TWI609082B - 一種無污染而快速檢測沙門菌的方法 - Google Patents
一種無污染而快速檢測沙門菌的方法 Download PDFInfo
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- TWI609082B TWI609082B TW100108448A TW100108448A TWI609082B TW I609082 B TWI609082 B TW I609082B TW 100108448 A TW100108448 A TW 100108448A TW 100108448 A TW100108448 A TW 100108448A TW I609082 B TWI609082 B TW I609082B
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- Prior art keywords
- salmonella
- cotton swab
- patients
- dna
- detecting
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- 238000000034 method Methods 0.000 title claims description 14
- 241000607142 Salmonella Species 0.000 title claims description 7
- 238000012216 screening Methods 0.000 title 1
- 239000001974 tryptic soy broth Substances 0.000 claims description 7
- 108010050327 trypticase-soy broth Proteins 0.000 claims description 7
- 229920000742 Cotton Polymers 0.000 claims description 6
- 238000001514 detection method Methods 0.000 claims description 6
- 238000007400 DNA extraction Methods 0.000 claims description 4
- 238000012360 testing method Methods 0.000 claims description 4
- 241000894006 Bacteria Species 0.000 claims description 3
- 239000002609 medium Substances 0.000 claims description 3
- 229920001817 Agar Polymers 0.000 claims description 2
- 108020000946 Bacterial DNA Proteins 0.000 claims description 2
- 108020004414 DNA Proteins 0.000 claims description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 claims description 2
- 238000006243 chemical reaction Methods 0.000 claims description 2
- 238000006116 polymerization reaction Methods 0.000 claims description 2
- 210000000664 rectum Anatomy 0.000 claims description 2
- 238000000605 extraction Methods 0.000 claims 1
- 239000004576 sand Substances 0.000 claims 1
- 238000005070 sampling Methods 0.000 description 8
- 230000001580 bacterial effect Effects 0.000 description 3
- 238000003912 environmental pollution Methods 0.000 description 3
- 238000007689 inspection Methods 0.000 description 3
- 230000035755 proliferation Effects 0.000 description 3
- 230000035945 sensitivity Effects 0.000 description 3
- 206010012735 Diarrhoea Diseases 0.000 description 2
- 206010039438 Salmonella Infections Diseases 0.000 description 2
- 210000003608 fece Anatomy 0.000 description 2
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 2
- 238000003753 real-time PCR Methods 0.000 description 2
- 206010039447 salmonellosis Diseases 0.000 description 2
- 238000010561 standard procedure Methods 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- 238000010998 test method Methods 0.000 description 2
- 230000004543 DNA replication Effects 0.000 description 1
- 206010017914 Gastroenteritis salmonella Diseases 0.000 description 1
- 208000025796 Salmonella gastroenteritis Diseases 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 238000002405 diagnostic procedure Methods 0.000 description 1
- 230000002550 fecal effect Effects 0.000 description 1
- 244000000036 gastrointestinal pathogen Species 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000000474 nursing effect Effects 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 239000004575 stone Substances 0.000 description 1
Classifications
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Description
本發明乃開發了直腸拭子採樣經細菌增殖培養為基礎的即時聚合脢鏈反應(real-time PCR)檢測方法來幀測和篩檢腹瀉兒童是否由沙門氏菌引起。
傳統的黃金標準方法，由病患或家屬及看護採樣糞便，需要3天以上時間；且採樣的品質不穩定性，更易造成環境污染。
傳統的黃金標準方法，但是培養過程耗時，需要經過3至4天才能得到檢驗結果，往往即使檢測出沙門氏菌感染，也已症狀加劇而藥石罔效。雖然目前已有一些分子檢測方法，但這些方法並不適用於直接從糞便或直腸拭子標本作為常規診斷方法來檢測胃腸道的病原體。
1.本發明係以直腸拭子採樣經細菌增殖培養為基礎的即時聚合脢鏈反應(real-time PCR)檢測方法來幀測和篩檢病患腹瀉症狀是否由沙門氏菌引起。
2.本發明因為使用5ml trypticase soy broth(TSB)medium讓檢體內菌種快速增生，僅需3小時即可完成培養，大幅縮短得到檢驗結果之時間，採檢完5~6小時內就可得到結果，以利儘早進行診斷及治療。
3.另外本發明以循環時間(CT,cycle time)小於38為陽性判斷之域，此CT值是由計算沙門桿菌DNA複製數而來，因為使用前述TSB讓細菌快速增生之情形下，能夠以小至DNA複製數之作為判斷基準，不但加快檢驗時間，同時提高精確性及靈敏度。
1.針對嬰幼兒疑似沙門菌感染之病患，也適用於成人病患。
2.相較於由病患或家屬及看護採樣糞便，採樣的品質不穩定性，且易造成環境污染；本發明由臨床醫師執行，可保障採樣的品質和穩定性，無糞便接觸及環境污染。
3.檢測過程快速，相較於傳統檢驗方法需時3天以上，本發明從採樣後約5-6小時內完成。
4.利用本方法針對92位臨床上被診斷為疑似沙門氏菌腸胃炎感染的
兒童病患進行沙門氏菌檢測，並且與傳統檢驗方法作交叉比較：
(1)在敏感性方面，本方法達到84.6%的幀測靈敏度，明顯高于傳統檢驗方法的56.4%。
(2)而在專一性方面，相較於傳統檢驗方法的100%，本方法達到97.1%的專一度，與傳統方法無明顯差異。
(1)採樣：用棉棒於直腸口(肛門口)輕輕擦抹，將棉棒放入含有charcoal-Amies agar gel之試管內送實驗室；(2)細菌增殖：經震蕩方法將棉棒上之樣本脫落，於5ml trypticase soy broth(TSB)medium內35℃之溫度培養3小時；(3)DNA萃取：以一般細菌DNA萃取方法萃取DNA；(4)檢測：以沙門菌DNA序列引子進行聚合脢連鎖反應及時定量方法檢測，並以CT值小於38為陽性判斷之域。
Claims (1)
- 一種無污染而快速檢測沙門菌的方法，包含以下步驟：(1)用棉棒於直腸口(肛門口)輕輕擦抹，將棉棒放入含有charcoal-Amies agar gel之試管內送實驗室；(2)經震蕩方法將棉棒上之樣本脫落，於5ml trypticase soy broth(TSB)medium內35℃之溫度培養3小時；(3)以一般細菌DNA萃取方法萃取DNA；(4)以沙門菌DNA序列引子進行聚合脢連鎖反應及時定量方法檢測，並以CT值小於38為陽性判斷之域。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| TW100108448A TWI609082B (zh) | 2011-03-14 | 2011-03-14 | 一種無污染而快速檢測沙門菌的方法 |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| TW100108448A TWI609082B (zh) | 2011-03-14 | 2011-03-14 | 一種無污染而快速檢測沙門菌的方法 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| TW201237171A TW201237171A (en) | 2012-09-16 |
| TWI609082B true TWI609082B (zh) | 2017-12-21 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| TW100108448A TWI609082B (zh) | 2011-03-14 | 2011-03-14 | 一種無污染而快速檢測沙門菌的方法 |
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| Country | Link |
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| TW (1) | TWI609082B (zh) |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010049489A1 (en) * | 2008-10-29 | 2010-05-06 | Check-Points Holding B.V. | Method for fast detection and identification of micro-organisms |
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2011
- 2011-03-14 TW TW100108448A patent/TWI609082B/zh not_active IP Right Cessation
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010049489A1 (en) * | 2008-10-29 | 2010-05-06 | Check-Points Holding B.V. | Method for fast detection and identification of micro-organisms |
Non-Patent Citations (1)
| Title |
|---|
| 疫情報導,第22卷第5期，2006年5月25日,「非傷寒類沙門氏菌與曲狀桿菌單管多重即時PCR快速檢測系統之建立」 * |
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| Publication number | Publication date |
|---|---|
| TW201237171A (en) | 2012-09-16 |
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