Source: UNIVERSITY OF CALIFORNIA, DAVIS submitted to NRP
OPTIMIZING BOVINE OCULAR ANTIBODY RESPONSES TO MORAXELLA BOVIS CYTOTOXIN
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
1008849
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
Dec 8, 2015
Project End Date
Sep 30, 2017
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671
Performing Department
Medicine & Epidemology
Non Technical Summary
Ocular immune responses of steers to two different formulations of an intranasal vaccine against Moraxella bovis (agent of infectious bovine keratoconjunctivitis; IBK; 'pinkeye') cytotoxin will be measured to determine whether one vaccine adjuvant is superior to another vaccine adjuvant in terms of stimulation of local ocular immune responses to M. bovis cytotoxin. To achieve this objective, a study will be conducted in which steers will be vaccinated. Serum and tear samples will be collected weekly for 8 weeks. At the end of the sample collection period, theserum and tear antibody responses over the course of 8 weeks will be measured and compared between vaccine groups to determine which group of steers has better ocular immune responses to M. bovis cytotoxin.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
31133101090100%
Knowledge Area
311 - Animal Diseases;

Subject Of Investigation
3310 - Beef cattle, live animal;

Field Of Science
1090 - Immunology;
Goals / Objectives
The major goals of this project are:#1) Vaccinate 3 groups of steers intranasally on days 0 and 21 with either: a) 500 µg MbxA adjuvanted with polyacrylic acid plus oil-in-water with DDA (Group 1); b) 500 µg MbxA adjuvanted with polyacrylic acid (Group 2); or c) water with polyacrylic acid plus oil-in-water with DDA (Group 3); and#2) Quantitate post-vaccinal fold changes over baseline in the following immune response variables: a) tear MbxA-specific IgA; b) tear MbxA-specific IgG; c) serum MbxA-specific IgG; d) tear cytotoxin neutralizing titer; and e) serum cytotoxin neutralizing antibody titer.
Project Methods
Design: Project goal#1: Fifteen healthy finishing steers maintained at the UC Davis feedlot will be used for this study.Based on results from our most recent intranasal vaccine study, we anticipate that 5 steers per group will be adequate for this study. Recombinant Moraxella bovis cytotoxin (MbxA) will be prepared and quantitated in Dr. Angelos' Laboratory.Either 500 µg MbxA (Group 1) or water (Group 3) will be adjuvanted according to manufacturer's instructions (MVP Technologies, Omaha, NE) with Carbigen™/Emulsigen™-D (1:1). For steers in Group 2, 500 µg MbxA will be adjuvanted with Carbigen™ according to manufacturer instructions. A 2 ml dose of vaccine will be administered intranasally (1 cc per nostril) to steers in each study group on days 0 and 21. Prior to vaccination, pooled tear samples from left and right eyes and a serum sample will be collectedand stored at -20oC until IgA and total IgG quantitation and completion of cytotoxin neutralizing assays. Tear and serum samples will be collected every 7 days for 8 weeks.Design: Project goal#2: The following immune response variables will be measured from tear and serum samples to quantitate the fold changes from prevaccination (day 0) to each subsequent weekly collection day: a) tear MbxA specific IgA; b) tear MbxA specific IgG; c) serum MbxA specific IgG; d) tear native M. bovis cytotoxin neutralizing antibody titer; and e) serum native M. bovis cytotoxin neutralizing antibody titer. Tear and serum cytotoxin neutralizing assays and ELISAs to quantitate the MbxA-specific tear IgA, tear IgG and serum IgG concentrations will be performedexcept that for the tear IgA ELISA, tear IgG ELISA, and tear cytotoxin neutralizing assays, the volume of tear samples entered into these assays will be standardized based on the sample total protein concentration (BCA Protein Assay; Pierce).Anticipated Results: We anticipate that steers vaccinated with MbxA adjuvanted with polyacrylic acid plus oil-in-water with DDA will have the highest fold changes in tear MbxA-specific IgA. We also predict that immune responses in steers vaccinated with MbxA adjuvanted with polyacrylic acid plus oil-in-water with DDA will be more sustained over the 8 week trial as compared to steers that receive MbxA plus polyacrylic acid alone. We do not anticipate increased MbxA responses in tears or sera of steers vaccinated with adjuvant alone (Group 3). We have already completed 2 studies with similar study design and have previously performed all of the procedures described above and do not anticipate any major pitfalls that would prevent us from completing the proposed research.Data Analysis: Differences in fold changes of MbxA specific tear IgA, tear IgG, serum IgG, tear cytotoxin neutralizing titer and serum cytotoxin neutralizing titer from day 0 to each weekly sample day will be evaluated using analysis of variance using the Friedman test. Post hoc analyses of differences between study groups will be determined using the Wilcoxon signed rank test with Bonferroni adjustment for multiple comparisons. P < 0.05 will be considered significant; for the Bonferroni adjustment a P < 0.0167 (0.05/3 comparisons) will be considered significant.

Progress 10/01/16 to 09/30/17

Outputs
Target Audience: Nothing Reported Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?During the next period our goal will be to write, submit, and publish a manuscript describing the results of this study.

Impacts
What was accomplished under these goals? We completed the above listed major goals of this project which were: #1) Vaccinate 3 groups of steers intranasally on days 0 and 21 with either: a) 500 µg Moraxella bovis recombinant cytotoxin subunit (MbxA) adjuvanted with polyacrylic acid plus oil-in-water containing dimethyldioctadecylammonium bromide (DDA) (Group 1); b) 500 µg MbxA adjuvanted with polyacrylic acid (Group 2); or c) water with polyacrylic acid plus oil-in-water with DDA (Group 3); and#2) Quantitate post-vaccinal fold changes over baseline in the following immune response variables: a) tear MbxA-specific IgA; b) tear MbxA-specific IgG; c) serum MbxA-specific IgG; d) tear cytotoxin neutralizing titer; and e) serum cytotoxin neutralizing antibody titer. Results of the immune response assays indicated that the Group 1 cattle (vaccinated with MbxA adjuvanted with polyacrylic acid plus oil-in-water containing DDA) had the highest mean fold changes in MbxA-specific tear IgA responses from study day 0 (day of primary vaccination) to day 14 (d0-d14), d0-d28, d0-d42, and d0-d49. These differences were significantly higher than the control group (Group 3; vaccinated intranasally with water with polyacrylic acid plus oil-in-water with DDA) at the d0-d14 and d0-d42 time intervals. The fold changes in the tear cytotoxin neutralizing titers at d0-d14, d0-d28, d0-d42, and d0-d49 were also highest in Group 1 cattle, however, these differences versus Group 1 or Group 2 animals were not significant. Mean fold changes in MbxA-specific tear IgG responses were higher in both Group 2 (group that received MbxA adjuvanted with polyacrylic acid without DDA) and Group 1 cattle versus the control group (Group 3), however, the highest fold changes were observed in Group 2; the differences were significantly higher than the control group at d0-d14, d0-d28, and d0-d42 time intervals. Mean fold changes in serum MbxA-specific IgG was also significantly higher in Group 2 versus Group 3 (control) at the d0-d14 and d0-d28 time intervals. Significant differences were not observed between groups in the fold changes in geometric mean serum cytotoxin neutralizing titers between groups at the d0-d14, d0-d28, d0-d42, d0-d49, and d0-d56 time intervals. In summary, tear IgA responses appeared highest in cattle that received an intranasal vaccine adjuvanted with polyacrylic acid plus DDA; it is likely that this difference accounted for the higher tear neutralizing responses that were observed in Group 1 animals versus the other 2 groups.

Publications


    Progress 12/08/15 to 09/30/17

    Outputs
    Target Audience:Results of this study were used to inform adjuvant selection for a subsequent experimental vaccine study (see CA-V-VME-4074-H "Intranasal Moraxella bovis/Moraxella bovoculi native cytotoxin vaccine to prevent naturally-occurring infectious bovine keratoconjunctivitis"). As such, the target audience was the investigation team who was conducting the study for CA-V-VME-4074-H). Following publication of study results, our target audience will be broader including the scientific and lay communities with interest in vaccine development for IBK. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?Data from this study has informed adjuvant selection for a subsequent study by the Principle Investigator. A manuscript describing the results of this study will be prepared and submitted for publication to the scientific and lay communities. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

    Impacts
    What was accomplished under these goals? We completed the above listed major goals of this project which were: #1) Vaccinate 3 groups of steers intranasally on days 0 and 21 with either: a) 500 μg Moraxella bovis recombinant cytotoxin subunit (MbxA) adjuvanted with polyacrylic acid plus oil-in-water containing dimethyldioctadecylammonium bromide (DDA) (Group 1); b) 500 μg MbxA adjuvanted with polyacrylic acid (Group 2); or c) water with polyacrylic acid plus oil-in-water with DDA (Group 3); and #2) Quantitate post-vaccinal fold changes over baseline in the following immune response variables: a) tear MbxA-specific IgA; b) tear MbxA-specific IgG; c) serum MbxA-specific IgG; d) tear cytotoxin neutralizing titer; and e) serum cytotoxin neutralizing antibody titer. Results of the immune response assays indicated that the Group 1 cattle (vaccinated with MbxA adjuvanted with polyacrylic acid plus oil-in-water containing DDA) had the highest mean fold changes in MbxA-specific tear IgA responses from study day 0 (day of primary vaccination) to day 14 (d0-d14), d0-d28, d0-d42, and d0-d49. These differences were significantly higher than the control group (Group 3; vaccinated intranasally with water with polyacrylic acid plus oil-in-water with DDA) at the d0-d14 and d0-d42 time intervals. The fold changes in the tear cytotoxin neutralizing titers at d0-d14, d0-d28, d0-d42, and d0-d49 were also highest in Group 1 cattle, however, these differences versus Group 1 or Group 2 animals were not significant. Mean fold changes in MbxA-specific tear IgG responses were higher in both Group 2 (group that received MbxA adjuvanted with polyacrylic acid without DDA) and Group 1 cattle versus the control group (Group 3), however, the highest fold changes were observed in Group 2; the differences were significantly higher than the control group at d0-d14, d0-d28, and d0-d42 time intervals. Mean fold changes in serum MbxA-specific IgG was also significantly higher in Group 2 versus Group 3 (control) at the d0-d14 and d0-d28 time intervals. Significant differences were not observed between groups in the fold changes in geometric mean serum cytotoxin neutralizing titers between groups at the d0-d14, d0-d28, d0-d42, d0-d49, and d0-d56 time intervals. In summary, tear IgA responses appeared highest in cattle that received an intranasal vaccine adjuvanted with polyacrylic acid plus DDA; it is likely that this difference accounted for the higher tear neutralizing responses that were observed in Group 1 animals versus the other 2 groups.

    Publications


      Progress 12/08/15 to 09/30/16

      Outputs
      Target Audience: Nothing Reported Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?We will be measuring the immune responses in the collected serum and tears and we will be preparing a manuscript describing results of this study.

      Impacts
      What was accomplished under these goals? We accomplished goal #1 above and are currently in the process of addressing goal #2.

      Publications