Bacterial cholecystitis and mucocele formation are considered the most common causes of gallbladder disease in dogs. Positive bacterial growth of bile samples from these patients is less common than is our suspicion for an infectious culprit. Studies to determine if bacteria reside in the gallbladder of healthy dogs or dogs with culture-negative gallbladder disease has not been reported. The present study performed next-generation sequencing of gallbladder bile and mucus from dogs with and without gallbladder disease to characterize the bacterial community composition for presence of a commensal microbiome or unrecognized pathogenic bacteria.
Bile samples were obtained by cholecystocentesis from 22 healthy dogs (CTRL) and 36 dogs with suspected bacterial cholecystitis (SBC). Gallbladder mucus was obtained after cholecystectomy in 22 dogs with mucocele formation (GBM). Presence of Eubacteria was examined by aerobic and anaerobic culture, cytology, and fluorescence in situ hybridization (FISH). All samples underwent DNA extraction, PCR amplification of the V3-V4 region of the 16S rRNA gene, and Illumina-based sequencing.
Dogs in the CTRL group were significantly younger (median 3, range, 1-4 yrs) than dogs in SBC (9.6, 1.2-15 yrs) or GBM (10.9, 4.8-16 yrs) groups (p < 0.001). GBM dogs had higher neutrophil counts (p = 0.006) and higher total serum bilirubin (p = 0.043) compared to SBC dogs. No CTRL dogs were positive for the presence of bacteria by means of culture, cytology, or FISH. Similar numbers of SBC and GBM dogs had positive results of bacterial culture, cytology, or FISH. Compared to conventional means to identify bacteria, a majority of CTRL (13/22; 59%), SBC (14/36; 39%) and GBM dogs (14/22; 64%) harbored a diverse microbiome. PCR targeting of the 16S rRNA gene was highly effective in identifying the same bacteria (i.e. E. coli and Enterococcus spp.) as were diagnosed by culture or revealing their presence in culture negative dogs with recent history of antibiotics. The remaining 16S rRNA gene positive, culture negative dogs demonstrated a diverse microbiome consisting predominantly of Proteobacteria (Neisseriaceae, Acinetobacter), Firmicutes (Staphylococcus, Streptococcus, Lactococcus), Actinobacteria (Corynebacterium, Propionibacterium), Cyanobacteria (Streptophyta), and Bacteroidetes. Obvious differences in the between CTRL and GBM dogs were not observed.
Results of these studies identify that a majority of normal and diseased dogs harbor a rich and diverse bile microbiome. This microbiota is likely to contain previously unrecognized beneficial as well as deleterious bacteria. Tailoring bile culture media to routinely identify these bacteria is likely to improve our understanding and diagnosis of canine hepatobiliary disease.
Resident in Small Animal Internal Medicine
North Carolina State University
Dr. Ashley Hartley earned her PhD in Infectious Diseases in 2013 and subsequently completed her DVM training in 2014 at the University of Georgia. Her graduate work focused on generating mutant Trypanosoma cruzi parasite lines and evaluating canine immune responses. After graduating from veterinary school, she completed a small animal medicine and surgery internship at the University of Tennessee. Currently, she is a small animal internal medicine resident at North Carolina State University. Her clinical and research interests are primarily in infectious disease, immunology, and hepatobiliary disease.
Friday, June 15
4:45 PM – 5:00 PM
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