Spectral imaging detection and counting of microbial cells in marine sediment

• Published in: Journal of microbiological methods 2003-04, Vol.53 (1), p.57-65
• Main Authors: Sunamura, Michinari, Maruyama, Akihiko, Tsuji, Takashi, Kurane, Ryuichiro
• Format: Article
• Language: English
• Subjects: Bacteria - classification; Bacteria - growth & development; Bacteria - isolation & purification; Biological and medical sciences; Colony Count, Microbial - methods; Direct counting; Earth Sciences; Fluorescent Dyes - chemistry; Fluorescent microscope; Fundamental and applied biological sciences. Psychology; Geologic Sediments - microbiology; Image analysis; Indoles - chemistry; Marine; Microbiology; Microscopy, Fluorescence; Oceanography; Sciences of the Universe; Seawater - microbiology; Spectral imaging; Spectrometry, Fluorescence - methods; SYBR Green II; Water Microbiology
• ISSN: 0167-7012; 1872-8359
• DOI: 10.1016/S0167-7012(02)00224-5

Semiautomated detection and counting techniques for microbial cells in soil and marine sediment using microscopic-spectral-imaging analysis were developed. Microbial cells in microscopic fields were selectively detected from other fluorescent particles by their fluorescent spectrum, based on the spectral shift between the conjunction and nonconjunction of DNA fluorochrome (SYBR Green II) with nucleic acids. Using this technique, microbial cells could be easily detected in soil and 30-cm deep sediment samples from Tokyo Bay, both of which contain particles other than microbial cells. Total cell density was semiautomatically estimated at 1–6×10 9 cells cm −3 of sediment sampled at different depths in Tokyo Bay, which corresponded to 65–106% (mean 88%) of visual direct counting. This technique may be useful for detecting microbial cells in soil and sediment samples from the deeper subsurface environment.