Use of restriction fragment length polymorphism analysis to differentiate strains of psychrophilic and psychrotrophic clostridia associated with 'blown pack' spoilage of vacuum-packed meats

Reference and meat strains of psychrophilic and psychrotrophic clostridia w ere differentiated using restriction fragment length polymorphism (RFLP) an alysis of genomic DNA (DNA-RFLP) and the polymerase chain reaction-amplifie d 16S rDNA gene (PCR-RFLP). Groupings obtained with PCR-RFLP were confirmed with 16S rDNA gene sequencing. DNA-RFLP resolved 19 of the 22 meat strains into 11 groups. Three meat strains were untypable using this method. All r eference strains representing different genotypic species could be distingu ished by the restriction patterns of 16S rDNA genes. With PCR-RFLP, the 22 meat strains produced eight distinct genotypes. 16S rDNA gene sequencing co nfirmed that each genotype was represented by a distinct sequence. PCR-RFLP restriction patterns of 15 meat strains matched those of one of two of the seven reference strains used. Seven meat strains whose RFLP restriction pa tterns of 16S rDNA genes differed from those of any reference strains proba bly represent four previously undescribed species. Although RFLP analysis o f the amplified 16S rDNA gene allowed differentiation of psychrophilic and psychrotrophic clostridia at the genotypic species level and below, compari son of PCR-RFLP patterns and 16S rDNA sequences of unknown clostridial isol ates with patterns and sequences of reference strains may not effect ready identification of these micro-organisms. The results of this study will be useful in diagnosis of the cause of premature spoilage of chilled vacuum-pa cked meats and in tracing spoilage-causing clostridia to their source(s) in the abattoir.