Rapid Detection of Staphylococcus aureus in Milk and Pork via Immunomagnetic Separation and Recombinase Polymerase Amplification

Separation processes using immunomagnetic beads (IMBs) are advantageous for the rapid detection of Staphylococcus aureus (S. aureus). Herein, a novel method, based on immunomagnetic separation using IMBs and recombinase polymerase amplification (RPA), was employed to detect S. aureus strains in milk and pork. IMBs were formed by the carbon diimide method using rabbit anti-S. aureus polyclonal antibodies and superparamagnetic carboxyl-Fe3O4 MBs. The average capture efficiency for 2.5 to 2.5 x 10(5) (CFU)/mL gradient dilution of S. aureus with 6mg of IMBs within 60min were a range of 62.74 to 92.75%. The detection sensitivity of the IMBs-RPA method in artificially contaminated samples was 2.5 x 10(1) CFU/mL. The entire detection process was completed within 2.5 h, including bacteria capture, DNA extraction, amplification, and electrophoresis. Among 20 actual samples, one case of raw milk sample and two cases of pork samples were tested positive using the established IMBs-RPA method, which were verified by the standard S. aureus inspection procedure. Therefore, the novel method shows potential for food safety supervision owing to its short detection time, higher sensitivity, and high specificity. IMPORTANCE Our study established IMBs-RPA method, which simplified the steps of bacteria separation, shortened the detection time, and realized the convenient detection of S. aureus in milk and pork samples. IMBs-RPA method was also suitable for the detection of other pathogens, providing a new method for food safety monitoring and a favorable basis for rapid and early diagnosis of diseases.

Automated summary

In this study, a novel method based on immunomagnetic separation using immunomagnetic beads (IMBs) and recombinase polymerase amplification (RPA) was developed for the rapid detection of Staphylococcus aureus (S. aureus) in milk and pork. The IMBs-RPA method showed potential for food safety supervision and rapid early diagnosis of diseases due to its short detection time, high sensitivity, and specificity.