Abstract
Marine mammals are top predators that are essential for stabilizing coastal marine community, structure, and function.4 However, incidents of unusual mortality events (UMEs) result in significant die offs that may negatively impact community structure. Since the 1990s NOAA has continuously documented yearly UMEs of marine mammals with cetaceans making up 55% percent of deaths.5 Surprisingly, about 50% of UMEs are yet to be determined and are not further researched.2 For example, NOAA conducted brain necropsies of the 1999 and 2000 UMEs of grey whales, and suggested the etiology to be "an unknown virus," but no further efforts were made to evaluate viral types.3 This lack of understanding of the pathogens responsible for significant mortality events in our seas is not limited to this example; little effort has been made to thoroughly examine the basis of stranding mortalities in cetaceans nationwide.1,10 Even in the few studies that have assessed cetacean and pinniped stranding mortalities, up to 20% still had an unknown cause of death.1,7 Importantly, research efforts constantly find that marine mammals are susceptible to neurological viral infections, such as Phocine Distemper Virus and Morbillivirus;2,7 therefore we hypothesis that undetected and/or emerging neurotropic viruses contribute to cetacean stranding events.
Currently, the discovery of novel viruses using standard techniques (e.g., marker genes degenerate PCR, microarrays, or microscopy) requires some prior knowledge of viral genome sequence or capsid morphology. To circumvent from these limitations we used a high throughput shotgun sequencing approach.8,9 In order to evaluate possible unknown or undetected viruses, we used a male harbor porpoise (Phocoena phocoena) found stranded along the Oregon Coast in June of 2011. The animal was collected by the Marine Mammal Institute in Oregon and was diagnosed by Oregon State University (OSU) veterinarians with encephalitis of possible viral origin. A subsample of brain tissue was used for RNA extraction and sent to OSU's Center for Genome Research and Bicomputing for cDNA library construction and then metagenomic sequencing on the Illumina HiSeq 2000 platform. Using a whole RNA shotgun sequencing approach we discovered, characterized, and annotated novel retroviral reads and Phycodnaviridae DNA viral transcripts (signifying an active viral infection) in Phocoena phocoena. The presence of these viruses confirms that marine mammal stranding events need to be monitored since these viruses may lead to future epidemics in marine mammals.
Acknowledgements
The authors would like to thank Jim Rice the coordinator of Oregon State University Marine Mammal Institute, Dr. Robert Bildfell at Oregon State University College of Veterinary Medicine. Mark Dasenko at Oregon State University Center for Genome Research and Biocomputing, Dr. Jesse Zaneveld at Oregon State University Department of Microbiology. The authors thank Oregon Sea Grant and the Oregon State University Diversity Advancement Pipeline Graduate fellowship for their financial support of this project.
* Presenting author
Literature Cited
1. Bogomolni A. 2010. Mortality trends of stranded marine mammals on Cape Cod and southeastern Massachusetts, USA, 2000 to 2006. Diseases of Aquatic Organisms 88: 143–155.
2. Goldstein T, Mazet J, Gill V.A, Doroff A, et al. 2009. Phocine Distemper Virus in Northern Sea Otters in the Pacific Ocean Alaska, USA. Emerging Infectious Diseases 15: 925–927.
3. Gulland F, Pérez-Cortés H, Urbán JR, Rojas-Bracho J, et al. 2005. Eastern North Pacific Gray Whale (Eschrichtius robustus) Unusual Mortality Event, 1999–2000. NOAA Technical Memorandum NMFS-AFSC-150.
4. Heiathus M, Frid A, Wirsing E, and Worm B. 2008. Predicting ecological consequences of marine top predator declines. Trends in Ecology and Evolution 23: 202–210.
5. NOAA Fisheries Office of Protected Resources. August 22, 2012. NOAA. www.nmfs.noaa.gov/pr/health/mmume/.
6. Reidarson T, McBain J, House C, King D, Stott J, et al. 1998. Morbillivirus Infection in Stranded Common Dolphins from the Pacific Ocean. Journal of Wildlife Diseases 34:771–776.
7. Stroud R, Roffe J. 1979. Causes of death in marine mammals stranded along the Oregon coast. Wildlife Diseases 15: 91–97.
8. Vega Thurber R, Hayne M, Breitbart M, Wegle L, et al. 2009. Laboratory procedures to generate viral metagenomes. Nature Protocols 4: 470–483.
9. Vega Thurber R. 2011. Methods in Viral Metagenomics. In: Bruijn FJ, editors. Handbook of Molecular Microbial Ecology II: Metagenomics in Different Habitats. Hoboken (NJ): John Wiley & Sons, Inc. p 15–24.
10. Zagzebski A, Gulland F, Haulena M, Lander, et al. 2006. Twenty-five years of rehabilitation of odontocetes stranded in central and northern California, 1977 to 2002. Aquatic Mammals 32: 334–345.