Zhao Y, Zhu A, Tang J, Tang C, Chen J
College of Life Science and Technology, Southwest University for Nationalities, Chengdu, China.
Lett Appl Microbiol. 2017 Jul;65(1):27-34. doi: 10.1111/lam.12751. Epub 2017 May 30.
Staphylococcus aureus produces a wide variety of staphylococcal enterotoxins (SEs, SEA to SEX), which are responsible for staphylococcal food poisoning. This study is aimed to establish a system to detect staphylococcal enterotoxin M (SEM) protein in food matrixes. In the present study, sem gene was characterized in a S. aureus isolate H4 associated with food poisoning. The amino acid sequence of the deduced SEM protein was same as that of previously identified SEM from S. aureus 04-02981. Subsequently, mature SEM protein was expressed in Escherichia coli BL21 (DE3) cells and purified with a Ni-NTA spin column. The polyclonal and monoclonal antibody against it were prepared. Using these antibodies, a highly sensitive, specific sandwich enzyme-linked immunosorbent assay (ELISA) system was developed capable of detecting SEM in milk, meat and rice. Cross-reactivity with SEB, SEI and SEK in this method was insignificant. Quantification of SEM secretion in vitro using this novel capture ELISA revealed that SEM was mainly secreted during the transition from the exponential to the stationary phase. Furthermore, sem gene and SEM protein production were screened by PCR and the developed ELISA system. The results indicated that there were two SEM+ strains of 19 S. aureus isolates originating in cold dishes and humans suffering from food poisoning. The investigations make it possible to assess SEM in food hygiene supervision in near future.
Staphylococcal enterotoxins (SE) are the main causative agents of staphylococcal food poisoning. Unlike classical SEs (SEA to SEE), the relationship between newly identified SEs (SEG to SEX) and staphylococcal food poisoning has not been clearly elucidated. Recently, mild emetic potential of staphylococcal enterotoxin M (SEM) has been demonstrated, which indicated that SEM might be associated with food poisoning. However, there is currently no commercial enzyme-linked immunosorbent assay (ELISA) kit available for immunological detection of it. Therefore, we developed a highly sensitive, specific sandwich ELISA system and assayed SEM in food matrixes. This assay facilitates the study of SEM expression in vitro and in vivo. In addition, the investigations would be helpful in addressing the relative incidence of SEM+ strains in near future.
金黄色葡萄球菌可产生多种葡萄球菌肠毒素（SEs，从SEA到SEX），这些毒素是导致葡萄球菌食物中毒的原因。本研究旨在建立一种在食品基质中检测葡萄球菌肠毒素M（SEM）蛋白的系统。在本研究中，对一株与食物中毒相关的金黄色葡萄球菌分离株H4中的sem基因进行了鉴定。推导的SEM蛋白的氨基酸序列与先前从金黄色葡萄球菌04 - 02981中鉴定出的SEM相同。随后，在大肠杆菌BL21（DE3）细胞中表达成熟的SEM蛋白，并使用镍 - 氮川三乙酸（Ni - NTA）旋转柱进行纯化。制备了针对它的多克隆抗体和单克隆抗体。利用这些抗体，开发了一种高灵敏度、特异性的夹心酶联免疫吸附测定（ELISA）系统，该系统能够检测牛奶、肉类和大米中的SEM。该方法与SEB、SEI和SEK的交叉反应不明显。使用这种新型捕获ELISA对体外SEM分泌进行定量分析表明，SEM主要在从指数生长期向稳定期转变期间分泌。此外，通过PCR和开发的ELISA系统对sem基因和SEM蛋白的产生进行了筛选。结果表明，在源自冷盘和食物中毒患者的19株金黄色葡萄球菌分离株中有两株SEM阳性菌株。这些研究使得在不久的将来对食品卫生监督中的SEM进行评估成为可能。
葡萄球菌肠毒素（SE）是葡萄球菌食物中毒的主要致病因素。与经典的SEs（从SEA到SEE）不同，新鉴定的SEs（从SEG到SEX）与葡萄球菌食物中毒之间的关系尚未明确阐明。最近，已证明葡萄球菌肠毒素M（SEM）具有轻微的催吐潜力，这表明SEM可能与食物中毒有关。然而，目前尚无用于其免疫检测的商业酶联免疫吸附测定（ELISA）试剂盒。因此，我们开发了一种高灵敏度、特异性的夹心ELISA系统，并在食品基质中检测SEM。该测定有助于研究SEM在体外和体内的表达。此外，这些研究将有助于在不久的将来解决SEM阳性菌株的相对发生率问题。
