Parasitic nematode (PN) infections remain a major threat to human health worldwide, with more than 1 billion people infected. Children, pregnant women, and the elderly are particularly susceptible to morbidity from nematode infection. Control strategies are restricted to periodic de-worming of infected individuals, which is limited by rapid re-infection rates and the development of drug resistant worm populations. There are no vaccines available for PN infections in humans. Development of new drugs and vaccines will require a better understanding of PN biology, particularly the infective process and the host's immune response to infection. The requirement of an obligate host and the lack of good animal models have limited investigations into mechanisms by which PNs infect and usurp the host immune response. While the free living nematode Caenorhabditis elegans is an excellent model for nematode development, it is not a parasite. The insect parasitic nematode Heterorhabditis bacteriophora (Hb) offers potential as a tractable model of PN infection that has many of the advantages of C. elegans, including easy culture and manipulation of all life history stages. However, many of the powerful genetic tools, like transgenesis and RNA interference, have not been fully developed in Hb, restricting its use as a widespread model. We propose to develop these genetic tools in Hb with a focus on targeting the infective juvenile and early parasitic stages in order to investigate infection mechanisms. We will take two parallel but independent approaches to develop these tools that will provide a foundation for future investigation into the mechanism of PN infection.
In Aim 1, we propose to develop a transfection protocol for Hb via germ-line microinjection of a piggybac retrotransposon-based integrating vector to introduce the reporter transgenes into Hb chromosomes.
In Aim 2, we will develop a robust RNAi protocol for 2nd generation Hb hermaphrodites. By focusing on 2nd generation hermaphrodites we are ideally positioned to probe the biology of PN infection, as the progeny of this stage are the infective stage. Development of methods for gene knock down (reverse genetics) and transfection in this novel model nematode would represent a significant advance for PN research, and will allow investigation of PN gene function during infection for the first time.

Public Health Relevance

Parasitic nematodes pose an important health threat, infecting over 1 billion people. This project will develop the first tools for genetic manipulationof a novel model parasitic nematode Heterorhabditis bacteriophora. These tools will allow the effective use of this model for functional genetic investigations and a better understanding of parasitic nematode infections that will result in improved control strategies.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21AI109517-02
Application #
8849839
Study Section
Innate Immunity and Inflammation Study Section (III)
Program Officer
Wali, Tonu M
Project Start
2014-05-16
Project End
2016-04-30
Budget Start
2015-05-01
Budget End
2016-04-30
Support Year
2
Fiscal Year
2015
Total Cost
$234,554
Indirect Cost
$84,554
Name
George Washington University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
043990498
City
Washington
State
DC
Country
United States
Zip Code
20052
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Vadnal, Jonathan; Ratnappan, Ramesh; Keaney, Melissa et al. (2017) Identification of candidate infection genes from the model entomopathogenic nematode Heterorhabditis bacteriophora. BMC Genomics 18:8
Shokal, Upasana; Eleftherianos, Ioannis (2017) Evolution and Function of Thioester-Containing Proteins and the Complement System in the Innate Immune Response. Front Immunol 8:759
Cooper, Dustin; Eleftherianos, Ioannis (2016) Parasitic Nematode Immunomodulatory Strategies: Recent Advances and Perspectives. Pathogens 5:
Ratnappan, Ramesh; Vadnal, Jonathan; Keaney, Melissa et al. (2016) RNAi-mediated gene knockdown by microinjection in the model entomopathogenic nematode Heterorhabditis bacteriophora. Parasit Vectors 9:160
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Yadav, Shruti; Shokal, Upasana; Forst, Steven et al. (2015) An improved method for generating axenic entomopathogenic nematodes. BMC Res Notes 8:461