Presentation Title

ABC-Type Multidrug Efflux Pump Affects Motility and EPS Production in Paraburkholderia unamae

Faculty Mentor

Shelley Thai

Start Date

17-11-2018 8:30 AM

End Date

17-11-2018 10:30 AM

Location

CREVELING 32

Session

POSTER 1

Type of Presentation

Poster

Subject Area

biological_agricultural_sciences

Abstract

Paraburkholderia unamae is a non-pathogenic, nitrogen-fixing bacterium and has a symbiotic relationship with its host plant. Two characteristics, exopolysaccharide (EPS) production and motility, are what allows bacteria to move from the soil to the plant root and establish symbiotic relationship with its plant host. It promotes plant growth and shows promise for possible use as enhanced fertilizers. The purpose of the study was to identify the genes involved in motility and exopolysaccharide (EPS) production, both of which are important factors in plant colonization. In order to conduct a genetic analysis of P.unamae, transposon mutagenesis method was employed to isolate and identify the genes responsible for exopolysaccharide (EPS) production and motility. During biparental conjugation, the pRL27 vector carrying Tn5-RL27 transposon was transferred from Escherichia coli (BW20767) into P. unamae. The hyperactive transposon randomly inserted itself into the chromosome. Mutants containing the transposable element (Tn5-RL27) were selected on M9 media, which contained rifampicin and kanamycin. Furthermore, 412 isolated colonies were selected and streaked to assess phenotypic characterizations of EPS and motility. Mutants LB 2.47 and PM 1.29 showed over-expressed exopolysaccharide (EPS) production and significantly reduced motility diameter compared to the wildtype, 83%, and 75%, respectively. Genomic DNA of these mutants were isolated and digested with Sac II restriction enzyme to identify the DNA region where the transposon inserted itself. Sequence analysis revealed that the transposon disrupted the ATP-binding (ABC) cassette gene that codes for the multidrug resistance-like ATP-binding protein. The primary function of this protein is to efflux out drugs by coupling energy derived from the hydrolysis of ATP. Present research demonstrates that bacterial species inhabiting plants rhizospheres have a larger number of genes encoding efflux pumps. It is hypothesized that besides resistance to antibiotics, efflux pumps also play a crucial role in natural ecosystems, especially in bacteria-plant interactions.

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Nov 17th, 8:30 AM Nov 17th, 10:30 AM

ABC-Type Multidrug Efflux Pump Affects Motility and EPS Production in Paraburkholderia unamae

CREVELING 32

Paraburkholderia unamae is a non-pathogenic, nitrogen-fixing bacterium and has a symbiotic relationship with its host plant. Two characteristics, exopolysaccharide (EPS) production and motility, are what allows bacteria to move from the soil to the plant root and establish symbiotic relationship with its plant host. It promotes plant growth and shows promise for possible use as enhanced fertilizers. The purpose of the study was to identify the genes involved in motility and exopolysaccharide (EPS) production, both of which are important factors in plant colonization. In order to conduct a genetic analysis of P.unamae, transposon mutagenesis method was employed to isolate and identify the genes responsible for exopolysaccharide (EPS) production and motility. During biparental conjugation, the pRL27 vector carrying Tn5-RL27 transposon was transferred from Escherichia coli (BW20767) into P. unamae. The hyperactive transposon randomly inserted itself into the chromosome. Mutants containing the transposable element (Tn5-RL27) were selected on M9 media, which contained rifampicin and kanamycin. Furthermore, 412 isolated colonies were selected and streaked to assess phenotypic characterizations of EPS and motility. Mutants LB 2.47 and PM 1.29 showed over-expressed exopolysaccharide (EPS) production and significantly reduced motility diameter compared to the wildtype, 83%, and 75%, respectively. Genomic DNA of these mutants were isolated and digested with Sac II restriction enzyme to identify the DNA region where the transposon inserted itself. Sequence analysis revealed that the transposon disrupted the ATP-binding (ABC) cassette gene that codes for the multidrug resistance-like ATP-binding protein. The primary function of this protein is to efflux out drugs by coupling energy derived from the hydrolysis of ATP. Present research demonstrates that bacterial species inhabiting plants rhizospheres have a larger number of genes encoding efflux pumps. It is hypothesized that besides resistance to antibiotics, efflux pumps also play a crucial role in natural ecosystems, especially in bacteria-plant interactions.