Physiological diversity among sympatric, conspecific endosymbionts of coral (Cladocopium C1acro) from the Great Barrier Reef

Strains of the common coral endosymbiont, Cladocopium C1acro were isolated from four Acropora corals collected from inshore sites on the Great Barrier Reef (Nelly Bay, Magnetic Island, and South Molle Island in the Whitsunday Islands). Colonies of Accropora Tenuis and Acropora millepora were growing at similar depths and photic habitats, between `~3-4m.

Before the culturing of the symbionts, freshly isolated cells from the host were genotyped. Clonal cultures were established using methods described in Beltrán, et.al. (2021).

For the generation of clonal cultures, two methods were used. Three of the four clonal cultures (Aten-MI-1, Aten-WSY, and Amil-MI) were obtained after plating the heterogeneous population onto IMK − 1% agar plus antibiotics and incubating them under the conditions as above and picking a single colony-forming unit. The fourth clonal culture (Aten-MI-2) was produced from a single cell isolated using a micro-manipulator (MN-153, Narishige) under an inverted microscope at 20X magnification (Leica DM IRB/E, Germany).

DNA was extracted from exponentially growing culture and sequence data from the chloroplast marker were deposited at the genbank with the accession numbers MW691103-06 for Aten-MI-2, Aten-MI-1, Amil-MI and Aten-WSY, respectively.

Genetic characterisation of Symbiodiniaceae cultures and growth kinetics were compared under different thermal scenarios. After ~ 30 days incubation, four replicates of each strain were selleted and divided over 16 tubes. For full details, see Beltran et al., (2021).

常见珊瑚内共生菌Cladocopium C1acro（Cladocopium C1acro）菌株，从采集自大堡礁近岸海域（惠森迪群岛的内利湾、磁岛及南莫尔岛）的4株轴孔珊瑚属（Acropora）珊瑚中分离得到。供试的细枝轴孔珊瑚（Acropora Tenuis）与米氏轴孔珊瑚（Acropora millepora）均生长于水深约3~4米的相似深度与光照生境中。
在培养共生菌之前，先对从宿主中新鲜分离的细胞进行了基因分型。克隆培养体系的建立参照Beltrán等人2021年发表的实验方法。
本次克隆培养的构建采用两种方法：其中3株克隆培养物（Aten-MI-1、Aten-WSY及Amil-MI）通过将异质性菌群接种于添加抗生素的IMK-1%琼脂培养基上，按照前述培养条件进行孵育后，挑取单菌落形成单位获得。第4株克隆培养物（Aten-MI-2）则通过显微操作器（MN-153，Narishige），在20倍放大倍率的倒置显微镜（Leica DM IRB/E，德国）下分离单个细胞得到。
从指数生长期的培养物中提取DNA，叶绿体标记基因的测序数据已提交至基因数据库（GenBank），对应登录号依次为：Aten-MI-2为MW691103、Aten-MI-1为MW691104、Amil-MI为MW691105、Aten-WSY为MW691106。
对虫黄藻科（Symbiodiniaceae）培养物的遗传特征及生长动力学开展了不同热胁迫场景下的比较分析。经过约30天的孵育后，将每个菌株的4个生物学重复样本选取后分装至16支试管中。完整实验细节详见Beltran等人2021年的研究。