Long-term preservation of eri and ailanthus silkworms using frozen gonads

Abstract

Cryopreservation of eri and ailanthus silkworms using frozen gonads was investigated. First, we evaluated the freeze tolerance of ovary and testis in the eri silkworm, which showed high tolerance. Mating between frozen ovary-transplanted females and frozen testis-transplanted males produced 163.0 eggs, yielding 105.7 larvae per moth. In a second experiment, we tested the use of the eri silkworm as a host insect for gonad transplantation from ailanthus silkworm donors. A high success ratio for laid and hatched eggs was demonstrated for ovary transplantation (97.8 and 51.3 eggs per moth, respectively). For testis transplantation, however, the average number of hatched larvae was low (12.0). Mating between host eri females and males in which both frozen ovary and testis of the ailanthus silkworm had been transplanted produced 6.4 fertilized eggs per host moth. Our success in using cross subspecies cryopreservation between these wild silkworms could lead to the alternative use of hosts between species in other insects.

Introduction

Long-term and secure preservation are required for bio-resources. In mammals, such as cows, mice, and humans, cryopreservation methods have been developed and are used as routine tools [2], [4]. On the other hand, the development of similar methods was rather late in insects. Recently, we succeeded in establishing a technique for using frozen ovaries in Bombyx mori [1]. However, this method has not yet been evaluated in any other insect nor has it been applied to testis. In the present study, we first investigated the possibility of applying our established technique to the eri silkworm. The eri silkworm (Samia cynthia ricini), which is primarily cultivated in India, Thailand, and South China, is one of the main domesticated silkworms used for producing fiber material. This insect is easy to handle because of its long-term selective breeding; thus, it is also used as an experimental animal. The eri silkworm is, however, multivoltine; therefore, continuous rearing and reproduction are needed for its preservation. For these reasons, we selected the eri silkworm for our first experiments. We also evaluated the suitability of frozen testes, in addition to ovaries, in this species.

Secondly, we investigated the possibility of using the eri silkworm as a host insect for transplantation of gonads from ailanthus silkworm (S. cynthia pryeri) donors. This species, which inhabits a broad territory from South to East Asia, is considered an ancestor of the eri silkworm. Further, taxonomically, they are considered a subspecies. Although the two insects have a very close taxonomic relationship, the ailanthus silkworm has not been domesticated and thus, is difficult to rear in captivity or under controlled conditions. Hence, it is problematic to apply a cryopreservation method to the ailanthus silkworm. If the eri silkworm could be used as a host for the ailanthus silkworm, cryopreservation of the latter might be possible.