JP2015500016A - ロイテリン産生ラクトバチルス・ブレビス - Google Patents
ロイテリン産生ラクトバチルス・ブレビス Download PDFInfo
- Publication number
- JP2015500016A JP2015500016A JP2014543986A JP2014543986A JP2015500016A JP 2015500016 A JP2015500016 A JP 2015500016A JP 2014543986 A JP2014543986 A JP 2014543986A JP 2014543986 A JP2014543986 A JP 2014543986A JP 2015500016 A JP2015500016 A JP 2015500016A
- Authority
- JP
- Japan
- Prior art keywords
- strain
- pylori
- reuterin
- brevis
- lactobacillus brevis
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
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- A—HUMAN NECESSITIES
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- A—HUMAN NECESSITIES
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- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
- A23C9/1234—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt characterised by using a Lactobacillus sp. other than Lactobacillus Bulgaricus, including Bificlobacterium sp.
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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Abstract
Description
形態:小鎖で集団化する均一な(homogen)着色を有する桿菌;
代謝:ヘテロ型発酵性(heterofermentative);
以下の糖の発酵(API MRS培地、37℃、48時間のApi 50 CHストリップ上で得られる結果):L−アラビノース、D−リボース、D−キシロース、D−グルコース、D−フルクトース、D−マルトース、D−メリビオース、グルコン酸カリウム、5−ケトグルコン酸(正:5-Ketogluconate)カリウム。
上で定義した本発明のL.ブレビス株、好ましくはCNCM I−4431株をグリセロールの存在下で培養する工程と、
培養によって産生されたロイテリンを回収する工程と、
を含む、方法である。
材料及び方法
ラクトバチルス・ブレビス株:
L.ブレビスCNCM I−4431
L.ブレビス80:従来のL.ブレビス株
3つのH.ピロリ参照株及び5つの臨床分離株を使用した。
26695:TOMBS et al. (Nature 388, 539-547, 1997)に記載;
HPAG1:OH et al. (Proc Natl Acad Sci U S A., 103, 9999-10004, 2006)に記載;
TN2GF4:日本で胃潰瘍患者から単離され、スナネズミに適合させた株(WATANABE, et al. Gastroenterol 1998; vol 115, pp642-648に記載の株)
CR113及びCR114:ヒト前がん胃病変生検材料から単離された株;
Axcan 342及びAxcan 374:それぞれ、H.ピロリの除菌のための4剤併用療法を試験する臨床研究に参加した胃炎患者及び食道炎患者から単離された株(MALFERTHEINER et al., Lancet, 377, 905-913, 2011)。
GC3C:胃がん患者から単離された株。
L.ブレビス株(正:strains)を、MRS Broth(pH6.2)中で17時間成長させた。細菌懸濁液をKOH溶液でpH6.8に中和した。
結果を下記表1に示す。
材料及び方法
ロイテリン産生に関与する酵素であるグリセロールデヒドラターゼの大サブユニット(GldC)をコードする遺伝子のL.ブレビスCNCM I−4431における存在を、非特許文献2(上掲)に記載の方法に従って試験した。
GDCRev:GCRGCYTTSATATCTKSAACCAT(配列番号1)及び
GDCFor:GCMTAYGCWGAAACCATTTCAGTTTA(配列番号2)
(どちらも非特許文献2に記載されている)。増幅されたgldCフラグメントの期待サイズは728bpである。
16SFor:ACTCCTACGGGAGGCAGCAG(配列番号3)及び
16SRev:GTATTACCGCGGCTGCTGGCAC(配列番号4)
(どちらも非特許文献2に記載されている)。
PCR反応の結果を図1に示す。L.ブレビスCNCM I−4431株、L.ブレビス80株及び(C+)ロイテリン産生L.ロイテリATCC 55730株は期待される728bp産物について陽性であった。したがって、L.ブレビスCNCM I−4431株は、ロイテリン産生を可能にするgldC遺伝子を有する。
L.ブレビス及びH.ピロリの培養
L.ブレビスCNCM I−4431株を実施例1に開示のようにMRS Broth中で成長させた。
野生型C.エレガンス株(N2)を用いて感染アッセイを行った。H.ピロリのNCTC11637株を線虫感染に使用した。実験は、同調幼若成虫(3日齢の線虫)を以下の異なる培養条件に移すことによって開始した:
NG培地+E.コリOP50(対照)
NG培地+E.コリOP50+100μLのLAB培養培地(対照培養培地)
NG培地+E.コリOP50+100μLの(H.ピロリNCTC 11637T+LAB)。
本研究に使用するプライマーはvacA遺伝子を対象とする(NAYAK & ROSE、J Appl Microbiol, 103, 1931-41, 2007)。プライマーはHpylF(5’ CAA TAG CAA TCA AGT GGC TTT G 3’、配列番号5)及びHpylR(5’ GCG CGC TTC CAC ATT AGC 3’、配列番号6)と名付けられている。特異性は既にBLASTオンラインツールによってコンピューターで、更にはH.ピロリ種の株、種々のプロバイオティクス株及びC.エレガンスの採餌に使用するE.コリOP50を用いてQ−PCRによってin vitroで確認されている(以前の最適化の報告を参照されたい)。「SYBR(商標)Green PCR Master Mix」(Applied Biosystem)及びStepOne Real−Timeサーモサイクラー(Applied Biosystem)を使用した。下記表2にH.ピロリのQ−PCR反応の最適化条件をまとめる。
ゲノム数=DNA(g)×NA/Mm
H.ピロリのゲノムサイズ=1.6Mpb
NA=6.023×1023
ゲノム数=DNA(g)×6.023×1023/1.6×106×2×309
Claims (8)
- ラクトバチルス・ブレビスCNCM I−4431株、
CNCM I−4431株の突然変異誘発又は遺伝子形質転換によって得られるラクトバチルス・ブレビス株、
から選択されるロイテリン蓄積ラクトバチルス・ブレビス株。 - ロイテリン産生への請求項1に記載のラクトバチルス・ブレビス株の使用。
- 請求項1に記載のラクトバチルス株を含む組成物。
- 乾燥重量1g当たり少なくとも105cfu、好ましくは少なくとも106cfuの前記ラクトバチルス・ブレビス株を含むことを特徴とする、請求項3に記載の組成物。
- 栄養組成物であることを特徴とする、請求項3又は4に記載の組成物。
- 乳製品であることを特徴とする、請求項5に記載の組成物。
- 薬剤として使用される、請求項1に記載のラクトバチルス・ブレビス株又は請求項3〜6のいずれか一項に記載の組成物。
- 前記薬剤がH.ピロリ感染を治療又は予防することを特徴とする、請求項7に記載の薬剤として使用されるラクトバチルス・ブレビス株又は組成物。
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JP2021516054A (ja) * | 2018-03-05 | 2021-07-01 | エムディー ヘルスケア インコーポレイテッドMd Healthcare Inc. | ラクトバチルス属細菌由来のナノ小胞及びその用途 |
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