The sperm storage capacity in hens was correlated with the morphological differences of the oviduct and uterus-vagina junction.

The fertilization rate is an important index to evaluate the reproductive capacity of hens, which is mainly affected by semen quality, timing of artificial insemination (AI), and the ability to store sperm. A high sperm storage (SS) capacity can extend the interval, reduce the frequency, and decrease the labor costs of AI. However, relatively few studies have investigated the SS capacity of hens. Therefore, the aims of the present study were to identify factors influencing the SS capacity of Guangxi partridge chickens and to explore the impact of the sperm count in different sections of the oviduct and sperm storage tubules (SSTs), in addition to the number and surface area of SSTs on SS capacity at different time points after AI. We found that SS capacity was positively correlated to the egg production rate (P < 0.01) and body length (P < 0.05). On post-AI days 5, 10, and 15, the sperm count was higher in uterus-vagina junction (UVJ) than the magnum, isthmus, and infundibulum (P < 0.01), but gradually decreased over time. Also, the duration of SS and the sperm count of the UVJ was greater in the high SS group than the low SS group (P < 0.05). Histopathological analysis of the UVJ showed that the number and surface area of the SSTs (P < 0.01), as well as the proportion of SSTs containing sperm, were greater in the high SS group at all time points post AI (P < 0.01), while the proportion of SSTs containing sperm gradually decreased over time. Collectively, these results highlight the potential for selective breeding of SS capacity and show that SS capacity is related to laying performance and body length of Guangxi partridge hens. In addition, SS capacity was positively correlated to the surface area of SSTs and the proportion containing sperm. A greater sperm count stored in the UVJ was correlated to more sperm transported to the infundibulum and subsequent greater SS capacity of hens.

Identification of genes associated with sperm storage capacity in hens at different times after insemination by RNA-seq and Ribo-seq.

BACKGROUND: Sperm storage capacity (SSC) determines the duration of fertility in hens and is an important reproduction trait that cannot be ignored in production. Currently, the genetic mechanism of SSC is still unclear in hens. Therefore, to explore the genetic basis of SSC, we analyzed the uterus-vagina junction (UVJ) of hens with different SSC at different times after insemination by RNA-seq and Ribo-seq. RESULTS: Our results showed that 589, 596, and 527 differentially expressed genes (DEGs), 730, 783, and 324 differentially translated genes (DTGs), and 804, 625, and 467 differential translation efficiency genes (DTEGs) were detected on the 5th, 10th, and 15th days after insemination, respectively. In transcription levels, we found that the differences of SSC at different times after insemination were mainly reflected in the transmission of information between cells, the composition of intercellular adhesion complexes, the regulation of ion channels, the regulation of cellular physiological activities, the composition of cells, and the composition of cell membranes. In translation efficiency (TE) levels, the differences of SSC were mainly related to the physiological and metabolic activities in the cell, the composition of the organelle membrane, the physiological activities of oxidation, cell components, and cell growth processes. According to pathway analysis, SSC was related to neuroactive ligand-receptor interaction, histidine metabolism, and PPAR signaling pathway at the transcriptional level and glutathione metabolism, oxidative phosphorylation, calcium signaling pathway, cell adhesion molecules, galactose metabolism, and Wnt signaling pathway at the TE level. We screened candidate genes affecting SSC at transcriptional levels (COL4A4, MUC6, MCHR2, TACR1, AVPR1A, COL1A1, HK2, RB1, VIPR2, HMGCS2) and TE levels(COL4A4, MUC6, CYCS, NDUFA13, CYTB, RRM2, CAMK4, HRH2, LCT, GCK, GALT). Among them, COL4A4 and MUC6 were the key candidate genes differing in transcription, translation, and translation efficiency. CONCLUSIONS: Our study used the combined analysis of RNA-seq and Ribo-seq for the first time to investigate the SSC and reveal the physiological processes associated with SSC. The key candidate genes affecting SSC were screened, and the theoretical basis was provided for the analysis of the molecular regulation mechanism of SSC.

Phylogenetic and taxonomic updates of Agaricales, with an emphasis on Tricholomopsis.

The order Agaricales was divided into eight suborders. However, the phylogenetic relationships among some suborders are largely unresolved, and the phylogenetic positions and delimitations of some taxa, such as Sarcomyxaceae and Tricholomopsis, remain unsettled. In this study, sequence data of 38 genomes were generated through genome skimming on an Illumina sequencing system. To anchor the systematic position of Sarcomyxaceae and Tricholomopsis, a phylogenetic analysis based on 555 single-copy orthologous genes from the aforementioned genomes and 126 publicly accessible genomes was performed. The results fully supported the clustering of Tricholomopsis with Phyllotopsis and Pleurocybella within Phyllotopsidaceae, which formed a divergent monophyletic major lineage together with Pterulaceae, Radulomycetaceae, and Macrotyphula in Agaricales. The analysis also revealed that Sarcomyxaceae formed a unique major clade. Therefore, two new suborders, Phyllotopsidineae and Sarcomyxineae, are proposed for the two major lineages. Analyses of 450 single-copy orthologous genes and four loci suggested that Tricholomopsis consisted of at least four clades. Tricholomopsis is subsequently subdivided into four distinct sections. Seventeen Tricholomopsis species in China, including six new species, are reported. Conoloma is established to accommodate T. mucronata. The substrate preference of Tricholomopsis species and the transitions of the pileate ornamentations among the species within the genus are discussed.

Molecular phylogeny and morphology of Sporodiniella sinensis sp. nov. (Syzygitaceae, Mucorales), an invertebrate-associated species from Yunnan, China.

During investigations of invertebrate-associated fungi in Yunnan Province of China, a new species, Sporodiniella sinensis sp. nov., was collected. Morphologically, S. sinensis is similar to Sporodiniella umbellata; however, it is distinguished from S. umbellata by its greater number of sporangiophore branches, longer sporangiophores, larger sporangiospores, and columellae. The novel species exhibits similarities of 91.62 % for internal transcribed spacer (ITS), 98.66-99.10 % for ribosomal small subunit (nrSSU), and 96.36-98.22 % for ribosomal large subunit (nrLSU) sequences, respectively, compared to S. umbellata. Furthermore, phylogenetic analyses based on combined sequences of ITS, nrLSU and nrSSU show that it forms a separate clade in Sporodiniella, and clusters closely with S. umbellata with high statistical support. The phylogenetic and morphological evidence support S. sinensis as a distinct species. Here, it is formally described and illustrated, and compared with other relatives.

Four New Species and One New Record of Thelephora from China.

Species of the genus Thelephora (Thelephorales, Thelephoraceae) are ectomycorrhizal symbionts of coniferous and broad-leaved plants, and some of them are well-known edible mushrooms, making it an exceptionally important group ecologically and economically. However, the diversity of the species from China has not been fully elucidated. In this study, we conducted a phylogenetic analysis based on the internal transcribed spacer (ITS) regions, using Maximum Likelihood and Bayesian analyses, along with morphological observations of this genus. Four new species from China are proposed, viz., T. dactyliophora, T. lacunosa, T. petaloides, and T. pinnatifida. In addition, T. sikkimensis originally described from India is reported for the first time from China. Thelephora dactyliophora, T. pinnatifida, and T. sikkimensis are distributed in subtropical forests and mainly associated with plants of the families Fagaceae and Pinaceae. Thelephora lacunosa and T. petaloides are distributed in tropical to subtropical forests. Thelephora lacunosa is mainly associated with plants of the families Fagaceae and Pinaceae, while T. petaloides is mainly associated with plants of the family Fagaceae. Line drawings of microstructures, color pictures of fresh basidiomes, and detailed descriptions of these five species are provided.

Molecular phylogeny and morphology reveal two new entomopathogenic species of Ophiocordyceps (Ophiocordycipitaceae, Hypocreales) parasitic on termites from China.

Two new termite-pathogenic species, Ophiocordycepsglobiperitheciata and O.longistipes, are described from Yunnan Province, China. Six-locus (ITS, nrSSU, nrLSU, tef-1α, rpb1 and rpb2) phylogenetic analyses in combination with morphological observations were employed to characterize these two species. Phylogenetically, O.globiperitheciata is most closely related to Hirsutellacryptosclerotium and O.communis, whereas O.longistipes shares a sister relationship with O.fusiformis. However, O.globiperitheciata differs from H.cryptosclerotium by parasitizing Blattodea and producing clavate, unbifurcated stromata. Ophiocordycepsglobiperitheciata is distinguished from O.communis by multiple stromata, shorter asci and ascospores. Ophiocordycepslongistipes differs from O.fusiformis in producing larger stromata, perithecia, asci and ascospores, as well as smaller citriform or oval conidia. Morphological descriptions of the two new species and a dichotomous key to the 19 termite-pathogenic Ophiocordyceps species are presented.

Molecular phylogenetics of the Ophiocordyceps sinensis-species complex lineage (Ascomycota, Hypocreales), with the discovery of new species and predictions of species distribution.

Ophiocordyceps sinensis is a famous traditional Chinese medicine adapted to the alpine environment of the Qinghai-Tibet Plateau and adjacent regions. Clarification of the species diversity of Ophiocordyceps sinensis and its relatives could expand the traditional medicinal resources and provide insights into the speciation and adaptation. The study is prompted by the discovery of a new species, O. megala, described here from a biodiversity hotspot in the Hengduan Mountains, China. Combined morphological, ecological, and phylogenetic evidence supports its distinctiveness from O. sinensis, O. xuefengensis, and O. macroacicularis. Additionally, based on the phylogenetic construction of Ophiocordyceps, a special clade was focused phylogenetically on the more closely related O. sinensis complex, which was defined as the O. sinensis- species complex lineage. A total of 10 species were currently confirmed in this lineage. We made a comprehensive comparison of the sexual/asexual morphological structures among this species complex, distinguishing their common and distinctive features. Furthermore, using the method of species distribution modelling, we studied the species ocurrences in relation to climatic, edaphic, and altitudinal variables for the eight species in the O. sinensis-species complex, and determined that their potential distribution could extend from the southeastern Qinghai-Tibet Plateau to the Xuefeng Mountains without isolating barrier. Thus, the biodiversity corridor hypothesis was proposed around the O. sinensis-species complex. Our study highlights the phylogeny, species diversity, and suitable distribution of the O. sinensis-species complex lineage, which should have a positive implication for the resource discovery and adaptive evolution of this unique and valuable group.

Stable functional structure despite high taxonomic variability across fungal communities in soils of old-growth montane forests.

BACKGROUND: Major advances over the past decade in molecular ecology are providing access to soil fungal diversity in forest ecosystems worldwide, but the diverse functions and metabolic capabilities of this microbial community remain largely elusive. We conducted a field survey in montane old-growth broadleaved and conifer forests, to investigate the relationship between soil fungal diversity and functional genetic traits. To assess the extent to which variation in community composition was associated with dominant tree species (oak, spruce, and fir) and environmental variations in the old-growth forests in the Jade Dragon Snow Mountain in Yunnan Province, we applied rDNA metabarcoding. We also assessed fungal gene expression in soil using mRNA sequencing and specifically assessed the expression of genes related to organic matter decomposition and nutrient acquisition in ectomycorrhizal and saprotrophic fungi. RESULTS: Our taxonomic profiling revealed striking shifts in the composition of the saprotrophic and ectomycorrhizal guilds among the oak-, fir-, and spruce-dominated forests. The core fungal microbiome comprised only ~ 20% of the total OTUs across all soil samples, although the overlap between conifer-associated communities was substantial. In contrast, seasonality and soil layer explained only a small proportion of the variation in community structure. However, despite their highly variable taxonomic composition, fungal guilds exhibited remarkably similar functional traits for growth-related and core metabolic pathways across forest associations, suggesting ecological redundancy. However, we found that the expression profiles of genes related to polysaccharide and protein degradation and nutrient transport notably varied between and within the fungal guilds, suggesting niche adaptation. CONCLUSIONS: Overall, our metatranscriptomic analyses revealed the functional potential of soil fungal communities in montane old-growth forests, including a suite of specialized genes and taxa involved in organic matter decomposition. By linking genes to ecological traits, this study provides insights into fungal adaptation strategies to biotic and environmental factors, and sheds light on the importance of understanding functional gene expression patterns in predicting ecosystem functioning. Video Abstract.

Transcriptome profiling analysis of uterus during chicken laying periods.

The avian eggshell is formed in the uterus. Changes in uterine function may have a significant effect on eggshell quality. To identify the vital genes impacting uterine functional maintenance in the chicken, uteri in three different periods (22W, 31W, 51W) were selected for RNA sequencing and bioinformatics analysis. In our study, 520, 706 and 736 differentially expressed genes (DEGs) were respectively detected in the W31 vs W22 group, W51 vs W31 group and W51 vs W22 group. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated DEGs were enriched in the extracellular matrix, extracellular region part, extracellular region, extracellular matrix structural constituent, ECM receptor interaction, collagen-containing extracellular matrix and collagen trimer in the uterus (P < 0.05). Protein-protein interaction analysis revealed that FN1, LOX, THBS2, COL1A1, COL1A2, COL5A1, COL5A2, POSTN, MMP13, VANGL2, RAD54B, SPP1, SDC1, BTC, ANGPTL3 might be key candidate genes for uterine functional maintenance in chicken. This study discovered dominant genes and pathways which enhanced our knowledge of chicken uterine functional maintenance.

Identification of genes involved in regulating the development of feathered feet in chicken embryo.

The genetic and developmental factors driving the diverse distribution and morphogenesis of feathers and scales on bird feet are yet unclear. Within a single species, Guangxi domestic chickens exhibit dramatic variety in feathered feet, making them an accessible model for research into the molecular basis of variations in skin appendages. In this study, we used H&E staining to observe the morphogenesis of feathered feet, scaled feet and wings skin at different embryonic stages in Longsheng-Feng chickens and Guangxi Partridge chickens. We selected 4 periods (E6, E7, E8, and E12) that play an important role in feather development and performed transcriptome sequencing to screen for candidate genes associated with feathered feet. Through comparison and analysis of transcriptome data, we identified a set of differently expressed genes (DGEs), which were enriched in appendage organ development, hindlimb morphogenesis, activation of transcription factor binding, and binding of sequence-specific DNA in the cis-regulatory region. In addition, we identified some feathered feet-related genes by analyzing the classical signaling pathways that regulate feather development. Finally, we identified candidate genes that regulate feathered feet formation, which include TBX5, PITX1, ZIC1, FGF20, WNT11, WNT7A, WNT16, and SHH. Interestingly, we found that TBX5 was significantly overexpressed in the skin of the feathered feet and had the highest expression at E7 (P < 0.01), whereas PITX1 expression was significantly reduced at E7(P < 0.01). It is hypothesized that TBX5 and PITX1 regulate the development of hair follicles through the Wnt/ß-catenin signaling pathway at E7. Our results provide a theoretical basis for investigating the molecular regulatory mechanisms underlying the formation of chicken feathered feet.

Phylogeny, Taxonomy and Evolutionary Trade-Offs in Reproductive Traits of Gomphoid Fungi (Gomphaceae, Gomphales).

Although functional ecology is a well-established field, our understanding of the evolutionary and ecological significance of the reproductive traits in macrofungi is still limited. Here, we reconstructed a phylogeny tree of gomphoid fungi in the narrower sense, including the species of the genera Gomphus and Turbinellus and used it to uncover the evolution of reproductive traits. Our analyses indicated that fungal fruit bodies and spores did not enlarge at a steady rate over time. Early gomphoid fungi essentially maintained their fruit body size, spore size and spore shape through the Mesozoic. In the Cenozoic, gomphoid fungi acquired significantly larger and more spherical spores by simultaneously expanding in length and width, with the fruit body size first decreasing and then enlarging. We argue that these trade-offs were driven by the effect of biological extinction and the dramatic climate changes of the Cenozoic. Gomphoid fungi initially increased in spore size and fruit body number as extinction survivors filled vacant niches. Both fruit bodies and spores eventually became larger as ecosystems saturated and competition intensified. One new species of Gomphus and nine new species of Turbinellus are described.

Phylogeny and Taxonomic Revision of the Family Discinaceae (Pezizales, Ascomycota).

Species of Discinaceae are common macrofungi with a worldwide distribution. Some of them are commercially consumed, while a few others are reported as poisonous. Two genera were accepted in the family: the epigeous Gyromitra with discoid, cerebriform to saddle-shaped ascomata and the hypogeous Hydnotrya with globose or tuberous ascomata. However, due to discrepancies in their ecological behaviors, a comprehensive investigation of their relationship was not thoroughly explored. In this study, phylogenies of Discinaceae were reconstructed using sequence analyses of combined and separate three gene partitions (internal transcribed spacer [ITS], large subunit ribosomal DNA [LSU], and translation elongation factor [TEF]) with a matrix containing 116 samples. As a result, the taxonomy of the family was renewed. Eight genera were recognized: two of them (Gyromitra and Hydnotrya) were retained, three (Discina, Paradiscina, and Pseudorhizina) were revived, and three (Paragyromitra, Pseudodiscina, and Pseudoverpa) were newly established. Nine new combinations were made in four genera. Two new species in Paragyromitra and Pseudodiscina and an un-named taxon of Discina were described and illustrated in detail based on the materials collected from China. Furthermore, a key to the genera of the family was also provided. IMPORTANCE Taxonomy of the fungal family Discinaceae (Pezizales, Ascomycota) was significantly renewed on the basis of sequence analyses of internal transcribed spacer (ITS), large subunit ribosomal DNA (LSU), and translation elongation factor (TEF). Eight genera were accepted, including three new genera; two new species were described; and nine new combinations were made. A key to the accepted genera of the family is provided. The aim of this study is to deepen the understanding of the phylogenetic relationships among genera of the group, as well as the associated generic concepts.

Whole-transcriptome RNA sequencing reveals the global molecular responses and circRNA/lncRNA-miRNA-mRNA ceRNA regulatory network in chicken fat deposition.

Fat deposition is a vital factor affecting the economics of poultry production. Numerous studies on fat deposition have been done. However, the molecular regulatory mechanism is still unclear. In the present study, the whole-transcriptome RNA sequencing in abdominal fat, back skin, and liver both high- and low-abdominal fat groups was used to uncover the competitive endogenous RNA (ceRNA) regulation network related to chicken fat deposition. The results showed that differentially expressed (DE) genes in abdominal fat, back skin, liver were 1207(784 mRNAs, 330 lncRNAs, 41 circRNAs, 52 miRNAs), 860 (607 mRNAs, 166 lncRNAs, 26 circRNAs, 61 miRNAs), and 923 (501 mRNAs, 262 lncRNAs, 15 circRNAs, 145 miRNAs), respectively. The ceRNA regulatory network analysis indicated that the fatty acid metabolic process, monocarboxylic acid metabolic process, carboxylic acid metabolic process, glycerolipid metabolism, fatty acid metabolism, and peroxisome proliferator-activated receptor (PPAR) signaling pathway took part in chicken fat deposition. Meanwhile, we scan the important genes, FADS2, HSD17B12, ELOVL5, AKR1E2, DGKQ, GPAM, PLIN2, which were regulated by gga-miR-460b-5p, gga-miR-199-5p, gga-miR-7470-3p, gga-miR-6595-5p, gga-miR-101-2-5p. While these miRNAs were competitive combined by lncRNAs including MSTRG.18043, MSTRG.7738, MSTRG.21310, MSTRG.19577, and circRNAs including novel_circ_PTPN2, novel_circ_CTNNA1, novel_circ_PTPRD. This finding provides new insights into the regulatory mechanism of mRNA, miRNA, lncRNA, and circRNA in chicken fat deposition.

Grade follicles transcriptional profiling analysis in different laying stages in chicken.

During follicular development, a series of key events such as follicular recruitment and selection are crucially governed by strict complex regulation. However, its molecular mechanisms remain obscure. To identify the dominant genes controlling chicken follicular development, the small white follicle (SWF), the small yellow follicle (SYF), and the large yellow follicle (LYF) in different laying stages (W22, W31, W51) were collected for RNA sequencing and bioinformatics analysis. There were 1866, 1211, and 1515 differentially expressed genes (DEGs) between SWF and SYF in W22, W31, and W51, respectively. 4021, 2295, and 2902 DEGs were respectively identified between SYF and LYF in W22, W31, and W51. 5618, 4016, and 4809 DEGs were respectively identified between SWF and LYF in W22, W31, and W51. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that extracellular matrix, extracellular region, extracellular region part, ECM-receptor interaction, collagen extracellular matrix, and collagen trimer were significantly enriched (P < 0.05). Protein-protein interaction analysis revealed that COL4A2, COL1A2, COL4A1, COL5A2, COL12A1, ELN, ALB, and MMP10 might be key candidate genes for follicular development in chicken. The current study identified dominant genes and pathways contributing to our understanding of chicken follicular development.

Gomphocantharellus, a new genus of Gomphales.

The genus Gomphocantharellus and species Gomphocantharellus cylindrosporus are proposed as new based on morphological assessments and molecular phylogenetic evidence inferred from nucleotide sequences of mitochondrial (mt) adenosine triphosphate (ATPase) subunit 6 (atp6) and mt small subunit rDNA (mtSSU). Basidiomes of G. cylindrosporus are characterized by the peach to pinkish orange color, cantharelloid habit with a gill-like hymenophore with obtuse edges, smooth and cylindrical to allantoid basidiospores, and cylindrical to narrowly clavate flexuous pleurocystidia. The species resembles a species of Cantharellus but differs from the latter by the cylindrical basidiospores. Phylogenetic analyses confirm the placement of Gomphocantharellus as an independent lineage within the order Gomphales.

Ecological Drivers of the Soil Microbial Diversity and Composition in Primary Old-Growth Forest and Secondary Woodland in a Subtropical Evergreen Broad-Leaved Forest Biome in the Ailao Mountains, China.

Replacement of primary old-growth forests by secondary woodlands in threatened subtropical biomes drives important changes at the level of the overstory, understory and forest floor, but the impact on belowground microbial biodiversity is yet poorly documented. In the present study, we surveyed by metabarcoding sequencing, the diversity and composition of soil bacteria and fungi in the old-growth forest, dominated by stone oaks (Lithocarpus spp.) and in the secondary Yunnan pine woodland of an iconic site for biodiversity research, the Ailaoshan National Nature Reserve (Ailao Mountains, Yunnan province, China). We assessed the effect of forest replacement and other environmental factors, including soil horizons, soil physicochemical characteristics and seasonality (monsoon vs. dry seasons). We showed that tree composition and variation in soil properties were major drivers for both bacterial and fungal communities, with a significant influence from seasonality. Ectomycorrhizal Operational Taxonomic Units (OTUs) dominated the functional fungal guilds. Species richness and diversity of the bacterial and fungal communities were higher in the pine woodland compared to the primary Lithocarpus forest, although prominent OTUs were different. The slightly lower complexity of the microbiome in the primary forest stands likely resulted from environmental filtering under relatively stable conditions over centuries, when compared to the secondary pine woodlands. In the old-growth forest, we found a higher number of species, but that communities were homogeneously distributed, whereas in the pine woodlands, there is a slightly lower number of species present but the communities are heterogeneously distributed. The present surveys of the bacterial and fungal diversity will serve as references in future studies aiming to assess the impact of the climate change on soil microbial diversity in both old-growth forests and secondary woodlands in Ailaoshan.

Evolutionary innovations through gain and loss of genes in the ectomycorrhizal Boletales.

We aimed to identify genomic traits of transitions to ectomycorrhizal ecology within the Boletales by comparing the genomes of 21 symbiotrophic species with their saprotrophic brown-rot relatives. Gene duplication rate is constant along the backbone of Boletales phylogeny with large loss events in several lineages, while gene family expansion sharply increased in the late Miocene, mostly in the Boletaceae. Ectomycorrhizal Boletales have a reduced set of plant cell-wall-degrading enzymes (PCWDEs) compared with their brown-rot relatives. However, the various lineages retain distinct sets of PCWDEs, suggesting that, over their evolutionary history, symbiotic Boletales have become functionally diverse. A smaller PCWDE repertoire was found in Sclerodermatineae. The gene repertoire of several lignocellulose oxidoreductases (e.g. laccases) is similar in brown-rot and ectomycorrhizal species, suggesting that symbiotic Boletales are capable of mild lignocellulose decomposition. Transposable element (TE) proliferation contributed to the higher evolutionary rate of genes encoding effector-like small secreted proteins, proteases, and lipases. On the other hand, we showed that the loss of secreted CAZymes was not related to TE activity but to DNA decay. This study provides novel insights on our understanding of the mechanisms influencing the evolutionary diversification of symbiotic boletes.

Transcriptome analysis reveals key genes and pathways associated with egg production in Nandan-Yao domestic chicken.

BACKGROUND: Egg production is a very important economic trait in chicken breeding, but its molecular mechanism is unclear until now. Nandan-Yao chicken (Gallus gallus domesticus) is a native breed in Guangxi province, China, which is famous for good meet quality, but with low egg production. METHODS: To explore the molecular regulation related to egg production, high egg production (HEP) and low egg production (LEP) were divided according to the total egg number at 55 weeks, and the concentration of serum sex hormones was tested to evaluate the physiological function of ovary and uterus. RNA sequencing (RNA-Seq) was used to explore the transcriptome from the ovary and uterus of Nandan-Yao chicken. RESULTS: The levels of serum sex hormone showed that concentrations of estradiol (E2), follicle-stimulating hormone (FSH), and luteotropic hormone (LH) were significantly higher in HEP than those in LEP (P < 0.01), while the concentration of testosterone (T) was significantly lower in HEP (P < 0.01). RNA-Seq analysis identified 901 and 2763 differentially expressed genes (DEGs) in ovary and uterus, respectively. Enrichment analysis showed that DEGs were significantly involved in the regulation of tight junction in the ovary (P < 0.05), while in uterus, DEGs were mainly enriched in the phagosome, ECM-receptor interaction, cell adhesion molecules (CAMs), focal adhesion, cardiac muscle contraction, cytokine-cytokine receptor interaction, and the regulation of MAPK signaling pathway (P < 0.05). Protein network interaction and function analyses revealed that FN1, FGF7, SOX2 identified from the ovary, and UQCRH, COX5A, FN1 from the uterus might be key candidate genes for egg production in Nandan-Yao chicken. CONCLUSIONS: Our study provided key candidate genes and pathways involved in the egg-laying process of Nandan-Yao chicken and could help to further understand the molecular mechanisms of chicken reproduction.

Genome-Wide Association Study Using Whole-Genome Sequencing Identifies a Genomic Region on Chromosome 6 Associated With Comb Traits in Nandan-Yao Chicken.

Comb traits have potential economic value in the breeding of indigenous chickens in China. Identifying and understanding relevant molecular markers for comb traits can be beneficial for genetic improvement. The purpose of this study was to utilize genome-wide association studies (GWAS) to detect promising loci and candidate genes related to comb traits, namely, comb thickness (CT), comb weight (CW), comb height, comb length (CL), and comb area. Genome-wide single-nucleotide polymorphisms (SNPs) and small insertions/deletions (INDELs) in 300 Nandan-Yao chickens were detected using whole-genome sequencing. In total, we identified 134 SNPs and 25 INDELs that were strongly associated with the five comb traits. A remarkable region spanning from 29.6 to 31.4 Mb on chromosome 6 was found to be significantly associated with comb traits in both SNP- and INDEL-based GWAS. In this region, two lead SNPs (6:30,354,876 for CW and CT and 6:30,264,318 for CL) and one lead INDEL (a deletion from 30,376,404 to 30,376,405 bp for CL and CT) were identified. Additionally, two genes were identified as potential candidates for comb development. The nearby gene fibroblast growth factor receptor 2 (FGFR2)-associated with epithelial cell migration and proliferation-and the gene cytochrome b5 reductase 2 (CYB5R2)-identified on chromosome 5 from INDEL-based GWAS-are significantly correlated with collagen maturation. The findings of this study could provide promising genes and biomarkers to accelerate genetic improvement of comb development based on molecular marker-assisted breeding in Nandan-Yao chickens.

Differential Expression of Amanitin Biosynthetic Genes and Novel Cyclic Peptides in Amanita molliuscula.

Amanita molliuscula is a basal species of lethal Amanita and intrigues the field because it does not produce discernable α-amanitin when inspected by High Performance Liquid Chromatography (HPLC), which sets it apart from all known amanitin-producing (lethal) Amanita species. In order to study the underlining genetic basis of the phenotype, we sequenced this species through PacBio and Illumina RNA-Seq platforms. In total, 17 genes of the "MSDIN" family (named after the first five amino acid residues of the precursor peptides) were found in the genome and 11 of them were expressed at the transcription level. The expression pattern was not even but in a differential fashion: two of the MSDINs were highly expressed (FPKM value > 100), while the majority were expressed at low levels (FPKM value < 1). Prolyl oligopeptidease B (POPB) is the key enzyme in the amanitin biosynthetic pathway, and high expression of this enzyme was also discovered (FPKM value > 100). The two MSDINs with highest transcription further translated into two novel cyclic peptides, the structure of which is distinctive from all known cyclic peptides. The result illustrates the correlation between the expression and the final peptide products. In contrast to previous HPLC result, the genome of A. molliuscula harbors α-amanitin genes (three copies), but the product was in trace amount indicated by MS. Overall, transcription of MSDINs encoding major toxins (α-amanitin, ß-amanitin, phallacidin and phalloidin) were low, showing that these toxins were not actively synthesized at the stage. Collectively, our results indicate that the amanitin biosynthetic pathway is highly active at the mature fruiting body stage in A. molliuscula, and due to the differential expression of MSDIN genes, the pathway produces only a few cyclic peptides at the time.