Canine cholecystitis in the presence of Hammondia spp. infection

Biliary protozoal infections are becoming increasingly recognized in dogs. Sarcocystidae protozoal infections (i.e Toxoplasma, Neospora, Hammondia, Besnoitia, and Sarcocystis spp.) are uncommon and host-specific. Diagnosis is traditionally dependent on tissue or serologic evaluation; sensitivity and specificity for these techniques can vary. PCR amplification and genetic sequencing provide a technique to expedite diagnosis and species identification in clinical biliary protozoal infections. Bile samples were collected percutaneously from two dogs presented to a referral hospital with mixed hepatopathies and hyperbilirubinemia. Diagnostic imaging revealed changes consistent with hepatobiliary inflammation, and tachyzoites were identified via bile cytology. Sera were positive for anti-Toxoplasma sp. (n = 1/2) and anti-Neospora sp. (n = 2/2) antibodies. Genomic isolation from bile and pan-Sarcocystidae (18S rRNA) PCR amplification were consistent with Hammondia spp. Additional 28S rRNA (LSU), alpha-tubulin (aTUB) and cytochrome b (CytB) gene-specific fragments were PCR amplified and sequenced. Phylogenetic analysis of LSU DNA fragment (584 bp) suggested both patients were infected with Hammondia spp. closely related to H. heydorni. Sequencing of aTUB (234 bp) and CytB (344 bp) revealed Patient 1 Hammondia spp. was more closely related to H. triffittae, while Patient 2 was more similar to H. heydorni. Treatment with clindamycin, and in one case additional enrofloxacin, resolved clinical and clinicopathologic changes in both dogs. This report highlights the importance of multilocus sequencing in protozoal identification in atypical infections, clinical manifestations of canine hepatobiliary hammondiasis, and need for regional Sarcocystidae prevalence studies.