Affiliation:
1. Nestlé Research Center, Vers-Chez-Les-Blanc, 1000 Lausanne 26, Switzerland
2. Alimentary Pharmabiotic Centre
3. Department of Microbiology
4. National Food Biotechnology Centre, National University of Ireland, Cork, Ireland
Abstract
ABSTRACT
The
atp
operon is highly conserved among eubacteria, and it has been considered a molecular marker as an alternative to the 16S rRNA gene. PCR primers were designed from the consensus sequences of the
atpD
gene to amplify partial
atpD
sequences from 12
Bifidobacterium
species and nine
Lactobacillus
species. All PCR products were sequenced and aligned with other
atpD
sequences retrieved from public databases. Genes encoding the subunits of the F
1
F
0
-ATPase of
Bifidobacterium lactis
DSM 10140 (
atpBEFHAGDC
) were cloned and sequenced. The deduced amino acid sequences of these subunits showed significant homology with the sequences of other organisms. We identified specific sequence signatures for the genus
Bifidobacterium
and for the closely related taxa
Bifidobacterium lactis
and
Bifidobacterium animalis
and
Lactobacillus gasseri
and
Lactobacillus johnsonii
, which could provide an alternative to current methods for identification of lactic acid bacterial species. Northern blot analysis showed that there was a transcript at approximately 7.3 kb, which corresponded to the size of the
atp
operon, and a transcript at 4.5 kb, which corresponded to the
atpC
,
atpD
,
atpG
, and
atpA
genes. The transcription initiation sites of these two mRNAs were mapped by primer extension, and the results revealed no consensus promoter sequences. Phylogenetic analysis of the
atpD
genes demonstrated that the
Lactobacillus atpD
gene clustered with the genera
Listeria
,
Lactococcus
,
Streptococcus
, and
Enterococcus
and that the higher G+C content and highly biased codon usage with respect to the genome average support the hypothesis that there was probably horizontal gene transfer. The acid inducibility of the
atp
operon of
B. lactis
DSM 10140 was verified by slot blot hybridization by using RNA isolated from acid-treated cultures of
B. lactis
DSM 10140. The rapid increase in the level of
atp
operon transcripts upon exposure to low pH suggested that the ATPase complex of
B. lactis
DSM 10140 was regulated at the level of transcription and not at the enzyme assembly step.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
Cited by
87 articles.
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