Effects of heat stress on antioxidant defense system,inflammatory injury,and heat shock proteins of Muscovy and Pekin ducks: evidence for differential thermal sensitivities

Rising temperatures are severely affecting the mortality, laying performance, and meat quality of duck. Our aim was to investigate the effect of acute heat stress on the expression of heat shock proteins (HSPs: HSP90, 70, 60, 40, and 10) and inflammatory factors (nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2)) and antioxidant enzyme activity (superoxide dismutase (SOD), malondialdehybe (MDA), catalase (CAT), total antioxidant capacity (T-AOC)) in livers of ducks and to compare the thermal tolerance of Pekin and Muscovy ducks exposed to acute heat stress. Ducks were exposed to heat at 39 ± 0.5 °C for 1 h and then returned to 20 °C for 1 h followed by a 3-h recovery period. The liver and other tissues were collected from each individual for analysis. The mRNA levels of HSPs (70, 60, and 40) increased in both species, except for HSP10, which was upregulated in Muscovy ducks and had no difference in Pekin ducks after heat stress. Simultaneously, the mRNA level of HSP90 decreased in the stress group in both species. Morphological analysis indicated that heat stress induced tissue injury in both species, and the liver of Pekin ducks was severely damaged. The activities of several antioxidant enzymes increased in Muscovy duck liver, but decreased in Pekin duck. The mRNA levels of inflammatory factors were increased after heat stress in both duck species. These results suggested that heat stress could influence HSPs, inflammatory factors expression, and the activities of antioxidant enzymes. Moreover, the differential response to heat stress indicated that the Muscovy duck has a better thermal tolerance than does the Pekin duck.     

Complete Genomic Sequence of a Reovirus Isolate from Pekin Ducklings in China

The complete genomic sequence of a Pekin duck origin reovirus (DRV) from China was determined. The genome comprises 23,419 bp, with segments ranging from 1,191 bp (S4) to 3,959 bp (L1). Pairwise comparisons and phylogenetic analysis indicate that the Pekin duck origin reovirus is more closely related to the new type of Muscovy duck origin reovirus (N-MDRV) identified recently than to the chicken origin avian orthoreovirus (ARV) and the originally described Muscovy duck origin reovirus (ARV-Md).     

Hormonal and metabolic responses to overfeeding in three genotypes of ducks

Muscovy, Pekin and Mule duck are different in their body weight. To make a valid comparison in the lipid metabolism between these three genotypes, overfeeding was carried out by providing the animals with amounts of food in proportion to their body weight. Under these conditions, Muscovy ducks developed a strong liver steatosis, whereas it was not very pronounced in the Mule ducks and even less in the Pekin ducks. On the contrary, Pekin ducks showed a much marked extrahepatic fattening. At the beginning of overfeeding, there was a similarity in the three genotypes as regards the post-heparin lipoprotein-lipase (LPL) activity and the insulin and glucagon concentrations. After 10 days of overfeeding, the LPL activity dramatically fell in Muscovy and in Mule ducks, whereas it remained steady in Pekin ducks. Compared to values found at the beginning of the overfeeding period, plasma glucagon and insulin shown no evolution, except for the insulin of Pekin ducks which was dramatically higher. Those data suggest that high plasma insulin concentrations measured in Pekin ducks after 10 days of overfeeding can be responsible for the maintenance of the LPL activity, which favors the extrahepatic fattening to the detriment of liver steatosis.     

Is the hepatic metabolism of glucose and linoleic acid influenced by species in overfed ducks?

There are genetic differences in the hepatic glucose and linoleic acid metabolisms between Muscovy and Pekin ducks ad libitum-fed. To understand the effect of overfeeding on the hepatic metabolisms in these two species of ducks, we compared the different pathways of glucose and linoleic acid reaching the liver of Muscovy (Cairina moschata) (n = 6) and Pekin (Anas platyrhynchos) (n = 6) ducks overfed for 1 week and sacrificed 2–4 h after their last meal by using the ex vivo method of liver slices incubated for 16 h with [U-¹⁴C]-glucose, [1-¹⁴C]-linoleic acid and [³⁵S]-methionine added to the survival medium. The glucose was the main precursor of triacylglycerol synthesis in the liver of these two species and its hepatic metabolism was similar between species. The hepatic uptake of linoleic acid was 1.7-fold higher (P = 0.020) in the Muscovy duck than in the Pekin duck leading to a 1.9-fold higher (P = 0.017) esterification of this fatty acid in the liver of the Muscovy duck than in that of the Pekin duck. Finally, both species after 1 week of overfeeding exhibited the same capacity to secrete VLDL remaining insufficient to avoid hepatic steatosis.     

Genomics of antiviral defenses in the duck, a natural host of influenza and hepatitis B viruses

We review our progress using genomics approaches to examine key antiviral defenses of the White Pekin mallard duck, Anas platyrhynchos. Our interest stems from the fact that ducks are the natural host of avian influenza, and are an important animal model for hepatitis B research. First, we have conducted an expressed sequence tag (EST) project and identified more than 200 immune relevant genes in the duck. Our analysis of these genes allows us to evaluate the homology between ducks and their closest genetic model organism, the chicken. We have also constructed genomic and cDNA libraries from the same individual duck, allowing us to directly compare expressed sequences with those present in the genome. These resources allow us to determine the organization and expression of regions of the genome important in antiviral defenses. Here we examine the organization of the immunoglobulin heavy chain locus, the Major Histocompatibility Complex class I region, the lectin immunoreceptors and Toll-like receptor 7. We discuss our research-in-progress in the context of the immune defense against viruses, particularly influenza.     

Does overfeeding enhance genotype effects on energy metabolism and lipid deposition in breast muscle of ducks?

We evaluated the effects of genotype (Muscovy, Pekin and their crossbreed hinny and mule ducks) and feeding levels (overfeeding between 12 and 14 weeks of age vs ad libitum feeding) on energy metabolism and lipid deposition in breast muscle of ducks. Samples of breast muscle (Pectoralis major) were collected at 14 weeks of age from 8 birds per group. Overfeeding induced an accumulation of lipids in breast muscle (1.5- to 1.7-fold, depending on genotype) mainly induced by triglyceride deposition. It also induced a considerable increase in the amounts (expressed as g/100 g of tissue) of saturated and mono-unsaturated fatty acids (SFA, MUFA), while the amounts of poly-unsaturated fatty acids (PUFA) remained unchanged in hinny and Muscovy ducks or slightly increased in Pekin and mule ducks. In breast muscle, overfeeding decreased the activity of the main enzymes involved in lipogenesis from glucose (glucose-6-phosphate dehydrogenase, G6PDH, malic enzyme, ME, acetyl CoA carboxylase, ACX). Lipoprotein lipase (LPL) activity in Pectoralis major muscle was also significantly decreased (-21%). The ability of muscle tissues to catabolize long-chain fatty acids, as assessed by beta-hydroxyacyl CoA dehydrogenase (HAD) activity, was increased in Pectoralis major muscle, as was cytochrome-c oxidase (COX) activity. Hybrid and Pekin ducks exhibited higher levels of ACX and LPL activity in Pectoralis major muscle than Muscovy ducks, suggesting a greater ability to synthesise lipids in situ, and to take up circulating lipids. Total lipid content in breast muscle of hybrid and Pekin ducks was higher than in that of Muscovy ducks. In hybrid and Pekin ducks, lipid composition of breast muscle was characterized by higher amounts of triglycerides, SFA and MUFA than in Muscovy ducks. Finally, oxidative metabolism was greater in Pectoralis major muscles of hybrid and Pekin ducks than in Muscovy ducks, suggesting an adaptative strategy of muscle energy metabolism according to lipid level.     

Duck hepatitis B virus infection of Muscovy duck hepatocytes and nature of virus resistance in vivo.

To test the hypothesis that in vivo resistance to hepadnavirus infection was due to resistance of host hepatocytes, we isolated hepatocytes from Muscovy ducklings and chickens, birds that have been shown to be resistant to duck hepatitis B virus (DHBV) infection, and attempted to infect them in vitro with virus from congenitally infected Pekin ducks. Chicken hepatocytes were resistant to infection, but we were able to infect approximately 1% of Muscovy duck hepatocytes in culture. Infection requires prolonged incubation with virus at 37 degrees C. Virus spread occurs in the Muscovy cultures, resulting in 5 to 10% DHBV-infected hepatocytes by 3 weeks after infection. The relatively low rate of accumulation of DHBV DNA in infected Muscovy hepatocyte cultures is most likely due to inefficient spread of virus infection; in the absence of virus spread, the rates of DHBV replication in Pekin and Muscovy hepatocyte cultures are similar. 5-Azacytidine treatment can induce susceptibility to DHBV infection in resistant primary Pekin hepatocytes but appears to have no similar effect in Muscovy cultures. The relatively inefficient infection of Muscovy duck hepatocytes that we have described may account for the absence of a detectable viremia in Muscovy ducklings experimentally infected with DHBV.     

Behavioural and physiological fear responses in ducks: genetic cross effects

Mule duck, a cross between a Muscovy drake and a Pekin female, is reported by the farmers to frequently express fear behaviours, such as man avoidance. The genetic basis of fear responses in mule ducks was therefore investigated in this study. According to a previous experiment, the dominant effect of Pekin genotype was hypothesised; however, due to the absence of birds from the reciprocal cross, a superiority of the Pekin in additive effect could not be distinguished from a direct maternal additive effect. In order to clarify this, ducks from the mule genotype, the two parental genotypes (Pekin and Muscovy) and the reciprocal intercross (hinny) underwent a set of physiological and individual behavioural tests of fear. Both parental genotypes were highly fearful but exhibited responses of different patterns: Pekin ducks manifested a higher locomotor activity, whereas the Muscovy ducks showed a higher avoidance to man. Hybrids expressed higher panic responses and specific fear of man than the two parent breeds. Both hybrids expressed similar patterns and the maternal effects were not significant. Significant heterosis effects were found for most of the behavioural responses, in agreement with the fact that higher fear responses were expressed by the hybrids compared to the parental genotypes. A significant heterosis effect was also found for basal adrenal activity; hybrids having higher basal level than parental genotypes. Maximum capacity of adrenal response appeared to be determined by direct additive effects with a superiority of the Pekin genotype.     

Is ability to hepatic steatosis influenced by age at the beginning of the overfeeding period in Muscovy and Pekin ducks?

The aim of this study was to analyse the effects of species (Muscovy and Pekin ducks) and age at the beginning of the overfeeding period on fatty liver production, carcass composition and lipid and moisture content of the liver and breast muscle. We reared four groups of 40 ducks per species for the study, starting at 2-week intervals in order to have four different ages together at the beginning of the overfeeding period (10, 12, 14 and 16 weeks). At the end of the overfeeding period, all ducks were slaughtered. Our results confirmed the high levels of difference in carcass composition and lipid content in the plasma, liver and breast muscle between Muscovy and Pekin ducks at all ages. Pekin ducks were not able to develop a high degree of hepatic steatosis, but had increased lipid storage in peripheral adipose and muscle tissues than Muscovy ducks. However, the fatty liver weight of Pekin ducks increased with age, with lipid deposition in the liver and peripheral tissues. The ability of Muscovy ducks to produce fatty livers remained unchanged with age in line, with lipid deposition in the liver and peripheral tissues. The sites of lipid deposition thus depend on species and not on the physiological maturity of ducks.     

Comparative Proteomic Analysis of the Hepatic Response to Heat Stress in Muscovy and Pekin Ducks: Insight into Thermal Tolerance Related to Energy Metabolism

The Pekin duck, bred from the mallard (Anas platyrhynchos) in china, is one of the most famous meat duck species in the world. However, it is more sensitive to heat stress than Muscovy duck, which is believed to have originated in South America. With temperature raising, mortality, laying performance, and meat quality of the Pekin duck are severely affected. This study aims to uncover the temperature-dependent proteins of two duck species using comparative proteomic approach. Duck was cultured under 39°C ± 0.5°C for 1 h, and then immediately returned to 20°C for a 3 h recovery period, the liver proteins were extracted and electrophoresed in two-dimensional mode. After analysis of gel images, 61 differentially expressed proteins were detected, 54 were clearly identified by MALDI TOF/TOF MS. Of the 54 differentially expressed protein spots identified, 7 were found in both species, whereas 47 were species specific (25 in Muscovy duck and 22 in Pekin duck). As is well known, chaperone proteins, such as heat shock protein (HSP) 70 and HSP10, were abundantly up-regulated in both species in response to heat stress. However, we also found that several proteins, such as α-enolase, and S-adenosylmethionine synthetase, showed different expression patterns in the 2 duck species. The enriched biological processes were grouped into 3 main categories according to gene ontology analysis: cell death and apoptosis (20.93%), amino acid metabolism (13.95%) and oxidation reduction (20.93%). The mRNA levels of several differentially expressed protein were investigated by real-time RT-PCR. To our knowledge, this study is the first to provide insights into the differential expression of proteins following heat stress in ducks and enables better understanding of possible heat stress response mechanisms in animals.     

Identification of histone H1.z components in a Muscovy duck (Cairina moschata L.) population

The general patterns of histone H1 proteins from erythrocyte nuclei of Muscovy duck individuals were similar to those of Pekin type ducks both in acetic acid-urea and 2D polyacrylamide gels. We show here that Muscovy duck histone H1.z in the tested population was represented by three different electromorphs, each presumably encoded by a distinct allelic gene. Accordingly, we have identified six phenotypes consisting of the homodimeric and heterodimeric combinations of the three isoforms. The frequency of the presumptive alleles ranged from 0.506 for the main allele z1 to 0.379 for allele z2 and only 0.113 for the rarest allele z3. In addition to a standard set of somatic H1 variants, an unusual protein X, absent in other avian species, was also revealed.     

Genes involved in the establishment of hepatic steatosis in Muscovy,Pekin and mule ducks

Our main objectives were to determine the genes involved in the establishment of hepatic steatosis in three genotypes of palmipeds. To respond to this question, we have compared Muscovy ducks, Pekin ducks and their crossbreed the mule duck fed ad libitum or overfed. We have shown a hepatic overexpression of fatty acid synthase (FAS) and di-acyl glycerol acyl transferase 2 (DGAT2) in overfed individuals, where DGAT2 seemed to be more regulated. This increase in lipogenesis genes is associated with a decrease of lipoprotein formation in Muscovy and mule ducks, especially apolipoprotein B (ApoB) and Microsomal Triglyceride Transfer Protein (MTTP), leading to lipid accumulation in liver. In Pekin ducks, MTTP expression is upregulated suggesting a better hepatic lipids exportation. Regarding lipids re-uptake, fatty acid-binding protein 4 and very-low-density-lipoprotein receptor are overexpressed in liver of mule ducks at the end of the overfeeding period. This phenomenon puts light on a mechanism unknown until today. In fact, mule can incorporate more lipids in liver than the two other genotypes leading to an intensified hepatic steatosis. To conclude, our results confirmed the genotype variability to overfeeding. Furthermore, similar observations are already described in non-alcoholic fatty liver disease in human, and ask if ducks could be an animal model to study hepatic triglyceride accumulation.     

Complete genome sequence analysis of duck circovirus strains from Cherry Valley duck

To investigate molecular epidemiology of DuCV in Cherry Valley ducks in China,the complete genomes of six DuCV strains,which were detected from Cherry Valley ducks in China between 2007 and 2008,were s...     

PCR detection and sequence analysis of duck circovirus in sick Muscovy ducks

The duck circovirus (DuCV) infection in sick ducks from Fujian Province was investigated. The liver samples of 43 sick Muscovy ducks with infectious serositis were collected from 12 duck farms in Fujian Province.Based on the published sequences of DuCV, two primers were designed for the detection of DuCV and four pairs of primers were designed to amplify four overlapping fragments that cover the complete genome of DuCV. The specific PCR products were amplified from positive samples. The fragments were then cloned into pMD18-T vector and sequenced, and the full length genomic sequence of the FJ0601 isolate of DuCV was obtained. PCR analysis showed that the proportion of ducks which were positive for circovirus was 79% and 10 out of the 12 farms were positive. Sequence analysis showed that the complete genome of DuCV-FJ0601 was 1988 bp and possessed features common to the family Circoviridae which included a stem-loop structure and the Rep protein motifs. Homology analysis showed that FJ0601 isolate of DuCV had 97.3%～97.5% nucleotide sequence identity to all the four Taiwan isolates (TC1/2002, TC2/2002, TC3/2002, TC4/2002), 82.9% identity to the America (33753-52) isolate and 82.3% identity to the Germany isolate. Phylogenetic analysis with Clustal W, however,showed that FJ0601 isolate of DuCV was on a common branch with Taiwan isolates, and Germany and America isolates belonged to the other branch.     

An outbreak of duck virus enteritis (duck plague) in Alberta.

Duck plague (Duck virus enteritis) was disgnosed in a resident population of Muscovy ducks (Cairina moschata) on a small game farm in Alberta. This disease has not been reported previously in Canada. Clinical signs consisted of cyanosis, depression and acute death. Necropsy of two Muscovy ducks revealed lesions typical of the disease. There were ulcerations with pseudomembranes in the small intestine, ulcerations with caseous plaques in the esophagus and eosinophilic intranuclear inclusion bodies in the spleen. Clinical disease with mortality was reproduced in young ducklings injected with tissue homogenates from field cases. All surviving inoculated ducklings seroconverted to highly significant titres of neutralizing antibodies to duck virus enteritis (DVE) virus. All attempts to isolate the agent in embryonating duck eggs or primary tissues cultures of duck and chicken kidney were negative. Identification of the DVE virus was accomplished by serum neutralization with ducklings as the host system.     

Does overfeeding enhance genotype effects on liver ability for lipogenesis and lipid secretion in ducks?

We evaluated the effects of genotype (Muscovy, Pekin and their crossbreed hinny and mule ducks) and feeding levels (overfeeding between 12 and 14 weeks of age vs ad libitum feeding) on liver ability for lipogenesis and lipid secretion in ducks. Samples of liver and blood were collected at 14 weeks of age from 8 birds per group. Plasma levels of insulin was considerably increased in overfed ducks (1.9-fold), stimulating the hepatic activity of the main enzymes involved in lipogenesis from glucose (glucokinase, GK, glucose-6-phosphate dehydrogenase, G6PDH, malic enzyme, ME, acetyl CoA carboxylase, ACX), while cytochrome-c oxidase (COX) activity, indicating overall oxidation ability of energy-yielding substrates, remained unchanged. Plasma levels of triglycerides, phospholipids and total cholesterol were therefore increased (1.9, 3.7, 1.6 and 1.6-fold, respectively). Glycaemia also significantly increased (+8%). Pekin ducks exhibited higher levels of GK and G6PDH activity in the liver than Muscovy ducks, suggesting a greater ability to use glucose consistent with their lower glycaemia. Muscovy ducks had greater ACX activity, suggesting greater ability to synthesise lipids. However, plasma lipid levels were much higher in Pekin ducks than in Muscovy ducks, suggesting a greater ability to export lipids from the liver. Values for the different criteria measured in this study were intermediate or similar in hinny and mule ducks to those of parental species. The high values for GK, G6PDH, ME and ACX activity in hybrid ducks enabled them to produce heavy fatty livers with the same chemical and lipid composition as Muscovy ducks and characterised by high amounts of triglycerides (around 96% of total lipids), and saturated and mono-unsaturated fatty acids.     

Proteomic analysis of liver development of lean Pekin duck (Anas platyrhynchos domestica)

The liver plays vital roles in digestion, metabolism and immune defense. To elucidate the molecular mechanism of nutrient metabolism and antioxidation of lean Pekin duck liver from hatching to slaughter, the proteome changes were investigated using 2-DE, MS, quantitative real-time PCR and bioinformatics. A total of 59 differentially expressed proteins were identified. Proteins involved in transportation were highly up-regulated in newborn ducks whereas 37 proteins associated with metabolism, defense and antioxidation were up-regulated in adult ducks. The over-expression of proteins at the last developmental stage presumably occurs to fulfill the needs of multiple functions of the liver. However, the over-expressed proteins related to transportation during the first developmental stage are involved in maintaining the high basal metabolism of newborn ducks. The functional enrichment analysis also confirmed these results. Furthermore, the protein interaction network predicted 28 proteins acting as key nodes for liver development. The validated expression between proteins and genes provides us target genes for future genetic analyses to improve the health and performance of these ducks. These significant advanced proteome data expand our knowledge on the physiology of the duck liver, thereby providing a potentially valuable foundation for molecular breeding to enhance feed efficiency and immunity and for optimizing the feeding strategy.