Korean Journal of Food Science and Technology (한국식품과학회지)

Korean Society of Food Science and Technology (한국식품과학회)
권호 표지
DOI QR코드

DOI QR Code

Enumeration of Weissella cibaria phage with cytometry, epifluorescence microscopy, and plaque assay

유세포분석기, 형광현미경, 용균반검사 분석을 이용한 Weissella cibaria 박테리오파지 정량분석 및 상관관계분석

  • Park, Won Jeong (Department of Food Science and Biotechnology, Gachon University) ;
  • Lim, Ga-Yeon (Department of Food Science and Biotechnology, Gachon University) ;
  • Park, Jong-Hyun (Department of Food Science and Biotechnology, Gachon University)
  • 박원정 (가천대학교 바이오나노대학 식품생물공학과) ;
  • 임가연 (가천대학교 바이오나노대학 식품생물공학과) ;
  • 박종현 (가천대학교 바이오나노대학 식품생물공학과)
  • Received : 2018.02.02
  • Accepted : 2018.02.21
  • Published : 2018.04.30
Download PDF
⟨ Previous Next ⟩

Abstract

Quantitative analysis for non-host infection bacteriophage was conducted for their enumeration. Flow cytometry and epifluorescence microscopy (EPM) were selected as counting methods. Correlation analysis was performed based on the plaque assay method on the existing host infection and consisted of Pearson correlation statistical analysis, regression analysis, and difference analysis. Analyses of 12 samples with flow cytometry and plaque assay methods showed that there was a correlation of 96.7% with Pearson correlation value r=0.967, $R^2$ 0.9352, and difference value of 1.063. Analyses of 12 samples with EPM and plaque assay methods showed that there was a correlation of 99.0% with Pearson correlation value r=0.990, $R^2$ 0.9811, and difference value of 1.605. Therefore, flow cytometry and epifluorescence microscopy would be effective for enumeration of Weissella cibaria bacteriophage with plaque assay.

유세포분석기, 형광현미경 계수법과 현재 주로 활용하는 숙주감염에 의한 용균반검사법에 의한 박테리오파지를 계수하는 방법을 Weissella cibaria 박테리오파지로 비교 연구하였다. 상관관계 분석은 피어슨 상관 통계분석, 회귀분석, 차이검정등으로 하였다. 12개의 시료로 유세포분석기와 용균반검사에 의한 계수법을 비교한 결과는 피어슨 분석으로는 96.7%의 상관관계(상관값 r=0.967, $R^2=0.9352$)와 차이검정은 1.063을 보였다. 형광현미경과 용균반검사에 의한 계수법 비교결과는 피어슨 분석으로는 99.0% (상관값 r=0.990, $R^2=0.9811$) 상관관계와 차이검정은 1.605이였다. 그러므로 유세포분석기와 형광현미경 계수법은 용균반검사 계수법과 같이 W. cibaria 박테리오파지 계수에 적합할 것으로 사료된다.

Keywords

References

  1. BerghO, BOrsheim KY, Bratbak G, Heldal M. High abundance of viruses found in aquatic environments. Nature 340: 467-468 (1989) https://doi.org/10.1038/340467a0
  2. Chen F, Lu JR, Binder BJ, Liu YC, Hodson RE. Application of digital image analysis and flow cytometry to enumerate marine viruses stained with SYBR Gold. Appl. Environ. Microbiol. 67: 539-5450 (2001) https://doi.org/10.1128/AEM.67.2.539-545.2001
  3. Chisholm SW, Olson RJ, Zettler ER, Goericke R, Waterbury JB, Welschmeyer NA. A novel free-living prochlorophyte abundant in the oceanic euphotic zone. Nature 334: 340-343 (1988) https://doi.org/10.1038/334340a0
  4. Duhamel S, Stephan J. Flow cytometric analysis of bacteria-and virus-like particles in lake sediments. J. Microbiol. Meth. 64: 316-332 (2006) https://doi.org/10.1016/j.mimet.2005.05.008
  5. Ferris MM, Stoffel CL, Maurer TT, Rowlen KL. Quantitative intercomparison of transmission electron microscopy, flow cytometry, and epifluorescence microscopy for nanometric particle analysis. Anal. Biochem. 304: 249-256 (2002) https://doi.org/10.1006/abio.2002.5616
  6. Fuhrma JA, Suttle CA. Viruses in marine planktonic systems. Oceanography. 6: 51-63 (1993) https://doi.org/10.5670/oceanog.1993.14
  7. Hara S, Terauchi K, Koike I. Abundance of viruses in marine waters: assessment by epifluorescence and transmission electron microscopy. Appl. Environ. Microbiol. 57: 2731-2734 (1991)
  8. Hennes KP, Suttle CA. Direct counts of viruses in natural waters and laboratory cultures by epifluorescence microscopy. Limnol. Oceanogr. 40: 1050-1055 (1995) https://doi.org/10.4319/lo.1995.40.6.1050
  9. Kropinski AM, Mazzocco A, Waddell TE, Lingohr E, Johnson RP. Enumeration of bacteriophages by double agar overlay plaque assay. Bacteriophages: Methods and Protocols, Vol. 1: Isolation, Characterization, and Interactions, Humana Press. New York. USA. pp. 69-76 (2009)
  10. Lu Z, Perez-Diaz IM, Hayes JS, Breidt F. Bacteriophage ecology in a commercial cucumber fermentation. Appl. Environ. Microbiol. 78: 8571-8578 (2012) https://doi.org/10.1128/AEM.01914-12
  11. Martha RJC, Andrew MK. Bacteriophages Methods and Protocols. Vol. 1, Isolation, Characterization, and Interactions. In: Alice C. Ortmann & Curtis A. Suttle. Determination of Virus Abundance by Epifluorescence Microsopy. Humana Press. New York. USA. pp. 87-95 (2009a)
  12. Martha RJC, Andrew MK. Bacteriophages Methods and Protocols Vol. 1, Isolation, Characterization, and Interactions. In: Elizabeth Kutter. Phage Host Range and Efficiency of Plating. Humana Press. New York. USA. pp. 141-149 (2009b)
  13. Noble RT, Fuhrman JA. Use of SYBR Green I for rapid epifluorescence counts of marine viruses and bacteria. Aquat. Microb. Ecol. 14: 113-118 (1998) https://doi.org/10.3354/ame014113
  14. Ortmann AC, Suttle CA. Determination of virus abundance by epifluorescence microscopy. Bacteriophages: Methods and Protocols, Vol. 1, Isolation, Characterization, and Interactions, Humana Press. New York. USA. pp. 87-95 (2009)
  15. Proctor LM, Fuhrman JA. Viral mortality of marine bacteria and cyanobacteria. Nature 343: 60-62 (1990) https://doi.org/10.1038/343060a0