Comparative Transcriptomic Analysis Of Common Carp Spleen In Response To Aeromonas Hydrophila Infection & Genome Wide Characterization Of Scavenger Receptors Class A In Common Carp

The common carp is one of the most important economic fishery in China, also, it is the most important freshwater aquaculture and aquarium fish in the world, which is cultured in more than 100 countries,accounting for 10% of freshwater aquaculture production. Aeromonas hydrophila is a Gram-negative bacterium and widely distributed in freshwater habitats such as pond, river and lakes and so on, it’s a pathogenic bacteria that can infect vertebrate such as human, reptile and fish and cause a series of sepsis. During the process of common carp breeding, the situation always happens that the A. hydrophila infect the common carp and induce the huge damage for the production. In order to better understand the common carp immune response process against bacteria at the transcriptomic level, we examined transcriptional profiles of the common carp spleen at three time point following experimental infection with A. hydrophila, as a result, a total of 545 million reads were obtained and each library generated more than 23 million reads. Afterremoving ambiguous nucleotides, low-quality sequences(Q<20) and short reads(length<20 bp), there are 536 million high-quality reads(98.3%) left for further analysis. A total of 2900 differentially expressed genes were identified in three different stage(4 h, 12 h, and 24 h post infection) by using the tools of Cufflinks and Tophat. There are 739up-regulated genes and 248 down-regulated genes differentially expressed at 4 h post infection compared to control, 953 up-regulated genes and 481 down-regulated genes differentially expressed at 12 h post infection compared to control, and 941 up-regulated genes and 704down-regulated genes differentially expressed at 24 h post infection comparing to control, while 243 up-regulated genes and 18 down-regulate genes were differentially expressed in all three tested time points. We randomly selected 23 genes for q RT-PCR confirmation of differential expression, and the result showed that all 23 randomly selected genes were in agreement with the results of the comparative transcriptome analysis with only slight differences in expression levels, indicating that there was no consistent bias in the expression patterns. We attempted to categorize the differentially expressed genes by using gene ontology enrichment, the result showed that all differentially expressed genes were classified into 29 differentially category, which including verity of categories that related with immune process such as immune response,response to external stimulus, oxidoreductase activity, signal transduceractivity, cofactor binding, response to stress. We also highlight several important genes that may relate to the immune response according to gene annotation and pathway analysis, and we suppose these genes most probably function on the immune response after the A. hydrophila infection in common carp.In order to systematically analyze the immune related genes, we chose the scavenger receptors class A for the deep study, which is the important pattern recognition receptors(PRRs). Phylogenetic and syntenic analysis, expression patterns in early developmental stages,healthy tissues and after A. hydrophila infection were conducted in our study. We identified 7 SCARA genes according to the genome resource,which including SCARA2-1, SCARA2-2, SCARA3-1, SCARA3-2,SACAR4-1, SCARA4-2 and SCARA5. The phylogenetic and syntenic analysis reveal gene duplication and lose may happened in SCARAs evolution. We also found SCARAs were widely distribute in early developmental stages, while duplicated SCARA members had a similar expression pattern at different developmental stages. SCARA genes were also observed widely expressed in all examined common carp healthy tissues, but exhibited unique tissue-specific expression pattern. We further determined the expression patterns of SCARA genes after bacteria A. hydrophila infection, spleen and intestine were selected as candidate tissues for investigate the temporal expression profile, as we all know, thespleen and intestine were the important immune tissue and mucosal tissue.Our results found that the SCARAs showed immediately up-regulated in4 hours in intestine, while the situation occurred in 12 hours in spleen, we inferred that the SCARA genes in mucosal tissue might mainly be involved in pathogen recognition, while in spleen they mainly functioned on pathogen clearence. We hope that the study would contribute to a better understanding in the immune response,and provide us with further molecular guidance in breeding process.