Prospective Evaluation for Bacterial Infection in Hepatic Tissue and Bile of Cats with Diffuse Hepatobiliary Disease
The goals of this study are to determine the prevalence of bacterial infection in cats with Feline Inflammatory Hepatobiliary Disease (FIHD), to evaluate the diagnostic utility of hepatic tissue versus bile for the identification of bacterial pathogens in cats with FIHD, and to assess the utility of real-time PCR (RT-PCR) for detection of bacterial organisms in frozen hepatic tissue and bile.
Cases in which hepatic biopsy was performed as part of a complete diagnostic evaluation of diffuse hepatobiliary disease were included. Cases were excluded for the following reasons: antibiotic therapy prior to sampling, untreated hyperthyroidism, or the presence of a hepatic mass. Liver tissue was acquired using the method desired by the attending clinician, and bile was collected via cholecystocentesis. Control samples were obtained at the time of euthanasia from normal cats that were part of a breeding colony. Aerobic and anaerobic bacterial culture was performed on all samples. RT-PCR was performed on batched, frozen samples within 6 months of their acquisition. Cases were categorized as having acute neutrophilic cholangitis (ANC), chronic neutrophilic cholangitis (CNC), acute lymphocytic cholangitis (ALC), chronic lymphocytic cholangitis (CLC), hepatic lipidosis (HL), or other hepatic disease by one pathologist (TVW). DNA was extracted from samples using the QiaAmp DNA Mini Kit, (Qiagen) as described by the manufacturer. Amplification and DNA detection were carried out in a Smart Cycler (Cepheid) using 16S rRNA primers.
Thirty cases, including 7 cats with CNC, 4 cats with CLC, 1 cat with ALC, 4 cats with HL, 6 cats with other types of hepatic disease, and 8 control cats were included. Bile culture was positive in 5 cases, including 4 cats with CNC, and 1 cat with HL and biliary carcinoma. The following bacterial organisms were isolated from bile: E.coli (3), α-hemolytic Streptococcus (1), P. aeruginosa (1) and an unidentified anaerobic gram-positive rod. Hepatic tissue culture was positive in 2 cases. In both cases E.coli was isolated from both hepatic tissue and bile. RT-PCR of the bile was positive in 4 cases, all of which had positive bile cultures. RT-PCR of hepatic tissue was negative in all cases. In 3 of the 4 cases in which RT-PCR was positive, DNA sequencing identified the same organism that had been found using traditional bacterial culture on the bile. In one case, DNA sequencing identified B.fragilis, which had not been identified using bacterial culture. Culture and RT-PCR revealed no evidence of bacterial infection in any control samples.
We conclude that: biliary bacterial infection is more common in cats with CNC than in cats with other forms of FIHD and other types of hepatobiliary disease; bile is more useful than hepatic tissue for the purpose of bacterial identification; and RT-PCR appears to be a viable method to rapidly and accurately detect bacterial DNA in bile, but not in hepatic tissue.