Crystal structures of Bbp from Staphylococcus aureus reveal the ligand binding mechanism with Fibrinogen α

Bone sialoprotein-binding protein (Bbp), a MSCRAMMs (Microbial Surface Components Recognizing Adhesive Matrix Molecules) family protein expressed on the surface of Staphylococcus aureus (S. aureus), mediates adherence to fibrinogen α (Fg α), a component in the extracellular matrix of the host cell and is important for infection and pathogenesis. In this study, we solved the crystal structures of apo-Bbp^273−598 and Bbp^273−598-Fg α^561−575 complex at a resolution of 2.03 Å and 1.45 Å, respectively. Apo-Bbp^273−598 contained the ligand binding region N2 and N3 domains, both of which followed a DE variant IgG fold characterized by an additional D1 strand in N2 domain and D1′ and D2′ strands in N3 domain. The peptide mapped to the Fg α^561−575 bond to Bbp^273−598 on the open groove between the N2 and N3 domains. Strikingly, the disordered C-terminus in the apo-form reorganized into a highly-ordered loop and a β-strand G′′ covering the ligand upon ligand binding. Bbp^{Ala298−Gly301} in the N2 domain of the Bbp^273−598-Fg α^561−575 complex, which is a loop in the apo-form, formed a short α-helix to interact tightly with the peptide. In addition, Bbp^{Ser547−Gln561} in the N3 domain moved toward the binding groove to make contact directly with the peptide, while Bbp^{Asp338−Gly355} and Bbp^{Thr365−Tyr387} in N2 domain shifted their configurations to stabilize the reorganized C-terminus mainly through strong hydrogen bonds. Altogether, our results revealed the molecular basis for Bbp-ligand interaction and advanced our understanding of S. aureus infection process.     

The investigation of Staphylococcus aureus and coagulase-negative staphylococci nasal carriage among patients undergoing haemodialysis

The frequency of nasal staphylococcal colonization among haemodialysed patients was investigated. The swabs were collected in 1998 and 2004 from 28 and 43 patients, respectively.

Staphylococcus aureus colonization rates were 57.1% and 27.9% in 1998 and 2004, respectively. Twenty-six coagulase-negative staphylococci (CNS) isolates were cultured: S. epidermidis (21), S. lugdunensis (2), single S. haemolyticus, S. warneri, and S. capitits isolates. One S. aureus and 10 CNS isolates were methicillin resistant. The methicillin-resistant S. aureus (MRSA) was resistant to β-lactams, tetracycline, and harbored the pvl gene encoding the Panton-Valentine leukocidin.

The decrease in S. aureus colonization at 6-year interval was observed. The presence of the pvl gene and a favorable antibiotic susceptibility pattern of the MRSA suggest that the isolate was a member of community-acquired MRSA (CA-MRSA). Concluding, screening of haemodialysed patients for staphylococcal colonization accompanied by characterization of cultured isolates is important to understand its epidemiology and to develop infection prevention measures and treatment strategies.     

两株链霉菌对非洲菊生长和生理生化指标的影响

本研究以非洲菊(Gerbera jamesonii)为试材,采用不同浓度的脱叶链霉菌FT05W和深蓝链霉菌ZEA17I孢子悬浮液浇灌盆栽非洲菊,筛选出非洲菊的最佳施用浓度,探索两株链霉菌对非洲菊生长和生理生化指标的影响,为非洲菊生产上科学合理施用链霉菌提供理论依据。结果表明: 脱叶链霉菌FT05W和深蓝链霉菌ZEA17I不同浓度孢子悬浮液均能有效促进非洲菊的生长,前者对非洲菊的作用效果优于后者,以浓度为1×10⁹ CFU·mL^-1的脱叶链霉菌FT05W效果最好。与蒸馏水对照相比,该处理显著促进了非洲菊株高(30.2%)和冠幅(41.5%)的生长,并在一定程度上促进了花茎的增长和增粗;能显著提高非洲菊植株叶片的叶绿素含量(65.2%)、根系活力(103.3%)和超氧化物歧化酶活性(84.4%),使丙二醛含量维持在较低的水平。因此在一定的浓度范围内,脱叶链霉菌FT05W和深蓝链霉菌ZEA17I均能有效促进非洲菊的生长,有利于非洲菊植株体内同化物的转运和积累,增强非洲菊植株抗逆性,特别是脱叶链霉菌FT05W具有在未来作为生物肥料解决非洲菊连作障碍的应用潜力。     

家蚕幼虫BmToll9基因对肽聚糖和金黄色葡萄球菌的响应表达

【目的】Toll信号通路是昆虫中重要的免疫信号通路,其中Toll受体在保持Toll通路的正常免疫应答、抵抗外源病原体中起到关键的作用。本研究旨在探究肽聚糖和金黄色葡萄球菌Staphylococcus aureus对家蚕Bombyx mori Toll受体基因BmToll9-1和BmToll9-2表达的影响。【方法】将革兰氏阳性菌细胞壁主要成分肽聚糖和革兰氏阳性菌金黄色葡萄球菌分别注射感染家蚕5龄第1天幼虫,诱导其发生免疫反应;采用实时荧光定量PCR分析注射后不同感染时间点BmToll9-2和BmToll9-1基因在家蚕幼虫中肠、表皮、脂肪体和丝腺中的相对表达水平。【结果】往家蚕5龄幼虫中注射肽聚糖或金黄色葡萄球菌后,BmToll9-2基因出现了时间和组织的差异性表达。注射肽聚糖和金黄色葡萄球菌均能诱导5龄幼虫中肠BmToll9-2基因的表达上调,注射肽聚糖和金黄色葡萄球菌分别在3和6 h时对基因表达的诱导效果最好,且注射金黄色葡萄球菌比注射肽聚糖对基因表达的诱导效果更好。注射金黄色葡萄球菌能引起5龄幼虫表皮、脂肪体和丝腺中BmToll9-2基因的表达上调,分别于注射后24, 6和24 h时诱导效果最好。注射金黄色葡萄球菌亦能诱导同源的BmToll9-1基因的上调表达。【结论】家蚕幼虫BmToll9基因在肽聚糖或金黄色葡萄球菌注射处理后均能在不同组织中发生上调表达,推测BmToll9基因参与了家蚕对肽聚糖和金黄色葡萄球菌的免疫反应。     

Synthesis and evaluation of anticancer benzoxazoles and benzimidazoles related to UK-1

UK-1 is a structurally unique bis(benzoxazole) natural product isolated from a strain of Streptomyces. UK-1 has been reported to possess anticancer activity but no activity against bacteria, yeast, or fungi. Previous work has also demonstrated the ability of UK-1 to bind a variety of di- and tri-valent metal ions, particularly Mg²⁺ ions, and to form complexes with double-stranded DNA in the presence of Mg²⁺ ions. Here we report the activity of UK-1 against a wide range of human cancer cell lines. UK-1 displays a wide spectrum of potent anticancer activity against leukemia, lymphoma, and certain solid tumor-derived cell lines, with IC₅₀ values as low as 20 nM, but is inactive against Staphylococcus aureus, a methicillin-resistant strain of S. aureus, or Pseudomonas aeruginosa. A series of analogues of the bis(benzoxazole) natural product UK-1 in which the carbomethoxy-substituted benzoxazole ring of the natural product was modified were prepared and evaluated for their anticancer and antibacterial properties. An analogue of UK-1 in which the carbomethoxy-substituted benzoxazole ring was replaced with a carbomethoxy-substituted benzimidazole ring was inactive against human cancer cell lines and the two strains of S. aureus. In contrast, a simplified analogue in which the carbomethoxy-substituted benzoxazole ring was replaced with a carbomethoxy group was almost as active as UK-1 against the four cancer cell lines examined but lacked activity against S. aureus. Metal ion binding studies of these analogues demonstrate that they both bind Zn²⁺ and Ca²⁺ ions about as well as UK-1. The non-cytotoxic benzimidazole UK-1 analogue binds Mg²⁺ ions 50-fold weaker than UK-1, whereas the simple benzoxazole analogue binds Mg²⁺ ions nearly as well as UK-1. These results support a role of Mg²⁺ ion binding in the selective cytotoxicity of UK-1 and provide a minimal pharmacophore for the selective cytotoxic activity of the natural product.     

Assessment of the antibacterial activity of selected flavonoids and consideration of discrepancies between previous reports

Activity of the flavonoids apigenin, baicalin and galangin against sensitive and antibiotic resistant strains of Staphylococcus aureus, Enterococcus faecalis, E. faecium, Escherichia coli and Pseudomonas aeruginosa was investigated. Using an agar dilution assay, galangin was shown to have a minimum inhibitory concentration (MIC) of 25 to 50 μg/mL against all six strains of S. aureus but negligible activity against the other species. Apigenin displayed only marginal activity against S. aureus and no activity was detected from baicalin. In inhibition curve studies, galangin caused a 100,000-fold decrease in the viability of a growing population of S. aureus NCTC 6571 within the first two hours of treatment. Decreases in viability of S. aureus NCTC 11561 and NCIMB 9968 populations were also observed.     

PCR fingerprinting for epidemiological studies of Staphylococcus aureus

Staphylococcus aureus isolates (n = 126), collected during two different periods from patients hospitalised in pediatric wards, were analysed using polymerase chain reaction (PCR) mediated genotyping. These isolates were compared with 29 isolates from individuals attending the out-patient clinic of the same hospital and 13 isolates from pediatric hospital personnel. Within a group of 99 isolates gathered from 48 individuals during surveillance period I, 22 distinct genotypes were identified by application of two PCR assays. Among the 58 isolates collected in surveillance period II from pediatric and out-clinic patients, 25 genotypes were detected by a single PCR assay only. Based on these results it was demonstrated that patients can be colonised with multiple strains that may persist in a certain anatomical location for prolonged periods of time. It is shown that persistence of a S. aureus strain in a pediatric ward can be deduced from the PCR genotyping studies. As such PCR can be used for longitudinal monitoring of bacterial infections in hospital departments, analysis of patient-to-patient and personnel-to-patient transmission and for detection of genetic variation in general in S. aureus. Also, isolate-specific DNA probes can be generated for S. aureus by PCR genotyping. The probes can be used for the recognition of re-emerging S. aureus epidemics.     

Field application of an insect virus in the Mekong Delta: Effects of a Vietnamese nucleopolyhedrovirus on Spodoptera litura (Lepidoptera: Noctuidae) and its parasitic natural enemies

A new Vietnamese isolate of Spodoptera litura nucleopolyhedrovirus (NPV) was applied in local soybean fields, and the effect of its application on S. litura larvae was examined. The virus was propagated in vivo and a crude extract was prepared for spraying at high and low doses (1.7×10⁸ and 3.3×10⁷ occlusion bodies/m², respectively). The percentage of larvae infected with NPV increased from 22.2% on the day before NPV application (Day 0) to 50.8% (Day 6) in the high dose treatment plot, and from 7.9% (Day 0) to 35.7% (Day 6) in the low-dose plot. Microsporidium sp. was observed as another major pathogen of S. litura larvae. Three dominant parasitic natural enemies were found in S. litura larvae: Microplitis manilae (Braconidae), Chelonus sp. (Braconidae), and Peribaea orbata (Tachinidae). The fate of parasitoids developing within virus- and Microsporidium- infected hosts differed between these three parasitoids; more Chelonus sp. emerging from infected hosts died during their larval stage before spinning cocoons, or failed to reach adult eclosion, than did P. orbata. This suggests that the impact of virus application on the survival of parasitoids varies from species to species.     

Genotyping of Staphylococcus aureus isolated from humans, bovine subclinical mastitis and food samples in Argentina

The aim of the present study was to characterize genotypically 45 Staphylococcus aureus strains isolated from humans, bovine subclinical mastitis and food samples in Argentina by rep-PCR and PCR amplification of virulence genes. Resistances to various antibiotics could be observed for the human S. aureus, less pronounced for the bovine strains, but not for the eight S. aureus isolated from food samples. The strains could be classified genotypically by rep-PCR and by amplification of the genes encoding protein A, coagulase, clumping factor, the collagen adhesin domains A and B, capsular polysaccharide 5 and 8, the accessory gene regulator agr classes I to III, and the S. aureus gene regulator sae. rep-PCR analyses and the different gene patterns revealed that the strains could be divided into seven groups mostly matching with the origin of the isolates. The present study describes genotypic variations of S. aureus strains isolated from different origins in Argentina. The study provides a valuable insight into molecular specificities of this important pathogen.     

Potential of Serratia marcescens strain B2 for biological control of rice sheath blight

Serratia marcescens strain B2 inhibited mycelial growth of the rice sheath blight pathogen Rhizoctonia solani AG-1 IA. Rice plants were treated with bacterial suspension and then challenge inoculated with the pathogen. Application of S. marcescens effectively reduced the incidence of sheath blight. S. marcescens survived in soil under glasshouse conditions at ca. 10⁸ colony forming units g^-1 of soil for 4 weeks after application. These results suggest that S. marcescens has potential as an effective and persistent biological control agent for rice sheath blight.     

Rapid detection of Staphylococcus aureus strains having reduced susceptibility to vancomycin using a chemiluminescence-based drug-susceptibility test

Current drug-susceptibility tests used routinely in clinical laboratories sometimes fail to identify strains of Staphylococcus aureus with reduced susceptibility to vancomycin. To solve this problem, we have developed a more sensitive and rapid method that measures bacterial metabolic activity by a chemiluminescence-based technique. This method is able to discriminate such strains from vancomycin-susceptible S. aureus with a sensitivity and specificity of > 95%. This rapid and reliable method appears to be promising for detection of vancomycin-intermediate S. aureus strains in clinical laboratories, and may supersede classical susceptibility testing.     

Multifactorial aspects of antimicrobial activity of propolis

We investigated the antibacterial activity of sub-inhibitory concentrations of ethanolic extract of propolis (EEP), and its effect on the antibacterial activity of some antibiotics. Some clinically isolated Gram-positive strains were used.

Moreover, sub-inhibitory concentrations of EEP were used to value its action on some important virulence factors like lipase and coagulase enzymes, and on biofilm formation in Staphylococcus aureus.

Our results indicated that EEP had a significant antimicrobial activity towards all tested clinical strains.

Adding EEP to antibacterial tested drugs, it drastically increased the antimicrobial effect of ampicillin, gentamycin and streptomycin, moderately the one of chloramphenicol, ceftriaxon and vancomycin, while there was no effect with erithromycin.

Moreover, our results pointed out an inhibitory action of EEP on lipase activity of 18 Staphylococcus spp. strains and an inhibitory effect on coagulase of 11 S. aureus tested strains.

The same EEP concentrations showed a negative interaction with adhesion and consequent biofilm formation in S. aureus ATCC 6538P.     

入侵植物加拿大一枝黄花根际解钾菌多样性及解钾活性

入侵植物加拿大一枝黄花(Solidago canadensis)具有较强的钾(K)富集能力, 这可能和其对土壤微生物群落的改变有关。根际解钾菌能够将植物难以利用的矿物态钾转化为植物可以利用的可溶性钾, 而加拿大一枝黄花如何影响根际解钾菌多样性和解钾活性尚未明了。该研究以浙江省杭州湾湿地围垦区内自然生长的加拿大一枝黄花和其伴生本地植物白茅(Imperata cylindrica)为研究对象, 比较了加拿大一枝黄花和白茅体内及土壤中的钾含量水平, 钾供给水平对生物量积累的影响, 以及根际解钾菌的数量、多样性和解钾活性的差异。结果表明, 加拿大一枝黄花茎、叶中的钾含量均显著高于白茅, 分别是白茅的1.59和7.33倍; 加拿大一枝黄花和白茅的土壤全钾含量差异不显著, 速效钾含量在0-10 cm土层中差异显著、在10-20 cm土层中差异不显著。随着钾供应水平提高, 加拿大一枝黄花和白茅的生物量均显著增加。利用解钾培养基计数培养后发现, 加拿大一枝黄花根际解钾菌的数量是白茅的3.51倍。分离培养后将出现解钾圈的菌株进行鉴定, 利用解钾液体培养实验测定其解钾量, 发现从加拿大一枝黄花根际土中分离得到的15个解钾菌株中, 有9个具有高效解钾能力, 其处理液中K ⁺含量较空白对照高出85.11%-192.54%, 其中菌株H2-20解钾能力最强, 解钾量为10.657 mg·L ^-1。加拿大一枝黄花根际解钾菌解钾作用显著高于白茅。经16S rDNA鉴定发现, 加拿大一枝黄花15个根际解钾菌株分属11个属, 其中有6个属已经被报道证实具有明显解钾能力。这些结果表明加拿大一枝黄花根际解钾菌数量较为丰富, 且大多具有较高解钾活性, 可能对其钾富集具有重要贡献。     

Biodiversity of Saccharomyces cerevisiae isolated from a survey of pito production sites in various parts of Ghana

Biodiversity among Saccharomyces cerevisiae predominating the spontaneous fermentation of Dagarti pito in Ghana was assessed. Two hundred and forty-nine isolates obtained from samples of dried yeast taken from commercial pito production sites in eight geographical regions of Ghana were characterized phenotypically by colony and cell morphology as well as carbohydrate assimilation profiling. Yeast populations ranged between 10⁶ and 10⁸ cfug⁻¹. Ninety-nine percent of the isolates (247) investigated showed macro-and micro morphological characteristics typical of S. cerevisiae. Of these, 72% (179) had assimilation profiles similar to S. cerevisiae while 28% (68) had assimilation profiles atypical of S. cerevisiae or any other member of the Saccharomyces sensu stricto complex. Amplification of the region spanning the two intergenic transcribed spacers (ITS) and the 5.8S ribosomal gene (ITS1-5.8S rDNA-ITS2), followed by restriction analysis, as well as determination of chromosome length polymorphism by pulsed field gel electrophoresis (PFGE) of 25 representative isolates strongly indicated that all belonged to S. cerevisiae, notwithstanding the phenotypic differences. Sequencing of the mitochondrial cytochrome-c oxidase II gene (COX 2) and the actin-encoding gene (ACT1) of four isolates, confirmed their close relatedness to S. cerevisiae, particularly to the type strain CBS1171 (98.7%), as well as other members of the Saccharomyces sensu stricto complex. Twenty isolates selected from eight geographical regions of Ghana and investigated for their technological properties, showed different patterns of growth and flocculation but otherwise similar technological characteristica. Most of the isolates produced pito having sensory attributes, which compared favourably with commercially produced pito.     

外源茉莉酸甲酯对干旱胁迫下狭叶黄芩光合和生理特性的影响

以狭叶黄芩为试验材料,测定了不同浓度(0.1、0.5、1、2 mmol·L^-1)茉莉酸甲酯对干旱胁迫下(15%,PEG-6000)狭叶黄芩表型特征、光合以及生理特性的影响。结果外源施加茉莉酸甲酯增强了狭叶黄芩的净光合速率、气孔导度和蒸腾速率。有效降低了狭叶黄芩受伤害等级。可以有效缓解降低了胞间CO2浓度,其中施用2 mmol·L^-1茉莉酸甲酯9 d后对狭叶黄芩的净光合速率、蒸腾速率、气孔导度、胞间CO2浓度的影响最为显著。处理组的游离脯氨酸含量低于CK。可溶性蛋白含量在前6 d的处理中低于CK,而在6 d以后的处理中显著高于CK。外源施加茉莉酸甲酯能够提高SOD、POD、CAT活性。对狭叶黄芩的光合参数和生理指标的研究结果表明,经外源施加的茉莉酸甲酯处理能够有效减缓干旱胁迫对狭叶黄芩的不良影响。     

苦参的染色体核型

苦参(Sophora flavescens Ait)是豆科槐属(Sophora L.)的多年生草本植物,我国南北各省区均产,苏联、朝鲜和日本亦有分布。在细胞学方面,Kawakami^[3]、Hexob^[8]和Kodama^[4]都报道过苦参的染色体数目为2n=18,但Nagl^[8]却报道2n=28。本文作者在对苦参作染色体基数的基础上,进行了核型的描述和分析。     

柳树新无性系(A42)对富营养水体不同浓度磷的吸收和净化机制

磷是评价水体富营养化的因子之一,也是植物生长必需营养元素,研究速生木本植物对富营养水体中磷的吸收具有重要意义.本研究以旱柳新无性系A42为对象,于2017年7—9月在温室大棚中进行浮床水培试验,研究了旱柳对不同磷营养水平水体(低磷0.1、0.2 mg·L^-1;中磷1.0、2.0 mg·L^-1;高磷10.0 mg·L^-1)的吸收和净化机制.结果表明:旱柳能有效净化水体中的磷营养(21 d达到79%以上),去除量与水体磷浓度呈正相关,但去除率随水体磷浓度增加呈先升高后降低趋势.旱柳可于7 d内将磷浓度为0.1～1 mg·L^-1的富营养水体中磷浓度降低至富营养阈值(0.016～0.032 mg·L^-1).旱柳富集同化的磷含量占水体磷总输入量的29.0%～66.9%,富集同化量与富集同化率分别与水体磷浓度呈正相关和负相关.旱柳在不同磷浓度水体中均能正常生长,根冠比随水体磷浓度下降而显著增加.氮、磷在旱柳体内积累均表现为茎>叶>根,旱柳的氮、磷转运系数均大于3,在高磷浓度水体中,氮磷营养在旱柳的茎部大量积累,氮、磷转运系数分别显著增加至4.53±0.24和4.92±0.62.表明旱柳在不同磷浓度富营养水体中均能正常生长且有良好的净化能力,能够通过富集转运磷营养至地上部来减少二次污染.实际应用中,对于低磷浓度水体,适合做短期净化;对于高磷浓度水体,适合做长期净化.     

缢蛏(Sinonovacula constricta)对海水中镉富集规律的研究

论文利用微波消解-石墨炉原子吸收法,研究在不同Cd²⁺浓度(0.005 mg·L^-1、0.025 mg·L^-1、0.05 mg·L^-1、0.1 mg·L^-1)暴露下缢蛏对水体Cd²⁺的富集规律。结果表明,缢蛏对重金属Cd²⁺有一定的富集能力,其软体组织对Cd²⁺的富集量、富集速率随暴露剂量增加而升高,并在168 h的暴露时间内,体内Cd含量及其富集倍数(BCM)与暴露时间呈显著的正相关关系(P<0.01);缢蛏不同组织对Cd的富集能力(富集量、富集速率、富集倍数)存在显著差异,其消化腺、鳃和肌肉组织对Cd富集能力的大小表现为:消化腺>鳃>肌肉。与其他贝类对Cd的富集情况相比,缢蛏不是Cd的强净积累者。     

Influence of culture medium composition, dissolved oxygen concentration and harvesting time on the production of Serratia entomophila, a microbial control agent of the New Zealand grass grub

The bacterium Serratia entomophila (Enterobacteriaceae) has been developed as a commercially available biopesticide for control of the pasture pest Costelytra zealandica. The influence of culture medium composition, dissolved oxygen (DO) concentration and harvesting time were investigated in order to optimise the production of S. entomophila. In batch fermentations, highest yields were achieved using sucrose (40 g L^-1) as the carbon source, followed closely by fructose and molasses. The effect of yeast extract (YE), marmite and bakery yeast as cell growth enhancers was also examined in both batch and fed-batch mode. Culture medium containing 20 g L^-1 of YE (fed-batch) produced the highest cell density. No significant effect on cell yield was detected when cultures were supplemented with bakery yeast or marmite. The DO concentration influenced biomass production: a 5-fold increase in cell density was achieved when the concentration of DO was maintained in the range of 20-50% (5.7×10¹⁰ CFUs mL^-1) in comparison with 1% (1.2×10¹⁰ CFUs mL^-1). In cultures maintained at 1 and 20% DO concentration, cells harvested from the exponential growth phase survived for less than 2 weeks when stored at 4°C. In contrast, high cell survival (85-100%) was achieved when cells were harvested after they had entered the stationary growth phase. Recommendations are provided for the production of robust, high cell density cultures of S. entomophila.     

Biological control of Strophosoma spp. (Coleoptera: Curculionidae) in greenery (Abies procera) plantations using Hyphomycetes

In Abies procera plantations Strophosoma melanogrammum and S. capitatum cause economic damage due to the adult stage feeding on the needles. No chemical treatments of these weevils are allowed in Denmark, so the potential of biological control was evaluated. We studied pathogenicity of thirteen isolates of entomopathogenic fungi and the field effect of soil application of Metarhizium anisopliae against Strophosoma spp. All of the tested isolates were capable of causing infections under laboratory conditions and average survival time at 20°C ranged between 13 and 23 days for S. melanogrammum and between 23 and 28 days for S. capitatum when dipped in a fungal suspension adjusted to 1×10⁷ conidia mL^-1. Under field conditions up to 90% of the living collected individuals died due to M. anisopliae infection in the treated plots, whereas less than 1% died of M. anisopliae infection in control plots. In accordance, the population of Strophosoma spp. was reduced in plots where the fungus was applied compared to control plots.