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For commercial aquatic animals, hypoxia phenomenon often occurs in live transport and aquaculture. In previous studies, much interest has been focused on antioxidant enzyme activities and could not present the complexities. The multifaceted responses, especially considering physiological indexes, histological structure, cell apoptosis, and immune pathways, are still unknown. In this study, we investigated the comprehensive hypoxic responses of Marsupenaeus japonicus. The results showed that the physiological indexes showed time-dependent changes upon hypoxia stress. Hypoxia stress led to significant tissue damage and cell apoptosis in the gill and hepatopancreas. Compared with the control group, the apoptosis index (AI) of the 12 h hypoxic treatment increased significantly (p < 0.05) in the gills and hepatopancreas. Comparative transcriptome analysis identified 900 and 1400 differentially expressed genes (DEGs) in the gill and hepatopancreas, respectively. Several DEGs were related to the lysosome, glycolysis/gluconeogenesis, citrate cycle, and apoptosis, and seven of them were validated using quantitative real-time PCR. This study provided valuable clues to understanding the mechanisms underlying the hypoxic responses of M. japonicus. • The physiological indexes showed time-dependent changes upon hypoxia stress. • Hypoxia stress led to significant tissue damage and cell apoptosis. • A total of 900 and 1400 DEGs were detected in the gill and hepatopancreas. • Glycolysis, anaerobic metabolism and TCA cycle were affected by hypoxia stress. [ABSTRACT FROM AUTHOR]

As invertebrates, shrimp are generally thought to solely rely on their innate immune system to combat invading pathogens. Recently, an increasing number of studies have revealed that the innate immune response of invertebrates exhibits diversity and specificity based on their diverse immune molecules. Herein, a full-length transcriptome analysis of several immune-related tissues (hepatopancreas, gill, hemocytes, stomach and intestine) in the kuruma shrimp (Marsupenaeus japonicus) was conducted to identify immune-related molecules with a focus on transcript variations. In total, 11,222 nonredundant full-length transcripts with an N50 length of 5174 were obtained, and most of these transcripts (94.84%) were successfully annotated. In addition, a total of 147 long noncoding RNAs (lncRNAs) were also predicted. Importantly, transcript variants of several vital immune-related genes were observed, including twenty-five alpha-2-macroglobulins (α2-Ms), ten Toll-like receptors (TLRs), six C-type lectins (CTLs), five M-type lectins (MTLs) and three Down syndrome cell adhesion molecules (Dscams). Furthermore, 509 nonredundant full-length transcripts were predicted to be generated from alternative splicing (AS) events, which contribute to the diversity of immune molecules. Overall, our study provides valuable data on the full-length transcripts of M. japonicus , which will facilitate the exploration of immune molecules in this species. Moreover, numerous transcript variants of immune molecules detected in this study provide clues for further investigating the diversity and specificity of the innate immune response in shrimp. • Full-length transcriptome analysis of immune-related tissues in Marsupenaeus japonicus was conducted. • 11,222 full-length transcripts enrich the transcriptomic information of M. japonicus. • Numerous transcript variations of immune molecules provide new insights into the innate immune system of shrimp. [ABSTRACT FROM AUTHOR]

Kuruma shrimp, a major farmed shrimp species in the world, includes two cryptic or sibling species, Form I (Marsupenaeus japonicus) and Form II (Marsupenaeus pulchricaudatus). Due to the lack of genomic resources, little is known about the molecular mechanisms associated with immune defense and hypoxia tolerance. Here, we sequenced the transcriptomes of two closely related Marsupenaeus species and compared genomic divergence. This study obtained 77049 and 84561 unigenes with N50 values of 1281bp and 1244bp for M. japonicus and M. pulchricaudatus , respectively, and 5036 pairs of putative orthologs were identified between two Marsupenaeus species. Estimation of Ka/Ks ratios indicated that 165 orthologous genes may be under positive selection (Ka/Ks > 0.5), including 49 pairs with a Ka/Ks ratio >1. According to the peak of synonymous rates, the divergence time between M. japonicus and M. pulchricaudatus was about 0.26–0.69 Mya. These positively selected orthologous genes related to the immune process mainly comprised single VWC domain protein, legumain , ras-related C3 botulinum, caspase , C-type lectin and were enriched in functions related to immune (Toll-like receptor and PI3K-Akt signaling) and hypoxia signaling (HIF-1 signaling and VEGF signaling). In this study, dozens of caspase-like unigenes were screened from two Marsupenaeus transcriptomes. Among these, the PjCaspase orthologous gene was subjected to positive selection (Ka/Ks = 1.22), which had different secondary and three-dimensional structure prediction. Based on the single copy caspase gene, eight populations of Marsupenaeus species were divided into two phylogeographic lineages from the East and South China. We characterized the transcriptomes of the two Marsupenaeus species and obtained several key orthologs associated with immune defense and hypoxia tolerance, which provides new insights into the immunity and genetic divergence of the two varieties. Moreover, this study will facilitate further comparative genomic studies of the two varieties. • This study firstly performed a de novo transcriptome comparison between two Marsupenaeus species. • Estimation of Ka/Ks ratios indicated that 165 orthologous genes may be under positive selection (ka/ks > 0.5) • These positively selected orthologous genes mainly comprised single VWC domain protein, Caspase , and C-type lectin. • This study found dozens of caspase-like unigenes from two Marsupenaeus transcriptomes. • The expression profiles of hypoxia tolerance related genes showed significant difference between the two varieties. [ABSTRACT FROM AUTHOR]

Anti-lipopolysaccharide factor (ALF), which belongs to the antimicrobial peptide (AMP) family, has become a relatively new weapon to combat severe infections and has been demonstrated to be active against bacteria, fungi and some viruses. In the present study, a new ALF of group D (MjALF-D; GenBank accession No. MN416688) from Marsupenaeus japonicus was detected. MjALF-D encodes a polypeptide with 124 aa, and the peptide contains a 26-residue signal peptide and a lipopolysaccharide-binding domain (LBD). The structure of MjALF-D was found to consist of three α-helices, four β-sheets and random coils. qRT-PCR analysis revealed that MjALF-D expression was primarily observed in the stomach and was universally upregulated in both the gill and stomach after challenge by lipopolysaccharide (LPS) and Vibrio parahaemolyticus. Moreover, rMjALF-D can inhibit the growth of V. parahaemolyticus. rMjALF-D could destroy the bacterial membrane and lead to cytoplasmic leakage investigated by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), which may be the mechanism by which rMjALF-D inhibits V. parahaemolyticus. Additionally, rMjALF-D showed distinct binding or antibacterial ability after direct incubation with V. parahaemolyticus or bacterial genomic DNA and a certain effect on the protein expression of it. Together, these results indicated that rMjALF-D possessed the antibacterial activity against V. parahaemolyticus and the potential involvement in the innate immune response of M. japonicus. • A group D anti-lipopolysaccharide factor (ALF) was identified in M. japonicus. • It could be upregulated after LPS and Vibrio parahaemolyticus challenge. • rMjALF-D could inhibit the growth of V. parahaemolyticus and destroy the bacterial membrane, leading to cytoplasmic leakage. • rMjALF-D could bind to bacteria directly and genomic DNA. • rMjALF-D played a multiple role in the bactericidal process. [ABSTRACT FROM AUTHOR]