Study of nano-hydroxyapatite tagged alkaline protease isolated from Himalayan sub-alpine Forest soil bacteria and role in recalcitrant feather waste degradation.

Purified calcium serine metalloprotease from Stenotrophomonas maltophilia strain SMPB12 exhibits highest enzyme activity at pH 9 and temperature range between 15 °C-25 °C. Enzyme supplemented with 40 µM Ca-Hap-NP (NP-protease) showed maximum elevated activity of 17.29 µmole/min/ml (1.9-fold of original protease activity). The thermostability of the enzyme was maintained for 1 h at 60 °C over an alkaline pH range 7.5-10, as compared to the NP untreated enzyme whose activity was of 8.97 µmole/min/ml. A significant loss of activity with EDTA (1.05 µmole/min/ml, 11.75 %), PMSF (0.93 µmole/min/ml, 10.46 %) and Hg2+ (3.81 µmole/min/ml, 42.49 %) was also observed. Kinetics study of NP-protease showed maximum decreases in Km (28.11 %) from 0.28 mM (NP untreated enzyme) to 0.22 mM (NP-protease) along with maximum increase in Vmax (42.88 %) from 1.25 µmole/min/ml to 1.79 µmole/min/ml at varying temperatures. The enhanced activity of NP-protease was able to efficiently degrade recalcitrant solid wastes like feather to produce value-added products like amino acids and helps in declogging recalcitrant solid wastes. The nano-enabled protease may be utilized in a smaller amount for degrading in bulk recalcitrant solid proteinaceous waste at 15 °C temperature as declogging agents providing an eco-friendly efficient process.

Chryseobacterium aquifrigidense keratinase liberated essential and nonessential amino acids from chicken feather degradation.

Keratinous biomass valorization for value-added products presents a high prospect in ecological management and the advancement of the bio-economy. Consequently, soil samples from the poultry dumpsite were collected. The bacteria isolated on the basal salt medium were screened for keratinolytic activity. The potent chicken feathers degrading bacteria were identified through 16S rRNA gene sequencing and phylogenetic analysis. Fermentation process conditions were optimized, and the amino acid compositions of the feather hydrolysate were likewise quantified. Ten (10) proteolytic bacteria evaluated on skimmed milk agar showed intact chicken feather degradation ranging from 33% (WDS-03) to 88% (FPS-09). The extracellular keratinase activity ranged from 224.52 ± 42.46 U/mL (WDS-03) to 834.55 ± 66.86 U/mL (FPS-07). Based on 16S rRNA gene sequencing and phylogenetic analysis, the most potent keratinolytic isolates coded as FPS-07, FPS-09, FPS-01, and WDS-06 were identified as Chryseobacterium aquifrigidense FANN1, Chryseobacterium aquifrigidense FANN2, Stenotrophomonas maltophilia ANNb, and Bacillus sp. ANNa, respectively. C aquifrigidense FANN2 maximally produced keratinase (1460.90 ± 26.99 U/mL) at 72 h of incubation under optimal process conditions of pH (6), inoculum side (5%; v/v), temperature (30°C), and chicken feather (25 g/L). The feather hydrolysate showed a protein value of 67.54%, with a relative abundance of arginine (2.84%), serine (3.14%), aspartic acid (3.33%), glutamic acid (3.73%), and glycine (2.81%). C. aquifrigidense FANN2 yielded high keratinase titre and dismembered chicken feathers into amino acids-rich hydrolysate, highlighting its significance in the beneficiation of recalcitrant keratinous wastes into dietary proteins as potential livestock feed supplements.

Understanding Transcriptomic and Serological Differences between Forced Molting and Natural Molting in Laying Hens.

Molting is natural adaptation to climate change in all birds, including chickens. Forced molting (FM) can rejuvenate and reactivate the reproductive potential of aged hens, but the effect of natural molting (NM) on older chickens is not clear. To explore why FM has a dramatically different effect on chickens compared with NM, the transcriptome analyses of the hypothalamus and ovary in forced molted and natural molted hens at two periods with feathers fallen and regrown were performed. Additionally, each experimental chicken was tested for serological indices. The results of serological indices showed that growth hormone, thyroid stimulating hormone, and thyroxine levels were significantly higher (p < 0.05) in forced molted hens than in natural molted hens, and calcitonin concentrations were lower in the forced molted than in the natural molted hens. Furthermore, the transcriptomic analysis revealed a large number of genes related to disease resistance and anti-aging in the two different FM and NM periods. These regulatory genes and serological indices promote reproductive function during FM. This study systematically revealed the transcriptomic and serological differences between FM and NM, which could broaden our understanding of aging, rejuvenation, egg production, and welfare issues related to FM in chickens.

Sustainable production, biochemical and molecular characterization of thermo-and-solvent stable alkaline serine keratinase from novel Bacillus pumilus AR57 for promising poultry solid waste management.

The increasing amount of recalcitrant keratinous wastes generated from the poultry industry poses a serious threat to the environment. Keratinase have gained much attention to convert these wastes into valuable products. Ever since primitive feathers first appeared on dinosaurs, microorganisms have evolved to degrade this most recalcitrant keratin. In this study, we identified a promising keratinolytic bacterial strain for bioconversion of poultry solid wastes. A true keratinolytic bacterium was isolated from the slaughterhouse soil and was identified and designated as Bacillus pumilus AR57 by 16S rRNA sequencing. For enhanced keratinase production and rapid keratin degradation, the media components and substrate concentration were optimized through shake flask culture. White chicken feather (1% w/v) was found to be the good substrate concentration for high keratinase production when supplemented with simple medium ingredients. The biochemical characterization reveals astounding results which makes the B. pumilus AR57 keratinase as a novel and unique protease. Optimum activity of the crude enzyme was exhibited at pH 9 and 45 °C. The crude extracellular keratinase was characterized as thermo-and-solvent (DMSO) stable serine keratinase. Bacillus pumilus AR57 showed complete degradation (100%) of white chicken feather (1% w/v) within 18 h when incubated in modified minimal medium supplemented with DMSO (1% v/v) at 150 rpm at 37 °C. Keratinase from modified minimal medium supplemented with DMSO exhibits a half-life of 4 days. Whereas, keratinase from the modified minimal medium fortified with white chicken feather (1% w/v) was stable for 3 h only. Feather meal produced by B. pumilus AR57 was found to be rich in essential amino acids. Hence, we proposed B. pumilus AR57 as a potential candidate for the future application in eco-friendly bioconversion of poultry waste and the keratinase could play a pivotal role in the detergent industry. While feather meal may serve as an alternative to produce animal feed and biofertilizers.

Do glucocorticoids or carotenoids mediate plumage coloration in parrots? An experiment in Platycercus elegans.

Conspicuous coloration can indicate phenotypic quality, and may reflect exposure or vulnerability to stress, or access to essential nutrients such as pigments. Although the production of pigmented colours is well understood, much less is known about how structural colours are affected by physiological state. In this study, we tested whether glucocorticoids (corticosterone) predicted expression of plumage coloration in an Australian parrot, the crimson rosella (Platycercus elegans). Parrots provide an interesting and unique test, as they possess conspicuous coloration produced by distinctive pigments known as psittacofulvins, in addition to structural coloration. We have previously documented that coloration in P. elegans is condition-dependent and responds to dietary manipulation. Here, n = 21 P. elegans underwent a dietary manipulation (including food restriction or carotenoid supplementation) during which they moulted, and the change in reflectance was measured for three structural and three pigmentary plumage patches. Stress-induced corticosterone (10 min after handling) measured at the start of the experiment predicted change in coloration in two pigmentary patches (crown and front). We also found that change in stress-induced corticosterone during the experiment was associated with the change in coloration of the crown and two structural patches (cheek and epaulette). Baseline corticosterone (<3 min after handling) was not associated with any measure of coloration. We found no effects of dietary manipulation on baseline or stress-induced corticosterone, but carotenoid supplementation was associated with an increase in a measure of chronic stress (heterophil/lymphocyte ratio), and the corticosterone response to handling decreased over the course of the study. Our results suggest that corticosterone may be linked to colour expression more broadly than previously recognised, including psittacofulvin and structural coloration in parrots, and they confirm the independence of plumage pigmentation in parrots from carotenoid accumulation. Moreover, our study provides new insight into the stress responses of Psittaciformes, one of the most highly threatened avian orders.

Strategical isolation of efficient chicken feather-degrading bacterial strains from tea plantation soil sample.

Chicken feather waste is generally insufficiently utilized despite its high content of protein, constituting an environmental issue. Biodegradation of the waste with enabling microbes provides an advantageous option among the available solutions. In this study, an efficient whole feather-degrading strain was strategically isolated from a soil sample taken from a local tea plantation that has little or nothing to do with feathers. The strain was identified as Bacillus thuringiensis (designated as FDB-10) according to the cloned complete 16S rRNA sequence. The FDB-10 could efficiently degrade briefly heat-treated whole feather (102 °C, 5 min; up to 90% of a maximum concentration of 30 g/L) in a salt medium supplemented with 0.1 g/L yeast extract within 24 h (37 °C, 150 rpm). Addition of carbon sources (glycerol, glucose, starch, Tween 20, Tween 80, 1.25 g/L as glycerol) to the fermentation medium could improve the degradation. However, significant inhibition could be observed when the added carbon source reached the amount usually adopted in the investigation of carbon source preference (1%). Nitrogen source (NH4Cl, (NH4)2SO4, peptone) adversely influenced the performance of the strain. When the molar concentrations of NH4+ were equal for the two salt, the inhibitory effect on degradation of whole feathers was similar. Entirely different from other reported feather-degrading strains showing a preference to melanin-free feather substrates, the strain isolated in this study could degrade melanin-containing feather equally efficiently, and higher protease activity could be detected in the digest mix. As a plus, the strain could degrade feathers in rice wash produced in daily cooking, indicating its potential use in the simultaneous treatment of rice cooker wastewater produced by a rice processing plant. All these results imply that the FDB-10 is a strain with great potential in the biodegradation of feather waste.

Mercury (Hg), Lead (Pb), Cadmium (Cd), Selenium (Se), and Arsenic (As) in Liver, Kidney, and Feathers of Gulls: A Review.

Mercury (Hg), lead (Pb), cadmium (Cd), selenium (Se), and arsenic (As) are metals or metalloids of high concern because of their effects on the environment and, specially, their potential toxicity on the animals inhabiting there. Due to their relevance, these elements have been object of several biomonitoring studies in different animal species around the world. Birds are widespread and, as species, are able to supply specific and relevant information about the regions where they live, being useful as bioindicators, as long as they are not birds with a strong migratory character. The main goal of this review is to summarize data collected from different studies using seabirds, paying special attention to gulls, in order to be helpful for coming studies and regulatory affairs.Several tissues have been used to evaluate Hg, Cd, Pb, Se, and As concentrations in seabirds, being focused the present review in those analyzing the liver, kidneys, and feathers. The most frequently analyzed tissue for Hg was the liver, followed by feathers, and finally kidney. For Cd levels, most of the studies were carried out in the liver, followed by feathers and kidneys. Pb, Se, and As levels were determined to a lesser extent. Feathers should be taken carefully as indicator of accumulation of pollutants, since procedure during analysis may lead to controversial results.Some authors reported that interspecific differences in the exposure of elements are determined by multiple factors, including properties of the contaminant, species, feeding habits, migratory status, sex, and age.The present review provides a comprehensive overview of the analyzed elements' occurrence in different species of seabirds, including gulls. Therefore, it can be a useful database providing for Hg, Pb, Cd, Se, and As levels in different tissues of seabirds.

Production and Characterization of Keratinolytic Proteases by a Chicken Feather-Degrading Thermophilic Strain, Thermoactinomyces sp. YT06.

Thermoactinomyces sp. strain YT06 was isolated from poultry compost and observed to degrade integral chicken feathers completely at 60°C, resulting in the formation of 3.24 mg/ml of free amino acids from 50 ml of culture containing 10 g/l chicken feathers. Strain YT06 could grow and secrete keratinase using feather as the only carbon and nitrogen sources without other supplement, but complementation of 10 g/l sucrose and 4 g/l NaNO3 increased the production of the keratinolytic enzyme. The maximum protease activity obtained was 110 U/ml and for keratinase was 42 U/ml. The keratinase maintained active status over a broad pH (pH 8-11) and temperature (60-75°C). It was inhibited by serine protease inhibitors and most metal ions; however, it could be stimulated by Mn²âº and the surfactant Tween-20. A reductive agent (ß-mercaptoethanol) was observed to cleave the disulfide bond of keratin and improve the access of the enzyme to the keratinaceous substrate. Zymogram analysis showed that strain YT06 primarily secreted keratinase with a molecular mass of approximately 35 kDa. The active band was assessed by MALDI-TOF mass spectrometry and was observed to be completely identical to an alkaline serine protease from Thermoactinomyces sp. Gus2-1. Thermoactinomyces sp. strain YT06 shows great potential as a novel candidate in enzymatic processing of hard-to-degrade proteins into high-value products, such as keratinous wastes.

Maternal transfer of contaminants in birds: Mercury and selenium concentrations in parents and their eggs.

We conducted a detailed assessment of the maternal transfer of mercury and selenium to eggs in three bird species (n = 107 parents and n = 339 eggs), and developed predictive equations linking contaminant concentrations in eggs to those in six tissues of the mother (blood, muscle, liver, kidney, breast feathers, and head feathers). Mercury concentrations in eggs were positively correlated with mercury concentrations in each of the mother's internal tissues (R(2) ≥ 0.95), but generally not with feathers. For each species, the proportion of mercury transferred to eggs decreased as mercury concentrations in the mother increased. At the same maternal mercury concentration, the proportion of mercury transferred to eggs differed among species, such that Forster's tern (Sterna forsteri) and black-necked stilt (Himantopus mexicanus) females transferred more methylmercury to their eggs than American avocet (Recurvirostra americana) females. Selenium concentrations in eggs also were correlated with selenium concentrations in the mother's liver (R(2) = 0.87). Furthermore, mercury and selenium concentrations in tern eggs were positively correlated with those in the father (R(2) = 0.84). Incubating male terns had 21% higher mercury concentrations in blood compared to incubating females at the same egg mercury concentration. We provide equations to predict contaminant concentrations in eggs from each of the commonly sampled bird tissues.

Selenium deposition kinetics of different selenium sources in muscle and feathers of broilers.

The objective of this study was to determine selenium (Se) deposition kinetics in muscles and feathers of broilers in order to develop a rapid method to compare bioavailability of selenium sources. Different Se sources such as 2-hydroxy-4-methylselenobutanoic acid (HMSeBA, SO), sodium selenite (SS) and seleno-yeast (SY) were compared for their kinetics on Se deposition in muscles and feathers in broiler chicks from 0 to 21 d of age. A total of 576 day-old broilers were divided into four treatments with 8 replicates of 18 birds per pen. The diets used in the experiment were a negative control (NC) not supplemented with Se and 3 diets supplemented with 0.2 mg Se/kg as SS, SY or SO. Total Se content in breast muscle and feathers were assessed on days 0, 7, 14 and 21. At 7 d of age, SO increased muscle Se content compared to D0 (P < 0.05), whereas with the other treatments, muscle Se concentration decreased (P < 0.05). After 21 days, organic Se sources maintained (SY) or increased (SO) (P < 0.05) breast muscle Se concentration compared to hatch value whereas inorganic source (SS) or non-supplemented group (NC) showed a significant decrease in tissue Se concentration (P < 0.05). At D21, Se contents of muscle and feathers were highly correlated (R(2) = 0.927; P < 0.0001). To conclude, these results indicate that efficiency of different Se sources can be discriminated through a 7 d using muscle Se content in broiler chickens. Muscle and feathers Se contents were highly correlated after 21 days. Also feather sampling at 21 days of age represents a reliable and non-invasive procedure for Se bioefficacy comparison.

Trace-elements, methylmercury and metallothionein levels in Magellanic penguin (Spheniscus magellanicus) found stranded on the Southern Brazilian coast.

Magellanic penguins have been reported as good biomonitors for several types of pollutants, including trace-elements. In this context, selenium (Se), total mercury, methylmercury, inorganic mercury (Hg(inorg)), cadmium (Cd) and lead (Pb), as well as metallothionein (MT) levels, were evaluated in the feathers, liver and kidney of juvenile Magellanic penguins found stranded along the coast of Southern Brazil. The highest concentrations of all trace-elements and methylmercury were found in internal organs. Concentrations of Cd and Se in feathers were extremely low in comparison with their concentrations in soft tissues. The results showed that both Se and MT are involved in the detoxification of trace-elements (Cd, Pb and Hg(inorg)) since statistically significant relationships were found in liver. Conversely, hepatic Se was shown to be the only detoxifying agent for methylmercury.

Emergence of differentially regulated pathways associated with the development of regional specificity in chicken skin.

BACKGROUND: Regional specificity allows different skin regions to exhibit different characteristics, enabling complementary functions to make effective use of the integumentary surface. Chickens exhibit a high degree of regional specificity in the skin and can serve as a good model for when and how these regional differences begin to emerge. RESULTS: We used developing feather and scale regions in embryonic chickens as a model to gauge the differences in their molecular pathways. We employed cosine similarity analysis to identify the differentially regulated and co-regulated genes. We applied low cell techniques for expression validation and chromatin immunoprecipitation (ChIP)-based enhancer identification to overcome limited cell availabilities from embryonic chicken skin. We identified a specific set of genes demonstrating a high correlation as being differentially expressed during feather and scale development and maturation. Some members of the WNT, TGF-beta/BMP, and Notch family known to be involved in feathering skin differentiation were found to be differentially regulated. Interestingly, we also found genes along calcium channel pathways that are differentially regulated. From the analysis of differentially regulated pathways, we used calcium signaling pathways as an example for further verification. Some voltage-gated calcium channel subunits, particularly CACNA1D, are expressed spatio-temporally in the skin epithelium. These calcium signaling pathway members may be involved in developmental decisions, morphogenesis, or epithelial maturation. We further characterized enhancers associated with histone modifications, including H3K4me1, H3K27ac, and H3K27me3, near calcium channel-related genes and identified signature intensive hotspots that may be correlated with certain voltage-gated calcium channel genes. CONCLUSION: We demonstrated the applicability of cosine similarity analysis for identifying novel regulatory pathways that are differentially regulated during development. Our study concerning the effects of signaling pathways and histone signatures on enhancers suggests that voltage-gated calcium signaling may be involved in early skin development. This work lays the foundation for studying the roles of these gene pathways and their genomic regulation during the establishment of skin regional specificity.

Evaluating the stress response as a bioindicator of sub-lethal effects of crude oil exposure in wild house sparrows (Passer domesticus).

Petroleum can disrupt endocrine function in humans and wildlife, and interacts in particularly complex ways with the hypothalamus-pituitary-adrenal (HPA) axis, responsible for the release of the stress hormones corticosterone and cortisol (hereafter CORT). Ingested petroleum can act in an additive fashion with other stressors to cause increased mortality, but it is not clear exactly why--does petroleum disrupt feedback mechanisms, stress hormone production, or both? This laboratory study aimed to quantify the effects of ingested Gulf of Mexico crude oil on the physiological stress response of house sparrows (Passer domesticus). We examined baseline and stress-induced CORT, negative feedback, and adrenal sensitivity in house sparrows given a 1% oil or control diet (n = 12 in each group). We found that four weeks on a 1% oil diet did not alter baseline CORT titers or efficacy of negative feedback, but significantly reduced sparrows' ability to secrete CORT in response to a standardized stressor and adrenocorticotropin hormone injection, suggesting that oil damages the steroid-synthesizing cells of the adrenal. In another group of animals on the same 1% oil (n = 9) or control diets (n = 8), we examined concentrations of eight different blood chemistry parameters, and CORT in feathers grown before and during the feeding experiments as other potential biomarkers of oil exposure. None of the blood chemistry parameters differed between birds on the oil and control diets after two or four weeks of feeding, nor did feather CORT differ between the two groups. Overall, this study suggests that the response of CORT to stressors, but not baseline HPA function, may be a particularly sensitive bioindicator of sub-lethal chronic effects of crude oil exposure.

Effects of carotenoid and vitamin E supplementation on oxidative stress and plumage coloration in house finches (Haemorhous mexicanus).

There has been much recent interest from both applied and basic scientists in the broad series of benefits that animals reap from acquiring high concentrations of dietary antioxidants, such as carotenoids and vitamins (e.g., vitamin E, or tocopherol). Most attention has been paid to separate effects of these compounds on, for example, coloration, health state, development, and vision, but because of possible interactions between these lipid-soluble molecules, we are in need of more studies that co-manipulate these substances and examine their possible synergistic impacts on animal physiology and phenotype. We capitalized on a model avian system (the house finch, Haemorhous mexicanus), where extensive information is available on the fitness roles of carotenoids, to test how variation in carotenoid and/or vitamin E concentrations in the diet impacts body accumulation of these compounds, factors related to oxidative damage (e.g., breast muscle and plasma oxidative-stress susceptibility, plasma nitric-oxide levels), and plumage color development. As in a previous study of ours on carotenoids and health in finches, we employed a 2×2 factorial experimental design on birds in both molting and non-molting conditions, to understand how seasonal shifts in carotenoid use (i.e., pigment incorporation into plumage) might alter the accumulation and roles of carotenoids and vitamins. As expected, lutein supplementation increased the level of circulating carotenoids in both experiments and the color of newly molted plumage. By contrast, vitamin E provisioning did not significantly affect plasma carotenoid levels or plumage coloration in either experiment. Interestingly, carotenoid provisioning decreased circulating vitamin E levels during molt, which suggests either molecular competition between carotenoids and tocopherol at the absorption/transport stages or that vitamin E serves as an antioxidant to offset harmful actions that carotenoids may have at very high concentrations. Finally, in both experiments, we found a reduction in breast-muscle oxidative damage for tocopherol-supplemented birds, which constitutes the first demonstration of a protective effect of vitamin E against oxidative stress in wild birds. Taken together, these findings provide an interesting contrast with our earlier work on season-specific physiological benefits of carotenoids in finches and point to complex associations between indicators of antioxidant and oxidative state in wild-caught animals.

Metal and selenium concentrations in blood and feathers of petrels of the genus Procellaria.

Concentrations of copper (Cu), zinc (Zn), cadmium (Cd), lead (Pb), mercury (Hg), and selenium (Se) were determined in blood and feathers of spectacled (Procellaria conspicillata) and white-chinned (Procellaria aequinoctialis) petrels, species that are phylogenetically related, but with distinct ecological niches. In winter, they feed on similar foods, indicated by an overlapping range of whole-blood stable isotopes values (δ(15) N; δ(13) C). No relation was found between blood metal concentration and stable isotope values. In spectacled petrels, metal concentrations appeared lower in blood (Cu = 0.79-20.77 µg/g; Zn = 10.95-28.02 µg/g; Cd = 1.73-10.11 µg/g; Pb = 5.02-26.03 µg/g; Hg = 0.84-9.86 µg/g) than in feathers (Cu = 1.05-21.57 µg/g; Zn = 45.30-81.49 µg/g; Cd = 3.76-10.44 µg/g; Pb = 16.53-59.00 µg/g; Hg = 4.24-24.03 µg/g). In white-chinned petrels, metal concentrations also appeared lower in blood (Cu = 0.62-10.4 µg/g; Zn = 10.73-24.69 µg/g; Cd = 2.00-6.31 µg/g; Pb = 5.72-24.03 µg/g) than in feathers (Cu = 2.68-23.92 µg/g; Zn = 48.96-93.54 µg/g; Cd = 5.72-24.03 µg/g; Pb = 18.62-55.51 µg/g), except for Hg (blood = 0.20-15.82 µg/g; feathers = 0.19-8.91 µg/g). Selenium (0.24-14.18 µg/g) and Hg (0.22-1.44 µg/g) concentrations showed a positive correlation in growing feathers of spectacled petrels. Blood and feather Hg levels were higher in spectacled petrels while feathers Cu and Zn concentrations were greater in white-chinned petrels. Juvenile white-chinned petrels exhibited greater blood Hg concentrations than adults. In the south Atlantic Ocean, discards from commercial fishing operations consumed by spectacled petrels year-round and by white-chinned petrels during the wintering period have elevated Hg concentrations. Because Hg toxicity is associated with behavioral and reproductive changes in birds, it could potentially have impacts on breeding of these seabirds, as both species are listed as threatened by extinction.

Independent sources of condition dependency and multiple pathways determine a composite trait: lessons from carotenoid-based plumage colouration.

Many colour ornaments are composite traits consisting of at least four components, which themselves may be more complex, determined by independent evolutionary pathways, and potentially being under different environmental control. To date, little evidence exists that several different components of colour elaboration are condition dependent and no direct evidence exists that different ornamental components are affected by different sources of variation. For example, in carotenoid-based plumage colouration, one of the best-known condition-dependent ornaments, colour elaboration stems from both condition-dependent pigment concentration and structural components. Some environmental flexibility of these components has been suggested, but specifically which and how they are affected remains unknown. Here, we tested whether multiple colour components may be condition dependent, by using a comprehensive 3 × 2 experimental design, in which we carotenoid supplemented and immune challenged great tit nestlings (Parus major) and quantified effects on different components of colouration. Plumage colouration was affected by an interaction between carotenoid availability and immune challenge. Path analyses showed that carotenoid supplementation increased plumage saturation via feather carotenoid concentration and via mechanisms unrelated to carotenoid deposition, while immune challenge affected feather length, but not carotenoid concentration. Thus, independent condition-dependent pathways, affected by different sources of variation, determine colour elaboration. This provides opportunities for the evolution of multiple signals within components of ornamental traits. This finding indicates that the selective forces shaping the evolution of different components of a composite trait and the trait's signal content may be more complex than believed so far, and that holistic approaches are required for drawing comprehensive evolutionary conclusions.

Toxicity of cadmium and lead in Gallus gallus domesticus assessment of body weight and metal content in tissues after metal dietary supplements.

The influence of dietary cadmium on the accumulation and effects of dietary lead, examined in chicken. This experiment was conducted to investigate the toxic effects of dietary Cd and Pb on chick's body weight and organ, content of the tissues of these two metals was also detected. One day age chicks of Gallus gallus domesticus fed diet supplemented with 25, 50, 100 ppm of Cd, second group exposure to 300, 500, 1000 ppm of Pb in feed daily during 4 weeks. The control groups were fed without supplementation of metals. The concentrations of Cd and Pb resulted in increased of Cd and Pb content in liver, gizzard and muscle. While Cd 100 ppm and Pb 1000 ppm were increased metals content in feather. Body weight of chicks was not influenced by Cd treatment. In contrary Pb treatment was significantly (p < 0.05) decreased body weight of chicks after dietary treatment. On the other hand, Liver weigh in chicks was significantly (p < 0.05) decreased after Cd and Pb treatments.

Effects of pre-treatment and bioaugmentation strategies on the anaerobic digestion of chicken feathers.

Anaerobic digestion of raw chicken feather waste and its co-digestion with poultry litter were assessed in batch assays. Following, two strategies were evaluated to improve methane production from chicken feathers: (i) waste pre-hydrolysis through thermochemical treatment using lime and sodium hydroxide, and (ii) amendment of digestion broth with the proteolytic bacterium Fervidobacterium pennivorans. Anaerobic digestion of the raw waste (2.5% total solids) allowed a specific methane production of 123 ± 3 L CH(4) kg(-1) VS. Pre-treatment and bioaugmentation strategies did not improve methane production from feather waste, despite the significant increase in waste solubilisation, from 45 ± 5% up to 64 ± 1% using F. pennivorans and up to 96% after pre-treatment with 2g NaOH g(-1) waste. These results indicate that conversion of soluble organic matter to methane, and not the hydrolysis rate, was the limiting step for the anaerobic digestion of chicken feather waste.

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.

Insight on trace element detoxification in the Black-tailed Godwit (Limosa limosa) through genetic, enzymatic and metallothionein analyses.

Trace element concentrations (Ag, As, Cd, Co, Cr, Cu, Fe, Hg, Mn, Ni, Pb, Se, Zn) were investigated in the liver, kidneys, muscle and feathers of 31 black-tailed godwits (Limosa limosa) accidentally killed during catches by mist net in the Pertuis Charentais, Atlantic coast of France. Analyses of carbon and nitrogen stable isotope ratios were carried out in liver, muscle and feathers in order to elucidate dietary patterns and to determine whether differences in diet explained the variation in elemental uptake. This study also aimed to have a preliminary assessment of sub-lethal effects triggered by trace elements through the investigation of gene expressions by quantitative real-time PCR, antioxidant enzyme activities (catalase, superoxide dismutase, glutathione peroxidase), and metallothionein (MT) levels. The results showed that Cr and Ni concentrations in tissues of adults were lower than in juveniles in part because adults may have eliminated these trace elements through moulting. Except for Cd and Ni, trace element concentrations were negatively correlated to the body mass of godwits. Ag, As, Hg and Se concentrations were positively linked with the trophic position of birds. The diet could be considered as a fundamental route of exposure for these elements demonstrating therefore the qualitative linkage between dietary habits of godwits and their contaminant concentrations. Our results strongly suggest that even though trace element concentrations were mostly below toxicity threshold level, the elevated concentrations of As, Ag, Cd, Cu, Fe and Se may however trigger sub-lethal effects. Trace elements appear to enhance expression of genes involved in oxidative stress defence, which indicates the production of reactive oxygen species. Moreover, birds with the highest concentrations appeared to have an increased mitochondrial metabolism suggesting that the fight against trace element toxicity requires additional energetic needs notably to produce detoxification mechanisms such as metallothioneins.