Successful Artificial Insemination in the Koala (Phascolarctos cinereus) Using Extended and Extended-Chilled Semen Collected by Electroejaculation1

Abstract

Artificial insemination in the koala using chilled, electroejaculated semen provides for a marked improvement in the reproductive and genetic management of captive koala colonies in Australia and internationally, and makes available the option of using semen collected from wild populations to expand restricted gene pools. Dilution of koala semen for artificial insemination is complicated because koalas are induced ovulators, and it is thought that ovulating factors are present in the semen, so that semen extension for preservation purposes might be anticipated to result in a failure to induce ovulation. The first two experiments of this study were designed to determine whether artificial insemination using undiluted, extended, and extended-chilled semen collected by electroejaculation was capable of inducing a luteal phase and/or the production of pouch young. In Experiment 1, 1 ml undiluted electroejaculated semen, 2 ml diluted (1:1) semen, and 1 ml diluted (1:1) semen resulted in seven of nine, six of nine, and six of nine koalas showing a luteal phase, respectively; four pouch young were produced in each treatment. A second artificial insemination experiment was conducted in which 2 ml diluted (1:1) semen was administered in three groups of nine koalas. The first group received semen that had been collected and diluted immediately without chilling, the second group was deposited with semen stored chilled for 24 h, and the final group received semen that had been chilled for 72 h. In the first group, five females had a luteal phase, but none became pregnant. In group 2, two of the five females that had a luteal phase gave birth, whereas in group 3, four of the six females that had a luteal phase produced pouch young. In addition, experiment 3 was conducted to determine whether it was possible to produce pouch young by naturally mating koalas that were in the latter stages of their behavioral estrus; this information is important to the logistics of transporting koala semen for artificial insemination by establishing the maximum time frame in which females might be expected to shed a fertile oocyte. Of the 12 females mated on Day 8 of estrus, 6 gave birth, whereas only 3 of the 10 females naturally mated on Day 10 of estrus produced pouch young. The majority of females (21 of 22) in experiment 3 showed evidence of a luteal phase. Together, these experiments have shown that it is possible to use undiluted, extended, or extended-chilled semen to produce koala offspring up to Day 8 of estrus at conception rates similar to those achieved following natural mating. These findings represent a significant advancement in the use of reproductive technology in marsupials and provide the basis for the shipment of koala semen over long distances. The pouch young produced in this study represent the first marsupials born following artificial insemination of extended-chilled semen and bring the total number of koalas produced by artificial insemination to 31.

Artificial insemination in marsupials

“…As discussed earlier, a contributing factor to the success of AI in the koala is that ovulation can be induced relatively simply [3,4,18]. By contrast, although ovulation cannot be induced by such manipulation in the tammar wallaby, birth and postpartum estrus can be tightly synchronized by removal of pouch young and ovulation is reasonably predictable, thus facilitating successful AI [6].…”

“…Koala sperm stored for 3 d at 4 8C has also been used for successful AI [3]. McCallum and Johnston [37] have also reported the successful storage of common wombat spermatozoa in the cauda epididymis.…”

“…Marsupial semen collection, handling and basic properties were the subject of a paper [1] at the 1977 Symposium of the Zoological Society of London, which led to Paul Watson's landmark book 'Artificial Breeding of Non-Domestic Animals' [2]. However, despite the three decades that have passed AI, and ART generally, have been demonstrated to have the potential to make a significant impact on conservation www.theriojournal.com Available online at www.sciencedirect.com Theriogenology 71 (2009) [176][177][178][179][180][181][182][183][184][185][186][187][188][189] and reproductive management of only one marsupial species, the koala (Phascolarctos cinereus; [3,4]). In the koala, 32 pouch young have been produced and birth rates following AI using extended and chilled semen are only slightly below those achieved by natural mating [3].…”

“…However, despite the three decades that have passed AI, and ART generally, have been demonstrated to have the potential to make a significant impact on conservation www.theriojournal.com Available online at www.sciencedirect.com Theriogenology 71 (2009) [176][177][178][179][180][181][182][183][184][185][186][187][188][189] and reproductive management of only one marsupial species, the koala (Phascolarctos cinereus; [3,4]). In the koala, 32 pouch young have been produced and birth rates following AI using extended and chilled semen are only slightly below those achieved by natural mating [3]. Furthermore, AI technology has now been incorporated in state government koala management policy [5].…”

“…Arguably the koala and tammar wallaby are special cases, because the timing of ovulation can be predicted with considerably more confidence than in any other marsupial. The koala is an induced ovulator and techniques to artificially induce ovulation have been successfully developed [3,4,14,[16][17][18] and the tammar has a highly predictable post-partum estrus, which can be tightly synchronized by removing pouch young, and a well-defined window of ovulation [19][20][21]. In a limited number of other marsupials, such precise timing of ovulation has only been achieved using hormonal treatment, which is likely to have undesirable effects.…”