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

被引:44
作者
Couesnon, A [1 ]
Raffestin, S [1 ]
Popoff, MR [1 ]
机构
[1] Inst Pasteur, Unite Bacteries Anaerobies & Toxines, Paris, France
来源
MICROBIOLOGY-SGM | 2006年 / 152卷
关键词
D O I
10.1099/mic.0.28561-0
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
Production of botulinum neurotoxin A (BoNT/A) and associated non-toxic proteins (ANTPs), which include a non-toxic non-haemagglutinin (NTNH/A) as well as haemagglutinins (HAs), was found previously to be dependent upon an RNA polymerase alternative sigma factor (BotR/A). Expression of the botR/A, bont/A and antp genes, monitored by reverse transcription and real-time PCR analysis, occurred concomitantly at the transition between the exponential and stationary growth phases of Clostridium botulinum A. The botR/A expression level was about 100-fold less than those of the bont/A and antp genes. Therefore, BotR/A is an alternative sigma factor controlling the botulinum A locus genes during the transition phase. The highest toxin concentration was released into the culture supernatant 12 h after maximum expression of the botR/A, bont/A and antp genes, without any apparent bacterial lysis. Toxin levels were then stable over 5 days in cultures at 37 degrees C, whereas a dramatic decrease in lethal activity was observed between 24 and 48 h in cultures at 44 degrees C. High temperature did inhibit transcription, since expression levels of the botR/A, bont/A and antp genes were similar in cultures at 37 and 44 degrees C. However, incubation at 44 degrees C triggered a calcium-dependent protease that degraded BoNT/A and NTNH/A, but not HAs. In C. botulinum E, which contains no gene related to botR, the bont/E and p47 genes were also expressed during the transition phase, and no protease activation at 44 degrees C was evident.
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页码:759 / 770
页数:12
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