CN103497907A - Application of bacillus altitudinis in prawn culture - Google Patents
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Abstract
The invention discloses an application of bacillus altitudinis in the prawn culture, and relates to the aqua culture. The preservation number of the bacillus altitudinis 1A08373 is CCTCC No. M2013292. The sediment from the surface layer in the deep India Ocean is separated to prepare a bacillus altitudinis preparation, wherein the bacillus altitudinis preparation comprises one component selected from the following components: a pathogenic vibrio bacteriostatic preparation, a prawn growth promoter, a prawn feed addictive, and a prawn culture water modifier. The number of active bacterium contained in the prepared bacillus altitudinis preparation is not less than 1*109 CFU/mL, and the maximum number can reach 1*1010 CFU/mL or more. The bacillus altitudinis has the functions of increasing the enzyme activity of the digestive tract of prawns, promoting the growth of prawns, improving the disease resistant performance of prawns, and reducing the occurrence of prawn diseases, and especially has a very good effect on prevention and treatment of prawn vibrio diseases and increasing of prawn output.
Description
Technical field
The present invention relates to aquaculture, especially relate to a kind of preparation that is applied to prawn culturing, the preparation of specifically being made by the high-altitude genus bacillus is suppressing vibrios, promotes the prawn growth, strengthens the prawn disease resistance, the application in the control vibriosis penaeus.
Background technology
The fast development of intensive, high-density aquiculture, be seriously damaged the breeding ecological environment, and aquiculture disease becomes increasingly complex, and sickness rate is more and more higher, endangers more and more seriously, and the usage quantity of antibiotic feed additive increases day by day.2010,7,640,000 hectares of national aquaculture areas, wherein had 240,000 hectares to suffer disease, nearly 300,000 tons of production loss.In 95% cultured area, in order to prevent disease, occur, in production blindly, throw in all kinds of microbiotic on a large scale, as paraxin, terramycin, Nifurazolidone etc.Paraxin directly is added to the water, and the terramycin mix fodder is thrown something and fed or directly splashed aquaculture water.Within 2000, antibiotic dosage is about 16200 tons, and approximately 70% for animal husbandry and aquaculture.Because the problems such as drug abuse, drug residue and bacterial drug resistance are on the rise, caused the concern of people to food and environmental safety.Finding a kind of novel, safe, green Substitutes For Antibiotic becomes one of focus of current aquatic products research, and wherein probiotic bacterium has the effects such as antibacterial, growth promotion, raising immunity and day by day comes into one's own because of it.
Probiotics is to reach the order ground of control aquatic animal disease by " with bacterium, controlling bacterium ".On the one hand, produce meta-bolites and physiologically active substance by beneficial microorganism and suppress or kill pathogenic bacteria, simultaneously in aquaculture water and cultivated animals enteron aisle with pathogenic bacteria competition nutrition, occupy ecological niche, form dominant microflora, reach the purpose of preventing and treating disease.On the other hand, beneficial microorganism, by producing multiple digestive ferment, helps the metabolism of protein, carbohydrate, fat, improves utilization ratio and the transformation efficiency of feed, reaches the purpose that reduces resource consumption and volume increase.
Chinese patent CN101993843A discloses a kind of Bacillus firmus strain and application thereof.Bacillus firmus (Bacillus firmus) SSL065 bacterial strain, deposit number is: CCTCC NO:M2010192.Bacillus firmus SSL065 bacterium powder prepared by this bacterial strain, can be used for strengthening the immune level of prawn cell and body fluid.The bacillus firmus SSL065 bacterial strain screening of this invention is from the prawn digestive tube, it is a kind of endogenic bacterial strain, there is obvious raising cultured prawn innate immune activity, greatly strengthen cultured prawn anti-microbial pathogen infectivity, particularly can significantly improve prawn and resist the various biological functions such as virus causing disease infection ability.
Chinese patent CN102586144A discloses a kind of bacillus pumilus, probiotics preparation and its preparation method and application.Bacillus pumilus (Bacillus pumilus) LV149 is preserved in Chinese Typical Representative culture collection center (CCTCC), deposit number: CCTCC NO:M2011411 on November 23rd, 2011.The bacillus pumilus of this invention (Bacillus pumilus) LV149 has strong extracellular protease, lipase, amylase activity, vibrios is had to inhibition widely, and do not have hemolytic activity.Using this bacterium as fermentation strain, through solid state fermentation, drying and crushing makes usings the bacillus pumilus probiotics preparation of bacillus pumilus (Bacillus pumilus) LV149 as activeconstituents, it is added in prawn feed, the prawn of feeding, not only can play and suppress the growth of gut of shrimp pathogenic vibrio, reduce the generation of vibriosis penaeus, promote the prawn growth simultaneously, reduce feed coefficient, improve commercial quality, Shortening culturing period, thereby reduce the cultivation risk, reduce aquaculture cost, and biological safety is high, in aquaculture, be with a wide range of applications.
Summary of the invention
The object of the present invention is to provide and a kind ofly for the vibriosis penaeus evil, there is better prevention effect, and can promote the high-altitude bacillus altitudinis1A08373 of prawn growth.
Another object of the present invention is to provide the application of high-altitude genus bacillus in prawn culturing.
Described high-altitude bacillus altitudinis1A08373 has been preserved in Chinese Typical Representative culture collection center, address: China, Wuhan, Wuhan University on 06 25th, 2013; Deposit number: CCTCC NO:M2013292.
High-altitude bacillus altitudinis1A08373 separates and obtains from Indian Ocean deep sea surface layer deposit sediment, and high-altitude bacillus altitudinis1A08373 has following characteristics:
1, morphological specificity: high-altitude bacillus altitudinis1A08373CCTCC NO:M2013292 is shaft-like, and size is (0.3～0.6) μ m * (1.2～1.9) μ m, and Gram-positive, can move, and the adnation flagellum is arranged, and produces gemma.Cultivate 48h for 28 ℃ on the 2216E solid medium, bacterium colony is white in color opaque, and smooth surface is moistening, regular edges, and the colony diameter size is about 2～3mm.
2, physiological and biochemical property: the strain growth salinity range is 0～15%, more than 18%, does not grow, and the suitableeest scope is 1%～5%; Growth pH scope is 5～12, and the suitableeest scope is 6～8.Oxydase (+), catalase (+), nitrate reduction (-), indole test (-), unfermentable D-Glucose produces acid, gelatine liquefication (+), urease (-), beta-glucosidase (+), beta-galactosidase enzymes (+), D-Glucose, L-arabinose, D-MANNOSE, PEARLITOL 25C, N-acetyl-glucosamine, oxysuccinic acid, citric acid can be utilized, maltose, capric acid, hexanodioic acid, toluylic acid can not be utilized.
API ZYM result shows, high-altitude bacillus altitudinis1A08373 has alkaline phosphatase, esterase, lipoid esterase, leucine aminopeptidase(LAP), alpha-chymotrypsin, acid phosphatase, β-Portugal (grape) Glycosylase, alpha-Mannosidase, do not have the enzymic activitys such as lipase, α-amino-isovaleric acid aminopeptidase, Gelucystine aminopeptidase, trypsinase, alpha-galactosidase, beta-galactosidase enzymes, β-Portugal (grape) glycuronide enzyme, α-Portugal (grape) Glycosylase, N-acetyl-glucosaminidase, Alpha-Fucosidase.
3,16S rRNA, pyrE and aroE gene sequencing: according to the genomic dna of ordinary method extracting high-altitude genus bacillus (Bacillus altitudinis) 1A08373, and 16S rRNA, pyrE and aroE gene order are measured, 16S rRNA, the pyrE of high-altitude genus bacillus 1A08373 and aroE gene order are as shown in SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3.
Through the BLAST compare of analysis, the 16S rDNA sequence homology of high-altitude bacillus altitudinis1A08373 and Bacillusaltitudini41KF2b (T) is 99.93%, with the 16S rDNA sequence homology of B.pumilus ATCC7061 (T) be 99.67%, with the 16S rDNA sequence homology of B.safensis FO-036b (T) be 99.54%.The comparison result of pyrE gene order shows, the homology of high-altitude bacillus altitudinis1A08373 and B.altitudini41KF2b (T), B.pumilus ATCC7061 (T) and B.safensis FO-036b (T) three strain type species is respectively 98.53%, 85.53% and 83.70%.The comparison result of aroE gene order shows, the homology of high-altitude bacillus altitudinis1A08373 and B.altitudini41KF2b (T), B.pumilus ATCC7061 (T) and B.safensis FO-036b (T) three strain type species is respectively 98.53%, 85.89% and 86.67%.
Morphological specificity, physio-biochemical characteristics and 16S rRNA, pyrE and the aroE gene sequencing result of comprehensive bacterial strain, bacterial strain 1A08373 of the present invention is accredited as the high-altitude genus bacillus, is specially high-altitude genus bacillus 1A08373(CCTCC NO:M2013292).
High-altitude of the present invention genus bacillus (Bacillus altitudinis) 1A08373 has significant fungistatic effect to kinds of pathogenic vibrio, and the prawn growth is had to obvious promoter action.High-altitude bacillus altitudinis1A08373 of the present invention can produce proteolytic enzyme, lipase and cellulase etc., and has stronger activity.
High-altitude bacillus altitudinis1A08373 of the present invention has inhibition widely to Vibrio harveyi SF1, Kan Beishi vibrios VH1, Vibrio anguillarum W1, Vibrio parahaemolyticus 1937, the prawn kinds of pathogenic vibrio such as Ou Wenshi vibrios VH2, Vibrio natriegen FS-1.
High-altitude bacillus altitudinis1A08373 of the present invention is improving the prawn disease resistance, is promoting, aspect the prawn growth, unusual effect is arranged.In the experiment with pathogenic Kan Beishi vibrios VH1 immersion infection prawn, find, the experimental group prawn survival rate of adding high-altitude genus bacillus 1A08373 is significantly higher than control group, has improved 32%.High-altitude genus bacillus 1A08373 is carried out to liquid fermenting, with 10
7the viable bacteria content of the every g feed of cfu/g() concentration is added in feed the Penaeus vannamei of throwing something and feeding, simultaneously with 10
5the viable bacteria content of every milliliter of seawater of cfu/mL() concentration is splashed to aquaculture water, found that, raising through 30 days, the experimental group of having added high-altitude genus bacillus 1A08373 feed of throwing something and feeding is compared with the control group that does not add probiotic bacterium, prawn mean body weight rate of increase has improved 18.7%, and the gut of shrimp trypsinase of probiotic bacterium 1A08373 experimental group and lipase all are significantly higher than control group (P<0.05).Vibrios content the dynamic monitor result shows, in the breeding seawater of 1A08373 experimental group, vibrios content is lower than control group; Bacterial strain 1A08373 can be survived in breeding seawater and gut of shrimp, and the viable bacteria content detected in breeding seawater is 10
4more than cfu/mL, in gut of shrimp, genus bacillus content is 10
4more than cfu/g.
Described high-altitude bacillus altitudinis1A08373 can be used for preparing the high-altitude bacillus preparation, and described high-altitude bacillus preparation comprises a kind of in kinds of pathogenic vibrio inhibiting-bacteria preparation, prawn growth stimulant, prawn feed additive, prawn culturing improver of water quality.
The preparation method's of described high-altitude bacillus preparation concrete steps are: the order of high-altitude genus bacillus (Bacillus altitudinis) 1A08373 being pressed to inclined-plane, liquid seeds, liquid fermenting is cultivated, and inclined-plane and seed culture medium are conventional nutrient agar or broth culture; Fermentative medium formula (W/V): soybean cake powder 3%, starch 1.5%, corn steep liquor 1%, glucose 0.5%, (NH
4)
2 sO
41%, NH
4cl0.1%, KH
2pO
40.05%, NaH
2pO
40.5%, MgSO
40.05%, MnSO
40.05%.
The viable count that prepared high-altitude bacillus preparation comprises is not less than 1 * 10
9cFU/mL, can reach 1 * 10
10more than CFU/mL.
Described high-altitude bacillus preparation is pressed 0.5～5% of prawn feed weight and is added.
The described high-altitude inoculum density of bacillus preparation in aquaculture water is 10
3～10
4cfu/mL.
High-altitude of the present invention genus bacillus (Bacillus altitudinis) 1A08373 has extensive inhibition to the aquaculture pathogenic vibrio.Using high-altitude genus bacillus 1A08373 fermented liquid as active bacteria formulation, splash to aquaculture water/add in the feed prawn of feeding, can play the breeding that suppresses pathogenic vibrio in aquaculture water, improving the prawn digestive enzyme lives, promote the prawn growth, improve the prawn disease resistance, reduce shrimp disease and occur, particularly for the control of vibriosis penaeus evil and the increase of prawn output, there is good effect.
The accompanying drawing explanation
The extracellular enzyme biopsy mapping that Fig. 1 is high-altitude bacillus altitudinis1A08373.1, proteolytic enzyme; 2, lipase; 3, cellulase.
Fig. 2 is high-altitude bacillus altitudinis1A08373 bacteriostatic activity detection figure.1, indicator: Kan Beishi vibrios VH1; 2, indicator: Vibrio harveyi SF1.
Fig. 3 is the inhibition of high-altitude bacillus altitudinis1A08373 fermented supernatant fluid to Kan Beishi vibrios VH1 growth.In Fig. 3, ◆ be contrast: VH1, ■ is for processing: VH1+1A08373.
The dynamic monitoring that Fig. 4 is vibrios content in aquaculture water.In Fig. 4, mark ◆ be 1A08373, ■ is blank.
Embodiment
The present invention is further illustrated in connection with accompanying drawing for following examples, but therefore do not limit the present invention among described scope of embodiments.Method in following embodiment, if no special instructions, be ordinary method.
The separation of embodiment 1 high-altitude genus bacillus, evaluation and antimicrobial spectrum are measured
(1) strains separation: get Indian Ocean pelagic deposit matter sample 1g in aseptic super clean bench, add 9ml stroke-physiological saline solution (0.9%NaCl) to make suspension, successively 10 times of gradient dilutions to 10
-2, 10
-3, 10
-4, get gradient dilution liquid 100 μ L coating Marine Agar solid mediums (2216E substratum), after 28 ℃ of cultivation 48h, picking list bacterium colony line purifying, separate and obtain bacterial strain BS1, preservation is to Chinese Sea microbial strains preservation administrative center (MCCC), and deposit number is MCCC1A08373.
(2) physiological and biochemical property of bacterial strain and Molecular Identification
Morphological specificity: high-altitude genus bacillus (Bacillus altitudinis) 1A08373CCTCC NO:M2013292 is shaft-like, size is (0.3～0.6) μ m * (1.2～1.9) μ m, and Gram-positive, can move, the adnation flagellum is arranged, produce gemma.Cultivate 48h for 28 ℃ on the 2216E solid medium, bacterium colony is white in color opaque, and smooth surface is moistening, regular edges, and the colony diameter size is about 2～3mm.
Physiological and biochemical property: the strain growth salinity range is 0～15%, more than 18%, does not grow, and the suitableeest scope is 1%～5%; Growth pH scope is 5～12, and the suitableeest scope is 6～8.Oxydase (+), catalase (+), nitrate reduction (-), indole test (-), V.P tests (+), unfermentable D-Glucose produces acid, gelatine liquefication (+), urease (-), beta-glucosidase (+), beta-galactosidase enzymes (+), can utilize D-Glucose, L-arabinose, D-MANNOSE, PEARLITOL 25C, N-acetyl-glucosamine, oxysuccinic acid, citric acid, can not utilize maltose, capric acid, hexanodioic acid, toluylic acid.Bacterial strain 1A08373 and type species Bacillus altitudini41KF2b (T) some physiological-biochemical characteristics are in Table 1.
API ZYM result shows, bacterial strain 1A08373 has alkaline phosphatase, esterase, lipoid esterase, leucine aminopeptidase(LAP), alpha-chymotrypsin, acid phosphatase, β-Portugal (grape) Glycosylase, alpha-Mannosidase, do not have the enzymic activitys such as lipase, α-amino-isovaleric acid aminopeptidase, Gelucystine aminopeptidase, trypsinase, alpha-galactosidase, beta-galactosidase enzymes, β-Portugal (grape) glycuronide enzyme, α-Portugal (grape) Glycosylase, N-acetyl-glucosaminidase, Alpha-Fucosidase.
Molecular Identification: according to the genomic dna of ordinary method extracting high-altitude genus bacillus (Bacillus altitudini) 1A08373, and 16S rRNA, pyrE and aroE gene order are measured, 16S rDNA, the pyrE of high-altitude genus bacillus 1A08373 and aroE gene order are as shown in SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3.
Through the BLAST compare of analysis, the 16S rDNA sequence homology of high-altitude genus bacillus (Bacillus altitudini) 1A08373 and Bacillusaltitudini41KF2b (T) is 99.93%, with the 16S rDNA sequence homology of B.pumilus ATCC7061 (T) be 99.67%, with the 16S rDNA sequence homology of B.safensis FO-036b (T) be 99.54%.The comparison result of pyrE gene order shows, the homology of 1A08373 and B.altitudini41KF2b (T), B.pumilus ATCC7061 (T) and B.safensis FO-036b (T) three strain type species is respectively 98.53%, 85.53% and 83.70%.The comparison result of aroE gene order shows, the homology of 1A08373 and B.altitudini41KF2b (T), B.pumilus ATCC7061 (T) and B.safensis FO-036b (T) three strain type species is respectively 98.53%, 85.89% and 86.67%.
Morphological specificity, physio-biochemical characteristics and 16S rRNA, pyrE and the aroE gene sequencing result of comprehensive bacterial strain, bacterial strain 1A08373 of the present invention is accredited as the high-altitude genus bacillus, be preserved in Chinese Typical Representative culture collection center (CCTCC), deposit number is respectively CCTCC NO:M2013292.16S rDNA, the pyrE of high-altitude genus bacillus 1A08373 and aroE gene gene order are as SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3.
Table 1: the physio-biochemical characteristics of husky good fortune genus bacillus 1A07588
Annotate: "+" means that positive reaction maybe can utilize; "-", the expression negative reaction maybe can not be utilized
(3) producing enzyme and bacteriostatic activity detects
The extracellular enzyme biopsy is surveyed:
Husky high-altitude genus bacillus 1A08373 is carried out to extracellular protease, amylase, lipase and cellulase activity and detect, found that, high-altitude genus bacillus 1A08373 has extracellular protease, lipase and cellulase activity, but does not have amylase activity.
Bacteriostatic activity detects:
Adopt lysoplate assay to be measured bacteriostatic activity and the antimicrobial spectrum of high-altitude genus bacillus 1A08373.Used medium is seawater 216L substratum, and indicator comprises that kinds of pathogenic vibrio is as Vibrio harveyi SF1, Kan Beishi vibrios VH1, Vibrio anguillarum W1, Vibrio parahaemolyticus 1937, Ou Wenshi vibrios VH2, Vibrio natriegen FS-1 and micrococcus luteus CGMCC1.2299, streptococcus aureus FDA209, intestinal bacteria CGMCC1.2389, Aeromonas hydrophila SHI and Elastolyticenzyme of pseudomonas aeruginosa 1A06466.Indicator and tested bacterium be incubated overnight in the 216L substratum respectively, gets appropriate indicator and is diluted to 10 by stroke-physiological saline solution
7cfu/ml, draw 100 μ L coating 216L flat boards, and on flat board, punching (aperture 6mm) adds tested fermented liquid 50 μ L in hole, cultivates 24h for 30 ℃, and observations, use the vernier caliper measurement antibacterial circle diameter.Result shows, high-altitude genus bacillus 1A08373 all has restraining effect in various degree to 11 strain indicators, and result is as shown in table 2.
High-altitude genus bacillus 1A08373 Extracellular enzyme activity detection figure and bacteriostatic activity are shown in Fig. 1 and 2.
Table 2: the bacteriostatic activity of bacterial strain 1A08373
Annotate: 1～5 is kinds of pathogenic vibrio, Penaeus vannamei is had stronger pathogenic.In form, numerical value represents antibacterial circle diameter (mm), and numerical value is larger, and fungistatic effect is stronger.-, mean not observe inhibition zone.
The inhibition of embodiment 2 high-altitude genus bacillus 1A08373 fermented supernatant fluids to the vibrios growth
By high-altitude genus bacillus 1A08373 and indicator Kan Beishi vibrios VH1 respectively in the 216L liquid nutrient medium 28 ℃ cultivate 24h.Then get 0.22um membrane filtration degerming for the 1A08373 fermented liquid, get the 1mL filtering supernatant and add in the 216L substratum that 4mL is fresh, then inoculate the indicator VH1 of 50 μ l in test tube.Inoculate indicator VH1(50 μ l simultaneously) in the 5mL216L liquid nutrient medium in contrast.Every group of three of test is parallel.Respectively in the growth of 3,6,9,12,15,18,21,24h sampling and measuring indicator VH1 (with OD
600mean).As shown in Figure 3, the experimental group indicator VH1 growth of adding the 1A08373 fermented supernatant fluid obviously is weaker than control group to experimental result, illustrates that growth has restraining effect to 1A08373 to vibrios VH1.The biological assay of embodiment 3 high-altitude genus bacillus 1A08373 to the Penaeus vannamei disease resistance
By high-altitude genus bacillus 1A08373 and indicator Kan Beishi vibrios VH1 respectively in the 216L liquid nutrient medium 28 ℃ cultivate 24h.Get each fermented liquid centrifugal 5min under 5000rpm, remove supernatant liquor, collect thalline, by stroke-physiological saline solution, be prepared into bacteria suspension, measure thalline OD
600, according to experiment institute expense, add in aquaculture water.
Experiment is carried out in the 30L aquarium, puts into the 10L fresh seawater in each aquarium and 50 tail Penaeus vannamei seedling PL-40(body weight are about 0.1g), guarantee enough inflations.Experiment arranges blank group (not adding any bacterium in aquaculture water), and control group (inoculates 10 in aquaculture water
6the Kan Beishi vibrios VH1 of cfu/ml) and experimental group (in aquaculture water, inoculate 10 simultaneously
6cfu/ml isocyatic high-altitude genus bacillus and Kan Beishi vibrios VH1).Experimental session does not change water, adds up the prawn death condition every day.Result is as shown in table 3, and through the observation of 7 days, result showed, high-altitude genus bacillus 1A08373 can significantly improve the prawn survival rate, reduces the mortality ratio of vibrio infection prawn.
Table 3: the high-altitude genus bacillus infects the provide protection of prawn to Kan Beishi vibrios VH1
The impact of embodiment 4 high-altitude genus bacillus 1A08373 on the Penaeus vannamei growth
The cultivation of high-altitude genus bacillus: the order of starting strain high-altitude genus bacillus (Bacillus altitudini1A08373) deposit number CCTCC NO:M2013292 being pressed to inclined-plane, liquid seeds, liquid fermenting is cultivated, and inclined-plane and seed culture medium are conventional nutrient agar or broth culture; Fermentative medium formula (W/V): soybean cake powder 3%, starch 1.5%, corn steep liquor 1%, glucose 0.5%, (NH
4)
2 sO
41%, NH
4cl0.1%, KH
2pO
40.05%, NaH
2pO
40.5%, MgSO
40.05%, MnSO
40.05%.After the high-altitude fermentation of bacillus liquid of above-mentioned acquisition is carried out to 10 times of gradient dilutions with physiological saline, get 100 μ L coating MA flat boards, cultivate 24h for 28 ℃, calculating viable bacteria content is 7.2 * 10
9cfu/mL.
Test Penaeus vannamei seedling used and buy from tall building, Xiamen gold dragon company of growing seedlings, the long 0.8cm of body left and right, support in laboratory temporarily to about the long 4cm of body, is transferred to culturing pool about body weight 1.3g and carries out experiment.The culturing pool volume is 500L, and effectively water body is 375L, and each culturing pool is put prawn 100 tails in a suitable place to breed.Experiment is divided into control group and experimental group, and three every group parallel.The prawn commercial feed (No. 2 feeds of " lucky star " board Penaeus vannamei) that the experimental group prawn is thrown something and fed and is added with the high-altitude bacillus preparation every day, control group is is only thrown something and fed and is not added the above-mentioned prawn commercial feed of bacterium; Within every 7 days, in the experimental group water body, splash 10 simultaneously
5the high-altitude genus bacillus of cfu/mL.Experimental group prawn feed preparation method: get the above-mentioned high-altitude of 10mL fermentation of bacillus liquid, by stroke-physiological saline solution, suitably after dilution, evenly be sprayed onto 500g feed surface, stir, guarantee in feed that viable bacteria concentration is 1 * 10
7cfu/g.Test 30 days by a definite date, the feed of throwing something and feeding every day 2 times, change water 20% every day.Add up prawn body weight gain amount after 30 days, measure intestinal protease, amylase and lipase activity and detect water body and enteron aisle genus bacillus number, vibrios number etc. simultaneously.
Table 4: the impact of high-altitude genus bacillus 1A08373 on the Penaeus vannamei growth
Annotate: * relative body weight rate of increase=(final body weight-initial body weight)/initial body weight.The different letters of subscript represent significant difference (P<0.05)
Table 5: the impact of high-altitude genus bacillus 1A08373 on the Penaeus vannamei Activity of Digestive Enzymes in Intestine
From table 4 and table 5, through 30 days feed, result showed, high-altitude genus bacillus 1A08373 can significantly improve the mean body weight increment of prawn, than control group, has improved 18.7%.In addition, the gut of shrimp trypsinase of 1A08373 probiotic bacterium treatment group and enteron aisle lipase are significantly higher than control group (P<0.05).In addition, in 1A08373 experimental group water body genus bacillus content 10
4more than cfu/mL, gut of shrimp genus bacillus number is 10
4more than cfu/g.Water vibrios content the dynamic monitor result shows (Fig. 4), and in the experimental group aquaculture water, vibrios content, far below control group, illustrates that high-altitude genus bacillus 1A08373 has the effect that suppresses the water vibrios breeding.To sum up, high-altitude genus bacillus 1A08373 has the effect that promotes the prawn growth, suppresses vibrios.
According to embodiment 2, bacterial strain 1,A08,373 28 ℃ of cultivation 24h in the 216L liquid nutrient medium are prepared to bacterium liquid.Get SPF level KM mouse, body weight 18～22g, each 10 of male and female, carry out gavage with bacterium liquid, every each gavage 0.4ml, in 24h, gavage is 5 times, and 2ml/ is only altogether.Continuous Observation 7 days, weigh on the 7th day.Result shows, abnormal conditions all do not appear in experiment mice, and body weight gain is normal.Illustrate that high-altitude genus bacillus 1A08373 is safety non-toxic.
Claims (4)
1. the high-altitude genus bacillus, is characterized in that for high-altitude bacillus altitudinis1A08373, on 06 25th, 2013, has been preserved in Chinese Typical Representative culture collection center, address: China, Wuhan, Wuhan University; Deposit number: CCTCC NO:M2013292.
2. the application of high-altitude genus bacillus in the bacillus preparation of preparation high-altitude as claimed in claim 1, described high-altitude genus bacillus is high-altitude bacillus altitudinis1A08373.
3. application as claimed in claim 2, is characterized in that described high-altitude bacillus preparation comprises a kind of in kinds of pathogenic vibrio inhibiting-bacteria preparation, prawn growth stimulant, prawn feed additive, prawn culturing improver of water quality.
4. application as claimed in claim 2, the concrete steps that it is characterized in that the preparation method of described high-altitude bacillus preparation are: the order of high-altitude bacillus altitudinis1A08373 being pressed to inclined-plane, liquid seeds, liquid fermenting is cultivated, and inclined-plane and seed culture medium are conventional nutrient agar or broth culture; Fermentative medium formula, W/V: soybean cake powder 3%, starch 1.5%, corn steep liquor 1%, glucose 0.5%, (NH
4)
2sO
41%, NH
4cl0.1%, KH
2pO
40.05%, NaH
2pO
40.5%, MgSO
40.05%, MnSO
40.05%.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108740289A (en) * | 2018-08-08 | 2018-11-06 | 广东绿百多生物开发有限公司 | A kind of fermented feed applied to prawn culturing |
| CN109055251A (en) * | 2017-10-31 | 2018-12-21 | 上海大学 | A kind of bacillus and its application |
| CN109880767A (en) * | 2019-03-22 | 2019-06-14 | 中国海洋大学 | A strain used to prevent or treat vibriosis in prawns |
| CN112662582A (en) * | 2020-12-28 | 2021-04-16 | 海南大学 | Marine bacterium HNB 15 for producing cellulase, microbial preparation thereof and degradation method of natural cellulose |
| CN112725226A (en) * | 2020-12-28 | 2021-04-30 | 海南大学 | Screening of cellulase-producing bacterium C18 and natural cellulose degradation method of microbial preparation thereof |
| CN114075519A (en) * | 2020-08-14 | 2022-02-22 | 广东百时康生物科技有限公司 | Paenibacillus graveolens and application thereof in prawn culture and medicine preparation |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106754559A (en) * | 2017-02-06 | 2017-05-31 | 青岛农业大学 | A kind of probiotics preparation method for effectively purifying water |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109055251A (en) * | 2017-10-31 | 2018-12-21 | 上海大学 | A kind of bacillus and its application |
| CN109055251B (en) * | 2017-10-31 | 2021-08-10 | 上海大学 | Bacillus and application thereof |
| CN108740289A (en) * | 2018-08-08 | 2018-11-06 | 广东绿百多生物开发有限公司 | A kind of fermented feed applied to prawn culturing |
| CN109880767A (en) * | 2019-03-22 | 2019-06-14 | 中国海洋大学 | A strain used to prevent or treat vibriosis in prawns |
| CN114075519A (en) * | 2020-08-14 | 2022-02-22 | 广东百时康生物科技有限公司 | Paenibacillus graveolens and application thereof in prawn culture and medicine preparation |
| CN114075519B (en) * | 2020-08-14 | 2023-08-08 | 广东百时康生物科技有限公司 | Bacillus megatherium tomb and application thereof in prawn culture and preparation of medicines |
| CN112662582A (en) * | 2020-12-28 | 2021-04-16 | 海南大学 | Marine bacterium HNB 15 for producing cellulase, microbial preparation thereof and degradation method of natural cellulose |
| CN112725226A (en) * | 2020-12-28 | 2021-04-30 | 海南大学 | Screening of cellulase-producing bacterium C18 and natural cellulose degradation method of microbial preparation thereof |
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