extraction-hemocytes-from-drosophila-melanogaster-larvae-for

Extraction of Hemocytes from <em>Drosophila melanogaster</em> Larvae for Microbial Infection and Analysis

During the pathogenic infection of Drosophila melanogaster, hemocytes play an important role in the immune response throughout the infection. Thus, the ...

Washington State University

Insects are not only major vectors of mammalian viruses, but are also host to insect-restricted viruses that can potentially be transmitted to mammals. While mammalian innate immune responses to arboviruses are well studied, less is known about how mammalian cells respond to viruses that are restricted to infect only invertebrates. Here we demonstrate that IIV-6, a DNA virus of the family Iridoviridae, is able to induce a type I interferon-dependent antiviral immune response in mammalian cells. Although IIV-6 is a DNA virus, we demonstrate that the immune response activated during IIV-6 infection is mediated by the RIG-I-like receptor (RLR) pathway, and not the canonical DNA sensing pathway via cGAS/STING. We further show that RNA polymerase III is required for maximal IFN-β secretion, suggesting that viral DNA is transcribed by this enzyme into an RNA species capable of activating the RLR pathway. Finally, we demonstrate that the RLR-driven mammalian innate immune response to IIV-6 is functionally capable of protecting cells from subsequent infection with the arboviruses Vesicular Stomatitis virus and Kunjin virus. These results represent a novel example of an invertebrate DNA virus activating a canonically RNA sensing pathway in the mammalian innate immune response, which reduces viral load of ensuing arboviral infection.

Invertebrate Iridescent Virus 6, a DNA Virus, Stimulates a Mammalian Innate Immune Response through RIG-I-Like Receptors.

 is the causative agent of Q fever, a zoonotic disease that threatens both human and animal health. Due to the paucity of experimental animal models, little is known about how host factors interface with bacterial components and affect pathogenesis. Here, we used , in conjunction with the biosafety level 2 (BSL2) Nine Mile phase II (NMII) clone 4 strain of , as a model to investigate host and bacterial components implicated in infection. We demonstrate that adult  flies are susceptible to  NMII infection and that this bacterial strain, which activates the immune deficiency (IMD) pathway, is able to replicate and cause mortality in the animals. We show that in the absence of Eiger, the only known tumor necrosis factor (TNF) superfamily homolog in , -infected flies exhibit reduced mortality from infection. We also demonstrate that the  type 4 secretion system (T4SS) is critical for the formation of the -containing vacuole and establishment of infection in  Altogether, our data reveal that the  TNF homolog Eiger and the  T4SS are implicated in the pathogenesis of  in flies. The /NMII model mimics relevant aspects of the infection in mammals, such as a critical role of host TNF and the bacterial T4SS in pathogenesis. Our work also demonstrates the usefulness of this BSL2 model to investigate both host and  components implicated in infection.

Aoi Hiroyasu

School of Molecular Biosciences,
College of Veterinary Medicine,
School of Molecular Biosciences, College of Veterinary Medicine

Washington State University

Extraction of Hemocytes from Drosophila melanogaster Larvae for Microbial Infection and Analysis

Aoi Hiroyasu

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Washington State University

Extraction of Hemocytes from Drosophila melanogaster Larvae for Microbial Infection and Analysis

School of Molecular Biosciences,
College of Veterinary Medicine,
School of Molecular Biosciences, College of Veterinary Medicine