Jian-Fang Gao

Neonate-to-adult transition of snake venomics in the short-tailed pit viper, Gloydius brevicaudus.

UNLABELLED Snake venoms undergo ontogenetic shifts in biochemical and pharmacological activities. This may be related to variation in venom components associated with the ontogenetic shift in diet. We used the short-tailed pit viper Gloydius brevicaudus that displays ontogenetic shifts in diet to examine whether the species displays a neonate-to-adult transition of snake venomics. Venoms from neonates and adults were pooled separately and then analyzed by 2-DE, MALDI-TOF-MS/MS and iTRAQ technologies. The 2-DE profiles showed that the main components in both types of venoms were acidic proteins, and that neonates and adults differed in snake venomics. The proteins with molecular masses/pI of ~12-39kDa/4.0-4.6, ~36-57kDa/5.6-7.0 and ~65-92kDa/4.5-5.8 were more abundant in the neonate venom, while the proteins with molecular masses/pI of ~12-19kDa/4.6-6.4, ~23-30kDa/5.4-6.3 and ~35-62kDa/4.6-5.4 were more abundant in the adult venom. The iTRAQ analysis showed quantitative changes in various toxin families, including mainly metalloproteinases, serine proteinases, phospholipase A2s and C-type lectins. The N-deglycosylation analysis demonstrated that glycosylation was an important post-translational modification of snake venom. Our results show a neonate-to-adult transition of snake venomics in G. brevicaudus. Such a transition might be driven by the divergence in dietary habits between neonates and adults. BIOLOGICAL SIGNIFICANCE This study is first to demonstrate a neonate-to-adult transition of snake venomics in G. brevicaudus, and the results will be helpful in predicting and treating clinical pathologic symptoms caused by the snake at different developmental stages.

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

UNLABELLED Bungarus 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. BIOLOGICAL SIGNIFICANCE The 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.

Immunoreactivity between venoms and commercial antiserums in four Chinese snakes and venom identification by species-specific antibody.

We studied the immunoreactivity between venoms and commercial antiserums in four Chinese venomous snakes, Bungarus multicinctus, Naja atra, Deinagkistrodon acutus and Gloydius brevicaudus. Venoms from the four snakes shared common antigenic components, and most venom components expressed antigenicity in the immunological reaction between venoms and antiserums. Antiserums cross-reacted with heterologous venoms. Homologous venom and antiserum expressed the highest reaction activity in all cross-reactions. Species-specific antibodies (SSAbs) were obtained from four antiserums by immunoaffinity chromatography: the whole antiserum against each species was gradually passed through a medium system coated with heterologous venoms, and the cross-reacting components in antiserum were immunoabsorbed by the common antigens in heterologous venoms; the unbound components (i.e., SSAbs) were collected, and passed through Hitrap G protein column and concentrated. The SSAbs were found to have high specificity by western blot and enzyme-linked immunosorbent assay (ELISA). A 6-well ELISA strip coated with SSAbs was used to assign a venom sample and blood and urine samples from the envenomed rats to a given snake species. Our detections could differentiate positive and negative samples, and identify venoms of a snake species in about 35 min. The ELISA strips developed in this study are clinically useful in rapid and reliable identification of venoms from the above four snake species.

Does the oviparity-viviparity transition alter the partitioning of yolk in embryonic snakes?

BackgroundThe oviparity-viviparity transition is a major evolutionary event, likely altering the reproductive process of the organisms involved. Residual yolk, a portion of yolk remaining unutilized at hatching or birth as parental investment in care, has been investigated in many oviparous amniotes but remained largely unknown in viviparous species. Here, we used data from 20 (12 oviparous and 8 viviparous) species of snakes to see if the oviparity-viviparity transition alters the partitioning of yolk in embryonic snakes. We used ANCOVA to test whether offspring size, mass and components at hatching or birth differed between the sexes in each species. We used both ordinary least squares and phylogenetic generalized least squares regressions to test whether relationships between selected pairs of offspring components were significant. We used phylogenetic ANOVA to test whether offspring components differed between oviparous and viviparous species and, more specifically, the hypothesis that viviparous snakes invest more in the yolk as parental investment in embryogenesis to produce more well developed offspring that are larger in linear size.ResultsIn none of the 20 species was sex a significant source of variation in any offspring component examined. Newborn viviparous snakes on average contained proportionally more water and, after accounting for body dry mass, had larger carcasses but smaller residual yolks than did newly hatched oviparous snakes. The rates at which carcass dry mass (CDM) and fat body dry mass (FDM) increased with residual yolk dry mass (YDM) did not differ between newborn oviparous and viviparous snakes. Neither CDM nor FDM differed between newborn oviparous and viviparous snakes after accounting for YDM.ConclusionsOur results are not consistent with the hypothesis that the partitioning of yolk between embryonic and post-embryonic stages differs between snakes that differ in parity mode, but instead show that the partitioning of yolk in embryonic snakes is species-specific or phylogenetically related. We conclude that the oviparity-viviparity transition does not alter yolk partitioning in embryonic snakes.