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The Molecular and Genetic Basis of Repeatable Coevolution between Escherichia coli and Bacteriophage T3 in a Laboratory Microcosm.

Perry EB, Barrick JE, Bohannan BJ - PLoS ONE (2015)

Bottom Line: Phage also showed repeatable evolution, with each chemostat producing host-range mutant phage with mutations in the phage tail fiber gene T3p48 which binds to the bacterial LPS during adsorption.Although a wide range of mutations occurred in the bacterial waaG gene, mutations in the phage tail fiber were restricted to a single codon, and several phage showed convergent evolution at the nucleotide level.Our data are also consistent with the expectation that adaptation via loss-of-function mutations is less constrained than adaptation via gain-of-function mutations.

View Article: PubMed Central - PubMed

Affiliation: Institute of Ecology and Evolution, University of Oregon, Eugene, Oregon, United States of America; Memorial Sloan Kettering Cancer Center, New York, New York, United States of America.

ABSTRACT
The objective of this study was to determine the genomic changes that underlie coevolution between Escherichia coli B and bacteriophage T3 when grown together in a laboratory microcosm. We also sought to evaluate the repeatability of their evolution by studying replicate coevolution experiments inoculated with the same ancestral strains. We performed the coevolution experiments by growing Escherichia coli B and the lytic bacteriophage T3 in seven parallel continuous culture devices (chemostats) for 30 days. In each of the chemostats, we observed three rounds of coevolution. First, bacteria evolved resistance to infection by the ancestral phage. Then, a new phage type evolved that was capable of infecting the resistant bacteria as well as the sensitive bacterial ancestor. Finally, we observed second-order resistant bacteria evolve that were resistant to infection by both phage types. To identify the genetic changes underlying coevolution, we isolated first- and second-order resistant bacteria as well as a host-range mutant phage from each chemostat and sequenced their genomes. We found that first-order resistant bacteria consistently evolved resistance to phage via mutations in the gene, waaG, which codes for a glucosyltransferase required for assembly of the bacterial lipopolysaccharide (LPS). Phage also showed repeatable evolution, with each chemostat producing host-range mutant phage with mutations in the phage tail fiber gene T3p48 which binds to the bacterial LPS during adsorption. Two second-order resistant bacteria evolved via mutations in different genes involved in the phage interaction. Although a wide range of mutations occurred in the bacterial waaG gene, mutations in the phage tail fiber were restricted to a single codon, and several phage showed convergent evolution at the nucleotide level. These results are consistent with previous studies in other systems that have documented repeatable evolution in bacteria at the level of pathways or genes and repeatable evolution in viruses at the nucleotide level. Our data are also consistent with the expectation that adaptation via loss-of-function mutations is less constrained than adaptation via gain-of-function mutations.

No MeSH data available.


Related in: MedlinePlus

A complete list of genomic mutations distinguishing derived phenotypes from their ancestors.The positions of mutations are indicated for regions of the genome in which mutations have been shown to be sufficient to confer the derived phenotype. (A) Mutations distinguishing first-order resistant B1 bacteria from the sensitive B0 ancestor. (B) Mutations distinguishing host-range mutant T31 phage from the wild-type T30 ancestor. (C) Mutations distinguishing second-order resistant B2 bacteria from the B0 ancestor. The “waa” prefix has been omitted from LPS biosynthesis genes to conserve space. The white area indicates a deletion that spans several genes. Detailed information about each mutation is provided in S1–S3 Tables.
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pone.0130639.g002: A complete list of genomic mutations distinguishing derived phenotypes from their ancestors.The positions of mutations are indicated for regions of the genome in which mutations have been shown to be sufficient to confer the derived phenotype. (A) Mutations distinguishing first-order resistant B1 bacteria from the sensitive B0 ancestor. (B) Mutations distinguishing host-range mutant T31 phage from the wild-type T30 ancestor. (C) Mutations distinguishing second-order resistant B2 bacteria from the B0 ancestor. The “waa” prefix has been omitted from LPS biosynthesis genes to conserve space. The white area indicates a deletion that spans several genes. Detailed information about each mutation is provided in S1–S3 Tables.

Mentions: B1 bacteria showed a strong signal of repeatable evolution at the gene level. Six of the B1 resistant genomes are distinguished from the ancestor by just a single mutation, and the seventh strain has two mutations. All but one of the mutations (and all of the non-synonymous mutations) occurred within a single gene (waaG) in the genome. (Fig 2) (S1 Table). The probability that every strain would have at least one mutation in this gene (a target of 1,125 base pairs in the 4.6 million base pair genome) by chance is extremely low (p = 1×10−25). The waaG gene codes for glucosyltransferase I. This enzyme is involved in the synthesis of the E. coli lipopolysaccharide (LPS) [31], an important outer membrane component of Gram-negative bacteria. The glucosyltransferase I enzyme links outer core glucose residues to the inner core saccharides in the LPS, and loss of this enzyme results in truncated LPS structures lacking outer core sugars (Fig 1B). The bacterial LPS is a target for selection in this system because it is the surface structure to which bacteriophage T3 binds when infecting E. coli [32, 33]. The six resistant strains that are distinguished from the sensitive ancestor by just a single mutation in waaG demonstrate that one mutation in this gene is sufficient to confer the resistant phenotype. Based on the molecular structure of the glucosyltransferase I enzyme [34], it is likely that every strain has severely disrupted the function of the enzyme, but that they do so in different ways. Two of the strains have deletions or insertions that shift the remainder of the protein coding sequence out of frame, two have in-frame deletions of two or five amino acids, and one has a mutation that introduces a stop codon early in the protein reading frame. The non-synonymous substitutions that occurred in two strains are also predicted to have a large effect on waaG structure and function, by altering key interactions in the ligand-binding pocket (F13V) or by introducing a charged amino acid into the hydrophobic core (L287R) [34].


The Molecular and Genetic Basis of Repeatable Coevolution between Escherichia coli and Bacteriophage T3 in a Laboratory Microcosm.

Perry EB, Barrick JE, Bohannan BJ - PLoS ONE (2015)

A complete list of genomic mutations distinguishing derived phenotypes from their ancestors.The positions of mutations are indicated for regions of the genome in which mutations have been shown to be sufficient to confer the derived phenotype. (A) Mutations distinguishing first-order resistant B1 bacteria from the sensitive B0 ancestor. (B) Mutations distinguishing host-range mutant T31 phage from the wild-type T30 ancestor. (C) Mutations distinguishing second-order resistant B2 bacteria from the B0 ancestor. The “waa” prefix has been omitted from LPS biosynthesis genes to conserve space. The white area indicates a deletion that spans several genes. Detailed information about each mutation is provided in S1–S3 Tables.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4482675&req=5

pone.0130639.g002: A complete list of genomic mutations distinguishing derived phenotypes from their ancestors.The positions of mutations are indicated for regions of the genome in which mutations have been shown to be sufficient to confer the derived phenotype. (A) Mutations distinguishing first-order resistant B1 bacteria from the sensitive B0 ancestor. (B) Mutations distinguishing host-range mutant T31 phage from the wild-type T30 ancestor. (C) Mutations distinguishing second-order resistant B2 bacteria from the B0 ancestor. The “waa” prefix has been omitted from LPS biosynthesis genes to conserve space. The white area indicates a deletion that spans several genes. Detailed information about each mutation is provided in S1–S3 Tables.
Mentions: B1 bacteria showed a strong signal of repeatable evolution at the gene level. Six of the B1 resistant genomes are distinguished from the ancestor by just a single mutation, and the seventh strain has two mutations. All but one of the mutations (and all of the non-synonymous mutations) occurred within a single gene (waaG) in the genome. (Fig 2) (S1 Table). The probability that every strain would have at least one mutation in this gene (a target of 1,125 base pairs in the 4.6 million base pair genome) by chance is extremely low (p = 1×10−25). The waaG gene codes for glucosyltransferase I. This enzyme is involved in the synthesis of the E. coli lipopolysaccharide (LPS) [31], an important outer membrane component of Gram-negative bacteria. The glucosyltransferase I enzyme links outer core glucose residues to the inner core saccharides in the LPS, and loss of this enzyme results in truncated LPS structures lacking outer core sugars (Fig 1B). The bacterial LPS is a target for selection in this system because it is the surface structure to which bacteriophage T3 binds when infecting E. coli [32, 33]. The six resistant strains that are distinguished from the sensitive ancestor by just a single mutation in waaG demonstrate that one mutation in this gene is sufficient to confer the resistant phenotype. Based on the molecular structure of the glucosyltransferase I enzyme [34], it is likely that every strain has severely disrupted the function of the enzyme, but that they do so in different ways. Two of the strains have deletions or insertions that shift the remainder of the protein coding sequence out of frame, two have in-frame deletions of two or five amino acids, and one has a mutation that introduces a stop codon early in the protein reading frame. The non-synonymous substitutions that occurred in two strains are also predicted to have a large effect on waaG structure and function, by altering key interactions in the ligand-binding pocket (F13V) or by introducing a charged amino acid into the hydrophobic core (L287R) [34].

Bottom Line: Phage also showed repeatable evolution, with each chemostat producing host-range mutant phage with mutations in the phage tail fiber gene T3p48 which binds to the bacterial LPS during adsorption.Although a wide range of mutations occurred in the bacterial waaG gene, mutations in the phage tail fiber were restricted to a single codon, and several phage showed convergent evolution at the nucleotide level.Our data are also consistent with the expectation that adaptation via loss-of-function mutations is less constrained than adaptation via gain-of-function mutations.

View Article: PubMed Central - PubMed

Affiliation: Institute of Ecology and Evolution, University of Oregon, Eugene, Oregon, United States of America; Memorial Sloan Kettering Cancer Center, New York, New York, United States of America.

ABSTRACT
The objective of this study was to determine the genomic changes that underlie coevolution between Escherichia coli B and bacteriophage T3 when grown together in a laboratory microcosm. We also sought to evaluate the repeatability of their evolution by studying replicate coevolution experiments inoculated with the same ancestral strains. We performed the coevolution experiments by growing Escherichia coli B and the lytic bacteriophage T3 in seven parallel continuous culture devices (chemostats) for 30 days. In each of the chemostats, we observed three rounds of coevolution. First, bacteria evolved resistance to infection by the ancestral phage. Then, a new phage type evolved that was capable of infecting the resistant bacteria as well as the sensitive bacterial ancestor. Finally, we observed second-order resistant bacteria evolve that were resistant to infection by both phage types. To identify the genetic changes underlying coevolution, we isolated first- and second-order resistant bacteria as well as a host-range mutant phage from each chemostat and sequenced their genomes. We found that first-order resistant bacteria consistently evolved resistance to phage via mutations in the gene, waaG, which codes for a glucosyltransferase required for assembly of the bacterial lipopolysaccharide (LPS). Phage also showed repeatable evolution, with each chemostat producing host-range mutant phage with mutations in the phage tail fiber gene T3p48 which binds to the bacterial LPS during adsorption. Two second-order resistant bacteria evolved via mutations in different genes involved in the phage interaction. Although a wide range of mutations occurred in the bacterial waaG gene, mutations in the phage tail fiber were restricted to a single codon, and several phage showed convergent evolution at the nucleotide level. These results are consistent with previous studies in other systems that have documented repeatable evolution in bacteria at the level of pathways or genes and repeatable evolution in viruses at the nucleotide level. Our data are also consistent with the expectation that adaptation via loss-of-function mutations is less constrained than adaptation via gain-of-function mutations.

No MeSH data available.


Related in: MedlinePlus