Spectroscopic head for identifying microoganisms in a biological sample

Abstract

The invention relates to laboratory equipment for carrying out microbiology testing. A system for identifying microorganisms contains a digital optical unit comprising: a housing with a platform for receiving a container with a biological sample; a camera disposed in the upper part of the housing above the platform for receiving a container; a spectroscopic head; and two lamps arranged to provide uniform illumination of the container with a biological sample. The spectroscopic head comprises a holder and, disposed therein, two optical systems designed to permit the incoupling and outcoupling of optical radiation in orthogonal planes, one of said systems being intended to supply excitation radiation, and the other being intended to detect a fluorescent response signal. Each of the optical systems contains an optical fibre disposed in lightguide channels. The system provides more accurate taxonomic identification of microorganisms and a more reliable and compact digital optical unit for this purpose.

Description

[0001] SPECTROSCOPIC HEAD FOR IDENTIFICATION OF MICROORGANISMS IN A BIOLOGICAL SAMPLE

[0002] Field of technology to which the invention relates

[0003] The invention relates to laboratory equipment for conducting microbiological research, namely to devices and systems for the spectroscopic study of biological samples placed in containers, for example, in Petri dishes, including recording the growth of microorganism colonies in real time, classifying microorganisms and their taxonomic identification based on optical response data, identifying the microbial agent in the sample, determining its resistance to antibacterial drugs, etc.

[0004] In laboratory equipment, a spectroscopic head is designed to deliver exciting radiation and detect the fluorescent response signal from a colony of microorganisms. A device for identifying microorganisms in a biological sample, containing a digital optical unit, can act as a stand-alone product or can be used in conjunction with a biological sample incubation chamber (see, for example, patent RU 2813445), which, in turn, can be part of an automated microbiological laboratory or any other system with a built-in biological sample photography function (see, for example, patent RU2811721). The digital optical unit in the device for identifying microorganisms can be designed in accordance with patent RU2813210, supplemented with a spectroscopic sample analysis function.To implement the function of spectroscopic examination of samples, the declared digital optical unit of the device and system for identifying microorganisms in the biological sample under study contains a movable spectroscopic head used to supply exciting radiation and detect the fluorescent response signal from a colony of microorganisms.

[0005] State of the art

[0006] The microbiology industry is currently experiencing rapid development, with one of its goals being the implementation of automated systems and equipment for incubating microorganisms and cellular structures and conducting microbiological research, including solving problems of microorganism classification and taxonomic identification. With the growing number of microbiological studies, the development of high-performance devices, increasing the speed of research, while ensuring high-quality results, including by eliminating human error, is becoming increasingly important.One of the challenges in automating microbiological research involves developing modules or units for biological sample analysis based on optical tools and methods, including digital imaging of biological samples and spectroscopy, ensuring highly accurate microorganism analysis, including their classification and taxonomic identification. These modules and units can be integrated into or integrated with a biological sample incubation chamber, enabling the incubation and analysis of biological samples within a closed volume.

[0007] A spectroscopic head for scanning a biological sample is known from the prior art (US 2018 / 0149594 A1). This optical head comprises a channel for transmitting excited optical radiation to the biological sample, which contains a lens, and a channel for transmitting reflected light from the biological sample. The output channel for reflected radiation from the biological sample, which contains a collecting lens and an optical fiber, is located outside the optical head. However, in this device, the optical components cannot be compactly housed in a single, movable housing, preventing the spectroscopic head from being moved to any point above the sample being studied. Conducting the study requires moving the sample beneath the spectroscopic head.

[0008] A coaxial optical head for scanning a biological sample is known, capable of generating optical radiation and recording reflected light from the scanned sample (WO 2006 / 009754 A1). The design of said optical head is shown in Fig. 3. Optical radiation enters through an optical fiber and is collimated by a lens system. The collimated light passes through an aperture and is directed into a beam splitter, which transmits part of this light to a reference photodetector aligned in the direction of the collimated light. The remaining light is directed at an angle of 45 degrees into the optical fiber, which transmits light to the analyzed biological sample. Diffuse and specular light returns from the analyzing section through the optical fiber to the beam splitter, which directs the returning light to a spatial frequency filter and to a lens, which focuses the diffuse light in the optical fiber leading to the photodetector.

[0009] However, the use of a beam splitter in the optical circuit leads to additional losses in the power of the emitted radiation and sets higher requirements for the power of the light source compared to the claimed invention.

[0010] A spectroscopic head, a digital optical unit and a system for identifying microorganisms are known from the prior art (US2011299071 A1), wherein the digital optical unit includes: a housing with a platform for placing a container with a biological sample; a digital camera located in the upper part of the housing; two optical systems, one of which is intended for supplying exciting radiation, the second for detecting a fluorescent response signal from the biological sample; an upper lamp located for illuminating the container with the biological sample. The device for optical identification of microorganisms contains a digital optical unit connected to a light source configured to select the required set of wavelengths, and a spectrometer.The system for optical identification of microorganisms comprises a device for optical identification of microorganisms, connected to a control module for the movement of a spectroscopic head, including control of the navigation of the optical head according to images obtained from a digital camera, with modules for moving the digital camera and container, as well as with a control module for optical measurements and a module for analyzing the obtained images.

[0011] However, in the described device, the radiation source and receiver are located separately, not integrated into a single housing, which increases the device's size. Furthermore, microorganism identification relies on Raman spectroscopy, which requires complex and expensive equipment for detecting Raman scattering optical response spectra—a spectrometer with high spectral resolution and high matrix sensitivity.

[0012] The closest spectroscopic head to the claimed one for identifying microorganisms in a biological sample is the spectroscopic head presented in US 2004 / 0160601 A1. The spectroscopic head comprises a holder (head housing) shaped like a parallelepiped and two optical systems housed within the holder. One of these systems is designed to deliver exciting radiation to the biological sample, and the other is designed to detect the response signal from the biological sample. Each optical system contains an optical fiber—an input and an output fiber, respectively—a convex lens, and a light filter. A flat beam-splitting mirror is installed between the lenses of the exciting and detecting optical systems. The optical axes of the optical system elements are positioned perpendicular to each other and at an angle of 45 degrees to the beam-splitting mirror.The spectroscopic head contains a switch, designed as a movable shutter capable of blocking optical radiation from the input optical fiber, functionally connected to a spring and a solenoid. A collimating lens is positioned between the beam-splitting mirror and the biological sample. Optical radiation from the input optical fiber passes through a convex lens, a chamber when the switch is in the "open" position, and a light filter (low-pass filter) of the first optical system. It is reflected from the beam-splitting mirror and passes through the collimating lens, which focuses the excitation radiation at a specific point on the biological sample. Radiation reflected from the sample also passes through the collimating lens, which now serves to collimate the scattered radiation and transmit it to the output optical fiber via the beam-splitting mirror, light filter, and convex lens of the second optical system.

[0013] However, the spectroscopic head contains moving parts, which complicate the device's design, reduce its reliability, and generate mechanical vibrations during operation, which introduce errors in the formation of the focal spot on the biological sample surface, reducing the accuracy of the detected optical signal. Furthermore, the placement of the optical systems in the holder, with their optical axes at right angles, increases the dimensions of the optical head compared to a parallel arrangement. Furthermore, detecting Raman scattering optical response spectra requires the use of a spectrometer with high spectral resolution and high matrix sensitivity. The holder's design as a single, one-piece housing does not ensure easy assembly and maintenance during operation.

[0014] The closest analogs to the claimed digital optical unit, device, and system for identifying microorganisms in a biological sample are the photography module unit and device provided as part of the biological culture incubation system (US 11041871 B2). The digital optical unit of the photography module is housed in a separate housing, which communicates with the working volume of the incubation chamber. The digital optical unit contains a photography unit and a container transport unit. The container transport unit is a rotating disk capable of horizontally moving the container from the working area of ​​the incubation chamber to the digital imaging zone and back. The rotating disk, by 90 degrees, places the sample in the photographing zone under the camera lens, after which, by another 90-degree turn, the container is transferred to a conveyor for return to the incubation chamber.

[0015] However, this system only produces optical images of cell cultures in the visible spectrum and does not measure the cell culture's fluorescence spectra under ultraviolet radiation, limiting the system's analytical capabilities. Furthermore, the design of its analogs and prototype lacks the ability to integrate fluorescence and photographic data for the taxonomic identification of microorganisms during biological sample analysis, reducing the accuracy of the results.

[0016] The technical problem, the solution of which is provided by the present invention, is the development of a device for identifying microorganisms in a biological sample being studied, eliminating the shortcomings of analogues and a prototype, capable of ensuring high accuracy in solving problems of classification and taxonomic identification of microorganisms in a biological sample being studied.

[0017] Disclosure of the essence of the invention

[0018] The technical result of the claimed group of inventions is to increase the accuracy of microorganism testing in biological samples placed in containers, such as Petri dishes, including their classification and taxonomic identification. This enhances the reliability and compactness of the digital optical unit used for these purposes, which includes a digital camera and a spectroscopic head. The spectroscopic head also facilitates easy assembly, generating high-quality input and output optical radiation, ensuring the coincidence of focal spots generated by the optical systems on the surface of the biological sample being tested. The spectroscopic head is reliable and compact, allowing it to be used in a single optical unit alongside other devices, such as a digital camera.

[0019] The technical result is achieved by implementing a spectroscopic head of a digital optical unit of a device for identifying microorganisms in a biological sample being studied, including a holder and two optical systems placed in the holder, configured with the possibility of inputting and outputting optical radiation in orthogonal planes, one of which is intended for supplying exciting radiation, the second - for detecting a fluorescent response signal from a biological sample, wherein the holder is made as a prefabricated assembly of at least three parts having adjacent profiled surfaces configured with the possibility of forming, when the parts are combined, two light-guide channels with the placement of the above-mentioned two optical systems in them.

[0020] Each of the optical systems of the spectroscopic head, in one embodiment, comprises an optical fiber, two convex (collimating) lenses—an input and an output lens—and a flat mirror located in the optical path between the lenses. These optical system elements are housed in the holder's lightguide channels, providing input and output of optical radiation in orthogonal planes. The optical fibers are preferably positioned in a plane parallel to the surface of the container support area. The optical systems are arranged so as to form an angle between their optical axes in the vertical plane, ensuring the coincidence of focal spots on the surface of the biological sample being examined. This angle between the optical axes of the optical systems can range from 30 to 42 degrees, for example, 36 degrees.

[0021] The spectroscopic head is capable of detecting the fluorescence response signal at a distance of up to 2 mm from the biological sample being studied. The spectroscopic head can employ: lenses with a focal length of 15 to 20 mm and an aperture (diameter) of 10 mm, wherein the lenses are made of KU-1 fused quartz; mirrors with a coating characterized by maximum reflectivity in the wavelength range from 280 to 800 nm; multimode optical fiber with a core diameter of d = 200 - 600 μm, and a numerical aperture of NA = 0.22; wherein the ends of the optical fibers are located at a distance Ax from 0 to 5 mm from the lens focus.

[0022] In one embodiment of the invention, the holder in the spectroscopic head is made in the form of a parallelepiped of three parts - a central element and two side elements located symmetrically relative to the central element; the elements are formed by dissecting the parallelepiped with two sectional planes located at an angle (up to 21 degrees) to the vertical longitudinal plane of the parallelepiped, wherein the holder elements preferably contain 1 / 2 parts of light guide channels in the cross section, configured with the possibility of forming light guide channels when combining the corresponding side elements with the central element, where one of the light guide channels is configured with the possibility of accommodating an optical system intended for supplying exciting radiation, the second light guide channel - with the possibility of accommodating a second optical system intended for detecting a fluorescent response signal from a biological sample.The light guide channels are provided with slots for accommodating lenses and mirrors of the optical systems. The light guide channel may comprise a section with a diameter dl corresponding to the diameter of the optical fiber for its fixed placement within the channel. This section of the channel is connected to a section of larger diameter d2, designed to propagate radiation from the optical fiber for the first optical system, or to the optical fiber for the second optical system, with the direction of radiation propagation being changed by 90 degrees using a mirror installed in the channel. The holder in the spectroscopic head, in the preferred embodiment, has a height of 25 mm or less, and a width and length of 75 mm or less.

[0023] The technical result is achieved by implementing a digital optical unit of a device for identifying microorganisms in a biological sample being tested, comprising a housing containing a platform for placing a container with a biological sample; a digital camera located in the upper part of the housing above the platform for placing the container; a spectroscopic head, the description of which is presented above, configured to move in at least a horizontal plane above the platform with the container; at least two lamps - an upper and a lower one, located to ensure uniform illumination of the container with the biological sample.

[0024] The movement of the spectroscopic head can be realized using an electromechanical movement drive equipped with a belt transmission, while the electromechanical drive is located on the bottom side of the digital optical unit housing.

[0025] The optical system elements—optical fiber, input and output convex lenses, and a flat mirror—are housed in the holder's light-guide channels and configured to input and output optical radiation in orthogonal planes. The digital camera can be equipped with a vertical focus drive.

[0026] In one embodiment, the digital optical unit is equipped with a container transport device, which includes a transport basket for the container. The container transport device may be equipped with a shutter capable of horizontal reciprocating movement and positioned in the photographic area beneath the transport basket, blocking the light from the lower luminaires. The transport device may also be equipped with separate horizontally oriented guide rails for the shutter to move along.

[0027] Additionally, the spectroscopic head can be mounted on a movable platform, which can be moved by an electromechanical drive comprising two stepper motors with pulleys and a cross-belt drive. The cross-belt drive is achieved using four rollers mounted on the movable platform, with the belt travel along each of the X and Y axes limited by mechanical stops and limit switches based on optical sensors.

[0028] The technical result is achieved by implementing a device for identifying microorganisms, containing a spectroscopic head located in a digital optical unit, the description of which is presented above, connected to a light source, configured to select the required set of wavelengths, and a spectrometer.

[0029] In a specific embodiment of the invention, the following may be used: a plasma light source capable of emitting at wavelengths in the range of 280-500 nm and a spectral brightness of 1 mW / (mm*sr*nm) to 35 mW / (mm*sr*nm), wherein the plasma light source is equipped with a monochromator; a spectrometer (detector) with a radiation detection range of 300-800 nm with a spectral resolution of no worse than 5 nm and having a scattered light detection level of no more than 0.5%; a monochromator having a spectral radiation tuning range of 280-500 nm, with an allocated spectral bandwidth of no more than 10 nm in the said range.

[0030] The technical result is achieved by implementing a system for identifying microorganisms, comprising a device for optical identification of microorganisms, the description of which is presented above, connected to a module for controlling the movement of a spectroscopic head, including control of the positioning of the optical head according to images obtained from a digital camera, modules for controlling the movement of a digital camera and a container, as well as a module for controlling optical measurements and a module for analyzing the obtained images and fluorescence spectroscopy data.

[0031] In various embodiments of the invention, the image analysis module may be configured to preprocess images to visualize the region associated with the nutrient medium and the microorganisms cultured therein. It may also include computer vision tools and a convolutional neural network used for microorganism classification. In a specific embodiment, the image analysis module may utilize a model with a ConvNeXt-type architecture, pretrained on the ImageNet database.

[0032] Furthermore, the image analysis module may include microorganism classification capabilities based on fluorescence spectroscopy results using machine learning tools, including fully connected neural networks trained using the amplitudes of the principal components of fluorescence spectra during the preprocessing of optical spectroscopy data and optical imaging data. The optical measurement control module is preferably configured to excite emission at multiple excitation wavelengths from 280 to 500 nm and detect at wavelengths from 300 to 800 nm.

[0033] The design of the claimed device utilizes an improved optical system holder. This compact, collapsible design allows for easy assembly and maintenance of the spectral head. The holder combines the receiving and transmitting channels, housing optical system elements that generate high-quality input and output optical radiation, ensuring the focal spots generated by the optical systems coincide with the surface of the biological sample being examined. The spectroscopic head's configuration and compact size allow it to be used in the optical unit implemented under patent RU2813210 without increasing the unit's dimensions.

[0034] The design of the claimed device also uses an improved digital optical unit, including a housing that has a modular design and is compatible with the automatic bacterial incubation system, while in one unit two systems of the optical and spectral research method are combined, allowing the formation of both a visible photographic image of a microbiological sample and fluorescence spectra in the ultraviolet spectrum, expanding the analytical capabilities of the device; a spectroscopic head, combining the receiving and emitting channels, which has a compact design with reliable placement of the elements of the optical system in it; a mechanism for rapid and precise movement and positioning of the spectroscopic head during optical measurements, and, as a result, high-precision guidance of the emitted light on the object, incl.by using an XY drive with a cross-shaped belt transmission and support rollers, which allows the head to move along an arbitrary trajectory with an accuracy of up to 50 µm.

[0035] In the claimed invention, the obtained accuracy, combining the results of optical studies of fluorescence data and digital photographic data on stratified samples, taking into account the distribution of the balance of microorganism species in a microbiological laboratory, is F = 0.828 ± 0.013, which exceeds the accuracy values ​​obtained separately on the basis of fluorescence data or digital photographic data.

[0036] Thus, the proposed spectroscopic head and optical unit can be integrated into a high-performance automated system for incubating and analyzing biological samples in a flow of incubated samples (at least 100 measurements per hour) with the ability to monitor incubated cultures during cultivation at a specified frequency. The spectroscopic head's dimensions allow it to be used in conjunction with a digital camera without increasing the module's size, while ensuring high-precision positioning of the applied excitation radiation relative to the sample under study and detection of the fluorescence response from the biological sample.Combining two systems—optical and spectral research methods—in a single unit allows for the formation of both a visible photographic image of a microbiological sample and fluorescence spectra in the ultraviolet spectrum, expanding the analytical capabilities of the device, which is extremely important for improving the quality of research conducted on biological samples, including as part of a "smart incubator."

[0037] The device is reliable, ergonomic, easy to manufacture, use, and maintain, and reduces the likelihood of operator errors that can lead to misdiagnosis, delayed diagnosis, and loss or damage of samples. Such errors can potentially negatively impact the quality of diagnostics and the effectiveness of patient treatment.

[0038] Brief description of the drawings

[0039] The group of inventions is explained by drawings, where Figure 1 shows an image of the claimed spectroscopic head built into the digital optical unit of the device for identifying microorganisms in the biological sample being studied, a general view, Figures 2 - 6 show the digital optical unit, a back view, a top view, a sectional view, a side view, a bottom view, respectively; Figure 7 shows a general view of the digital optical unit combined with a container movement unit; Figure 8 shows an incubation chamber with two 3-coordinate manipulators - upper and lower, in which two digital optical units are installed, a general view from the side of the container loading / unloading window;Figure 9 schematically shows an example of the use of a digital optical unit of a device for identifying microorganisms in a biological sample being studied in an automated microbiological laboratory, where, in addition to the aforementioned unit built into the incubation chamber, the laboratory also contains a module for seeding biomaterial into containers, a conveyor system, a module for loading and unloading containers from the incubation chamber, a laboratory assistant's workstation with a PC for conducting studies of biological samples; Figure 10 shows a basic block diagram of performing optical measurements and identifying microorganisms based on optical response data, implemented in a system containing a device with a digital optical unit, modules for controlling the movement of the spectroscopic head, the movement of the container and camera, optical measurements and image analysis;Figure 11 shows a diagram of the assembly of the spectroscopic head, the holder of which consists of three parts (components) - a central one and two lateral ones; Figure 12 shows a photograph of the parts of the holder of two optical systems of the spectroscopic head, manufactured using the 3D printing method; Figures 13 and 14 show a diagram of the placement of the spectroscopic head and a diagram of the location of the detecting and exciting parts of the spectroscopic head relative to each other and the Petri dish during measurements; Figure 15 shows the optical diagram of the excitation / detection radiation of the optical system, where AX is the distance of the end of the optical fiber from the focal plane of the lens, AZ is the height of the location of the object under study above the plane characterized by the maximum radiation intensity;Figure 16 shows graphs comparing A) the intensities of the fluorescence response depending on the location of the emitting end of the transmitting-receiving fiber AX, and B) the spectral band of the fluorescence emission detected from a model object using a two-fiber transmitting-receiving circuit and the spectroscopic head of the device; Figure 17 shows graphs of A) the fluorescence spectra of the Escherichia coli bacterium upon excitation at 380 nm with different configurations of the optical head; B) the fluorescence spectra of the Escherichia coli bacterium normalized to the maxima upon excitation at 380 nm with different configurations of the spectroscopic head; Figure 18 shows the distribution of the number of objects of different taxonomic categories used for testing and assessing the accuracy of the device; Figure 19 shows an example of an optical image and fluorescence spectra obtained using the claimed device;Figure 20 shows the values ​​of the quality metrics for the taxonomic identification of microorganism classes based on optical imaging data, fluorescence spectroscopy, and their combination, obtained during testing of the device; Figure 21 shows a basic block diagram for combining classification models based on fluorescence response data and images;

[0040] The following items are designated by positions in the drawings: 1 - digital optical unit, 2 - spectroscopic head, 3 - digital camera, 4 - upper lamp, 5 - lower lamp, 6 - side lamp, 7 - digital optical unit housing, 8 - housing platform for placing the container, 9 - first optical system - for supplying exciting radiation, 10 - second optical system - for detecting the fluorescent response signal from the biological sample, 11 - optical system holder (or spectroscopic head housing), 12 - spectroscopic head console, 13 - optical fiber (optic fibers), 14 - convex lens, 15 - flat mirror, 16 - light guide channel of the holder, 17 - electromechanical drive of the spectroscopic head, 18 - movable platform on which the spectroscopic head is fixed, 19 - motor with a drive pulley of the spectroscopic head, 20 - belt drive spectroscopic head drive,21 - guides for moving the spectroscopic head, 22 - limit switch of the spectroscopic head drive, 23 - central element of the holder, 24 - side elements of the holder, 25 - drive for moving the digital camera, 26 - screw gear, 27 - limit switch of the digital camera drive, 28 - means for moving the container, 29 - transport basket for the container, 30 - shutter, 31 - guide rails for moving the basket, 32 - guide rails for moving the shutter, 33 - light source, 34 - spectrometer, 35 - monochromator, 36 - incubation chamber, 37 - three-coordinate manipulator, 38 - control unit, 39 - container movement unit.,

[0041] Implementation of the invention

[0042] A more detailed description of the claimed invention is provided below.

[0043] The following terms, definitions and abbreviations are used in this description.

[0044] When describing the claimed invention, the term “system for incubation and digital imaging of biological samples” may be abbreviated as “system,” “incubation system,” “incubation system,” or “automated system.”

[0045] "Container" means a container that can accommodate a test sample, including by manual and / or automated inoculation. The container typically contains a substrate or medium with nutrients for the growth of target microorganisms. According to the present invention, containers such as Petri dishes (hereinafter also referred to as dishes) containing cultured medium, test tubes containing broth, and slides containing biological samples, etc., can be used in an automated testing system.

[0046] “Biological sample” (hereinafter also referred to as sample) is a sample of fluid and / or tissue of the human body, as well as any other material sample potentially containing microbiological objects, taken for laboratory testing.

[0047] "Taxonomic identification of microorganisms" is the determination of the microorganism's belonging to a specific genus, species, and other taxonomic categories using the optical response of the microorganism's colony.

[0048] The claimed spectroscopic head may be part of a digital optical unit of a device included in a photography module, the design solution of which is presented in the description of the invention according to patent RU 2813210; the claimed digital optical unit of the device and system for identifying microorganisms in a biological sample under study may be implemented on the basis of a digital optical unit included in a photography module according to patent RU 2813210. In this case, the optical unit 1 is additionally equipped with a movable spectroscopic head 2, used for supplying exciting radiation and detecting a fluorescent response signal from a biological sample (for example, a colony of microorganisms).

[0049] The digital optical unit 1 (Figs. 1 - 6) of the device for identifying microorganisms in a biological sample being studied has a modular design and includes a housing 7 with a platform 8 for placing a container with a biological sample; located in the housing 7: a spectroscopic head 2, configured to move at least in a horizontal plane above the platform 8 with the container; a digital camera 3, fixed in the upper part of the housing 7 above the platform 8 for placing the container, and configured to move at least in the vertical direction; at least two lamps - an upper 4 and a lower 5, fixed in the housing to ensure uniform illumination of the container with the biological sample.

[0050] The housing 7 of the digital optical unit can have various configurations, preferably in the form of a rectangular box, facilitating assembly and maintenance. The housing comprises a top, bottom, front, side, and rear wall, and is equipped with electrical connectors, brackets, access holes and recesses, mounting locations, and adapter pads with quick-release eccentric fasteners for installation and connection to the automated bacterial incubation system, as well as for ease of disassembly and maintenance. The housing can be made of sheet metal with an anti-reflective (absorbing) matte coating on the interior.

[0051] The spectroscopic head 2 (Figs. 11 - 15) contains two optical systems 9 and 10, located in the holder 11. One of the optical systems, 9, is intended for supplying exciting radiation, while the second 10 is intended for detecting the fluorescent response signal from the biological sample. Each of the optical systems contains an optical fiber 13, two convex lenses 14 - an input and an output one, and a flat mirror 15, located on the optical path between the lenses 14, wherein the said elements of the optical systems are located in the light-guide channels 16 of the holder 11, configured with the possibility of inputting and outputting optical radiation in orthogonal planes, wherein the optical fibers 13 are located parallel to the surface of the platform 8 for placing the container. This arrangement of the optical fibers ensures unimpeded movement of the optical head relative to other electromechanical parts of the device.In certain embodiments of the invention, the spectroscopic head 2 comprises a multimode optical fiber 13 with a core diameter of d = 200 - 600 μm and a numerical aperture NA = 0.22, wherein the ends of the optical fibers 13 are located at a distance Ax from 0 to 5 mm from the focus of the collimating lens 14; the lenses 14 are used with a focal length of 15 to 20 mm and an aperture (diameter) of 10 mm, wherein the lenses are preferably made of fused quartz of the KU-1 brand; the mirrors 15 have a coating characterized by maximum reflectivity in the wavelength range from 280 to 800 nm.

[0052] Optical systems 9 and 10 are positioned in holder 11 so that the focal spots they form can align on the surface of the biological sample being examined. This is achieved by creating an angle between the optical axes of these optical systems in the vertical plane, which in a specific embodiment can be 36 degrees.

[0053] Spectroscopic head 2 is mounted on console 12 at a predetermined distance from the container platform, capable of detecting the fluorescent response signal from the biological sample. In a specific embodiment of the invention, the distance (height H) from the head to the biological sample being tested is preferably between 2 and 5 mm.

[0054] The holder 11 of the optical systems in one of the embodiments of the invention is made in the form of a parallelepiped containing three elements - a central 23 and two lateral 24 elements located symmetrically relative to the central element; the elements are formed by dissecting the parallelepiped with two section planes located at an angle to the vertical longitudinal plane of the parallelepiped, wherein the elements of the holder contain parts of light guide channels (preferably 1 / 2 part in cross section), configured with the possibility of forming light channels 16 when combining the corresponding lateral elements 24 with the central element 23, where one of the light guide channels is configured with the possibility of placing an optical system intended for supplying exciting radiation, the second light guide channel - with the possibility of placing a second optical system intended for detecting a fluorescent response signal from a biological sample.The holder's channels are grooved to accommodate lenses and mirrors of the optical systems. This design of the spectroscopic head holder ensures ease of manufacture, ease of maintenance, and reliability of the optical system components housed within.

[0055] Lightguide channel 16 comprises a section with a diameter dl corresponding to the diameter of optical fiber 13 for its fixed placement within the channel. This section of the channel is connected to a section of larger diameter d2, designed to propagate radiation from the optical fiber (for the first optical system) or to the optical fiber (for the second optical system), with a 90-degree change in the direction of radiation propagation achieved by mirror 15 installed within the channel.

[0056] Holder 11 in spectroscopic head 2 preferably has a height (A) no greater than 25 mm, width (B) and length (C) no greater than 75 mm (Figs. 13, 14). These dimensions of the holder allow the spectroscopic head to be integrated into the design of the optical unit, implemented according to patent RU 2813210, without increasing the dimensions of the optical unit.

[0057] The movement of the spectroscopic head 2 is realized with the help of an electromechanical movement drive 17, equipped with a belt transmission 20 with a cross-shaped arrangement of belts, while the spectroscopic head 2 is fixed on a movable platform 18, the drive is located on the lower side of the housing of the digital optical block.

[0058] The electromechanical drive for the spectroscopic head movement comprises two stepper motors 19 with pulleys mounted on fixed supports. Cylindrical elements with plain bearings serve as guides 21 for the spectroscopic head movement. Four rollers, providing a cross-shaped belt configuration, are also located on the moving platform 18. A fastening for securing the loose ends of the belt is located at the rear wall of the digital optical unit (the position corresponding to the retracted head). The travel of the spectroscopic head along each of the X and Y axes is limited by mechanical stops and limit switches 22 based on optical sensors.The specified configuration allows for an arbitrary trajectory of the head movement, makes the spectroscopic head movement unit more compact (compared to drives with screw gears) and allows both motors 19 to be fixed in place, which eliminates the need for movable wiring and increases the reliability of the movement unit.

[0059] The lighting system in the digital optical unit comprises at least two luminaires—an upper luminaire (4) and a lower luminaire (5)—mounted within the housing (7) to provide uniform illumination of the container holding the biological sample. A side luminaire (6) can be added for more uniform illumination of the sample being examined. The luminaires can be equipped with cylindrical, conical, parabolic, or spherical diffusers, allowing light to be directed toward the sample container and the digital camera mounted in the upper portion of the digital unit housing. In one embodiment, the luminaires are ring-shaped or circular flat printed circuit boards with an array of LEDs mounted thereon. The boards are secured to metal plates located within the housing (7) of the unit.

[0060] Digital camera 3 is equipped with a vertical focusing drive 25, comprising a screw transmission 26 and a limit switch 27. The camera lens is positioned in an opening in the housing, above the platform 8 for holding the sample container, coaxial with the upper 4 and lower 5 lamps. Camera focusing can be performed either manually or using a separate electric drive, including a stepper motor, a screw transmission, guides, and optical or inductive limit switches.

[0061] The digital optical unit 1 is provided with a means for moving the container 28 in the horizontal direction (or in the horizontal plane), containing a transport basket 29 for the container (Fig. 7). The transport basket 29 can be cantilevered onto a movable carriage, which, in turn, is secured to guide rails 31. The transport basket 29 is provided with its own electric motor with a belt transmission (drive), providing for the reciprocating movement of the basket in the horizontal direction along the guide rails 31 from the container receiving area to the photography area - under the lens of the camera 3. The transport basket 29, in one embodiment of the invention, can be made in the form of a support ring having a configuration that ensures the placement of a container in it.

[0062] The means for moving the container 28 in one embodiment may comprise a shutter 30, designed with the possibility of reciprocating movement in the horizontal direction and placement in the photographing zone under the transport basket 29 with the possibility of blocking the light flux from the lower lamps 5. In this case, the means for moving the container 28 is preferably provided with separate horizontally oriented guide rails 32 for moving the shutter along them and its own drive (an electric motor with a belt transmission), providing reciprocating movement of the shutter 30 along the guide rails 32 under the lens of the camera 3.Shutter 30 is positioned below the level of the transport basket so that when the basket is positioned in the digital optical unit beneath the camera lens, the shutter is positioned beneath the transport basket (under the container), blocking the light from the lower luminaires when used in this unit's design. Container transport means 28 are also equipped with shutter position sensors—"closed" or "open"—for when it is positioned beneath the camera and outside this area, respectively. The shutter is designed for capturing digital images against a dark background; if it is absent, photographs are captured against a light background.

[0063] The device for identifying microorganisms (Fig. 10) contains a digital optical unit 1 connected to a light source capable of selecting the required set of wavelengths and a spectrometer.

[0064] A specific embodiment of the invention may utilize a plasma light source with an emission range of 280-500 nm and a spectral radiance ranging from 1 mW / (mm*sr*nm) to 35 mW / (mm*sr*nm). Monochromators constructed using any known design, such as Czerny-Turner, Ebert-Fastie, and others, may be used for spectral excitation selection. The primary characteristics of a monochromator are its spectral range and the bandwidth it extracts from a broad spectrum of radiation. The device's spectral range should lie between 300 and 800 nm, covering the entire visible region of the optical spectrum. Specifically, the XWS-65 lamp from Troitsky Engineering Center, combined with an OceanOptics Monoscan 2000 monochromator, meets these requirements. This monochromator is optimal, as it isolates bands of approximately 5-7 nm width from the plasma source spectrum entering it.Alternatively, near-ultraviolet and visible LEDs can be used as excitation sources. Dielectric filters, which isolate the desired spectral range, can be used for additional spectral selection. The device preferably uses a spectrometer (detector) with a detection range of 300-800 nm, a spectral resolution of at least 5 nm, and a scattered light detection level of no more than 0.5%. The OceanOptics USB2000 device possesses these characteristics, combining high performance, good resolution, and optimal signal-to-noise ratio. The device also preferably uses a monochromator with a spectral tuning range of at least 300-500 nm, with a spectral bandwidth of no more than 10 nm in this range.

[0065] The use of quartz optical fibers 13 to detect the fluorescent response from bacterial colonies increases the intensity of the detected emission signal by about 30-40% in the region from 280 to 380 nm.

[0066] During studies using the device, it was found that a set of 5-6 excitation wavelengths concentrated in the 300-420 nm range is sufficient to achieve classification accuracies comparable to those obtained using the fluorescence response at 10 excitation wavelengths in the 280-460 nm range. It was established that solving the classification problem requires detection of the fluorescence response with a resolution of no worse than ~5 nm. The influence of spectral resolution significantly affects the accuracy of low-level classification and is less pronounced when solving the mid-level classification problem. The influence of various components of the experimental device on the ability to detect the fluorescence response upon excitation in the near ultraviolet and visible ranges was experimentally assessed.It was demonstrated that the intensity of the fluorescence response can be increased by using fused silica fibers with increased near-UV transmittance.

[0067] The microorganism identification system (Fig. 10) comprises a device for optical identification of microorganisms, connected to a control unit comprising at least two modules: a module for controlling the movement of the spectroscopic head (a control module for the electromechanical part of the device) and the navigation of the spectroscopic head according to images obtained from a digital camera (a control module for the approach of the spectroscopic head), and a control module for optical measurements. The control unit may be additionally equipped with modules for moving the digital camera and container, as well as a module for analyzing the obtained images and fluorescence spectroscopy data.

[0068] The image and fluorescence spectroscopy data analysis module contains computer vision tools for analyzing optical images and can be configured to preprocess images to visualize the region associated with the nutrient medium and the microorganisms cultured therein. The image analysis module contains a convolutional neural network used for microorganism classification, which can be a model with a ConvNeXt-type architecture (Woo, Sanghyun, et al. "Convnext v2: Co-designing and scaling convnets with masked autoencoders." Proceedings of the IEEE / CVF Conference on Computer Vision and Pattern Recognition. 2023). Other predictive model architectures with a similar or more complex architecture can be used if this improves the accuracy of microorganism taxonomic identification.

[0069] To analyze the obtained fluorescence response data, the image and fluorescence spectroscopy data analysis module contains tools for classifying microorganisms based on the results of fluorescence spectroscopy using machine learning tools, including fully connected neural networks trained using the amplitudes of the principal components of the fluorescence spectra at the stage of preprocessing the optical spectroscopy data and optical image data, as well as based on the original fluorescence response spectra obtained at various fluorescence excitation wavelengths.

[0070] The optical measurement control module is designed with the ability to excite emission at several excitation wavelengths from 280 to 500 nm, and detection from 300 to 800 nm.

[0071] The listed modules allow for the semi-automatic movement of the optical head to a specified location on the image of a dish with a nutrient medium, control of the optical signal measurement, and the transfer of data to algorithms for the taxonomic identification of microorganisms based on optical response data.

[0072] These modules can be implemented as computer programs, using a graphical interface displaying an image of a Petri dish containing the biological specimen being studied, as well as spectroscopic head controls allowing the user to select the geometric region of the image to which the spectroscopic head should be positioned. The software can be configured to select coordinates for positioning the spectroscopic head by clicking on the corresponding image region, or by manually controlling the vertical and horizontal movement of the spectroscopic head within the image. The resulting images can be saved for further analysis and use in classification algorithms. After positioning the spectroscopic head at the specified location within the dish, the module controlling the optical measurements can be launched.The specified module, among other things, controls the change in the excitation wavelength by controlling the monochromator, which provides spectral selection of the radiation of the broadband plasma source, as well as collecting data from the spectrometer, which detects the optical response.

[0073] When performing a measurement, the user sees the current fluorescence response spectrum at a given excitation wavelength, as well as all fluorescence spectra at multiple excitation wavelengths obtained during the current measurement. The user can configure measurement settings, including selecting excitation wavelengths, signal accumulation times, emission, and graphical display parameters within the module window. After clicking the "Start Measurement" button, a set of fluorescence spectra is measured at various excitation wavelengths, the data is automatically recorded, and fed to the taxonomic identification algorithm.

[0074] Thus, with the help of these modules, it is possible to carry out high-level control of image acquisition from the digital camera of the unit used for targeting a colony of microorganisms, control of the approach of the spectroscopic head to the required measurement location, control of the measurement of the optical signal from colonies of microorganisms, and, as a result, control of algorithms for the taxonomic identification of microorganisms based on optical response data.

[0075] The digital optical unit with the claimed spectroscopic head and the device for identifying microorganisms using this unit can be integrated into an incubation chamber (Fig. 8) for incubating containers with samples under optimal conditions and conducting real-time sample testing at a specified (or required) frequency (e.g., every 60 minutes). Sample testing can be implemented using software containing algorithms based on artificial intelligence. The incubation chamber, in turn, can be integrated into the structure of a microbiological laboratory (Fig. 9), which can include modules and / or units for automatic seeding of biomaterial into containers, such as Petri dishes, with the function of loading containers into cassettes, labeling containers; preparing samples for microfluidic tests; selecting colonies; determining antibiotic resistance, etc.In particular, a device for identifying microorganisms in a biological sample under study, containing a digital optical unit 1, can be incorporated into an automated microbiological laboratory or any other system with a built-in biological sample photography function, as implemented, for example, in accordance with invention patent RU2811721. Furthermore, a device for optically identifying microorganisms can be used as an additional module within a smart incubator—a "spectroscopy module," for example, within a module and system for generating digital images of biological samples, as presented in invention patent RU2813210.

[0076] Incubation chamber 36 (Fig. 8) may be constructed in accordance with invention patent RU 2813445. The working volume of the chamber contains shelves with cells (nests) for accommodating Petri dishes with biological samples. The chamber houses engineering systems for creating optimal conditions for microorganism growth within its working volume, including means for establishing the required incubation regime (heating, humidification, CO2 concentration maintenance, and air circulation) within the working volume. The incubation chamber may be equipped with sensors for monitoring temperature, humidity, and medium composition, as well as means for automatically maintaining the required parameters. The incubation chamber is equipped with a window for loading and unloading containers, as well as one or two windows for communication with the corresponding digital optical units of the device for identifying microorganisms in the biological sample being tested.It is preferable to implement the incubation chamber with two digital optical units 1 - upper and lower, functioning independently of each other, one of which is located above the window for loading and unloading containers, the second unit - below the said window. Such an arrangement of units in the incubation chamber assumes the use of two three-coordinate manipulators 37, which are located in the chamber one above the other with the ability to service with each manipulator its own zone containing a digital unit - upper and lower, respectively, and a common loading and unloading zone. In this case, the upper manipulator 37 ensures the movement of containers between the window for loading and unloading containers, the upper optical unit and the corresponding (upper) cells of the incubation chamber; the lower manipulator 37 ensures the movement of containers between the window for loading and unloading containers, the lower optical unit and the corresponding (lower) cells of the incubation chamber.The manipulators in the incubation chamber can be moved independently according to control programs. The incubation chamber housing can have separate compartments for the upper and lower optical units. The shelves with cells can be arranged in various ways. Each cell is adapted to receive and hold a sample container during incubation. The incubation chamber can contain a variable number of cells, depending on the needs of the microbiology laboratory. Each cell is characterized by its spatial coordinates. The digital optical unit 1 is housed in its housing 7 and can be integrated into the working volume of the incubation chamber or external to it.

[0077] When using a microorganism identification system in conjunction with an incubation chamber, the control unit is a hardware and software system. It includes local storage for the software required to control system elements and independent local storage for optical measurement results and biological sample analysis. The control unit may contain a microprocessor and a set of control boards or microcontrollers that manage system functions and ensure operation according to a specified algorithm for the incubation chamber, manipulators within the chamber, and the digital optical unit. The control unit consists of power supplies, voltage converters, circuit breakers, stepper motor control boards, DC motor control boards, control boards for lamps, heaters, and other incubation chamber equipment.

[0078] Example of implementation of the invention.

[0079] The experimental prototype of the device for identifying microorganisms in the biological sample under study contains a digital optical unit 1 with a spectroscopic head 2, a plasma light source XWS-65, manufactured by Troitsky Engineering Center LLC, an OceanOptics Monoscan2000 monochromator, an OceanOptics USB2000 spectrometer (detector), optical fibers d=600 μm, material - glass, numerical aperture - 0.22.

[0080] Several variants of the spectroscopic head 2 were developed and manufactured, each with dimensions that satisfied the specified constraints. A three-dimensional model of the spectroscopic head is shown in Fig. 11. The positions of the mirrors and lenses relative to the optical fibers were fixed and selected so that, at a given distance from the object, the radiation spot of the excitation portion of the optical system overlapped the recording region of the detection portion of the system. Fig. 12 shows the component parts of the optical head, manufactured using 3D printing.

[0081] Three variants of the spectroscopic head were manufactured, differing in the distance from the fiber end to the focal plane of the lens (dimension "X") (Fig. 15), while the angle between the detecting and excitation parts (Fig. 14) was selected so as to maximize the intersection of the excitation spot and the recording area in a given configuration of the optical scheme (the angle between the optical axes of the optical systems was 30, 36, and 42 degrees). Each of the optical head variants was tested on a model object for the possibility of detecting a fluorescent signal in comparison with a two-fiber transmit-receive scheme, in which the radiation was directly supplied to the studied object, located at a distance of 1 mm from the transmit-receive fiber.The fluorescence of the model object was excited in a wide spectral range (from 280 to 500 nm), the signal accumulation time on the spectrometer varied depending on the type of detection system (two-fiber scheme, scheme with a spectroscopic head).

[0082] The tests performed showed good results. With comparable accumulation times (0.1 s for the dual-fiber setup and 1 s for the spectroscopic / optical head), the differences in the intensity of the detected signal were less than 2-fold for one of the optical head options (Fig. 16A), while the signal-to-noise ratio in both configurations was the same and amounted to approximately 50. The shape of the fluorescence spectra of the model object matched for both detection system configurations (Fig. 16B).

[0083] Next, a comparison of the considered spectroscopic head configurations was conducted on a model microorganism—the bacterium Escherichia coli (see Fig. 17A). For the manufactured variants of the spectroscopic head on bacteria, it was possible to obtain a detectable signal comparable to that of the fiber-optic circuit (the "signal-to-noise" for all configurations was approximately 25) within a reasonable signal accumulation time (1 second for the spectroscopic heads versus 0.1 s for the fiber-optic circuit). Moreover, no significant difference was observed between the shape of the emission band detected using the transmitting-receiving fiber and the optical heads (see Fig. 17B).

[0084] The optical systems of the spectroscopic head, the first and second, were completely placed in a parallelepiped with a height (A) of no more than < 25 mm, and a width and depth (B and C) of no more than < 74.5 mm. The resulting spectroscopic head was located in the digital optical block at a height H of 2 to 5 mm above the surface of the Petri dish with the nutrient medium. The introduction of excitation radiation and the recording of the optical response were implemented using transceiver optical fibers. The optical scheme allowed the detection of the fluorescence response in the spectral ranges of excitation (280-500 nm) and detection (300-800 nm). The introduction of excitation radiation in this scheme was carried out through a multimode fiber with a core diameter of d = 550 μm and a numerical aperture of NA = 0.22.

[0085] To obtain a signal equivalent to measuring the fluorescence response from a microbial colony using a dual-fiber array located at a distance of ~1-2 mm from the measured object, lenses with a large diameter and short focal length were used so that the angular size of the detecting lens located at the focal length was comparable in order of magnitude to the angular size of the dual-fiber array fiber end face at a distance of approximately 1-2 mm from the measured object. Given the limitations on the optical array height, lenses with a focal length of 15 to 20 mm and an aperture (diameter) of 10 mm were used. To enhance the spectrometer's sensitivity in the near UV range, lenses made of KU-1 fused quartz, which has high transmittance in the range >280 nm, were used. Mirrors were coated with an aluminum coating, providing high reflectivity in the range of 280-1000 nm.

[0086] The microorganism identification device was then used to measure the optical response and subsequently identify microorganisms belonging to clinically significant species important for medical decision-making. For this purpose, the device was used to perform measurements on Petri dishes containing Columbia agar medium for microorganism colonies incubated for 24 hours under standard conditions. The microorganism species, independent of the optical measurements performed by the device, was determined using time-of-flight mass spectrometry to determine the microorganism's taxonomic category.In total, to determine the final accuracy of taxonomic identification of microorganisms using the device, 10,000 measurements were carried out on samples with 25 clinically significant categories of microorganisms - microorganisms of the genus Lactobacillus, Neisseria, Corynebacterium, anaerobic microorganisms, as well as microorganisms of the species Enterococcus faecalis, Enterococcus faecium, Staphylococcus aureus, Staphylococcus lugdunensis, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus pneumonia, Pseudomonas aeruginosa, Acinetobacter baumannii, Stenotrophomonas maltophilia, Candida albicans, Candida auris, Moraxella catarrhalis, Haemophilus influenzae, Gardnerella vaginalis. The distribution of the number of objects of different types used to test the device is shown in Figs. 18–20.For each measurement, the "true" microorganism class, identified using time-of-flight mass spectrometry, was known, as was the device's prediction based on optical imaging, fluorescence spectroscopy, and a combination of these methods. An example of an image and fluorescence spectra obtained by the device for colonies of Candida auris is shown in Fig. 19.

[0087] To determine the accuracy of the device's taxonomic identification, the match between the prediction obtained by the device and the "true" class label was determined for each measured sample. The number of true positive (TP), false positive (FP), and false negative (FN) device responses was then calculated. This was used to calculate the F-measure classification metric, as F = 2*TP / (2*TP + FP + FN), for each of the classes under consideration. The F-metric values ​​obtained for each class were then averaged, which characterized the final classification quality. The values ​​of the per-class classification metrics obtained using the two methods are shown in Fig. 20. Device testing revealed that taxonomic identification of microorganisms is possible with F = 0.78 using the optical visualization method implemented within the device.Using fluorescence spectroscopy, the accuracy of taxonomic identification is F = 0.83, and when combining two optical methods, the accuracy was F = 0.85, with the data of the two optical modalities determined according to the scheme shown in Fig. 21. That is, each of the used methods - optical visualization using a digital camera of the digital optical unit, and fluorescence diagnostics carried out using an optical head - had high taxonomic identification accuracies (F = 0.78 and F = 0.83, respectively). Moreover, when combining two optical methods in the implemented identification system, an increase in the accuracy of taxonomic identification of microorganisms to F = 0.85 was achieved.

[0088] Thus, the developed spectroscopic head and optical module are simple to manufacture and maintain, easily integrating into existing smart incubator modules and units containing a digital camera. The developed and tested optical circuit of the spectroscopic head, used to deliver excitation radiation and detect the fluorescence response signal from a microorganism colony, demonstrated the ability to obtain fluorescence signals equivalent to those obtained using a dual-fiber transceiver probe, which was used to collect optical response statistics. The developed circuit is fully compatible with the electromechanical module for moving the spectroscopic head, ensuring high-precision microorganism identification based on optical response data obtained using the proposed device.

Claims

CLAUSES OF THE INVENTION 1. A spectroscopic head for a digital optical unit of a device for identifying microorganisms in a biological sample under study, comprising a holder and two optical systems placed in the holder, configured to input and output optical radiation in orthogonal planes, one of which is intended to supply exciting radiation, the second - to detect a fluorescent response signal from the biological sample; each of the optical systems contains an optical fiber placed in light guide channels, two convex lenses - an input and an output one, and a flat mirror located on the optical path between the lenses, and the holder is made as an assembly of at least three parts having adjacent profiled surfaces configured with the possibility of forming, upon combining parts of two light guide channels with the placement of the said optical systems therein.

2. A spectroscopic head according to claim 1, characterized in that the optical fibers are located in a plane parallel to the surface of the container placement area.

3. A spectroscopic head according to claim 1, characterized in that it is configured to form an angle between the optical axes of the optical systems in a vertical plane, ensuring the possibility of coincidence of the focal spots formed by the optical systems on the surface of the biological sample being studied.

4. The spectroscopic head according to item 1, characterized in that the angle between the optical axes of the optical systems is from 30 to 42 degrees.

5. The spectroscopic head according to claim 1, characterized in that it is designed with the ability to detect a fluorescent response signal at a distance of up to 2 mm from the biological sample being studied.

6. A spectroscopic head according to I.1, characterized by the fact that lenses with a focal length of 15 to 20 mm and an aperture with a diameter of 10 mm are used, while the lenses are made of fused quartz of the KU-1 brand.

7. The spectroscopic head according to claim 1, characterized in that the mirrors have a coating characterized by maximum reflectivity in the wavelength range from 280 to 800 nm.

8. A spectroscopic head according to I.1, characterized by the fact that a multimode optical fiber with a core diameter of d = 200 - 600 µm and a numerical aperture of NA = 0.22 is used.

9. The spectroscopic head according to item 1, characterized in that the ends of the optical fibers are located at a distance Ax from 0 to 5 mm from the focus of the lens.

10. A spectroscopic head according to claim 1, characterized in that grooves are made in the light guide channels for accommodating lenses and mirrors of optical systems.

11. The spectroscopic head according to i.1, characterized in that the holder is made in the form of a parallelepiped of three parts - a central element and two side elements located on opposite sides relative to the central element; the elements are formed by dissecting the parallelepiped with two sectional planes located at an angle to the vertical longitudinal plane of the parallelepiped, wherein the elements of the holder contain parts of light-guide channels in cross-section, configured with the possibility of forming light-guide channels when combining the corresponding side elements with the central element, where one of the light-guide channels is configured with the possibility of placing an optical system intended for supplying exciting radiation, the second light-guide channel - with the possibility of placing a second optical system intended for detecting a fluorescent response signal from a biological sample.

12. The spectroscopic head according to claim 1, characterized in that the light guide channel contains a section having a diameter dl corresponding to the diameter of the optical fiber for its fixed placement in the channel, this section of the channel is connected to a section of a larger diameter d2, intended for the propagation of radiation from the optical fiber - for the first optical system, or to the optical fiber - for the second optical system, with a change in the direction of propagation of radiation by 90 degrees using a mirror installed in the channel.

13. The spectroscopic head according to claim 1, characterized in that the holder in the spectroscopic head has a height of 25 mm or less, a width and length of 75 mm or less.

14. A digital optical unit of a device for identifying microorganisms in a biological sample being studied, including: - a housing containing a platform for placing a container with a biological sample; - a digital camera located in the upper part of the body above the platform for placing the container, - a spectroscopic head, made according to paragraph 1 with the possibility of movement, at least in a horizontal plane above the platform with container; - at least two lamps - upper and lower, located to ensure uniform illumination of the container with the biological sample.

15. A digital optical unit according to claim 14, characterized in that the movement of the spectroscopic head is realized with the help of an electromechanical movement drive equipped with a belt transmission, wherein the electromechanical drive is located on the bottom side of the housing of the digital optical unit.

16. A digital optical unit according to item 14, characterized in that the elements of the optical systems, placed in the light-guide channels of the holder, are configured with the possibility of inputting and outputting optical radiation in orthogonal planes.

17. A digital optical unit according to item 14, characterized in that the digital camera is equipped with a drive for its vertical movement for focusing.

18. A digital optical unit according to item 14, characterized in that it is equipped with a means for moving the container, containing a transport basket for the container.

19. A digital optical unit according to item 18, characterized in that the means for moving the container is provided with a shutter, designed with the possibility of reciprocating movement in the horizontal direction and placement in the photography zone under the transport basket with the possibility of blocking the light flow from the lower lamps.

20. A digital optical unit according to item 18, characterized in that the means for moving the container in the horizontal direction is provided with separate horizontally oriented guide rails for moving the curtain along them.

21. A digital optical unit according to item 14, characterized in that the spectroscopic head is fixed on a movable platform, wherein the electromechanical drive contains two stepper motors with pulleys and a belt drive with a cross-shaped arrangement of belts.

22. A digital optical unit according to claim 21, characterized in that the cross-shaped arrangement of the belts is realized using four rollers placed on a movable platform, wherein the movement of the belts along each of the X and Y axes is limited by mechanical stops and limit switches based on optical sensors.

23. A device for identifying microorganisms, comprising a spectroscopic head according to claim 1, located in a digital optical unit according to claim 14, connected to a light source configured to select the required set of wavelengths, and a spectrometer.

24. The device according to item 23, characterized in that a plasma light source is used with the ability to emit at wavelengths in the range of 280-500 nm and a spectral brightness from 1 mW / (mm*sr*nm) to 35 mW / (mm*sr*nm), wherein the plasma light source is equipped with a monochromator.

25. The device according to item 23, characterized in that a spectrometer (detector) is used with a radiation detection range of 300-800 nm with a spectral resolution of no worse than 5 nm and having a scattered light detection level of no more than 0.5%.

26. The device according to item 24, characterized in that a monochromator is used that has a spectral range of radiation tuning of 280-500 nm, with an allocated spectral bandwidth of no more than 10 nm in the specified range.

27. A system for identifying microorganisms, comprising a device for optical identification of microorganisms, made according to claim 23, connected to a module for controlling the movement of a spectroscopic head, including control of the positioning of the optical head according to images obtained from a digital camera, modules for controlling the movement of a digital camera and a container, as well as a module for controlling optical measurements and a module for analyzing the obtained images and fluorescence spectroscopy data.

28. The system according to claim 27, characterized in that the image analysis module contains computer vision tools.

29. The system according to item 27, characterized in that the image analysis module is configured to pre-process images, ensuring visualization of the area associated with the nutrient medium and the microorganisms cultivated on them.

30. The system according to I.27, characterized in that the image analysis module contains a convolutional neural network used for classifying microorganisms.

31. The system according to I.30, characterized by the fact that a model with a ConvNeXt-type architecture, pre-trained on the ImageNet database, is used.

32. The system according to I.27, characterized in that the image analysis module contains means for classifying microorganisms based on the results fluorescence spectroscopy using machine learning tools, including fully connected neural networks trained using the amplitudes of the principal components of the fluorescence spectra at the stage of pre-processing optical spectroscopy data and optical image data.

33. The system according to claim 27, characterized in that the optical measurement control module is configured to excite emission at several excitation wavelengths from 280 to 500 nm, and detection from 300 to 800 nm.

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