Molecular detection of toxins A, B and E of Clostridium botulinum in botulism suspected specimens

BACKGROUND AND OBJECTIVESBotulism is caused by Clostridium botulinum toxins (BoNT), which is produced by improper use of canned goods, dairy products, and fish. C. botulinum toxin has seven distinct serotypes (A-B-C-D-E-F-G) that have a similar structure but differ antigenically. Serotypes A, B, and E are associated with human clinical cases. Today, in Iran, a definitive diagnosis of botulism is made by identifying the bacterial toxin using the laboratory mice (mouse bioassay). As botulism is a life-threatening condition, a rapid diagnosis is required for successful therapy. The rapid and accurate disease diagnosis techniques and methods play an effective role in saving the lives of those affected. In addition to being fast, accurate, and economical, these diagnostic methods must have high specificity and sensitivity. The purpose of this research is to use the Real-time PCR molecular method to detect C. botulinum bacteria in a clinical sample.MATERIALS AND METHODSIn ۶ months, ۵۰ suspected clinical samples (feces, patient's stomach secretions, and food samples), sent to the Pasteur Institute of Iran, were collected. After ۴۸ hours of cultivation in Cook-meat medium and under anaerobic conditions, DNA was extracted and SYBR Green Real-time PCR was done for the genes of toxins A, B, and E and the results were analyzed.RESULTS AND DISCUSSIONIn this research, out of the ۵۰ samples, ۶۸% were genetically positive. The frequency of toxins B, E, BE, A, and AB of C. botulinum was ۴۴%, ۳۲%, ۲۲%, ۱۶%, and ۱۰%, respectively. Mouse bioassay was positive for ۶ specimens. The difference between the results of the SYBR Green Real-time test compared to the Mouse bioassay can be due to the high sensitivity of the molecular test to the phenotype as well as the lack of expression of toxin genes in some samples.CONCLUSIONDue to the high sensitivity and accuracy of the SYBR Green Real-time method and the rapid diagnosis of this method, it is recommended to use this method along with the Mouse bioassay method to detect botulism toxin.