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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

Transgenic phage efficacy on short term biofilms. Each panel represents a different media (full strength LB,  LB, and M9 glucose) treated with the same phage stocks (T7+ is wild type T7, T7v is the empty T7 vector, and T7dsp is T7dsp+trx 10B). The vertical axis is the absorbance (560 nm) of the eluent after crystal violet staining. The bars give mean absorbance with 1 std error. Across each media, there is significant heterogeneity when and only when treatment with T7dsp is included (only marginally so for M9). There is also significant heterogeneity among the T7dsp treatments across the 3 media due to the M9 glucose experiment. Lower signal indicates greater biofilm clearing.
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Figure 2: Transgenic phage efficacy on short term biofilms. Each panel represents a different media (full strength LB, LB, and M9 glucose) treated with the same phage stocks (T7+ is wild type T7, T7v is the empty T7 vector, and T7dsp is T7dsp+trx 10B). The vertical axis is the absorbance (560 nm) of the eluent after crystal violet staining. The bars give mean absorbance with 1 std error. Across each media, there is significant heterogeneity when and only when treatment with T7dsp is included (only marginally so for M9). There is also significant heterogeneity among the T7dsp treatments across the 3 media due to the M9 glucose experiment. Lower signal indicates greater biofilm clearing.

Mentions: Short-term biofilms were grown as overnight cultures at 37° in 24-well, treated polystyrene microtiter plates, then incubated with phages for 5 h, and finally assayed for clearing with crystal violet (CV, Figure 2). Three types of media were used, full strength broth (LB), dilute broth ( LB), and minimal (M9 glucose). With such short-term bacterial growth, a strong effect of media on biofilm mass is expected and was in fact observed: across the 3 types of media, the untreated biofilms grown in full strength broth (LB) yielded the highest CV signal. For different phage treatments within the same media, the lowest CV signal was invariably with T7dsp; heterogeneity within each media was significant when all treatments were included but vanished when the T7dsp treatment was removed (when T7dsp was included, heterogeneity significance levels were P <0.0001 for LB, P <0.02 for LB, and P <0.04 for M9 glucose). However, the CV signal from T7dsp was significantly higher in M9 glucose media than in the other media (P <0.0001). For all 3 media, treatments with phages lacking transgenic dispersin were statistically indistinguishable from phage-free controls in the same media. These results support the main observations of Lu and Collins [4] but they further suggest that the magnitude of effect is media-dependent.


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)

Transgenic phage efficacy on short term biofilms. Each panel represents a different media (full strength LB,  LB, and M9 glucose) treated with the same phage stocks (T7+ is wild type T7, T7v is the empty T7 vector, and T7dsp is T7dsp+trx 10B). The vertical axis is the absorbance (560 nm) of the eluent after crystal violet staining. The bars give mean absorbance with 1 std error. Across each media, there is significant heterogeneity when and only when treatment with T7dsp is included (only marginally so for M9). There is also significant heterogeneity among the T7dsp treatments across the 3 media due to the M9 glucose experiment. Lower signal indicates greater biofilm clearing.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 2: Transgenic phage efficacy on short term biofilms. Each panel represents a different media (full strength LB, LB, and M9 glucose) treated with the same phage stocks (T7+ is wild type T7, T7v is the empty T7 vector, and T7dsp is T7dsp+trx 10B). The vertical axis is the absorbance (560 nm) of the eluent after crystal violet staining. The bars give mean absorbance with 1 std error. Across each media, there is significant heterogeneity when and only when treatment with T7dsp is included (only marginally so for M9). There is also significant heterogeneity among the T7dsp treatments across the 3 media due to the M9 glucose experiment. Lower signal indicates greater biofilm clearing.
Mentions: Short-term biofilms were grown as overnight cultures at 37° in 24-well, treated polystyrene microtiter plates, then incubated with phages for 5 h, and finally assayed for clearing with crystal violet (CV, Figure 2). Three types of media were used, full strength broth (LB), dilute broth ( LB), and minimal (M9 glucose). With such short-term bacterial growth, a strong effect of media on biofilm mass is expected and was in fact observed: across the 3 types of media, the untreated biofilms grown in full strength broth (LB) yielded the highest CV signal. For different phage treatments within the same media, the lowest CV signal was invariably with T7dsp; heterogeneity within each media was significant when all treatments were included but vanished when the T7dsp treatment was removed (when T7dsp was included, heterogeneity significance levels were P <0.0001 for LB, P <0.02 for LB, and P <0.04 for M9 glucose). However, the CV signal from T7dsp was significantly higher in M9 glucose media than in the other media (P <0.0001). For all 3 media, treatments with phages lacking transgenic dispersin were statistically indistinguishable from phage-free controls in the same media. These results support the main observations of Lu and Collins [4] but they further suggest that the magnitude of effect is media-dependent.

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