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Detection of Bacteria in Phagocyte‐Smears from Septicemia‐Suspected Blood by In Situ Hybridization Using Biotinylated Probes
Author(s) -
Matsuhisa Akio,
Saito Yoshihiro,
Sakamoto Yoshimasa,
Keshi Hiroyuki,
Ueyama Hiroshi,
Aikawa Youko,
Kishi Youichiro,
Ohno Tsuneya
Publication year - 1994
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.1994.tb01816.x
Subject(s) - biology , in situ hybridization , bacteria , microbiology and biotechnology , hybridization probe , phagocyte , enterococcus faecalis , oligomer restriction , enterobacter , escherichia coli , phagocytosis , dna , oligonucleotide , staphylococcus aureus , biochemistry , gene expression , genetics , gene
We report herein the detection of intracellular bacteria in phagocyte‐smears obtained from septicemia‐suspected blood samples by in situ hybridization. This was obtained by using nick‐translated biotin‐11‐dUTP‐labeled DNA probes and streptavidin‐alkaline phosphatase conjugates for visualization of the hybridized signals. The probes were made from random genomic DNA clones of bacteria which are frequently the causative agents of bacteremia, such as Staphylococcus spp., Pseudomonas aeruginosa, Enterococcus faecalis, Escherichia coli, Klebsiella spp. and Enterobacter spp. When our in situ hybridization method was compared with conventional culture protocols for the ability to detect bacteria from the blood of patients suspected of having septicemia, 30 positive results were obtained in 50 specimens by in situ hybridization methods. In contrast, only 7 positive results were obtained by blood cultures. Thus, even if bacteria cannot be detected by conventional blood cultures and histology, our in situ hybridization method allows for direct observation of bacterial foci in circulating phagocytes and identification of the bacteria. Our investigations suggest that in septicemia, circulating polymorphonuclear neutrophils carry some surviving bacteria as well as metabolized bacterial DNA and RNA for a considerable period of time. Thus, our in situ hybridization method using the phagocyte‐smears have diagnostic value for detecting most bacteria which cause septicemia.