De novo assembly of Sockeye salmon kidney transcriptomes reveal a limited early response to piscine reovirus with or without infectious hematopoietic necrosis virus superinfection

Background Piscine reovirus (PRV) has been associated with the serious disease known as Heart and Skeletal Muscle Inflammation (HSMI) in cultured Atlantic salmon Salmo salar in Norway. PRV is also prevalent in wild and farmed salmon without overt disease manifestations, suggesting multifactorial triggers or PRV variant-specific factors are required to initiate disease. In this study, we explore the head kidney transcriptome of Sockeye salmon Oncorhynchus nerka during early PRV infection to identify host responses in the absence of disease in hopes of elucidating mechanisms by which PRV may directly alter host functions and contribute to the development of a disease state. We further investigate the role of PRV as a coinfecting agent following superinfection with infectious hematopoietic necrosis virus (IHNV) – a highly pathogenic rhabdovirus endemic to the west coast of North America. Results Challenge of Sockeye salmon with PRV resulted in high quantities of viral transcripts to become present in the blood and kidney of infected fish without manifestations of disease. De novo transcriptome assembly of over 2.3 billion paired RNA-seq reads from the head kidneys of 36 fish identified more than 320,000 putative unigenes, of which less than 20 were suggested to be differentially expressed in response to PRV at either 2 or 3 weeks post challenge by DESeq2 and edgeR analysis. Of these, only one, Ependymin, was confirmed to be differentially expressed by qPCR in an expanded sample set. In contrast, IHNV induced substantial transcriptional changes (differential expression of > 20,000 unigenes) which included transcripts involved in antiviral and inflammatory response pathways. Prior infection with PRV had no significant effect on host responses to superinfecting IHNV, nor did host responses initiated by IHNV exposure influence increasing PRV loads. Conclusions PRV does not substantially alter the head kidney transcriptome of Sockeye salmon during early (2 to 3 week) infection and dissemination in a period of significant increasing viral load, nor does the presence of PRV change the host transcriptional response to an IHNV superinfection. Further, concurrent infections of PRV and IHNV do not appear to significantly influence the infectivity or severity of IHNV associated disease, or conversely, PRV load. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-3196-y) contains supplementary material, which is available to authorized users.


265
Mid kidney, with tubuli and glomeruli: Sparse -moderate amount of MMF and melaninfoci, like 216. Multifocally mild circulatory disturbances (blood interstitially and in vessels). Many tubuli with eosinphilic material in lumen, a few with small basophilic grains. Some tubuli with enlarged and/or non-similar epithelial nuclei/nuclei in different levels. Some PMN leucocytes interstitially.
One small focus of red muscle degenerative changes as in 211 and 213. One small focus with possible inflammation and lack of red muscle fibers close to epidermis.

195
Mid kidney, small parts of head kidney. Few MMF/melanin foci. Small amounts of eosinophilic material in lumen of some tubuli.
No tissue found.
Small area of large clear vacuoles in red muscle fibers. Degenerative changes like 211 and 213 in red muscle.

197
Head kidney with a few tubuli and a few foci of endocrine tissue. Very few MMF/few foci of melanin.
Epidermis and dermis is lacking (prob. sampling artefact). Focally possible exudation/oedema between some fibers of both red and white muscle. Mild degeneration, exudation and hypercellularity in red muscle.

272
Mid kidney. Few MMF/melanin foci. Small amounts of eosinophilic material in lumen of some tubuli.
Focus of red muscle with degenerative changes like in 211 and 213.

203
Head kidney, with some tubuli in 1/5 of the tissue. One foci of endocrine tissue. Few MMF/foci of melanin.
Mild degeneration in mid line red muscle (as in 199, 935 and others).
Small amounts of red muscle. One small focus with loss of structure (mild form of degeneration).  Very small amount of red muscle, midline red muscle and large parts of epidermis/dermis is lacking (prob. artefact). Multifocally possible exudation/oedema between fibers of white muscle.
Mid line red muscle with degenerative changes like in 211 and 213.
Very small sample, no red muscle for evaluation, NPLF.
Moderately large focus with some degenerative red muscle changes.

285
Mid kidney. Interstitially multifocally few MMF/melanin (distribution as in 269 And 283). Some tubuli with eosinophilic material in lumen, multifocally some with eosinophilic granula in tubuliepithelial cells. Focally mild circ. disturbances and a few PMN-leucoccytes in the interstitium. Few tubuli with nuclei like 265 (diff. levels etc).
Small sample, small part only with epidermis, dermis and red muscle: A few red muscle fibers with large, clear vacuoles.
Some minor vacuolization of a few red m. fibres. Focally degenerative red muscle changes similar to 211 and 213.

288
Mid kidney. Interstitially few -some MMF/melanindeposits (distribution as in 269 and 283 + others) mild circ. disturbances, some -many PMNleucocytes interstitially. Some tubuli with eosinophilic material in lumen. Few tubuli with nuclei like 265 (diff. levels etc). Relatively high proportion of blood cells in vessels consists of leucocytes, could be post-mortem artefact.

214
Head kidney, one focus of endocrine tissue. Blood in larger vessels and easily observed/"standing out " sinusoids: Possible mild circulatory disturbances. A few MMF/melaninfoci. One small inflammatory focus and a vessel with sparse perivascular inflammatory cell infiltration and sparse oedema in white muscle. One small area of possible degeneration and inflammation of red muscle, degenerative changes similar to changes in 211 and 213.

216
Head kidney, small piece: A few tubuli, a few MMF/melanin foci. Some areas w. sp.-mod. MMF/melanin foci intersititially, between tubuli. Small amounts of eosinophilic material in tubuli, probably epithelial debris. Relatively high proportion of PMN leucocytes interstitially (personal impression). Small pieces, a lot of possibilities for «edge artefacts», difficult to interpret.
A minor focus of inflammation in white muscle. Focal degenerative changes of red muscle red muscle, like in 211 and 213.