Effect of Arabinoxylan from Wastewater Generated during Vital Wheat Gluten Production on Liver Metabolism in Type 2 Diabetic Mice.

Arabinoxylan (AX) is a dietary fiber that has been proven to have a significant antidiabetic effect. Liver metabolic disorders frequently coincide with the development of type 2 diabetes, but research on the hepatoprotective effects of AX in type 2 diabetic mice is lacking. As AX is abundant in the wastewater produced during vital wheat gluten protein production, this study used it as a raw material to evaluate its protective effect on liver function. The study employed an AX intervention in type 2 diabetic mice induced by a high-fat diet combined with streptozotocin and collected serum and liver tissue samples after 4 weeks. Serum and liver function indicators were measured using an automatic biochemistry analysis apparatus, and liver fat accumulation was observed using oil red O staining. Nontargeted metabolomics analysis of liver tissues was conducted using UHPLC-MS/MS. The results showed that AX significantly improved liver function indicators and histopathological damage, and regulated liver metabolic disorders by improving the differential metabolites of pantothenate and CoA biosynthesis, as well as purine metabolism. This study demonstrated that AX may exert a significant hepatoprotective effect by regulating metabolic disorders.

BaeR overexpression enhances the susceptibility of acrB deleted Salmonella enterica serovar Typhimurium to polymyxin.

The mechanism of polymyxin resistance is complex, and the modification of lipopolysaccharide mediated by two-component system is one of the main cause of polymyxin resistance. To date, no studies have reported the contribution of the BaeSR two-component system to the polymyxin resistance of Salmonella. In this study, baeR, acrB single and double gene deletion strains of Salmonella typhimurium (AT-P128) which induced polymyxin resistance in vitro were constructed by using CRISPR/Cas9 gene editing technology, and the baeR gene was overexpressed in the acrB single gene deletion strain by the pUC19 plasmid. The susceptibility of different strains to polymyxin was determined by broth dilution method. Time-kill assay was carried out with different concentrations of polymyxin. The difference of gene expression among strains was compared by transcriptome sequencing (RNA-seq) and RT-qPCR. As a result, the MIC of the BaeR overexpression strain (AT-P128ΔacrB/pbaeR) to polymyxin was significantly reduced by 8-fold compared with the other tested strains. The growth curve results showed no significant change in the growth rate of the strain before and after gene deletion and overexpression. The time-kill assay showed that AT-P128ΔacrB/pbaeR was more susceptible under different concentrations of polymyxin. RNA-seq and RT-qPCR results showed that the expression levels of several polymyxin resistance-related genes including phoPQ, pmrD, pmrAB, arnT, eptB, lpxD, pagC and pagL changed significantly. These results indicate that overexpression of baeR in the context of the acrB gene deletion increases the polymyxin susceptibility of the strain and affects the expression level of polymyxin resistance genes, providing insight into the polymyxin resistance mechanism of S. typhimurium.

Quantitative phosphoproteomics reveals the effect of baeSR and acrB genes on protein phosphorylation in Salmonella enterica serovar typhimurium.

The BaeSR two-component system and the AcrB efflux pump are closely associated with Salmonella resistance to antibiotics. However, the relationship between the two-component system, efflux pumps and protein phosphorylation of Salmonella is poorly understood. In this study, Salmonella typhimurium ciprofloxacin-resistant strain CR, baeSR gene deletion strain CRΔbaeSR, acrB gene deletion strain CRΔacrB, and double gene deletion strain CRΔbaeSRΔacrB were used to explore phosphorylated proteins with significant difference, based on non-marker, quantitative phosphorylation modified proteomics technique. Consequently, 363 phosphosites of 213 phosphoproteins were identified in the four strains. More than 70% of the phosphosites were serine phosphorylation. In the CRΔbaeSR/CR, CRΔacrB/CR and CRΔbaeSRΔacrB/CR comparison groups, 36, 37 and 49 phosphosites were significantly altered, respectively. Bioinformatic analysis revealed that the main enrichment pathways of these differentially phosphorylated proteins were metabolic pathways, biosynthesis of antibiotics, phosphotransferase system (PTS), ABC transporters, and lipopolysaccharide biosynthesis. Furthermore, 21 differentially phosphorylated proteins were identified to be associated with antibiotic resistance. These results suggest that the BaeSR two-component system and the AcrB efflux pump affect the phosphorylation of proteins in S. typhimurium and may influence the drug resistance and virulence of S. typhimurium by affecting protein phosphorylation, providing a new idea to explore the mechanism of drug resistance in Salmonella.

Mechanisms of polymyxin resistance induced by Salmonella typhimurium in vitro.

The increase incidence of multi-drug resistant (MDR) Salmonella has become a major global health concern. Polymyxin, an ancient polypeptide antibiotic, has been given renewed attention over recent years, resulting in resistance of Gram-negative bacteria to polymyxin, but its resistance mechanism is not completely clear. Thus, it is important to study its resistance mechanisms. In this study, an in vitro induced polymyxin-resistant strain of Salmonella typhimurium in the laboratory were constructed to investigate the mechanism of resistance of Salmonella to polymyxin. Gradual induction of Salmonella typhimurium ATCC13311 (AT) by concentration increment was used to screen for a highly polymyxin-resistant strain AT-P128. The broth dilution technique was used to compare the sensitivity of the two strains to different antimicrobial drugs. Single nucleotide polymorphisms (SNPs) were then identified by whole genome sequencing, and differences in gene expression between the two strains were compared by transcriptome sequencing and reverse transcription-quantitative PCR (RT-qPCR). Finally, for the first time, the CRISPR/Cas9 gene-editing system was used to construct gene deletion mutants in Salmonella to knock out the phoP gene of AT-P128. The results showed that strain AT-P128 was significantly more resistant to amoxicillin, ceftiofur, ampicillin, fluphenazine, and chloramphenicol and significantly less resistant to sulfamethoxazole than the parental strain AT. The growth curve results showed no significant change in the growth rate between AT-P128 and AT. Motility and biofilm formation assays showed a significant decrease in AT-P128. Additionally, the WGS results showed that AT-P128 had mutations in 9 genes involving 14 SNPs. RNA-seq and RT-qPCR results showed increased expression of phoPQ. The loss of the phoP gene decreased AT-P128ΔphoP resistance to polymyxin by 32-fold. These results suggested that polymyxin resistance affected the biology, genome components, and gene expression levels of Salmonella and that the PhoPQ two-component system played a key role in polymyxin resistance in Salmonella, providing insights into the diversity and complexity of polymyxin resistance in Salmonella.