Research in Shelly Michalski's Lab

The laboratory of Dr. Michelle Michalski will be employing DNA microarrays to gain insight into the maturation process of filarial nematodes: arthropod-borne, threadlike worms that cause a variety of diseases in vertebrates. The life cycles of these nematodes are fascinating and complex. The adult worms live and mate in the tissues of vertebrate hosts and produce tiny offspring called microfilariae. In many species, the microfilariae circulate in the peripheral blood and are transferred to the arthropod host via a bloodmeal. Once taken up into the arthropod midgut, microfilariae undergo a series of developmental changes that allow them to become infective for the next vertebrate. Little is understood about the transition of microfilariae from the physiology of the vertebrate host to that of the arthropod. For example, it has been shown in an animal model that mosquitoes are refractory to infection with newly-borne microfilariae of Brugia spp., but are susceptible to older ones, suggesting that a maturation process occurs within the vertebrate host prior to transfer. Thus, Michalski hypothesizes that changes in gene expression occur during the maturation process. The REU student engaged in this project will use an animal model and microarrays to test this hypothesis in the filarial nematode Brugia malayi. Advantages to using B. malayi over other filarial nematodes are: (a) B. malayi is the causative agent of lymphatic filariasis in humans and there is a large body of knowledge concerning its biology, (b) it is the model organism for the WHO-funded Filarial Genome Project (http://www.nematodes.org/fgn/filgen1.html), and (c) its entire life cycle can be maintained in the laboratory within gerbils and mosquitoes. Microarray chips containing 65-70mer oligonucleotides representing approximately 4,000 B. malayi genes are available through collaborators at the Filarial Genome Project. Infected gerbils will be obtained from the Filariasis Repository located at the University of Georgia.

The student involved with this project will obtain gerbils that have been experimentally infected with male and female B. malayi. Gravid female worms will be isolated from infected gerbils and implanted into uninfected gerbils so that the age of released microfilariae can be accurately estimated. Young (1-2 day) and mature (>6 day) microfilariae will be collected, and mRNA will be isolated from both populations. These mRNA samples will be converted to cDNA, labeled with spectrally-distinguishable dyes (e.g. Cy3 and Cy5), then co-hybridized onto the microarray chips. This technique will allow the student to identify cDNAs that are expressed differentially between immature and mature microfilariae. These preliminary data can then be used to identify candidate genes whose function is related to maturation of microfilariae, and the differential expression of these genes will be verified by RT-PCR. Future students will correlate these data with protein expression profiles of both populations and will pursue gene knockout studies to identify those genes/proteins that are essential in the maturation process.