Molecular Characterization of Rickettsial Diseases in Dogs

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Title: Molecular Characterization of Rickettsial Diseases in Dogs
Author: Kidd, Linda Benjamin
Advisors: Robert Kelly, Committee Member
Paul Orndorff, Committee Member
Ed Breitschwerdt, Committee Chair
Wayne Tompkins, Committee Member
Scott Laster, Committee Member
Abstract: Spotted Fever Group Rickettsia are important causes of morbidity and mortality worldwide. These arthropod borne, obligately intracellular organisms are notoriously difficult to detect in blood samples from infected patients. Standard diagnostic techniques do not differentiate among SFG members, thus the species of infecting Rickettsia is presumed based on geographic location. With the advent of molecular biology, newly discovered and previously known pathogenic and "non-pathogenic" SFG Rickettsia have been associated with disease in people in expanding regions of the world. R. rickettsii, the cause of Rocky Mountain Spotted Fever, is arguably the most well known and well characterized SFG Rickettsia The manifestations of this disease are similar in dogs and people, and because infection can precede or occur simultaneously with infection in their human companions, dogs are considered sentinels for the disease. Despite this, the species of Rickettsia infecting dogs with RMSF had never been characterized using molecular techniques. Using PCR, we amplified and sequenced portions of three genes from SFG Rickettsia isolated from dogs and people with RMSF in an endemic region. Gene sequence based criteria were applied and the isolates were identified as R. rickettsii. This study provides support that naturally occurring RMSF in dogs is comparable to the disease in people. Rickettsia rickettsii has been declared a Select Agent by the CDC and a Category C priority pathogen by the NIAID due to concerns that it is amenable for use in a bioterrorist attack. As a result of this study, 15 consensus sequences of genes amplified from R. rickettsii naturally infecting dogs and people in North Carolina have been deposited in GENBANK (Accesion numbersDQ15680-DQ15694). Knowledge of sequences of naturally occurring isolates may help identify aberrant strains intentionally released as a result of an act of bioterrorism. Amplifying DNA from cultures of Rickettsia does not require a particularly sensitive PCR. Due to the nature of these organisms, sensitive assays are required in the clinical setting. No diagnostic tool that is sensitive, and can differentiate among species of infecting Rickettsia has been validated for use in infected dogs. We have created a PCR that amplifies a portion of the ompA gene from infected dog blood with a limit of detection of 1.5-30 copies of SFG Rickettsia. Sequencing of the product differentiates a number of different species. Using this tool we documented the presence of R. conorii ssp conorii DNA in the blood of three male Sicilian Yorkshire Terriers. These dogs had clinical illness compatible with acute rickettsiosis. This provides unique evidence that R. conorii may infect dogs and cause disease in this host. Future studies should investigate the role of R. conorii and other SFG Rickettsia as disease causing agents in dogs. Identifying SFG Rickettsia in naturally infected dogs may have implications for their human companions due to their potential role as sentinels for this group of illnesses.
Date: 2006-07-26
Degree: PhD
Discipline: Immunology

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