Biomedical subjects

T T Koivula

Publications and source records attributed to T T Koivula.

6 recordsLinked to original sources

PubMed2006

Characterization of the brewery spoilage bacterium Obesumbacterium proteus by automated ribotyping and development of PCR methods for its biotype 1.

AIMS: 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. METHODS AND 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. SIGNIFICANCE AND IMPACT OF THE STUDY: Automated ribotyping and PCR could improve microbiological quality control in breweries by facilitating the detection, identification and tracing of spoilage bacteria.

Beer↗

PubMed1997

Diversity within bacterial isolates hybridizing with Comamonas probe ppT.

Diversity within bacteria isolated from activated sludge and the Baltic sea and recognized by the rRNA targeted oligonucleotide probe ppT (designed by BRAUN-HOWLAND et al., 1993 to recognize C. testosteroni) was studied. The partial nucleotide sequences of 16S genes of the isolates revealed that the activated sludge and Baltic sea isolates each formed a separate group distinct from C. testosteroni. The analysis of phage and bacteriocin sensitivity as well as whole cell protein patterns revealed the same grouping, but in these characters also intragroup variation was observed. As a conclusion, neither of the studied groups were C. testosteroni, but they probably belong to two previously unknown species common in activated sludge or the Baltic sea.

Bacterial Proteins↗

PubMed1997

Comparison of partial 16S rRNA gene sequences obtained from activated sludge bacteria.

The cultivated and uncultivated bacterial communities of an activated sludge plant were studied. Two samples were taken and a total of 516 bacterial isolates were classified into groups using their whole-cell protein patterns. The distribution of bacteria into protein-pattern groups differed significantly between the two samples, suggesting variation in culturable bacterial flora. Partial 16S rRNA gene sequences were determined for representatives of the commonest protein-pattern groups. Most of the sequences obtained were previously unknown, but relatively closely related to known sequences of organisms belonging to the alpha, beta or gamma subclasses of the proteobacteria, the first two subclasses being predominant. This classification of bacteria isolated on a diluted nutrient-rich medium differed from recent culture-dependent studies using nutrient-rich media. The uncultivated bacterial community was studied by analyzing ten partial 16S rRNA gene sequences cloned directly from activated sludge. None of the cloned sequences was identical to those determined for culturable organisms; or to those in the GenBank database. They were, however, related to the alpha or beta subclasses of the proteobacteria, or to the gram-positive bacteria with a high G + C DNA content.

Bacteria↗

PubMed1996

Bacterial diversity at surface water in three locations within the Baltic sea as revealed by culture-dependent molecular techniques.

Diversity of culturable bacteria inhabiting the Baltic sea surface waters was studied in three separate locations. Based on electrophoretically separated whole cell proteins the number of operational taxonomic units (OTU) within each sampling location was high. Most of the OTUs were unique to single locations. Within each sampling location 8-22% of isolates belonged to a single OTU. Rarefaction analysis revealed that the bacterial community was more divergent at a polluted location than at clean areas. Also the most common OTUs were different in clean locations compared to the polluted site suggesting that both diversity and species composition of the bacterial community is greatly affected by pollution. The partial 16S rRNA gene sequences of the isolates of the most common OTUs are unique. Intragroup variation and an OTU-specific bacteriocin system was observed among the isolates of the second common OTU. The bacteriocin activity was linked to restriction fragment length polymorphism grouping, although additional variation correlating to geographic origin of isolates was observed.

Bacteria↗

PubMed1994

Hormone-sensitive lipase is closely related to several bacterial proteins, and distantly related to acetylcholinesterase and lipoprotein lipase: identification of a superfamily of esterases and lipases.

We have sequenced a gene from Bacillus acidocaldarius which encodes an open reading frame (ORF3) of 310 amino acids. The ORF3 was found to be related to the mammalian hormone-sensitive lipase (HSL). Searching the protein data base revealed five other bacterial proteins related to the HSL. Upon further sequence comparisons this HSL-group was found to be related to the family of carboxylesterases, and to a family of lipases (lipoprotein, hepatic and pancreatic lipases). The evolutionary relationship of these serine-dependent hydrolytic enzymes has not been studied previously, and it has not been known that these proteins belong to the same superfamily. Finally, the alignment of the HSL with the bacterial proteins allowed us to infer the location of the hormone-sensitive regulatory domain of the HSL-protein.

Acetylcholinesterase↗

PubMed1993

Cloning and sequencing of a gene encoding acidophilic amylase from Bacillus acidocaldarius.

Two starch-degrading enzymes produced by Bacillus acidocaldarius (renamed as Alicyclobacillus acidocaldarius) were identified. According to SDS-PAGE, the apparent molecular masses of the enzymes were 90 and 160 kDa. Eight peptide fragments and the N-terminal end of the 90 kDa polypeptide were sequenced. An oligonucleotide, based on the amino acid sequence of a peptide fragment of the 90 kDa protein, was used to screen a lambda gt10 bank of B. acidocaldarius, and the region encoding the 90 kDa protein was cloned. Unexpectedly, the ORF continued upstream of the N terminus of the 90 kDa protein. The entire ORF was 1301 amino acids (aa) long (calculated molecular mass 140 kDa) and it was preceded by a putative ribosomal binding site and a promoter. Computer analysis showed that the 1301 aa protein was closely related to an alpha-amylase-pullulanase of Clostridium thermohydrosulfuricum. We suggest that the starch-degrading 160 kDa protein of B. acidocaldarius is an alpha-amylase-pullulanase, and the 90 kDa protein is a cleavage product of the 160 kDa protein. Another ORF, apparently in the same transcription unit, was found downstream from the amylase gene. It encoded a protein that was closely related to the maltose-binding protein of Escherichia coli.

ATP-Binding Cassette Transporters↗