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Complete genome determination and analysis of Acholeplasma oculi strain 19L, highlighting the loss of basic genetic features in the Acholeplasmataceae.

Siewert C, Hess WR, Duduk B, Huettel B, Reinhardt R, Büttner C, Kube M - BMC Genomics (2014)

Bottom Line: Sequencing by synthesis resulted in six large genome fragments, while the single molecule real time sequencing approach yielded one circular chromosome sequence.Comparative genome analyses revealed that the process of losing particular basic genetic features during genome reduction occurs in both genera, as indicated for several phytoplasma strains and at least A. oculi.The loss of the F1FO-type Na+ ATPase system may separate Acholeplasmataceae from other Mollicutes, while the loss of those genes encoding the chaperone GroEL/ES is not a rare exception in this bacterial class.

View Article: PubMed Central - PubMed

Affiliation: Humboldt-Universität zu Berlin, Faculty of Life Science, Thaer-Institute, Division Phytomedicine, Lentzeallee 55/57, 14195 Berlin, Germany. Michael.Kube@agrar.hu-berlin.de.

ABSTRACT

Background: Acholeplasma oculi belongs to the Acholeplasmataceae family, comprising the genera Acholeplasma and 'Candidatus Phytoplasma'. Acholeplasmas are ubiquitous saprophytic bacteria. Several isolates are derived from plants or animals, whereas phytoplasmas are characterised as intracellular parasitic pathogens of plant phloem and depend on insect vectors for their spread. The complete genome sequences for eight strains of this family have been resolved so far, all of which were determined depending on clone-based sequencing.

Results: The A. oculi strain 19L chromosome was sequenced using two independent approaches. The first approach comprised sequencing by synthesis (Illumina) in combination with Sanger sequencing, while single molecule real time sequencing (PacBio) was used in the second. The genome was determined to be 1,587,120 bp in size. Sequencing by synthesis resulted in six large genome fragments, while the single molecule real time sequencing approach yielded one circular chromosome sequence. High-quality sequences were obtained by both strategies differing in six positions, which are interpreted as reliable variations present in the culture population. Our genome analysis revealed 1,471 protein-coding genes and highlighted the absence of the F1FO-type Na+ ATPase system and GroEL/ES chaperone. Comparison of the four available Acholeplasma sequences revealed a core-genome encoding 703 proteins and a pan-genome of 2,867 proteins.

Conclusions: The application of two state-of-the-art sequencing technologies highlights the potential of single molecule real time sequencing for complete genome determination. Comparative genome analyses revealed that the process of losing particular basic genetic features during genome reduction occurs in both genera, as indicated for several phytoplasma strains and at least A. oculi. The loss of the F1FO-type Na+ ATPase system may separate Acholeplasmataceae from other Mollicutes, while the loss of those genes encoding the chaperone GroEL/ES is not a rare exception in this bacterial class.

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Phylogenetic tree based on 16SrRNAgene sequences of acholeplasmas (orange) and phytoplasmas (green). The tree was calculated by employing the maximum likelihood algorithm and bootstrap calculation for 1,000 replicates (only values of at least 70% are shown). The bar indicates 0.05 substitutions per nucleotide. The accession numbers are given in parentheses. Mycoplasma genitalium strain G37 is set as an out-group. Species with complete chromosomes available are shown in bold. Roman numerals are given according to acholeplasma clades [14]. The coloured boxes indicate that gene encoding F1F0 Na+ ATP synthase (light blue), V1VO Na+ ATP synthase (yellow), V1VO H+ ATP synthase (dark blue), GroEL (red) or SecB (violet) are present (limited to complete genome sequences).
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Fig2: Phylogenetic tree based on 16SrRNAgene sequences of acholeplasmas (orange) and phytoplasmas (green). The tree was calculated by employing the maximum likelihood algorithm and bootstrap calculation for 1,000 replicates (only values of at least 70% are shown). The bar indicates 0.05 substitutions per nucleotide. The accession numbers are given in parentheses. Mycoplasma genitalium strain G37 is set as an out-group. Species with complete chromosomes available are shown in bold. Roman numerals are given according to acholeplasma clades [14]. The coloured boxes indicate that gene encoding F1F0 Na+ ATP synthase (light blue), V1VO Na+ ATP synthase (yellow), V1VO H+ ATP synthase (dark blue), GroEL (red) or SecB (violet) are present (limited to complete genome sequences).

Mentions: The finished consensus sequence of A. oculi strain 19L consists of 1,587,120 bp encoding two rRNA operons, 34 tRNA genes and 1,471 predicted protein coding genes (Table 3). A. laidlawii strain PG-8A is the closest known relative of A. oculi strain 19L, which is supported by the construction of the phylogenetic tree (Figure 2). This close relationship is also reflected by the prediction of 1,068 shared proteins (77%) compared to 866 (60%) and 973 (57%) proteins shared with A. brassicae strain O502 and A. palmae strain J233 (Figure 3). The predicted core of the four Acholeplasma spp. consists of 703 proteins and the calculated pan-genome comprises 2,867 proteins in total (Figure 4). The highest number of unique proteins (570) possesses A. brassicae, which also exhibits the largest genome in this family (1,877,792 bp, 1,704 protein coding genes, Table 3).Table 3


Complete genome determination and analysis of Acholeplasma oculi strain 19L, highlighting the loss of basic genetic features in the Acholeplasmataceae.

Siewert C, Hess WR, Duduk B, Huettel B, Reinhardt R, Büttner C, Kube M - BMC Genomics (2014)

Phylogenetic tree based on 16SrRNAgene sequences of acholeplasmas (orange) and phytoplasmas (green). The tree was calculated by employing the maximum likelihood algorithm and bootstrap calculation for 1,000 replicates (only values of at least 70% are shown). The bar indicates 0.05 substitutions per nucleotide. The accession numbers are given in parentheses. Mycoplasma genitalium strain G37 is set as an out-group. Species with complete chromosomes available are shown in bold. Roman numerals are given according to acholeplasma clades [14]. The coloured boxes indicate that gene encoding F1F0 Na+ ATP synthase (light blue), V1VO Na+ ATP synthase (yellow), V1VO H+ ATP synthase (dark blue), GroEL (red) or SecB (violet) are present (limited to complete genome sequences).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4221730&req=5

Fig2: Phylogenetic tree based on 16SrRNAgene sequences of acholeplasmas (orange) and phytoplasmas (green). The tree was calculated by employing the maximum likelihood algorithm and bootstrap calculation for 1,000 replicates (only values of at least 70% are shown). The bar indicates 0.05 substitutions per nucleotide. The accession numbers are given in parentheses. Mycoplasma genitalium strain G37 is set as an out-group. Species with complete chromosomes available are shown in bold. Roman numerals are given according to acholeplasma clades [14]. The coloured boxes indicate that gene encoding F1F0 Na+ ATP synthase (light blue), V1VO Na+ ATP synthase (yellow), V1VO H+ ATP synthase (dark blue), GroEL (red) or SecB (violet) are present (limited to complete genome sequences).
Mentions: The finished consensus sequence of A. oculi strain 19L consists of 1,587,120 bp encoding two rRNA operons, 34 tRNA genes and 1,471 predicted protein coding genes (Table 3). A. laidlawii strain PG-8A is the closest known relative of A. oculi strain 19L, which is supported by the construction of the phylogenetic tree (Figure 2). This close relationship is also reflected by the prediction of 1,068 shared proteins (77%) compared to 866 (60%) and 973 (57%) proteins shared with A. brassicae strain O502 and A. palmae strain J233 (Figure 3). The predicted core of the four Acholeplasma spp. consists of 703 proteins and the calculated pan-genome comprises 2,867 proteins in total (Figure 4). The highest number of unique proteins (570) possesses A. brassicae, which also exhibits the largest genome in this family (1,877,792 bp, 1,704 protein coding genes, Table 3).Table 3

Bottom Line: Sequencing by synthesis resulted in six large genome fragments, while the single molecule real time sequencing approach yielded one circular chromosome sequence.Comparative genome analyses revealed that the process of losing particular basic genetic features during genome reduction occurs in both genera, as indicated for several phytoplasma strains and at least A. oculi.The loss of the F1FO-type Na+ ATPase system may separate Acholeplasmataceae from other Mollicutes, while the loss of those genes encoding the chaperone GroEL/ES is not a rare exception in this bacterial class.

View Article: PubMed Central - PubMed

Affiliation: Humboldt-Universität zu Berlin, Faculty of Life Science, Thaer-Institute, Division Phytomedicine, Lentzeallee 55/57, 14195 Berlin, Germany. Michael.Kube@agrar.hu-berlin.de.

ABSTRACT

Background: Acholeplasma oculi belongs to the Acholeplasmataceae family, comprising the genera Acholeplasma and 'Candidatus Phytoplasma'. Acholeplasmas are ubiquitous saprophytic bacteria. Several isolates are derived from plants or animals, whereas phytoplasmas are characterised as intracellular parasitic pathogens of plant phloem and depend on insect vectors for their spread. The complete genome sequences for eight strains of this family have been resolved so far, all of which were determined depending on clone-based sequencing.

Results: The A. oculi strain 19L chromosome was sequenced using two independent approaches. The first approach comprised sequencing by synthesis (Illumina) in combination with Sanger sequencing, while single molecule real time sequencing (PacBio) was used in the second. The genome was determined to be 1,587,120 bp in size. Sequencing by synthesis resulted in six large genome fragments, while the single molecule real time sequencing approach yielded one circular chromosome sequence. High-quality sequences were obtained by both strategies differing in six positions, which are interpreted as reliable variations present in the culture population. Our genome analysis revealed 1,471 protein-coding genes and highlighted the absence of the F1FO-type Na+ ATPase system and GroEL/ES chaperone. Comparison of the four available Acholeplasma sequences revealed a core-genome encoding 703 proteins and a pan-genome of 2,867 proteins.

Conclusions: The application of two state-of-the-art sequencing technologies highlights the potential of single molecule real time sequencing for complete genome determination. Comparative genome analyses revealed that the process of losing particular basic genetic features during genome reduction occurs in both genera, as indicated for several phytoplasma strains and at least A. oculi. The loss of the F1FO-type Na+ ATPase system may separate Acholeplasmataceae from other Mollicutes, while the loss of those genes encoding the chaperone GroEL/ES is not a rare exception in this bacterial class.

Show MeSH
Related in: MedlinePlus