Characteristics of Extended-Spectrum ß-Lactamase-Producing Escherichia coli Derived from Food and Humans in Northern Xinjiang, China.

This study aimed to investigate the drug resistance, molecular characteristics, and genetic relationship of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli isolated from food and human stool samples in northern Xinjiang. From 2015 to 2016, a total of 431 samples (meats and vegetables) were collected from retail markets and supermarkets located in the regions of Urumqi, Shihezi, and Kuitun in Xinjiang, China, and 20 human stool samples from the Shihezi Hospital. The PCR method was used to detect E. coli, and the presence of ESBL-producing E. coli was confirmed using the K-B disk diffusion confirmatory method. The susceptibility to ESBL-producing E. coli was tested by the microdilution broth method, and the minimum inhibitory concentration was determined. PCR was used to detect the resistance and virulence genes of ESBL-producing E. coli, and phylogenetics, plasmid replicon typing, screening of three integrons, and multilocus sequence typing (MLST) were performed. The results showed that 127 E. coli strains (15 human stool and 112 food samples) were isolated. Out of the 127 E. coli strains, 38 strains (6 human stool and 32 food 34 samples) of ESBL-producing E. coli were identified through screening. These 38 strains showed resistance to cefotaxime (94.74%) and cefepime (94.74%), and were sensitive to meropenem (0.00%). The most detected resistance genes were blaTEM (47.37%), and the most detected virulence genes were fimH (97.73%), ompA (97.73%), hlyE (97.73%), and crl (97.37%). The isolates belonged to phylogroups B1 (42.11%), C (23.68%), and A (21.05%). Among the plasmid replicon subtypes, IncFIB was the main type (42.11%). The integrons detected were of the first type (47.37%) and the third type (26.32%). The 38 E. coli strains had 19 different sequence-type (ST) strains. These 38 strains of ESBL-producing E. coli were analyzed using MLST and STs are varied.

The tainted altruism effect: a successful pre-registered replication.

Newman and Cain (Newman, Cain 2014 Psychol. Sci. 25, 648-655 (doi:10.1177/0956797613504785)) reported that observers view a person's choices as less ethical when that person has acted in response to both altruistic and selfish (commercial) motivations, as compared with purely selfish interests. The altruistic component reduces the observers' approval rather than raising it. This puzzling phenomenon termed the 'tainted altruism' effect, has attracted considerable interest but no direct replications in prior research. We report direct replications of Newman and Cain's Experiments 2 and 3, using a larger sample (n = 501) intended to be fairly representative of the US population. The results confirm the original findings in considerable detail.

A method based on amino-modified magnetic nanoparticles to extract DNA for PCR-based analysis.

DNA extraction using magnetic particles (MPs) is a simple and rapid process. MPs directly combine with DNA, thereby facilitating removal of impurities in complex samples by magnetic attraction. Amino-modified magnetic nanoparticles (AMNPs) have been described as poor substrates for DNA extraction because it was difficult to desorb DNA from the AMNPs for downstream assays. In this study, we proposed to use the complexes of AMNPs and DNA directly as templates for PCR, thereby bypassing the difficulty of desorbing DNA from AMNPs. At first, we compared three distinct chemical modifications for magnetic nanoparticles (hydroxyl modified-, carboxyl modified-, and amino modified-magnetic nanoparticles) for the extraction of DNA under experimental conditions compatible with downstream assays (e.g., PCR). Under such conditions, we demonstrated that only the AMNPs had a powerful adsorption capability for DNA with 98% separation rate. We directly employed the complexes of AMNPs and extracted DNA as templates for PCR, which reduced the separation steps and minimized the loss of DNA. Notably, we minimized AMNPs-caused inhibition of the PCR by adding BSA to the PCR mixture. The AMNPs extraction was successfully applied for the detection of a genetically modified organism, species identification, and an allergen in a heterogenous mixture. The AMNPs-based extraction method is a simple and rapid process to extract DNA and even trace amounts of DNA for PCR-based analysis.

Enhancement of PCR Sensitivity and Yield Using Thiol-modified Primers.

Various additives can enhance the quality of PCR amplification, but these generally require considerable optimization to achieve peak performance. Here, we demonstrate that the use of thiol-modified primers can enhance both PCR sensitivity and yield. In experiments with V. parahaemolyticus genomic DNA, this primer modification enhances PCR sensitivity by more than 100-fold, with accompanying improvements in amplicon yield. Then, an artificial plasmid with the same primer binding regions and different internal amplification sequence was designed. The result showed that the amplification also be improved by using the same thiol-modified primers. It indicated the enhancement was not caused by the effect of the thiol-modified primers on the second structure of amplification sequence. Subsequent experiments demonstrate that the effects of this modification are potentially due to altered interaction between the primers and proteins in the reaction mixture. Amplification with thiol-modified primers was strongly inhibited by the presence of extraneous proteins relative to standard DNA primers, which indicates that thiol-modified primers may be inhibited due to interaction with these proteins. In contaminant-free reactions, however, the thiol-modified primers might interact more strongly with DNA polymerase, which could in turn improve PCR amplification.

A long time low drift integrator with temperature control.

The output of an operational amplifier always contains signals that could not have been predicted, even with knowledge of the input and an accurately determined closed-loop transfer function. These signals lead to integrator zero-drift over time. A new type of integrator system with a long-term low-drift characteristic has therefore been designed. The integrator system is composed of a temperature control module and an integrator module. The aluminum printed circuit board of the integrator is glued to a thermoelectric cooler to maintain the electronic components at a stable temperature. The integration drift is automatically compensated using an analog-to-digital converter/proportional integration/digital-to-analog converter control circuit. Performance testing in a standard magnet shows that the proposed integrator, which has an integration time constant of 10 ms, has a low integration drift (<5 mV) over 1000 s after repeated measurements. The integrator can be used for magnetic flux measurements in most tokamaks and in the wire rope nondestructive test.

The Construction of an Aqueous Two-Phase System to Solve Weak-Aggregation of Gigaporous Poly(Styrene-Divinyl Benzene) Microspheres.

Gigaporous poly(styrene-divinyl benzene) microspheres made via the surfactant reverse micelles swelling method had a controllable pore size of 100⁻500 nm. These microspheres had unique advantages in biomacromolecule separation and enzymes immobilization. However, the obtained microspheres adhered to each other in the preparation process. Though the weak aggregation could be re-dispersed easily by mechanical force, it will be difficult to scale up. By analyzing the formation mechanism of the aggregates, a method was presented to rebuild the interface between the internal aqueous channel and the external continuous phase by constructing an aqueous two-phase system (ATPS). Based on the ATPS, the method of emulsification, stirring speed, and surfactant concentration in oil phase were optimized. Under the optimum condition (screen emulsification method, 120 rpm for polymerization and 55% surfactant), the microspheres with a controllable particle size of 10⁻40 µm and a pore size of about 150 nm were obtained. This new method could significantly decrease the weak-aggregation of microspheres.

Overexpression of specific proton motive force-dependent transporters facilitate the export of surfactin in Bacillus subtilis.

A novel surfactin producer, Bacillus subtilis THY-7, was isolated from soil. Using liposomes and transmembrane transport inhibitors, the surfactin efflux in THY-7 was determined to be mainly dependent on proton motive force (PMF), not ATP hydrolysis. YcxA, KrsE and YerP, three putative lipopeptide transporters with PMF as energy source, were then highlighted in this work. A mutant YcxA named as YcxAmt, with 2 transmembrane helices deletion due to a code-shift mutation of the encoding gene, was identified in THY-7. This truncated YcxAmt was confirmed unable to transfer surfactin; on the contrary, overexpression of the natural full-lengthYcxA enhanced the secretion of surfactin by 89 %. KrsE, a putative kurstakin transporter, was found also capable of transporting surfactin. Overexpression of KrsE increased the production of surfactin by 52 %. In the culture of YerP-overexpressing strain at 24 h, surfactin titer reached 1.58 g L(-1), which was 145 % higher than that of the control. This indicated that YerP acted as the major surfactin exporter in B. subtilis THY-7.

Synergistic effects and physiological responses of selected bacterial isolates from animal feed to four natural antimicrobials and two antibiotics.

In this study, 20 samples from three different sources of animal feed were investigated and six bacterial isolates were identified. The susceptibility of four natural antimicrobials, namely, eugenol, cinnamaldehyde, thymol, and carvacrol, against six of these isolates was determined. Carvacrol and eugenol showed better inhibitory effects with larger zones of inhibition. The minimal inhibitory concentration for a range of antibiotics on the susceptibility of two isolates (namely, Sphingomonas paucimobilis and Klebsiella oxytoca) was investigated using the VITEK® 2 microbiological identification system. Both isolates showed a variety of resistance to 18 antibiotics. The minimal inhibitory concentration and fractional inhibitory concentration index of those two isolates for ampicillin and nitrofurantoin in combination with four phenolic compounds was determined. Synergistic interactions were found for most antimicrobial/antibiotic combinations; thymol and carvacrol were very effective (fractional inhibitory concentration ≤0.5) in combination with all antibiotics tested against S. paucimobilis and K. oxytoca, respectively. Ultra performance liquid chromatography techniques were used to investigate the physiological effects of the four natural antimicrobials against those two isolates. Two identical peaks were found to be systematically different between cinnamaldehyde-treated and -untreated cells. The identity of the peaks is unknown and further investigation is needed.

The influence of non-lethal temperature on the rate of inactivation of vegetative bacteria in inimical environments may be independent of bacterial species.

The influence of non-lethal temperature on the survival of two species of food-borne bacteria under growth-preventing pH and water activity conditions was investigated. Specifically, inactivation rates of four strains of Escherichia coli and three strains of Listeria monocytogenes were determined in culture broth adjusted to pH 3.5 and water activity 0.90, to prevent growth of both species, and for temperatures in the range 5-45 degrees C at 5 degrees C intervals. Sixty-three inactivation rates were obtained, plotted on Arrhenius co-ordinates, and lines of best-fit determined by simple linear regression. Differences in the mean inactivation rate of each species at a given temperature were not significant (p < 0.05) with the exception of the rates at 25 degrees C. The inactivation rate responses of both species were comparable to those reported by McQuestin et al. (Appl. Environ. Microbiol., 75:6963-6972, 2009) for a variety of E. coli strains under a wide range of growth-preventing pH and water activity conditions. The results support the hypothesis that non-lethal temperature is a key factor governing the rate of inactivation of vegetative bacteria in foods when other hurdles prevent their growth and indicate that the temperature effect may also be independent of bacterial species.

Temperature governs the inactivation rate of vegetative bacteria under growth-preventing conditions.

Novel studies, in combination with a meta-analysis of available data, were undertaken to explore the kinetics of non-thermal inactivation of Escherichia coli with particular attention to inactivation in fermented meats and including analogous broth-based model systems. The analyses were based on rates of inactivation and specifically investigated the influence of temperature, pH and water activity at levels that alone, or in combination, prevented growth. When independently-derived inactivation data, obtained using different test conditions and diverse E. coli strains, were presented as Arrhenius plots, temperature was found to have a strong effect on the rate of inactivation, explaining 60% of the variance in the data. The slope of the Arrhenius plot changed, however, at temperatures above approximately 47 degrees C, corresponding to the maximum for growth of E. coli. A strong and consistent effect of pH or water activity on inactivation rate was not observed upon meta-analysis of collated data, but the relative effect of both factors was quantified in an analogous broth-based system. We also observed that inactivation rates of three strains of Listeria monocytogenes in the range 5 to 40 degrees C did not differ systematically from those of four strains of E. coli when growth was prevented by low pH and water activity. The observations of a consistent slope of Arrhenius plots for non-thermal inactivation rate of bacteria under diverse environmental conditions and for different strains and species, but which differ from slopes associated with thermal inactivation, raise the intriguing possibility of a mechanism of inactivation at sub-lethal temperatures, distinct from thermal inactivation, that is common to many vegetative bacteria.