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rpoX, a Vibrio alginolyticus specific stress regulating gene, was used to detect this fish pathogen by SYBR Green I-based real-time PCR. The specificity of the detection was confirmed in different samples. The minimum level of detection was 10(3) cells from pure culture and 10(2) cells from seawater.
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Proteínas Bacterianas/genética , Enfermedades de los Peces/microbiología , Compuestos Orgánicos/química , Reacción en Cadena de la Polimerasa/métodos , Vibrio alginolyticus/genética , Animales , Benzotiazoles , Cartilla de ADN/genética , ADN Bacteriano/química , ADN Bacteriano/genética , Diaminas , Enfermedades de los Peces/diagnóstico , Perciformes , Quinolinas , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Factor sigma/genéticaRESUMEN
A previous study has shown that Vibrio alginolyticus ZJ-51 undergoes colony phase variation between opaque/rugose (Op) and translucent/smooth (Tr). The AI-2 quorum-sensing master regulator ValR, a homolog to V. harveyi LuxR, was suggested to be involved in the transition. To investigate the role of ValR in the variation and in biofilm formation, an in-frame deletion of valR in both Op and Tr backgrounds was carried out. The mutants in both backgrounds showed an intermediate colony morphotype, where the colonies were less opaque/rugose but not fully translucent/smooth either. They also showed an intermediate level of motility. However, biofilm formation was severely decreased in both mutants and polar flagella were depleted also. Quantitative PCR showed that most of the genes related to flagellar and polysaccharide biosynthesis were upregulated in the mutant of Op background (Delta valR/Op) but downregulated in the mutant of Tr background (Delta valR/Tr) compared with their parental wild-type strains. This suggests that ValR may control biofilm formation by regulating flagellar biosynthesis and affect the expression of the genes involved in colony phase variation in V. alginolyticus.
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Eliminación de Gen , Regulación Bacteriana de la Expresión Génica , Percepción de Quorum , Proteínas Represoras/genética , Transactivadores/genética , Vibrio alginolyticus/crecimiento & desarrollo , Biopelículas/crecimiento & desarrollo , Variación Genética , Datos de Secuencia Molecular , Fenotipo , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Vibrio alginolyticus/genética , Vibrio alginolyticus/metabolismoRESUMEN
Objective To prepare a thymopentin-contained supramolecular hydrogel, and characterize its micromorphology and mechanical property, and further investigate its effects on the cellular uptake and the immunomodulatory performance in vitro . Methods The self-assembling peptide containing thymopentin was prepared by solid phase synthesis method and identified using LC-MS after being purified by high performance liquid chromatography (HPLC). Supramolecular hydrogel was prepared through a heating-cooling process, and its micromorphology and mechanical property were characterized using transmission electron microscopy (TEM) and rheology. The cellular uptake efficiencies of free thymopentin and thymopentin nanofibers were observed by inverted fluorescence microscope after FITC-labeling. The abilities of free thymopentin and thymopentin nanofibers to stimulate RAW 264.7 cells to secrete tumor necrosis factor (TNF-α) were studied by enzyme-linked immunosorbent assay (ELISA). Results A macroscopic visible supramolecular hydrogel was obtained by a heating-cooling process, composing with long cross-linked nanofibers and possessing good ductility of mechanics. Compared with free thymopentin, thymopentin nanofibers showed much more enhanced cellular uptake, and better immunomodulation property as stimulating the RAW 264.7 cells to produce much higher concentration of TNF-α in vitro . Conclusion After rational structural modification, the cellular uptake and immunomodulatory activity of thymopentin, which formed nanomedicine, were significantly enhanced. This study can provide new methods and guidance for improving the therapeutic effect of thymopentin in clinical application.
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Objective To design and synthesize a novel type of combined anti-tumor drug-doxorubicin modified silver nanoparticles(DOX-Ag NPs)with pH response,characterize its physical and chemical properties,and investigate its drug responsive release and anti-tumor activity in vitro.Methods DOX-Ag NPs were prepared by coupling silver nanoparticles (Ag NPs) with doxorubicin (DOX) via a LA-NHNH2linker. The structure of LA-NHN=DOX was confirmed by nuclear magnetic resonance(1H NMR)and high resolution mass spectrometry(HRMS).The particle size and micromorphology of the nanoparticles were detected by dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. The optical properties of the nanoparticles were characterized by UV-vis absorption spectroscopy and fluorescence spectroscopy.The DOX release kinetics of DOX-Ag NPs under different pH conditions were examined by dialysis method combined with fluorescence spectroscopy. The in vitro anti-tumor effects of DOX-Ag NPs were evaluated by MTT assay. Results DOX-Ag NPs were spherical nanoparticles with a particle size of (40.4 ± 3.8) nm. DOX-Ag NPs could rapidly release DOX under weak acid condition.DOX-Ag NPs significantly inhibited the proliferation and cell viability of HepG2 cells in concentration dependent manner.When DOX concentration was 0.5-20 mg/L(Ag concentration was 0.45-18 mg/L), the cell survival rate was significantly lower in DOX-Ag NPs group than that of DOX group and Ag NPs group(P<0.05). Conclusion DOX-Ag NPs are a combined anti-tumor nano-drug with pH-responsive ability, which can release DOX rapidly in tumor tissues and play an anti-tumor effect through synergistic treatment with Ag NPs in vitro.
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Objective To investigate the value of serum?-human chorionic gonadotropin(?-HCG) and progesterone monitoring in medicinal conservative treatment for ectopic pregnancy. Methods Sixty patients with ectopic pregnancy treated with methotrexate were divideded into group A(n=39,?-HCG0.05).The time of serum progesterone level declining to normal level was significantly shorter than that of ?-HCG in both group A and group B(P