In vivo detection of endogenous toxic phenolic compounds of intestine.

Phenol and p-cresol are two common toxic small molecules related to various diseases. Existing reports confirmed that high L-tyrosine in the daily diet can increase the concentration of phenolic compounds in blood and urine. L-tyrosine is a common component of protein-rich foods. Some anaerobic bacteria in the gut can convert non-toxic l-tyrosine into these two toxic phenolic compounds, phenol and p-cresol. Existing methods have been constructed for measuring the concentration of phenolic compound in feces. However, there is still a lack of direct visual evidence to measure the phenolic compounds in the intestine. In this study, we aimed to construct a whole-cell biosensor for phenolic compounds detection based on the dmpR, the regulator from the phenol metabolism cluster. The commensal bacterium Citrobacter amalonaticus PS01 was selected and used as the chassis. Compared with the biosensor based on ECN1917, the biosensor PS01[dmpR] could better implant into the mouse gut through gavage and showed a higher sensitive to phenolic compound. And the concentration of phenolic compounds in the intestines could be observed with the help of in vivo imaging system using PS01[dmpR]. This paper demonstrated endogenous phenol synthesis in the gut and the strategy of using commensal bacteria to construct whole-cell biosensors for detecting small molecule compounds in the intestines.

Bacterial load in meconium.

The spike-in plasmid method was utilized to perform an analysis on meconium and second-pass feces, yielding both relative and absolute quantitative results. With the absolute quantitative data, the abundance of bacteria in 17 meconium samples and 17 second-pass fecal samples were found to be 1.14 × 107 and 1.59 × 109 copies/g, respectively. The mode of delivery can significantly influence the alterations and compositions of gut bacteria in a newborn within 72 h.

Identification and expression analysis of sex biased miRNAs in chinese hook snout carp Opsariichthys bidens.

As an economically important fish, Opsariichthys bidens has obvious sexual dimorphism and strong reproductive capacity, but no epigenetics study can well explain its phenotypic variations. In recent years, many microRNAs involved in the regulation of reproductive development have been explored. In this study, the small RNA libraries of O. bidens on the testis and ovary were constructed and sequenced. A total of 295 known miRNAs were obtained and 100 novel miRNAs were predicted. By comparing testis and ovary libraries, 115 differentially expressed (DE) miRNAs were selected, of which 53 were up-regulated and 62 were down-regulated. A total of 64 GO items (padj < 0.01) and 206 KEGG pathways (padj < 0.01) were enriched in the target gene of miRNA. After that, the expression levels of nine DE miRNAs, including let-7a, miR-146b, miR-18c, miR-202-5p, miR-135c, miR-9-5p, miR-34c-3p, miR-460-5p and miR-338 were verified by qRT-PCR. Furthermore, bidirectional prediction of DE miRNAs and sex-related genes was carried out and the targeting correlation between miR-9-5p and nanos1 was verified by Dual-Luciferase reporter assay. Our findings identified the differentially expressed miRNA and paved the way to new possibilities for the follow-up study on the mechanism of miRNA-mRNA interaction in the gonads of O. bidens.

Low-Cost and Rapid Method of DNA Extraction from Scaled Fish Blood and Skin Mucus.

PCR-based DNA amplification has been one of the major methods in aquaculture research for decades, although its use outside the modern laboratory environment is limited due to the relatively complex methods and high costs. To this end, we investigated a swabbing and disc protocol for the collection of DNA samples from fish which could extract DNA from fish skin mucus by a non-invasion technique costing only $0.02 (USD) and requiring less than 30 seconds. The disc method that we chose could use the cheap filter paper to extract DNA from above 104 crucian carp blood cells, which is comparable to the commercial kit. By using skin mucus swabbing and the disc method, we can obtain amplification-ready DNA from mucus to distinguish different species from our smallest fish (medaka, ~2.5 cm and crucian carp, ~7 cm) to our biggest fish (tilapia, ~15 cm). Furthermore, the viral pathogen Carassius auratus herpesvirus (CaHV) of crucian carp was detected using our method, which would make performing molecular diagnostic assays achievable in limited-resource settings including aquafarms and aqua stores outside the laboratory environment.