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Applied and Environmental Microbiology, August 2002, p. 3750-3758, Vol. 68, No. 8
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.8.3750-3758.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Culture-Based and Non-Growth-Dependent Detection of the Burkholderia cepacia Complex in Soil Environments

Suzanne C. M. Miller,1 John J. LiPuma,2 and Jennifer L. Parke1,3*

Department of Crop and Soil Science,1 Department of Botany and Plant Pathology, Oregon State University, Corvallis, Oregon 97331-7306,3 Department of Pediatrics and Communicable Diseases, University of Michigan Medical School, Ann Arbor, Michigan 48109-06462

Received 10 December 2001/ Accepted 25 April 2002

Burkholderia cepacia complex (Bcc) bacteria reside in soil, plant rhizospheres, and water, but their prevalence and distribution in outdoor environments is not clear. We sampled a variety of soil and rhizosphere environments with which people may have contact: playgrounds, athletic fields, parks, hiking trails, residential yards, and gardens. A total of 91 sites was sampled in three large U.S. cities. In the first phase of the study, putative Bcc isolates were recovered on Burkholderia cepacia selective agar and trypan blue tetracycline medium and subsequently examined for biochemical reactivity and growth at 32 and 22°C. Isolates were further examined by PCR assays targeting Bcc-specific ribosomal DNA and recA gene sequences. Among the 1,013 bacterial isolates examined, 68 were identified as Bcc; 14 (15%) of 91 sampled sites yielded Bcc isolates. In the second phase, DNA was extracted directly from soil samples and examined with PCR assays targeting Bcc 16S rRNA gene sequences. Either 82 or 93% of the soil samples were positive for at least one Bcc genomovar, depending on the PCR assay system used. Cloning and sequencing were performed to check the specificity of the PCR assays. Sequence analysis of the 463-bp 16S rRNA inserts from eight clones indicated that all were from members of the Bcc. The four soil samples from which these clones were generated did not yield isolates identified as Bcc. Based on PCR detection, Bcc appears to be prevalent in soil from urban and suburban environments. Culture-based recovery of Bcc may underestimate environmental populations.

* Corresponding author. Mailing address: Department of Crop and Soil Science, Oregon State University, ALS 3017, Corvallis, OR 97331-7106. Phone: (541) 737-8170. Fax: (541) 737-5725. E-mail: Jennifer.Parke{at}orst.edu.

Applied and Environmental Microbiology, August 2002, p. 3750-3758, Vol. 68, No. 8
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.8.3750-3758.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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