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Molting in Salmonella Enteritidis-Challenged Laying Hens Fed Alfalfa Crumbles. I. Salmonella Enteritidis Colonization and Virulence Gene hilA Response

K. D. Dunkley^*, J. L. McReynolds^*,,1, M. E. Hume, C. S. Dunkley^*,2, T. R. Callaway, L. F. Kubena, D. J. Nisbet and S. C. Ricke^*,3

^* Department of Poultry Science, Texas A&M University, College Station 77843; and Southern Plains Agricultural Research Center, Food and Feed Safety Research Unit, Agricultural Research Service, USDA, College Station, TX 77845

¹ Corresponding author: mcreynolds{at}ffsru.tamu.edu

The objectives of this study were to enumerate Salmonella enterica serovar Enteritidis colonization in fecal, cecal, and internal organs, and to compare the level of virulence gene expression (hilA) of experimentally challenged laying hens fed different dietary molt-induction regimens. Twelve Salmonella-free Single Comb Leghorn hens (>50 wk old) hens were randomly assigned to each of 6 treatment groups designated based on diet in 2 trials: 1) feed withdrawal Salmonella Enteritidis-positive (FW+), 2) fully fed Salmonella Enteritidis-positive (FF+), 3) 100% alfalfa crumble Salmonella Enteritidis-positive (ALC+), 4) feed withdrawal Salmonella Enteritidis-negative, 5) fully fed Salmonella Enteritidis-negative, and 6) 100% alfalfa crumble Salmonella Enteritidis-negative. A forced molt was induced by a 12-d alfalfa diet and a feed-withdrawal regimen. On d 4 of the molt, all hens in groups 1, 2, and 3 were challenged by crop gavage with 1 mL of inocula containing approximately 10⁶ cfu of nalidixic acid- and novobiocin-resistant Salmonella Enteritidis (phage type 13A). At the conclusion of both trials, all hens were euthanized and Salmonella Enteritidis colonization was enumerated in the cecal contents, liver, spleen, and ovaries. In addition, fecal (d 4 and 8) and cecal samples (necropsy at d 12) were collected postchallenge from treatment groups 1, 2, and 3 (Salmonella Enteritidis-positive) to quantify hilA expression by PCR. In both trials, all nonchallenged birds were Salmonella Enteritidis-negative; therefore, no further analysis was done. In trial 1, a 2-fold reduction in Salmonella Enteritidis colonization was observed in the ALC+ hens (log10 Salmonella Enteritidis of 1.99) compared with the FW+ hens (log₁₀ Salmonella Enteritidis of 3.89). In trial 2, a 4-fold reduction in Salmonella Enteritidis colonization was observed in the ALC+ hens (log₁₀ Salmonella Enteritidis of 1.27) compared with the FW+ hens (log₁₀ Salmonella Enteritidis of 5.12). In trial 2, Salmonella Enteritidis colonization in spleens was higher (P 0.05) in FW+ hens compared with ALC+ and FF+ hens. Relative expression of hilA was higher (P 0.05) in FW+ compared with FF+ hens, whereas the FF+ and ALC+ groups were not different (P > 0.05). In trial 2, hilA expression in FW+ hens was higher (P 0.05) for d 6, 11, and 12, respectively, when compared with ALC+ and FF+ hens. The results of these studies support the concept that changes in the gastrointestinal tract microenvironment, such as those created during feed deprivation, encourage Salmonella Enteritidis virulence and susceptibility in molted hens.

Key Words: Salmonella • hilA • alfalfa • molting • virulence

² Current address: Dept. of Poultry Science, University of Georgia, PO Box 748, Tifton, GA 31793-0478.

³ Current address: Center for Food Safety and Microbiology, IFSE, University of Arkansas, 2650 N. Young Ave., Fayetteville, AR 72704.

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