Characterisation of bacteria in ascites--reporting the potential of culture-independent, molecular analysis.
Spontaneous bacterial peritonitis (SBP) is a severe complication of liver disease. A significant proportion of patients have culture-negative ascites, despite having similar signs, symptoms and mortality to those with SBP. Therefore, empirical antibiotic treatment for infection is often started without knowledge of the causative organisms. Here, we investigated the potential of molecular techniques to provide rapid and accurate characterisation of the bacteria present in ascitic fluid. Ascites samples were obtained from 29 cirrhotic patients undergoing clinically indicated therapeutic paracentesis. Bacterial content was determined by terminal restriction fragment length polymorphism (T-RFLP) analysis, quantitative polymerase chain reaction (PCR) and 16S ribosomal clone sequence analysis. Bacterial signal was detected in all samples, compared to three out of ten using standard methods. Bacterial loads ranged from 5.5 x 10(2) to 5.4 x 10(7) cfu/ml, with a mean value of 1.9 x 10(6) cfu/ml (standard deviation +/- 9.6 x 10(6) cfu/ml). In all but one instance, bacterial species identified by culture were also confirmed by molecular analyses. Preliminary data presented here suggests that culture-independent, molecular analyses could provide rapid characterisation of the bacterial content of ascites fluid, providing a basis for the investigation of SBP development and allowing early and targeted antibiotic intervention.
Molecular Microbiology Research Group, PSD, King's College London, London, UK. geraint.rogers@kcl.ac.uk
G B
GB
Rogers
L E
LE
Russell
P G
PG
Preston
P
P
Marsh
J E
JE
Collins
J
J
Saunders
J
J
Sutton
D
D
Fine
K D
KD
Bruce
M
M
Wright
D000328:Adult; D000368:Aged; D001201:Ascites / Q000382:microbiology*; D001419:Bacteria / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D015373:Bacterial Typing Techniques / Q000379:methods; D046508:Culture Techniques / Q000379:methods; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D005260:Female; D006801:Humans; D008103:Liver Cirrhosis / Q000382:microbiology* / Q000601:surgery; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods; D019152:Paracentesis; D010538:Peritonitis / Q000382:microbiology / Q000517:prevention & control; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D012336:RNA, Ribosomal, 16S / Q000032:analysis / Q000235:genetics
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S