Electronic one-tube automatic detection system and detection method for streptococcus agalactiae or gonococcus

Abstract

The invention provides an electronic one-tube automatic detection system and a detection method for streptococcus agalactiae or gonococci, through an LAMP and CRISPR-Cas12a combined mediated one-tube visual streptococcus agalactiae or gonococci detection system, in combination with an electronic automation technology, an LAMP reaction mixture and a CRISPR-Cas12a reaction mixture are mixed to form a one-tube reaction system, and the one-tube reaction system is used for detecting the streptococcus agalactiae or gonococci. Automatic mixing, preheating reaction and fluorescence detection of various detection reagents and samples are realized, and visual display is performed on detection results by adopting visual equipment. Visual detection of bacteria is achieved by detecting fluorescence signals, rapid positive or quantity detection results can be provided for patients and other users, and clinical detection efficiency is improved.

Description

Technical Field

[0001] The present invention relates to the field of biological detection technology, and in particular to an electronic and automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae, a detection method for Streptococcus agalactiae or Neisseria gonorrhoeae, an electronic device and a computer-readable storage medium. Background Art

[0002] LAMP (loop-mediated isothermal amplification) and CRISPR-Cas12a together constitute a highly sensitive and specific detection method in bacterial detection.

[0003] ‌The role of LAMP (loop-mediated isothermal amplification) in bacterial detection‌: LAMP is a simple, rapid, accurate and inexpensive gene amplification method, which is particularly suitable for pathogen detection.

[0004] It uses 4-6 primers that recognize specific regions of the target DNA and amplify them under constant temperature conditions to produce a large amount of DNA products, thereby achieving rapid detection of bacteria.

[0005] LAMP technology has the characteristics of high sensitivity and specificity, and can amplify extremely small amounts of DNA in a short period of time, making the detection more accurate.

[0006] ‌Role of CRISPR-Cas12a in Bacterial Detection‌: CRISPR-Cas12a is a powerful detection and identification tool that originates from the adaptive immune system of bacteria and archaea and can resist attacks from foreign mobile genetic factors. In bacterial detection, CRISPR-Cas12a can bind to specific crRNA, identify and cut target DNA, thereby achieving accurate detection of bacteria.

[0007] ‌Combination application of LAMP and CRISPR-Cas12a‌: Combining LAMP and CRISPR-Cas12a technology can achieve high sensitivity and high specificity detection of bacteria. For example, by amplifying the target DNA of bacteria through LAMP and then using CRISPR-Cas12a to cut and identify them, accurate detection of bacteria can be achieved.

[0008] This method can not only detect highly pathogenic Helicobacter pylori in saliva samples, but can also be applied to the detection of other bacteria, such as African swine fever virus. The combined use can significantly improve the sensitivity and accuracy of detection, while achieving rapid and on-site detection, which is of great significance for the early detection and prevention and control of diseases.

[0009] Streptococcus agalactiae and gonococci play an important role in fertility. At present, the detection of Streptococcus agalactiae and gonococci is mainly blood or secretion culture. Biochemical tests such as catalase test and CAMP test can also help to distinguish them from other streptococci. However, clinical laboratory cultivation takes a lot of time and operation steps, involving the circulation management of many culture, experimental and other reaction equipment, so it is inconvenient to use, and each operation is scattered and difficult to centrally manage.

[0010] LAMP and CRISPR-Cas12a play an important role in bacterial detection. Their combined application provides a new technical means for the rapid and accurate detection of bacteria, and can bring new and rapid detection methods for Streptococcus agalactiae or Neisseria gonorrhoeae, achieving the effect of negative detection or rapid detection of quantity distribution; however, there is currently no relevant mature reaction system to show that it can be used in actual clinical detection of Streptococcus agalactiae or Neisseria gonorrhoeae. Summary of the invention

[0011] In order to solve the technical problems existing in the prior art, the present invention provides the following technical solutions: On the one hand, an electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae is provided, the system comprising a support 1, on which the following are arranged: Controller 3, used for logic calculation and control; A LAMP primer module, used for providing a LAMP primer mixed solution to a bacterial reaction module; A CRISPR-Cas12a primer module, used to provide a CRISPR-Cas12a primer mixed solution to a bacterial reaction module; A bacterial reaction module, for preparing a LAMP-CRISPR-Cas12a reaction system according to the LAMP primer mixed solution and the CRISPR-Cas12a primer mixed solution; and, after the Streptococcus agalactiae or gonococci are injected, providing a mixed reaction environment for the Streptococcus agalactiae or gonococci and the LAMP-CRISPR-Cas12a reaction system, and outputting the corresponding bacterial reaction solution to the detection dish 17; A fluorescence detector 12, used for detecting the fluorescence signal of the bacterial reaction liquid in the detection dish 17 and feeding it back to the controller 3, wherein the controller 3 processes the fluorescence signal and generates a detection value corresponding to the Streptococcus agalactiae or Neisseria gonorrhoeae and sends and displays the detection value on an electronic screen; Power supply, used for power supply; The LAMP primer module, the CRISPR-Cas12a primer module, the bacterial reaction module, the fluorescence detector 12 and the power supply are electrically connected to the controller 3 respectively.

[0012] Preferably, the LAMP primer module comprises a first reaction tube 2, a heater A4 and an electronic valve A5, wherein: The first reaction tube 2 is vertically arranged on the support 1, and a LAMP primer mixed solution is prepared therein; The heater A4 is arranged on the outer side of the first reaction tube 2 to provide preheating service; The electronic valve A5 is disposed at the bottom of the first reaction tube 2 and is used to control the dosage of the LAMP primer mixed solution delivered to the bacterial reaction module; The heater A4 and the electronic valve A5 are electrically connected to the controller 3 respectively.

[0013] Preferably, the CRISPR-Cas12a primer module comprises a second reaction tube 4, a heater B9 and an electronic valve B10, wherein: The second reaction tube 4 is vertically arranged on the support 1, and a CRISPR-Cas12a primer mixed solution is prepared therein; The heater B9 is arranged on the outer side of the second reaction tube 4 to provide preheating service; The electronic valve B10 is disposed at the bottom of the second reaction tube 4 and is used to control the dosage of the CRISPR-Cas12a primer mixture solution delivered to the bacterial reaction module; The heater B9 and the electronic valve B10 are electrically connected to the controller 3 respectively.

[0014] Preferably, it is characterized in that the bacterial reaction module comprises a third reaction tube 7, an injection port 8, a heater C13 and an electronic valve C14, wherein: The injection port 8 is provided at the top of the third reaction tube 7 and is used for injecting the extract of Streptococcus agalactiae or Neisseria gonorrhoeae; The third reaction tube 7 is vertically arranged on the support 1, and a pH 7.0 sterile sodium chloride-peptone buffer, a 0.9% sterile sodium chloride solution or a pH 7.0 sterile phosphate buffer is prepared therein; and the installation height of the third reaction tube 7 is lower than the LAMP primer module and the CRISPR-Cas12a primer module; The heater C13 is arranged on the outer side of the third reaction tube 7 to provide preheating service; The electronic valve C14 is disposed at the bottom of the third reaction tube 7 and is used to control the dosage of the bacterial reaction solution delivered to the detection dish 17; The heater C13 and the electronic valve C14 are electrically connected to the controller 3 , respectively.

[0015] Preferably, the testing dish 17 is provided with a first sterility testing platform 15 and a second sterility testing platform 16, which are used to hold the bacterial reaction solution and the comparison solution respectively.

[0016] Preferably, the fluorescence detector 12 is disposed on the support 1 via a flexible support 11 and is located above the detection dish 17 .

[0017] Preferably, two fluorescence detectors 12 are provided, which are respectively arranged directly above the first sterility detection platform 15 and the second sterility detection platform 16 , and the two fluorescence detectors 12 are isolated by a shading plate.

[0018] On the other hand, a method for detecting Streptococcus agalactiae or Neisseria gonorrhoeae is provided, which is implemented based on the above-mentioned electronic and tube-based automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae, and the method comprises: The operating parameters of each heater and electronic valve are pre-set in the controller 3; After the extract of Streptococcus agalactiae or Neisseria gonorrhoeae is injected, the system is activated and the system reaction is controlled by the controller 3; When the preset time is reached, the fluorescence detector 12 is controlled to detect the fluorescence signal of the bacterial reaction solution and feed it back to the controller 3, and the controller 3 processes the fluorescence signal and generates a detection value corresponding to the Streptococcus agalactiae or Neisseria gonorrhoeae; The detected value is sent and displayed on an electronic screen.

[0019] On the other hand, an electronic device is provided, comprising: a processor; a memory, wherein the memory stores computer-readable instructions, and when the computer-readable instructions are executed by the processor, any one of the above-mentioned electronically controlled automatic detection systems and methods for Streptococcus agalactiae or Neisseria gonorrhoeae is implemented.

[0020] On the other hand, a computer-readable storage medium is provided, wherein at least one instruction is stored in the storage medium, and the at least one instruction is loaded and executed by a processor to implement any one of the above-mentioned electronic and automatic detection systems and methods for Streptococcus agalactiae or Neisseria gonorrhoeae.

[0021] The beneficial effects brought about by the technical solution provided by the embodiment of the present invention include at least: The present invention combines the electronic automation technology to mix the LAMP reaction mixture and the CRISPR-Cas12a reaction mixture to form a "one-tube" reaction system through the "one-tube" visual Streptococcus agalactiae or gonococcus detection system mediated by LAMP and CRISPR-Cas12a. Through the designed "one-tube" electronic automation reaction system, through electronic automation control, the automatic mixing, preheating reaction and fluorescence detection of various detection reagents and samples are realized, and the detection results are visualized using a visualization device. The whole process can be controlled by electronic automation, and the changes in the fluorescence signal can be automatically monitored in real time, and the presence or absence of Streptococcus agalactiae and / or gonococci can be judged according to the changes in fluorescence intensity. The analysis results are realized. If the fluorescence signal is significantly enhanced, it indicates that the sample contains Streptococcus agalactiae and / or gonococci; if the fluorescence signal does not change significantly, it indicates that the sample does not contain target bacteria. The system automatically records and reports the test results.

[0022] The present invention utilizes a "one-tube" visual bacterial detection system mediated by LAMP (loop-mediated isothermal amplification) and CRISPR-Cas12a (CRISPR-associated nuclease), which is an efficient and rapid bacterial detection method. The system combines the isothermal amplification capability of LAMP and the specific cutting activity of CRISPR-Cas12a, and realizes visual detection of bacteria by detecting fluorescent signals, which can provide patients and other users with rapid positive or quantitative detection results, thereby improving clinical detection efficiency. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0024] Figure 1 This is an application structure diagram of an electronic one-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae provided by an embodiment of the present invention; Figure 2 is a schematic diagram of a system control circuit provided by an embodiment of the present invention; Figure 3 is a flow chart of a detection method provided by an embodiment of the present invention; Figure 4 It is a structural schematic diagram of an electronic device provided by an embodiment of the present invention. DETAILED DESCRIPTION

[0025] The technical solution of the present invention is described below in conjunction with the accompanying drawings.

[0026] In the embodiments of the present invention, words such as "exemplarily" and "for example" are used to indicate examples, illustrations or explanations. Any embodiment or design described as "example" in the present invention should not be interpreted as being more preferred or more advantageous than other embodiments or designs. Specifically, the use of the word "example" is intended to present the concept in a specific way. In addition, in the embodiments of the present invention, the meaning expressed by "and / or" can be both, or it can be either of the two.

[0027] In the embodiments of the present invention, "image" and "picture" can sometimes be used interchangeably. It should be noted that when the difference between them is not emphasized, the meanings they intend to express are the same. "of", "corresponding, relevant" and "corresponding" can sometimes be used interchangeably. It should be noted that when the difference between them is not emphasized, the meanings they intend to express are the same.

[0028] In the embodiments of the present invention, sometimes a subscript such as W1 may be mistakenly written as a non-subscript such as W1. When the difference is not emphasized, the meanings to be expressed are consistent.

[0029] In order to make the technical problems, technical solutions and advantages to be solved by the present invention more clear, a detailed description will be given below with reference to the accompanying drawings and specific embodiments.

[0030] In this embodiment, the experimental reagents, equipment, etc. involved can be provided by a biological laboratory or purchased on the market, and this embodiment does not limit this.

[0031] The design of LAMP primers and CRISPR-Cas12aLAMP primers and the preparation of corresponding reagents can be designed and prepared by the user according to the experimental requirements and conventional biochemical reaction steps. The primer sequences for the detection of Streptococcus agalactiae or Neisseria gonorrhoeae can be prepared by the biochemical laboratory. This scheme does not involve the description of biological primer probes, but only proposes a biological detection system and method.

[0032] The extraction and preparation of Streptococcus agalactiae or Neisseria gonorrhoeae can be collected from animal models, etc., or the corresponding bacterial cultures can be purchased.

[0033] The embodiment of the present invention provides an electronic and automatic detection system and method for Streptococcus agalactiae or Neisseria gonorrhoeae. The method can be implemented by an electronic device, which can be a terminal or a server. Figure 1 The structure diagram of the electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae shown in the figure, the system comprises a bracket 1, and the bracket 1 is provided with: Controller 3, used for logic calculation and control; A LAMP primer module, used for providing a LAMP primer mixed solution to a bacterial reaction module; A CRISPR-Cas12a primer module, used to provide a CRISPR-Cas12a primer mixed solution to a bacterial reaction module; A bacterial reaction module, for preparing a LAMP-CRISPR-Cas12a reaction system according to the LAMP primer mixed solution and the CRISPR-Cas12a primer mixed solution; and, after the Streptococcus agalactiae or gonococci are injected, providing a mixed reaction environment for the Streptococcus agalactiae or gonococci and the LAMP-CRISPR-Cas12a reaction system, and outputting the corresponding bacterial reaction solution to the detection dish 17; A fluorescence detector 12, used for detecting the fluorescence signal of the bacterial reaction liquid in the detection dish 17 and feeding it back to the controller 3, wherein the controller 3 processes the fluorescence signal and generates a detection value corresponding to the Streptococcus agalactiae or Neisseria gonorrhoeae and sends and displays the detection value on an electronic screen; Power supply, used for power supply; The LAMP primer module, the CRISPR-Cas12a primer module, the bacterial reaction module, the fluorescence detector 12 and the power supply are electrically connected to the controller 3 respectively.

[0034] The present invention is provided with a plurality of reaction detection modules on a bracket, which can realize a mixed reaction of Streptococcus agalactiae or Neisseria gonorrhoeae bacteria and their primer solutions, and output the reaction solution, and detect the fluorescence signal of the bacteria after the amplification reaction by a fluorescence detector, and then the fluorescence signal is processed by a chip in a controller, such as signal A / D conversion, filtering, and operational amplifier (these circuits are common integrated circuits on the control circuit board, so this embodiment does not describe the processing circuit in the controller in detail), etc., so as to obtain the corresponding fluorescence value, and reflect the concentration or quantity of the corresponding bacteria by the fluorescence value.

[0035] Each reaction tube can be fixed on the bracket by means of a buckle, a hoop or a strap. Its installation position, quantity, etc. can be configured according to the requirements of this experiment or the laboratory. The controller uses an STM32 / 64-bit series of single-chip microcomputer MCU chips, etc.

[0036] The specific system design is as follows: 1. Design primers: According to the specific gene sequence of the target bacteria, the internal and external primers required for LAMP amplification are designed.

[0037] At the same time, the crRNA of CRISPR-Cas12a is designed so that it can specifically recognize and cut specific sequences in the LAMP amplification product.

[0038] 2. Prepare the reaction system: Mix together the components required for LAMP amplification, including enzymes, primers, dNTPs, buffer, etc.

[0039] Add CRISPR-Cas12a protein, crRNA, and a fluorescent reporter system (such as a fluorescent dye-labeled DNA probe or ssDNA-FQ).

[0040] The conversion of the number of bacteria based on the detection value of the fluorescent marker mainly depends on the relationship between the intensity of the fluorescent signal and the ATP content in the bacteria. The following is a description of the conversion: ‌Relationship between fluorescence signal and ATP content‌: Fluorescent markers (such as luciferase) can react with ATP in bacteria to generate luciferase complexes and emit fluorescent signals.

[0041] The intensity of the fluorescence signal is proportional to the ATP content, that is, the more ATP there is, the stronger the fluorescence signal‌1.

[0042] ‌Conversion steps‌: First, the fluorescence signal intensity of the sample is measured using a fluorescence detector.

[0043] Next, based on the known relationship between fluorescence signal intensity and ATP content (usually established through a standard curve or mathematical model), the fluorescence signal intensity is converted into ATP content.

[0044] Finally, since living cells such as bacteria contain the same amount of ATP, the number of bacteria can be indirectly calculated based on the ATP content.

[0045] 3. Optimize reaction conditions: Determine the optimal temperature and time for LAMP amplification and CRISPR-Cas12a cleavage.

[0046] Ensure that the reaction system can proceed stably under "one-tube" conditions.

[0047] 4. Testing process 4.1 Sample processing: Extract bacterial DNA or RNA from the sample to be tested.

[0048] Add the extracted nucleic acid directly to the prepared reaction system.

[0049] 4.2 Isothermal amplification and cleavage: At a constant temperature (e.g., 60-65°C), LAMP primers guide isothermal amplification of nucleic acids.

[0050] The amplified product is recognized and cut by CRISPR-Cas12a, releasing the signal of the fluorescent reporter system.

[0051] 4.3 Fluorescence detection: Use a fluorescence detector (such as a fluorescence spectrophotometer, fluorescence microscope, or portable fluorescence detector) to detect the fluorescence signal in the reaction system.

[0052] The intensity of the fluorescent signal is proportional to the number of target bacteria in the sample, thus achieving quantitative detection.

[0053] 4.4 Results Analysis: The presence and quantity of target bacteria in the sample can be determined based on the strength of the fluorescent signal.

[0054] A threshold value can be set to distinguish between positive and negative results, thereby improving the accuracy of the test (the present invention sets a comparison group (fluorescence value of the comparison fluid)).

[0055] The controller is integrated with an electronic screen that can be used to display the test value. The contrast coefficient between the intensity of the corresponding fluorescent signal and the number of target bacteria in the sample, the bacterial number threshold for distinguishing positive and negative results, etc. can be determined according to the clinical diagnostic criteria. It is also possible to configure a (wired or wireless) communication module or interface to transmit the MCU calculation results to the laboratory PC (terminal), report and record the corresponding bacterial test results, and use them for subsequent bacterial negative analysis, quantity comparison analysis, etc.

[0056] Therefore, the present invention combines the electronic automation technology to mix the LAMP reaction mixture and the CRISPR-Cas12a reaction mixture through the "one-tube" visualization of Streptococcus agalactiae or gonococcus detection system mediated by LAMP and CRISPR-Cas12a to form a "one-tube" reaction system. Through the designed "one-tube" electronic automation reaction system, various detection reagents and samples are automatically mixed, preheated and reacted, and fluorescence detection is performed through electronic automation control, and the detection results are visualized using a visualization device. The whole process can be controlled by electronic automation, and the changes in the fluorescence signal can be automatically monitored in real time, and the presence or absence of Streptococcus agalactiae and / or gonococci can be judged according to the changes in fluorescence intensity. The analysis results are realized. If the fluorescence signal is significantly enhanced, it indicates that the sample contains Streptococcus agalactiae and / or gonococci; if the fluorescence signal does not change significantly, it indicates that the sample does not contain target bacteria. The system automatically records and reports the test results.

[0057] The composition structure and principle of the present invention will be further described below.

[0058] The modules below are basically composed of a reaction tube, a heater and a solenoid valve. The corresponding solution can be injected into the reaction tube by injection or other means; and a solenoid valve is set at the bottom of the reaction tube, which is controlled by the controller to control the output. Heating and preheating services are also provided. On the outer side of the reaction tube near the bottom, a corresponding heater is installed. The heater is a ring-shaped deployment structure, which can be heated under the control of the controller to preheat the solution in the reaction tube.

[0059] The size and structure of each reaction tube are not limited in this embodiment. The input and output ports, etc., can be designed or purchased according to the needs. The specific models and installation structures of the heater and electronic valve can be selected by yourself. The corresponding sealing and fixed installation methods can be provided by the equipment provider or manufacturer.

[0060] There are many types of modules that can provide heating. Here are some common types of heating modules: Instant heating module: This type of heating module is usually used in water dispensers, pipeline machines and other equipment. It adopts flat-plate wire-wrapped heating element or rare earth thick film wire-wrapped rapid heating technology, which can quickly heat water or fluid.

[0061] Infrared heating module: The infrared heating module uses the thermal effect of infrared rays for heating and is widely used in various occasions that require fast and uniform heating, such as drying, curing, heat treatment, etc.

[0062] Industrial heating module: This type of heating module is usually used in industrial production equipment, such as kilns, heating furnaces, etc. It is made of high-temperature resistant materials such as zirconium-containing aluminum silicate fiber, can withstand high temperature environment and provide stable heating effect.

[0063] ‌PTC heating module‌: PTC heating module is a heating module that uses PTC thermistor as the heating element. It has the characteristics of automatic temperature control, safety and reliability, energy saving and high efficiency. It is widely used in automobiles, home appliances, industrial equipment and other fields.

[0064] Induction heating module: Induction heating technology is a non-contact heating method that converts electrical energy into thermal energy through the principle of electromagnetic induction. It is usually used in processes such as heating, smelting, and welding of metal materials. The induction heating module is the key component to realize this technology.

[0065] The controller can control the corresponding working time of the heater and electronic valve of each reaction tube according to the preset program instructions. For example, at the beginning of work, the controller controls the heaters of the two primer modules to work for 1 to 3 minutes, preheats the primer mixture solution therein, and then controls the electronic valve to work for 3 to 5 seconds before and after, so as to output the corresponding dose of primer mixture solution to the total reaction tube. In the total reaction tube, the preheating can also be controlled by the controller in the above manner. The preheating temperature, time, etc. can be confirmed according to the working parameters of the selected heater. The specific amount of reaction to be added can be calculated according to the demand, and the opening and closing of the electronic valve is controlled by the controller. Finally, part of the bacterial culture solution after the amplification reaction in the total reaction tube is discharged onto the test dish. After a certain period of time (such as 3 minutes of finalization), the fluorescence detector is controlled to start, perform fluorescence detection, and feedback the corresponding detection signal.

[0066] Using the above method, the solution in this system can also be divided into several batches. By detecting the fluorescence detection value of each batch, the average is finally taken as the final bacterial detection value, so as to improve the accuracy. For example, 10ml of solution is configured in the reaction tube of the primer module. The electronic valve can be controlled to output 2ml of solution each time according to 5 batches. After reaction and detection, a value is recorded by the MCU controller. After receiving the fluorescent signal, the administrator can clean or replace the test dish, and then the controller will delay for 5 minutes before conducting the next batch of tests. During this period, the administrator will replace or clean the test dish.

[0067] Preferably, the LAMP primer module comprises a first reaction tube 2, a heater A4 and an electronic valve A5, wherein: The first reaction tube 2 is vertically arranged on the support 1, and a LAMP primer mixed solution is prepared therein; The heater A4 is arranged on the outer side of the first reaction tube 2 to provide preheating service; The electronic valve A5 is disposed at the bottom of the first reaction tube 2 and is used to control the dosage of the LAMP primer mixed solution delivered to the bacterial reaction module; The heater A4 and the electronic valve A5 are electrically connected to the controller 3 respectively.

[0068] Combined with Figure 2 As shown in the figure, this is the schematic diagram of the control circuit of this system (each electronic device is in Figure 1 (Referring to the reference numerals in the figure, not marked here).

[0069] Preferably, the CRISPR-Cas12a primer module comprises a second reaction tube 4, a heater B9 and an electronic valve B10, wherein: The second reaction tube 4 is vertically arranged on the support 1, and a CRISPR-Cas12a primer mixed solution is prepared therein; The heater B9 is arranged on the outer side of the second reaction tube 4 to provide preheating service; The electronic valve B10 is disposed at the bottom of the second reaction tube 4 and is used to control the dosage of the CRISPR-Cas12a primer mixture solution delivered to the bacterial reaction module; The heater B9 and the electronic valve B10 are electrically connected to the controller 3 respectively.

[0070] Preferably, it is characterized in that the bacterial reaction module comprises a third reaction tube 7, an injection port 8, a heater C13 and an electronic valve C14, wherein: The injection port 8 is provided at the top of the third reaction tube 7 and is used for injecting the extract of Streptococcus agalactiae or Neisseria gonorrhoeae; The third reaction tube 7 is vertically arranged on the support 1, and a pH 7.0 sterile sodium chloride-peptone buffer, a 0.9% sterile sodium chloride solution or a pH 7.0 sterile phosphate buffer is prepared therein; and the installation height of the third reaction tube 7 is lower than the LAMP primer module and the CRISPR-Cas12a primer module; The heater C13 is arranged on the outer side of the third reaction tube 7 to provide preheating service; The electronic valve C14 is disposed at the bottom of the third reaction tube 7 and is used to control the dosage of the bacterial reaction solution delivered to the detection dish 17; The heater C13 and the electronic valve C14 are electrically connected to the controller 3 , respectively.

[0071] In bacterial testing, the preparation solution before mixing the bacterial extract can be pH 7.0 sterile sodium chloride-peptone buffer, 0.9% sterile sodium chloride solution or pH 7.0 sterile phosphate buffer, etc. These solutions are usually used as diluents. In microbial limit tests, they can help dilute the antibacterial product by appropriate multiples to reduce the concentration of antibacterial components in the product, thereby weakening or eliminating its inhibitory effect on microorganisms, making subsequent bacterial testing more accurate and reliable. The specific choice of preparation solution may need to be determined based on the specific requirements of the experiment, the type of bacteria, and the characteristics of the antibacterial product.

[0072] Preferably, the testing dish 17 is provided with a first sterility testing platform 15 and a second sterility testing platform 16, which are used to hold the bacterial reaction solution and the comparison solution respectively.

[0073] By setting the contrast solution, the current bacterial detection and analysis results can be quickly distinguished by fluorescence signal detection, calculation and comparison, and by referring to the detection value of the contrast solution. For example, the contrast solution uses a sterile solution of standard concentration or a bacterial solution with a known positive structure, which can quickly compare the test results and distinguish between positive and negative.

[0074] When analyzing bacterial counts, you can also take the following steps: ‌Establish a standard curve‌: Before the experiment, a standard curve of fluorescence signal intensity and bacterial number needs to be established. This is usually done by using a bacterial sample of known concentration for amplification and detection, and then plotting the relationship between fluorescence signal intensity and bacterial number.

[0075] ‌Calculate the number of bacteria‌: Based on the measured fluorescence signal intensity, find the corresponding number of bacteria on the standard curve. This usually involves converting the fluorescence signal intensity to ATP content, and then calculating the number of bacteria based on the relationship between ATP content and bacterial number.

[0076] ‌Calibration and Validation‌: The fluorescence detector is calibrated regularly to ensure the accuracy of the test results. At the same time, bacterial samples with known concentrations are used for validation to evaluate the reliability of the detection method.

[0077] Preferably, the fluorescence detector 12 is disposed on the support 1 via a flexible support 11 and is located above the detection dish 17 .

[0078] Preferably, two fluorescence detectors 12 are provided, which are respectively arranged directly above the first sterility detection platform 15 and the second sterility detection platform 16 , and the two fluorescence detectors 12 are isolated by a shading plate.

[0079] Here, two fluorescence detectors 12 are respectively shielded to avoid light interference and improve accuracy. The flexible bracket can be a multi-axis connecting rod, which can rotate in three dimensions to adjust the height and angle of the fluorescence detector 12 for easy operation. The flexible bracket can be hinged on the bracket 1.

[0080] Therefore, the present invention utilizes a "one-tube" visual bacterial detection system mediated by LAMP (loop-mediated isothermal amplification) and CRISPR-Cas12a (CRISPR-associated nuclease), which is an efficient and rapid bacterial detection method. The system combines the isothermal amplification capability of LAMP and the specific cutting activity of CRISPR-Cas12a, and realizes visual detection of bacteria by detecting fluorescent signals, which can provide patients and other users with rapid positive or quantitative detection results, thereby improving clinical detection efficiency.

[0081] like Figure 3As shown, on the other hand, a method for detecting Streptococcus agalactiae or Neisseria gonorrhoeae is provided, which is implemented based on the above-mentioned electronic and tube-based automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae, and the method comprises: S1, pre-set the working parameters of each heater and electronic valve in the controller 3; S2, after the extract of Streptococcus agalactiae or Neisseria gonorrhoeae is injected, the system is activated and the system reaction is controlled by the controller 3; S3, when the preset time is reached, the fluorescence detector 12 is controlled to detect the fluorescence signal of the bacterial reaction solution and feed it back to the controller 3, and the controller 3 processes the fluorescence signal and generates a detection value corresponding to the Streptococcus agalactiae or Neisseria gonorrhoeae; S4. Sending and displaying the detected value on an electronic screen.

[0082] Please implement the above steps in conjunction with the above system description, and will not be repeated here.

[0083] Figure 4 is a schematic diagram of the structure of an electronic device provided by an embodiment of the present invention, such as Figure 4 As shown, the electronic device 410 may include a first processor 2001 .

[0084] Optionally, the electronic device 410 may further include a memory 2002 and a transceiver 2003 .

[0085] The first processor 2001, the memory 2002 and the transceiver 2003 may be connected via a communication bus.

[0086] Combine the following Figure 4 The components of the electronic device 410 are described in detail: The first processor 2001 is the control center of the electronic device 410, and may be a processor or a general term for multiple processing elements. For example, the first processor 2001 is one or more central processing units (CPUs), or may be application specific integrated circuits (ASICs), or may be one or more integrated circuits configured to implement the embodiments of the present invention, such as one or more microprocessors (digital signal processors, DSPs), or one or more field programmable gate arrays (field programmable gate arrays, FPGAs).

[0087] Optionally, the first processor 2001 can execute various functions of the electronic device 410 by running or executing a software program stored in the memory 2002 and calling data stored in the memory 2002 .

[0088] In a specific implementation, as an embodiment, the first processor 2001 may include one or more CPUs, such as Figure 4 CPU0 and CPU1 are shown in FIG.

[0089] In a specific implementation, as an embodiment, the electronic device 410 may also include multiple processors, such as Figure 4 The first processor 2001 and the second processor 2004 are shown in FIG. Each of these processors may be a single-core processor (single-CPU) or a multi-core processor (multi-CPU). The processor here may refer to one or more devices, circuits, and / or processing cores for processing data (e.g., computer program instructions).

[0090] The memory 2002 is used to store the software program for executing the solution of the present invention, and is controlled to be executed by the first processor 2001. The specific implementation method can refer to the above method embodiment, which will not be repeated here.

[0091] Optionally, the memory 2002 may be a read-only memory (ROM) or other types of static storage devices that can store static information and instructions, a random access memory (RAM) or other types of dynamic storage devices that can store information and instructions, or an electrically erasable programmable read-only memory (EEPROM), a compact disc read-only memory (CD-ROM) or other optical disc storage, optical disc storage (including compressed optical disc, laser disc, optical disc, digital versatile disc, Blu-ray disc, etc.), a magnetic disk storage medium or other magnetic storage device, or any other medium that can be used to carry or store the desired program code in the form of instructions or data structures and can be accessed by a computer, but is not limited thereto. The memory 2002 may be integrated with the first processor 2001, or may exist independently and access the first processor 2001 through the interface circuit ( Figure 4 (not shown) is coupled to the first processor 2001, which is not specifically limited in this embodiment of the present invention.

[0092] The transceiver 2003 is used to communicate with a network device or a terminal device.

[0093] Optionally, the transceiver 2003 may include a receiver and a transmitter ( Figure 4 The receiver is used to implement a receiving function, and the transmitter is used to implement a sending function.

[0094] Optionally, the transceiver 2003 may be integrated with the first processor 2001, or may exist independently and communicate with the first processor 2001 through the interface circuit ( Figure 4 (not shown) is coupled to the first processor 2001, which is not specifically limited in this embodiment of the present invention.

[0095] It should be noted that Figure 4 The structure of the electronic device 410 shown in the figure does not constitute a limitation on the router, and the actual knowledge structure recognition device may include more or fewer components than those shown in the figure, or combine certain components, or arrange the components differently.

[0096] In addition, the technical effects of the electronic device 410 can refer to the technical effects of the electronic integrated automatic detection system and detection method for Streptococcus agalactiae or Neisseria gonorrhoeae described in the above method embodiments, which will not be repeated here.

[0097] It should be understood that the first processor 2001 in the embodiment of the present invention may be a central processing unit (CPU), and the processor may also be other general-purpose processors, digital signal processors (DSP), application specific integrated circuits (ASIC), field programmable gate arrays (FPGA) or other programmable logic devices, discrete gate or transistor logic devices, discrete hardware components, etc. The general-purpose processor may be a microprocessor or the processor may also be any conventional processor, etc.

[0098] It should also be understood that the memory in the embodiments of the present invention may be a volatile memory or a non-volatile memory, or may include both volatile and non-volatile memories. Among them, the non-volatile memory may be a read-only memory (ROM), a programmable read-only memory (PROM), an erasable programmable read-only memory (EPROM), an electrically erasable programmable read-only memory (EEPROM), or a flash memory. The volatile memory may be a random access memory (RAM), which is used as an external cache. By way of example and not limitation, many forms of random access memory (RAM) are available, such as static RAM (SRAM), dynamic random access memory (DRAM), synchronous DRAM (SDRAM), double data rate synchronous dynamic random access memory (DDR SDRAM), enhanced synchronous dynamic random access memory (ESDRAM), synchronous link DRAM (SLDRAM), and direct rambus RAM (DR RAM).

[0099] The above embodiments can be implemented in whole or in part by software, hardware (such as circuits), firmware or any other combination. When implemented by software, the above embodiments can be implemented in whole or in part in the form of a computer program product. The computer program product includes one or more computer instructions or computer programs. When the computer instructions or computer programs are loaded or executed on a computer, the process or function described in the embodiment of the present invention is generated in whole or in part. The computer can be a general-purpose computer, a special-purpose computer, a computer network, or other programmable device. The computer instructions can be stored in a computer-readable storage medium, or transmitted from one computer-readable storage medium to another computer-readable storage medium. For example, the computer instructions can be transmitted from one website, computer, server or data center to another website, computer, server or data center by wired (such as infrared, wireless, microwave, etc.). The computer-readable storage medium can be any available medium that can be accessed by a computer or a data storage device such as a server or data center that contains one or more available media sets. The available medium can be a magnetic medium (for example, a floppy disk, a hard disk, a tape), an optical medium (for example, a DVD), or a semiconductor medium. The semiconductor medium can be a solid-state hard disk.

[0100] It should be understood that the term "and / or" in this article is only a description of the association relationship of associated objects, indicating that there can be three relationships. For example, A and / or B can represent: A exists alone, A and B exist at the same time, and B exists alone. A and B can be singular or plural. In addition, the character " / " in this article generally indicates that the associated objects before and after are in an "or" relationship, but it may also indicate an "and / or" relationship. Please refer to the context for specific understanding.

[0101] In the present invention, "at least one" means one or more, and "plurality" means two or more. "At least one of the following" or similar expressions refers to any combination of these items, including any combination of single items or plural items. For example, at least one of a, b, or c can mean: a, b, c, ab, ac, bc, or abc, where a, b, and c can be single or multiple.

[0102] It should be understood that in various embodiments of the present invention, the size of the serial numbers of the above-mentioned processes does not mean the order of execution. The execution order of each process should be determined by its function and internal logic, and should not constitute any limitation on the implementation process of the embodiments of the present invention.

[0103] Those skilled in the art will appreciate that the units and algorithm steps of each example described in conjunction with the embodiments disclosed herein can be implemented in electronic hardware, or a combination of computer software and electronic hardware. Whether these functions are performed in hardware or software depends on the specific application and design constraints of the technical solution. Professional and technical personnel can use different methods to implement the described functions for each specific application, but such implementation should not be considered to be beyond the scope of the present invention.

[0104] Those skilled in the art can clearly understand that, for the convenience and brevity of description, the specific working processes of the above-described equipment, devices and units can refer to the corresponding processes in the aforementioned method embodiments and will not be repeated here.

[0105] In the several embodiments provided by the present invention, it should be understood that the disclosed devices, apparatuses and methods can be implemented in other ways. For example, the device embodiments described above are only schematic. For example, the division of the units is only a logical function division. There may be other division methods in actual implementation, such as multiple units or components can be combined or integrated into another device, or some features can be ignored or not executed. Another point is that the mutual coupling or direct coupling or communication connection shown or discussed can be through some interfaces, indirect coupling or communication connection of devices or units, which can be electrical, mechanical or other forms.

[0106] The units described as separate components may or may not be physically separated, and the components shown as units may or may not be physical units, that is, they may be located in one place or distributed on multiple network units. Some or all of the units may be selected according to actual needs to achieve the purpose of the solution of this embodiment.

[0107] In addition, each functional unit in each embodiment of the present invention may be integrated into one processing unit, or each unit may exist physically separately, or two or more units may be integrated into one unit.

[0108] If the functions are implemented in the form of software functional units and sold or used as independent products, they can be stored in a computer-readable storage medium. Based on this understanding, the technical solution of the present invention, or the part that contributes to the prior art, or the part of the technical solution, can be embodied in the form of a software product. The computer software product is stored in a storage medium, including several instructions for enabling a computer device (which can be a personal computer, server, or network device, etc.) to perform all or part of the steps of the methods described in various embodiments of the present invention. The aforementioned storage medium includes: U disk, mobile hard disk, read-only memory (ROM), random access memory (RAM), disk or optical disk, and other media that can store program codes.

[0109] The above is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art who is familiar with the technical field can easily think of changes or substitutions within the technical scope disclosed by the present invention, which should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention should be based on the protection scope of the claims.

Claims

1. An electronic and automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae, characterized in that: The system comprises a support (1), on which is arranged: Controller (3), used for logic calculation and control; A LAMP primer module, used for providing a LAMP primer mixed solution to a bacterial reaction module; A CRISPR-Cas12a primer module, used to provide a CRISPR-Cas12a primer mixed solution to a bacterial reaction module; A bacterial reaction module, used for preparing a LAMP-CRISPR-Cas12a reaction system according to the LAMP primer mixed solution and the CRISPR-Cas12a primer mixed solution; and, after the Streptococcus agalactiae or Neisseria gonorrhoeae is injected, providing a mixed reaction environment for the Streptococcus agalactiae or Neisseria gonorrhoeae and the LAMP-CRISPR-Cas12a reaction system, and outputting the corresponding bacterial reaction solution to the detection dish (17); A fluorescence detector (12) for detecting the fluorescence signal of the bacterial reaction liquid in the detection dish (17) and feeding it back to the controller (3); the controller (3) processes the fluorescence signal and generates a detection value corresponding to the Streptococcus agalactiae or Neisseria gonorrhoeae, and transmits and displays the detection value on an electronic screen; Power supply, used for power supply; The LAMP primer module, the CRISPR-Cas12a primer module, the bacterial reaction module, the fluorescence detector (12) and the power supply are electrically connected to the controller (3) respectively.

2. The electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to claim 1, characterized in that: The LAMP primer module comprises a first reaction tube (2), a heater A (4) and an electronic valve A (5), wherein: The first reaction tube (2) is vertically arranged on the support (1), and contains a LAMP primer mixed solution; The heater A (4) is arranged on the outer side of the first reaction tube (2) to provide preheating service; The electronic valve A (5) is arranged at the bottom of the first reaction tube (2) and is used to control the dosage of the LAMP primer mixed solution delivered to the bacterial reaction module; The heater A (4) and the electronic valve A (5) are electrically connected to the controller (3) respectively.

3. The electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to claim 1, characterized in that: The CRISPR-Cas12a primer module comprises a second reaction tube (4), a heater B (9) and an electronic valve B (10), wherein: The second reaction tube (4) is vertically arranged on the support (1), and contains a CRISPR-Cas12a primer mixed solution; The heater B (9) is arranged on the outer side of the second reaction tube (4) to provide preheating service; The electronic valve B (10) is disposed at the bottom of the second reaction tube (4) and is used to control the dosage of the CRISPR-Cas12a primer mixture solution delivered to the bacterial reaction module; The heater B (9) and the electronic valve B (10) are electrically connected to the controller (3) respectively.

4. The electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to claim 1, characterized in that: The bacterial reaction module comprises a third reaction tube (7), an injection port (8), a heater C (13) and an electronic valve C (14), wherein: The injection port (8) is disposed at the top of the third reaction tube (7) and is used for injecting the extract of Streptococcus agalactiae or Neisseria gonorrhoeae; The third reaction tube (7) is vertically arranged on the support (1), and contains a pH 7.0 sterile sodium chloride-peptone buffer, a 0.9% sterile sodium chloride solution, or a pH 7.0 sterile phosphate buffer; and the installation height of the third reaction tube (7) is lower than that of the LAMP primer module and the CRISPR-Cas12a primer module; The heater C (13) is arranged on the outer side of the third reaction tube (7) to provide preheating service; The electronic valve C (14) is arranged at the bottom of the third reaction tube (7) and is used to control the dosage of the bacterial reaction liquid delivered to the detection dish (17); The heater C (13) and the electronic valve C (14) are electrically connected to the controller (3) respectively.

5. The electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to claim 1, characterized in that: The detection dish (17) is provided with a first sterile detection platform (15) and a second sterile detection platform (16), which are used to hold the bacterial reaction solution and the comparison solution respectively.

6. The electronic-tube automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to claim 5, characterized in that: The fluorescence detector (12) is arranged on the support (1) via a flexible support (11) and is located above the detection dish (17).

7. The electronic and tube-based automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to claim 6, characterized in that: Two fluorescence detectors (12) are provided, which are respectively arranged directly above the first sterility detection platform (15) and the second sterility detection platform (16), and the two fluorescence detectors (12) are isolated by a light shielding plate.

8. A method for detecting Streptococcus agalactiae or Neisseria gonorrhoeae, which is implemented based on the electronic and automatic detection system for Streptococcus agalactiae or Neisseria gonorrhoeae according to any one of claims 1 to 7, characterized in that: The method comprises: The operating parameters of each heater and electronic valve are pre-set in the controller (3); After the extract of Streptococcus agalactiae or Neisseria gonorrhoeae is injected, the system is activated and the system reaction is controlled by the controller (3); When a preset time is reached, the fluorescence detector (12) is controlled to detect the fluorescence signal of the bacterial reaction liquid and feed it back to the controller (3), and the controller (3) processes the fluorescence signal and generates a detection value corresponding to the Streptococcus agalactiae or Neisseria gonorrhoeae; The detected value is sent and displayed on an electronic screen.

9. An electronic device, characterized in that: The electronic device comprises: processor; A memory having computer-readable instructions stored thereon, wherein when the computer-readable instructions are executed by the processor, the method according to any one of claims 1 to 7 is implemented.

10. A computer-readable storage medium, characterized in that: The computer-readable storage medium stores program codes, which can be called by a processor to execute the method according to any one of claims 1 to 7.

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