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Background: MicroRNAs can regulate various biological functions including cell proliferation, differentiation, embryo formation, and implantation. The giant panda exhibits embryonic diapause, with embryo development resuming in late pregnancy. However, the changes in microRNAs during late pregnancy remain poorly understand. Methods and results: After mating, plasma samples were collected on day 40 of early pregnancy (EP; n = 3) and 30 days before delivery of late pregnancy (LP; n = 3). Following microRNAs screening, a total of 120 microRNAs were detected in the plasma exosomes of pregnant pandas. Nine differentially expressed microRNAs (DEmicroRNAs) were identified in LP compared to EP, including three that were upregulated and six that were downregulated. Notably, miR-25b and miR-47 were significantly downregulated in LP group. All DEmicroRNAs were predicted to target a total of 2,675 genes. Pathway enrichment analysis of these target genes revealed significant enrichment in the MAPK and Rap1 signaling pathways, which are closely related to cell proliferation, differentiation, and cell-cell and cell-matrix interactions. Analysis of protein-protein interaction networks showed that most of the hub genes (five out of eight), including Fgfr1, Fgf2, Fgf18, Erbb4, and Kras within the MAPK and Rap1 pathways are associated with the cell proliferation and differentiation. Significantly, Erbb4 was regulated by significantly differentially expressed miRNA-47. Conclusions: We suggest that plasma exosomal microRNAs are involved in cell proliferation and differentiation during embryonic development by regulating key hub genes within MAPK and Rap1 pathways. These findings provided new insights into the development of giant panda embryos. [ABSTRACT FROM AUTHOR]

Simple Summary: Red pandas have evolved to become specialized bamboo eaters within Carnivora. Probably due to the difficulty in obtaining materials, reports on genes related to digestion and metabolism at expression and regulation levels in red pandas are rare. Red pandas and carnivorous mammal ferrets have a close phylogenetic relationship. They both consume highly nutritious milk during the suckling period, but consume low-energy bamboo and high-energy meat during the adult period, respectively. In order to explore the molecular mechanisms of dietary changes and nutrient utilization in red pandas, we discussed (1) the differences in expression changes of some genes related to digestion and metabolism in the stomach of these two species after food changes, as well as the expression adaptation of genes related to digestion and metabolism in both species during different feeding periods, and (2) the regulatory effects of lncRNAs and miRNAs on adaptive expressions of genes related to digestion and metabolism in both species from suckling to adult. Red pandas evolved from carnivores to herbivores and are unique within Carnivora. Red pandas and carnivorous mammals consume milk during the suckling period, while they consume bamboo and meat during the adult period, respectively. Red pandas and carnivorous mammal ferrets have a close phylogenetic relationship. To further investigate the molecular mechanisms of dietary changes and nutrient utilization in red pandas from suckling to adult, comparative analysis of the whole transcriptome was performed on stomach tissues from red pandas and ferrets during the suckling and adult periods. The main results are as follows: (1) we identified ncRNAs for the first time in stomach tissues of both species, and found significant expression changes of 109 lncRNAs and 106 miRNAs in red pandas and 756 lncRNAs and 109 miRNAs in ferrets between the two periods; (2) up-regulated genes related to amino acid transport regulated by lncRNA-miRNA-mRNA networks may efficiently utilize limited bamboo amino acids in adult red pandas, while up-regulated genes related to amino acid degradation regulated by lncRNAs may maintain the balance of amino acid metabolism due to larger daily intakes in adult ferrets; and (3) some up-regulated genes related to lipid digestion may contribute to the utilization of rich nutrients in milk for the rapid growth and development of suckling red pandas, while up-regulated genes associated with linoleic acid metabolism regulated by lncRNA-miRNA-mRNA networks may promote cholesterol decomposition to reduce health risks for carnivorous adult ferrets. Collectively, our study offers evidence of gene expression adaptation and ncRNA regulation in response to specific dietary changes and nutrient utilization in red pandas during suckling and adult periods. [ABSTRACT FROM AUTHOR]

Epigenetic regulation plays an important role in the evolution of species adaptations, yet little information is available on the epigenetic mechanisms underlying the adaptive evolution of bamboo‐eating in both giant pandas (Ailuropoda melanoleuca) and red pandas (Ailurus fulgens). To investigate the potential contribution of epigenetic to the adaptive evolution of bamboo‐eating in giant and red pandas, we performed hepatic comparative transcriptome and methylome analyses between bamboo‐eating pandas and carnivorous polar bears (Ursus maritimus). We found that genes involved in carbohydrate, lipid, amino acid, and protein metabolism showed significant differences in methylation and expression levels between the two panda species and polar bears. Clustering analysis of gene expression revealed that giant pandas did not form a sister group with the more closely related polar bears, suggesting that the expression pattern of genes in livers of giant pandas and red pandas have evolved convergently driven by their similar diets. Compared to polar bears, some key genes involved in carbohydrate metabolism and biological oxidation and cholesterol synthesis showed hypomethylation and higher expression in giant and red pandas, while genes involved in fat digestion and absorption, fatty acid metabolism, lysine degradation, resistance to lipid peroxidation and detoxification showed hypermethylation and low expression. Our study elucidates the special nutrient utilization mechanism of giant pandas and red pandas and provides some insights into the molecular mechanism of their adaptive evolution of bamboo feeding. This has important implications for the breeding and conservation of giant pandas and red pandas. [ABSTRACT FROM AUTHOR]

A non-invasive, fast, and reliable method for sex identification of the giant panda (Ailuropoda melanoleuca) was developed by genotyping five SNPs located at the sex chromosome amelogenin (AMEL) paralogs. The giant panda's sex can be accurately distinguished by the homozygous or heterozygous SNP genotypes. Mass spectrometry assay and Sanger sequencing both successfully genotyped tissue and fresh fecal samples collected from known-sex giant pandas. Additionally, we simulated the conditions of wild fecal samples, which were naturally degraded for up to two weeks, and successfully genotyped these samples by mass spectrometry assay in parallel. These results suggest that this is a powerful method for sex determination in highly degraded and lower concentrated DNA samples. [ABSTRACT FROM AUTHOR]

Simple Summary: In this study, 471 microsatellite loci which are common among 8 pheasant species were screened based on genome data, and 119 loci were selected to develop microsatellite markers. After PCR amplifications and reaction condition optimizations, and validation of microsatellite loci in 14 species of 11 genera within Phasianidae. Finally, 49 potentially universal microsatellite markers in the pheasant species were obtained. The 49 potentially universal microsatellite loci were successfully applied to a genetic diversity assessment and comparison for three pheasant birds, the Sichuan Hill partridge, blood pheasant and buff-throated partridge. These 49 microsatellite loci are potentially universal microsatellite loci for pheasants and are of great significance to establish a shared platform in population genetics study of pheasants. Pheasants are widely distributed in the southwest of China, but many of them are endangered due to habitat fragmentation and environmental changes. Genetic diversity is crucial for species to maintain their evolutionary potential, and thus it is important to develop universal genetic markers for facilitating the assessment of genetic diversity and planning effective conservation actions in these endangered species. In this study, 471 microsatellite loci which are common among eight pheasant species were screened based on genome data, and 119 loci were selected to develop microsatellite markers. After PCR amplifications and reaction condition optimizations, and validation of microsatellite loci in 14 species of 11 genera within Phasianidae. Finally, 49 potentially universal microsatellite markers in pheasant species were obtained. These microsatellite markers were successfully applied to assess the genetic diversity of 3 pheasant species. The Sichuan hill partridge (Arborophila rufipectus), blood pheasant (Ithaginis cruentus), buff-throated partridge (Tetraophasis szechenyii) and Sichuan hill partridge had a relatively low genetic diversity level. These 49 microsatellite loci are potentially universal microsatellite loci for pheasants and are of great significance to establish a shared platform in population genetics study of pheasants. [ABSTRACT FROM AUTHOR]

Simple Summary: To explore a safe and effective way to potentiate the systemic immunity of animals against infectious diseases in an economical way, the interleukin-7 (IL-7) gene of Tibetan pig was utilized to construct a recombinant eukaryotic plasmid. We first detected the bioactivity of expressed IL-7 on porcine lymphocytes in vitro and then encapsulated the IL-7 gene with different chitosan (CS) and its two modified derivatives, CS-PEG-PEI and CS-PEG-GAL nanoparticles. Then, we injected mice intramuscularly or intraperitoneally with various nanoparticles containing the IL-7 gene to evaluate their immune effects in vivo. We observed a significant increase in neutralizing antibodies and specific IgG levels in response to the rabies vaccine in the treated mice compared to the controls. Treated mice also manifested elevated leukocytes, CD4+ and CD8+ T cells, and mRNA levels of toll-like receptors (TLR1/4/6/9), IL-1, IL-2, IL-4, IL-6, IL-7, IL-23, and transforming growth factor-beta (TGF-β). Notably, the recombinant IL-7 gene encapsulated in CS-PEG-PEI provoked the best increases of immunoglobulins, CD4+ and CD8+ T cells, TLRs, and cytokines in the blood of the mice, implying that chitosan-PEG-PEI can become a promising carrier for in vivo IL-7 gene expression to enhance the innate and adaptive immunity for the prevention of animal diseases. To investigate a safe and effective approach for enhancing the in vivo expression of recombinant genes and improving the systemic immunity of animals against infectious diseases, we employed the interleukin-7 (IL-7) gene from Tibetan pigs to construct a recombinant eukaryotic plasmid (VRTPIL-7). We first examined VRTPIL-7's bioactivity on porcine lymphocytes in vitro and then encapsulated it with polyethylenimine (PEI), chitosan copolymer (CS), PEG-modified galactosylated chitosan (CS-PEG-GAL) and methoxy poly (ethylene glycol) (PEG) and PEI-modified CS (CS-PEG-PEI) nanoparticles using the ionotropic gelation technique. Next, we intramuscularly or intraperitoneally injected mice with various nanoparticles containing VRTPIL-7 to evaluate their immunoregulatory effects in vivo. We observed a significant increase in neutralizing antibodies and specific IgG levels in response to the rabies vaccine in the treated mice compared to the controls. Treated mice also exhibited increased leukocytes, CD8+ and CD4+ T lymphocytes, and elevated mRNA levels of toll-like receptors (TLR1/4/6/9), IL-1, IL-2, IL-4, IL-6, IL-7, IL-23, and transforming growth factor-beta (TGF-β). Notably, the recombinant IL-7 gene encapsulated in CS-PEG-PEI induced the highest levels of immunoglobulins, CD4+ and CD8+ T cells, TLRs, and cytokines in the mice's blood, suggesting that chitosan-PEG-PEI may be a promising carrier for in vivo IL-7 gene expression and enhanced innate and adaptive immunity for the prevention of animal diseases. [ABSTRACT FROM AUTHOR]

Simple Summary: Olfaction in animals plays important roles in many aspects, such as food recognition, mate detection and risk avoidance and social communication. Compared to other Ursidae species, the obligate bamboo feeder, giant panda, shows special diet, and how the diet transformation affects the olfactory system remains little known. In this study, we identified the olfactory receptor (OR) genes of the giant panda based on the chromosome-level genome and conducted comparative analysis of OR genes among Ursidae species. The giant panda had 639 OR genes, and chromosome 8 had the most OR genes. The giant panda had 31 unique OR gene subfamilies (containing 35 OR genes), of which 10 genes were clustered into 8 subfamilies with 10 known human OR genes (OR8J3, OR51I1, OR10AC1, OR1S2, OR1S1, OR51S1, OR4M1, OR4M2, OR51T1 and OR5W2). Compared to other Ursidae species, the giant panda lacked OR genes similar to OR2B1, OR10G3, OR11H6 and OR11H7P, which may be related to the diet transformation from carnivore to herbivore. Hence, these results may shed light on the olfactory function and variation of the giant panda. The giant panda (Ailuropoda melanoleuca) is the epitome of a flagship species for wildlife conservation and also an ideal model of adaptive evolution. As an obligate bamboo feeder, the giant panda relies on the olfaction for food recognition. The number of olfactory receptor (OR) genes and the rate of pseudogenes are the main factors affecting the olfactory ability of animals. In this study, we used the chromosome-level genome of the giant panda to identify OR genes and compared the genome sequences of OR genes with five other Ursidae species (spectacled bear (Tremarctos ornatus), American black bear (Ursus americanus), brown bear (Ursus arctos), polar bear (Ursus maritimus) and Asian black bear (Ursus thibetanus)). The giant panda had 639 OR genes, including 408 functional genes, 94 partial OR genes and 137 pseudogenes. Among them, 222 OR genes were detected and distributed on 18 chromosomes, and chromosome 8 had the most OR genes. A total of 448, 617, 582, 521 and 792 OR genes were identified in the spectacled bear, American black bear, brown bear, polar bear and Asian black bear, respectively. Clustering analysis based on the OR protein sequences of the six species showed that the OR genes distributed in 69 families and 438 subfamilies based on sequence similarity, and the six mammals shared 72 OR gene subfamilies, while the giant panda had 31 unique OR gene subfamilies (containing 35 genes). Among the 35 genes, there are 10 genes clustered into 8 clusters with 10 known human OR genes (OR8J3, OR51I1, OR10AC1, OR1S2, OR1S1, OR51S1, OR4M1, OR4M2, OR51T1 and OR5W2). However, the kind of odor molecules can be recognized by the 10 known human OR genes separately, which needs further research. The phylogenetic tree showed that 345 (about 84.56%) functional OR genes were clustered as Class-II, while only 63 (about 15.44%) functional OR genes were clustered as Class-I, which required further and more in-depth research. The potential odor specificity of some giant panda OR genes was identified through the similarity to human protein sequences. Sequences similar to OR2B1, OR10G3, OR11H6 and OR11H7P were giant panda-specific lacking, which may be related to the transformation and specialization from carnivore to herbivore of the giant panda. Since our reference to flavoring agents comes from human research, the possible flavoring agents from giant panda-specific OR genes need further investigation. Moreover, the conserved motifs of OR genes were highly conserved in Ursidae species. This systematic study of OR genes in the giant panda will provide a solid foundation for further research on the olfactory function and variation of the giant panda. [ABSTRACT FROM AUTHOR]

There are only six isolated living giant panda populations, and a comprehensive understanding of their genetic health status is crucial for the conservation of this vulnerable species. Liangshan Mountains is one of the main distribution areas of living giant pandas and is outside the newly established Giant panda national park. In this study, 971 giant panda fecal samples were collected in the heartland of Liangshan Mountains (Mabian Dafengding Nature Reserve: MB; Meigu Dafengding Nature Reserve: MG; and Heizhugou Nature Reserve: HZG). Microsatellite markers and mitochondrial D‐loop sequences were used to estimate population size and genetic diversity. We identified 92 individuals (MB: 27, MG: 22, HZG: 43) from the three reserves. Our results showed that: (1) genetic diversity of three giant panda populations was moderate; (2) several loci deviated significantly from the Hardy–Weinberg equilibrium and almost all these deviated loci showed significant heterozygote deficiencies and inbreeding; (3) three giant panda populations have substantial genetic differentiation with the most differentiation between MB and the two other populations; and (4) a large amount of giant panda feces outside the three reserves were found, implying the existence of protection gap. These results indicated that under stochastic events, the giant panda populations in Liangshan Mountains are at risk of genetic decline or extinction and urgent need of human management. This study revealed that high attention should be paid to the protection of these giant panda populations outside the Giant panda national park, to ensure their survival in their distribution areas. [ABSTRACT FROM AUTHOR]

Background: It is inevitable to change the function or expression of genes during the environmental adaption of species. Both the giant panda (Ailuropoda melanoleuca) and red panda (Ailurus fulgens) belong to Carnivora and have developed similar adaptations to the same dietary switch to bamboos at the morphological and genomic levels. However, the genetic adaptation at the gene expression level is unclear. Therefore, we aimed to examine the gene expression patterns of giant and red panda convergent specialized bamboo-diets. We examined differences in liver and pancreas transcriptomes between the two panda species and other non-herbivorous species. Results: The clustering and PCA plots suggested that the specialized bamboo diet may drive similar expression shifts in these two species of pandas. Therefore, we focused on shared liver and pancreas DEGs (differentially expressed genes) in the giant and red panda relative to other non-herbivorous species. Genetic convergence occurred at multiple levels spanning carbohydrate metabolism, lipid metabolism, and lysine degradation. The shared adaptive convergence DEGs in both organs probably be an evolutionary response to the high carbohydrate, low lipid and lysine bamboo diet. Convergent expression of those nutrient metabolism-related genes in both pandas was an intricate process and subjected to multi-level regulation, including DNA methylation and transcription factor. A large number of lysine degradation and lipid metabolism related genes were hypermethylated in promoter regions in the red panda. Most genes related to carbohydrate metabolism had reduced DNA methylation with increased mRNA expression in giant pandas. Unlike the red panda, the core gene of the lysine degradation pathway (AASS) doesn't exhibit hypermethylation modification in the giant panda, and dual-luciferase reporter assay showed that transcription factor, NR3C1, functions as a transcriptional activator in AASS transcription through the binding to AASS promoter region. Conclusions: Our results revealed the adaptive expressions and regulations of the metabolism-related genes responding to the unique nutrients in bamboo food and provided data accumulation and research hints for the future revelation of complex mechanism of two pandas underlying convergent adaptation to a specialized bamboo diet. [ABSTRACT FROM AUTHOR]

Background: The malignant behaviors of lung cancers are affected by not only cancer cells but also many kinds of stromal cells in tumor microenvironment (TME), including macrophages. Macrophages have been proven to extensively influence tumor progression through several mechanisms, among which switching of macrophages from pro-inflammatory phenotypes (M1-like) to anti-inflammatory phenotypes (M2-like) mediated by transcription factors such as nuclear factor κB (NF-κB) is the most crucial event. The regulation of NF-κB has been well studied, however some details remain fuzzy. Methods: Mouse primary bone marrow-derived macrophages (BMDMs) were cultured in Lewis lung carcinoma cell line LL-2-derived conditioned medium (LL-2-CM). Proliferation, migration, and polarization of BMDMs were tested by CCK8, scratch test, transwell, and flow cytometry. Secretion of several cytokines were detected by ELISA or cytometric bead array. To further explore the underlying mechanisms, BMDMs cultured in LL-2-CM were harvested for RNA-seq. Cytosolic calcium was detected by calcium probe Fluo-4-AM. Western blot was applied to exam the activation of NF-κB signal. BAPTA-AM was applied to sequestrate cytosolic calcium to further investigate the relationship between calcium and NF-κB signal. The polarization, calcium alteration, and NF-κB signal activation were further validated in BMDMs treated by CMT-64-derived conditioned medium (CMT-64-CM). Results: LL-2-CM promoted proliferation, migration, and M2-like polarization of BMDMs and inhibited M1-like polarization of BMDMs. However two pro-inflammatory cytokines, interleukin-6 (IL-6) and tumor necrosis factor- α (TNF- α ) were secreted. RNA-seq indicated that LL-2-CM activated both canonical and non-canonical NF-κB signal in BMDMs. Western blot showed that canonical NF-κB was temporarily elicited and attenuated at 24 h, while non-canonical NF-κB was consistently activated. At the same time, expression of genes that regulate cytosolic calcium ion concentration were down regulated, which caused diminution of cytosolic calcium in BMDMs treated with LL-2-CM. The decreased cytosolic calcium, M2-like polarization, and NF-κB activation was also observed in CMT-64-CM treated BMDMs. On the contrary, elevated cytosolic calcium was observed during M1-like polarization of BMDMs elicited by lipopolysaccharide (LPS). Interestingly, administration of calcium chelator, BAPTA-AM, impeded activation of canonical NF-κB and expression of M1-like marker induced by LPS, which further confirmed the relationship between cytosolic calcium and canonical NF-κB signal. Conclusions: In summary, lung cancer cell-derived conditioned medium promoted migration, proliferation, and M2-like polarization of BMDMs. The suppressed M1-like polarization was achieved through mitigating canonical NF-κB pathway via diminishing cytosolic calcium concentration. As far as we know, our work firstly revealed that cytosolic calcium is the key during inhibition of canonical NF-κB and M1-like polarization in macrophages by tumor cells. [ABSTRACT FROM AUTHOR]

Background: The giant panda (Ailuropoda melanoleuca) is a threatened species endemic to China. Alopecia, characterized by thinning and broken hair, mostly occurs in breeding males. Alopecia significantly affects the health and public image of the giant panda and the cause of alopecia is unclear. Results: Here, we researched gene expression profiles of four alopecia giant pandas and seven healthy giant pandas. All pandas were approximately ten years old and their blood samples collected during the breeding season. A total of 458 up-regulated DEGs and 211 down-regulated DEGs were identified. KEGG pathway enrichment identified that upregulated genes were enriched in the Notch signaling pathway and downregulated genes were enriched in ribosome, oxidative phosphorylation, and thermogenesis pathways. We obtained 28 hair growth-related DEGs, and identified three hub genes NOTCH1, SMAD3, and TGFB1 in PPI analysis. Five hair growth-related signaling pathways were identified with abnormal expression, these were Notch, Wnt, TGF-β, Mapk, and PI3K-Akt. The overexpression of NOTCH1 delays inner root sheath differentiation and results in hair shaft abnormalities. The delayed hair regression was associated with a significant decrease in the expression levels of TGFB1. Conclusions: Our data confirmed the abnormal expression of several hair-related genes and pathways and identified alopecia candidate genes in the giant panda. Results of this study provide theoretical basis for the establishment of prevention and treatment strategies for giant pandas with alopecia. [ABSTRACT FROM AUTHOR]

Short Tandem repeats (STRs) often occur within coding regions and adaptive selection could play a vital role in shaping the landscape of coding STRs. Here, we identified 849, 1282 and 1501 genes that contained 966, 1565 and 1921 STRs in the coding regions of the giant panda, polar bear and brown bear genomes, respectively. The results showed that coding STRs were subject to strong selection on STR type, motif, repetition and mode of evolution. Coding STRs were primarily found in regulatory genes. Of the three ursids studied, we found 585 differential genes in the giant panda. Gene Ontology analysis showed that the significant enrichment term (insulin-like growth factor receptor signaling pathway) exerted direct carbohydrate metabolic effects in vivo in this species. The enrichment of this pathway suggested that the giant panda's ability to absorb carbohydrates (starch) and adapt to a bamboo diet might be enhanced by variable coding STRs. We also identified 377 conserved coding STRs located in 377 genes across the three species. Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed that these genes were significantly enriched in two pathway involved in key physiological processes, including cardiovascular function and energy metabolism regulation. This study provides an important resource for future studies on the regulation of rapid diet and environmental adaptation of species by coding STRs. [ABSTRACT FROM AUTHOR]

We investigated the temporal dynamics of population genetics of the Chinese longsnout catfish (Leiocassis longirostris) given wild population decline due to anthropogenic disturbances such as overfishing in the upper Yangtze River. We compared genetic variation and structure between five age classes (1, 2, 3, 4 and 5 years old) using the mitochondrial DNA (mtDNA) control region and 12 microsatellite markers. The five age classes had high haplotype diversity (h = 0.8743) and nucleotide diversity (π = 0.0082) inferred from the control region, and observed heterozygosity (HO = 0.7342) and polymorphism information content (PIC = 0.6764) inferred from microsatellites. Low genetic distances based on the control region and the median-joining network representing a widespread distribution of haplotypes identified close genetic relationships between age classes. The results of AMOVA and pairwise fixation index (FST) using microsatellites demonstrated the significant genetic differentiation between age classes and FST was mostly low but significantly different between age classes (P < 0.05). Overall, L. longirostris in the upper Yangtze River has maintained considerable genetic diversity, but temporal genetic instability was detected between age classes. With the implementation of a ten-year fishing ban in the Yangtze River from 2021, it is expected that genetic differentiation between age classes will gradually decrease with the recovery of fish resources, and future genetic monitoring of L. longirostris populations is recommended. [ABSTRACT FROM AUTHOR]

Both the giant panda (Ailuropoda melanoleuca) and red panda (Ailurus fulgens) belong to the order Carnivora, but have changed their dietary habits to eating bamboo exclusively. The convergent evolution characteristics of their morphology, genome and gut flora have been found in the two pandas. However, the research on the convergent adaptation of their digestion and metabolism to the bamboo diet, mediated by the dietary shift of the two pandas at the gene-expression and epigenetic regulation levels, is still lacking. We therefore used RNA sequencing among five species (two pandas and three non-herbivore mammals) and bisulfite sequencing among three species (two pandas and a carnivore ferret) to sequence key digestion and metabolism tissues (stomach and small intestine). Our results provide evidence that the convergent differentially expressed genes (related to carbohydrate utilization, bile secretion, Lys and Arg metabolism, vitamin B12 utilization and cyanide detoxification) of the two pandas are adaptive responses to the bamboo diet containing low lipids, low Lys and Arg, low vitamin B12 and high cyanide. We also profiled the genome-wide methylome maps of giant panda, red panda and ferret, and the results indicated that the promoter methylation of the two pandas may regulate digestive and metabolic genes to adapt to sudden environmental changes, and then, transmit genetic information to future generations to evolve into bamboo eaters. Taken together, our study provides new insights into the molecular mechanisms of the dietary shift and the adaptation to a strict bamboo diet in both pandas using comparative transcriptomics and methylomics. [ABSTRACT FROM AUTHOR]

Trueperella pyogenes (T. pyogenes) is a crucial opportunistic pathogen normally causing mastitis, abscesses and pneumonia in economically important ruminants. Although only one commercial vaccine of T. pyogenes is currently obtainable, its immunoprotective effect is limited. Pyolysin (PLO) is the most predominant virulence factor highly expressed in T. pyogenes and is an excellent target for the development of novel vaccines against T. pyogenes. In this study, we designed a heterologous prime-boost vaccination scheme combining a DNA vaccine pVAX1-PLO and a subunit vaccine His-PLO to maximize host responses in mice. Humoral and cellular immune responses and protective effects were evaluated in mice to compare the immunogenicity induced by different immunization schemes. Compared to the PBS-control group, in vivo immunization results showed that better immune responses of mice immunized with the pVAX1-PLO plasmids and His-PLO proteins were induced. The residual bacterial burdens from the liver and peritoneal fluid were remarkably decreased in the immunized mice compared with the PBS group. Notably, the heterologous prime-boost vaccination groups significantly enhanced host humoral and cellular immune responses and protected mice from different virulent T. pyogenes strains infection. Conclusively, this study provides a favorable strategy for the further development of next-generation vaccines against T. pyogenes infections. [ABSTRACT FROM AUTHOR]

The complete mitochondrial genome of Episymploce splendens, 15,802 bp in length, was determined and annotated in this study. The mito-genome included 13 PCGs, 20 tRNAs and 2 rRNAs. Unlike most typical mito-genomes with conservative gene arrangement and exceptional economic organization, E. splendens mito-genome has two tRNAs (tRNA-Gln and tRNA-Met) absence and a long intergenic spacer sequence (93 bp) between tRNA-Val and srRNA, showing the diversified features of insect mito-genomes. This is the first report of the tRNAs deletion in blattarian mito-genomes and we supported the duplication/random loss model as the origin mechanism of the long intergenic spacer. Two Numts, Numt-1 (557 bp) and Numt-2 (975 bp) transferred to the nucleus at about 14.15 Ma to 22.34 Ma, and 19.19 Ma to 24.06 Ma respectively, were found in E. splendens. They can be used as molecular fossils in insect phylogenetic relationship inference. Our study provided useful data for further studies on the evolution of insect mito-genome. [ABSTRACT FROM AUTHOR]

Norovirus is a major cause of acute gastroenteritis worldwide, and no vaccine is currently available. The genetic and antigenic diversity of Norovirus presents challenges for providing broad immune protection, which calls for a multivalent vaccine application. In this study, we investigated the possibility of developing a virus-like particle (VLP)-based 6-valent Norovirus vaccine candidate (Hexa-VLPs) that covers GI.1, GII.2, GII.3, GII.4, GII.6, and GII.17 genotypes. Hexa-VLPs (30 µg) adjuvanted with 500 µg of aluminum hydroxide (alum) were selected as the optimal immunization dose after a dose-escalation study. Potent and long-lasting blockade antibody responses were induced by 2-or 3-shot Hexa-VLPs, especially for the emerging GII.P16-GII.2 and GII.17 (Kawasaki 2014) genotypes. Hexa-VLPs plus alum elicited Th1/Th2 mixed yet Th2-skewed immune responses, characterized by an IgG1-biased subclass profile and significant IL-4+ T-cell activation. Notably, simultaneous immunization with a mixture of six VLPs revealed no immunological interference among the component antigens. These results demonstrate that Hexa-VLPs are promising broad-spectrum vaccines to provide immunoprotection against major GI/GII epidemic strains in the future. [ABSTRACT FROM AUTHOR]

Background: Trueperella pyogenes and Pseudomonas aeruginosa are two important bacterial pathogens closely relating to the occurrence and development of forest musk deer respiratory purulent disease. Although T. pyogenes is the causative agent of the disease, the subsequently invaded P. aeruginosa will predominate the infection by producing a substantial amount of quorum-sensing (QS)-controlled virulence factors, and co-infection of them usually creates serious difficulties for veterinary treatment. In order to find a potential compound that targets both T. pyogenes and P. aeruginosa, the antibacterial and anti-virulence capacities of 55 compounds, which have similar core structure to the signal molecules of P. aeruginosa QS system, were tested in this study by performing a series of in vitro screening experiments. Results: We identified that furazolidone could significantly reduce the cell densities of T. pyogenes in mono-culture or in the co-culture with P. aeruginosa. Although the growth of P. aeruginosa could also be moderately inhibited by furazolidone, the results of phenotypic identification and transcriptomic analysis further revealed that sub-inhibitory furazolidone had remarkable inhibitory effect on the biofilm production, motility, and QS system of P. aeruginosa. Moreover, furazolidone could efficiently protect Caenorhabditis elegans models from P. aeruginosa infection under both fast-killing and slow-killing conditions. Conclusions: This study reports the antibacterial and anti-virulence abilities of furazolidone on T. pyogenes and P. aeruginosa, and provides a promising strategy and molecular basis for the development of novel anti-infectious drugs to dealing with forest musk deer purulent disease, or other diseases caused by T. pyogenes and P. aeruginosa co-infection. [ABSTRACT FROM AUTHOR]

Edible insects are alternative sources of high protein. Southeast Asia, South America, Africa, and Europe have recorded edible insects and, especially in Europe and America, there are factories that produce insects as food. In some areas of China, there has also been the habit of eating insects since ancient times. Periplaneta americana is an insect with the homology of medicine and food. In recent years, its medicinal and nutritional functions have attracted extensive attention and research. Its adult powder has been certified as a health product. It not only contains a variety of proteins but also is rich in fatty acids. The composition and antioxidant function of adult powder extract was analyzed in this study. Using adult P. americana powder as the raw material, it was extracted with n‐Hexane, dichloromethane, and ethyl acetate, respectively. Gas chromatography–mass spectrometry (GC–MS) removed a total of 60 compounds, and many active components were extracted from P. americana powder for the first time. The parts and relative content of each extracted sample were obtained, and the total antioxidant capacity (T‐AOC) of each extracted piece was determined, among which the antioxidant activity of ethyl acetate was the highest. [ABSTRACT FROM AUTHOR]

The giant panda (Ailuropoda melanoleuca) is a global flagship species for biodiversity conservation. As the time for captive giant pandas to be released into the wild matures, wildness training is provided to allow adaptation to their natural environment. It is assumed that changes in the immune system would be integral in this adaptation from captive to wild, where many more pathogens would be encountered in their natural habitats. Therefore, this study aims to determine the expression changes of immune-related genes and their potential as immunoassay markers for adaptation monitoring in wildness training giant pandas, and then to understand the adaptation strategy of wildness training giant pandas to the wild environment, thereby improving the success rate of panda reintroduction. We obtained 300 differentially expressed genes (DEGs) by RNA-seq, with 239 up-regulated and 61 down-regulated DEGs in wildness training giant pandas compared to captive pandas. Functional enrichment analysis indicated that up-regulated DEGs were enriched in several immune-related terms and pathways. There were 21 immune-related DEGs, in which most of them were up-regulated in wildness training giant pandas, including several critical innate and cellular immune genes. IL1R2 was the most significantly up-regulated gene and is a signature of homeostasis within the immune system. In the protein–protein interaction (PPI) analysis, CXCL8, CXCL10, and CCL5 were identified as the hub immune genes. Our results suggested that wildness training giant pandas have stronger innate and cellular immunity than captive giant pandas, and we proposed that a gene set of CXCL8, CXCL10, CCL5, CD3D, NFKBIA, TBX21, IL12RB2, and IL1R2 may serve as potential immunoassay markers to monitor and assess the immune status of wildness training giant pandas. Our study offers the first insight into immune alterations of wildness training giant pandas, paving the way for monitoring and evaluating the immune status of giant pandas when reintroducing them into the wild. [ABSTRACT FROM AUTHOR]

Thymosin is a physiologically active polypeptide hormone which has many functions involved in promoting wound healing, promoting hair growth, and accelerating tissue and organ regeneration. American cockroach (Periplaneta americana) has three subtypes thymosin (THY1, THY2, THY3) which are different splices from the same thymosin gene. Real‐time PCR was used to detect and analyze the expression differences of THY1 and THY2 in different stages, sexes, tissues and after stimulation by bacteria. Thymosins were expressed in adult, eclosion, and ootheca, and their expression levels were significantly up‐regulated during the eclosion period, showing the thymosin might be involved in the eclosion process. The expression levels of thymosins in females were higher than males, and the differences in eclosion stages were more significant (P < 0.01), indicating that they were closely related to development and female reproduction. The expressions of THY1 and THY2 in hemolymph were significantly higher (P < 0.01) than that in other tissues. The up‐regulated expression of thymosin in two immune‐related tissues (Hemolymph and adipose) after Escherichia coli stimulation further confirmed its involvement in the immune response. Our research would provide a basis for screening of novel pesticides target genes. [ABSTRACT FROM AUTHOR]

FASTA and FASTQ are the most widely used biological data formats that have become the de facto standard to exchange sequence data between bioinformatics tools. With the avalanche of next-generation sequencing data, the amount of sequence data being deposited and accessed in FASTA/Q formats is increasing dramatically. However, the existing tools have very low efficiency at random retrieval of subsequences due to the requirement of loading the entire index into memory. In addition, most existing tools have no capability to build index for large FASTA/Q files because of the limited memory. Furthermore, the tools do not provide support to randomly accessing sequences from FASTA/Q files compressed by gzip, which is extensively adopted by most public databases to compress data for saving storage. In this study, we developed pyfastx as a versatile Python package with commonly used command-line tools to overcome the above limitations. Compared to other tools, pyfastx yielded the highest performance in terms of building index and random access to sequences, particularly when dealing with large FASTA/Q files with hundreds of millions of sequences. A key advantage of pyfastx over other tools is that it offers an efficient way to randomly extract subsequences directly from gzip compressed FASTA/Q files without needing to uncompress beforehand. Pyfastx can easily be installed from PyPI (https://pypi.org/project/pyfastx) and the source code is freely available at https://github.com/lmdu/pyfastx. [ABSTRACT FROM AUTHOR]

Canine distemper (CD) is a significant threat to wild and captive giant panda populations. Captive giant pandas are inoculated with canine distemper virus (CDV) vaccination to prevent the infection with the CDV. As an important regulator, microRNA (miRNA) plays a crucial role in regulating gene expression, including in disease immunity. To understand the role of miRNA in immune response to CDV vaccination, we investigated the miRNA expression profile in five giant panda cubs after two inoculations, 21 days apart. A total of 187 conserved miRNAs and 96 novel miRNAs were identified. Among the 187 conserved miRNAs, 29 differentially expressed miRNAs were found postinoculation. The upregulation of miR-16, miR-182, miR-30b, and miR-101 indicated that the innate immune may be enhanced, whereas the upregulation of miR-142 and miR-19a are probably involved in the enhanced cellular immune response. However, the downregulated miR-155 and miR-181a might indicate the giant panda has weak ability to produce antibodies and memory B cells. Integrated analysis of miRNA-messenger RNA (mRNA) found 20 negatively regulated miRNA-mRNA pairs, where downregulated miR-204 might enhance giant panda cub innate immunity by increasing TLR6 expression, and downregulated miR-330 might activate macrophages and regulate the immune response by increasing TMEM106A expression. Our research provides key information for future development to enhance the immune response of giant pandas and potentially improve the survival of captive and wild giant panda populations threatened by CD. [ABSTRACT FROM AUTHOR]

Background: The giant panda (Ailuropoda melanoleuca) is a threatened endemic Chinese species and a flagship species of national and global conservation concern. Life history theory proposes that reproduction and immunity can be mutually constraining and interrelated. Knowledge of immunity changes of male giant pandas during the breeding season is limited. Results: Here, we researched peripheral blood gene expression profiles associated with immunity. Thirteen captive giant pandas, ranging from 9 to 11 years old, were divided into two groups based on their reproductive status. We identified 318 up-regulated DEGs and 43 down-regulated DEGs, which were enriched in 87 GO terms and 6 KEGG pathways. Additionally, we obtained 45 immune-related genes with altered expression, mostly up-regulated, and identified four hub genes HSPA4, SUGT1, SOD1, and IL1B in PPI analysis. These 45 genes were related to pattern recognition receptors, autophagy, peroxisome, proteasome, natural killer cell, antigen processing and presentation. SUGT1 and IL1B were related to pattern recognition receptors. HSP90AA1 was the most up-regulated gene and is a member of heat shock protein 90 family. HSP90 contributes to the translocation of extracellular antigen. KLRD1 encodes CD94, whose complex is an inhibitor of the cytotoxic activity of NK cells, was down-regulated. IGIP, which has the capability of inducing IgA production by B cells, was down-regulated, suggesting low concentration of IgA in male giant pandas. Our results suggest that most immune-related genes were up-regulated and more related to innate immune than adaptive immune. Conclusions: Our results indicated that breeding male giant pandas presented an immunoenhancement in innate immunity, enhanced antigen presentation and processing in cellular immunity compared to non-breeding males. The humoral immunity of male giant pandas may show a tendency to decrease during the breeding season. This study will provide a foundation for further studies of immunity and reproduction in male giant pandas. [ABSTRACT FROM AUTHOR]

Mitophagy has recently been implicated in bacterial infection but the underlying mechanism remains largely unknown. Here, we uncover a role of microRNA-302/367 cluster in regulating mitophagy and its associated host response against bacterial infection. We demonstrate that miR-302/367 cluster expression was significantly increased after Pseudomonas aeruginosa infection. Enhanced expression of miR-302/367 cluster accelerated the mitophagic response in macrophages, thus increasing clearance of invading P. aeruginosa by regulating production of reactive oxygen species (ROS), while application of miR-302/367 cluster inhibitors decreased bacterial clearance. Blocking mitophagy with siRNA against mitophagy receptor prohibitin 2 (PHB2) reduced the effect of miR-302/367 cluster on induction of mitophagy and its-associated P. aeruginosa elimination. In addition, we found that miR-302/367 cluster also increased bacterial clearance in mouse model. Mechanistically, we illustrate that miR-302/367 cluster binds to the 3′-untranslated region of nuclear factor kappa B (NF-κB), a negative regulator of mitophagy, accelerated the process of mitophagy by inhibiting NF-κB. Furthermore, inhibition of NF-κB in macrophages attenuated the ROS or cytokines production and may reduce cell injury by P. aeruginosa infection to maintain cellular homeostasis. Collectively, our findings elucidate that miR-302/367 cluster functions as potent regulators in mitophagy-mediated P. aeruginosa elimination and pinpoint an unexpected functional link between miRNAs and mitophagy. [ABSTRACT FROM AUTHOR]

Diet analysis is essential to fully understanding the biology of a species and its function within the ecosystem, and is also key in identifying food web interactions and the population dynamics of predators and prey. There is currently little information on the diet of small to mid-sized carnivores due to their elusive behavior. We used a DNA-based method to analyze the vertebrate prey of the Pallas's cat (Otocolobus manul) in the Gongga Mountain Nature Reserve of southwestern China, a global biodiversity hotspot region. We confirmed 14 scat samples containing the Pallas's cat DNA from 100 small carnivore scats collected at altitudes between 3200 and 4900 m, and then performed polymerase chain reaction (PCR) amplifications using primers targeting the vertebrate mitochondrial 12S rRNA gene with a blocking oligonucleotide specific to the predator sequence. We identified a total of 18 prey taxa, including nine mammals, eight birds and one fish. Our results show that pikas (in 79% of the scats) and birds (in 50% of the scats) are the predominant prey consumed by the Pallas's cat in our investigation area. The most important finding in our study is that Cypriniformes species were found in the Pallas's cat's diet, exhibiting dietary diversification consistent with the accessible fauna of the surrounding environment. In order to increase the detectability of both typical and rare vertebrate prey items, we constructed a blocking primer to inhibit the amplification of the Pallas's cat DNA. [ABSTRACT FROM AUTHOR]

The genus Schizothorax is one of the most diverse groups of schizothoracine fish. Many species within this genus possess highly similar morphological characters and are very difficult to be identified accurately only based on morphology. The present study aims to test the effectiveness of mitochondrial cytochrome c oxidase subunit I (COI) gene and cytochrome b (Cytb) gene for discriminating the Schizothorax fish. A total of 185 individuals of 11 species for COI gene and 264 individuals of 23 species for Cytb gene were used for analyzing, respectively. According to the genetic distances, only one species based on COI gene and five species based on Cytb gene had "barcoding gaps," respectively. The tree-based analysis displayed that four species based on COI gene and six species based on Cytb gene clustered monophyletic group with strong support, respectively. The optimal threshold value of Schizothorax is 0.005 based on COI gene and 0.008 based on Cytb gene. The results of genetic similarity tests performed through online BLAST showed that 108 of 185 similarity searches succeeded in identifying conspecific sequences based on COI gene and 199 of 264 succeeded in identifying conspecific sequences based on Cytb gene. Considering greater interspecific genetic distance in Kimura 2-parameter (K2P) analysis and many clades with higher supporting values in tree-based analysis, we suggest that Cytb gene has better resolution in discrimination of Schizothorax species than COI gene. However, there are still many confused clustering relationships based on molecular data currently available. Incomplete lineage sorting, the existence of possible cryptic species and problematic morphological identification, etc. might have greatly weakened the resolution of Cytb gene in discrimination of Schizothorax species. [ABSTRACT FROM AUTHOR]

The family Bagridae is a collection of species widely distributed in Africa and Asia with a high diversity of morphology. The species identification and phylogenetic relationship in this family have been confused and controversial. In order to explore the effectiveness of DNA barcoding in species identification of Bagridae, sequences of mitochondrial cytochrome c oxidase I (COI) gene of 20 species in four genera of Bagridae were used to analyse barcoding gap and to reconstruct phylogenetic relationship. Both the barcoding gap and the phylogenetic tree analysis showed that the COI gene-based DNA barcoding is an effective molecular technique for most species recognition of Chinese Bagridae. However, the rapid speciation and incomplete lineage sorting may affect the accuracy of DNA barcoding in species identification in certain species, and adding additional genes, such as nuclear gene, may help to achieve accurate identification of these species. The phylogenetic tree showed that the monophyly of genera Pelteobagrus, Leiocassis and Pseudobagrus did not exist, which supports that the species of genera Pelteobagrus, Pseudobagrus and Leiocassis distributed in China should be revised into one genus. [ABSTRACT FROM AUTHOR]

The Indian Cuckoo, Cuculus micropterus, belongs to the family Cuculidae. In this paper, we sequenced and analysized the complete mitochondrial genome of C. micropterus. The complete mitochondrial genome of C. micropterus is 17,541 bp in length, which was submitted to the NCBI database under the accession number MZ048030. It contains 13 protein-coding genes, 22 transfer RNA genes, two ribosome RNA genes, and one non-coding control region. The overall base composition of the mitochondrial DNA is 33.2% for A, 24.2% for T, 29.8% for C, and 12.8% for G, with a GC content of 42.6%. In order to explore the molecular phylogenetics evolution of Cuculidae, the nucleotide sequence data of 13 PCGs of C. micropterus and other 11 Cuculiformes were used for the phylogenetic analysis. The result shows that C. micropterus is closely related to Cuculus canorus bakeri. The study contributes to illuminating the taxonomic status of C. micropterus, and may facilitate further investigation of the evolution of Cuculidae. [ABSTRACT FROM AUTHOR]

Macaques are the most widely used non-human primates in biomedical research. The genetic divergence between these animal models is responsible for their phenotypic differences in response to certain diseases. However, the macaque single nucleotide polymorphism resources mainly focused on rhesus macaque (Macaca mulatta), which hinders the broad research and biomedical application of other macaques. In order to overcome these limitations, we constructed a database named MACSNVdb that focuses on the interspecies genetic diversity among macaque genomes. MACSNVdb is a web-enabled database comprising ~74.51 million high-quality non-redundant single nucleotide variants (SNVs) identified among 20 macaque individuals from six species groups (muttla , fascicularis , sinica , arctoides , silenus , sylvanus). In addition to individual SNVs, MACSNVdb also allows users to browse and retrieve groups of user-defined SNVs. In particular, users can retrieve non-synonymous SNVs that may have deleterious effects on protein structure or function within macaque orthologs of human disease and drug-target genes. Besides position, alleles and flanking sequences, MACSNVdb integrated additional genomic information including SNV annotations and gene functional annotations. MACSNVdb will facilitate biomedical researchers to discover molecular mechanisms of diverse responses to diseases as well as primatologist to perform population genetic studies. We will continue updating MACSNVdb with newly available sequencing data and annotation to keep the resource up to date. Database URL : http://big.cdu.edu.cn/macsnvdb/ [ABSTRACT FROM AUTHOR]

Microsatellites are widely distributed throughout nearly all genomes which have been extensively exploited as powerful genetic markers for diverse applications due to their high polymorphisms. Their length variations are involved in gene regulation and implicated in numerous genetic diseases even in cancers. Although much effort has been devoted in microsatellite database construction, the existing microsatellite databases still had some drawbacks, such as limited number of species, unfriendly export format, missing marker development, lack of compound microsatellites and absence of gene annotation, which seriously restricted researchers to perform downstream analysis. In order to overcome the above limitations, we developed PSMD (Pan‐Species Microsatellite Database, http://big.cdu.edu.cn/psmd/) as a web‐based database to facilitate researchers to easily identify microsatellites, exploit reliable molecular markers and compare microsatellite distribution pattern on genome‐wide scale. In current release, PSMD comprises 678,106,741 perfect microsatellites and 43,848,943 compound microsatellites from 18,408 organisms, which covered almost all species with available genomic data. In addition to interactive browse interface, PSMD also offers a flexible filter function for users to quickly gain desired microsatellites from large data sets. PSMD allows users to export GFF3 formatted file and CSV formatted statistical file for downstream analysis. We also implemented an online tool for analysing occurrence of microsatellites with user‐defined parameters. Furthermore, Primer3 was embedded to help users to design high‐quality primers with customizable settings. To our knowledge, PSMD is the most extensive resource which is likely to be adopted by scientists engaged in biological, medical, environmental and agricultural research. [ABSTRACT FROM AUTHOR]

The American cockroach (Periplaneta americana) is a globally invasive pest that can cause significant economic loss and threaten human health. Although it is abundant and lives in close proximity to humans, few studies have investigated the genetic diversity of P. americana. Our study analyzed 1,053 P. americana and other Periplaneta species' samples from different locations in China and the United States. A traditional tree‐based method using 17 unique mitochondrial COI haplotypes of P. americana and 20 haplotypes of the other Periplaneta species accurately identified P. americana with a barcoding threshold of 5.1%. To identify the population genetic structure of P. americana, we investigated wingless gene and pooled them with obtained mtDNA data for a combined analysis. Although the genetic diversity of the USA group was relatively higher than the China group, the number of haplotypes and alleles of both groups was small. The analysis of molecular variance (AMOVA), intraspecific phylogeny, and haplotype networks indicated that P. americana had very little global genetic differentiation. The weak geographic genetic structure might reflect the human‐mediated dispersal of P. americana. Despite no apparent phylogeographic assignment of mtDNA and nuclear lineages was observed in both BI trees, the integrated COI sequence data identified four distinct P. americana haplotype groups, showing four ancient maternal lineages of P. americana in China and the United States. [ABSTRACT FROM AUTHOR]

The globally distributed American cockroach (Periplaneta americana) is considered a pest, but it has been widely used in traditional Chinese medicine. In the past, the American cockroach's genome and transcriptomes were sequenced, but the differential expression transcripts between developmental stages were unavailable. We performed the de novo assembly and analysis of American cockroach transcriptomes from four developmental stages. Approximately 200 million high-quality paired-end reads were generated by using Illumina Hiseq 2000 sequencer. The assembly produced 291,250 transcripts with an average length of 714 bp. In addition, 38,052 microsatellites and 11,060,020 transposable elements were identified. Based on sequence homology, 53,262 transcripts were annotated. After calculating the expression levels of all the transcripts, we found that 13 transcripts were highly expressed in all the samples and at least two, p10 and actin-related protein 1, played important roles during development. A total of 7954 differentially expressed transcripts (DETs) were identified. The adult had the largest number of DETs when compared to other samples (4818), while the 3rd and 8th larva had the least number of DETs (1332). We performed gene enrichment analysis with the DETs, and some interesting results were detected in the different groups. For example, chitin is the major component of the insect exoskeleton, and the chitin-related genes in larvae and new molted samples had higher expression levels than in adults. In addition, the enrichment analysis detected many chitin-related pathways. Our study performed the first large-scale comparative transcriptomics between the developmental stages of American cockroach, which could provide useful gene expression data for future studies. [ABSTRACT FROM AUTHOR]

A total of 45 tetranucleotide chromosome-specific microsatellite markers with polymorphism were developed successfully based on three reference rhesus monkey genomes and on In-silico PCR prescreening. The polymorphic information content (PIC) values of 45 polymorphic microsatellite loci ranged from 0.487 to 0.879, with an average of 0.715, which were proven to be moderate to highly polymorphic. We detected 315 alleles on 45 microsatellite loci in 24 Rhesus monkeys. The number of alleles ranged from 3 to 15 and the mean number of alleles was 7 for each locus. Accordingly, the observed and expected heterozygosities obtained were between 0.417 and 1.0 and between 0.550 and 0.908, with an average value of 0.736 and 0.767, respectively. Genetic information demonstrated that 10 loci significantly deviated from Hardy–Weinberg equilibrium (P < 0.05). All 45 primers were not significant with regard to linkage disequilibrium (P > 0.001). Pearson correlation indicated that the PIC value exhibited a significant negative correlation with the loci number (r = − 0.741, P = 0.022), whereas the positive correlation with the number of the samples (r = 0.847, P = 0.070) was not significant. This may be attributed to the presence of random particularities within the loci. The T test of the sample groups indicated that the PIC difference was not significant when the number of samples was set at 10 and/or ≥ 15 (P = 0.7472 ~ 0.8564). These polymorphic and valuable microsatellite loci will facilitate further conservation genetics studies for rhesus monkeys and can be further applied to develop novel genetic markers for other species. [ABSTRACT FROM AUTHOR]

Repeat number variations of coding simple sequence repeats (coding SSRs) generate phenotype diversity for environmental adaptation and rapid adaptive evolution. Despite the positive potential of coding SSR mutations, the repeat variations have also been shown to cause hereditary disorders and severe diseases. However, there have been few reports on the analysis of SSR polymorphism in genome-wide coding regions. We identified 1016 coding SSRs, of which 283 SSRs (27.9%) were polymorphic with 2 to 8 alleles, using 33 re-sequenced giant panda genomes. The 283 polymorphic coding SSRs were mapped on 260 genes, which were assigned into 547 GO terms. Functional analysis of 260 genes showed that four significantly enriched pathways: thyroid hormone signaling pathway, pancreatic secretion, gastric acid secretion, and thyroid hormone synthesis, were relevant to digestion and metabolism. The comparison analysis of gene functions exhibited positive regulation of transcription, DNA-templated term (GO:0045893), the poly(A) RNA-binding term (GO:0008270), and zinc ion–binding term (GO:0044822) which were only significantly enriched in genes containing polymorphic coding SSRs. This indicates coding SSR polymorphism may be involved in binding and regulation functions that can affect the expression of their downstream genes and contribute to diverse phenotypes. Our results provide a solid foundation for further study regarding the effect of coding SSR polymorphism on the adaptation, adaptive evolution, and abnormal traits of the giant panda. [ABSTRACT FROM AUTHOR]

The Chinese forest musk deer (Moschus berezovskii ; FMD) is an artiodactyl mammal and is both economically valuable and highly endangered. To investigate the genetic mechanisms of musk secretion and adaptive immunity in FMD, we compared its genome to nine other artiodactyl genomes. Comparative genomics demonstrated that eight positively selected genes (PSGs) in FMD were annotated in three KEGG pathways that were related to metabolic and synthetic activity of musk, similar to previous transcriptome studies. Functional enrichment analysis indicated that many PSGs were involved in the regulation of immune system processes, implying important reorganization of the immune system in FMD. FMD-specific missense mutations were found in two PSGs (MHC class II antigen DRA and ADA) that were classified as deleterious by PolyPhen-2, possibly contributing to immune adaptation to infectious diseases. Functional assessment showed that the FMD-specific mutation enhanced the ADA activity, which was likely to strengthen the immune defense against pathogenic invasion. Single nucleotide polymorphism-based inference showed the recent demographic trajectory for FMD. Our data and findings provide valuable genomic resources not only for studying the genetic mechanisms of musk secretion and adaptive immunity, but also for facilitating more effective management of the captive breeding programs for this endangered species. [ABSTRACT FROM AUTHOR]

Background: Trueperella pyogenes is a commensal and opportunistic pathogen that normally causes mastitis, liver abscesses and pneumonia of economically important livestock. To develop efficacious and potent vaccine against T. pyogenes, chimeric gene DNA vaccines were constructed and encapsulated in chitosan nanoparticles (pPCFN-CpG-CS-NPs). Results: The pPCFN-CpG-CS-NPs consists of the plo, cbpA, fimA, and nanH gene of T. pyogenes and CpG ODN1826. It was produced with good morphology, high stability, a mean diameter of 93.58 nm, and a zeta potential of + 5.27 mV. Additionally, chitosan encapsulation was confirmed to protect the DNA plasmid from DNase I digestion. The immunofluorescence assay indicated that the four-chimeric gene could synchronously express in HEK293T cells and maintain good bioactivity. Compared to the mice immunized with the control plasmid, in vivo immunization showed that mice immunized with the pPCFN-CpG-CS-NPs had better immune responses, and release of the plasmid DNA was prolonged. Importantly, immunization with pPCFN-CpG-CS-NPs could significantly protect mice from highly virulent T. pyogenes TP7 infection. Conclusions: This study indicates that chitosan-DNA nanoparticles are potent immunization candidates against T. pyogenes infection and provides strategies for the further development of novel vaccines encapsulated in chitosan nanoparticles. [ABSTRACT FROM AUTHOR]

The lineage of the Bar-backed Partridge () was investigated to determine the phylogenetic relationships within Arborophila as the species is centrally distributed within an area covered by the distributions of 22 South-east Asian hill partridge species. The complete mitochondrial genome (mitogenome) of A. brunneopectus was determined and compared with four other hill partridge species mitogenomes. NADH subunit genes are radical in hill partridge mitogenomes and contain the most potential positive selective sites around where variable sites are abundant. Together with 44 other mitogenomes of closely related species, we reconstructed highly resolved phylogenetic trees using maximum likelihood (ML) and Bayesian inference (BI) analyses and calculated the divergence and dispersal history of Arborophila using combined datasets composed of their 13-protein coding sequences. Arborophila is reportedly be the oldest group in Phasianidae whose ancestors probably originated in Asia. A. rufipectus shares a closer relationship with A. ardens and A. brunneopectus compared to A. gingica and A. rufogularis, and such relationships were supported and profiled by NADH dehydrogenase subunit 5 (ND5). The intragenus divergence of all five Arborophila species occurred in the Miocene (16.84~5.69 Mya) when there were periods of climate cooling. We propose that these cooling events in the Miocene forced hill partridges from higher to lower altitudes, which led to geographic isolation and speciation. We demonstrated that the apparently deleterious +1 frameshift mutation in NADH dehydrogenase subunit 3 (ND3) found in all Arborophila is an ancient trait that has been eliminated in some younger lineages, such as Passeriformes. It is unclear of the biological advantages of this elimination for the relevant taxa and this requires further investigation. [ABSTRACT FROM AUTHOR]

Trueperella pyogenes is a prevalent opportunistic bacterium that normally causes diverse suppurative lesions, endometritis and pneumonia in various economically important animals. Although the genomic information of this species has been announced, little is known about its functional profiles. In this study, by performing a comparative transcriptome analysis between the highly and moderately virulent T. pyogenes isolates, we found the expression of a LuxR-type DNA-binding response regulator, PloR, was significantly up-regulated in the highly virulent T. pyogenes. Protein crystal structure prediction and primary functional assessment suggested that, the quorum-sensing signal molecules of Gram-negative bacteria such as Pseudomonas aeruginosa and Escherichia coli could significantly inhibit the growth, biofilm production and hemolysis of T. pyogenes by binding to the upstream sensor histidine kinase, PloS. Therefore, the PloS/PlosR two-component regulatory system might dominate the virulence of T. pyogenes. Our findings provide a major advance in understanding the pathogenesis of T. pyogenes, and may shed new light on the development of novel therapeutic strategies to control T. pyogenes infection. [ABSTRACT FROM AUTHOR]

Complete mitochondrial genomes (mitogenomes) of two cockroach species, Periplaneta australasiae and Neostylopyga rhombifolia, 15,605 bp and 15,711 bp in length, respectively, were determined. As reported for other cockroach mitogenomes, the two mitogenomes possessed typical ancestral insect mitogenome gene composition and arrangement. Only several small intergenic spacers were found: one, which was common in all sequenced cockroach mitogenomes except for the genus Cryptocercus, was between tRNA-Ser (UCN) and ND1 and contained a 7bp highly conserved motif (WACTTAA). Three different types of short tandem repeats in the N. rhombifolia control region (CR) were observed. The homologous alignments of these tandem repeats with other six cockroach mitogenome CRs revealed a low similarity. Three conserved sequence blocks (CSB) were detected in both cockroach mitochondrial CRs. CSB1 was specific for blattinine mitogenomes and was highly conserved with 95% similarity, speculating that this block was a possible molecular synapomorphy for this subfamily. CSB3 located nearby downstream of CSB1 and has more variations within blattinine mitogenomes compared with CSB1. The CSB3 was capable of forming stable stem-loop structure with a small T-stretch in the loop portion. We assessed the influence of four datasets and two inference methods on topology within Orthopteroidea. All genes excluding the third codon positions of PCGs could generate more stable topology, and higher posterior probabilities than bootstrap values were presented at some branch nodes. The phylogenetic analysis with different datasets and analytical methods supported the monophyly of Dictyoptera and supported strongly the proposal that Isoptera should be classified as a family (Termitidae) of the Blattaria. Specifically, Shelfordella lateralis was inserted in the clade Periplaneta. Considering the K2P genetic distance, morphological characters, and the phylogenetic trees, we suggested that S. lateralis should be placed in the genus Periplaneta. [ABSTRACT FROM AUTHOR]

Chicken repeat 1 ( CR1), a member of non- LTR retroposon, is an important phylogenetic marker in avian systematics. In this study, we reported several characteristics of CR1 elements in a draft genome of Arborophila rufipectus (Sichuan partridge). According to the analyses of RepeatMasker, approximately 254 966 CR1 elements were identified in A. rufipectus, covering 6.7% of the genome. Subsequently, we selected eighteen novel CR1 elements by comparing the chicken genome, turkey genome and assembled A. rufipectus scaffolds. Here, a combined data set comprising of 22 CR1 loci, mitochondrial genomes and eight unlinked introns was analysed to infer the evolutionary relationships of twelve Phasianidae species. The applicability of CR1 sequences for inferring avian phylogeny relative to mt DNA and intron sequences was investigated as well. Our results elucidated the position of A. rufipectus in Phasianidae with robust supports that it presented a sister clade to Arborophila ardens/ Arborophila brunneopectus, and implied that genus Arborophila was in a basal phylogenetic position within Phasianidae and a phylogenetic affinity between Meleagris gallopavo and Pucrasia macrolopha. Therefore, this work not only resolved some of the confounding relationships among Phasianidae, but also suggested CR1 sequences could provide powerful complementary data for phylogeny reconstruction. [ABSTRACT FROM AUTHOR]

Microsatellites are found to be related with various diseases and widely used in population genetics as genetic markers. However, it remains a challenge to identify microsatellite from large genome and screen microsatellites for primer design from a huge result dataset. Here, we present Krait, a robust and flexible tool for fast investigation of microsatellites in DNA sequences. Krait is designed to identify all types of perfect or imperfect microsatellites on a whole genomic sequence, and is also applicable to identification of compound microsatellites. Primer3 was seamlessly integrated into Krait so that users can design primer for microsatellite amplification in an efficient way. Additionally, Krait can export microsatellite results in FASTA or GFF3 format for further analysis and generate statistical report as well as plotting. Availability and implementation: Krait is freely available at https://github.com/lmdu/krait under GPL2 License, implemented in C and Python, and supported on Windows, Linux andMac operating systems. [ABSTRACT FROM AUTHOR]

The giant panda (Ailuropoda melanoleuca) is one of the world’s most beloved endangered mammals. Although the draft genome of this species had been assembled, little was known about the composition of its microRNAs (miRNAs) or their functional profiles. Recent studies demonstrated that changes in the expression of miRNAs are associated with immunity. In this study, miRNAs were extracted from the blood of four healthy giant pandas and sequenced by Illumina next generation sequencing technology. As determined by miRNA screening, a total of 276 conserved miRNAs and 51 novel putative miRNAs candidates were detected. After differential expression analysis, we noticed that the expressions of 7 miRNAs were significantly up-regulated in young giant pandas compared with that of adults. Moreover, 2 miRNAs were up-regulated in female giant pandas and 1 in the male individuals. Target gene prediction suggested that the miRNAs of giant panda might be relevant to the expressions of 4,602 downstream genes. Subseuqently, the predicted target genes were conducted to KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis and we found that these genes were mainly involved in host immunity, including the Ras signaling pathway, the PI3K-Akt signaling pathway, and the MAPK signaling pathway. In conclusion, our results provide the first miRNA profiles of giant panda blood, and the predicted functional analyses may open an avenue for further study of giant panda immunity. [ABSTRACT FROM AUTHOR]

Both Dabry's sturgeon ( Acipenser dabryanus) and Chinese sturgeon ( A. sinensis) are rare fish and listed as first class protected animals in China. Nowadays, an artificial release program has been established as a main way to maintain and restore their wild populations. Unintentional interspecific hybridization of these two sturgeons might occur because of their close relationship. To ensure the purity of germplasm, Dabry's sturgeon and Chinese sturgeon must be identified before releasing. Owing to their highly similar morphological characteristics, it is difficult to discriminate the two sturgeon species based on morphology in their larval and juvenile stages. Moreover, it is almost impossible to discriminate their hybrids based on morphology. Therefore, we established a new molecular approach to discriminate Dabry's sturgeon and Chinese sturgeon based on mitochondrial DNA barcode and six nuclear markers (three nuclear genes (TYR, GHR, and RAG2) and three SNP markers (SNP21564, SNP28822, and SNP12974) in this study. These two sturgeons were explicitly distinguished from other sturgeon species using the DNA barcode and three nuclear genes. Further, the two species were accurately discriminated using the three SNP markers. We concluded through blind laboratory trials involving 95 individuals that our method is 100% reliable in distinguishing between the two species. [ABSTRACT FROM AUTHOR]

As an endangered animal group in China, musk deer (genus Moschus) have attracted the attention of deer biologists and wildlife conservationists. Clarifying the taxonomic status and distribution of musk deer species is important to determine the conservation status for each species and establish appropriate conservation strategies. There remains some uncertainty about the species determination of the musk deer in the Guandi Forest District of Shanxi Province, China. The musk deer in Shanxi would appear to represent an extension of the geographical distribution of either the Forest Musk Deer from the southwest or the Siberian Musk Deer from the northeast, or possibly both. The musk deer population in Shanxi Province provides an interesting and significant case to test the value of applying molecular methods to make a genetic species identification. In order to clarify the species status of the Shanxi musk deer, we sequenced 627 bp of the COI gene and ≈723 bp of the D-loop gene in 12 musk deer samples collected from the Guandi Forest District, and the two reference samples collected from Sichuan. Genetic analyses from the data suggest that all of the samples from the Guandi Forest District are M. berezovskii rather than M. moschiferus. It is most likely that the most previous studies had wrong species identification. And it is the first time we use DNA barcoding to prove that Shanxi is a new distribution of M. berezovskii. [ABSTRACT FROM AUTHOR]

The giant panda ( Ailuropoda melanoleuca) is one of the most famous flagship species for conservation, and its draft genome has recently been assembled. However, the transcriptome is not yet available. In this study, the blood transcriptomes of three pandas were characterized and about 160 million sequencing reads were generated using Illumina Hi Seq 2000 paired-end sequencing technology. The assembly yielded 92 598 transcripts with an average length of 1626 bp and N50 length of 2842 bp. Based on a sequence similarity search against nonredundant (nr) protein database, a total of 38 522 (41.6%) transcripts were annotated. Of these annotated transcripts, 25 142 and 8272 transcripts were assigned to gene ontology terms and clusters of orthologous group, respectively. A search against the Kyoto Encyclopedia of Genes and Genomes Pathway database ( KEGG) indicated that 9098 (9.83%) transcripts mapped to 324 KEGG pathways, and the best represented functional categories of pathways were signal transduction and immune system. We have also identified 23 460 microsatellites, 43 560 SNPs as well as 21 456 alternative splicing events in the assembly. Additionally, a total of 24 341 complete open reading frames ( ORFs) were detected from the assembly where 1492 ORFs were found to be novel gene loci as these have not been annotated so far in any public database. [ABSTRACT FROM AUTHOR]

Asiatic shrewlike moles are distributed almost entirely in south-west China; four of the five species of the genus Uropsilus, Uropsilus aequodonenia, U. andersoni, U. investigator and U. soricipes are endemic to China. Excluding the five species, three cryptic species ( U. sp. 1, U. sp. 2 and U. sp. 3) and two putative species, U. nivatus and U. atronates, are recognized. The phylogenetic relationships among the species remain unclear and these preclude investigations of their potential adaptations for living in high altitudes. We sequenced the complete mitochondrial DNA genomes of three species of Asiatic shrewlike moles ( U. aequodonenia, U. andersoni and U. nivatus). Phylogenetic analyses of 16 published and our de novo mitogenomes yield single, robust trees with the relationships being ( U. soricipes ( U. sp. 1 ( U. nivatus ( U. andersoni, U. aequodonenia)))). Further, the tree verifies the validity of recently described U. aequodonenia. Analyses of selection pressure suggest that the 13 mt DNA-encoding genes of species in the genus Uropsilus all have experienced strong purifying selection, although ATP8 accumulated a higher ratio of non-synonymous substitutions than the other loci, which might reflect adaptation of the genus Uropsilus to different environments/elevations. [ABSTRACT FROM AUTHOR]

The Tibetan Plateau underwent dramatic geological and climatic changes, which had important implications for genetic divergence and population dynamics of freshwater fish populations. Fluctuations of the ecogeographical environment and major hydrographic formations might have promoted the formation of new subspecies or species. In order to understand the impact of plateau uplift on freshwater fish evolutionary history, we estimated the genetic diversity and population structure in two subspecies of Schizopygopsis chengi ( S. c. chengi and S. c. baoxingensis) in upper Yangtze River in Tibetan Plateau area using mitochondrial DNA control region and eight microsatellite markers, which suggested that there was a close genetic relationship. S. chengi showed some significant genetic structure that did not correlate with geographic distance. Bayesian assignment tests indicated that S. chengi samples in the study could be divided into four populations: upstream population, midstream population, tributary population and S. c. baoxingensis population. S. c. chengi and S. c. baoxingensis showed significant genetic divergence. However, phylogenetic analysis, population structure analysis and historical gene flow estimation suggested that there was close genetic relationship between S. c. baoxingensis and the Dawei population which belongs to populations of S. c. chengi. The time that Dawei population suffered from a bottleneck and S. c. baoxingensis underwent population expansion was congruent with the last glacial period on the Tibetan Plateau. The results confirmed the hypothesis that the Dawei River and Baoxing River were once connected, and the Dawei and Baoxing populations originated from a single population, but were isolated into separate populations because of crustal movements and the Baoxing population evolved as S. c. baoxingensis. [ABSTRACT FROM AUTHOR]