Innate Immune Responses and Permissiveness to Ranavirus Infection of Peritoneal Leukocytes in the FrogXenopus laevis

Abstract

Ranaviruses such as frog virus 3 ([FV3]family Iridoviridae) are increasingly prevalent pathogens that infect reptiles, amphibians, and fish worldwide. Whereas studies in the frog Xenopus laevis have revealed the critical involvement of CD8 T-cell and antibody responses in host resistance to FV3, little is known about the role played by innate immunity to infection with this virus. We have investigated the occurrence, composition, activation status, and permissiveness to infection of peritoneal leukocytes (PLs) in Xenopus adults during FV3 infection by microscopy, flow cytometry, and reverse transcription-PCR. The total number of PLs and the relative fraction of activated mononucleated macrophage-like cells significantly increase as early as 1 day postinfection (dpi), followed by NK cells at 3 dpi, before the peak of the T-cell response at 6 dpi. FV3 infection also induces a rapid upregulation of proinflammatory genes including arginase 1, interleukin-1␤, and tumor necrosis factor alpha. Although PLs are susceptible to FV3 infection, as evidenced by apoptotic cells, active FV3 transcription, and the detection of viral particles by electron microscopy, the infection is weaker (fewer infectious particles), more transitory, and involves a smaller fraction (less than 1%) of PLs than the kidney, the main site of infection. However, viral DNA remains detectable in PLs for at least 3 weeks postinfection, past the point of viral clearance observed in the kidneys. This suggests that although PLs are actively involved in anti-FV3 immune responses, some of these cells can be permissive and harbor quiescent, asymptomatic FV3.

Reliability of non-lethal surveillance methods for detecting ranavirus infection

“…In Xenopus laevis, the liver often becomes infected after other organs (Robert et al 2005). It also is possible that qPCR detected quiescent ranavirus in circulating macrophages (Morales et al 2010 The trend of decreasing ranavirus prevalence over time suggests that bullfrog tadpoles have the capability of clearing this pathogen, which is the first direct evidence for this species. Clearing of ranavirus infections has been reported in Xenopus laevis (Gantress et al 2003).…”

“…Inasmuch as target organs may differ for different types of virus and host species, our results may be limited to American bullfrog tadpoles and the isolates used in our study. Also, a positive qPCR result indicated that ranavirus DNA was present (Green et al 2010); however, distinction be tween active and quiescent infections (Morales et al 2010) could not be made. Other techniques, e.g.…”

Ranavirus Ecology and Evolution: From Epidemiology to Extinction

“…Additionally, Robert et al ( 2007 ) demonstrated that X. laevis , which is generally resistant to FV3 infections, can be asymptomatic carriers. In a subsequent study, Morales et al ( 2010 ) showed that peritoneal macrophages sometimes harbor quiescent FV3 infections for at least 3 weeks. Asymptomatic infections can be reactivated in animals that are immunocompromised by γ-irradiation (Robert et al 2007 ).…”

Characterization of Chinese giant salamander iridovirus tissue tropism and inflammatory response after infection

“…TNF-α and IL-1β are common proinflammatory cytokines released from cells prior to the in flammation. Increased ex pression of TNF-α and IL-1β were reported in FV3-infected Xenopus and EPC cells and EHNV-infected EPC cells (Morales et al 2010, Holopainen et al 2012). In our study, GSIV infection triggered rapid and high expression of MIF, TNF-α and IL-1β in the spleen at 3 dpi, which may subsequently influence viral replication.…”

“…Recent ly, the Chinese giant salamander MIF gene sequence was identified, and its encoded protein was shown to have enzymatic redox and tautomerase activity, and was also capable of inhibiting the migration of macrophages (Wang et al 2013). Rana virus infection can up-regulate pro-inflammatory cytokines such as TNF-α and interleukin-1β (IL-1β) in fish and frogs (Morales et al 2010, Holopainen et al 2012, but the inflammatory responses to rana virus infection have not been characterized in amphi bian hosts, such as the Chinese giant salamander.…”