Email updates

Keep up to date with the latest news and content from BMC Bioinformatics and BioMed Central.

This article is part of the supplement: UT-ORNL-KBRIN Bioinformatics Summit 2010

Open Access Poster presentation

Molecular analysis of Trypanosoma cruzi isolates obtained from raccoons in Warren and Barren counties of Kentucky

LiPeng Bi1, Chad Groce2 and Cheryl Davis1*

Author Affiliations

1 Department of Biology, Western Kentucky University, Bowling Green, KY 42101, USA

2 College of Veterinary Medicine, Auburn University, Auburn, AL 36849, USA

For all author emails, please log on.

BMC Bioinformatics 2010, 11(Suppl 4):P3  doi:10.1186/1471-2105-11-S4-P3


The electronic version of this article is the complete one and can be found online at: http://www.biomedcentral.com/1471-2105/11/S4/P3


Published:23 July 2010

© 2010 Davis et al; licensee BioMed Central Ltd.

Background

Trypanosoma cruzi, the etiologic agent of Chagas disease, infects a variety of wild mammals in the southern United States, but it has only recently been isolated from raccoons trapped in the state of Kentucky. The purpose of the present study was to use a molecular typing approach [1,2], followed by DNA sequencing to determine the genotypes (type I, or types IIa–IIe) of 15 of the Kentucky isolates.

Methods

DNA samples were prepared from 15 T. cruzi- isolates using a Qiagen mini kit, and PCR amplification was performed using published primers for the 24S a rDNA sequence (D71 and D72), the non–transcribed spacer of the mini–exon genes (TC, TC1, and TC2), the 18S rDNA sequence (V1 and V2), and TCZ1 and TCZ2 primers that amplify a 188–base pair segment of the repetitive 195–bp nuclear DNA sequence of T. cruzi. DNA sequencing (ABI 3130 Genetic Analyzer) was performed on all amplification products obtained from the PCR analysis of the RW2 and RB12 isolates (randomly selected to represent both Warren and Barren counties of Kentucky). The resulting sequences were edited before analysis using the BLAST database of the National Center for Biotechnology Information (NCBI) Genbank.

Results

All 15 isolates were positively confirmed as T. cruzi based upon PCR amplification of a195 bp repetitive genomic DNA sequence, and all 15 isolates showed identical PCR amplification results with all 4 sets of T. cruzi–specific primers. Two positive PCR samples were randomly selected for further DNA sequence analysis, and all samples were positively identified as the type IIa genotype of T. cruzi with max identities ranging from 94%-99%.

Conclusions

The results of this study confirmed that all hemoculture isolates obtained from raccoons trapped in Warren and Barren counties of Kentucky are T. cruzi. Furthermore, all BLAST comparisons of amplicon DNA sequences showed high sequence identity to Type IIa strains of T. cruzi. The Type IIa strain of T. cruzi is the most commonly reported genotype reported from raccoons trapped in the U.S.A.

Acknowledgements

Author C. Davis gratefully acknowledges administrative support from NIH Grant Number 2 P20 RR–16481 from the National Center for Research Resources.

References

  1. Brisse S, Verhoef J, Tibayrenc M: Characterisation of large and small subunit rRNA and mini-exon genes further supports the distinction of six Trypanosoma cruzi lineages.

    Int J Parasitol 2001, 31:1218-1226. PubMed Abstract | Publisher Full Text OpenURL

  2. Roeling D, Brown E, Barnabe C, Tibayrenc M, Steurer F, Yabsley M: Molecular typing of Trypanosoma cruzi isolates, United States.

    Emerg Infect Dis 2008, 14:1123-1125. PubMed Abstract | Publisher Full Text | PubMed Central Full Text OpenURL