Model study on the clinical signs and residue concentrations of sublethal carbofuran poisoning in birds

The incidence of fatal poisoning of birds of prey caused by carbofuran has increased markedly in Hungary since 2007. An experimental model with broiler chickens was used to study clinical signs of sublethal carbofuran poisoning in birds and to measure the residue concentrations of carbamate in tissues after exposure. Eight chickens were treated with a carbofuran-containing insecticide orally by gastric tube at a single dose of 2.5 mg/kg body weight, and clinical signs of poisoning were observed. Gas chromatography was used to determine carbofuran concentrations in the blood, muscle, and liver samples, and in stomach contents. Poisoning was characterized by typical muscarinic and nicotinic clinical signs without mortality. Carbofuran in the stomach and edible tissues of acutely poisoned birds may lead to secondary poisoning of predators and may also present risks to human health.     

Pesticide poisoning events in wild birds in Korea from 1998 to 2002

We describe cases of pesticide poisoning of wild birds diagnosed at the National Veterinary Research and Quarantine Service (Kyunggi, Korea) from 1998 to 2002. Forty-one mortality events (759 birds) of 87 incidents (2,464 birds) investigated were associated with pesticide poisoning, and six organophosphates or carbamates were identified as being responsible for the poisoning. Phosphamidon was most frequently identified as the cause of poisoning, accounting for 23 mortality events. Other pesticides identified as poisons for birds were organophosphates monocrotophus, fenthion, parathion, EPN, and diazinon, and the carbamate carbofuran. Pesticide poisoning is a problem in wild birds in Korea.     

Oxidative Stress and Metabolic Perturbations in Wooden Breast Disorder in Chickens

This study was conducted to characterize metabolic features of the breast muscle (pectoralis major) in chickens affected with the Wooden Breast myopathy. Live birds from two purebred chicken lines and one crossbred commercial broiler population were clinically examined by manual palpation of the breast muscle (pectoralis major) at 47–48 days of age. Metabolite abundance was determined by gas chromatography/mass spectrometry (GC/MS) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) using breast muscle tissue samples from 16 affected and 16 unaffected chickens. Muscle glycogen content was also quantified in breast muscle tissue samples from affected and unaffected chickens. In total, levels of 140 biochemicals were significantly different (FDR < 0.1 and fold-change A/U > 1.3 or < 0.77) between affected and unaffected chickens. Glycogen content measurements were considerably lower (1.7-fold) in samples taken from Wooden Breast affected birds when compared with samples from unaffected birds. Affected tissues exhibited biomarkers related to increased oxidative stress, elevated protein levels, muscle degradation, and altered glucose utilization. Affected muscle also showed elevated levels of hypoxanthine, xanthine, and urate molecules, the generation of which can contribute to altered redox homeostasis. In conclusion, our findings show that Wooden Breast affected tissues possess a unique metabolic signature. This unique profile may identify candidate biomarkers for diagnostic utilization and provide mechanistic insight into altered biochemical processes contributing to tissue hardening associated with the Wooden Breast myopathy in commercial chickens.     

Case report: secondary poisoning in a white-tailed sea eagle caused by carbofuran

A juvenile white-tailed sea eagle found alive on an island in the Baltic Sea died due to a secondary carbofuran poisoning. At necropsy, parts of a raven were found in the eagle’s gizzard, including the stomach containing seeds with carbofuran residues. Illegal persecution using poisoned baits often kill scavenging birds of prey. However, secondary poisoning of birds of prey has been rarely reported in Europe. The report aims to raise awareness to include toxicological analysis in forensic examinations in cases of otherwise healthy and well-feed birds found dead with no obvious cause of mortality.     

Muscle fat composition of pheasants (<Emphasis Type="Italic">Phasianus colchicus)</Emphasis>, wild ducks (<Emphasis Type="Italic">Anas platyrhynchos</Emphasis>) and black coots (<Emphasis Type="Italic">Fulica atra</Emphasis>)

The aim of this investigation was to characterise the fatty acid composition within intramuscular fat (IMF) of two muscles (breast and thigh) from 28 pheasants, ten wild ducks and 27 black coots from Slovakia. A high variability for all single fatty acids (FA) and the total fat concentration in muscles of wild birds was identified. Black coots deposited the highest fat in breast muscle whereas wild ducks and pheasants accumulated more lipids in thigh muscle. In general, the content of polyunsaturated fatty acids (PUFA) of the IMF in wild bird muscles was high, and the saturated FA concentration was lower compared with muscles of domestic farm animals. The ratio between PUFA and saturated fatty acids (PSQ) ranked between 0.6 and 1.2, and the ratio of n-6/n-3 fatty acid was favourably low in black coot and wild pheasants (3.2 and 2.9, respectively). Farmed pheasants had increased IMF and more saturated and n-6 fatty acids deposited in thigh muscle but not in breast muscle.     

The effect of treating method of game on the content of biogenic amines in wild duck (Anas platyrhynchos) meat during the course of storage

The aim of present study was to assess the correlation between the method of treating the carcasses of shot wild ducks (Anas platyrhynchos) and the formation of biogenic amines in the their muscles. The carcasses of wild ducks (n?=?180) were divided into three groups of 60 carcasses according to the method of treatment: eviscerated, drawn, and left untreated. Each group was further divided into three subgroups of 20 duck carcasses on the basis of the storage temperature (0, 7, and 15 °C) and stored for 21 days. Samples of breast and thigh muscles were taken at regular weekly intervals. Biogenic amines (cadaverine, putrescine, tyramine, histamine, phenylethylamine, and tryptamine) in samples of breast and thigh muscles were separated by reverse-phase liquid chromatography and detected by tandem mass spectrometry. The sum of biogenic amines was compared with a value of 5 mg/kg, indicating the critical content for fresh meat of high hygienic quality. The results of this study indicated that the sum of biogenic amines in wild duck meat exceeded this limit in an extremely short period of time after the commencement of storage (during the first week of storage). Higher content of biogenic amines were recorded in thigh muscle compared to breast muscle of drawn ducks and untreated ducks. According to our results, the generally recommended method for treating the carcasses of feathered game after hunting (evisceration) does not represent a method that would ensure a longer period of freshness or higher hygiene quality of the game than the other two possible methods of treatment from the biogenic amines point of view.     

Sex and storage affect cholinesterase activity in blood plasma of Japanese quail

Blood plasma cholinesterase (ChE) activity is a sensitive indicator of exposure to organophosphorus and carbamate insecticides. Effects of sex and storage of samples were studied as sources of variability by treating breeding Japanese quail (Coturnix japonica) with 3 mg of dicrotophos or carbofuran per kg of body weight and comparing blood plasma ChE activities for samples collected at 1 hr postdosage and assayed fresh, after 1 and 2 days of refrigeration (4 C), and after 1, 7 and 28 days of freezing (-25 C). ChE activity of fresh control plasma was 34% (P less than 0.01) higher in males than females. Male ChE activity remained essentially unchanged during storage while female ChE activity increased (P less than 0.05) gradually over time under both storage conditions. In contrast, when plasma ChE activity was inhibited by either antiChE, male plasma ChE activity was depressed further than female ChE (P less than 0.01) and remained constant during storage while female ChE activity continued to decrease (P less than 0.05). These divergent effects of exposure to antiChE compounds and sample storage indicate extreme care should be exercised in use of blood plasma for evaluation of antiChE exposure in wild birds.     

The level of major proteins and minerals in the blood serum of chickens fed diets with pure cellulose

The aim of the research was to determine the concentration of total protein and its fractions as well as the concentration of selected mineral components in the blood serum of male broiler chickens Cobb 500 fed diets with different cellulose content. Blood samples were collected for examination from the birds' pterygoid canal veins on their 42 day of age. There was no influence of cellulose preparation on the content of total protein and its fractions: albumins, alfa1-, alfa2-, beta-, gamma-globulins, albumin to globulin ratio, inorganic phosphorus, sodium, potassium, chloride and iron concentrations in blood serum. The highest calcium concentration (P < 0.05) was detected in the blood serum of males fed a diet with the highest cellulose content (0.75-0.95%). Simultaneously, a tendency of increased calcium content was observed along with an increased amount of pure cellulose in diets. The lowest magnesium level (P < 0.05) was observed in the blood of birds fed diets with the lowest amount of cellulose (0.25-0.45%). The magnesium level in the blood of birds fed a diet with higher amounts of cellulose did not considerably differ from the control group. Results from the current study suggest that introduction of a limited amount of pure cellulose into the diet of broiler chickens does not affect total protein concentration and protein fractions but can influence the mineral content in the blood serum.     

Thyroidal influence on body growth

This review of thyroid influence on body growth in poultry is organized around the following parameters of growth: increase in body weight and skeletal size, muscle growth, and growth of cartilage and bone. The greatest effect of goitrogens on growth of embryos occurs during late embryogenesis at a time when normal thyroid hormone levels are increasing. Posthatching growth is reduced in severely hypothyroid animals, and body weight gain is affected more than bone growth. Thyroid hormone replacement restores body growth of thyroidectomized chickens, but supplemental hormone in normal animals has no beneficial effect on growth. Excessive T3 (fed at 1 ppm) is detrimental to growth and feed efficiency. No clear correlation between thyroid hormone concentration and growth rate of normal chickens has been identified. Growth depression in sex-linked dwarf birds is at least partially reversed by supplemental T3. Muscle growth is reduced in goitrogen-treated chickens and the growth reduction is reversed by supplemental thyroxine. Total DNA accumulation is reduced in hypothyroid chickens, but muscle mass relative to DNA content is normal following long-term treatment; this suggests some regulation of muscle mass relative to DNA content. T3 increases the number of muscle fiber nuclei in hypothyroid chickens and the uptake of 3H-thymidine into nuclei within the basal lamina. T3 directly stimulates growth and maturation of embryonic chick cartilage and enhances the in vitro action of somatomedins on cartilage growth. There is little information concerning the role of the thyroid in posthatching cartilage and bone growth in poultry.     

Impaired development of broiler chickens by stress mimicked by corticosterone exposure

The effects of corticosterone (CORT) administration on the development of muscular tissues of broiler chickens (Gallus gallus domesticus) fed with diets differing in lipid content were investigated. The experimental chickens were given one of two experimental diets: high lipid diet (9.9% crude fat) or control diet, from 21 d of age. At 28 d of age, half of the chickens in each dietary treatment were exposed to CORT treatment, supplemented with 30 mg CORT/kg diet for 12 days, while the other half continued to consume the former diet. The zootechnical parameters were recorded at 21, 28, 35 and 39 d, and a blood sample was obtained from 8 birds of each group, respectively. The growth performance of broiler chickens was significantly depressed by CORT administration, but not by dietary treatment. Corticosterone treatment resulted in enhanced energy expenditure. The results indicate that the development of breast muscle was more susceptible to stress mimicked by CORT administration. The results suggest that corticosterone administration enhanced hepatic fatty acid synthesis and resulted in the redistribution of energy to abdominal store from peripheral tissues. Diet rich in lipid content was favorable to the central fat deposit in stressed broiler chickens.     

Toxicity of some insecticides to the haemocytes of giant honeybee,Apis dorsata F. under laboratory conditions

Quantitative studies concerning total and differential haemocyte counts and abnormalities were performed under laboratory conditions for larvae, pupae and adults collected from a wild Apis dorsata colony. Haemolymph samples were observed immediately, thirty and sixty minutes after field recommended concentration exposure of five different insecticides. Total haemocyte counts were significantly higher for larvae and pupae but less for adult bees, however, differential haemocyte counts insignificantly different. Exposure of insecticides showed variable response for tested insecticides with immediate increased change with ethofenprox, diafenthiuron and imidacloprid but decreased for all tested insecticides after sixty minutes. For differential haemocyte counts, plasmatocytes and granulocytes increased with exposure of insecticides. Immune response of haemocytes against insecticides showed different degrees of abnormalities like agglutination, denucleation and cell shape distortion. Such studies may help in possible identification of insect defense mechanisms against their exposure to external hazards for instance insecticide exposure.     

Validation and novel applications of the whole-blood chemiluminescence assay of innate immune function in wild vertebrates and domestic chickens

The whole-blood chemiluminescence (WBCL) assay is a simple and rapid method of measuring production of reactive oxygen species by circulating leukocytes, particularly heterophils (birds) and neutrophils (other vertebrates). In the interest of substantiating a broadly applicable measure of innate immunity, we investigated the microplate WBCL method for several wildlife species as well as domestic broiler chickens. Lucigenin as a light enhancer was used for all avian blood, wild and domestic, while luminol was used in bear and frog blood. Use of ethylenedinitrilotetraacetic acid (EDTA) as the anticoagulant caused hemolysis of frog blood and decreased WBCL responses in all animals tested. Heparin, even in high concentrations, caused modest to no decrease of WBCL responses. The WBCL response correlated highly with heterophil or neutrophil numbers in all species tested. The WBCL response was tested in freshly collected blood as well as in blood one to five days postcollection in order to determine the utility of this assay for field studies when immediate access to laboratory facilities is not possible. One to three days of delay in performing the test after blood collection caused no, or only a slight, decay of the chemiluminescence response in most animals. Using domestic chickens, we tested the sensitivity of the WBCL method to detect differences between treatment groups and looked for loss of chemiluminescence response over several days using the original blood samples. Significant differences in the WBCL response between experimental groups of broiler chicken were detectable in freshly collected blood, as well as one- to four-day-old blood. Our results show that the innate immune response of populations of wild animals may be successfully compared using this assay, even when blood cannot be tested until a few days after collection.     

Breast muscle and plasma metabolomics profile of broiler chickens exposed to chronic heat stress conditions

Understanding the variations of muscle and plasma metabolites in response to high environmental temperature can provide important information on the molecular mechanisms related to body energy homeostasis in heat-stressed broiler chickens. In this study, we investigated the effect of chronic heat stress conditions on the breast muscle (Pectoralis major) and plasma metabolomics profile of broiler chickens by means of an innovative, high-throughput analytical approach such as the proton nuclear magnetic resonance (¹H NMR) spectrometry. A total of 300 Ross 308 male chicks were split into two experimental groups and raised in either thermoneutral conditions for the entire rearing cycle (0–41 days) (TNT group; six replicates of 25 birds/each) or exposed to chronic heat stress conditions (30 °C for 24 h/day) from 35 to 41 days (CHS group; six replicates of 25 birds/each). At processing (41 days), plasma and breast muscle samples were obtained from 12 birds/experimental group and then subjected to ¹H NMR analysis. The reduction of BW and feed intake as well as the increase in rectal temperature and heterophil: lymphocyte ratio confirmed that our experimental model was able to stimulate a thermal stress response without significantly affecting mortality. The ¹H NMR analysis revealed that a total of 26 and 19 molecules, mostly related to energy and protein metabolism as well as antioxidant response, showed significantly different concentrations respectively in the breast muscle and plasma in response to the thermal challenge. In conclusion, the results obtained in this study indicated that chronic heat stress significantly modulates the breast muscle and plasma metabolome in fast-growing broiler chickens, allowing to delineate potential metabolic changes that can have important implications in terms of body energy homeostasis, growth performance and product quality.     

Glutamine metabolism in uricotelic species: variation in skeletal muscle glutamine synthetase, glutaminase, glutamine levels and rates of protein synthesis

High intracellular glutamine levels have been implicated in promoting net protein synthesis and accretion in mammalian skeletal muscle. Little is known regarding glutamine metabolism in uricotelic species but chicken breast muscle exhibits high rates of protein accretion and would be predicted to maintain high glutamine levels. However, chicken breast muscle expresses high glutaminase activity and here we report that chicken breast muscle also expresses low glutamine synthetase activity (0.07±0.01 U/g) when compared to leg muscle (0.50±0.04 U/g). Free glutamine levels were 1.38±0.09 and 9.69±0.12 nmol/mg wet weight in breast and leg muscles of fed chickens, respectively. Glutamine levels were also lower in dove breast muscle (4.82±0.35 nmol/mg wet weight) when compared to leg muscle (16.2±1.0 nmol/mg wet weight) and much lower (1.80±0.46 nmol/mg wet weight) in lizard leg muscle. In fed chickens, rates of fractional protein synthesis were higher in leg than in breast muscle, and starvation (48 h) resulted in a decrease in both glutamine content and rate of protein synthesis in leg muscle. Thus, although tissue-specific glutamine metabolism in uricotelic species differs markedly from that in ureotelic animals, differences in rates of skeletal muscle protein synthesis are associated with corresponding differences in intramuscular glutamine content.     

Preserving avian blood and DNA sampled in the wild: A survey of personal experiences

Collecting and storing biological material from wild animals in a way that does not deteriorate DNA quality for subsequent analyses is instrumental for research in ecology and evolution. Our aims were to gather reports on the effectiveness of methods commonly used by researchers for the field collection and long‐term storage of blood samples and DNA extracts from wild birds. Personal experiences were collected with an online survey targeted specifically at researchers sampling wild birds. Many researchers experienced problems with blood sample storage but not with DNA extract storage. Storage issues generated problems with obtaining adequate DNA quality and sufficient DNA quantity for the targeted molecular analyses but were not related to season of blood sampling, access to equipment, transporting samples, temperature, and method of blood storage. Final DNA quality and quantity were also not affected by storage time before DNA extraction or the methods used to extract DNA. We discuss practical aspects of field collection and storage and provide some general recommendations, with a list of pros and cons of different preservation methods of avian blood samples and DNA extracts.     

Histological alterations of intestinal villi in chickens fed dried Bacillus subtilis var. natto

Two experiments were conducted. In experiment 1, chickens were fed dried Bacillus subtilis var. natto for 3 or 28 days. Growth performance and internal organs were not different from controls, but feed efficiency tended to be improved in the 28-day feeding. In these birds, blood ammonia concentration was decreased (P<0.05). Blood glucose concentration, and amylase and lipase activity in the intestinal content were not significantly different among dietary groups. These results suggest that the B. subtilis natto depressed ammonia concentration. In experiment 2, chickens were fed dietary B. subtilis natto for 28 days. These birds had a tendency to display greater growth performance and intestinal histologies, such as villus height, cell area and cell mitosis, than the controls. Flat cell outline on the duodenal villus surface in controls developed large, protruded cell clusters and cell protuberances after feeding of dietary B. subtilis natto. These results indicate that intestinal function was activated by the depressed blood ammonia concentration in the body of the chicken. The present results may suggest that the B. subtilis natto has the potential to be a beneficial microorganism in chickens.     

Blood lead levels in wintering and moulting Icelandic whooper swans over two decades

Lead poisoning of waterfowl through the ingestion of spent gunshot and discarded anglers' weights continues to be a problem worldwide. We took blood samples from 363 whooper swans Cygnus cygnus at wintering sites in Britain and Ireland and at moulting sites in Iceland during 2001–2005, and analysed them for total blood lead. Lead levels were generally low in swans in Iceland; 6% of samples exceeded 1.21  μ mol L⁻¹, the level indicative of elevated lead and above background levels. The proportion of swans with elevated lead was much higher in the wintering range, varying between 43 and 70% at three sites monitored over the winters 2003/2004–2005/2006, and with blood lead levels ranging up to 19.6  μ mol L⁻¹. The highest levels were in samples taken from swans in Scotland, with a mean value of 3.0  μ mol L⁻¹, but nevertheless they indicated a marked decrease compared with blood lead levels measured for whooper swans at the same site 20 years ago. There also appeared to be a significant long-term decrease in the proportion of swans with elevated blood lead in Iceland, from 60% of birds in 1984 to 8% in 2005, although this may be due to the birds being caught in different parts of Iceland. The most likely source of elevated lead in whooper swans is spent gunshot. Management measures to reduce the impact of lead in the environment on the birds are discussed.     

Effect of serum from chickens treated with clenbuterol on myosin accumulation, β-adrenergic receptor population, and cyclic AMP synthesis in embryonic chicken skeletal muscle cell cultures

Broiler chickens at 35 d of age were fed 1 ppm clenbuterol for 14 d. This level of dietary clenbuterol led to 5-7% increases in the weights of leg and breast muscle tissue. At the end of the 14-d period, serum was prepared from both control and clenbuterol-treated chickens, and was then employed as a component of cell culture media at a final concentration of 20% (v/v). Muscle cell cultures were prepared from both the leg and the breast muscle groups of 12-d chick embryos. Treatment groups included control chicken serum to which 10 nM, 50 nM, and 1 microM clenbuterol had been added, as well as cells grown in media containing 10% horse serum. Cultures were subjected to each treatment for 3 d, beginning on the seventh d in culture. Neither the percent fusion nor the number of nuclei in myotubes was significantly affected by any of the treatments. The quantity of myosin heavy chains (MHCs) was not increased by serum from clenbuterol-treated chickens in either breast or leg muscle cultures; however, the MHC quantity was 50-150% higher in cultures grown in control chicken serum to which 10 and 50 nM clenbuterol had also been added. The beta-adrenergic receptor (betaAR) population was 4000-7000 betaARs per cell in cultures grown in chicken serum, with leg muscle cultures having approximately 25-30% more receptors than breast muscle cultures. Receptor population was not significantly affected by the presence of clenbuterol or by the presence of serum from clenbuterol-treated chickens. In contrast, the betaAR population in leg and breast muscle cultures grown in the presence of 10% horse serum was 16,000-18,000 betaARs per cell. Basal concentration of cyclic adenosine 3':5'monophosphate (cAMP) was not significantly affected by the treatments. When cultures grown in chicken serum were stimulated for 10 min with 1 microM isoproterenol, limited increases of 12-20% in cAMP concentration above the basal levels were observed. However, when cultures grown in the presence of horse serum were stimulated with 1 microM isoproterenol, cAMP concentration was stimulated 5- to 9-fold above the basal levels. Thus, not only did cells grown in horse serum have a higher betaAR population, but also each receptor had a higher capacity for cAMP synthesis following isoproterenol stimulation. Finally, the hypothesis that clenbuterol exerts its action on muscle protein content by changes in cAMP concentration was tested. No correlation was apparent between basal cAMP concentration and MHC content.