Use of a species-specific multiplex PCR for the identification of pediococci.
In this study, the 23S rRNA genes of nine different Pediococcus type strains were sequenced. By using a multiple sequence alignment with 23S rDNA sequences of related lactic acid bacteria two primer pairs were constructed, one for the general identification of the genus Pediococcus and one for the identification of the atypical species, P. dextrinicus. Furthermore, a primer set for a rapid multiplex PCR identification of the eight typical Pediococcus species was developed. With this technique, the species P. damnosus, P. parvulus, P. inopinatus, P. cellicola, P. pentosaceus, P. acidilactici, P. claussenii, and P. stilesii could be discriminated simultaneously in a single PCR. Experiments with inoculated grape musts showed that the detection limit was 10 cells ml(-1). The multiplex PCR assay was tested by the usage of 62 Pediococcus strains from different culture collections and 47 strains recently isolated from German wines and musts. In addition, contaminations with P. parvulus and P. damnosus could be detected after purification of DNA from spoilt wine samples. The method demonstrates a rapid and easy to handle tool for the species affiliation of pediococci in beverages and food samples.
Institute of Microbiology and Wine Research, Johannes Gutenberg-University of Mainz, Becherweg 15, D-55099 Mainz, Germany. pfannebe@uni-mainz.de
Jens
J
Pfannebecker
Jürgen
J
Fröhlich
D001483:Base Sequence; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D005506:Food Contamination / Q000032:analysis*; D010376:Pediococcus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012338:RNA, Ribosomal, 23S / Q000235:genetics*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D014920:Wine / Q000382:microbiology*
D004269:DNA, Bacterial; D012338:RNA, Ribosomal, 23S