Synergistic Interaction Between Paired Combinations of Natural Antimicrobials Against Poultry-Borne Pathogens.

Natural antimicrobials (NAM) are promising candidates for the successful control of poultry-borne bacteria, carrying potent antimicrobial activity (AMA) against a wide range of multidrug-resistant pathogens. Individual activities of carvacrol, eugenol, trans-cinnamaldehyde, oregano, and thymol, along with the combined activity of paired compounds, were examined using broth microdilution and checkerboard techniques. The characteristic interactions between the compounds were calculated using an improved method, based on combination index (CI) values. The bacteria examined herein were selected due to their known genetic resistance to at least one antibiotic. Our results indicated that thymol was most effective, exhibiting the lowest minimum inhibitory concentration (MIC) value against Salmonella pullorum, Escherichia coli, and Klebsiella pneumoniae, establishing the order of antimicrobial efficacy as: thymol > oregano > carvacrol > trans-cinnamaldehyde > eugenol. In the interaction study, the paired combination of carvacrol and thymol showed synergistic effects and was highly effective in reducing the antibiotic resistance of all the evaluated pathogens. Notably, all CI values were <1.0 in evaluations of S. pullorum, indicating the absence of antagonism between eugenol and thymol (or oregano). In K. pneumoniae, majority of CI values, which had a few concentration points, were smaller than 1.0, indicating a synergistic effect between eugenol and carvacrol (oregano or thymol), and trans-cinnamaldehyde and carvacrol. In E. coli, apart from some concentration points, some CI values were smaller than 1.0, demonstrating a synergistic effect between eugenol and carvacrol, and thymol and carvacrol (eugenol or oregano). It is therefore of great significance to investigate and illuminate the minimal effect concentration of these five components when they are used in combination as feed additives. Moreover, the improved evaluation method of this study provides a precise and extensive means to assess the synergistic effects of NAM.

Effect of Bacterial Resistance of Escherichia coli From Swine in Large-Scale Pig Farms in Beijing.

With widespread use of antibiotics in the aquaculture industry, bacterial resistance has recently attracted increasing attention. Continuous emergence of multi-resistant bacteria has greatly threatened human and animal health, as well as the quality and safety of livestock products. To control bacterial resistance, the effect of bacterial resistance needs to be well understood. The purpose of this study was to explore the factors influencing Escherichia coli (E. coli) drug resistance in large-scale pig farms. In this study, 296 strains of E. coli isolated and identified from large-scale pig farms in Beijing were used as the research objects. In vitro drug sensitivity tests were used to determine the sensitivity to 10 antibiotics of pig-derived E. coli. SPSS logistic regression was employed to analyze the effects of the season, pig type, sampling point (medication type) and sampling location on resistance and multi-drug resistance of E. coli from pigs. The degrees of drug resistance to 10 antibiotics of the 296 strains of pig-derived E. coli were varied, their resistance rates were between 4.05 and 97.64%, and their multi-drug resistance was appalling, with the highest resistance to six antibiotics being 26.35%. The isolated strains were proven more resistant to tetracyclines, penicillin and chloramphenicol, which are commonly used for disease prevention in pig farms, and less resistant to quinolones and aminoglycosides, which are not used in pig farms. The resistance of the isolated strains in spring and summer was generally higher than that in winter. E. coli resistance in piglets, fattening pigs and sows was more serious than that in nursery and sick pigs. The results showed that the season, type of medication and type of pig had an influence on the pig-derived E. coli resistance, among which the type of medication was the most influencing factor.

Isolation of an acidic polysaccharide from the flowers of Leucosceptrum canum Smith and its immunomodulatory activity evaluation.

A water-soluble polysaccharide (LCP-05) was isolated from the flowers of Leucosceptrum canum Smith. LCP-05 was an acidic polysaccharide with a molecular weight of approximately 8.9 kDa. Monosaccharide composition analysis indicated that LCP-05 was composed of Man, Rha, GlcA, GalA, Glc, Gal and Ara in a molar ratio of 0.83:1.68:0.33:2.15:1.00:1.45:1.22. The framework of LCP-05 was speculated to be a branched rhamnogalacturonan with the backbone consisting of α-1,2,4-linked Rhap and α-1,4-linked GalAp, and bearing branches at the O-4 position of the Rha residues. The side chains are terminated primarily with the Araf and Glcp residues. LCP-05 was found to be able to significantly induce the production of NO, IL-6, and TNF-α in RAW 264.7 cells, and to induce RAW 264.7 cell's suppressive effect on both cell growth and cell migration of 4 T1 mammary breast cancer cells.

Determination of the levels of tryptophan and 12 metabolites in milk by liquid chromatography-tandem mass spectrometry with the QuEChERS method.

Tryptophan and metabolites have important biological functions in humans. Milk is an important source of tryptophan intake. In this study, we developed a method to detect levels of tryptophan and 12 metabolites in milk. The analytes were extracted by using the QuEChERS (quick, easy, cheap, effective, rugged, and safe) procedure and analyzed by liquid chromatography-tandem mass spectrometry with electrospray ionization. The proposed method resulted in suitable accuracy (standard deviation ≤10.31%) and high sensitivity (the limits of quantification were between 0.05 and 5 ng/mL). Recoveries were in the range of 44 to 126%. Finally, the developed method was successfully applied to compare the content of tryptophan and metabolites in 4 milk products produced by different processes: pasteurized milk, UHT milk, milk powder, and yogurt. The results of partial least squares-discriminant analysis (PLS-DA) showed that different types of processed milk could be distinguished clearly according to the method used here. The determined tryptophan and metabolites levels in milk can provide a new reference for evaluation of milk.

Characterization of Salmonella enterica Isolates from Diseased Poultry in Northern China between 2014 and 2018.

Salmonella infection not only causes acute and chronic diseases in poultry flocks, but the infected poultry are among the most important reservoirs for a variety of Salmonella serovars frequently transmitted to humans. This study aimed to investigate the occurrence of Salmonella spp. in local poultry farms in China. Samples (n = 4255), including dead-in-shell embryos, culled day-old-hatchings and 1- to 4-week-old diseased birds, were collected for Salmonella culture from broiler chicken, meat-type duck and pigeon farms in northern China between 2014 and 2018. A total of 103 Salmonella were isolated. S. enterica serovar Enteritidis and S. Typhimurium were the most prevalent serovars, representing 53.4% and 34.9% of the isolates, respectively. Serovar diversity was the highest in ducks, with the S. Apeyeme being isolated for the first time from duck tissues. All isolates were characterized by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). MLST showed that all S. Enteritidis isolates shared the same sequence type (ST11), and Typhimurium showed several rare STs in addition to ST19. In comparison, PFGE showed better discrimination for S. Enteritidis and S. Typhimurium isolates, with nine distinct pulsotypes being observed. The isolates exhibited varying degrees of resistance to 15 tested antimicrobials and identified S. Enteritidis isolates (98.18%) with multiple antimicrobial resistance were a cause for concern. Our data on invasive Salmonella infection in meat-type poultry in local farms can be used to identify sources and factors associated with Salmonella spread in poultry and the associated food chain.

Comparison of the quality attributes of coconut waters by high-pressure processing and high-temperature short time during the refrigerated storage.

This study compared the shelf life and quality of high-pressure processing (HPP) and high-temperature short time (HTST)-treated coconut water at 4°C. HPP of 500 MPa (5 min) and HTST of 72°C (15 s) treatments could ensure microbial safety of coconut water during refrigerated storage of 25 and 15 days, respectively. At the end of 15 days of storage, loss of 51.54% amino acids and 32.37% protein, and retention of 65.0% total sugars, 64.51% ascorbic acid, and 74.34% total phenols were found in HTST group. More nutrient contents, 76.85% amino acids, 76.76% total protein, and 93.17% total phenols, were retained in HPP groups at the end of 25 days of storage. HPP-treated fresh-like product could provide an effective approach of extending shelf life of coconut water.

Identification of transcriptional biomarkers by RNA-sequencing for improved detection of ß2-agonists abuse in goat skeletal muscle.

In this paper, high-throughput RNA-sequencing (RNA-seq) was used to search for transcriptional biomarkers for ß2-agonists. In combination with drug mechanisms, a smaller group of genes with higher detection accuracy was screened out. Unknown samples were first predicted by this group of genes, and liquid chromatograph tandem mass spectrometer (LC-MS/MS) was applied to positive samples to validate the biomarkers. The results of principal component analysis (PCA), hierarchical cluster analysis (HCA) and discriminant analysis (DA) indicated that the eight genes screened by high-throughput RNA-seq were able to distinguish samples in the experimental group and control group. Compared with the nine genes selected from an earlier literature, 17 genes including these nine genes were proven to have a more satisfactory effect, which validated the accuracy of gene selection by RNA-seq. Then, six key genes were selected from the 17 genes according to the variable importance in projection (VIP) value of greater than 1. The test results using the six genes and 17 genes were similar, revealing that the six genes were critical genes. By using the six genes, three positive samples possibly treated with drugs were screened out from 25 unknown samples through DA and partial least squares discriminant analysis (PLS-DA). Then, the three samples were verified by a standard method, and mapenterol was detected in a sample. Therefore, the six genes can be used as biomarkers to detect ß2-agonists. Compared with the previous study, accurate detection of ß2-agonists abuse using six key genes is an improvement method, which show great significance in the monitoring of ß2-agonists abuse in animal husbandry.

New Analytical Tool for the Detection of Ractopamine Abuse in Goat Skeletal Muscle by Potential Gene Expression Biomarkers.

In this study, quantification of mRNA gene expression was examined as biomarkers to detect ractopamaine abuse and ractopamaine residues in cashmere goats. It was focused on the identification of potential gene expression biomarkers and describing the coreletionship between gene expression and residue level by 58 animals for 49 days. The results showed that administration periods and residue levels significantly influenced mRNA expressions of the ß2-adrenergic receptor (ß2AR), the enzymes PRKACB, ADCY3, ATP1A3, ATP2A3, PTH, and MYLK, and the immune factors IL-1ß and TNF-α. Statistical analysis like principal components analysis (PCA), hierarchical cluster analysis (HCA), and discriminant analysis (DA) showed that these genes can serve as potential biomarkers for ractopamine in skeletal muscle and that they are also suitable for describing different residue levels separately.

Comparative study between macrolide regulatory proteins MphR(A) and MphR(E) in ligand identification and DNA binding based on the rapid in vitro detection system.

The macrolide regulatory protein MphR(A) has been widely studied and used in various aspects such as metabolism monitoring, exogenous gene expression, and in vivo and in vitro macrolide antibiotic screening. Another macrolide regulatory protein, MphR(E), has rarely been reported. In this study, in vitro ELISA-type systems were established for MphR(A) and MphR(E) to study their correlation. The reactivity of 14 macrolide antibiotics and pseudo-macrolide antibiotics was tested in the systems. The results indicated that the ligand identification spectra of MphR(A) and MphR(E) were basically consistent. The binding characteristics of MphR(A) and MphR(E) with three corresponding promoter DNA sequences were preliminarily studied. According to the ELISA-type analysis results, MphR(A) and MphR(E) have consistent DNA binding properties, which bind to A-DNA/B-DNA more easily than to C-DNA. This study has confirmed that MphR(E) can bind to the promoter DNA sequences mrx(E) and mph(E) in plasmid pRSB111, and different DNAs can affect the sensitivity of the in vitro detection systems.