Multi-path switching deep sea microorganism in-situ culture device and culture method

Abstract

The invention discloses a multi-path switching deep sea microorganism in-situ culture device, which comprises a rack, and at least one microorganism culture cabin, a seawater exchange assembly, a power supply and a control module which are arranged on the rack, the microorganism culture cabin is provided with a water inlet flow channel and a water outlet flow channel, and a movable piston is arranged in the microorganism culture cabin; the water outlet circulation is opened and closed through the movement of the piston; solid or colloidal culture media with different types or components are placed in the different microorganism culture cabins and are used for in-situ culture of different types of microorganisms, so that in-situ parallel or contrast culture of various microorganisms can be realized through one-time distribution; and the seawater exchange assembly is used for selectively pumping deep sea in-situ seawater to the at least one microorganism culture cabin and driving the piston to move. According to the device, a programmable control seawater active exchange system and a plurality of independent piston type culture cabins are integrated, so that multi-path, anti-pollution and controllable in-situ culture is realized.

Description

Technical Field

[0001] This invention relates to the field of deep-sea equipment, specifically a multi-channel switching in-situ culture device and method for deep-sea microorganisms. Background Technology

[0002] The deep-sea environment harbors abundant extremophile resources, holding significant value in research on the origin of life and applications of biotechnology. Microbial culture is a crucial means of revealing their ecological functions and metabolic mechanisms. Traditional laboratory culture methods have significant limitations: for example, drastic environmental changes during deep-sea sample transfer can easily lead to microbial inactivation or dormancy, resulting in resource "capture loss"; laboratory conditions cannot completely simulate the in-situ environment, making it impossible to culture the vast majority of microorganisms, which become "microbial dark matter." In-situ culture technology is a microbial culture method that maintains chemical signal exchange and population interactions among microorganisms, and can improve the yield of difficult-to-culture microorganisms. It is currently a research hotspot, but existing deep-sea in-situ culture devices have shortcomings in multi-condition parallel culture and sample recovery and contamination prevention.

[0003] Most existing deep-sea in-situ culture devices still employ a passive culture design based on diffusion principles. These devices typically have an open structure, which, while relatively simple to deploy and retrieve, also presents significant drawbacks: during descent and ascent, they are highly susceptible to contamination by microorganisms from the upper and middle water layers. Once these invasive organisms enter the culture environment, they significantly alter the original structure and composition of the seabed in-situ microbial community, affecting the accuracy of experimental results. Some improved devices attempt to use a "seabed opening and retrieval sealing" mechanical structure to reduce the risk of contamination during deployment. However, in practical use, this structure often fails to achieve sufficient convection exchange between the culture medium surface and the surrounding seawater, thus failing to effectively simulate the dynamic material and energy exchange processes in the natural deep-sea environment, limiting the accuracy of ecological research and biological resource extraction.

[0004] In multiple parallel or comparative experiments, existing devices generally suffer from insufficient precision in controlling culture conditions such as the fluid environment, making it difficult to guarantee consistency of conditions between experimental groups. More seriously, due to inadequate flow path design or physical isolation, cross-contamination of culture medium or microorganisms can easily occur between different culture units, introducing uncontrolled interference variables, thereby significantly increasing inter-group errors and reducing the reliability and repeatability of experimental results.

[0005] On the other hand, many in-situ culture devices are designed for single culture conditions or a limited number of samples, resulting in relatively limited functionality. When research requires multivariate regulation (such as comparative experiments with different nutrient substrates), multiple independent culture devices typically need to be deployed simultaneously. This strategy not only significantly increases the equipment cost, overall volume, and weight of deep-sea scientific expeditions but also introduces greater complexity and risk to deployment, manipulation, and retrieval operations. Furthermore, even when multiple devices are deployed in the same area, it is difficult to ensure that the macroscopic environment of each device is completely consistent, thus affecting the scientific validity and comparability of experimental conclusions.

[0006] Therefore, there is an urgent need to develop a microbial culture device capable of flexibly switching and independently controlling multiple culture units in an in-situ deep-sea environment. Key technical challenges include precise regulation of in-situ seawater exchange, simultaneous control of multiple comparative experiments, and sample recovery with contamination prevention. This will meet the needs of parallel comparative studies, targeted high-throughput screening, and environmental factor responses, while ensuring the device's stability and operational reliability under high-pressure extreme environments. The successful development of this device will significantly improve the efficiency and scientific rigor of in-situ deep-sea microbial culture experiments, providing strong technical support for in-depth exploration of the deep-sea microbial world. Summary of the Invention

[0007] The purpose of this invention is to overcome the shortcomings of the prior art and propose a multi-channel switching deep-sea microbial in-situ culture device and culture method. Its core lies in integrating a programmable controllable active seawater exchange system with multiple independent piston-type culture chambers to achieve multi-channel, pollution-proof, and controllable in-situ culture.

[0008] To achieve the above objectives, the technical solution specifically adopted by the present invention is as follows: A multi-channel switching in-situ culture device for deep-sea microorganisms includes a frame and at least one microbial culture chamber, a seawater exchange component, and a power supply and control module mounted on the frame. The microbial culture chamber is equipped with an inlet channel and an outlet channel. The microbial culture chamber is equipped with a movable piston. The outlet channel is opened and closed by the movement of the piston. Different types or components of solid or colloidal culture media are placed in different microbial culture chambers for in-situ culture of different types of microorganisms, so as to realize the in-situ parallel or comparative culture of multiple microorganisms with one placement. The seawater exchange assembly is used to selectively pump deep-sea in-situ seawater to the at least one microbial culture chamber and drive the movement of the piston. A power supply and control module is used to supply power to the seawater exchange component and control its operation.

[0009] Preferably, the microbial culture chamber includes a culture chamber cylinder, a front end cap and a rear end cap detachably and sealed to both ends of the culture chamber cylinder, a large piston movably disposed within the culture chamber cylinder, and a small piston penetrating the large piston; the large piston is sealed to the inner wall of the culture chamber cylinder, and its end face facing the front end cap has an annular groove for containing culture medium; the small piston includes a piston rod and a limiting disk, the piston rod is a hollow structure and its side wall has a through hole, the small piston penetrates the large piston such that the limiting disk is located on the side of the large piston facing the front end cap, and one end of the piston rod extends out of the large piston on the side facing the rear end cap.

[0010] Preferably, the large piston has a magnetic ring on its end face facing the rear end cover, and the rear end cover has a magnetic ring plate on its end face facing the large piston that can attract the magnetic ring.

[0011] Preferably, the front cover is provided with a water inlet, which is connected to the seawater exchange component through a fluid pipeline to form the water inlet channel of the microbial culture chamber.

[0012] Preferably, the front cover is also provided with a cleaning discharge port that communicates with the interior of the microbial culture chamber, and the cleaning discharge port is provided with a shut-off valve.

[0013] Preferably, the seawater exchange assembly includes a deep-sea gear pump and a multi-channel switching valve; the multi-channel switching valve has an inlet connected to the outlet of the deep-sea gear pump and multiple outlets, each of the outlets being selectively fluidly connected to an inlet of one of the microbial culture chambers.

[0014] Preferably, the multi-channel switching valve includes a stepper motor and a one-inlet-multiple-outlet valve head. The one-inlet-multiple-outlet valve head includes a valve sleeve, a central rotor rotatably disposed within the valve sleeve, and a sealing gasket disposed within the valve sleeve. The valve sleeve has a cavity for the central rotor to rotate. A water inlet hole is provided at the center of the top side of the valve sleeve, and the water inlet hole communicates with the cavity. Several water outlet holes are provided around the water inlet hole on the top side of the valve sleeve, and the water outlet holes communicate with the cavity. The central rotor has an L-shaped flow channel. The inlet of the L-shaped flow channel is connected to the water inlet hole. The outlet of the L-shaped flow channel can be connected to different radial channels on the valve sleeve by its rotation, thereby realizing the switching of the flow path.

[0015] Preferably, the motor of the deep-sea gear pump and the stepper motor of the multi-channel switching valve are both located in a separate motor compartment, which is pressure balanced with the external environment through a pressure balancing oil bladder.

[0016] Preferably, the pressure balancing oil bladder is connected to the interior of the motor compartment through a hollow titanium alloy tube, the top of the pressure balancing oil bladder is provided with an exhaust port with a one-way valve, and the motor compartment is provided with an oil inlet for filling insulating oil.

[0017] Preferably, the power supply and control module includes a control circuit, a battery, and a signal receiver housed in the power supply compartment. The control circuit is configured to control the operating timing of the seawater exchange component according to a preset program. The signal receiver is used to receive external trigger signals and instruct the control circuit to perform preset operations.

[0018] This invention also provides a method for in situ culture of deep-sea microorganisms, comprising: The large piston containing the culture medium was moved to the front cover of the microbial culture chamber, and the device was deployed to the target deep-sea environment. The power supply and control module controls the seawater exchange component to selectively pump deep-sea in-situ seawater into at least one of the microbial culture chambers. The pumped in-situ seawater drives a large piston in the microbial culture chamber to move from the front cover to the rear cover, thereby creating a culture space in the chamber and bringing the culture medium into contact with the in-situ seawater for in-situ microbial culture. Meanwhile, the inlet and outlet channels of the microbial culture chamber are connected, and seawater is pumped in intermittently according to the preset program to ensure that the seawater in the microbial culture chamber is fully exchanged with the outside seawater during the culture process. After the culture is completed, the device is recovered and the cultured sample is obtained.

[0019] First, this invention proposes a method for in-situ microbial culture to prevent sample contamination. Through a unique piston-type structure design of the microbial culture chamber, and utilizing in-situ seawater to drive the piston movement and create the culture space, it solves the problem of microorganisms from the upper layer of seawater being introduced into the seabed for contamination. It also addresses the issue of seawater circulation during the culture process, ensuring sufficient exchange of seawater on the culture medium surface, maintaining the characteristics of the in-situ seabed environment, and guaranteeing the stability of the microbial community. Second, a signal receiver is incorporated to determine the termination of culture or the removal of the equipment from the seabed, and to shut off the motor and power supply, preventing external microorganisms from entering the sample chamber during retrieval. This effectively ensures the quality of the in-situ cultured microbial samples.

[0020] Another significant advantage of this invention is that it proposes a highly efficient and sealed deep-sea multi-channel switching valve to ensure the uniformity of seawater exchange in each culture chamber during the deep-sea culture process. This not only enables the acquisition of multiple in-situ culture samples of seabed microorganisms with a single deployment, greatly improving efficiency and reducing the cost of oceanographic research, but also allows for the conduct of multi-dimensional scientific experiments that meet the needs of comparative experimental research, targeted high-throughput screening, and environmental factor response, thus expanding the scope of application of the equipment. Attached Figure Description

[0021] Figure 1 This is a schematic diagram of a multi-channel switching in-situ culture device for deep-sea microorganisms in an embodiment of the present invention.

[0022] Figure 2 for Figure 1 A schematic diagram of the layout and structure of the microbial culture chamber.

[0023] Figure 3 for Figure 1 A schematic diagram of the working structure of the microbial culture chamber.

[0024] Figure 4 for Figure 1 Schematic diagram of a gear pump for deep-sea operations.

[0025] Figure 5 for Figure 1 Schematic diagram of a multi-channel switching valve for deep-sea applications.

[0026] Figure 6 for Figure 1 Schematic diagram of the valve head structure of a multi-channel switching valve for deep-sea applications.

[0027] Figure 7 for Figure 1 Schematic diagram of the power supply and control module.

[0028] Figure label: 1-Microbial culture chamber, 2-Seawater exchange assembly, 3-Power supply and control module, 4-Rack; 11-Cultivation chamber body, 12-Front end cover, 13-Rear end cover, 14-Stop valve, 15-Large piston, 131-Magnetic ring plate, 151-Annular groove, 152-Magnetic ring, 16-Small piston, 161-Limiting disc, 162-Piston column, 163-Round hole; 21-Deep-sea gear pump, 211-Gear pump motor compartment, 212-DC brushless motor, 213-Thrust bearing, 214-Deep groove ball bearing, 215-Chec seal ring, 216-Gear pump head; 23-Pressure balance oil bladder, 231-Exhaust port, 232-Check valve, 233-Hollow titanium alloy tube, 234-Oil inlet; 22-Multi-channel switching valve, 221-Switching valve motor compartment, 222-Stepper motor, 223-Motor mounting base, 224-One-inlet-multiple-outlet valve head, 2241-Center rotor, 2242-Valve sleeve, 2243-Mushroom-shaped sealing gasket; 31-Battery compartment, 32-Control circuit, 33-Signal receiver. Detailed Implementation

[0029] The present invention will now be described in detail with reference to specific embodiments. These embodiments will help those skilled in the art to further understand the present invention, but do not limit the invention in any way. It should be noted that, unless otherwise specified, the embodiments and features described in the present invention can be combined with each other. Example

[0030] This embodiment provides a multi-channel switching in-situ culture device for deep-sea microorganisms, such as... Figure 1 As shown, its overall structure mainly includes a frame 4, as well as multiple microbial culture chambers 1, a seawater exchange assembly 2, and a power supply and control module 3 integrated and installed on the frame 4. The frame 4 provides mechanical support and overall rigidity for the entire device, facilitating assembly, debugging on the deck of a research vessel, and deployment and retrieval via deep-sea cables or carriers.

[0031] Specifically, refer to Figure 2 and Figure 3 The microbial culture chamber 1 is the core unit that directly contains the culture medium and conducts in-situ microbial culture. Each microbial culture chamber 1 is set up independently and is not interconnected to prevent cross-contamination.

[0032] The microbial culture chamber 1 mainly consists of a culture chamber cylinder 11, a front end cover 12, and a rear end cover 13. The cylinder 11 is a hollow cylindrical structure with a smooth surface and high-precision machined inner wall, typically made of seawater-resistant, high-strength materials such as titanium alloy or special stainless steel. The front end cover 12 and the rear end cover 13 are tightly fixed to both ends of the cylinder 11 by circumferentially distributed bolts. O-rings or similar radial sealing elements are installed on the contact surfaces between the end covers and the cylinder, as well as on other static sealing parts of the end covers, to ensure a reliable sealed space inside the chamber under the high pressure environment of the deep sea.

[0033] Furthermore, the front cover 12 is provided with two flow channel holes. One of them serves as a water inlet, with a two-way compression fitting installed on its outer side. This inlet is connected to the outlet of the multi-channel switching valve 22 of the seawater exchange assembly 2 via a flexible, pressure-resistant pipe, such as a silicone tube, forming the water inlet flow channel. The other flow channel hole serves as a cleaning discharge port, on which a shut-off valve 14 is installed. Its function and principle are as follows: after the device is recovered to the deck, samples need to be removed and the chamber cleaned. At this time, the shut-off valve 14 can be opened to connect to an external freshwater source for rinsing. Wastewater is discharged from this port, avoiding the cumbersome process of disassembling the front cover for cleaning, improving operational efficiency, and protecting the main sealing surface of the end cover.

[0034] In a further embodiment, the culture chamber is equipped with a movable piston assembly, including a large piston 15 and a small piston 16 passing through it.

[0035] Specifically, the large piston 15 has an outer diameter that precisely matches the inner diameter of the cylinder 11 and is equipped with a sealing ring (such as a Glyd ring or Step seal) to ensure both flexible movement and effective sealing during axial sliding within the cylinder. An annular groove 151 is machined on the end face of the large piston 15 facing the front cover 12. The principle and function of this design is to allow pre-prepared solid (such as agar) or colloidal culture media of different compositions to be stably placed in this groove during preparation before deployment. The groove structure prevents the culture media from shifting, falling off, or deforming under piston movement or seawater erosion, ensuring the stability and consistency of the culture interface. A magnetic ring 152 made of permanent magnet material is embedded on the end face of the large piston 15 facing the rear cover 13.

[0036] Small piston 16: Includes piston rod 162 and a limiting disk 161 located at one end of piston rod 162. Piston rod 162 is a hollow tubular structure, and its side wall near the limiting disk 161 has one or more radial through holes 163. Small piston 16 passes through the through hole in the center of large piston 15, so that the limiting disk 161 is located on the side of large piston 15 facing the front end cap, while the end of piston rod 162 extends towards the rear end of large piston 15.

[0037] Magnetic Adsorption Structure: A magnetic ring 131 made of a magnetically conductive material (such as soft iron) is fixedly installed on the end face of the rear end cover 13 facing the interior of the tank. Its working principle and effect are as follows: When in-situ seawater is pumped into the tank, pushing the large piston 15 towards the rear end cover 13 and approaching it to a certain distance, a strong magnetic attraction is generated between the magnetic ring 152 on the large piston 15 and the magnetic ring 131 on the rear end cover 13. This force can overcome the inertia of the piston movement and possible minor water flow disturbances, ensuring that the large piston 15 quickly, stably, and tightly adheres to the rear end cover 13. This precise positioning determines the maximum culture space (i.e., the volume between the inner side of the front cover and the front face of the large piston) in the working state of the culture chamber. On the other hand, when the device completes the culture and is ready for recovery, even if the external water pump stops working, the magnetic adsorption can "lock" the large piston 15 in the position of the rear cover, preventing it from moving unexpectedly due to changes in external pressure or shaking during the recovery and floating process. This maintains the preset open state of the water outlet channel. At the same time, since there is no active pump pressure, it effectively suppresses the active backflow of seawater from the upper layer, forming a key link in pollution prevention.

[0038] Flow channel formation principle: such as Figure 3As shown, in the initial state before the device is deployed, the large piston 15 is typically located near the front cover 12. After operation begins, seawater is pumped in through the inlet, acting on the front face of the large piston 15 and pushing it towards the rear cover 13. When the large piston 15 finally adheres to the rear cover 13 under magnetic force, the through-hole 163 on the side wall of the small piston 16 is fully exposed within the culture space enclosed by the front face of the large piston, the inner wall of the cylinder, and the front cover. At this point, the seawater flow path is as follows: seawater enters through the inlet → flows through the culture space and fully contacts the culture medium in the annular groove 151 → enters the hollow interior of the piston rod 162 through the through-hole 163 on the side wall of the small piston → flows out from the opening at the end of the piston rod and is discharged back to the external environment through the corresponding flow channel hole on the rear cover 13. This design cleverly utilizes the movement of the piston to automatically open and form a dual "inlet-out" flow path, achieving continuous seawater exchange during the culture process.

[0039] Further settings in this embodiment are described in reference to... Figure 4 , Figure 5 and Figure 6 The seawater exchange component 2 is responsible for drawing in-situ seawater from the deep-sea environment and distributing it to designated microbial culture chambers according to a predetermined procedure. It is accomplished by the coordinated operation of a deep-sea gear pump 21 and a multi-channel switching valve 22.

[0040] Deep-sea gear pump 21: Provides stable power for seawater pumping. Its core feature is the isolation of the power unit from the high-pressure seawater. A DC brushless motor 212 is sealed within the gear pump motor housing 211. The motor shaft 212 is stably supported by a combination of a thrust bearing 213 (primarily bearing axial load and providing axial positioning) and a deep groove ball bearing 214 (primarily bearing radial load), and protrudes stably from the front wall of the motor housing. At the shaft protrusion point, a Cherry lip seal 215 provides dynamic sealing; this sealing method offers excellent sealing performance and lifespan under high-pressure, low-speed conditions. The shaft drives the external gear pump head 216 to rotate, generating suction to pump seawater in place.

[0041] Multi-channel switching valve 22: Enables one-to-many flow path selection and control. Stepper motor 222 is sealed inside the switching valve motor compartment 221 and fixed by motor mounting bracket 223. Its shaft also passes through the compartment wall with two O-ring seals, driving the one-inlet-multiple-outlet valve head 224.

[0042] Valve head structure principle: refer to Figure 6 The valve head mainly comprises a valve sleeve 2242, a central rotor 2241, and a mushroom-shaped sealing gasket 2243. The valve sleeve 2242 has a cylindrical cavity at its center, with a main water inlet at the top center (connecting to the gear pump outlet), and multiple independent water outlets evenly distributed around the top (each connecting to the inlet of a culture chamber). The central rotor 2241 can be precisely rotatably positioned within the cavity, and its interior is machined with an L-shaped flow channel (the vertical inlet and horizontal outlet are perpendicular).

[0043] Switching Process and Anti-Crossflow Design: Stepper motor 222 receives control signals and drives the central rotor 2241 to rotate at a specific angle. When water needs to be supplied to a certain culture chamber, the rotor is controlled to rotate until the horizontal outlet of its L-shaped flow channel aligns with the corresponding channel position on the side wall of valve sleeve 2242 for the target culture chamber. At this time, the seawater flows through the following path: main inlet → vertical inlet of rotor L-shaped flow channel → horizontal outlet → corresponding channel on valve sleeve side wall → corresponding outlet at top of valve sleeve → target culture chamber. Mushroom-shaped sealing gasket 2243 is made of elastic materials such as nitrile rubber and is installed on the inside of each channel opening on the side wall of valve sleeve. Its protruding head design ensures that it can elastically press against the outer wall of the rotor to close the channel when the rotor is not aligned; when the rotor is aligned, the outer wall of the rotor pushes the sealing gasket head to deform and open the flow channel. This design effectively prevents seawater leakage from non-working channels or crossflow between different channels, ensuring the independence of each culture chamber and precise control of seawater exchange.

[0044] Pressure balancing system: To ensure the safety of the motor room under the water pressure of thousands of meters deep in the ocean, pressure balancing oil bladders 23 are provided for both the gear pump motor room 211 and the switching valve motor room 221.

[0045] Connection and Balancing Principle: The pressure balancing oil bladder 23 is connected and sealed to the corresponding motor compartment via a hollow titanium alloy tube 233 using threads (O-rings are provided on the contact surfaces). The compartment is filled with insulating oil (such as transformer oil). The oil bladder 23 is made of flexible material and is directly exposed to the deep-sea environment. Its core function is: as the device's diving depth increases and the external water pressure increases, the seawater pressure compresses the oil bladder 23, reducing its volume. This pressure is then transmitted to the insulating oil inside the compartment through the titanium alloy tube 233, ensuring real-time balance between the oil pressure inside the compartment and the external water pressure. This prevents the motor compartment shell from deforming or rupturing due to a large pressure difference.

[0046] Oil Filling and Venting: Each engine compartment is equipped with an oil filling port 234 for filling the compartment with insulating oil from land. The top of the oil bladder 23 has a vent 231, on which a one-way valve 232 is installed (allowing only gas to escape from inside the oil bladder). The operation and principle are as follows: When filling with insulating oil, open the plugs on the oil filling port and the vent vent. Oil is injected through the oil filling port, pushing the air inside the compartment and oil bladder out through the vent vent. When the oil is full and oil overflows from the vent vent, it indicates that the air has been completely vented. At this point, tighten the vent vent plug (one-way valve 232 prevents seawater backflow), and then seal the oil filling port. This process ensures that there is no gas present in the pressure balance system, because the compressibility of gas can severely affect the pressure balance effect and even lead to danger.

[0047] Further settings in this embodiment are described in reference to... Figure 7 The power supply and control module 3 provides energy to the device and implements intelligent control.

[0048] Understandably, all electrical components are housed in a separate deep-sea power compartment 31. This compartment is equipped with a high-capacity rechargeable lithium battery pack to power the DC brushless motor 212, the stepper motor 222, and the control circuit 32 itself. The power compartment 31 is connected to the two motor compartments via deep-sea pressure-resistant watertight cables and connectors.

[0049] It should be noted that the control circuit 32 (usually based on a microcontroller) is the control center. Before deployment, technicians can program the control circuit 32 via a dedicated interface to set complex operating sequences, such as: the initial start-up delay time after the device is deployed to the seabed; the daily start-up time and duration of each operation of the deep-sea gear pump 21; and the sequence and dwell time of the stepper motor 222 of the multi-channel switching valve 22 when switching to different channels in each cycle (i.e., the water injection duration and cycle of each culture chamber). This allows multiple culture chambers to receive automatic, cyclical seawater injection according to preset, potentially different, "time-flow" programs, completely simulating the predetermined culture conditions.

[0050] Understandably, signal reception and safe termination are achieved through a signal receiver 33 installed externally in the power compartment 31. This receiver can be configured to respond to specific signals, such as flashes of light at a certain frequency, changes in magnetic fields of a certain intensity, or the action of a mechanical trigger mechanism. Its core function and resulting safety effect are as follows: when the cultivation experiment ends or emergency recovery is required, researchers can transmit a light signal from the ship to the underwater environment (or apply a magnetic signal via the unmanned submersible). Upon receiving this "termination" command, the signal receiver 33 immediately sends an interrupt signal to the control circuit 32. The control circuit 32 then executes a preset safety procedure: immediately stopping all motors and cutting off the main power output. This mechanism ensures that the seawater exchange components completely cease operation during the critical stage of the device's ascent and recovery, completely eliminating the possibility of introducing contaminated seawater from the upper layers due to accidental pump operation, while also conserving battery power and ensuring safe recovery. In addition, the control circuit 32 can also be programmed to monitor the pressure sensor data inside the cabin or in the environment. When the pressure value remains stable, it can be determined that the device has successfully landed. When the pressure value suddenly drops to a certain value (such as 98 kPa, which means the device has risen 10 meters), it can automatically determine that the device has left the seabed and is being recovered, and autonomously execute the above-mentioned termination procedure as a backup safety mechanism in case of signal trigger failure. Example

[0051] This embodiment provides a method for in-situ culture of deep-sea microorganisms with multiple switching channels. The method uses a device for in-situ culture of deep-sea microorganisms with multiple switching channels provided in Embodiment 1, and the method is as follows: (1) Land preparation: According to the experimental design, different solid / colloidal culture media were placed in the annular grooves 151 of the large piston 15 of multiple microbial culture chambers 1. The large piston 15 was moved to the front cover 12 and fitted to it to assemble the culture chambers. The system consisting of the gear pump motor chamber 211, the switching valve motor chamber 221 and the pressure balancing oil bladder 23 was filled with insulating oil and the system was vented and sealed. The two motor chambers were filled with insulating oil through the oil inlet 234. The water inlets of each culture chamber were connected to the outlets of the multi-channel switching valve 22 through hoses. The culture program was set in the control circuit 32 (e.g., water was injected twice a day, and each time the valve body supplied water to each chamber for 10 minutes).

[0052] (2) Deployment and Start-up: The entire device is deployed to the target seabed using a deep-sea lander or underwater robot. After the preset first start-up time is reached, the control circuit 32 is automatically powered on, performs a system self-check, and then starts the deep-sea gear pump 21 and the multi-channel switching valve 22 according to the program. Seawater is pumped into the currently selected culture chamber, which drives the large piston to move, eventually forming a stable culture space and starting seawater circulation.

[0053] (3) In-situ culture: Over the next few days to weeks, the device operates completely autonomously. The control circuit 32 strictly follows the preset timing sequence, cyclically switching the multi-channel switching valve 22 so that each culture chamber can obtain fresh in-situ seawater exchange regularly and quantitatively. The seawater flows over the surface of the culture medium, bringing nutrients and carrying away metabolic products. At the same time, the reciprocating motion of the piston also promotes interface renewal, realizing dynamic and active simulation of the deep-sea in-situ environment, which greatly improves the survival and growth probability of difficult-to-culture microorganisms.

[0054] (4) Termination and Recovery: After the experiment, a light signal is sent from the ship (or the preset incubation time is waited for). The signal receiver 33 triggers the safety termination procedure, all motors stop, and the power is cut off. The device is recovered and floats to the surface. During this process, since the water pump has stopped and the large piston is magnetically attracted to the rear end cover, the passive aspiration of microorganisms in the water layer during the ascent is effectively prevented, thus protecting the precious in-situ sample.

[0055] (5) Sample acquisition: After the device is recovered to the deck, push the large piston forward to squeeze out the culture medium, bottle it for storage, and then remove the front cover 12 of the culture chamber. Take out the culture medium block that has completed in situ culture from the annular groove 151 of the large piston 15 for subsequent analysis. Open the shut-off valve 14 to easily flush the chamber.

[0056] In summary, this specific embodiment clearly and completely discloses all the key technical features, connection relationships, working principles, and operating steps of the device of the present invention. Based on this description, and combined with known knowledge of deep-sea equipment manufacturing, sealing, material selection, and control programming, those skilled in the art are fully capable of manufacturing and using this device to achieve multi-channel, pollution-proof, and program-controllable in-situ culture of deep-sea microorganisms, achieving the expected technical effects of the present invention. The present invention effectively solves the problems mentioned in the background art, such as pollution, single path, insufficient exchange, and poor environmental adaptability, through its innovative piston flow control structure, highly reliable multi-channel switching valve, and intelligent program control.

Claims

1. A multi-channel switching in-situ culture device for deep-sea microorganisms, characterized in that, It includes a frame (4) and at least one microbial culture chamber (1) mounted on the frame (4), a seawater exchange assembly (2), and a power supply and control module (3). The microbial culture chamber (1) is provided with an inlet channel and an outlet channel. The microbial culture chamber (1) is equipped with a movable piston. The outlet channel is opened and closed by the movement of the piston. Different microbial culture chambers (1) contain different types or components of solid or colloidal culture media for in-situ culture of different types of microorganisms. The seawater exchange assembly (2) is used to selectively pump deep-sea in-situ seawater to at least one microbial culture chamber (1) and drive the movement of the piston; The power supply and control module (3) is used to supply power to the seawater exchange component (2) and control its operation.

2. The multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 1, characterized in that, The microbial culture chamber (1) includes a culture chamber cylinder (11), a front end cap (12) and a rear end cap (13) detachably and sealed to both ends of the culture chamber cylinder (11), a large piston (15) movably disposed inside the culture chamber cylinder (11), and a small piston (16) penetrating the large piston (15); the large piston (15) is sealed to the inner wall of the culture chamber cylinder (11), and its end face facing the front end cap (12) is provided with an annular groove (151) for accommodating culture medium; the small piston (16) includes a piston rod (162) and a limiting disk (161), the piston rod (162) is a hollow structure and its side wall is provided with a through hole (163), the small piston (16) penetrates the large piston (15) so that the limiting disk (161) is located on the side of the large piston (15) facing the front end cap (12), and one end of the piston rod (162) extends out of the large piston (15) on the side facing the rear end cap (13).

3. The multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 2, characterized in that, The large piston (15) has a magnetic ring (152) on its end face facing the rear end cover (13), and the rear end cover (13) has a magnetic ring plate (131) on its end face facing the large piston (15) that can attract the magnetic ring (152).

4. The multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 2, characterized in that, The front cover (12) is provided with a water inlet, which is connected to the seawater exchange component (2) through a fluid pipeline to form the water inlet channel of the microbial culture chamber (1).

5. The multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 4, characterized in that, The front cover (12) is also provided with a cleaning discharge port that communicates with the interior of the microbial culture chamber (1), and the cleaning discharge port is provided with a shut-off valve (14).

6. The multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 1, characterized in that, The seawater exchange assembly (2) includes a deep-sea gear pump (21) and a multi-channel switching valve (22); the multi-channel switching valve (22) has an inlet connected to the outlet of the deep-sea gear pump (21) and multiple outlets, each of the outlets being used to selectively fluidly communicate with an inlet of one of the microbial culture chambers (1).

7. A multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 6, characterized in that, The multi-channel switching valve (22) includes a stepper motor (222) and a single-inlet multi-outlet valve head (224). The single-inlet multi-outlet valve head (224) includes a valve sleeve (2242), a central rotor (2241) rotatably disposed within the valve sleeve (2242), and a sealing gasket (2243) disposed within the valve sleeve (2242). The valve sleeve (2242) has a cavity for the central rotor (2241) to rotate. The valve sleeve (2242) has a water inlet hole at the center of its top side, which is connected to the cavity. The valve sleeve (2242) has several water outlet holes around the water inlet hole on its top side, which are connected to the cavity. The central rotor (2241) has an L-shaped flow channel inside. The inlet of the L-shaped flow channel is connected to the water inlet hole. The outlet of the L-shaped flow channel can be connected to different radial channels on the valve sleeve (2242) by its rotation, thereby realizing the switching of the flow path.

8. A multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 6, characterized in that, The motor of the deep-sea gear pump (21) and the stepper motor (222) of the multi-channel switching valve (22) are both located in an independent motor compartment, which is pressure balanced with the external environment through a pressure balancing oil bladder (23).

9. A multi-channel switching in-situ culture device for deep-sea microorganisms according to claim 8, characterized in that, The pressure balancing oil bladder (23) is connected to the interior of the motor compartment through a hollow titanium alloy tube (233). The top of the pressure balancing oil bladder (23) is provided with an exhaust port (231) with a one-way valve (232). The motor compartment is provided with an oil inlet (234) for filling insulating oil.

10. The multi-channel switching deep-sea microbial in-situ culture device according to claim 1, characterized in that, The power supply and control module (3) includes a control circuit (32), a battery and a signal receiver (33) housed in the power supply compartment (31). The control circuit (32) is configured to control the operation sequence of the seawater exchange component (2) according to a preset program. The signal receiver (33) is used to receive external trigger signals and instruct the control circuit (32) to perform preset operations.

11. A method for in-situ culture of deep-sea microorganisms, characterized in that, The method using the multi-channel switching deep-sea microbial in-situ culture device as described in any one of claims 2 to 10 includes: The large piston (15) containing the culture medium was moved to the front cover (12) of the microbial culture chamber (1), and the device was deployed to the deep-sea target environment. The power supply and control module (3) controls the seawater exchange component (2) to selectively pump deep-sea in-situ seawater into at least one of the microbial culture chambers (1). The pumped in-situ seawater drives the large piston in the microbial culture chamber (1) to move from the front cover to the rear cover, so as to form a culture space in the chamber, so that the culture medium comes into contact with the in-situ seawater for in-situ microbial culture; Meanwhile, the inlet and outlet channels of the microbial culture chamber are connected, and seawater is pumped in intermittently according to the preset program, so that the seawater in the microbial culture chamber (1) can be fully exchanged with the external environment during the culture process. After the culture is completed, the device is recovered and the cultured sample is obtained.

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