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Applied and Environmental Microbiology, October 2007, p. 6584-6594, Vol. 73, No. 20
0099-2240/07/$08.00+0     doi:10.1128/AEM.00537-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Localization and Visualization of a Coxiella-Type Symbiont within the Lone Star Tick, Amblyomma americanum{triangledown} ,{dagger}

Olga Klyachko, Barry D. Stein, Nathan Grindle, Keith Clay, and Clay Fuqua*

Department of Biology, Indiana University, Bloomington, Indiana 47405

Received 8 March 2007/ Accepted 1 August 2007

A Coxiella-type microbe occurs at 100% frequency in all Amblyomma americanum ticks thus far tested. Using laboratory-reared ticks free of other microbes, we identified the Amblyomma-associated Coxiella microbe in several types of tissue and at various stages of the life cycle of A. americanum by 16S rRNA gene sequencing and diagnostic PCR. We visualized Amblyomma-associated Coxiella through the use of a diagnostic fluorescence in situ hybridization (FISH) assay supplemented with PCR-based detection, nucleic acid fluorescent staining, wide-field epifluorescence and confocal microscopy, and transmission electron microscopy (TEM). Specific fluorescent foci were observed in several tick tissues, including the midgut and the Malpighian tubules, but particularly bright signals were observed in the granular acini of salivary gland clusters and in both small and large oocytes. TEM confirmed intracellular bacterial structures in the same tissues. The presence of Amblyomma-associated Coxiella within oocytes is consistent with the vertical transmission of these endosymbionts. Further, the presence of the Amblyomma-associated Coxiella symbiont in other tissues such as salivary glands could potentially lead to interactions with horizontally acquired pathogens.

* Corresponding author. Mailing address: Department of Biology, Indiana University, 1001 E. 3rd St., Jordan Hall 142, Bloomington, IN 47405. Phone: (812) 856-6005. Fax: (812) 855-6705. E-mail: cfuqua{at}indiana.edu

{triangledown} Published ahead of print on 24 August 2007.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

Applied and Environmental Microbiology, October 2007, p. 6584-6594, Vol. 73, No. 20
0099-2240/07/$08.00+0     doi:10.1128/AEM.00537-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.





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