Isolation of SARS-CoV-2-related coronavirus from Malayan pangolins.

The current outbreak of coronavirus disease-2019 (COVID-19) poses unprecedented challenges to global health1. The new coronavirus responsible for this outbreak-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-shares high sequence identity to SARS-CoV and a bat coronavirus, RaTG132. Although bats may be the reservoir host for a variety of coronaviruses3,4, it remains unknown whether SARS-CoV-2 has additional host species. Here we show that a coronavirus, which we name pangolin-CoV, isolated from a Malayan pangolin has 100%, 98.6%, 97.8% and 90.7% amino acid identity with SARS-CoV-2 in the E, M, N and S proteins, respectively. In particular, the receptor-binding domain of the S protein of pangolin-CoV is almost identical to that of SARS-CoV-2, with one difference in a noncritical amino acid. Our comparative genomic analysis suggests that SARS-CoV-2 may have originated in the recombination of a virus similar to pangolin-CoV with one similar to RaTG13. Pangolin-CoV was detected in 17 out of the 25 Malayan pangolins that we analysed. Infected pangolins showed clinical signs and histological changes, and circulating antibodies against pangolin-CoV reacted with the S protein of SARS-CoV-2. The isolation of a coronavirus from pangolins that is closely related to SARS-CoV-2 suggests that these animals have the potential to act as an intermediate host of SARS-CoV-2. This newly identified coronavirus from pangolins-the most-trafficked mammal in the illegal wildlife trade-could represent a future threat to public health if wildlife trade is not effectively controlled.

A SARS-CoV-2 protein interaction map reveals targets for drug repurposing.

A newly described coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the causative agent of coronavirus disease 2019 (COVID-19), has infected over 2.3 million people, led to the death of more than 160,000 individuals and caused worldwide social and economic disruption1,2. There are no antiviral drugs with proven clinical efficacy for the treatment of COVID-19, nor are there any vaccines that prevent infection with SARS-CoV-2, and efforts to develop drugs and vaccines are hampered by the limited knowledge of the molecular details of how SARS-CoV-2 infects cells. Here we cloned, tagged and expressed 26 of the 29 SARS-CoV-2 proteins in human cells and identified the human proteins that physically associated with each of the SARS-CoV-2 proteins using affinity-purification mass spectrometry, identifying 332 high-confidence protein-protein interactions between SARS-CoV-2 and human proteins. Among these, we identify 66 druggable human proteins or host factors targeted by 69 compounds (of which, 29 drugs are approved by the US Food and Drug Administration, 12 are in clinical trials and 28 are preclinical compounds). We screened a subset of these in multiple viral assays and found two sets of pharmacological agents that displayed antiviral activity: inhibitors of mRNA translation and predicted regulators of the sigma-1 and sigma-2 receptors. Further studies of these host-factor-targeting agents, including their combination with drugs that directly target viral enzymes, could lead to a therapeutic regimen to treat COVID-19.

Pathogenicity, tissue tropism and potential vertical transmission of SARSr-CoV-2 in Malayan pangolins.

Malayan pangolin SARS-CoV-2-related coronavirus (SARSr-CoV-2) is closely related to SARS-CoV-2. However, little is known about its pathogenicity in pangolins. Using CT scans we show that SARSr-CoV-2 positive Malayan pangolins are characterized by bilateral ground-glass opacities in lungs in a similar manner to COVID-19 patients. Histological examination and blood gas tests are indicative of dyspnea. SARSr-CoV-2 infected multiple organs in pangolins, with the lungs the major target, and histological expression data revealed that ACE2 and TMPRSS2 were co-expressed with viral RNA. Transcriptome analysis indicated that virus-positive pangolins were likely to have inadequate interferon responses, with relative greater cytokine and chemokine activity in the lung and spleen. Notably, both viral RNA and viral proteins were detected in three pangolin fetuses, providing initial evidence for vertical virus transmission. In sum, our study outlines the biological framework of SARSr-CoV-2 in pangolins, revealing striking similarities to COVID-19 in humans.

A newly identified secretory phospholipase A2 group XIIA homolog (LcPLA2XIIA) in Larimichthys crocea exhibits antimicrobial and antitumor activities.

The phospholipase A2 (PLA2) superfamily has attracted increasing attention in recent years due to the multiple physiological and pathological functions exerted by its members. Up to date, the knowledge about the biological role of PLA2XIIA subfamily members remains limited. In this study, a new member of PLA2XIIA subfamily, LcPLA2XIIA, was characterized in large yellow croaker. Different from most members of the PLA2 superfamily with positive charge, LcPLA2XIIA encodes an anionic protein, which is similar to other members of PLA2XIIA subfamily. LcPLA2XIIA is highly expressed in the intestine, and afterwards, it is up-regulated after with Pseudomonas plecoglossicida or Staphylococcus aureus. LcPLA2XIIA exhibits strong inhibitory activity against these two bacteria. The results indicate that LcPLA2XIIA plays an important role in the antimicrobial immune responses of large yellow croaker. LcPLA2XIIA displays strong binding activity to all the tested bacteria. It specifically interacts with LTA, a unique component on the surface of Gram-positive bacteria. It also significantly promotes bacterial agglutination in the presence of Ca2+. These findings reveal that the binding and agglutinating abilities of LcPLA2XIIA to bacteria contribute greatly to its antibacterial activity. In addition, LcPLA2XIIA significantly inhibits the proliferation of infectious hematopoietic necrosis virus instead of recombinant human adenovirus type 5. It also suppresses the growth of human colorectal adenocarcinoma cells by inducing apoptosis, but it has no obvious inhibitory effect on the growth of epithelioma papulosum cyprinid cells. This study provides new insights into the antibacterial activity, and the mechanism of LcPLA2XIIA in large yellow croaker, and antiviral and antitumor functions of PLA2XIIA subfamily members.

Population genomic analysis provides evidence of the past success and future potential of South China tiger captive conservation.

BACKGROUND: Among six extant tiger subspecies, the South China tiger (Panthera tigris amoyensis) once was widely distributed but is now the rarest one and extinct in the wild. All living South China tigers are descendants of only two male and four female wild-caught tigers and they survive solely in zoos after 60 years of effective conservation efforts. Inbreeding depression and hybridization with other tiger subspecies were believed to have occurred within the small, captive South China tiger population. It is therefore urgently needed to examine the genomic landscape of existing genetic variation among the South China tigers. RESULTS: In this study, we assembled a high-quality chromosome-level genome using long-read sequences and re-sequenced 29 high-depth genomes of the South China tigers. By combining and comparing our data with the other 40 genomes of six tiger subspecies, we identified two significantly differentiated genomic lineages among the South China tigers, which harbored some rare genetic variants introgressed from other tiger subspecies and thus maintained a moderate genetic diversity. We noticed that the South China tiger had higher FROH values for longer runs of homozygosity (ROH > 1 Mb), an indication of recent inbreeding/founder events. We also observed that the South China tiger had the least frequent homozygous genotypes of both high- and moderate-impact deleterious mutations, and lower mutation loads than both Amur and Sumatran tigers. Altogether, our analyses indicated an effective genetic purging of deleterious mutations in homozygous states from the South China tiger, following its population contraction with a controlled increase in inbreeding based on its pedigree records. CONCLUSIONS: The identification of two unique founder/genomic lineages coupled with active genetic purging of deleterious mutations in homozygous states and the genomic resources generated in our study pave the way for a genomics-informed conservation, following the real-time monitoring and rational exchange of reproductive South China tigers among zoos.

Unveiling the Molecular Mechanisms Regulating Muscle Elasticity in the Large Yellow Croaker: Insights from Transcriptomics and Metabolomics.

The large yellow croaker (Larimichthys crocea) is an important economic fish in China. However, intensive farming practices, such as high stocking densities, suboptimal water quality, and imbalanced nutrition, have led to a decline in muscle quality. Muscle elasticity is a key texture property influencing muscle quality. Herein, transcriptomic and metabolomic analyses were performed on four groups: male high muscle elasticity (MEHM), female high muscle elasticity (MEHF), male low muscle elasticity (MELM), and female low muscle elasticity (MELF), to explore the molecular regulation underlying muscle elasticity in the large yellow croaker. Transcriptomics identified 2594 differentially expressed genes (DEGs) across the four groups, while metabolomics revealed 969 differentially expressed metabolites (DEMs). Association analysis indicated that the valine, leucine, and isoleucine biosynthesis pathways were significantly enriched between the MELF and MEHF groups; 2-Oxoisovalerate and L-Valine were DEMs; and the gene encoding L-threonine ammonia-lyase was a DEG. In the MELM and MEHM groups, pathways such as arginine biosynthesis; arginine and proline metabolism; and valine, leucine, and isoleucine degradation were significantly enriched. 4-guanidinobutanoate, L-aspartate, N-acetylornithine, and L-leucine were among the DEMs, while the DEGs included glul, gls, srm, hmgcs, and aacs. These findings provide insights into the molecular mechanisms controlling muscle elasticity, representing a theoretical foundation to breed high-quality large yellow croakers.

Characterization of TLR5 and TLR9 from silver pomfret (Pampus argenteus) and expression profiling in response to bacterial components.

Toll like receptor (TLR) 5 and 9 are important members of the TLR family that play key roles in innate immunity in all vertebrates. In this study, paTLR5 and paTLR9 were identified in silver pomfret (Pampus argenteus), a marine teleost of great economic value. Open reading frames (ORFs) of paTLR5 and paTLR9 are 2646 and 3225 bp, encoding polypeptides of 881 and 1074 amino acids, respectively. Sequence analysis revealed several conserved characteristic features, including signal peptides, leucine-rich repeat (LRR) motifs, and a Toll/interleukin-I receptor (TIR) domain. Sequence, phylogenetic and synteny analysis revealed high sequence identity with counterparts in other teleosts, confirming their correct nomenclature and conservation during evolution. Quantitative real-time PCR revealed that the that both TLRs were ubiquitously expressed in all investigated tissues, most abundantly in liver, kidney, spleen, intestine and gill, but lower in muscle and skin. In vitro immunostimulation experiments revealed that Aeromonas hydrophila lipopolysaccharide (LPS) and Vibrio anguillarum flagellin induced higher levels of paTLR9 and paTLR5 mRNA expression in isolated fish intestinal epithelial cells (FIECs) than Lactobacillus plantarum lipoteichoic acid (LTA), but all increased the secretion of IL-6 and TNF-α and induced cell apoptosis and necrosis. Together, these results indicate that paTLR5 and paTLR9 may function in the response to bacterial pathogens. Our findings enhance our understanding of the function of TLRs in the innate immune system of silver pomfret and other teleosts.

Molecular characterization and expression analysis of toll-like receptor 2 in response to bacteria in silvery pomfret intestinal epithelial cells.

Toll-like receptor 2 (TLR2) has been shown to play a crucial role in the host defense of pathogenic microbes in innate immunity. In this study, the full-length cDNA of TLR2 in silvery pomfret (Pampus argenteus) was cloned by homology cloning and the rapid amplification of cDNA ends (RACE) technique. The complete cDNA sequence of TLR2 was 2932 bp, containing an open reading frame (ORF) of 2469 bp encoding 822 amino acids. A multiple alignment analysis of the silvery pomfret TLR2 protein-coding sequence with other known TLR2 sequences from Oplegnathus fasciatus, Epinephelus coioides, Larimichthys crocea, Miichthys miiuy, Oreochromis niloticus, Paralichthys olivaceus, Trematomus bernacchii, Sparus aurata, and Chionodraco hamatus exhibited a high degree of homology of 78.83%, 75.91%, 74.21%, 74.94%, 71.95%, 72.57%, 73.68%, 75%, and 72.52 respectively, between these fish. Analysis of the TLR2 domain structures indicated that TLR2 from the silvery pomfret has the typical structural features of proteins that belong to the TLR family, including one transmembrane domain, eleven leucine-rich repeats (LRRs), and one Toll/IL-1 receptor homology domain (TIR). In vitro immunostimulation experiments revealed that Lactobacillus plantarum and Clostridium butyricum induce high levels of TLR2 mRNA and protein expression, but they induce only moderate levels of IL-8 and TNF-α production compared to Vibrio anguillarum. This suggests that TLR2 might play a vital role in the L. plantarum and C. butyricum-mediated immune response. In contrast, V. anguillarum significantly increased the secretion of IL-8 and TNF-α and induced cell apoptosis and necrosis. Due to the lower expression of TLR2 and higher levels of IL-8 and TNF-α induced by V. anguillarum, we hypothesize that a V. anguillarum infection is independent of the TLR2-induced production of pro-inflammatory cytokines. These results indicate that TLR2 may be involved in molecular interactions between the host and commensal bacteria, that exist in the silvery pomfret intestinal tract.

Ability of Lactobacillus plantarum lipoteichoic acid to inhibit Vibrio anguillarum-induced inflammation and apoptosis in silvery pomfret (Pampus argenteus) intestinal epithelial cells.

Lipoteichoic acid (LTA) is a major constituent of the cell wall of Gram-positive bacteria. The structure and immunomodulation of LTA vary greatly between different species. LTA from Lactobacillus plantarum has been shown to exert anti-pathogenic effects. Vibrio anguillarum is a major causative agent of vibriosis, one of the most prevalent fish diseases. The purpose of this study was to examine the effects of L. plantarum LTA on V. anguillarum growth, adhesion, and induced inflammation and apoptosis in intestinal epithelial cells of silvery pomfret (Pampus argenteus). Our results showed that L. plantarum LTA was unable to inhibit V. anguillarum growth; however, it significantly inhibited adhesion of V. anguillarum. It also showed significant inhibitory effects on EHEC-induced inflammation and apoptosis by modulating the expression of NF-κB (nuclear factor kappa B), IκB (inhibitor of NF-κB), Bcl2 (B-cell leukemia/lymphoma-2), BAX (Bcl-2-associated X protein), IL-8 (interleukin 8) and TNF-α (tumor necrosis factor-α), and via inhibition of caspase-9 and caspase-3 activation. These data extend our understanding of the beneficial effects of L. plantarum LTA, which is related to the inhibition of V. anguillarum, and suggest that L. plantarum LTA has potential as a new therapeutic agent against V. anguillarum-caused vibriosis in fish.

[Effect of Transplanting Time on Growth,Dry Matter Accumulation and Distribution,and Yield of Ligusticum chuanxiong].

Objective: To study the effect of transplanting time on growth, dry matter accumulation and distribution,and yield of Ligusticum chuanxiong,in order to provide theoretical and practical basis for the choice of cultivation methods. Methods: Ligusticum chuanxiong which planted in Pengshan were used as materials, and rearched in the field plot experiment. By the single factor randomized block design, the effects on growth development,dry matter accumulation and distribution of different periods of Ligusticum chuanxiong were analyzed. Results: Transplanting time has an important influence on the development of Ligusticum chuanxiong after lodging, and the lodging time was positively related to transplanting time; dry matter translocated to the underground part in January of the second year, and the dry matter translocated to the aerial parts in February, the dry matter secondary translocated to the underground part during March and April. Conclusion: Transplanting time affects the growth of Ligusticum chuanxiong,and affects the accumulation and distribution of every dry matter during the process of growth. Ligusticum chuanxiong transplant on August 31,and distribute more manure to improve the growth of aerial parts in the early stages,and spread additional manure to improve the growth of root stock at second year in shoot growth period,which can make a good harvest.

[Effects of Chemical Fertilizers and Organic Fertilizer on Yield of Ligusticum chuanxiong Rhizome].

OBJECTIVE: To study the effects of different N, P, K and organic fertilizer (OF) on yield of Ligusticum chuanxiong rhizome, in order to provide the theoretical foundation for the establishment of standardization cultivation techniques. METHODS: The field plot experiments used Ligusticum chuanxiong rhizome which planted in Pengshan as material, and were studied by the four factors and five levels with quadratic regression rotation-orthogonal combination design. According to the data obtained, a function model which could predict the fertilization and yield of Ligusticum chuanxiong rhizome accurately was established. RESULTS: The model analysis showed that the yields of Ligusticum chuanxiong rhizome were significantly influenced by the N, P, K and OF applications. Among these factors, the order of increase rates by the fertilizers was K > OF > N > P; The effect of interaction between N and K, N and OF, K and OF on the yield of Ligusticum chuanxiong rhizome were significantly different. High levels of N and P, N and organic fertilizer, K and organic fertilizer were conducive to improve the yield of Ligusticum chuanxiong rhizome. The results showed that the optimal fertilizer application rates of N was 148.20 - 172.28 kg/hm2, P was 511.92 - 599.40 kg/hm2, K was 249.70 - 282.37 kg/hm2, and OF was 940.00 - 1 104.00 kg/hm2. CONCLUSION: N, P, K and OF obviously affect the yield of Ligusticum chuanxiong rhizome. K and OF can significantly increase the yield of Ligusticum chuanxiong rhizome. Thus it is suggested that properly high mount of K and OF and appropriate increasing N are two favorable factors for cultivating Ligusticum chuanxiong.

[Quadratic Orthogonal Rotation Combination Design on Alisma orientalis of Fertilization].

OBJECTIVE: To study the effects of combined N, P, K and micronutrient fertilizers on the yield of Alisma orientalis tuber, and to optimize the fertilizer application rate. METHODS: Four factors five levels quadratic orthogonal rotation combination design was used. A function was established on nitrogen, phosphor, potassium and microelement fertilizer application rate with the yield of Alisma orientalis tuber. RESULTS: The established mathematical model was of high reliability for prediction with quadratic regression equation of R2 = 0. 8980. The order of increasing Alisma orientalis tuber yield was nitrogen > micronutrient fertilizer > potassium > phosphor. The results of the frequency analysis showed that for the target yield over 8 250 kg/hm2 and the confidence interval of 95%, the optimal fertilizer application rates were as follows :nitrogen of 241. 45 - 283. 55 kg/hm2, phosphor of 81. 14 - 208. 44 kg/hm2, potassium of 95. 57 - 239. 42 kg/hm2, and zinc fertilizer of 14. 32 - 16. 18 kg/hm2, boron fertilizer of 18. 84 - 19. 86 kg/hm2, and molybdenum fertilizer of 0. 151 -0. 159 kg/hm2 in micronutrient fertilizer. CONCLUSION: Nitrogen is related to the growth of Alisma orientalis, potassium promotes tuber bulking, micronutrient fertilizer consisted of zinc, boron and molybdenum fertilizer promotes Alisma orientalis growth and the absorption of nitrogen, phosphor and potassium. Moderate application of nitrogen, phosphorus, potassium, zinc, boron and molybdenum fertilizer can promote Alisma orientalis tuber yield. The nitrogen has the best effect.

Incorporating replacement free energy of binding-site waters in molecular docking.

Binding-site water molecules play a crucial role in protein-ligand recognition, either being displaced upon ligand binding or forming water bridges to stabilize the complex. However, rigorously treating explicit binding-site waters is challenging in molecular docking, which requires to fully sample ensembles of waters and to consider the free energy cost of replacing waters. Here, we describe a method to incorporate structural and energetic properties of binding-site waters into molecular docking. We first developed a solvent property analysis (SPA) program to compute the replacement free energies of binding-site water molecules by post-processing molecular dynamics trajectories obtained from ligand-free protein structure simulation in explicit water. Next, we implemented a distance-dependent scoring term into DOCK scoring function to take account of the water replacement free energy cost upon ligand binding. We assessed this approach in protein targets containing important binding-site waters, and we demonstrated that our approach is reliable in reproducing the crystal binding geometries of protein-ligand-water complexes, as well as moderately improving the ligand docking enrichment performance. In addition, SPA program (free available to academic users upon request) may be applied in identifying hot-spot binding-site residues and structure-based lead optimization.

Growth, feed intake and immune responses of orange-spotted grouper (Epinephelus coioides) exposed to low infectious doses of ectoparasite (Cryptocaryon irritans).

To explore the effect of low-dose Cryptocaryon irritans infection on growth, feeding and antiparasitic immunity of orange-spotted grouper (Epinephelus coioides), this study utilized C. irritans at concentrations of 5500 theronts/fish (Group I, 1/10 of 96 h LC50) or 11,000 theronts/fish (Group II) to infect E. coioides weighing 38 g on average at week 0, 2 and 4, respectively. Food consumption was recorded daily; the fish were weighed weekly; serum immobilizing titer (SIT), and acid phosphatase (ACP), alkaline phosphatase (AKP), superoxide dismutase (SOD), lysozyme (LZM) activity were recorded every 2 weeks; the fish were treated with lethal dose (70,000 theronts/fish) of C. irritans in the 8th week and death number were recorded. The result shows that in the 1st week after the first infection, the fish's weight gain (WG), length gain (LG), and specific growth rate (SGR) dropped as parasite dose increased, and WG, SGR values were negative; while, after the 2nd and the 3rd infection, no significant differences were detected among the three groups. These results indicated that the 1st infection affected the fish most, while the following infections were protected by some immunity. In the 3rd, 7th, and 8th week, condition factor (CF) increased with the increased infectious dose, indicating that the parasite affected body length more than body weight. As the experiment went on, accumulated food consumption (AFC) of all three groups steadily grew (control > Group I > Group II). But on the 2nd day after the first infection, daily food consumption (DFC) of Group I and II significantly dropped, the decline of Group II was greater than that of Group I, DFC recovered in the following week, with Group I earlier than Group II. After the 2nd infection, DFC of Group I and II dropped again, Group II still dropped more than Group I, and both groups recovered on the 3rd day after infection. The 3rd infection caused no significant difference in week food consumption (WFC). These results indicated that a higher dose of infection causes a greater drop in FC and a slower recovery. Weekly feed conversion ratio (WFCR) values of Group I and II in the 1st week was negative; in the 2nd week, WFCR was lower in the group infected by a higher dose of parasite; while in the 3rd and following weeks, no significant pattern was observed. Accumulate feed conversion ratio (AFCR) dropped as the infectious dose increased (control > Group I > Group II), AFCR of Group I and II reached above 0 in the 2nd and 4th week, respectively. From the 4th week on, the inter-group AFCR of the 3 groups still took on a declining trend with the increased infectious dose but the gap became smaller. One week after the first infection, SIT of Group I and Group II were 0; one week after the 2nd infection, SIT reached up to 8 (Group I) and 16 (Group II) respectively; and after the 3rd infection, SIT further increased and peaked in the 7th week. When challenged by lethal dose of C. irritans, fish of all 3 groups began to die since the 3rd day after infection, and the final deaths were 14, 12 and 8 for the control group, Group I and Group II, respectively. ACP activity in the 1st, 5th, 7th but the 3rd week was higher in the experiment group than that in the control group, but no significant difference was detected between Group I and II throughout the experiment. AKP activity increased as the infectious dose increased, but the difference among the three groups gradually became less obvious in latter infections, and no significant difference can be detected in the end. SOD activity increased with infection dose at each time point, while both group I and group II had their SOD activities first increased and then decreased as times of infection increased. The LZM activity of the two infection groups increased as the infectious times increased. Combining the results on growth and feeding, we speculated that the fish's physiological condition stabilized after 3 rounds of infection. To sum up, low-dose infection by C. irritans can induce the fish's immunity, but at the cost of decreasing food intake, decreased food conversion, and lagged growth.

Effects of Cryptocaryon irritans infection on the survival, feeding, respiratory rate and ionic regulation of the marbled rockfish Sebastiscus marmoratus.

To clarify the effects of a Cryptocaryon irritans infection on the physiological functions of the marbled rockfish Sebastiscus marmoratus, this study utilized C. irritans at concentrations of 2500; 5000; 7500; 10,000; 20,000; and 30,000 theronts/fish to infect marbled rockfish weighing 45 ± 3 g. The survival rate, food intake, respiratory rate, serum ion concentrations and gill Na+/K+-ATPase activity were determined. With the increase of the infection concentration and the passage of time, the survival rate of the rockfish gradually decreased. The groups infected with more than 5000 theronts/fish had stopped feeding within 4 days. The respiratory rates of the fish in the groups infected with 2500 and 5000 theronts/fish initially increased and then decreased. In contrast, the respiratory rate of the fish in the groups infected with more than 7500 theronts/fish was elevated to levels significantly higher than the control group after 12 h. The Na+/K+-ATPase activity and serum Na+ and Cl- concentrations increased with increasing infection concentration. In conclusion, the physiological functions of the fish infected with low concentrations of C. irritans can be effectively restored, whereas a high concentration infection induced severe stress. The declined food intake and accelerated respiratory rate could be useful for an early warning system as important indicators.

Transcriptomic Analysis of the Hepatopancreas in the Sex-Related Size Differences of Macrobrachium nipponense.

Macrobrachium nipponense, a commercially popular crustacean species within the Chinese context, is recognized for its exceptional nutritional composition and palatability. There are significant differences in growth between male and female M. nipponense. Herein, transcriptomics was used to determine the hepatopancreas transcriptome differences between sex-related size differences in M. nipponense. We identified 974 differentially expressed genes (DEGs) between the SHE (female) and BHE (male) groups, which were validated using RT-qPCR. The genes encoding matrix metalloproteinase-9 (MM9), Ribosome-binding protein 1 (RBP1), Aly/REF export factor 2, and hematological and neurological expressed 1 (HN1) may play a role in modulating the sex-related size differences observed in M. nipponense. Clusters of orthologous groups and gene ontology functional analysis demonstrated that the DEGs for sex-related size in M.nipponense were associated with various biological functions. The Kyoto Encyclopedia of Genes and Genomes pathways analysis demonstrated that upregulated DEGs were mainly enriched in lysine biosynthesis, tryptophan metabolism, and lysine degradation pathways, whereas the downregulated DEGs were mainly enriched in ascorbate and aldarate metabolism, retinol metabolism, and drug metabolism-cytochrome P450 pathways. The results indicated the molecular mechanism underlying the sex-related size differences and identified key genes. This data will be invaluable to support explanations of individual differences between male and female prawns.

Metagenomic and Metabolomic Analyses Reveal the Role of Gut Microbiome-Associated Metabolites in the Muscle Elasticity of the Large Yellow Croaker (Larimichthys crocea).

The large yellow croaker (LYC, Larimichthys crocea) is highly regarded for its delicious taste and unique flavor. The gut microbiota has the ability to affect the host muscle performance and elasticity by regulating nutrient metabolism. The purpose of this study is to establish the relationship between muscle quality and intestinal flora in order to provide reference for the improvement of the muscle elasticity of LYC. In this study, the intestinal contents of high muscle elasticity males (IEHM), females (IEHF), and low muscle elasticity males (IELM) and females (IELF) were collected and subjected to metagenomic and metabolomic analyses. Metagenomic sequencing results showed that the intestinal flora structures of LYCs with different muscle elasticities were significantly different. The abundance of Streptophyta in the IELM (24.63%) and IELF (29.68%) groups was significantly higher than that in the IEHM and IEHF groups. The abundance of Vibrio scophthalmi (66.66%) in the IEHF group was the highest. Based on metabolomic analysis by liquid chromatograph-mass spectrometry, 107 differentially abundant metabolites were identified between the IEHM and IELM groups, and 100 differentially abundant metabolites were identified between the IEHF and IELF groups. Based on these metabolites, a large number of enriched metabolic pathways related to muscle elasticity were identified. Significant differences in the intestinal metabolism between groups with different muscle elasticities were identified. Moreover, the model of the relationship between the intestinal flora and metabolites was constructed, and the molecular mechanism of intestinal flora regulation of the nutrient metabolism was further revealed. The results help to understand the molecular mechanism of different muscle elasticities of LYC and provide an important reference for the study of the mechanism of the effects of LYC intestinal symbiotic bacteria on muscle development, and the development and application of probiotics in LYC.

Recombinase-aid amplification combined with lateral flow detection assay for sex identification of the great white pelican (Pelecanus onocrotalus).

Sex identification in avian species is essential for biodiversity conservation and ecological studies. However, the sex of nearly half of the birds could not be identified based on their external appearance. It is difficult to visually identify sex to monitor the ecology and conservation of wild populations. In this study, we designed primer pairs for large white pelican using recombinase-based isothermal amplification combined with a lateral flow dipstick (RAA-LFD) assay for chromo-helicase-DNA binding protein (CHD) genes mapped to W chromosomes and an ultra-conserved element (UCE) located on chromosome 6, respectively. Our result showed that the raaW4-RAA-LFD can detect up to 0.1 ng of genomic DNA (gDNA) templates of female pelicans in 30 min at 39 ℃ and accurately distinguish female from male without any cross reactivity. RaaUCE2-RAA-LFD can amplify both male and female pelicans with a detection limit of 25 pg. To further evaluate the assay, 15 white pelicans of unknown sex were tested using the RAA-LFD assay and conventional polymerase chain reaction (PCR). The results of the raaW4-RAA-LFD assay were consistent with those of the conventional PCR. The developed RAA-LFD assay is equipped with field-deployable instruments and offers a field platform for rapid and reliable sex identification in pelicans.

Genomic evolution of island birds from the view of the Swinhoe's pheasant (Lophura swinhoii).

Island endemic birds account for the majority of extinct vertebrates in the past few centuries. To date, the evolutionary characteristics of island endemic bird's is poorly known. In this research, we de novo assembled a high-quality chromosome-level reference genome for the Swinhoe's pheasant, which is a typical endemic island bird. Results of collinearity tests suggest rapid ancient chromosome rearrangement that may have contributed to the initial species radiation within Phasianidae, and a role for the insertions of CR1 transposable elements in rearranging chromosomes in Phasianidae. During the evolution of the Swinhoe's pheasant, natural selection positively selected genes involved in fecundity and body size functions, at both the species and population levels, which reflect genetic variation associated with island adaptation. We further tested for variation in population genomic traits between the Swinhoe's pheasant and its phylogenetically closely related mainland relative the silver pheasant, and found higher levels of genetic drift and inbreeding in the Swinhoe's pheasant genome. Divergent demographic histories of insular and mainland bird species during the last glacial period may reflect the differing impact of insular and continental climates on the evolution of species. Our research interprets the natural history and population genetic characteristics of the insular endemic bird the Swinhoe's pheasant, at a genome-wide scale, provides a broader perspective on insular speciation, and adaptive evolution and contributes to the genetic conservation of island endemic birds.