Nucleotide sequence analysis of the complement resistance gene from plasmid R100

Area of Science:

• Microbiology
• Molecular Biology
• Genetics

Background:

• The R100 plasmid confers multiple antibiotic resistances to Escherichia coli.
• Some plasmids can also provide resistance to host defense mechanisms like serum complement.
• Understanding these resistance mechanisms is crucial for combating bacterial infections.

Purpose of the Study:

• To investigate the genetic basis of serum complement resistance conferred by the R100 plasmid in Escherichia coli.
• To characterize the specific DNA fragment and potential protein product responsible for this resistance.

Main Methods:

• Construction of a hybrid plasmid (pOW3) by combining fragments from R100 and pBR322.
• Assessing serum sensitivity of E. coli strains harboring different plasmids (pOW3, R100, pBR322, or no plasmid).
• Nucleotide sequencing of the relevant R100 fragment and Southern blot analysis.

Main Results:

• E. coli strains with R100 or pOW3 showed significantly reduced sensitivity (up to 10,000-fold) to serum complement killing.
• The R100 fragment contained an insertion sequence (IS2) and a large open reading frame encoding a protein with a signal sequence.
• The predicted protein showed high amino acid similarity to the traT protein of the E. coli F factor.
• The R100 serum resistance gene is distinct from the one found in ColV,I-K94.

Conclusions:

• The R100 plasmid encodes a specific gene that confers resistance to serum complement in Escherichia coli.
• This resistance mechanism appears to involve a protein similar to the traT protein.
• The identified serum resistance gene is novel and distinct from previously characterized genes.