Application of a functional marker for the effect of cryoprotectant agents on gorgonian coral (Junceela juncea and J. fragilis) sperm sacs

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ABSTRACT

BACKGROUND:

The establishment of coral sperm repositories which retain good post-rewarming viability and fertility play a vital role in species conservation.

OBJECTIVE:

This study aimed at obtaining baseline information regarding the effects of cryoprotectant agents (CPAs) on gorgonian coral (Junceella juncea and J. fragilis) sperm sacs.

METHODS:

The adenosine triphosphate assay was used to determine the energy level of the gorgonian sperm sacs as an indicator of sperm viability after exposure to cryoprotectants.

RESULTS:

The ‘no observed effect concentrations’ (NOECs) of methanol, dimethyl sulfoxide (DMSO), polypropylene glycol (PG), ethylene glycol (EG) and glycerol for J. juncea sperm sacs were 3 M, 3 M, 1 M, 2 M and 1 M respectively after 20 min exposure; whilst the NOECs for J. fragilisoocytes were 2 M, 3 M, 1 M, 2 M and 2 M, respectively. Methanol and DMSO had the least impact. PG was the most toxic CPA after 10 min exposure. ATP content of J. juncea and J. fragilis sperm sacs did not differ significantly from the control with incubation times of 10-20 min with 2 M EG. However, ATP content dropped significantly after exposing sperm sacs to 2 M EG for 40 min with average values of 2.34 ± 0.12 and 1.97 ± 0.48 μg/ml respectively. ATP content for J. juncea and J. fragilis sperm sacs was significantly decreased to 1.79 ± 0.31 and 2.40 ± 0.36 μg/ml after 20 min incubation in 2 M PG when compared to the control with 2.98 ± 0.16 and 4.14 ± 0.42 μg/ml respectively. Normalized ATP content for sperm sacs of two different gorgonian coral after incubation in methanol, DMSO, PG, EG and glycerol showed that J. juncea sperm sacs were slightly less tolerant to CPAs compared to J. fragilis sperm sacs.

CONCLUSIONS:

DMSO or methanol can be considered as efficient CPAs for gorgonian sperm sacs cryopreservation. The ATP luminescence assay provided sensitive and rapid quantification of mitochondrial activity in gorgonian coral sperm sacs. The study on the impact of CPA will contribute to the development of a cryopreservation protocol for coral sperm conservation.