Targeted next generation sequencing panel for comprehensive testing of vector-borne pathogens

Diagnosing vector-borne disease (VBD) in dogs can be difficult for a number of reasons. First, there are many different disease-causing agents that can be transmitted from ticks/fleas, and the clinical signs caused by these agents in dogs can overlap. Additionally, because ticks/fleas can harbor more than one agent at a time, multiple pathogens may be passed to a dog with a single vector bite, resulting in co-infections.

VBD infections can initially present with non-specific signs, such as fever, lethargy, vomiting, diarrhea, and/or respiratory signs. Severe cases can be associated with neurologic signs. These signs can be a diagnostic conundrum. While initial blood work can be helpful and suggest VBD, it does not determine the infecting agent.

This study will develop a comprehensive next generation sequencing panel to detect and identify major VBD agents known to cause disease in dogs and to aid in diagnosis of active infections. Additionally, through parallel sequencing with this method, this panel will incorporate testing for additional infectious diseases that may cause GI, respiratory, or neurologic signs in dogs. The comprehensive nature of this sequencing panel should be a useful tool for surveillance of infectious diseases in the canine population for rapid identification of VBD in dogs and protection of pet owners from such zoonotic diseases.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02553

RESEARCHERS

Rebecca Wilkes, DVM; Purdue University
Amount: $2,500 (May 2019) $7,500 (November 2019)

Developing a Next Generation Sequencing Diagnostic Platform for Tick-Borne Diseases

Diagnostic tests based on the detection of DNA from harmful organisms in clinical samples have revolutionized veterinary medicine in the last decades. Currently, diagnostic panels for several vector-borne organisms are available through universities and private labs in the USA and abroad. However, the vast majority of results from sick dogs are negative, which frustrates veterinarians and dog owners trying to reach a definitive diagnosis. These panels are based on the detection of previously known DNA sequences of each pathogen, which limits their ability to detect novel organisms.

In this study, the investigators will adapt high-throughput nextgeneration sequencing (NGS) to the detection of tick-borne bacteria in dog blood in an effort to overcome the limitations of current diagnostics for tick-borne diseases. If successful, increasing the capabilities of NGS to detect infected dogs and to accurately determine which bacteria are responsible for disease will support the development of a better diagnostic tool to simultaneously advance canine and human health. This work expands on Dr. Diniz’s previous CHF-funded study #02292.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02528

RESEARCHERS

Pedro Diniz, DVM, PhD; Western University of Health Sciences
Amount: $5,000

Identifying Cellular Mechanisms of Inflammation During Canine Tick-Borne Diseases

Tick-borne diseases are found in all 50 states of the United States and are the most common vector-borne disease diagnosed in people in the US. The predominant disease is Lyme disease, caused by Borrelia burgdorferi and related species (sensu lato). Other important canine tick-borne diseases include those caused by Anaplasma platys, Anaplasma phagocytophilum (Anaplasmosis), Babesia canis, Babesia conradea and Babesia gibsonii (Babesiosis), and Ehrlichia canis, Ehrlichia chaffiensis and Ehrlichia ewingii (Ehrlichiosis).

Many of these diseases also affect people. Dogs can serve as sentinel species for human disease and there are many areas where the immune responses and disease outcomes are very similar in people and dogs, meaning that important lessons can be learned by sharing information between human and animal health (One Health).

The researchers will further investigate the dog’s immune system to determine which immune cells are responsible for the cure or creation of canine tick-borne disease. Through understanding which cells are responsible for causing disease, the goal is to then specifically target the molecules they produce using immunotherapy or immune modulation to improve treatment of tick-borne diseases in all dogs.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02383

RESEARCHERS

Christine A Petersen DVM PhD
University of Iowa
Amount: $5,000

Enhanced Testing for the Diagnosis of Bartonellosis in Dogs

Bartonellosis, a zoonotic bacterial disease of worldwide distribution, is caused by approximately 10 different Bartonella species. Bartonella are transmitted to canines and humans by ticks, fleas, lice, mites, and sand flies. Dr. Breitschwerdt’s laboratory demonstrated the first evidence for Bartonella infections in dogs in 1993.

Bartonella species have been associated with an expanding spectrum of important disease manifestations including anemia, endocarditis, hepatitis, lymphadenitis, myocarditis, thrombocytopenia and vascular tumor-like lesions. Infections can be life-threatening. Due to a lack of sensitive and reliable diagnostic assays, definitive diagnosis of bartonellosis in dogs remains a significant problem. Because these bacteria invade cells and infect tissues throughout the body, this chronic intracellular infection is difficult to cure with currently used antibiotic regimens.

This study will develop improved serodiagnostic tests for bartonellosis in dogs. These assays can also be used for world-wide sero-epidemiological prevalence studies, and to establish early and accurate diagnosis. Dr. Breitschwerdt’s research group has described concurrent infection in dogs, their owners and veterinary workers; this allows for a One Health approach to this important emerging infectious disease.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02287

RESEARCHERS

Dr. Edward B Breitschwerdt, DVM
North Carolina State University
Amount: $3,000

Broad-Range Detection of Canine Tick-Borne Disease and Improved Diagnostics Using Next-Generation Sequencing

Diagnostic tests based on the detection of DNA of infectious organisms from clinical samples have revolutionized veterinary medicine in the last decades. Currently, diagnostic panels for several tickborne organisms are available through universities and private laboratories in the USA and abroad. However, the vast majority of results from clinically ill dogs are negative for tick-borne diseases, which frustrates veterinarians and dog owners trying to reach a definitive diagnosis and improve treatment options.

These panels are based on the detection of previously known DNA sequences of each pathogen, with little room for detecting new organisms. Consequently, the current assays may suffer from “myopia”: a self-fulfilling effect that prevents the detection of new or emerging organisms.

Using an innovative approach, the investigators will employ next-generation sequencing (NGS) to overcome the limitations of current diagnostic technology. With NGS, the investigators can generate millions of individual gene sequencing reads from each clinical sample, allowing for the identification and characterization of multiple organisms from a single sample. Testing samples from dogs naturally exposed to tick-borne diseases, NGS will detect not only new organisms but also characterize genetic differences among known organisms.

The resulting dataset of a large number of DNA sequences of known tick-borne organisms and previously undetected organisms in naturally-infected dogs will support the development of diagnostic tools to simultaneously advance canine and human health.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02292

RESEARCHERS

Dr. Pedro Paul Diniz, DVM, PhD
Western University of Health Sciences
Amount: $3,000