Discovery of novel molecular immune mediators in the American lobster (Homarus americanus) during bacterial, eukaryotic parasitic and viral challenges

Abstract

The American lobster (Homarus americanus) is the most economically significant commercial fishery species in Canada. The Canadian lobster fishery provides tens of thousands of jobs, and is the economic driver of hundreds of rural communities in Atlantic Canada and Quebec. The adult lobster is susceptible to relatively few pathogens but very little is known about how its immune system is capable of mediating this pathogen resistance. Additionally, there is no definitive clinical biomarker that is capable of assessing overall lobster health, something that would be valuable to a fishery where lobsters are the most lucrative when they are sold as a live product. The purpose of this project was to discover how the H. americanus humoral immune system responds to bacterial, eukaryotic parasitic and viral pathogens. This was performed by using high-throughput transcriptomics, in the form of microarray gene-expression analysis, to monitor the expression of classical immune molecules, as well as discover novel immune mediators. This project discovered hundreds of new H. americanus genes that have not previously been associated with either lobster or crustacean immunity. The expression of these novel immune-related genes is especially interesting because over half of them have no similarity to proteins in GenBank, little if any functional characterization, and pathogen class-specific expression. The conventional immune paradigm for any crustacean innate immune response is that it is non-specific and responds similarly to all microorganisms, pathogenic or not. These studies have determined that this is not the case. There is consistent and unique differential expression of genes for each of the pathogen classes examined, and even differential expression of isoforms within gene families that is pathogen dependent. Among the most interesting genes that have been discovered are the six isoforms of anti-lipopolysaccharide factor family (ALFHa-1, ALFHa-2, ALFHa-3, ALFHa-4, ALFHa-6 and ALFHa-7), acute phase serum amyloid protein A (SAA) and trypsin 1b. Four ALFHa family isoforms are differentially expressed during bacterial, Aerococcus viridans var. homari, and parasitic, Anophryoides haemophila, infections, where the differential expression of ALFHa-4 and ALFHa-2 isoforms are the most significant during bacterial and parasitic infections respectively. However, none of the six ALFHa genes are differentially expressed during viral, White Spot Syndrome Virus, infection. SAA is important because its expression is significantly increased in moribund lobsters during bacterial and parasitic infections, although not in viral infections. The protein sequence of SAA is highly conserved between humans and H. americanus suggesting the conservation of an important biological function such as innate immune activation. Trypsin 1b is the only gene that was differentially expressed during all three pathogen challenges. It is unclear what immunological role this gene plays in lobster or crustacean immunity but it is likely pivotal to immune activation, and a promising lobster health biomarker. These findings have made significant advances to our understanding of lobster immunity which can be applied to crustacean immunology as a whole. The lobster immune response can no longer be thought of a generalized non-specific response, but as one tailored to the invading pathogen