Divergent Synthesis of 2-Chromonyl-3-hydrazono-chromones and 2-Alkoxy-3-hydrazono-chromones through Switchable Annulation Reactions of o-Hydroxyphenylenaminones with Aryldiazonium Salts.

An unprecedented selective chromone annulation reaction controlled by solvent for the divergent synthesis of two types of 2,3-disubstituted chromone skeletons has been developed. A variety of 2-chromonyl-3-hydrazono-chromones and 2-alkoxy-3-hydrazono-chromones were constructed efficiently from readily available o-hydroxyphenylenaminones (o-HPEs) and aryldiazonium salts at room temperature. This strategy is highly chemoselective and features mild reaction conditions, broad substrate scope, broad functional group tolerance, easy gram-scale preparation, and simple filtration to obtain the pure products without tedious column chromatography.

Comprehensive characterization of the bacterial community structure and metabolite composition of food waste fermentation products via microbiome and metabolome analyses.

Few studies have characterized the microbial community and metabolite profile of solid food waste fermented products from centralized treatment facilities, which could potentially be processed into safe animal feeds. In this study, 16S rRNA gene sequencing and liquid/gas chromatography-mass spectrometry were conducted to investigate the bacterial community structure and metabolite profile of food waste samples inoculated with or without 0.18% of a commercial bacterial agent consisting of multiple unknown strains and 2% of a laboratory-made bacterial agent consisting of Enterococcus faecalis, Bacillus subtilis and Candida utilis. Our findings indicated that microbial inoculation increased the crude protein content of food waste while reducing the pH value, increasing lactic acid production, and enhancing aerobic stability. Microbial inoculation affected the community richness, community diversity, and the microbiota structure (the genera with abundances above 1.5% in the fermentation products included Lactobacillus (82.28%) and Leuconostoc (1.88%) in the uninoculated group, Lactobacillus (91.85%) and Acetobacter (2.01%) in the group inoculated with commercial bacterial agents, and Lactobacillus (37.11%) and Enterococcus (53.81%) in the group inoculated with homemade laboratory agents). Microbial inoculation reduced the abundance of potentially pathogenic bacteria. In the metabolome, a total of 929 substances were detected, 853 by LC-MS and 76 by GC-MS. Our results indicated that inoculation increased the abundance of many beneficial metabolites and aroma-conferring substances but also increased the abundance of undesirable odors and some harmful compounds such as phenol. Correlation analyses suggested that Leuconostoc, Lactococcus, and Weissella would be promising candidates to improve the quality of fermentation products. Taken together, these results indicated that inoculation could improve food waste quality to some extent; however, additional studies are required to optimize the selection of inoculation agents.

Whole-genome sequencing reveals sex determination and liver high-fat storage mechanisms of yellowstripe goby (Mugilogobius chulae).

As a promising novel marine fish model for future research on marine ecotoxicology as well as an animal model of human disease, the genome information of yellowstripe goby (Mugilogobius chulae) remains unknown. Here we report the first annotated chromosome-level reference genome assembly for yellowstripe goby. A 20.67-cM sex determination region was discovered on chromosome 5 and seven potential sex-determining genes were identified. Based on combined genome and transcriptome data, we identified three key lipid metabolic pathways for high-fat accumulation in the liver of yellowstripe goby. The changes in the expression patterns of MGLL and CPT1 at different development stage of the liver, and the expansion of the ABCA1 gene, innate immune gene TLR23, and TRIM family genes may help in balancing high-fat storage in hepatocytes and steatohepatitis. These results may provide insights into understanding the molecular mechanisms of sex determination and high-fat storage in the liver of marine fishes.

Characterization of transcriptional responses mediated by benzo[a]pyrene stress in a new marine fish model of goby, Mugilogobius chulae.

Benzo[a]pyrene (BaP) is one of the most studied targets among polycyclic aromatic hydrocarbons (PAHs). Because of the complexity of the toxicity mechanism in BaP, little is known about the molecular mechanism at the level of transcription of BaP in marine fishes. The primary objective of this study was to investigate the molecular basis of the effects of BaP on marine fish, using Mugilogobius chulae (Smith 1932) as the model. A closed colony of M. chulae was used for the BaP toxicity test. Two fish liver samples per replicate from each group were excised and blended into one sample by pooling an equal amount of liver tissue. Total RNA of all samples was extracted separately. Equal quantities of total RNA from the three replicates of the two groups were pooled for sequencing. The sequencing cDNA libraries were sequenced using Illumina HiSeq 2000 system. Differentially expressed genes were detected with the DEGSeq R package. In total, 52,364,032 and 53,771,748 clean nucleotide reads were obtained in the control and BaP-exposed libraries, respectively, with N50 lengths of 1277 and 1288 bp, respectively. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed a significant enrichment of genes related to detoxification, transportation, and lipid metabolism. We also identified, for the first time, an association between endoplasmic reticulum dysfunction and lipid metabolism resulting from BaP exposure. Using quantitative real-time PCR, some effective molecular biomarkers for monitoring of BaP-polluted seawater were identified. The results demonstrate that BaP enhanced the expression of genes involved in detoxification in M. chulae and inhibited that of genes related to lipid metabolism, possibly by suppressing the expression of numerous ER-related genes involved in fat digestion and absorption.

Interaction of Group B Streptococcus sialylated capsular polysaccharides with host Siglec-like molecules dampens the inflammatory response in tilapia.

Group B Streptococcus (GBS, S. agalactiae) infection in tilapia (Oreochromis niloticus) causes widespread death of this species and is a significant issue for the aquaculture industry. The major virulence factor for GBS is its sialylated capsular polysaccharides (CPs). These CPs interact with sialic acid-binding immunoglobulin-like lectins (Siglecs) on the host immune cells to regulate the downstream inflammatory response and evade detection. Previously, we cloned multiple Siglec-like molecules from an O. niloticus cDNA library, all of which were shown to interact with the sialylated CPs of GBS. In the present study, we investigated the effects of GBS infection on the expression of pro- and anti-inflammatory cytokines in O. niloticus as well as OnSiglec-like-transfected macrophage cells. Eukaryotic expression vectors containing full-length OnSiglec-1-like, -4b-like, -14-like were constructed and used to transfect RAW264 macrophages in vitro as well as live tilapia in vivo prior to GBS infection. The expression of the anti-inflammatory cytokine interleukin (IL)-10 and the pro-inflammatory cytokines tumor necrosis factor (TNF)-α, IL-6, and interferon (INF)-ß were then analyzed by qPCR. Our results indicate that as infection progressed, IL-10 expression was significantly upregulated, while that of TNF-α and IL-6 were significantly downregulated in the OnSiglec-like-transfected cells. INF-ß expression was also downregulated in cells transfected with OnSiglec-1-like and -4b-like, but was not significantly effected in OnSiglec-14-like-transfected cells. Notably, the magnitude of these cytokine expression changes was greatly decreased when a ΔneuA GBS mutant was used to infect the OnSiglec-1-like-transfected cells. In GBS-infected tilapia, IL-10 expression was significantly upregulated in all tissues, whereas INF-ß expression in the spleen, kidney, and gills was significantly downregulated at 12 hpi. While the expression of TNF-α was slightly upregulated, this change was not significant. In GBS ΔneuA mutant-infected O. niloticus, IL-10 expression in all of the tissues was significantly lower than that observed for the wild-type GBS group, while TNF-α expression was higher in the mutant infected group. There was no significant difference in INF-ß expression between the two groups. Taken together, sialylated CPs on GBS appear to interact with host OnSiglec-like molecules to transmit negative regulatory signals via enhanced anti-inflammatory cytokine IL-10 production and reduced pro-inflammatory cytokine production, ultimately leading to dampening of the host immune response. The results of this study further elucidate the molecular mechanism underlying GBS infection in tilapia and also provide candidate drug target molecules.

De novo transcriptome assembly of the new marine fish model of goby, Mugilogobius chulae.

The yellowstripe goby (Mugilogobius chulae) is an ideal experimental marine fish model in the field of marine environmental toxicology. To clarify the mechanisms of molecular toxicity of benzo[a]pyrene (BaP) in standard laboratory fish, we carried out a genome-wide analysis of transcriptional profiles in M. chulae by RNA sequencing. A total of 47,979 unigenes were assembled de novo, with N50 lengths of 1658 bp. These results provide an important resource for future studies on the effects of BaP on marine animals.

Discovery and expression of 3 siglecs-like in Oreochromis niloticus neutrophil, and their interaction with group B streptococcal sialylated capsular polysaccharides.

Sialic acid - binding immunoglobulin - like lectins (Siglecs) are members of the largest superfamily of immune receptors; they recognize sialic acid and are mainly expressed in immune cells. Studies on mammals indicate that Streptococcus agalactiae (GBS) evade immune reactions by interacting with the host immune cells via the sialic acid of sialylated capsular polysaccharides. However, it is currently unknown if fish-derived GBS can interact with Siglecs to evade host immunity. In this study, we examined the binding of FITC-GBS with neutrophils to determine the presence of receptors that binds with GBS. Furthermore, 3 Siglec-like genes, (OnSiglec-1-like/-4b-like/-14-like) from the neutrophils cDNA were screened by PCR. All the genes had specific domains (immunostimulation and immunosuppression domains), conserved amino acid residues, and sialic acid polysaccharide binding sites that are found in mammalian Siglecs. Flow cytometry of Siglecs-like/COS-7 cells and ELISA of Siglecs/Ex-Fc fusion proteins confirmed that 3 Siglecs-like have high binding activity with GBS. Erythrocytes adhesion assays and sialylated glycans binding assay confirmed that 3 Siglecs-like bind to sialic acid polysaccharides. Siglecs-like had high expression levels in the spleen, gill, and kidney in Oreochromis niloticus by qPCR. After experimental infection, Siglec-1-like/-14-like showed a significant upregulated initially and later downregulated in liver, spleen, kidney, and gill. However, Siglec-4b-like was downregulated in most tissues, except that in liver. The results indicate that 3 OnSiglecs-like may recognize GBS sialylated capsular polysaccharides. GBS infections led to significant changes in Siglecs-like expression in immune-related tissues. However, immunostimulation or immunosuppression via the recognition of GBS by different Siglecs-like molecules requires additional studies.