title: Development of a new in situ hybridization method for the detection of global bacterial DNA to provide early evidence of a bacterial infection in spontaneous bacterial peritonitis.
abstract: OBJECTIVE
  Despite the importance of identifying the causative pathogen(s), ascitic fluid cultures are occasionally negative in patients with spontaneous bacterial peritonitis (SBP). A novel strategy using the in situ hybridization (ISH) method was introduced to detect the bacterial genomic DNA phagocytized in the blood of patients with sepsis. In the present study, we developed a new ISH probe to detect global bacterial DNA (named as GB probe) and evaluated its utility for detecting the phagocytized bacterial DNA in SBP ascites.
  
  
  METHODS
  Hybridization of bacterial DNA with the GB probe was examined by dot-blot and ISH tests. In addition, the utility of the ISH method to detect the bacterial DNA in the leukocytes of SBP ascites was evaluated.
  
  
  RESULTS
  The GB probe hybridized with the genomic DNA of all 59 bacterial strains tested (59 species of 36 genus). Eleven of 51 patients with ascites (out of total 542 cirrhotic inpatients) were categorized as SBP. The ISH tests showed positive results in 10 of 11 SBP cases. However, the ISH tests all showed negative results in the 40 non-SBP ascitic samples. Therefore, the ISH tests yielded highly sensitive and specific results for detecting the phagocytized bacterial DNA in the leukocytes of SBP ascites. Moreover, all of the ISH test results were obtained within only one day.
  
  
  CONCLUSIONS
  Our newly established ISH method was found to provide both a rapid and sensitive detection of bacterial DNA in SBP ascites, thus suggesting its utility for providing early and direct evidence of bacterial infection in SBP ascites.
authors.affiliation: Division of Hepatobiliary and Pancreatic Disease, Department of Internal Medicine, Hyogo College of Medicine, Mukogawa-cho 1-1, Nishinomiya, Hyogo 663-8501, Japan.
authors.forename: Hirayuki
authors.initials: H
authors.lastname: Enomoto
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Inoue
authors.forename: Akio
authors.initials: A
authors.lastname: Matsuhisa
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Aizawa
authors.forename: Hiroyasu
authors.initials: H
authors.lastname: Imanishi
authors.forename: Masaki
authors.initials: M
authors.lastname: Saito
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Iwata
authors.forename: Hironori
authors.initials: H
authors.lastname: Tanaka
authors.forename: Naoto
authors.initials: N
authors.lastname: Ikeda
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Sakai
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Takashima
authors.forename: Soji
authors.initials: S
authors.lastname: Shimomura
authors.forename: Hiroko
authors.initials: H
authors.lastname: Iijima
authors.forename: Hideji
authors.initials: H
authors.lastname: Nakamura
authors.forename: Shuhei
authors.initials: S
authors.lastname: Nishiguchi
mesh_terms: D001201:Ascites / Q000150:complications; D001424:Bacterial Infections / Q000175:diagnosis* / Q000209:etiology / Q000382:microbiology*; D015331:Cohort Studies; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics*; D005260:Female; D006801:Humans; D017403:In Situ Hybridization / Q000379:methods*; D007962:Leukocytes / Q000382:microbiology; D008103:Liver Cirrhosis / Q000150:complications; D008297:Male; D008875:Middle Aged; D010538:Peritonitis / Q000175:diagnosis* / Q000209:etiology / Q000382:microbiology*; D010587:Phagocytosis; D011446:Prospective Studies
chemical_list: D004269:DNA, Bacterial