Cross-Phosphorylation between AgrC Histidine Kinase and the Noncognate Response Regulator Lmo1172 in Listeria monocytogenes under Benzalkonium Chloride Stress.

Benzalkonium chloride (BC) is widely used for disinfection in the food industry. However, Listeria monocytogenes strains with resistance to BC have been reported recently. In L. monocytogenes, the Agr communication system consists of a membrane-bound peptidase AgrB, a precursor peptide AgrD, a histidine kinase (HK) AgrC, and a response regulator (RR) AgrA. Our previous study showed that the agr genes are significantly upregulated by BC adaptation. This study aimed to investigate the role of the Agr system in BC resistance in L. monocytogenes. Our results showed that the Agr system was involved in BC resistance. However, a direct interaction between BC and AgrC was not observed, nor between BC and AgrA. These results indicated that BC could induce the Agr system via an indirect action. Both AgrBD and AgrC were required for growth under BC stress. Nevertheless, when exposed to BC, the gene deletion mutant ∆agrA strain exhibited better growth performance than its parental strain. The RR Lmo1172 played a role in BC resistance in the ∆agrA strain, suggesting that Lmo1172 may be an alternative to AgrA in the phosphotransfer pathway. Phosphorylation of Lmo1172 by AgrC was observed in vitro. The cognate HK Lmo1173 of Lmo1172 was not involved in BC stress, regardless of whether it was as the wild-type or the ∆agrA mutant strain. Our evidence suggests that the HK AgrC cross-phosphorylates its noncognate RR Lmo1172 to cope with BC stress when the cognate RR AgrA is absent. In vivo, further studies will be required to detect phosphotransfer of AgrC/AgrA and AgrC/Lmo1172.

Influence of Fe2O3 on the Properties and Structure of Iron Coke.

Iron cokes were produced in an electrical furnace from a coal blend containing varying levels of added Fe2O3. The effects of Fe2O3 on the properties and structure of the iron coke were then investigated using the coke for metallurgy determination of mechanical strength, determination of coke reactivity and coke strength after the reaction, X-ray diffraction, Raman spectroscopy, and a method for quantitative analysis of the minerals in coal and coke. Further, the relationships between the properties and structures of iron coke samples were established. The results show that the addition of Fe2O3 can reduce the tumble strength, coke strength after the reaction, aromaticity, microcrystalline size, graphitization degree, crystalline volume, and carbon order degree of the iron coke and increase the abrasion resistance, coke reactivity index, and pulverization rate. Moreover, the degree of influence increases with increasing levels of added Fe2O3. The Fe2O3 is mainly transformed into metallic iron during the coking process, and part of metallic iron is converted into Fe3O4 during iron coke gasification. With an increasing Fe2O3 content, the trend of the change in the minerals from Fe to Fe3O4 becomes much more obvious, resulting in deeper influences on the iron coke thermal properties. There are obvious correlations among the iron coke reactivity and iron coke strength after the reaction and the crystalline volume, carbon order degree, and metallic iron content. It is concluded that the addition of Fe2O3 decreases the crystalline volume and carbon order degree and increases the metallic iron content, resulting in increases in abrasion resistance and coke reactivity and decreases in tumble strength and coke strength after the reaction.

Novel heterostructured metal doped MgFe2O4@g-C3N4 nanocomposites with superior photo-Fenton preformance for antibiotics removal: One-step synthesis and synergistic mechanism.

A novel metal doped MgFe2O4@g-C3N4 (m-MF@CN) nanocomposite was synthesized by one-pot method using saprolite laterite nickel ore and urea as raw materials. The heterostructure was verified as an effective heterogeneous Fenton-like catalyst for degrading antibiotics including tetracycline, oxytetracycline and chlortetracycline hydrochloride, and the related catalytic mechanism was elaborated in detail. Under the optimum conditions, the m-MF@CN/H2O2/vis system exhibited superior photo-Fenton property (degradation efficiency of 93.15% within 30 min, TOC removal efficiency was as high as 60.54% within 120 min) and cycle stability for tetracycline removal. The combination of MgFe2O4 and g-C3N4 enhanced the absorption of visible light, and the energy level matched heterojunction promoted the separation of photogenerated electron-holes to accelerate the redox cycle of ≡Fe3+/≡Fe2+. Free radical quenching and electron spin resonance (ESR) analysis confirmed that O2- was the main active species, h+ and OH also played a synergistic role in the degrading reactions. Notably, a possible degradation pathway of tetracycline was proposed according to the intermediates produced in the reaction process. The one-step synthesized m-MF@CN nanocomposite catalysts possessed high catalytic performance, good stability and recoverability, which not only realized the high-value utilization of ore raw materials, but also provided a potential practical way for efficient treatment of antibiotic wastewater.

Benzalkonium Chloride Adaptation Increases Expression of the Agr System, Biofilm Formation, and Virulence in Listeria monocytogenes.

Benzalkonium chloride (BC) is widely used for disinfection in food industry. However, prolonged exposure to BC may lead to the emergence of BC adapted strains of Listeria monocytogenes, an important foodborne pathogen. Until now, two communication systems, the LuxS/AI-2 system and the Agr system, have been identified in L. monocytogenes. This study aimed to investigate the role of communication systems in BC adaptation and the effect of BC adaptation on two communication systems and the communication-controlled behaviors in L. monocytogenes. Results demonstrated that the Agr system rather than the LuxS system plays an important role in BC adaptation of L. monocytogenes. Neither luxS expression nor AI-2 production was affected by BC adaptation. On the other hand, the expression of the agr operon and the activity of the agr promoter were significantly increased after BC adaptation. BC adaptation enhanced biofilm formation of L. monocytogenes. However, swarming motility was reduced by BC adaptation. Data from qRT-PCR showed that flagella-mediated motility-related genes (flaA, motA, and motB) were downregulated in BC adapted strains. BC adaptation increased the ability of L. monocytogenes to adhere to and invade Caco-2 cells but did not affect the hemolytic activity. Compared with the wild-type strains, the expression levels of virulence genes prfA, plcA, mpl, actA, and plcB increased more than 2-fold in BC adapted strains; however, lower than 2-fold changes in the expression of hemolysis-associated gene hly were observed. Our study suggests that BC adaptation could increase the expression of the Agr system and enhance biofilm formation, invasion, and virulence of L. monocytogenes, which brings about threats to food safety and public health. Therefore, effective measures should be taken to avoid the emergence of BC adapted strains of L. monocytogenes.

An efficient and low-cost magnetic heterogenous Fenton-like catalyst for degrading antibiotics in wastewater: Mechanism, pathway and stability.

Metal-doped MgFe2O4 spinel ferrite synthesized from saprolite laterite nickel ore was verified as an efficient heterogeneous Fenton-like catalyst for degrading antibiotics including tetracycline (TC) and metronidazole (MNZ) in a "catalyst/oxalic acid (H2C2O4)/visible light (vis)" system. The degradation efficiencies reached over 95% and total organic carbon (TOC) removal efficiencies were nearly 50% of the two antibiotics within 210 min, under the optimal conditions, especially 90% catalytic activity of the fresh catalyst was maintained after five cycles, suggesting the ferrite possessed excellent degrading performance, cycling stability and applicability. Moreover, the degradation mechanism and pathway of TC were elucidated in detail. Results revealed that the [≡Fe(C2O4)3]3- complex ions formed by octahedral Fe3+ in spinel ferrite with oxalate ions on the surface of MgFe2O4, played the key role in production of ·OH radicals which decomposed antibiotic TC into small molecules even mineralized in three pathways. Cost-effective preparation, high catalytic performance and long cycle life may accelerate the practical application of the heterogeneous Fenton-like catalyst.

The effect of sublethal concentrations of benzalkonium chloride on the LuxS/AI-2 quorum sensing system, biofilm formation and motility of Escherichia coli.

Escherichia coli can survive improper disinfection processes, which is a potential source of contamination of food products. Benzalkonium chloride (BC) is a common disinfectant widely used in food industry. Bacterial quorum sensing (QS) plays a major role in food spoilage, biofilm formation and food-related pathogenesis. Understanding QS can help to control the growth of undesirable food-related bacteria. The LuxS/AI-2 QS system of E. coli has been confirmed to regulate many important phenotypes including biofilm formation and motility. In the current study, we aimed to investigate the effect of sublethal concentrations of BC on the LuxS/AI-2 system of E. coli isolates from retail meat samples, as well as bacterial biofilm formation and motility. Our results showed that sublethal concentrations of BC promoted AI-2 production in four test E. coli isolates. The results from microplate assay and confocal laser scanning microscopy (CLSM) analysis indicated that sublethal concentrations of BC enhanced biofilm formation of E. coli. When treated with sublethal concentrations of BC, exopolysaccharides (EPS) production during biofilm development increased significantly and swimming motility of tested isolates was also promoted. The expression levels of luxS, biofilm-associated genes and flagellar motility genes were increased by BC at sublethal concentrations. Our findings underline the importance of proper use of the disinfectant BC in food processing environments to control food contamination by E. coli.

Production and characterization of monoclonal antibodies against GII.6 norovirus virus-like particles.

In this study we generated and characterized a series of monoclonal antibodies (mAbs) against GII.6 norovirus (NoV) virus like particles (VLPs). Mice were immunized with purified GII.6 NoV VLPs and peptide-bovine serum albumin (BSA) conjugates with the peptide sequence (31 aa) derived from the trypsin cleavage region. An indirect enzyme-linked immunosorbent assay was used to identify positive cell clones during cloning and subcloning, and an in vitro VLP-histo-blood group antigens (HBGAs) binding blockade assay was used to identify mAbs with blocking ability. A total of seven mAbs comprising five (1F7, 1F11, 2B6, 2C4, and 2E10) reactive with major capsid proteins (VP1) and two (1E5 and 2B2) reactive with both VP1 proteins and the peptide were identified. mAb 1F7, 1F11, and 2B6 were identified as blocking antibodies. Sandwich ELISA indicated that all these mAbs recognized soluble GII.6 NoV VLPs. Cross-reactivities with GI.7, GII.3, and GII.4 NoV VLPs were observed in indirect and sandwich ELISA. Western blot analysis indicated that all non-blocking mAbs recognized denatured GII.6 VP1 proteins and blocking mAbs only recognized non-denatured proteins. The in vitro VLP-HBGA binding blockade assay indicated that the three blocking antibodies exhibited blocking effects against GII.6 NoV VLPs, but not GI.7, GII.3, and GII.4 NoV VLPs. Epitope mapping and HBGA blocking assay indicated that mAbs targeting the predicted surface-exposed loop region did not have blocking effects, suggesting a possible important role of this region in regulating NoV-HBGA interactions. This is the first report regarding the characterization of mAbs with blocking ability against GII.6 NoV VLPs. These mAbs might be useful in facilitating our understanding of this group of viruses.

Transformation of organic sulfur and its functional groups in nantong and laigang coal under microwave irradiation.

The microwave irradiation experiment of Nantong coal (NTC) and Laigang coal (LGC) has been carried out in a microwave oven and the transformation of organic sulfur and its functional groups was investigated via compact sulfur tester and Fourier transform infrared spectra. The dielectric properties of coal sample are also measured by an Agilent N5244A network analyzer. The result shows that a certain amount of organic sulfur in both NTC and LGC is transformed into iron sulfide, sulfate, and sulfur-containing gas after irradiation of microwave. The retention proportion of the three typical sulfur-containing functional groups in coal is ranking as: disulfide bond (S-S) > sulfinyl (SO) > sulfydryl (-SH), and such phenomenon has been explained by the first principle calculation based on the density functional theory. The calculation results of Fukui indices, electrostatic potential, and frontier orbital analysis indicate the reactivity of the SO and SS is lower than that of the -SH. The calculated bond dissociation enthalpies and bond dissociation times indicate the chemical bonds of -SH and S-S need more time to rupture than SO when microwave power is 1000 W. © 2019 Wiley Periodicals, Inc.

Effects of poplar addition on tar formation during the co-pyrolysis of fat coal and poplar at high temperature.

Three-stage absorption by butyl acetate was used to obtain tar components during the co-pyrolysis of fat coal and poplar at high temperature. The resulting tar yields were calculated relative to the fat coal and poplar blends. The tar components were characterized by gas chromatography-mass spectrometry, Fourier transform-infrared spectroscopy and 1H nuclear magnetic resonance spectroscopy. The effects of the added poplar on tar formation were then considered. The results show that the poplar-fat coal tar yield rose slightly when the poplar addition levels ranged from 4% to 12% and then increased much more at higher poplar addition levels. Oxygenated and aromatic compounds contributed greatly to the poplar-fat coal tar yield. The quantity of oxygenated components increased in the poplar blending ratio range from 4% to 12% and decreased as the ratio increased further, while the quantity of aromatic components showed the opposite trend. The influences of poplar addition levels on tar formation could be divided into two stages: (a) lighten the tar by stabilizing radicals at low poplar addition levels; (b) form heavier tar due to cross-linking reactions of the remaining radicals at high poplar addition levels. When the poplar addition levels ranged from 4% to 12%, due to synergistic effects, large amounts of free radicals and hydrogen from the co-pyrolysis of coal and poplar formed lighter stable compounds, which were then transported into the tar. Further, cross-linking reactions could be decreased because fewer free radicals and less hydrogen remained. As a result, the amount of PAHs declined, the tar yield rose slightly, the hydrocarbon-generating potential improved, the aliphatic chain length shortened, and the aromatic protons decreased. At higher blending ratios, excess radicals existed after stabilization due to the increasing poplar addition levels. These radicals underwent cross-linking reactions and produced PAHs, resulting in heavily increased tar yields, weakened hydrocarbon-generating potential, extended aliphatic chain lengths and increased aromatic protons.

Prevalence and molecular identification of Cryptosporidium spp. in pigs in Henan, China.

The distribution and public health significance of Cryptosporidium species/genotypes in pigs differ among geographic areas and studies. To characterize the prevalence of cryptosporidiosis in pigs in Henan, China, a total of 1,350 fecal samples from 14 farms in ten prefectures in Henan Province were examined. The overall prevalence of Cryptosporidium was 8.2% (111/1,350), with the highest infection rate (79/383 or 20.6%) in 1-2-month-old piglets and the lowest infection rates in 3-6-month-old pigs. Cryptosporidium-positive samples from 108 animals were analyzed by polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis of the small subunit rRNA gene, and 35 were further analyzed by DNA sequencing of the PCR products. Two Cryptosporidium species/genotype were identified, including Cryptosporidium suis (94/108) and the Cryptosporidium pig genotype II (14/108). C. suis infection was more common in younger piglets whereas the pig genotype II was relatively common in older pigs. These findings suggest that pigs are not a major source of zoonotic Cryptosporidium in the study area.