Optimization of bacteriocin production by Lactobacillus sp. MSU3IR against shrimp bacterial pathogens

BACKGROUND: Aquaculture is one amongst the growing and major food producing sectors. Shrimp culture is one of the subsectors of aquaculture that attracts more attention because of the economic interest. However, the shrimp culture systems have been facing severe consequences and economical losses due to disease outbreaks. Risk of disease outbreak can be combated with the application of probiotics. For economically viable production of such probiotic products, the present study provides information on the optimization and partial purification of bacteriocin produced by a goat milk isolate Lactobacillus sp. MSU3IR against the shrimp bacterial pathogens. RESULTS: Bacteriocin production was estimated as a measure of bactericidal activity (arbitrary Unit/ml) over the test strains. The optimum culture conditions and media components for maximum bacteriocin production by Lactobacillus sp. MSU3IR were: pH: 5.0, temperature: 30°C, carbon source: lactose; nitrogen source: ammonium acetate; NaCl: 3.0% and surfactant: Tween 80. MRS medium was found to extend better bacteriocin production than other tested media. Upon partial purification of bacteriocin, the SDS-PAGE analysis had manifested the presence of two peptide bands with the molecular weight of 39.26 and 6.38 kDa, respectively. CONCLUSION: The present results provide baseline trend for the statistical optimization, scale up process and efficient production of bacteriocin by the candidate bacterial strain Lactobacillus sp. MSU3IR which could be used to replace the usage of conventional chemotherapeutics in shrimp culture systems

Proizvodnja lipaze s pomoću bakterije Serratia rubidaea izolirane iz mlijeka

Production of extracellular lipase in submerged culture of Serratia rubidaea has been investigated. The lipase production was optimized in shake flask experiments. The observed pH and temperature range optimum for maximum lipase production were 7–8 and 30–40 °C, respectively. With a selected nitrogen source, casein ((6.5±0.015) U/mL) and soytone ((9.4±0.02) U/mL) were suitable substrates for accelerating lipase production. The optimized concentration of casein and soytone was 24 g/L ((9.95±0.02) U/mL) and 5 g/L ((14.8±0.03) U/mL), respectively. The effect of carbon source on lipase production indicated that starch was suitable substrate to maximize lipase production ((15.60±0.20) U/mL) and the optimum concentration registered was 4 g/L ((17.46±0.20) U/mL). Investigating the effect of lipids and surfactants showed that the gingily oil ((20.52±0.20) U/mL) and Tween 20 ((27.10±0.01) U/mL) were suitable substrates for maximizing lipase production, and the optimum concentrations registered were 15 mL/L ((23.15±0.24) U/mL) and 6 mL/L ((34.20±0.01) U/mL), respectively. Partial purification of lipase indicated that the molecular mass of partially purified enzyme was 54 kDa.U radu je istražena proizvodnja ekstracelularne lipaze pri submerznom uzgoju bakterije Serratia rubidaea, optimirana pokusima na tresilici. Za maksimalnu proizvodnju lipaze optimalna pH-vrijednost bila je 7-8, a optimalna temperatura 30-40 °C. Primjenom kazeina ((6,5±0,015) U/mL) i peptona iz soje ((9,4±0,02) U/mL) kao izvora dušika, pospješena je proizvodnja lipaze. Optimirana koncentracija kazeina iznosila je 24 g/L ((9,95±0,02) U/mL), a peptona iz soje 5 g/L ((14,8±0,03) U/mL). Za maksimalnu proizvodnju lipaze ((15,60±0,20) U/mL) najbolji izvor ugljika bio je škrob, pri optimalnoj koncentraciji od 4 g/L ((17,46±0,20) U/mL). Ispitivanje utjecaja lipida i površinski aktivnih tvari pokazalo je da su sezamovo ulje ((20,52±0,20) U/mL) i Tween 20 ((27,10±0,01) U/mL) najprikladniji supstrati za maksimalnu proizvodnju lipaze. Optimalni volumni udio sezamovog ulja bio je 15 mL/L ((23,15±0,24) U/mL), a Tweena 20 je bio 6 mL/L ((34,20±0,01) U/mL). Djelomično pročišćavanje lipaze pokazalo je da je njezina molekularna masa 54 kDa

IMMUNOMODULATORY AND GROWTH-PROMOTING POTENTIAL OF LOWCOST PROBIOTIC PRODUCT IN Penaeus monodon CULTURE SYSTEM

Sposobnost probiotičkog proizvoda s niskim troškovima procijenjena je pri uzgoju vrste Penaeus monodon u vanjskom sustavu uzgoja tijekom 90 dana. Probiotski proizvod niske cijene formuliran je miješanjem bujonske kulture Bacillus cereus zajedno s komercijalnom praškastom kredom u omjeru 1: 1. Pripremljeni probiotički produkt uz dodatak praha inkorporiran je zajedno s hranom za rakove u obroke različite koncentracije (1 do 5%). Održivost B. cereusa u eksperimentalnoj prehrani testirana je jednom u 30 dana do 90 dana tijekom trajanja eksperimenta. Nakon 90 dana od početka hranidbenog istraživanja, određene su performanse rasta rakova. Utvrđena je maksimalna proizvodnja od 11,98 ± 0,367 g, SGR od 5,030 ± 0,171% i FCE od 86,63 ± 1,300% u P. monodon koji su hranjeni s D4 (4% dodatak probiotika). Imunološki parametri određeni na kraju eksperimenta pokazali su varijaciju među ishranama. Ukupni broj hemocita (273,66 ± 3,09 x 105 stanica / ml), aktivnost fenoloksidaze (0,132 ± 0,007 OD) i aktivnost respiratornog sustava (0,291 ± 0,020 OD) je bila visoka kod P. monodon koji su hranjeni s D4. Slični efekti zabilježeni su za koncentraciju proteina u plazmi (68,00 ± 1,41 mg / ml), aktivnost lizozima (0,483 ± 0,014 U / ml) i baktericidnu aktivnost (81,0 ± 1,33%) kod rakova hranjenih s D4. Zaključci ovog istraživanja su da je probiotički produkt u koncentraciji od 4% učinkovit za povećanje rasta i imunološke učinke kod rakova P. monodon.The capability of a low-cost probiotic product was evaluated in Penaeus monodon reared under an outdoor culture system for a period of 90 days. The low-cost probiotic product was formulated by mixing the broth culture of Bacillus cereus along with commercial chalk powder in 1:1 ratio. The prepared probiotic product as a powdered supplement was incorporated along with the shrimp diet at various concentrations (1 to 5%). The viability of B. cereus in the experimental diet was tested once in 30 days up to 90 days during the experimentation. After 90 days of feeding experiments, the growth performance of shrimp was determined with a maximum production of 11.98 ± 0.367 g, SGR of 5.030 ± 0.171% and FCE of 86.63 ± 1.300% in P. monodon fed D4 diet (4% probiotic supplement). The immunological parameters determined at the end of the culture experiment showed variation among diets. The total haemocyte count (273.66 ± 3.09 x 105 cells/ml), phenoloxidase activity (0.132 ± 0.007 OD) and respiratory burst activity (0.291 ± 0.020 O.D) were high in P. monodon fed D4 diet. Similar effects were observed for plasma protein concentration (68.00 ± 1.41 mg/ ml), lysozyme activity (0.483 ± 0.014 U/ml) and bactericidal activity (81.0 ± 1.33%) in shrimp fed D4 diet. The results of the present investigation conclude that the probiotic product at a concentration of 4% was effective in elevating the growth and immune performances in shrimp P. monodon

Antifouling and toxic properties of the bioactive metabolites from the seagrasses Syringodium isoetifolium and Cymodocea serrulata

International audienceThe present study documents the antifouling and toxic properties of seagrasses Syringodium isoetifolium and Cymodocea serrulata. For that, the seagrasses S. isoetifolium and C. serrulata were extracted individually using organic solvents viz. dichloromethane, acetone and methanol. Amongst the extracts, the maximum antimicrofouling and antimacrofouling activities were exhibited by methanol extracts of both the seagrasses. The Minimal Inhibitory Concentration (MIC) of methanolic extracts of seagrasses was ranged from 1.0 to 10 mu g/ml against test biofilm bacteria and microalgal strains. Similarly, 100% fouling inhibition of limpet Patella vulgata was found at 6.0 mg/ml of methanolic extracts of seagrasses. The mussel Perna indica showed 50% of byssal production and attachment inhibition at 21.51 +/- 2.03, 17.82 +/- 1.07 mu g/ml and the anticrustaecean activity for 50% mortality of Anemia sauna was recorded at 732.14 +/- 921 and 394.16 +/- 5.16 mu g/ml respectively for methanolic extracts of S. isoetifolium and C serrulata. The minimal inhibitory and higher lethal concentrations of active methanol extracts shows it's less toxic nature. Based on the prolific results, methanol extracts of S. isoetifolium and C serrulata were subjected to purification using silica gel column and thin layer chromatography. Then the active compounds of the bioassay guided fractions were partially characterized using gas chromatography coupled with mass spectroscopy (GC-MS) and keyed out that fatty acids (C-16 to C-24) were the major components which responsible for the antifouling properties of the candidate seagrasse

Immunomodulatory effect of alginic acid from brown seaweed sargassum wightii on disease resistance in penaeus monodon

In recent days, bacterial diseases are very common among all the aquatic organisms. Particularly in grow out ponds they cause heavy loss in aquaculture. To overcome this problem, the use of antibiotics become a common practice, thus leads to the development of antibiotic resistant strains. In this regard, a new attempt has been made to study the immunostimulatory effect of alginic acid, a sulphated polysaccharide derived from brown seaweed Sargassum wightii, on the disease resistance in Penaeus monodon. The immunological parameters, such as Total Haemocyte Count (THC), Prophenoloxidase activity, Respiratory burst (NBT assay), Superoxide dismutase activity and Phagocytic activity were monitored after challenging against Vibrio parahaemolyticus. Before the challenge study, the test animals were fed with different concentrations (1,2,3 g/kg as A1, A2, A3) of alginic acid fed diet for 45 days. All the immunological parameters showed a significant increase with an increasing concentration of alginic acid, in the test animals compared to the control throughout the experimental study period

Deproteinization potential and antioxidant property of haloalkalophilic organic solvent tolerant protease from marine Bacillus sp. APCMST-RS3 using marine shell wastes

The current increase in the vast amount of marine crustacean shell waste produced by the fish processing industries has led to the need to find new methods for its disposal. Hence, the present study was carried out via marine shell wastes as substrate for protease production. The maximum production (4000.65 U/ml) from Bacillus sp. APCMST-RS3 was noticed in 3:1% shrimp and oyster shell powder (SOSP) as substrate. Purified protease showed 53.22% and 22.66% enzyme yield; 3.48 and 8.49 fold purity with 40 kDa molecular weight; whereas, its Km and Vmax values were 0.6666 g/l, 1111.11 U/ml. This enzyme showed optimum activity at pH 9 and 60 °C temperature. Also, it retained maximum protease activity in the presence of NaCl (2.5 M), surfactants (Tween 20, 40, 60, 80 and SDS) and metal ions (MnCl2, CaCl2, HgCl2 and BaCl2) and solvents. The candidate bacterium effectively deproteinized (84.35%) shrimp shell and its antioxidant potentials

Immunomodulatory and Growth-Promoting Potential of Lowcost Probiotic Product in Penaeus Monodon Culture System

The capability of a low-cost probiotic product was evaluated in Penaeus monodon reared under an outdoor culture system for a period of 90 days. The low-cost probiotic product was formulated by mixing the broth culture of Bacillus cereus along with commercial chalk powder in 1:1 ratio. The prepared probiotic product as a powdered supplement was incorporated along with the shrimp diet at various concentrations (1 to 5%). The viability of B. cereus in the experimental diet was tested once in 30 days up to 90 days during the experimentation. After 90 days of feeding experiments, the growth performance of shrimp was determined with a maximum production of 11.98 ± 0.367 g, SGR of 5.030 ± 0.171% and FCE of 86.63 ± 1.300% in P. monodon fed D4 diet (4% probiotic supplement). The immunological parameters determined at the end of the culture experiment showed variation among diets. The total haemocyte count (273.66 ± 3.09 x 105 cells/ml), phenoloxidase activity (0.132 ± 0.007 OD) and respiratory burst activity (0.291 ± 0.020 O.D) were high in P. monodon fed D4 diet. Similar effects were observed for plasma protein concentration (68.00 ± 1.41 mg/ ml), lysozyme activity (0.483 ± 0.014 U/ml) and bactericidal activity (81.0 ± 1.33%) in shrimp fed D4 diet. The results of the present investigation conclude that the probiotic product at a concentration of 4% was effective in elevating the growth and immune performances in shrimp P. monodon

Investigation of Lipase Production by Milk Isolate Serratia rubidaea

Production of extracellular lipase in submerged culture of Serratia rubidaea has been investigated. The lipase production was optimized in shake flask experiments. The observed pH and temperature range optimum for maximum lipase production were 7–8 and 30–40 °C, respectively. With a selected nitrogen source, casein ((6.5±0.015) U/mL) and soytone ((9.4±0.02) U/mL) were suitable substrates for accelerating lipase production. The optimized concentration of casein and soytone was 24 g/L ((9.95±0.02) U/mL) and 5 g/L ((14.8±0.03) U/mL), respectively. The effect of carbon source on lipase production indicated that starch was suitable substrate to maximize lipase production ((15.60±0.20) U/mL) and the optimum concentration registered was 4 g/L ((17.46±0.20) U/mL). Investigating the effect of lipids and surfactants showed that the gingily oil ((20.52±0.20) U/mL) and Tween 20 ((27.10±0.01) U/mL) were suitable substrates for maximizing lipase production, and the optimum concentrations registered were 15 mL/L ((23.15±0.24) U/mL) and 6 mL/L ((34.20±0.01) U/mL), respectively. Partial purification of lipase indicated that the molecular mass of partially purified enzyme was 54 kDa

Influence of selected Indian immunostimulant herbs against white spot syndrome virus (WSSV) infection in black tiger shrimp, Penaeus monodon with reference to haematological, biochemical and immunological changes

Immunostimulants are the substances, which enhance the non-specific defence mechanism and provide resistance against the invading pathogenic micro-organism. In order to increase the immunity of shrimps against the WSSV, the methanolic extracts of five different herbal medicinal plants like Cyanodon dactylon, Aegle marmelos, Tinospora cordifolia, Picrorhiza kurooa and Eclipta alba were selected and mixed thoroughly in equal proportion. The mixed extract was supplemented with various concentrations viz. 100 (A), 200 (B), 400 (C), and 800 (D) mg kg(-1) through artificial diets individually. The prepared diets (A-D) were fed individually to WSSV free healthy shrimp Penaeus monodon with an average weight of 8.0 +/- 0.5 g for 25 days. Control diet (E), devoid of herbal extract was also fed to shrimps simultaneously. After 25 days of feeding experiment, the shrimps were challenged with WSSV, which were isolated and propagated from the infected crustaceans. The shrimps succumbed to death within 7 days when fed on no herbal immunostimulant diet (E). Among the different concentrations of herbal immunostimulant supplemented diets, the shrimps fed on diet D (800 mg kg(-1)) significantly (P < 0.0001) had more survival (74%) and reduction in the viral load. Also the better performance of haematological, biochemical and immunological parameters was found in the immunostimulant incorporated diets fed shrimps. The present work revealed that the application of herbal immunostimulants will be effective against shrimp viral pathogenesis and they can be recommended for shrimp culture. (c) 2006 Published by Elsevier Ltd.Immunostimulants are the substances, which enhance the non-specific defence mechanism and provide resistance against the invading pathogenic micro-organism. In order to increase the immunity of shrimps against the WSSV, the methanolic extracts of five different herbal medicinal plants like Cyanodon dactylon, Aegle marmelos, Tinospora cordifolia, Picrorhiza kurooa and Eclipta alba were selected and mixed thoroughly in equal proportion. The mixed extract was supplemented with various concentrations viz. 100 (A), 200 (B), 400 (C), and 800 (D) mg kg(-1) through artificial diets individually. The prepared diets (A-D) were fed individually to WSSV free healthy shrimp Penaeus monodon with an average weight of 8.0 +/- 0.5 g for 25 days. Control diet (E), devoid of herbal extract was also fed to shrimps simultaneously. After 25 days of feeding experiment, the shrimps were challenged with WSSV, which were isolated and propagated from the infected crustaceans. The shrimps succumbed to death within 7 days when fed on no herbal immunostimulant diet (E). Among the different concentrations of herbal immunostimulant supplemented diets, the shrimps fed on diet D (800 mg kg(-1)) significantly (P < 0.0001) had more survival (74%) and reduction in the viral load. Also the better performance of haematological, biochemical and immunological parameters was found in the immunostimulant incorporated diets fed shrimps. The present work revealed that the application of herbal immunostimulants will be effective against shrimp viral pathogenesis and they can be recommended for shrimp culture. (c) 2006 Published by Elsevier Ltd