The Use of Gel Electrophoresis to Separate Multiplex Polymerase Chain Reaction Amplicons Allows for the Easy Identification and Assessment of the Spread of Toxigenic Clostridioides difficile Strains.

Clostridioides difficile is a common etiological factor of hospital infections, which, in extreme cases, can lead to the death of patients. Most strains belonging to this bacterium species synthesize very dangerous toxins: toxin A (TcdA) and B (TcdB) and binary toxin (CDT). The aim of this study was to assess the suitability of agarose gel electrophoresis separation of multiplex PCR amplicons to investigate the toxinogenic potential of C. difficile strains. Additionally, the frequency of C. difficile toxin genes and the genotypes of toxin-producing strains were determined. Ninety-nine C. difficile strains were used in the detection of the presence of genes encoding all of these toxins using the multiplex PCR method. In 85 (85.9%) strains, the presence of tcdA genes encoding enterotoxin A was detected. In turn, in 66 (66.7%) isolates, the gene encoding toxin B (tcdB) was present. The lowest number of strains tested was positive for genes encoding a binary toxin. Only 31 (31.3%) strains possessed the cdtB gene and 22 (22.2%) contained both genes for the binary toxin subunits (the cdtB and cdtA genes). A relatively large number of the strains tested had genes encoding toxins, whose presence may result in a severe course of disease. Therefore, the accurate diagnosis of patients, including the detection of all known C. difficile toxin genes, is very important. The multiplex PCR method allows for the quick and accurate determination of whether the tested strains of this bacterium contain toxin genes. Agarose gel electrophoresis is a useful tool for visualizing amplification products, allowing one to confirm the presence of specific C. difficile toxin genes as well as investigate their dissemination for epidemiological purposes.