Toll-like receptor 4 (TLR4) recognizes pathogen-associated molecular patterns in some animals and has been shown to be closely associated with several diseases such as tumors, atherosclerosis, and asthma. However, its function in ducks is not clear. Alternative splicing of the TLR4 gene has been identified in pigs, sheep, mice, and other species, but has not yet been reported in the duck. In this study, alternative splicing of the duck TLR4 gene was investigated using reverse transcription-polymerase chain reaction (RT-PCR). Duck TLR4 gene (duTLR4, accession number: KF278109) was found to consist of 3367 nucleotides of coding sequence. An alternative splice form, TLR4-b, was identified and shown by alignment to retain the intron between exons 1 and 2. Real-time quantitative polymerase chain reaction (qPCR) analyses suggested that duTLR4-a (wild-type) mRNA is widely expressed in various healthy tissues, whereas TLR4-b is expressed at only low levels. Following stimulation of normal duck embryo fibroblasts with lipopolysaccharide, the expression of both isoforms initially increased and then decreased. Expression of the wild-type isoform subsequently increased again, while that of the variant remained low. The expression levels of wild-type TLR4 were further analyzed by transient transfection of a pcDNA3.1(+)-TLR4-a overexpression vector into duck embryo fibroblasts. qRT-PCR analyses showed that after stimulation with LPS and poly(I:C) the expression levels of IL-1β, IL6, and MHC II increased with a response-efficacy relationship. Our experimental results indicate that TLR4 plays an important role in resistance to both bacterial and viral infections in the duck.

Mating behavior is a critically important component of poultry reproduction. Here, a total of 135 geese were selected, specifically, Sichuan white geese (Anser cygnoides), Zhedong white geese (Anser cygnoides), and Hungarian geese (Anser anser) (300-day-old), and the mating behavior was monitored daily from 6:00 a.m. to 6:00 p.m. during the 20-day observation period. The results showed that the mating process included mounting, female cooperation, and successful copulation. Overall, the three breeds preferred mating on land. More than thirty percent of the mating time was primarily concentrated from 4:00 p.m. to 6:00 p.m. in domesticated geese from Anser cygnoides, the corresponding values for Sichuan white geese and Zhedong white geese were 32.0% and 33.3%, respectively. The mating of the Hungarian geese usually took place in the morning. In addition, the frequency of successful copulation of Sichuan white geese and Zhedong white geese were 2.31 and 1.94 times per day, significantly greater than that of Hungarian geese (0.89 times). Furthermore, a significant positive correlation between successful copulation and laying rates (r = 0.985) or fertilization rates (r = 0.992) was observed in Hungarian geese. Taken together, the mating behaviors among the different breeds were mainly reflected in time preference and successful copulation frequency.