Source: PRAIRIE VIEW A&M UNIVERSITY submitted to NRP
IMPROVEMENT OF DAIRY PRODUCTION AND QUALITY BY A LACTOFERRIN-METHYL-SULFONYL-METHANE COMPLEX AS IMMUNOMODULATOR TO TARGET SUB-CLINICAL MASTITIS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
ACTIVE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1029893
Grant No.
2023-67018-39181
Cumulative Award Amt.
$300,000.00
Proposal No.
2022-09018
Multistate No.
(N/A)
Project Start Date
Apr 1, 2023
Project End Date
Nov 30, 2025
Grant Year
2023
Program Code
[A1332]- Food Safety and Defense
Recipient Organization
PRAIRIE VIEW A&M UNIVERSITY
P.O. Box 519, MS 2001
PRAIRIE VIEW,TX 77446
Performing Department
(N/A)
Non Technical Summary
Udder health affects the yield and quality of milk and, consequently, cheese production and quality. In milk with high somatic cell count (SCC), the deterioration of cheese-making properties occurs. The relationship between bacterial infection, inflammatory response, proteolysis of casein and, consequently, cheese yield and quality are complicated. Milk from mastitic udders exhibits increased proteolytic activity. Our goal is to measure the immune cell profiles and expression of inflammatory cytokines in milk leucocytes using flow cytometry and ELISA. Our central hypothesis is that intramammary infusion with a novel micronized lactoferrin-sulfonyl-methane complex (GRAS) might help in the elimination and/or prevention of new infection by E. coli, Staph. aureus, Staph. chromogenes, Strep. uberis, and/or Strep. agalactiae sub-clinical mastitis, thereby resulting in enhanced milk quantity and quality. The work proposed may reduce the use of antibiotics in milk production systems and advance the science in the important area of reducing intramammary infections and problematic lesions.
Animal Health Component
70%
Research Effort Categories
Basic
10%
Applied
70%
Developmental
20%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
31134501090100%
Knowledge Area
311 - Animal Diseases;

Subject Of Investigation
3450 - Milk;

Field Of Science
1090 - Immunology;
Goals / Objectives
The goal of this project is to measure the immune cell profiles and expression of inflammatory cytokines in milk leucocytes in order to testthe eficciency of the micronized lactoferrin-methylsulfonyl-methane complex for curing/preventing bacterial infection responsible for sub-clinical mastitis. The specific objectives of this study are to: (a) examinethe effect of four major udder pathogenes: Staphylococcus aureus, Staphylococcuschromogenes, Escherichia coli and Streptococcus agalactiae which are commonly associated with sub-clinical mastitis; b) study the relationship between bacterial infection, milk composition, proteolysis of casein in milk, milk lipolysis,and curd firmness; and c) evaluate sensory characteristics of milks from dairy goats treated with the micronized lactoferrin-methyl-sulfonyl-methane complex (GRAS).
Project Methods
The micronized lactoferrin-methyl-sulfonyl-methane complex (GRAS) will be prepared according to methods described by Mora-Gutierrez et al. (2022a). Duplicate quarter milk samples will be taken aseptically and submitted to the laboratory within one hour. Bacteriological analysis will be performed according to accepted standards of microbiological procedurs as described by the National Mastitis Council. All samples will be taken during milking. Somatic Cell Count (SCC) in the milk samples will be measured using aflow cytometer. Monoclonal antibodies will be used to measure plasma cytokine levels usingEnzyme-Linked Immuno-Sorbant Assay (ELISA). Milk gross composition (protein, fat and lactose contents) will be measured using the Milkoscan 6000. Milk lipolysis will be measured by GC-MS. Milk oxidation will be measured by TBARS. Proteolysis of casein will be measured by measuring the absorbance at 650 nm in a UV/Vis spectrophotometer. The level of tyrosine will be determined by comparing the absorbance values with a standard curve of known tyrosine concentration. The extent of proteolysisis proportional to the level of tyrosine released during the hydrolysis of proteins. Curd firmness will be determined by Optigraph. Sensory evaluation of milk samples will be performed by trained panelists.

Progress 04/01/24 to 03/31/25

Outputs
Target Audience:Dairy Scientists, farmers, and industry will benefit of our research project. Changes/Problems:We have not encountered any major problems that will require major changes in the planned approaches for the accomplishment of this research project. What opportunities for training and professional development has the project provided?A Research Specialist is conducting some of the laboratory experiments and has been assigned duties at 45% of this research project. One undergraduate student has been assigned to this research project. This student received training in our laboratory. This project has provided a unique professional development opportunity for this student, who is majoring in Animal Science at Prairie View A&M University (PVAMU). How have the results been disseminated to communities of interest?The results of this research project are being disseminated to the scientific community via publications in scientific journals. What do you plan to do during the next reporting period to accomplish the goals?Phase 1: To study the potential therapeutic effects of the intramammary infusion of vitamin D3 encapsulated by complex coacervates with lactoferrin-alginate and zinc oxide nanoparticles in synergy with guava (Psidium guajava L.) leaf extract against Escherichia coli in dairy goats. The aim of the study is to promote sustained mastitis treatment. Phase II: To study the effect of the oral administration of vitamin D3 delivered by an emulsion gel comprised of soy protein isolate and the oligosaccharide stachyose on immune response and performance in dairy goats. The aim of the study is to alter the composition of the rumen microbiota, thereby having a beneficial effect on immune response and performace in dairy goats.

Impacts
What was accomplished under these goals? This project is to provide dairy farm animals natural immunity against infectious diseases that have plagued U.S. dairy farmers for decades and are estimated to cost them billions of dollars annually. Mastitis, an acute inflammation of the mammary glands caused by pathogens, reduces the productive life of the affected dairy farm animal and the quality of the milk. The poor hygiene in dairy farm animals contributes to the proliferation of the pathogen Staphylococcus aureus. However, other pathogens may be involved in infectious mastitis, such as Escherichia coli, Staphylococcus chromogenes, Streptococcus uberis, and Streptococcus agalactiae. The animals infected by such pathogens are clinic dependent on the capacity and virulence of the strain and the host responsiveness. One of the main concerns of the dairy industry is bacterial resistance, as milk is the main substrate of highly consumed dairy food products such as cottage cheese, ice-cream, and yogurt. This makes antibiotics ineffective. Careful and responsible management of food-producing animals is an important aspect of the fight against antibiotic resistance. The use of vaccines reduces bacterial and viral disease prevalence in farm animals, thereby, reducing the need for antibiotics. USDA-NIFA funding has supported basic research on immune systems across species to optimize vaccine and adjuvant development. In this research project, a recombinant protein vaccine (rYidR) containing vaccine adjuvant (aluminum hydroxide gel) against coliform infection was used in 6 dairy goats (Group 1). The same vaccine (rYidR), but with the vaccine adjuvant lactoferrin-methyl-sulfonyl-methane complex was used in 6 dairy goats (Group 2). Another group of 6 dairy goats was used as placebo after injecting them with a physiological saline solution at pH 7.2 (Group 3). All animals in the study received a total of five injections of the rYidR vaccines. All injections were given subcutaneously in the neck region using automatic self-filling syringes. The experimental animals were selected from the dairy herd based on the number of Somatic Cell Counts (SCC). Mastitis-causing bacteria increase SCC. Therefore, animals with SCC ≥ 150,000cells/mL were selected for this study. Coliforms such as Escherichia coli and Klebsiella spp. are 'naturally' occurring in milks with high SCC. Immunized dairy goats were assessed by antibody serum titers (IgM and IgG). Milk production was also evaluated.The results will be reported at the scientific meetings and to the dairy industry. Dietary interventions can be used to beneficially affect immunity in commercially important dairy farm animals such as cow, goat, and sheep. Further research on nutrients and phytochemicals is required to define efficacious interventions appropriate to the goals of developing diets that can maintain normal immune defenses in sick animals and promote optimal immune function in healthy animals to reduce their risk of infectious diseases.

Publications

  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Influence of Deliverable Form of Dietary Vitamin D3 on the Immune Response in Late-Lactating Dairy Goats. Dairy 2024, 5, 308-315


Progress 04/01/23 to 03/31/24

Outputs
Target Audience:Dairy scientists, farmers, and industry will benefit of our research project. Changes/Problems:We have not encountered any major problems that will require major changes in the planned approaches for the accomplishment of this research project. What opportunities for training and professional development has the project provided?A Research Specialist, Dr. Maryuri. T. Nunez de Gonzalez, is conducting some of the laboratory experiments and currently has been assigned at 50% of this research project. Additionally, an undergraduate student has been assigned to this research project and is currently undergoing training.This project has provided a unique professional development opportunity for this student. How have the results been disseminated to communities of interest?The results of this research project have not been disseminated to the scientific community yet; however, we are planning to present our data at the Association of 1890 Research Directors (ARD) in April 6-9, 2024 and the American Dairy Science Association (ADSA) in June 16-19, 2024. A research paper is being submitted for consideration as a publication to theJ. Anim. Phys. Anim. Nutr. March 29, 2024. What do you plan to do during the next reporting period to accomplish the goals?Currently, we are developing a vaccine containing the recombinant protein (YidR) combined withsulfur-saturated bovine lactoferrin-alginate complex coacervatesadjuvants against clinical mastitis.Immunized dairy goats will be assessed by antibody serum titers (IgM and IgG). Milk production will also be evaluated.The results will be reported at the scientific meetings and to the dairy industry.

Impacts
What was accomplished under these goals? Mastitis-causing bacteria are able to establish persistent infections in the mammary gland of commercially important dairy animals despite the presence of strong specific humoral and cellular immune mechanisms. The novel vitamin D3encapsulated by complex coacervation with sulfur-saturated bovine lactoferrin-alginate was investigated as an immunomodulator in late-lactating dairy goats. Blood serum biomarkers of the immune response were assessed. Dairy goats (n = 18) in late lactation were randomly assigned to three experimental groups (n = 6), after balancing for weight. Dairy goats were orally administered with 0.35 mg vitamin D3/day in the unencapsulated form and 0.35 mg vitamin D3/day in the encapsulated powder form. Another group received the basal diet. The experimental period lasted 6 weeks.The blood serum concentrations of 25-hydroxyvitamin D3[25-(OH)-D3], lactoferrin, immunoglobulin A (IgA), and interferon-gamma (INF-γ) were measured. The delivery of vitamin D3to dairy goats resulted in a marked increase of 25-(OH)-D3concentration in serum while the serum level of lactoferrin also increased. Alternatively, the serum levels of IgA and the immunomodulatory cytokine INF-γ were elevated following supplementation with the encapsulated vitamin D3. Overall, the capsules in the powder form used in this study kept vitamin D3highly bioavailable.

Publications

  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Title: Encapsulation of vitamin D3 and curcumin combination by sulfur-saturated bovine lactoferrin-alginate complex coacervates for enhancing the immune response: In-vitro study. 2024 American Dairy Science Association (ADSA) Annual Meeting, West Palm Beach, FL., June 16-19, 2024
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2023 Citation: Title: Sulfur-Saturated Bovine Lactoferrin: In-vitro Digestibility and Digestion Behavior. 2024 Association of 1890 Research Directors (ARD) Symposium, Nashville, TN., April 6-9, 2024
  • Type: Journal Articles Status: Submitted Year Published: 2024 Citation: Title: Impact of vitamin D3 encapsulated by sulfur-saturated bovine lactoferrin-alginate complex coacervates on the immune response of late-lactating dairy goats. Journal of Animal Physiology and Animal Nutrition (WILLEY)