The effect of rearing density on immune responses of hepatopancreas and intestine in Litopenaeus vananmei against Vibrio paraheamolyticus E1 challenge.

Rearing density and disease management are considered as pivotal factors determining shrimp farm productivity and profitability. To systematically investigate the potential mechanisms for density-related differences between disease susceptibility and rearing densities, we conducted comparative transcriptome analysis of the molecular differences between hepatopancreas and intestine of Litopenaeus vannamei under two different rearing densities (800- and 400- shrimp/m3) for 15 d and further analyzed the differences in immune response to Vibrio parahaemolyticus E1 (VPE1) raised under two density conditions. Totally 45 different expression genes (DEGs) were identified in the hepatopancreas under two different rearing densities, the DEGs were grouped into four processes or pathways related to animal immune system. Then, exposure to the VPE1 resulted in 639 DEGs, involved into fourteen immune related processes or pathways. In the intestine, seventeen processes or pathways related to the immune system were identified among the 5470 DEGs under two different rearing densities. 279 DEGs were identified post VPE1 challenge, classified into five processes or pathways associated with the immune system. Meanwhile, the results of growth performance, histopathology and the activities of antioxidant enzymes in the hepatopancreas and intestines of shrimp showed that high density decreased weight gain rate (63.20 ± 1.67% and 18.73 ± 3.35% in the high and low rearing density groups, respectively), severely destroyed the histopathology and inhibited the antioxidant enzymes activities. This study demonstrated that rearing density in L. vannamei significantly impacts susceptibility to the VPE1, via altered transcriptional challenge responses, and thus higher mortality due to disease.

Wang Y ，Wang B ，Shao X ，Shao J ，Liu M ，Wang M ，Wang L ... -  《-》

Response of the Litopenaeus vananmei intestinal bacteria and antioxidant system to rearing density and exposure to Vibrio paraheamolyticus E1.

Intestinal bacteria and bacterial metabolic products are indispensable components of both invertebrate and vertebrate physiology, directly influencing many functions including host energy absorption and metabolism, intestinal barrier integrity and immune function. To investigate the influence of rearing density on shrimp intestinal health, antioxidant responses and disease susceptibility, we simultaneously monitored the dynamic changes of intestinal bacteria and antioxidant enzymes activities in Litopenaeus vananmei under two different rearing densities (400 and 800 shrimp/m3) and further investigated the difference in response to Vibrio paraheamolyticus E1 (VPE1) challenge. We showed that the phyla Proteobacteria, Bacteroidetes, Planctomycetes and Firmicutes were the predominant microflora in all treatment groups. Rearing L. vannamei at high density for 15 days resulted in the reduction of Bacteroidetes and Firmicutes and increase of Planctomycetes. At the genus level, high rearing density induced reduction of Pseudoalteromonas and Blastopirellula, and an increase of Photobacterium and Vibrio. Notably, the relative abundance of Ascidiaceihabitans and Flavobacteria NS10_marine_group increased in the low rearing density groups after VPE1 challenge, suggesting that these two types of bacteria might have an important role in resisting to VPE1 infection. High density stress caused suppression of total superoxide dismutase (T-SOD), catalase (CAT), and glutathione peroxidase (GPX) activities in shrimp. Hence, high density stress altered the functional composition of shrimp intestinal bacteria and damaged the antioxidant system, which increased pathogen susceptibility.

Wang Y ，Liu M ，Wang B ，Jiang K ，Wang M ，Wang L ... -  《-》

Transcriptomic and morphological analyses of Litopenaeus vannamei intestinal barrier in response to Vibrio paraheamolyticus infection reveals immune response signatures and structural disruption.

The white shrimp Litopenaeus vannamei has been greatly impacted by Vibrio infection. In this study, we investigated the intestinal barrier response of L vannamei following challenge with Vibrio parahaemolyticus E1, by examining morphological changes and transcriptome expression levels. A total of 16,4420 unigenes were obtained from RNAseq data after quality control and assembly, and 4646 differentially expressed genes (DEGs) were identified following Vibrio challenge, of which 2469 unigenes were significantly up-regulated and 2177 were significantly down-regulated. DEGs were determined to be involved in various physical, chemical and immunological intestinal barrier functions, including peritrophin, cytoskeleton and cell junction, pattern recognition receptors, antimicrobial peptide and immune signaling pathways, serine protease/protease inhibitor and prophenoloxidase system, apoptosis and phagocytosis, and antioxidant systems. Fifteen DEGs were randomly selected for validation by real-time quantitative PCR (RT-qPCR) and showed results consistent with the RNA-seq data. Intestinal epithelial cell morphology was also affected by Vibrio challenge, showing epithelial detachment, nuclear pyknosis, and destruction of cell junctions. These results improve our current understanding of the intestinal barrier function in the shrimp response to bacterial infection.

Qi C ，Wang L ，Liu M ，Jiang K ，Wang M ，Zhao W ，Wang B ... -  《-》

Transcriptome analysis of Pacific white shrimp (Litopenaeus vannamei) challenged by Vibrio parahaemolyticus reveals unique immune-related genes.

Pacific white shrimp (Litopenaeus vannamei) is an important cultural species worldwide. However, Vibrio spp. infections have caused a great economic loss in Pacific white shrimp culture industry. The immune responses of Pacific white shrimp to the Vibrio spp. is not fully characterized. In this study, the transcriptomic profiles of L. vannamei hemocytes were explored by injecting with or without Vibrio parahaemolyticus. Totally, 42,632 high-quality unigenes were obtained from RNAseq data. Comparative genome analysis showed 2258 differentially expressed genes (DEGs) following the Vibrio challenge, including 1017 up-regulated and 1241 down-regulated genes. Eight DEGs were randomly selected for further validation by quantitative real-time RT-PCR (qRT-PCR) and the results showed that are consistent with the RNA-seq data. Due to the lack of predictable adaptive immunity, shrimps rely on an innate immune system to defend themselves against invading microbes by recognizing and clearing them through humoral and cellular immune responses. Here we focused our studies on the humoral immunity, five genes (SR, MNK, CTL3, GILT, and ALFP) were selected from the transcriptomic data, which were significantly up-regulated by V. parahaemolyticus infection. These genes were widely expressed in six different tissues and were up-regulated by both Gram negative bacteria (V. parahaemolyticus) and Gram positive bacteria (Staphylococcus aureus). To further extend our studies, we knock-down those five genes by dsRNA in L. vannamei and analyzed the functions of specific genes against V. parahaemolyticus and S. aureus by bacterial clearance analysis. We found that the ability of L. vannamei was significantly reduced in bacterial clearance when treated with those specific dsRNA. These results indicate that those five genes play essential roles in antibacterial immunity and have its specific functions against different types of pathogens. The obtained data will shed a new light on the immunity of L. vannamei and pave a new way for fighting against the bacterial infection in Pacific white shrimp.

Qin Z ，Babu VS ，Wan Q ，Zhou M ，Liang R ，Muhammad A ，Zhao L ，Li J ，Lan J ，Lin L ... -  《-》

Comparative transcriptome analysis reveals the different roles between hepatopancreas and intestine of Litopenaeus vannamei in immune response to aflatoxin B1 (AFB1) challenge.

Aflatoxin B1 (AFB1) is a mycotoxin mainly produced by Aspergillus flavus and Aspergillus parasiticus contaminating food, feed ingredients and products of animal origin. In mammals, this toxin causes widespread organ-specific damage; it is immunotoxicity and could promote hepatotoxicity, alter intestinal functions and so on. In this study, we conducted transcriptome and histomorphology analyses of hepatopancreas and intestinal in Litopenaeus vannamei (L. vannamei) challenged with AFB1. Totally 12,014 and 1387 differentially expression genes (DEGs) were identified in the hepatopancreas and intestine, respectively. In hepatopancreas, a total of 1995 DEGs were mainly annotated and grouped into 18 processes or pathways related to animal immune system. With respect to intestine, a total of 152 DEGs were mainly annotated to 7 processes or pathways related to animal immune system. Meanwhile, we determined the relative mRNA expression of several crucial representative immune genes including Toll, immune deficiency (IMD), prophenoloxidase (proPO), Rab and glutathione S-transferase (GST) in the hepatopancreas and intestines of shrimp at 3-, 6-, 12-, 18-, 24- and 30-d after challenged by AFB1. Exposure to AFB1 increased mortality, decrease weight gain rate, severely destroyed the histomorphology of hepatopancreas and intestine, and resulted in the damaged of immune system of shrimp. The present data reveals the different roles between hepatopancreas and intestine of L. vannamei in immune response to AFB1 challenge, and provides insight into the molecular basis of the relationship between hepatopancreas and intestinal immunity during either homeostasis or inflammation.

Wang Y ，Wang B ，Liu M ，Jiang K ，Wang M ，Wang L ... -  《COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY C-TOXICOLOGY & PHARMACOLOGY》