JP2008185518A - Device and method for detecting bacterium - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a device and method for detecting bacteria capable of determining the existence of bacteria lighted by fluorescence from a plurality of fields of view at a high speed in image processing of fluorescent observation. <P>SOLUTION: This bacteria detecting device for detecting the existence of fluorescence-labeled bacteria includes an optical means 11 for receiving the light obtained by irradiating a specimen with light of specific wavelength and outputting it as light information, a visual field moving means 20 for moving the visual field of the optical means 11, an imaging means 12 for imaging the light information as an image in an exposed state during moving the visual field, a position specifying means 14 that extracts brightness information on a linear or curved scan line crossing the moving direction from the image photographed by the imaging means 12, determines the existence of fluorescence, and specifies the position of the visual field including the fluorescence, when the visual field moving means 20 moves from the target visual field to the adjacent next visual field, and a bacteria detecting means 17 for detecting the fluorescence-labeled bacteria from the image of the visual field including the fluorescence. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to bacterial fluorescence observation, and more particularly to a technique for identifying a visual field in which bacteria that fluoresce from a plurality of visual fields are present.

  The agar culture method is the most common method for classifying bacteria. This is an inspection method in which colonies formed by applying a sample to an agar medium and culturing for a predetermined time are classified by an observer using a microscope, with or without staining. However, since this agar culture method is basically performed manually, the process is troublesome, and since it is cultured, it takes time to determine the type of bacteria.

Therefore, there is a fluorescence observation method for observation without culturing. This is an inspection method in which a specimen is adjusted so that only bacteria fluoresce, and an observer observes the fluorescence using optical means such as a falling-down fluorescence microscope. In addition, the detection of fluorescent bacteria has been performed manually by an observer, but in recent years, it has been devised to perform it automatically. As a normal automatic process, the captured image for one field of view is scanned all over, a bacteria detection process is performed, and it is performed for the entire field of view. However, processing time is required to perform the bacteria detection process on the entire visual field. Therefore, as a method for detecting at high speed an acid-fast bacterium with a low appearance frequency of fluorescent bacteria, the image for the entire visual field is added to the image for one visual field. A technique is disclosed in which an acid-fast bacilli detection process is performed on a generated image for one field of view (see, for example, Patent Document 1).
JP 2004-333151 A

  However, in the configuration of Patent Document 1, since all pixels in the field of view are scanned regardless of the presence or absence of acid-fast bacteria, processing time is required for detection of acid-fast bacteria.

  In addition, it is assumed that there is no noise in the photographed image, but there is actually noise. For example, when observing 1000 fields per plate, adding an image for 1000 fields to a single image also increases the noise by 1000, and detects a difference in brightness value from the portion where the acid-fast bacteria are fluorescent. This makes it difficult to make accurate judgments.

  The present invention solves the above-mentioned conventional problem, and observes a plurality of fields in fluorescence observation of bacteria, and specifies a field where bacteria that rapidly fluoresce exist when the appearance frequency of the bacteria that fluoresce is low An object of the present invention is to provide a detection apparatus and a bacteria detection method.

  In order to solve the above-described conventional problems, the bacteria detection apparatus of the present invention receives light obtained by irradiating a specimen with light of a specific wavelength in a bacteria detection apparatus that detects the presence or absence of fluorescently labeled bacteria. Optical means for outputting as optical information, visual field moving means for moving the visual field of the optical means, imaging means for imaging the optical information as an image while being exposed while moving the visual field, and the visual field moving means When moving from the target field of view to the next field of view adjacent to the target, the luminance information on the scanning line of a straight line or a curve that crosses the moving direction is extracted from the image captured by the imaging unit to determine the presence or absence of fluorescence, And a bacteria detection means for detecting the fluorescently labeled bacteria from the image of the visual field including the fluorescence.

  Furthermore, in the bacteria detection apparatus, in the bacteria detection apparatus for detecting the presence or absence of fluorescently labeled bacteria, optical means for receiving light obtained by irradiating a specimen with light of a specific wavelength and outputting it as optical information; A visual field moving means for moving the visual field of the means, an imaging means for capturing the optical information as an image while being exposed while moving the visual field, and when the visual field moving means moves in a wider range than one visual field. A range specifying means for extracting luminance information on a scanning line of a straight line or a curve crossing the moving direction from an image taken by the imaging means and determining the presence or absence of fluorescence, and the visual field moving means moves by one visual field or less. Position specifying means for specifying the position of the field of view containing fluorescence from the luminance information of a scanning line of a straight line or a curve that traverses the moving direction in the one field of view image taken by the imaging means, It is characterized in that the field of view of the image that contains the fluorescent including bacterial detection means for detecting the fluorescence-labeled bacteria.

  Furthermore, in the bacteria detection apparatus, the position specifying unit specifies a region in a direction perpendicular to a moving direction of a visual field including fluorescence based on a location where the luminance information of the scanning line has a feature of fluorescence. Is.

  Further, in the bacteria detection apparatus, in the position specifying unit, the visual field including the fluorescence is an area in the moving direction based on the feature that the luminance information of the plurality of scanning lines fluoresces and the area indicating the trajectory of the plurality of scanning lines. It is characterized by specifying.

  Further, in the bacteria detection apparatus, in the position specifying unit, a region having a fluorescent feature of the luminance information of the scanning line, a fluorescent feature of the luminance information of the plurality of scanning lines, and a region indicating the trajectory of the plurality of scanning lines Based on the above, the region is specified in the vertical direction and the moving direction with respect to the moving direction of the visual field including the fluorescence.

  Furthermore, in the bacteria detection apparatus, when the region is specified by the position specifying means, bacteria are detected from an image obtained by photographing the region specified in the bacteria detection step in detail.

  Furthermore, in the bacteria detection method of the present invention, in the observation of the fluorescence reaction of the specimen, the field of view observed by the imaging means is photographed while being exposed while moving from the currently observed field of view by a movement amount of one field or less. The exposure photographing step for acquiring a single field image, and the position of the field of view in which fluorescence is included in the field of view observed by the imaging means from the luminance information of a scanning line of a straight line or a curve that crosses the moving direction of the field of view in the single field of view image And a bacteria detection step of detecting fluorescent bacteria in the specimen from the image of the visual field containing the fluorescence.

  Further, in the method for detecting bacteria, in the observation of the fluorescence reaction of the specimen, the field of view observed by the imaging means is photographed while being exposed while moving from the field of view currently observed to a range wider than one field of view. A wide-field exposure photographing step of acquiring a range, and a range specifying step of determining whether or not fluorescence is included in the wide range from luminance information of a scanning line of a straight line or a curved line crossing the moving direction of the visual field in the wide-field image, In the range including the fluorescence, exposure shooting is performed in which the field of view observed by the imaging unit moves while moving from the currently observed field of view by an amount of movement equal to or less than one field of view and is captured while being exposed. A position of a visual field including fluorescence in the visual field observed by the imaging unit, based on luminance information of a scanning line that is a straight line or a curved line that traverses the moving direction of the visual field in the one-field image Is characterized in that comprises a position specifying step of specifying, a bacteria detection step of detecting the fluorescence bacteria in the sample from the field of view of the image that contains the fluorescent.

  Further, in the method for detecting bacteria, in the position specifying step, a region perpendicular to the moving direction of the visual field including the fluorescence is specified based on a location where the luminance information of the scanning line has a fluorescent feature. Is.

  Further, in the bacteria detection method, in the position specifying step, the field of view including the fluorescence is a region in the moving direction based on the feature that the luminance information of the plurality of scanning lines fluoresces and the region indicating the trajectory of the plurality of scanning lines. It is characterized by specifying.

  Further, in the bacteria detection method, in the position specifying step, a region having a fluorescent feature of the luminance information of the scanning line, a fluorescent feature of the luminance information of the plurality of scanning lines, and a region indicating the trajectory of the plurality of scanning lines Based on the above, the region is specified in the vertical direction and the moving direction with respect to the moving direction of the visual field including the fluorescence.

  Furthermore, in the bacteria detection method, when an area is specified in the position specifying step, bacteria are detected from an image obtained by photographing the area specified in the bacteria detecting step in detail.

  According to the bacteria detection apparatus and the bacteria detection method of the present invention, a plurality of visual fields are observed in the fluorescence observation of bacteria, and when the frequency of appearance of fluorescent bacteria is low, the field of appearance of fluorescent bacteria is identified at high speed. Can do.

  Embodiments of the bacteria detection apparatus and bacteria detection method of the present invention will be described below in detail with reference to the drawings.

  FIG. 1 shows a block diagram of a bacteria detection apparatus in the first embodiment of the present invention. In FIG. 1, a plate 1 is an object to be inspected. The optical unit 11 irradiates the plate 1 with excitation light and outputs the light from the plate 1 to the imaging unit 12.

  The imaging unit 12 acquires a signal for controlling imaging from the control unit 18. The imaging unit 12 captures an area under fluorescence observation, generates an image, and transmits the image to the control unit 18. The exposure photographing means 13 performs exposure while moving by one field of view, and transmits a signal for photographing to the control means 18. The position specifying unit 14 includes a possibility determining unit 15 and a visual field specifying unit 16. The possibility determination unit 15 acquires a photographed image from the control unit 18, determines the possibility that fluorescent bacteria exist, and transmits the determined result to the control unit 18. The visual field specifying unit 16 acquires information on the scanning line from the control unit 18, specifies a visual field in which fluorescent bacteria may exist, and transmits the result of the specified visual field to the control unit 18. The bacteria detection means 17 acquires the image of the specified visual field from the control means 18, detects bacteria from the specified visual field image, and transmits the bacteria detection result to the control means 18. The control means 18 is an image photographed from the imaging means 12, a signal for exposure photographing from the exposure photographing means 13, a result judged from the possibility judging means 15, a result of the visual field identified from the visual field identifying means 16, and bacteria. A detection result is acquired from the detection means 17. Further, the control means 18 controls the imaging means 12 to control photographing, the image obtained by exposure to the possibility judging means 15, the scanning line information to the visual field specifying means 16, and the visual field image specified to the bacteria detecting means 17. And a signal for controlling movement to the moving means 20 and a detection result to the display means 19. The display means 19 acquires the detection result and displays the detection result. The moving means 20 acquires a signal for controlling movement from the control means 18 and moves the field of view being observed.

  Next, the plate 1 will be described in detail. The plate 1 uses a slide glass and is coated with a specimen that is adjusted so that bacteria are fluorescent. If necessary, a heat treatment or the like is performed. Examples of the specimen include human sputum and stool. Note that any plate 1 such as a petri dish may be adopted as the plate 1.

  Next, the optical means 11 will be described in detail with reference to FIG. The optical means 11 performs fluorescence observation using a fluorescence microscope. FIG. 2 is an explanatory view showing an example of processing of the optical means 11 in fluorescence observation. First, a mercury lamp or the like is used as the light source 21 in order to irradiate the plate 1 with light 22 from the light source 21 including ultraviolet rays. Light 22 from the light source 21 passes through the excitation filter 23. The excitation filter 23 transmits light having a wavelength necessary for the bacteria to fluoresce (hereinafter referred to as excitation light 24), and shields unnecessary light. Next, the excitation light 24 is guided to the objective lens 25 by the dichroic mirror 29 and irradiated onto the plate 1 via the objective lens 25. The dichroic mirror 29 is installed at an angle of 45 degrees with respect to the light source 21 and reflects only a specific wavelength. The specimen is fluorescent with the irradiated excitation light 24, and the fluorescence 27 passes the objective lens 25, passes through the absorption filter 28, and guides the fluorescence to the imaging unit 12. The absorption filter 28 transmits the fluorescence 27 and blocks light 26 other than fluorescence, such as reflected excitation light 24 and external light. Note that any optical means 11 such as a micrometer capable of fluorescence observation may be employed as the optical means 11.

  Next, the imaging means 12 will be described in detail. The imaging means 12 uses a monochrome CCD camera and is joined to a fluorescence microscope as the optical means 11 with a C mount. Note that any imaging means 12 such as a color CCD camera may be employed as the imaging means 12. The fluorescence 27 from the specimen guided by the optical means 11 is photographed by a CCD camera, and the photographed image is transmitted to the control means 18.

  Next, the exposure photographing means 13 will be described in detail with reference to FIGS. FIG. 3 is an explanatory diagram showing an example of movement in one shooting. FIG. 3A is a diagram illustrating a state in which a region 33 to be photographed is in the current visual field 31. FIG. 3B is a diagram showing a state in which the area 33 to be photographed is moved from the current visual field 31 to the next visual field 32 by one visual field.

  FIG. 4 is an explanatory diagram showing an example of the one-field image 41 and the scanning line 42 taken while performing exposure. 4A shows a one-field image 41 taken while performing exposure while moving from the current field of view 31 to the next field of view 32 as shown in FIG. 3, and a linear scanning line 42 perpendicular to the moving direction. FIG. The scanning line 42 is predetermined as a straight line or a curve that crosses the moving direction. Further, the position of the scanning line 42 is determined in advance, for example, at the center of the captured image. FIG. 4B is a graph showing the luminance value of each pixel in the scanning line 42 that sequentially acquires the luminance value pixel by pixel from the top to the bottom of FIG.

  First, the exposure photographing means 13 transmits a control signal for the image pickup means 12 to start exposure to the control means 18 in a state where there is a photographing region 33 at the position of FIG. Next, the exposure photographing means 13 transmits a control signal for moving from the current visual field 31 to the next visual field 32 to the control means 18. When the imaging region 33 moves to the position shown in FIG. 3B, a control signal for ending the exposure of the imaging unit 12 is transmitted to the control unit 18. When the captured one-field image 41 includes fluorescence 27 such as bacteria 44 that fluoresce, the fluorescence locus 43 is photographed in a line.

  Next, the possibility determination means 15 will be described in detail with reference to FIG. First, the possibility determination unit 15 acquires the one-view image 41 taken from the control unit 18. Next, the possibility determination means 15 determines the possibility that the bacteria 44 that fluoresce exist in the visual field from the luminance information of the scanning line 42 by using the captured one visual field image 41. As a method for determining the possibility of the presence of the fluorescent bacteria 44, the fluorescent bacteria 44 may exist when the range 46 in which the luminance value of the predetermined value 45 or more continues in the predetermined range in FIG. 4B. Judge that there is sex. An optimum predetermined value 45 is determined between the luminance values of the background region and the fluorescent region. Since the optimum value varies depending on the system configuration, photographing conditions, and the like, the observer performs evaluation based on images of a plurality of samples and sets a predetermined value 45 in advance. For example, the average value of the luminance values of the background region and the fluorescent region of a plurality of samples is calculated, and the median value is set to the predetermined value 45. Further, the minimum and maximum sizes of the fluorescence 27 are determined in advance, and the range is determined as a predetermined range. Since the optimum value varies depending on the system configuration, imaging conditions, and the like, the observer performs evaluation based on images of a plurality of samples and sets a predetermined range in advance. For example, the minimum value and the maximum value of the size of the fluorescence 27 are calculated for a plurality of samples, and the range is set as a predetermined range. Next, the possibility determination unit 15 transmits the determined result to the control unit 18.

  Next, the visual field specifying means 16 will be described in detail with reference to FIG. FIG. 5 is an explanatory diagram showing an example of specifying the visual field. FIG. 5A is a diagram showing that the fluorescent object 51 is present in the next field of view 32. FIG. 5B is a diagram showing a locus region 53 of the scanning line 42 when moving from left to right as shown in FIG. FIG. 5C is a view showing the image 54 and the fluorescent object 51 of the identified visual field when the region 53 of the trajectory of the scanning line 42 is photographed.

  If there is a possibility in the above-described possibility determination means 15, the visual field identification means 16 acquires position information such as the current position 31 and the position of the scanning line 42 from the control means 18, and sets the locus region 53 of the scanning line 42. Identify the field of view to include.

  An example of a method for specifying the field of view will be described. First, consider the case of movement as shown in FIG. If the fluorescent object 51 is present at the position shown in FIG. 5A, the one-field image 41 taken as shown in FIG. 4A is taken. As shown in FIG. 4A, the scanning line 42 is a straight line perpendicular to the moving direction, and is defined as the center position of the image. The region 33 to be photographed is moved from the position of the next visual field 32 to a position including the region 53 of the trajectory of the scanning line 42. For example, from the left end of the next visual field 32 to the position 52 of the scanning line 42 in the current visual field 31, the next visual field 32 returns to the current visual field 31 in the direction opposite to the moving direction. When photographing is performed in this state, an image 54 of the specified visual field as shown in FIG. 5C is photographed. It is specified that the fluorescent object 51 is present in the image 54 of the specified field of view. The visual field specifying means 16 transmits the specified result to the control means 18.

  Next, the bacteria detection means 17 will be described in detail. The bacteria detecting means 17 determines whether or not there is bacteria in the visual field image 54 specified by the control means 18 based on luminance information and the like. For example, it is determined whether or not the area of connected pixels whose luminance value in each pixel is a predetermined value 45 or more is within the range of the size of bacteria. Since the range of the size of bacteria varies depending on the system configuration, imaging conditions, etc., the observer performs an evaluation based on images of a plurality of samples, and determines the range of the size of bacteria in advance. Further, the bacteria detection means 17 transmits the result of bacteria detection to the control means 18.

  Next, the control means 18 will be described in detail with reference to FIG. First, the movement of the area 33 to be photographed in the entire visual field will be described with reference to FIG. FIG. 6 is a diagram illustrating an example of a state in which the area 33 to be photographed moves through the entire field of view 61 before the exposure field is completed. FIG. 6A is a diagram illustrating an example of a state in which the region 33 to be photographed moves through the entire visual field 61 before the exposure photographing is completed. FIG. 6B shows a region of the trajectory of the scanning line 42 when an area 62 that is not a scanning target is prepared at both ends in FIG. FIG. 53 is a diagram illustrating an example in which 53 includes the entire field of view 61 region.

  The control means 18 controls the scanning of the entire visual field 61 as follows. In the scanning of the entire visual field 61, for example, as shown in FIG. 6A, if the start position is in the first upper left line in the entire visual field 61, the image is moved from the left end to the right end by one visual field. After moving to the right end and completing the first line, the second line is moved to the left end and the same operation is performed for the second line. When the process is finished for the last line, the process is terminated. Assuming that the scanning line 42 is positioned at the center of the linear observation region perpendicular to the moving direction, the region of the half of one field at the left end and the right end in the movement of the entire visual field 61 is changed to the locus region 53 of the scanning line 42. Since it is not included, the regions at both ends cannot be scanned. For this reason, the region 53 of the trajectory of the scanning line 42 includes the region of the entire visual field 61. For example, as shown in FIG. 6B, areas 62 that are not to be scanned are prepared in advance at both ends of the entire visual field 61, and the locus area 53 of the scanning line 42 includes the entire visual field 61 area.

  The control means 18 controls the following information and performs processing. The control means 18 is an image photographed from the imaging means 12, a signal for exposure photographing from the exposure photographing means 13, a result judged from the possibility judging means 15, a result of the visual field identified from the visual field identifying means 16, and bacteria. The detection result is obtained from the detection means 17, and the control means 18 controls the imaging means 12 to control imaging, the image obtained by exposure exposure to the possibility determination means 15, the information on the scanning line 42 to the visual field identification means 16, and bacteria. The visual field image specified by the detection means 17 and a signal for controlling movement are transmitted to the movement means 20. Further, the control means 18 transmits the detection result to the display means 19 when the scanning of the entire visual field 61 is completed.

  As the display means 19, for example, a CRT monitor is used. As the display means 19, any display means 19 such as a touch panel may be adopted.

  As the moving means 20, for example, an automatic XY stage for a microscope is used. The automatic XY stage is attached as a stage of a fluorescence microscope of the optical means 11. Thereafter, the plate 1 is set on the automatic XY stage. Although the field of view is moved this time by moving the plate 1, for example, the field of view may be moved by moving the imaging unit 12.

  An example of the operation of the bacteria detection apparatus configured as described above will be described with reference to FIG. FIG. 7 is a flowchart showing an example of each step in the bacteria detection method.

  First, in step S1, initial settings such as moving the visual field to the start position are performed. Next, in the exposure photographing process of step S2, the exposure photographing means 13 performs exposure while moving by one field of view, and transmits a control signal for photographing to the control means 18. The control unit 18 acquires a control signal for exposure shooting from the exposure shooting unit 13 and transmits a control signal for starting exposure to the imaging unit 12. Further, the optical means 11 performs fluorescence observation of the plate 1 and outputs the observed fluorescence 27 to the imaging means 12. The imaging unit 12 acquires the fluorescence 27 being observed from the optical unit 11, acquires a control signal for starting exposure from the control unit 18, and starts exposure. Subsequently, the control unit 18 transmits a signal for controlling movement to the moving unit 20. The moving unit 20 obtains a signal for controlling the movement from the control unit 18 and moves by one field of view. Subsequently, the control unit 18 transmits a control signal for ending the exposure to the imaging unit 12. The imaging unit 12 acquires a control signal for ending the exposure from the control unit 18, ends the exposure, and transmits the captured one-field image 41 to the control unit 18.

  Next, in acquiring luminance information of the scanning line 42 in step S <b> 3, the control unit 18 acquires the captured one-field image 41 from the imaging unit 12 and transmits it to the possibility determination unit 15. The possibility determination unit 15 acquires the captured one-field image 41 from the control unit 18, and acquires, for example, a luminance value pixel by pixel from the upper part to the lower part in the captured one-field image 41.

  Next, in the step of examining the fluorescence feature in step S <b> 4, the control unit 18 acquires the result of the possibility determination unit 15 determining whether or not there is a fluorescence feature from the possibility determination unit 15. As a result of the determination, if there is a possibility that the fluorescent bacteria 44 are present in the captured one-field image 41, the control means 18 obtains the result of the possibility determination from the possibility determination means 15, and the process proceeds to step S5. . If there is no possibility, the process proceeds to step S7.

  Next, step S <b> 5 is a visual field specifying step, and the control unit 18 transmits information on the scanning line 42 as a result of determining the possibility to the visual field specifying unit 16. The visual field specifying means 16 acquires information on the scanning line 42 from the control means 18 and transmits the result of specifying the visual field to the control means 18. The control means 18 acquires the result of specifying the visual field from the visual field specifying means 16. The control means 18 transmits a control signal for moving to the specified visual field to the moving means 20 from the result of specifying the visual field. The moving means 20 acquires a control signal for moving to the specified visual field from the control signal 18, and moves to the specified visual field. Subsequently, the control means 18 transmits a control signal for photographing the specified field of view to the imaging means 12. The imaging unit 12 acquires a control signal for photographing the specified field of view from the control unit 18, and images the specified field of view. In addition, the imaging unit 12 transmits the image 54 of the specified visual field to the control unit 18.

  Next, step S <b> 6 is a bacteria detection step, and the control unit 18 acquires the image 54 of the specified visual field from the imaging unit 12 and transmits the image 54 of the specified visual field to the bacteria detection unit 17. The bacteria detection means 17 acquires the image 54 of the specified visual field from the control means 18 and transmits the detection result of the fluorescent bacteria 44 to the control means 18. The control means 18 acquires a detection result.

  Next, in step S7, when the entire visual field 61 to be determined is completed, the control unit 18 outputs the result to the display unit 19 in step S8, and the process is terminated. On the other hand, if there is the next visual field 32, the process returns to step S2 and steps S3 to S6 are repeated.

  As described above, in the first embodiment, exposure is performed while moving for one field of view, and the possibility of the presence of fluorescent bacteria 44 from the captured one field image 41 is determined. Moreover, the visual field which has a possibility is specified and it is determined whether the image 54 of the specified visual field contains bacteria. When the appearance frequency of the bacteria 44 that fluoresce in one visual field is small, it is possible to determine one field of view by partial scanning, so that the bacterial fluorescence 27 can be determined at high speed.

  Further, in the visual field specifying step, which is step S5 in FIG. 7 of the first embodiment, there is the following method as a method for narrowing the region in the moving direction with the image 54 of the specified visual field. FIG. 8 shows that the fluorescent object 51 may exist in a region in the moving direction within the range 82 including the range 46 in which the luminance value equal to or greater than the predetermined value 45 in FIG. It is explanatory drawing which shows narrowing down as the characteristic area | region 81. FIG. The processing time can be shortened by narrowing the target region in the bacteria detection as shown in FIG.

  Further, in the visual field specifying step, which is step S5 of the control means 18 of the first embodiment, there is the following method as a method for narrowing the region in the direction perpendicular to the moving direction in the image 54 of the specified visual field. FIG. 9 is an explanatory diagram showing a method of narrowing the region in the direction perpendicular to the moving direction in the image 54 of the identified visual field. FIG. 9A is a diagram showing that two scanning lines 42 are acquired at the left end and the center of the one-field image 41 taken while being exposed. FIG. 9B shows the locus region 53a of the scanning line 42a drawn when the scanning line 42a moves to move by one visual field and the locus region of the scanning line 42b drawn when the scanning line 42b moves to move by one visual field. It is explanatory drawing which shows 53b. In FIG. 9C, for example, when the scanning line 42a has no fluorescent feature and the scanning line 42b has a fluorescent feature, the region 53b of the scanning line 42b overlaps the region 53a of the scanning line 42a. It is explanatory drawing which shows narrowing down as the area | region 91 in which the fluorescent object 51 except for may exist. The processing time can be shortened by narrowing the target region in the bacteria detection as shown in FIG.

  Further, in the visual field specifying step, which is step S5 in FIG. 7 of the first embodiment, the following method is used as a method of narrowing down the region in both the direction perpendicular to the moving direction and the moving direction in the image 54 of the specified visual field. There is. FIG. 10 is an explanatory diagram showing a region 101 in which there is a possibility that a fluorescent object 51 having a region narrowed in the moving direction as shown in FIG. 8 and further narrowed in a direction perpendicular to the moving direction as shown in FIG. is there. The processing time can be further shortened by narrowing down the target region for bacteria detection as shown in FIG.

  In the first embodiment, the bacteria are detected using the image 54 of the visual field specified by the bacteria detecting means 17, but the region where the fluorescent bacteria 44 are present by the above-described method is detected from the specified visual field. For example, the aperture and the optical means 11 are used to take an image of the area 101 where the fluorescent object 51 may exist by increasing the magnification, and use the image of the area 101 where the fluorescent object 51 may exist in detail. Thus, bacteria can be detected with high accuracy by detecting bacteria.

  In the first embodiment, the image capturing unit 12 is captured by a monochrome CCD. However, in the image captured using the image capturing unit 12 capable of capturing in color such as RGB, specific color information is present in the fluorescent bacteria 44, for example. When there is a feature such as no feature, the feature of the fluorescence can be accurately detected by adding a condition such as the absence of specific color information when calculating the feature of the fluorescence in the scanning line 42.

  Only differences from the first embodiment will be described with reference to FIG. In FIG. 11, the wide-field exposure photographing unit 111 performs exposure while moving one row in the entire visual field 61 and transmits a signal for photographing to the control unit 18. The range specifying unit 112 acquires information on the scanning line 42 from the control unit 18, specifies a range in which the fluorescent bacteria 44 may exist, and transmits the result of the specified range to the control unit 18. The control unit 18 further obtains a signal for exposing and photographing a wide field of view from the wide field exposure photographing unit 111 and a result of the range specified by the range specifying unit 112. Further, the control unit 18 further transmits the information of the scanning line 42 to the range specifying unit 112.

  Next, the wide-field exposure photographing unit 111 will be described in detail with reference to FIG. FIG. 12 is an explanatory diagram showing that a region 33 to be imaged is present at the left end in the entire visual field 61 and exposure is performed while moving to the right end.

  First, the wide-field exposure photographing unit 111 transmits a control signal for starting exposure to the control unit 18 in a state where it is at the left end in the entire visual field 61. Next, the wide-field exposure photographing unit 111 transmits a control signal for moving the region 33 to be photographed from the left end to the right end in the entire visual field 61 to the control unit 18. When the imaging region 33 moves to the rightmost position in the entire visual field 61, a control signal for ending the exposure of the imaging unit 12 is transmitted to the control unit 18. That is, while the exposure photographing means 13 is moved by one field of view, photographing is performed while performing exposure, whereas the wide field exposure photographing means 111 is 1 from the left end to the right end in the entire field 61 as shown in FIG. While moving by line, take a picture while performing exposure. At this time, similarly to the first embodiment, a region not to be scanned similar to that shown in FIG. 6B may be provided so as to include the regions at both ends.

  Next, the range specifying means 112 will be described in detail with reference to FIG. First, the range specifying unit 112 acquires a wide-field image taken from the control unit 18. Next, the range specifying means 112 uses the captured wide-field image to determine the possibility of the presence of fluorescent bacteria 44 in the visual field from the luminance information of the scanning line 42. As a method for determining the possibility of the presence of the fluorescent bacteria 44, the fluorescent bacteria 44 may exist when the range 46 in which the luminance value of the predetermined value 45 or more continues in the predetermined range in FIG. 4B. Judge that there is sex. Next, the range specifying unit 112 transmits the determined result to the control unit 18.

  An example of the operation of the bacteria detection apparatus configured as described above will be described with reference to FIG. FIG. 13 is a flowchart showing an example of each step in the bacteria detection method.

  First, in step S21, initial settings such as moving the visual field to the start position are performed. Next, in the wide-field exposure photographing process in step S22, the wide-field exposure photographing means 111 performs exposure while moving by one line, and transmits a control signal for photographing to the control means 18. The control unit 18 acquires a control signal for exposure shooting from the wide-field exposure imaging unit 111 and transmits a control signal for starting exposure to the imaging unit 12. Further, the optical means 11 performs fluorescence observation of the plate 1 and outputs the observed fluorescence 27 to the imaging means 12. The imaging unit 12 acquires the fluorescence 27 being observed from the optical unit 11, acquires a control signal for starting exposure from the control unit 18, and starts exposure. Subsequently, the control unit 18 transmits a signal for controlling movement to the moving unit 20. The moving unit 20 acquires a signal for controlling the movement from the control unit 18 and moves by one line. Subsequently, the control unit 18 transmits a control signal for ending the exposure to the imaging unit 12. The imaging unit 12 acquires a control signal for ending the exposure from the control unit 18, ends the exposure, and transmits the captured wide-field image to the control unit 18. In addition, the control unit 18 acquires a captured wide-field image from the imaging unit 12.

  Next, in acquiring the luminance information of the scanning line 42 in step S23, the control unit 18 transmits the captured wide-field image to the range specifying unit 112. The range specifying unit 112 acquires a captured wide-field image from the control unit 18, and acquires, for example, a luminance value pixel by pixel from the top to the bottom in the captured one-field image 41.

  Next, in the step of examining the fluorescence characteristics in step S24, the range specifying unit 112 determines the possibility that the fluorescent object 51 is included in the captured wide-field image. Further, the result of determining the possibility is transmitted to the control means 18. From the result of determining the possibility, the control means 18 transmits a control signal for moving to the start position of the moved line to the movement means 20 if there is a possibility. The moving unit 20 acquires a control signal for moving to the start position of the row from the control unit 18, moves the position of the imaging region 33 at the right end to the left end, and proceeds to step S25. If there is no possibility, the process proceeds to step S31.

  Next, in the exposure photographing process of step S25, the exposure photographing means 13 performs exposure while moving by one field of view, and transmits a control signal for photographing to the control means 18. The control unit 18 acquires a control signal for exposure shooting from the exposure shooting unit 13 and transmits a control signal for starting exposure to the imaging unit 12. Further, the optical means 11 performs fluorescence observation of the plate 1 and outputs the observed fluorescence 27 to the imaging means 12. The imaging unit 12 acquires the fluorescence 27 being observed from the optical unit 11, acquires a control signal for starting exposure from the control unit 18, and starts exposure. Subsequently, the control unit 18 transmits a signal for controlling movement to the moving unit 20. The moving unit 20 obtains a signal for controlling the movement from the control unit 18 and moves by one field of view. Subsequently, the control unit 18 transmits a control signal for ending the exposure to the imaging unit 12. The imaging unit 12 acquires a control signal for ending the exposure from the control unit 18, ends the exposure, and transmits the captured one-field image 41 to the control unit 18.

  Next, in acquiring the luminance information of the scanning line 42 in step S <b> 26, the control unit 18 acquires the captured one-field image 41 from the imaging unit 12 and transmits it to the possibility determination unit 15. The possibility determination unit 15 acquires the captured one-field image 41 from the control unit 18, and acquires, for example, a luminance value pixel by pixel from the upper part to the lower part in the captured one-field image 41.

  Next, in the step of examining the fluorescence characteristics in step S <b> 27, the control means 18 acquires the result of the possibility determination means 15 determining whether or not there is a fluorescence characteristic from the possibility determination means 15. As a result of the determination, if there is a possibility that the fluorescent bacteria 44 are present in the captured one-field image 41, the control means 18 obtains the result of the possibility determination from the possibility determination means 15, and the process proceeds to step S28. . If there is no possibility, the process proceeds to step S30.

  Next, step S28 is a visual field specifying step, and the control unit 18 transmits information on the scanning line 42 as a result of determining the possibility to the visual field specifying unit 16. The visual field specifying means 16 acquires information on the scanning line 42 from the control means 18 and transmits the result of specifying the visual field to the control means 18. The control means 18 acquires the result of specifying the visual field from the visual field specifying means 16. The control means 18 transmits a control signal for moving to the specified visual field to the moving means 20 from the result of specifying the visual field. The moving means 20 acquires a control signal for moving to the specified visual field from the control means 18 and moves to the specified visual field. Subsequently, the control means 18 transmits a control signal for photographing the specified field of view to the imaging means 12. The imaging unit 12 acquires a control signal for photographing the specified field of view from the control unit 18, and images the specified field of view. In addition, the imaging unit 12 transmits the image 54 of the specified visual field to the control unit 18.

  Next, step S <b> 6 is a bacteria detection step, and the control unit 18 acquires the image 54 of the specified visual field from the imaging unit 12 and transmits the image 54 of the specified visual field to the bacteria detection unit 17. The bacteria detection means 17 acquires the image 54 of the specified visual field from the control means 18 and transmits the detection result of the fluorescent bacteria 44 to the control means 18. The control means 18 acquires a detection result.

  Next, in step S30, when it complete | finishes about 1 line which performs determination, the control means 18 transfers to step S31. On the other hand, if there is the next visual field 32, the process returns to step S25, and steps S26 to S29 are repeated.

  Next, in step S31, when the entire visual field 61 to be determined is finished, the control means 18 outputs the result to the display means 19 in step S32, and the process is finished. On the other hand, if there is a next line, the control means 18 transmits a control signal for moving to the next line to the movement means 20. Further, the moving means 20 acquires a control signal for moving to the next line, and moves to the next line. Then, it returns to step S22 and repeats steps S23-S30.

  As described above, while moving by one line in the second embodiment, exposure is performed to photograph, and the possibility of the presence of fluorescent bacteria 44 is determined from the photographed wide-field image. In addition, exposure is performed while the target line is moved by one field of view, and the possibility of the presence of fluorescent bacteria 44 from the captured one-field image 41 is determined. Moreover, the visual field which has a possibility is specified and it is determined whether the image 54 of the specified visual field contains bacteria. When the frequency of appearance of the bacteria 44 that fluoresce in one line is low, the fluorescence 27 of the bacteria can be determined at a higher speed than in the first embodiment by performing the determination for one line by partial scanning.

  According to the bacteria detection apparatus and the bacteria detection method of the present invention, a luminescent object is present in a moved area by exposing and photographing a luminescent object while moving and scanning a part from the characteristics of the photographed image. It has a function of determining possibility and is useful as an image recognition technique.

The block diagram of the bacteria detection apparatus in Example 1 of this invention. Explanatory drawing of the fluorescence observation in the optical means 11 of Example 1 and 2 of this invention Explanatory drawing which shows an example of the movement in exposure imaging | photography of Example 1 and 2 of this invention Explanatory drawing which shows an example of the image image | photographed by exposure imaging | photography of Example 1 and 2 of this invention. Explanatory drawing which shows an example which specifies the visual field of Example 1 and 2 of this invention Explanatory drawing which shows an example of the imaging | photography area | region of Example 1 and 2 of this invention which moves a full visual field. The flowchart of the bacteria detection method in Example 1 of this invention Explanatory drawing which shows an example which restrict | squeezes an area | region with the specific visual field of Example 1 and 2 of this invention Explanatory drawing which shows an example which restrict | squeezes an area | region with the specific visual field of Example 1 and 2 of this invention Explanatory drawing which shows an example which restrict | squeezes an area | region with the specific visual field of Example 1 and 2 of this invention The block diagram of the bacteria detection apparatus in Example 2 of this invention. Explanatory drawing which shows an example of the movement in wide view exposure imaging | photography of Example 2 of this invention. Flowchart of bacteria detection method in Example 2 of the present invention

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Plate 11 Optical means 12 Imaging means 13 Exposure imaging means 14 Position identification means 15 Possibility determination means 16 View field identification means 17 Bacteria detection means 18 Control means 19 Display means 20 Moving means 21 Light source 22 Light from light source 21 Excitation filter 24 Excitation light 25 Objective lens 26 Light other than fluorescence 27 Fluorescence 28 Absorption filter 29 Dichroic mirror 31 Current field of view 32 Next field of view 33 Imaged area 41 Single field of view image 42 Scanning line 43 Fluorescence locus 44 Fluorescent bacteria 45 Predetermined value 46 Continuous Scope 51 Fluorescent object 52 Position of scanning line 42 in current visual field 31 from left end of next visual field 32 53 Trajectory region of scanning line 42 Image of specified visual field 61 Whole field 62 Non-scan target region 81 Fluorescent object Territory where 51 may exist 82 Range 91 potential region 111 wide viewing exposure shooting means 112 range specifying means region 101 fluorescent substance 51 that may fluorescent object 51 is present is present

Claims (12)

In a bacteria detection apparatus for detecting the presence or absence of fluorescently labeled bacteria,
Optical means for receiving light obtained by irradiating a specimen with light of a specific wavelength and outputting it as optical information;
Visual field moving means for moving the visual field of the optical means;
An imaging means for imaging the optical information as an image while being exposed while moving the field of view;
When the visual field moving means moves from the target visual field to the next adjacent visual field, the luminance information on a scanning line of a straight line or a curved line crossing the moving direction is extracted from the image captured by the imaging means to determine the presence or absence of fluorescence. A position specifying means for determining and specifying a position of a visual field including fluorescence;
Bacteria detection means for detecting the fluorescently labeled bacteria from an image of a field of view containing the fluorescence,
Bacterial detection device characterized by that. In a bacteria detection apparatus for detecting the presence or absence of fluorescently labeled bacteria,
Optical means for receiving light obtained by irradiating a specimen with light of a specific wavelength and outputting it as optical information;
Visual field moving means for moving the visual field of the optical means;
An imaging means for imaging the optical information as an image while being exposed while moving the field of view;
Specifying a range for determining the presence or absence of fluorescence by extracting luminance information on a scanning line of a straight line or a curve crossing the moving direction from an image taken by the imaging unit when the visual field moving unit moves in a wider range than one visual field Means,
The position of the visual field including the fluorescence is specified from the luminance information of a scanning line of a straight line or a curve that crosses the moving direction in the one visual field image captured by the imaging unit when the visual field moving unit moves by one field of view or less. Positioning means;
Bacteria detection means for detecting the fluorescently labeled bacteria from an image of a field of view containing the fluorescence,
Bacterial detection device characterized by that. 3. The position specifying unit specifies a region in a direction perpendicular to a moving direction of a visual field including fluorescence based on a location where the luminance information of the scanning line has a fluorescent feature. Bacteria detection device. The position specifying means specifies a region in a moving direction of a visual field including fluorescence based on a feature that the luminance information of the plurality of scanning lines fluoresces and a region indicating a locus of the plurality of scanning lines. The bacteria detection apparatus according to claim 1 or 2. In the position specifying means, the fluorescence is generated based on a location where the luminance information of the scanning line has a fluorescent feature, a fluorescent feature of the luminance information of the plurality of scanning lines, and a region indicating a trajectory of the plurality of scanning lines. The bacteria detection apparatus according to claim 1 or 2, wherein the region is specified in a direction perpendicular to a moving direction of the visual field included and a moving direction. 3. The bacteria detection apparatus according to claim 1, wherein when the region is specified by the position specifying means, bacteria are detected from an image obtained by photographing the region specified in the bacteria detection step in detail. In the observation of the fluorescence reaction of the specimen, an exposure photographing step of obtaining a single field image by photographing while moving while moving the field of view observed by the imaging means by a moving amount of one field or less from the field of view currently observed When,
In the one-field image, a position specifying step for specifying a position of a visual field including fluorescence in a visual field observed by the imaging unit from luminance information of a scanning line of a straight line or a curve that traverses the moving direction of the visual field,
Including a bacteria detecting step of detecting fluorescent bacteria in a specimen from an image of a field of view containing the fluorescence,
A method for detecting bacteria. In the observation of the fluorescence reaction of the specimen, a wide-field exposure is performed in which the field of view observed by the imaging means moves from a field currently being observed to a range wider than one field of view, and a wide-field image is obtained while being exposed. Shooting process,
In the wide-field image, a range specifying step for determining whether fluorescence is included in the wide range from luminance information of a straight line or a curved scanning line that crosses the moving direction of the visual field, and
An exposure photographing step in which a field of view observed by the imaging means is moved by a moving amount of one field of view or less from the field of view currently observed in the range including the fluorescence to capture a single field of view image while being exposed. When,
In the one-field image, a position specifying step for specifying a position of a visual field including fluorescence in a visual field observed by the imaging unit from luminance information of a scanning line of a straight line or a curve that traverses the moving direction of the visual field,
Including a bacteria detecting step of detecting fluorescent bacteria in a specimen from an image of a field of view containing the fluorescence,
A method for detecting bacteria. 9. The position specifying step specifies a region in a direction perpendicular to a moving direction of a visual field including fluorescence based on a location where the luminance information of the scanning line has a fluorescent feature. Bacteria detection method. In the position specifying step, based on the feature that the luminance information of the plurality of scan lines fluoresces and the region that indicates the trajectory of the plurality of scan lines, the region that specifies the field of view including the fluorescence in the moving direction is specified. The method for detecting bacteria according to claim 7 and 8. In the position specifying step, the fluorescence is generated based on a location where the luminance information of the scanning line has a fluorescent feature, a fluorescent feature of the luminance information of the plurality of scanning lines, and a region indicating the trajectory of the plurality of scanning lines. The bacteria detection method according to claim 7 or 8, wherein regions are specified in a direction perpendicular to a moving direction of the visual field included and a moving direction. The bacteria detection method according to claim 7 or 8, wherein when the region is specified in the position specifying step, bacteria are detected from an image obtained by photographing the region specified in the bacteria detection step in detail.

Images