Extreme evolutionary conservation of CaSox4 gene in alligator Alligator sinensis

The SOX genes form a gene family related by homology to the high-mobility-group (HMG) box region of the testis-determining gene, SRY. Using degenerate primer PCR, we have cloned and sequenced the HMG-box motif of CaSox4 gene from genomic DNA of the Chinese alligator, Alligator sinensis, a species closely related to American alligator, Alligator mississippiensis. Sequence comparison showed that CaSox4 gene shared 97% homology at the nucleotide level and complete identity at the amino acid level, respectively, with AES4 gene of Alligator mississippiensis. It indicates that the Sox4 gene has been extremely conserved during evolution. The result obtained in the present study strongly suggests that the Sox4 gene did not diverge during the differentiation of two closely related alligators, which are thought to have originated from a common ancestor and have existed largely unchanged for 200-250 Myr.     

扬子鳄的CaSox4基因的分子克隆和进化分析

The completely identical HMG-box motif of CaSox4 gene from both male and female genomic DNA of the Chinese alligator (alligator sinensis) was cloned and sequenced by degenerate primer PCR.Compared with the human and mouse SRY,CaSox4 revealed 51% and 57% nucleotide homology respectively and 49% and 55% amino acid identity respectively,CaSox4 belongs to subgroup C of the Sox gene family.The GC content is 86% in the HMG-box region of the CaSox4 gene,Blast analysis showed that the CaSox4 gene shares 100^ amino acid identity with human Sox4.bird SoxLF4,turtle Sra4 and lizard CvSox4 genes.Casox4 may be orthologous with the human SOx4 gene.This indicates that CaSox4 gene shows the remarkable evolutionary conservation during the evolution of Alligator Sinensis,The extensive sequence conservation of the Sox4 gene between reptiles,mammals and birds suggests major functional constraints[Acta Zoologica Sinica 49(3):404-407,2003].     

扬子鳄的起源

介绍了现存鳄彼此间的亲缘关系、扬子鳄起源的时间和地点。主要结论可归纳如下：①现存鳄类在科、属、种分类上都存在不同意见;②扬子鳄与密河鳄被分在同一属内,但两者之间的亲缘关系存在争议,两者在形态、生化、细胞、分子生物学上差异很大;③Stell认为扬子鳄在中新世开始出现,作者认为它可能起源于渐新世或中新世早期;④Stell认为分布于北美的汤氏鳄（A.thomsoni)可能是扬子鳄的祖先类型,而另一些学者认为扬子鳄起源于亚洲的可能性更大些。     

Modelling the spatial patterning of teeth primordia in the alligator

We propose a model mechanism for the initiation and spatial positioning of teeth primordia in the alligator, Alligator mississippiensis. Detailed embryological studies by Westergaard and Ferguson (1986, 1987, 1990) have shown that jaw growth plays a crucial role in the developmental patterning of the tooth initiation process. Based on biological data we develop a dynamic patterning mechanism, which crucially includes domain growth. The mechanism can reproduce the spatial pattern development of the first seven teeth primordia in each half jaw of A. mississippiensis. The results for the precise spatio-temporal sequence compare well with experiment. Simulation of the model also predicts that certain transplantations can alter the spatial sequence of teeth primordia initiation.     

Growth rates of Chinese and American alligators

Growth rates in two closely related species, Alligator mississippiensis (American alligator) and Alligator sinensis (Chinese alligator), were compared under identical conditions for at least 1 year after hatching. When hatched, Chinese alligators were approximately 2/3 the length and approximately 1/2 the weight of American alligator hatchlings. At the end of 1 year of growth in captivity in heated chambers, the Chinese alligators were approximately 1/2 as long and weighed approximately 1/10 as much as American alligator yearlings. When the animals were maintained at 31 degrees C, Chinese alligator food consumption and length gain rates dropped to near zero during autumn and winter and body weights decreased slightly, apparently in response to the change in day length. At constant temperature (31 degrees C), food consumption by American alligators remained high throughout the year. Length gain rates in American alligators decreased slowly as size increased, but were not affected by photoperiod. Daily weight gains in American alligators increased steadily throughout the year. In autumn, provision of artificial light for 18 h a day initially stimulated both length and weight gain in Chinese alligators, but did not affect growth in American alligators. Continuation of the artificial light regimen seemed to cause deleterious effects in the Chinese alligators after several months, however, so that animals exposed to the normal light cycle caught up to and then surpassed the extra-light group in size. Even after removal of the artificial light, it was several months before these extra-light animals reverted to a normal growth pattern. These findings may be of interest to those institutions engaged in captive growth programs intended to provide animals for reintroduction to the wild or to protected habitat.     

基于BDNF和NT-3基因部分序列探讨爬行动物系统发生中的若干问题及扬子鳄分类地位

通过两个核基因NT-3基因(约740 bp)和BDNF基因部分序列(约720 bp)的分析,对爬行动物的系统发生关系中鳄类与鸟类和哺乳类系统发生关系、龟类在爬行动物系统发生中的位置以及扬子鳄属的划分等问题进行探讨.结果表明,在NT-3基因序列中,有307个位点存在变异(约为总位点数的47.3%),BDNF基因序列有256个变异位点(约为38.79%);构建的分子系统树显示,NT-3和BDNF基因以及两序列合并数据后所得系统树的拓扑结构均支持鳄类和鸟类聚为一支构成姐妹群,鳄类与蜥蜴类尽管在形态上非常相似,但它们的亲缘关系仍然较远;同时支持把龟鳖类作为鳄类和鸟类支系的姐妹群,支持把蜥蜴类(有鳞类)放在爬行类系统发生树的基部,而不是龟鳖类作为现代爬行类中最基部的1支.对扬子鳄分类地位的研究结果支持在现存的鳄类中扬子鳄与密西西比鳄的亲缘关系较近的结论.     

Gender differences in haemogregarine infections in American alligators (Alligator mississippiensis) at Savannah River, South Carolina, USA

We report a host gender bias in haemogregarine infection characteristics in the American alligator (Alligator mississippiensis) at the Savannah River Site, South Carolina, USA. Prevalence and severity in female alligators was higher than it was in males. The reason for this pattern is not clear.     

The complete mitochondrial genome of Alligator mississippiensis and the separation between recent archosauria (birds and crocodiles)

The complete mitochondrial genome of the alligator, Alligator mississippiensis, was sequenced. The size of the molecule is 16,642 nucleotides. Previously reported rearrangements of tRNAs in crocodile mitochondrial genomes were confirmed and, relative to mammals, no other deviations of gene order were observed. The analysis of protein-coding genes of the alligator showed an evolutionary rate that is roughly the same as in mammals. Thus, the evolutionary rate in the alligator is faster than that in birds as well as that in cold-blooded vertebrates. This contradicts hypotheses of constant body temperatures or high metabolic rate being correlated with elevated molecular evolutionary rates. It is commonly acknowledged that birds are the closest living relatives to crocodiles. Birds and crocodiles represent the only archosaurian survivors of the mass extinction at the Cretaceous/Tertiary boundary. On the basis of mitochondrial protein- coding genes, the Haemothermia hypothesis, which defines birds and mammals as sister groups and thus challenges the traditional view, could be rejected. Maximum-likelihood branch length data of amino acid sequences suggest that the divergence between the avian and crocodilian lineages took place at approximately equal to 254 MYA.      

Proteome analysis of the leukocytes from the American alligator (Alligator mississippiensis) using mass spectrometry

Mass spectrometry was used in conjunction with gel electrophoresis and liquid chromatography, to determine peptide sequences from American alligator (Alligator mississippiensis) leukocytes and to identify similar proteins based on homology. The goal of the study was to generate an initial database of proteins related to the alligator immune system. We have adopted a typical proteomics approach for this study. Proteins from leukocyte extracts were separated using two-dimensional gel electrophoresis and the major bands were excised, digested and analyzed by on-line nano-LC MS/MS to generate peptide sequences. The sequences generated were used to identify proteins and characterize their functions. The protein identity and characterization of the protein function were based on matching two or more peptides to the same protein by searching against the NCBI database using MASCOT and Basic Local Alignment Search Tool (BLAST). For those proteins with only one peptide matching, the phylum of the matched protein was considered. Forty-three proteins were identified that exhibit sequence similarities to proteins from other vertebrates. Proteins related to the cytoskeletal system were the most abundant proteins identified. These proteins are known to regulate cell mobility and phagocytosis. Several other peptides were matched to proteins that potentially have immune-related function.     

The karyotype of Alligator mississippiensis,and chromosomal mapping of the ZFY/X homologue,Zfc

Comparative mapping studies of X-linked genes in mammals have provided insights into the evolution of the X chromosome. Many reptiles including the American alligator, Alligator mississippiensis, do not appear to possess heteromorphic sex chromosomes, and sex is determined by the incubation temperature of the egg during embryonic development. Mapping of homologues of mammalian X-linked genes in reptiles could lead to a greater understanding of the evolution of vertebrate sex chromosomes. One of the genes used in the mammalian mapping studies was ZFX, an X-linked copy of the human ZFY gene which was originally isolated as a candidate for the mammalian testis-determining factor (TDF). ZFX is X-linked in eutherians, but maps to two autosomal locations in marsupials and monotremes, close to other genes associated with the eutherian X. The alligator homologue of the ZFY/ZFX genes, Zfc, has been isolated and described previously. A detailed karyotype of A. mississippiensis is presented, together with chromosomal in situ hybridisation data localising the Zfc gene to chromosome 3. Further chromosomal mapping studies using eutherian X-linked genes may reveal conserved chromosomal regions in the alligator that have become part of the eutherian X chromosome during evolution.     

Fluctuating water depths affect American alligator (Alligator mississippiensis) body condition in the Everglades,Florida, USA

Successful restoration of wetland ecosystems requires knowledge of wetland hydrologic patterns and an understanding of how those patterns affect wetland plant and animal populations. Within the Everglades, Florida, USA restoration, an applied science strategy including conceptual ecological models linking drivers to indicators is being used to organize current scientific understanding to support restoration efforts. A key driver of the ecosystem affecting the distribution and abundance of organisms is the timing, distribution, and volume of water flows that result in water depth patterns across the landscape. American alligators (Alligator mississippiensis) are one of the ecological indicators being used to assess Everglades restoration because they are a keystone species and integrate biological impacts of hydrological operations through all life stages. Alligator body condition (the relative fatness of an animal) is one of the metrics being used and targets have been set to allow us to track progress. We examined trends in alligator body condition using Fulton's K over a 15 year period (2000–2014) at seven different wetland areas within the Everglades ecosystem, assessed patterns and trends relative to restoration targets, and related those trends to hydrologic variables. We developed a series of 17 a priori hypotheses that we tested with an information theoretic approach to identify which hydrologic factors affect alligator body condition. Alligator body condition was highest throughout the Everglades during the early 2000s and is approximately 5–10% lower now (2014). Values have varied by year, area, and hydrology. Body condition was positively correlated with range in water depth and fall water depth. Our top model was the “Current” model and included variables that describe current year hydrology (spring depth, fall depth, hydroperiod, range, interaction of range and fall depth, interaction of range and hydroperiod). Across all models, interaction between range and fall water depth was the most important variable (relative weight of 1.0) followed by spring and fall water depths (0.99), range (0.96), hydroperiod (0.95) and interaction between range and hydroperiod (0.95). Our work provides additional evidence that restoring a greater range in annual water depths is important for improvement of alligator body condition and ecosystem function. This information can be incorporated into both planning and operations to assist in reaching Everglades restoration goals.     

Primary structure of two forms of gonadotropin-releasing hormone from brains of the American alligator (Alligator mississippiensis)

Two forms of gonadotropin-releasing hormone (GnRH) have been purified from brains of the American alligator, Alligator mississippiensis, using reverse-phase high-pressure liquid chromatography (HPLC). The concentration of total GnRH was 8.8 ng/g of frozen brain tissue or 21.1 ng per brain. The amino acid sequence of each form of GnRH was determined using automated Edman degradation. The presence of the N-terminal pGlu residue was established by digestion studies with bovine pyroglutamyl aminopeptidase and coelution with synthetic forms of the native peptide. The primary structure of alligator GnRH I is pGlu-His-Trp-Ser-Tyr-Gly-Leu-Gln-Pro-Gly-NH2 and alligator GnRH II is pGlu-His-Trp-Ser-His-Gly-Trp-Tyr-Pro-Gly-NH2.     

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described.     

Egg yolk serositis in an American alligator (Alligator mississippiensis)

An adult female American alligator (Alligator mississippiensis) had diffuse, yellow, granular serosal thickening at necropsy. Light microscopic examination of affected stomach, small intestine and spleen revealed a chronic proliferative serositis associated with 3 to 15 micron eosinophilic extracellular globules identified histochemically and morphologically as egg yolk. The intracoelomic egg yolk was considered to be the cause of the serosal reaction.     

扬子鳄4个Sox基因保守区的克隆及序列分析

参考人SRY基因HMG－box的保守区序列,设计一对简并引物,用PCR扩增了扬子鳄Sox基因的HMG－box,并对PCR产物进行了亚克隆和测序。结果在雌雄个体中均筛选到4个不同的Sox基因,无性别差异。其序列与人相应的SOX基因保守区编码序列的相似性分别为91％、96％、100％、96％,分别命名为AS－Sox1,ASSox2,ASSox11,ASSox22。与其他动物相关的Sox/SOX基因的聚类分析结果表明,扬子鳄Sox基因编码的氨基酸序列与进化位置各异的其他动物的Sox/SOX基因编码的氨基酸序列存在高度的同源性,显示出Sox基因在系统进化上的高度保守性。     

Changes in thyrotropin-releasing hormone levels in alligator stomach during fasting

Stomach tissue of the American alligator, Alligator mississippiensis, contains substantial levels of thyrotropin-releasing hormone (TRH) which behaves identically to the synthetic hormone on radioimmunoassay (RIA) and high performance liquid chromatography (HPLC). Fasting induces a marked increase in gastric tissue levels of this hypophysiotropic hormone, but is without effect on hypothalamic content, suggesting a physiological role for TRH in gastric function of this vertebrate.