Expression of bacterial beta-glucuronidase in human bile: an in vitro study

Background. Bacterial beta -glucuronidase causes deconjugation of bilirubin diglucuronide resulting in the precipitation of calcium bilirubinate, whic h contributes to biliary sludge and stone formation. This process is attrib uted to enzyme activity produced by the aerobic enterobacteriaceae such as Escherichia coli and Klebsiella sp. The presence of Clostridium sp. was det ected in 48 of 56 intrahepatic stones by using polymerase chain reaction te chniques and cultured Clostridium perfringens from 14 of 18 unblocked bilia ry stents. Such bacteria are reported to produce beta -glucuronidase activi ty. The aim of this study was to determine the proportion of biliary bacter ia isolated from pigment stones and stents that produce beta -glucuronidase and to compare the enzyme activity expressed by the different bacteria in human bile. Methods; A total of 202 bacteria were isolated from blocked and unblocked b iliary stents and pigment ductal stones recovered from patients. Of these, 61 bacteria expressed beta -glucuronidase activity in brain heart infusion broth. These 61 bacteria were subsequently grown in human bile under aerobi c or anaerobic conditions to the early stationary phase and assayed for bet a -glucuronidase activity by using rho -nitrophenyl beta -D glucuronide as substrate. Results were normalized and reported as units of enzyme activity per milligram protein of the bacteria. Results: C perfringens produced beta -glucuronidase enzyme activity that wa s 34-fold higher than that for E coli, Staphylococcus, Corynebacterium sp., Bacillus sp., Enterococcus sp., Acinetobacter sp., Streptococcus sp., and Klebsiella sp. Conclusion: C perfringens with its higher enzyme activity is more important in the deconjugation of bilirubin diglucuronide than E coli and Klebsiella sp.