Development of a qPCR detection approach for pathogenic Burkholderia cenocepacia associated with fresh vegetables

Aixin Liu; Kate Phillips; Jiayuan Jia; Peng Deng; Dunhua Zhang; Sam Chang; Shi-En Lu

doi:10.1016/j.fm.2023.104333

Development of a qPCR detection approach for pathogenic Burkholderia cenocepacia associated with fresh vegetables

Food Microbiol. 2023 Oct:115:104333. doi: 10.1016/j.fm.2023.104333. Epub 2023 Jul 6.

Authors

Aixin Liu¹, Kate Phillips², Jiayuan Jia², Peng Deng², Dunhua Zhang³, Sam Chang⁴, Shi-En Lu⁵

Affiliations

¹ Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, MS, 39762, USA; College of Plant Protection, Shandong Agricultural University, Shandong, 271018, China.
² Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, MS, 39762, USA.
³ Aquatic Animal Health Research Unit, USDA-ARS, 990 Wire Road, Auburn, AL, 36832, USA.
⁴ Coastal Research and Extension Center, Seafood Processing Laboratory, Department of Food Science, Nutrition and Health Promotion, Mississippi State University, Pascagoula, MS, 39567, USA.
⁵ Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, MS, 39762, USA. Electronic address: sl332@msstate.edu.

PMID: 37567623
DOI: 10.1016/j.fm.2023.104333

Abstract

Natural environment serves as a reservoir for Burkholderia cepacia complex organisms, including the highly transmissible opportunistic human pathogen B. cenocepacia. Currently, there is a lack of an effective and quantitative method for B. cenocepacia detection in fresh food and other environmental niches. A quantitative real-time PCR (qPCR) detection method for B. cenocepacia bacteria was established in this study and validated using artificially inoculated fresh vegetable samples. Genome-wide comparative methods were applied to identify target regions for the design of species-specific primers. Assay specificity was measured with 12 strains of closely related Burkholderia bacteria and demonstrated the primer pair BCF6/R6 were 100% specific for detection of B. cenocepacia. The described qPCR assay evaluated B. cenocepacia with a 2 pg μl^-1 limit of detection and appropriate linearity (R² = 0.999). In 50 samples of experimentally infected produce (lettuce, onion, and celery), the assay could detect B. cenocepacia as low as 2.6 × 10² cells in each sample equal to 1 g. The established qPCR method quantitatively detects B. cenocepacia with high sensitivity and specificity, making it a promising technique for B. cenocepacia detection and epidemiological research on B. cepacia complex organisms from fresh vegetables.

Keywords: Burkholderia cenocepacia; Detection; Food safety; Fresh vegetables; Quantitative real-time PCR.

Published by Elsevier Ltd.

MeSH terms

Burkholderia cenocepacia*
Burkholderia cepacia complex*
Humans
Vegetables