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Journal of Clinical Microbiology, March 2006, p. 1065-1073, Vol. 44, No. 3
0095-1137/06/$08.00+0     doi:10.1128/JCM.44.3.1065-1073.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Molecular Subtyping of Poultry-Associated Type A Clostridium perfringens Isolates by Repetitive-Element PCR

G. R. Siragusa,1* M. D. Danyluk,2,{dagger} K. L. Hiett,1 M. G. Wise,1 and S. E. Craven1

Agricultural Research Service, U.S. Department of Agriculture, Russell Research Center, Athens, Georgia,1 Department of Food Science and Technology, University of Georgia, Athens, Georgia2

Received 18 August 2005/ Returned for modification 23 October 2005/ Accepted 27 October 2005

Clostridium perfringens strains (type A) isolated from an integrated poultry operation were subtyped using repetitive-element PCR with Dt primers. Isolates were obtained from fecal, egg shell, fluff, and carcass rinse samples as part of a previously reported temporally linked epidemiological survey. A total of 48 isolates of C. perfringens were obtained from different stages of the broiler chicken production chain from two separate breeder farms that supplied a single hatchery that in turn provided chicks to a single grow-out farm whose flocks were processed at a single plant. All 48 isolates were typeable (100% typeability) by repetitive-element PCR with Dt primers. This subtyping method was highly reproducible and discriminatory. By repetitive-element PCR with Dt primers, isolates were classified into four major branches with 12 subgroups or clades. The Simpson's index of discrimination was calculated to be 0.96 for groupings of >95% correlation. Toxin gene profiles of the isolates indicated that all of the isolates were C. perfringens alpha-toxin gene positive and 46 of 48 isolates were beta2-toxin gene positive. All strains were negative for beta- and epsilon-toxin genes. Repetitive sequence-based PCR was found to be a technically practical and reproducible means of subtyping C. perfringens libraries from specific epidemiological or production environment settings.

* Corresponding author. Mailing address: Agricultural Research Service, United States Department of Agriculture, Russell Research Center, Athens, GA 30604-5677. Phone: (706) 546-3596. Fax: (706) 546-3772. E-mail: siragusa{at}saa.ars.usda.gov.

{dagger} Present address: Department of Food Science, University of California—Davis, Davis, Calif.

Journal of Clinical Microbiology, March 2006, p. 1065-1073, Vol. 44, No. 3
0095-1137/06/$08.00+0     doi:10.1128/JCM.44.3.1065-1073.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.





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