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J Bacteriol, March 1998, p. 1533-1539, Vol. 180, No. 6
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Characterization of the Gene Cassette Required for Biosynthesis of the (alpha 1right-arrow 6)-Linked N-Acetyl-D-Mannosamine-1-Phosphate Capsule of Serogroup A Neisseria meningitidis

John S. Swartley,1,2,3 Li-Jun Liu,3,1,3 Yoon K. Miller,1,3 Larry E. Martin,1,3 Srilatha Edupuganti,1,3 and David S. Stephens1,2,3,*

Departments of Medicine1 and Microbiology and Immunology,2 Emory University School of Medicine, and Department of Veterans Affairs Medical Center,3 Atlanta, Georgia

Received 21 August 1997/Accepted 13 December 1997

The (alpha 1right-arrow6)-linked N-acetyl-D-mannosamine-1-phosphate meningococcal capsule of serogroup A Neisseria meningitidis is biochemically distinct from the sialic acid-containing capsules produced by other disease-associated meningococcal serogroups (e.g., B, C, Y, and W-135). We defined the genetic cassette responsible for expression of the serogroup A capsule. The cassette comprised a 4,701-bp nucleotide sequence located between the outer membrane capsule transporter gene, ctrA, and galE, encoding the UDP-glucose-4-epimerase. Four open reading frames (ORFs) not found in the genomes of the other meningococcal serogroups were identified. The first serogroup A ORF was separated from ctrA by a 218-bp intergenic region. Reverse transcriptase (RT) PCR and primer extension studies of serogroup A mRNA showed that all four ORFs were cotranscribed in the opposite orientation to ctrA and that transcription of the ORFs was initiated from the intergenic region by a sigma -70-type promoter that overlapped the ctrA promoter. The first ORF exhibited 58% amino acid identity with the UDP-N-acetyl-D-glucosamine (UDP-GlcNAc) 2-epimerase of Escherichia coli, which is responsible for the conversion of UDP-GlcNAc into UDP-N-acetyl-D-mannosamine. Polar or nonpolar mutagenesis of each of the ORFs resulted in an abrogation of serogroup A capsule production as determined by colony immunoblots and enzyme-linked immunosorbent assay. Replacement of the serogroup A biosynthetic gene cassette with a serogroup B cassette by transformation resulted in capsule switching from a serogroup A capsule to a serogroup B capsule. These data indicate that assembly of the serogroup A capsule likely begins with monomeric UDP-GlcNAc and requires proteins encoded by three other genes found in the serogroup A N. meningitidis-specific operon located between ctrA and galE.

* Corresponding author. Mailing address: Division of Infectious Diseases, Emory University School of Medicine, 69 Butler Street, SE, Atlanta, GA 30303. Phone: (404) 728-7688. Fax: (404) 329-2210. E-mail: DSTEP01{at}Emory.edu.




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