Location: Ruminant Diseases and Immunology Research
2017 Annual Report
Objectives
Objective 1. Develop non-antibiotic interventions to prevent and control mastitis, including developing and testing non-antibiotic immune modulators to prevent periparturient dairy cows from developing mastitis, and developing and testing dry cow therapy(s) that use natural, non-antibiotic strategies that accelerate the development of the cow’s natural antimicrobial dry secretions to prevent mastitis infections in subsequent lactations.
Sub-objective 1.1: Develop and test non-antibiotic immune modulators to prevent periparturient dairy cows from developing mastitis.
Sub-objective 1.2: Develop and test a dry cow therapy that uses natural, non-antibiotic strategies that accelerates the development of the cow’s natural antimicrobial dry secretions to prevent mastitis infections in the subsequent lactation.
Objective 2: Determine the interactions between mastitis-causing pathogens and the host innate immune mechanisms in the mammary gland, starting with determining the host-pathogen interaction associated with Escherichia coli strains linked to persistent mammary gland infections, and determining the host pathogen interaction associated with Staphylococcus aureus persistent infections.
Sub-objective 2.1: Determine the host-pathogen interaction associated with Escherichia coli strains linked to persistent mammary gland infections.
Sub-objective 2.2: Determine the host pathogen interaction associated with Staphylococcus aureus persistent infections.
Approach
Mastitis is the most prevalent infectious disease in dairy herds and the most costly disease for dairy producers. Older cost estimates for mastitis are in the neighborhood of $2 billion per year for producers. Newer estimates of the economic impact of mastitis on the dairy industry calculate the cost of a single case of clinical mastitis to be approximately $586 due to mammary gland damage, loss of milk production, discarded milk, and the costs of treatment and labor. Antibiotics are the mainstay for mastitis treatment and control and dairy cattle with mastitis receive more antibiotic therapy for its prevention and treatment than for all other dairy cattle diseases combined. Valid concerns by consumers regarding antibiotic usage need to be addressed by research on non-antibiotic alternatives. To achieve the goal of reducing the use of antibiotics we need a better understanding of how the immune system is failing to completely eliminate mastitis infections. Progress towards this goal can be achieved in two ways. First, is to manipulate the host in a way that optimizes the immune response to pathogens. Second, to gain a better understanding of the various mechanisms that allow bacteria to evade the host’s immune system. To achieve the goal of manipulating the immune system to optimize its response to pathogens we plan to develop non-antibiotic interventions to prevent and control mastitis. This approach would include developing and testing non-antibiotic immune modulators to prevent periparturient dairy cows from developing mastitis, and developing and testing dry cow therapy(s) that use natural, non-antibiotic strategies that accelerate the development of the cow’s natural antimicrobial dry secretions to prevent mastitis infections in subsequent lactations. To achieve the second goal of understanding the mechanisms of how bacteria can evade the immune system by studying the mechanisms that allow for persistent mammary gland infections. Knowledge of how bacteria escape the immune system and establish persistent infections is a necessary precursor to any therapeutic for these persistent infections. Successful manipulation of the host immune system that targets the pathogen at the site of the infection holds the potential of clearing an infection without the use of antibiotics.
Progress Report
Sub-objective 1.1: Our first sub-objective was to optimize the dose of an expression vector designed to deliver an immune stimulator to increase the number of immune cells in the blood of cattle. The immune stimulator, called granulocyte colony-stimulating factor (G-CSF), is a natural protein that stimulates release of immune cells from the bone marrow into the blood. The genetic material that produces bovine G-CSF was cloned into an expression vector and given to cows. Initial studies demonstrated a need for a much higher dose of the viral vector to achieve the desired outcome. In 2016, a commercial version of bovine G-CSF with sustained activity was approved by the Food and Drug Administration. We have determined that this commercially-available product is more convenient to use than the G-CSF delivered in the virus vector. Our future G-CSF experiments will use the commercial version.
Sub-objective 1.2: The first dry cow experiment is complete but the dose of fatty acids used did not accelerate the development of the cow’s natural antimicrobial dry secretions as expected. We plan to repeat this experiment with higher doses in the coming year. We did however discover that some dry secretion dogma is questionable. The increase in lactoferrin dry secretions is associated with dry secretion antimicrobial actions compared to milk. However, what is not appreciated and found by our research is that among day 21 dry secretions lactoferrin is not at all correlated with antimicrobial effects on 6 bacterial strains among 12 cows. These day 21 dry secretions have antimicrobial activities that range from robust to none at all. The key antimicrobial factor(s) in dry secretions remains a mystery which we hope to uncover with proteomics and other approaches during the next 4 years of the project.
Sub-objective 2.1: We have sequenced all the messenger RNA (transcriptome) from several strains of the same bacteria Escherichia (E.) coli. Some of these bacteria cause persistent infections while others just a transient. Our goal is to determine what genes from very similar bacteria lead to some causing a more damaging and costly persistent infection. We have started that process by obtaining the data to begin our analysis.
Accomplishments
1. Genes controlling bacterial motility vary among different growth conditions. ARS researchers at Ames, Iowa have reported work this year that showed the differences in gene expression when bacteria are grown in conditions that cause bacteria to move in different ways. It has been shown that bacteria motility is related to virulence and therefore research leading to discoveries affecting bacterial motility are hypothesized to affect virulence. Furthermore, our previous research has indicated that there are important expression differences in proteins associated with bacterial motility when comparing Escherichia (E.) coli strains that cause transient or persistent intramammary infections. Our work has led to a new hypothesis of how to treat persistent intramammary infections, namely, to inhibit bacterial motility. In addition, we showed that bacterial iron regulation plays a critical role in bacterial motility. Our demonstration that bacterial motility is an important component of the pathogenesis of bacteria in causing mastitis in dairy cattle is critical knowledge required as part of the quest to develop non-antibiotic therapies.
