JP2011101624A - Anaerobic bacterium-culturing kit, anaerobic bacterium-inspecting device and method for inspecting anaerobic bacterium - Google Patents
Anaerobic bacterium-culturing kit, anaerobic bacterium-inspecting device and method for inspecting anaerobic bacterium Download PDFInfo
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- 238000000034 method Methods 0.000 title claims abstract description 18
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- 241001148471 unidentified anaerobic bacterium Species 0.000 claims description 97
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- 238000001514 detection method Methods 0.000 description 8
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- 241001112696 Clostridia Species 0.000 description 4
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 4
- 239000001301 oxygen Substances 0.000 description 4
- 229910052760 oxygen Inorganic materials 0.000 description 4
- 238000005192 partition Methods 0.000 description 4
- 241000178985 Moorella Species 0.000 description 2
- 241001603561 Thermoanaerobacter mathranii Species 0.000 description 2
- 241000193447 Thermoanaerobacter thermohydrosulfuricus Species 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 229920003002 synthetic resin Polymers 0.000 description 2
- 239000000057 synthetic resin Substances 0.000 description 2
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 241000193459 Moorella thermoacetica Species 0.000 description 1
- 229940123973 Oxygen scavenger Drugs 0.000 description 1
- QABCGOSYZHCPGN-UHFFFAOYSA-N chloro(dimethyl)silicon Chemical compound C[Si](C)Cl QABCGOSYZHCPGN-UHFFFAOYSA-N 0.000 description 1
- 238000012136 culture method Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 230000014670 detection of bacterium Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
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Abstract
Description
本発明は、嫌気性菌培養キット、嫌気性菌検査装置および嫌気性菌検査方法に関する。 The present invention relates to an anaerobic bacteria culture kit, an anaerobic bacteria testing apparatus, and an anaerobic bacteria testing method.
従来の嫌気性菌の検査では、試料を添加した寒天培地を、嫌気チャンバーや脱酸素剤とともに嫌気ジャーに入れて培養し、培養により***増殖した嫌気性菌を確認することにより、嫌気性菌が試料中に存在しているかどうかを判定している（例えば、特許文献１、２または３参照）。細菌の検出に関し、寒天培地を透過する光による投影像に基づいて、細菌を正確かつ迅速に検出することができる投影検出法が本発明者等により提案されている（例えば、特許文献４参照）。 In the conventional anaerobic bacteria test, the agar medium with the sample added is cultured in an anaerobic jar together with an anaerobic chamber and an oxygen scavenger, and the anaerobic bacteria are confirmed by dividing and growing in the culture. It is determined whether or not it exists in the sample (for example, see Patent Documents 1, 2, or 3). Regarding the detection of bacteria, the present inventors have proposed a projection detection method capable of accurately and quickly detecting bacteria based on a projection image by light transmitted through an agar medium (see, for example, Patent Document 4). .
しかしながら、特許文献１、２または３に記載の方法では、嫌気性菌を培養するための設備が大型となり、操作も煩雑であるという課題があった。また、嫌気性菌の成長検出を確認するたびに、各寒天培地を嫌気チャンバーや嫌気ジャーから取り出さなければならず、手間がかかり煩雑であるという課題もあった。 However, the methods described in Patent Documents 1, 2, and 3 have a problem that the equipment for culturing anaerobic bacteria becomes large and the operation is complicated. Moreover, each time the growth detection of anaerobic bacteria is confirmed, each agar medium must be taken out from the anaerobic chamber or anaerobic jar, which is troublesome and cumbersome.
本発明は、このような課題に着目してなされたもので、嫌気性菌を容易かつ正確に検出することができる嫌気性菌培養キット、嫌気性菌検査装置および嫌気性菌検査方法を提供することを目的としている。 The present invention has been made paying attention to such problems, and provides an anaerobic bacteria culture kit, an anaerobic bacteria testing apparatus, and an anaerobic bacteria testing method that can easily and accurately detect anaerobic bacteria. The purpose is that.
上記目的を達成するために、本発明に係る嫌気性菌培養キットは、培地を収納して密封される透明袋と、前記透明袋の収納部を有する培養ケースとを有し、前記収納部は、前記透明袋を保持して収納するよう所定の間隔をあけて互いに対向する１対の透明な側壁板の間に形成され、前記透明袋を入れるための開口を有することを、特徴とする。 In order to achieve the above object, an anaerobic bacterium culture kit according to the present invention has a transparent bag that contains a culture medium and is sealed, and a culture case that has a storage part for the transparent bag. The transparent bag is formed between a pair of transparent side wall plates facing each other at a predetermined interval so as to hold and store the transparent bag, and has an opening for receiving the transparent bag.
本発明に係る嫌気性菌培養キットは、試料の中に嫌気性菌が存在するかどうかを調べるために使用される。透明袋を密封するときは、酸素および空気が入らないようにすることが好ましい。透明袋は、酸素および空気を通さない気密性の袋から成ることが好ましい。なお、側壁板および透明袋は、全体が透明のほか、一部のみが透明であってもよい。 The anaerobic bacteria culture kit according to the present invention is used for examining whether or not anaerobic bacteria are present in a sample. When sealing the transparent bag, it is preferable that oxygen and air do not enter. The transparent bag is preferably made of an airtight bag that is impermeable to oxygen and air. Note that the side wall plate and the transparent bag may be entirely transparent or only partially transparent.
本発明に係る嫌気性菌培養キットは、培養ケースの収納部の各側壁板の間に、培地を入れた透明袋を配置することにより、培地を、各側壁板の所定の間隔と同じ厚みにすることができる。この状態で、培養ケースごとインキュベータなどの培養装置に入れることができる。透明な側壁板を通して培地を観察し、嫌気性菌が試料中に存在しているかどうかを容易かつ正確に確認することができる。 In the anaerobic bacteria culture kit according to the present invention, a transparent bag containing a culture medium is disposed between the side wall plates of the storage part of the culture case, so that the culture medium has the same thickness as a predetermined interval between the side wall plates. Can do. In this state, the entire culture case can be placed in a culture apparatus such as an incubator. It is possible to easily and accurately confirm whether or not anaerobic bacteria are present in the sample by observing the medium through the transparent side wall plate.
培地を収納した透明袋は、各側壁板の間に保持して収納されるため、位置が固定され、経時的に培地の同じ位置を観察することができる。透明袋は、両側の側壁板に密着して保持されることが好ましい。透明袋は、側壁板に密着しやすい合成樹脂の袋から成ることが好ましい。一方の側壁板の側から光を当てることにより、側壁板および透明袋を透過した光による投影像に基づいて、嫌気性菌を正確かつ迅速に検出することができる。 Since the transparent bag containing the medium is held and stored between the side wall plates, the position is fixed, and the same position of the medium can be observed over time. The transparent bag is preferably held in close contact with the side wall plates on both sides. The transparent bag is preferably made of a synthetic resin bag that easily adheres to the side wall plate. By applying light from the side of the one side wall plate, anaerobic bacteria can be detected accurately and quickly based on the projection image of the light transmitted through the side wall plate and the transparent bag.
本発明に係る嫌気性菌培養キットによれば、嫌気チャンバーや嫌気ジャーなどの大型の設備が不要となり、簡単な構成で、嫌気性菌の培養および検出を容易に行うことができる。なお、培地には、透明袋に入れるときに流動性がある寒天培地が好ましい。また、培地には、嫌気性菌の培養に適した培地が好ましく、圧力検出を優先させるときは微生物の利用する糖分配合などを工夫してガスを発生し易くした培地が好ましい。各側壁板の所定の間隔は、各側壁板の間の培地を観察しやすい間隔が好ましく、例えば、シャーレの培地の厚みと同じ程度の間隔が好ましい。 The anaerobic bacteria culture kit according to the present invention eliminates the need for large facilities such as an anaerobic chamber and an anaerobic jar, and can easily culture and detect anaerobic bacteria with a simple configuration. The medium is preferably an agar medium that has fluidity when placed in a transparent bag. Further, the medium is preferably a medium suitable for culturing anaerobic bacteria, and when priority is given to pressure detection, a medium in which gas is easily generated by devising a sugar composition used by microorganisms is preferable. The predetermined interval between the side wall plates is preferably an interval at which the medium between the side wall plates can be easily observed.
前記収納部は、例えば、各側壁板の間に、各側壁板に沿った方向の三方を塞ぐ壁材を有し、各側壁板に沿った方向の残りの一方が前記開口を成し、各側壁板に沿った方向の奥行きおよび幅よりも各側壁板の間隔を狭く形成した簡単な構成でよい。 The storage portion includes, for example, a wall material that covers three sides in the direction along each side wall plate between the side wall plates, and the other one in the direction along each side wall plate forms the opening. A simple configuration in which the interval between the side wall plates is formed narrower than the depth and width in the direction along the line.
前記培養ケースは、前記収納部を複数有し、各収納部は各側壁板に沿った方向に並んで配置されていることが好ましい。この場合、複数の試料を同時に検査することができる。
本発明に係る嫌気性菌培養キットは、各側壁板を挟む位置に前記側壁板にかかる応力を光弾性効果により検出する１対の偏光板を有していてもよい。この場合、偏光板、側壁板、透明袋および培地を透過した光による投影像から、側壁板にかかる応力を光弾性効果により検出することによって、嫌気性菌を検出することができる。各偏光板は、各側壁板の外面に貼り付けられていてもよい。
The culture case preferably includes a plurality of the storage units, and the storage units are arranged side by side in a direction along each side wall plate. In this case, a plurality of samples can be inspected simultaneously.
The anaerobic bacteria culture kit according to the present invention may have a pair of polarizing plates that detect a stress applied to the side wall plate by a photoelastic effect at a position between each side wall plate. In this case, anaerobic bacteria can be detected by detecting the stress applied to the side wall plate by the photoelastic effect from the projection image of the light transmitted through the polarizing plate, the side wall plate, the transparent bag and the culture medium. Each polarizing plate may be affixed to the outer surface of each side wall plate.
本発明に係る嫌気性菌検査装置は、本発明に係る嫌気性菌培養キットと、各側壁板に向けて光を透過させるよう設けられた光源と、前記光源からの前記側壁板、前記透明袋および前記培地を透過した光による投影像を取得可能に設けられた画像取得手段とを、有することを特徴とする。 The anaerobic bacteria testing apparatus according to the present invention includes an anaerobic bacteria culture kit according to the present invention, a light source provided to transmit light toward each side wall plate, the side wall plate from the light source, and the transparent bag. And image acquisition means provided so as to be able to acquire a projection image by light transmitted through the culture medium.
本発明に係る嫌気性菌検査装置は、光源により一方の側壁板に向けて光を照射すると、画像取得手段が側壁板、透明袋および培地を透過した光による投影像を取得する。これにより、培地のコロニーの投影像に基づいて、嫌気性菌を正確かつ迅速に検出することができる。 In the anaerobic bacteria testing device according to the present invention, when light is irradiated toward one side wall plate by a light source, the image acquisition means acquires a projection image by light transmitted through the side wall plate, the transparent bag, and the culture medium. Thereby, an anaerobic microbe can be detected correctly and rapidly based on the projection image of the colony of a culture medium.
本発明に係る嫌気性菌検査装置は、本発明に係る嫌気性菌培養キットと、各側壁板を挟む位置に設けられ、前記側壁板にかかる応力を光弾性効果により検出する１対の偏光板と、各偏光板および各側壁板に向けて光を透過させるよう設けられた光源と、前記光源からの前記偏光板、前記側壁板、前記透明袋および前記培地を透過した光による投影像を取得可能に設けられた画像取得手段とを、有するものであってもよい。 The anaerobic bacteria testing apparatus according to the present invention is provided at a position sandwiching each side wall plate with the anaerobic bacteria culture kit according to the present invention, and detects a stress applied to the side wall plate by a photoelastic effect. And a light source provided to transmit light toward each polarizing plate and each side wall plate, and a projection image by light transmitted through the polarizing plate, the side wall plate, the transparent bag, and the medium from the light source is obtained. The image acquisition means may be provided as possible.
この場合、１対の偏光板を透過した光による投影像を画像取得手段で取得することにより、側壁板にかかる応力を光弾性効果により検出することができる。培地の試料が嫌気性菌を含んでいると、培養された菌が増殖してガスを発生し、透明袋がそのガスで膨らんで各側壁板を外側に向かって押す。このときの側壁板にかかる応力を検出することにより、嫌気性菌の存在を検出することができる。このため、コロニーの検出が難しい場合にも、嫌気性菌の検出が容易である。 In this case, the stress applied to the side wall plate can be detected by the photoelastic effect by acquiring the projection image by the light transmitted through the pair of polarizing plates by the image acquisition means. If the sample of the medium contains anaerobic bacteria, the cultured bacteria grow and generate gas, and the transparent bag expands with the gas and pushes each side wall plate outward. By detecting the stress applied to the side wall plate at this time, the presence of anaerobic bacteria can be detected. For this reason, anaerobic bacteria can be easily detected even when colonies are difficult to detect.
本発明に係る嫌気性菌検査方法は、本発明に係る嫌気性菌培養キットの前記収納部に、嫌気性菌検査用の試料を入れた培地を収納し密封した前記透明袋を配置して嫌気性菌の所定の培養温度で保ち、前記側壁板、前記透明袋および前記培地を透過する光を照射し、透過した光によるコロニーの投影像を検出することを、特徴とする。 In the anaerobic bacteria testing method according to the present invention, the anaerobic bacteria culture kit according to the present invention is provided with the transparent bag in which the medium containing a sample for anaerobic bacteria testing is stored and sealed. It is characterized in that it is maintained at a predetermined culture temperature of the sex bacteria, irradiates light that passes through the side wall plate, the transparent bag and the medium, and detects a projected image of a colony by the transmitted light.
本発明に係る嫌気性菌検査方法は、本発明に係る嫌気性菌培養キットを使用するため、嫌気チャンバーや嫌気ジャーなどの大型の設備が不要となり、簡単な構成で、嫌気性菌の培養および検出を容易に行うことができる。また、均一な厚みの培地を観察することができ、嫌気性菌を容易かつ正確に検出することができる。培地のコロニーの投影像に基づいて、嫌気性菌を正確かつ迅速に検出することができる。 Since the anaerobic bacteria testing method according to the present invention uses the anaerobic bacteria culture kit according to the present invention, large-scale equipment such as an anaerobic chamber and an anaerobic jar is not required. Detection can be performed easily. In addition, a medium with a uniform thickness can be observed, and anaerobic bacteria can be detected easily and accurately. An anaerobic bacterium can be detected accurately and rapidly based on the projection image of the colony of the medium.
他の、本発明に係る嫌気性菌検査方法は、本発明に係る嫌気性菌培養キットの前記収納部に、嫌気性菌検査用の試料を入れた培地を収納し密封した前記透明袋を配置して嫌気性菌の所定の培養温度で保ち、各側壁板を挟む位置に１対の偏光板を設け、前記偏光板、前記側壁板、前記透明袋および前記培地を透過する光を照射し、透過した光による投影像から、前記側壁板にかかる応力を光弾性効果により検出することを、特徴とする。 In another method for testing anaerobic bacteria according to the present invention, the storage bag of the anaerobic bacteria culture kit according to the present invention contains the transparent bag containing a sealed medium containing a sample for anaerobic bacteria testing. And maintaining a predetermined culture temperature of anaerobic bacteria, providing a pair of polarizing plates at a position sandwiching each side wall plate, irradiating light passing through the polarizing plate, the side wall plate, the transparent bag and the medium, The stress applied to the side wall plate is detected by a photoelastic effect from a projected image by transmitted light.
他の本発明に係る嫌気性菌検査方法では、培地の試料が嫌気性菌を含んでいると、培養された菌が増殖してガスを発生し、透明袋がそのガスで膨らんで各側壁板を外側に向かって押す。このときの側壁板にかかる応力を検出することにより、嫌気性菌の存在を検出することができる。このため、培地がくずれたりしてコロニーの検出が難しい場合にも、嫌気性菌の検出が容易である。なお、所定の培養温度としては、至適培養温度が好ましい。 In other anaerobic bacteria testing methods according to the present invention, when the sample of the medium contains anaerobic bacteria, the cultured bacteria grow to generate gas, and the transparent bag is inflated with the gas, and each side wall plate Press outward. By detecting the stress applied to the side wall plate at this time, the presence of anaerobic bacteria can be detected. For this reason, anaerobic bacteria can be easily detected even when it is difficult to detect colonies due to the collapse of the culture medium. The predetermined culture temperature is preferably an optimum culture temperature.
本発明に係る嫌気性菌検査方法では、透明袋に培地を入れ、透明袋を収納部に収納した後、透明袋を密封しても、透明袋に培地を入れ、透明袋を密封した後、透明袋を収納部に収納しても、いずれでもよい。また、透明袋を収納部に収納してから培地を注入しても、透明袋に培地を注入してから収納部に収納しても、いずれでもよい。 In the anaerobic bacteria testing method according to the present invention, after putting the medium in the transparent bag, storing the transparent bag in the storage unit, sealing the transparent bag, putting the medium in the transparent bag, and sealing the transparent bag, The transparent bag may be stored in the storage unit or any of them. The medium may be injected after the transparent bag is stored in the storage unit, or may be stored in the storage unit after the medium is injected into the transparent bag.
本発明によれば、嫌気性菌を容易かつ正確に検出することができる嫌気性菌培養キット、嫌気性菌検査装置および嫌気性菌検査方法を提供することができる。 ADVANTAGE OF THE INVENTION According to this invention, the anaerobic microbe culture | cultivation kit, the anaerobic microbe inspection apparatus, and the anaerobic microbe inspection method which can detect anaerobic microbe easily and correctly can be provided.
以下、図面に基づき本発明の実施の形態について説明する。
図１乃至図７は、本発明の実施の形態の嫌気性菌培養キット、嫌気性菌検査装置および嫌気性菌検査方法を示している。
図１に示すように、嫌気性菌培養キット１０は、培養ケース１１と透明袋１２とを有している。
Hereinafter, embodiments of the present invention will be described with reference to the drawings.
1 to 7 show an anaerobic bacteria culture kit, an anaerobic bacteria testing apparatus, and an anaerobic bacteria testing method according to an embodiment of the present invention.
As shown in FIG. 1, the anaerobic bacteria culture kit 10 includes a culture case 11 and a transparent bag 12.
図１（ａ）および（ｂ）に示すように、培養ケース１１は、基板２１と１対の側壁板２２，２３とを有している。基板２１は、外形が矩形状の薄い不透明の板から成っている。基板２１は、一方の側縁２１ａから他方の側縁２１ｂに向かって矩形状に切り込んで形成された５つの凹部２１ｃを有している。各凹部２１ｃは、基板２１の一方の側縁２１ａに沿って並んで形成されている。基板２１は、各凹部２１ｃの間に細長く残された４つの隔壁部２１ｄを有している。また、基板２１は、表面２１ｅが平坦で、裏面２１ｆが両端部２１ｇより中央部２１ｈが窪んだ形状を成している。基板２１は、両端部２１ｇが厚く、各凹部２１ｃおよび各隔壁部２１ｄを含む中央部２１ｈが両端部２１ｇよりも薄く形成されている。基板２１は、中央部２１ｈが、シャーレで培養する場合の培地の厚みと同じ程度の厚みを有している。 As shown in FIGS. 1A and 1B, the culture case 11 includes a substrate 21 and a pair of side wall plates 22 and 23. The substrate 21 is made of a thin opaque plate having a rectangular outer shape. The substrate 21 has five recesses 21c formed by cutting in a rectangular shape from one side edge 21a to the other side edge 21b. Each recess 21 c is formed side by side along one side edge 21 a of the substrate 21. The substrate 21 has four partition walls 21d that are left elongated between the recesses 21c. The substrate 21 has a flat surface 21e and a back surface 21f in which a central portion 21h is recessed from both end portions 21g. The substrate 21 has thicker end portions 21g, and a central portion 21h including the recesses 21c and the partition walls 21d is formed thinner than the end portions 21g. The substrate 21 has a thickness about the same as the thickness of the medium when the central portion 21h is cultured in a petri dish.
各側壁板２２，２３は、透明な矩形状の板から成り、それぞれ基板２１の裏面２１ｆおよび表面２１ｅに取り付けられている。基板２１の裏面２１ｆの側壁板２２は、基板２１の幅と同じ幅を有し、基板２１の裏面２１ｆの中央部２１ｈと同じ長さを有している。また、側壁板２２は、基板２１の裏面２１ｆの両端部２１ｇと中央部２１ｈとの厚みの差と同じ厚みを有している。側壁板２２は、基板２１の裏面２１ｆの中央部２１ｈに嵌め込んで固定されている。これにより、培養ケース１１は、基板２１の裏面２１ｆの側が、基板２１の両端部２１ｇと側壁板２２とにより、平坦になっている。
側壁板２３は、基板２１の裏面２１ｆの側壁板２２の幅および長さと同じ幅および長さを有し、側壁板２２よりも薄く形成されている。側壁板２３は、側壁板２２と対応する位置で、側壁板２２と平行になるよう基板２１の表面２１ｅに固定されている。
Each of the side wall plates 22 and 23 is made of a transparent rectangular plate and is attached to the back surface 21f and the front surface 21e of the substrate 21, respectively. The side wall plate 22 of the back surface 21f of the substrate 21 has the same width as the width of the substrate 21 and has the same length as the central portion 21h of the back surface 21f of the substrate 21. Further, the side wall plate 22 has the same thickness as the difference in thickness between both end portions 21g and the central portion 21h of the back surface 21f of the substrate 21. The side wall plate 22 is fixed by being fitted into the central portion 21 h of the back surface 21 f of the substrate 21. Thereby, the culture case 11 is flat on the back surface 21 f side of the substrate 21 by the both end portions 21 g of the substrate 21 and the side wall plate 22.
The side wall plate 23 has the same width and length as the side wall plate 22 on the back surface 21 f of the substrate 21, and is formed thinner than the side wall plate 22. The side wall plate 23 is fixed to the surface 21 e of the substrate 21 at a position corresponding to the side wall plate 22 so as to be parallel to the side wall plate 22.
培養ケース１１は、基板２１の各凹部２１ｃと、透明な各側壁板２２，２３とで囲まれた５つの収納部２４を有している。各側壁板２２，２３は、シャーレで培養する場合の培地の厚みと同じ程度の間隔をあけて、互いに対向するよう配置されている。各側壁板２２，２３の間の各隔壁部２１ｄ、基板２１の両端部２１ｇおよび基板２１の他方の側縁２１ｂは、各側壁板２２，２３に沿った方向の三方を塞ぐ壁材を成している。各収納部２４は、各側壁板２２，２３に沿った方向の残りの一方、すなわち基板２１の一方の側縁２１ａの側に開口２４ａを有している。各収納部２４は、各側壁板２２，２３に沿った方向の奥行きおよび幅よりも各側壁板２２，２３の間隔が狭く形成されている。各収納部２４は、各側壁板２２，２３および基板２１の一方の側縁２１ａに沿った方向に並んで配置されている。 The culture case 11 has five storage portions 24 surrounded by the concave portions 21 c of the substrate 21 and the transparent side wall plates 22 and 23. The side wall plates 22 and 23 are arranged so as to face each other with an interval of the same degree as the thickness of the medium when culturing in a petri dish. Each partition wall portion 21d between the side wall plates 22 and 23, both end portions 21g of the substrate 21 and the other side edge 21b of the substrate 21 form a wall material that covers three sides in the direction along the side wall plates 22 and 23. ing. Each storage portion 24 has an opening 24 a on the other side in the direction along the side wall plates 22, 23, that is, on the side of one side edge 21 a of the substrate 21. Each storage portion 24 is formed such that the interval between the side wall plates 22 and 23 is narrower than the depth and width in the direction along the side wall plates 22 and 23. Each storage portion 24 is arranged side by side in a direction along each side wall plate 22, 23 and one side edge 21 a of the substrate 21.
図１（ｃ）に示すように、透明袋１２は、酸素および空気を通さない気密性の透明な合成樹脂の袋から成っている。透明袋１２は、矩形状を成し、周囲の三辺が閉じられて、残りの一辺が開口１２ａを成している。透明袋１２は、培養ケース１１の収納部２４の奥行きおよび幅と同じ長さおよび幅を有している。透明袋１２には、開口１２ａから嫌気性菌を検査するための培地を収納可能である。また、透明袋１２は、培養ケース１１の収納部２４の開口２４ａから収納部２４の内部に収納可能になっている。 As shown in FIG.1 (c), the transparent bag 12 consists of the bag of the airtight transparent synthetic resin which does not let oxygen and air pass. The transparent bag 12 has a rectangular shape, the three sides are closed, and the other side forms an opening 12a. The transparent bag 12 has the same length and width as the depth and width of the storage portion 24 of the culture case 11. The transparent bag 12 can store a medium for inspecting anaerobic bacteria from the opening 12a. The transparent bag 12 can be stored in the storage unit 24 through the opening 24 a of the storage unit 24 of the culture case 11.
嫌気性菌培養キット１０は、試料の中に嫌気性菌が存在するかどうかを調べるため、本発明の実施の形態の嫌気性菌検査方法で好適に使用される。まず、嫌気性菌を検査するための複数の試料を、それぞれ別の培地に添加して、混合する。培地は、大腸菌や硫酸塩還元細菌などの嫌気性菌を培養可能な寒天培地から成り、透明袋１２に入れるとき流動性を有している。各培地をそれぞれ別の透明袋１２に注入し、各透明袋１２を培養ケース１１の各収納部２４に開口２４ａから収納する。このとき、透明袋１２が、収納部２４の奥行きおよび幅と同じ長さおよび幅を有しているため、収納部２４への透明袋１２の収納が容易である。 The anaerobic bacteria culture kit 10 is suitably used in the anaerobic bacteria testing method of the embodiment of the present invention in order to check whether anaerobic bacteria are present in a sample. First, a plurality of samples for inspecting anaerobic bacteria are added to different media and mixed. The medium is composed of an agar medium capable of culturing anaerobic bacteria such as Escherichia coli and sulfate-reducing bacteria, and has fluidity when placed in the transparent bag 12. Each culture medium is poured into a separate transparent bag 12, and each transparent bag 12 is stored in each storage portion 24 of the culture case 11 from the opening 24 a. At this time, since the transparent bag 12 has the same length and width as the depth and width of the storage unit 24, the transparent bag 12 can be easily stored in the storage unit 24.
透明袋１２を収納部２４に収納したならば、空気が入らないように透明袋１２をヒートシール加工で密封する。このとき、空気が入らないようにするため、培地をシール部１２ｂから若干はみ出すようにして、ヒートシール加工してもよい。こうして、透明袋１２の中に酸素が入らないようにし、嫌気性菌の培養に適した条件にすることができる。培養ケース１１の収納部２４の各側壁板２２，２３の間に、培地を入れた透明袋１２を収納し、培地を各側壁板２２，２３の間隔と同じ厚みで、均一の厚みにすることができる。培地は固まる前には透明袋１２の内部で比較的自由に流動できるため、各側壁板２２，２３の間隔で均一の厚みに調整しやすい。培地は、透明袋１２の中で次第に固まる。透明袋１２は、両側の側壁板２２，２３に密着して保持される。 If the transparent bag 12 is stored in the storage unit 24, the transparent bag 12 is sealed by heat sealing so that air does not enter. At this time, in order to prevent air from entering, the medium may protrude slightly from the seal portion 12b and heat-sealed. In this way, oxygen can be prevented from entering the transparent bag 12, and conditions suitable for anaerobic bacteria culture can be achieved. The transparent bag 12 containing the culture medium is stored between the side wall plates 22 and 23 of the storage unit 24 of the culture case 11, and the culture medium has the same thickness as the interval between the side wall plates 22 and 23 and has a uniform thickness. Can do. Since the medium can flow relatively freely inside the transparent bag 12 before it hardens, it can be easily adjusted to a uniform thickness at the interval between the side wall plates 22 and 23. The medium gradually solidifies in the transparent bag 12. The transparent bag 12 is held in close contact with the side wall plates 22 and 23 on both sides.
透明袋１２は、各側壁板２２，２３の間に保持して収納されるため、位置が固定され、経時的に培地の同じ位置を観察することができる。透明袋１２の開口１２ａを上に向けて培地の注入および透明袋１２の密封を行うことにより、重力で培地が下方に下がり、各側壁板２２，２３に向かって押す力が働くため、培地を収納した透明袋１２と各側壁板２２，２３の内側面との間に働く摩擦力を高めることができる。このため、透明袋１２は、各側壁板２２，２３の間で位置がずれにくくなっている。 Since the transparent bag 12 is held and accommodated between the side wall plates 22 and 23, the position is fixed, and the same position of the culture medium can be observed over time. By injecting the culture medium with the opening 12a of the transparent bag 12 facing upward and sealing the transparent bag 12, the culture medium is lowered by gravity and a pressing force acts on the side wall plates 22 and 23. The frictional force acting between the stored transparent bag 12 and the inner side surfaces of the side wall plates 22 and 23 can be increased. For this reason, the position of the transparent bag 12 is difficult to shift between the side wall plates 22 and 23.
密封した透明袋１２を収納部２４に収納した状態で、培養ケース１１ごとインキュベータなどに入れ、嫌気性菌の所定の培養温度で保つ。所定の培養温度としては、至適培養温度が好ましい。所定時間ごとに、透明な各側壁板２２，２３を通して培地を観察する。これにより、嫌気性菌が試料中に存在しているかどうかを容易かつ正確に確認することができる。また、複数の試料を同時に検査することができる。嫌気性菌培養キット１０により、嫌気チャンバーや嫌気ジャーなどの大型の設備が不要となり、簡単な構成で、嫌気性菌の培養および検出を容易に行うことができる。 In a state where the sealed transparent bag 12 is stored in the storage unit 24, the culture case 11 is placed in an incubator or the like and kept at a predetermined culture temperature for anaerobic bacteria. As the predetermined culture temperature, an optimum culture temperature is preferable. The medium is observed through the transparent side wall plates 22 and 23 at predetermined time intervals. Thereby, it can be confirmed easily and correctly whether anaerobic bacteria exist in a sample. In addition, a plurality of samples can be inspected simultaneously. The anaerobic bacteria culture kit 10 eliminates the need for large equipment such as an anaerobic chamber and an anaerobic jar, and allows easy culture and detection of anaerobic bacteria with a simple configuration.
なお、透明袋１２を収納部２４に収納した後、培地を透明袋１２へ注入してもよい。また、透明袋１２の内部に嫌気性菌検査用の培地を入れて透明袋１２を密封した後、透明袋１２を開口２４ａから培養ケース１１の収納部２４に収納し、所定の培養温度で保ってもよい。この場合、透明袋１２への培地の密封および透明袋１２の収納部２４への収納の各操作が独立しており、容易である。 Note that the medium may be injected into the transparent bag 12 after the transparent bag 12 is stored in the storage unit 24. Further, after a medium for anaerobic bacteria test is put inside the transparent bag 12 and the transparent bag 12 is sealed, the transparent bag 12 is stored in the storage unit 24 of the culture case 11 through the opening 24a and kept at a predetermined culture temperature. May be. In this case, the operations of sealing the culture medium in the transparent bag 12 and storing the transparent bag 12 in the storage unit 24 are independent and easy.
図２に示すように、嫌気性菌が試料中に存在しているかどうかを確認するのに、本発明の実施の形態の嫌気性菌検査装置を使用することができる。嫌気性菌検査装置５０は、光源５１と画像取得手段５２とを有している。光源５１は、嫌気性菌培養キット１０の一方の側壁板２２の外側に設置され、側壁板２２，２３および収納部２４の透明袋１２に向けて光を照射し、光を透過させるようになっている。画像取得手段５２は、スキャナー装置またはデジタルカメラから成り、他方の側壁板２３の外側に設置されている。画像取得手段５２は、光源５１からの側壁板２２，２３、透明袋１２および培地を透過した光による投影像を取得するよう設けられている。嫌気性菌検査装置５０を用いて、画像取得手段５２により取得した培地のコロニーの投影像に基づき、嫌気性菌を正確かつ迅速に検出することができる。なお、嫌気性菌検査装置は、培養ケース１１を嫌気性菌の至適培養温度に保つインキュベータを備えていることが好ましい。 As shown in FIG. 2, the anaerobic bacteria testing apparatus according to the embodiment of the present invention can be used to check whether anaerobic bacteria are present in the sample. The anaerobic bacteria testing apparatus 50 includes a light source 51 and an image acquisition unit 52. The light source 51 is installed outside the one side wall plate 22 of the anaerobic bacteria culture kit 10, and irradiates light toward the side walls 22, 23 and the transparent bag 12 of the storage unit 24 to transmit light. ing. The image acquisition means 52 is composed of a scanner device or a digital camera, and is installed outside the other side wall plate 23. The image acquisition means 52 is provided so as to acquire a projection image by light transmitted through the side wall plates 22 and 23, the transparent bag 12 and the medium from the light source 51. Anaerobic bacteria can be detected accurately and rapidly based on the projection image of the colony of the medium acquired by the image acquisition means 52 using the anaerobic bacteria testing apparatus 50. In addition, it is preferable that the anaerobic bacteria inspection apparatus includes an incubator that maintains the culture case 11 at an optimum culture temperature for anaerobic bacteria.
図３に示すように、嫌気性菌検査装置５０は、各側壁板２２，２３の外側に、各側壁板２２，２３を挟む位置に設けられた１対の偏光板５３，５４を有していてもよい。この場合、１対の偏光板５３，５４を通した投影像を、画像取得手段５２で取得することにより、各側壁板２２，２３にかかる応力を光弾性効果により検出することができる。１対の偏光板５３，５４は、各側壁板２２，２３を挟むよう各側壁板２２，２３の外側に取り付けられていてもよい。 As shown in FIG. 3, the anaerobic bacteria testing apparatus 50 has a pair of polarizing plates 53 and 54 provided at positions sandwiching the side wall plates 22 and 23 outside the side wall plates 22 and 23. May be. In this case, by acquiring the projection image that has passed through the pair of polarizing plates 53 and 54 by the image acquisition means 52, the stress applied to the side wall plates 22 and 23 can be detected by the photoelastic effect. The pair of polarizing plates 53 and 54 may be attached to the outside of the side wall plates 22 and 23 so as to sandwich the side wall plates 22 and 23.
図４に示すように、培地の試料が嫌気性菌を含んでいると、培養された菌が増殖してガスを発生し、透明袋１２がそのガスで膨らんで側壁板２２，２３が外側に向かって押される。これにより、側壁板２２，２３の屈折率が変化するため、１対の偏光板５３，５４を通した投影像に、側壁板２２，２３の歪みに応じた干渉縞ができる。この干渉縞に基づいて、側壁板２２，２３にかかる応力を検出することができる。このため、ガスにより菌を直接観察できない場合であっても、菌によるガスの発生を検出することができ、嫌気性菌の存在を検出することができる。 As shown in FIG. 4, when the medium sample contains anaerobic bacteria, the cultured bacteria grow to generate gas, and the transparent bag 12 expands with the gas so that the side wall plates 22 and 23 are outward. It is pushed toward. Thereby, since the refractive index of the side wall plates 22 and 23 changes, an interference fringe corresponding to the distortion of the side wall plates 22 and 23 is formed on the projected image that passes through the pair of polarizing plates 53 and 54. Based on the interference fringes, the stress applied to the side wall plates 22 and 23 can be detected. For this reason, even if it is a case where bacteria cannot be observed directly with gas, generation of gas by bacteria can be detected and presence of anaerobic bacteria can be detected.
なお、初期段階から投影像に干渉縞が認められても、初期画像との比較によりキャンセルし、培養によって発生したガスによる変化だけを画像の変化として捉えることができる。寒天培地がくずれたりしてコロニーの検出が難しい場合にも、ガスを利用して検出することにより、嫌気性菌を検出することができる。なお、実験によれば、菌の培養によるガスの発生は、ＭＲＳ寒天培地で最も多く、TGC寒天培地、GAM寒天培地、標準寒天培地（SPC）の順に少なくなっている。 Even if an interference fringe is recognized in the projected image from the initial stage, it can be canceled by comparison with the initial image, and only the change due to the gas generated by the culture can be regarded as the image change. Even when it is difficult to detect colonies due to breakage of the agar medium, anaerobic bacteria can be detected by detection using gas. According to the experiment, the generation of gas due to the culture of the bacteria is the largest in the MRS agar medium, and decreases in the order of the TGC agar medium, the GAM agar medium, and the standard agar medium (SPC).
嫌気性菌培養キット１０および本発明の実施の形態の嫌気性菌の培養方法（以下、「パウチ法」）により、３種類の菌株を使用して培養および観察を行った。培養および観察は、嫌気性菌検査装置５０であるデジタル顕微鏡方式細菌検出装置（Digital Microscope Culturing System；製品名「Biomatic（登録商標） DMCS
FA-100（マイクロバイオ株式会社製）」）で行った。使用菌株は、サーモアナエロバクター・サーモハイドロスルフリカス（Thermoanaerobacter thermohydrosulfuricus；ATCC35045）、サーモアナエロバクター・マスラニ（Thermoanaerobacter mathranii；DSM11426）、モーレラ・サーモアセティカ（Moorella
thermoacetica）である。使用した培地は、クロストリジア寒天培地（CLO）である。培養温度は、６０℃または５５℃である。
The anaerobic bacterium culture kit 10 and the anaerobic bacterium culture method of the embodiment of the present invention (hereinafter referred to as “pouch method”) were used for culture and observation. Cultivation and observation are performed using an anaerobic bacteria testing apparatus 50, a digital microscopic culture system (product name “Biomatic (registered trademark) DMCS”).
FA-100 (manufactured by Microbio Inc.) ”). The strains used are Thermoanaerobacter thermohydrosulfuricus (ATCC35045), Thermoanaerobacter mathranii (DSM11426), Moorella Thermosetica (Moorella)
thermoacetica). The medium used is Clostridia agar (CLO). The culture temperature is 60 ° C or 55 ° C.
また、比較のために、寒天の平板培地を用いた一般的な寒天平板培地法（ｉＰＰＭ）により、同じ菌株を使用して培養および観察を行った。使用した培地は、標準寒天培地（SPC）、GAM寒天培地（GAM）、変法TGC寒天培地（mTGC）、クロストリジア寒天培地（CLO）である。使用装置、使用菌株および培養条件は、パウチ法と同じである。 For comparison, the same strain was used for culture and observation by a general agar plate medium method (iPPM) using an agar plate medium. The media used were standard agar medium (SPC), GAM agar medium (GAM), modified TGC agar medium (mTGC), and Clostridia agar medium (CLO). The device used, strain used, and culture conditions are the same as in the pouch method.
パウチ法による各菌株の培養後の観察結果を、図５乃至図７に示す。また、パウチ法および寒天平板培地法（ｉＰＰＭ）による培養後の観察結果をまとめ、表１に示す。培養後のコロニー数は、培地を透過した光による投影像に基づいて、自動的にカウントしている。 The observation results after culturing each strain by the pouch method are shown in FIGS. Moreover, the observation results after the culture by the pouch method and the agar plate medium method (iPPM) are summarized and shown in Table 1. The number of colonies after culturing is automatically counted based on a projection image of light transmitted through the medium.
表１に示すように、パウチ法は、寒天平板培地法（ｉＰＰＭ）のどの培地のものよりも、菌の検出速度が速いことが確認された。これは、寒天平板培地法（ｉＰＰＭ）に比べ、大気への暴露時間が短いためであると考えられる。また、パウチ法のクロストリジア培地（CLO）に比べ、寒天平板培地法（ｉＰＰＭ）のクロストリジア培地（CLO）では、細菌が産生する硫化水素が空間へ拡散するため、コロニーが黒色化しにくく、検出が遅れるということも考えられる。なお、パウチ法は、寒天平板培地法（ｉＰＰＭ）よりも操作が容易であることが確認できた。 As shown in Table 1, it was confirmed that the pouch method has a faster detection rate of bacteria than any medium of the agar plate medium method (iPPM). This is considered to be because the exposure time to the atmosphere is shorter than that of the agar plate medium method (iPPM). In addition, compared to the pouch-type Clostridia medium (CLO), the agar plate method (iPPM) Clostridia medium (CLO) diffuses hydrogen sulfide produced by bacteria into the space, so that colonies are less likely to be blackened and detection is delayed. It can be considered. The pouch method was confirmed to be easier to operate than the agar plate medium method (iPPM).
１０ 嫌気性菌培養キット
１１ 培養ケース
１２ 透明袋
１２ａ 開口
１２ｂ シール部
２１ 基板
２１ａ 一方の側縁
２１ｂ 他方の側縁
２１ｃ 凹部
２１ｄ 隔壁部
２１ｅ 表面
２１ｆ 裏面
２１ｇ 端部
２１ｈ 中央部
２２，２３ 側壁板
２４ 収納部
２４ａ 開口
５０ 嫌気性菌検査装置
５１ 光源
５２ 画像取得手段
５３，５４ 偏光板
DESCRIPTION OF SYMBOLS 10 Anaerobic bacteria culture kit 11 Culture case 12 Transparent bag 12a Opening 12b Seal part 21 Substrate 21a One side edge 21b The other side edge 21c Recessed part 21d Partition part 21e Surface 21f Back surface 21g End part 21h Central part 22,23 Side wall plate 24 Storage unit 24a Opening 50 Anaerobic bacteria inspection device 51 Light source 52 Image acquisition means 53, 54 Polarizing plate
Claims (6)
前記透明袋の収納部を有する培養ケースとを有し、
前記収納部は、前記透明袋を保持して収納するよう所定の間隔をあけて互いに対向する１対の透明な側壁板の間に形成され、前記透明袋を入れるための開口を有することを、
特徴とする嫌気性菌培養キット。 A transparent bag that contains the medium and is sealed;
A culture case having a storage part for the transparent bag,
The storage portion is formed between a pair of transparent side wall plates facing each other at a predetermined interval so as to hold and store the transparent bag, and has an opening for storing the transparent bag.
Anaerobic bacteria culture kit characterized.
各側壁板に向けて光を透過させるよう設けられた光源と、
前記光源からの前記側壁板、前記透明袋および前記培地を透過した光による投影像を取得可能に設けられた画像取得手段とを、
有することを特徴とする嫌気性菌検査装置。 An anaerobic bacteria culture kit according to claim 1 or 2,
A light source provided to transmit light toward each side wall plate;
Image acquisition means provided so as to be able to acquire a projection image by light transmitted through the side wall plate, the transparent bag and the medium from the light source;
An anaerobic bacteria testing apparatus characterized by having.
各側壁板を挟む位置に設けられ、前記側壁板にかかる応力を光弾性効果により検出する１対の偏光板と、
各偏光板および各側壁板に向けて光を透過させるよう設けられた光源と、
前記光源からの前記偏光板、前記側壁板、前記透明袋および前記培地を透過した光による投影像を取得可能に設けられた画像取得手段とを、
有することを特徴とする嫌気性菌検査装置。 An anaerobic bacteria culture kit according to claim 1;
A pair of polarizing plates provided at positions sandwiching each side wall plate, and detecting a stress applied to the side wall plate by a photoelastic effect;
A light source provided to transmit light toward each polarizing plate and each side wall plate;
Image acquisition means provided so as to be able to acquire a projection image by light transmitted through the polarizing plate, the side wall plate, the transparent bag and the medium from the light source;
An anaerobic bacteria testing apparatus characterized by having.
In the storage part of the anaerobic bacteria culture kit according to claim 1, the transparent bag in which the medium containing the sample for anaerobic bacteria test is stored and sealed is arranged and kept at a predetermined culture temperature of the anaerobic bacteria, A pair of polarizing plates is provided at a position sandwiching each side wall plate, irradiated with light that passes through the polarizing plate, the side wall plate, the transparent bag, and the culture medium, and applied to the side wall plate from a projected image by the transmitted light An anaerobic test method characterized by detecting stress by a photoelastic effect.
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