Pacific Center for Emerging Infectious Diseases Research
John A. Burns School of Medicine, University of Hawai'i at Manoa
Director: Dr. Richard Yanagihara

PILOT PROJECTS

Saguna Verma, Ph.D.
Junior Researcher, Retrovirology Research Laboratory,
John A. Burns School of Medicine

Effects of Selenium Deficiency on Genomic Mutation of RNA Viruses
Nutritional deficiencies in antioxidants, such as selenium and vitamin E, lead to increased oxidative stress in humans. In animal models, increased oxidative stress during viral infection causes point mutations in certain RNA viruses, which are associated with heightened virulence. The central hypotheses of this proposal are that selenium deficiency causes genomic mutations in a wide variety of RNA viruses and that these mutations, which lead to increased virulence, can be monitored in vitro. The primary objective of this project is to investigate how RNA viruses may be affected by selenium deficiency within the cells in which they replicate.

Specific aims:

1. Develop an in vitro system to determine if selenium deficiency leads to increased mutations of viral genomic RNA for West Nile virus and Coxsackievirus B.
2. Identify which specific nucleotides within the genomes of these RNA viruses are highly susceptible to mutations caused by selenium deficiency.

To achieve these specific aims, we will develop an in vitro tissue culture system that allows the growth of cells under selenium-deficient conditions. Vero (African green monkey kidney) cells, which are highly susceptible to a wide variety of RNA viruses, will be depleted of selenium following inoculation with less virulent strains of coxsackievirus B and West Nile virus. At least three different gene regions of progeny virus, collected after each of three passages, will be amplified by reverse transcriptase-polymerase chain reaction (RT-PCR), then cloned. Approximately 20 randomly selected clones of each virus from each passage level will be sequenced to ascertain the frequency and nucleotide position of mutations. Comparisons will be made with point mutations associated with increased viral pathogenicity in selenium-deficient animals. The newfound knowledge from this project will provide insights into the emergence of pathogenic viruses.

James Holm-Kennedy, Ph.D.
Professor, Department of Electrical Engineering, College of Engineering

Development of a Novel Si-Based Biosensor Platform for Rapid Human Disease Diagnostics
A proprietary novel silicon-based biosensing platform for low-cost, rapid identification of human pathogens is under development and innovation based on prior proof of concept work. In this grant, fundamental studies are directed to building a solid pathogen diagnostics system foundation for future human pathogen sensing applications. The current activities focus on:

1. Commercial level sensor fabrication: Production of the next proprietary sensor prototype.
2. Sensor Sensitivity Characterization.
3. Prototype Pathogen detection.

These “fundamental” studies are intended and expected to lead to broad pathogen (human, animal and plant) diagnostic applications of this new, Silicon-based, biochemical sensoring platform. Extended future applications are planned. These include, but are not limited to, addressing nucleic acid diagnostics, proteomics and genetics applications, incorporation of sensor arrays, automated testing and related instrumentation development.

Tung T. Hoang , Ph.D.
Assistant Professor, Department of Microbiology, College of Natural Sciences

Characterization of Lung Surfactant Lipid Degradation in Pseudomonas aeruginosa
Pseudomonas aeruginosa, an opportunistic pathogen which causes life-threatening infections in wounds, burns and in particular the lung of cystic fibrosis patients, is a bacterium which is extremely difficult to eradicate with antibiotics due to resistance mechanisms and biofilm formation. The pathogenesis of P. aeruginosa lung infection in cystic fibrosis patients is not clear. A better understanding of the pathogenesis and mechanisms of growth of P. aeruginoa in the lung may lead to identification of new ways to eradicate the organism in these chronic lung infections.

Specific aims:

1. Compare the in vitro versus in vivo expression profile of P. aeruginosa, using Affymetrix GeneChip.
2. Perform real-time PCR analyses of a selected group of genes for the in vitro condition (grown on PC) and the in vivo condition (CF sputum).