Effect of water temperature on the immune response of American lobster (Homarus americanus) experimentally infected with white spot syndrome virus

Water temperature influences basic life history traits of the American lobster (Homarus americanus) such as survival, growth and reproduction. Yet, relatively little is known about the effects of water temperature on the immune response of H. americanus. White spot syndrome virus (WSSV) is currently one of the largest impediments to the shrimp aquaculture industry worldwide. The World Organization for Animal Health (Office International de Epizootic; OIE) lists WSSV as a notifiable disease with the potential to infect all crustacean decapods. This project investigates the constraints imposed by a range of temperatures (10 oC, 15 oC, 17.5 oC, 20 oC) on the clinical, tissue and molecular immune responses of H. americanus experimentally infected with WSSV. Haemocyte concentration and qPCR testing of haemolymph were used to monitor the host’s clinical response over time and to determine presence of viral infection, respectively. A decrease in clinical condition of H. americanus was observed at 17.5 oC and 20 oC but not at 10 oC and 15 oC. WSSV-qPCR revealed viral amplification was associated with higher ambient temperature. A combination of light and electron microscopy, as well as qPCR testing of various tissues was used to elucidate changes within infected H. americanus at the various temperatures. Of the 11 host tissues examined, hypertrophied nuclei (indicative of WSSV infection) were most easily identifiable within the antennal gland and intestine of infected animals at warmer temperatures (17.5 oC and 20 oC). At colder temperatures (10 oC) no WSSV associated histopathological changes were identified in any of the examined tissues. Electron microscopy was used to explore the general ultrastructure of the antennal gland and the confirmed presence of WSSV rod-shaped and enveloped virions. In infected animals the two main regions of the antennal gland, the coelomosac and labyrinth, contained randomly distributed infected nuclei with marginated host chromatin and WSSV virions in various stages of replication. In marine decapods, the antennal gland is involved in osmoregulation and excretion suggesting that the antennal gland may be a preferred site for WSSV viral replication and inclusion within the excretory product. Lastly, a lobster specific microarray was used to monitor transcriptomic changes across 14,592 genes during viral infection at the different temperatures. Using one-way ANOVA analyses, a total of 717 genes were identified as significantly differentially expressed between infected and control H. americanus at the various temperatures. Differentially expressed genes included, ribosomal proteins (L27a, L13, L11, and L39), mitogen-activated protein kinase organizer 1, prolyl-4-hydroxylase-alpha, laminin subunit gamma-3, short-chain dehydrogenase and acute phase serum amyloid A. Microarray results were verified using RT-qPCR. Clinical, tissue and molecular immune results from this study provide a thorough assessment of the impact of temperature and WSSV in H. americanus. The use of a range of approaches and experimental temperatures is critical for broadening our understanding of how temperature influences host pathogen interactions in the economically important American lobster H. americanus