Xiang Ji

Does body-volume constrain reproductive output in lizards?

The numbers and sizes of eggs produced by adult females ultimately determine the viability of populations, as well as the evolutionary fitness of the females themselves. Despite an enormous amount of literature on the adaptive significance of fecundity variation within and among populations, simpler questions—such as the proximate mechanisms by which a female determines her clutch size—have attracted less attention. Our surgical manipulations show that the amount of space available to hold eggs within a females abdomen influences her total reproductive allocation, enabling her to flexibly modify her reproductive output as she grows larger.

Evolution of viviparity in warm‐climate lizards: an experimental test of the maternal manipulation hypothesis

The maternal manipulation hypothesis for the evolution of reptilian viviparity has been claimed to apply to any situation where gravid females are able to maintain body temperatures different from those available in external nests, but empirical data that support this hypothesis are very limited. Here, we tested this hypothesis using gravid females of a warm‐climate lizard, Mabuya multifasciata, by subjecting them to five thermal regimes for the whole gestation period. We found gravid females selected lower body temperatures and thermoregulated more precisely than did nongravid females. Offspring produced in different treatments differed in head size, limb length and sprint speed, but not in overall body size or mass. Variation in morphological traits of offspring was induced primarily by extreme temperatures. Sprint speed of offspring was more likely affected by the mean but not by the variance of gestation temperatures. Gravid females maintained more stable body temperatures than did nongravid females not because these temperatures resulted in the optimization of offspring phenotypes but because the range of temperatures optimal for embryonic development was relatively narrow. Our data conform to the main predictions from the maternal manipulation hypothesis that females should adjust thermoregulation during pregnancy to provide optimal thermal conditions for developing embryos and that phenotypic traits forged by maternal thermoregulation should enhance offspring fitness.

Proteomic and biochemical analyses of short-tailed pit viper (Gloydius brevicaudus) venom: Age-related variation and composition–activity correlation

UNLABELLEDnWe conducted an in-depth analysis of the proteomic and biochemical profiles of the venom of neonate and adult short-tailed pit vipers (Gloydius brevicaudus). Identified proteins were assigned to a few main toxin families. Disintegrin, phospholipase A2 (PLA2), serine proteinase, cysteine-rich secretory protein, C-type lectin-like protein, l-amino acid oxidase and snake venom metalloproteinase (SVMP) were detected in both venoms, while 5-nucleotidase was detected only in the adult venom. SVMP was the predominant protein family in both venoms (neonate: 65.7%; adult: 64.4%), followed by PLA2 (neonate: 13.4%; adult: 25.0%). Antivenomic analysis revealed that commercial G. brevicaudus antivenom almost neutralized the chromatographic peaks with medium and high molecular masses in both venoms, but did not completely recognize peaks with low molecular mass. Toxicological and enzymatic activities show remarkable age-related variation in G. brevicaudus venom, probably resulting from variation in venom composition. Our data demonstrate age-related variation across venomics, antivenomics and biochemical profiles of G. brevicaudus venom, and have implications for the management of G. brevicaudus bites, including improving antivenom preparation by combining both venoms.nnnBIOLOGICAL SIGNIFICANCEnThis study investigates the composition and biochemical activity of neonate and adult Gloydius brevicaudus venoms. We found remarkable age-related variation in venom biological activity, likely the result of variation in venom composition. Antivenomics analysis was used to explore difference in neonate and adult G. brevicaudus venoms. Our findings have implications for the diagnosis and clinical management of G. brevicaudus bites, and the design of venom mixtures that will increase the efficacy of commercial antivenom. This article is part of a Special Issue entitled: Proteomics of non-model organisms.

Clutch Frequency Affects the Offspring Size-Number Trade-Off in Lizards

Background Studies of lizards have shown that offspring size cannot be altered by manipulating clutch size in species with a high clutch frequency. This raises a question of whether clutch frequency has a key role in influencing the offspring size-number trade-off in lizards. Methodology/Principal Findings To test the hypothesis that females reproducing more frequently are less likely to tradeoff offspring size against offspring number, we applied the follicle ablation technique to female Eremias argus (Lacertidae) from Handan (HD) and Gonghe (GH), the two populations that differ in clutch frequency. Follicle ablation resulted in enlargement of egg size in GH females, but not in HD females. GH females switched from producing a larger number of smaller eggs in the first clutch to a smaller number of larger eggs in the second clutch; HD females showed a similar pattern of seasonal shifts in egg size, but kept clutch size constant between the first two clutches. Thus, the egg size-number trade-off was evident in GH females, but not in HD females. Conclusions/Significance As HD females (mean u200a=u200a3.1 clutches per year) reproduce more frequently than do GH females (mean u200a=u200a1.6 clutches per year), our data therefore validate the hypothesis tested. Our data also provide an inference that maximization of maternal fitness could be achieved in females by diverting a large enough, rather than a higher-than-usual, fraction of the available energy to individual offspring in a given reproductive episode.

Proteomic characterization and comparison of venoms from two elapid snakes (Bungarus multicinctus and Naja atra) from China.

UNLABELLEDnBungarus multicinctus (many-banded krait) and Naja atra (Chinese cobra) are widely distributed and medically important venomous snakes in China; however, their venom proteomic profiles have not been fully compared. Here, we fractionated crude venoms and analyzed them using a combination of proteomic techniques. Three-finger toxins (3-FTx) and phospholipase A2 (PLA2) were most abundant in both species, respectively accounting for 32.6% and 66.4% of total B. multicinctus venom, and 84.3% and 12.2% of total N. atra venom. Venoms from these two species contained one common protein family and six less abundant species-specific protein families. The proteomic profiles of B. multicinctus and N. atra venoms and analysis of toxicological activity in mice suggested that 3-FTx and PLA2 are the major contributors to clinical symptoms caused by envenomation. The venoms differed in enzymatic activity, likely the result of inter-specific variation in the amount of related venom components. Antivenomics assessment revealed that a small number of venom components (3-FTxs and PLA2s in B. multicinctus, and 3-FTxs in N. atra) could not be immunocaptured completely, suggesting that we should pay attention to enhancing the immune response of these components in designing commercial antivenoms for B. multicinctus and N. atra.nnnBIOLOGICAL SIGNIFICANCEnThe proteomic profiles of venoms from two medically important snake species - B. multicinctus and N. atra - have been explored. Quantitative and qualitative differences are evident in both venoms when proteomic profiles and transcriptomic results are compared; this is a reminder that combined approaches are needed to explore the precise composition of snake venom. Two protein families (3-FTx and PLA2) of high abundance in these snake venoms are major players in the biochemical and pharmacological effects of envenomation. Elucidation of the proteomic profiles of these snake venoms is helpful in understanding composition-function relationships and will facilitate the clinical application of antivenoms.

Viviparity in high‑altitude Phrynocephalus lizards is adaptive because embryos cannot fully develop without maternal thermoregulation

Viviparous Phrynocephalus lizards (Agamidae) are mainly restricted to the Qinghai-Tibet Plateau of China. In this study, we used Phrynocephalus vlangalii females kept under seven thermal regimes for the whole gestation period to test the hypothesis that viviparity in high-altitude Phrynocephalus lizards is adaptive because embryos cannot fully develop without maternal thermoregulation. All females at 24xa0°C and 93xa0% of the females at 28xa0°C failed to give birth or produced stillborns, and proportionally fewer females gave birth at 29 or 35xa0°C than at 32xa0°C. Though the daily temperatures encountered were unsuitable for embryonic development, 95xa0% of the females in nature and 89xa0% of the females thermoregulating in the laboratory gave birth. There was no shift in the thermal preferences of females when they were pregnant. Although thermal conditions inside natural burrows were unsuitable for embryonic development, mass and sprint speed were both greater in neonates produced in nature. Our data show that (1) long-term exposure of P. vlangalii embryos to temperatures outside the range of 29–35xa0°C may result in the failure of development, but daily or short-term exposure may not necessarily increase embryonic mortality; (2) low gestation temperatures slow but do not arrest embryonic development, and females produce high-quality offspring in the shortest possible time by maintaining gestation temperatures close to the upper thermal limit for embryonic development; and (3) viviparity is currently adaptive at high elevations because embryos in nature cannot fully develop without relying on maternal thermoregulation. Our data validate the hypothesis tested.

Patterns of Interspecific Variation in the Heart Rates of Embryonic Reptiles

New non-invasive technologies allow direct measurement of heart rates (and thus, developmental rates) of embryos. We applied these methods to a diverse array of oviparous reptiles (24 species of lizards, 18 snakes, 11 turtles, 1 crocodilian), to identify general influences on cardiac rates during embryogenesis. Heart rates increased with ambient temperature in all lineages, but (at the same temperature) were faster in lizards and turtles than in snakes and crocodilians. We analysed these data within a phylogenetic framework. Embryonic heart rates were faster in species with smaller adult sizes, smaller egg sizes, and shorter incubation periods. Phylogenetic changes in heart rates were negatively correlated with concurrent changes in adult body mass and residual incubation period among the lizards, snakes (especially within pythons) and crocodilians. The total number of embryonic heart beats between oviposition and hatching was lower in squamates than in turtles or the crocodilian. Within squamates, embryonic iguanians and gekkonids required more heartbeats to complete development than did embryos of the other squamate families that we tested. These differences plausibly reflect phylogenetic divergence in the proportion of embryogenesis completed before versus after laying.

Combined venomics, antivenomics and venom gland transcriptome analysis of the monocoled cobra (Naja kaouthia) from China

We conducted an omics-analysis of the venom of Naja kaouthia from China. Proteomics analysis revealed six protein families [three-finger toxins (3-FTx), phospholipase A2 (PLA2), nerve growth factor, snake venom metalloproteinase (SVMP), cysteine-rich secretory protein and ohanin], and venom-gland transcriptomics analysis revealed 28 protein families from 79 unigenes. 3-FTx (56.5% in proteome/82.0% in transcriptome) and PLA2 (26.9%/13.6%) were identified as the most abundant families in venom proteome and venom-gland transcriptome. Furthermore, N. kaouthia venom expressed strong lethality (i.p. LD50: 0.79μg/g) and myotoxicity (CK: 5939U/l) in mice, and showed notable activity in PLA2 but weak activity in SVMP, l-amino acid oxidase or 5 nucleotidase. Antivenomic assessment revealed that several venom components (nearly 17.5% of total venom) from N. kaouthia could not be thoroughly immunocaptured by commercial Naja atra antivenom. ELISA analysis revealed that there was no difference in the cross-reaction between N. kaouthia and N. atra venoms against the N. atra antivenom. The use of commercial N. atra antivenom in treatment of snakebites caused by N. kaouthia is reasonable, but design of novel antivenom with the attention on enhancing the immune response of non-immunocaptured components should be encouraged.nnnBIOLOGICAL SIGNIFICANCEnThe venomics, antivenomics and venom-gland transcriptome of the monocoled cobra (Naja kaouthia) from China have been elucidated. Quantitative and qualitative differences are evident when venom proteomic and venom-gland transcriptomic profiles are compared. Two protein families (3-FTx and PLA2) are found to be the predominated components in N. kaouthia venom, and considered as the major players in functional role of venom. Other protein families with relatively low abundance appear to be minor in the functional significance. Antivenomics and ELISA evaluation reveal that the N. kaouthia venom can be effectively immunorecognized by commercial N. atra antivenom, but still a small number of venom components could not be thoroughly immunocaptured. The findings indicate that exploring the precise composition of snake venom should be executed by an integrated omics-approach, and elucidating the venom composition is helpful in understanding composition-function relationships and will facilitate the clinical application of antivenoms.

Genetic Structure and Demographic History Should Inform Conservation: Chinese Cobras Currently Treated as Homogenous Show Population Divergence

An understanding of population structure and genetic diversity is crucial for wildlife conservation and for determining the integrity of wildlife populations. The vulnerable Chinese cobra (Naja atra) has a distribution from the mouth of the Yangtze River down to northern Vietnam and Laos, within which several large mountain ranges and water bodies may influence population structure. We combined 12 microsatellite loci and 1117 bp of the mitochondrial cytochrome b gene to explore genetic structure and demographic history in this species, using 269 individuals from various localities in Mainland China and Vietnam. High levels of genetic variation were identified for both mtDNA and microsatellites. mtDNA data revealed two main (Vietnam + southern China + southwestern China; eastern + southeastern China) and one minor (comprising only two individuals from the westernmost site) clades. Microsatellite data divided the eastern + southeastern China clade further into two genetic clusters, which include individuals from the eastern and southeastern regions, respectively. The Luoxiao and Nanling Mountains may be important barriers affecting the diversification of lineages. In the haplotype network of cytchrome b, many haplotypes were represented within a “star” cluster and this and other tests suggest recent expansion. However, microsatellite analyses did not yield strong evidence for a recent bottleneck for any population or genetic cluster. The three main clusters identified here should be considered as independent management units for conservation purposes. The release of Chinese cobras into the wild should cease unless their origin can be determined, and this will avoid problems arising from unnatural homogenization.

The complete mitochondrial genome of Leiolepis reevesii (Sauria, Agamidae).

Abstract In this paper, we report the complete mitochondrial genome of Leiolepis reevesii (Sauria, Agamidae), which is a circular molecule of 16,908u2009bp in size and consists of 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs and a control region. The A+T content of overall base composition of H-strand is 59.8% (T: 25.1%, C: 27.5%, A: 34.7%, G: 12.7%). Some short microsatellite-like repeat regions (polyA and polyT) are scattered in the control region. All the results provide powerful data to further study of the molecular systematics, species identification and conservation genetics.