Development and Evaluation of a Loop-mediated Isothermal Amplification Assay for the Rapid Detection and Identification of Pectobacterium carotovorum on Celery in the Field

Abstract

Pectobacterium carotovorum is the causal agent of bacterial soft rot in a wide range of vegetable host species. Once P. carotovorum infects the plant, the spread of the disease is difficult to control. In this study, a rapid and sensitive method based on loop-mediated isothermal amplification (LAMP) was developed for detecting P. carotovorum in celery with soft rot using a primer set designed from the pmrA conserved sequence of P. carotovorum. The specificity of the LAMP primer set for P. carotovorum was extensively validated on both P. carotovorum strains and non-target strains. The sensitivity was 1 pg of P. carotovorum genomic DNA, which demonstrated 10 times more sensitive than the conventional PCR assay. LAMP was also used to detect P. carotovorum in bacterial suspension. The lowest detection concentration was 104 CFU · mL−1. In addition, a LAMP assay, in conjunction with a crude DNA extraction method, was successfully performed on P. carotovorum-infected samples derived from both artificially and naturally infected plants. In summary, the LAMP assay established in this study constitutes a simple, sensitive, and rapid method for the detection of P. carotovorum, and has potential application in the control of celery soft rot disease through early detection.