Crocodilian perivitelline membrane‐bound sperm detection

Abstract

Advanced reproductive technologies (ART's) are often employed with various taxa to enhance captive breeding programs and maintain genetic diversity. Perivitelline membrane-bound (PVM-bound) sperm detection has previously been demonstrated in avian and chelonian species as a useful technique for breeding management. In the absence of embryotic development within an egg, this technique can detect the presence of sperm trapped on the oocyte membrane confirming breeding, male reproductive status, and pair compatibility. PVM-bound sperm were successfully detected in three clutches of Cuban crocodile (Crocodylus rhombifer) eggs at the Smithsonian's National Zoological Park (NZP) for the first time in any crocodilian species. PVM-bound sperm were detected in fresh and incubated C. rhombifer eggs, as well as eggs that were developing (banded) and those that were not (not banded). The results of this study showed significant differences in average sperm densities per egg between clutches (p = 0.001). Additionally, there was not a significant difference within clutches between eggs that banded and those that did not band (Clutch A, p = 0.505; Clutch B, p = 0.665; Clutch C, p = 0.266). The results of this study demonstrate the necessity to microscopically examine eggs that do not develop (do not band), to determine if sperm is present, which can help animal managers problem solve reproductive shortcomings. PVM-bound sperm detection could be a useful technique in assessing crocodilian breeding programs, as well as have potential uses in studies assessing sperm storage, artificial insemination, and artificial incubation.

Understanding early reproductive failure in turtles and tortoises

“…relocations), and other potential drivers of reproductive failure such as pollutants and disease exposure, to monitor the impact of environmental change on early reproductive processes. We also anticipate that this approach will be applicable to other reptile groups, such as crocodiles (Augustine 2017), allowing scope to investigate similarities across birds and reptiles. The methods outlined here may therefore equip researchers and conservationists working across broad taxonomic groups with a tool to inform conservation and breeding management in the face of global change.…”

“…In published studies to date, undeveloped eggs are typically classified as unfertilised without further examination (e.g., Langer et al, 2020;Gane et al, 2020a), and where attempts are made to determine fertilisation success, potentially inaccurate macroscopic methods are typically used (Gárriz et al, 2020;Phillott and Godfrey, 2020). Recently, a small number of reptilian captive breeding programmes have trialled microscopic techniques, originally developed for birds (Birkhead et al, 2008;Hemmings et al, 2012), to help investigate infertility (Croyle et al, 2015;Croyle et al, 2016;Augustine, 2017), and the application of these methods has also been recommended for assessing fertility of wild sea turtle eggs (Phillott and Godfrey, 2020;Phillott, et al, 2021). The techniques allow detection of sperm on the perivitelline membrane (PVM) surrounding the yolk and embryonic nuclei in the germinal disc, thereby providing unequivocal evidence of fertilisation.…”

Modification and Clinical Application of the Inner Perivitelline Membrane Test in Different Avian Species

“…This technique avoids the problem of separating different parts of the perivitelline membrane, as no separation is necessary. Results in various species, including birds [9,23,24,25], chelonians [26], and crocodiles [27], prove this test to be highly useful.…”