Behaviour of bacteriophage Mu in Acinetobacter calcoaceticus EBF65/65

Area of Science:

• Microbiology
• Molecular Biology
• Bacteriology

Background:

• Plasmid transfer efficiency between bacterial species is often limited by host defense mechanisms.
• Acinetobacter calcoaceticus possesses cryptic plasmids, such as pAV2, potentially encoding restriction/modification systems.
• Bacteriophage Mu is a versatile genetic tool used in bacterial research.

Purpose of the Study:

• To investigate the inefficiency of RP4::Mu plasmid transfer into Acinetobacter calcoaceticus EBF65/65.
• To identify and characterize restriction/modification systems in Acinetobacter calcoaceticus.
• To explore the potential of 'plasmid suicide by Mu' for directed chromosome mobilization in Acinetobacter calcoaceticus.

Main Methods:

• Conjugative transfer experiments using RP4 and RP4::Mu plasmids between Escherichia coli K12 and Acinetobacter calcoaceticus EBF65/65 strains.
• Analysis of plasmid-borne defective Mu prophages and their ability to produce phage particles.
• Utilizing a mutant Mu prophage (Mu cts62 r23) to generate non-defective RP4::Mu plasmids.
• Thermoinduction experiments and plaque assays on different Escherichia coli K12 strains.

Main Results:

• RP4::Mu plasmid transfer to Acinetobacter calcoaceticus was inefficient, particularly in strains harboring pAV2.
• Transferred plasmids carried defective Mu prophages, indicating host-induced defects.
• Evidence for a second restriction/modification system in Acinetobacter calcoaceticus primarily affecting Mu DNA was found.
• Non-defective Mu plasmids were obtained using a mutant prophage, but Mu expression in Acinetobacter calcoaceticus remained poor.

Conclusions:

• Acinetobacter calcoaceticus possesses a novel restriction system that significantly impacts Mu DNA.
• The observed 'plasmid suicide by Mu' phenomenon suggests a potential for developing directed chromosome mobilization strategies in Acinetobacter calcoaceticus.
• Further research into this restriction system could enhance genetic manipulation techniques in Acinetobacter species.