Influence of age,sex and rearing systems on Toll-like receptor 7 (TLR7) expression pattern in gut,lung and lymphoid tissues of indigenous ducks

1. The objective of the experiment was to determine the influence of age, sex and rearing system on Toll-like receptor 7 (TLR7) gene expression in gut, lung and lymphoid tissues and physiological responses to stress in male and female indigenous ducks of Tamil Nadu, India.

2. A total of 36 ducks (12 males and 24 females) were obtained from local farmers and tissue samples of gut tissues (duodenum, jejunum, ileum and caecum), lymphoid organs (spleen and bursa) and lungs were collected in RNAlater solution followed by RNA extraction.

3. After normalisation to β-actin (endogenous control) qPCR analysis identified a significant effect of age, sex and rearing system on TLR7 expression in the ducks.

4. A significant up-regulation of TLR7 expression was observed in lungs, duodenum, jejunum, ileum and caecum of sexually mature (45 wk) compared with that of immature ducks (16 wk). Among sexes, male ducks had significantly higher TLR7 expression than female ducks.

5. Age and sex interactions were significant in lungs, duodenum, jejunum and caecum. Ducks reared in an extensive housing system showed significantly higher TLR7 expression in bursa, lungs, duodenum, ileum and caecum compared to intensively reared ducks. There were no effects of age, sex and rearing systems on TLR7 expression in the spleen.

6. The heterophil-to-lymphocyte ratio and serum corticosterone were higher in ducks reared on an intensive system compared with ducks from an extensive rearing system.     

Survivability,immunity, growth and production traits in indigenous and White Leghorn breeds of chicken

ABSTRACT

1. The survivability, innate and adaptive immunity, growth and production traits up to 72 weeks of age were determined in Ghagus, Nicobari (unimproved indigenous) and White Leghorn (WLH) breeds and the study investigated links between innate and adaptive immunity and survivability and production traits.

2. At 20 and 40 weeks of age, there was a significant effect of breed on innate immunity assessed by measuring titres of natural antibody (NAb) binding to rabbit red blood cells (RRBC) and adaptive immunity assessed by measuring specific antibody titre (SpAb) to Newcastle disease virus.

3. Highest survivability was in WLH (91.6%) followed by Nicobari (87.1%) and Ghagus (82.9%) breeds. Growth traits at different ages were higher (P< 0.001) in Ghagus followed by WLH and Nicobari breeds. Egg production up to 72 weeks was higher (P < 0.001) in WLH followed by Nicobari and Ghagus breeds, whereas egg weight at different ages was higher (P < 0.001) in WLH than Ghagus and Nicobari breeds.

4. NAb titres measured at 20 weeks were significantly (P = 0.002) associated with the survivability of hens during 20 to 72 weeks of age. Breed-wise analysis showed a significant (P = 0.019) association between NAb titres at 20 weeks and survivability in the Ghagus breed. Furthermore, NAb titres at 20 weeks were higher in hens which survived to 72 weeks compared with those that died (P = 0.002).

5. Measuring NAb titres to RRBC is quick, economical and simple. This method has potential to be used in a breeding programme to increase survivability of laying hens.     

Toll-like receptor mRNA expression, <Emphasis Type="Italic">iNOS</Emphasis> gene polymorphism and serum nitric oxide levels in indigenous chickens

Toll-like receptor (TLR) family is one of the important members of innate immune system that recognizes conserved microbial patterns and induces innate immune response. They also act as a link to adaptive immune response. Nitric oxide (NO) is a multi-functional mediator with diverse physiological and immunological roles. In the present study TLR mRNA expression in heterophils, serum nitric oxide level and iNOS (inducible Nitric Oxide Synthase) gene polymorphism were investigated in cockerels of two Indian native chicken breeds, Aseel and Kadaknath. TLR (4 and 5) mRNA expression as quantified by real time RT-PCR revealed Kadaknath males expressed significantly (P < 0.01) higher TLR4 mRNA than Aseel males. iNOS gene polymorphism analyzed by PCR-RFLP method revealed difference in allele frequency. Kadaknath males had higher allele B frequency (0.81) than Aseel males (0.56). However, there were no genotype and breed effect on serum nitric oxide level. Based on the present study we conclude that Kadaknath has comparatively higher innate immunity levels than Aseel, however further investigations are needed.     

Co-administration of toll-like receptor (TLR)-3 and 4 ligands augments immune response to Newcastle disease virus (NDV) vaccine in chicken

Toll-like receptors (TLRs) are pattern recognition receptors (PRRs) that mediate first line of host defence to pathogens. TLR agonists are potent immunostimulatory agents that help to prime a robust adaptive immune response. In the present study, adjuvant potential of Poly I:C and lipopolysaccharide (LPS) were evaluated with live Newcastle disease virus (NDV) vaccine. Cornish chickens were immunized with live Newcastle disease virus (NDV) vaccine (R2B-mesogenic strain) adjuvanted either with Poly I:C (TLR3 agonist) or LPS-TLR4 agonist and both. Humoral Immune response to ND vaccine was evaluated through haemagglutination inhibition (HI) test and ELISA, while the cellular immune response (CMI) was quantified by lymphocyte transformation test (LTT). IL-1β cytokine mRNA levels in spleen tissue were also quantified by real time PCR. The results suggest that TLR3 and TLR4 agonists are an efficient immune-stimulators separately, as LPS co-administered group has shown significantly higher serum titre on second week post-immunization and Poly I:C group on third week post-immunization both by HI and ELISA (P?<?0.01), however, the combined administration of both LPS and Poly I:C did not give any complementary effect on serum titre. There were no significant differences in stimulation indices (SI) and IL-1β cytokine levels between groups at different intervals post-immunization. Hence, TLR agonists LPS followed by Poly I:C could be used as adjuvant to enhance the immune response to NDV vaccine in chicken.

     

Differential gene expression of antimicrobial peptides β defensins in the gastrointestinal tract of Salmonella serovar Pullorum infected broiler chickens

Salmonella enterica serovar Pullorum causes substantial mortality in chicks as well as results in persistent infection and vertical transmission in layer birds. An effective innate immune response in the early stages of infection could reduce bacterial colonization and mortality in chicks and persistency of infection in later stages. β Defensins (AvBDs) are now considered as one of the key components of innate immunity in avian species. In the present study, we quantified the mRNA expression levels of AvBDs (1–14) by real-time PCR in the gastrointestinal (GI) tissues (duodenum, jejunum, ileum and caecum) of 3-day-old broiler chicks after 24 h of oral infection with Salmonella Pullorum. Quantitative real-time PCR analysis revealed significant (P < 0.05) upregulation of AvBD3, 4, 5, 6 and 12 and a significant (P < 0.05) down regulation in the expressions of AvBD10, 11, 13 and 14 in one or few GI tissues, while no significant changes were observed for AvBD1, 2, 7, 8 and 9 gene expressions in any of the GI tissues investigated upon infection with S. Pullorum. Most substantial change in gene expression was found for AvBD5, being significantly (P < 0.01) upregulated in most of the GI tissues investigated. The differential expression levels of β defensins shed light on tailored innate immune response induced by S. Pullorum during the early stages of infection in chicks.     

Differential expression of Toll-like receptor mRNA in White Leghorn and indigenous chicken of India

In the present experiment, the expression profile of Toll-like receptor mRNA in indigenous and pure line chickens was studied. The expression of TLR3, TLR4, TLR5 and TLR7 were quantified in heterophils of Aseel, Kadaknath, Naked neck, Dwarf and White Leghorn lines by Quantitative Real-time PCR. White Leghorns expressed significantly (P < 0.01) higher levels of TLR3 mRNA compared to other lines. TLR4 and TLR5 mRNA were significantly highly expressed in Kadaknath line. Among the TLRs investigated TLR5 was more expressed in all lines studied. TLR7 was highly expressed in indigenous chicken Aseel and Kadaknath than other lines. Dwarf chicken expressed significantly (P < 0.01) lower levels of all TLRs investigated. On the basis of the present study we conclude that the differential expression of TLR mRNA in the heterophils of indigenous and other chicken breeds might contribute to their variable disease resistance/susceptibility.