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Infection and Immunity, June 2003, p. 3320-3328, Vol. 71, No. 6
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.6.3320-3328.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

Aloka B. Bandara,1 Mark L. Lawrence,2 Hugo P. Veit,1 and Thomas J. Inzana1*

Center for Molecular Medicine and Infectious Diseases, Virginia-Maryland Regional College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061,1 Department of Basic Sciences, College of Veterinary Medicine, Mississippi State University, Mississippi State, Mississippi 397622

Received 7 October 2002/ Returned for modification 18 December 2002/ Accepted 10 March 2003

The capsular polysaccharide (CP) of Actinobacillus pleuropneumoniae is required for virulence of the bacteria in swine. However, a molecular investigation of whether the type or quantity of CP affects A. pleuropneumoniae virulence has not been reported. To initiate this investigation, a DNA region downstream of conserved genes required for CP export in A. pleuropneumoniae serotype 1 was cloned and sequenced. Three open reading frames, designated cps1A, cps1B, and cps1C, were identified that had amino acid homology to bacterial carbohydrate biosynthesis genes. A kanamycin resistance cassette (Kanr) was inserted into a 750-bp deletion spanning cps1AB or into a 512-bp deletion in cps1B only, and the constructs were cloned in a suicide vector. The Kanr gene was then transferred into the chromosome of strain 4074 by homologous recombination to produce strain 4074{Delta}cps1N and strain 4074{Delta}cps1B, respectively. Strain 4074{Delta}cps1N produced no detectable CP, but strain 4074{Delta}cps1B made 15% of the serotype 1 CP made by the parent strain, 4074, as determined by enzyme-linked immunosorbent assay and precipitation of free CP. The cps1ABC genes of strain 4074 and the cps5ABC and cps5ABCDE genes of serotype 5a strain J45 were cloned into the shuttle vector pLS88 and electroporated into 4074{Delta}cps1N to produce 4074{Delta}cps1N(pABcps101), 4074{Delta}cps1N(pJMLcps53), and 4074{Delta}cps1N(pABcps55), respectively. Strain 4074{Delta}cps1N(pABcps101) produced about 33% of the serotype 1 CP produced by strain 4074. Strains 4074{Delta}cps1N(pJMLcps53) and 4074{Delta}cps1N(pABcps55) produced serotype 5a CP in similar quantity or in fourfold excess, respectively, to that produced by strain 4074. With intratracheal challenge in pigs at similar dosages, the order of virulence of strains producing serotype 1 CP (assessed by mortality, lung consolidation, hemorrhage, and fibrinous pleuritis) was the following: strain 4074 > strain 4074{Delta}cps1N(pABcps101) >= strain 4074{Delta}cps1N > strain 4074{Delta}cps1B. Strain 4074{Delta}cps1N(pJMLcps53) was less virulent than strain 4074{Delta}cps1N(pABcps55). However, both strains produced serotype 5a CP in similar or greater quantities than was observed for production of serotype 1 CP by the parent strain, 4074, but were less virulent than the parent strain. Therefore, the amount of serotype 1 or 5a CP produced by isogenic strains of A. pleuropneumoniae correlated with the virulence of the bacteria in pigs. However, virulence was also influenced by the type of CP produced or by its mechanism of expression.

* Corresponding author. Mailing address: 1410 Prices Fork Rd., CMMID, VA-MD Regional College of Veterinary Medicine, Virginia Tech, Blacksburg, VA 24061. Phone: (540) 231-4692. Fax: (540) 231-3426. E-mail: tinzana{at}vt.edu.

Editor: F. C. Fang

Infection and Immunity, June 2003, p. 3320-3328, Vol. 71, No. 6
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.6.3320-3328.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Bosse, J. T., Durham, A. L., Rycroft, A. N., Kroll, J. S., Langford, P. R. (2009). New Plasmid Tools for Genetic Analysis of Actinobacillus pleuropneumoniae and Other Pasteurellaceae. Appl. Environ. Microbiol. 75: 6124-6131 [Abstract] [Full Text]  
  • Ali, T., Oldfield, N. J., Wooldridge, K. G., Turner, D. P., Ala'Aldeen, D. A. A. (2008). Functional Characterization of AasP, a Maturation Protease Autotransporter Protein of Actinobacillus pleuropneumoniae. Infect. Immun. 76: 5608-5614 [Abstract] [Full Text]  


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