Ultrasonic-microwave synergistic supramolecular solvent liquid-liquid microextraction of trace biogenic amines in fish and beer based on solidification of floating organic droplet.

It is important to detect the presence of biogenic amines (BAs) as indicators of food freshness. The purpose of this study was to develop a novel ultrasonic-microwave synergistic supramolecular solvent liquid-liquid microextraction based on solidification of floating organic droplet (UMS-SUPRAS-SFO-LLME) combined with high-performance liquid chromatography for the determination of BAs. The physical properties and microstructure of SUPRAS based on 1-dodecanol and tetrahydrofuran were studied, and the extraction conditions such as the SUPRAS volume, the UMS process, and the centrifugal conditions were optimized. The results for the extraction kinetics and thermodynamics showed that UMS-SUPRAS-SFO-LLME is a spontaneous, endothermic diffusion process. The linear ranges of this method are 0.1-2.0 × 105 ng·mL-1 (R2 > 0.994), the limits of detection are 4.0 × 10-3-6.0 × 10-2 ng·mL-1, and the recoveries were 96.28-103.15%. Compared with existing analysis methods, UMS-SUPRAS-SFO-LLME is a sensitive, green and economical sample pretreatment method for analyzing the enrichment of BAs in beer and fish.

Efficient imaging based on P - and N-codoped carbon dots for tracking division and viability assessment of lactic acid bacteria.

Assessment of lactic acid bacteria (LAB) activity plays a key role in the fermented food industry. Fluorescence imaging method based on dye is facile to detect LAB viability. However, it is difficult to obtain stable fluorescence, non-toxic and low-cost dyes. In this study, we prepare P- and N-doped carbon dots (PN-CDs) via microwave-assisted hydrothermal synthesis. The properties of high quantum yield (60.36%) and excitation dependence allowed for multicolor imaging of LAB (Lactobacillus plantarum [L.p] and Streptococcus thermophilus [S.t]). The abundant functional groups and positive charges (+2.34 mV) on the surface of PN-CDs facilitated their quickly integrated into cell wall of live LAB with obvious fluorescence or into dead cells. As a result, PN-CDs can not only be used to rapidly and efficiently monitor bacterial viability (one minute), but can also be used to visualize LAB division using fluorescence imaging. Importantly, the PN-CDs have potential to rapidly detect LAB activity in LAB-fermented juices.

Effects of carbon concentration, oxygen, and controlled pH on the engineering strain Lactiplantibacillus casei E1 in the production of bioethanol from sugarcane molasses.

Sugarcane molasses are considered a potential source for bioethanol's commercial production because of its availability and low market price. It contains high concentrations of fermentable sugars that can be directly metabolized by microbial fermentation. Heterofermentative lactic acid bacteria, especially Lactiplantibacillus casei, have a high potential to be a biocatalyst in ethanol production that they are characterized by strong abilities of carbohydrate metabolism, ethanol synthesis, and high alcohol tolerance. This study aimed to evaluate the feasibility of producing ethanol by Lactiplantibacillus casei used the ethanologen engineering strain L. casei E1 as a starter culture and cane molasses as substrate medium. The effects of environmental factors on the metabolism of L. casei E1 were analyzed by high-performance liquid chromatography (HPLC) system, and the gene expression of key enzymes in carbon source metabolism was detected using quantitative real-time PCR (RT-qPCR). Results showed that the strain could grow well, ferment sugar quickly in cane molasses. By fermenting this bacterium anaerobically at 37 °C for 36 h incubation in 5 °BX molasses when the fermenter's pH was controlled at 6.0, ethanol yield reached 13.77 g/L, and carbohydrate utilization percentage was 78.60%. RT-qPCR results verified the strain preferentially ferment glucose and fructose of molasses to ethanol at the molecular level. In addition, the metabolism of sugars, especially fructose, would be inhibited by elevating acidity. Our findings support the theoretical basis for exploring Lactic acid bacteria as a starter culture for converting sugarcane molasses into ethanol.

MicroRNA-216b inhibits heat stress-induced cell apoptosis by targeting Fas in bovine mammary epithelial cells.

Heat stress affects milk yield and quality in lactating dairy cows in summer. Bovine mammary epithelial cells (bMECs) play a key role in milk secretion, and microRNAs (miRNAs) regulate numerous functions of bMEC. Previous reports have verified that miR-216b regulated cell apoptosis through repressing target genes in several cancer cells. So, our purpose was to explore the potential involvement of miR-216b in heat stress-induced cell apoptosis in bMECs. Firstly, the heat stress model was constructed and we found that apoptotic rates of bMECs significantly increased under heat stress. The expression of miR-216b, Bax mRNA, and caspase-3 mRNA was upregulated. However, Bcl-2 mRNA level was detected to differentially downregulated. Overexpression of miR-216b remarkably downregulated the expression of caspase-3 and Bax mRNA and protein, and the mRNA and protein level of Bcl-2 was increased. Inhibition of miR-216b increased the activity of caspase-3 and Bax, and the level of Bcl-2 was inhibited. Moreover, Fas was identified as a target gene of miR-216b through bioinformatic analysis and dual-luciferase reporter assay. Fas activity was significantly inhibited and enhanced respectively after transfecting miRNA mimics and inhibitor. Finally, inhibition of Fas via the small interfering RNA (siRNA) also inhibited cell apoptosis induced by heat stress. Taken together, our results indicated that miR-216b exerted as an anti-apoptotic effect under heat stress in bMECs by targeting Fas.

Landscape of long non-coding RNAs in Trichophyton mentagrophytes-induced rabbit dermatophytosis lesional skin and normal skin.

Emerging evidences suggest that long non-coding RNAs (lncRNAs) play important role in disease development. However, the role of rabbit lncRNAs in the pathogenesis of dermatophytosis remains elusive. The present study aimed to study and characterize lncRNA transcriptome in 8 T. mentagrophytes-induced female rabbit dermatophytosis lesional (TM) and 4 normal saline-infected (NS) skin biopsies using RNAseq. We identified 5883 lncRNAs in 12 strand-specific RNA-seq libraries and found 64 differentially expressed lncRNAs (q < 0.05) in TM relative to NS. As in other mammalian counterparts, rabbit lncRNAs were distributed in all chromosomes except the Y chromosome and were generally smaller in size and fewer in exon numbers compared to protein coding genes. Next, co-expression analysis revealed that 107 pairs between 32 DE lncRNAs and 96 protein coding genes showed a highly correlated expression (|r| > 0.8). Moreover, miRPara analysis of the lncRNAs revealed 173 lncRNAs with precursor sequences for 9561 probable novel miRNAs. Finally, q-PCR results validated the RNA-seq results with eight randomly selected lncRNAs. To the best of our knowledge, this is the first report on rabbit lncRNAs, and our results highlighted the potential role of lncRNAs in the pathogenesis of dermatophytosis.

Genome-wide microRNA profiling of bovine milk-derived exosomes infected with Staphylococcus aureus.

Bovine milk is rich in exosomes, which contain abundant miRNAs and play important roles in the regulation of neonatal growth and development of adaptive immunity. Here, we analyzed miRNA expression profiles of bovine milk exosomes from three healthy and three mastitic cows, and then six miRNA libraries were constructed. Interestingly, we detected no scRNAs and few snRNAs in milk exosomes; this result indicated a potential preference for RNA packaging in milk exosomes. A total of 492 known and 980 novel exosomal miRNAs were detected, and the 10 most expressed miRNAs in the six samples accounted for 80-90% of total miRNA-associated reads. Expression analyses identified 18 miRNAs with significantly different expression between healthy and infected animals; the predicted target genes of differentially expressed miRNAs were significantly enriched in immune system process, response to stimulus, growth, etc. Moreover, target genes were significantly enriched in several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways including inflammatory, immune, and cancer pathways. Our survey provided comprehensive information about milk exosomes and exosomal miRNAs involved in mastitis. Moreover, the differentially expressed miRNAs, especially miR-223 and miR-142-5p, could be considered as potential candidates for mastitis.

miR-148a-3p promotes rabbit preadipocyte differentiation by targeting PTEN.

Although emerging data support crucial roles for microRNAs (miRNAs) during adipogenesis, the detailed mechanisms remain largely unknown. In this study, it was shown that in rabbits, levels of miR-148a-3p not only increased in white adipose tissue during early stages of growth but also during in vitro cultured preadipocyte differentiation. Furthermore, overexpression of miR-148a-3p significantly upregulated the mRNA levels of PPARγ, C/EBPα, and FABP4, as well as the protein levels of PPARγ, as indicated by qPCR and western blotting analyses. Overexpression of miR-148a-3p also promoted intracellular triglyceride accumulation. In contrast, downregulation of miR-148a-3p inhibited the differentiation of rabbit preadipocytes. Next, based on target gene prediction and a luciferase reporter assay, we further demonstrated that miR-148a-3p directly targeted one of the 3' untranslated regions of PTEN. Finally, it was observed inhibition of PTEN by siRNA promoted rabbit preadipocyte differentiation. Taken together, our results suggested that miR-148a-3p could be involved in regulating rabbit preadipocyte differentiation through inhibiting expression of PTEN, which further highlighted the importance of miRNAs during adipogenesis.

miR-221 modulates skeletal muscle satellite cells proliferation and differentiation.

MicroRNAs (miRNAs) are a class of small non-coding RNA molecules, which play important roles in animals by targeting mRNA transcripts for translational repression. Many recent studies have shown that miRNAs are involved in the control of muscle development. In this study, the expression levels of miR-221 in different tissues and during rabbit skeletal muscle satellite cells (SMSCs) differentiation were detected. Gene ontology term enrichment was used to predict the potential biological roles of miR-221. A synthetic miR-221 mimic and a miR-221 inhibitor were used to investigate the functions of miR-221 during SMSCs proliferation and differentiation to further verify the functions of miR-221 in muscle development. In this report, we compared the expression levels of miR-221 in different tissues. The expression levels of miR-221 were upregulated after the induction of differentiation, and then were gradually downregulated during SMSCs differentiation. Overexpression of miR-221 promoted SMSCs proliferation, whereas inhibiting expression restrained proliferation in the EdU and CCK-8 assays. In addition, overexpression of miR-221 led to a decline in the expression levels of the differentiation marker genes MyoG and MHC. miR-221 overexpression suppressed SMSCs myotube formation. On the contrary, inhibition of miR-221 promoted myotube formation. Our data showed that miR-221 increased SMSCs proliferation and decreased differentiation.