在大肠杆菌K99(中国QH株)菌毛形成亚单位基因(fanC)的克隆和测序中发现IS1 插入序列

目的 构建大肠杆菌K99菌毛表达载体。方法 通过逐级亚克隆的方法克隆k99菌毛形成亚单位基因（fanC），并进行序列测定和fanC亚单位的抗原决定簇模拟，以确定突变部位。结果 克隆了包括K99菌毛形成亚单位基因（fanC）和部分fanD基因在内的一段基因，该基因与国外报道相比明显偏大。序列测定表明：在fanC尾部多出 24bp，在fanC与 fanD，的结合部有一 800bp左右的新基因。经与基因库中大肠杆菌基因作同源性比较发现，该新基因为768bp的插入序列IS1。其他基因为K99自身fonC与fanD片段之间的过渡基因。IS1的插入未改变fanD，基因的阅读框架，只是使fanD片段末端增加了 8个氨基酸。fanC亚单位的抗原决定簇模拟结果显示，K99 fanC片段具有一个非常突出的亲水区域，根据经验它应是抗原决定簇所在的部位。结论 首次发现大肠杆菌K99基因中含有插入序列IS1。     

Steric hindrance in immunolabelling

In this paper we describe immunocytochemical detection of PhoE pore protein in the outer membrane of E. coli K-12 cells in dependence of a variety of labelling approaches. Immuno-gold labelling on ultrathin cryosections showed a uniform, dense labelling of the outer membrane of all cells. However, using immunofluorescence, whole-mount or freeze-etch labelling methods, labelling was limited to less than 5% of the cell population. In order to understand this phenomenon, immunoincubated cells exhibiting less than 5% labelling were processed for cryo-ultramicrotomy. Reincubation of the sections with antibody and probe resulted in labelling of all of the cells. If an E. coli Gal-U mutant strain, defective in the lipopolysaccharide (LPS) carbohydrate chain length, was used, each approach rendered 100% labelling. From these results it is concluded that the antigenic sites of the PhoE pore protein are not accessible in intact ‘wild-type’ cells due to steric hindrance caused by the LPS carbohydrate chains. In cryosections this steric hindrance is partly precluded since antigenic determinants are exposed at the section surface in transversely cut membranes. Our results emphasize the necessity to compare results obtained by means of several, basically different approaches.     

Green and gentle synthesis of Cu2O nanoparticles using lignin as reducing and capping reagent with antibacterial properties

In this work, Cu₂O nanoparticles of a particular shape were prepared by an eco-friendly, gentle and low-cost synthetic method using lignin as a reducing and capping reagent. Structure and morphology of the Cu₂O nanoparticles were characterised by high-resolution transmission electron microscopy, X-ray photoelectron spectroscopy, X-ray diffraction (XRD) and Fourier-transform (FT-IR) spectroscopy. The results established that Cu₂O nanoparticles coated by lignin showed a particular shape. The morphology of Cu₂O nanoparticles presented as some loose accumulation of particles just like broccoli, and the particle size range was between 100 and 200 nm. And, the XRD revealed the structure of crystalline of the Cu₂O nanoparticles. In addition, the sterilisation of Cu₂O nanoparticles on Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) was also investigated. The Cu₂O nanoparticles showed effective bactericidal activity against E. coli and S. aureus. The antibacterial rate could get 100% after 30 min with 4.0 g/L Cu₂O nanoparticles. Furthermore, the Cu₂O nanoparticles were confirmed to have low cytotoxicity.     

Expression of Aspergillus nidulans phy Gene in Nicotiana benthamiana Produces Active Phytase with Broad Specificities

A full-length phytase gene (phy) of Aspergillus nidulans was amplified from the cDNA library by polymerase chain reaction (PCR), and it was introduced into a bacterial expression vector, pET-28a. The recombinant protein (rPhy-E, 56 kDa) was overexpressed in the insoluble fraction of Escherichia coli culture, purified by Ni-NTA resin under denaturing conditions and injected into rats as an immunogen. To express A. nidulans phytase in a plant, the full-length of phy was cloned into a plant expression binary vector, pPZP212. The resultant construct was tested for its transient expression by Agrobacterium-infiltration into Nicotiana benthamiana leaves. Compared with a control, the agro-infiltrated leaf tissues showed the presence of phy mRNA and its high expression level in N. benthamiana. The recombinant phytase (rPhy-P, 62 kDa) was strongly reacted with the polyclonal antibody against the nonglycosylated rPhy-E. The rPhy-P showed glycosylation, two pH optima (pH 4.5 and pH 5.5), an optimum temperature at 45~55 °C, thermostability and broad substrate specificities. After deglycosylation by peptide-N-glycosidase F (PNGase-F), the rPhy-P significantly lost the phytase activity and retained 1/9 of the original activity after 10 min of incubation at 45 °C. Therefore, the deglycosylation caused a significant reduction in enzyme thermostability. In animal experiments, oral administration of the rPhy-P at 1500 U/kg body weight/day for seven days caused a significant reduction of phosphorus excretion by 16% in rat feces. Besides, the rPhy-P did not result in any toxicological changes and clinical signs.     

猕猴桃汁的紫外线杀菌效果研究

研究了猕猴桃果汁膜厚度、辐射距离、时间和温度4个因素对紫外线杀灭接种在猕猴桃清汁中的大肠杆菌的影响。并通过正交试验得出各因素的最佳组合。结果表明:处理时间和果汁厚度对紫外线的杀菌效果影响最为显著。当果汁厚度为0.5mm,辐射时间为5min,辐射距离为100mm,杀菌温度为30℃时,杀菌率可达99.97%。     

Oxygen-dependent enhancement of hydrogen production by engineering bacterial hemoglobin in Escherichia coli

H₂ production under aerobic conditions has been proposed as an alternative method to overcome the fundamentally low yield of H₂ production by fermentative bacteria by maximizing the number of electrons that are available for H₂. Here, we engineered Vitreoscilla hemoglobin (VHb) in Escherichia coli to study the effects of this versatile oxygen (O₂)-binding protein on oxic H₂ production in a closed batch system that was supplemented with glucose. The H₂ yields that were obtained with the VHb-expressing E. coli were greatly enhanced in comparison to the negative control cells in culture that started with high O₂ tensions. The formate hydrogen lyase (FHL) activity of oxically cultured, VHb-expressing cells was also much higher than that of the negative control cells. Through inhibitor studies and time-course experiments, VHb was shown to contribute to the improved H₂ yield primarily by increasing the efficiency of cellular metabolism during the aerobic phase before the onset of H₂ production and not by working as an O₂-scavenger during H₂ production. This new approach allowed more substrate to remain to be further utilized for the production of more H₂ from limited resources. We expect that VHb can be successfully engineered in potential aerobic H₂-producing microbial systems to enhance the overall H₂ production yield. In addition, the remarkably high FHL activity of oxically grown, VHb-expressing cells may make this engineered strain an attractive whole-cell biocatalyst for converting formate to H₂.     

Escherichia coli contamination of the river Thames in different seasons and weather conditions

Contamination of public water ways with sewage represents a serious environmental and health risk. We monitored pollution of the river Thames by enumerating the indicator organism Escherichia coli. Samples were taken from a site in central London near Waterloo Bridge in different seasons. E. coli were quantified using a membrane filtration method, and correlated with the tidal variations of the river and meteorological data on rainfall and temperature. More frequent and severe incidents of pollution occurred in the autumn. Heavy rainfall resulted in sharp peaks of E. coli contamination that implies a potential increase of numbers of pathogenic micro‐organisms. Sixty percent of all samples were found to be in excess of the accepted upper limit of pollution set by European Union (EU) legislation for bathing water. This study demonstrated that frequent sewage pollution of the Thames results in high numbers of E. coli and incidents of detectable levels of pathogenic bacteria demonstrating the need for regular monitoring of bacterial pollution.     

Modelling concentration gradients in fed-batch cultivations of E. coli – towards the flexible design of scale-down experiments

The cover image is based on the Research Article Modelling concentration gradients in fed-batch cultivations of E. coli - towards the flexible design of scale-down experiments by Emmanuel Anane et al., DOI: 10.1002/jctb.5798 .

     

Chemical warfare and survival strategies in bacterial range expansions

Dispersal of species is a fundamental ecological process in the evolution and maintenance of biodiversity. Limited control over ecological parameters has hindered progress in understanding of what enables species to colonize new areas, as well as the importance of interspecies interactions. Such control is necessary to construct reliable mathematical models of ecosystems. In our work, we studied dispersal in the context of bacterial range expansions and identified the major determinants of species coexistence for a bacterial model system of three Escherichia coli strains (toxin-producing, sensitive and resistant). Genetic engineering allowed us to tune strain growth rates and to design different ecological scenarios (cyclic and hierarchical). We found that coexistence of all strains depended on three strongly interdependent factors: composition of inoculum, relative strain growth rates and effective toxin range. Robust agreement between our experiments and a thoroughly calibrated computational model enabled us to extrapolate these intricate interdependencies in terms of phenomenological biodiversity laws. Our mathematical analysis also suggested that cyclic dominance between strains is not a prerequisite for coexistence in competitive range expansions. Instead, robust three-strain coexistence required a balance between growth rates and either a reduced initial ratio of the toxin-producing strain, or a sufficiently short toxin range.     

Antibacterial effects of chitosan coating containing Mentha aquatica L. essence against Escherichia coli,Staphylococcus aureus and Listeria monocytogenes in Iranian white cheese

The effect of a chitosan coating and Mentha aquatica L. essence on Iranian white cheese was investigated. Results showed 100% inhibition of Escherichia coli growth using 1.5% essence after 10 days. After 15 days of incubation, the Staphylococcus aureus population was reduced by 44.2%, 70.0%, and 88.5% using 0.5, 1.0 and 1.5% essence, respectively. After 15 days, Listeria monocytogenes growth was inhibited by 63.84%, 70.12%, and 85.9% using 0.5, 1.0, and 1.5% essence, respectively. Inhibition zone diameter studies also confirmed the antibacterial effects of applied coating against all the above‐mentioned bacteria in Iranian white cheese.