Imaging incubator capable of recording movement of bacterial cluster in real time

Abstract

The invention belongs to the field of bacterial movement monitoring, and particularly relates to an imaging incubator capable of recording bacterial cluster movement in real time, the incubator comprises a box body, a delay shooting system, a temperature control system and a humidity control system; the time-delay shooting system comprises a camera, a shading plate and a flash lamp, the camera automatically captures an image according to preset exposure time at each preset interval, and the flash lamp is turned on at each preset interval and corresponds to the time when the camera automatically captures the image. The periodic lighting of the flash lamp is precisely synchronized with the shooting time of the camera, so that the long-time high brightness is avoided, and the possible inhibition effect of light intensity on the growth of microorganisms is prevented. According to the device, the illumination requirement during shooting is guaranteed, the influence on the culture environment is reduced to the greatest extent, clear images are captured regularly under the condition that the growth of microorganisms is not interfered, and valuable data support is provided for subsequent research.

Description

Technical Field

[0001] The invention belongs to the field of bacterial movement monitoring, and in particular relates to an imaging incubator capable of recording bacterial cluster movement in real time. Background Art

[0002] Bacteria's motility is an important feature of their survival and adaptation to the environment. They use different modes of movement to explore and utilize the surrounding resources. These modes of movement include swimming, swarming, and sliding, each of which corresponds to the adaptation strategy of bacteria in a specific environment. For example, swimming allows bacteria to move in a liquid environment through the rotation of flagella, while sliding involves the movement of bacteria on solid surfaces, usually relying on the secretion of mucus or other surfactants. Among these modes of movement, swarming movement is particularly eye-catching. Swarming movement refers to the coordinated multicellular migration of bacteria on a wet surface. This process involves the collective behavior of bacterial groups, which achieve rapid expansion through the secretion of mucus and intercellular signaling. This mode of movement not only demonstrates the social interaction of bacteria, but is also essential for their spread in the natural environment and the infection process in the host. To better understand the relationship between bacterial movement and host health, scientists are exploring the speed and morphological patterns of colony expansion. These studies involve real-time imaging of bacterial group behavior and how they respond to challenges from the host immune system. Through these studies, we can reveal the molecular mechanisms of bacterial movement and how they affect the host's immune response and disease progression.

[0003] When studying bacterial swarming, a homogenized stool sample solution may take 10-48 hours to grow a typical clonal morphology of swarming bacteria, while a purified and identified typical swarming bacterial solution may take 8-16 hours to grow a full 9 cm diameter culture dish. In addition to monitoring the colony coverage time, we sometimes need to accurately understand the diffusion length of the colony edge, the specific time of branch formation, and the specific moment of cell elongation at the microscopic level during the lag phase of colony growth. In order to obtain these data, researchers must check the culture dish frequently. If the experiment is started during the day, the experimenter needs to perform continuous scanning for about 48 hours, especially at a frequency of 10-30 min / time. The manual operation is complicated, tedious, time-consuming and labor-intensive. At the same time, the existing preparation, imaging and quantitative analysis of bacterial swarming often rely on commercial equipment, such as Bruker in vivo imaging station. However, this equipment has been discontinued and is expensive in the second-hand market. For most microbiology laboratories, the complex functions provided by this equipment, including X-ray imaging and in vivo fluorescence imaging, are often unnecessary. These labs may have limited budgets or be unwilling to pay high prices for advanced features beyond their needs. Summary of the invention

[0004] In order to solve the problem that the observation process of bacterial cluster movement is cumbersome and the existing observation equipment is expensive.

[0005] The present invention provides an imaging incubator capable of recording the movement of bacterial clusters in real time, the incubator comprising a box body, a time-lapse shooting system, a temperature control system and a humidity control system; The time-lapse shooting system includes a camera, a sunshade and a flash. The camera automatically captures an image at a preset interval according to a preset exposure time. The flash lights up at a preset interval and corresponds to the time when the camera automatically captures the image.

[0006] Furthermore, the preset interval period is 10 minutes, and the preset exposure time is 100 ms.

[0007] In one embodiment, the temperature control system includes a thermal insulation tent, a temperature sensor, a heating plate and a temperature control unit. The box is arranged in the thermal insulation tent. The temperature sensor is used to obtain the temperature of the thermal insulation tent. The temperature control unit is used to obtain user setting information and oscillate the heating plate according to the user setting information and the data obtained by the temperature sensor. The heating plate is used to heat and place several groups of culture dishes.

[0008] In one embodiment, the humidity control system also includes a humidity control unit, a humidity sensor, a humidifier, a fan and a spray pipe, one end of the spray pipe extends into the interior of the box through a hole in the thermal insulation tent, and the other end of the spray pipe is connected to the humidifier; and a water tank is provided below one end of the spray pipe located inside the box.

[0009] In one embodiment, the humidity sensor is used to obtain the humidity of the thermal insulation tent, and the humidity control unit is used to obtain user setting information and dynamically oscillate the humidifier according to the data obtained by the humidity sensor.

[0010] In one embodiment, the flash lamps surround the heating plate.

[0011] The beneficial effects of the present invention are: (1) By periodically synchronizing the flash with the camera's shooting time, the control logic of the lighting system avoids maintaining high brightness for a long time to prevent the possible inhibition of microbial growth by light intensity. This design not only ensures the lighting requirements during shooting, but also minimizes the impact on the culture environment. In this way, clear images can be captured regularly without interfering with microbial growth, providing valuable data support for subsequent research; at the same time, the cost of this set of equipment is low, allowing laboratories with limited budgets to conduct high-quality bacterial cluster movement research.

[0012] (2) Side lighting by using LED light strips. Using a light shield can improve image quality, while LED lights can improve lighting efficiency by illuminating a larger area at the same time.

[0013] (3) Accurate real-time temperature monitoring and control are achieved. The heating plate is placed on the support of the microbial culture plate. By minimizing the presence of the air layer, we ensure that heat transfer is completely carried out through solid conduction, thereby improving the efficiency and uniformity of temperature control. It can ensure that when the culture dish is placed upside down, the temperature of the culture dish cover in contact with the heating platform is slightly higher than that of the agar, thereby effectively preventing the formation of condensation water.

[0014] (4) The humidifier is oscillated and adjusted with real-time feedback through the humidity control unit. At the same time, the humidity control unit can obtain user setting information, so the user can set the parameters before the experiment starts, so as to adjust the threshold according to the specific needs of microbial culture. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a front view of the overall structure of the imaging incubator.

[0016] Figure 2 Schematic diagram of part of the structure of the imaging incubator.

[0017] Figure 3 This is a real shot of the structure of the imaging incubator.

[0018] Figure 4 Cultivate and capture images of bacterial colony growth in culture dishes for imaging incubators.

[0019] Figure 5 A comparison of the morphologies of bacteria at different stages of swarming movement. DETAILED DESCRIPTION

[0020] The technical scheme of the present invention is described in detail below in conjunction with the accompanying drawings and embodiments, but the present invention is not limited to the scope of the embodiments. The process parameters not specified in the embodiments of the present application can be carried out according to conventional methods, and the raw materials used can be obtained through commercial channels.

[0021] Example 1 See also Figure 1-5 As shown, this embodiment provides an imaging incubator that can record the movement of bacterial clusters in real time. The incubator includes a box, a time-lapse shooting system, a temperature control system, and a humidity control system. The time-lapse shooting system includes a camera, a visor, and a flash. The camera uses a Hikvision MV-CS200-10UM camera and is equipped with an MVL-KF1224M-25MP lens to achieve a field of view of 350mm×230mm at a working distance of about 300mm. The image is previewed through the MVS visual platform provided by Hikvision. At the same time, the camera automatically captures an image at a preset exposure time at each preset interval. The flash lights up at each preset interval, corresponding to the time when the camera automatically captures the image. The flash lights up periodically and is precisely synchronized with the camera's shooting time. The control logic of the lighting system avoids maintaining high brightness for a long time to prevent the possible light intensity from inhibiting the growth of microorganisms. Such a design not only ensures the lighting requirements during shooting, but also minimizes the impact on the culture environment. In this way, clear images can be captured regularly without disturbing the growth of microorganisms, providing valuable data support for subsequent research. At the same time, the low cost of this equipment enables laboratories with limited budgets to conduct high-quality bacterial swarm movement research.

[0022] As an embodiment, the image processing software used may be ImageJ, which is a mature open source software with powerful image processing and analysis functions.

[0023] The entire imaging incubator is carried out in a stable indoor heat-insulating tent (hydroponic tent), which serves as the basis of the entire experimental device. The heat-insulating tent (hydroponic tent) achieves a relatively sealed environment (all holes in the tent are closed, and the tent zipper is tightened to achieve sealing), ensuring the stability of the experimental environment and preventing external air from entering and causing interference. The environment inside the heat-insulating tent (hydroponic tent) can maintain stable humidity and temperature, providing an ideal experimental environment for bacterial cultivation and monitoring, and improving the reliability and repeatability of the experiment.

[0024] In addition, during the test, an image is automatically captured every 10 minutes, and the exposure time is adjusted to 100,000 microseconds (i.e. 100 milliseconds) to obtain the best imaging effect.

[0025] Please refer to Figure 1 As shown in the figure, the incubator is made of durable 3030 aluminum profile. The aluminum profile has good structural strength and flexibility, and can be adjusted in height and angle according to the needs of the experiment. The imaging frame provides reliable support for each component, while ensuring the stability and accuracy of each system.

[0026] At the bottom of the frame, a 24 cm × 24 cm custom water tank is embedded. The main function of the water tank is to collect excess water droplets generated by the humidifier. This design not only prevents excess water from leaking out, but also keeps the experimental environment clean and safe, and prevents water droplets from affecting the operation of other electronic equipment.

[0027] A custom-made fixing plate with holes is installed above the water tank. This plate can not only firmly fix the fan, but also ensure that the water vapor generated by the humidifier is smoothly transported upward to maintain the humidity of the entire bacterial culture layer. This design not only improves air circulation, but also effectively prevents water vapor from accumulating at the bottom, ensuring uniform temperature and humidity in the culture environment.

[0028] The petri dish is placed on a black heating plate with holes on its side, which can provide stable heating for the petri dish and allow water vapor to pass through the holes on the side of the heating plate, so that humidity can be transferred smoothly. At the same time, the black background of the heating plate helps to optimize the imaging effect and make the image contrast clearer.

[0029] Flashlights are arranged around the heating plate. The flashlights use customized LED light strips. Through the external Arduino control board, the LED light strips are precisely synchronized with the camera timing shooting system to ensure that a consistent light source is provided during each shooting. In this way, the light can be stabilized during long-term time-lapse photography, and the quality and consistency of imaging can be improved. In addition, since the clarity of the photos of clustered movement of colonies on the culture dish is affected by many factors, such as the lid, agar surface and bottom of the culture dish, they will affect the optical quality of the imaging. When using a camera flash or an auxiliary front light source, part of the light will be reflected by the lid and bottom of the culture dish to form a light spot in the photo. In the present invention, for experiments involving multiple culture dish plates (up to 9), LED light strips are used for side lighting. The use of a light mask can improve image quality, and LED lights can improve lighting efficiency because they can illuminate a larger area at the same time.

[0030] Above the heating plate and light strip, there are precisely calculated visors installed. These visors are designed to ensure the best lighting conditions while avoiding interference from external light. By controlling the direction and intensity of light, the visors can effectively optimize the imaging effect, making the pictures taken clearer and more contrasty.

[0031] A high-resolution CMOS camera is fixed at the top of the entire culture rack. As the most important imaging component in the entire system, the CMOS camera is responsible for taking images of the bacterial culture process at regular intervals. It works closely with the underlying LED light strip system and Arduino control board to achieve precise time-lapse photography, which can record subtle changes in the culture process and ultimately obtain high-quality experimental data.

[0032] Example 2 The incubator includes a box, a time-lapse shooting system, a temperature control system and a humidity control system; the temperature control system includes a heat preservation tent, a temperature sensor, a heating plate and a temperature control unit, the box is arranged in the heat preservation tent, the temperature sensor is used to obtain the temperature of the heat preservation tent, the temperature control unit is used to obtain user setting information and oscillate the heating plate according to the user setting information and the data obtained by the temperature sensor, and the heating plate is used to heat and place several groups of culture dishes. Since agar gel usually contains more than 97% of water, condensation water is easily formed on the cover of the culture dish, which will hinder the shooting of the image. The incubator of the present invention controls the temperature of the heating plate by the temperature control unit, so that when the culture dish is placed upside down, the temperature of the culture dish cover in contact with the heating platform is slightly higher than that of agar, thereby effectively preventing the formation of condensation water. At the same time, the heating plate adopts a constant temperature for heating, and relies on the temperature sensor combined with the oscillation control of real-time feedback. The oscillation regulation is to control the start and power-off of the heating plate and the humidifier. If the set value is exceeded, the power is cut off, and if it is not exceeded, it works normally, thereby realizing accurate real-time temperature monitoring and control. The heating plate is placed on the support of the microbial culture plate. By minimizing the presence of air layers, we ensure that heat transfer is entirely through solid conduction, thereby improving the efficiency and uniformity of temperature control.

[0033] It is important to note that agar surface structure may vary depending on pouring method, thickness, and agar percentage, factors that need to be optimized for specific bacteria.

[0034] In addition, since the temperature control unit can obtain user setting information, the user can set parameters before the experiment begins and dynamically adjust the environmental conditions through the temperature control unit, where the temperature control unit can be implemented through a digital sensor and a feedback controller.

[0035] In addition, the heating plate in the present invention can simultaneously place multiple culture dishes with a diameter of 9 cm, which can efficiently utilize the space and energy in the incubator, and the geometric structure of the incubator and the photographic light field has been carefully calibrated to ensure that when multiple culture dishes are placed at the same time, each culture dish can obtain the best lighting effect, thereby ensuring the uniformity and quality of imaging. This design not only improves the efficiency of the experiment, but also ensures the consistency of data when comparing different culture dishes.

[0036] Example 3 The incubator includes a box, a time-lapse shooting system, a temperature control system and a humidity control system; the temperature control system also includes a humidity control unit, a humidity sensor, a humidifier, a fan and a spray pipe, one end of the spray pipe extends to the inside of the box through the hole on the insulation tent, and the other end of the spray pipe is connected to the humidifier; and a water tank is provided below one end of the spray pipe located inside the box. The humidity sensor is used to obtain the humidity of the insulation tent, and the humidity control unit is used to obtain user setting information and dynamically oscillate the humidifier according to the data obtained by the humidity sensor. The incubator of the present invention performs real-time feedback oscillation control on the humidifier through the humidity control unit, and the humidity control unit can obtain user setting information, so the user can set parameters before the experiment starts, so as to adjust the threshold according to the specific needs of microbial culture. The retractable plastic spray pipe is extended to the inside of the tent through the hole on the tent wall and is located below the sample platform. In this way, we can accurately control the humidity environment in the tent and provide ideal conditions for microbial culture.

[0037] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. An imaging incubator capable of recording bacterial cluster movement in real time, characterized in that: The incubator comprises a box body, a time-lapse shooting system, a temperature control system and a humidity control system; The time-lapse shooting system includes a camera, a sunshade and a flash, wherein the camera automatically captures an image at a preset interval according to a preset exposure time, and the flash lights up at a preset interval corresponding to the time when the camera automatically captures the image; The temperature control system includes a heat preservation tent, a temperature sensor, a heating plate and a temperature control unit. The box is arranged in the heat preservation tent. The temperature sensor is used to obtain the temperature of the heat preservation tent. The temperature control unit is used to obtain user setting information and oscillate the heating plate according to the user setting information and the data obtained by the temperature sensor. The heating plate is used to heat and place several groups of culture dishes.

2. The imaging incubator capable of recording bacterial cluster movement in real time according to claim 1, characterized in that: The preset interval period is 10 minutes, and the preset exposure time is 100 ms.

3. The imaging incubator capable of recording bacterial cluster movement in real time according to claim 1, characterized in that: The humidity control system also includes a humidity control unit, a humidity sensor, a humidifier, a fan and a spray pipe. One end of the spray pipe extends into the interior of the box through a hole in the heat-insulating tent, and the other end of the spray pipe is connected to the humidifier. A water tank is provided below one end of the spray pipe located inside the box.

4. The imaging incubator capable of recording bacterial cluster movement in real time according to claim 3, characterized in that: The humidity sensor is used to obtain the humidity of the heat preservation tent, and the humidity control unit is used to obtain user setting information and dynamically oscillate the humidifier according to the data obtained by the humidity sensor.

5. The imaging incubator capable of recording bacterial cluster movement in real time according to claim 1, characterized in that: The flash lamps surround the heating plate.

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