Characterization of an Indigenous Isolate, Dunaliella tertiolecta ABRIINW-G3, from Gavkhooni Salt Marsh in Iran Based on Molecular and Some Morpho-physiological Attributes

Authors
N. Hosseinzadeh Gharajeh 1
M. A. Hejazi 1
S. Nazeri 2
A. Barzegari 1
1 Branch of the Northwest and West region, Agricultural Biotechnology Research Institute of Iran (ABRII), Tabriz, Islamic Republic Iran.
2 Department of Biotechnology, Faculty of Agriculture, Bu Ali Sina University, Hamedan, Islamic Republic Iran.
Abstract
Dunaliella is a green halotolerant microalga, which has several industrial applications e.g. β-carotene production. Identification of different Dunaliella species has been carried out by morpho-physiological and recently molecular studies. To achieve an improved understanding of taxonomy, these studies are required to be in linkage. The present study describes molecular and specific morpho-physiological properties of a Dunaliella isolate obtained from Gavkhooni salt marsh in Iran. Phylogenetic analysis of Internal Transcribed Spacer region demonstrated that the isolate was associated with different species except D. salina (CCAP 19/18 and 19/30) and D.viridis. 18S rDNA size of the isolate was identical to that of D. tertiolecta and intron-lacking strains of D. salina. 18S rDNA fingerprint profile and phylogenetic analysis revealed D. tertiolecta as the closest taxon to the isolate. Features of optimum growth salinity (1.5-3% w/v) and maximum carotenoid per cell (0.7 pg cell-1) were comparable with reported data for D. terrtiolecta. Morphological characteristics including the size and color of the cells, presence and location of stigma and refractile granules were similar to those of D. tertiolecta. Totally, considering molecular and morpho-physiological properties, the isolate was attributed to the species D. tertiolecta and was named as Dunaliella tertiolecta ABRIINW-G3.

Keywords

1. Azúa-Bustos, A., González-Silva, C., Salas, L., Palma R. E. and Vicuña R. 2010. A Novel Subaerial Dunaliella Species Growing on Cave Spiderwebs in the Atacama Desert. Extremophiles, 14(5): 443-452.
2. Barzegari, A., Hejazi, M. A., Hosseinzadeh, N., Eslami, S., Mehdizadeh Aghdam, E. Cfor Molecular Farming. Mol. Biol. Rep., 37(7): 3427-3430.
3. Ben-Amotz, A. 1995. New Mode of Dunaliella Biotechnology: Two Phase Growth for β-carotene Production. J. Appl. Phycol., 7(1): 65-68.
4. Ben-Amotz, A. and Avron, M. 1973. The Role of Glycerol in the Osmotic Regulation in the Halophilic Alga, Dunaliella parva. Pl. Physiol., 51(5): 875-878.
5. Ben-Amotz, A. and Avron, M. 1982. Accumulation of β-carotene in Halotolerant Algae: Purification and Characterization of β-carotene-rich Globules from Dunaliella bardawil (Chlorophyceae). J. Phycol., 18(4): 529-537.
6. Ben-Amotz, A. and Avron, M. 1983. On the Factors which Determine Massive ß-carotene Accumulation in the Halotolerant Alga Dunaliella bardawil. Pl. Physiol., 72(3): 593-597.
7. Ben-Amotz, A. and Avron, M. 1987. On the Mechanism of Osmoregulation in Dunaliella. In: "Energetic and Structure of Halophilic Microorganism" (Eds.): Caplan, S. R. and Gizburg M.. Elsevier, Amesterdam, PP. 529-541.
8. Ben-Amotz, A., Shaish, A. and Mordhay, A. 1989. Mode of Action of the Massively Accumulated ß-carotene of Dunaliella bardawil in Protecting the Alga against Damage by Excess Irradiation. Pl. Physiol., 91(3): 1040-1043.
9. Ben-Amotz, A., Polle, J. E. W. and Subba Rao, D. V., Eds. 2009. The Alga Dunaliella: Biodiversity, Physiology, Genomics and Biotechnology. Science Publishers, Enfield, New Hampshire, 555 PP.
10. Borowitzka, L. J., Borowitzka, M. A. and Moulton T. P. 1984. The Mass Culture of Dunaliella salina for Fine Chemicals: From Laboratory to Pilot Plant. Hydrobiologia, 116/117(1): 115-121.
11. Borowitzka, L. J. and Brown, A. D. 1974. The Salt Relations of Marine and Halophilic Species of the Unicellular Green Algae, Dunaliella. Arch. Microbiol., 96(1): 37-52.
12. Borowitzka, M. A. and Siva, C. 2007. The Taxonomy of the Genus Dunaliella (Chlorophyta, Dunaliellales) with Emphasis on the Marine and Halophilic Species. J. Appl. Phycol., 19(5): 567-590.
13. Coleman, A. W. and Mai, J. C. 1997. Ribosomal DNA and ITS-2 Sequence Comparisons as a Tool for Predicting Genetic Relatedness. J. Mol. Evol., 45(2): 168-177.
14. Brown, R. M. and Arnott, H. J. 1970. Structure and Function of the Algal Pyrenoid. I. Ultrastructure and Cytochemistry during Zoosporogenesis of Tetracystis excentrica. J. Phycol., 6(1): 14-22.
15. Duncan, D. B. 1995. Multiple Range and Multiple F Tests. Biometrics, 11(1): 1-5.
16. Fazeli, M. R., Tofighi, H., Samadi, N. and Jamalifar, H. 2006. Effect of Salinity on β-Carotene Production by Dunaliella salina DCCBC26 Isolated from the Urmia Salt Lake, North of Iran. Biores. Tech., 97(18): 2453-2456.
17. Gómez, P. and Gonzaléz, M. 2001. Genetic Polymorphism in Eight Chilean Strains of the Carotenogenic Microalga Dunaliella salina Teodoresco (Chlorophyta). Biol. Res., 34(1): 23-30.
18. Gómez, P. I., Barriga, A., Cifuentes, A. S. and González, M. A. 2003. Effect of Salinity on the Quantity and Quality of Carotenoids Accumulated by Dunaliella salina (strain CONC-007) and Dunaliella bardawil (Strain ATCC 30861) Chlorophyta. Biol. Res., 36(2): 185-192.
19. Gómez, P. I. and Gonzaléz, M. A. 2004. Genetic Variation among Seven Strains of Dunaliella salina (Chlorophyta) with Industrial Potential, Based on RAPD Banding Patterns and on Nuclear ITS rDNA Sequences. Aquaculture, 233(1-4): 149-162.
20. Gómez-Pinchetti, J. L., Ramazanov, Z. M., Fontes, A. G. and García Reina, G. 1992. Photosynthetic Characteristics of Dunaliella salina (Chlorophyceae, Dunaliella) in Relation to ß-carotene Content. J. Appl. Phycol. 4(1): 11-15.
21. González, F., Coleman, A. W., Gómez, P. I. and Montoya, R. 2001. Phylogenetic Relationship among Various Strains of Dunaliella (Chlorophyceae) Based on Nuclear ITS rDNA Sequences. J. Phycol., 37(4): 604-611.
22. González, M. A., Gómez, P. I. and Montoya, R. 1999. Comparison of PCR-RLFP Analysis of the ITS Region with Morphological Criteria of Various Strains of Dunaliella. J. Appl. Phycol., 10(6): 573-580.
23. Hadi, M. R., Shariati, M. and Afsharzadeh, S. 2008. Microalgal Biotechnology: Carotenoid and Glycerol Production by the Green Algae Dunaliella Isolated from the Gave-Khooni Salt Marsh, Iran. Biotechnol. Bioprocess Eng., 13(5): 540-544.
24. Hejazi, M. A., Barzegari, A., Hosseinzadeh Gharajeh, N. and Hejazi, M. S. 2010. Introduction of a Novel 18S rDNA Gene arrangement along with Distinct ITS Region in the Saline Water Microalga Dunaliella. Saline Sys., 6(1): 4-14.
25. Hejazi, M. A., de Lamarliere, C., Rocha, J. M. S., Vermuë, M., Tramper, J. and Wijffels, R. H. 2002. Selective Extraction of Carotenoids from the Alga Dunaliella salina with Retention of the Viability. Biotech. Bioeng., 79(1): 29-36.
26. Hejazi, M. A. and Wijffels, R. H. 2003. Effect of Light Intensity on ß-carotene Production and Extraction by Dunaliella salina in Two-phase Bioreactors. Biomol. Eng., 20(4-6): 171-175.
27. Jahnke, L. S. and White, L. A. 2003. Long-term Hyposaline and Hypersaline Stresses Produce Distinct Antioxidant Responses in the Marine Alga Dunaliella tertiolecta. J. Plant Physiol., 160(10): 1193-1202.
28. Librado, P. and Rozas, J. 2009. DnaSP v5: A Software for Comprehensive Analysis of DNA Polymorphism Data. Bioinformatics, 25(11): 1451-1452.
29. Olmos, J., Ochoa, L., Paniagua-Michel, J. and Contreras, R. 2009. DNA Fingerprinting Differentiation between ß-carotene Hyperproducer Strains of Dunaliella from around the World. Saline Sys.,5(1): 5-14.
30. Olmos, J., Paniagua, J. and Contreras, R. 2000. Molecular Identification of Dunaliella sp. Utilizing the 18S rDNA Gene. Lett. Appl. Microbiol., 30(1): 80-84.
31. Olmos-Soto, J., Paniagua-Michel, J., Contreras, P. R. and Trujillo, L. 2002. Molecular Identification of ß-carotene Hyper-producing Strains of Dunaliella from Saline Environments Using Species-specific Oligonucleotides. Biotech. Lett., 24(5): 365-369.
32. Raja, R., Iswarya, S. H., Balasubramanyam, D. and Rengasamy, R. 2007. PCR-identification of Dunaliella salina (Volvocales, Chlorophyta) and Its Growth Characteristics. Microbiol. Res., 162(2): 168-176.
33. Sciandra, A., Gostan, J., Collos, Y., Descolas-Gros, C., Le Boulanger, C., Martin-Jézéquel, V., Denis, M., Lefèvre, D., Copin-Montégut, G. and Avril, B. 1997. Growth-compensating Phenomena in Continuous Cultures of Dunaliella tertiolecta Limited Simultaneously by Light and Nitrate. Limnol. Oceanogr., 42(6): 1325-1339.
34. Tamura, K., Dudley, J., Nei, M. and Kumar, S. 2007. MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24(8): 1596-1599.
35. Zamani, H., Moradshahi, A. and Karbalaei-Heidari H. R. 2011. Characterization of a New Dunaliella salina Strain MSI-1 Based on Nuclear rDNA ITS Sequences and Its Physiological Response to Changes in Composition of Growth Media. Hydrobiol., 658(1): 67-75.
Journal of Agricultural Science and Technology
Volume 14, Issue 7
November and December 2012
Pages 1579-1590