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Journal of Clinical Microbiology, April 2002, p. 1487-1492, Vol. 40, No. 4
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.4.1487-1492.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Identification of Two Phylogenetically Related Organisms from Feces by PCR for Detection of Salmonella spp.

Claudia Gentry-Weeks,1,2* H. Joel Hutcheson,2,3 Lisa Marie Kim,4,{dagger} Denise Bolte,1 Josie Traub-Dargatz,4 Paul Morley,4 Barbara Powers,1 and Michael Jessen1,{ddagger}

Colorado State University Veterinary Diagnostic Laboratory,1 Department of Microbiology, Immunology, and Pathology,2 Arthropod-Borne and Infectious Diseases Laboratories,3 Department of Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado 805234

Received 12 July 2001/ Returned for modification 18 October 2001/ Accepted 30 December 2001

Two previously reported PCR methods were evaluated to determine whether they are as sensitive and specific as conventional culture methods in detecting Salmonella spp. from feces. Bovine and equine feces were enriched overnight in brain heart infusion broth and assayed using PCR methods and primer sets described by other investigators. A total of 774 fecal specimens were tested using a primer set (invE-A primer set) that amplifies a region spanning the invasin E and A genes of Salmonella enterica serovar Typhimurium. A subset of these fecal specimens (306 of the 774 total) were tested using primers (hisJ primer set) that amplify a portion of the histidine transport J gene. The PCR required 24 h to obtain results, whereas it took 5 to 7 days to identify Salmonella spp. by culture. PCR detection of Salmonella spp. using the hisJ primers and the invE-A primers had a sensitivity of 93.3 and 80%, respectively, and a specificity of 85.6 and 98.6%, respectively, compared with bacterial culture. Amplification of 42 culture-negative fecal specimens (of 306 total specimens) generated a DNA fragment that corresponded to the molecular weight of the amplified hisJ gene. The hisJ-generated amplicons from six culture-negative and six culture-positive specimens were sequenced and analyzed using DNA sequence alignment and phylogenetic analysis software. A neighbor-joining dendrogram of the DNA sequences of both sets of hisJ amplicons revealed two distinct groups—one group of amplicons from culture-positive specimens identical to the hisJ gene of S. enterica serovar Typhimurium and a second group of amplicons from culture-negative specimens that were more closely related to hisJ of S. enterica serovar Typhimurium than to other hisJ sequences present in nucleotide databases.

* Corresponding author: Department of Microbiology, Immunology, and Pathology, Fort Collins, CO 80523. Phone: (970) 491-5411. Fax: (970) 491-1815. E-mail: Claudia.Gentry-Weeks{at}Colostate.edu.

{dagger} Present address: USDA, Agricultural Research Service, Athens, GA 30605.

{ddagger} Present address: ISC BioExpress, Kaysville, UT 84037.

Journal of Clinical Microbiology, April 2002, p. 1487-1492, Vol. 40, No. 4
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.4.1487-1492.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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