Assessment of the macroalgal diversity of Kuwait by using the Germling Emergence Method

Cryptic stages of diverse macroalgae present in natural substrata, “the bank of microscopic forms”, were isolated into clonal cultures and identified based on both morphological characteristics and DNA barcoding. Approximately 120 clonal isolates from 308 natural substratum samples were collected from the entire coastline of Kuwait. Amongst these isolates, 77 (64%) were identified through DNA barcoding using the nuclear ribosomal small subunit, RuBisCO spacer (ITS2, tufa, rbcL, psaA, and psbA) and sequencing. Twenty-six isolates (34%) were identified in the division Chlorophyta, 18 (23%) as Phaeophyceae, and 33 (43%) as Rhodophyta. For all DNA sequences in this study, species-level cut off applied was ≥98% homology which depend entirely on the markers used. Three putative new records of Chlorophyta new for the Arabian Gulf were made: Cladophora laetevirens (Dillwyn) Kützing, Ulva torta (Mertens) Trevisan and Ulvella leptochaete (Huber) R. Nielsen, C. J. O′Kelly & B. Wysor in Nielsen, while Cladophora gracilis Kützing and Ulva ohnoi M. Hiraoka & S. Shimada are new records for Kuwait. For Phaeophyceae, Ectocarpus subulatus Kützing and Elachista stellaris Areschoug were new records for the Gulf and Kuwait. In the Rhodophyta, Acrochaetium secundatum (Lyngbye) Nägeli in Nägeli & Cramer, Ceramium affine Setchell & N. L. Gardner, Gelidium pusillum var. pakistanicum Afaq-Husain & Shameel and Dasya caraibica Børgesen are new records for the Gulf and Kuwait, while the red alga Stylonema alsidii (Zanardini) K. Drew is a new record for Kuwait. Several isolates identified corresponded to genera not previously reported in Kuwait and / or the Arabian Gulf, such as Porphyrostromium Trevisan, a new genus from the Bangiales, and two unidentified species for the Planophilaceae Škaloud & Leliaert. The isolates cultivated from substrata enhance understanding of the marine macroalgal diversity in the region and confirmed that the Germling Emergence Method is suitable for determining the actual diversity of a given study area through isolation from cryptic life-history phases.