Abstract
Over one hundred bacterial species have been determined to comprise the human microbiota in a healthy individual. Bacteria including Escherichia coli, Bacillus cereus, Clostridioides difficile, and Vibrio parahaemolyticus are found inside of the human body and B. cereus and E. coli are also found on the skin. These bacteria can act as human pathogens upon ingestion of contaminated food or water, if they enter an open wound, or antibiotics, and environment or stress can alter the microbiome. In this study, we present new polymerase chain reaction (PCR) high-resolution melt (HRM) assays to detect and identify the above microorganisms. Amplified DNA from C. difficile, E. coli, B. cereus, and V. parahaemolyticus melted at 80.37 ± 0.45 °C, 82.15 ± 0.37 °C, 84.43 ± 0.50 °C, and 86.74 ± 0.65 °C, respectively. A triplex PCR assay was developed to simultaneously detect and identify E. coli, B. cereus, and V. parahaemolyticus, and cultured microorganisms were successfully amplified, detected, and identified. The assays demonstrated sensitivity, specificity, reproducibility, and robustness in testing.
Highlights
Healthy individuals have a diverse microbiota and contain over one hundred bacterial species [1]
Escherichia coli, Bacillus cereus, Clostridioides difficile, and Vibrio parahaemolyticus are found inside of the human body and B. cereus and E. coli are found on the skin
These and other bacteria are listed by the United States Center for Disease Control and Prevention (CDC) as notable foodborne pathogens that cause the majority of such illness, along with other pathogens [2]
Summary
Healthy individuals have a diverse microbiota and contain over one hundred bacterial species [1]. Escherichia coli, Bacillus cereus, Clostridioides difficile, and Vibrio parahaemolyticus are found inside of the human body and B. cereus and E. coli are found on the skin. C. difficile has been most problematic in individuals with dysbiosis or in hospital settings, but recent research indicates that contaminated food products with spores may lead to C. difficile infection [4,5]. These and other bacteria are listed by the United States Center for Disease Control and Prevention (CDC) as notable foodborne pathogens that cause the majority of such illness, along with other pathogens [2]
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