Single-cell multi-omics profiling reveals key regulatory mechanisms that poise germinal vesicle oocytes for maturation in pigs.

The molecular mechanisms controlling the transition from meiotic arrest to meiotic resumption in mammalian oocytes have not been fully elucidated. Single-cell omics technology provides a new opportunity to decipher the early molecular events of oocyte growth in mammals. Here we focused on analyzing oocytes that were collected from antral follicles in different diameters of porcine pubertal ovaries, and used single-cell M&T-seq technology to analyze the nuclear DNA methylome and cytoplasmic transcriptome in parallel for 62 oocytes. 10× Genomics single-cell transcriptomic analyses were also performed to explore the bi-directional cell-cell communications within antral follicles. A new pipeline, methyConcerto, was developed to specifically and comprehensively characterize the methylation profile and allele-specific methylation events for a single-cell methylome. We characterized the gene expressions and DNA methylations of individual oocyte in porcine antral follicle, and both active and inactive gene's bodies displayed high methylation levels, thereby enabled defining two distinct types of oocytes. Although the methylation levels of Type II were higher than that of Type I, Type II contained nearly two times more of cytoplasmic transcripts than Type I. Moreover, the imprinting methylation patterns of Type II were more dramatically divergent than Type I, and the gene expressions and DNA methylations of Type II were more similar with that of MII oocytes. The crosstalk between granulosa cells and Type II oocytes was active, and these observations revealed that Type II was more poised for maturation. We further confirmed Insulin Receptor Substrate-1 in insulin signaling pathway is a key regulator on maturation by in vitro maturation experiments. Our study provides new insights into the regulatory mechanisms between meiotic arrest and meiotic resumption in mammalian oocytes. We also provide a new analytical package for future single-cell methylomics study.

Exposure to nicotine regulates prostaglandin E2 secretion and autophagy of granulosa cells to retard follicular maturation in mammals.

Exposure to nicotine by cigarette smoking have shown strongly defectives on the physiological function of ovaries, which in turn leads to disorders of fertility in women. However, the potential molecular mechanisms remain to be elucidated. In this study, we notably found that nicotine was likely to specifically raise the expression of histone deacetylase 3 (HDAC3) to promote the apoptosis and autophagy of granulosa cells (GCs) and block follicular maturation. Moreover, prostaglandin E2 (PGE2) inhibited the apoptosis of GCs and facilitated follicular maturation, and nicotine appeared to inhibit PGE2 secretion by freezing the expression of cyclooxygenase 1 (COX1), which was the rate-limiting and essential enzyme for PGE2 synthesis. Epigenetically, the nicotine was observed to diminish the histone H3 lysine 9 acetylation (H3K9ac) level and compact the chromatin accessibility in -1776/-1499 bp region of COX1 by evoking the expression of HDAC3, with the deactivated Cas9-HDAC3/sgRNA system. Mechanistically, the COX1 protein was found to pick up and degrade the autophagy related protein beclin 1 (BECN1) to control the autophagy of GCs. These results provided a potential new molecular therapy to recover the damage of female fertility induced by nicotine from cigarette smoking.

Knockdown of DNMT1 Induces SLCO3A1 to Promote Follicular Growth by Enhancing the Proliferation of Granulosa Cells in Mammals.

In female mammals, the proliferation and apoptosis of granulosa cells (GCs) have been shown to determine the fate of follicles. DNA methyltransferases (DNMTs) and SLCO3A1 have been reported to be involved in the survival of GCs and follicular growth. However, the molecular mechanisms enabling DNMTs to regulate the expression of SLCO3A1 to participate in follicular growth are unclear. In this study, we found that the knockdown of DNMT1 enhanced the mRNA and protein levels of SLCO3A1 by regulating the chromatin accessibility probably. Moreover, SLCO3A1 upregulated the mRNA and protein levels of MCL1, PCNA, and STAR to promote the proliferation of GCs and facilitated cell cycle progression by increasing the mRNA and protein levels of CCNE1, CDK2, and CCND1, but it decreased apoptosis by downregulating the mRNA and protein levels of CASP3 and CASP8. Moreover, SLCO3A1 promoted the growth of porcine follicles and development of mice follicles. In conclusion, the knockdown of DNMT1 upregulated the mRNA and protein levels of SLCO3A1, thereby promoting the proliferation of GCs to facilitate the growth and development of ovarian follicles, and these results provide new insights into investigations of female reproductive diseases.

Integrating Multiple Database Resources to Elucidate the Gene Flow in Southeast Asian Pig Populations.

The domestic pig (Sus scrofa) and its subfamilies have experienced long-term and extensive gene flow, particularly in Southeast Asia. Here, we analyzed 236 pigs, focusing on Yunnan indigenous, European commercial, East Asian, and Southeast Asian breeds, using the Pig Genomics Reference Panel (PGRP v1) of Pig Genotype-Tissue Expression (PigGTEx) to investigate gene flow and associated complex traits by integrating multiple database resources. In this study, we discovered evidence of admixtures from European pigs into the genome of Yunnan indigenous pigs. Additionally, we hypothesized that a potential conceptual gene flow route that may have contributed to the genetic composition of the Diannan small-ear pig is a gene exchange from the Vietnamese pig. Based on the most stringent gene introgression scan using the fd statistic, we identified three specific loci on chromosome 8, ranging from 51.65 to 52.45 Mb, which exhibited strong signatures of selection and harbored the NAF1, NPY1R, and NPY5R genes. These genes are associated with complex traits, such as fat mass, immunity, and litter weight, in pigs, as supported by multiple bio-functionalization databases. We utilized multiple databases to explore the potential dynamics of genetic exchange in Southeast Asian pig populations and elucidated specific gene functionalities.

Genome-Wide Identification and Characterization of the CCT Gene Family in Rapeseed (Brassica napus L.).

The CCT gene family is present in plants and is involved in biological processes such as flowering, circadian rhythm regulation, plant growth and development, and stress resistance. We identified 87, 62, 46, and 40 CCTs at the whole-genome level in B. napus, B. rapa, B. oleracea, and A. thaliana, respectively. The CCTs can be classified into five groups based on evolutionary relationships, and each of these groups can be further subdivided into three subfamilies (COL, CMF, and PRR) based on function. Our analysis of chromosome localization, gene structure, collinearity, cis-acting elements, and expression patterns in B. napus revealed that the distribution of the 87 BnaCCTs on the chromosomes of B. napus was uneven. Analysis of gene structure and conserved motifs revealed that, with the exception of a few genes that may have lost structural domains, the majority of genes within the same group exhibited similar structures and conserved domains. The gene collinearity analysis identified 72 orthologous genes, indicating gene duplication and expansion during the evolution of BnaCCTs. Analysis of cis-acting elements identified several elements related to abiotic and biotic stress, plant hormone response, and plant growth and development in the promoter regions of BnaCCTs. Expression pattern and protein interaction network analysis showed that BnaCCTs are differentially expressed in various tissues and under stress conditions. The PRR subfamily genes have the highest number of interacting proteins, indicating their significant role in the growth, development, and response to abiotic stress of B. napus.

Evaluation of six machine learning classification algorithms in pig breed identification using SNPs array data.

Breed identification utilizing multiple information sources and methods is widely applicated in the field of animal genetics and breeding. Simultaneously, with the development of artificial intelligence, the integration of high-throughput genomic data and machine learning techniques is increasingly used for breed identification. In this context, we used 654 individuals from 15 pig breeds, evaluating the performance of machine learning and stacking ensemble learning classifiers, as well as the function of feature selection and anomaly detection in different scenarios. Our results showed that, when using a training set of 16 individuals per breed and 32 features (SNPs), the accuracy of breed identification with feature selection (eXtreme Gradient Boosting, XGBoost) could exceed 95.00% (nine breeds), and was improved by 7.04% over the results with random selection. For stacking ensemble learning, feature selection methods (including random selection method) were used before different base learners. When these base learners' training set had 16 individuals per breed and 32 features, the accuracy of stacking ensemble learning improved by 9.24% over the best base learner (nine breeds), but did not significantly increase the advantage over the models with XGBoost feature selection. When using a training set of 16 individuals and 512 features per breed, breed identification with anomaly detection (local outlier factor, LOF) and random selection could achieve an accuracy of 89.06% (15 breeds). These results show that machine learning could be an effective tool for breed identification and this study will also provide useful information for the application of machine learning in animal genetics and breeding.

Application of Nanomaterials in the Production of Biomolecules in Microalgae: A Review.

Nanomaterials (NMs) are becoming more commonly used in microalgal biotechnology to empower the production of algal biomass and valuable metabolites, such as lipids, proteins, and exopolysaccharides. It provides an effective and promising supplement to the existing algal biotechnology. In this review, the potential for NMs to enhance microalgal growth by improving photosynthetic utilization efficiency and removing reactive oxygen species is first summarized. Then, their positive roles in accumulation, bioactivity modification, and extraction of valuable microalgal metabolites are presented. After the application of NMs in microalgae cultivation, the extracted metabolites, particularly exopolysaccharides, contain trace amounts of NM residues, and thus, the impact of these residues on the functional properties of the metabolites is also evaluated. Finally, the methods for removing NM residues from the extracted metabolites are summarized. This review provides insights into the application of nanotechnology for sustainable production of valuable metabolites in microalgae and will contribute useful information for ongoing and future practice.

Advances in Oocyte Maturation In Vivo and In Vitro in Mammals.

The quality and maturation of an oocyte not only play decisive roles in fertilization and embryo success, but also have long-term impacts on the later growth and development of the fetus. Female fertility declines with age, reflecting a decline in oocyte quantity. However, the meiosis of oocytes involves a complex and orderly regulatory process whose mechanisms have not yet been fully elucidated. This review therefore mainly focuses on the regulation mechanism of oocyte maturation, including folliculogenesis, oogenesis, and the interactions between granulosa cells and oocytes, plus in vitro technology and nuclear/cytoplasm maturation in oocytes. Additionally, we have reviewed advances made in the single-cell mRNA sequencing technology related to oocyte maturation in order to improve our understanding of the mechanism of oocyte maturation and to provide a theoretical basis for subsequent research into oocyte maturation.

Comprehensive Analysis of CircRNA Expression Profiles in Multiple Tissues of Pigs.

Circular RNAs (circRNAs) are a class of non-coding RNAs with diverse functions, and previous studies have reported that circRNAs are involved in the growth and development of pigs. However, studies about porcine circRNAs over the past few years have focused on a limited number of tissues. Based on 215 publicly available RNA sequencing (RNA-seq) samples, we conducted a comprehensive analysis of circRNAs in nine pig tissues, namely, the gallbladder, heart, liver, longissimus dorsi, lung, ovary, pituitary, skeletal muscle, and spleen. Here, we identified a total of 82,528 circRNAs and discovered 3818 novel circRNAs that were not reported in the CircAtlas database. Moreover, we obtained 492 housekeeping circRNAs and 3489 tissue-specific circRNAs. The housekeeping circRNAs were enriched in signaling pathways regulating basic biological tissue activities, such as chromatin remodeling, nuclear-transcribed mRNA catabolic process, and protein methylation. The tissue-specific circRNAs were enriched in signaling pathways related to tissue-specific functions, such as muscle system process in skeletal muscle, cilium organization in pituitary, and cortical cytoskeleton in ovary. Through weighted gene co-expression network analysis, we identified 14 modules comprising 1377 hub circRNAs. Additionally, we explored circRNA-miRNA-mRNA networks to elucidate the interaction relationships between tissue-specific circRNAs and tissue-specific genes. Furthermore, our conservation analysis revealed that 19.29% of circRNAs in pigs shared homologous positions with their counterparts in humans. In summary, this extensive profiling of housekeeping, tissue-specific, and co-expressed circRNAs provides valuable insights into understanding the molecular mechanisms of pig transcriptional expression, ultimately deepening our understanding of genetic and biological processes.

Study on detection of CNVs using human whole genome bisulfite sequencing data.

It has been reported that the aberrant DNA methylation may result in copy number variations (CNVs), and the CNVs may alter the levels of DNA methylation. Whole genome bisulfite sequencing (WGBS) is able to generate the sequencing data of DNAs, and shows the potential ability to detect CNVs. However, the evaluations and performances on the detections of CNVs using WGBS data is still unclear. In this study, five software with different strategies for CNV detections, e.g., BreakDancer, cn.mops, CNVnator, DELLY and Pindel, were selected to explore and benchmark the performances of CNV detections with WGBS data. Based on the real (2.62 billion reads) and simulated (12.35 billion reads) WGBS data of humans, we calculated the number, precision, recall, relative ability, memory usage, and running time of CNV detections by 150 times, and tried to figure out the optimal strategy for CNV detections with WGBS data. Based on the real WGBS data, Pindel detected the most deletions and duplications, CNVnator detected the deletions with the highest precision, cn.mops detected the duplications with the highest precision, Pindel detected the deletions with the highest recall, and cn.mops detected the duplications with the highest recall. Based on the simulated WGBS data, BreakDancer detected the most deletions, and cn.mops detected the most duplications. The CNVnator showed the highest precision and recall for both deletions and duplications. In real and simulated WGBS data, the ability of CNVnator to detect CNVs was likely to overtake that in the whole genome sequencing data. Additionally, DELLY and BreakDancer displayed the lowest peak of memory usage and the lest CPU runtime, while CNVnator expressed the highest peak of memory usage and the most CPU runtime. Taken together, CNVnator and cn.mops showed the excellent performances of CNV detections with WGBS data. These results suggested that it was feasible to detect CNVs using WGBS data, and provided the useful information to further investigate both CNVs and DNA methylation using WGBS data alone.

Ovary-derived circular RNAs profile analysis during the onset of puberty in gilts.

BACKGROUND: In mammals, the ovary is the essential system of female reproduction for the onset of puberty, and the abnormal puberty has negative outcomes on health. CircRNA is a non-coding RNA produced by non-canonical alternative splicing (AS). Several studies have reported that circRNA is involved in the gene regulation and plays an important role in some human diseases. However, the contribution of circRNA has received little known within the onset of puberty in ovary. RESULTS: Here, the profiles of ovarian circRNAs across pre-, in- and post-pubertal stages were established by RNA-sEq. In total, 972 circRNAs were identified, including 631 stage-specific circRNAs and 8 tissue-specific circRNAs. The biological functions of parental genes of circRNAs were enriched in steroid biosynthesis, autophagy-animal, MAPK signaling pathway, progesterone-mediated oocyte maturation and ras signaling pathway. Moreover, 5 circRNAs derived from 4 puberty-related genes (ESR1, JAK2, NF1 and ARNT) were found in this study. The A3SS events were the most alternative splicing, but IR events were likely to be arose in post-pubertal ovaries. Besides, the circRNA-miRNA-gene networks were explored for 10 differentially expressed circRNAs. Furthermore, the head-to-tail exon as well as the expressions of 10 circRNAs were validated by the divergent RT-qPCR and sanger sequencing. CONCLUSIONS: In summary, the profiles of ovarian circRNAs were provided during pubertal transition in gilts, and these results provided useful information for the investigation on the onset of puberty at the ovarian-circRNAs-level in mammals.

The Overlooked Genetic Diversity in the Dryland Soil Surface-Dwelling Cyanobacterium Nostoc flagelliforme as Revealed by the Marker Gene wspA.

Biodiversity is recognized to be relatively low in the dryland ecosystem. However, we might overlook the accumulating genetic variation in those dryland micro-populations, which should eventually increase the dryland biodiversity. In the xeric steppes of western and northwestern China, there are two soil surface-dwelling and genetically close cyanobacterial species, Nostoc commune and Nostoc flagelliforme. They respectively exhibit lamellate and filamentous colony shapes. Their individual colony is consisted of hundreds of trichomes and the common exopolysaccharide matrix. N. flagelliforme is exclusively distributed in the dryland and supposed to be evolved from N. commune. We previously reported that the morphological diversity of N. flagelliforme colonies was very limited, being either cylindrical or strip-like. In this communication, we performed single-nucleotide polymorphism (SNP) analysis of the marker gene wspA as well as phylogenetic analysis of the WspA protein in N. flagelliforme colonies to gain insights into its genetic diversity. SNP analysis suggested that there existed plentiful nucleotide variations in the individual colonies and meanwhile these variations shared certain evolutionary regularity. Phylogenetic analysis of the deduced proteins from the cloned wspA sequences suggested that the relatively regular variations were possibly dispersed in the N. flagelliforme populations of different regions. Thus, these results presented a scenario of the underestimated genetic diversity hidden behind the limited morphotype of dryland cyanobacteria. Maybe, we can consider the individual cyanobacterial colony as a potential biodiversity pool in the drylands.

FoxA2 and p53 regulate the transcription of HSD17B1 in ovarian granulosa cells of pigs.

The oestrogens have been highly implicated in the fertility of female animals. It is widely known that the oestrogens are primarily synthetized by the ovarian granulosa cells (GCs), and the final and essential step of this process is to catalyse the oestrone to the more active oestradiol by the protein coded by hydroxysteroid 17-beta dehydrogenase 1 (HSD17B1) gene. However, the molecular mechanism regarding the transcription of HSD17B1 remains to be fully elucidated in ovarian GCs. In this study, the 5'-deletion, luciferase assay and chromatin immunoprecipitation (ChIP) were utilized to explore the molecular regulation of transcription of HSD17B1 with the porcine ovarian GCs as the cellular model. After the deletions with -2105 to -1754 bp, -1753 to -1429 bp, -1430 to -1081 bp and -1082 to -730 bp, the relative luciferase activity of HSD17B1 promoter did not change significantly, but the deletion of -731 to -332 bp significantly increased the relative luciferase activity of HSD17B1 promoter, and an insertion (GTTT) that might raise the transcription of HSD17B1 was identified at -401 bp of HSD17B1. These findings suggested the region from -731 to +38 bp was the core promoter of HSD17B1, and the region between -731 to -332 bp might be a silence element for HSD17B1. Furthermore, the forkhead box A2 (FoxA2) directly bound at -412 to -401 bp to negatively but p53 bound at -383 to -374 bp to positively regulate the transcription and translation of HSD17B1 in ovarian GCs. These findings will improve our understanding on HSD17B1-mediated oestrogens and provide useful information for further investigations into fertility of females.

MiR-590-3p regulates proliferation, migration and collagen synthesis of cardiac fibroblast by targeting ZEB1.

Previous studies have implicated the attractive and promising role of miR-590-3p to restore the cardiac function following myocardial infarction (MI). However, the molecular mechanisms for how miR-590-3p involves in cardiac fibrosis remain largely unexplored. Using human cardiac fibroblasts (HCFs) as the cellular model, luciferase report assay, mutation, EdU assay and transwell migration assay were applied to investigate the biological effects of miR-590-3p on the proliferation, differentiation, migration and collagen synthesis of cardiac fibroblasts. We found that miR-590-3p significantly suppressed cell proliferation and migration of HCFs. The mRNA and protein expression levels of α-SMA, Col1A1 and Col3A were significantly decreased by miR-590-3p. Moreover, miR-590-3p directly targeted at the 3'UTR of ZEB1 to repress the translation of ZEB1. Interfering with the expression of ZEB1 significantly decreased the cell proliferation, migration activity, mRNA and protein expressions of α-SMA, Col1A1 and Col3A. Furthermore, the expressions of miR-590-3p and ZEB1 were identified in infarct area of MI model in pigs. Collectively, miR-590-3p suppresses the cell proliferation, differentiation, migration and collagen synthesis of cardiac fibroblasts by targeting ZEB1. These works will provide useful biological information for future studies on potential roles of miR-590-3p as the therapeutic target to recover cardiac function following MI.

Newly reported alkaloids produced by marine-derived Penicillium species (covering 2014-2018).

Alkaloids, especially heterocyclic alkaloids, have received remarkable attention due to their intriguing structures and potential pharmacological activities. The marine fungi residing in extreme environmental conditions are among the richest sources of these basic nitrogen-containing compounds. Fungal species belonging to the genus Penicillium have been studied worldwide for their biosynthetic potential for generating bioactive alkaloids. This paper offers a systematic review of the newly reported alkaloids produced by marine-derived Penicillium species over the past five years (covering the literature from the beginning of 2014 through the end of 2018) and describes the structural diversity, biological activities, and plausible biosynthetic pathway of the reported compounds. A total of 106 alkaloids and 81 references are included in this review, which is expected to be beneficial for drug development and biosynthesis in the near future.

Cadinane- and drimane-type sesquiterpenoids produced by Paecilomyces sp. TE-540, an endophyte from Nicotiana tabacum L., are acetylcholinesterase inhibitors.

Sesquiterpenoids with diverse skeleton types are regarded as potential lead compounds in pharmacological and other applications. Herein, we report the discovery of two new cadinane-type sesquiterpenoids, paecilacadinol A (1) and B (2); two new drimane-type sesquiterpenoids, ustusol D (3) and ustusol E (4); and six known analogs (5-10) from the endophytic fungus Paecilomyces sp. TE-540, enriching the structural diversity of naturally occurring sesquiterpenoids. Their planar structures were determined on the basis of detailed interpretation of 1D and 2D NMR spectroscopy and HRESIMS data, while their stereochemical structures were established by X-ray crystallographic analyses for 1 and 3-8 and theoretical calculations for 2. Notably, compounds 1 and 2 represent novel examples of cadinane-type sesquiterpenoids with ether bonds formed by intramolecular dehydration. Compounds 5 and 6 showed moderate activities against acetylcholinesterase (AChE), with IC50 values of 43.02 ± 6.01 and 35.97 ± 2.12 µM, respectively. Docking analysis predicted that 5 bound well in the catalytic pocket of AChE via hydrophobic interactions with Trp84, Gly117, Ser122, and Tyr121 residues, while 6 was located with Asp72 and Ser122 residues.

Recent Discovery of Heterocyclic Alkaloids from Marine-Derived Aspergillus Species.

Nitrogen heterocycles have drawn considerable attention due to of their significant biological activities. The marine fungi residing in extreme environments are among the richest sources of these basic nitrogen-containing secondary metabolites. As one of the most well-known universal groups of filamentous fungi, marine-derived Aspergillus species produce a large number of structurally unique heterocyclic alkaloids. This review attempts to provide a comprehensive summary of the structural diversity and biological activities of heterocyclic alkaloids that are produced by marine-derived Aspergillus species. Herein, a total of 130 such structures that were reported from the beginning of 2014 through the end of 2018 are included, and 75 references are cited in this review, which will benefit future drug development and innovation.

Optimizing genomic prediction model given causal genes in a dairy cattle population.

As genotypic data are moving from SNP chip toward whole-genome sequence, the accuracy of genomic prediction (GP) exhibits a marginal gain, although all genetic variation, including causal genes, are contained in whole-genome sequence data. Meanwhile, genetic analyses on complex traits, such as genome-wide association studies, have identified an increasing number of genomic regions, including potential causal genes, which would be reliable prior knowledge for GP. Many studies have tried to improve the performance of GP by modifying the prediction model to incorporate prior knowledge. Although several plausible results have been obtained from model modification or strategy optimization, most of them were validated in a specific empirical population with a limited variety of genetic architecture for complex traits. An alternative approach is to use simulated genetic architecture with known causal genes (e.g., simulated causative SNP) to evaluate different GP models with given causal genes. Our objectives were to (1) evaluate the performance of GP under a variety of genetic architectures with a subset of known causal genes and (2) compare different GP models modified by highlighting causal genes and different strategies to weight causal genes. In this study, we simulated pseudo-phenotypes under a variety of genetic architectures based on the real genotypes and phenotypes of a dairy cattle population. Besides classical genomic best linear unbiased prediction, we evaluated 3 modified GP models that highlight causal genes as follows: (1) by treating them as fixed effects, (2) by treating them as a separate random component, and (3) by combining them into the genomic relationship matrix as random effects. Our results showed that highlighting the known causal genes, which explained a considerable proportion of genetic variance in the GP models, increased the predictive accuracy. Combining all given causal genes into the genomic relationship matrix was the optimal strategy under all the scenarios validated, and treating causal genes as a separate random component is also recommended, when more than 20% of genetic variance was explained by known causal genes. Moreover, assigning differential weights to each causal gene further improved the predictive accuracy.

Consumption of Wild Rice (Zizania latifolia) Prevents Metabolic Associated Fatty Liver Disease through the Modulation of the Gut Microbiota in Mice Model.

Metabolic associated fatty liver disease (MAFLD) due to excess weight and obesity threatens public health worldwide. Gut microbiota dysbiosis contributes to obesity and related diseases. The cholesterol-lowering, anti-inflammatory, and antioxidant effects of wild rice have been reported in several studies; however, whether it has beneficial effects on the gut microbiota is unknown. Here, we show that wild rice reduces body weight, liver steatosis, and low-grade inflammation, and improves insulin resistance in high-fat diet (HFD)-fed mice. High-throughput 16S rRNA pyrosequencing demonstrated that wild rice treatment significantly changed the gut microbiota composition in mice fed an HFD. The richness and diversity of the gut microbiota were notably decreased upon wild rice consumption. Compared with a normal chow diet (NCD), HFD feeding altered 117 operational taxonomic units (OTUs), and wild rice supplementation reversed 90 OTUs to the configuration in the NCD group. Overall, our results suggest that wild rice may be used as a probiotic agent to reverse HFD-induced MAFLD through the modulation of the gut microbiota.

Characterization of Nuclear and Mitochondrial Genomes of Two Tobacco Endophytic Fungi Leptosphaerulina chartarum and Curvularia trifolii and Their Contributions to Phylogenetic Implications in the Pleosporales.

The symbiont endophytic fungi in tobacco are highly diverse and difficult to classify. Here, we sequenced the genomes of Curvularia trifolii and Leptosphaerulina chartarum isolated from tobacco plants. Finally, 41.68 Mb and 37.95 Mb nuclear genomes were sequenced for C. trifolii and L. chartarum with the scaffold N50, accounting for 638.94 Kb and 284.12 Kb, respectively. Meanwhile, we obtained 68,926 bp and 59,100 bp for their mitochondrial genomes. To more accurately classify C. trifolii and L. chartarum, we extracted seven nuclear genes and 12 mitochondrial genes from these two genomes and their closely related species. The genes were then used for calculation of evolutionary rates and for phylogenetic analysis. Results showed that it was difficult to achieve consistent results using a single gene due to their different evolutionary rates, while the phylogenetic trees obtained by combining datasets showed stable topologies. It is, therefore, more accurate to construct phylogenetic relationships for endophytic fungi based on multi-gene datasets. This study provides new insights into the distribution and characteristics of endophytic fungi in tobacco.