Baojie Wang

Purification and characterization of a natural lectin from the plasma of the shrimp Fenneropenaeus chinensis

A natural lectin from the plasma of the shrimp Fenneropenaeus chinensis was purified by singlestep affinity chromatography using fetuin-coupled agarose. The purified plasma lectin showed a strong affinity for human A/B/O erythrocytes (RBC), mouse RBC and chicken RBC. The hemagglutinating (HA) activity of the lectin was dependent on Ca(2+) and reversibly sensitive to EDTA. This lectin was named FC-L and its inactive form had a molecular mass estimate of 168 kDa. Fifteen N-terminal amino acid sequences of this protein were determined. We performed HA-inhibition assays with several carbohydrates and glycoproteins. FC-L showed a distinct and unique specificity to N-acetylated sugars, particularly sialic acid and sialoproteins. The FC-L also has binding activity to some Gram-negative bacteria which caused disease in shrimp and fish. The activity of FC-L was inhibited at temperatures greater than 75 degrees C and at a pH less than 7 or greater than 11. These results suggest that FC-L may play a role as pattern recognition proteins in the reorganization and clearance of invaders in shrimp F. chinensis.

De Novo Characterization of Japanese Scallop Mizuhopecten yessoensis Transcriptome and Analysis of Its Gene Expression following Cadmium Exposure

Background Japanese scallop has been cultured on a large-scale in China for many years. However, serious marine pollution in recent years has resulted in considerable loss to this industry. Moreover, due to the lack of genomic resources, limited research has been carried out on this species. To facilitate the understanding at molecular level immune and stress response mechanism, an extensive transcriptomic profiling and digital gene expression (DGE) database of Japanese scallop upon cadmium exposure was carried out using the Illumina sequencing platform. Results RNA-seq produced about 112 million sequencing reads from the tissues of adult Japanese scallops. These reads were assembled into 194,839 non-redundant sequences with open reading frame (ORF), of which 14,240 putative amino acid sequences were assigned biological function annotation and were annotated with gene ontology and eukaryotic orthologous group terms. In addition, we identified 720 genes involved in response to stimulus and 302 genes involved in immune-response pathways. Furthermore, we investigated the transcriptomic changes in the gill and digestive gland of Japanese scallops following cadmium exposure using a tag-based DGE system. A total of 7,556 and 3,002 differentially expressed genes were detected, respectively, and functionally annotated with KEGG pathway annotations. Conclusion This study provides a comprehensive transcripts sequence resource for the Japanese scallop and presents a survey of gene expression in response to heavy metal exposure in a non-model marine invertebrate via the Illumina sequencing platform. These results may contribute to the in-depth elucidation of the molecular mechanisms involved in bivalve responses to marine pollutants.

Sensitive and Rapid Detection of Genetic Modified Soybean (Roundup Ready) by Loop-Mediated Isothermal Amplification

Using the LAMP method, a highly specific and sensitive detection system for genetically modified soybean (Roundup Ready) was designed. In this detection system, a set of four primers was designed by targeting the exogenous 35S epsps gene. Target DNA was amplified and visualized on agarose gel within 45 min under isothermal conditions at 65 °C. Without gel electrophoresis, the LAMP amplicon was visualized directly in the reaction tube by the addition of SYBR Green I for naked-eye inspection. The detection sensitivity of LAMP was 10-fold higher than the nested PCR established in our laboratory. Moreover, the LAMP method was much quicker, taking only 70 min, as compared with 300 min for nested PCR to complete the analysis of the GM soybean. Compared with traditional PCR approaches, the LAMP procedure is faster and more sensitive, and there is no need for a special PCR machine or electrophoresis equipment. Hence, this method can be a very useful tool for GMO detection and is particularly convenient for fast screening.

The transcriptomic response to copper exposure in the digestive gland of Japanese scallops (Mizuhopecten yessoensis)

The present study was conducted to elucidate the effects of copper exposure on the immune system and lipid metabolism of the Japanese scallop, Mizuhopecten yessoensis. Transcriptional levels of differentially expressed genes (DEGs)in M. yessoensis digestive gland tissue were analyzed using the deep-sequencing platform Illumina HiSeq™ 2000. In total, 841 and 877 genes were identified as significantly up- or down-regulated, respectively. In addition, significant enrichment analysis identified 3 gene ontology terms and 15 pathways involved in the response to copper exposure. Analysis of transcripts related to the immune response revealed a complex pattern of innate recognition receptors, including toll-like receptors, NOD-like receptors and downstream pathway effectors, including those involved in apoptosis. Furthermore, genomic analysis revealed that genes involved in extracellular matrix (ECM)-receptor interactions were enriched in Cu-exposed scallop glands. These results will provide a resource for subsequent gene expression studies regarding heavy metal exposure and the identification of copper-sensitive biomarkers for the aquaculture of M. yessoensis.

Transcriptome, antioxidant enzyme activity and histopathology analysis of hepatopancreas from the white shrimp Litopenaeus vannamei fed with aflatoxin B1(AFB1)

&NA; Aflatoxin produced by Aspergillus flavus or Aspergillus parasiticus fungi during grain and feed processing and storage. Aflatoxins cause severe health problems reducing the yield and profitability of shrimp cultures. We sought to understand the interaction between shrimp immunity and aflatoxin B1 (AFB1), analyzing transcriptome expression, antioxidant enzyme activity, and histological features of the hepatopancreas of shrimp fed with AFB1. From over 4 million high‐quality reads, de novo unigene assembly produced 103,644 fully annotated genes. A total of 1024 genes were differentially expressed in shrimp fed with AFB1, being involved in functions, such as peroxidase metabolism, signal transduction, transcriptional control, apoptosis, proteolysis, endocytosis, and cell adhesion and cell junction. Upon AFB1 challenge, there were severe histological alterations in shrimp hepatopancreas. AFB1 challenge increased the activity of several antioxidant enzymes. Our data contribute to improve the current understanding of host‐AFB1 interaction, providing an abundant source for identification of novel genes. HighlightsThis study provides valuable information how the L. vannamei respond to AFB1 toxicity.The transcriptome information of L. vanname challenged AFB1 is presented for the first time.Provides evidence and analysis in physiological, biochemical and molecular levels.

Bacterial Population in Intestines of Litopenaeus vannamei Fed Different Probiotics or Probiotic Supernatant.

The interactions of microbiota in the gut play an important role in promoting or maintaining the health of hosts. In this study, in order to investigate and compare the effects of dietary supplementation with Lactobacillus pentosus HC-2 (HC-2), Enterococcus faecium NRW-2, or the bacteria-free supernatant of a HC-2 culture on the bacterial composition of Litopenaeus vannamei, Illumina sequencing of the V1-V2 region of the 16S rRNA gene was used. The results showed that unique species exclusively existed in specific dietary groups, and the abundance of Actinobacteria was significantly increased in the intestinal bacterial community of shrimp fed with the bacteria-free supernatant of an HC-2 culture compared with the control. In addition, the histology of intestines of the shrimp from the four dietary groups was also described, but no obvious improvements in the intestinal histology were observed. The findings in this work will help to promote the understanding of the roles of intestinal bacteria in shrimps when fed with probiotics or probiotic supernatant.

Synthesis and characterization of biodegradable thermoresponsive N-maleyl gelatin-co-P(N-isopropylacrylamide) hydrogel cross-linked with Bis-acrylamide for control release

Biodegradable N-maleyl gelatin (N-MAGEL) was synthesized with gelatin (GEL) and maleic anhydride (MAH) by acylation. With Bis-acrylamide (Bis) cross-linker, a series of cross-linked poly (N-isopropylacrylamide-co-N-maleyl gelatin) [P(NIPAAm-co-N-MAGEL)] hydrogels were prepared, and their lower critical solution temperature (LCST), water content, swelling/deswelling kinetics, drug delivery behaviors, and enzymatic degradation properties were investigated. As weight ratios of N-MAGEL/NIPAAm in hydrogel were increasing, water content increased, and it had no significant influence on LCST of hydrogel in spite of the weight ratios reaching 30:70. In the swelling/deswelling kinetics, hydrogels with higher weight ratios of N-MAGEL/NIPAAm exhibited faster swelling rate and slower deswelling rate. Drug delivery for bovine serum albumin (BSA) research indicated that increasing weight ratios of N-MAGEL/NIPAAm in hydrogels contribute to raising release time and cumulative release ratios. Lastly, hydrogels with higher weight ratios of N-MAGEL/NIPAAm degraded rapidly in phosphate-buffered saline (PBS) containing papain.

Gut bacterial diversity of farmed sea cucumbers Apostichopus japonicus with different growth rates

Farmed sea cucumbers (Apostichopus japonicus) of the same age displayed significantly different body sizes and weights under the same farming conditions. To examine the gut bacterial diversity of sea cucumbers and whether the growth differences between them were related to intestinal microbiota, the bacterial species from intestinal samples of 30 farmed A. japonicus from the same tank (15 large sea cucumbers vs. 15 small sea cucumbers) were characterized based on 16S rRNA gene analysis by means of high throughout sequencing. The results showed that bacterial phylotypes in both sizes of sea cucumbers were closely related to Proteobacteria, Verrucomicrobia, Bacteroidetes, Actinobacteria, Firmicutes, Cyanobacteria, Planctomycete, and Spirochaetes, of which Proteobacteria were predominant (>50%). There were no significant differences in the relative abundances of each bacterial phylotype between the two groups, except for Actinobacteria (P < 0.05). In addition, different species uniquely belonging to all three tested samples in the large group and the small group were found. It was interestingly that Vibrio were absent from both groups. It is likely that the differences in the abundances of Actinobacteria and different species in the two groups may be related to their remarkable disparities in body sizes.

Effect of dietary supplementation of vitamin C on growth, reactive oxygen species, and antioxidant enzyme activity of Apostichopus japonicus (Selenka) juveniles exposed to nitrite

Different amounts of vitamin C were added to diets fed to juveniles (2.5 ± 0.15 g) of sea cucumber Apostichopus japonic u s (Selenka) in an attempt to reduce the stress response of specimens exposed to nitrite stress. A commercial feed was used as the control diet and three experimental diets were made by supplementing 1 000, 1 500, or 2 000 mg vitamin C/kg diet to control diet separately in a 45-day experiment. Sea cucumbers were exposed to three different levels (0.5, 1.0, and 1.5 mg/L) of nitrite stress for 4, 8, and 12 h at four time intervals (0, 15, 30, and 45 d). Growth of the animals was recorded during the experiment. Reactive oxygen species (ROS) (i.e. hydroxyl free radical (-OH), malondialdehyde (MDA) and total antioxidant capacity (T-AOC)) and antioxidant enzyme activities (i.e., superoxide dismutase (SOD) and catalase (CAT)) were measured. Response surface methodology (RSM) was used to analyze the effect of multiple factors on ROS indices and enzyme activities. Weight gain (WG) and special growth rate (SGR) of vitamin C supplementation groups were significantly higher than those of control group (P < 0.05). The levels of -OH and MDA increased under exposure time extending and nitrite concentration increasing, whereas T-AOC level decreased. SOD and CAT activities increased at 4 h and 8 h and decreased at 12 h. During the days in which the animal consumed experimental diets, the levels of -OH and MDA decreased and that of T-AOC increased. This result suggests that diets containing vitamin C could reduce the nitrite stress response in the animals and increase their antioxidant capacity. The multifactor regression equation of growth performance, ROS indices, and duration of feeding results suggest that vitamin C supplementation of 1 400–2 000 mg/kg diet for 29–35 days could reduce effectively the effects of nitrite exposure.

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

ABSTRACT 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. HighlightsProvides valuable information about how the L. vannamei intestinal barrier respond to V.parahaemolyticus infection.The first simultaneous transcriptome and morphological analyses of changes of the intestinal barrier in L. vannamei.The transcriptome information of L. vanname challenged by V.parahaemolyticus. is presented for the first time.