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Infection and Immunity, May 1999, p. 2464-2474, Vol. 67, No. 5
Oral Infection and Immunity Branch, National
Institute of Dental and Craniofacial Research, National Institutes of
Health, Bethesda, Maryland 20892,1 and
Department of Biochemistry, Molecular Biology, and Cell
Biology, Northwestern University, Evanston, Illinois
602082
Received 10 November 1998/Returned for modification 6 January
1999/Accepted 25 February 1999
Most human oral viridans streptococci participate in intrageneric
coaggregations, the cell-to-cell adherence among genetically distinct
streptococci. Two genes relevant to these intrageneric coaggregations
were identified by transposon Tn916 mutagenesis of
Streptococcus gordonii DL1 (Challis). A 626-bp sequence
flanking the left end of the transposon was homologous to
dltA and dltB of Lactobacillus
rhamnosus ATCC 7469 (formerly called Lactobacillus casei). A 60-kb probe based on this flanking sequence was used to
identify the homologous DNA in a fosmid library of S. gordonii DL1. This DNA encoded
D-alanine-D-alanyl carrier protein ligase that
was expressed in Escherichia coli from the fosmid clone. The cloned streptococcal dltA was disrupted by inserting an
ermAM cassette, and then it was linearized and transformed
into S. gordonii DL1 for allelic replacement.
Erythromycin-resistant transformants containing a single insertion in
dltA exhibited a loss of D-alanyl esters in
lipoteichoic acid (LTA) and a loss of intrageneric coaggregation. This
phenotype was correlated with the loss of a 100-kDa surface protein
reported previously to be involved in mediating intrageneric coaggregation (C. J. Whittaker, D. L. Clemans, and P. E. Kolenbrander, Infect. Immun. 64:4137-4142, 1996). The mutants retained
the parental ability to participate in intergeneric coaggregation with
human oral actinomyces, indicating the specificity of the mutation in altering intrageneric coaggregations. The mutants were altered morphologically and exhibited aberrant cell septa in a variety of
pleomorphs. The natural DNA transformation frequency was reduced 10-fold in these mutants. Southern analysis of chromosomal DNAs from
various streptococcal species with the dltA probe revealed the presence of this gene in most viridans streptococci. Thus, it is
hypothesized that D-alanyl LTA may provide binding sites for the putative 100-kDa adhesin and scaffolding for the proper presentation of this adhesin to mediate intrageneric coaggregation.
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Insertional Inactivation of Genes Responsible for the
D-Alanylation of Lipoteichoic Acid in
Streptococcus gordonii DL1 (Challis) Affects
Intrageneric Coaggregations


and
*
Corresponding author. Mailing address: National
Institutes of Health, Bldg. 30, Rm. 310, 30 Convent Dr., MSC 4350, Bethesda, MD 20892-4350. Phone: (301) 496-1497. Fax: (301) 402-0396. E-mail: pkolenbrander{at}dir.nidcr.nih.gov.
Present address: University of Michigan, Department of Pediatrics
and Communicable Diseases, Ann Arbor, MI 48109-2029.
Present address: Antiinfectives Research, Smith-Kline Beecham
Pharmaceuticals, Collegeville, PA 19426.
§
Present address: Institute for Animal Health, Compton Laboratory,
Compton, Newbury, Berks RG20 7NN, United Kingdom.
Present address: USDA, ARS, Roman L. Hruska U.S. Meat Animal
Research Center (MARC), Clay Center, NE 68933.
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