project is working toward removing these pollutants. Piscivore birds like the purple heron (Ardea purpurea) may be useful bioindicators, so eggs and nestling feathers were sampled during the 2006-2008 breeding seasons at three localities: a reference site situated upstream and two potentially affected by the toxic muds; one at the focal area and one at a distal area, the Ebro Delta. The samples were analyzed for isotopic signatures of ¹5N and ¹³C and concentrations of heavy metals and selenium. Baseline nitrogen signatures were higher in riverine sites than in the delta. Nitrogen together with carbon signatures adequately discriminated riverine and deltaic ecosystems. Mercury levels are highly influenced by the polluted sediments at Flix and pose potential risks for the birds, as they are among the highest ever recorded in heron species. Selenium and copper concentrations probably derive from other sources. Except for mercury, heavy metals and selenium levels were below toxic levels. Purple heron eggs and nestling feathers have demonstrated their usefulness as bioindicators for pollution in the river biota; feathers in particular show pollutant impacts on a strict local basis. A long series of study years is necessary in dynamic ecosystems such as this, so continued monitoring of the heron population at Flix is advisable to trace the effects of the toxic muds, particularly during their removal, because of the high levels of mercury detected.

Tracing waterbird exposure to total mercury and selenium: a case study at the solar saltworks of Thyna (sfax, Tunisia).

Saltworks have emerged as important alternative/complementary feeding habitats for avifauna. However, the consequences of such habitat shifts in terms of changes in exposure to contaminants are poorly understood. We evaluated the exposure of the waterbird community breeding at the saltworks of Thyna (Tunisia) to total Hg (THg) and Se according to their differential use of saltworks dietary resources, as revealed by δ¹³C and δ¹5N values in their eggs (included species [n] -sorted according to increasing reliance on saltworks resources: Yellow-legged Gull Larus michahellis [12], Common Tern Sterna hirundo [12], Slender-billed Gull Larus genei [15], Little Egret Egretta garzetta [20], and Pied Avocet Recurvirostra avosetta [22]). Concentrations of THg and Se were under the threshold points for deleterious effects. Egg THg concentrations significantly decreased as the dietary contribution of saltworks resources increased (mean: 3.23, 1.66, 0.76, 0.4, and 0.27 µg/g dw, respectively). Conversely, egg Se concentrations did not vary according to foraging habitats (2.49, 2.96, 2.61, 3.27, and 1.5 µg/g dw, respectively). Tracing waterbird exposure to THg and Se at saltworks was feasible through the use stable isotopic assays of eggs. Birds using saltworks are not exposed to higher concentrations of THg and Se than in adjacent marine habitats.

Little Egret (Egretta garzetta) as a bioindicator of trace element pollution in Tunisian aquatic ecosystems.

The Little Egret, Egretta garzetta, has breeding colonies in the island of Chikly (in the lake of Tunis) and in Thyna saltpans (in the gulf of Gabès), two important Tunisian wetlands that are strongly affected by anthropogenic activity. Here, we used E. garzetta chick feathers for environmental monitoring of breeding grounds of this species. Since trophic ecology is fundamental when interpreting contamination levels, our approach combined both trace-element (Hg, Pb, Cd, and Se) and stable-isotope analysis of δ13C, δ15N, and δ34S. Hg, Se, and δ15N levels were higher in specimens collected on Chikly than in Thyna. These observations highlight the degree of eutrophication of the lake of Tunis. Yellow-legged gulls (Larus michahellis) breeding in the same areas also feeds in the lake and attains similar concentrations of Hg and Se. In Thyna, egrets and gulls exploit distinct foraging habitats, as demonstrated by stable isotope analysis. The highest Hg and Se concentrations were found in Thyna. This result is consistent with greater exploitation of marine resources from the gulf of Gabès.

Influence of refuse sites on the prevalence of Campylobacter spp. and Salmonella serovars in seagulls.

Wild animals are well-known reservoirs of Campylobacter and Salmonella. We investigated the influence of insalubrious diets on the prevalence of both enterobacteria in seagulls. Campylobacter occurrence in gull chicks sampled along the northeastern Iberian coast was directly related to the degree of refuse consumption. High Salmonella values from the sampling sites did not reflect any dietary relationship.

Evaluating cleansing effects on trace elements and stable isotope values in feathers of oiled birds.

Feathers of seabirds are widely used as a nondestructive tissue for pollution monitoring of trace elements, as well as convenient samples for trophic ecology studies by means of stable isotope analysis (SIA). Nevertheless, feathers can be occasionally impregnated with oil from deliberate ship discharges and from massive oil spill accidents. The feather structure makes them effective traps for particles and are subject to external contamination. It is unknown to what extent the oil adhered to feathers can change trace element concentrations or stable isotope signatures. This study has two primary objectives: (1) to assess if there are differences between trace element concentrations and stable isotope signatures of oiled and clean feathers, and (2) to determine if the cleansing of oiled feathers using commonly applied techniques such as sodium hydroxide (NaOH) washes in combination with an organic solvent (hexane) is more effective than using NaOH alone. In order to do this, we analysed trace elements (Se, Hg, Pb, Cu and Zn) and stable isotopes (delta(13)C and delta(15)N) of individual feathers of yellow-legged gulls (Larus michahellis) which were affected by the 2002 Prestige oil spill in Galicia (NW Spain). Two sets of feathers were analysed, one group were oil-free (Control group) and the other had oil adhered to its surface (Oiled group). We expected to find differences between control and oiled feathers when cleaning exclusively with NaOH and no differences when using hexane. Our results did not show significant differences between Control and Oiled groups as a consequence of the cleansing method used. Unexpectedly, the additional cleansing with hexane resulted in decreasing selenium concentrations and increasing zinc and delta(15)N values in all groups of feathers.

Estimation of the diagnostic threshold accounting for decision costs and sampling uncertainty.

Medical diagnostic tests are used to classify subjects as non-diseased or diseased. The classification rule usually consists of classifying subjects using the values of a continuous marker that is dichotomised by means of a threshold. Here, the optimum threshold estimate is found by minimising a cost function that accounts for both decision costs and sampling uncertainty. The cost function is optimised either analytically in a normal distribution setting or empirically in a free-distribution setting when the underlying probability distributions of diseased and non-diseased subjects are unknown. Inference of the threshold estimates is based on approximate analytically standard errors and bootstrap-based approaches. The performance of the proposed methodology is assessed by means of a simulation study, and the sample size required for a given confidence interval precision and sample size ratio is also calculated. Finally, a case example based on previously published data concerning the diagnosis of Alzheimer's patients is provided in order to illustrate the procedure.

High Trophic Niche Overlap between a Native and Invasive Mink Does Not Drive Trophic Displacement of the Native Mink during an Invasion Process.

The pressure elicited by invasive species on native species significantly increases with the increase of the overlap of their ecological niches. Still, the specific mechanisms of the trophic displacement of native species during the invasion process are unclear. The effects of the invasive American mink (Neovison vison) on the critically endangered European mink (Mustela lutreola) was assessed by analyses of diet and niche overlap during the invasion process. To do this, the isotopic composition (δ13C and δ15N) of both species of mink and their four main types of prey was analysed. Significant trophic overlap between the native European mink and invasive American mink was found when they coexisted in sympatry. Furthermore, both mink species were characterised by significant individual variation in diet and no obvious change in diet of the native species in response to the arrival of the introduced species was observed. High niche overlap registered between both species in sympatry with no displacement in diet of the native mink in response to the arrival of the invasive mink is expected to have important consequences for the viability and conservation of the native mink populations, as it suggests high competitive pressure.

Probe-specific mixed-model approach to detect copy number differences using multiplex ligation-dependent probe amplification (MLPA).

BACKGROUND: MLPA method is a potentially useful semi-quantitative method to detect copy number alterations in targeted regions. In this paper, we propose a method for the normalization procedure based on a non-linear mixed-model, as well as a new approach for determining the statistical significance of altered probes based on linear mixed-model. This method establishes a threshold by using different tolerance intervals that accommodates the specific random error variability observed in each test sample. RESULTS: Through simulation studies we have shown that our proposed method outperforms two existing methods that are based on simple threshold rules or iterative regression. We have illustrated the method using a controlled MLPA assay in which targeted regions are variable in copy number in individuals suffering from different disorders such as Prader-Willi, DiGeorge or Autism showing the best performace. CONCLUSION: Using the proposed mixed-model, we are able to determine thresholds to decide whether a region is altered. These threholds are specific for each individual, incorporating experimental variability, resulting in improved sensitivity and specificity as the examples with real data have revealed.

Assessing the effects of the Prestige oil spill on the European shag (Phalacrocorax aristotelis): trace elements and stable isotopes.

The Prestige oil spill resulted in the mortality of several seabird species on the Atlantic NW coast of Spain. Shag casualties were particularly relevant, since populations are resident in the area the whole year round and because of several features which make them highly vulnerable to environmental hazards. Ecological catastrophes give us the opportunity of collecting samples which, otherwise, would be difficult to obtain. We examine the potential of shag corpses as bioindicators of inorganic pollution and the possible factors of variability, such as biological traits (sex, age) or nutrition status. We determined trace elements (Hg, Se, Cr, Pb, Zn and Cu) and isotopic signatures (15N, 13C) in soft tissues (muscle, liver) and in primary feathers formed at different times (before and after the Prestige) in individuals of known sex and age, collected at the time of the Prestige disaster. These were compared with data from another group of shags trapped accidentally in fishing gear in 2005. Our results did not seem to be affected by sex or age on any of the analysed variables. The higher nitrogen isotopic signatures in the soft tissues of the Prestige shags may be related to the nutrition stress caused by a poorer body condition, which is also reflected in increasing levels of some metals in the liver. This isotopic enrichment was also observed in newly forming feathers when compared to the old ones. On the other hand, the lower delta15N and Hg values in shag feathers from 2005 point to a shift in feeding resources to prey of lower trophic levels. We found that feather features (being an inert tissue and having a conservative composition), if combined with careful dating and chemical analysis, offer a very useful tool to evaluate temporal and spatial changes in seabird ecology in relation to pollution events.

Analysis of micronucleated erythrocytes in heron nestlings from reference and impacted sites in the Ebro basin (N.E. Spain).

The frequency of micronuclei (MN) in peripheral erythrocytes was tested for 59 heron nestlings (Ardea purpurea, Egretta garzetta and Bubulcus ibis) sampled at two areas (polluted and reference) on the River Ebro (NE Spain) and at its Delta during Spring 2006. Flow-cytometry analysis revealed higher (three- to six-fold) MN counts in samples from the most polluted site relative to samples from the reference area. Samples from the Delta showed intermediate values. Age, morphometric parameters (weight, tarsus size and bill-head length) and maturation status showed no significant differences among the different populations for each species; nor were they correlated with MN levels. The data suggest that elevated levels of MN in chicks in impacted areas reflected the chemical pollution of their nesting sites. The use of nestlings for this assay appears to be a convenient, non-destructive method to assess the impact of pollution in natural bird populations.

Analysis of the ectoenzymes ADA, ALP, ENPP1, and ENPP3, in the contents of ovarian endometriomas as candidate biomarkers of endometriosis.

PROBLEM: The diagnosis of endometriosis, a prevalent chronic disease with a strong inflammatory component, is usually delayed due to the lack of noninvasive diagnostic tests. Purinergic signaling, a key cell pathway, is altered in many inflammatory disorders. The aim of the present work was to evaluate the levels of adenosine deaminase (ADA), alkaline phosphatase (ALP), ecto-nucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1), and ENPP3, elements of purinergic signaling, as biomarker candidates for endometriosis. METHOD OF STUDY: A case-control comparative study was conducted to determine ADA, ALP, ENPP1 and ENPP3 levels in echo-guided aspirated fluids of endometriomas (case group) and simple ovarian cysts (control group) using the ELISA technique. RESULTS: Adenosine deaminase, ALP, ENPP1, and ENPP3 were present and quantifiable in the contents of endometriomas and simple cysts. There were significant differences in ADA and ENPP1 levels in endometriomas in comparison with simple cysts (2787 U/L and 103.9 ng/mL more in endometriomas, for ADA and ENPP1, respectively). Comparisons of ALP and ENPP3 levels between the two groups did not reveal significant differences. CONCLUSION: The ectoenzymes ADA and ENPP1 are biomarker candidates for endometriosis.

Imbalance between endothelial damage and repair capacity in chronic obstructive pulmonary disease.

BACKGROUND: Circulating endothelial microparticles (EMPs) and progenitor cells (PCs) are biological markers of endothelial function and endogenous repair capacity. The study was aimed to investigate whether COPD patients have an imbalance between EMPs to PCs compared to controls and to evaluate the effect of cigarette smoke on these circulating markers. METHODS: Circulating EMPs and PCs were determined by flow cytometry in 27 nonsmokers, 20 smokers and 61 COPD patients with moderate to severe airflow obstruction. We compared total EMPs (CD31+CD42b-), apoptotic if they co-expressed Annexin-V+ or activated if they co-expressed CD62E+, circulating PCs (CD34+CD133+CD45+) and the EMPs/PCs ratio between groups. RESULTS: COPD patients presented increased levels of total and apoptotic circulating EMPs, and an increased EMPs/PCs ratio, compared with nonsmokers. Women had less circulating PCs than men through all groups and those with COPD showed lower levels of PCs than both control groups. In smokers, circulating EMPs and PCs did not differ from nonsmokers, being the EMPs/PCs ratio in an intermediate position between COPD and nonsmokers. CONCLUSIONS: We conclude that COPD patients present an imbalance between endothelial damage and repair capacity that might explain the frequent concurrence of cardiovascular disorders. Factors related to the disease itself and gender, rather than cigarette smoking, may account for this imbalance.

Audouin's gull chicks as bioindicators of mercury pollution at different breeding locations in the western Mediterranean.

Mercury levels of Audouin's gull are amongst the highest for Mediterranean seabirds, and have been mainly attributed to its piscivorous habits in these naturally Hg rich waters. Moreover, two additional factors could enhance its mercury intake: the consumption of discarded fish (which attain higher concentrations) and/or feeding in areas receiving Hg anthropogenic inputs. In order to differentiate the relevance of both sources we analysed Hg and stable isotopes of chick feathers from different breeding locations in western Mediterranean: one in its northern part (Ebro Delta) and two southern (Chafarinas Isl. and Alborán Isl.). The results from stable isotopes indicate that consumption of discards is higher at Alborán Isl., followed by the Ebro Delta and Chafarinas Isl. Thus, the higher mercury levels found in the Ebro Delta cannot be explained uniquely by the contribution of discarded fish to diet, but local pollution caused by the river Ebro waters accounts for Hg differences observed.

Influence of diet in the accumulation of organochlorine compounds in herons breeding in remote riverine environments.

The composition of organochlorine compounds (OCs), pentachlorobenzene (PeCB), hexachlorobenzene (HCB), hexachlorocyclohexanes (HCHs), DDTs and polychlorobiphenyls (PCBs), has been analyzed in eggs from cattle egret (Bubulcus ibis) and little egret (Egretta garzetta), two species of herons (family Ardeidae), nesting at the same remote riverine environment (Aiguabarreig, Ebro River). These two species were selected to evaluate the importance of diet in the accumulation of OCs. Cattle egret essentially feeds on dry grassy habitats and follow cattle or other large animals whereas little egret feeds on fish, amphibians and crustaceans captured in shallow waters. The δ(15)N and δ(13)C isotopic composition of the sampled eggs was studied and the results were consistent with these species feeding habits. In both species, the compounds accumulated the most were the less volatile and more lipophilic, e.g. PCB congeners of higher chlorination, DDT and metabolites. The distinct foraging species preferences were reflected in significant higher concentrations in little egret than cattle egret of all pollutant groups analysed. These differences were statistically significant for DDTs and PCBs (p < 0.015 and p < 0.047, respectively), e.g. the p,p'-DDE and PCB concentrations were 6 and 4.5 times higher, respectively, in the former than the latter. This strong contrast indicates that in remote environments aquatic riverine ecosystems are more efficient OC reservoirs than the terrestrial ecosystem.