Development of gold nanoparticle-based visual assay for rapid detection of Escherichia coli specific DNA in milk of cows affected with mastitis

Abstract

For early diagnosis of infections, a rapid, sensitive and precise method to identify microbial DNA coming from veterinary clinical samples, such as milk of cows affected with mastitis, is fascinating. We developed an amplification-free visual assay for rapid and sensitive detection of specific DNA from Escherichia coli, particularly uid A gene encoding for beta-glucuronidase, isolated in milk of cows affected with mastitis, based on multiple gold nanoparticles (AuNPs) captured on a magnetic microbead surface, leading to plasmonic signal enhancement, thereby improving overall sensitivity of the assay. This test can be performed in 1–1.5 h after post template DNA preparation and can be visualized with naked eyes without the use of any expensive equipment. The visual assay detects a minimum microbial load of 102 CFU/μl and can identify bacterial DNA as low as 1 pg. This test provides a precise detection tool for E coli in clinical samples, such as milk of cows affected with mastitis, as a quick and user-friendly molecular detection approach.