乙醇促进副溶血性弧菌直接耐热溶血素的基因表达

研究乙醇对副溶血性弧菌(Vibrio parahaemolyticus)直接耐热溶血素(thermostable direct hemolysin,TDH)产生和它的编码基因tdh表达的影响。以0.5%、1.0%、2.0%、4.0%、8.0%乙醇处理2株带有tdh副溶血性弧菌ATCC33847和SZ32,研究对副溶血性弧菌有氧或者无氧生长的影响。选取1.0%乙醇处理来研究TDH产量和tdh表达变化。使用TDH抗血清试剂盒测定副溶血性弧菌培养液上清中TDH水平;使用荧光定量PCR方法分析tdh基因表达状况。低浓度(0.5%、1.0%、2.0%)乙醇存在时,副溶血弧菌菌株的生长未受到显著影响,乙醇浓度4.0%时副溶血性弧菌生长受到明显抑制,8%时未见有细菌生长。1.0%乙醇处理副溶血性弧菌培养液上清中TDH水平较未处理显著上升。胞内tdh表达水平升高,ATCC33847在有氧和无氧时分别升高到6.6倍和5.7倍,SZ32在有氧和无氧时分别升高到5.9倍和8.6倍。乙醇能够促进tdh基因表达从而使得TDH蛋白产量升高。本研究还比较了甲醇、乙醇、正丙醇对tdh表达的影响,发现它们对tdh表达均具有促进作用但相互之间没有明显差异。

Ethanol Treatment Enhances Expression of Thermostable Direct Hemolysin Gene by Vibrio parahaemolyticus

The effect of ethanol on thermostable direct hemolysin (TDH) production and corresponding tdh gene expression by Vibrio parahaemolyticus was studied. Two tdh-positive V. parahaemolyticus strains, ATCC33847 and SZ32 were separately treated with 0.5%, 1.0%, 2.0%, 4.0%, and 8.0% ethanol solutions. The effects on aerobic or anaerobic growth of V. parahaemolyticus were monitored, while changes in TDH production and tdh gene expression were studied with 1.0% ethanol treatment. TDH concentration in V. parahaemolyticus culture supernatant was quantitatively analyzed using a TDH detection kit (KAP-RPLA, Denka Seiken Co., Japan), while tdh gene expression was analyzed by real-time fluorescence-based quantitative polymerase chain reaction. The results showed that V. parahaemolyticus growth was not influenced by the presence of ethanol at low concentrations (0.5%, 1.0%, and 2.0%). On the other hand, 4.0% and 8.0% ethanol caused significant and total inhibition of bacterial growth, respectively. With ethanol treatment, TDH titers of culture supernatants showed a significant increase under aerobic as well as anaerobic conditions and intracellular tdh expression was also elevated. Moreover, ATCC33847 strain showed an increase in tdh expression by 6.6- and 5.7-fold, while SZ32 strain showed an increase by 5.9- and 8.6-fold under aerobic and anaerobic conditions, respectively. Thus, ethanol treatment enhanced tdh gene expression, which may have resulted in an increase in TDH production. The effects of methanol, ethanol, and n-propanol on tdh gene expression were also compared and the results showed increased tdh gene expression by all three alcohols, however, the effects were not significantly different.