Variations dans ľexpression du pouvoir oncogène d'Agrobacterium tumefaciens

Variations in the expression of the oncogenic power of Agrobacterium tumefaciens due to interactions between bacteria. The effects of the virulent strain A6 of Agrobacterium tumefaciens after co-culture with non-virulent variants of the same bacterium, or even with other bacterial species (e.g. Escherichia coli), showed that interactions between different strains of bacteria exist. These interactions are expressed as a phenomenon of either enhancement or inhibition, as shown by an increase or a decrease in the weights of tumors induced in decapitated stems of Pisum sativum L. cv. Annonay. These two phenomena depend on the contact time between the bacteria in mixed cultures (type I). With a short contact time between the two bacterial types (one or two generations), infections in decapitated pea stems produced by mixed inocula caused an increase in tumor weight compared with infections induced by inocula of virulent bacteria only. If the contact time was increased to the end of the log phase, a decrease in tumor weight was observed. Clarified supernatant fluids of spent media were also used as culture media (type II) for the virulent A6 bacteria. The stimulatory or inhibitory activity of (a) substance(s) present in these supernatant fluids depended on two variables: culture time of the bacteria A6 in the supernatant fluids and the “age” of the bacterial culture used to prepare them. Weight increase of the tumors was obtained if the proliferating time of the A6 bacteria in the supernatant fluids was short (4 h), or if the super natants were obtained from “young” cultures. Inhibition of the tumor expression occurred if the contact time of A6 bacteria in the supernatant fluids was increased or if the supernatants originated from bacterial cultures at the end of their growth. The in vitro interactions between the two bacterial strains in a mixed culture (type I or type II) were suppressed in the presence of pancreatic ribonuclease. Deoxyribonuclease had no effect. This provides indirect evidence for the action of a ribonucleic acid in the expression of the oncogenic power of a bacterial population of Agrobacterium tumefaciens.     

Antibiofilm action of a toluidine blue O-silver nanoparticle conjugate on Streptococcus mutans: a mechanism of type I photodynamic therapy

The objective of this study was to evaluate the anti-biofilm efficacy of photodynamic therapy by conjugating a photosensitizer (TBO) with silver nanoparticles (AgNP). Streptococcus mutans was exposed to laser light (630 nm) for 70 s (9.1 J cm^?2) in the presence of a toluidine blue O–silver nanoparticle conjugate (TBO–AgNP). The results showed a reduction in the viability of bacterial cells by 4 log₁₀. The crystal violet assay, confocal laser scanning microscopy and scanning electron microscopy revealed that the TBO–AgNP conjugates inhibited biofilm formation, increased the uptake of propidium iodide and leakage of the cellular constituents, respectively. Fluorescence spectroscopic studies confirmed the generation of OH^? as a major reactive oxygen species, indicating type I phototoxicity. Both the conjugates down-regulated the expression of biofilm related genes compared to TBO alone. Hence TBO–AgNP conjugates were found to be more phototoxic against S. mutans biofilm than TBO alone.     

Phagocytosis and nodule formation by hemocytes of Manduca sexta larvae following injection of Pseudomonas aeruginosa

The time course of clearance of an injected dose of 10⁶ CFU ml^?1 hemolymph of Pseudomonas aeruginosa ATCC 9027 in larvae of the tobacco hornworm, Manduca sexta, has been examined in detail. The clearance process has been subdivided into three stages during which the rates of reduction in concentration of circulating viable bacteria were clearly different. Contributions of hemocyte reactions to bacterial clearance were examined during stages I and II. During stage I (0–2 hr postinoculation (PI), nodule formation produced a dramatic reduction in circulating bacteria by entrapping over 90% of the injected dose in the first 30 min. Phagocytosis of bacteria by circulating hemocytes and subsequent intracellular digestion contributed significantly to reductions in circulating bacteria during stage II (2–8 hr PI). Viable cells of the virulent P. aeruginosa P11-1 were trapped in nodules as efficiently as the less virulent 9027 during the first 30 min after injection into M. sexta. Bacteria of strain P11-1 were also phagocytosed by hemocytes during stage II, however, phagocytosed bacteria were observed less frequently in P11-1-treated insects and intracellular digestion of these bacteria was only rarely observed. The increased virulence of P11-1 in larvae of M. sexta may be due to less efficient phagocytosis by circulating hemocytes and to insensitivity of this strain to killing reactions in nodules and following phagocytosis.     

Photodynamic effects of deuteroporphyrin on Gram-positive bacteria

The photodynamic effects of deuteroporphyrin (DT) on the growth and viability ofStaphylococcus aureus, Streptococcus faecalis, andBacillus cereus are described. By exposure to light and DT, the growth ofSta. aureus andStr. Faecalis strains was markedly inhibited, whereasB. cereus was undergoing lysis. Counting the viable bacteria during the DT treatment revealed that more than 99% of the initial bacterial population was killed within the first 30 min of treatment. The pH dependence of the photodynamic inhibitory effect shows that it is best obtained at pH 6.5. No temperature dependence in the range of 37°–44°C could be detected. The best photodynamic effect was achieved when the culture was in the mid-log phase. DT-treated bacteria, when grown in the dark or in the presence of horse serum albumin, were unaffected by the porphyrin action. Electron microscopic examinations ofSta. aureus andStr. faecalis showed the appearance of well-developed mesosomes within the cell cytoplasm.B. cereus preparations have not shown any mesosome formation but showed some lysed cells as well as some spores. None of the mentioned effects by DT and light could be observed on Gramnegative bacteria such asEscherchia coli orPseudomonas aeruginosa.     

Apoptosis of sea bass (Dicentrarchus labrax L.) neutrophils and macrophages induced by experimental infection with Photobacterium damselae subsp. piscicida

The infection of sea bass (Dicentrarchus labrax L.) by intraperitoneal (i.p.) injection of the agent of fish pasteurellosis Photobacterium damselae subsp. piscicida resulted in the apoptosis of peritoneal neutrophils and macrophages. All the eight virulent and none of the two non-virulent strains tested exhibited apoptogenic activity. A secreted bacterial protein(s) is a likely candidate as the factor(s) responsible for this activity, since no apoptosis was induced by i.p. injected UV-killed virulent strains and the virulent culture supernatants exhibited a thermo-labile apoptogenic activity identical to that of live bacteria. The apoptotic process was characterized by the occurrence of DNA fragmentation detected by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) staining and DNA electrophoresis, and of typical ultrastructural alterations namely cell shrinkage, chromatin condensation, nuclear fragmentation and production of blebs with shedding of apoptotic bodies. In the apoptotic process induced by lethal doses of virulent bacteria or culture supernatants both peritoneal macrophages and neutrophils were extensively affected, the majority of these cells being apoptotic and reaching values around 10(7)per peritoneal cavity for each cell type at 24h post-injection. Moreover, the number of non-apoptotic macrophages was always below the initial number in the resting peritoneal cavity. Since macrophages are key cells in the elimination of both bacteria and apoptotic moribund cells and apoptotic bodies, the induction by Ph. damselae subsp. piscicida of simultaneous macrophage and neutrophil apoptosis results, on the one hand, in the destruction of the two phagocytic cell types involved in the restriction of multiplication of the bacteria and, on the other hand, in the uncontrolled progression of the apoptotic process towards secondary necrosis and eventual lysis of high numbers of moribund neutrophils and of neutrophilic apoptotic bodies, with the consequent extensive release of their highly cytotoxic components. Abundant apoptotic cells were also seen in sections of head-kidney from fish dying from experimental pasteurellosis. In contrast, no apoptosis was seen in vitro after the treatment with virulent culture supernatants of sea bass head-kidney macrophage cultures or after the treatment ex vivo of peritoneal exudate leukocytes with virulent bacteria or culture supernatants. The apoptotic process described here appears as a novel and very powerful microbial pathogenic strategy.     

Human probiotic bacteria attenuate Pseudomonas aeruginosa biofilm and virulence by quorum-sensing inhibition

Abstract

This work investigated chloroform extracts from culture supernatants of two human probiotic bacteria, Lactobacillus casei CRL 431 and Lactobacillus acidophilus CRL 730 for the production of virulence factors and quorum sensing (QS) interference against three Pseudomonas aeruginosa strains. Both extracts inhibited biofilm biomass (up to 50%), biofilm metabolic activity (up to 39%), the production of the enzyme elastase (up to 63%) and pyocyanin (up to 77%), and decreased QS, without presenting any antibacterial acgivity. In addition, the chloroform extracts of both strains disrupted preformed biofilms of the three strains of P. aeruginosa analyzed (up to 40%). GC-MS analysis revealed that the major compounds detected in the bioactive extracts were four diketopiperazines. This study suggests that the metabolites of L. casei and L. acidophilus could be a promising alternative to combat the pathogenicity of P. aeruginosa.     

Surface sensing triggers a broad-spectrum antimicrobial response in Pseudomonas aeruginosa

Interspecies bacterial competition may occur via cell-associated or secreted determinants and is key to successful niche colonization. We previously evolved Pseudomonas aeruginosa in the presence of Staphylococcus aureus and identified mutations in the Wsp surface-sensing signalling system. Surprisingly, a ΔwspF mutant, characterized by increased c-di-GMP levels and biofilm formation capacity, showed potent killing activity towards S. aureus in its culture supernatant. Here, we used an unbiased metabolomic analysis of culture supernatants to identify rhamnolipids, alkyl quinoline N-oxides and two siderophores as members of four chemical clusters, which were more abundant in the ΔwspF mutant supernatants. Killing activities were quorum-sensing controlled but independent of c-di-GMP levels. Based on the metabolomic analysis, we formulated a synthetic cocktail of four compounds, showing broad-spectrum anti-bacterial killing, including both Gram-positive and Gram-negative bacteria. The combination of quorum-sensing-controlled killing and Wsp-system mediated biofilm formation endows P. aeruginosa with capacities essential for niche establishment and host colonization.     

Efficiency of using green algae as biological controllers against toxic algal taxa in cultured Nile tilapia Oreochromis niloticus,based on histopathological examinations

The incidence of harmful cyanobacterial blooms in surface waters has increased in frequency and outbreaks have become more severe. This research aimed at studying the effect of a culture of two green algal species as biological control of the growth of toxic blue-green algae. Nile tilapia of an initial mean weight of 55 g fish^?1 (SE 5) were used for each of four treatments in triplicate. All algal seedings were done at 4 × 10³ cells ml^?1. Treatment I (untreated) served as a control, Treatment II was seeded with Microcystis aeruginosa, Treatment III was seeded with green algae Chlorella ellipsoidea and Scenedesmus bijuga, and Treatment IV was seeded with a mixture of M. aeruginosa and C. ellipsoidea and S. bijuga. After 10 days, Treatment IV showed 3.4% viable cell survival, compared to 35% and 55% in Treatments II and III, respectively. Histopathological examination revealed mild degenerative changes and focal necrosis, as well as a depletion of haematopoietic tissues in Treatment IV compared to Treatment II. These findings suggest the efficacy of C. ellipsoidea and S. bijuga in controlling the growth of M. aeruginosa and minimising its side effects on cultured Nile tilapia.     

Modulation of the mitogenic PHA response of carp, Cyprinus carpio L., by extracellular products of Aeromonas salmonicida

The influence of bacteria-free supernatants from cultures of atypical virulent (V234/81, auto-agglutinating. A-layer positive) and avirulent (126/68, non-agglutinating, A-layer negative) strains of A. salmonicida , obtained after different culture times in yeast-tryptone broth at 20°C, was tested on the PHA response of carp pronephric leucocytes in vitro . Supernatants from virulent cultures modulated the response, whereas avirulent supernatants had no effect. The response was enhanced (400%) by supernatant from early virulent cultures (20 h), but severely depressed (<3%) by later ones (96 h). The effects were dose-dependent. Inhibitory activity of 96-h supernatant was lost by heating (70°C, 30 min), suggesting that the inhibiting factors are all proteinaceous.
Heated 96-h supernatant was as stimulatory as early supernatant. Stimulation of leucocytes also occurred in the absence of PHA with early and heat-treated 96-h supernatants, but at a tenth of the level, suggesting that only stimulated cells (blasts) might respond to the substance(s) present in supernatants. Membrane fragments from virulent and avirulent bacteria, and purified LPS from virulent bacteria, were stimulatory with or without PHA. Endotoxin-free, heat-treated, 96-h culture supernatants were also stimulatory, suggesting that an additional mitogenic factor(s), other than LPS, is present in the supernatants. The modulating in vitro effects of extracellular products from A. salmonicida might explain the immunosuppression seen during later stages of erythrodermatitis in vivo.     

应用siRNA技术探讨MCF-7乳腺癌细胞分泌的VEGF对树突状细胞的影响

为探讨MCF-7乳腺癌细胞分泌的血管内皮生长因子（ vascular endothelial growth factor, VEGF）对树突状细胞（dendritic cell, DC）功能及其分化的影响,针对VEGF基因设计siRNA（small interfering RNA, siRNA),采用脂质体转染法以100 nmol/L最佳转染浓度导入MCF-7乳腺癌细胞（siRNA组）,以脂质体Lipofectamine　2000TM转染MCF-7 乳腺癌细胞培养上清培养正常DC作为对照（对照组）,采用ELISA法检测经siRNA 干扰VEGF基因后的MCF-7 乳腺癌细胞分泌的VEGF因子含量, Western 印迹检测VEGF蛋白表达,以探讨siRNA的基因沉默效果;以siRNA组和对照组培养上清分别培养外周血单个核细胞,用流式细胞仪检测所诱导DC表型CD1a、CD80、CD83、CD86和HLA-DR的表达,用MTT法检测转染前后两组DC 诱导的细胞毒性T淋巴细胞（cytotoxic T lymphocyte, CTL）对MCF-7细胞的细胞毒作用.结果显示,MCF-7 乳腺癌细胞培养上清能明显抑制正常DC分化成熟及抗原递呈能力,干扰VEGF基因后MCF-7 乳腺癌细胞培养上清对DC的影响明显降低,CD80、CD83、CD86和HLA-DR的表达较对照组显著升高,而CD1a表达下降（P＜0.01）.转染前后DC 诱导的CTL对MCF-7细胞的杀伤活性有明显差异（P＜0.01）.由此可见,siRNA可靶向抑制MCF-7乳腺癌细胞VEGF的表达,下调VEGF后的MCF-7 细胞上清对DC分化成熟及功能的抑制作用明显降低,从而推测VEGF在肿瘤的发生、发展和免疫抑制方面可能起着重要的作用.     

Bacterial weathering of rapakivi granite

Rapakivi granite samples were incubated with Pseudomonas aeruginosa culture solutions in order to elucidate the possible role of bacteria in rapakivi (crumbling stone) disintegration. SEM micrographs showed micromorphological alterations on the incubated rapakivi surface at 21 to 23°C for 20 days. Elevated concentrations of Na, Ca, K, Fe, and Mg were detected in the culture solutions after incubation. Elemental oxide ratios [K₂O : (Na₂O + CaO)] in culture solutions were similar to those in rapakivi ovoids, suggesting a proportional dissolution pattern of these elements.     

Pseudomonas aeruginosa-derived rhamnolipids subvert the host innate immune response through manipulation of the human beta-defensin-2 expression

Pseudomonas aeruginosa is a well‐known cause of infections especially in compromised patients. To neutralize this pathogen, the expression of antimicrobial factors in epithelial cells is crucial. In particular the human beta‐defensin hBD‐2 is especially active against P. aeruginosa. In this study, we identified rhamnolipids in P. aeruginosa culture supernatants that are able to prevent the pathogen‐induced hBD‐2 response in keratinocytes. The presence of rhamnolipids within the host cells and inhibition assays suggest that calcium‐regulated pathways and protein kinase C activation are impaired by rhamnolipids. In consequence, the induction of hBD‐2 in keratinocytes by P. aeruginosa‐derived flagellin as well as the host's own hBD‐2 mediator interleukin IL‐1β is inhibited. Strikingly, rhamnolipids did not affect the release of the proinflammatory mediator interleukin IL‐8 by flagellin. Thus, in addition to their function in establishment and persistence of P. aeruginosa infections, rhamnolipids can be engaged by P. aeruginosa for a targeted attenuation of the innate immunity to manage its survival and colonization on compromised epithelia.     

Bacteriophage phi297, a new species of <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> temperate phages with a mosaic genome: Potential use in phage therapy

The genome of halo-forming temperate Pseudomonas aeruginosa phage phi297 and lytic activity of its virulent mutant were studied. A mosaic structure was revealed for phi297 genome by its complete sequencing. The phi297 genome was partly homologous to the genomes of phages D3 and F116. High lytic activity was assumed for temperate P. aeruginosa bacteriophage phi297 on the basis of morphological features of negative colonies. Virulent mutant phi297vir, which was capable of lysing the wild-type phage bacteria, was isolated. Lytic activity was compared for phi297 and the phages from commercial mixtures of two manufacturers (facilities of Nizhnii Novgorod and Perm’). Phage phi297 caused lysis of the mutant PAO1 bacteria that were resistant to the phages from commercial preparations, but the lytic activity spectrum of phi297 was narrower that the spectra of the commercial phages. The use of nonreverting virulent mutants of certain temperate bacteriophages was proposed for the treatment of P. aeruginosa infections.     

Conditioned medium promotes the attachment ofAgrobacterium tumefaciens strain NT 1 to carrot cells

Summary Wild-typeAgrobacterium tumefaciens bind to carrot suspension culture cells. Avirulent strain NT 1 did not bind to carrot cells when they were incubated together in Murashige and Skoog medium. Conditioned medium was prepared by incubatingA. tumefaciens virulent strain C 58 with carrot cells and removing the bacteria and carrot cells using filter sterilization. This conditioned medium promoted the binding of NT 1 to carrot cells. Conditioned medium did not promote the nonspecific attachment ofEscherichia coli to carrot cells. These results suggest that when wild-typeA. tumefaciens are incubated with plant host cells, some substance(s) involved in bacterial attachment are released into the medium. Filter-sterilized medium from the incubation of the nonattachingchvB mutant A 1045 with carrot cells promoted the attachment of strain NT 1 even though A 1045 bacteria did not bind to the carrot cells. However, filter-sterilized medium from the incubation of the non-attachingatt mutant Att-B 123 with carrot cells was unable to promote the binding of strain NT 1. This suggests that nonattaching mutants ofA. tumefaciens can be divided into two groups on the basis of the properties of the substances released into the medium when the bacteria are incubated with carrot cells.Abbreviations MS Murashige and Skoog tissue culture medium Dedicated to the memory of Professor John G. Torrey     

Human dendritic cells process and present Listeria antigens for in vitro priming of autologous CD4+ T lymphocytes

The role of human dendritic cells (DC) in the immune response toward intracellularly growing Listeria was analyzed under in vitro conditions using several morphological and functional methods. DC incubated with Listeria innocua and L. monocytogenes, respectively, readily phagocytosed the bacteria. Listeria did not impair viability and immunogenic potential of human DC. Listerial antigens were found to be processed within the lysosomal compartment of DC and colocalized with major histocompatibility complex (MHC) class II molecules, as shown by fluorescence and transmission electron microscopy. DC challenged with apathogenic L. innocua were highly effective in priming autologous naïve T cells (mainly CD4+) in vitro. The T cells strongly proliferated in the presence of DC incubated with L. innocua, which could be significantly inhibited by anti-MHC II mAb. L. innocua-primed T cells were also successfully stimulated by DC harboring the pathogenic L. monocytogenes, either the wild-type strain EGD or the p60 reduced mutant strain RIII. From our results, we conclude that human DC infected with nonpathogenic intracellular bacteria are able to efficiently prime naïve T cells, which are then suitable for recognition of antigens derived from related virulent bacterial species. This in vitro human model provides an interesting tool for basic research in infectious immunology and possibly for a new immunotherapy.     

Insecticidal bacterium isolated from an ant lion larva from Munakata,Japan

Twenty bacteria were isolated from four ant lion larvae. The isolates were classified into three groups by biological characteristics. Since Group I, Group II and Group III were isolated from individual larvae Kuo1, Kuo3, 4 and Kuo2, respectively, with exception of one isolate Kuo2‐1, each ant lion tested had its own dominant bacterial flora. Groups I and II were closer to Serratia liquefaciens and Enterobacter cloacae, respectively, whereas Group III could not be identified by the test used. The phylogenetic analysis of GroEL amino acid sequences revealed that Group I, II and III were related to those of Serratia spp., E. cloacae and Salmonella spp. –Escherichia/Shigella spp., respectively. Among these groups, Group I was highly virulent against Bombyx mori and Periplaneta americana, and caused 100% mortality within 24 h. The other two groups (Group II and III) were avirulent to these insect species. The culture filtrate of Group I caused killing activity to B. mori larvae and the insecticidal substance was purified from culture filtrate of Group I bacterium. Since the insecticidal activity highly correlated with proteolytic activity in the chromatographies, Group I bacterium may secret insecticidal proteinase in vitro.     

巨噬细胞集落刺激因子经PI3K/AKT信号途径影响人乳腺癌MCF-7细胞糖代谢

本文探讨巨噬细胞集落刺激因子（M-CSF）对人乳腺癌MCF-7细胞糖代谢的影响及其机制. 构建胞质稳定转染 M-CSF的MCF-7细胞（MCF-7-M）;ATP检测试剂盒检测MCF-7和MCF-7-M细胞的ATP生成;葡萄糖测定试剂盒、乳酸测试盒检测MCF-7和MCF-7-M细胞的葡萄糖摄取和乳酸分泌情况;蛋白质印迹法检测在糖酵解抑制剂2-脱氧葡萄糖（2-DG）和氧化磷酸化抑制剂OLIG处理后,M-CSF对MCF-7细胞的糖酵解关键酶：己糖激酶2（HK2）、丙酮酸激酶M2（PKM2）及葡萄糖转运体1（GLUT-1）表达的影响;MTT法检测在ATP消耗剂3-溴丙酮酸（3-BrPA）处理后,MCF-7和MCF-7-M细胞对5-FU敏感性的变化. 结果发现：MCF-7-M细胞的ATP水平显著高于MCF-7细胞（P<0.05）;2-DG降低了MCF-7和MCF-7-M细胞的ATP水平,并且降低MCF-7-M细胞ATP的效果更明显（P<0.01）;MCF-7-M细胞的糖摄取能力和乳酸分泌量显著高于MCF-7细胞（P<0.01）,经API-2处理后,MCF-7和MCF-7-M细胞葡萄糖消耗和乳酸分泌量均显著减少（P<0.01）;MCF-7-M细胞GLUT-1、HK2和PKM2的表达显著高于MCF-7细胞（P<0.01）;LY294002和API-2均可抑制MCF-7-M细胞GLUT-1的表达（P<0.05）;用3-BrPA处理后,MCF-7-M和MCF-7细胞对5-FU的药物敏感性显著增强（P<0.01). 综上,得出结论： 胞质M-CSF通过诱导GLUT-1、HK2和PKM2的表达,活化MCF-7细胞糖酵解途径;PI3K/AKT信号通路参与胞质M-CSF活化MCF-7细胞的糖酵解途径.     

Assessment of Photodynamic Destruction of Escherichia coli O157:H7 and Listeria monocytogenes by Using ATP Bioluminescence

Antimicrobial photodynamic therapy was shown to be effective against a wide range of bacterial cells, as well as for fungi, yeasts, and viruses. It was shown previously that photodestruction of yeast cells treated with photosensitizers resulted in cell destruction and leakage of ATP. Three photosensitizers were used in this study: tetra(N-methyl-4-pyridyl)porphine tetratosylate salt (TMPyP), toluidine blue O (TBO), and methylene blue trihydrate (MB). A microdilution method was used to determine MICs of the photosensitizers against both Escherichia coli O157:H7 and Listeria monocytogenes. To evaluate the effects of photodestruction on E. coli and L. monocytogenes cells, a bioluminescence method for detection of ATP leakage and a colony-forming assay were used. All tested photosensitizers were effective for photodynamic destruction of both bacteria. The effectiveness of photosensitizers (in microgram-per-milliliter equivalents) decreased in the order TBO > MB > TMPyP for both organisms. The MICs were two- to fourfold higher for E. coli O157:H7 than for L. monocytogenes. The primary effects of all of the photosensitizers tested on live bacterial cells were a decrease in intracellular ATP and an increase in extracellular ATP, accompanied by elimination of viable cells from the sample. The time courses of photodestruction and intracellular ATP leakage were different for E. coli and L. monocytogenes. These results show that bioluminescent ATP-metry can be used for investigation of the first stages of bacterial photodestruction.

     

TatC-dependent translocation of pyoverdine is responsible for the microbial growth suppression

Infections are often not caused by a colonization of Pseudomonas aeruginosa alone but by a consortium of other bacteria. Little is known about the impact of P. aeruginosa on the growth of other bacteria upon coinfection. Here, cellree culture supernatants obtained from P. aeruginosa suppressed the growth of a number of bacterial strains such as Corynebacterium glutamicum, Bacillus subtilis, Staphylococcus aureus, and Agrobacterium tumefaciens, but had little effect on the growth of Escherichia coli and Salmonella Typhimurium. The growth suppression effect was obvious when P. aeruginosa was cultivated in M9 minimal media, and the suppression was not due to pyocyanin, a well-known antimicrobial toxin secreted by P. aeruginosa. By performing transposon mutagenesis, PA5070 encoding TatC was identified, and the culture supernatant of its mutant did not suppress the growth. HPLC analysis of supernatants showed that pyoverdine was a secondary metabolite present in culture supernatants of the wild-type strain, but not in those of the PA5070 mutant. Supplementation of FeCl₂ as a source of iron compromised the growth suppression effect of supernatants and also recovered biofilm formation of S. aureus, indicating that pyoverdine-mediated iron acquisition is responsible for the growth suppression. Thus, this study provides the action of TatC-dependent pyoverdine translocation for the growth suppression of other bacteria, and it might aid understanding of the impact of P. aeruginosa in the complex community of bacterial species upon coinfection.     

Effect of curcumin-nanoemulsion associated with photodynamic therapy in breast adenocarcinoma cell line

Curcumin, a natural compound has several antineoplastic activities and is a promising natural photosensitizer used in photodynamic therapy. However, its low solubility in physiological medium limit the clinical use of curcumin. This study aimed to analyze the action of curcumin-nanoemulsion, a new and well-designed Drug Delivery System (DDS+) molecule, used as a photosensitizing agent in photodynamic therapy in an in vitro breast cancer model, MCF-7 cells. The empty nanoemulsion fulfils all necessary requirements to be an excellent DDS. Furthermore, the use of curcumin-nanoemulsion in photodynamic therapy resulted in a high phototoxic effect after activation at 440?nm, decreasing to <10% viable tumor cells after two irradiations and increasing the reactive oxygen species (ROS) production. The use of curcumin-nanoemulsion associated with photodynamic therapy resulted in an increase in the levels of caspase 3/7 activity for the studied MCF-7 cell model, indicating that this therapy triggers a cascade of events that lead to cell death, such as cellular apoptosis. In conclusion, curcumin-nanoemulsion proved to be efficient as a photosensitizing agent, had phototoxic effects, significantly decreased the proliferation of MCF-7 cells and stimulating the ROS production in combination with photodynamic therapy, so, this formulation has a great potential for use in treatment of breast cancer.