Molecular analysis of the 16S rDNA of the intestinal microbiota of whiteleg shrimp
Litopenaeus vannamei
was examined to investigate the effect of a
Bacillus
mix (
Bacillus endophyticus
YC3-b,
...Bacillus endophyticus
C2-2,
Bacillus tequilensis
YC5-2) and the commercial probiotic (Alibio
®
) on intestinal bacterial communities and resistance to
Vibrio
infection. PCR and single strain conformation polymorphism (SSCP) analyses were then performed on DNA extracted directly from guts. Injection of shrimp with
V. parahaemolyticus
at 2.5 × 10
5
CFU g
−1
per shrimp followed 168 h after inoculation with
Bacillus
mix or the Alibio probiotic or the positive control. Diversity analyses showed that the bacterial community resulting from the
Bacillus
mix had the highest diversity and evenness and the bacterial community of the control had the lowest diversity. The bacterial community treated with probiotics mainly consisted of α- and γ-proteobacteria, fusobacteria, sphingobacteria, and flavobacteria, while the control mainly consisted of α-proteobacteria and flavobacteria. Differences were grouped using principal component analyses of PCR-SSCP of the microbiota, according to the time of inoculation. In
Vibrio parahaemolyticus
-infected shrimp, the
Bacillus
mix (~33 %) induced a significant increase in survival compared to Alibio (~21 %) and the control (~9 %). We conclude that administration of the
Bacillus
mix induced modulation of the intestinal microbiota of
L. vannamei
and increased its resistance to
V. parahaemolyticus
.
In bivalve aquaculture, selecting suitable probiotic treatments can be crucial for improving hatchery‐rearing of larvae and juveniles. We assessed the potential of five bacterial strains, previously ...selected in vitro, to improve survival, growth and resistance of catarina scallop Argopecten ventricosus during early and late larval and juvenile developmental stages, as well as during exposure to the pathogen Vibrio alginolyticus. Hatchery‐reared larvae and juveniles were treated with eight treatments of single or combined strains of Bacillus and Lactobacillus at 1 × 106 CFU mL−1 every 48 h for 9 days (larvae) and 21 days (juveniles). Compared with the control, significantly higher survival and growth in size and weight of early veliger larvae occurred with the antibiotic and the RL5 (Lactobacillus graminis) treatments. Significantly enhanced settlement of pediveliger larvae occurred with a different probiotic strain, the mix of Lactobacillus and Bacillus (MIX‐LB), while higher survival and growth of early juveniles occurred with C3 (Lactobacillus plantarum). The mix of Bacillus (MIX‐B) significantly increased survival of juveniles from V. alginolyticus after 120‐h infection, consistent with maximum activity of superoxide dismutase enzyme. In contrast, all untreated and infected scallops died by 96 h. The three Bacillus strains performed poorly when used as single treatments and when given to early developing larvae. Our results indicate that the action mechanism of probiotic strains is stage specific and strain specific, generating different responses by the host, including improved survival and growth (likely from better nutrient assimilation) and higher resistance against pathogens (possibly from strengthening the immune system).
Antibiotic usage to control infectious diseases in shrimp aquaculture has led to serious problems on antimicrobial resistance. An alternative to mitigate this issue is the use of probiotics, which ...can be easily administered by feed and water. This study examines immunomodulatory and protective effects of the marine yeasts Debaryomyces hansenii CBS8339 (Dh) and Yarrowia lipolytica Yl-N6 (Yl) -alone and mixed-in white shrimp Penaeus vannamei post-larvae. Administration routes (fed and water alone or in combination), supplementation frequency and time elapsed after the last dietary supplement were tested on growth and gene expression of penaeidin, lectin, lysozyme, superoxide dismutase, catalase, and peroxidase, as well as survival upon Vibrio parahaemolyticus IPNGS16 challenge. Penaeidin and lectin genes were upregulated in post-larvae fed orally with Yl or combined Dh + Yl. Higher growth and survival for yeast supplementation treatments were observed compared to the control group, mainly when yeasts (Dh + Yl) and administration routes (feed and water) were combined. In conclusion, mixed yeast and combined administration routes improved growth and immunity against V. parahaemolyticus.
•Individually or combined yeast administration increase growth of shrimp postlarvae.•Combined yeast routes induce higher survival in shrimp postlarvae after infection.•Oral yeast supplementation was the most efficient route on immune response in shrimp postlarvae.•Daily administration of combined yeast and route protect shirmp against bacterial infection at least for seven days.
The immunostimulatory potential of the marine yeast Yarrowia lipolytica (D1 and N6 strains) administered orally was evaluated in the white shrimp Litopenaeus vannamei. Yeasts and commercial glucans ...were mixed with a commercial feed to formulate diets with a 1.1% concentration of immunostimulants. The shrimp were fed daily for a period of 21 days. Weekly determinations were performed for immunological parameters in hemolymph, such as total hemocyte count (THC), lysozyme activity (LYZ), prophenoloxidase activity, antioxidant enzymatic activities (superoxide dismutase SOD, catalase CAT, and peroxidases), and bactericidal activity against Vibrio parahaemolyticus. Expression profiles of penaeidin (PEN), lysozyme (LYZ), and prophenoloxidase (proPO) immune genes were evaluated in hemocytes. In general, an increase in the immune parameters was observed in shrimp fed yeast diet compared to glucan and the control diets. Yarrowia lipolytica, especially strain N6, provided maximum immunostimulatory effects evidenced by the increase of immune parameters (THC, LYZ, SOD, CAT) and gene expression profile. In conclusion, this study demonstrated that Y. lipolytica had immunostimulatory effects and increased bactericidal activity in L. vannamei hemocytes against V. parahaemolyticus. These findings open the path for the potential application of Y. lipolytica-based immunostimulant for shrimp aquaculture.
•This is the first report on the immunological response to Yarrowia lipolytica in shrimp.•Oral administration of Y. lipolytica stimulates innate immune responses.•Y. lipolytica increased bactericidal activity in shrimp.•Circulating hemocytes and immune-related enzymatic activities increased in shrimp fed Y. lipolytica.•Immune-related gene expression up-regulated in hemocytes upon yeast administration.
We studied the effects of four strains of
Bacillus isolated from the guts of healthy wild adult shrimp on survival and rate of development of whiteleg shrimp
Litopenaeus vannamei larvae to understand ...how endemic
Bacillus probiotic strains improve the health of larvae. This included bacterial isolation, assays of hemolytic activity, antagonism against pathogenic
Vibrio species, growth, and adhesion to shrimp intestinal mucus, and molecular identification.
Bacillus strains were tested on larval shrimp using a daily concentration of 1
×
10
5
CFU
mL
–1, starting each bioassay at nauplii V at a density of 225 nauplii L
−
1
. Inoculations of four natural, commercial products and antibiotic oxytetracycline were added directly to the water. All treatments induced a significant increase in survival compared to the control, with strain YC5-2 showing the highest survival (~
67%), followed by Alibio
MR (~
57%). A mix of two strains induced the highest rate of development (7.00), followed by Alibio
MR (6.35). The results showed remarkable antagonistic activity by the four non-hemolytic
Bacillus strains against
Vibrio campbelli,
V. vulnificus,
V. parahaemolyticus, and
V. alginolyticus and the potential and efficiency of probiotics isolated from shrimp gut to improve survival of shrimp larvae.
► Four strains of
Bacillus isolated from adult shrimp
L. vannamei, enhanced survival and rate of development of shrimp larvae. ►
Bacillus strains isolated from the shrimp
L. vannamei, showed antagonism against
Vibrio campbelli,
V. vulnificus,
V. parahaemolyticus, and
V. alginolyticus. ► Probiotic strains adhered and grow in shrimp intestinal mucosa. ► The highest larval survival was registered with the strain YC5-2 followed by a commercial probiotic.
We isolated microbiota from the digestive tract of Nodipecten subnodosus and Pteria sterna and determined in vitro their haemolytic activity, antagonism against Vibrio spp., bacterial hydrophobicity, ...production of extracellular enzymes and molecular identification. Five bacterial strains were selected: RL5 and C3 (Lactobacillus spp.) and PB1‐1, PB1‐5 and PB1‐6 (Bacillus spp.). The RL5 and C3 isolates showed antimicrobial activity against Vibrio spp. and the PB1‐1, PB1‐5 and PB1‐6 isolates showed enzymatic activity for amylase, protease, lipase and cellulose; the C3 and PB1‐5 isolates were highly hydrophobic. The selected strains of bacteria were tested in vivo as probiotics, together with a treatment of ampicillin and a control without bacteria on juvenile Kumamoto oysters Crassostrea sikamea. The strains were provided individually and as mixes of isolates. Survival, growth and biochemical composition of the juveniles were determined as in vivo indicators. Juveniles grew significantly larger and faster when treated with a specific mix of isolates (MIX‐B), compared with the control. The protein, lipid and carbohydrate concentrations were also significantly higher in oysters exposed to probiotic treatments, compared with the control and the antibiotic treatment. The selected microbiota showed probiotic proprieties for cultivating C. sikamea juveniles.
This study evaluated low salinity effect on acute Hg toxicity and bioconcentration capacity in L. vannamei juveniles (8.4 ± 0.7 g), because this species is frequently exposed to hypo-osmotic ...environments in its natural habitat, and in farming ponds. Hg LC50 values were 1723, 1272, 983 and 536 μg L−1 at salinity of 5 ppt (parts per thousand); 2203, 1740, 1340 and 873 μg L−1 at 10 ppt; and 7013, 5693, 1759 and 1534 μg L−1 at 25 ppt for 24, 48, 72 and 96 h, respectively. After 96 h, acute Hg toxicity in shrimp was significantly higher in salinity of 5 ppt than in 25 ppt; likewise, Hg bioconcentration in shrimp exposed to different Hg treatments was statistically greater in salinity of 5 ppt than in 25 ppt. The chemical speciation calculated in experimental waters suggested that neutral chemical Hg species (HgCl2 and HgClOH) were the most bioavailable because their fractions (51–62%) increased when salinity decreased. Therefore, the inverse relationship between Hg toxicity and salinity was due to osmotic stress and the neutral chemical Hg species fractions that increased at lower salinities. Hg bioconcentration factors indicated that the higher Hg waterborne concentrations were the most saturated uptake and storage mechanisms in shrimp. Thus, Hg concentrations in organisms did not increase in proportion to waterborne Hg concentrations in the three salinities. These results support the hypothesis of an effect of low salinity on Hg toxicity and bioconcentration capacity in L. vannamei. The safe Hg concentrations 5.4, 8.7 and 15.3 μg L−1 were proposed for shrimp exposed to salinity of 5, 10 and 25 ppt, respectively. This information allows recognizing risky environments for both wild and cultured healthy growth of these shrimp, which can help decision makers on coastal management and shrimp pond managers to have better water quality.
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•Hg toxicity and bioconcentration in shrimp were tested at 5, 10 and 25 ppt salinity.•Hg LC50–96 h values were 536, 873 and 1534 μg L−1 for 5, 10 and 25 ppt salinity.•Acute Hg toxicity in Litopenaeus vannamei juveniles was higher at lower salinities.•Hg bioconcentration in whole-body of shrimp was greater at lower salinities.•Osmotic stress and most bioavailable Hg chemical species increased at low salinity.
Vibrio parahaemolyticus is the main etiological agent of human gastroenteritis by seafood consumption and some strains from this species causing the Acute Hepatopancreatic Necrosis Disease in shrimp ...have been recently reported. The PirA-like toxin from V. parahaemolyticus (ToxA) has been recently reported as an attractive antigen implicated in subunit vaccine development. Since plants are attractive hosts for the production and delivery of vaccines in the present study plants expressing ToxA were developed to account with a low cost platform for the production and oral delivery of ToxA. Tobacco plants were genetically engineered by Agrobacterium-mediated transformation to stably integrate the ToxA-coding gene into the nuclear genome. Transgenic lines were rescued in kanamycin-containing medium and analyzed by ELISA to determine ToxA yields observing levels up to 9 μg g−1 FW leaf tissues. Western blot analysis confirmed the presence of the ToxA protein in plant extracts. Immunogenicity assessment of the plant-made ToxA was performed in mice, comprising a 4-dose oral immunization scheme; revealing the induction of anti-ToxA humoral responses (IgG in serum and IgA in feces). This study opens the path for the development of low cost plant-based vaccines against Vibrio parahaemolyticus.
To evaluate the antioxidant activity and oxidative damage by relaxing, wounding, and seeding of a saibo of different origin on Pteria sterna hosts, five oyster treatments were included: (1) relaxed ...(REL) but neither wounded nor seeded; (2) relaxed and wounded (WOU) but not seeded; (3) relaxed, wounded, and seeded with an allograft (ALL); (4) relaxed, wounded, and seeded with an autograft (AUT); and (5) unrelaxed, unwounded, and unseeded as control (CTR). Superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and thiobarbituric acid (TBARS) activity were quantified between 3 and 24 h post-seeding. Compared to the CTR oysters, which did not suffer oxidative stress, SOD activity significantly decreased in the gonad and digestive gland in all treatments and decreased in mantle tissue in AUT oysters; this indicates that the entire process of preparing oysters for pearl culture (relaxing, wounding, and seeding) generates oxidative stress in the host. CAT was not a sensitive enzyme for measuring the short-term response of oysters to the wounding–seeding processes but rather a more prolonged or chronic stress. Similar to SOD, the lowest GPx and TBARS activity in seeded oysters evidenced their susceptibility to oxidative stress and damage, particularly in the WOU treatment. Evidence from this study indicates that SOD is a more sensitive enzyme for measuring the short-term response of the host oyster to the wounding and seeding of a saibo. It is also clear that the host undergoes stress at all stages of the pearl culture process, mostly during gonad wounding and regardless of the origin of saibo.
Survival and growth were investigated in Litopenaeus vannamei fed with powdered whole leaf of Aloe vera and co-infected with white spot syndrome virus (WSSV) and Vibrio parahaemolyticus by ingestion ...and immersion, respectively. A. vera was added to commercial feed. The infection with both pathogens was made at the same time. A. vera did not affect shrimp growth. Shrimps fed with the plant (1gkgfeed−1) every two days showed higher survival as compared to the control group not fed without A. vera. A. vera has the potential to be used prophylactically as antiviral and antibacterial agent in cultured shrimps.
•This study evaluated the protective effect of aloe against pathogens in L. vannamei.•Aloe protects shrimp against WSSV and V. parahaemolyticus infections.•Aloe can be used as prophylactic treatment against diseases in cultured shrimp.
