Abstract
Acute hepatopancreatic necrosis disease (AHPND), caused by marine bacteria Vibrio Parahaemolyticus, is a huge problem in shrimp farms. The V. parahaemolyticus infecting material is contained in a plasmid which encodes for the lethal toxins PirABVp, whose primary target tissue is the hepatopancreas, causing sloughing of epithelial cells, necrosis, and massive hemocyte infiltration. To get a better understanding of the hepatopancreas response during AHPND, juvenile shrimp Litopenaeus vannamei were infected by immersion with V. parahaemolyticus. We performed transcriptomic mRNA sequencing of infected shrimp hepatopancreas, at 24 hours post-infection, to identify novel differentially expressed genes a total of 174,098 transcripts were examined of which 915 transcripts were found differentially expressed after comparative transcriptomic analysis: 442 up-regulated and 473 down-regulated transcripts. Gene Ontology term enrichment analysis for up-regulated transcripts includes metabolic process, regulation of programmed cell death, carbohydrate metabolic process, and biological adhesion, whereas for down-regulated transcripts include, microtubule-based process, cell activation, and chitin metabolic process. The analysis of protein- protein network between up and down-regulated genes indicates that the first gene interactions are connected to oxidation-processes and sarcomere organization. Additionally, protein-protein networks analysis identified 20-top highly connected hub nodes. Based on their immunological or metabolic function, ten candidate transcripts were selected to measure their mRNA relative expression levels in AHPND infected shrimp hepatopancreas by RT-qPCR. Our results indicate a close connection between the immune and metabolism systems during AHPND infection. Our RNA-Seq and RT-qPCR data provide the possible immunological and physiological scenario as well as the molecular pathways that take place in the shrimp hepatopancreas in response to an infectious disease.
Highlights
Aquaculture of Litopenaeus vannamei along with Penaeus monodon, is one of the most important cultivated shrimp species with a significant economic value that generates billions of USD every year
To confirm that we were using the correct bacterial strain for our infection experiments, an end-point PCR was performed to screen for the PHP and PirB gene sequences from two different V. parahaemolyticus strains: the IPNGS16 strain which tested positive for the two gene sequences and the CAIM170, a non-Acute hepatopancreatic necrosis disease (AHPND) causing strain which is negative for PHP and PirB sequences (S1 Fig)
Ge et al (2017) which used the ridgetail white prawn (E. carinicauda) and V. parahaemolyticus to challenge the specimens by intramuscular injection found up- regulation of the hypoxia upregulated gene (HCY) and chymotrypsin (CTR), and down- regulation of the Cathepsin L protein (CTSH) gene [15], these data agree with our results, we found that these three genes have the same gene expression dynamics after bacteria challenge
Summary
Aquaculture of Litopenaeus vannamei (whiteleg shrimp) along with Penaeus monodon, is one of the most important cultivated shrimp species with a significant economic value that generates billions of USD every year. Despite the success of shrimp production worldwide, this industry has had several setbacks due to viral and bacterial diseases which have caused until 2014, 118 million USD in losses [2,3]. The etiological agent causing AHPND are bacteria of the genus Vibrio, being V. parahaemolyticus the primary infection vector. This bacteria species contains a plasmid that encodes for the lethal toxins PirABVp that cause sloughing of hepatopancreas epithelial tubule cells (F, R, and B cells) in early infection stages, followed by necrosis, atrophy, and massive hemocytes infiltration [5,6,7,8]
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