An alternative real-time PCR method for the detection of thermotolerant Bacillus sensu lato contaminants in naturally-contaminated gelatine.
OBJECTIVE
Comparison of an internally-controlled real-time PCR assay with the current plate-based assay for the detection of Bacillus sensu lato contaminants in gelatine.


RESULTS
A comprehensive TaqMan probe was designed allowing the real-time PCR assay to be fully inclusive for the gelatine-contaminating Bacillus s.l. species. An internal amplification control was implemented at 500 copies per reaction without impact on target detection. Specific and selective detection of target cells was achieved with a quick and simple DNA preparation procedure. No significant difference (Kappa value = 0.94) was observed between the performance of the real-time PCR and the current plate-based method on naturally contaminated gelatines (n = 162). Relative accuracy, relative sensitivity and relative specificity were 97.5%.


CONCLUSIONS
The real-time PCR assay is an adequate alternative of the current plate-based assay.


CONCLUSIONS
The real-time PCR assay decreased the time between sample collection and result from 2 days to 2 h. The gelatine-producing industry can ensure gelatine quality in a much faster way.
Laboratory of Microbiology, Department of Biochemistry and Microbiology, Ghent University, Ghent, Belgium. Rieka.Reekmans@UGent.be
R
R
Reekmans
C
C
Van den Plas
P
P
Stevens
T
T
Vervust
P
P
De Vos
D000818:Animals; D001407:Bacillus / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D005506:Food Contamination / Q000032:analysis*; D005780:Gelatin / Q000032:analysis*; D006358:Hot Temperature; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D005780:Gelatin