Location: Warmwater Aquaculture Research Unit
2018 Annual Report
Objectives
1. Develop and adapt existing imaging and photon emitting technologies toward describing the intrauterine environment, and sperm qualities such as motility, and develop strategies to enhance and improve reproductive health and fertility in food animal reproduction systems.
1.A. Examine in vivo uteroplacental hemodynamics following acute maternal infusions with vasoactive supplements.
1.B. Development of in vitro and ex-vivo approaches for cellular and tissue biophotonic imaging using nanoparticles.
1.C. Development of specific molecular-based approaches for in vivo biophotonic imaging.
1.D. Development of alternate biophotonic animal models and the use of digital infrared thermal imaging and near infrared (NIR) spectroscopic approaches to study specific reproductive health, environmental, and/or physiological processes in livestock and poultry.
2. Use novel imaging and related technologies for the tracking of relevant pathogens (disease stressors; e.g. Salmonella, Mycobacterium avian paratuberculosis) in avian and livestock hosts using photon emitting sentinels in the animal system and/or environment (e.g., nanoparticles, energy transfer systems, transformed bacterium) to address bacterial abundance and persistence related to livestock well-being and production performance, and develop mitigation strategies.
2.A. Use of biophotonics imaging strategies to establish potentially unknown infection sites of Mycobacterium avium subsp. Paratuberculosis in a rodent model.
2.B. Development of alternate biophotonic animal models and the use of digital infrared thermal imaging approaches to study various disease states in livestock and poultry.
Approach
There is a critical need for technological innovations that will permit production-based questions to be asked and answered in the context of the living animal. The overall goal of this project is to develop technologies that can perform in situ time-lapse and in vivo bio-imaging of cellular and molecular events and biological processes in livestock and poultry, in real-time. Specifically, this basic and applied research will target the improvement of reproductive efficiency and the mitigation of disease, which are both essential for efficient food and fiber production. Novel technologies will utilize photonic and/or thermal signatures, spectroscopy and/or fluorescence, ultrasonography, and nanotechnology in adaptive research aimed at facilitating physiological assessments related to reproduction and disease monitoring in livestock and poultry. To this end, this project is designed to cover a broad range of research in the organismal, cellular and molecular life sciences aimed at understanding production performance end-points through the development of new life-science research models. With these new models in place, translational research can then be applied in livestock and poultry production-management settings for application to the real-world enviroment.
Progress Report
Mississippi State University scientists have made additional progress in all of the planned milestones through the continued efforts of the project's Principal Investigator, an Agriculture Research Service (ARS) scientist at Stoneville, Mississippi, in concert with existing and new collaborations, and through the support of graduate students and research staff. Significant progress of note have been made specifically in the following areas: (1) Enhancing imaging in animal reproduction and disease states using bioluminescent and nanotechnology approaches: Specifically, scientists have developed a highly bioliuminescent Mycobacterium avium subsp. paratuberculosis (MAP) strain and have validated its kinetics and luminescent properties for future in vivo studies; with some preliminary work already accomplished in a chicken embryo model. Initial studies in poultry have evaluated the establishment of bioluminescent bacteria in the broiler embryo after in ovo injection. These studies have determined that in ovo injection into the amnion is a quicker method to administer probiotics to the embryo which reaches multiple tissues. Moreover, a real-time investigation of the antibacterial effect of single walled carbon nanotubes coated with pegylated silver has been completed for a better characterization of their novel actions on bacteria. In addition, antimicrobial properties of silver magnetic nanoparticles (Ag-MNP) against various foodborne pathogens (E. Coli, Salmonella Typhimurium, and Salmonella Anatum) that were rendered bioluminescent for real-time monitoring of bacterial growth inhibition and survival were tested. These studies will now be extended to assess the mitigation effects in infectious disease models; (2) Use of near infrared reflectance spectroscopy (NIR) and Doppler technologies for monitoring animal physiology: Specifically, NIR has been used for the rapid and non-destructive analysis of biofluids in numerous species for monitoring both physiological processes and reproductive health. In vivo uteroplacental dynamics are also being examined in both cattle and sheep using Doppler ultrasonography and laser Doppler perfusion techniques. Moreover, a novel fluorescent perfusion technique has been established to examine macroscopic blood vessel density of the placentome. These techniques are being validated by sampling placental tissue and comparing in vivo results versus those of molecular markers of placental blood perfusion and angiogenesis; (3) Research has expanded to include more poultry emphasis through applications of digital infrared thermal imaging in broiler production environments and have revealed, through serial imaging over the past year in production houses, potential deficiencies in housing insulation and construction that may influence environmental controls; and (4) Expansion of our imaging systems to poultry disease models have also resulted in a new model for tracking in ovo-injected migration of bacterial progression. Challenges have been experienced in transformation of our bioluminescent constructs into gram-positive anaerobic bacteria for poultry disease model development, but these short-comings are being addressed to achieve progress in this aspect of our objectives for the coming year. Studies have revealed in poultry models new methods for tracking bacterial progression in eggs, and these systems will be used to identify sites of colonization by both harmful and probiotic bacteria for further study. In addition, the use of thermal imaging in production broiler poultry houses have revealed structural changes that may need to be made to improve environmental control systems. Experimental types of insulation are being monitored in breeder houses that have begun to reveal “hot spots” of inconsistency within the insulated structure. These may directly impact the efficiency of environmental controls as well as the inputs and costs associated with optimal boiler/breeder housing, and research is ongoing to define these anomalies discovered through this research. These, and our other projects in this initiative in 2017-2018, supported directly (in full or in part) the research programs of three postdoctoral fellows and two Assistant Research Professors, in addition to the project PI’s. In addition, the project supports directly or indirectly the work, in part or in full, of two PhD students, three Mississippi State students, and five undergraduate students. A total of 11 peer-reviewed journal publications were produced, transmitting the research results to peers via the scientific literature.
Accomplishments
1. New method for rapid screening health and reproductive status in livestock. Real-time screening in livestock would improve the efficiency of detection of reproductive status and health status. Scientists at Mississippi State University through cooperative agreement with ARS scientists at Stoneville, Mississippi, have used ultrasonography to screen novel therapeutic supplements to increase uteroplacental blood flow in sheep and cattle. Several antioxidant supplements have been shown to have acute effects on in vivo placentome blood perfusion, which corresponds to changes in tissue angiogenic factor expression. These early validation results are expected to have a significant impact on the development of effective therapeutics to improve fetal development via increased placental blood flow.
2. Silver coated magnetic particles reveal potential novel biomarkers for inhibition of bacterial growth. Foodborne pathogens increasingly exhibit drug resistance against antibiotics, and despite the broad-spectrum of developed antibiotics during the last decades, bacteria still show multidrug-resistance. Advances in nanotechnology and microbiology provide promising applications in the inhibition of bacterial growth. Scientists at Mississippi State University through cooperative agreement with ARS scientists at Stoneville, Mississippi, tested the antimicrobial properties of silver magnetic nanoparticles (Ag-MNP) against various foodborne pathogens (E. Coli, Salmonella Typhimurium, and Salmonella anatum) that were rendered bioluminescent for real-time monitoring of bacterial growth inhibition and survival. Molecular changes were also evaluated. The results indicated the antimicrobial effects of Ag-MNP that was likely exerted through protein changes affecting critical bacterial functions. These proteins can serve as novel biomarkers for further targeting of bacterial contamination with potential impacts in livestock production.
Review Publications
Lemley, C.O. 2017. Investigating reproductive organ blood flow and blood perfusion to ensure healthy offspring. Animal Frontiers. 7(3):18-24.
Hart, C., Voelz, B., Brockus, K., Lemley, C. 2018. Hepatic steroid inactivating enzymes, hepatic portal blood flow and corpus luteum blood perfusion in cattle. Reproduction of Domestic Animals. 53:751-758.
Lemley, C., Camacho, L., Hallford, D., Vonnahme, K. 2018. Uteroplacental secretion of progesterone and estradiol-17ß in an ovine model of intrauterine growth restriction. Animal Reproduction Science. 193:68-78.
Park, S., Steadman, C., Chaudhari, A., Pillai, S., Singh, S., Ryan, P., Willard, S., Feugang, J. 2018. Proteomic analysis of antimicrobial effects of pegylated silver coated carbon nanotubes in Salmonella enterica serovar Typhimurium. Journal of Nanobiotechnology (Biomed Central Open Access). http://doi.org/10.1186/s12951-018-0355-0.
Feugang, J., Liao, S., Willard, S., Ryan, P. 2018. In-depth proteomic analysis of boar spermatozoa through shotgun and gel-based methods. BMC Genomics. http://doi.org/10.1186/s12864-018-4442-2.
Gastal, G., Aguiar, F., Rodrigues, A., Scimeca, J., Apgar, G., Banz, W., Feugang, J., Gastal, G. 2018. Cryopreservation and in vitro culture of white-tailed deer ovarian tissue. Theriogenology. 113:253-260.
Lewis, M., Durfey, C., Hartung, S., Steadman, C., Park, S., Clemente, H., Willard, S., Ryan, P., Feugang, J. 2017. Investigating the Cryotolerance of Boar Spermatozoa Subjected to Prior Selection. Journal Of Reproduction, Fertility And Development. 30:244.
Durfey, C., Burnett, D., Liao, S., Steadman, C., Crenshaw, M., Clemente, H., Willard, S., Ryan, P., Feugang, J. 2017. Nanotechnology-based selection of boar spermatozoa: growth development and health assessments of produced offspring. Horizons in Livestock Sciences Conference. 205:137-142.
Owen, M., McCarty, K., Hart, C., Steadman, C., Lemley, C. 2018. Endometrial blood perfusion as assessed using a novel laser doppler technique in Angus cows. Animal Reproduction Science. 190:119-126.
Owen, M., Ferjak, E., Cavinder, C., McCarty, K., Yankey, K., Hart, C., Burnett, D., Dinh, T., Lemley, C. 2017. Effects of body condition score (BCS) on steroid-and eicosanoid-metabolizing enzyme activity in various mare tissues during winter anoestrus. Reproduction of Domestic Animals. 53:296-303.
Jain, S., Park, S., Pillai, S., Ryan, P., Willard, S., Feugang, J. 2018. Applications of fluorescent quantum dots for reporductive medicine and disease detection. Intech. http://doi.org/10.5772.intech.72978.
