Analysis of the genomes of two distantly related bird species, chicken and zebra finch (divergence of about 100 million years), indicate that there are ten avian toll-like receptors and that five of these, TLR2a, 2b, 3, 4, 5 and 7, are clear orthologs to TLRs found in mammals. Duplication of genes has led to TLR1La and 1Lb, TLR2a and 2b, and two TLR7s in the zebra finch. Avian TLR21 may be orthologous to TLR21 found in fish and amphibians, and avian TLR15, which is phylogenetically related to the TLR2 family, appears to be unique to avian species. While TLR2 is conserved between mammalian and avian species, the other TLR2 family members evolved independently. Dimerization between either of the two avian TLR2 species and TLR1La or 1Lb permits recognition of the same broad range of molecules as recognized by mammalian TLR2 dimerized with either TLR1, 6 and 10. Similarly, while TLR9 has been lost from the avian genome, DNA high in unmethylated CpG motifs is immunostimulatory through avian TLR21 which is absent in mammals. Thus, while some TLR members were commonly retained in both mammals and birds, others were separately lost or gained, or diverged independently; but broadly speaking the TLRs of the two classes of vertebrates evolved to recognize very similar spectra of microbial products. Components of downstream TLR signaling are also mostly conserved but with some losses in avian species; notably, TRAM is absent in avian genomes and, hence, the TRIF/TRAM-dependent signaling pathway utilized by mammals in LPS activation appears to be absent in birds.

Avian toll-like receptors.

Vertebrates vary in resistance and resilience to infectious diseases, and the mechanisms that regulate the trade-off between these often opposing protective processes are not well understood. Variability in the sensitivity of species to the induction of damaging inflammation in response to equivalent pathogen loads (resilience) complicates the use of animal models that reflect human disease. We found that induction of proinflammatory cytokines from macrophages in response to inflammatory stimuli in vitro is regulated by proteins in the sera of species in inverse proportion to their in vivo resilience to lethal doses of bacterial lipopolysaccharide over a range of 10,000-fold. This finding suggests that proteins in serum rather than intrinsic cellular differences may play a role in regulating variations in resilience to microbe-associated molecular patterns between species. The involvement of circulating proteins as key molecules raises hope that the process might be manipulated to create better animal models and potentially new drug targets.

Resilience to Bacterial Infection: Difference between Species Could Be Due to Proteins in Serum

During evolution, mammals have evolved a powerful innate immune response to LPS. Chickens are much more resistant to LPS-induced septic shock. Herein we report that chickens sense LPS via orthologs of mammalian TLR4 and myeloid differentiation protein-2 (MD-2) rather than the previously implicated chicken TLR2 isoform type 2 (chTLR2t2) receptor. Cloning and expression of recombinant chTLR4 and chMD-2 in HeLa 57A cells activated NF-kappaB at concentrations of LPS as low as 100 pg/ml. Differential pairing of chicken and mammalian TLR4 and MD-2 indicated that the protein interaction was species-specific in contrast to the formation of functional human and murine chimeric complexes. The chicken LPS receptor responded to a wide variety of LPS derivatives and to the synthetic lipid A compounds 406 and 506. The LPS specificity resembled the functionality of the murine rather than the human TLR4/MD-2 complex. Polymorphism in chTLR4 (Tyr(383)His and Gln(611)Arg) did not influence the LPS response. Interestingly, LPS consistently failed to activate the MyD88-independent induction of IFN-beta in chicken cells, in contrast to the TLR3 agonist poly(I:C) that yielded a potent IFN-beta response. These results suggest that chicken lack a functional LPS-specific TRAM-TRIF (TRIF-related adapter molecule/TIR-domain-containing adapter-inducing IFN-beta) signaling pathway, which may explain their aberrant response to LPS compared with the mammalian species.

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Unique properties of the chicken TLR4/MD-2 complex: selective lipopolysaccharide activation of the MyD88-dependent pathway.

Toll-like receptors (TLRs) are a major class of innate immune pattern recognition receptors that have a key role in immune homeostasis and the defense against infections. The research explosion that followed the discovery of TLRs more than a decade ago has boosted fundamental knowledge on the function of the immune system and the resistance against disease, providing a rational for clinical modulation of the immune response. In addition, the conserved nature of the ancient TLR system throughout the animal kingdom has enabled a comparative biology approach to understand the evolution, structural architecture, and function of TLRs. In the present review we focus on TLR biology in the avian species, and, especially, on the unique functional properties of the chicken TLR repertoire.

Unique features of chicken Toll-like receptors

Effect of Dietary Fat Source on Antibody Production and Lymphocyte Proliferation in Chickens

Large doses of purified lipopolysaccharide (LPS) purified from Escherichia coli induced clinical signs but no mortality in chicks. Five chicks survived a mean dose of 517 mg/kg. One individual that received LPS at 577 mg/kg recovered from clinical manifestations within two days. Attempts failed to produce a generalized Shwartzman-like reaction with two intravenous inoculations of LPS at about 24-hour intervals. Prior injection of uric acid did not protect chicks from LPS by intravenous exposure.

H E Adler

Papers

Toxicity of Endotoxin to Chicks