CN114391494B - Screening method of comprehensive stress-resistant buried bivalve shellfish - Google Patents
Screening method of comprehensive stress-resistant buried bivalve shellfish Download PDFInfo
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- 238000012216 screening Methods 0.000 title claims abstract description 74
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- 239000004576 sand Substances 0.000 claims abstract description 44
- 230000004060 metabolic process Effects 0.000 claims abstract description 42
- 230000000241 respiratory effect Effects 0.000 claims abstract description 40
- 238000011084 recovery Methods 0.000 claims abstract description 24
- 230000004083 survival effect Effects 0.000 claims abstract description 19
- 238000012544 monitoring process Methods 0.000 claims abstract description 18
- 230000037328 acute stress Effects 0.000 claims abstract description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 66
- 239000013535 sea water Substances 0.000 claims description 31
- 238000002474 experimental method Methods 0.000 claims description 25
- 230000037406 food intake Effects 0.000 claims description 19
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- 241000195649 Chlorella <Chlorellales> Species 0.000 claims description 13
- 238000011010 flushing procedure Methods 0.000 claims description 13
- 239000011521 glass Substances 0.000 claims description 12
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims description 10
- 239000001301 oxygen Substances 0.000 claims description 10
- 229910052760 oxygen Inorganic materials 0.000 claims description 10
- 241000894006 Bacteria Species 0.000 claims description 8
- 229940041514 candida albicans extract Drugs 0.000 claims description 7
- 230000007613 environmental effect Effects 0.000 claims description 7
- 230000001546 nitrifying effect Effects 0.000 claims description 7
- 239000000843 powder Substances 0.000 claims description 7
- 238000002791 soaking Methods 0.000 claims description 7
- 239000012138 yeast extract Substances 0.000 claims description 7
- 238000009395 breeding Methods 0.000 claims description 6
- 230000001488 breeding effect Effects 0.000 claims description 6
- 206010020880 Hypertrophy Diseases 0.000 claims description 5
- JVMRPSJZNHXORP-UHFFFAOYSA-N ON=O.ON=O.ON=O.N Chemical compound ON=O.ON=O.ON=O.N JVMRPSJZNHXORP-UHFFFAOYSA-N 0.000 claims description 5
- UCKMPCXJQFINFW-UHFFFAOYSA-N Sulphide Chemical compound [S-2] UCKMPCXJQFINFW-UHFFFAOYSA-N 0.000 claims description 5
- XKMRRTOUMJRJIA-UHFFFAOYSA-N ammonia nh3 Chemical compound N.N XKMRRTOUMJRJIA-UHFFFAOYSA-N 0.000 claims description 5
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- 230000001737 promoting effect Effects 0.000 abstract description 2
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- 241000237519 Bivalvia Species 0.000 description 71
- 241001339782 Scapharca broughtonii Species 0.000 description 32
- 230000029058 respiratory gaseous exchange Effects 0.000 description 14
- 238000012360 testing method Methods 0.000 description 12
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- 239000013505 freshwater Substances 0.000 description 8
- ZKQDCIXGCQPQNV-UHFFFAOYSA-N Calcium hypochlorite Chemical group [Ca+2].Cl[O-].Cl[O-] ZKQDCIXGCQPQNV-UHFFFAOYSA-N 0.000 description 7
- 239000007844 bleaching agent Substances 0.000 description 7
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- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/50—Culture of aquatic animals of shellfish
- A01K61/54—Culture of aquatic animals of shellfish of bivalves, e.g. oysters or mussels
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/90—Sorting, grading, counting or marking live aquatic animals, e.g. sex determination
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
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- Life Sciences & Earth Sciences (AREA)
- Environmental Sciences (AREA)
- Zoology (AREA)
- Marine Sciences & Fisheries (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Farming Of Fish And Shellfish (AREA)
Abstract
The invention provides a screening method of comprehensive stress-resistant buried bivalve shellfish, which comprises the following steps: preparing a culture system; monitoring a culture environment, and checking whether the environment index of the survival of the cultured organisms is met; after the detection is qualified, the buried bivalve shellfish is put into a culture system; sequentially screening appearance form observation, shell opening recovery behavior after acute stress, feeding behavior, sand burying behavior and respiratory metabolism, and screening out high-quality shellfish parents for propagation. According to the invention, by utilizing the nonspecific property of stress and based on the principle that the individual stress recovery time is different after the bivalve shellfish is exposed to the stress, namely, the opening time, feeding intensity, sand burying time and degree and respiratory metabolism rate of the bivalve shellfish after the stress are used as screening indexes of the comprehensive stress-resistant buried bivalve shellfish, shellfish individuals with strong stress resistance are screened out by adopting a multi-round screening and comprehensive evaluation method, the health of shellfish groups is ensured, the culture yield and survival rate are improved, and the method has great significance in promoting the sustainable development of natural resources.
Description
Technical Field
The invention belongs to the technical field of aquaculture, and particularly relates to a screening method of comprehensive stress-resistant buried bivalve shellfish.
Background
In recent years, the aquaculture industry in China develops rapidly, is the only main fishery country in which the total amount of the cultured aquatic products exceeds the fishing total amount, and the culture yield accounts for more than 70% of the global aquaculture yield. The method has the advantages that huge achievement is achieved in the aquaculture industry in China, and meanwhile the problems of pollution around the aquaculture water area, unreasonable aquaculture layout, lag in aquaculture management and breeding theory and the like are faced. The buried bivalve shellfish is a marine product with important economic value in China, is widely distributed in coastal areas in China, inevitably encounters various environmental changes and artificial stress, causes the death rate of the shellfish to be increased, and simultaneously pollutes the water quality, thereby inducing a series of vicious cycles in the cultivation process.
Disclosure of Invention
Aiming at the defects in the prior art, the invention aims to provide a simple, convenient and efficient screening method for comprehensive stress-resistant buried bivalve shellfish.
To achieve the above object, the solution of the present invention is:
a screening method of comprehensive stress-resistant buried bivalve shellfish comprises the following steps:
(1) Sterilizing the culture system before culture, then injecting seawater and continuously aerating, and adding yeast extract powder and nitrifying bacteria into a circulating water system of the culture system to ensure that the circulating water system runs for more than 2 weeks;
(2) Continuously monitoring the circulating water system until the standard environmental index is reached;
(3) Aerating and flushing the buried bivalve shellfish in advance, soaking in the seawater with the aeration in advance, and then placing in a circulating water system reaching the standard in the step (2);
(4) Selecting healthy individuals with hypertrophy individuals and fresh shell surfaces from the buried bivalve shellfish of the same age, weighing, arranging the healthy individuals from large to small according to the weight, and selecting 1/3 of bivalve shellfish before the weighing to put the bivalve shellfish into a culture system for temporary culture;
(5) Screening out the active healthy buried bivalve shellfish by sequentially utilizing the behavior of opening and recovering after acute stress, ingestion, sand burying and respiratory metabolism;
(6) And (3) selecting shellfish individuals with low respiratory metabolism rate and strong stress resistance as parent shellfish for breeding from the active buried bivalve shellfish obtained in the step (5) until the screening is finished.
In the step (5), as a preferred embodiment of the present invention, the screening method for the recovery behavior of open shell after acute stress is: stopping feeding the buried bivalve shellfish for one day, taking out, exposing in air for 10min, and placing in a glass jar filled with seawater; shooting the buried bivalve shellfish horizontally for 30min by using a camera monitoring system, and recording the shell opening time of the bivalve shellfish; arranging the numerical value of the shell opening time from short to long, selecting the buried bivalve shellfish of the first 1/2 of the number for subsequent screening, and recovering the bivalve shellfish for a plurality of hours after screening.
Specifically, experimental bivalve shellfish is fed 4×10 daily 4 mL -1 The chlorella keeps continuously aerated water, and water is changed for 1 time every day, each timeWater was changed 1/2 and soil was sucked 20:00 a day. Filling 3/4 of seawater into a glass jar for a shell opening experiment, wherein the salinity is 23-25 per mill; taking out the buried bivalve shellfish from the culture jar after exposing for 10min, wiping seawater on the surface with clean filter paper, and exposing on a dry table for 10min.
The horizontal shooting is specifically to keep the shell opening direction of the bivalve shell facing the camera, select the superposition of the bottom of the picture and the cylinder bottom, and the superposition of the top of the picture and the water surface.
The shell opening action is specifically that the bivalve shell opens and stretches out of the water outlet pipe hole to perform water filtering action; the shell opening time is the time for recording the time for opening the bivalve shell to extend out of the water inlet pipe hole after the buried bivalve shell recovers from stress.
In step (5), as a preferred embodiment of the present invention, the screening method of feeding behavior is as follows: the ingestion experiment is measured by a still water system: placing buried bivalve shellfish into a feeding bottle filled with equal amount of chlorella after exposing the bivalve shellfish to air for 10min before starting the experiment, continuously feeding the whole experiment for 2h, taking water samples every 20min after starting the experiment to measure the chlorella concentration, and calculating the feeding rate of the bivalve shellfish; arranging the ingestion rate from small to large, selecting 1/2 of the buried bivalve shellfish for subsequent screening, and recovering for several hours after screening.
Specifically, chlorella concentration of 4×10 is added in feeding experiment 4 mL -1 Taking 200 mu L of water sample every 20min after the experiment starts, measuring the concentration of chlorella by a counting method, and measuring and recording the ingestion rate of the buried bivalve shellfish.
In the step (5), as a preferred embodiment of the present invention, the screening method of the sand burying behavior is as follows: taking out the buried bivalve shellfish, exposing for 10min, and placing into a glass jar filled with seawater and fine sand to ensure clean sand and spreading; establishing a camera monitoring system, placing the buried bivalve shellfish vertical to the water surface with the opening direction facing downwards, and recording the time of the buried bivalve shellfish completely buried in sand, wherein the shooting time is limited to 30 minutes; arranging the sand burying time values from short to long, selecting the front 1/2 of buried bivalve shellfish for subsequent screening, and recovering for several hours after screening.
Specifically, the sand burying experiment is carried out in a glass cylinder filled with fine sand, wherein the depth of the fine sand is about 8-10cm, and the flat sand surface and the clear water body are ensured; placing the buried bivalve shellfish on sand, keeping the opening direction of the bivalve shellfish downward, selecting the bottom of the picture to coincide with the bottom of the cylinder, and the top of the picture to coincide with the water surface; the time for completely burying the shell body of the buried bivalve shellfish is recorded as the time for the water inlet and outlet pipe holes to extend out of the sand surface.
As a preferred embodiment of the present invention, in step (5), the screening method of respiratory metabolism comprises: measuring respiratory metabolism rate (MO) of buried bivalve by automatic intermittent flow respiratory measuring instrument 2 ；mgO 2 h -1 ) In order to avoid the increase of the background oxygen consumption value caused by bacteria breeding before the experiment starts, disinfectant is used for disinfection and clean water is used for washing; after cleaning, the buried bivalve shellfish is placed in a breathing chamber, the temperature during breathing measurement is kept constant, a starting program (namely, one program set by an intermittent breathing measuring instrument comprises 240s flushing, 60s waiting and 300s measuring three parts, wherein three parts are one cycle, each cycle is 10 min), the breathing metabolism rate of the bivalve shellfish is recorded every 10min, the measuring time lasts for 2h, the breathing metabolism rate values are arranged from small to large, and buried bivalve shellfish individuals with the breathing metabolism rate of 1/2 are selected.
In practice, the disinfectant is a bleaching powder solution, and the available residual chlorine in the disinfectant is 30-35ppm.
As a preferred embodiment of the present invention, in the step (1), the disinfectant at the time of sterilization is bleaching powder; the adding amount of yeast extract powder and nitrifying bacteria is 0.01-0.03g/L and 0.1-0.3ml/L respectively.
As a preferred embodiment of the present invention, in the step (2), the required environmental indicators are: the water temperature in the circulating water system is 18-25 ℃, natural illumination is kept, the illumination time and the dark time are respectively 12 hours, the illumination intensity is 35-45lx, the pH value is 7.5-8.5, and the circulating water system is slightly alkaline; the dissolved oxygen concentration is more than or equal to 6.8mg/L, the ammonia nitrogen concentration is less than or equal to 0.1mg/L, the nitrite nitrogen concentration is less than or equal to 0.1mg/L, and the sulfide concentration is less than or equal to 0.1mg/L.
As a preferred embodiment of the invention, in the step (3), the flushing times of the buried bivalve shellfish are more than 3 times, and the dissolved oxygen concentration of the immersed aeration seawater is more than or equal to 6.8mg/L.
As a preferred embodiment of the present invention, in step (3), the buried bivalve shellfish is selected from all bivalve shellfish of class lamellida, such as razor clam, etc., and may preferably be clam and clam.
As a preferred embodiment of the present invention, in the step (6), stress resistance means that the buried bivalve shellfish has a stronger physiological recovery ability, a high survival rate and appetite, and a low respiratory metabolism rate after being subjected to air exposure stress, and can exhibit active viability such as sand burying. Physiological recovery, survival and appetite, and respiratory metabolism rates cannot be quantified due to strain differences.
By adopting the scheme, the invention has the beneficial effects that:
according to the method, firstly, the nonspecific property of stress is utilized, and according to the principle that after the bivalve shellfish is exposed to the stress, the individual stress recovery time is different, namely, the opening time, ingestion intensity, sand burying time and degree and respiratory metabolism rate of the bivalve shellfish after the stress are utilized as screening indexes of the comprehensive stress-resistant buried bivalve shellfish, and a screening method of multi-round screening and comprehensive evaluation is adopted, so that buried bivalve shellfish with strong stress resistance is screened out, the health of the population of the screened buried bivalve shellfish is ensured, and the culture yield and survival rate are further improved.
Secondly, the screening method is simple on the premise of not damaging the shellfish, and the buried bivalve shellfish individuals with strong stress resistance are finally selected through meticulous multiple screening, and the stress resistance and the disease resistance of the buried bivalve shellfish are closely related, so that the occurrence of diseases is reduced in the cultivation process, the integral disease resistance of the buried bivalve shellfish is improved, and the survival rate of the buried bivalve shellfish is greatly enhanced; in addition, the method has the advantages that the required instrument is simple, the operation is convenient and quick, the adopted screening method can be implemented in a common buried bivalve shellfish culture field, the practicability is high, and the method can be widely applied to the field of large-scale tidal flat bivalve shellfish aquaculture.
In a word, the invention screens out the buried bivalve seashell individuals with strong stress resistance, and then the raising can avoid the occurrence of raising disasters. The comprehensive stress-resistant buried bivalve shellfish screening method can further improve the culture yield and the culture quality of buried bivalve shellfish, improve the fine seed rate of breeding, reduce the culture cost of farmers, reduce environmental pollution to a certain extent, and has great significance for promoting sustainable development of natural resources and developing high-quality tidal flat bivalve shellfish artificial culture and seedling culture.
Drawings
FIG. 1 is a schematic representation of the recovery of shells from air exposure of the offspring of the screened clams in example 1.
FIG. 2 is a schematic representation of the post-air exposure feeding recovery of the offspring of the screened clams of example 1.
FIG. 3 is a schematic representation of the recovery of sand burial after air exposure of the offspring of clams screened in example 1.
Fig. 4 is a schematic representation of respiratory recovery after air exposure of the screened clam offspring of example 1.
FIG. 5 is a schematic representation of survival of the screened clam offspring after air exposure in example 1.
Detailed Description
The invention provides a screening method of comprehensive stress-resistant buried bivalve shellfish.
The screening method of the comprehensive stress-resistant buried bivalve shellfish comprises the following steps:
(1) Sterilizing the buried bivalve shellfish culture system by using a disinfectant before culturing, flushing and soaking the bivalve shellfish culture system by using fresh water, discharging the fresh water, and exposing the culture system to sunlight; the culture system is connected with a circulating water pump, then seawater with the salinity of 18-25 per mill (preferably 23-25 per mill) is injected, and gas stones are put into the culture system for continuous aeration, and a proper amount of yeast extract powder and nitrifying bacteria are put into a circulating water system of the culture system, so that the circulating water system is ensured to run for more than 2 weeks, and a biological film suitable for buried bivalve shellfish is cultivated;
(2) Continuously monitoring the culture circulating water system to ensure that the culture system reaches the standard environmental index, meet the survival requirement standard of the buried bivalve shellfish, and reduce the occurrence of diseases;
(3) Flushing the buried bivalve shellfish to be put into a culture system with fresh water which is aerated in advance, soaking the bivalve shellfish in the seawater which is aerated in advance, and then putting the bivalve shellfish into a circulating water system reaching the standard in the step (2);
(4) Selecting healthy individuals with hypertrophy individuals and fresh shell surfaces from the buried bivalve shellfish of the same age, weighing, arranging the healthy individuals from large to small according to the weight, and selecting 1/3 of bivalve shellfish before the weighing to put the bivalve shellfish into a circulating water system for temporary culture;
(5) The buried bivalve shellfish in the step (4) is subjected to stress resistance screening, and active healthy buried bivalve shellfish is screened out by sequentially utilizing the behavior of opening and recovering after acute stress, the feeding behavior, the sand burying behavior and respiratory metabolism;
(6) And (3) selecting shellfish individuals with low respiratory metabolism rate and strong stress resistance as parent shellfish for breeding from the active buried bivalve shellfish obtained in the step (5) until the screening is finished.
Wherein in the step (1), the disinfectant in the disinfection is bleaching powder; the number of times of fresh water flushing is more than 3; the soaking time is more than 1 day; the cultivation system is placed in the sun for 3 days; the adding amount of yeast extract powder and nitrifying bacteria is 0.01-0.03g/L and 0.1-0.3ml/L respectively.
In step (2), the required environmental indicators are: the water temperature in the circulating water system is 18-25 ℃, natural illumination is kept, the illumination time and the dark time are respectively 12 hours, the illumination intensity is 35-45lx, the pH value is 7.5-8.5, and the circulating water system is slightly alkaline; the dissolved oxygen concentration is more than or equal to 6.8mg/L, the ammonia nitrogen concentration is less than or equal to 0.1mg/L, the nitrite nitrogen concentration is less than or equal to 0.1mg/L, and the sulfide concentration is less than or equal to 0.1mg/L.
In the step (3), the flushing times of the buried bivalve shellfish are more than 3 times, and the dissolved oxygen concentration of the soaked aerated seawater is more than or equal to 6.8mg/L.
In the step (5), the screening method of the recovery behavior of opening the shell after acute stress comprises the following steps: after feeding of the cultured buried bivalve shellfish is stopped for one day, taking the shellfish out of a culture jar, placing the shellfish on a tabletop for exposure for 10min, and then placing the shellfish in a glass jar filled with seawater; shooting the shellfish horizontally for 30min by using a camera monitoring system, and recording the shell opening time of the buried bivalve shellfish; arranging the numerical values of the shell opening time from small to large, selecting the buried bivalve shellfish of the first 1/2, and recovering the bivalve shellfish for a plurality of hours after screening.
In the step (5), the screening method of ingestion behavior comprises the following steps: the experiment is measured by a still water system; after feeding of the cultured buried bivalve shellfish for one day is stopped before the experiment starts, taking out the bivalve shellfish from a culture cylinder, exposing the bivalve shellfish for 10min, respectively putting the bivalve shellfish into a feeding bottle filled with equal amount of chlorella, taking 200 mu L of water sample every 20min after the experiment starts, measuring the concentration of the chlorella, and calculating the feeding rate of the bivalve shellfish; arranging the ingestion rate from small to large, selecting 1/2 of the buried bivalve shellfish, and recovering for several hours after screening.
In the step (5), the screening method of the sand burying behavior comprises the following steps: taking out the buried bivalve shellfish, exposing for 10min, and placing into a glass jar filled with 3/4 seawater and about 10cm high fine sand to ensure clean sand and spreading; establishing a shooting monitoring system, placing the buried bivalve shellfish vertical to the water surface with the opening direction downward, recording the sand burying time of the bivalve shellfish, and limiting the shooting time to 30min; arranging the sand burying time values from small to large, selecting the buried bivalve shellfish of the first 1/2, and recovering for several hours after screening.
In step (5), the screening method of respiratory metabolism comprises the following steps: measurement of respiratory metabolism rate (MO) of buried bivalve individuals using automated intermittent flow respirometer (AutoRespTM, logo Systems, denmark) 2 ；mgO 2 h -1 ) Before the experiment starts, in order to avoid the increase of the background oxygen consumption value caused by bacterial growth, the breathing chamber is disinfected and cleaned in advance, the shellfish is put into the breathing chamber after the completion of the experiment, the respiratory metabolism rate of the bivalve shellfish is recorded every 10min after the starting procedure, and the measurement time lasts for 2h; arranging the values of the respiratory metabolism rate from small to large, selecting buried bivalve shellfish individuals with the respiratory metabolism rate of 1/2 of the previous respiratory metabolism rate, and recovering the bivalve shellfish individuals for a plurality of hours after screening.
In the step (6), the stress resistance is that the buried bivalve shellfish has stronger physiological recovery capability, high survival rate and appetite, low respiratory metabolism rate and can show active viability such as sand burying after being stressed by air exposure. Physiological recovery, survival and appetite, and respiratory metabolism rates cannot be quantified due to strain differences.
The technical scheme of the present invention will be clearly and completely described in the following in connection with the specific embodiments of the present invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The invention utilizes the non-specificity of stress, and based on the principle that the stress performance and recovery time are different after the buried bivalve shellfish with different physique and individuality is stimulated by various external stimuli: after the buried bivalve shellfish is stimulated by air exposure, the buried bivalve shellfish with different individualities has different stress recovery time to the stimulation from the outside. The active buried bivalve shellfish can recover normal physiological behaviors faster after being stimulated, and the surrounding is explored more 'brave'; and after being stimulated, the passive buried bivalve shellfish needs to recover normal physiological behaviors for a longer time, and the passive buried bivalve shellfish is in a self-closing and biliary tract behavior mode. However, a scientific and reasonable detection method is needed during screening, so that death of the buried bivalve shellfish due to overlarge screening strength is avoided, and buried bivalve shellfish individuals with strong stress resistance can be accurately screened out, so that a reasonable screening method is needed.
The present invention will be described in detail with reference to examples.
Example 1:
the clam screening method of the embodiment comprises the following steps:
(1) Preparing a culture system:
after the cultivation system is thoroughly disinfected by using bleaching powder, the cultivation system is washed 3 times by fresh water and soaked for 1 day, seawater with the salinity of about 18 per mill is injected, and yeast extract powder with the salinity of 0.02g/L and nitrifying bacteria with the concentration of 0.2ml/L are put into a circulating water system of the cultivation system, so that the circulating system is ensured to run for more than 2 weeks, and a biological membrane suitable for clam survival is cultivated.
(2) And monitoring the culture circulating water system, ensuring that the culture system reaches standard indexes and meeting the survival requirement standard of clams.
Wherein, the water temperature in the circulating water system is about 18 ℃, natural illumination is kept, the illumination time and the darkness time are respectively 12 hours, the illumination intensity is 35lx, the pH value is 7.5, and the circulating water system is alkalescent; the dissolved oxygen concentration is 6.8mg/L, the ammonia nitrogen concentration is 0.1mg/L, the nitrite nitrogen concentration is 0.1mg/L, and the sulfide concentration is 0.1mg/L.
(3) And (3) after the clams are detected to be qualified in the step (2), flushing clams which are required to be put into the circulating water system with fresh water which is aerated in advance for 3 times, then putting the clams into the seawater which is aerated in advance for soaking for 30 minutes, and then putting the clams into the circulating water system which reaches the standard in the step (2).
Wherein the salinity of the soaked seawater is 25 per mill, and the concentration of dissolved oxygen is 6.8mg/L. The density of clams introduced into the culture system is 2.2kg/m 2 。
(4) And selecting healthy individuals with hypertrophy individuals and fresh shell surfaces from the clams of the same age, weighing, arranging the healthy individuals from large to small according to the weight, and placing 1/3 of clams in a circulating water system for temporary culture.
(5) Feeding of food was stopped until clams were screened the first day. A camera monitoring system is built in advance, clams are taken out of a culture tank by net fishing, sea water on the surfaces of the clams is wiped by clean filter paper, the clams are placed on a dry tabletop to be exposed for 10 minutes, and then the clams are placed in a glass tank filled with sea water; horizontally shooting clams by using a camera monitoring system, recording the shell opening time of clams, and limiting the shooting time to 30min; arranging the numerical value of the shell opening time from small to large, selecting the front 1/2 of active clams, recovering the clams for a plurality of hours after screening, and using the clams as the passive clams 1/2 of the active clams.
Feeding and screening the clams at the same time on the next day: the ingestion experiment is measured by a still water system; taking out the active clams from the culture tank by using a net, wiping the seawater on the surface of the active clams with clean filter paper, putting the active clams on a dry tabletop for exposure for 10min, and respectively putting the active clams into ingestion bottles containing equal amount of chlorella; taking 200 mu L of water sample every 20min after the experiment starts to measure the chlorella concentration, and calculating the ingestion rate of clams; the ingestion rate is arranged from small to large, 1/2 of the active clams are selected, and the clams are recovered for a plurality of hours after screening, and the first 1/2 of the clams are used as passive clams.
And (3) carrying out sand burying screening on clams in the same group at the same time on the following third day: clam is taken out and placed in a glass jar filled with seawater and 10cm high fine sand after being exposed to the desktop air for 10min, so that the sand is ensured to be clean and paved; establishing a camera monitoring system, placing clams vertically on a water surface with an opening facing downwards, recording the time of the clams to completely embed sand, and limiting the shooting time to 30min; arranging the sand burying time values from short to long, selecting 1/2 of active clams before the sand burying time, recovering the active clams for a plurality of hours after screening, and then using 1/2 of the active clams as passive clams.
Respiratory screening of the clams was performed on the fourth day at the same time: breath test measurement of respiratory Metabolism (MO) of Meretrix individuals using automated intermittent flow breath measuring apparatus (AutoRespTM, logo Systems, denmark) 2 ；mgO 2 h -1 ) In order to avoid the increase of the background oxygen consumption value caused by bacterial growth before the experiment starts, the bleaching powder solution needs to be used for disinfection and clear water flushing in advance. After the completion, the clams are placed into a breathing chamber, a program is started, the breathing metabolism rate of the clams is recorded every 10min, and the measurement time lasts for 2h; arranging the values of the respiratory metabolism rate from small to large, selecting 1/2 of high stress resistance clam individuals in front of the respiratory metabolism rate as parent clams for propagation, and then 1/2 of clams as low stress resistance clams, so as to finish screening.
Example 2:
the screening method of the arca subcrenata in the embodiment comprises the following steps:
(1) Preparing a culture system:
after the cultivation system is thoroughly disinfected by bleaching powder, the cultivation system is washed 3 times by fresh water and soaked for 1 day, seawater with the salinity of about 25 per mill is injected, and yeast extract powder with the salinity of 0.02g/L and nitrifying bacteria with the concentration of 0.2ml/L are put into a circulating water system of the cultivation system, so that the circulating system is ensured to run for more than 2 weeks, and a biological film suitable for survival of the arca subcrenata is cultivated.
(2) And monitoring the culture circulating water system, ensuring that the culture system reaches standard indexes, and meeting the survival requirement standard of the arca subcrenata.
Wherein, the water temperature in the circulating water system is about 20 ℃, natural illumination is kept, the illumination time and the dark time are respectively 12 hours, the illumination intensity is 45lx, and the pH value is 8.5; the dissolved oxygen concentration is 6.8mg/L, the ammonia nitrogen concentration is 0.1mg/L, the nitrite nitrogen concentration is 0.1mg/L, and the sulfide concentration is 0.1mg/L.
(3) And (3) after the step (2) is detected to be qualified, flushing the blood clam needing to be put into the circulating water system with fresh water which is aerated in advance for 3 times, then putting the blood clam into the sea water which is aerated in advance for soaking for 30 minutes, and then putting the blood clam into the circulating water system which meets the standard in the step (2).
Wherein the salinity of the soaked seawater is 22 per mill, and the concentration of dissolved oxygen is 6.8mg/L. The density of the blood clam introduced into the culture system is 1.8kg/m 2 。
(4) And selecting healthy individuals with hypertrophy individuals and fresh shell surfaces from the blood clam of the same age, weighing, arranging the individuals according to the weight from large to small, and selecting the blood clam of the first 1/3 of the individuals to be put into a circulating water system for temporary culture.
(5) And continuously stopping feeding food before the blood clam is screened on the first day. A camera monitoring system is built in advance, the clam is taken out of a culture jar by using a net, sea water on the surface of the clam is wiped by clean filter paper, the clam is placed on a dry tabletop for exposure for 10min, and then the clam is placed in a glass jar filled with sea water; using a camera monitoring system to horizontally shoot the arca subcrenata, recording the shell opening time of the arca subcrenata, and limiting the shooting time to 30 minutes; arranging the numerical value of the shell opening time from small to large, selecting the front 1/2 of active blood clam, recovering the active blood clam for a plurality of hours after screening, and then taking the active blood clam as the passive blood clam 1/2.
Feeding and screening the blood clam at the same time the next day: the ingestion experiment is measured by a still water system; taking out the blood clam from the culture jar by using a net, wiping the seawater on the surface of the blood clam with clean filter paper, putting the blood clam on a dry tabletop for exposure for 10min, and respectively putting the blood clam into a feeding bottle filled with equal amount of chlorella; taking 200 mu L of water sample every 20min after the experiment starts to measure the chlorella concentration and calculate the feeding rate of the arca subcrenata; the feeding rate values are arranged from small to large, 1/2 of active blood clam individuals are selected, the active blood clam individuals are recovered for a plurality of hours after screening, and the first 1/2 of active blood clam individuals serve as passive blood clam.
And (3) carrying out sand burying screening on the blood clam in the same group at the same time on the following third day: taking out the blood clam, exposing the blood clam on a desktop for 10min, and putting the blood clam into a glass jar filled with seawater and 10cm high fine sand to ensure that sand is clean and paved; establishing a camera monitoring system, placing the arca subcrenata vertical to the water surface, enabling the opening direction to be downward, recording the time for completely burying sand of the arca subcrenata, and limiting the shooting time to 30 minutes; arranging the sand burying time values from short to long, selecting 1/2 of active blood clam individuals before the sand burying time, recovering the active blood clam individuals for a plurality of hours after screening, and then using the active blood clam individuals as passive blood clam individuals 1/2.
And carrying out respiration screening on the blood clam at the same time on the fourth day: breath test measurement of respiratory Metabolism (MO) of individual blood clam by automatic intermittent flow breath measuring instrument (AutoRespTM, logo Systems, denmark) 2 ；mgO 2 h -1 ) In order to avoid the increase of the background oxygen consumption value caused by bacterial growth before the experiment starts, the bleaching powder solution needs to be used for disinfection and clear water flushing in advance. After the completion, the arca subcrenata is placed into a breathing chamber, a program is started, the breathing metabolic rate of the arca subcrenata is recorded every 10min, and the measurement time lasts for 2h; arranging the values of the respiratory metabolism rate from small to large, selecting 1/2 of high stress resistance blood clam individuals in front of the respiratory metabolism rate as parent shellfish for propagation, and then 1/2 of the high stress resistance blood clam individuals as parent shellfish for propagation until the screening is finished.
Test results:
with example 1 as the test sample, other examples were similar in effect to example 1.
Twenty active clams and passive clams offspring screened by the method are respectively selected for stress resistance test.
(1) Environmental stress resistance test of clam offspring:
1) Feeding rate (cell L) within 2h after stress -1 h -1 )；
2) Nanometer TiO 2 Survival rate (survivin rate,%) after 30 days exposure.
| feeding rate(cell L -1 h -1 ) | survival rate(％) | |
| Active clam | 11.48*10 5 | 83 |
| Passive clam | 3.36*10 5 | 46 |
From the above table, it can be seen that the environmental stress resistance index possessed by the offspring of the screened active clams is better.
(2) The recovery condition of the shell opening time after the stress of the screened active clams and passive clams:
the test results are shown in fig. 1, and the screened active clams can recover the shell after being stressed far faster than the passive clams.
(3) Feeding recovery after stress of the screened active clams and passive clams:
the test results are shown in fig. 2, and the ingestion rate of the screened active clams is obviously higher than that of the passive clams in 20min, 40min and 80min, and then the ingestion rate and the ingestion rate of the screened active clams are not obviously changed.
(4) The sand burying recovery condition after the stress of the screened active clams and passive clams:
the test results are shown in fig. 3, and the test results show that the sand burying recovery speed of the active clams screened out after stress is far faster than that of the passive clams.
(5) The respiration recovery condition of the screened active clams and passive clams after stress is as follows:
the test results are shown in fig. 4, and the breath recovery of the active clams screened after stress is obviously lower than that of the passive clams in 20min, 40min and 60min, and no obvious change is caused after the breath recovery, so that the stress resistance of the active clams is stronger than that of the passive clams.
(6) Survival of the selected active and passive clams:
the test results are shown in fig. 5, and the survival rate of the active clams screened after stress is obviously higher than that of the passive clams.
Finally, it should be noted that: while the invention has been described in detail with reference to the foregoing embodiments, it will be apparent to one skilled in the art that the invention is not limited to the preferred embodiments, and modifications and variations can be made to the embodiments described herein without departing from the spirit and principles of the invention.
Claims (3)
1. A screening method of comprehensive stress-resistant buried bivalve shellfish is characterized in that: the method comprises the following steps:
(1) Sterilizing a culture system before culture, then injecting seawater and continuously aerating, and adding yeast extract powder and nitrifying bacteria into a circulating water system of the culture system to ensure that the circulating water system runs for more than 2 weeks;
(2) Monitoring the water quality of the culture system until the environmental index of the culture standard is reached;
(3) Aerating and flushing buried bivalve shellfish in advance, soaking in the seawater aerated in advance, and then placing in a circulating water system reaching the standard in the step (2);
(4) Selecting healthy individuals with hypertrophy individuals and fresh shell surfaces from the buried bivalve shellfish of the same age, arranging the healthy individuals from large to small according to the weight, and selecting 1/3 of the bivalve shellfish to be put into a culture system for temporary culture;
(5) The buried bivalve shellfish screened in the step (4) is screened out by sequentially utilizing the behavior of opening the shell after acute stress, ingestion behavior, sand burying behavior and respiratory metabolism;
(6) Selecting shellfish individuals with low respiratory metabolism rate and strong stress resistance as parent shellfish for breeding from the active healthy buried bivalve shellfish obtained in the step (5) until the screening is finished;
in the step (5), the screening method of the recovery behavior of opening the shell after acute stress comprises the following steps: after stopping feeding the buried bivalve shellfish for one day, taking out, exposing the bivalve shellfish with air, and placing the bivalve shellfish into a glass jar filled with seawater; using a camera monitoring system to horizontally shoot the buried bivalve shellfish and recording the shell opening time of the bivalve shellfish; arranging the numerical value of the shell opening time from short to long, selecting the front 1/2 of buried bivalve shellfish for subsequent screening, and recovering the bivalve shellfish for a plurality of hours after screening;
the screening method of the ingestion behavior comprises the following steps: the ingestion experiment is measured by a still water system: placing buried bivalve shellfish into an air-exposed feeding bottle containing equal amount of chlorella before the experiment starts, taking water samples every 20min after the experiment starts to measure the chlorella concentration, and calculating the feeding rate of the bivalve shellfish; arranging the ingestion rate values from small to large, selecting 1/2 of the buried bivalve shellfish for subsequent screening, and recovering the bivalve shellfish for several hours after screening;
the screening method of the sand burying behavior comprises the following steps: taking out the buried bivalve shellfish, exposing the bivalve shellfish in air, and putting the bivalve shellfish into a glass jar filled with seawater and fine sand to ensure that sand is clean and paved; establishing a camera monitoring system, placing the buried bivalve shellfish vertical to the water surface, and recording the time of completely burying the bivalve shellfish in sand, wherein the opening direction is downward; arranging the sand burying time values from short to long, selecting the front 1/2 of buried bivalve shellfish for subsequent screening, and recovering for several hours after screening;
the screening method of respiratory metabolism comprises the following steps: measuring the respiratory metabolism rate of the buried bivalve shellfish by using an automatic intermittent flow respiratory measuring instrument, sterilizing and cleaning the respiratory measuring instrument before the experiment is started, putting the buried bivalve shellfish into a respiratory chamber after the experiment is finished, recording the respiratory metabolism rate of the bivalve shellfish after a starting program, arranging the respiratory metabolism rate values from small to large, and selecting buried bivalve shellfish individuals with the respiratory metabolism rate 1/2 of that before the respiratory metabolism rate;
the stress resistance is that the buried bivalve shellfish has strong physiological recovery capability, high survival rate, appetite and low respiratory metabolism rate after being subjected to air exposure stress;
the buried bivalve shellfish is clam.
2. The method for screening comprehensive stress-resistant buried bivalve shellfish according to claim 1, wherein the method comprises the steps of: in the step (2), the environmental index is: the water temperature in the culture system is 18-25 ℃, the illumination time and the darkness time are respectively 12 hours, the illumination intensity is 35-45lx, and the pH value is 7.5-8.5; the dissolved oxygen concentration is more than or equal to 6.8mg/L, the ammonia nitrogen concentration is less than or equal to 0.1mg/L, the nitrite nitrogen concentration is less than or equal to 0.1mg/L, and the sulfide concentration is less than or equal to 0.1mg/L.
3. The method for screening comprehensive stress-resistant buried bivalve shellfish according to claim 1, wherein the method comprises the steps of: in the step (3), the flushing times of the buried bivalve shellfish are more than 3 times, and the dissolved oxygen concentration of the soaked aerated seawater is more than or equal to 6.8mg/L.
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| CN112931311A (en) * | 2021-02-09 | 2021-06-11 | 上海海洋大学 | Flounder health condition evaluation method |
| CN113940294A (en) * | 2021-10-13 | 2022-01-18 | 上海海洋大学 | Breeding method for negative correlation characters of fishes |
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| CN107568131A (en) * | 2017-09-11 | 2018-01-12 | 上海海洋大学 | The comprehensive degeneration-resistant flounder sole screening technique of one kind |
| CN112931311A (en) * | 2021-02-09 | 2021-06-11 | 上海海洋大学 | Flounder health condition evaluation method |
| CN113940294A (en) * | 2021-10-13 | 2022-01-18 | 上海海洋大学 | Breeding method for negative correlation characters of fishes |
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