دانشگاه اصفهان زیست شناسی میکروبی 3060-7647 12 47 2024 01 21 A Study on the Diversity of Unicellular Algae Genus in Lake Urmia with an Emphasis on Dunaliella Species مطالعه تنوع جنس‌های جلبک‌های تک‌سلولی دریاچه ارومیه با تأکید بر گونه‌های جنس دونالیلا 39 62 27476 10.22108/bjm.2023.135417.1502 FA رامین مناف فر گروه شیلات، دانشکدۀ منابع طبیعی، دانشگاه ارومیه، ارومیه، ایران سکینه مرادخانی عضو هیات علمی دانشگاه پیام نور تهران هدیه یزدانی گروه شیلات، دانشکده منابع طبیعی، دانشگاه ارومیه، ارومیه، ایران. Journal Article 2022 10 13 Introduction: The simplest living things containing chlorophyll are algae, which use photosynthesis to create and release oxygen. The environment with the highest concentration of algae is water. These organisms contain vital chemicals and are rich in different types of proteins and carbohydrates. Their cultivation and purification have been developed as a result. It is feasible to research growing methods, habitats and the nutritional value of algae as well as to purify beneficial species for use in aquaculture by correctly identifying algae. One phylum of green algae is Dunaliella. The cells are biflagellate and motile. Blue-green algae are thought to be the earliest photoautotrophic animals. They are unicellular or filamentous creatures that contain phycocyanin and chlorophyll. Fish breeding ponds frequently use these kinds of creatures. Diatoms are eukaryotic microalgae that play a significant role in the ocean's food chain and can fix carbon dioxide. Diatoms are used by aquatic species as food as well. Additionally, they serve as unique sources of several fuels and oils. Because of their high sensitivity to salinity, acidity and pollution, they are referred to as biological indicators. One of Iran's oligotrophic lakes, Lake Urmia is situated in West Azerbaijan province. The population of Artemia has declined as a result of the rise in salinity of the water and the fall in Dunaliella algae. Thus, one of the main objectives of this study was to examine the variety of unicellular algae present in Lake Urmia Materials and Methods: Lake water was sampled in the final month of each of the four seasons that were conducted. The pH and salinity were measured, simultaneously. Microalgae were tallied using a hemocytometer slide. Water samples from each station were collected independently and they were cultured in 250 ml flasks by Walne culture media for unicellular algae at two salinities of 30 and 100 grams per liter. There was a constant flow of aeration and 100 candelas of fluorescent light. Samples were detected using a light microscope after an algal bloom that lasted for roughly ten days. Two direct approaches were employed to purify the algae: streak culture (using agar culture medium) and inverted microscope (using Johnson's culture medium). Dunaliella species were identified molecularly using the ITS sequence and the 18srDNA gene. The CTAB method was also utilized for DNA extraction. A pair of specific primers was used to amplify the 18srDNA gene, containing the forward sequence 5'-cgg gat ccg tag tca tat gct tgt ctc-3' and the reverse sequence 5'-cgg aat tcc ttc tgc agg ttc acc-3'. Additionally, a pair of forward specific primers was used to amplify the region ITS, consisting of the sequence 5-aat cta tca ata acc aca ccg-3 and the sequence 5-ttt cat tcg cca tta cta agg-3. Amplification of the 18srDNA gene and ITS region requires PCR reagents, which are as follows: 4.2 µl of deionized distilled water, 1 µl buffer, 1.4 µl MgCl2, 0.8 µl dNTP and 0.2 µl primer for each reaction. Next, 0.2 µl of reverse primer, 0.2 µl of Tag DNA polymerase and 2 µl of sample-extracted DNA were taken into consideration and a total volume of 25 µl was used for the PCR process. The temperature cycle that was employed similarly consists of initial annealing at 95°C for one minute, final annealing at 95°C for one minute, primer binding at 52°C for one minute, new strand production at 72°C for two minutes and twenty seconds and final extension at 72°C for four minutes. This cycle was repeated in 33 cycles. The final PCR product was detected by electrophoresis technique. Primer binding took place at 57°C for 50 seconds, first annealing took place at 95°C for 5 minutes, final annealing took place at 94°C for 1 minute and 72°C It was utilized for one minute to create a new strand and for 10 minutes at 72 degrees Celsius to complete the 35 cycles of final strand expansion. Ultimately, a comparison was made between the sequencing results and the sequences found in the GenBank gene database. Results: Comparing freshwater and ocean waters to Lake Urmia, the latter two have more diversity and abundance of unicellular algae due to their oligotrophic nature. Identification of microalgae by morphological and physiological traits is insufficiently precise. Seven species of the Dunaliella genus are identified by a study based on the morphological traits of Dunaliella isolated from eleven distinct areas of Urmia Lake. Morphological analyses revealed that D. salina was the predominant species, with D. acidophilia and D. parva species isolated exclusively in the Shahid Kalantari transit area. D. salina and D. tertiolecta were the species found in the Hydar-Abad region. Due to the region's declining rainfall, retreating sea level and rising salinity, a study of Dunaliella species in the area reveals a low species diversity. The algae species isolated from Golmankhane port are D. bardawil, D. salina and D. tertiolecta. Research revealed that the Agh-Gol region is home to the isolated species D. bardawill, D. salina and D. tertiolecta. Two species of Dunaliella were isolated in the Zanbil mountain area. D. primulecta, D. salina and more species found in the Bari region. In the vicinity of Shahid Kalantari, six species of Dunaliella were isolated: D. salina, D. tertiolecta, D. bardawil, D. parva, D. ruineniana and D. acidophilia. Discussion and Conclusion: The findings indicated that the Shahid Kalantari neighborhood of Dunaliella has the highest level of diversity. Algal diversity was higher in the spring than it was in the summer. The drop in diversity during the summer may have been brought on by rising temperatures, which also increased salinity and exacerbated the drought. The links between several strains of this species of algae have been better understood through the application of molecular techniques. It is conclusively known that at least four species of D. salina, D. tertiolecta, D. viridis and D. bardawil, as well as four new strains, have been identified in Lake Urmia by concurrent investigations of morphological and molecular traits. مقدمه: دریاچه ارومیه یکی از دریاچه‌های بسیار شور دنیا است؛ با این حال، برخی از جلبک‌های تک‌سلولی در این دریاچه یافت می‌شوند که تنوع زیادی دارند و برخلاف شرایط بحرانی حاکم بر دریاچه ارومیه گاهی شکوفایی بسیار شدیدی از این نوع از جلبک‌ها در نقاط مختلف دریاچه مشاهده می‌شود. به‌دلیل افزایش شوری و کاهش شدید میزان آب دریاچه مطالعه‌شده و افزایش درجه شوری تا حد اشباع، پیش‌بینی شد تنوع جلبک‌های تک‌سلولی در این دریاچه به شدت کاهش یافته است. مواد و روش‏‏ها: در این تحقیق سعی شد شناسایی جنس‌های مختلف جلبک‌های تک‌سلولی موجود در دریاچه ارومیه با استفاده از روش‌های ریخت‌شناسی بررسی شود. با توجه به تنوع گونه‌ای جنس دونالیلا در این مطالعه، این کار با استفاده از تکنیک‌های ریخت‌شناسی و مولکولی انجام شد. نتایج: بررسی‌های مولکولی نشان‌دهندة وجود تنوعی از جنس‌های مختلف از 3 رده کلروفیسه، دیاتومه‌ها و سیانوفیسه‌ها بود. در بررسی تنوع گونه‌ای در جنس دونالیلا و مقایسه این نتایج با نمونه‌های موجود در بانک ژنی نشان‌دهندة تغییر تدریجی جمعیت این جلبک‌ها در اثر تغییر شرایط اقلیمی دریاچه ارومیه بود. بحث و نتیجه‏گیری: تنوع جلبک‌های جداشده از نواحی مختلف دریاچه نشان داد با وجود بحران خشکسالی و کاهش شدید تراکم این جلبک‌ها، همچنان چندین جنس و گونه مختلف از جلبک‌های تک‌سلولی در ایستگاه‌های مورد مطالعه دریاچه وجود دارد.

https://bjm.ui.ac.ir/article_27476_9ac676e9e0b1bd366e59c049a82d5300.pdf