Hidden hotspots of amphibian biodiversity in China.

Identifying and protecting hotspots of endemism and species richness is crucial for mitigating the global biodiversity crisis. However, our understanding of spatial diversity patterns is far from complete, which severely limits our ability to conserve biodiversity hotspots. Here, we report a comprehensive analysis of amphibian species diversity in China, one of the most species-rich countries on Earth. Our study combines 20 y of field surveys with new molecular analyses of 521 described species and also identifies 100 potential cryptic species. We identify 10 hotspots of amphibian diversity in China, each with exceptional species richness and endemism and with exceptional phylogenetic diversity and phylogenetic endemism (based on a new time-calibrated, species-level phylogeny for Chinese amphibians). These 10 hotspots encompass 59.6% of China's described amphibian species, 49.0% of cryptic species, and 55.6% of species endemic to China. Only four of these 10 hotspots correspond to previously recognized biodiversity hotspots. The six new hotspots include the Nanling Mountains and other mountain ranges in South China. Among the 186 species in the six new hotspots, only 9.7% are well covered by protected areas and most (88.2%) are exposed to high human impacts. Five of the six new hotspots are under very high human pressure and are in urgent need of protection. We also find that patterns of richness in cryptic species are significantly related to those in described species but are not identical.

Early results of the integrative epigenomic-transcriptomic landscape of colorectal adenoma and cancer.

BACKGROUND: Aberrant methylation is common during the initiation and progression of colorectal cancer (CRC), and detecting these changes that occur during early adenoma (ADE) formation and CRC progression has clinical value. AIM: To identify potential DNA methylation markers specific to ADE and CRC. METHODS: Here, we performed SeqCap targeted bisulfite sequencing and RNA-seq analysis of colorectal ADE and CRC samples to profile the epigenomic-transcriptomic landscape. RESULTS: Comparing 22 CRC and 25 ADE samples, global methylation was higher in the former, but both showed similar methylation patterns regarding differentially methylated gene positions, chromatin signatures, and repeated elements. High-grade CRC tended to exhibit elevated methylation levels in gene promoter regions compared to those in low-grade CRC. Combined with RNA-seq gene expression data, we identified 14 methylation-regulated differentially expressed genes, of which only AGTR1 and NECAB1 methylation had prognostic significance. CONCLUSION: Our results suggest that genome-wide alterations in DNA methylation occur during the early stages of CRC and demonstrate the methylation signatures associated with colorectal ADEs and CRC, suggesting prognostic biomarkers for CRC.

Ultrasonic-Assisted Surface Finishing of STAVAX Mold Steel Using Lab-Made Polishing Balls on a 5-Axis CNC Machining Center.

The inconvenience of conventional wool ball polishing is that the surface finishing process should be equipped with a slurry container. The main objective of this research is to develop an ultrasonic-assisted surface finishing process for STAVAX mold steel on a 5-axis CNC machining center, by using new lab-made rubber polishing balls containing the abrasive aluminum oxide instead of the traditional wool ball polishing. In total, five types (type A to type E) of new rubber-matrixed polishing balls with a composite of nitrile butadiene rubber (NBR), an abrasive of aluminum oxide, and an additive of silicon dioxide have been developed. The performance of the composites with different grain sizes (0.05 µm to 3 µm) and concentrations of the abrasive of aluminum oxide have been investigated. The effects of multiple polishing passes on the surface roughness improvement for the lab-made polishing balls have also been investigated in this study. A surface roughness of Ra 0.027 µm on average was achieved by using the multiple polishing process of E-C-B-A. The volumetric wear of the lab-made polishing balls, using ultrasonic vibration-assisted polishing, can be improved from about 12.64% (type A) to 65.48% (type E) compared with the non-vibration-assisted polishing. The suitable combination of the ultrasonic vibration-assisted polishing parameters were an amplitude of 10 µm, a frequency of 23 kHz, a spindle speed of 5000 rpm, a feed rate of 60 mm/min, a stepover of 20 µm, a penetration depth of 180 µm, and a polishing pass of E-C-B-A, based on the experimental results. The surface roughness improvement on a test carrier with a saddle surface has also been presented by using the ultrasonic vibration-assisted polishing with the lab-made polishing balls.

A single-nucleus transcriptomic atlas of the dog hippocampus reveals the potential relationship between specific cell types and domestication.

The process of domestication has led to dramatic differences in behavioral traits between domestic dogs and gray wolves. Whole-genome research found that a class of putative positively selected genes were related to various aspects of learning and memory, such as long-term potentiation and long-term depression. In this study, we constructed a single-nucleus transcriptomic atlas of the dog hippocampus to illustrate its cell types, cell lineage and molecular features. Using the transcriptomes of 105 057 nuclei from the hippocampus of a Beagle dog, we identified 26 cell clusters and a putative trajectory of oligodendrocyte development. Comparative analysis revealed a significant convergence between dog differentially expressed genes (DEGs) and putative positively selected genes (PSGs). Forty putative PSGs were DEGs in glutamatergic neurons, especially in Cluster 14, which is related to the regulation of nervous system development. In summary, this study provides a blueprint to understand the cellular mechanism of dog domestication.

Joint profiling of gene expression and chromatin accessibility during amphioxus development at single-cell resolution.

Vertebrate evolution was accompanied by two rounds of whole-genome duplication followed by functional divergence in terms of regulatory circuits and gene expression patterns. As a basal and slow-evolving chordate species, amphioxus is an ideal paradigm for exploring the origin and evolution of vertebrates. Single-cell sequencing has been widely used to construct the developmental cell atlas of several representative species of vertebrates (human, mouse, zebrafish, and frog) and tunicates (sea squirts). Here, we perform single-nucleus RNA sequencing (snRNA-seq) and single-cell assay for transposase accessible chromatin sequencing (scATAC-seq) for different stages of amphioxus (covering embryogenesis and adult tissues). With the datasets generated, we constructed a developmental tree for amphioxus cell fate commitment and lineage specification and characterize the underlying key regulators and genetic regulatory networks. The data are publicly available on the online platform AmphioxusAtlas.

Longitudinal transcriptome analyses show robust T cell immunity during recovery from COVID-19.

Understanding the processes of immune regulation in patients infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is crucial for improving treatment. Here, we performed longitudinal whole-transcriptome RNA sequencing on peripheral blood mononuclear cell (PBMC) samples from 18 patients with coronavirus disease 2019 (COVID-19) during their treatment, convalescence, and rehabilitation. After analyzing the regulatory networks of differentially expressed messenger RNAs (mRNAs), microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) between the different clinical stages, we found that humoral immunity and type I interferon response were significantly downregulated, while robust T-cell activation and differentiation at the whole transcriptome level constituted the main events that occurred during recovery from COVID-19. The formation of this T cell immune response might be driven by the activation of activating protein-1 (AP-1) related signaling pathway and was weakly affected by other clinical features. These findings uncovered the dynamic pattern of immune responses and indicated the key role of T cell immunity in the creation of immune protection against this disease.

Capture Hybridization of Long-Range DNA Fragments for High-Throughput Sequencing.

Capture hybridization coupled with high-throughput sequencing (HTS) has become one of the most popular approaches to address some scientific problems not only for fundamental evolution but also for ecology and human disease in recent years. However, the technical problem of limited probe capture ability affects its widespread application. Here, we propose to capture hybridize long-range DNA fragments for HTS (termed LR-LCH). We provide a case of three amphibian samples to examine LR-LCH with 2 kb libraries and comparison of standard capture hybridization with 480 bp libraries. Capture sensitivity increased from an average 13.57% of standard capture hybridization to an average 19.80% of LR-LCH; capture efficiency also increased from an average 72.56% of standard capture hybridization to an average 97.71% of LR-LCH. These indicate that longer fragments in the library generally contain both relatively variable regions and relatively conservative regions. The divergent parts of target DNA are enriched along with conservative parts of DNA sequence that effectively captured during hybridization. We present a protocol that allows users to overcome the low capture sensitivity problem for high divergent regions.

Identification of a novel mtDNA lineage B3 in chicken (Gallus gallus domesticus).

In this study, we sequenced the complete mitochondrial DNA genome (mitogenome) of the Zhengyang Yellow chicken (Gallus gallus domesticus) by next-generation sequencing technology. Samples were taken from Zhumadian city, Henan Province, China. The complete mitogenome was 16 785 bp in size, and had a nucleotide composition of 30.3% (A), 23.7% (T), 32.5% (C), and 13.5% (G), with a high AT content of 54.0%. The assembled mitogenome exhibited typical mitochondrial DNA (mtDNA) structure, including a non-coding control region, two rRNA genes, 13 protein-coding genes, and 22 tRNA genes. Phylogenetic analysis indicated that this mitogenome defined a novel sub-haplogroup B3 within haplogroup B. These results should provide essential information for chicken domestication and insight into the evolution of genomes.

Association of a common variant of SYNPO2 gene with increased risk of serous epithelial ovarian cancer.

In China, the majority of ovarian cancer patients (80%-90%) are women who are diagnosed with epithelial ovarian cancer. The SYNPO2 gene has recently been reported to be associated with epithelial ovarian cancer in Europeans. To investigate the association of common variants of SYNPO2 gene with epithelial ovarian cancer in Han Chinese individuals, we designed a case-control study with 719 epithelial ovarian cancer patients and 1568 unrelated healthy controls of Han Chinese descent. A total of 49 tagging single-nucleotide polymorphisms were genotyped; single-single-nucleotide polymorphism association, imputation, and haplotypic association analyses were performed. The single-nucleotide polymorphism rs17329882 was found to be strongly associated with serous epithelial ovarian cancer and with ages ≤49 years, consistent with the pre-menopausal status of analyzed epithelial ovarian cancer cases. Odds ratios and 95% confidence intervals provided evidence of the risk effects of the C allele of the single-nucleotide polymorphism on epithelial ovarian cancer. Imputation analyses also confirmed the results with a similar pattern. Additionally, haplotype analyses indicated that the haplotype block that contained rs17329882 was significantly associated with epithelial ovarian cancer risk, specifically with the serous epithelial ovarian cancer subtype. In conclusion, our results show that SYNPO2 gene plays an important role in the etiology of epithelial ovarian cancer, suggesting that this gene may be a potential genetic modifier for developing epithelial ovarian cancer.

Identification and validation of colorectal neoplasia-specific methylation biomarkers based on CTCF-binding sites.

To date, the sensitivity of currently available biomarkers based on the methylation of gene promoters is suboptimal for detecting adenomas and early-stage colorectal cancer (CRC). We aimed to develop biomarkers with methylated DNA binding sites of the multifunctional transcriptional factor CTCF for early detection of CRC. Using combined analyses of genome-wide occupation and the methylation profile of CTCF-binding sites, we identified candidate CTCF-binding sites. Then, we applied methylation-sensitive high-resolution melting (MS-HRM) and mass spectrometry analysis to screen and validate these candidate sites in diverse sample sets. We identified a set of colorectal neoplasia-specific biomarkers with robust performance. The top five biomarkers were selected and recommended for early detection of colorectal neoplasia. All of the five novel biomarkers exhibited a more robust discriminatory performance than that by BMP3 and NDRG4, two currently acknowledged robust methylation biomarkers. When the five new biomarkers were considered as a marker panel and tumor-positive was defined as having two or more (of the five) positive biomarkers, the marker panel could achieve a sensitivity of 91.67% for adenomas, 97.44% for Stage I CRC, 94.06% for Stage II CRC, 93.62% for Stage III CRC, and 93.54% for total colorectal tumors with a specificity of 94.05%. To our knowledge, this is the first study for colorectal neoplasia-specific methylation biomarkers based on CTCF-binding sites. Using a similar strategy, CTCF-binding sites could be potentially developed into biomarkers for other tumors. In summary, this study opens a new area in developing biomarkers for tumor prevention and treatment.

Genomic incompatibilities in the diploid and tetraploid offspring of the goldfish × common carp cross.

Polyploidy is much rarer in animals than in plants but it is not known why. The outcome of combining two genomes in vertebrates remains unpredictable, especially because polyploidization seldom shows positive effects and more often results in lethal consequences because viable gametes fail to form during meiosis. Fortunately, the goldfish (maternal) × common carp (paternal) hybrids have reproduced successfully up to generation 22, and this hybrid lineage permits an investigation into the genomics of hybridization and tetraploidization. The first two generations of these hybrids are diploids, and subsequent generations are tetraploids. Liver transcriptomes from four generations and their progenitors reveal chimeric genes (>9%) and mutations of orthologous genes. Characterizations of 18 randomly chosen genes from genomic DNA and cDNA confirm the chimera. Some of the chimeric and differentially expressed genes relate to mutagenesis, repair, and cancer-related pathways in 2nF1. Erroneous DNA excision between homologous parental genes may drive the high percentage of chimeric genes, or even more potential mechanisms may result in this phenomenon. Meanwhile, diploid offspring show paternal-biased expression, yet tetraploids show maternal-biased expression. These discoveries reveal that fast and unstable changes are mainly deleterious at the level of transcriptomes although some offspring still survive their genomic abnormalities. In addition, the synthetic effect of genome shock might have resulted in greatly reduced viability of 2nF2 hybrid offspring. The goldfish × common carp hybrids constitute an ideal system for unveiling the consequences of intergenomic interactions in hybrid vertebrate genomes and their fertility.

The Genome of Dendrobium officinale Illuminates the Biology of the Important Traditional Chinese Orchid Herb.

Dendrobium officinale Kimura et Migo is a traditional Chinese orchid herb that has both ornamental value and a broad range of therapeutic effects. Here, we report the first de novo assembled 1.35 Gb genome sequences for D. officinale by combining the second-generation Illumina Hiseq 2000 and third-generation PacBio sequencing technologies. We found that orchids have a complete inflorescence gene set and have some specific inflorescence genes. We observed gene expansion in gene families related to fungus symbiosis and drought resistance. We analyzed biosynthesis pathways of medicinal components of D. officinale and found extensive duplication of SPS and SuSy genes, which are related to polysaccharide generation, and that the pathway of D. officinale alkaloid synthesis could be extended to generate 16-epivellosimine. The D. officinale genome assembly demonstrates a new approach to deciphering large complex genomes and, as an important orchid species and a traditional Chinese medicine, the D. officinale genome will facilitate future research on the evolution of orchid plants, as well as the study of medicinal components and potential genetic breeding of the dendrobe.

Resequencing 302 wild and cultivated accessions identifies genes related to domestication and improvement in soybean.

Understanding soybean (Glycine max) domestication and improvement at a genetic level is important to inform future efforts to further improve a crop that provides the world's main source of oilseed. We detect 230 selective sweeps and 162 selected copy number variants by analysis of 302 resequenced wild, landrace and improved soybean accessions at >11× depth. A genome-wide association study using these new sequences reveals associations between 10 selected regions and 9 domestication or improvement traits, and identifies 13 previously uncharacterized loci for agronomic traits including oil content, plant height and pubescence form. Combined with previous quantitative trait loci (QTL) information, we find that, of the 230 selected regions, 96 correlate with reported oil QTLs and 21 contain fatty acid biosynthesis genes. Moreover, we observe that some traits and loci are associated with geographical regions, which shows that soybean populations are structured geographically. This study provides resources for genomics-enabled improvements in soybean breeding.

Genome-wide SNP and haplotype analyses reveal a rich history underlying dog domestication.

Advances in genome technology have facilitated a new understanding of the historical and genetic processes crucial to rapid phenotypic evolution under domestication. To understand the process of dog diversification better, we conducted an extensive genome-wide survey of more than 48,000 single nucleotide polymorphisms in dogs and their wild progenitor, the grey wolf. Here we show that dog breeds share a higher proportion of multi-locus haplotypes unique to grey wolves from the Middle East, indicating that they are a dominant source of genetic diversity for dogs rather than wolves from east Asia, as suggested by mitochondrial DNA sequence data. Furthermore, we find a surprising correspondence between genetic and phenotypic/functional breed groupings but there are exceptions that suggest phenotypic diversification depended in part on the repeated crossing of individuals with novel phenotypes. Our results show that Middle Eastern wolves were a critical source of genome diversity, although interbreeding with local wolf populations clearly occurred elsewhere in the early history of specific lineages. More recently, the evolution of modern dog breeds seems to have been an iterative process that drew on a limited genetic toolkit to create remarkable phenotypic diversity.

Population phylogenomic analysis of mitochondrial DNA in wild boars and domestic pigs revealed multiple domestication events in East Asia.

BACKGROUND: Previously reported evidence indicates that pigs were independently domesticated in multiple places throughout the world. However, a detailed picture of the origin and dispersal of domestic pigs in East Asia has not yet been reported. RESULTS: Population phylogenomic analysis was conducted in domestic pigs and wild boars by screening the haplogroup-specific mutation motifs inferred from a phylogenetic tree of pig complete mitochondrial DNA (mtDNA) sequences. All domestic pigs are clustered into single clade D (which contains subclades D1, D2, D3, and D4), with wild boars from East Asia being interspersed. Three haplogroups within D1 are dominant in the Mekong region (D1a2 and D1b) and the middle and downstream regions of the Yangtze River (D1a1a), and may represent independent founders of domestic pigs. None of the domestic pig samples from North East Asia, the Yellow River region, and the upstream region of the Yangtze River share the same haplogroup status with the local wild boars. The limited regional distributions of haplogroups D1 (including its subhaplogroups), D2, D3, and D4 in domestic pigs suggest at least two different in situ domestication events. CONCLUSION: The use of fine-grained mtDNA phylogenomic analysis of wild boars and domestic pigs is a powerful tool with which to discern the origin of domestic pigs. Our findings show that pig domestication in East Asia mainly occurred in the Mekong region and the middle and downstream regions of the Yangtze River.

Multiple maternal origins of chickens: out of the Asian jungles.

Domestic chickens have long been important to human societies for food, religion, entertainment, and decorative uses, yet the origins and phylogeography of chickens through Eurasia remain uncertain. Here, we assessed their origins and phylogeographic history by analyzing the mitochondrial DNA hypervariable segment I (HVS-I) for 834 domestic chickens (Gallus gallus domesticus) across Eurasia as well as 66 wild red jungle fowls (Gallus gallus) from Southeast Asia and China. Phylogenetic analyses revealed nine highly divergent mtDNA clades (A-I) in which seven clades contained both the red jungle fowls and domestic chickens. There was no breed-specific clade in the chickens. The clades A, B, and E are distributed ubiquitously in Eurasia, while the other clades were restricted to South and Southeast Asia. Clade C was mainly distributed in Japan and Southeast China, while clades F and G were exclusive to Yunnan, China. The geographic distribution of clade D was closely related to the distribution of the pastime of cock fighting. Statistical tests detect population expansion within each subclade. These distinct distribution patterns and expansion signatures suggest that different clades may originate from different regions, such as Yunnan, South and Southwest China and/or surrounding areas (i.e., Vietnam, Burma, and Thailand), and the Indian subcontinent, respectively, which support the theory of multiple origins in South and Southeast Asia.