Jean Hall, DVM, Ph.D. is a professor in the Department of Biomedical Sciences.

Dr. Hall’s laboratory is performing research studies to determine how nutrition affects immunity. They are interested in nutrigenomic technology, or the study of how nutraceuticals affect the expression of genes involved in the immune response. They have several dog studies in progress to determine whether dietary changes can alter innate immune responses via changes in gene expression. So far, they have shown that phagocytosis by peripheral blood neutrophils and transcript levels of genes involved in neutrophil-mediated functions are decreased in older dogs compared with dogs less than 1 year of age, which may contribute to increased morbidity and mortality with aging. Currently, they are trying to determine if dietary modifications can enhance neutrophil functions, and thus innate immunity, particularly in older dogs.

Dr. Hall’s other research projects involve sheep and cows supplemented with selenium (Se) and its effects on the immune response. Selenium has been known to function as a nutrient for over 50 years. However, the dietary requirements of selenium for optimal immune function remain to be determined. In addition, the efficacy of organic versus inorganic sources of selenium has not been thoroughly investigated. Dr. Hall’s goals are to determine if supplementing selenium at levels above those currently recommended can improve innate and adaptive immune responses, and whether organic selenium has increased bioavailability compared to inorganic selenium. Agronomic biofortification is defined as increasing the bioavailable concentration of an essential element in edible portions of crop plants through the use of fertilizers. They are also investigating the potential for using Se-containing fertilizers to increase crop Se concentrations.

Mahfuz Sarker, Ph.D. is an associate professor with joint appointment in the Department of Biomedical Sciences, College of Veterinary Medicine and Department of Microbiology, College of Science.

Overview of current research projects: Dr. Sarker’s group conducting research on the molecular pathogenesis of C. perfringens type A isolates associated with C. perfringens type A food poisoning and non-food-borne gastrointestinal (GI) diseases in humans, and GI diseases in domestic animals. Specifically, they investigate the molecular mechanisms of i) C. perfringens sporulation ii) sporulation-regulated synthesis of C. perfringens enterotoxin (CPE), a major virulence factor of C. perfringens pathogenesis iii) spore germination, and iv) spore resistance to various stress factors such as, chemicals, heat and high pressure processing.

The gram-positive, spore-forming, anaerobic C. perfringens bacteria is an important cause of both histotoxic and GI diseases in humans and domestic animals. C. perfringens type A food poisoning currently ranks as the third most commonly reported food-borne disease in the United States. In the U.S. alone C. perfringens type A food poisoning results in annual economic losses of over $120 million dollars. The development of preventive measures against C. perfringens-mediated diseases has been hampered by the lack of understanding of the molecular mechanisms of C. perfringens spore-formation, -germination and -resistance. A unique feature of C. perfringens pathogenesis is that it can produce toxins/virulence factors in two stages of its life cycle (Fig. 1). C. perfringens can remain as vegetative cells or initiate the sporulation process if environmental conditions are not favorable. During vegetative growth, C. perfringens produces and secretes many toxins including, a-toxin that is responsible for gas gangrene. Typically, vegetative cells continue to grow until they sense environmental signals that triggers sporulation. At the later stage of sporulation process, the mother cells lyses to release the mature dormant spores and CPE, which has been identified as an essential virulence factor for GI diseases. C. perfringens dormant spores are highly resistant to heat and other environmental insults, and can survive for long periods in the environment. Once conditions are favorable, these spores undergo germination, outgrowth and return to vegetative cells. Spore germination is an irreversible process in which a highly resistant, dormant spore is transformed into a metabolically active cell.

Sarker lab conducted a five-year study on the mechanism of C. perfringens spore resistance through grants from US Department of Agriculture and Army Research Office. They have shown that C. perfringens spores are highly resistant to heat, chemicals, UV-radiation and high hydrostatic pressure processing. Their studies have found that the main factors involved in these resistance processes are: i) the small-acid soluble proteins (SASPs) that saturate the spore’s DNA; ii) a low spore core water content, that protects essential enzymes and DNA in the spore core from environmental insults, is controlled in part by SpmA-B and DacB proteins; iii) the spore’s large depot of pyridine-2,6-dicarboxylic acid (dipicolinic acid (DPA)) which is pumped into the spore core during sporulation at least in part by the SpoVA proteins.

One approach to develop efficient therapies against C. perfringens diseases is to block or induce spore germination. Blocking spore germination would block the resumption of growing vegetative cells, while inducing germination would yield C. perfringens spores that have lost their resistance properties to conventional treatments applied in the food industry and in clinical settings, and thus becoming more sensitive to inactivation by milder treatments. Therefore, basic knowledge on the mechanism of spore germination is warranted. Dr. Sarker was recently awarded a MURI (Multi-Disciplinary University Research Initiative) award from Department of Defense for five years to study the Mechanism of Clostridium perfringens Spore Germination and its Heterogeneity.

Sarker lab identified for the first time the nutrient germinants (KCl and a mixture of L-asparagine and KCl) and their cognate receptors (GerKA and/or GerKC) required for C. perfringens spore germination. In a following project, they showed that in C. perfringens: i) SpoVA proteins are essential for Ca-DPA uptake by the developing spore during C. perfringens sporulation; and ii) SpoVA proteins and Ca-DPA release, required for Bacillus subtilis signal transduction, are not required for the transduction of the germination signal from the receptors to the cortex-lytic enzymes (CLEs) during C. perfringens spore germination. Recently, they have also found that SleC is an essential CLE for cortex peptidoglycan (PG) hydrolysis during germination of spores of C. perfringens. The hydrolysis of cortex PG is the culminating event in the germination of bacterial spores, and is essential for resumption of enzymatic activity in the spore core and eventual vegetative growth. As a consequence, cortex hydrolysis is essential for spores of any pathogenic organism such as C. perfringens to cause disease. The identification of SleC as the major essential CLE in C. perfringens spores thus makes this enzyme of potential interest for development of inhibitors, since such compounds would block spore germination and thus the ability of spores to cause disease. Cortex hydrolysis also makes the now fully germinated spore much less resistant to common decontamination procedures. Consequently, a drug that could rapidly activate SleC in spores would also be useful, since such a drug would allow decontamination of the now germinated C. perfringens spores under less harsh conditions than needed for destruction of the much more resistant dormant spores. Sarker group anticipate that their research will facilitate the designing and developing a preventive measure against Clostridial diseases.

Dr. Manoj Pastey, Ph.D, is an assistant professor in the Department of Biomedical Sciences.

Dr. Manoj Pastey’s laboratory is conducting research work on the pathogenesis of,HIV, influenza, and respiratory syncytial virus (RSV).

HIV Research Study: Our laboratory is testing a polyherbal vaginal microbicide named “BASANT” that has been shown to inhibit a wide range of sexually transmitted pathogens including HIV. Preliminary studies have also shown safety and acceptability in Phase I (acceptability and toxicity study) human trials in India. Therefore, the next step is to verify the effectiveness of the BASANT in preventing HIV transmission in vivo. The central goal of these studies is to understand the mechanism of microbicide anti-HIV action and to determine the efficacy of BASANT in preventing intravaginal/intrarectal HIV transmission in humanized mouse model. In addition, the efficacy of BASANT will be evaluated against six major globally prevalent strains of genetically and biologically characterized HIV-1 isolates.

Influenza Research Study: Each year, influenza kills approximately 36,000 people in the United States. These deaths are mainly due to secondary bacterial infection. Therefore, we are focusing our research on identifying biomarkers in blood and urine for respiratory tract dysfunction caused by co-infection of Staphylococcus aureus and influenza virus. Accomplishments of the proposed goals will help us predict the evolution of S. aureus super-infection in patients with H1N1 influenza virus disease. Using mice co-infected with influenza virus and S. aureus, gene expression changes obtained from DNA-microarrays and proteomic changes obtained from mass spectrometry will aid in identification of early and clinically relevant diagnostic and prognostic bio-markers. This knowledge will allow development of a predictive statistical model to afford a better understanding of virulence mechanisms and pathogenesis of respiratory tract co-infections by these microorganisms, such that risk of pneumonia can be assessed and effective preventative or treatment regimens can be initiated in infected individuals.

RSV Research Study: Respiratory Syncytial Virus (RSV) is a leading cause of bronchopneumonia in infants and the elderly. There are no vaccines or effective treatment available. Knowledge of viral and host protein interactions is important for better understanding of the viral pathogenesis and may lead to development of novel therapeutic drugs. In our lab, we have shown that Respiratory Syncytial Virus Matrix (M) protein interacts with cellular adaptor protein complex (AP)-3 and its medium (µ) subunit. A yeast two-hybrid assay indicated a novel protein-protein interaction that was then further confirmed in a mammalian system by co-localization between the RSV M and AP3 µ 1 proteins in a cytoplasmic defined region via Confocal Laser Scanning Microscopy (CLSM) analysis. Further evidence of this novel interaction was indicated via the presence of a known adaptor protein µ subunit sorting signal sequence, YXXL that is conserved across various animal RSV M proteins. Subsequent Western blot studies also showed a specific upregulation in the amount of AP3 µ 1 protein found in the cell during RSV infection, while corresponding subunits of the AP3 complex were unaffected. The interaction of AP3 µ 1 with RSV M represents a critical insight into the life cycle of this important virus and may represent a novel drug target.