Kuan-Fu Liu

Effects of the probiotic, Bacillus subtilis E20, on the survival, development, stress tolerance, and immune status of white shrimp, Litopenaeus vannamei larvae.

In this study, the probiotic, Bacillus subtilis E20, isolated from the human health food, natto, was used for white shrimp, Litopenaeus vannamei, larvae breeding to improve the larval survival rate and development by adding probiotic to the rearing water at (control), 10(8), and 10(9) cfu L(-1) salt water once every 3 days during the 14 days of breeding experiment. Thereafter, stress tolerance and immune status of postlarvae were evaluated. Shrimp larval development was significantly accelerated after adding the probiotic to the larval rearing water at a level of 10(9) cfu L(-1). The survival rate of larvae was significantly higher in the treatment with 10(9) cfu L(-1) compared to the control and the treatment with 10(8) cfu L(-1) after all larvae had metamorphosed to postlarvae. Adding the probiotic to the shrimp larvae rearing water produced a weak inhibition of bacterial growth by an analysis of the total bacterial count and presumptive Vibrio count. For stress tests, no postlarvae died when they were reared in water in which the temperature was decreased from 30 to 2 degrees C at a rate of 0.1 degrees C min(-1). Postlarvae had significantly lower cumulate mortality in the treatments with 10(8) and 10(9) cfu L(-1) compared to the control when they were suddenly exposed to fresh water and 60 per thousand salt water. A significant decrease in the cumulative mortality of postlarvae treated with the probiotic at a level of 10(9) cfu L(-1) was recorded after the sudden transfer to 300 mg L(-1) nitrite-N compared to the control and treatment with 10(8) cfu L(-1). The analysis of immune-related gene expressions showed that the gene expression of prophenoloxidase I, prophenoloxidase II, and lysozyme of larvae were significantly increased after being reared in probiotic-containing water at the levels of 10(8) and 10(9) cfu L(-1). However, no significant difference in serine proteinase or glutathione peroxidase gene expressions was recorded in this study. It is therefore suggested that 10(9) cfu L(-1) of probiotic, B. subtilis E20 adding to rearing water for shrimp larva breeding.

Hepatopancreas and ovary are sites of vitellogenin synthesis as determined from partial cDNA encoding of vitellogenin in the marine shrimp, Penaeus vannamei

Summary The site of yolk protein synthesis in crustaceans has long been a subject of controversy. A portion of the vitellogenin gene structure was reported recently in a freshwater giant prawn (Macrobrachium rosenbergii) and black tiger shrimp (Penaeus monodori), in which the hepatopancreas was confirmed to be the extraovarian site of vitellogenin synthesis. The ovary is also frequently reported to be the site of yolk protein synthesis in penaeid shrimp. The same PCR product was obtained using cDNA from the hepatopancreas or the ovary as a template. The deduced amino acid sequence of Vg in P. vannamei showed high identities of 57% and 78% with those from M. rosenbergii and P. monodon, respectively. The same location of the intron in the sequenced region of genomic DNA was also found between these three species. We therefore concluded that the hepatopancreas and ovary are sites of vitellogenin synthesis in P. vannamei. The partial structure of the vitellogenin gene is further presented.

Silencing tyrosine hydroxylase retards depression of immunocompetence of Litopenaeus vannamei under hypothermal stress

ABSTRACT Tyrosine hydroxylase (TH), the first and rate‐limiting step in the synthesis of catecholamines, is required in catecholamine synthesis of the neuroendocrine regulatory network against stress in shrimp. The immunocompetence, catecholamine biosynthesis, and carbohydrate metabolites were evaluated in Litopenaeus vannamei received L. vannamei TH (LvTH) double‐stranded (ds)RNA, diethyl pyrocarbonate‐water, or non‐targeted dsRNA for 3 days then transferred from 28 to 20 or 28 °C. The immunocompetence of LvTH‐depleted shrimp held at 28 °C was promoted, and those were downregulated under hypothermal stress and revealed higher level than the other two dsRNA treatments. Meanwhile, the decrease of catecholamine biosynthesis was observed in LvTH‐depleted shrimp held at 28 °C, and those were elevated under hypothermal stress and revealed lower levels, compared to two dsRNA treatments. The reduced carbohydrate metabolites was observed in LvTH‐depleted shrimp held at 28 °C, and those were upregulated under hypothermal stress and showed lower levels than the other two dsRNA treatments. It was therefore concluded that LvTH‐depleted shrimp revealed enhanced immunocompetence and reduced carbohydrate metabolites when exposed to a hypothermal stress condition, and in the meantime, even though catecholamine biosynthesis was downregulated, no significant difference was observed in DA or NE levels. HighlightsLvTH‐depleted shrimp exhibited enhanced immunocompetence and reduced carbohydrate metabolite.Depressed immunocompetence under hypothermal stress was retarded in LvTH‐depleted shrimp.TH modulated physiological and immunological responses under normal and hypothermal conditions.

L-3,4-Dihydroxyphenylalanine ( l -DOPA) induces neuroendocrinological, physiological, and immunological regulation in white shrimp, Litopenaeus vannamei

ABSTRACT L‐3,4‐Dihydroxyphenylalanine (l‐DOPA) is a precursor for dopamine (DA) synthesis. Assessments were conducted to analyze the effects of l‐DOPA on mediating regulation of neuroendocrinological, immunological, and physiological parameters in the shrimp, Litopenaeus vannamei when they were individually injected with 0.01N HCl or l‐DOPA at 0.5 or 1.0&mgr;mol shrimp−1 for 60, 120, and 240min. For catecholamine synthesis evaluation, tyrosine hydroxylase (TH) and DA beta hydroxylase (DBH) activities, l‐DOPA, DA, and norepinephrine (NE) levels in hemolymph were determined. The total hemocyte count (THC), differential hemocyte count (DHC), phenoloxidase (PO) activity, respiratory bursts (RBs), superoxide dismutase (SOD) activity, phagocytic activity, and clearance efficiency in response to the pathogen, Vibrio alginolyticus were assessed for immune responses, and plasma glucose and lactate levels were for physiological response. Results showed that the TH activity, THC, hyaline cells (HCs), and semigranular cells (SGCs) at 120 min, DA levels at 60–240 min, PO activity in hemocytes per 50 &mgr;L of hemolymph at 60–120 min, and PO activity per granulocyte (granular cells (GCs) + SGCs) at 60 min significantly increased, but TH activity, l‐DOPA levels, GCs, SGCs, and respiratory bursts in hemocytes per 10&mgr;L of hemolymph at 60min, respiratory bursts per hemocyte and SOD activity at 120min, phagocytic activity at 60–240min, and the clearance efficiency at 60–120min significantly decreased in shrimp injected with l‐DOPA at 1.0&mgr;mol shrimp−1. In another experiment, 60min after shrimp had received l‐DOPA at 0.5 or 1.0&mgr;mol shrimp−1, they were challenged with an injection of V. alginolyticus at 2×105colony‐forming units (cfu) shrimp−1. The injection of l‐DOPA at 1.0&mgr;mol shrimp−1 also significantly increased the cumulative mortality of shrimp by 16.7%, compared to the HCl‐challenged control after 120h. These results suggest that l‐DOPA administration at 1.0&mgr;mol shrimp−1 can mediate the transient regulation of neuroendocrinological, immunological, and physiologic responses resulting in immunosuppression, which in turn promoted the susceptibility of L. vannamei to V. alginolyticus. HIGHLIGHTSCatecholamine biosynthesis was mediated after the shrimp receiving l‐DOPA.l‐DOPA induced the immunological and physiologic responses of shrimp.The decreased survival rate against infection revealed in shrimp after receiving l‐DOPA.