Evaluation of euthanasia methods using injectable agents in leopard geckos (Eublepharis macularius)

Reptile euthanasia techniques are poorly described. This non-blinded randomized study compared pentobarbital, lidocaine, and potassium chloride (KCl) for leopard geckos (Eublepharis macularius) euthanasia at 20° Celsius and evaluated for post-mortem artifacts. Fifty-three non-clinical adult leopard gecko were scheduled for euthanasia due to Cryptosporidium varanii exposure. Based on pilot study results, geckos randomly received one of six treatments: intracoelomic (ICo) pentobarbital 400mg/kg (ICo-P4, n = 9) or 800mg/kg (ICo-P8, n = 9), intramuscular (IM) alfaxalone 20mg/kg followed by intracardiac (ICa) pentobarbital 800mg/kg (ICa-P, n = 9), KCl 10mEq/kg (ICa-K, n = 9), or lidocaine 100mg/kg (ICa-L, n = 9) or IM alfaxalone 20mg/kg (IM-A, n = 8, control). Serial Doppler heart rate assessments were performed until cardiac arrest, 30 minutes (ICa groups), or 60 minutes (ICo groups). Cardiac arrest occurred in 7/9, 9/9, 9/9, 8/9, and 8/9 geckos in ICo-P4, ICo-P8, ICa-P, ICa-K, and ICa-L, respectively, with median (range) times of 35 (25 - 45), 30 (15-60), 0 (0 - 6.33), 0 (0 - 0.05), and 0 (0 - 0.03) minutes. Side effects were noted in a subset of ICo-P (arched posture) and ICa-K (muscle spasms) geckos. Six geckos in ICa-L had Doppler sound resumption several hours following cessation, with three displaying spontaneous movement. All geckos in IM-A recovered. Intracardiac pentobarbital following IM alfaxalone caused rapid, permanent loss of heartbeat in all tested geckos. Intracoelomic pentobarbital was also effective but had a prolonged time to cardiac arrest. Intracardiac lidocaine is not recommended for leopard gecko euthanasia.

Goniurosaurus chengzheng sp. nov., a new species of Leopard Gecko from Guangxi, China (Squamata: Eublepharidae)

Six species of geckos in the genus Goniurosaurus have been recorded from Guangxi, China. Here we describe a new species, Goniurosaurus chengzheng sp. nov. The new species is similar to allied species from Guangxi, but unique in a combination of the following characters: (1) four body bands with three between limb insertions; (2) precloacal pores 20; (3) body color reddish- brown; (4) snout to eye distance: eye to ear distance < 1. We used the mitochondrial genes 16S and cytb to confirm the distinctiveness of the species and place it within a molecular phylogeny of Goniurosaurus. The type specimens are deposited in the Museum of Biology, East China Normal University (ECNU).

Genetics of white color and iridophoroma in “Lemon Frost” leopard geckos

The squamates (lizards and snakes) are close relatives of birds and mammals, with more than 10,000 described species that display extensive variation in a number of important biological traits, including coloration, venom production, and regeneration. Due to a lack of genomic tools, few genetic studies in squamates have been carried out. The leopard gecko, Eublepharis macularius, is a popular companion animal, and displays a variety of coloration patterns. We took advantage of a large breeding colony and used linkage analysis, synteny, and homozygosity mapping to investigate a spontaneous semi-dominant mutation, “Lemon Frost”, that produces white coloration and causes skin tumors (iridophoroma). We localized the mutation to a single locus which contains a strong candidate gene, SPINT1, a tumor suppressor implicated in human skin cutaneous melanoma (SKCM) and over-proliferation of epithelial cells in mice and zebrafish. Our work establishes the leopard gecko as a tractable genetic system and suggests that a tumor suppressor in melanocytes in humans can also suppress tumor development in iridophores in lizards.

Tooth Removal in the Leopard Gecko and the de novo Formation of Replacement Teeth

Many reptiles are able to continuously replace their teeth through life, an ability attributed to the existence of epithelial stem cells. Tooth replacement occurs in a spatially and temporally regulated manner, suggesting the involvement of diffusible factors, potentially over long distances. Here, we locally disrupted tooth replacement in the leopard gecko (Eublepharis macularius) and followed the recovery of the dentition. We looked at the effects on local patterning and functionally tested whether putative epithelial stem cells can give rise to multiple cell types in the enamel organs of new teeth. Second generation teeth with enamel and dentine were removed from adult geckos. The dental lamina was either left intact or disrupted in order to interfere with local patterning cues. The dentition began to reform by 1 month and was nearly recovered by 2–3 months as shown in μCT scans and eruption of teeth labeled with fluorescent markers. Microscopic analysis showed that the dental lamina was fully healed by 1 month. The deepest parts of the dental lamina retained odontogenic identity as shown by PITX2 staining. A pulse-chase was carried out to label cells that were stimulated to enter the cell cycle and then would carry BrdU forward into subsequent tooth generations. Initially we labeled 70–78% of PCNA cells with BrdU. After a 1-month chase, the percentage of BrdU + PCNA labeled cells in the dental lamina had dropped to 10%, consistent with the dilution of the label. There was also a population of single, BrdU-labeled cells present up to 2 months post surgery. These BrdU-labeled cells were almost entirely located in the dental lamina and were the likely progenitor/stem cells because they had not entered the cell cycle. In contrast fragmented BrdU was seen in the PCNA-positive, proliferating enamel organs. Homeostasis and recovery of the gecko dentition was therefore mediated by a stable population of epithelial stem cells in the dental lamina.