Location: Livestock Issues Research
Project Number: 3096-32000-008-10-T
Project Type: Trust Fund Cooperative Agreement
Start Date: Jan 18, 2016
End Date: Jan 17, 2021
BASE AGREEMENT: The objective of this study is to evaluate the effect of CLOSTAT, an active microbial with GRAS status, supplementation on innate immunity and pathogen migration, translocation, and microbial ecology shifts in weaned dairy calves following an oral Salmonella challenge. AMENDMENT 1: The specific objective of this experiment is to determine if feeding CLOSTAT will mitigate some of the negative effects associated with Bovine Respiratory Disease following an oral Salmonella infection in weaned dairy calves. The scope of work is similar to the scope of work in the existing agreement with the exception that the immune challenge will be a combination of an oral Salmonella Typhimurium challenge followed by the Bovine Respiratory Disease challenge that consists of an intranasal Bovine Herpesvirus-1 inoculation followed by an intra-tracheal Mannheimia haemolytica challenge. Additionally, there will be a preliminary study conducted to assess this dual challenge model prior to initiation of the full study to ensure that the model will provide the appropriate immune, endocrine, and physiological responses. Tissue and serum samples to be collected and analyzed will be similar to that in the original agreement with the exception of samples collected for the Bovine respiratory disease challenge such as antibody titers and nasal swabs, and the increased overall number of samples to be collected.
Weaned dairy calves (n >100; ~200 lbs; all same sex) will be sourced from a commercial calf ranch in the Texas Panhandle area. Calves will be housed and processed in accordance to the prescribed procedures of the calf ranch. Calves will be assigned to one of two treatment groups: 1) basal starter diet or 2) basal starter diet with the inclusion of CLOSTAT. Calves will remain on their respective treatment diets for 28 to 35 d. On d – 14, all calves will be weighed and a rectal fecal swab will be collected to determine the Salmonella status of the calves. Based on the Salmonella status and body weight, a subset of 40 calves will be selected and transported to Lubbock, TX (d -2). Calves will be individually housed in an environmentally-controlled facility with ad libitum access to treatment diets and water for the duration of the study. Upon arrival, a body weight will be obtained and a rectal swab will be collected to ensure animals are not shedding Salmonella. Additionally, calves will be assigned to 1 of 4 treatments (10 calves/treatment): 1) control diet – Salmonella (negative control), 2) control diet + Salmonella, 3) CLOSTAT – Salmonella, or 4) CLOSTAT + Salmonella. On d -1, indwelling jugular catheters and rectal temperature recording devices will be inserted in all calves. Calves will be returned to their pens and allowed to acclimate overnight. On d 0, calves will be orally inoculated with 1.0 x 108 CFU of Salmonella Typhimurium via a bottle containing bacterial culture combined with milk replacer. Whole blood samples will be collected from the catheter every 6 h for 48 or 96 h to evaluate whole blood hematology and serum cortisol and pro-inflammatory cytokines [tumor necrosis factor alpha (TNF-a), Interleukin-6, (IL-6) and Interferon gamma (IFN-')]. Rectal swabs will be collected daily to monitor fecal shedding of Salmonella. Also, visual evaluation of sickness will occur daily by a trained observer on a 4 point scale (1 = normal maintenance behaviors; 2 = calm, but head is distended and increased respiration; 3 = clinical signs of sickness and increase respiration with drool; 4 = lying on side with labored breathing and frothing at the mouth). Feed and water disappearance will be recorded daily. The Salmonella Typhimurium strain will be labeled with an antibiotic resistance transposon and will be cultured on selective media containing antibiotic to ensure that all Salmonella cultured matches the orally inoculated strain. At 48 h (d 2) post-inoculation, 5 calves from each treatment will be humanely euthanized using Phenobarbital under the supervision of a veterinarian. Animals will be weighed and tissues samples will be collected during necropsy. Tissues to be collected include: mesenteric lymph node, subiliac lymph node, ileum, jejunum, duodenum, transverse colon, cecum, liver, kidney, rumen fluid and synovial fluid. Each tissue will be homogenized, weighed, and Salmonella will be quantified utilizing selective media. Each tissue will also be enriched to identify Salmonella present below the limit of quantification. The necropsy process will be repeated for the remaining calves at 96 h post-inoculation.