Ultrasensitive Detection of Pathogenic Bacteria by Targeting High Copy Signature Genes

Abstract

Bacterial load in clinical samples is relatively low and difficult to detect. Improvements in assay sensitivity will greatly reduce false negative results and contribute to more accurate diagnoses. In the present study, we present a new strategy to improve the sensitivity of a nucleic acid assay by detecting the presence of a multi-copy gene. By using Brucella as a test model, we screened the genome and identified IS711 as a multiple copy gene. Distribution analysis of insertion sequence IS711 among different species and strains showed that each of the strains have 5 to 13 copies of IS711. Compared with the BMEI1001, BMEI0775 and BMEI0027, the assays of high copy genes IS711 showed higher sensitivity and is an ideal high copy signature gene for Brucella. Detection of clinical samples with assays targeting the signature genes showed that IS711 exist in higher concentrations than BMEI1001, BMEI0775 and BMEI0027. In addition, IS711 assay is more sensitive than other signature genes assay. Analysis of several other pathogenic bacteria successfully identified high copy number genes that could be used as signature genes. Therefore, this strategy of targeting high copy signature genes represents a universal strategy for the ultrasensitive detection of bacteria.