Fig 1.
A male tuatara (Sphenodon punctatus) stands outside his burrow.
Tuatara are sexually dimorphic at maturity, with males being on average larger, having a wider jaw, more triangular head, and a crest with larger, more closely arranged spikes.
Fig 2.
Tuatara (Sphenodon punctatus) mating.
Tuatara mating on the porch of a Department of Conservation ranger house on Takapourewa. The male is postured on top of the female, with his hind limbs and tail twisting to oppose their cloaca. Tuatara are unique among reptiles for their lack of intromittent organ.
Fig 3.
Sperm visible on a female’s cloaca after mating and collection.
(A) Sperm is visible on the female’s cloaca after separating a mating pair. (B) Sperm being collected from the female’s cloaca with a microcapillary tube.
Table 1.
Tuatara (Sphenodon punctatus) sperm sample details.
Fig 4.
Tuatara (Sphenodon punctatus) sperm viability images.
(A + C) Sphenodon punctatus sperm stained with Eosin Nigrosin to assess viability (membrane integrity). Sperm whose heads are penetrated with stain are non-viable (A + C), while sperm heads that reject stain, and thus appear white, are considered viable (B + D). Various artefacts from the cloaca, in addition to crystals formed by the stain cracking as it dried, are also visible.
Fig 5.
Tuatara (Sphenodon punctatus) sperm visualized using SEM.
Photographs (A–C) of whole Sphenodon punctatus sperm captured using scanning electron microscopy. Sperm had been previously treated with Lake’s solution and flash frozen, before being thawed and photographed. Note the scale difference on the first image. T = tail end.
Fig 6.
Tuatara sperm tail tips and midpiece damage (Sphenodon punctatus), visualized using SEM.
Photographs of Sphenodon punctatus sperm treated with Lake’s solution and viewed using scanning electron microscopy, showing (A + B) reduced tail tips and (C + D) different types of midpiece damage, likely due to ineffective cryoprotectant during freezing and thawing.
Table 2.
Viability analysis results for tuatara (Sphenodon punctatus) sperm.
Fig 7.
Velocity analyses of tuatara (Sphenodon punctatus) sperm.
(A) Intraclass correlation coefficients (ICC) and between and within individual variance in Sphenodon punctatus sperm speed (curvilinear velocity, μm × s−1) of different groups of fastest sperm cells. The lowest within male variance (0.27), highest between male variance (0.76), and highest ICC (0.74) were all achieved by grouping only the five fastest sperm cells from each sample. (B) Results of movement analysis (curvilinear velocity, VCL) for the five fastest sperm cells per S. punctatus sample, grouped by individual and treatment. All values presented are in μm × s−1 and bars represent 95% confidence intervals. All samples without the added effect of time are shown. (C) S. punctatus VCL values (μm × s−1) for all paired samples showing the effect of time on sample velocity. Error bars represent 95% confidence intervals.