Abstract
Simple and rapid detection of Burkholderia cepacia complex (BCC) bacteria, a common cause of pharmaceutical product recalls, is essential for consumer safety. In this study, we developed and evaluated a ribB-based colorimetric loop-mediated isothermal amplification (LAMP) assay for the detection of BCC in (i) nuclease-free water after 361 days, (ii) 10 μg/mL chlorhexidine gluconate (CHX) solutions, and (iii) 50 μg/mL benzalkonium chloride (BZK) solutions after 184 days. The RibB 5 primer specifically detected 20 strains of BCC but not 36 non-BCC strains. The limit of detection of the LAMP assay was 1 pg/μL for Burkholderia cenocepacia strain J2315. Comparison of LAMP with a qPCR assay using 1440 test sets showed higher sensitivity: 60.6% in nuclease-free water and 42.4% in CHX solution with LAMP vs. 51.3% and 31.1%, respectively, with qPCR. These results demonstrate the potential of the ribB-based LAMP assay for the rapid and sensitive detection of BCC in pharmaceutical manufacturing.
Highlights
The genus Burkholderia comprises >100 species of Gram-negative, non-spore-forming β-proteobacteria
We demonstrated the potential of droplet digital polymerase chain reaction and flow cytometry as more sensitive alternatives to culture-based methods to detect Burkholderia cepacia complex (BCC) in autoclaved nuclease-free water and antiseptic samples [12]
Using GGM, we identified 206 BCCspecific clusters which are only present in BCC genomes
Summary
The genus Burkholderia comprises >100 species of Gram-negative, non-spore-forming β-proteobacteria. Within this genus, the Burkholderia cepacia complex (BCC) includes 24 closely related species of opportunistic pathogens that are commonly found in natural environments [1,2,3,4]. The Burkholderia cepacia complex (BCC) includes 24 closely related species of opportunistic pathogens that are commonly found in natural environments [1,2,3,4] These species may contaminate various drug products and are a public health concern. This led the United States Pharmacopeia (USP) to publish General Chapter (60) (Microbiological examination of non-sterile products: Tests for Burkholderia cepacia complex) (effective date: 1 December 2019) to detect the presence of BCC species in pharmaceutical substances and non-sterile pharmaceutical products [8]
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