JP2006280219A - Selective culture medium for determining legionella, and method for culturing legionella bacterium - Google Patents
Selective culture medium for determining legionella, and method for culturing legionella bacterium Download PDFInfo
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Abstract
Description
本発明は、レジオネラ検査用選択培地に関する。 The present invention relates to a selective medium for Legionella examination.
レジオネラ属菌はレジオネラ症の原因微生物として知られており、自然界や人工の水環境に広く生息しているグラム陰性の細菌である。レジオネラ属菌に汚染された冷却塔水や温泉水、循環風呂水などのエアロゾルを吸い込むことによって、レジオネラ属菌に感染し、レジオネラ症を引き起こす場合がある。そのため、「新版 レジオネラ症防止指針」（非特許文献１）が定められ、これらの環境水中のレジオネラ属菌のコントロールを行うことが求められている。 Legionella is known as the causative microorganism of Legionella disease, and is a Gram-negative bacterium widely inhabiting the natural world and artificial water environment. Inhalation of aerosols such as cooling tower water, hot spring water, and circulating bath water contaminated with Legionella spp. May infect Legionella spp. And cause Legionellosis. Therefore, “new edition legionellosis prevention guideline” (Non-patent Document 1) is established, and it is required to control Legionella spp. In these environmental waters.
これら環境水からレジオネラ属菌を検出する方法にはさまざまな方法があるが、そのなかでも平板培養法はレジオネラ属菌検出の標準的方法として主要な方法である。ここで上記「新版 レジオネラ症防止指針」に記載された冷却遠心濃縮法を用いた平板培養法のフローの大略について図１に示す。 There are various methods for detecting Legionella spp. From these environmental waters. Among them, the plate culture method is the main method as a standard method for detecting Legionella spp. Here, FIG. 1 shows an outline of the flow of the plate culture method using the cooling centrifugal concentration method described in the above-mentioned “New Edition Legionellosis Prevention Guidelines”.
レジオネラ属菌を培養して検出する場合、レジオネラ属菌以外の微生物の影響を除くために試料の前処理を行なう。前処理方法としては検査対象水あるいは検査対象水の冷却遠心沈渣に滅菌水を加えたものに対し、ＨＣｌ・ＫＣｌ緩衝液を加える酸処理を行って、レジオネラ属菌以外の菌を殺菌する方法が広く用いられている。 When Legionella spp. Are cultured and detected, the sample is pretreated to remove the influence of microorganisms other than Legionella spp. As a pretreatment method, there is a method of sterilizing bacteria other than Legionella spp. By performing an acid treatment to which HCl / KCl buffer solution is added to water to be inspected or sterilized water added to a cooled centrifugal sediment of water to be inspected. Widely used.
しかしながら、このような酸処理によってもレジオネラ属菌以外の菌が残留することが多い。 However, bacteria other than Legionella spp. Often remain after such acid treatment.
これらレジオネラ属菌以外の菌（雑菌）は一般にレジオネラ属菌よりもコロニー形成及びその拡大が早いので、レジオネラ属菌コロニー計数の妨げとなり、また、それら雑菌によっては、近接するレジオネラ属菌コロニーの成長を妨げるなどの障害をもたらすため、レジオネラ属菌に対して障害を引き起こさず、かつ、レジオネラ属菌以外のこれら雑菌に対して有効な、抗菌剤を添加した培地を用いて、それらの繁殖を抑制する。 Since these bacteria (miscellaneous bacteria) other than Legionella spp. Generally colonize and expand faster than Legionella spp., They interfere with the count of Legionella spp., And depending on the bacteria, the growth of nearby Legionella spp. Inhibiting the growth of Legionella spp. By using an antibacterial medium that does not cause damage to Legionella spp. And is effective against these other bacteria than Legionella spp. To do.
このような抗菌剤を添加した培地として、ＧＶＰ培地が挙げられる。ＧＶＰ培地はレジオネラ属菌を培養可能な培地であるＢＣＹＥα培地に対しレジオネラ属菌に対して選択性を付与するために、抗菌剤である、グリシン、バンコマイシン及びポリミキシンＢが添加されたものであり、雑菌の影響が排除されている。 An example of a medium supplemented with such an antibacterial agent is GVP medium. The GVP medium is an antibacterial agent added with glycine, vancomycin and polymyxin B in order to give selectivity to Legionella bacteria over BCYEα medium, which is a medium capable of culturing Legionella bacteria. The effects of germs are eliminated.
しかし、ＧＶＰ培地は真菌に対しては対策が取られていないため、レジオネラ属菌がコロニーを形成するのに必要な５日程度の期間に培地に真菌が繁殖し、上記雑菌同様に、レジオネラ属菌の計数精度に大きな影響を及ぼす。 However, since the GVP medium does not take measures against fungi, the fungus propagates in the medium for about 5 days necessary for the Legionella genus to form colonies. It greatly affects the counting accuracy of bacteria.
そのため、ＢＣＹＥα培地にグリシン、バンコマイシン、及び、ポリミキシンＢに加え抗真菌剤であるアニソマイシンを添加したＭＷＹ培地、または、ＢＣＹＥα培地にグリシン、バンコマイシン、及び、ポリミキシンＢに加え、抗真菌剤であるシクロヘキシミドを添加したＧＶＰＣ培地、あるいは、ＢＣＹＥα培地にグリシン、バンコマイシン、及び、ポリミキシンＢに加え抗真菌剤であるアンホテリシンＢを添加したＷＹＯ培地などが、真菌の影響をある程度排除できる選択培地として知られている（特開昭６０−１９９３９８号公報（特許文献１））。 Therefore, in addition to glycine, vancomycin, and polymyxin B in addition to glycine, vancomycin, and polymyxin B, in addition to glycine, vancomycin, and polymyxin B, in addition to glycine, vancomycin, and polymyxin B, an antimycotic agent is added to BCYEα medium. Is known as a selective medium that can eliminate the effects of fungi to some extent, such as GVPC medium supplemented with glycine, vancomycin, and BMYE medium supplemented with antimycotic amphotericin B in addition to polymyxin B (Japanese Unexamined Patent Publication No. 60-199398 (Patent Document 1)).
しかしながら、真菌に対策が取られたこれら培地であっても、充分な効果が確実に得られるのではなく、これら抗真菌剤に対して耐性を有する糸状菌や酵母の繁殖により測定ができなくなったり、あるいは、レジオネラ属菌の計数精度に不安が生じるなどの問題が依然としてあった。
本発明は、上記した従来の問題点を改善する、すなわち、真菌とレジオネラ属菌とが併存する場合であっても、真菌による影響を受けずに信頼性が高いレジオネラ属菌検査を可能とするレジオネラ検査用選択培地を提供することを目的とする。 The present invention improves the above-described conventional problems, that is, even when fungi and Legionella coexist, it enables a highly reliable Legionella test without being affected by the fungus. An object is to provide a selective medium for Legionella examination.
本発明のレジオネラ検査用選択培地は上記課題を解決するため、請求項１に記載の通り、レジオネラ属菌を培養可能な培地であって、シクロヘキシミド、アンホテリシンＢ、及び、チアベンダゾールを含有することを特徴とするレジオネラ検査用選択培地である。 In order to solve the above-mentioned problem, the selective medium for Legionella test of the present invention is a medium capable of culturing Legionella spp., And contains cycloheximide, amphotericin B, and thiabendazole. It is a selective medium for Legionella inspection.
本発明のレジオネラ検査用選択培地は請求項２に記載の通り、請求項１に記載のレジオネラ検査用選択培地において、ＧＶＰ培地に、シクロヘキシミド、アンホテリシンＢ、及び、チアベンダゾールを含有させたことを特徴とするレジオネラ検査用選択培地である。
The selective medium for Legionella test according to the present invention is characterized in that, as described in Claim 2, the selective medium for Legionella test according to
本発明のレジオネラ属菌の培養方法は、請求項４に記載の通り、レジオネラ属菌を培養可能な培地であって、シクロヘキシミド、アンホテリシンＢ、及び、チアベンダゾールを含有する培地を用いて培養するレジオネラ属菌の培養方法である。 The method of culturing Legionella according to the present invention is a culture medium capable of culturing Legionella as described in claim 4, wherein the culture is performed using a medium containing cycloheximide, amphotericin B, and thiabendazole. This is a method for culturing fungi.
本発明のレジオネラ検査用選択培地によれば、細菌の影響の排除は勿論、真菌による影響も極めて少なく、信頼性が高いレジオネラ属菌検査が可能となる。 According to the selective medium for Legionella test of the present invention, not only the influence of bacteria is excluded, but also the influence of fungi is extremely small, and a highly reliable Legionella bacterium test can be performed.
本発明において、レジオネラ属菌を培養可能な培地とは、レジオネラ属菌の培養に必要な成分を、レジオネラ属菌の培養に適した濃度で有し、かつ、その培地によりレジオネラ属菌のコロニーが形成可能な培地を云う。このようなレジオネラ属菌を培養可能な培地としては例えば、ＢＣＹＥα寒天培地が知られている。代表的な組成例（新版レジオネラ症防止指針記載のもの。必要により使用者により各成分の配合量は適宜変更される場合がある）を表１に示す。 In the present invention, the medium capable of cultivating Legionella spp. Has components necessary for culturing Legionella sp. At a concentration suitable for culturing Legionella spp., And colonies of Legionella spp. A medium that can be formed. As a medium capable of culturing such Legionella spp., For example, a BCYEα agar medium is known. Table 1 shows typical composition examples (as described in the new edition of Legionellosis prevention guideline. The amount of each component may be appropriately changed by the user if necessary).
しかし、ＢＣＹＥα寒天培地には選択性がないので、レジオネラ属菌と他の細菌（一般にレジオネラ属菌より増殖が早い。「雑菌」と云う）とが併存する場合には、レジオネラ属菌のコロニー計数時の妨げとなったり、あるいはレジオネラ属菌のコロニー形成が阻害される。このため、本発明ではこれら雑菌を抑制する抗菌剤を上記培地に配合することもできる。このような抗菌剤としてグリシン、バンコマイシン及びポリミキシンＢの３種の抗菌剤の組合せが適している。 However, since there is no selectivity in BCYEα agar medium, if Legionella spp. And other bacteria (generally faster growth than Legionella spp., Called “miscellaneous bacteria”) coexist, colony count of Legionella spp. Time hindrance, or Legionella colonization is inhibited. For this reason, in this invention, the antibacterial agent which suppresses these miscellaneous bacteria can also be mix | blended with the said culture medium. As such an antibacterial agent, a combination of three antibacterial agents, glycine, vancomycin and polymyxin B, is suitable.
ここで、グリシン、バンコマイシン及びポリミキシンＢが含有された、レジオネラ属菌を培養可能な培地として、ＧＶＰ培地が知られている。ＧＶＰ培地は表１に示した組成で代表されるようなＢＣＹＥα寒天培地にこれらグリシン、バンコマイシン及びポリミキシンＢを加えたものであり、その代表的な組成はＢＣＹＥα寒天培地の各成分にグリシン３ｇ、バンコマイシン５ｍｇ及びポリミキシンＢ１０００００ＩＵを添加して計１Ｌ（ｐＨ：６．９±０．１）となるようにしたものである（新版レジオネラ症防止指針記載のもの。必要により使用者により各成分の配合量は適宜変更される場合があるが、その場合も本発明におけるＧＶＰ培地に含まれる）。 Here, a GVP medium is known as a medium containing glycine, vancomycin and polymyxin B and capable of culturing Legionella spp. The GVP medium is obtained by adding these glycine, vancomycin and polymyxin B to a BCYEα agar medium represented by the composition shown in Table 1, and a typical composition thereof is 3 g of glycine and vancomycin for each component of the BCYEα agar medium. 5 mg and polymyxin B 100000 IU added to a total of 1 L (pH: 6.9 ± 0.1) (as described in the new edition of Legionellosis prevention guideline. If necessary, the amount of each component is determined by the user) Although it may be changed as appropriate, it is also included in the GVP medium in the present invention).
本発明のレジオネラ検査用選択培地には、上記のレジオネラ属菌を培養可能な培地に、真菌（糸状菌や酵母）に対して有効な抗真菌剤としてシクロヘキシミド、アンホテリシンＢ、及び、チアベンダゾールを配合する必要がある。これら３成分の配合により、真菌による障害を極めて効果的に防止することができ、レジオネラ属菌の高精度で確実な検出が可能となる。 In the selective medium for Legionella test of the present invention, cycloheximide, amphotericin B, and thiabendazole are blended as an antifungal agent effective against fungi (filamentous fungi and yeast) in a culture medium capable of culturing the above Legionella spp. There is a need. By combining these three components, fungal damage can be prevented very effectively, and Legionella spp. Can be accurately detected with high accuracy.
これら、シクロヘキシミド、アンホテリシンＢ、及び、チアベンダゾールの含有量は、培地１Ｌあたりそれぞれ、４０ｍｇ以上４００ｍｇ以下、４０ｍｇ以上４００ｍｇ以下、及び、２０ｍｇ以上２００ｍｇ以下であるとレジオネラ属菌の培養を阻害せず、同時に、真菌による障害をより効果的に防止することができ、レジオネラ属菌のより高精度な検出が可能となる。より好ましいチアベンダゾールの添加量は２０〜８０ｍｇ／Ｌである。 The content of these cycloheximide, amphotericin B and thiabendazole is 40 mg or more, 400 mg or less, 40 mg or more and 400 mg or less, and 20 mg or more and 200 mg or less per liter of the medium respectively, without inhibiting the culture of Legionella sp. In addition, it is possible to more effectively prevent fungal damage and to detect Legionella spp. With higher accuracy. A more preferable addition amount of thiabendazole is 20 to 80 mg / L.
以下に本発明のレジオネラ検査用選択培地の実施例について具体的に説明する。 Examples of the selective medium for Legionella test of the present invention will be specifically described below.
＜レジオネラ属菌に対する各抗真菌剤の最小発育阻止濃度の検討＞
抗真菌剤として、シクロヘキシミド（略号：ＣＨ）、アンホテリシンＢ（略号：ＡＭＰＨ）及び、チアベンダゾール（略号：ＴＢＺ）について、レジオネラ属菌に対する最小発育阻止濃度の検討を行った。
<Examination of minimum inhibitory concentration of each antifungal agent against Legionella spp.>
As antifungal agents, cycloheximide (abbreviation: CH), amphotericin B (abbreviation: AMPH) and thiabendazole (abbreviation: TBZ) were examined for minimum growth inhibitory concentrations against Legionella spp.
レジオネラの各菌株（表２参照）はあらかじめＢＹＥ液体培地（表３参照。ＢＣＹＥα寒天培地から、寒天、活性炭を除いたもの）を用い、３７℃の温度下で、２４時間培養した。各抗真菌剤をジメチルスルホキシド（ＤＭＳＯ）に４０ｍｇ／ｍＬとなるように溶解させ、所定濃度まで２倍の希釈列を作製した。４８穴マイクロプレートに溶解・希釈した各抗真菌剤および陽性コントロールとしてＤＭＳＯをそれぞれ１０μＬずつ添加した。レジオネラの培養液を新しいＢＹＥ液体培地で１００倍に希釈し、上記４８穴マイクロプレートの各ウェルに９９０μＬずつ加えていく（このときＤＭＳＯの濃度は１％となりレジオネラの培養に影響がないものとなる）。次いで、３７℃のインキュベーター内に３日間静置して培養する。その後、各ウェルのレジオネラ属菌の増殖による濁度上昇が目視で認められない最小濃度を求めた。結果を表２に併せて示す（表中”＞４００”は４００μｇ／ｍＬの添加でも発育阻止が見られなかったことを示す）。 Each strain of Legionella (see Table 2) was cultured in advance using a BYE liquid medium (see Table 3. BCYEα agar medium excluding agar and activated carbon) at a temperature of 37 ° C. for 24 hours. Each antifungal agent was dissolved in dimethyl sulfoxide (DMSO) to a concentration of 40 mg / mL, and a 2-fold dilution series was prepared to a predetermined concentration. 10 μL of each antifungal agent dissolved and diluted in a 48-well microplate and DMSO as a positive control were added. The Legionella culture solution is diluted 100-fold with fresh BYE liquid medium, and 990 μL is added to each well of the 48-well microplate (at this time, the concentration of DMSO is 1% and there is no effect on Legionella culture). ). Subsequently, it culture | cultivates by leaving still for 3 days in a 37 degreeC incubator. Thereafter, the minimum concentration at which no increase in turbidity due to growth of Legionella in each well was visually observed was determined. The results are shown in Table 2 (“> 400” in the table indicates that growth inhibition was not observed even when 400 μg / mL was added).
表２よりシクロヘキシミド、アンホテリシンＢは、４００ｍｇ／Ｌ以下の濃度で、チアベンダゾールは１００〜２００ｍｇ／Ｌ未満の濃度でレジオネラ属菌の生育に影響を与えないことが確認された。 From Table 2, it was confirmed that cycloheximide and amphotericin B did not affect the growth of Legionella at concentrations of 400 mg / L or less and thiabendazole at concentrations of 100 to less than 200 mg / L.
＜各選択培地の抗真菌効果＞
それぞれ異なるビルの冷却塔水から採取したサンプル水を真菌培養用の一般的な培地であるＰＤＡ培地に接種したときに発育した糸状菌Ｍ−１〜Ｍ−４、それぞれ異なるビルの冷却塔水から採取したサンプル水をＧＶＰＣ培地（抗真菌剤としてシクロヘキシミドが配合された培地）に接種したときに発育した糸状菌Ｍ−５〜Ｍ−１０（これらはシクロヘキシミドに対して耐性を有する）を準備した。
<Antifungal effect of each selective medium>
Filamentous fungi M-1 to M-4 grown when inoculating PDA medium, which is a general medium for fungal culture, with sample water collected from cooling tower water of different buildings, respectively, from cooling tower water of different buildings Filamentous fungi M-5 to M-10 (which are resistant to cycloheximide) were prepared when the collected sample water was inoculated into GVPC medium (medium containing cycloheximide as an antifungal agent).
また培地としては「新版レジオネラ症防止指針」に記載された培地、及び、それら培地にさらに成分を添加したいくつかの培地を準備した。 In addition, as the culture medium, the culture medium described in “New Edition Legionellosis Prevention Guidelines” and several culture media in which components were further added to these culture media were prepared.
すなわち、
・ＧＶＰ培地：ＢＣＹＥα培地を基礎培地とし、これにグリシンを３ｍｇ／ｍＬ、バンコマイシンを５μｇ／ｍＬ、ポリミキシンＢを１００ＩＵ／ｍＬを添加した培地（表４に組成を示す）、
・ＧＶＰＣ培地：上記ＧＶＰ培地を基礎培地とし、シクロヘキシミドを８０μｇ／ｍＬ添加した培地、
・ＧＶＰＡ培地：上記ＧＶＰ培地を基礎培地とし、アンホテリシンＢを８０μｇ／ｍＬ添加した培地（ＷＹＯα培地）、
・ＧＶＰＣＡ培地：上記ＧＶＰ培地を基礎培地とし、シクロヘキシミドを８０μｇ／ｍＬ、アンホテリシンＢを８０μｇ／ｍＬそれぞれ添加した培地、
・ＧＶＰＴ培地：上記ＧＶＰ培地を基礎培地とし、チアベンダゾールを４０μｇ／ｍＬ添加した培地、
・ＣＡＴ培地：上記ＧＶＰ培地を基礎培地とし、シクロヘキシミドを８０μｇ／ｍＬ、アンホテリシンＢを８０μｇ／ｍＬ、チアベンダゾールを４０μｇ／ｍＬそれぞれ添加した培地（本発明実施例：培地１Ｌ当たりの組成を表５に示す。）、
の６種類の培地を準備した。
That is,
GVP medium: BCYEα medium as a basal medium, to which glycine was added at 3 mg / mL, vancomycin at 5 μg / mL, and polymyxin B at 100 IU / mL (the composition is shown in Table 4),
GVPC medium: a medium supplemented with 80 μg / mL of cycloheximide, using the above GVP medium as a basal medium,
GVPA medium: a medium (WYOα medium) supplemented with 80 μg / mL amphotericin B using the GVP medium as a basal medium,
GVPCA medium: a medium supplemented with 80 μg / mL of cycloheximide and 80 μg / mL of amphotericin B using the above GVP medium as a basal medium,
GVPT medium: a medium supplemented with 40 μg / mL thiabendazole using the above GVP medium as a basal medium,
CAT medium: medium containing the above GVP medium as a basal medium, cycloheximide 80 μg / mL, amphotericin B 80 μg / mL, and thiabendazole 40 μg / mL (Example of the present invention: composition per liter of medium is shown in Table 5) ),
6 types of media were prepared.
実験は次のように行った。上記Ｍ−１〜Ｍ−１０の糸状菌を滅菌脱イオン水に懸濁させ、これらそれぞれに、レジオネラ ニューモフィラ（Legionella pneumophila）ＡＴＣＣ３３１５２株を別の滅菌脱イオン水に懸濁させた液を混合して、上記の選択培地に接種した。その後３７℃で５日間培養して、各培地の糸状菌の発育状況を観察した。抗真菌効果検討結果を表６に示す（表６中”＋＋＋”は糸状菌が培地に広く発育したことを、”＋＋”は”＋＋＋”よりは小さく発育したことを、”＋”は”＋＋”より小さく殆ど発育しなかったことを、”−”は発育が見られなかったことをそれぞれ示す）。 The experiment was performed as follows. The M-1 to M-10 filamentous fungi are suspended in sterilized deionized water, and a solution obtained by suspending Legionella pneumophila ATCC33152 in another sterilized deionized water is mixed with each. And inoculated into the above selective medium. Thereafter, the cells were cultured at 37 ° C. for 5 days, and the growth of filamentous fungi in each medium was observed. The antifungal effect examination results are shown in Table 6 (in Table 6, “++++” indicates that the filamentous fungus has grown widely in the medium, “++” indicates that it has grown smaller than “++++”, and “+” indicates “++”. “It is smaller and hardly developed, and“-”indicates that no growth was observed).
また同時に、各培地でのレジオネラ検出状況（レジオネラ ニューモフィラのコロニー数）を調べた。このときブランクとしてレジオネラ属菌は添加したものの糸状菌を添加しなかった系についても併せて調べた。これら結果を表７に示す（表７中、”検出不能”は、糸状菌の生育によりレジオネラのコロニーが計数不能であったことを示す）。 At the same time, the detection status of Legionella in each medium (the number of Legionella pneumophila colonies) was examined. At this time, a system in which Legionella was added as a blank but no filamentous fungus was added was also examined. These results are shown in Table 7 (in Table 7, “Undetectable” indicates that Legionella colonies could not be counted due to the growth of filamentous fungi).
表６及び表７より本発明に係るレジオネラ検査用選択培地によれば、糸状菌の生育を効果的にかつ、確実に防止することができ、そのとき、レジオネラ属菌生菌数を正確に調べることができることが判る。 From Table 6 and Table 7, according to the selective medium for Legionella test according to the present invention, the growth of filamentous fungi can be effectively and reliably prevented, and at that time, the number of viable Legionella bacteria is accurately examined. I can see that
なお、別途表５における本発明に係る培地と同様の組成で、ただし、培地１Ｌ当たり、チアベンダゾールの添加量を２００ｍｇとした培地（実施例）についても、別途、表５の組成の本発明の培地同様に検討を行ったが、そのときの結果ではレジオネラ属菌のコロニー形成に若干時間がかかったものの、表５の組成の培地での結果との間に有意差が認められなかった。このことからチアベンダゾールを２００ｍｇ／Ｌ添加した培地でも、レジオネラ検査用選択培地として使用可能であることが判った。 In addition, it is the composition similar to the culture medium which concerns on this invention in Table 5 separately, However, The culture medium of this invention of the composition of Table 5 separately also about the culture medium (Example) which added 200 mg of thiabendazole per 1L of culture media. Although examination was conducted in the same manner, the results at that time took some time for colonization of Legionella spp., But no significant difference was found between the results in the medium having the composition shown in Table 5. From this, it was found that even a medium supplemented with 200 mg / L of thiabendazole can be used as a selective medium for Legionella test.
本発明によれば真菌の生育を効果的にかつ、確実に防止することができ、その結果、真菌とレジオネラ属菌とが併存する場合であっても、レジオネラ属菌生菌数を正確に調べることができるため、再検査等が不要となり、例えば浴槽水など、レジオネラ属菌の有無が短期間で確実に検出される必要がある用途にも対応できる。 According to the present invention, fungal growth can be effectively and reliably prevented, and as a result, even when the fungus and Legionella spp. Coexist, the number of Legionella spp. Therefore, re-examination or the like is unnecessary, and for example, it is possible to cope with applications such as bath water where presence or absence of Legionella spp.
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| WO2011151793A1 (en) * | 2010-06-03 | 2011-12-08 | Basf Se | Detection and enumeration of microorganisms |
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| JP2021108573A (en) * | 2020-01-09 | 2021-08-02 | 栄研化学株式会社 | Coloring culture medium for differentiation of legionella bacteria |
| JP2021533802A (en) * | 2018-08-24 | 2021-12-09 | セクレタリー オブ ステート フォー ヘルス アンド ソーシャル ケアSecretary Of State For Health And Social Care | Charcoal-free medium for Legionella |
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| JP2011516052A (en) * | 2008-04-04 | 2011-05-26 | ビーエーエスエフ ソシエタス・ヨーロピア | Microbe detection and counting |
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| WO2011151793A1 (en) * | 2010-06-03 | 2011-12-08 | Basf Se | Detection and enumeration of microorganisms |
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| US9057093B2 (en) | 2010-06-03 | 2015-06-16 | Basf Se | Detection and enumeration of microorganisms |
| JP2019058088A (en) * | 2017-09-25 | 2019-04-18 | アクアス株式会社 | Method of examination of legionella bacteria |
| JP7009137B2 (en) | 2017-09-25 | 2022-01-25 | アクアス株式会社 | How to test for Legionella spp. |
| JP2021533802A (en) * | 2018-08-24 | 2021-12-09 | セクレタリー オブ ステート フォー ヘルス アンド ソーシャル ケアSecretary Of State For Health And Social Care | Charcoal-free medium for Legionella |
| JP7401526B2 (en) | 2018-08-24 | 2023-12-19 | セクレタリー オブ ステート フォー ヘルス アンド ソーシャル ケア | Charcoal-free medium for Legionella |
| JP2021108573A (en) * | 2020-01-09 | 2021-08-02 | 栄研化学株式会社 | Coloring culture medium for differentiation of legionella bacteria |
| JP7503384B2 (en) | 2020-01-09 | 2024-06-20 | 栄研化学株式会社 | Chromogenic medium for Legionella species identification |
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