Australian Poultry Cooperative Research Center

Armidale, Australia

Australian Poultry Cooperative Research Center

Armidale, Australia
Time filter
Source Type

Bannam T.L.,Monash University | Bannam T.L.,Australian Poultry Cooperative Research Center | Yan X.-X.,Monash University | Harrison P.F.,Monash University | And 16 more authors.
mBio | Year: 2011

The pathogenesis of avian necrotic enteritis involves NetB, a pore-forming toxin produced by virulent avian isolates of Clostridium perfringens type A. To determine the location and mobility of the netB structural gene, we examined a derivative of the tetracycline-resistant necrotic enteritis strain EHE-NE18, in which netB was insertionally inactivated by the chloramphen-icol and thiamphenicol resistance gene catP. Both tetracycline and thiamphenicol resistance could be transferred either together or separately to a recipient strain in plate matings. The separate transconjugants could act as donors in subsequent matings, which demonstrated that the tetracycline resistance determinant and the netB gene were present on different conjugative elements. Large plasmids were isolated from the transconjugants and analyzed by high-throughput sequencing. Analysis of the resultant data indicated that there were actually three large conjugative plasmids present in the original strain, each with its own toxin or antibiotic resistance locus. Each plasmid contained a highly conserved 40-kb region that included plasmid replication and transfer regions that were closely related to the 47-kb conjugative tetracycline resistance plasmid pCW3 from C. perfringens. The plasmids were as follows: (i) a conjugative 49-kb tetracycline resistance plasmid that was very similar to pCW3, (ii) a conju-gative 82-kb plasmid that contained the netB gene and other potential virulence genes, and (iii) a 70-kb plasmid that carried the cpb2 gene, which encodes a different pore-forming toxin, beta2 toxin. IMPORTANCE: The anaerobic bacterium Clostridium perfringens can cause an avian gastrointestinal disease known as necrotic enteritis. Disease pathogenesis is not well understood, although the plasmid-encoded pore-forming toxin NetB, is an important virulence factor. In this work, we have shown that the plasmid that carries the netB gene is conjugative and has a 40-kb region that is very similar to replication and transfer regions found within each of the sequenced conjugative plasmids from C. perfringens. We also showed that this strain contained two additional large plasmids that were also conjugative and carried a similar 40-kb region. One of these plasmids encoded beta2 toxin, and the other encoded tetracycline resistance. To our knowledge, this is the first report of a bacterial strain that carries three closely related but different independently conjugative plasmids. These results have significant implications for our understanding of the transmission of virulence and antibiotic resistance genes in pathogenic bacteria. © 2011 Bannam et al.

Bao Y.M.,University of New England of Australia | Choct M.,University of New England of Australia | Choct M.,Australian Poultry Cooperative Research Center | Iji P.A.,University of New England of Australia | Bruerton K.,University of New England of Australia
Asian-Australasian Journal of Animal Sciences | Year: 2010

An experiment was conducted to evaluate the effects of low concentrations of organic and inorganic dietary trace minerals on broiler performance and trace mineral digestibility along the small intestine of 35-day-old broiler chickens reared under floor-pen conditions. Eight hundred male, day-old Cobb broiler chickens were randomly allocated to 4 dietary treatments (25 birds per pen with 8 replicates per treatment). Broilers fed diets supplemented with 4, 20, 40 and 30 mg/kg, respectively, of Cu, Fe, Mn and Zn from organic chelates and inorganic salts achieved the same body weight gain as those supplemented at the NRC levels (8 mg Cu, 40 mg Fe, 60 mg Mn and 40 mg Zn/kg, respectively) from inorganic salts. However, birds fed a control diet without any supplementation at dietary levels of 7.4-8.8, 60.1-69.2, 14.6-15.4 and 19.1-20.6 mg/kg of Cu, Fe, Mn and Zn, respectively, had decreased feed intake and growth rate. There was no significant difference in the digestibility of Cu in all regions of the small intestine. Throughout the small intestine the apparent absorption of Mn from both organic and inorganic sources was small, whereas the digestibility of Zn seemed to be more complex, exhibiting differences in the apparent absorption due to both mineral source and intestinal site. Therefore, the digestibility of organic Zn was improved (p<0.01) in the ileum compared to inorganic Zn. The digestibility of Zn in the duodenum was smaller (p<0.05) than that in the ileum.

Bao Y.M.,University of New England of Australia | Bao Y.M.,Pacific Biosciences | Choct M.,Australian Poultry Cooperative Research Center | Iji P.A.,University of New England of Australia | Bruerton K.,Protea Park Nutrition
British Poultry Science | Year: 2010

1. The aim of the present study was to demonstrate trace mineral interactions among organic copper, iron, manganese and zinc (Cu, Fe, Mn and Zn) in broiler chickens. 2. Three experiments were conducted using a control diet which was deficient in Cu, Fe, Mn and Zn. 3. In experiment 1, the control diet, supplemental organic Cu, Fe alone and combined diets, were randomly fed to 4 groups of one-day-old Cobb broilers (each group had 6 replicates of 4 birds). 4. In experiment 2, the control diet, supplemental organic Mn and Zn alone or combined with Cu, Fe diets and corresponding inorganic combined diet, were randomly fed to 6 groups (each group had 8 replicates of 6 birds). 5. In experiment 3, the depletion of organic Zn, the depletion of inorganic Zn and normal Zn treatments were carried out in three groups of one-day-old Cobb broilers (each group had 8 replicates of 6 birds). 6. Adding organic Cu, Fe and Mn alone or combined to Zn deficient diets did not significantly improve bird performance and were mostly excreted. Supplemental organic Zn alone or combined with other elements significantly increased feed intake, body weight gain and tibia bone length. However, supplemental organic Fe alone or combined with Cu significantly increased feed intake but had no obvious effect on body weight gain. The organic Fe supplementation resulted in a wider tibia. 7. Depletion of organic and inorganic Zn resulted in decreased feed intake, body weight gain and total tibia bone Zn content. Zinc deficiency did not affect the uptake of organic Fe by tibia bone but reduced its total Fe content. 8. Zinc is the first limiting element among these 4 trace minerals. Adding Mn, Cu and Fe to Zn deficient diets did not stimulate bird performance. Surplus organic Fe and Cu resulted in increased feed intake and increased tibia bone Fe content but did not contribute to bird performance. © 2010 British Poultry Science Ltd.

Cheung J.K.,Monash University | Keyburn A.L.,Monash University | Keyburn A.L.,CSIRO | Keyburn A.L.,Australian Poultry Cooperative Research Center | And 8 more authors.
Infection and Immunity | Year: 2010

Clostridium perfringens causes several diseases in domestic livestock, including necrotic enteritis in chickens, which is of concern to the poultry industry due to its health implications and associated economic cost. The novel pore-forming toxin NetB is a critical virulence factor in the pathogenesis of this disease. In this study, we have examined the regulation of NetB toxin production. In C. perfringens, the quorum sensing-dependent VirSR two-component signal transduction system regulates genes encoding several toxins and extracellular enzymes. Analysis of the sequence upstream of the netB gene revealed the presence of potential DNA binding sites, or VirR boxes, that are recognized by the VirR response regulator. In vitro binding experiments showed that purified VirR was able to recognize and bind to these netB-associated VirR boxes. Furthermore, using a reporter gene assay, the netB VirR boxes were shown to be functional. Mutation of the virR gene in two avian C. perfringens strains was shown to significantly reduce the production of the NetB toxin; culture supernatants derived from these strains were no longer cytotoxic to Leghorn male hepatoma cells. Complementation with the virRS operon restored the toxin phenotypes to wild type. The results also showed that the VirSR two-component system regulates the expression of netB at the level of transcription. We postulate that in the gastrointestinal tract of infected birds, NetB production is upregulated when the population of C. perfringens cells reaches a threshold level that leads to activation of the VirSR system. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Svihus B.,Norwegian University of Life Sciences | Sacranie A.,Australian Poultry Cooperative Research Center | Denstadli V.,Norwegian University of Life Sciences | Choct M.,Australian Poultry Cooperative Research Center
Poultry Science | Year: 2010

Two experiments were carried out to study the effect of intermittent feeding, diet structure, and their interaction on the performance, feeding pattern, and digestive adaptation of broiler chickens. In experiment 1, 40 Cobb broiler chickens were fed, either ad libitum or intermittently, a commercial starter diet diluted with 150 g/kg of ground or whole wheat in a 2 × 2 factorial arrangement. Ad libitum feeding consisted of continuous access to feed in a room with 18 h of light and 6 h of complete darkness, whereas birds on intermittent feeding had restricted access to feed from 7 d of age, with 4 one-hour feeding bouts/d and one 2-h feeding bout/d from d 14. No interaction was found between feed structure and feeding regimen for any of the measurements. Although BW gain was not significantly different among any of the treatments, birds given whole wheat or fed intermittently had significantly better feed efficiency than those given ground wheat and fed ad libitum. Inclusion of whole wheat instead of ground wheat also markedly increased (P < 0.001) the AMEn value and fecal starch digestibility. In experiment 2, 60 Ross broiler chickens were trained to meal feeding on a wheat-based diet containing a commercial phytase, and 5 birds were killed every 15 min after having access to feed, to collect crop contents. Phytate was gradually degraded in the crop, with a 50% reduction in inositol 6-phosphate after a 100-min retention time. In conclusion, these results indicate that broiler chickens have a remarkable ability to maintain growth rate during intermittent feeding because the crop is used as an intermediate storage organ, and that improvements in feed efficiency may occur, among others through increased efficacy of exogenous enzymes. Even though stimulation of the gizzard through a coarse feed structure improves feed efficiency, it does not appear to affect the ability of the bird to handle intermittent feeding. © 2010 Poultry Science Association Inc.

Cummins D.M.,CSIRO | Cummins D.M.,Australian Poultry Cooperative Research Center | Tyack S.G.,CSIRO | Doran T.J.,CSIRO
Biochemical and Biophysical Research Communications | Year: 2011

The U6 and 7SK RNA polymerase III promoters are widely used in RNAi research for the expression of shRNAs. However, with their increasing use in vitro and in vivo, issues associated with cytotoxicity have become apparent with their use. Therefore, alternative promoters such as the weaker H1 promoter are becoming a popular choice. With interest in the chicken as a model organism, we aimed to identify and characterise the chicken H1 promoter for the expression of shRNAs for the purpose of RNAi.The chicken H1 promoter was isolated and sequence analysis identified conserved RNA polymerase III promoter elements. A shRNA expression cassette containing the chicken H1 promoter and shRNA targeting enhanced green fluorescent protein (EGFP) was developed. An RNAse protection assay confirmed activity of the promoter determined by the detection of expressed shRNAs. Comparison of the H1 promoter to the chicken RNA polymerase III 7SK and U6 promoters demonstrated that expressed shRNAs from the H1 promoter induced gene specific silencing, albeit to lower levels in comparison to both 7SK and U6 promoters.Here we have identified a new tool for RNAi research with specific applications to the chicken. The availability of a RNA polymerase III promoter that drives shRNA expression to reduced levels will greatly benefit in ovo/. in vivo applications where there are concerns of cytotoxicity resulting from overexpression of an shRNA. © 2011 Elsevier Inc.

Loading Australian Poultry Cooperative Research Center collaborators
Loading Australian Poultry Cooperative Research Center collaborators