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Title: DETECTION OF FRANCISELLA TULARENSIS IN INFECTED MAMMALS AND VECTORS USING PROBE-BASED PCR

Author
item HIGGINS, JAMES
item HUBALEK, ZDENEK - MED ZOOLOGY, CZECH REP.
item HALOUZKA, JIRI - MED ZOOLOGY, CZECH REP.
item ELKINS, KAREN - ROCKVILLE, MD
item SJOSTEDT, ANDERS - UMEA UNIV., SWEDEN
item SHIPLEY, MICHELLE - FORT DETRICK, MD
item IBRAHIM, SOFI - FORT DETRICK, MD

Submitted to: Journal of Clinical Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/29/1999
Publication Date: 2/1/2000
Citation: N/A

Interpretive Summary: Tularemia is a serious bacterial disease of humans and companion animals in the United States and in Europe. The infection is usually transmited by tick bite. The authors of this paper used the Polymerase Chain Reaction (PCR) assay, in conjuction with oligonucleotide DNA probes, to detect tularemia bacteria in experimentally infected mice, and in ticks. The PCR assays can be performed in a short time, on large numbers of samples, which may be advantageous when detection is needed for suspected bioterrorism or biowarfare incidents.

Technical Abstract: We investigated the use of a TaqMan 5 nuclease assay (5NA) directed against the tul 4 gene, for detection of this organism in experimentally infected mice, and in field-collected tick vectors. We also evaluated the use of specially formulated filter paper (FTA) for rapid sample preparation The 5NA had a detection limit of 1 pg genomic DNA (<100 cfu), and could be completed within several hours. The PCR-EIA had detection limits of 1 pg genomic DNA, and 10 ag (22 copies) of cloned insert, but takes longer to perform. Both assays were genus-specific, and successfully detected F tularensis in mouse tisues (5NA) and in tick extracts (PCR-EIA). The FTA paper provided inexpensive, rapid, template preparation for the tick extracts, mouse tissues, and DNA obtained from clinical specimens. These probe-based assays have the potential to provide rapid, real-time/high- throughput molecular diagnostics in field situations.