Effects of Hanseniaspora opuntiae C21 on the growth and digestive enzyme activity of juvenile sea cucumber Apostichopus japonicas

The effects of a diet containing Hanseniaspora opuntiae C21 on growth and digestive enzyme activity were estimated in juvenile Apostichopus japonicus. Groups of sea cucumbers were fed diets containing H. opuntiae C21 at 0(control), 10 4, 10 5, and 10 6 CFU(colony-forming units)/g feed. Results showed that after 45 d the specific growth rate(SGR) of sea cucumbers fed a C21-supplemented diet at 10 4 CFU/g feed was significantly higher than that of the control( P < 0.05). Intestinal trypsin and lipase activities were significantly enhanced by C21 administration at 10 4 and 10 5 CFU/g feed compared with the control( P < 0.05). After feeding for 23–42 d, C21 was demonstrated by denaturing gradient gel electrophoresis to be present in the intestine of sea cucumbers. In addition, after feeding the C21-supplemented diets for 15 d, the sea cucumbers were switched to an unsupplemented diet and C21 was confirmed to be capable of colonizing the intestine for at least 31 d after cessation of feeding. In conclusion, C21 was shown to successfully colonize the intestine of juvenile A. japonicus via dietary supplementation, and improve growth and digestive enzyme activity.

Relationships between body weight and other morphological traits in young sea cucumbers Apostichopus japonicas

Correlation, multiple regression, and path analyses were used to investigate the relationships between body weight and three other morphological traits in juvenile Japanese sea cucumbers Apostichopus japonicus . We measured live body weight (BW), body length (BL), numbers of papillae (NP), and numbers of tube feet (NF) at 60, 80, 100, and 130 days post-hatching (dph). We calculated path correlation coefficients, correlation indices ( R 2 ), and coefficients of determination with BW as the dependent variable and the other morphological traits as independent variables. The coefficient of variation for BW was high across all age groups, and all measured morphological traits were significantly correlated ( P < 0.01). BL had the greatest direct eff ect on BW across all age groups (60 dph, 0.526;80 dph, 0.404;100 dph, 0.620;and 130 dph, 0.681), while NF had the greatest indirect eff ect on BW across all age groups (60 dph, 0.528;80 dph, 0.452;100 dph, 0.666;and 130 dph, 0.603). Regression analyses between morphological traits and BW indicated that R 2 was greater than 0.85 only in the 100-dph specimens. The indirect eff ects of the other measured morphological traits on BW were age-dependent. The optimal regression equations, as determined with stepwise regression, were, for 60-dph specimens: BW 60 =10 (-3.04+0.092BL+0.014NP+0.014NF)( R 2 =0.632);for 80-dph specimens: BW 80 =10 (- 3.035+0.056BL+0.017NP+0.02NF)( R 2 =0.686);for 100-dph specimens: BW 100 =10 (- 3.742+0.069BL+0. 633 · lg (NP)+0. 464 · lg (NF))( R 2 =0.893);and for 130-dph specimens: BW 130 =10 (- 2.472+0.065BL+0.012NP)( R 2 =0.774). Our work clarified the correlation between various morphological traits and body weight of a commercially-important sea cucumber species ( A . japonicus ). Our predictive models for body weight might be useful for the aquaculture and selective breeding of A . japonicus . These models might also provide theoretical support for the indirect selection of traits that are difficult to select directly.

Isolation of a New PAK1 Gene from Sea Cucumber(Apostichopus japonicus) and Its Expression Analysis and Function Characterization

The p21-activated kinase 1(PAK1)is a downstream serine/threonine kinase effector of Rac1/Cdc42 that regulates various biological processes including those associating with pathological inflammation.To investigate the function of PAK1 in echinoderms,we isolated a new PAK1 from sea cucumber Apostichopus japonicus(AjPAK1)by transcriptome database mining and with rapid amplification of cDNA ends(RACE).The full-length cDNA AjPAK1 was 2303 bp in length,containing a 1587 bp ORF encoding 528 amino acid residues.The deduced AjPAK1 contained a p21-Rho-binding domain(PBD)and a serine/threonine protein kinase catalytic domain(S_TKc),which was similar to the PAK1 of crown-of-thorns starfish Acanthaster planci and other eukaryotes.AjPAK1 expressed in all tissues of adult A.japonicus analyzed with the highest transcript anumdance detected in coelomocytes.Significant change in AjPAK1 abundance was observed at 4,24,48 and 72 h after Vibrio splendidus infection.Silencing AjPAK1 induced a significant reduction of lysozyme content in coelomic fluid and relative transcript abundances of AjRac1 and AjMKK3/6 in A.japonicus coelomocytes.These results should aid to characterize PAK1 of sea cucumber and decipher its immune function.

Characterization and Expression Analysis of Protein Kinase C Gene from Dunaliella salina

Protein kinase C (PKC) has a crucial role in signal transduction for a variety of biologically active substances which activate cellular functions and proliferation. We previously isolated the full-length PKC gene from Dunaliella salina (DsPKC) using rapid amplification of cDNA ends (RACE) and RT-PCR methods. And we submitted the mRNA sequence of DsPKC gene to NCBI (Genbank No. JN625213). In the present paper, the DsPKC gene open reading frame obtained by PCR was cloned into pGS-21a vector and transformed into Escherichia coli to generate the fusion protein. Bioinformatics analysis revealed that DsPKC gene was a member of serine/threonine kinase with two conserved domains and highly conserved motifs. The DsPKC was highly expressed upon induction with isopropyl-β-d-thiogalactoside (IPTG) at a final concentration of 0.2 mmol L 1 at 37℃. Under salt stress, the fu- sion protein Green Fluorescent Protein (GFP)-DsPKC was transferred from the cytoplasm to the cell membrane. The expression pat- tern of DsPKC gene was analyzed using real-time quantitative PCR, and indicated that DsPKC gene was up-regulated by 3.0 mol L 1 NaCl at 12 h, which was significantly higher than in control values (P < 0.05). These results suggest that the DsPKC gene plays an important role in response to salt stress in D. salina.

Temperature-control-induced masculinization in tiger puffer Takifugu rubripes

This study used tiger puffer Takifugu rubripes to explore new,simple methods for lowtemperature-induced masculinization in a cultured fish without the use of ultraviolet irradiation or sex hormones.An orthogonal test L9(3^4)design was used to consider three factors at three levels:treatment starting times(days post-hatch,dph:factor A)of 20,50 and 80 dph;treatment temperatures(factor B)of 13℃,15℃and 17℃;and treatment durations(factor C)of 30,45 and 60 days.A control group was reared at 21±1℃.The experiments were repeated twice.At 230 dph,the gonads were removed from thirty randomly sampled fish in each group.Histological observations and analysis of single nucleotide polymorphisms(SNP)were used to identified pseudo males,which biological sex was male and genetic sex were female(XX).Treatment group 4(A2B1C2)resulted in the highest proportion of males(75%).According to the intuitive analysis of the orthogonal-array experiments,the optimal combination of lowtemperature-induced masculinization of T.rubripes was A2B1C2.The population sex ratio depended on the three factors in the sequence B→A→C.A comparison of the daily increases in length and weight during and after the low-temperature conditions showed that the absolute daily increases in weight and length were significantly less during treatment than after treatment.Daily increases in weight and length did not significantly differ between the treatment groups and the controls(P>0.05),demonstrating that the growth rate could return to normal after the low-temperature conditions.This study establishes a low-temperatureinduced masculinization technology for T.rubripes and demonstrates that although the growth rate(length and weight)decreased in an array of nine treatment groups during the processing time,it returned to a normal level after processing.The results should serve as a guide for achieving the masculinization of T.rubripes in production.