Differentiation of Clostridium perfringens and Clostridium botulinum from non-toxigenic clostridia, isolated from prepared and frozen foods by PCR-DAN based methods.
During the elaboration process of prepared and frozen foods, Clostridium sp. have been reported. From those microorganisms, C. perfringens and C. botulinum may pose a high risk for the consumers. To avoid these pathogenic organisms an HACCP program should be implemented, but in addition sensitive and moderately time consuming microbiological methods for monitoring C. perfringens and C. boulinum should be established. In this work, an RFLP analysis of the 16S rDNA will be developed to differentiate C. perfringens from other Clostridium sp. In addition, a PCR protocol, will be assayed to detect C. botulinum. Both two methods will be compared with biochemical characterization by API system. The restriction analysis of the 16S rDNA with Taq I and Rsa I showed at least the same sensitivity to differentiate C. perfringens from clostridial isolates as biochemical identification. However, the former method takes only 8-10 h of analysis as compared with 24-48 h required for biochemical characterization. With the specific PCR protocol to detect C. botulinum a band of 1.1 kbp was obtained derived from the specific amplification of BoNT genes, taking 6-8 h for analysis. Both two molecular DNA based methods should be considered as verification techniques of pathogenic clostridia in the HACCP program.
Córdoba
M G
MG
Universidad de Extremadura, Escuela de Ingenierfas Agrarias, Ciencia y Tecnología de los Alimentos, Ctra. Cáceres s/n, E-06071 Badajoz, Spain. mdeguia@unex.es
Aranda
E
E
Medina
L M
LM
Jordano
R
R
Córdoba
J J
JJ
D003014:Clostridium botulinum / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D003016:Clostridium perfringens / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D004275:DNA, Ribosomal / Q000032:analysis*; D004587:Electrophoresis, Agar Gel; D005516:Food Microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D015183:Restriction Mapping; D013997:Time Factors
D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S