Shuliang Liu

Prevalence and characterization of Salmonella species isolated from pigs, ducks and chickens in Sichuan Province, China

This study aimed to analyze the prevalence of Salmonella isolated from different parts of the food production chain, and to characterize these isolates. A total of 165 Salmonella enterica isolates were identified from 1382 samples taken from conventional farms, abattoirs and retail markets from 2010 to 2011 in Sichuan, China. The Salmonella isolates were assayed for serotype, antimicrobial susceptibility, prevalence of class 1 integrons and β-lactamase genes, and subtyped using pulsed-field gel electrophoresis. Among these isolates, S. enterica serotypes Derby (76 isolates, 46%) and Typhimurium (16 isolates, 10%) were the most prevalent, and high antimicrobial resistance rates were observed for tetracycline (77%), sulfamethoxazole/trimethoprim (43%), nalidixic acid (41%) and spectinomycin (41%). Class 1 integrons were detected in 21% of these isolates, and contained gene cassettes dfrA12-aadA2, dfrA1-aadA1, dfrA1, blaPSE-1 and dfrA1/aadA2. blaOXA-1 was the most commonly identified β-lactamase gene (n=14), followed by blaTEM-1 (n=6), blaPSE-1 (n=4) and blaCMY-2 (n=1). A S. enterica serotype Indiana isolate derived from chicken from a market was positive for both blaOXA-1 and blaCMY-2, and resistant to nine tested antibiotics. The PFGE patterns were diverse. Our findings indicated that most isolates from different sampling sites were phenotypically and genetically diverse, and Salmonella was widespread and may transmit along the food production chain from farm to market. Isolates with decreased susceptibility to fluoroquinolones and extended-spectrum cephalosporins, which are used to fight foodborne Salmonella, pose a serious threat to public health.

Antimicrobial resistance and resistance genes in Salmonella strains isolated from broiler chickens along the slaughtering process in China

A total of 189 Salmonella isolates were recovered from 627 samples which were collected from cecal contents of broilers, chicken carcasses, chicken meat after cutting step and frozen broiler chicken products along the slaughtering process at a slaughterhouse in Sichuan province of China. The Salmonella isolates were subjected to antimicrobial susceptibility testing to 10 categories of antimicrobial agents using the Kirby-Bauer disk diffusion method. Those antibiotics-resistant isolates were further investigated for the occurrence of resistance genes, the presence of class 1 integron as well as the associated gene cassettes, and the mutations within the gyrA and parC genes. Consequently, the prevalence of Salmonella was 30.14% (47.96% for cecal content, 18.78% for chicken carcasses, 31.33% for cutting meat and 14.00% for frozen meat, respectively). The predominant serotypes were S. Typhimurium (15.34%) and S. Enteritidis (69.84%). High resistance rates to the following drugs were observed: nalidixic acid (99.5%), ampicillin (87.8%), tetracycline (51.9%), ciprofloxacin (48.7%), trimethoprim/sulfamethoxazole (48.1%), and spectinomycin (34.4%). Antimicrobial resistance profiling showed that 60.8% of isolates were multidrug resistant (MDR), and MDR strains increased from 44.7% to 78.6% along the slaughtering line. 94.6% (n=157) of beta-lactam-resistant isolates harbored at least one resistance gene of blaTEM or blaCTX-M. The relatively low prevalence of aminoglycoside resistance genes (aac(3)-II, aac(3)-IV, and ant(2″)-I) was found in 49 (66.2%) of antibiotic-resistant isolates. The tetracycline resistance genes (tet(A), tet(B), tet(C), and tet(G) and sulfonamide resistance genes (sul1, sul2, and sul3) were identified in 84 (85.7%) and 89 (97.8%) antibiotic-resistant isolates respectively. floR was identified in 44 (97.8%) florfenicol-resistant isolates. Class 1 integron was detected in 37.4% (n=43) of the MDR isolates. Two different gene cassettes, blaOXA-30-aadA1 (19 isolates) and blaOXA-30-aadA1/drfA1-orfC (2 isolates), were identified in class 1 integron-positive isolates. 97.9% (184/188) of quinolone-resistant isolates had at least one mutation within gyrA or parC. Overall, antimicrobial resistance showed an increasing trend along the slaughtering process. The results showed that broiler chicken products in the slaughterhouse were contaminated with MDR Salmonella, which might originate from food producing animals to some extent, and cross-contamination during slaughter, and facilitate the dissemination of the resistance genes to consumers along the production chain, which suggests importance of controlling Salmonella during slaughter for public health, underlying strict hygiene method and HACCP management to reduce cross-contamination.

Effect of oxidized lipids stored under different temperatures on muscle protein oxidation in Sichuan-style sausages during ripening

This study was conceived to research muscle protein oxidation under the influence of four different degrees of oxidized lipids during the ripening of Sichuan-style sausages. Lipids were stored at different temperatures to obtain different oxidation degrees. To elucidate the relationship between lipid oxidation and protein oxidation, the indicators of lipid oxidation, protein oxidation and protein degradation were analysed. During ripening, the carbonyl, SH, SS and free amino acid contents changed significantly. The carbonyl and SS contents increased first in all samples, then decreased, whereas the SH content showed the opposite results. These results showed a positive correlation between protein oxidation and lipid oxidation. Lipids with a higher oxidation degree induced a stronger oxidation reaction to protein. Meanwhile, the results of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that the influence of lipid oxidation on myofibrillar proteins was much more intense than on sarcoplasmic proteins.

Exopolysaccharides produced by yogurt-texture improving Lactobacillus plantarum RS20D and the immunoregulatory activity

The strain RS20D capable of significantly improving yogurt texture was isolated from traditional fermented vegetable products, and identified as Lactobacillus plantarum RS20D. The total exopolysaccharides (EPS) were prepared from reconstituted skim milk fermentation by RS20D, and purified through DEAE-Sepharose CL-6B and Sephadex G-100, and consequently the purified fraction designated as RS-r2 was obtained. The further work aimed to elucidate the structural features of RS-r2 via FT-IR spectrum, HPSEC and monosaccharide composition analysis was carried out. The results showed that RS-r2 was a novel acidic heteropolysaccharide mainly consisted of glucose, galactose and glucosamine in a molar ratio of 2.0:1.5:1. The molecular weight was estimated to be 1.69u202f×u202f106u202fDa. The EPS had a high degradation temperature (250u202f°C), suggesting its high thermal stability. SEM and AFM analysis of EPS further revealed chain microstructure anchored with many regular spherical shape in aqueous solution. In vitro test showed that total EPS secreted by RS20D could stimulate macrophage RAW264.7 to release NO significantly and up-regulated the gene expression of pro-inflammatory cytokines at the mRNA level. Current study suggested that RS20D could be a potential source of immunoregulatory polysaccharide and may be applied as a functional starter culture to improve yogurt texture in the dairy industry.

Preparation of magnetic molecularly imprinted polymers with double functional monomers for the extraction and detection of chloramphenicol in food

In this study, an efficient, selective, and simple analytical method for the extraction of chloramphenicol (CAP) from food using magnetic molecularly imprinted polymers (MMIPs) as the solid-phase extraction (SPE) sorbent was successfully developed. MMIPs with varying ratios of methacrylic acid to acrylamide were prepared by suspension polymerization on the surface of double-bond-modified Fe3O4 magnetic nanoparticles. Further, these MMIPs were characterized by Fourier transform infrared spectroscopy and scanning electron microscopy, as well as using a vibrating sample magnetometer. Furthermore, the adsorption capacities of MMIPs and MNIPs were investigated by binding experiments. Methodology evaluation for the detection of CAP from food was carried out using MMIPs as the SPE sorbent. By using an external magnetic field, MMIPs were separated by a simple and rapid method. The diameter of the so-obtained MMIPs, exhibiting good monodispersity, was 400-700u202fnm. The MMIPs exhibited the maximum apparent adsorption capacity of up to 42.60u202fmgu202fg-1 with good selectivity. For the detection of food samples, the linear response range was 0.02-10.00u202fmgu202fL-1, with a detection limit of 10u202fμgu202fL-1, and intra- and inter-day stabilities ranged from 1.34% to 1.89% and from 1.76% to 2.77%, respectively, with good recoveries (95.31%-106.89%) and satisfactory relative standard deviations (1.21%-2.60%).

Purification, characterization and antioxidant activity of the exopolysaccharide from Weissella cibaria SJ14 isolated from Sichuan paocai

In the present study, an exopolysaccharide (EPS)-producing strain SJ14 isolated from Sichuan paocai was identified as Weissella cibaria, with a typical ropy phenotype. W. cibaria SJ14 possessed good capabilities of acid production, salt tolerance, and nitrite depletion. The crude polysaccharides were obtained from the culture supernatant of strain SJ14 and further fractionated by DEAE-Sepharose Fast Flow ion-exchange and Sephadex G-100 size-exclusion chromatography. Consequently, two acidic EPS fractions (EPS-1 and EPS-3) were obtained with the average molecular weights of 7.12u202f×u202f104 and 3.01u202f×u202f104u202fDa, respectively. They were heteropolysaccharides, among which EPS-1 were rich in mannose, and composed of mannose, glucose, galactose, arabinose, xylose, and rhamnose in a molar ratio of 23.79: 4.80: 1.66: 1.00: 0.21: 0.09, whereas EPS-3 consisted of galactose, mannose, glucose, and arabinose in a molar ratio of 7.47: 3.69: 1.00: 0.85 were rich in galactose. Two EPS fractions also exhibited potential antioxidant properties in vitro, showing strong scavenging activities on three kinds of free radicals and reducing power, and the antioxidant activities of EPS-1 were significantly stronger than that of EPS-3.

A Minireview of the Methods for Listeria monocytogenes Detection

Listeria monocytogenes (LM) is recognized as an opportunistic, foodborne pathogen that leads to the disease listeriosis. Although the incidence of listeriosis is low, listeriosis has a high mortality rate. LM can survive the most common stresses present during food processing steps, and it causes contamination in many food products. Consequently, most countries have endorsed strong restrictions on LM in food products, especially in ready-to-eat products. Conventional culture-based methods are currently the gold standard for testing, but their inefficiency can no longer meet the needs of batch inspection. As a result, a sensitive, fast, and reliable method for LM detection is required. There are many rapid detection methods for LM that are based on different principles, and an overview of the methods of LM detection is addressed here, with an emphasis on chromatographic, immunological, and aptamer-based techniques. Additionally, the prospect of developing novel LM detection methods is discussed.

Label-free and enzyme-free sensitive fluorescent method for detection of viable Escherichia coli O157:H7

We have developed a label-free, enzyme-free, modification-free and DNA extraction-free fluorescent aptasensing (LEFA) method for detection of E. coli O157:H7 based on G-quadruplex formation using two ingeniously designed hairpin probes (GHP1 and GHP2). In the presence of E. coli O157:H7, it released the single stranded initiation sequence (IS) resulting in the toehold strand displacement between GHP1 and GHP2, which in turn led to the cyclic reuse of the production of DNA assemblies with numerous G-quadruplex structures and initiation sequences. Then these G-quadruplex structures can be recognized quickly by N-methyl mesoporphyrin IX (NMM) resulting in significantly enhanced fluorescence. The LEFA method was successfully implemented for detecting E. coli O157:H7 with a detection limit of 66u202fCFU/mL in pure culture, 10u202fCFU/mL and 1u202fCFU/mL after pre-incubation of the milk and tap water for 4 and 8u202fh, respectively. Moreover, the strategy could distinguish viable E. coli O157:H7 from dead E. coli O157:H7 and other species of pathogen cells. Furthermore, the whole process of the strategy is accomplished within 100u202fmin. The results indicated that the approach may be used to effectively control potential microbial hazards in human health, food safety, and animal husbandry.

Purification and characterization of neutral protease from Aspergillus oryzae Y1 isolated from naturally fermented broad beans

The strain Y1, with a notably high production of neutral protease, was isolated from naturally fermented broad beans and subsequently identified as Aspergillus oryzae, through the analysis of its morphology characteristics and 18S rDNA sequence. Naturally fermented broad beans are the main raw material in Sichuan broad-bean sauce. The neutral protease from Aspergillus oryzae Y1 was purified using ammonium sulphate precipitation and DEAE-Sepharose Fast Flow chromatography, which resulted in a 10.0-fold increase in the specific activity (2264.3xa0U/mg) and a recovery rate of 21%. The estimated molecular mass of the purified protease was approximately 45xa0kDa. The optimal pH and temperature of the purified protease were 7.0 and 55xa0°C, respectively. The heat resistance of the purified protease was significantly higher than the commercial protease. The effect of metal ions on the activity of the purified protease approximated that of commercial neutral protease. Furthermore, the maximum hydrolysis rate (Vmax) and apparent Michaelis–Menten constant (Km) values of the purified protease were 256.4103xa0μg/mLxa0min and 20.0769xa0mg/mL, respectively. The purified protease had a higher affinity for the substrate than the commercial neutral protease. All the results suggest that this neutral protease exhibits the potential for application in industry due to its good resistance to high temperatures and wide range of acids and bases.

Microbial BOD sensors based on Zr (IV)-loaded collagen fiber

Biochemical oxygen demand (BOD) sensors based on Zr (IV)-loaded collagen fiber (ZrCF), a novel material with great porous structure, were developed. This novel material shows adsorbability by microorganisms. Saccharomyces cerevisiae and Escherichia coli were used for the construction of BOD sensors. Factors affecting BOD sensor performance were examined. The ZrCF-based BOD sensor showed different sensitivities and linear response ranges with different biofilm densities. The amount of microorganisms strongly affected the performance of the BOD sensor. Poor permeability of previously reported immobilization carriers were greatly circumvented by ZrCF. The service life of the ZrCF-based BOD sensor was more than 42 days. The immobilized microorganisms can be stored for more than 6 months under 4°C in PB solution. There was good correlation between the results of the sensor method and the standard 5-day BOD method in the determination of pure organic substrates and real water samples.