Actinoplanes sandaracinus sp. nov., a novel ligninase- producing and cellulose-degrading actinobacterium isolated from soil.

A novel ligninase-producing and cellulose-degrading actinobacterium, designated strain NEAU-A12T, was isolated from a soil sample collected from Aohan banner, Chifeng City, Inner Mongolia Autonomous Region, PR China. A polyphasic taxonomic study was used to establish the status of strain NEAU-A12T. 16S rRNA gene sequence analysis revealed that strain NEAU-A12T belonged to the genus Actinoplanes and showed the highest similarity (98.3 %) to Actinoplanes palleronii DSM 43940T, while showing less than 98.3 % similarity to other members of the genus Actinoplanes. The phospholipid profile contained diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol and glycosylphosphatidylinositol. The diagnostic sugars in cell hydrolysates were determined to be arabinose, glucose and xylose. The cell wall contained meso-diaminopimelic acid as the diagnostic diamino acid. The predominant menaquinones were MK-9(H4), MK-9(H6) and MK-9(H2). The major fatty acids were C15 : 0, C16 : 0, C16 : 1 ω7c and C17 : 0. Meanwhile, genomic analysis revealed a genome size of 10 192 524 bp and a DNA G+C content of 70.6 mol%, and indicated that strain NEAU-A12T had the potential to degrade lignin and cellulose, as well as produce bioactive compounds. In addition, the average nucleotide identity values between strain NEAU-A12T and its reference strains A. palleronii DSM 43940T, Actinoplanes regularis DSM 43151T, Actinoplanes philippinensis DSM 43019T, Actinoplanes xinjiangensis DSM 45184T and Actinoplanes italicus DSM 43146T were 80.3, 80.3, 84.1, 84.3 and 84.0 %, respectively. The levels of digital DNA-DNA hybridization between them were found to be 23.6 % (21.3-26.1 %), 23.8 % (21.5-26.3 %), 28.3 % (25.9-30.8 %), 28.6 % (26.0-30.9 %) and 28.4 % (26.2-31.1 %), respectively. Based on phenotypic, chemotaxonomic and genotypic data, strain NEAU-A12T is considered to represent a novel species of the genus Actinoplanes, for which the name Actinoplanes sandaracinus sp. nov. is proposed, with NEAU-A12T (=CCTCC AA 2020039T=DSM 112043T) as the type strain.

Comprehensive analysis on the regulation of differentially expressed of mRNA and ncRNA in different ovarian stages of ark shell Scapharca broughtonii.

BACKGROUND: Ovarian development is an important prerequisite and basis for animal reproduction. In many vertebrates, it is regulated by multiple genes and influenced by sex steroid hormones and environmental factors. However, relative information is limited in shellfish. To explore the biological functions and molecular mechanisms of mRNA and non-coding RNA that regulate ovarian development in Scapharca broughtonii, we performed whole transcriptome sequencing analysis on ovaries at three developmental stages. Furthermore, the biological processes involved in the differential expression of mRNA and ncRNA were analyzed. RESULTS: A total of 11,342 mRNAs, 6897 lncRNAs, 135 circRNAs, and 275 miRNAs were differentially expressed. By mapping the differentially expressed RNAs from the three developmental stages of Venn diagram, multiple groups of shared mRNAs and lncRNAs were found to be associated with ovarian development, with some mRNA and ncRNA functions associated with steroid hormone. In addition, we constructed and visualized the lncRNA/circRNA-miRNA-mRNA network based on ceRNA targeting relationships. CONCLUSIONS: These findings may facilitate our further understanding the mRNA and ncRNAs roles in the regulation of shellfish reproduction.

MicroRNA expression signature in the striated and smooth adductor muscles of Yesso scallop Patinopecten yessoensis.

Increasing evidences point to the potential role of microRNAs (miRNAs) in muscle growth and development in animals. However, knowledge on the identity of miRNAs and their targets in molluscs remains largely unknown. Scallops have one large adductor muscle, composed of fast (striated) and slow (smooth) muscle types, which display great differences in muscle fibers, meat quality, cell types and molecular components. In the present study, we performed a comprehensive investigation of miRNA transcriptomes in fast and slow adductor muscles of Yesso scallop Patinopecten yessoensis. As a result, 47 differentially expressed miRNAs representing ten miRNA families were identified between the striated and smooth adductor muscles. The KEGG enrichment analysis of their target genes were mainly associated with amino acid metabolism, energy metabolism and glycan biosynthesis. The target genes of miR-133 and miR-71 were validated by the dual-luciferase reporter assays and miRNA antagomir treatment in vivo. The identification and functional validation of these different miRNAs in scallops will greatly help our understanding of miRNA regulatory mechanism that achieves the unique muscle phenotypes in scallops. The present findings provide the direct evidences for muscle-specific miRNAs involved in muscle growth and differentiation in molluscs.

Mice and Rats Exhibit Striking Inter-species Differences in Gene Response to Acute Stroke.

Neuroprotection in acute stroke has not been successfully translated from animals to humans. Animal research on promising agents continues largely in rats and mice which are commonly available to researchers. However, controversies continue on the most suitable species to model the human situation. Generally, putative agents seem less effective in mice as compared with rats. We hypothesized that this may be due to inter-species differences in stroke response and that this might be manifest at a genetic level. Here we used whole-genome microarrays to examine the differential gene regulation in the ischemic penumbra of mice and rats at 2 and 6 h after permanent middle cerebral artery occlusion (pMCAO; Raw microarray CEL data files are available in the GEO database with an accession number GSE163654). Differentially expressed genes (adj. p ≤ 0.05) were organized by hierarchical clustering, correlation plots, Venn diagrams and pathway analyses in each species and at each time-point. Emphasis was placed on genes already known to be associated with stroke, including validation by RT-PCR. Gene expression patterns in the ischemic penumbra differed strikingly between the species at both 2 h and 6 h. Nearly 90% of significantly regulated genes and most pathways modulated by ischemia differed between mice and rats. These differences were evident globally, among stroke-associated genes, immediate early genes, genes implicated in stress response, inflammation, neuroprotection, ion channels, and signal transduction. The findings of this study may have significant implications for the choice of species for screening putative stroke therapies.

Nocardia rosealba sp. nov., a novel ligninase-producing Actinobacterium isolated from soil.

A novel ligninase-producing actinomycete, designated strain NEAU-G4T, was isolated from a soil sample and subjected to a polyphasic taxonomic study to establish its status. According to 16S rRNA gene sequence comparisons, the isolate was identified as a member of the genus Nocardia, with the highest sequence similarity to Nocardia ignorata DSM 44496T (99.2 %). The whole-cell sugars contained galactose and arabinose. The amino acid of the cell wall was determined to be meso-diaminopimelic acid. The major fatty acids (>10 %) were C16 : 0, C18 : 1 ω9c, C18 : 0 and C16 : 1 ω7c. The predominant menaquinone was identified as MK-8(H6, ω-cycl). The major polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylinositol. Strain NEAU-G4T had a draft genome size of 6 405 167 bp, annotated with 5815 protein-coding genes. The DNA G+C content was 67.6 mol%. Phylogenetic analysis using the 16S rRNA gene and whole-genome sequences showed that strain NEAU-G4T formed a stable phyletic line with N. ignorata DSM 44496T. The digital DNA-DNA hybridization and average nucleotide identity values between them were 63.7 % (60.8-66.5 %) and 95.5 %, respectively. Moreover, genomic analysis indicated that strain NEAU-G4T had the potential to degrade lignin and produce bioactive compounds. On the basis of genotypic analysis, physiological data, as well as phenotypic and chemotaxonomic characterizations, it is concluded that the organism be classified as representing a novel species of the genus Nocardia, for which the name Nocardia rosealba sp. nov. is proposed. The type strain is NEAU-G4T (=CCTCC AA 2020038T=DSM 111936T).

Saccharothrix luteola sp. nov., a novel cellulose-degrading actinobacterium isolated from soil and emended description of the genus Saccharothrix.

A novel cellulose-degrading actinobacterium, designated strain NEAU-S10T, was isolated from soil collected from Chifeng, Inner Mongolia Autonomous Region, PR China, and characterized using a polyphasic approach. Pairwise similarity of the 16S rRNA gene sequence showed that strain NEAU-S10T was a representative of Saccharothrix and was closely related to Saccharothrix carnea NEAU-yn17T (99.2 %), Saccharothrix saharensis SA152T (99.0 %), Saccharothrix texasensis DSM 44231T (98.5 %) and Saccharothrix xinjiangensis NBRC 101911T (98.5 %). Physiological and chemotaxonomic characteristics of the strain further supported its affiliation to the genus Saccharothrix. The whole-cell sugars contained galactose, ribose and mannose. The polar lipids contained diphosphatidylglycerol, phosphatidylmonomethylethanolamine, phosphatidylethanolamine, phosphatidylinositol and phosphatidylinositol mannoside. The predominant menaquinones were MK-9(H0), MK-9(H2), MK-9(H4) and MK-10(H4). The major fatty acids were iso-C16 : 0, C16 : 0, anteiso-C17 : 0, iso-C15 : 0 and iso-C17 : 0. The genomic DNA G+C content was 71.8 mol%. The levels of digital DNA-DNA hybridization between isolate and S. carnea NEAU-yn17T, S. saharensis SA152T and S. texasensis DSM 44231T were 40.1 % (37.6-42.6 %), 38.soap8 % (36.3-41.3 %) and 44.8 % (42.2-47.3 %) and the ANI values between them were determined to be 90.2, 89.8 and 91.7 %, the results indicated that strain NEAU-S10T could be distinguished from its reference strains. The assembled genome sequence of strain NEAU-S10T was found to be 10 305 394 bp long. The NCBI Prokaryotic Genome Annotation Pipeline (PGAP) revealed 8 994 protein-coding genes. Genomic analysis and Congo red staining test indicated that strain NEAU-S10T had the potential to degrade cellulose. The genomic and phenotypic results indicate that strain NEAU-S10T represents a novel species of the genus Saccharothrix, for which the name Saccharothrix luteola sp. nov. is proposed, with NEAU-S10T (=CCTCC AA 2020037T=JCM 34800T) as the type strain.

Cell type diversity in scallop adductor muscles revealed by single-cell RNA-Seq.

Studies on cell atlas in marine invertebrates provide a better understanding of cell types, stem cell maintenance, and lineages of cell differentiation. To investigate the molecular features of various cell types in molluscan muscles, we performed single-cell RNA sequencing (scRNA-seq) to map cell types in scallop adductor muscles. We uncovered the cell type-specific features of 20 cell clusters defined by the expression of multiple specific molecular markers. These cell clusters are mainly classified into four broad classes, including mesenchymal stem cells, muscle cells, neurons, and haemolymph cells. In particular, we identified a diverse repertoire of neurons in the striated adductor muscle, but not in the smooth muscle. We further reconstructed the cell differentiation events using all the cell clusters by single-cell pseudotemporal trajectories. By integrating dual BrdU-PCNA immunodetection, neuron-specific staining and electron microscopy observation, we showed the spatial distribution of mesenchymal stem cells and neurons in striated adductor muscle of scallops. The present findings will not only be useful to address the cell type-specific gene expression profiles in scallop muscles, but also provide valuable resources for cross-species comparison of marine organisms.

Actinoplanes flavus sp. nov., a novel cellulase-producing actinobacterium isolated from coconut palm rhizosphere soil.

A novel cellulase-producing actinomycete, designated strain NEAU-H7T, was isolated from coconut palm rhizosphere soil collected from Wenchang City, Hainan Province, PR China. A polyphasic taxonomic study was carried out to establish the status of this strain. Results of 16S rRNA gene sequence analysis indicated that strain NEAU-H7T belonged to the genus Actinoplanes, with highest similarity to Actinoplanes hulinensis NEAU-M9T (99.2 % 16S rRNA gene sequence similarity). The diagnostic sugars in cell hydrolysates were determined to be ribose, galactose and mannose. The major fatty acids (>10%) were C16 : 0, C18 : 1 ω9c and C18 : 0. The predominant menaquinones were identified as MK-9(H4) and MK-9(H6). The major polar lipids were phosphatidylethanolamine, phosphatidylinositol and two phosphatidylinositol mannosides. The amino acid of the cell-wall peptidoglycan was determined to be meso-diaminopimelic acid. The DNA G+C content was 71.2 mol%. Phylogenetic analysis using 16S rRNA gene sequences showed that strain NEAU-H7T formed a stable phyletic line with A. hulinensis NEAU-M9T. However, whole-genome phylogeny showed strain NEAU-H7T formed a stable phyletic line with A. hulinensis NEAU-M9T (99.2%), Actinoplanes campanulatus DSM 43148T (98.6%), Actinoplanes capillaceus DSM 44859T (98.3%) and Actinoplanes lobatus DSM 43150T (97.6%). The digital DNA-DNA hybridization (dDDH) results between them were 53.6 (50.9-56.2), 54.1 (51.3-56.9), 53.1 (50.3-55.9) and 52.9 % (50.1-55.6 %), and whole-genome average nucleotide identity (ANI) values between them were 93.7, 93.6, 93.5 and 93.5 %. The low dDDH and ANI values demonstrated that strain NEAU-H7T could be distinguished from its reference strains. Moreover, genomic analysis indicated that the strain NEAU-H7T had the potential to decompose cellulose and produce bioactive compounds. On the basis of morphological, chemotaxonomic and phylogenetic characteristics, strain NEAU-H7T is proposed to represent a novel species of the genus Actinoplanes, with the name Actinoplanes flavus sp. nov. The type strain is NEAU-H7T (=CCTCC AA 2020034T=DSM 112042T).

Lentzea alba sp. nov., a novel actinobacterium isolated from soil.

A novel actinobacterium, designated strain NEAU-D13T, was isolated from soil collected from Aohan Banner, Chifeng, Inner Mongolia Autonomous Region, China and characterized using a polyphasic approach. On the basis of 16S rRNA gene sequence analysis, strain NEAU-D13T belonged to the genus Lentzea and shared the highest sequence similarity with Lentzea kentuckyensisJCM 14913T (99.17 %). Morphological and chemotaxonomic characteristics of the strain also supported its assignment to the genus Lentzea. Cell walls contained meso-diaminopimelic acid as the diagnostic diamino acid and the whole-cell sugars were ribose and mannose. The phospholipid profile contained diphosphatidylglycerol, phosphatidylethanolamine, hydroxyphosphatidylethanolamine and phosphatidylinositol. The menaquinone was only MK-9(H4). The major fatty acids were iso-C16:0, C16:0, anteiso-C17:0, iso-C15:0 and anteiso-C15:0. DNA G+C content was 68.71 mol%. Phylogenetic analysis using the 16S rRNA gene sequences showed that the strain formed a stable clade with L. kentuckyensisJCM 14913T in the genus Lentzea. Meanwhile, a combination of digital DNA-DNA hybridization results and some phenotypic characteristics demonstrated that strain NEAU-D13T could be distinguished from its closely related strain. Therefore, it is concluded that strain NEAU-D13T represents a novel species of the genus Lentzea, for which the name Lentzea alba sp. nov. is proposed, with NEAU-D13T (=CCTCC AA 2019089T=JCM 33970T) as the type strain.

Streptomyces botrytidirepellens sp. nov., a novel actinomycete with antifungal activity against Botrytis cinerea.

The fungal pathogen Botrytis cinerea is the causal agent of devastating gray mold diseases in many economically important fruits, vegetables, and flowers, leading to serious economic losses worldwide. In this study, a novel actinomycete NEAU-LD23T exhibiting antifungal activity against B. cinerea was isolated, and its taxonomic position was evaluated using a polyphasic approach. Based on the genotypic, phenotypic and chemotaxonomic data, it is concluded that the strain represents a novel species within the genus Streptomyces, for which the name Streptomyces botrytidirepellens sp. nov. is proposed. The type strain is NEAU-LD23T (=CCTCC AA 2019029T=DSM 109824T). In addition, strain NEAU-LD23T showed a strong antagonistic effect against B. cinerea (82.6±2.5%) and varying degrees of inhibition on nine other phytopathogenic fungi. Both cell-free filtrate and methanol extract of mycelia of strain NEAU-LD23T significantly inhibited mycelial growth of B. cinerea. To preliminarily explore the antifungal mechanisms, the genome of strain NEAU-LD23T was sequenced and analyzed. AntiSMASH analysis led to the identification of several gene clusters responsible for the biosynthesis of bioactive secondary metabolites with antifungal activity, including 9-methylstreptimidone, echosides, anisomycin, coelichelin and desferrioxamine B. Overall, this research provided us an excellent strain with considerable potential to use for biological control of tomato gray mold.

Actinoplanes aureus sp. nov., a novel protease-producing actinobacterium isolated from soil.

A novel protease-producing actinobacterium, designated strain NEAU-A11T, was isolated from soil collected from Aohan banner, Chifeng, Inner Mongolia Autonomous Region, China, and characterised using a polyphasic approach. The hydrolytic enzymes, such as proteases, played critical roles in destruction of fungi by degrading the protein linkages to disrupt integrity in the cell wall. This suggested that the isolate could be a good biocontrol candidate against pathogens to control fungal diseases. On the basis of 16S rRNA gene sequence analysis, strain NEAU-A11T was indicated to belong to the genus Actinoplanes and was most closely related to Actinoplanes rectilineatus JCM 3194 T (98.9%). Cell walls contained meso-diaminopimelic acid as the diagnostic diamino acid and the whole-cell sugars were arabinose, xylose and glucose. The phospholipid profile contained diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol and two phosphatidylinositol mannosides. The predominant menaquinones were MK-9(H4), MK-9(H6) and MK-9(H8). The major fatty acids were C18:0, C16:0, C18:1 ω9c, C17:0 and C15:0. Genome sequencing revealed a genome size of 10,742,096 bp, a G + C content of 70.5% and 9,514 protein-coding genes (CDS), including 102 genes coding for protease. Moreover, Genome analysis showed that strain NEAU-A11T contained 255 glycoside hydrolases (GHs), 152 glycosyl transferases (GTs), 40 carbohydrate esterases (CEs), 26 polysaccharide lyases (PLs), and 12 auxiliary activities (AAs) genes. Genome mining analysis using antiSMASH 5.0 led to the identification of 20 putative gene clusters responsible for the production of diverse secondary metabolites. Phylogenetic analysis using the 16S rRNA gene sequences showed that the strain formed a stable clade with A. rectilineatus JCM 3194 T in the genus Actinoplanes. Whole-genome phylogeny showed strain NEAU-A11T formed a stable phyletic line with Actinoplanes lutulentus DSM 45883 T (97.6%). However, whole-genome average nucleotide identity value between strain NEAU-A11T and its reference strains A. rectilineatus JCM 3194 T and A. lutulentus DSM 45883 T were found to be 81.1% and 81.6%, respectively. The levels of digital DNA-DNA hybridization between them were 24.6% (22.2-27.0%) and 24.8% (22.5-27.3%), respectively. The values were well below the criteria for species delineation of 70% for dDDH and 95-96% for ANI, suggesting that the isolate differed genetically from its closely related type strain. The content of G + C in genomic DNA was 70.5%, within the range of 67-76%. In addition, evidences from phenotypic, chemotaxonomic and genotypic studies indicated that strain NEAU-A11T represents a novel species of the genus Actinoplanes, for which the name Actinoplanes aureus sp. nov. is proposed, with NEAU-A11T (= CCTCC AA 2019063 T = JCM 33971 T) as the type strain.

Micromonospora rubida sp. nov., a novel actinobacterium isolated from soil of Harbin.

A novel actinobacterium, designated strain NEAU-HG-1T, was isolated from soil collected from Harbin, Heilongjiang Province, Northeast China and characterised using a polyphasic approach. On the basis of 16S rRNA gene sequence analysis, strain NEAU-HG-1T belonged to the genus Micromonospora, and shared high sequence similarities with Micromonospora auratinigra DSM 44815T (98.9%) and Micromonospora coerulea DSM 43143T (98.7%). Morphological and chemotaxonomic characteristics of the strain also supported its assignment to the genus Micromonospora. Cell wall contained meso-diaminopimelic acid and the whole-cell sugars were arabinose and xylose. The polar lipid contained diphosphatidylglycerol, phosphatidylethanolamine, glycolipid and phosphatidylinositol. The predominant menaquinones were MK-10(H2), MK-10(H4) and MK-10(H6). The major fatty acids were C17:0 cycle, iso-C15:0, and iso-C16:0. Furthermore, strain NEAU-HG-1T displayed a DNA-DNA relatedness of 33.8 ± 2.2% with M. coerulea DSM 43143T. The level of digital DNA-DNA hybridization between strain NEAU-HG-1T and M. auratinigra DSM 44815T was 27.2% (24.8-29.7%). The value was well below the criteria for species delineation of 70% for dDDH. Whole-genome average nucleotide identity analyses result also indicated that the isolate should be assigned to a new species under the genus Micromonospora. Therefore, it is concluded that strain NEAU-HG-1T represents a novel species of the genus Micromonospora, for which the name Micromonospora rubida sp. nov. is proposed, with NEAU-HG-1T (= CGMCC 4.7479T = JCM 32386T) as the type strain.

Transcriptomics analysis revealing candidate genes and networks for sex differentiation of yesso scallop (Patinopecten yessoensis).

BACKGROUND: The Yesso scallop, Patinopecten (Mizuhopecten) yessoensis, is a commercially important bivalve in the coastal countries of Northeast Asia. It has complex modes of sex differentiation, but knowledge of the mechanisms underlying this sex determination and differentiation is limited. RESULTS: In this study, the gonad tissues from females and males at three developmental stages were used to investigate candidate genes and networks for sex differentiation via RNA-Req. A total of 901,980,606 high quality clean reads were obtained from 18 libraries, of which 417 expressed male-specific genes and 754 expressed female-specific genes. Totally, 10,074 genes differentially expressed in females and males were identified. Weighted gene co-expression network analysis (WGCNA) revealed that turquoise and green gene modules were significantly positively correlated with male gonads, while coral1 and black modules were significantly associated with female gonads. The most important gene for sex determination and differentiation was Pydmrt 1, which was the only gene discovered that determined the male sex phenotype during early gonadal differentiation. Enrichment analyses of GO terms and KEGG pathways revealed that genes involved in metabolism, genetic and environmental information processes or pathways are sex-biased. Forty-nine genes in the five modules involved in sex differentiation or determination were identified and selected to construct a gene co-expression network and a hypothesized sex differentiation pathway. CONCLUSIONS: The current study focused on screening genes of sex differentiation in Yesso scallop, highlighting the potential regulatory mechanisms of gonadal development in P. yessoensis. Our data suggested that WCGNA can facilitate identification of key genes for sex differentiation and determination. Using this method, a hypothesized P. yessoensis sex determination and differentiation pathway was constructed. In this pathway, Pydmrt 1 may have a leading function.

Ark shell Scapharca broughtonii hemocyte response against Vibrio anguillarum challenge.

Scapharca broughtonii is one of the most important Arcidae aquaculture species in the Asia-Pacific region. We aimed to investigate the immune responses of hemocytes from ark shell S. broughtonii hemolymph against pathogens. Hemocyte ultrastructure and immunological activity in response to Vibrio anguillarum challenge were observed by scanning and transmission electron microscopy. Before ultrastructure observation, we used the API ZYM semi-quantitative kit to evaluate the levels of hydrolytic enzymes in the plasma and hemocytes following V. anguillarum infection. An enzyme-linked immunosorbent assay kit was used to investigate the variation in the lysozyme activity and hemocytes following bacterial infection. The results showed that hemocytes were the main defense cells against bacterial infection, whereas plasma played a role in the transport and support of hemocytes. It was presumed that an important function of lysozymes and hydrolytic enzymes in lysosomes was for bacterial digestion. Three major types of hemocytes were observed, namely, red blood cells (RBCs), white blood cells (WBCs), and thrombocytes (TCs). Scanning electron microscopy showed that the normal RBCs appeared pie-shaped with 10 µm diameter and 4 µm central thickness, whereas WBCs were spherical in shape with varying sizes, 4-8 µm diameter, and included small lymphocytes. TCs were long, spindle-shaped, and 12-20 µm in length. The cell membrane surface was smooth and even for all cells before pathogen challenge. Under transmission electron microscopy, RBCs displayed a limited ability to devour and digest bacteria adherent to the cell surface following infection. Many hemoglobin particles were observed in the RBC cytoplasm. WBCs were very active against bacterial invasion and showed a strong ability to digest and decompose infected and wrapped V. anguillarum through phagocytosis and lysosome fusion. Digestive vacuoles rapidly became transparent and were thought to contain increasing quantities of pathogen-induced lysozymes. WBCs that devoured pathogenic bacteria were prone to deformation as well as adhesion to each other. TCs were rich in endoplasmic reticulum (ER) content in their cytoplasm and were widely connected in a net-shaped structure. Mitochondria in TCs formed clusters upon invasion of V. anguillarum in the hemolymph. TCs disintegrated to release the ER into the plasma to form a mesh that facilitated clotting. The ability of circulating hemocytes to quickly modify their morphologies and stainability suggests that S. broughtonii is endowed with highly dynamic hemocyte populations capable of coping with environmental changes and rapidly growing pathogens.

Pseudonocardia tritici sp. nov., a novel actinomycete isolated from rhizosphere soil of wheat (Triticum aestivum L.).

An aerobic, non-motile, Gram-stain positive actinomycete, designated strain NEAU-YY211T, was isolated from the rhizosphere soil of wheat (Triticum aestivum L.) collected from Zhumadian, Henan Province, mid-eastern China, and characterised taxonomically using a polyphasic approach. Comparative analysis of the 16S rRNA gene sequence indicated that strain NEAU-YY211T belongs to the genus Pseudonocardia, showing high similarities with respect to Pseudonocardia ammonioxydans H9T (99.1%) and Pseudonocardia antitumoralis SCSIO 01299T (99.0%), respectively. The cell wall was found to contain meso-diaminopimelic acid and the whole cell sugars were identified as arabinose and galactose. The predominant menaquinone of strain NEAU-YY211T was identified as MK-8(H4) and the major fatty acids were identified as iso-C16:0, C17:1ω8c and iso-C16:1. The phospholipid profile was found to consist of diphosphatidylglycerol, phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, phosphatidylinositol and an unidentified phospholipid. The G+C content of the genomic DNA was determined to be 72.6 mol%. Levels of DNA-DNA relatedness with P. ammonioxydans JCM 12462T and P. antitumoralis DSM 45322T were 54.5 ± 3.5% and 49.8 ± 2.5% (mean ± SD), respectively. Based on phylogenetic analysis, phenotypic and genotypic data, it is concluded that the isolate can be distinguished from closely related type strains and thus represents a novel species of the genus Pseudonocardia, for which the name Pseudonocardia tritici sp. nov. is proposed. The type strain is NEAU-YY211T (= DSM 106068T = CGMCC 4.7474T).

Cryobacterium tepidiphilum sp. nov., isolated from rhizosphere soil of lettuce (var. ramosa Hort.).

A Gram-stain positive, aerobic, rod-shaped bacterium, designated strain NEAU-85T, was isolated from rhizosphere soil of lettuce and characterised using a polyphasic approach. Strain NEAU-85T was found to be catalase positive, motile and able to grow at between 10 and 30 °C. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain NEAU-85T belongs to the genus Cryobacterium and is closely related to the type strain Cryobacterium psychrotolerans JCM 13925T (98.5%) and also to Leifsonia kafniensis NCCB 100216T (97.6%). Multilocus sequence analysis using the concatenated gene sequences of the atpD, dnaK, recA, rpoB, sevY and ychF genes showed that strain NEAU-85T belongs to the genus Cryobacterium. The digital DNA-DNA hybridization values revealed that strain NEAU-85T is distinct from its close phylogenetic neighbour C. psychrotolerans. The menaquinones were identified as MK-10 and MK-11. The phospholipid profile was found to consist of diphosphatidylglycerol, phosphatidylglycerol, phospholipid, an unidentified glycolipid and an unidentified lipid. The major fatty acids were identified as anteiso-C15:0, anteiso-C15:1 and iso-C16:0. The genomic DNA G + C content of strain NEAU-85T was determined to be 68.9 mol%. The DNA-DNA hybridization value between them was less than 70%. On the basis of phenotypic, genotypic and phylogenetic data, strain NEAU-85T can be concluded to represent a novel species of the genus Cryobacterium, for which the name Cryobacterium tepidiphilum sp. nov. is proposed. The type strain is NEAU-85T (= CCTCC AA 2018035T = JCM 32545T).

Differences between fast and slow muscles in scallops revealed through proteomics and transcriptomics.

BACKGROUND: Scallops possess striated and catch adductor muscles, which have different structure and contractile properties. The striated muscle contracts very quickly for swimming, whereas the smooth catch muscle can keep the shells closed for long periods with little expenditure of energy. In this study, we performed proteomic and transcriptomic analyses of differences between the striated (fast) and catch (slow) adductor muscles in Yesso scallop Patinopecten yessoensis. RESULTS: Transcriptomic analysis reveals 1316 upregulated and 8239 downregulated genes in slow compared to fast adductor muscle. For the same comparison, iTRAQ-based proteomics reveals 474 differentially expressed proteins (DEPs), 198 up- and 276 downregulated. These DEPs mainly comprise muscle-specific proteins of the sarcoplasmic reticulum, extracellular matrix, and metabolic pathways. A group of conventional muscle proteins-myosin heavy chain, myosin regulatory light chain, myosin essential light chain, and troponin-are enriched in fast muscle. In contrast, paramyosin, twitchin, and catchin are preferentially expressed in slow muscle. The association analysis of proteomic and transcriptomic data provides the evidences of regulatory events at the transcriptional and posttranscriptional levels in fast and slow muscles. Among 1236 differentially expressed unigenes, 22.7% show a similar regulation of mRNA levels and protein abundances. In contrast, more unigenes (53.2%) exhibit striking differences between gene expression and protein abundances in the two muscles, which indicates the existence of fiber-type specific, posttranscriptional regulatory events in most of myofibrillar proteins, such as myosin heavy chain, titin, troponin, and twitchin. CONCLUSIONS: This first, global view of protein and mRNA expression levels in scallop fast and slow muscles reveal that regulatory mechanisms at the transcriptional and posttranscriptional levels are essential in the maintenance of muscle structure and function. The existence of fiber-type specific, posttranscriptional regulatory mechanisms in myofibrillar proteins will greatly improve our understanding of the molecular basis of muscle contraction and its regulation in non-model invertebrates.

Molecular cloning, expression and biochemical characterization of hemoglobin gene from ark shell Scapharca broughtonii.

Hemoglobin, the main component of haemolymph, is widely distributed in animals. Although its important oxygen transport functions has been extensively reported, studies on the immune function of hemoglobin in mollusc are few. Research on immune of hemoglobin of ark shell Scapharca broughtonii attracted more and more attention due to its ownership of erythrocyte comparing with many other shellfish. In this study, the hemoglobin cDNA of S. broughtonii was cloned by EST and RACE methods (named as SbHb). Sequence analysis revealed that the cDNA was 946 bp in length, including an open reading frame (ORF) of 459 bp which encoded a polypeptide of 152 amino acid residues, and a 5'-untranslated region (UTR) of 313 bp, a 3'-UTR of 174 bp. Sequence and homology analysis showed that the SbHb shared similarity with that of other related species. The mRNA expression profiles of SbHb in tested tissues analyzed by quantitative real-time PCR (qRT-PCR) revealed that the mRNA of SbHb could be all detected in foot, gill, mantle, adductor muscle, haemocytes and hepatopancreas, and the highest level was found in the haemocytes, which is 163.2 times higher than that in adductor muscle. Vibrio anguillarum stimulation and hypoxia treatment both had significant impact on the expression of SbHb, which showed the same trends as increasing first to the highest at 16 h after treatment and then followed by declining. Recombinant protein of SbHb (rSbHb) was successfully obtained by prokaryotic expression, and further function analysis indicated obviously that the rSbHb had very strong phenoloxidase-like activity (PO-like activity) and it could remarkably inhibit growth of gram-negative bacteria V. anguillarum. All the data suggested that the SbHb plays a significant role in the process of antibacterial and anoxia tolerance reaction in S. broughtonii, providing the evidence that SbHb is a key immune factor.