Characterization of the brewery spoilage bacterium Obesumbacterium proteus by automated ribotyping and development of PCR methods for its biotype 1.
OBJECTIVE
To study the ability of automated ribotyping to characterize Obesumbacterium proteus and Hafnia alvei, to design primers and to evaluate standard end-point and real-time PCR for the detection of O. proteus biotype 1 in beer and in brewers's yeast-containing samples.


RESULTS
Automated ribotyping was carried out using the standard method with EcoRI and PvuII. The digestions with both enzymes clearly differentiated O. proteus biotypes 1 and 2 and H. alvei. PCR primers were designed according to the 16S rRNA gene sequence of the O. proteus type strain. Two primer sets (Obs137-Obs558 and Obs137-Obs617) detected O. proteus biotype 1 and H. alvei but not O. proteus biotype 2 or other tested beer spoilage bacteria (40 species) in the end-point and real-time PCR, indicating their high specificity. The detection limit for O. proteus was 160-1600 CFU 100 ml(-1) beer in the end-point PCR reactions and < or =160 CFU 100 ml(-1) beer in the real-time PCR reactions. More cells (from 16 to 3200) were needed for detection in the presence of brewer's yeast cells.


CONCLUSIONS
Automated ribotyping is a useful tool to characterize and identify O. proteus and H. alvei isolates. The designed primers are suitable for the rapid detection of O. proteus biotype 1 and H. alvei in brewery samples by PCR.


CONCLUSIONS
Automated ribotyping and PCR could improve microbiological quality control in breweries by facilitating the detection, identification and tracing of spoilage bacteria.
Koivula
T T
TT
VTT Biotechnology, Espoo, Finland.
Juvonen
R
R
Haikara
A
A
Suihko
M-L
ML
D001515:Beer / Q000382:microbiology; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis; D004755:Enterobacteriaceae / Q000235:genetics / Q000302:isolation & purification*; D005511:Food Handling; D005516:Food Microbiology; D020611:Hafnia alvei / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*; D021521:Ribotyping / Q000379:methods*; D012441:Saccharomyces cerevisiae
D017931:DNA Primers; D004269:DNA, Bacterial