Cloning of genes affecting capsule expression in Staphylococcus aureus strain M

Area of Science:

• Microbiology
• Molecular Biology
• Genetics

Background:

• Staphylococcus aureus strain M exhibits high capsular polysaccharide production.
• Capsule production can be lost, with increased frequency at elevated temperatures (43°C).
• Extrachromosomal elements (plasmid, prophage) were present but not essential for capsulation.

Purpose of the Study:

• To clone and characterize the capsular (cap) genes responsible for polysaccharide production in S. aureus.
• To identify the genetic basis for capsule stability and loss.

Main Methods:

• Construction of a plasmid library from S. aureus M in S. aureus RN4200.
• Phage transduction using phage 80 alpha to transfer the library.
• Complementation of capsular-deficient (Cap-) mutants with recombinant plasmids.
• Chromosomal walking to identify adjacent cap genes.

Main Results:

• A recombinant plasmid conferring a capsular-positive (Cap+) phenotype was obtained.
• A 19.4 kb DNA fragment was identified that complemented most Cap- mutants.
• Evidence suggests the majority of cap genes are clustered together on the chromosome.

Conclusions:

• The capsular (cap) genes in S. aureus M are located in a cluster.
• This cluster is approximately 19.4 kb in size and contains the majority of genes required for capsule production.
• Understanding this gene cluster provides insight into the regulation of S. aureus capsular polysaccharide synthesis.