Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches
ABSTRACT Mycoplasma species are responsible for several economically significant livestock diseases for which there is a need for new and improved vaccines. Most of the existing mycoplasma vaccines are attenuated strains that have been empirically obtained by serial passages or by chemical mutagenes...
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2019-06-01
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| Series: | mSphere |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mSphere.00030-19 |
| _version_ | 1818990178683846656 |
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| author | Carole Lartigue Yanina Valverde Timana Fabien Labroussaa Elise Schieck Anne Liljander Flavio Sacchini Horst Posthaus Brigitte Batailler Pascal Sirand-Pugnet Sanjay Vashee Joerg Jores Alain Blanchard |
| author_facet | Carole Lartigue Yanina Valverde Timana Fabien Labroussaa Elise Schieck Anne Liljander Flavio Sacchini Horst Posthaus Brigitte Batailler Pascal Sirand-Pugnet Sanjay Vashee Joerg Jores Alain Blanchard |
| author_sort | Carole Lartigue |
| collection | DOAJ |
| description | ABSTRACT Mycoplasma species are responsible for several economically significant livestock diseases for which there is a need for new and improved vaccines. Most of the existing mycoplasma vaccines are attenuated strains that have been empirically obtained by serial passages or by chemical mutagenesis. The recent development of synthetic biology approaches has opened the way for the engineering of live mycoplasma vaccines. Using these tools, the essential GTPase-encoding gene obg was modified directly on the Mycoplasma mycoides subsp. capri genome cloned in yeast, reproducing mutations suspected to induce a temperature-sensitive (TS+) phenotype. After transplantation of modified genomes into a recipient cell, the phenotype of the resulting M. mycoides subsp. capri mutants was characterized. Single-point obg mutations did not result in a strong TS+ phenotype in M. mycoides subsp. capri, but a clone presenting three obg mutations was shown to grow with difficulty at temperatures of ≥40°C. This particular mutant was then tested in a caprine septicemia model of M. mycoides subsp. capri infection. Five out of eight goats infected with the parental strain had to be euthanized, in contrast to one out of eight goats infected with the obg mutant, demonstrating an attenuation of virulence in the mutant. Moreover, the strain isolated from the euthanized animal in the group infected with the obg mutant was shown to carry a reversion in the obg gene associated with the loss of the TS+ phenotype. This study demonstrates the feasibility of building attenuated strains of mycoplasma that could contribute to the design of novel vaccines with improved safety. IMPORTANCE Animal diseases due to mycoplasmas are a major cause of morbidity and mortality associated with economic losses for farmers all over the world. Currently used mycoplasma vaccines exhibit several drawbacks, including low efficacy, short time of protection, adverse reactions, and difficulty in differentiating infected from vaccinated animals. Therefore, there is a need for improved vaccines to control animal mycoplasmoses. Here, we used genome engineering tools derived from synthetic biology approaches to produce targeted mutations in the essential GTPase-encoding obg gene of Mycoplasma mycoides subsp. capri. Some of the resulting mutants exhibited a marked temperature-sensitive phenotype. The virulence of one of the obg mutants was evaluated in a caprine septicemia model and found to be strongly reduced. Although the obg mutant reverted to a virulent phenotype in one infected animal, we believe that these results contribute to a strategy that should help in building new vaccines against animal mycoplasmoses. |
| first_indexed | 2024-12-20T19:50:15Z |
| format | Article |
| id | doaj.art-ed83cef1f72e4a6fb7a486e1ae7354d3 |
| institution | Directory Open Access Journal |
| issn | 2379-5042 |
| language | English |
| last_indexed | 2024-12-20T19:50:15Z |
| publishDate | 2019-06-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mSphere |
| spelling | doaj.art-ed83cef1f72e4a6fb7a486e1ae7354d32022-12-21T19:28:20ZengAmerican Society for MicrobiologymSphere2379-50422019-06-014310.1128/mSphere.00030-19Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology ApproachesCarole Lartigue0Yanina Valverde Timana1Fabien Labroussaa2Elise Schieck3Anne Liljander4Flavio Sacchini5Horst Posthaus6Brigitte Batailler7Pascal Sirand-Pugnet8Sanjay Vashee9Joerg Jores10Alain Blanchard11INRA, UMR 1332 de Biologie du Fruit et Pathologie, Villenave d’Ornon, FranceINRA, UMR 1332 de Biologie du Fruit et Pathologie, Villenave d’Ornon, FranceINRA, UMR 1332 de Biologie du Fruit et Pathologie, Villenave d’Ornon, FranceInternational Livestock Research Institute, Nairobi, KenyaInternational Livestock Research Institute, Nairobi, KenyaInternational Livestock Research Institute, Nairobi, KenyaInstitute of Veterinary Pathology, University of Bern, Bern, SwitzerlandINRA, UMR 1332 de Biologie du Fruit et Pathologie, Villenave d’Ornon, FranceINRA, UMR 1332 de Biologie du Fruit et Pathologie, Villenave d’Ornon, FranceJ. Craig Venter Institute, Rockville, Maryland, USAInstitute of Veterinary Bacteriology, University of Bern, Bern, SwitzerlandINRA, UMR 1332 de Biologie du Fruit et Pathologie, Villenave d’Ornon, FranceABSTRACT Mycoplasma species are responsible for several economically significant livestock diseases for which there is a need for new and improved vaccines. Most of the existing mycoplasma vaccines are attenuated strains that have been empirically obtained by serial passages or by chemical mutagenesis. The recent development of synthetic biology approaches has opened the way for the engineering of live mycoplasma vaccines. Using these tools, the essential GTPase-encoding gene obg was modified directly on the Mycoplasma mycoides subsp. capri genome cloned in yeast, reproducing mutations suspected to induce a temperature-sensitive (TS+) phenotype. After transplantation of modified genomes into a recipient cell, the phenotype of the resulting M. mycoides subsp. capri mutants was characterized. Single-point obg mutations did not result in a strong TS+ phenotype in M. mycoides subsp. capri, but a clone presenting three obg mutations was shown to grow with difficulty at temperatures of ≥40°C. This particular mutant was then tested in a caprine septicemia model of M. mycoides subsp. capri infection. Five out of eight goats infected with the parental strain had to be euthanized, in contrast to one out of eight goats infected with the obg mutant, demonstrating an attenuation of virulence in the mutant. Moreover, the strain isolated from the euthanized animal in the group infected with the obg mutant was shown to carry a reversion in the obg gene associated with the loss of the TS+ phenotype. This study demonstrates the feasibility of building attenuated strains of mycoplasma that could contribute to the design of novel vaccines with improved safety. IMPORTANCE Animal diseases due to mycoplasmas are a major cause of morbidity and mortality associated with economic losses for farmers all over the world. Currently used mycoplasma vaccines exhibit several drawbacks, including low efficacy, short time of protection, adverse reactions, and difficulty in differentiating infected from vaccinated animals. Therefore, there is a need for improved vaccines to control animal mycoplasmoses. Here, we used genome engineering tools derived from synthetic biology approaches to produce targeted mutations in the essential GTPase-encoding obg gene of Mycoplasma mycoides subsp. capri. Some of the resulting mutants exhibited a marked temperature-sensitive phenotype. The virulence of one of the obg mutants was evaluated in a caprine septicemia model and found to be strongly reduced. Although the obg mutant reverted to a virulent phenotype in one infected animal, we believe that these results contribute to a strategy that should help in building new vaccines against animal mycoplasmoses.https://journals.asm.org/doi/10.1128/mSphere.00030-19GTPase ObgMycoplasmaSaccharomyces cerevisiaeattenuated straingenome engineeringgenome transplantation |
| spellingShingle | Carole Lartigue Yanina Valverde Timana Fabien Labroussaa Elise Schieck Anne Liljander Flavio Sacchini Horst Posthaus Brigitte Batailler Pascal Sirand-Pugnet Sanjay Vashee Joerg Jores Alain Blanchard Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches mSphere GTPase Obg Mycoplasma Saccharomyces cerevisiae attenuated strain genome engineering genome transplantation |
| title | Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches |
| title_full | Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches |
| title_fullStr | Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches |
| title_full_unstemmed | Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches |
| title_short | Attenuation of a Pathogenic <italic toggle="yes">Mycoplasma</italic> Strain by Modification of the <italic toggle="yes">obg</italic> Gene by Using Synthetic Biology Approaches |
| title_sort | attenuation of a pathogenic italic toggle yes mycoplasma italic strain by modification of the italic toggle yes obg italic gene by using synthetic biology approaches |
| topic | GTPase Obg Mycoplasma Saccharomyces cerevisiae attenuated strain genome engineering genome transplantation |
| url | https://journals.asm.org/doi/10.1128/mSphere.00030-19 |
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