The number of bacteria that entered into COEC and the expression of selected AvBD genes were determined at 1 hpi. The results showed that ZM106 (pipB) was less invasive than ZM100 (wt) and introduction LY294002 of pPipB, a plasmid expressing the pipB gene, to ZM106 (pipB) complemented the invasion defect of this strain (Figure 6A). Although the number of ZM106 that entered into COEC was less than that of the wild type SE, ZM106 still induced the expression of AvBD2 and AvBD6 at levels higher than that induced by ZM100 (Figure 6B). Introduction of the cloned pipB gene into ZM106 weakened

the strain’s capacity to induce AvBD mRNA expression

(Figure 6B). Thus, differential induction of AvBDs by ZM100 and ZM106 was indeed associated with their genetic backgrounds, with or without a functional pipB. Figure 6 PipB-mediated entry of SE into COEC and suppression of AvBDs in SE-infected COEC. COEC in 48-well culture Daporinad molecular weight plates were infected with ZM100 (wt), ZM106 (pipB), or ZM106-C (pipB, pPipB) at MOI of 20:1 (bacteria:cell). Data shown are geometric means of three independent experiments ± standard deviation. 6A. Number of intracellular bacteria (log CFU/well) at 1 hpi. * indicates that the difference in the Ketotifen number of intracellular bacteria between ZM100 (wt) and ZM106 (pipB) is significant (p < 0.05). 6B. SE-induced changes in the mRNA expression of AvBDs in COEC at 1 hpi. * indicates that the difference between the amounts of AvBD transcripts in ZM100-infected COEC and ZM106-infected COEC is significant (p < 0.05). Discussion As a key component of innate

immune response, defensins are synthesized in many tissues, especially those constantly exposed to microbial pathogens [26–30]. For example, a number of AvBD genes are expressed in the vagina of laying hens and the amount of AvBD mRNA increases following LPS treatment [31]. Although the vagina is anatomically prone to exposure to intestinal or environmental pathogens, the isthmus is likely a critical site in terms of persistent reproductive tract colonization and egg membrane contamination by SE [32, 33]. In an attempt to understand the innate immune responses against SE colonization of chicken oviduct epithelium, we determined the AvBD expression profile in primary oviduct epithelial cells. Although the preparation of primary chicken oviduct epithelial cells is empirical, the COEC cultures used in this study consisted of a high percentage of epithelial cells and spontaneous apoptosis of COEC was minimal under the experimental conditions used.