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Gene transfer in enteric bacteria through the formation of R-prime plasmids by an RP4: :mini-Mu element
M R Sarker1, Z U Ahmed, M M Rahman
1International Centre for Diarrhoeal Disease Research, Bangladesh, Dhaka.
Abstract:
Gene transfer in seven pathogenic enteric bacteria was studied using an RP4: :mini-Mu element, the plasmid pULB113. From the E. coli K-12 host strain the plasmid could be efficiently transferred to these enteric bacteria, but its transfer back to E. coli K-12 was not as efficient, being detected only in Shigella dysenteriae 1, S. flexneri and the 'smooth' variant of S. sonnei. In these three species, transposition of chromosomal fragments into the plasmid to produce R-prime plasmid was also detected at a frequency of approximately 10(-5). Transposition was random as suggested by the recovery at approximately the same frequency (10(-5) to 10(-6)) of R-primes involving 20 different auxotrophic markers from widely separated chromosomal locations. Formation of R-prime plasmids expressing toxicity in the E. coli K-12 recipient strain was also efficient in S. dysenteriae 1 but the toxin-activity was rapidly lost from these R-primes. In our experiments, the plasmid pULB113 incorporated relatively small amounts of chromosomal DNA as determined by restriction endonuclease digestion. For a Thy+ R-prime that we analyzed, the amount of cloned DNA was approximately 15 kb.
Insights
This study explored gene transfer in pathogenic enteric bacteria using the plasmid pULB113. Researchers found efficient transfer to bacteria but limited return to the host, with R-prime plasmid formation observed in specific species.
Area of Science:
- Microbiology
- Bacterial Genetics
- Molecular Biology
Background:
- Gene transfer mechanisms are crucial for understanding bacterial evolution and pathogenicity.
- The plasmid pULB113, a composite of RP4 and mini-Mu elements, facilitates intergeneric gene transfer.
Purpose of the Study:
- To investigate the efficiency and mechanisms of gene transfer mediated by the plasmid pULB113 in pathogenic enteric bacteria.
- To characterize the formation and properties of R-prime plasmids generated during this process.
Main Methods:
- Utilized the RP4: :mini-Mu element, plasmid pULB113, for gene transfer experiments.
- Assessed plasmid transfer efficiency between Escherichia coli K-12 and seven pathogenic enteric bacterial species.
- Analyzed R-prime plasmid formation and characterized the cloned chromosomal DNA using restriction endonuclease digestion.
Main Results:
- Efficient transfer of pULB113 from E. coli K-12 to pathogenic enteric bacteria was observed.
- Return transfer of pULB113 to E. coli K-12 was limited, primarily occurring in Shigella dysenteriae 1, Shigella flexneri, and a variant of Shigella sonnei.
- R-prime plasmid formation occurred at a frequency of approximately 10(-5), involving diverse chromosomal markers, indicating random transposition.
- Toxicity expression from R-prime plasmids in E. coli K-12 was efficient in S. dysenteriae 1 but transient.
Conclusions:
- The plasmid pULB113 is a versatile tool for gene transfer into pathogenic enteric bacteria.
- R-prime plasmid formation is a significant outcome of pULB113 mediated transfer in certain Shigella species.
- The capacity for R-prime formation and the stability of transferred genetic material vary among enteric bacterial species.