Quantitative duplex PCR of Clostridium botulinum types A and B neurotoxin genes

被引：28

作者：

Kasai, Yoshiaki ^{[1
]}

Kimura, Bon ^{[1
]}

Tajima, Yosuke ^{[1
]}

Fujii, Tateo ^{[1
]}

机构：

[1] Tokyo Univ Marine Sci & Technol, Fac Marine Sci, Dept Food Sci & Technol, Minato Ku, Tokyo 1088477, Japan

来源：

JOURNAL OF THE FOOD HYGIENIC SOCIETY OF JAPAN | 2007年 / 48卷 / 01期

关键词：

Clostridium botulinum; quantitative PCR; duplex; TaqMan; POLYMERASE-CHAIN-REACTION; LINKED-IMMUNOSORBENT-ASSAY; REVERSE TRANSCRIPTION-PCR; REAL-TIME PCR; MOUSE BIOASSAY; SODIUM-NITRITE; FOOD SAMPLES; TOXIN GENES; IDENTIFICATION; EXPRESSION;

D O I：

10.3358/shokueishi.48.19

中图分类号：

TS2 [食品工业];

学科分类号：

0832 ;

摘要：

A duplex quantitative polymerase chain reaction (PCR) assay for Clostridium botulinum types A and B was developed. The sensitivity and specificity of the assay were verified by using 6 strains of type A, 7 strains of type 13, and 14 genera of 42 non-C. botulinum types A and B strains, including C. botulinum types C, D, E, F, and G. In pure culture, the detection limit was 10(2) CFU/mL for type A and 10(3) CFU/mL for type B. In mushroom broth, increases in the amounts of C. botulinum types A and B could be monitored separately (the quantifiable range was 10(2) to 10(6) for type A and 10(2) to 10(7) for type 13) from each sample that contained a large number of background bacteria, and toxin could be detected much earlier than with mouse assay. These results suggest that duplex quantitative PCR methods are useful to detect and quantify C. botulinum types A and/or B toxin genes.

引用

页码：19 / 26

页数：8

共 53 条

[1]

Akbulut D, 2004, Foodborne Pathog Dis, V1, P247, DOI 10.1089/fpd.2004.1.247

[2] Improvement in laboratory diagnosis of wound botulism and tetanus among injecting illicit-drug users by use of real-time PCR assays for neurotoxin gene fragments [J].

Akbulut, D ;

Grant, KA ;

McLauchlin, J .

JOURNAL OF CLINICAL MICROBIOLOGY, 2005, 43 (09) :4342-4348

[3] Selection of primers for specific detection of Clostridium botulinum types B and E neurotoxin genes using PCR method [J].

Alsallami, AA ;

Kotlowski, R .

INTERNATIONAL JOURNAL OF FOOD MICROBIOLOGY, 2001, 69 (03) :247-253

[4] IN-SITU CHARACTERIZATION OF CLOSTRIDIUM-BOTULINUM NEUROTOXIN SYNTHESIS AND EXPORT [J].

CALL, JE ;

COOKE, PH ;

MILLER, AJ .

JOURNAL OF APPLIED BACTERIOLOGY, 1995, 79 (03) :257-263

[5] GENE PROBES FOR IDENTIFICATION OF THE BOTULINAL NEUROTOXIN GENE AND SPECIFIC IDENTIFICATION OF NEUROTOXIN TYPE-B, TYPE-E, AND TYPE-F [J].

CAMPBELL, KD ;

COLLINS, MD ;

EAST, AK .

JOURNAL OF CLINICAL MICROBIOLOGY, 1993, 31 (09) :2255-2262

[6] A highly sensitive immuno-polymerase chain reaction assay for Clostridium botulinum neurotoxin type A [J].

Chao, HY ;

Wang, YC ;

Tang, SS ;

Liu, HW .

TOXICON, 2004, 43 (01) :27-34

[7] Detection of biological threat agents by real-time PCR: Comparison of assay performance on the RAPID, the LightCycler, and the smart cycler platforms [J].

Christensen, DR ;

Hartman, LJ ;

Loveless, BM ;

Frye, MS ;

Shipley, MA ;

Bridge, DL ;

Richards, MJ ;

Kaplan, RS ;

Garrison, J ;

Baldwin, CD ;

Kulesh, DA ;

Norwood, DA .

CLINICAL CHEMISTRY, 2006, 52 (01) :141-145

[8] A SIMPLE METHOD FOR DNA PURIFICATION FROM PERIPHERAL-BLOOD [J].

CIULLA, TA ;

SKLAR, RM ;

HAUSER, SL .

ANALYTICAL BIOCHEMISTRY, 1988, 174 (02) :485-488

[9] Expression of botulinum neurotoxins A and E, and associated non-toxin genes, during the transition phase and stability at high temperature: analysis by quantitative reverse transcription-PCR [J].

Couesnon, A ;

Raffestin, S ;

Popoff, MR .

MICROBIOLOGY-SGM, 2006, 152 :759-770

[10]

Craven KE, 2002, J AOAC INT, V85, P1025

← 1 2 3 4 5 6 →