Microbiologically influenced corrosion resistance enhancement of coppercontaining high entropy alloy Fe_(x)Cu_(1−x)CoNiCrMn against Pseudomonas aeruginosa

To enhance the microbiologically influenced corrosion(MIC)resistance of FeCoNiCrMn high entropy alloy(HEAs),a series of Fe_(x)Cu_((1−x))CoNiCrMn(x=1,0.75,0.5,and 0.25)HEAs were prepared.Microstructural characteristics,corrosion behavior(morphology observation and electrochemical properties),and antimicrobial performance of Fe_(x)Cu_((1−x))CoNiCrMn HEAs were evaluated in a medium inoculated with typical corrosive microorganism Pseudomonas aeruginosa.The aim was to identify copper-containing FeCoNiCrMn HEAs that balance corrosion resistance and antimicrobial properties.Results revealed that all Fe_(x)Cu_((1−x))CoNiCrMn(x=1,0.75,0.5,and 0.25)HEAs exhibited an FCC(face centered cubic)phase,with significant grain refinement observed in Fe_(0.75)Cu_(0.25)CoNiCrMn HEA.Electrochemical tests indicated that Fe_(0.75)Cu_(0.25)CoNiCrMn HEA demonstrated lower corrosion current density(i_(corr))and pitting potential(E_(pit))compared to other Fe_(x)Cu_((1−x))CoNiCrMn HEAs in P.aeruginosa-inoculated medium,exhibiting superior resistance to MIC.Anti-microbial tests showed that after 14 d of immersion,Fe_(0.75)Cu_(0.25)CoNiCrMn achieved an antibacterial rate of 89.5%,effectively inhibiting the adhesion and biofilm formation of P.aeruginosa,thereby achieving resistance to MIC.

Study on Distribution of Four Pseudomonas Species in Living Environment Using Multiplex PCR

Purpose: The genus Pseudomonas is a ubiquitous microorganism frequently detected from immunocompromised patients. The inherent resistance to numerous antimicrobial agents contributes to the opportunistic character of this pathogen exhaustive monitoring of this pathogen is considered of critical importance to public health organizations. The reliable identification method able to distinguish genetic close Pseudomonas species is needed, because these organisms are difficult to differentiate by phenotypic or biochemical methods. The purpose of the present study was to design species-specific primers in order to identify and detect four Pseudomonas species which are frequently detected from the human oral cavities, and to investigate the distribution of these organisms in the living environment using a multiplex PCR. Methods: Polymerase chain reaction (PCR) primers were designed based on partial sequences of the rpoD gene of four Pseudomonas species. Swab samples were collected from fifty washstands, and the distribution of Pseudomonas species was investigated using a conventional PCR at genus level and a multiplex PCR at species level. Results: Multiplex PCR method developed in this study was able to distinguish four Pseudomonas species clearly. The genus Pseudomonas was detected from all samples (100%), whereas P. putida, P, aeruginosa, P. stutzeri and P. fluorescens were detected at 44%, 8%, 4% and 2% in fifty swab samples, respectively. Conclusion: Our developed one-step multiplex PCR method is accurate, specific, cost-effective, time-saving, and works without requiring DNA extraction. It was indicated that washstands were the uninhabitable environment for P. putida, P, aeruginosa, P. stutzeri and P. fluorescens.

Thoracic spine infection caused by Pseudomonas fluorescens:A case report and review of literature

BACKGROUND The clinical incidence of spinal infection is gradually increasing,and its onset is insidious,easily leading to missed diagnosis and misdiagnosis,which may lead to serious complications such as nervous system dysfunction,spinal instability and/or deformity,and cause a huge burden on society and families.Early identification of the causative agent and precision medicine will greatly reduce the suffering of patients.At present,the main pathogenic bacteria that cause spinal infection are Staphylococcus aureus,Streptococcus,Pneumococcus,Escherichia coli,and Klebsiella.There are no reports of spinal infection caused by Pseudomonas fluorescens.CASE SUMMARY We report a 32-year-old female patient with spinal infection.She presented with flank pain,initially thought to be bone metastases or bone tuberculosis,and had a family background of tumors.Her clinical features and changes in imaging and laboratory tests led to the suspicion of thoracic spine infection.Histopathology of the lesion showed inflammation,tissue culture of the lesion was negative several times,and the possible pathogen-Pseudomonas fluorescens was found after gene sequencing of the lesion.The patient recovered completely after a full course of antibiotic treatment.CONCLUSION This report increases the range of pathogens involved in spinal infections,highlights the unique advantages of gene sequencing technology in difficult-todiagnose diseases,and validates conservative treatment with a full course of antibiotics for spinal infections without complications.

Molecular investigation of exoU and exoY virulence genes in Pseudomonas aeruginosa collected from hospitalized patients in North of Iran:A descriptive-analytical study

Objective:To investigate the frequency of exoU and exoY genes in patients with Pseudomonas aeruginosa infection.Methods:In this study,100 clinical isolates of Pseudomonas aeruginosa were collected from patients hospitalized in educational-therapeutic hospitals and were identified using standard microbiological tests.Then,the antibiotic resistance pattern of the isolates was determined by the disk agar diffusion method.The bacterial DNAs were extracted by the alkaline lysis method.Finally,the presence of exoU and exoY genes was evaluated by the PCR test.Results:In this study,47%,72%,29%,39%,40%,and 44%of the isolates were non-susceptible to piperacillin,aztreonam,ceftazidime,imipenem,tobramycin,and ciprofloxacin,respectively.In addition,95%and 93%of the clinical isolates carried the exoU and exoY genes.Blood and fecal isolates had both virulence genes,while only one wound isolate had neither genes.Meanwhile,all urinary isolates contained the exoY gene and only one isolate lacked the exoU gene.Also,88 isolates simultaneously had both exoU and exoY genes.Conclusions:High prevalence of exoU and exoY genes in this region indicates a significant role of typeⅢsecretion system in pathogenesis of Pseudomonas aeruginosa.The typeⅢsecretion system may be a suitable target to reduce the pathogenicity of this bacterium.

Efficacy of Pudilan Xiaoyan Oral Liquid in the treatment of pneumonia induced by drug-resistant Pseudomonas aeruginosa

Background:Pudilan Xiaoyan Oral Liquid(PDL)is a Chinese patent medicine with notable pharmacological properties,including anti-inflammatory and antibacterial effects.Drug-resistant Pseudomonas aeruginosa infection is a common and refractory bacterial infection in clinical practice.Due to its high drug resistance,it brings great challenges to treatment.This study aimed to assess the therapeutic efficacy of PDL in a murine model of pneumonia induced by drug-resistant Pseudomonas aeruginosa.Methods:Three different doses of PDL(11 mL/kg/d,5.5 mL/kg/d,2.75 mL/kg/d)were used to observe lung tissue pathology and inflammatory cytokine levels in pneumonia mouse models induced by multidrug-resistant Pseudomonas aeruginosa(MDR-PA).Additionally,the protective efficacy of PDL against mortality in infected mice was evaluated using a death model caused by MDR-PA.Finally sub-MIC concentration of levofloxacin was used to induce drug-resistant mice pneumonia model to evaluate the role of PDL in reversing drug resistance.Experimental data are expressed as mean±standard deviation.Statistical significance was determined by one-way analysis of variance followed by Tukey’s multiple-comparisons test.Results:Treatment effect of PDL on MDR-PA pneumonia:the medium and small doses of PDL can significantly reduce the lung index of multi-drug resistant bacteria infected pneumonia model mice(P<0.05),the lung index inhibition rates for these groups were 55.09%and 58.43%,and improve the degree of lung tissue lesions of mice;The expression of serum cytokines keratinocyte chemoattractant,tumor necrosis factor-αand monocyte chemoattractant protein-1 could be decreased in the three dosage groups of PDL(P<0.01).PDL treatment not only lowered the mortality but also extended the survival duration in mice infected with MDR-PA.It was found after sub-MIC concentration of levofloxacin induced resistance of Pseudomonas aeruginosa to pneumonia in mice.Compared with the model group,the lung index of mice in high and medium PDL doses was significantly reduced(P<0.05),with inhibition rates of 32.16%and 37.73%,respectively.Conclusion:PDL demonstrates protective effects against MDR-PA infection pneumonia,notably decreasing serum inflammatory factor levels.It shows promise in mitigating antibiotic resistance and offers potential for treating pneumonia resulting from Pseudomonas aeruginosa resistance.

荧光假单胞菌Pseudomonas fluorescens RN-88抑菌特性研究

本研究旨在利用拮抗菌防治植物病害,为桃的病害生物防治提供更多可行性的方案。以前期筛选获得的一株桃褐腐病拮抗细菌荧光假单胞菌(Pseudomonas fluorescens)RN-88为试验材料,采用杯碟法测定发酵液对桃褐腐菌(Monilinia fructicol)的抑菌活性,并结合抑菌圈法,研究了该菌在pH、温度、紫外线(UV)等不同环境中的稳定性。实验结果表明,细菌RN-88能显著降低桃褐腐病的发病率,其发酵液对褐腐病菌(M.fructicol)的抑制率可达44.69%。在过滤除菌后,相对于高温灭菌,发酵液中抑菌物质的损失较小。在pH 7.5~9.0的碱性溶液中,发酵液的抑菌活性有较好的稳定性,但在酸性溶液(3.0≤pH≤6.5)中处理后,其抑菌能力减弱。发酵液的抑菌率几乎不受温度和紫外线(UV)的影响。Pseudomonas fluorescens RN-88细菌具有较好的环境稳定性,具有开发成桃褐腐病生物拮抗剂的潜在价值,本研究为荧光假单胞菌作为新型生防菌的应用提供了理论依据,同时为实现桃病害的生物防治提供了更多可行性的方案。

外源甜菜碱提高恶臭假单胞菌(Pseudomonas putida)DLL-1耐盐性的研究

研究了外源甜菜碱对恶臭假单胞菌(Pseudomonas putida)DLL-1耐盐性的影响并对其渗透保护机制进行了初步的探讨;结果表明培养基中添加甜菜碱可以改善DLL-1细胞在高盐培养基中的生长情况,添加150mg/L的甜菜碱可以使DLL-1在1.2mol/L NaCl的基础盐培养基中生长,添加10mg/L的甜菜碱就足以显著缩短渗透胁迫条件下DLL-1细胞的延滞期和代时,增加生长量;和不添加对照相比,延滞期由24h缩短到6h,代时由60min缩短到35.7min,最大生长量OD610由1.29增长到1.57。在渗透胁迫条件下,细胞从外界快速吸收外源甜菜碱来代替自身相容性溶质的合成。

趋化信号转导基因cheA突变对Pseudomonas putida DLL-1甲基对硫磷的趋化性及原位降解的影响

Pseudomonas putida DLL-1是一株甲基对硫磷（MP）高效降解菌株,同时对MP具有趋化性.cheA基因是菌株趋化信号转导过程中负责编码组氨酸激酶的基因,为了研究菌株趋化性在农药原位降解中的作用,通过基因打靶的方式使P.putida DLL-1染色体上单拷贝的cheA基因失活,成功地获得了MP的趋化突变株P.putida DAK,突变株与野生菌株生长能力没有显著差异.通过土壤盆钵试验（MP浓度为50mg/kg）,发现在灭菌与未灭菌土壤中趋化突变株对MP的降解能力低于原始出发菌株DLL-1约20%～30%,说明菌株DLL-1趋化性的丧失会减慢其对农药的降解,趋化性在农药的原位降解过程中发挥重要作用.

固定化Pseudomonas putida CGMCC3830转化3-氰基吡啶制备烟酸

采用固定化Pseudomonas putida CGMCC3830转化3-氰基吡啶制备烟酸。选择海藻酸钠作为包埋材料进行固定化,考察固定化细胞的制备条件、转化条件以及批次稳定性。结果表明,优化的固定化条件为:海藻酸钠2 g/dL,氯化钙0.4 g/dL,固化时间6 h;最适转化条件为:温度35℃,pH 7.0,底物浓度100 mmol/L。批次转化实验结果显示,当底物浓度为100 mmol/L时,固定化细胞重复使用10次,酶活仍保留59.1%,产物烟酸的得率为91.8 g/g细胞干重,而游离细胞的使用3次后,酶活下降至45.6%。

不同生长期的Pseudomonas putida 5-x细胞对工业废水中Cu^(2+)的吸附

研究了不同生长阶段的Cu2 + 吸附细菌Pseudomonasputida 5 x对Cu2 + 的吸附容量 .结果表明在低硫培养基中 ,当P putida 5 x细胞生长刚进入对数生长期时 ,单位细胞对Cu2 + 的吸附溶量最低 .而当生长进入稳定生长期的后期或死亡期的早期时 ,单位细胞对Cu2 + 的吸附量最大 .实验结果还表明 ,高浓度的Cu2 + 对对数期细胞的吸附能力有一定的抑制 ,而对死亡期的细胞影响较小 .早期研究已证明细胞对重金属离子的吸附和细胞表面某些对重金属离子具有亲和性的功能基团 (如某些膜蛋白及羰基化合物 )有关 .利用透视电子显微镜观察了不同生长阶段的细胞表面结构及对铜离子的吸附效果 ,发现生长后期细胞表面吸附有较多的Cu2 + 。

Pseudomonas putida S1海藻糖合成酶基因在大肠杆菌中的克隆表达

利用PCR和TA克隆方法扩增和克隆得到了恶臭假单胞菌Pseudomonas putidaS1的海藻糖合成酶基因treS。对其进行序列分析表明,其编码区含有2067bp,编码含688个氨基酸残基的蛋白质,其核苷酸序列和蛋白质序列与来源于其它假单胞菌属细菌的海藻糖合成酶的序列表现出了较高同源性。将该基因序列与表达载体pQE30T连接,构建重组质粒pQE30T-TS,并将其转化至E.coliM15菌株中。重组菌株经诱导表达后SDS-聚丙烯酰胺凝胶电泳结果显示有明显的分子量约77.5kD的特异蛋白条带出现。经测定酶活力达19U/mL,约是原始菌株P.putidaS1的50倍。

Pseudomonas putida MnB1氧化Mn^2＋实验研究

Pseudomonas putida MnB1是存在于淡水和土壤中的锰氧化细菌。本文在中性条件下对MnB1锰氧化特性及锰氧化产物进行了研究。XANES结果显示,MnB1可将溶液中Mn(Ⅱ)氧化并形成Mn(Ⅳ)的锰氧化物;XRD结果进一步证实锰氧化物为δ-MnO2。MnB1的锰氧化性能受重金属离子的影响,其中Cd2+影响最大、Cu2+影响最小。在4μM Cd2+、6μM Pb2+、200μM Cu2+和20μmol/L Ni 2+的体系中,锰氧化速率和氧化量均有所降低,锰氧化量由208.5μM分别降至20.3μM、192.5μM、129.5μM和52.7μM。

Pseudomonas putida MnB1氧化分解菱锰矿的实验研究

自然界中,菱锰矿氧化形成锰的氧化物矿物是非常普遍的现象,在菱锰矿被氧化分解发生物相转变的过程中,碳酸盐溶解和锰的氧化往往同时发生,微生物可能起着催化作用。选取锰氧化模式菌株Pseudomonas putida Mn B1和广西梧州菱锰矿,通过菱锰矿在该细菌作用下发生转变的实验,利用场发射扫描电镜、扫描透射X射线显微成像等分析方法,研究了矿石表面形貌变化以及锰元素在细胞上的分布特征。结果表明细菌显著促进的菱锰矿的溶解,在此基础上,进一步探讨了细菌在菱锰矿氧化过程中的贡献,本实验结果丰富了次生锰矿床的微生物成因研究。

Pseudomonas putida KT2442制备中长链聚羟基烷酸酯

初步探讨了利用PseudomonasputidaKT2442制备中长链聚羟基烷酸酯(MCL PHAs)的工艺,考察了不同碳源对菌体生长和产物合成的影响。在2.5L发酵罐上采用高密度细胞培养制备:以油酸为碳源,细胞干重达120g L,MCL PHAs质量浓度达40g L,产物质量分数为34%,生产速率为0.965g·L-1·h-1;以玉米油水解物为碳源,细胞干重达105g L,MCL PHAs达28g L,产物质量分数为26.5%,生产速率为0.667g·L-1·h-1。以玉米油水解物为碳源所得细胞浓度、产物浓度与以油酸为碳源所得结果比较接近,有利于降低生产成本。

Pseudomonas putida ZWL73经龙胆酸途径分解代谢3-羟基苯甲酸

一株4-氯硝基苯(4-CNB)的完全降解菌Pseudomonas putidaZWL73能利用3-羟基苯甲酸(3-HBA)为惟一碳源和能源生长,而且ZWL73在对3-HBA降解的过程中,有中间产物龙胆酸生成;酶学分析的分光光度法证实,ZWL73通过GSH依赖型的龙胆酸途径代谢3-HBA,并且该途径需经3-HBA或龙胆酸诱导.

Pseudomonas putida S1海藻糖合成酶表达质粒的构建及诱导条件优化

采用PCR方法从Pseudomonas putidaS1中克隆出编码海藻糖合成酶的基因treS,并与质粒pQE30T相连,构建了表达质粒pQE-TS2。将此重组质粒转化宿主菌E.coliM15进行诱导表达。十二烷基磺酸钠-聚丙烯酰胺凝胶SDS-PAGE电泳结果表明,treS基因在大肠杆菌中获得了高效表达。通过对诱导温度、诱导剂浓度、加诱导剂时间和诱导时间的优化研究,在菌液生长至OD600值为0.6时,加入诱导剂IPTG至终浓度0.01 mmol/L,20℃诱导20 h,蛋白的表达量达到每克干细胞89 mg的蛋白,粗酶液酶活达到19 U/mL。

Pseudomonas putida细胞对微囊藻毒素-LR的胁迫响应

将1.0g/L的微囊藻毒素-LR(MC-LR)降解菌恶臭假单胞菌(Pseudomonas putida)置于含不同浓度MC-LR的体系中,研究了体系中菌体细胞完整性和生物量的变化,考察了MC-LR对细胞的氧化胁迫以及抗氧化酶的响应.结果表明,MC-LR能够增大P.putida细胞质膜通透性,造成膜损伤,导致胞内物质外流,使细胞完整性遭到破坏;同时,MC-LR能够引起P.putida细胞的氧化胁迫,随着毒素暴露时间的延长,活性氧自由基(ROS)和膜脂过氧化产物丙二醛(MDA)含量显著升高,具有明显的剂量效应.超氧化物歧化酶(SOD)活性在MC-LR的诱导下有一个先升后降的过程,表现为对低浓度污染物的主动响应,而高浓度(2.5mg/L)MC-LR作用5d后,ROS积累到相当高水平,对细胞代谢功能造成破坏,使SOD活性下降,并加速细胞的死亡,P.putida生物量与对照相比,下降了将近50%.

酚酸介导下连作茶树根际病原菌Alternaria sp.及其拮抗菌Pseudomonas sp.变化

茶树是我国重要的经济作物,然而长期宿根连作铁观音茶园存在土壤微生物群落结构失衡、病害加重等连作障碍问题,探究铁观音茶树连作障碍形成的分子机制对寻求有效的防控技术具有重要意义。采用微生物分离纯化、平板对峙等方法对铁观音茶树根际病原菌及其拮抗菌进行分离、鉴定,并对不同宿根连作年限(0、1、10、20 a)铁观音茶树根际土壤中的病原菌和拮抗菌数量进行定量分析;同时运用高效液相色谱(HPLC)技术分析不同连作年限根际土壤中酚酸含量变化,并模拟土壤中各酚酸配比,研究酚酸类物质对茶树根际病原菌及其拮抗菌的影响。结果显示,从连作20 a患病铁观音茶树根系分离鉴定到1株病原真菌链格孢菌(Alternaria sp.),并从根际土壤中筛选到1株拮抗菌假单胞菌(Pseudomonas sp.)。荧光定量PCR分析发现,与1 a茶园相比,20 a茶园土壤中的链格孢菌绝对含量显著偏高,而假单胞菌含量却显著偏低。连作茶园根际土壤中共检测到对羟基苯甲酸、香草酸、丁香酸、香兰素、阿魏酸5种酚酸类物质,其平均配比为38∶229∶11∶11∶3。连作条件下酚酸类物质并未在土壤中累积,而是随种植年限的延长呈现出先增加后降低的趋势。模拟试验发现,中低浓度(30~120μmol·L^(-1))的混合酚酸能够显著促进链格孢菌菌丝的生长,单一酚酸对羟基苯甲酸、香草酸和丁香酸在低浓度(30μmol·L^(-1)和60μmol·L^(-1))时也能够显著促进链格孢菌菌丝的生长。对羟基苯甲酸对假单胞菌的生长有抑制作用,并且随着浓度的增加抑制作用增强,混合酚酸以及其他单一酚酸对假单胞菌的生长无明显作用。由此可见,铁观音根系分泌物酚酸类物质对根际土壤关键微生物菌群具有不同的生态效应,是引起微生物群落结构失衡、病害增加等连作障碍问题的重要因素。研究结果为进一步揭示铁观音茶树连作障碍机理提供一些理论依据。

Degradation of o-chloronitrobenzene as the sole carbon and nitrogen sources by Pseudomonas putida OCNB-1

A bacterial strain that utilized o-chloronitrobenzene （o-CNB） as the sole carbon, nitrogen and energy sources was isolated from an activated sludge collected from an industrial waste treatment plant. It was identified as Pseudomonas putida based on its morphology, physiological, and biochemical characteristics with an automatic biometrical system and the 16S rRNA sequence analysis. Microcosm study showed that the biodegradation of o-CNB was optimized at culture medium pH 8.0 and 32℃. At these conditions, the strain degraded 85% of o-CNB at a starting concentration of 1.1 mmol/L in 42 h. o-Chloroaniline was identified as the major metabolite with high performance liquid chromatography （HPLC） and gas chromatography-mass spectrometry （GC-MS）. The study showed that o-CNB degradation by Pseudomonas putida OCNB-1 was initiated by aniline dioxyenase, nitrobenzene reductase and catechol-l,2- dioxygenase.

Enhanced Biological Phosphorus Removal with Pseudomonas putida GM6 from Activated Sludge

The enhanced biological phosphorus removal （EBPR） method is widely adopted for phosphorus removal from wastewater, yet little is known about its microbiological and molecular mechanisms. Therefore, it is difficult to predict and control the deterioration of the EBPR process in a large-scale municipal sewage treatment plant. This study used a novel strain isolated in the laboratory, Pseudomonas putida GM6, which had a high phosphate accumulating ability and could recover rapidly from the deteriorated system and enhance the capability of phosphorus removal in activated sludge. Strain GM6 marked with gfp gene, which was called GMTR, was delivered into a bench-scale sequencing batch reactor （SBR） of low efficiency, to investigate the colonization of GMTR and removal of phosphorus. After 21 days, the proportion of GMTR in the total bacteria of the sludge reached 9.2%, whereas the phosphorus removal rate was 96%, with an effluent concentration of about 0.2 mg L^-1. In the reactor with the addition of GMTR, phosphorus was removed quickly, in 1 h under anaerobic conditions, and in 2 h under aerobic conditions. These evidences were characteristic of EBPR processes. Field testing was conducted at a hospital sewage treatment facility with low phosphorus removal capability. Twentyone days after Pseudomonas putida GM6 was added, effluent phosphorus concentration remained around 0.3 mg L^-1, corresponding to a removal rate of 96.8%. It was therefore demonstrated that Pseudomonas putida GM6 could be used for a quick startup and enhancement of wastewater biological phosphorus removal, which provided a scientific basis for potential large-scale engineering application.