Chengdong Wang

Factors Affecting the Outcome of Artificial Insemination Using Cryopreserved Spermatozoa in the Giant Panda (Ailuropoda melanoleuca)

Artificial insemination (AI) is an important component of captive breeding programs for endangered species, such as the giant panda. The panda has been the subject of increasingly successful captive breeding programs involving a compilation of assisted breeding techniques, including AI using cryopreserved spermatozoa. AI implementation is currently hampered by a lack of understanding of the factors that may cause failure. We investigated factors influencing the probability of success of AI for 14 giant panda females housed at the China Center for Research and Conservation of the Giant Panda (CCRCGP) inseminated in a total of 20 instances using cryopreserved spermatozoa from 11 males currently residing in 6 different captive breeding institutions. One of the pandas was the oldest giant panda female to ever successfully conceive from AI (20.5 years old). The success of AI was significantly affected by the timing of AI in relationship to both timing of peak urinary estrogen of the female and percent decline in urinary estrogen between the peak level and the first AI attempt. Our results suggest that the window for successful AI in giant pandas may be narrower than previously suspected, although individual differences in rates of decline in urinary estrogen may reflect some degree of variation in this crucial window across females. Our results are consistent with recent research on pandas and other species that demonstrates the efficacy of cryopreserved spermatozoa for AI and highlights the need for more in-depth analysis of factors related to female physiology that may influence its success.

Influenza A(H1N1)pdm09 Virus Infection in Giant Pandas, China

We confirmed infection with influenza A(H1N1)pdm09 in giant pandas in China during 2009 by using virus isolation and serologic analysis methods. This finding extends the host range of influenza viruses and indicates a need for increased surveillance for and control of influenza viruses among giant pandas.

Analysis of Codon Usage Bias in Interferon Alpha Gene of the Giant Panda (Ailuropoda Melanoleuca)

The analysis on codon usage bias of IFN-a gene of giant panda (Ailuropoda melanoleuca) may provide a basis for understanding the evolution relationship of giant panda and for selecting appropriate host expression systems to improve the expression of target genes. In this paper, the codon usage bias in the mature IFN-a sequence of giant panda and 15 reference species have been analyzed. The results showed that the synonymous codons with G and C at the third codon position were widely used and the ENC-GC3S plot revealed that the genetic heterogeneity in IFN-a gene was main constrained by mutational bias. Contrastive analysis revealed that there were 40 codons showing distinct usage differences between GpIFN-a and Escherichia coli, 38 codons between GpIFN-a and yeast. and only 30 between GpIFN-a and Homo sapiens. Therefore the Homo expression system may be more suitable for the expression of GpIFN-a genes.

Sequence analysis of the ATP synthase of subunits (ATP8 and ATP6) genes of mitochondrial DNA genome from Ailuropoda melanoleuca

Abstract To explore the effects of the mutations of ATP6 and ATP8 genes on energy metabolism and genetic structure, we sequenced the ATP6 and ATP8 genes of Ailuropoda melanoleuca. Our results showed that ATP8 is a conserved gene and ATP6 gene is positively selected during the evolution of the giant panda population with a low genetic diversity. Population expansion was observed in the giant panda group. The T179C mutation on Haplotype7 made the production of a potential phosphorylation site. This non-synonymous mutation may occur at the post-translational modification site that have a potential effect on the function of ATP synthase, related to the maintenance of body temperature of pandas at low metabolic rates.

Transcriptome Sequencing Reveals the Differentially Expressed lncRNAs and mRNAs Involved in Cryoinjuries in Frozen-Thawed Giant Panda (Ailuropoda melanoleuca) Sperm

Sperm cryopreservation and artificial insemination are important methods for giant panda breeding and preservation of extant genetic diversity. Lower conception rates limit the use of artificial insemination with frozen-thawed giant panda sperm, due to the lack of understanding of the cryodamaging or cryoinjuring mechanisms in cryopreservation. Long non-coding RNAs (lncRNAs) are involved in regulating spermatogenesis. However, their roles during cryopreservation remain largely unexplored. Therefore, this study aimed to identify differentially expressed lncRNAs and mRNAs associated with cryodamage or freeze tolerance in frozen-thawed sperm through high throughput sequencing. A total of 61.05 Gb clean reads and 22,774 lncRNA transcripts were obtained. From the sequencing results, 1477 significantly up-regulated and 1,396 significantly down-regulated lncRNA transcripts from fresh and frozen-thawed sperm of giant panda were identified. GO and KEGG showed that the significantly dysregulated lncRNAs and mRNAs were mainly involved in regulating responses to cold stress and apoptosis, such as the integral component of membrane, calcium transport, and various signaling pathways including PI3K-Akt, p53 and cAMP. Our work is the first systematic profiling of lncRNA and mRNA in fresh and frozen-thawed giant panda sperm, and provides valuableinsights into the potential mechanism of cryodamage in sperm.

Complete mitochondrial genome sequence of Jianmen-guan phasianus colchicus (Aves, Gallifoemes, phasianidae) and its phylogenetic analysis

Abstract Jianmen-guan phasianus colchicusare one of the most important and be cherished uncultivated animals, although, origins of the most J. phasianus colchicus remain unknown. Therefore, the complete mitochondrial genome of the J. phasianus colchicus (also named seven-colours J. phasianus colchicus, one of the oldest wild birds in China known for their preciousness) was obtained for the first time, the mitogenome is 166,786 bp in length, and it harbours 2rRNA genes, 13 protein-coding genes, 22tRNA genes, and a D-loop region. According to the phylogenetic tree, we can assume that Jianmen-guan and Xianju are within the same lineage, and J. phasianus colchicus is a different group with Red Jungle.

Molecular Cloning and Sequence Analysis of the Gene Encoding Interferon Alpha of the Giant Panda (Ailuropoda melanoleuca)

nterferon-alpha (IFN-a) is a kind of cytokines that share antiviral, as well as immunomodulatory, and antiproliferative effects on cell functions. In this report, the cDNA for Ailuropoda melanoleucas interferon alpha was cloned from the ConA-stimulated giant panda peripheral blood mononuclear cells (PBMCs) by RT-PCR. Bioinformatics analysis was performed to predict the characteristics of this gene. Sequencing revealed that the fragment was composed of 495 nucleotides, intronless, encoding a mature polypeptide with 164 amino acids and exhibiting a molecular mass of 18.15 kDa. The analysis of the functional sites and antigenic determinants demonstrated that this protein has 27 functional sites and 9 antigenic determinants, And possesses typical characteristics of interferon alpha, beta and delta family. Compared with 10 corresponding IFN-α sequences. It revealed that the GpIFN-a gene had a close evolutionary relationship with mammalians IFN-a. Phylogenetic tree based on nucleotide sequences showed that giant panda, ferret, dog and cat clustered together and evolved into a distinct phylogenetic lineage. In conclusion, all the data and consequences will provide a basis for further functional study of the Ailuropoda melanoleuca IFN-a.