JPH0265798A - Method for detecting salmonellal bacteria, culture medium and detecting paper for salmonellal bacteria used therefor - Google Patents
Method for detecting salmonellal bacteria, culture medium and detecting paper for salmonellal bacteria used thereforInfo
- Publication number
- JPH0265798A JPH0265798A JP21581488A JP21581488A JPH0265798A JP H0265798 A JPH0265798 A JP H0265798A JP 21581488 A JP21581488 A JP 21581488A JP 21581488 A JP21581488 A JP 21581488A JP H0265798 A JPH0265798 A JP H0265798A
- Authority
- JP
- Japan
- Prior art keywords
- salmonella
- bacteria
- medium
- sodium
- salmonellal
- 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
- 239000001963 growth medium Substances 0.000 title claims abstract description 12
- 238000000034 method Methods 0.000 title claims description 17
- 241000894006 Bacteria Species 0.000 title abstract description 33
- 238000001514 detection method Methods 0.000 claims abstract description 59
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 claims abstract description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 15
- 229930195725 Mannitol Natural products 0.000 claims abstract description 14
- 239000000594 mannitol Substances 0.000 claims abstract description 14
- 235000010355 mannitol Nutrition 0.000 claims abstract description 14
- 239000001888 Peptone Substances 0.000 claims abstract description 13
- 108010080698 Peptones Proteins 0.000 claims abstract description 13
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims abstract description 13
- 235000019319 peptone Nutrition 0.000 claims abstract description 13
- 229910000365 copper sulfate Inorganic materials 0.000 claims abstract description 12
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 claims abstract description 12
- 239000000284 extract Substances 0.000 claims abstract description 9
- 229940046892 lead acetate Drugs 0.000 claims abstract description 9
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 claims abstract description 8
- BVTBRVFYZUCAKH-UHFFFAOYSA-L disodium selenite Chemical compound [Na+].[Na+].[O-][Se]([O-])=O BVTBRVFYZUCAKH-UHFFFAOYSA-L 0.000 claims abstract description 8
- 239000011781 sodium selenite Substances 0.000 claims abstract description 8
- 229960001471 sodium selenite Drugs 0.000 claims abstract description 8
- 235000015921 sodium selenite Nutrition 0.000 claims abstract description 8
- 229940041514 candida albicans extract Drugs 0.000 claims abstract description 7
- 238000012258 culturing Methods 0.000 claims abstract description 7
- 235000002639 sodium chloride Nutrition 0.000 claims abstract description 7
- 239000012138 yeast extract Substances 0.000 claims abstract description 7
- 239000004313 iron ammonium citrate Substances 0.000 claims abstract description 6
- 235000000011 iron ammonium citrate Nutrition 0.000 claims abstract description 6
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims abstract description 6
- 235000019796 monopotassium phosphate Nutrition 0.000 claims abstract description 6
- 239000011780 sodium chloride Substances 0.000 claims abstract description 6
- BVHLGVCQOALMSV-JEDNCBNOSA-N L-lysine hydrochloride Chemical compound Cl.NCCCC[C@H](N)C(O)=O BVHLGVCQOALMSV-JEDNCBNOSA-N 0.000 claims abstract description 5
- 108010009004 proteose-peptone Proteins 0.000 claims abstract description 5
- AKHNMLFCWUSKQB-UHFFFAOYSA-L sodium thiosulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=S AKHNMLFCWUSKQB-UHFFFAOYSA-L 0.000 claims abstract description 5
- 235000019345 sodium thiosulphate Nutrition 0.000 claims abstract description 5
- 241000607142 Salmonella Species 0.000 claims description 88
- 239000002609 medium Substances 0.000 claims description 63
- 239000007788 liquid Substances 0.000 claims description 16
- 239000003833 bile salt Substances 0.000 claims description 9
- 239000004615 ingredient Substances 0.000 claims description 9
- 229940082569 selenite Drugs 0.000 claims description 9
- MCAHWIHFGHIESP-UHFFFAOYSA-L selenite(2-) Chemical compound [O-][Se]([O-])=O MCAHWIHFGHIESP-UHFFFAOYSA-L 0.000 claims description 9
- GLMQHZPGHAPYIO-UHFFFAOYSA-L azanium;2-hydroxypropane-1,2,3-tricarboxylate;iron(2+) Chemical compound [NH4+].[Fe+2].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O GLMQHZPGHAPYIO-UHFFFAOYSA-L 0.000 claims description 5
- 229940093761 bile salts Drugs 0.000 claims description 5
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 5
- 229960001506 brilliant green Drugs 0.000 claims description 4
- HXCILVUBKWANLN-UHFFFAOYSA-N brilliant green cation Chemical compound C1=CC(N(CC)CC)=CC=C1C(C=1C=CC=CC=1)=C1C=CC(=[N+](CC)CC)C=C1 HXCILVUBKWANLN-UHFFFAOYSA-N 0.000 claims description 4
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 claims description 3
- 239000011734 sodium Substances 0.000 claims description 3
- 229910052708 sodium Inorganic materials 0.000 claims description 3
- 239000003613 bile acid Substances 0.000 claims 1
- 150000003839 salts Chemical class 0.000 claims 1
- 235000013305 food Nutrition 0.000 abstract description 13
- 238000009630 liquid culture Methods 0.000 abstract 3
- HSINOMROUCMIEA-FGVHQWLLSA-N (2s,4r)-4-[(3r,5s,6r,7r,8s,9s,10s,13r,14s,17r)-6-ethyl-3,7-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1h-cyclopenta[a]phenanthren-17-yl]-2-methylpentanoic acid Chemical class C([C@@]12C)C[C@@H](O)C[C@H]1[C@@H](CC)[C@@H](O)[C@@H]1[C@@H]2CC[C@]2(C)[C@@H]([C@H](C)C[C@H](C)C(O)=O)CC[C@H]21 HSINOMROUCMIEA-FGVHQWLLSA-N 0.000 abstract 1
- 239000007836 KH2PO4 Substances 0.000 abstract 1
- FRHBOQMZUOWXQL-UHFFFAOYSA-L ammonium ferric citrate Chemical compound [NH4+].[Fe+3].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O FRHBOQMZUOWXQL-UHFFFAOYSA-L 0.000 abstract 1
- 235000019800 disodium phosphate Nutrition 0.000 abstract 1
- 229910000397 disodium phosphate Inorganic materials 0.000 abstract 1
- 229960004642 ferric ammonium citrate Drugs 0.000 abstract 1
- 230000035987 intoxication Effects 0.000 abstract 1
- 231100000566 intoxication Toxicity 0.000 abstract 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 abstract 1
- 229940001474 sodium thiosulfate Drugs 0.000 abstract 1
- 238000012360 testing method Methods 0.000 description 13
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 description 10
- 230000001580 bacterial effect Effects 0.000 description 10
- 229910000037 hydrogen sulfide Inorganic materials 0.000 description 10
- 206010016952 Food poisoning Diseases 0.000 description 6
- 208000019331 Foodborne disease Diseases 0.000 description 6
- 235000013372 meat Nutrition 0.000 description 6
- 229920001817 Agar Polymers 0.000 description 5
- 239000008272 agar Substances 0.000 description 5
- 238000002474 experimental method Methods 0.000 description 4
- 239000003112 inhibitor Substances 0.000 description 4
- 235000015097 nutrients Nutrition 0.000 description 4
- 239000012085 test solution Substances 0.000 description 4
- 108091005804 Peptidases Proteins 0.000 description 3
- 239000004365 Protease Substances 0.000 description 3
- 241000588769 Proteus <enterobacteria> Species 0.000 description 3
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 3
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 description 3
- 238000012790 confirmation Methods 0.000 description 3
- 238000011109 contamination Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000007789 gas Substances 0.000 description 3
- 229910052739 hydrogen Inorganic materials 0.000 description 3
- 239000008213 purified water Substances 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 239000003381 stabilizer Substances 0.000 description 3
- 229910052717 sulfur Inorganic materials 0.000 description 3
- 241000293869 Salmonella enterica subsp. enterica serovar Typhimurium Species 0.000 description 2
- 241000607272 Vibrio parahaemolyticus Species 0.000 description 2
- 238000010411 cooking Methods 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 244000144977 poultry Species 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 238000002791 soaking Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- ZEWICQFXGKLTOP-UHFFFAOYSA-N 2-hydroxypropane-1,2,3-tricarboxylic acid;sulfuric acid Chemical compound OS(O)(=O)=O.OC(=O)CC(O)(C(O)=O)CC(O)=O ZEWICQFXGKLTOP-UHFFFAOYSA-N 0.000 description 1
- 206010004022 Bacterial food poisoning Diseases 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- 229920000742 Cotton Polymers 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 208000005577 Gastroenteritis Diseases 0.000 description 1
- 101000990986 Homo sapiens Myosin regulatory light chain 12A Proteins 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 102100030329 Myosin regulatory light chain 12A Human genes 0.000 description 1
- FNZSVEHJZREFPF-ZETCQYMHSA-N Nonate Chemical compound CCCCC[C@H](C(O)=O)CC(O)=O FNZSVEHJZREFPF-ZETCQYMHSA-N 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 241000588770 Proteus mirabilis Species 0.000 description 1
- 241000277263 Salmo Species 0.000 description 1
- 208000006731 Salmonella Food Poisoning Diseases 0.000 description 1
- 241001354013 Salmonella enterica subsp. enterica serovar Enteritidis Species 0.000 description 1
- 241000293871 Salmonella enterica subsp. enterica serovar Typhi Species 0.000 description 1
- 241000277331 Salmonidae Species 0.000 description 1
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 1
- 241000191967 Staphylococcus aureus Species 0.000 description 1
- 229910000831 Steel Inorganic materials 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- CIUQXNVNAZQKLO-UHFFFAOYSA-K [Cu+3].C(C)(=O)[O-].S(=O)(=O)([O-])[O-] Chemical compound [Cu+3].C(C)(=O)[O-].S(=O)(=O)([O-])[O-] CIUQXNVNAZQKLO-UHFFFAOYSA-K 0.000 description 1
- 208000012873 acute gastroenteritis Diseases 0.000 description 1
- 210000000941 bile Anatomy 0.000 description 1
- 239000003054 catalyst Substances 0.000 description 1
- 229910052802 copper Inorganic materials 0.000 description 1
- 239000010949 copper Substances 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 208000002925 dental caries Diseases 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 231100000676 disease causative agent Toxicity 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229960002413 ferric citrate Drugs 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 150000002431 hydrogen Chemical class 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- NPFOYSMITVOQOS-UHFFFAOYSA-K iron(III) citrate Chemical compound [Fe+3].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NPFOYSMITVOQOS-UHFFFAOYSA-K 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 229960005337 lysine hydrochloride Drugs 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 1
- 235000019799 monosodium phosphate Nutrition 0.000 description 1
- PGSADBUBUOPOJS-UHFFFAOYSA-N neutral red Chemical compound Cl.C1=C(C)C(N)=CC2=NC3=CC(N(C)C)=CC=C3N=C21 PGSADBUBUOPOJS-UHFFFAOYSA-N 0.000 description 1
- 230000007918 pathogenicity Effects 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 239000002574 poison Substances 0.000 description 1
- 231100000614 poison Toxicity 0.000 description 1
- -1 polypeptone Substances 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 238000003908 quality control method Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 210000000813 small intestine Anatomy 0.000 description 1
- 239000001509 sodium citrate Substances 0.000 description 1
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 1
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 1
- 239000001488 sodium phosphate Substances 0.000 description 1
- 229910000162 sodium phosphate Inorganic materials 0.000 description 1
- 235000011008 sodium phosphates Nutrition 0.000 description 1
- 229910052938 sodium sulfate Inorganic materials 0.000 description 1
- 235000011152 sodium sulphate Nutrition 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000010959 steel Substances 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 239000011593 sulfur Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
Description
【発明の詳細な説明】
〔産業上の利用分野〕
本発明は、食肉等に存在する食中毒原因菌であるサルモ
ネラ属の細菌を簡便に検出することができるサルモネラ
菌の検出方法並びに該方法に用いるサルモネラ菌用培地
及び検出紙に関する。Detailed Description of the Invention [Field of Industrial Application] The present invention provides a method for detecting Salmonella that can easily detect bacteria of the genus Salmonella that causes food poisoning present in meat, etc., and a method for detecting Salmonella used in the method. related to a medium for use and a detection paper.
我国で発生する細菌性食中毒のうち、発生件数の上位を
占める原因物質を見ると、第1位が腸炎ビブリオ、第2
位が黄色ブドウ球菌、第3位がサルモネラであり、サル
モネラは病原性の面から見て重要な食中毒細菌である。Looking at the causative agents that account for the highest number of bacterial food poisoning cases in Japan, the first is Vibrio parahaemolyticus, and the second is Vibrio parahaemolyticus.
Staphylococcus aureus ranks third, followed by Salmonella, and Salmonella is an important food-poisoning bacterium from the perspective of pathogenicity.
サルモネラによる急性胃腸炎は、食品中で増殖した大量
のサルモネラが経口的に摂取され、胃内を通過するとき
にある程度は殺菌されるが、なお生き残った菌が小腸内
に入り、再び増殖する結果発症するもので、その殆どは
7η染食品の摂取によって起こるものと考えられている
。特に、食肉類のサルモネラ汚染は世界各国がその防止
対策に苦慮している問題であり、我国においても、と殺
場及び食鳥処理場で生産される食肉類からサルモネラが
高率で検出され、サルモネラ食中毒の原因食品としては
これら食肉類及びその加工品による発生件数が最も高い
とされている。Acute gastroenteritis caused by Salmonella occurs when large amounts of Salmonella that have grown in food are ingested orally and are sterilized to some extent as they pass through the stomach, but the surviving bacteria enter the small intestine and multiply again. Most of the symptoms are thought to be caused by ingestion of 7η-dyed foods. In particular, salmonella contamination of meat is a problem that countries around the world are struggling to prevent, and even in Japan, salmonella has been detected at a high rate in meat produced at slaughterhouses and poultry processing plants. Among the foods that cause salmonella food poisoning, these meats and their processed products are said to cause the highest number of outbreaks.
従って、サルモネラによる食中毒を予防するためには、
まず第1に検査によりサルモネラに汚染された食品を早
く探知し、排除すること、第2に食品を低温に保存して
汚染サルモネラの増殖を抑制するか、或いは食品の加熱
によって汚染菌の殺菌を行なうかの対策を検査結果に応
じて講じることが重要であり、そのためにはサルモネラ
の検査を日常業務の中に取り入れ、品質管理を十分に行
なうことが必要である。Therefore, to prevent food poisoning caused by Salmonella,
Firstly, food contaminated with Salmonella should be quickly detected through inspection and eliminated.Secondly, foods should be stored at low temperatures to inhibit the growth of contaminated Salmonella, or food should be heated to sterilize the contaminated bacteria. It is important to take countermeasures depending on the test results, and for this purpose, it is necessary to incorporate Salmonella tests into daily work and to perform sufficient quality control.
この場合、サルモネラの検出方法としては、従来より以
下に述べる方法が一般に採用されている。In this case, as a method for detecting Salmonella, the method described below has conventionally been generally employed.
即ち、まず精製水12に下記成分
ペプトン i o、 o gマ
ンニット 5.0gリン酸一
水素ナトリウム 6.45 gリン酸二水素ナ
トリウム 2.0g精製生胆汁末
20.0 gブリリアントグリン
0.0135 gを溶解したpH7.2±0.1の2
2Mブイヨンに検体を加え、35℃において15〜18
時間前培養する。この22Mブイヨンはダラム陽性菌の
発育を阻止するものであるが、腸内細菌間ではマンニッ
ト非発酵性のプロテウスの発育がやや遅れるにすぎず、
従って本培地はサルモネラ検査の曲培養のみに用いられ
る。次に、精製水12中に下記成分ペプトン
5.0g乳糖 4.0g
リン酸ナトリウム 10.0 g亜セ
レン酸ナトリウム 4.0gを溶解した
pH7,0±0.1のセレナイト培地に上記培養後の2
2Mブイヨンを所定量加え、43℃で18〜24時間増
菌培養する。本培地は中等度の選択性をもつサルモネラ
の増菌培地で、大便中のサルモネラ・チフィやサルモネ
ラ・バラチフィAの増菌にも使用できるが、サルモネラ
・コレレシュイスは本培地に発育しない。その後、精製
水IP中に下記成分
肉エキス 5.0gペプトン
7.5g胆汁酸塩
9.0g乳 キ唐
1 0.
Ogクエン′酸ナトリウム 8.5
g千オ硫酸ナトリウム 5.5gクエ
ン酸第二鉄 1.0gニュートラル
レッド 0.025gブリリアントグリ
ン 0.33mg寒天 13.
5 g
を溶解して固めたpH7,0±0.1のSS寒天培地に
上記セレナイト培地を所定量移植し、35゛Cにおいて
18〜2・1時間直接分離培養を行なう。そして、その
後更にサルモネラの疑いのある集落をとってTSI寒天
、LIM培地等の確認培地に移植し、濫別培養を行なっ
た後、サルモネラの同定を行なうものである。That is, first, the following ingredients were added to purified water 12: peptone io, o g mannitol 5.0 g sodium monohydrogen phosphate 6.45 g sodium dihydrogen phosphate 2.0 g purified raw bile powder
20.0 g brilliant green
0.0135 g dissolved at pH 7.2 ± 0.1 2
Add the sample to 2M broth and incubate at 35°C for 15-18
Pre-incubate for an hour. This 22M broth inhibits the growth of Durham-positive bacteria, but among intestinal bacteria, the growth of Proteus, which does not ferment mannitol, is only slightly delayed.
Therefore, this medium is used only for culture for Salmonella testing. Next, in purified water 12, the following ingredients peptone were added.
5.0g Lactose 4.0g Sodium phosphate 10.0g Sodium selenite 4.0g dissolved in selenite medium at pH 7.0±0.1.
Add a predetermined amount of 2M broth and culture for enrichment at 43°C for 18 to 24 hours. This medium is an enrichment medium for Salmonella with moderate selectivity, and can be used to enrich Salmonella typhi and Salmonella baratifi A in stool, but Salmonella cholerethuis will not grow on this medium. After that, add the following ingredients to the purified water IP: 5.0g peptone, 7.5g bile salts.
9.0g milk Kikara
1 0.
Og Sodium Citrate 8.5
g Sodium 1,000 sulfate 5.5 g Ferric citrate 1.0 g Neutral Red 0.025 g Brilliant Green 0.33 mg Agar 13.
A predetermined amount of the above selenite medium is transferred to a pH 7.0±0.1 SS agar medium in which 5 g of the selenite medium has been dissolved and solidified, and direct separation culture is performed at 35°C for 18 to 2.1 hours. Thereafter, a colony suspected of containing Salmonella is taken and transplanted onto a confirmation medium such as TSI agar or LIM medium, followed by separate culture, followed by identification of Salmonella.
上述したように、サルモネラの検出には、EEMブイヨ
ンによる前培養、セレナイト培地による増菌培養、SS
寒天による直接分離培養、確認培地による鑑別培養とい
う4回の培養を行なう必要がある。これは、サルモネラ
のみを選択的に増菌させることが非常に困難なためであ
る。従って、サルモネラ汚染の検査を日常的に行なうに
は膨大な時間、設備、費用を要し、このような検査をな
かなか実施できないのが実情であった。As mentioned above, the detection of Salmonella requires pre-culture with EEM broth, enrichment culture with selenite medium, SS
It is necessary to perform four cultures: direct isolation culture using agar and differential culture using confirmation medium. This is because it is extremely difficult to selectively increase Salmonella alone. Therefore, routinely testing for Salmonella contamination requires enormous amounts of time, equipment, and expense, and the reality is that such testing is difficult to carry out.
本発明は、上記事情に鑑みなされたもので、サルモネラ
の検出を短時間で簡便かつ安価に行なうことができる検
出方法並びに該方法に用いる培地及び検出紙を提供する
ことを目的とする。The present invention was made in view of the above circumstances, and aims to provide a detection method that allows Salmonella detection to be carried out easily and inexpensively in a short time, as well as a culture medium and detection paper used in the method.
〔課題を解決するための手段及び作用〕本発明は、上記
目的を達成するため、水1000m6中に下記成分
ペプトン 9〜11gマンニット
4.5〜5,5gリン酸一水素ナトリ
ウム 6〜7gノン酸二水素カリウム 1,5
〜2.5g亜セレン酸ナトリウム 9〜ftg硫
酸洞 0.5〜1.5mgブリリア
ントグリン 0.1〜0.3mgを溶解したpH
7,2±0.2の前処理用液体培地にサルモネラを含む
検体を加え、33〜39℃において16〜26時間培養
を行なった後、水1000ml中に下記成分
ポリペプトン
プロテオーゼペプトン
酵母エキス
ハートエキス
マンニット
塩化ナトリウJ、
千オ硫酸ナトリウム
クエン酸1失アニ/モニウム
1、−リン;ン塩酸塩
胆汁酸塩
酢酸鉛
硫酸銅
9〜11g
4.5〜5.5g
4.5〜5.5g
1.5〜2.5g
2.5〜3.5g
3.5〜4.5g
3.5〜4.5g
0、5〜1,5g
4.5〜5.5g
2.5〜3.5g
O,1〜0.3g
1.5〜2.5mg
を溶解したpH6,8±0.2の検出用液体培地を含浸
してなる検出紙に上記培養後の前処理用液体培地を付着
させ、33〜39゛Cにおいて18〜26時間培養を行
なった後、該検出紙上にあらわれるスポットからサルモ
ネラを検出するようにしたサルモネラの検出方法を提供
する。[Means and effects for solving the problems] In order to achieve the above-mentioned object, the present invention contains 9 to 11 g of the following component peptone in 1000 m6 of water.
4.5-5.5g Sodium monohydrogen phosphate 6-7g Potassium dihydrogen nonate 1.5
~2.5g sodium selenite 9~ftg sulfate 0.5~1.5mg brilliant green 0.1~0.3mg dissolved pH
A sample containing Salmonella was added to a pretreatment liquid medium of 7.2 ± 0.2, and after culturing at 33 to 39°C for 16 to 26 hours, the following ingredients polypeptone protease, peptone yeast extract, and heart extract were added to 1000 ml of water. Mannitol sodium chloride J, sodium 1,000 sulfate citrate 1 loss/monium 1,-phosphorus; hydrochloride bile salt lead acetate copper sulfate 9-11 g 4.5-5.5 g 4.5-5.5 g 1 .5-2.5g 2.5-3.5g 3.5-4.5g 3.5-4.5g 0,5-1,5g 4.5-5.5g 2.5-3.5g O, The pretreatment liquid medium after culturing was attached to a detection paper impregnated with a detection liquid medium of pH 6.8 ± 0.2 in which 1 to 0.3 g of 1.5 to 2.5 mg was dissolved. A method for detecting Salmonella is provided, in which Salmonella is detected from spots appearing on the detection paper after culturing at 39°C for 18 to 26 hours.
また、本発明は、水1000+nj!中に下記成分ペプ
トン 9〜11gマンニット
4.5〜5,5gリン酸一水素ナトリウム
6〜7gリン酸二水素カリウム 1.5〜2
.5g亜セレン酸ナトリウム 9〜11g硫fJ
銅 0.5〜1.5mgブリリアン
トグリン 0.1〜0.3mgが溶解され、pt
+が7.2±0.2であるサルモネラ用培地を提供する
。In addition, the present invention uses water 1000+nj! Contains the following ingredients: peptone 9-11g mannitol
4.5-5.5g Sodium monohydrogen phosphate 6-7g Potassium dihydrogen phosphate 1.5-2
.. 5g Sodium selenite 9-11g Sulfur fJ
Copper 0.5-1.5 mg brilliant green 0.1-0.3 mg are dissolved, pt
Provided is a Salmonella medium in which + is 7.2±0.2.
更に本発明は、水100100O中に下記成分ポリペプ
トン 9〜11gプロテオーゼベプトン
4.5〜5.5g酵母エキス 4
,5〜5.5gハートエキス 1.5〜2
.5gマンニット 2.5〜3.5g
塩化ナトリウム 3.5〜4.58チオ硫
酸ナトリウム 3.5〜4.5gクエン酸鉄アン
モニウム 0.5〜1.5gL−リジン塩酸塩
4.5〜5.5g胆汁酸塩 2.
5〜3.5g酢酸鉛 0.1〜0.
3g硫酸銅 】、5〜2.5 m
gが溶解され、pHが6.8±0.2であるサルモネラ
用培地を提供する。Furthermore, the present invention contains the following components polypeptone 9 to 11 g proteose peptone 4.5 to 5.5 g yeast extract 4 in 100100 O of water.
, 5-5.5g heart extract 1.5-2
.. 5g Mannit 2.5-3.5g
Sodium chloride 3.5-4.58 Sodium thiosulfate 3.5-4.5g Iron ammonium citrate 0.5-1.5g L-lysine hydrochloride
4.5-5.5g bile salts 2.
5-3.5g Lead acetate 0.1-0.
3g copper sulfate], 5-2.5 m
Provide a Salmonella medium in which g is dissolved and the pH is 6.8±0.2.
また史に、本発明は、上記検出用液体培地を含浸してな
るサルモ不う用検出紙を提供する。Additionally, the present invention provides a detection paper for Salmonella impregnated with the above-mentioned liquid medium for detection.
本発明方法においては、まず前処理用液体培地によって
サルモネラを選択的に増菌培養し、次いでこのサルモネ
ラを検出紙に移植すると共に、検出用液体培地によって
更に培養を行ない、サルモネラが産生ずる硫化水素と検
出用培地中の酢酸鉛毒との反応によって検出紙にあらわ
れるスポットからサルモネラの定性試験を行なうもので
、これにより検体中の微量のサルモネラでも検出できる
ようにしたものである。即ち、本発明方法は、前処理用
液体培地で第1次の培面を行なうことにより、サルモネ
ラを増菌すると共に、サイトロハクター、プロテウス等
のサルモネラ以外の硫化水素産生菌の増菌を抑制し、次
いで上記第1次の選別終了後に検出用液体培地を含む検
出紙でサルモネラを第2次的に選択発育させ、検出紙に
スボノl−を形成させるようにしたもので、従って検出
紙」二に硫化水素と酢酸鉛との反応による赤銅色のスポ
ットがあられれれば、その検体はサルモネラ陽性と定性
的に判定することになる。In the method of the present invention, first, Salmonella is selectively enriched and cultured in a pretreatment liquid medium, and then this Salmonella is transplanted onto a detection paper, and further cultured in a detection liquid medium to produce hydrogen sulfide produced by Salmonella. This is a qualitative test for Salmonella from the spots that appear on the detection paper due to the reaction between the lead acetate poison in the detection medium, and this makes it possible to detect even minute amounts of Salmonella in the sample. That is, in the method of the present invention, by carrying out the first culture using a pretreatment liquid medium, Salmonella is enriched, and at the same time, the proliferation of hydrogen sulfide-producing bacteria other than Salmonella, such as Cytorrhacter and Proteus, is suppressed. Then, after the first selection is completed, Salmonella is selectively grown on a detection paper containing a liquid detection medium to form subonol-1 on the detection paper. Second, if a copper-colored spot appears due to the reaction between hydrogen sulfide and lead acetate, the sample is qualitatively determined to be Salmonella positive.
この場合、上記前処理用培地においては、ペプトン及び
マンニットが栄l素、リン酸一水素ナトリウム、リン酸
二水素カリウムがp++安定剤、亜セレン酸ナトリウム
及び硫酸銅がサルモネラ以外の菌の発育を抑制する抑制
剤、ブリリアン)・グリノがpn安定剤及び抑制剤とし
て作用するものであるが、本培地においては抑制剤とし
て亜セレン酸ナトリウムと従来この種の培地で使用され
ていなかった硫酸銅とを併用したことにより、サルモネ
ラを選択的に増菌し、他の細菌の増菌を確実に抑制する
ことができる。また、上記噴出用培地においてはポリペ
プトン、プロテオーゼベプトン、酵母エキス、ハートエ
キス(牛心臓抽出物)、マンニットが栄養素、塩化ナト
リウムがpH安定剤、チオ硫酸ナトリウム及びクエン酸
鉄アンモニウムがサルモネラによる硫化水素ガス発生の
原料、L−リジン塩酸塩がサルモネラが選択的に利用し
得る栄養素、胆汁酸塩及び硫酸鋼が抑制剤、酢酸鉛がサ
ルモネラが産生ずる硫化水素ガスと結合してスポットを
形成する指示薬として作用するものであるが、本培地に
おいては抑制剤として胆汁酸塩と従来使用されていなか
った硫酸銅とを併用したことによりサルモネラのみを確
実に選択増菌させることができると共に、栄養素として
プロテオーゼペプトン及びハートエキスを用いたことに
より、サルモ不うの硫化水素ガス発生が良好に促進され
、検出紙に明確なスポットを形成させることができる。In this case, in the above pretreatment medium, peptone and mannitol are used as nutrient, sodium monohydrogen phosphate and potassium dihydrogen phosphate are used as p++ stabilizers, and sodium selenite and copper sulfate are used to prevent the growth of bacteria other than Salmonella. Brillian) and Gulino act as pn stabilizers and inhibitors, but in this culture medium, sodium selenite and copper sulfate, which have not been used in this type of culture medium, are used as inhibitors. By using these in combination, Salmonella can be selectively enriched and proliferation of other bacteria can be reliably suppressed. In addition, in the above-mentioned eruption medium, polypeptone, protease peptone, yeast extract, heart extract (bovine heart extract), and mannitol are nutrients, sodium chloride is a pH stabilizer, and sodium thiosulfate and iron ammonium citrate are used to prevent salmonella. Raw material for hydrogen sulfide gas generation, L-lysine hydrochloride is a nutrient that Salmonella can selectively use, bile salts and steel sulfate are inhibitors, and lead acetate combines with hydrogen sulfide gas produced by Salmonella to form spots. However, in this medium, by using bile salts as an inhibitor together with copper sulfate, which has not been used in the past, it is possible to reliably selectively enrich only Salmonella, as well as provide nutrients. By using protease peptone and heart extract as the catalyst, the generation of hydrogen sulfide gas by Salmonella can be favorably promoted and a clear spot can be formed on the detection paper.
従って、これまではEEMブイヨンによってサルモネラ
及びその他の細菌を一緒に増菌させ、次にセレナイト培
地によってサルモネラの選択培養を行なっていたのに対
し、本発明前処理培地によれば1回の培養でサルモ不う
を確実に選択培養することができる。また、これまでは
直接分層培養及び鑑別培養を行なってサルモネラを検出
していたのに対し、本発明検出用培地によれば検出紙に
おいて培養を行ない、検出紙にあらわれるスポットを調
べるだけで而単にサルモネラの検出を行なうことができ
る。Therefore, whereas previously Salmonella and other bacteria were multiplied together using EEM broth and then selectively cultured for Salmonella using selenite medium, with the pretreatment medium of the present invention, only one culture is required. Salmonella can be reliably selectively cultured. In addition, whereas previously Salmonella was detected by direct layer culture and differential culture, with the detection medium of the present invention, it is possible to simply culture on detection paper and examine the spots that appear on the detection paper. One can simply detect Salmonella.
それ故、本発明によれば、選択性に優れた上記前処理用
培地及び検出用培地を用いてサルモネラを増菌培養する
ようにしたので、2回の培養によってサルモネラと他の
硫化水素産生菌とを選別でき、従って短時間でサルモネ
ラを検出できると共に、検出用培地による培養を検出紙
において行ない、この検出紙にあらわれるスポットによ
って判定するようにしたので、極めて簡便かつ安価にサ
ルモネラの検出を行なうことができるものである。Therefore, according to the present invention, since Salmonella is enriched and cultured using the pretreatment medium and detection medium with excellent selectivity, Salmonella and other hydrogen sulfide-producing bacteria can be cultured twice. Therefore, Salmonella can be detected in a short time. In addition, culture using a detection medium is carried out on a detection paper, and judgment is made by the spots that appear on this detection paper, making it possible to detect Salmonella very simply and at low cost. It is something that can be done.
以下、本発明につき更に詳しく説明する。The present invention will be explained in more detail below.
本発明検出方法においては、まずサルモネラを含む検体
を上記前処理用液体培地に加え、増菌培養を行なう。な
お、培養はできるだけ嫌気的条件下で行なうことが好ま
しい。In the detection method of the present invention, first, a sample containing Salmonella is added to the above-mentioned pretreatment liquid medium and cultured for enrichment. In addition, it is preferable to perform the culture under anaerobic conditions as much as possible.
この場合、培地に検体を加える手段に限定はないが、固
体検体、粉体検体の場合は直接加える方法、拭き取り検
体の場合は綿棒等で検体を拭き取った後、この2綿棒等
を培地中に浸漬し、洗い出す方法を採用することができ
る。なお、検体の添加量に特に制限はない。In this case, there are no limitations on the method of adding the specimen to the medium, but in the case of solid or powdered specimens, it may be added directly, or in the case of wiped specimens, after wiping the specimen with a cotton swab etc., the two swabs etc. are added to the medium. A soaking and washing method can be adopted. Note that there is no particular limit to the amount of sample added.
また、前処理用液体培地は、水1000ml中における
各成分の配合量を下記の通りとし、かつp++を7.2
とすることがサルモネラの増菌のために特に好ましい。In addition, the pretreatment liquid medium has the following amounts of each component in 1000 ml of water, and has a p++ of 7.2.
It is particularly preferable to increase Salmonella bacteria.
即ち、
ペプトン 10gマンニット
5gリン酸一水素ナトリ
ウム 6.45 gリン酸二水素カリウム
2g亜セレン酸ナトリウム
10g硫酸銅 low
ブリリアントグリン 0.2■更に、培養
温度は35〜37℃、培養時間は18〜20時間とする
ことがサルモネラの増菌の点で特に好適である。Namely: Peptone 10g Mannitol 5g Sodium monohydrogen phosphate 6.45 g Potassium dihydrogen phosphate
2g sodium selenite
10g copper sulfate low
Brilliant Grine 0.2 ■ Furthermore, it is particularly preferable to set the culture temperature to 35 to 37°C and the culture time to 18 to 20 hours from the viewpoint of increasing Salmonella bacteria.
本発明においては、次に上記増菌培養を行なった前処理
用培地を上記検出用液体培地を含浸した検出紙に付着さ
せ、更に培養を行なう。なお、培養はできるだけ嫌気的
条件下で行なうことが望ましい。In the present invention, next, the pretreatment medium subjected to the enrichment culture is attached to a detection paper impregnated with the liquid detection medium, and further culture is performed. Note that it is desirable to perform the culture under anaerobic conditions as much as possible.
この場合、検出紙に前処理用培地を付着させる手段に限
定はないが、検出紙を前処理用培地に浸漬し、検出紙に
前処理用培地をしみこませる方法が好適に採用される。In this case, there is no limitation on the means for attaching the pretreatment medium to the detection paper, but a method of immersing the detection paper in the pretreatment medium and soaking the pretreatment medium into the detection paper is preferably adopted.
なお、前処理用培地の検出紙への付着量に特に制限はな
い。Note that there is no particular limit to the amount of the pretreatment medium attached to the detection paper.
また、検出紙に含浸させる検出用液体培地は、水100
0ml中における各成分の配合量を下記の通りとし、か
つpHを6.8とすることがサルモネラの検出のために
特に好ましい。即ち、ポリペプトン
lOgプロテオーゼペプトン 5g
酵母エキス 5gハートエキ
ス 2gマンニット
3g塩化ナトリウム
4g千オ硫酸ナトリウム
4gクエン酸鉄アンモニウム 1gL
−リジン塩酸塩 5g胆汁酸塩
3g酢酸鉛
0.2g硫酸銅
0.002g更に、培養温度は35〜37℃1培養
時間は20〜24時間とすることがサルモネラの検出の
点で特に好適である。なお、胆汁酸塩としてはデイフコ
社製胆汁酸塩隘3を用いることが好ましい。In addition, the detection liquid medium to be impregnated into the detection paper is water 100%
It is particularly preferable for the detection of Salmonella that the amounts of each component in 0 ml are as follows and the pH is 6.8. i.e. polypeptone
lOg proteose peptone 5g
Yeast extract 5g Heart extract 2g Mannitol
3g sodium chloride
4g sodium sulfate
4g iron ammonium citrate 1gL
-Lysine hydrochloride 5g bile salt
3g lead acetate
0.2g copper sulfate
0.002g Further, it is particularly suitable for the detection of Salmonella that the culture temperature is 35 to 37°C and the culture time is 20 to 24 hours. As the bile salt, it is preferable to use Bile Salt No. 3 manufactured by Difco.
本発明は、上記培養後、検出紙上にあられれたスポット
からサルモ不うを検出する。この場合、検出紙の表面及
び裏面を観察し、赤銅色のスポットが見られた場合はサ
ルモネラ陽性と判定するものである。The present invention detects Salmonella from the spot formed on the detection paper after the above-mentioned culture. In this case, the front and back sides of the detection paper are observed, and if a copper-colored spot is seen, it is determined to be salmonella positive.
なお、本発明は、サルモネラ属に属するあらゆる硫化水
素産生菌の検出に使用し得る。Note that the present invention can be used to detect any hydrogen sulfide-producing bacteria belonging to the genus Salmonella.
以上説明したように、本発明によれば、食肉等に存在す
る食中毒細菌であるサルモネラを迅速、筒便かつ安価に
検出することができる。この場合、本発明前処理用培地
及び検出用培地によればサルモネラを確実に選択増菌さ
せることができ、本発明検出紙によればサルモネラに起
因するスポットを確実に発現させることができる。As explained above, according to the present invention, Salmonella, which is a food poisoning bacterium present in meat and the like, can be detected quickly, conveniently, and inexpensively. In this case, the pretreatment medium and detection medium of the present invention can reliably selectively enrich Salmonella, and the detection paper of the present invention can reliably cause spots caused by Salmonella to appear.
従って、本発明は、と斎場、食鳥処理場、食肉販売店、
給食センター、弁当仕出し店など、多くの食品を取り扱
う業界で食品や調理器具などがサルモ不うに汚染されて
いるか否かを常時検査するのに、簡易、迅速で、しかも
安価であるという点で利用価値が高い。また、DHL、
MLCB等の生培地では、サルモネラと他の産生菌が同
時に発育するので、分離後確認培地で鑑別をしなければ
ならないが、本発明検出紙はサルモネラのみが赤銅色の
スポットを形成するので、菌の分離、同定を省略しても
発色したものはほぼまちがいなくすルモネラであると判
定できる。但し、本発明による検出は前処理培地による
選択増菌という前処理を経由するため発色スポット数は
食品中のサルモネラ閑散ではなく、あくまで定性試験で
あり、このような性質を理解して本発明を利用すれば、
頻繁に食品や調理器具のサルモネラ汚染をチエツクする
ことができるので、本発明は食中毒予防の点で極めて有
用なものである。Therefore, the present invention is applicable to funeral homes, poultry slaughterhouses, meat shops,
It is used in industries that handle many foods, such as school lunch centers and lunch box caterers, because it is simple, quick, and inexpensive to constantly inspect food and cooking utensils to see if they are contaminated with salmonids. High value. Also, DHL,
In a live medium such as MLCB, Salmonella and other producing bacteria grow at the same time, so they must be identified using a confirmation medium after separation, but with the detection paper of the present invention, only Salmonella forms copper-colored spots, so Salmonella can be easily detected. Even if separation and identification are omitted, anything that develops color can be determined to be Lumonella without any doubt. However, since the detection by the present invention goes through a pretreatment of selective enrichment using a pretreatment medium, the number of colored spots does not indicate the presence of Salmonella in food, but is only a qualitative test. If you use it,
The present invention is extremely useful in preventing food poisoning, as food and cooking utensils can be frequently checked for Salmonella contamination.
以下、実験例により本発明の効果を具体的に示す。Hereinafter, the effects of the present invention will be specifically illustrated by experimental examples.
上記前処理用液体培地及び検出紙の性能を調べるため、
サルモ不う及びその他の硫化水素産生菌を用いて下記の
実験を行なった。In order to investigate the performance of the above pretreatment liquid medium and detection paper,
The following experiments were conducted using Salmo nigra and other hydrogen sulfide producing bacteria.
供試菌株
■ サルモネラ・チフィムリウム(Salmoneli
atyphimurium、 食中毒由来法):以下
Stと略す■ サルモネラ・エンテリチジス(Salm
onellaenLeritidis、 食中毒由来
法);以下Seと略す■ サイトロハクター・フロイン
デイ (Ci tro−bacLer freundi
i、食品由来法)二辺下crと略す■ プロテウス・ミ
ラビリス(Proteus mlrabilis食品由
来株)
供試菌液の調製
保存菌株を普通ブイヨンで3回継代培養して若返りをは
かったのち、普通寒天斜面培地に培養して試験菌とした
。Test strain ■ Salmonella typhimurium
atyphimurium, food poisoning origin method): Hereinafter abbreviated as St ■ Salmonella enteritidis (Salm.
onellaenLeritidis, food poisoning origin); hereinafter abbreviated as Se■ Citro-bacLer freundi
i, Food-derived method) Abbreviated as 2-sided cr ■ Proteus mirabilis (Proteus mlrabilis food-derived strain) Preparation of test bacterial solution The preserved bacterial strain was subcultured three times in ordinary broth to rejuvenate it, and then transferred to ordinary agar. It was cultured on a slant medium and used as a test fungus.
試験菌の調製には、StとSeを普通ブイヨンに一緒に
摂取し、また、Cfc!:Pmも普通ブイヨンに一緒に
トH取し、これらを37℃で24時間培養した後、Cf
+Pmはこれを試験液とし、St+36についてはこれ
を試験原液とし、更に滅菌生理食塩水で10−1及び1
0−2に希釈したものを実験に使用した。To prepare the test bacteria, St and Se were taken together in normal broth, and Cfc! : Pm was also collected in ordinary broth, and after culturing them at 37°C for 24 hours, Cf
For +Pm, this was used as the test solution, and for St+36, this was used as the test stock solution, and then 10-1 and 1 was added with sterile physiological saline.
A dilution of 0-2 was used in the experiment.
サル七名うと他の1(2S産生菌との徂合わせ混合菌量
比による実験群は次の通りである。The experimental groups according to the combined bacterial ratio of seven monkeys and one other 2S-producing bacteria are as follows.
St+3e : Cf+Pm
1群 1 ・ 1
■群 1 ・ 10
■群 1:]00
このような菌量比になるように調製した菌液を普通ブイ
ヨンにそれぞれIccづつ摂取し、第1図に示す1順序
に従って実験した。St+3e: Cf+Pm Group 1 1 ・ 1 ■ Group 1 ・ 10 Group ■ 1: ] 00 Ingest 1cc of each of the bacterial solutions prepared to have such a bacterial amount ratio in normal broth, and then ingest them in the order shown in Figure 1. The experiment was conducted according to the following.
実験結果
■ 菌憧別試験液に対する検出化の比較サルモネラと他
の硫化水素産生菌の各菌鼠別試験液を本発明前処理用培
地及びセレナイト培地で前処理した後、これら培地を本
発明検出紙に一定量吸着させ、37℃20時間培養し、
検出紙上に出現した発色スポット数を比較すると表−1
に見られる結果を得た。Experimental Results ■ Comparison of Detection for Test Solutions for Salmonella and Other Hydrogen Sulfide-Producing Bacteria After pre-treating the test solutions for Salmonella and other hydrogen sulfide-producing bacteria with the pretreatment medium of the present invention and the selenite medium, these media were tested for the detection of the present invention. Adsorb a certain amount onto paper and culture at 37°C for 20 hours.
Table 1 shows a comparison of the number of colored spots that appeared on the detection paper.
We obtained the results shown in .
表−1
(増菌培養後の試験紙上のスポット検出状況)うの菌量
より多い割合になるにしたがって減少する傾向が見られ
るが、試験液中に他のHzS産生閑の1/100iのサ
ルモネラの存在であっても検出紙上にスポットの発現は
見られた。Table 1 (Spot detection status on test paper after bacterial enrichment culture) There is a tendency to decrease as the proportion becomes higher than the amount of caries, but there are 1/100i of Salmonella in other HzS producing cells in the test solution. Spots were observed on the detection paper even in the presence of .
■ 検出紙上のスポット菌種同定
前処理用の増菌培養としての本発明前処理培地とセレナ
イト培地との選択性を比較するため、発現スポットから
菌の分離を行ない、TSI培地およびLIM培地で性状
を調べ、サルモネラ用の診断血清で接種菌の確認同定を
行なった。結果は表−2に見られるとうりで、本発明培
地で前処理した場合は各菌量混合比の1群から■群とも
検出紙上に発現したスポットはすべてサルモネラであっ
た。■ In order to compare the selectivity of the pretreatment medium of the present invention and the selenite medium as enrichment culture for pretreatment for identification of bacterial species spotted on the detection paper, bacteria were isolated from the expression spots, and their properties were determined using TSI medium and LIM medium. The inoculated bacteria were confirmed and identified using diagnostic serum for Salmonella. The results are shown in Table 2, and when pretreated with the culture medium of the present invention, all spots expressed on the detection paper in groups 1 to 2 of each bacterial amount mixing ratio were Salmonella.
発色スポット数はCf+Pmの菌量がサルモネしかるに
、セレナイト培地で前処理したものでは、1群の菌量比
の場合は発現スポットはすべてサルモネラであったが、
■群の1:10菌量比の場合は発現スポットの80%が
サルモネラ、20%が他の11□3産生菌であった。The number of colored spots was determined by the number of Salmonella bacteria in Cf + Pm.However, in the case of pre-treated with selenite medium, all the expressed spots were Salmonella when the bacterial amount ratio was 1 group.
In the case of the 1:10 bacterial load ratio in group (2), 80% of the expression spots were Salmonella and 20% were other 11□3-producing bacteria.
■群のt:ioo閏世比の場合は発現スポットの40%
がサルモネラで60%が他のH,S産生菌とサルモ不う
の菌量が少なくなるに従って検出紙上に発現するスポッ
トも他の11□S産生菌が多くなり選択性が弱くなるこ
とがわかる。■In the case of group t:ioo leap ratio, 40% of the expression spots
It can be seen that as the amount of Salmonella and 60% of other H, S-producing bacteria decreases, the number of spots expressed on the detection paper increases as other 11□S-producing bacteria become weaker.
この事から、本発明検出紙は、前処理用培地として本発
明前処理培地を用いた場合の方が選択性が高く、検出紙
上のスポットはサルモネラ以外のH,S産生菌は発現し
ないことが確認された。From this, the detection paper of the present invention has higher selectivity when the pretreatment medium of the present invention is used as the pretreatment medium, and the spots on the detection paper do not express H, S-producing bacteria other than Salmonella. confirmed.
第1図は実験例における培養の手順を示すフローチャー
トである。FIG. 1 is a flowchart showing the culture procedure in an experimental example.
Claims (1)
ルモネラ菌を含む検体を加え、33〜39℃において1
6〜26時間培養を行ない、次いで水1000ml中に
下記成分 ポリペプトン9〜11g プロテオーゼペプトン4.5〜5.5g 酵母エキス4.5〜5.5g ハートエキス1.5〜2.5g マンニット2.5〜3.5g 塩化ナトリウム3.5〜4.5g チオ硫酸ナトリウム3.5〜4.5g クエン酸鉄アンモニウム0.5〜1.5g L−リジン塩酸塩4.5〜5.5g 胆汁酸塩2.5〜3.5g 酢酸鉛0.1〜0.3g 硫酸銅1.5〜2.5mg を溶解したpH6.8±0.2の検出用液体培地を含浸
してなる検出紙に上記培養後の前処理用液体培地を付着
させ、33〜39℃において18〜26時間培養を行な
った後、該検出紙上にあらわれるスポットからサルモネ
ラ菌を検出するようにしたことを特徴とするサルモネラ
菌の検出方法。 2、水1000ml中に下記成分 ペプトン9〜11g マンニット4.5〜5.5g リン酸一水素ナトリウム6〜7g リン酸二水素カリウム1.5〜2.5g 亜セレン酸ナトリウム9〜11g 硫酸銅0.5〜1.5mg ブリリアントグリン0.1〜0.3mg が溶解され、pHが7.2±0.2であることを特徴と
するサルモネラ菌用培地。 3、水1000ml中に下記成分 ポリペプトン9〜11g プロテオーゼペプトン4.5〜5.5g 酵母エキス4.5〜5.5g ハートエキス1.5〜2.5g マンニット2.5〜3.5g 塩化ナトリウム3.5〜4.5g チオ硫酸ナトリウム3.5〜4.5g クエン酸鉄アンモニウム0.5〜1.5g L−リジン塩酸塩4.5〜5.5g 胆汁酸塩2.5〜3.5g 酢酸鉛0.1〜0.3g 硫酸銅1.5〜2.5mg が溶解され、pHが6.8±0.2であることを特徴と
するサルモネラ菌用培地。 4、請求項3記載のサルモネラ菌用培地を含浸してなる
ことを特徴とするサルモネラ菌用検出紙。[Claims] 1. In 1000 ml of water, the following ingredients: 9 to 11 g of peptone, 4.5 to 5.5 g of mannitol, 6 to 7 g of sodium monohydrogen phosphate, 1.5 to 2.5 g of potassium dihydrogen phosphate, selenite A sample containing salmonella was added to a pretreatment liquid medium of pH 7.2±0.2 in which 9 to 11 g of sodium, 0.5 to 1.5 mg of copper sulfate, and 0.1 to 0.3 mg of brilliant green were dissolved, and the sample was heated at 33 to 39°C. In 1
Cultivate for 6 to 26 hours, then add the following ingredients to 1000 ml of water: 9 to 11 g of polypeptone, 4.5 to 5.5 g of proteose peptone, 4.5 to 5.5 g of yeast extract, 1.5 to 2.5 g of heart extract, and Mannitol 2. .5-3.5g Sodium chloride 3.5-4.5g Sodium thiosulfate 3.5-4.5g Iron ammonium citrate 0.5-1.5g L-lysine hydrochloride 4.5-5.5g Bile acids The above detection paper is impregnated with a liquid detection medium of pH 6.8 ± 0.2 in which 2.5 to 3.5 g of salt, 0.1 to 0.3 g of lead acetate, and 1.5 to 2.5 mg of copper sulfate are dissolved. A method for detecting Salmonella, which comprises: attaching a liquid medium for post-culture pretreatment and culturing at 33 to 39°C for 18 to 26 hours, and then detecting Salmonella from spots appearing on the detection paper. . 2. In 1000 ml of water, the following ingredients: 9-11 g of peptone, 4.5-5.5 g of mannitol, 6-7 g of sodium monohydrogen phosphate, 1.5-2.5 g of potassium dihydrogen phosphate, 9-11 g of sodium selenite, copper sulfate A culture medium for Salmonella, characterized in that 0.5 to 1.5 mg and 0.1 to 0.3 mg of brilliantogrin are dissolved therein and the pH is 7.2±0.2. 3. In 1000 ml of water, the following ingredients: 9-11 g of polypeptone, 4.5-5.5 g of proteose peptone, 4.5-5.5 g of yeast extract, 1.5-2.5 g of heart extract, 2.5-3.5 g of mannitol, chloride Sodium 3.5-4.5g Sodium thiosulfate 3.5-4.5g Iron ammonium citrate 0.5-1.5g L-lysine hydrochloride 4.5-5.5g Bile salts 2.5-3. A culture medium for Salmonella, characterized in that 5g of lead acetate, 0.1 to 0.3g of copper sulfate, and 1.5 to 2.5mg of copper sulfate are dissolved therein, and the pH thereof is 6.8±0.2. 4. A Salmonella detection paper impregnated with the Salmonella culture medium according to claim 3.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP21581488A JPH0669395B2 (en) | 1988-08-30 | 1988-08-30 | Method for detecting salmonella, medium for primary selective enrichment used in the method, medium for secondary selective enrichment detection and detection paper |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP21581488A JPH0669395B2 (en) | 1988-08-30 | 1988-08-30 | Method for detecting salmonella, medium for primary selective enrichment used in the method, medium for secondary selective enrichment detection and detection paper |
Publications (2)
Publication Number | Publication Date |
---|---|
JPH0265798A true JPH0265798A (en) | 1990-03-06 |
JPH0669395B2 JPH0669395B2 (en) | 1994-09-07 |
Family
ID=16678697
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JP21581488A Expired - Lifetime JPH0669395B2 (en) | 1988-08-30 | 1988-08-30 | Method for detecting salmonella, medium for primary selective enrichment used in the method, medium for secondary selective enrichment detection and detection paper |
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Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6203996B1 (en) * | 1994-03-01 | 2001-03-20 | Teagasc, The Agriculture And Food Development Authority | Rapid detection of bacteria liquid cultures |
WO2000077242A3 (en) * | 1999-06-15 | 2001-05-03 | Internat Diagnostics Group Plc | Detection of microorganisms |
WO2010029360A1 (en) | 2008-09-10 | 2010-03-18 | Solus Scientific Solutions Limited | Compositions and methods for the rapid growth and detection of microorganisms |
CN110241170A (en) * | 2019-07-24 | 2019-09-17 | 山西省食品药品检验所(山西省药品包装材料监测中心) | A kind of pure chemistry synthetic proteins peptone water reference culture medium and its preparation method and application |
WO2021200927A1 (en) | 2020-03-31 | 2021-10-07 | 日水製薬株式会社 | Medium for bacillus cereus group detection |
-
1988
- 1988-08-30 JP JP21581488A patent/JPH0669395B2/en not_active Expired - Lifetime
Cited By (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6203996B1 (en) * | 1994-03-01 | 2001-03-20 | Teagasc, The Agriculture And Food Development Authority | Rapid detection of bacteria liquid cultures |
WO2000077242A3 (en) * | 1999-06-15 | 2001-05-03 | Internat Diagnostics Group Plc | Detection of microorganisms |
WO2010029360A1 (en) | 2008-09-10 | 2010-03-18 | Solus Scientific Solutions Limited | Compositions and methods for the rapid growth and detection of microorganisms |
US20110159515A1 (en) * | 2008-09-10 | 2011-06-30 | Solus Scientific Solutions Limited | Compositions and methods for the rapid growth and detection of microorganisms |
CN110241170A (en) * | 2019-07-24 | 2019-09-17 | 山西省食品药品检验所(山西省药品包装材料监测中心) | A kind of pure chemistry synthetic proteins peptone water reference culture medium and its preparation method and application |
WO2021200927A1 (en) | 2020-03-31 | 2021-10-07 | 日水製薬株式会社 | Medium for bacillus cereus group detection |
Also Published As
Publication number | Publication date |
---|---|
JPH0669395B2 (en) | 1994-09-07 |
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