• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    It is proposed a non-invasive device for determining the fertility and / or sex of an egg (1). This device comprises means (200) for obtaining a spectral response of at least a portion of at least one layer of the shell (10), including its surface, to an incident light signal, and means ( 300) for determining the fertility and / or the sex of the egg (1) as a function of the spectral response obtained.
    公开号:FR3023617A1
    申请号:FR1456744
    申请日:2014-07-11
    公开日:2016-01-15
    发明作者:Marc Schortgen
    申请人:TRONICO；
    IPC主号:
    专利说明:

    [0001] Non-invasive device for determining the fertility and / or sex of an egg, and corresponding method FIELD OF THE INVENTION The field of the invention is that of poultry farming.
    [0002] More specifically, the invention relates to a non-invasive technique for determining the fertility of an egg and / or the sex thereof (i.e. the genus, male or female, of the embryo contained in the egg if it has been fertilized). 2. TECHNOLOGICAL BACKGROUND For many years, the poultry field has been dealing with the recurrent problem of determining the fertility and sex of eggs before they hatch. Traditionally, sexing is done on the day-old chick by human intervention mainly via two techniques: the observation of the feathers and the observation of the cloaca.
    [0003] Although effective, a major disadvantage of these two techniques is that they intervene too late in the production cycle. For example, for laying hens, it takes 21 days of incubation to separate females from males (the latter being eliminated). Determining the genus of embryos at an early stage of incubation would improve poultry production, saving both time and money. As a result, a number of more or less invasive determination techniques have been developed to evaluate the fertility and sex of a non-hatched bird egg.
    [0004] A first known technique is described in patent US 6029080, which proposes a method of determination by nuclear magnetic resonance. This technique makes it possible to sort eggs into three categories: eggs containing a male embryo, those containing a female embryo and clear eggs (not fertilized). Sorting is done by observing the reproductive organs through the shell. More particularly, the observation is feasible only after cooling of the egg to minimize the movements of the embryo. A disadvantage of this technique is that the cooling of the egg causes disturbances in the future development of the embryo. Another disadvantage lies in the relatively expensive implementation of such a technique, inducing a brake on its industrial development. A second known technique is described in patent application US2011 / 0144473, which describes the use of UV spectroscopy. This technique consists in emitting a wave in the ultraviolet so as to cause a self-fluorescence of a region of the germ (blastoderm). To determine the sex of the embryo, a decay coefficient is computed and compared with a database, following the observation of the decay of the auto-fluorescence after stopping the emission of the UV wave. However, a disadvantage of this technique is that it requires access to the blastoderm by invasive sampling. A third known technique is based on infrared spectroscopy. This method, also invasive, consists of the introduction of a probe in the region of the germ, in order to characterize the constitutive cells by infrared spectroscopy.
    [0005] A major disadvantage of the second and third known techniques is that they are invasive: they require the introduction of instruments and the removal of cells in the germ. These operations are therefore not without risks for the future development of the embryo. Another disadvantage of these two techniques is the risk of contamination of the whole brood.
    [0006] In order to overcome this major drawback, a fourth known, non-invasive technique is described in patent application US2013 / 0044210. It consists of a hyper-spectral analysis (ie from the mid-infrared to the near-UV) of an optical spectrum in reflection of the egg, subtracting the spectral response of the shell and detecting the components biological of the egg. This technique makes it possible to determine, from the second day, whether the egg is fertilized and, on the twelfth day, the genus of the embryo. A major disadvantage of this technique is that it requires corrections to overcome the shell (subtraction of its spectral response) and environmental disturbances (light, moisture, etc.). For this purpose, each egg must be placed on a specific support, maintaining a known distance between the radiation source, the detector and the egg. This technique is therefore difficult to assimilate.
    [0007] Probably because of the aforementioned drawbacks, none of the known techniques mentioned above has really given rise to industrial development. OBJECTIVES OF THE INVENTION The invention, in at least one embodiment, has the particular objective of overcoming these various disadvantages of the state of the art. More specifically, in at least one embodiment of the invention, an objective is to provide a technique for determining the fertility and / or sex of an egg, which is non-invasive (no penetration of the shell by a any tool, nor any sampling in the egg) and simpler to implement, and therefore more easily industrialized, than the fourth known technique mentioned above. Another object in at least one embodiment of the invention is to provide such a technique which is reliable. 4. OBJECTIVES OF THE INVENTION These objectives, as well as others which will appear more clearly later, are achieved by means of a non-invasive device for determining the fertility and / or sex of an egg. comprising means for obtaining a spectral response of at least a portion of at least one layer of the shell, including its surface, to an incident light signal, and means for determining the fertility and / or sex of the egg according to the spectral response obtained.
    [0008] The general principle of the invention therefore consists in obtaining a spectral response of at least one layer of the shell to an incident light signal and in determining the fertility and / or sex of the egg according to this spectral response. Thus, this particular embodiment of the invention relies on a completely new and inventive approach to noninvasive determination of the fertility and / or sex of an egg, since it relies on surface spectroscopy. and / or inside the shell of the egg. Indeed, as detailed below, the inventors have speculated that the discriminating substances, existing in all the cells of the embryo and playing a role in sexual differentiation, are not concentrated exclusively in the gonads, but diffuse in the egg through the allantoic fluid and may be in part minute but measurable, be present on the surface and inside the shell.
    [0009] It is recalled that in the fourth known technique (the only noninvasive technique known to the inventors), on the contrary, it is sought to overcome the spectral response of the shell by subtracting it from that of the egg, to retain only the spectral response. the contents of the egg (including albumin, yolk and germinal disc). The incident light signal is either a single-wavelength light signal (ie having a single wavelength) or a multi-wavelength light signal having at least two wavelengths (successively or simultaneously) .
    [0010] According to a particular characteristic, the incident light signal comprises at least one wavelength of between 200 and 1100 nm. This range of wavelengths allows optimal operation. According to one particular aspect of the invention, the means for obtaining the spectral response comprise focussing means adapted to focus the incident light signal on a surface portion and / or an internal portion of said at least one layer of the shell. . Such means make it possible to focus the light signal on a precise layer of the shell to be examined. According to a particular aspect, the focusing means are adapted to focus the incident light signal on a portion of the outer surface of an outer layer of the shell. The advantage of focusing on the surface of the shell is that the reflected signal (for example a reflected Raman signal) is richer, the absorption being less. According to a particular aspect, the focusing means comprise a probe, having a focusing distance D relative to a lens, and a tip whose one end is integral with the probe and placed at a distance dl from the lens, and a second end is intended to be in contact with the egg, the tip having a distance d2 between the first and second ends, with d2 = D-dl.
    [0011] Such a tip thus allows the probe to always be at the same distance from the surface of the egg. In other words, the tip allows easy adaptability of the device to all sizes of eggs, without additional adjustment. The industrialization of such a device is therefore naturally conceivable. The distance between the lens of the probe and the egg being fixed by the nozzle calibrated in length, the focusing distance D can easily be adjusted by changing the wavelength of the incident light signal. Such an aspect also allows a high accuracy of the device. In addition, the adjustment and maintenance of such a positive are facilitated since only the wavelength is considered variable. According to a particular aspect, the means for obtaining the spectral response comprise a probe, or said probe, adapted to transmit the incident light signal towards the egg and to receive, with a zero reflection angle, a reflected light signal resulting from a reflection by the coffin of the incident light signal. In this way, even if the reflected light signal is relatively weak, such a probe makes it possible to ensure and optimize the recovery of the reflected signal. In a particular embodiment of the invention, the probe is a Raman type probe.
    [0012] Such a probe allows a high accuracy in the transmission of the incident signal and in the reception of the reflected signal. According to a particular aspect of the invention, the means for determining the fertility and / or sex of the egg comprise means for comparing the spectral response obtained with at least one reference spectral response.
    [0013] Such a comparison makes it easier to determine the fertility and / or sex of the egg. Indeed, the comparison means make it possible to avoid precisely determining which discriminating substances are present, but to compare the general appearance of the spectrum obtained with one or more reference spectra.
    [0014] According to a particular aspect of the invention, said at least one reference spectral response belongs to the group comprising: a first spectral reference response for unfertilized eggs; a second reference spectral response for fertilized eggs containing a dead embryo; a third reference spectral response for fertilized eggs containing a male embryo; and a fourth reference spectral response for fertilized eggs containing a female embryo. The invention also relates to a non-invasive method for determining the fertility and / or sex of an egg, comprising a step of obtaining a spectral response of at least a portion of at least one layer of the shell an incident light signal, and a step of determining the fertility and / or sex of the egg as a function of the spectral response obtained. 5. LIST OF FIGURES Other features and advantages of the invention will become apparent on reading the following description, given by way of indicative and nonlimiting example, and the appended drawings, in which: FIG. 1 shows the structure simplified of a fertilized egg; - Figure 2 is a sectional view of the shell structure of an egg; - Figure 3 shows a device for determining the fertility and / or sex of an egg, according to a particular embodiment of the invention; FIG. 4 shows a particular embodiment of the probe appearing in FIG. 3; and FIG. 5 illustrates an example of a spectrum obtained with the device of FIG. 3. DETAILED DESCRIPTION OF AN EMBODIMENT In all the figures of the present document, the elements and identical steps are designated by the same reference numeral. The inventors started from several observations, namely: 1. The sexual differentiation of the embryo does not essentially result from the action of hormones on the gonads. 2. Somatic cells have an autonomous sexual identity. 3. The main substances involved in the female determination include: a. proteins such as FOXL2 involved in maintaining ovarian function; b. estrogens such as estradiol secreted by the ovary of the embryo; vs. enzymes such as MHM or Aromatase (CYP19A1) that help convert steroid hormones into estrogen; d. genetic material such as the HINTW gene (or WPKCI) present in several tissues of the female embryo as well as the genes DAX1, WNT4, FET1. 4. The main substances involved in the male determination include: a. hormones, such as the AMH responsible for regression of the female genital tract; b. genetic material, such as the DMRT1 gene that participates in testicular formation, or SOX9 that participates in gonad formation. The aforementioned substances (hereinafter also referred to as "discriminating substances") are of different types: - degradation products resulting from the biosynthesis, such as the aromatase which indicates the presence of steroids, the hormone AMH present in the allantoic fluid in greater quantity for the female embryo than for the male, the MHM enzyme or the HINTW gene; - steroids, such as estradiol are also present in the allantoic fluid. The inventors hypothesized that these discriminant substances, existing in all embryo cells and playing a role in sexual differentiation, were not concentrated exclusively in the gonads, but spread in the egg via of the allantoic fluid and can in part small but measurable, either migrate to and through the shell (during the evacuation of water vapor or CO2), or become embedded in the shell. Depending on their size, they are found either on the surface of the shell or embedded in the shell (for example in the shell membranes). As illustrated in FIG. 1, an egg 1 comprises a shell 10 and an internal content that itself comprises albumin 20, yolk 30 and germinal disc 40.
    [0015] As illustrated in FIG. 2, the shell 10 comprises a superposition of six layers, namely (starting from the albumin 20): the inner shell membrane 11, the outer shell membrane 12, the mamillary layer 13, the palisade layer 14 traversed by the pores 17, a monolayer of vertical crystals (15) and finally the cuticle 16 enveloping all the mineral layers. As indicated above, and illustrated in FIG. 1, a plurality of discriminating substances 50 are present not only in the internal content (and more specifically in the albumin 20), but also on the surface and / or inside the the shell 10. Although weak, the presence and the quantity of these discriminating substances 50 on the surface and / or in the shell 10 are then measurable by spectroscopy. FIG. 3 shows a device 100 for determining the fertility and / or sex of an egg, according to a particular embodiment of the invention. Raman spectroscopy is a non-destructive analysis technique for characterizing the molecular composition and structure of a material. This technique is based on the detection of photons scattered inelastically following the interaction of the sample with a monochromatic light beam (Raman scattering). The difference in frequency between the exciter photon and the reflected photon informs about the chemical nature of the substance (molecule) at the origin of the diffusion.
    [0016] In other words, Raman scattering is the physical phenomenon by which a medium is able to slightly modify the frequency of the light circulating there. This frequency shift corresponds to an exchange of energy between the light beam and the medium. This exchange may have several causes: vibrations of the crystal or of the molecule, magnetic excitations, & c. The measurement of this shift makes it possible to go back to certain properties of the medium. In the particular embodiment described below, Raman spectroscopy is used to determine the fertility and / or sex of an egg. It is clear however that the present invention is not limited to this type of spectroscopy, but can be implemented with any technique making it possible to obtain the spectral response of at least a portion of at least one layer of the shell. . The general principle of the proposed technique is as follows: a small surface of the shell receives at least one incident electromagnetic wave of defined length. According to the technique employed, the generated spot (hereinafter also referred to as the "point or focusing zone") will have, for example, a diameter of a few microns to 150 microns. The incident wave is reflected at the same frequency for the most part, and at different frequencies for a small part. Depending on the type of spectroscopy used, one exploits one or the other of the reflected waves. In the embodiment shown, the device 100 comprises: means 200 for obtaining a spectral response of at least a portion of at least one layer of the shell 10 of an egg 1. These means 200 comprise a light source 101, a first optical fiber 102, a probe 110, a tip 104, a second optical fiber 105 and a spectrometer 106; means 300 for determining the fertility and / or sex of the egg 1 as a function of the spectral response obtained. These means 300 comprise a computer 107, a database 109 and a display 108. The light source 101 is for example a laser capable of providing an incident light signal. More particularly, the light signal provided by the laser 101 is for example an electromagnetic wave of single wavelength, preferably between 200 and 1100 nm (and even more preferably between 500 and 700 nm). In one variant (multi-wavelength spectroscopy), the incident light signal is multi-wavelength, with at least two wavelengths (successive or simultaneous) each ranging between 200 and 1100 nm. This variant makes it possible to obtain a more complete spectrum, and consequently more reliable results. It also makes it possible to overcome the fluorescence. The probe 110 and the tip 104 together form focusing means. The first optical fiber 102 is intended to transmit the incident light signal to the probe 110. The tip 104 is calibrated in length and integral with the probe 110. It allows the probe 110 to be located at a constant distance from the cuff 1. A first end of the tip 104 is integral with the probe 110, and a second end of the tip 104 is intended to come into contact with the egg 1.
    [0017] The response of the light signal reflected by the surface of the shell 10 of the egg 1 being relatively small, the probe 110 is, in a particular embodiment, adapted to recover the signal reflected in the same axis as the incident signal. This type of probe is more commonly known as the Raman probe.
    [0018] The operating principle of such a Raman probe 110 is illustrated in FIG. 4. The incident light signal coming from the first optical fiber 102 is collimated by means of a first lens 111. A collimated light is a light whose radiations are substantially parallel. and unfold slowly as they spread. In other words, it is a light that does not disperse with distance (in theory), or which will be very little dispersed (in practice). This collimated signal is then focused, via a second lens 112, at a point or a focusing zone 113, located on the surface or inside of the shell 10 to be examined. Focus point 113 is also called spot. Since the first and second lenses (111, 112) are fixed, the focusing distance can only be changed by varying the wavelength of the incident light signal. Here, the focusing distance D of the probe 110 is defined as the distance between the second lens 111 and the spot 113. In a particular implementation (illustrated in FIG. 3), the spot 113 is located on the outer surface of the layer 16 outer shell 10. For this purpose, the length d2 of the tip 104 is obtained according to the formula: d2 = D-dl, with dl the distance between a first end of the tip 104 (integral with the probe 104) and the lens 112. A second end of the tip 104, located at the distance d2 of the first end, is intended to be in contact with the outer surface of the egg. In this particular implementation, the incident light signal is reflected (by the portion of the surface of the egg that corresponds to the spot 113) and the reflected signal is collected / collimated by the same lens 112, then directed by means of a mirror dichroic 114 to the spectrometer 106. In order to optimize the signal received by the spectrometer 106, the signal directed by means of a dichroic mirror 114 passes firstly through a high-pass optical filter 115, intended to eliminate the Rayleigh scattering, is then focused by a third lens 116, and is finally transmitted to the spectrometer 106 by the second optical fiber 105.
    [0019] The spectrometer 106 analyzes the received signal and provides a spectral response to the means 300 for determining the fertility and / or sex of the egg 1. Within the means 300 for determining the fertility and / or sex of the egg 1, the calculator 107 and the database 109 form comparison means.
    [0020] The latter make it possible to exploit the spectral response transmitted by the spectrometer 106 and to compare it with a library of spectral reference responses. This comparison relates either to the amplitude of one or more lines of the spectral response, or to the pace of the whole of the spectral response. In a particular embodiment, the following four reference spectral responses are used: a first spectral reference response, for unfertilized eggs; a second spectral reference response for fertilized eggs containing a dead embryo; a third spectral reference response, for fertilized eggs containing a male embryo; a fourth reference spectral response, for fertilized eggs containing a female embryo. In a variant, the group of unfertilized eggs and that of fertilized eggs containing a dead embryo can be grouped together in one and the same group.
    [0021] In other words, the first and fourth reference spectral responses are one and the same reference spectral response. The display 108 makes it possible to display the spectral response obtained, as well as the result of comparing it with the database of the spectral reference responses.
    [0022] The obtaining of the database 109 comprises for example the following steps: Step 1: on a brood ("configuration brood"), a Raman spectrum (spectral response of the shell according to the technique described above in relation to the Figures 3 and 4) is taken from each egg at different days of incubation; - Step 2: a correlation is performed to determine the areas that differ from one spectrum to another (zones of determination). This method makes it possible to classify spectra into categories (spectral reference responses). Step 3: The sexing of the chicks then makes it possible to classify these categories, that is to say to associate with each category one of the following information: "male embryo", "female embryo" and "clear egg or embryo dead ". Steps 1 to 3 can be repeated on several "configuration clutches". The database 109 is thus enriched by the regular provision of new measures.
    [0023] The decision algorithm as to the fertility and / or sex of an egg comprises, for example, the following stages, for each egg of a given brood ("brood to be tested"): Step a: a Raman spectrum (response spectral shell according to the technique described above in relation to Figures 3 and 4) is noted on the test egg; Step b: the quality of the measurement is evaluated on each determination zone (see definition above), for example by calculating the first derivative and the second derivative of the spectrum. For example, the quality criterion is calculated by taking into account the measurement dynamic represented by the maximum deviation of the measurement in the zone, the variability of the data represented by the number of slope inversion in the zone given by the first derivative and the noise level given by the standard deviation of the second derivative in the zone. Step c: if the quality criterion is sufficient, the spectrum is compared, by correlation on the determination zones, with the various reference spectra of the database, in order to determine the category and therefore the type (among "Male embryo", "female embryo" and "light egg or dead embryo"). FIG. 5 illustrates an example of spectral response (spectrum) obtained with the device 100 of FIG. 3. This spectrum is representative of a combination of the responses of the substances 50 present on the surface and / or inside the shell 10 The comparison of these spectra with the spectra of biological samples referenced in the literature makes it possible to associate the visible bands with the presence of certain compounds or molecules. For example, bands A are associated with calcium carbonate, bands B with carotenoids, C with DNA, D with lipid and protein structures and E with phenylalanine. The comparison with the database is performed on these bands but also on intermediate bands where the signals are weaker as in the 800 to 900 cm 'zone.
    [0024] In yet another embodiment, the length d2 of the tip 104 is calculated so that the focus point 113 is located in one of the layers of the shell 10. In other words, the presence of the discriminating substances 50 no longer on the outer surface of the outer layer 16 of the shell 10, but inside one of the layers (11, 12, 13, 14, 15 and 16) forming the shell 10.
    权利要求:
    Claims (10)
    [0001]
    REVENDICATIONS1. Non-invasive device for determining the fertility and / or sex of an egg (1), characterized in that it comprises means (200) for obtaining a spectral response of at least one portion of at least one layer of the shell (10), including its surface, at an incident light signal, and means (300) for determining the fertility and / or sex of the egg (1) as a function of the spectral response obtained.
    [0002]
    2. Device according to claim 1, characterized in that the incident light signal comprises at least one wavelength of between 200 and 1100 nm.
    [0003]
    3. Device according to any one of claims 1 and 2, characterized in that the means (200) for obtaining the spectral response comprise focusing means (110, 104) adapted to focus the incident light signal on a portion. surface (113) and / or an inner portion of said at least one layer of the shell (10).
    [0004]
    4. Device according to claim 3, characterized in that the focusing means (110, 104) are adapted to focus the incident light signal on a portion of the outer surface of an outer layer (16) of the shell (10) .
    [0005]
    5. Device according to any one of claims 3 and 4, characterized in that the focusing means (110, 104) comprise a probe (110) having a focusing distance D relative to a lens (112), and a tip (104) whose first end is secured to the probe and placed at a distance dl from the lens (112), and a second end is intended to be in contact with the egg, the tip having a distance d2 between the first and second ends, with d2 = D-dl.
    [0006]
    6. Device according to any one of claims 1 to 5, characterized in that the means (200) for obtaining the spectral response comprise a probe (110), or said probe, adapted to transmit the light signal incident to the egg (1) and to receive, with a zero angle of reflection, a reflected light signal resulting from reflection by the egg (1) of the incident light signal.
    [0007]
    7. Device according to claim 6, characterized in that the probe (110) is a Raman type probe.
    [0008]
    8. Device according to any one of claims 1 to 7, characterized in that the means (300) for determining the fertility and / or sex of the egg (1) comprise comparison means (107, 109). the spectral response obtained with at least one reference spectral response.
    [0009]
    9. Device according to claim 8, characterized in that said at least one reference spectral response belongs to the group comprising: a first spectral reference response for unfertilized eggs; a second reference spectral response for fertilized eggs containing a dead embryo; a third reference spectral response for fertilized eggs containing a male embryo; and a fourth reference spectral response for fertilized eggs containing a female embryo.
    [0010]
    10. A non-invasive method for determining the fertility and / or sex of an egg (1), characterized in that it comprises a step of obtaining a spectral response of at least a portion of at least one a layer of the shell (10) to an incident light signal, and a step of determining the fertility and / or sex of the egg (1) according to the spectral response obtained.
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    优先权:
    申请号 | 申请日 | 专利标题
    FR1456744|2014-07-11|
    FR1456744A|FR3023617B1|2014-07-11|2014-07-11|NON - INVASIVE DEVICE FOR DETERMINING THE FERTILITY AND / OR SEX OF AN EGG AND CORRESPONDING METHOD.|FR1456744A| FR3023617B1|2014-07-11|2014-07-11|NON - INVASIVE DEVICE FOR DETERMINING THE FERTILITY AND / OR SEX OF AN EGG AND CORRESPONDING METHOD.|
    EP15735965.4A| EP3167285B1|2014-07-11|2015-07-09|Non-invasive device and method for determining fertility and/or sex of an egg|
    US15/325,646| US10060854B2|2014-07-11|2015-07-09|Non-invasive device for determining the fertility and/or the sex of an egg, and corresponding method|
    PCT/EP2015/065778| WO2016005539A1|2014-07-11|2015-07-09|Non-invasive device for determining the fertility and/or sex of an egg, and corresponding method|
    RU2017103534A| RU2017103534A3|2014-07-11|2015-07-09|
    TR2018/10666T| TR201810666T4|2014-07-11|2015-07-09|Non-invasive device and corresponding process for determining fertility and / or sex of an egg.|
    BR112017000073A| BR112017000073A2|2014-07-11|2015-07-09|"Noninvasive device for determining fertility and / or sex of an egg and its method"|
    PL15735965T| PL3167285T3|2014-07-11|2015-07-09|Non-invasive device and method for determining fertility and/or sex of an egg|
    CN201580037803.7A| CN106574892B|2014-07-11|2015-07-09|For determining the fertilization of egg and/or non-invasive device and the corresponding method of gender|
    ES15735965.4T| ES2681628T3|2014-07-11|2015-07-09|Non-invasive device for determining the fertility and / or sex of an egg, and corresponding procedure|
    AU2015286598A| AU2015286598A1|2014-07-11|2015-07-09|Non-invasive device for determining the fertility and/or sex of an egg, and corresponding method|
    CA2953271A| CA2953271A1|2014-07-11|2015-07-09|Non-invasive device for determining the fertility and/or sex of an egg,'and corresponding method|
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