Dataset Information

Rapid High-Resolution Melt Analysis of Cytauxzoon felis Cytochrome b To Aid in the Prognosis of Cytauxzoonosis.

ABSTRACT: Cytauxzoon felis is a virulent, tick-transmitted, protozoan parasite that infects felines. Cytauxzoonosis was previously thought to be uniformly fatal in domestic cats. Treatment combining atovaquone and azithromycin (A&A) has been associated with survival rates of over 60%. Atovaquone, a ubiquinone analogue, targets C. felis cytochrome b (cytb), of which 30 unique genotypes have been identified. The C. felis cytb genotype cytb1 is associated with increased survival rates in cats treated with A&A. The purpose of this study was to design a PCR panel that could distinguish C. felis cytb1 from other cytochrome b genotypes. Primer pairs were designed to span five different nucleotide positions at which single-nucleotide polymorphisms in the C. felis cytb gene had been identified. Through the use of high-resolution melt analysis, this panel was predicted to distinguish cytb1 from other cytb genotypes. Assays were validated using samples from 69 cats with cytauxzoonosis for which the C. felis cytb genotypes had been characterized previously. The PCR panel identified C. felis cytb1 with 100% sensitivity and 98.2% specificity. High-resolution melt analysis can rapidly provide prognostic information for clients considering A&A treatment in cats with cytauxzoonosis.

SUBMITTER: Schreeg ME

PROVIDER: S-EPMC4508407 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature

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Publications

Rapid High-Resolution Melt Analysis of Cytauxzoon felis Cytochrome b To Aid in the Prognosis of Cytauxzoonosis.

Schreeg Megan E ME Marr Henry S HS Tarigo Jaime L JL Cohn Leah A LA Levy Michael G MG Birkenheuer Adam J AJ

Journal of clinical microbiology 20150527 8

Cytauxzoon felis is a virulent, tick-transmitted, protozoan parasite that infects felines. Cytauxzoonosis was previously thought to be uniformly fatal in domestic cats. Treatment combining atovaquone and azithromycin (A&A) has been associated with survival rates of over 60%. Atovaquone, a ubiquinone analogue, targets C. felis cytochrome b (cytb), of which 30 unique genotypes have been identified. The C. felis cytb genotype cytb1 is associated with increased survival rates in cats treated with A& ...[more]

PMID: 26019197

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Project description:Cytauxzoon felis is a tick-borne hemoprotozoan parasite that causes life-threatening disease in domestic cats in the United States. Currently, the platforms for C. felis research are limited to natural or experimental infection of domestic cats. This study aims to develop an alternative model by infecting Amblyomma americanum ticks with C. felis via direct injection. Amblyomma americanum adults were injected with C. felis-infected feline erythrocytes through two routes: directly into the digestive tract through the anal pore (IA injection), or percutaneously into the tick hemocoel (IH injection). RNAscope® in situ hybridization (ISH) was used to visualize the parasites within the ticks at different time points after injection. Four months after injection, ticks were divided into 3 infestation groups based on injection methods and inoculum type and fed on 3 naïve cats to assess the ticks' ability to transmit C. felis. Prior to the transmission challenge, selected ticks from each infestation group were tested for C. felis RNA via reverse transcription-PCR (RT-PCR). In both IA- and IH-injected ticks, ISH signals were observed in ticks up to 3 weeks after injection. The number of hybridization signals notably decreased over time, and no signals were detected by 4 months after injection. Prior to the transmission challenge, 37-57% of the sampled ticks were positive for C. felis RNA via RT-PCR. While the majority of injected ticks successfully attached and fed to repletion on all 3 cats during the transmission challenge, none of the cats became infected with C. felis. These results suggest that injected C. felis remained alive in ticks but was unable to progress to infective sporozoites after injection. It is unclear why this infection technique had been successful for other closely related tick-borne hemoprotozoa and not for C. felis. This outcome may be associated with uncharacterized differences in the C. felis life cycle, the lack of the feeding or molting in our model or absence of gametocytes in the inoculum. Nonetheless, our study demonstrated the potential of using ticks as an alternative model to study C. felis. Future improvement of a tick model for C. felis should consider other tick species for the injection model or utilize infection methods that more closely emulate the natural infection process.

Dataset Information

Rapid High-Resolution Melt Analysis of Cytauxzoon felis Cytochrome b To Aid in the Prognosis of Cytauxzoonosis.

Publications

Rapid High-Resolution Melt Analysis of Cytauxzoon felis Cytochrome b To Aid in the Prognosis of Cytauxzoonosis.

Similar Datasets

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