Progress 10/01/05 to 09/30/10
Outputs
OUTPUTS: The objectives of this proposal were to identify host genes that were modulated upon pathogen infection, and to determine if these host genes were important for resistance to infection. We identified several candidate genes and will continue to characterize their role in the immune response. We disseminated our findings through peer-reviewed publications and state, national, and international scientific meetings. The work has been evaluated by our scientific peers in the form of proposals submitted to NSF, NIH, and the Maine Science and Technology Foundation. My lab has taught and mentored a number of undergraduate and graduate research assistants. In addition, my lab is distributing a detailed protocol developed in my lab that measures the overall innate immune health of the fish. PARTICIPANTS: Carol Kim, PhD, Principal Investigator - oversaw progress of the project, mentored undergraduate and graduate students, conceived of scientific experiments, wrote manuscripts The following students participated in the project: Master's Students: Akshata Nayak, Greg Giasson, Matthew Sullivan, Chad Stevens PhD student: Con Sullivan Postdoctoral research fellow: Christopher Lage Technician: Juyoung Shim TARGET AUDIENCES: The target audience was undergraduate and graduate students as well as associated faculty. There has been formal classroom and laboratory instruction associated with this project. PROJECT MODIFICATIONS: The original workplan included the exposure of zebrafish to a bacterial pathogen and subsequent microarray analysis. These studies were to be conducted in collaboration with Dr. Renae Malek at The Institute for Genomic Research (TIGR) in Rockville, Maryland. In 2006, Dr. Malek left TIGR for another job opportunity, and was therefore, unable to continue our collaboration. In July 2007, I was fortunate to travel to Singapore for a 6 month sabbatical leave at the National University of Singapore. During my sabbatical, I conducted the microarray experiments outlined in the Significant Findings section. I was only able to carry out exposures to virus and the environmental toxicant, arsenic in the zebrafish, but did not have the time or the funds to carry out the bacterial exposures. We have not completed Specific Aim 2 to characterize the capacity of the zebrafish to mount an immune response to viral and bacterial pathogens following the disruption of the expression of genes that have been identified as upregulated or downregulated. We have identified caveolin-1 as a gene that is modulated upon viral infection and have conducted gene knockdown studies in the zebrafish for this gene. However, there are many other genes that we plan to characterized and will perform experiments similar to the ones outlined in the proposal.
Impacts
In this project we were attempting to develop a model system for fish disease control and prevention. Although zebrafish are a recognized model for human disease, they can also be used as a model for diseases of fish. By understanding the immune response of our model species, we hope to develop better strategies to enhance the immune response to infectious diseases and to develop more effective vaccines for economically important Maine fish species, such as rainbow trout and Atlantic salmon. My group has published a number of manuscripts in high impact journals and has successfully sought federal funding.
Publications
- Hermann, A.C. and C.H. Kim. 2005. Effects of arsenic on the zebrafish innate immune system. Mar. Biotechnol. 7:494-505.
- Millard, P.J., L.E. Bickerstaff, S.E. LaPatra, and C.H. Kim. 2006. Detection of infectious hematopoietic necrosis virus and infectious salmon anemia virus by molecular padlock amplification. J. Fish. Dis. 29:201-213.
- Lage, C.R., A. Nayak, and C.H. Kim. 2006. Arsenic ecotoxicology and innate immunity. Integr. Comp. Biol. 2006; 46: 1040-1054
- Sullivan, C., J.H. Postlethwait, C.R. Lage, P.J. Millard, and C.H. Kim. 2007. Evidence for Evolving TICAM Function in Vertebrates. J. Immunol. 178:4517-4527.
- Nayak, A.N, C.R. Lage, and C.H. Kim. 2007. Effects of Low Concentrations of Arsenic on the Innate Immune System of the Zebrafish (Danio rerio). Toxicol. Sci. 98:118-24.
- Sullivan, C., and C. H. Kim. 2008. Innate Immune System of the Zebrafish, Danio rerio. In Innate Immunity of Plants, Animals, and Humans. H. Heine, ed. 113-133.
- Sullivan, C. and C.H. Kim. 2008. Zebrafish as a model for infectious disease and immune function. Fish Shellfish Immunol. 25:341-50.
- Singer, J. T., M. J. Sullivan, L. A. Porter, R. T. Phennicie, and C. H. Kim. 2009. Broad host range plasmids for red fluorescent protein labeling of Gram-negative bacteria for use in the zebrafish model system. Appl. Environ. Microbiol. 76:3467-74.
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Progress 10/01/08 to 09/30/09
Outputs
OUTPUTS: During this reporting period we conducted and analyzed experiments that were included in publications from our laboratory. In addition, we mentored several undergraduate students and a middle school student. The research was presented at national and international conferences that focused on fish disease and animal health. The research work was disseminated in the form of talks and posters presented by students and the PI. PARTICIPANTS: Carol Kim, PI of the project, designed the experiments, discussed results with students, and wrote the publications for this funded research TARGET AUDIENCES: Target audience is the aquaculture industry in Maine. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
As the fisheries and aquaculture industries continue to expand in the state of Maine, disease control methods and diagnostic tests will be desperately needed. Through our studies, we will determine the role of innate immunity, and then modulate this immune function to upregulate host response to infection. We propose to conduct these studies in zebrafish, because much of the information gained through this model system can be transferred to other fish species, such as rainbow trout and Atlantic salmon, economically important fish of Maine. The long term goals for this proposal will be A) to isolate and characterize cytokines and immune factors from fish, and B) to use these factors to upregulate immune function for better disease control and vaccine development. The proposed studies will enable the development prophylactic, diagnostic and therapeutic procedures to reduce mortality, and thereby save the Maine aquaculture industry millions of dollars of lost revenue.
Publications
- Sullivan, C., J. Charette, J. Catchen, C.R. Lage, G. Giasson, J.H. Postlethwait, P.J. Millard and C.H. Kim. 2009. The gene history of zebrafish tlr4a and tlr4b is predictive of their divergent functions. J. Immunol. 83:5896-908.
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Progress 10/01/07 to 09/30/08
Outputs
OUTPUTS: The results have been disseminated through scientific publications in peer reviewed journals and at national and international scientific meetings. In addition, magazine articles in publications such as "UMaine Today" have been published for the community, faculty, and alumni. We also provide tours of our zebrafish facilities to the community and provide an overview of our scientific research. PARTICIPANTS: Carol Kim, PI of the project, designed the experiments, discussed results with students, and wrote the publications for this funded research. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Our findings have shown that although the zebrafish have many of the same immune components as mammals, the mechanism and function of these immune pathways in fish are different. As such, the immune systems of fish need to be carefully characterized rather than simply extrapolated from mammalian systems. We have recently shown that homologs of the zebrafish TLR pathways, although highly conserved, do not necessarily function as they do in mammals. We are using co-immunoprecipitation and yeast two-hybrid libraries to determine unique signaling partners in the zebrafish. We also conducting infection studies in the zebrafish to determine if the immune factors are important for resistance to bacterial and viral pathogens. The results of characterization of the immune pathways of finfishes could provide crucial information for the development of effective vaccines and prophylactics against infectious diseases.
Publications
- Sullivan, C., and C. H. Kim. 2008. Innate Immune System of the Zebrafish, Danio rerio. In Innate Immunity of Plants, Animals, and Humans. H. Heine, ed. 113-133.
- Sullivan, C. and C.H. Kim. 2008. Zebrafish as a model for infectious disease and immune function. Fish Shellfish Immunol. 25:341-50.
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Progress 10/01/06 to 09/30/07
Outputs
The results have been disseminated through scientific publications in peer reviewed journals. In addition, magazine articles in publications such as "UMaine Today" have been published for the community, faculty, and alumni. We also provide tours of our zebrafish facilities to the community and provide an overview of our scientific research.
Impacts
Our findings have shown that although the zebrafish have many of the same immune components as mammals, the mechanism and function of these immune pathways in fish are different. As such, the immune systems of fish need to be carefully characterized rather than simply extrapolated from mammalian systems. The results of characterization of the immune pathways of finfishes could provide crucial information for the development of effective vaccines and prophylactics against infectious diseases.
Publications
- Nayak, A.N, C.R. Lage, and C.H. Kim. 2007. Effects of Low Concentrations of Arsenic on the Innate Immune System of the Zebrafish (Danio rerio). Toxicol. Sci. 98:118-24.
- Lage, C.R., A. Nayak, and C.H. Kim. 2006. Arsenic ecotoxicology and innate immunity. Integr. Comp. Biol. 2006; 46: 1040-1054
- Sullivan, C., J.H. Postlethwait, C.R. Lage, P.J. Millard, and C.H. Kim. 2007. Evidence for Evolving TICAM Function in Vertebrates. J. Immunol. 178:4517-4527.
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Progress 10/01/05 to 09/30/06
Outputs
Global expression patterns following infection with SHRV or E. tarda were determined in the zebrafish. To establish gene expression signatures, zebrafish embryos were exposed to pathogen by static immersion, and then collected for total RNA extraction for quantitative PCR (qPCR) and oligonucleotide microarray analyses. The Compugen zebrafish oligonucleotide set was used for gene expression profiling and several genes were upregulated upon pathogen infection, including SOCS-4 and Flotillin. Flotillins are integral membrane proteins that are present in several subcellular components, including caveolae, lipid rafts, and the Golgi apparatus. SOCS proteins are known to negatively regulate the Janus kinase signal transducer and activator of transcription (JAK-STAT) signaling cascade. We have cloned both SOCS-4 and Flotillin and are currently conducting qPCR analysis.
Impacts
As the fisheries and aquaculture industries continue to expand in the state of Maine, disease control methods and diagnostic tests will be desperately needed. Through the studies outlined in this proposal, we hope to first determine the role of innate immunity, and then to explore ways to modulate this immune function to upregulate host response to infection. Although we propose to conduct these studies in zebrafish, we feel that much of the information gained through this model system can be transferred to other fish species, such as rainbow trout and Atlantic salmon, economically important fish of Maine.
Publications
- Millard, P.J., L.E. Bickerstaff, S.E. LaPatra, and C.H. Kim. 2006. Detection of infectious hematopoietic necrosis virus and infectious salmon anemia virus by molecular padlock amplification. J. Fish. Dis. 29:201-213.
- Lage, C.R., A. Nayak, and C.H. Kim. 2006. Arsenic ecotoxicology and innate immunity. Integr. Comp. Biol. Doi:10.1093/icb/icl048. Published on line, print publication pending.
- Sullivan, C., J.H. Postlethwait, C.R. Lage, P.J. Millard, and C.H. Kim. 2006. Evidence for Evolving TICAM Function in Vertebrates. J. Immunol. In Press.
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