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Challenges in predicting the evolutionary maintenance of a phage transgene.

Schmerer M, Molineux IJ, Ally D, Tyerman J, Cecchini N, Bull JJ - J Biol Eng (2014)

Bottom Line: Consistent with the previous study, the dispersin phage was superior to unmodified phage at clearing short term biofilms grown in broth, shown here to be an effect attributable to free enzyme.There was little empirical support for the tragedy of the commons framework despite a strong theoretical foundation for its supposed relevance.Expressed from a different part of the genome, the transgene did behave as if intrinsically costly, but its maintenance did not benefit from spatially structured growth per se - violating the tragedy framework.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Computational Biology and Bioinformatics, The University of Texas at Austin, Austin, TX, USA.

ABSTRACT

Background: In prior work, a phage engineered with a biofilm-degrading enzyme (dispersin B) cleared artificial, short-term biofilms more fully than the phage lacking the enzyme. An unresolved question is whether the transgene will be lost or maintained during phage growth - its loss would limit the utility of the engineering. Broadly supported evolutionary theory suggests that transgenes will be lost through a 'tragedy of the commons' mechanism unless the ecology of growth in biofilms meets specific requirements. We test that theory here.

Results: Functional properties of the transgenic phage were identified. Consistent with the previous study, the dispersin phage was superior to unmodified phage at clearing short term biofilms grown in broth, shown here to be an effect attributable to free enzyme. However, the dispersin phage was only marginally better than control phages on short term biofilms in minimal media and was no better than control phages in clearing long term biofilms. There was little empirical support for the tragedy of the commons framework despite a strong theoretical foundation for its supposed relevance. The framework requires that the transgene imposes an intrinsic cost, yet the transgene was intrinsically neutral or beneficial when expressed from one part of the phage genome. Expressed from a different part of the genome, the transgene did behave as if intrinsically costly, but its maintenance did not benefit from spatially structured growth per se - violating the tragedy framework.

Conclusions: Overall, the transgene was beneficial under many conditions, but no insight to its maintenance was attributable to the established evolutionary framework. The failure likely resides in system details that would be used to parameterize the models. Our study cautions against naive applications of evolutionary theory to synthetic biology, even qualitatively.

No MeSH data available.


Related in: MedlinePlus

Assays of dispersin activity using a colorimetric substrate. Note that a high absorbance in this assay indicates high enzyme activity (in contrast to the CV assay). The Pure enzyme is a commercial preparation and shows high activity, reflecting its activity as an exoglycosidase on a substrate designed for that activity. The control is from a freeze-thaw lysate of BL21(DE3) cells carrying an empty pET15b plasmid. The bars for enzyme extract (Xtr) are from freeze-thaw lysates of BL21(DE3) cells carrying plasmid pET15b-dspB, with the volume of the extract in parentheses. Bars of 1 std. error are shown (sometimes too small to view).
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Figure 3: Assays of dispersin activity using a colorimetric substrate. Note that a high absorbance in this assay indicates high enzyme activity (in contrast to the CV assay). The Pure enzyme is a commercial preparation and shows high activity, reflecting its activity as an exoglycosidase on a substrate designed for that activity. The control is from a freeze-thaw lysate of BL21(DE3) cells carrying an empty pET15b plasmid. The bars for enzyme extract (Xtr) are from freeze-thaw lysates of BL21(DE3) cells carrying plasmid pET15b-dspB, with the volume of the extract in parentheses. Bars of 1 std. error are shown (sometimes too small to view).

Mentions: The model requires that the transgene benefit the phage. We have thus far merely conjectured from plausibility that a greater clearing ability translates into greater progeny production. However, Lu and Collins [4] provided assays that supported a link between biofilm clearance and phage amplification. The dispersin phage consistently showed greater bacterial killing than the control phage (their Figures 3 and 4) as well as a superiority in phage numbers soon after treatment (their Figure 4); the latter effect is visually small because it is plotted on a log scale. Most phage replication occurs on planktonic cells in the media, so the effect of killing biofilm cells is not expected to be easily noted under the conditions used.


Challenges in predicting the evolutionary maintenance of a phage transgene.

Schmerer M, Molineux IJ, Ally D, Tyerman J, Cecchini N, Bull JJ - J Biol Eng (2014)

Assays of dispersin activity using a colorimetric substrate. Note that a high absorbance in this assay indicates high enzyme activity (in contrast to the CV assay). The Pure enzyme is a commercial preparation and shows high activity, reflecting its activity as an exoglycosidase on a substrate designed for that activity. The control is from a freeze-thaw lysate of BL21(DE3) cells carrying an empty pET15b plasmid. The bars for enzyme extract (Xtr) are from freeze-thaw lysates of BL21(DE3) cells carrying plasmid pET15b-dspB, with the volume of the extract in parentheses. Bars of 1 std. error are shown (sometimes too small to view).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4128545&req=5

Figure 3: Assays of dispersin activity using a colorimetric substrate. Note that a high absorbance in this assay indicates high enzyme activity (in contrast to the CV assay). The Pure enzyme is a commercial preparation and shows high activity, reflecting its activity as an exoglycosidase on a substrate designed for that activity. The control is from a freeze-thaw lysate of BL21(DE3) cells carrying an empty pET15b plasmid. The bars for enzyme extract (Xtr) are from freeze-thaw lysates of BL21(DE3) cells carrying plasmid pET15b-dspB, with the volume of the extract in parentheses. Bars of 1 std. error are shown (sometimes too small to view).
Mentions: The model requires that the transgene benefit the phage. We have thus far merely conjectured from plausibility that a greater clearing ability translates into greater progeny production. However, Lu and Collins [4] provided assays that supported a link between biofilm clearance and phage amplification. The dispersin phage consistently showed greater bacterial killing than the control phage (their Figures 3 and 4) as well as a superiority in phage numbers soon after treatment (their Figure 4); the latter effect is visually small because it is plotted on a log scale. Most phage replication occurs on planktonic cells in the media, so the effect of killing biofilm cells is not expected to be easily noted under the conditions used.

Bottom Line: Consistent with the previous study, the dispersin phage was superior to unmodified phage at clearing short term biofilms grown in broth, shown here to be an effect attributable to free enzyme.There was little empirical support for the tragedy of the commons framework despite a strong theoretical foundation for its supposed relevance.Expressed from a different part of the genome, the transgene did behave as if intrinsically costly, but its maintenance did not benefit from spatially structured growth per se - violating the tragedy framework.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Computational Biology and Bioinformatics, The University of Texas at Austin, Austin, TX, USA.

ABSTRACT

Background: In prior work, a phage engineered with a biofilm-degrading enzyme (dispersin B) cleared artificial, short-term biofilms more fully than the phage lacking the enzyme. An unresolved question is whether the transgene will be lost or maintained during phage growth - its loss would limit the utility of the engineering. Broadly supported evolutionary theory suggests that transgenes will be lost through a 'tragedy of the commons' mechanism unless the ecology of growth in biofilms meets specific requirements. We test that theory here.

Results: Functional properties of the transgenic phage were identified. Consistent with the previous study, the dispersin phage was superior to unmodified phage at clearing short term biofilms grown in broth, shown here to be an effect attributable to free enzyme. However, the dispersin phage was only marginally better than control phages on short term biofilms in minimal media and was no better than control phages in clearing long term biofilms. There was little empirical support for the tragedy of the commons framework despite a strong theoretical foundation for its supposed relevance. The framework requires that the transgene imposes an intrinsic cost, yet the transgene was intrinsically neutral or beneficial when expressed from one part of the phage genome. Expressed from a different part of the genome, the transgene did behave as if intrinsically costly, but its maintenance did not benefit from spatially structured growth per se - violating the tragedy framework.

Conclusions: Overall, the transgene was beneficial under many conditions, but no insight to its maintenance was attributable to the established evolutionary framework. The failure likely resides in system details that would be used to parameterize the models. Our study cautions against naive applications of evolutionary theory to synthetic biology, even qualitatively.

No MeSH data available.


Related in: MedlinePlus