In vitro development of gametophytes and sporophytes of critically endangered fern Christella kendujharensis S.K. Behera & S.K. Barik (Thelypteridaceae)

Christella kendujharensis S.K. Behera & S.K. Barik (family: Thelypteridaceae) is a critically endangered fern endemic to India. We developed its mass propagation protocol through in vitro spore culture in a liquid suspended medium for germination of gametophytes that produced sporophytes subsequently. The spores were monolete, reniform (mean length 36.49 ± 3.6 × breadth 25.67 ± 4.2 μm) to spheroidal with fimbriate and broad-winged perispore. The maximum germination was in the Parker's and Thompson's (P&TV) medium with 0.1 mg L⁻¹ thiamin, 0.1 mg L⁻¹ riboflavin, 0.1 mg L⁻¹ biotin, 0.1 mg L⁻¹ pyridoxin, 0.5 mg L⁻¹ nicotinic acid, 100 mg L⁻¹ myoinositol, 0.01 mg L⁻¹ folic acid, and 3 mg L⁻¹ glycine. Spore germination was Vittaria type and gametophyte development was Aspidium type. The cordate gametophyte developed archegonia on the 38th day and antheridia on the 42nd day of spore sowing. Both isolate and composite cultures developed sporophytes indicating the presence of intra-gametophytic selfing. Based on the maximum number of regenerated gametophytes, two combinations, that is, (i) 2 mg L⁻¹ kinetin, and 0.01 mg L⁻¹ 2,4-dichlorophenoxyacetic acid, and (ii) 0.1 mg L⁻¹ naphthaleneacetic acid and 2 mg L⁻¹ kinetin in P&TV media were recommended for mass propagation of C. kendujharensis.