Abstract
Application and availability of real-time polymerase chain reaction (PCR) assay to detect and quantify the Noctiluca scintillans zoospore were investigated seasonally. Specific primer set for N. scintillans 18S rDNA was designed and applied to real-time PCR assay using the serial dilutions of N. scintillans zoospores. The real-time PCR assays with Ns63F and Ns260R primers were applied to sea water samples collected weekly in Manazuru Port of Sagami Bay, Japan from April 2005 to June 2006. We developed effective DNA preparation steps for collecting the template DNA of N. scintillans zoospore: size fraction and filter concentration of the water samples, fixation with Lugol solution, cell lysis, and purification. This method is useful for the monitoring of the zoospores of N. scintillans, and can also be used for other small and physiologically fragile planktonic cell. Variation in the density of zoospore was successfully detected in the field samples. The peak density of N. scintillans zoospore was observed to occur just before or at the same time as the peak of the vegetative cells. Moreover, zoospores were detected in seawater even when the vegetative cells were not observed. The presence of zoospore was found all year round in the present study. In this regards, this information is essential for the study of the life cycle and seasonal variation of N. scintillans in the coastal waters.
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Bowers HA, Tengs T, Glasgow HB Jr, Burkholder JM, Rublee PA, Oldach DW (2000) Development of real-time PCR assays for rapid detection of Pfiesteria piscicida and related dinoflagellates. Appl Environ Microbiol 66, 4641–4648
Dela-Cruz J, Ajani P, Lee R, Pritchard T, Suthers I (2002) Temporal abundance patterns of the red tide dinoflagellate Noctiluca scintillans along the southeast coast of Australia. Mar Ecol Prog Ser 236, 75–88
Elbrächter M, Qi YZ (1998) “Aspects of Noctiluca (Dinophyceae) population dynamics”. In: Physiological Ecology of Harmful Algal Blooms. NATO ASI Series, Vol. G 41, Anderson DM, Cembella AD, Hallegraeff MG eds. (Berlin: Springer-Verlag) pp 315–335
Fukuda Y, Endoh H (2006) New details from the complete life cycle of the red-tide dinoflagellate Noctiluca scintillans (Ehenberg) McCartney. European J Protistology 42, 209–219
Galluzzi L, Penna A, Bertozzini E, Vila M, Garces E, Magnani M (2004) Development of a real-time PCR assay for rapid detection and quantification of Alexandrium minutum (a dinoflagellate). Appl Environ Microbiol 70, 1199–1206
Guillard RP, Ryther HJ (1962) Studies of marine planktonic diatoms 1 Cyclotella nana Hustedt and Detonula confervacea (Cleave) Gran. Can J Microbiol 8, 229–239
Hallengraeff GM (1993) A review of harmful algal blooms and their apparent global increase. Phycologia 32, 79–99
Hosoi-Tanabe S, Sako Y (2005) Species-specific detection and quantification of toxic marine dinoflagellates Alexandrium tamarense and A catenella by real-time PCR assay. Mar Biotechnol 7, 506–514
Huang C, Qi Y (1997) The abundance cycle and influence factors on red tide phenomena of Noctiluca scintillans (Dinophyceae) in Dapeng Bay, the South China Sea. J Plankton Res 19, 303–318
Kiørboe T (1998) Intensive aggregate formation with low flux during an upwelling-induced diatom bloom. Limnol Oceanogr 43, 104–116
Miller PE, Scholin CA (1996) Identification of cultured Pseudo-nitzschia (Bacillariophyceae) using species-specific LSU rRNA-targeted fluorescent probes. J Phycol 32, 646–655
Miyaguchi H, Fujiki T, Kikuchi T, Kuwahara VS, Toda T (2006) Relationship between the bloom of Noctiluca scintillans and environmental factors in the coastal waters of Sagami Bay, Japan. J Plank Res 28, 313–324
Murray S, Suthers M (1999) Population ecology Noctiluca scintillans Macartney, a red-tide-forming dinoflagellate. Austral J Mar Fresh Res 50, 243–252
Omori M, Hamner MW (1982) Patchy distribution of zooplankton: behavior, population assessment and sampling problems. Mar Biol 72, 193–200
Onoue Y, Toda T, Ban S (2004) Morphological features and hatching pattern of eggs in Acartia steueri (Crustacea, Copepoda) from Sagami Bay, Japan. Hydrobiologia 511, 17–24
Popels CL, Cary CS, Hutchins AD, Forbes R, Pustizzi F, Gober JC, Coyne JK (2003) The use of quantitative polymerase chain reaction for the detection and enumeration of the harmful alga Aureococcus anophagefferens in environmental samples along the United States East Coast. Limnol Oceanogr Methods 1, 92–102
Sako Y, Hosoi-Tanabe S, Uchida A (2004) Fluorescence in situ hybridization using rRNA-targeted probes for simple and rapid identification of the toxic dinoflagellates Alexandrium tamarense and Alexandrium catenella. J Phycol 40, 598–605
Sato SM, Suzuki M, Hayashi H (1998) The density of homogeneous population of cells controls resetting of the program for swarmer formation in the unicellular marine microorganism Noctiluca scintillans. Exp Cell Res 245, 290–293
Satoh F, Hamasaki K, Toda T, Taguchi S (2000) Summer phytoplankton bloom in Manazuru Harbor, Sagami Bay, central Japan. Plank Biol Ecol 47, 73–79
Smayda JT (1997) What is a bloom? A commentary. Limnol Oceanogr 42, 1132–1136
Sugawara T, Hamasaki K, Toda T, Kikuchi T, Taguchi S (2003) Response of natural phytoplankton assemblages to solar ultraviolet radiation (UV-B) in the coastal water, Japan. Hydrobiologia 493, 17–26
Tada K, Pithakpol S, Montani S (2004) Seasonal variation in the abundance of Noctiluca scintillans in the Seto Inland Sea, Japan. Plank Biol Ecol 51, 7–14
Uhlig G, Sahling G (1990) Long-term studies on Noctiluca scintillans German Bight population dynamics and red tide phenomena 1968–1988. Neth J Sea Res 25, 101–112
Yin K (2003) Influence of monsoons and oceanographic processes on red tides in Hong Kong waters. Mar Ecol Prog Ser 262, 27–41
Zingmark RG (1970) Sexual reproduction in the dinoflagellate Noctiluca miliaris Suriray. J Phycol 6, 122–126
Acknowledgments
We thank Prof. T. Kikuchi and Mr. Y. Asakura at Manazuru Marine Laboratory for Science Education, Yokohama National University, for their support in collecting samples. We also thank Dr. S.V. Kuwahara and Dr. S.C.Y. Leong, Soka University for investing their time and effort in criticizing the manuscript. We also thank Prof. K. Tada, Kagawa University and Prof. K. Furuya, The University of Tokyo, for the supply and instruction on the culture of isolated N. scintillans and Prof. S. Taguchi, Soka University and Dr. K. Hamasaki, Ocean Research Institute, The University of Tokyo for supplying some isolated phytoplankton. We also thank Mr. N. Nagao, Soka University, Ms. K. Watanabe, Soka University, and Dr. S. Hosoi-Tanabe, Kobe University for many helpful comments that improved the manuscript. This research was partially funded by the University Joint Research Project for Private Universities: matching fund subsidy from MEXT (Ministry of Education, Culture Sports, Science and Technology), 2004-2008.
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Miyaguchi, H., Kurosawa, N. & Toda, T. Real-Time Polymerase Chain Reaction Assays for Rapid Detection and Quantification of Noctiluca scintillans Zoospore. Mar Biotechnol 10, 133–140 (2008). https://doi.org/10.1007/s10126-007-9031-3
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DOI: https://doi.org/10.1007/s10126-007-9031-3