Complementation of the Mycoplasma synoviae MS-H vaccine strain with wild-type oppF 1 influences its growth characteristics

• Published in: Avian pathology Vol. 49; no. 3; pp. 275 - 285
• Main Authors: Kordafshari, Somayeh, Marenda, Marc S, Agnew, Rebecca, Shil, Pollob, Shahid, Muhammad A, Marth, Christina, Konsak, Barbara M, Noormohammadi, Amir H
• Format: Journal Article
• Language: English
• Published: England 01-06-2020
• Subjects: Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacterial Vaccines; Genetic Complementation Test; Membrane Transport Proteins; Models, Molecular; Mutation; Mycoplasma synoviae - genetics; Protein Conformation; Vaccines, Attenuated; Whole Genome Sequencing; growth characteristics; MS-H vaccine strain; oligopeptide permease transporter; truncated oppF1; Mycoplasma synoviae; frameshift mutation; wild-type oppF1
• ISSN: 0307-9457; 1465-3338
• DOI: 10.1080/03079457.2020.1729957

The
(MS) vaccine strain MS-H harbours a frameshift mutation in
(oligopeptide permease transporter) which results in expression of a truncated OppF
. The effect of this mutation on growth and attenuation of the MS-H is unknown. In this study, the impact of the mutation on the vaccine phenotype was investigated
by introducing a wild-type copy of
gene in the MS-H genome. Wild-type
was cloned under the
promoter into an
vector carrying a tetracycline resistance gene. MS-H was successfully transformed with the final construct pMS-oppF
-tetM or with a similar vector lacking
coding sequence (pMS-tetM). The MS-H transformed with pMS-oppF
-tetM exhibited smaller colony size than MS-H transformed with pMS-tetM. Monospecific rabbit sera against C-terminus of OppF
detected bands of expected size for full-length OppF
in the 86079/7NS parental strain of MS-H and the MS-H transformed with pMS-oppF
-tetM, but not in MS-H and MS-H transformed with pMS-tetM. Comparison of the growth curve of MS-H transformants harvested from media with/without tetracycline was conducted using
Q-PCR which revealed that MS-H transformed with pMS-tetM had a higher growth rate than MS-H transformed with pMS-oppF
-tetM in the media with/without tetracycline. Lastly, the whole genome sequencing of MS-H transformed with pMS-oppF
-tetM (passage 27) showed that the chromosomal copy of the mutated
had been replaced with a wild-type version of the gene. This study reveals that the truncation of
impacts on growth characteristics of the MS-H and provides insight into the molecular pathogenesis of MS and perhaps broader mycoplasma species.
The full-length OppF
was expressed in
MS-H vaccine.Truncation of
impacts on growth characteristics of the MS-H.Chromosomal copy of the mutated
in MS-H was replaced with wild-type
.