**Recommended for Funding Spring 2021**

Metagenomic and community-level sequencing efforts have revealed an astonishing diversity of viruses associated with grossly normal marine invertebrates. The vast majority of detected viruses likely represent asymptomatic infections under typical conditions, but may generate pathology in hosts under changing environmental conditions. Recent work concluded that sea star wasting (SSW) was a sequela of suboxic conditions that set up within boundary layers that overlie respiratory tissues, possibly fueled be heterotrophic remineralization of phytoplankton- and macroalgal-derived dissolved organic matter. This condition was accompanied by the proliferation of viruses, which was hypothesized to be due to enhanced replication as a consequence of oxygen stress. Hence, there is significant need to understand cues which trigger the switch between non-pathogenic and pathogenic viral infection and subsequent impacts on host biology and ecology. Sea cucumbers (Holothuroidea; Echinodermata) are ecologically important constituents of marine benthic habitats. Recent reports of a wasting-like condition concomitant with SSW suggest that similar factors may be at play. At the same time, we recently discovered a representative of the insect-only Flaviviruses, a deeply branched clade of enveloped positive sense single-stranded RNA viruses that includes important mammalian pathogens, in wasting holothurians. This proposal will investigate the ecology of this newly-discovered group in Parastichopus californicus by addressing three hypotheses: H1) Aquatic insect-only Flaviviruses (aiFVs) do not cause gross pathology under typical conditions; H2) aiFVs proliferate and generate clinical and gross pathology under suboxic stress; and H3) Periodic increases in primary production and mean temperature excursions cause aiFV proliferation and subsequently exacerbate holothurian disease process. The proposed study will comprise a restricted survey of aiFV diversity via amplicon sequencing and their prevalence within and between populations, the development of an antibody-based approach for aiFV detection, and examination of aiFV behavior in concert with host transcription and veterinary pathology. The study will also examine the impacts of representative organic matter enrichment on aiFVs and their hosts, and follow seasonal progression of aiFV within populations through citizen science-driven efforts. Finally, we will examine the impacts of phytoplankton productivity on aiFV dynamics in Apostichopus californicus.

The proposed work will provide transformative and mechanistic knowledge on the role of environmental conditions in marine metazoan viral prevalence, replication and disease process. The proposed work does not represent a wide survey of correlative genomes/genes with environmental conditions, but rather will focus on key, specific questions about the interactions between host gene expression and transcription factors, viral replication, and biological oceanographic conditions. We will employ a time series survey to ground truth experimental manipulations of oxygen, temperature, organic matter and simulated phytoplankton bloom collapse, and thereby directly tie together experimental parameters, host and viral genomic elements, and field conditions. Since enveloped +ssRNA viruses are not frequently encountered during surveys of host-associated viruses, our focused suite of observations will also provide transformative information about this enigmatic group of viruses.