Acetobacter pasteurianus IFO 3283-32 (= NBRC 105190)
open/close allAbout this Microorganism
Photo by Dr. Muramatsu (NBRC, NITE)
Acetic acid bacteria (AAB) are a group of widely diverged bacteria within alpha-proteobacteria and also known for their genetic instability. Acetobacter pasteurianus is one of the popular AAB, which has been used for traditional vinegar production. A. pasteurianus IFO 3283 (= NBRC 3283) was isolated from a pellicle formed on the surface of fermentation of traditional Japanese rice vinegar. Since the deposit of the pure culture in IFO (Institute for Fermentation, Osaka) in 1954, the strain had been maintained by a series of passages at three-months intervals until 1974 when freeze-dry preservation method was established. As a result, a multi-phenotype cell complex with non-homogeneous colony textures was formed.
To avoid the loss of genetic information pertaining to useful features maintained in the cell complex, whole-genome shotgun libraries were constructed from genomic DNA extracted from the cell complex without any single colony isolation. After sequencing and assembly, 38 assembling gaps and sequence discrepancies remained as candidate loci of genomic variations. A total of 32 substrains were then isolated from the cell complex and their sequence variations were confirmed by resequencing of the candidate loci. In total, 4 transposon insertions, 3 SNPs and 3 Hyper-mutable Tandem Repeats (HTRs) were identified as chromosome variations, while a transposon insertion and a HTR were observed in the largest plasmid. This strategy allowed us to establish whole genome sequences, each being composed of a single chromosome and 6 plasmids, of 7 representative substrains (IFO 3283-01, 03, 07, 12, 22, 26 and 32). Three SNPs and 4 transposon insertions on the chromosome were used to deduce the generation order of genomic variations as shown in the Figure presented below. In addition, a temperature resistant mutant was obtained from one of the isolates, IFO 3283-01, after prolonged cultivation at 40 C. Genome sequencing of the temperature resistant mutant revealed 3 SNPs and a 92 kb deletion. Genome analysis of A. pasteurianus thus provided important insights into the genetic instability and flexibility of hyper-mutabe AAB.
For simplicity, this database deals only with one of the 7 representative substrains, IFO 3283-32, which is thought to bear the original phenotype of IFO 3283. Genomic variations found in other 6 substrains are summarized in and can be accessed from the following figures.
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| Fig. Genotype lineage (chromosome) of 32 substrains of the IFO 3283 cell complex |
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Seven representative substrains isolated from IFO 3283 cell comlex (IFO 3283-01, 03, 07, 12, 22, 26, 32) and the high temperature-resistant strain (IFO 3283-01-42C) were redeposited in NBRC with catalogue numbers shown in the "General Procedure" head. The original cell complex is also available as NBRC 3283.
Variation observed in IFO 3283 cell complex| 1 | type: Hyper mutable Tandem Repeats (HTR) releated ORFs: APA32_02730 product: Beta-Ig-H3/fasciclin repeat containing protein |
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Show Annotation |
| 2 | type: Single Nucleotide Polymorphism (SNP) releated ORFs: APA32_09550 product: DNA-directed RNA polymerase alpha subunit gene: rpoA |
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| 3 | type: Insertion Sequence (IS) releated ORFs: APA32_10210 product: Two component sensor histidine kinase |
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| 4 | type: Single Nucleotide Polymorphism (SNP) releated ORFs: APA32_12550 product: Glycerol kinase |
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| 5 | type: Hyper mutable Tandem Repeats (HTR) releated ORFs: APA32_16984 product: Transcriptional regulator MerR |
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| 6 | type: Single Nucleotide Polymorphism (SNP) releated ORFs: APA32_17930 product: Hypothetical protein |
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| 7 | type: Insertion Sequence (IS) releated ORFs: APA32_18050 product: Transposase |
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| 8 | type: Hyper mutable Tandem Repeats (HTR) releated ORFs: APA32_18746, APA32_18748 product: DNA helicase restriction enzyme Type III R subunit |
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Show Annotation 1 Show Annotation 2 |
| 9 | type: Hyper mutable Tandem Repeats (HTR) releated ORFs: APA32_20310 product: DNA polymerase III exonuclease epsilon subunit like protein |
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Show Annotation |
| 10 | type: Insertion Sequence (IS) releated ORFs: APA32_21760 product: Protein translocase subunit SecB |
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| 11 | type: Insertion Sequence (IS) releated ORFs: APA32_24440 product: Glycosyl transferase |
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| 12 | type: Insertion Sequence (IS) releated ORFs: none (Intergenic region) |
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| 13 | type: Hyper mutable Tandem Repeats (HTR) releated ORFs: APA32_40324 product: Hypothetical protein |
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Show Annotation |
Project history
2009-10-20 ..... 1
Release of the Acetobacter pasteurianus IFO 3283 genomic sequence data | We published the genome data of Acetobacter pasteurianus IFO 3283 including the information of DNA clones distributed from the NBRC |
Summary of the genomic data
| Genomic size | 3,337,040 bp | ||||||||||||||||||||||||||||||||||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| G+C content | 53.10 % | ||||||||||||||||||||||||||||||||||||||||||||||||||
| Number of ORFs assigned | 3,046 | ||||||||||||||||||||||||||||||||||||||||||||||||||
| Percentage of the coding regions | 89.27 % | ||||||||||||||||||||||||||||||||||||||||||||||||||
| Percentage of the intronic regions | 0.00 % | ||||||||||||||||||||||||||||||||||||||||||||||||||
| Number of rRNA genes |
15
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| Number of tRNA genes |
57
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| Number of other features (misc_RNA,misc_feature,repeat) |
0 |
General Procedure
Shotgun DNA sequencing was carried out using mixed genomic DNA extracted from the cell complex of A. pasteurianus IFO 3283.
General Procedure
- DNA shotgun library
DNA shotgun libraries with inserts of 1.5 and 5.0 kb in pUC118 vector (TAKARA) was constructed. - Cosmid library
A Cosmid library with inserts of 35 kb in the pCC1FOS fosmid vector was constructed (Epicentre). - Nucleotide sequencing
Plasmid clones were end-sequenced using dye-terminator chemistry on an Applied Biosystems 3730xl DNA Analyzer.
Fosmid DNA was extracted from E. coli transformants using the Montage BAC96 MiniPrep Kit (Millipore) and end-sequencing was carried out using dye-terminator chemistry on Applied Biosystems 3730xl DNA Analyzer.
Raw sequence data corresponding to approximately 9.3-fold coverage were assembled using PHRED/PHRAP/CONSED software (http://www.phrap.org). - Gap closing and substrain isolation
30 gaps and 84 low quality regions under 40 in Phrap score were remained after construction of seven supercontigs corresponding to a chromosome and 6 plasmids.
DNA fragments were produced to fill all the gaps by PCR from genomic DNA of the 32 isolates from the IFO 3283 cell complex. All regions with lower quality sequences than 25 in Phred value were re-sequenced by primer walking.
As a result, 13 variable loci were identified in the 32 isolates of A. pasteurianus IFO 3283 (see About this microbe page). - Validation of the assembled sequence data
The assembled sequence was validated by Pulse field gel electrophoresis (Bio-Rad), restriction enzyme digestion of fosmid clone and Optical Mapping system (OpGen).
When any inconsistency in assembling and co-existence of different nucleotides were detected, the sequence variations were confirmed by re-sequencing all 32 isolated clones.
Genome analysis and annotation
- rRNA and tRNA genes were predicted using programs, BLASTN and tRNAscan-SE, respectively.
- For the identification of protein-coding genes, candidates of ORFs were extracted by using GenomeGambler, GeneHackerplus and Glimmer 3.0 with length longer than 90 bp and manually inspected.
- The translated sequences of the predicted protein-coding genes were searched against non-redundant protein database using BLASTP and FASTA3.
- Transmembrane protein genes were predicted using SOSUI.
- The software, JSTRING, was used for the extraction of hyper-mutable tandem repeats (HTR) from genomic DNA sequences of IFO 3283 substrains.
Summary of the A. pasteurianus IFO 3283 substrains Genomic Data
Strain name
IFO 3283-32
IFO 3283-01
IFO 3283-07
IFO 3283-12
IFO 3283-22
IFO 3283-26
IFO 3283-03
IFO 3283-01-42C
Genomic size:
2,904,642 bp
2,907,495 bp
2,906,044 bp
2,904,624 bp
2,907,267 bp
2,907,309 bp
2,907,287 bp
2,815,241 bp
GC content:
53.03%
53.04%
53.04%
53.03%
53.04%
53.04%
53.04%
53.09%
Coding region coverage:
90.31%
90.17%
90.17%
90.31%
90.31%
90.32%
90.26%
90.25%
Protein coding genes:
2,625
2,628
2,626
2,625
2,627
2,627
2,627
2,562
5S, 16S and 23S rRNA genes:
5 each
5 each
5 each
5 each
5 each
5 each
5 each
4 each
tRNA genes:
57
57
57
57
57
57
57
54
Strain name:
IFO 3283-32
IFO 3283-01
IFO 3283-07
IFO 3283-12
IFO 3283-22
IFO 3283-26
IFO 3283-03
IFO 3283-01-42C
Plasmid name:
pAPA32-012
pAPA01-011
pAPA07-010
pAPA12-014
pAPA22-010
pAPA26-013
pAPA03-010
pAPA42C-011
Genomic size:
191,443 bp
191,799 bp
191,427 bp
191,411 bp
191,427 bp
191,419 bp
191,427 bp
191,799 bp
GC content:
53.37%
53.36%
53.37%
53.37%
53.37%
53.37%
53.37%
53.36%
Coding region coverage:
80.53%
80.54%
80.54%
80.56%
80.54%
80.52%
80.54%
80.54%
Protein coding genes:
177
178
177
177
177
177
177
178
Genomic size:
182,940 bp
GC content:
53.50%
Coding region coverage:
85.10%
Protein coding genes:
174
Genomic size:
49,961 bp
GC content:
53.88%
Coding region coverage:
82.62%
Protein coding genes:
63
Genomic size:
3,204 bp
GC content:
53.75%
Coding region coverage:
80.81%
Protein coding genes:
3
Genomic size:
3,035 bp
GC content:
55.78%
Coding region coverage:
36.38%
Protein coding genes:
2
Genomic size:
1,815 bp
GC content:
58.57%
Coding region coverage:
53.55%
Protein coding genes:
2
Related links of A. pasteurianus IFO 3283 substrains data
Strain name
IFO 3283-32
IFO 3283-01
IFO 3283-07
IFO 3283-12
IFO 3283-22
IFO 3283-26
IFO 3283-03
IFO 3283-01-42C
NBRC culture catalogue NBRC No.
NBRC 105190
NBRC 105184
NBRC 105187
NBRC 105191
NBRC 105188
NBRC 105189
NBRC 105186
NBRC 105185
INSD, RefSeq Chromosome
DDBJ
AP011156
AP011121
AP011135
AP011170
AP011142
AP011149
AP011128
AP011163
RefSeq
NC_013209
Plasmid 1
DDBJ
AP011157
AP011122
AP011136
AP011171
AP011143
AP011150
AP011129
AP011164
RefSeq
NC_013210
Plasmid 2
DDBJ
AP011158
AP011123
AP011137
AP011172
AP011144
AP011151
AP011130
AP011165
RefSeq
NC_013211
Plasmid 3
DDBJ
AP011159
AP011124
AP011138
AP011173
AP011145
AP011152
AP011131
AP011166
RefSeq
NC_013212
Plasmid 4
DDBJ
AP011160
AP011125
AP011139
AP011174
AP011146
AP011153
AP011132
AP011167
RefSeq
NC_013213
Plasmid 5
DDBJ
AP011161
AP011126
AP011140
AP011175
AP011147
AP011154
AP011133
AP011167
RefSeq
NC_013214
Plasmid 6
DDBJ
AP011162
AP011127
AP011141
AP011176
AP011148
AP011155
AP011134
AP011169
RefSeq
NC_013215
