Transportation method of *Pterygodon oryzae* for benthic organism restoration

Abstract

This invention discloses a method for transporting *Pterygodon spp.* for benthic bioremediation, including pre-transportation pretreatment, low-temperature transportation, and post-transportation adaptation and domestication. Before transportation, by withholding feed and stimulating the expulsion of mud and sand by low osmotic pressure, the *Pterygodon spp.* effectively induces the *Pterygodon spp.* to contract its body segments, actively expel mud and sand to clean its intestines, and achieve intestinal and body surface purification, avoiding mechanical damage and water quality deterioration risks caused by mud and sand residue in the body during transportation. After transportation, adaptation and domestication can significantly reduce the *Pterygodon spp.*'s transportation stress response, reduce body segment damage caused by violent twisting, and induce the *Pterygodon spp.* to burrow naturally by creating a suitable substrate environment and tidal environment, quickly restoring its burrowing and benthic lifestyle.

Description

Technical Field

[0001] This invention belongs to the field of biological transportation technology for ecological restoration, specifically, it relates to a method for transporting *Pterygota esculenta* for benthic biological restoration. Background Technology

[0002] The double-toothed sandworm (Perinereis aibuhitensis) belongs to the phylum Annelida, class Polychaeta, and genus Nereis. Commonly known as the "sea centipede," it is a typical large marine benthic organism that lives on the coastal mudflats of my country.

[0003] As a key species in coastal wetland ecosystems, the *Pterygodon spp.* (also known as the sandworm) is a preferred high-protein food source for long-distance migratory birds due to its large size and high biomass. Furthermore, it is an important food source for native birds during their brooding period. Simultaneously, *Pterygodon spp.* is also a crucial food source for many benthic fish (such as gobies and tongue soles) and crabs, and its biomass directly impacts the stability of upper-layer predator populations. Therefore, *Pterygodon spp.* is considered a "cornerstone for maintaining biodiversity and ecosystem stability in coastal wetlands."

[0004] However, due to the impact of human activities such as land reclamation, pollution, and overfishing throughout history, as well as threats from invasive alien species, the wild population of *Pteris vittata* has declined sharply. For example, in traditionally high-yield areas such as the Bohai Bay and the Jiangsu coast, many areas rich in *Pteris vittata* have disappeared or their biomass has decreased by 50%-80%. Therefore, in the ecological restoration of many coastal wetlands, especially in stopover and wintering grounds for migratory birds, *Pteris vittata* is being artificially propagated and released as an important restoration tool to help restore the ecological environment.

[0005] Due to the decline in natural populations, which cannot meet the huge market demand, *Pterygodium bispinosa* currently relies mainly on artificial breeding, with breeding areas concentrated in Dongying City, Shandong Province, Tangshan City, Hebei Province, and Yancheng City, Jiangsu Province. Currently, *Pterygodium bispinosa* is one of my country's most exported parsnip species, sold to Japan, South Korea, and other countries. In recent years, my country's annual parsnip production has reached 1,000 tons, with an annual output value of US$70 million, accounting for approximately 60-70% of the global total, firmly ranking first in the world.

[0006] Whether used for artificial propagation and release in ecological restoration, or for aquaculture and sea fishing bait, *Pterygium bisporum* requires high freshness after transportation, especially during the peak market season from June to September. High temperatures during transport often cause the sandworms' body segments to break, become infected, rot, and smell foul, rendering them unusable. Therefore, strict control of transportation conditions is crucial. Currently, temperature control is implemented during sandworm transportation, but significant problems remain: residual mud and sand within the sandworms during transport can easily cause mechanical damage between individuals, leading to water quality deterioration, high mortality rates, low survival rates, weak vitality, and severe stress responses and increased body segment damage rates after transportation.

[0007] Currently, patents related to *Pterygodium spp.* mainly focus on breeding methods and seedling cultivation methods. Patents concerning transportation methods are limited to "A method for manufacturing a storage and transportation pad for *Pterygodium spp.*" (Publication No.: 1594044A), "A method for preserving *Pterygodium spp.* with added anti-stress factors" (108967283A), and "A *Pterygodium spp.* rearing and transportation device" (213281177U). The first two patents mainly disclose the manufacturing parameters and methods for the storage and transportation pads and the biomimetic preservation paper with added anti-stress factors during the transportation process of *Pterygodium spp.* The last patent designs a set of ultrasonic rearing chamber devices for *Pterygodium spp.* for sorting *Pterygodium spp.* of different qualities. None of the three patents cover efficient pre-transportation treatment methods for *Pterygodium spp.*, environmental parameter control during transportation, or post-transportation adaptation treatment methods.

[0008] The information disclosed in this background section is only intended to enhance the understanding of the background technology of this application, and therefore may include prior art that is not known to those skilled in the art. Summary of the Invention

[0009] This invention proposes a method for transporting *Nematocystis jirovecii* for benthic organism remediation, in order to solve the technical problems of existing *Nematocystis jirovecii* transport, which often suffers from mechanical damage between individuals due to residual mud and sand in their bodies, leading to water quality deterioration, high mortality rate, low survival rate, weak vitality, and strong stress response and increased segment damage rate after transport.

[0010] To achieve the above-mentioned invention / design objectives, the present invention adopts the following technical solution:

[0011] A method for transporting *Pterygodon spp.* for benthic bioremediation, the method comprising:

[0012] Pre-transportation processing:

[0013] Stop feeding the animals;

[0014] Harvesting sandworms after a set time;

[0015] The sandworms are stimulated to expel mud and sand by low osmotic pressure: the cleaned sandworms are transferred to a temporary holding box with a moist mat at the bottom. Low osmotic pressure seawater is injected at regular intervals. The temperature of the low osmotic pressure seawater is higher than that of the seawater at the excavation site, and the salinity of the low osmotic pressure seawater is lower than that of the seawater at the excavation site.

[0016] Transportation process: The sandworms are packed into insulated boxes for transportation, and the temperature inside the insulated boxes is controlled at 10-20℃;

[0017] Post-transport acclimatization process:

[0018] The insulated box was moved to a low-temperature space and seawater was sprayed regularly. The seawater temperature was raised from 10-15℃ to 18-22℃ and the seawater salinity was adjusted from 15-25 to the salinity of the ecological restoration area.

[0019] After the adaptation period, screening and disinfection will be carried out.

[0020] Inducing burrowing: The disinfected sandworms are placed in a container with a sloping substrate. The container has a water inlet near the lower part of the substrate and a drain near the higher part of the substrate. The water inlet is connected to a water pump. The water pump simulates tides to induce the sandworms to actively burrow in the sand.

[0021] The above-described method for transporting *Pterygodon spp.* for benthic bioremediation involves screening and cleaning the harvested *Pterygodon spp.*

[0022] As described above, the method for transporting *Pterygodon spp.* for benthic bioremediation involves washing the *Pterygodon spp.* with seawater of the same temperature and salinity as the extraction pond.

[0023] The above-described method for transporting *Pterygodon spp.* for benthic organism restoration involves periodically injecting low-osmotic-pressure seawater into the holding tank, controlling the seawater depth inside the holding tank to 2-3 cm, and maintaining a dark environment within the holding tank.

[0024] The above-described method for transporting *Pterygodon spp.* for benthic organism restoration involves periodically changing the water in the temporary holding tank, with each water change involving 70%-100% of the total water volume.

[0025] The above-described method for transporting *Pterygota praecox* for benthic bioremediation involves classifying the *Pterygota praecox* by weight and then placing them in insulated boxes for transport, with the weight difference of *Pterygota praecox* within the same insulated box being less than 20%.

[0026] The above-described method for transporting *Pterygodon spp.* for benthic bioremediation involves loading pretreated *Pterygodon spp.* and seawater into an insulated box at a ratio of 1:1.3-1.8. The insulated box contains absorbent polymer resin.

[0027] The above-described method for transporting *Pterygodon spp.* for benthic bioremediation uses a substrate consisting of fine sand mixed with shell fragments and having a particle size of less than 1 mm, with a substrate thickness of 10-15 cm.

[0028] The method for transporting *Nematocystis jirovecii* for benthic bioremediation, as described above, uses a water pump with a timer, where a water inlet and outlet cycle is 5-8 hours.

[0029] The above-described method for transporting *Nereidum didentatum* for benthic bioremediation involves adding diatomaceous earth solution to the container at an algal density of 1 × 10⁻⁶. 3 -1×10 4 The cells / mL concentration is 2-5 ml / L.

[0030] Compared with the prior art, the advantages and positive effects of the present invention are:

[0031] This invention relates to a method for transporting *Pterygodon spp.* for benthic bioremediation, comprising pre-transport pretreatment, low-temperature transportation, and post-transport acclimatization. Before transportation, by withholding feed and stimulating the expulsion of mud and sand through low osmotic pressure, the *Pterygodon spp.* effectively induces the *Pterygodon spp.* to contract its body segments, actively expelling mud and sand to clean its intestines, thus achieving intestinal and body surface purification and avoiding mechanical damage and water quality deterioration risks caused by mud and sand residue remaining in the body during transportation. Post-transport acclimatization significantly reduces the *Pterygodon spp.*'s transportation stress response, reduces body segment damage caused by violent twisting, and induces natural burrowing by creating a suitable substrate and tidal environment, rapidly restoring its burrowing and benthic lifestyle.

[0032] Other features and advantages of the present invention will become clearer after reading the detailed embodiments of the invention in conjunction with the accompanying drawings. Attached Figure Description

[0033] To more clearly illustrate the technical solutions in the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0034] Figure 1 This is a schematic diagram of an insulated box according to a specific embodiment of the present invention.

[0035] Figure 2 This is a schematic diagram of a container according to a specific embodiment of the present invention.

[0036] Figure 3 , Figure 4 It is a sandworm with damaged body segments.

[0037] Figure 5 , Figure 6 These are healthy sandworms. Detailed Implementation

[0038] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0039] In the description of this invention, it should be understood that all other embodiments obtained by the terminologist without inventive effort are embodiments described herein. The orientations or positional relationships indicated herein are based on the orientations or positional relationships shown in the drawings and are only for the purpose of facilitating the description of this invention and simplifying the description, and are not intended to indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on this invention.

[0040] In the description of this invention, it should be noted that, unless otherwise explicitly specified and limited, the term "in the description" is used to indicate that, unless it is to be interpreted broadly, it can refer to a fixed connection, a detachable connection, or an integral connection. Those skilled in the art can understand the specific meaning of the above terms in this invention based on the specific circumstances. In the description of embodiments, specific features, structures, materials, or characteristics can be combined in any suitable manner in one or more embodiments or examples.

[0041] The term "feature" has a specific meaning. In practice, it is used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Therefore, the term "for descriptive purposes" is limited to the fact that a feature may explicitly or implicitly include one or more of that feature.

[0042] In the description of this invention, unless otherwise stated, "the meaning in the description is two or more".

[0043] A method for transporting *Nereidum bidentatum* for benthic organism restoration includes: a pre-transportation pretreatment process, a transportation process, and a post-transportation adaptation and domestication process. This method significantly reduces the damage rate of *Nereidum bidentatum* body segments and rapidly restores its vitality and physiological functions.

[0044] 1. Pre-transportation processing:

[0045] Stop feeding.

[0046] Sandworms are harvested after a set time.

[0047] The harvested sandworms are screened and cleaned. During the cleaning process, they are washed with seawater of the same temperature and salinity as the harvesting site.

[0048] Low osmotic pressure stimulation to induce sand expulsion and mud removal in sandworms: After cleaning, the sandworms are transferred to a temporary holding box with a moist mat at the bottom. Low osmotic pressure seawater is injected regularly. The temperature of the low osmotic pressure seawater is higher than that of the seawater in the excavation pond, and the salinity of the low osmotic pressure seawater is lower than that of the excavation pond.

[0049] Regularly inject low-osmotic-pressure seawater into the holding tank, maintaining a seawater depth of 2-3 cm inside the tank, and keep the holding tank environment dark.

[0050] Change the water in the temporary holding tank regularly, replacing 70%-100% of the total water volume each time.

[0051] After being graded by weight, the sandworms were placed in insulated boxes for transportation, with the weight difference of the sandworms in the same insulated box being less than 20%.

[0052] The core of pretreatment lies in stimulating the sandworm to expel mud and cleanse its intestines through conditions such as low osmotic pressure, thereby reducing the risk of mechanical damage and water quality deterioration caused by the presence of mud and sand in its body during transportation.

[0053] The specific steps are as follows:

[0054] (1) Stop feeding: Stop feeding 7 days before transportation and stop feeding any food into the sandworm digging pond to empty its intestines.

[0055] (2) Screening: After harvesting, screen out damaged sandworms, remove individuals with incomplete body walls (such as missing parapodia or broken segments), rough body surfaces (such as white ulcer spots), dull body color (too transparent or semi-transparent), or weak vitality (slow snout contraction), to ensure that the initial health of the transport group is good.

[0056] (3) Cleaning: Wash with seawater 2-3 times to remove the adhering substances on the body surface. The temperature and salinity of the seawater used for cleaning are the same as the temperature of the excavation pool.

[0057] (4) Low osmotic pressure stimulation to expel mud and sand: After cleaning, the sandworms are transferred to a temporary rearing box. The box is flat, with a damp gauze or sponge pad at the bottom. The sandworms are laid flat in a single layer on the pad, and then low osmotic pressure adjustment is performed: low osmotic pressure seawater is gently injected every 1 hour. The temperature of the low osmotic pressure seawater is 2-3℃ higher than that of the extraction pool, and the salinity is 4-6 lower. The depth of the seawater in the temporary rearing box is controlled at 2-3cm, and the box environment is kept dark. This process enhances metabolism by moderately raising the temperature, and the low osmotic pressure environment formed by low salinity and the dark conditions work together to effectively stimulate the sandworms to contract their body segments, actively expel mud and sand to clean their intestines, and achieve intestinal and body surface purification.

[0058] Change the water in the temporary holding tank once every 24 hours, replacing 70%-100% of the total water volume.

[0059] (5) Grading and Packing: The sandworms are graded and packed within 48 hours of pretreatment. The grading method is based on the weight of each sandworm, ensuring that the weight difference of sandworms in the same transport box is less than 20%, so as to avoid the sandworms attacking or squeezing each other due to excessive weight difference.

[0060] 2. Transportation process: The goods will be transported in an insulated box, with the temperature inside the box controlled at 10-20℃.

[0061] Pretreated sandworms and seawater are packed into an insulated box at a ratio of 1:1.3-1.8. The insulated box contains absorbent polymer resin cotton.

[0062] The specific steps are as follows:

[0063] (1) Packing: Quickly pack the pretreated sandworms and pre-cooled seawater into boxes. Figure 2 The incubator shown contains sandworms in a 1:1.5 ratio with seawater.

[0064] The insulated box includes a foam box body 1, with an internal partition 3. The inner side and top wall of the foam box body 1 are covered with a plastic film 2 to prevent sandworms from crawling out. The inner side of the foam box body 1 is lined with absorbent polymer resin 4. The absorbent polymer resin 4 prevents the sandworms from being squeezed and damaged during transportation. The absorbent polymer resin 4 is pre-cooled and soaked in seawater. Ice packs are placed inside the partition 3, but the ice packs do not come into direct contact with the sandworms.

[0065] (2) Transportation environment: The temperature should be controlled at 10-20℃ throughout the transportation process. The ice pack should be replaced every 48 hours to maintain the low temperature inside the box.

[0066] 3. Post-transportation acclimatization process:

[0067] The insulated box was moved to a low-temperature space and seawater was sprayed regularly to raise the seawater temperature from 10-15℃ to 18-22℃ and adjust the seawater salinity from 15-25 to the salinity of the ecological restoration area.

[0068] After the adaptation period, screening and disinfection will be carried out.

[0069] Inducing burrowing: The disinfected sandworms are placed in a container with a sloping substrate. The container has an inlet near the bottom and an outlet near the top. The inlet is connected to a water pump. The water pump simulates tides to induce the sandworms to actively burrow.

[0070] The substrate consists of fine sand mixed with shell fragments and particles smaller than 1 mm, with a thickness of 10-15 cm.

[0071] The water pump is equipped with a timer, with a water inlet and outlet cycle of 5-8 hours.

[0072] Add diatomaceous earth solution to the container, with an algae density of 1×10⁻⁶. 3 -1×10 4 The cells / mL concentration is 2-5 ml / L.

[0073] The key to the post-transport acclimatization process is to alleviate transportation stress and induce natural burrowing behavior by creating a suitable substrate environment, thereby quickly restoring burrowing and habitat behaviors. The specific steps are as follows:

[0074] (1) Adaptation and acclimatization: After transportation, the insulated box was moved to the low-temperature chamber, and the box lid was slowly opened. Seawater was sprayed every 2 hours for 6-8 hours, with the seawater temperature gradually increasing from 10-15℃ to 18-22℃ and the seawater salinity gradually adjusted from 15-25 to the salinity of the ecological restoration area. This temperature and salinity adaptation and acclimatization can significantly reduce the stress response of sandworms and reduce the damage to body segments caused by violent twisting.

[0075] (2) Screening and disinfection: After adaptation, remove damaged segments ( Figure 3 , Figure 4 Sandworms in good condition. Figure 5 , Figure 6 Transfer to a disinfection box for immersion for no more than 30 minutes. The disinfection box contains citric acid (1.2 g / L) or vibrio disinfectant (1.0 g / L).

[0076] (3) Inducing burrowing: After disinfection, place the sandworms in a container with a substrate. The substrate is fine sea sand with a particle size of less than 1 mm, mixed with a small amount of shell fragments to enhance air permeability. The sand layer is sloping and the thickness is controlled at 10-15 cm to provide suitable space for the sandworms to construct U-shaped burrows.

[0077] like Figure 2 As shown, container 5 has an inlet 51 near the lower part of sand layer 6 and an outlet 52 at the higher part of sand layer 6. Container 5 is connected to a water pump equipped with a timer. A 6-hour "dry-dry + soaking" cycle simulates tidal action, inducing sandworms to burrow and feed. Diatomaceous algae such as *Scirpus scoparia* and *Skeletonema stenoptera* (algae density 1×10⁻⁶) are added to the pumped water. 3 -1×10 4 (Addition of cells / mL, at a rate of 2-5 ml / L) to stabilize the microbial environment and provide initial food. These methods work together to induce and promote sandworms to actively burrow in the sand, restoring their natural benthic lifestyle.

[0078] The transportation method in this embodiment innovatively utilizes "low osmotic pressure stimulation" to achieve efficient mud and sand discharge, reducing the inherent risks during transportation. Furthermore, through optimized substrate preparation and ecological induction methods, it ensures that sandworms can quickly burrow into the sand and settle after release, ultimately providing a living organism guarantee for large-scale benthic biological restoration projects.

[0079] Using the transportation method of this embodiment, the body segment damage rate of *Pterygota bisporus* after transportation can be reduced to below 8%, and the burrowing behavior is rapid, with the burrowing time shortened to 2 minutes, significantly improving overall vitality and adaptability.

[0080] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions claimed by the present invention.

Claims

1. A method for transporting *Nematocystis jirovecii* for benthic bioremediation, characterized in that, The method includes: Pre-transportation processing: Stop feeding the animals; Harvesting sandworms after a set time; The sandworms are stimulated to expel mud and sand by low osmotic pressure: the cleaned sandworms are transferred to a temporary holding box with a moist mat at the bottom. Low osmotic pressure seawater is injected at regular intervals. The temperature of the low osmotic pressure seawater is higher than that of the seawater at the excavation site, and the salinity of the low osmotic pressure seawater is lower than that of the seawater at the excavation site. Transportation process: The sandworms are packed into insulated boxes for transportation, and the temperature inside the insulated boxes is controlled at 10-20℃; Post-transport acclimatization process: The insulated box was moved to a low-temperature space and seawater was sprayed regularly. The seawater temperature was raised from 10-15℃ to 18-22℃ and the seawater salinity was adjusted from 15-25 to the salinity of the ecological restoration area. After the adaptation period, screening and disinfection will be carried out. Inducing burrowing: The disinfected sandworms are placed in a container with a sloping substrate. The container has a water inlet near the lower part of the substrate and a drain near the higher part of the substrate. The water inlet is connected to a water pump. The water pump simulates tides to induce the sandworms to actively burrow in the sand.

2. The method for transporting *Nematocystis jirovecii* for benthic bioremediation according to claim 1, characterized in that, The harvested sandworms are screened and cleaned.

3. The method for transporting *Pterygodon spp.* for benthic bioremediation according to claim 2, characterized in that, During cleaning, the water is washed with seawater of the same temperature and salinity as the extraction pool.

4. The method for transporting *Nematocystis jirovecii* for benthic bioremediation according to claim 1, characterized in that, Regularly inject low-osmotic-pressure seawater into the holding tank, control the seawater depth in the holding tank to 2-3cm, and keep the holding tank environment dark.

5. The method for transporting *Nematocystis jirovecii* for benthic bioremediation according to claim 1, characterized in that, The water in the temporary holding tank should be changed regularly, with each change replacing 70%-100% of the total water volume.

6. The method for transporting *Pterygodon spp.* for benthic bioremediation according to claim 1, characterized in that, After being graded by weight, the sandworms were placed in insulated boxes for transportation, with the weight difference of the sandworms in the same insulated box being less than 20%.

7. The method for transporting *Nematocystis jirovecii* for benthic bioremediation according to claim 1, characterized in that, Pretreated sandworms and seawater are packed into an insulated box at a ratio of 1:1.3-1.

8. The insulated box contains absorbent polymer resin cotton.

8. The method for transporting *Nematocystis jirovecii* for benthic bioremediation according to claim 1, characterized in that, The substrate is fine sand mixed with shell fragments and with a particle size of less than 1 mm, and the thickness of the substrate is 10-15 cm.

9. The method for transporting *Pterygodon spp.* for benthic bioremediation according to claim 1, characterized in that, The water pump is a timer-equipped pump, with a water inlet and outlet cycle of 5-8 hours.

10. The method for transporting *Pterygodon spp.* for benthic bioremediation according to claim 1, characterized in that, Diatomaceous earth solution was added to the container, with an algae density of 1×10⁻⁶. 3 -1×10 4 The cells / mL concentration is 2-5 ml / L.

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