Isolation and Characterization of a Lytic Myoviridae Bacteriophage PAS-1 with Broad Infectivity in Aeromonas salmonicida

To search for candidate control agents against Aeromonas salmonicida subsp. salmonicida infections in aquaculture, one bacteriophage (phage), designated as PAS-1, was isolated from the sediment samples of the rainbow trout (Oncorhynchus mykiss) culture farm in Korea. The PAS-1 was morphologically classified as Myoviridae and possessed approximately 48 kb of double-strand genomic DNA. The phage showed broad host ranges to other subspecies of A. salmonicida as well as A. salmonicida subsp. salmonicida including antibiotic-resistant strains. Its latent period and burst size were estimated to be approximately 40 min and 116.7 PFU/cell, respectively. Furthermore, genomic and structural proteomic analysis of PAS-1 revealed that the phage was closely related to other Myoviridae phages infecting enterobacteria or Aeromonas species. The bacteriolytic activity of phage PAS-1 was evaluated using three subspecies of A. salmonicida strain at different doses of multiplicity of infection, and the results proved to be efficient for the reduction of bacterial growth. Based on these results, PAS-1 could be considered as a novel Aeromonas phage and might have potentiality to reduce the impacts of A. salmonicida infections in aquaculture.     

Seaweeds from Vietnam used for functional food, medicine and biofertilizer

Several Vietnamese seaweed species have economic importance as food for humans, as industrial materials, as ingredients in traditional medicine, and as biofertilizers. The nutritional values of nine representative Vietnamese seaweed species were analyzed. In this report, all of the species studied are rich in proteins, lipids (especially polyunsaturated fatty acids), vitamins, pigments, and macro- and micro-elements. The effect of the physiological activities of the green alga, Ulva reticulata, on hepatic fatty acid metabolism were examined in mice. The results indicate that Vietnamese seaweeds are abundant and have high quality materials for industrial and agricultural purposes. Presented at the 6th Meeting of the Asian Pacific Society of Applied Phycology, Manila, Philippines.     

Synergistic effects of sequential treatment with methyl jasmonate, salicylic acid and yeast extract on benzophenanthridine alkaloid accumulation and protein expression in Eschscholtzia californica suspension cultures

To develop an optimal bioprocess for secondary metabolite production and explain the bioprocess at the molecular level, we examine the synergistic effects of sequential treatment with methyl jasmonate (MJ), salicylic acid (SA) and yeast extract (YE) on benzophenanthridine alkaloid accumulation and protein expression in Eschscholtzia californica suspension cultures. Serial treatment of MJ, SA and YE at 24 h intervals enhanced the accumulation of dihydrosanguinarine (2.5 times) and sanguinarine (5.5 times). This sequential treatment using different signal elicitors was more effective than single elicitor or simultaneous treatment of the elicitors; it induced benzophenanthridine alkaloid accumulation to 917.7 ± 42.0 mg/L. Also, (S)-methylcoclaurine-3′-hydroxylase (CYP80B1) and 3′-hydroxy-(S)-N-methylcoclaurine-4′-O-methyltransferase (4′OMT) expressions among enzymes in sanguinarine biosynthetic pathway explained the synergistic effects by sequential treatment of the elicitors. The sequential treatment strategy using elicitors related to different signal transduction pathways can be used to design better processes to increase accumulation of secondary metabolites in plant cell culture. Analysis of protein expression provides the detailed information about metabolite accumulation through the correlated results.     

Flavobacterium kyungheensis sp. nov., isolated from soil of a ginseng field

A Gram-staining negative, strictly aerobic, motile by gliding, non-spore-forming, pale yellow pigmented and rod-shaped bacterium designated strain THG-107^T was isolated from soil of a ginseng field on Ganghwa Island in the Republic of Korea and its taxonomic position was investigated by using a polyphasic study. Growth of strain THG-107^T was found to occur at 4–37 °C (optimum, 20–30 °C), at pH 5.5–10 (optimum, pH 7.0) and in the presence of 0–1 % (w/v) NaCl (optimum, absence) on R2A agar. On the basis of 16S rRNA gene sequence similarity, strain THG-107^T was shown to belong to the family Flavobacteriaceae and was related to Flavobacterium denitrificans ED5^T (99.1 % similarity). The G+C content of the genomic DNA was determined to be 34.2 mol%. These results are consistent with characteristics of members of the genus Flavobacterium. The only isoprenoid quinone detected in strain THG-107^T was menaquinone-6 (MK-6) and the major polyamine was identified as homospermidine (82.9 %). The major polar lipid detected was phosphatidylethanolamine and the major cellular fatty acids were identified as iso-C_15:0 (26.3 %), iso-C_17:0 3OH (12.6 %) and summed feature 3 (comprising C_16:1 ω7c and/or C_16:1 ω6c; 11.6 %). Flexirubin-type pigments were found to be present. Strain THG-107^T has β-glucosidase activity to convert ginsenosides Rb₁ and Rd into Gyp17 and F₂. DNA-DNA hybridization with F. denitrificans ED5^T was 52 %. Strain THG-107^T could be distinguished from F. denitrificans ED5^T and the other species of the genus Flavobacterium by its phylogenetic and genetic distinctiveness and by several phenotypic properties. Therefore, strain THG-107^T is considered to represent a novel species in the genus Flavobacterium, for which the name Flavobacterium kyungheensis sp. nov. is proposed (type strain THG-107^T = KACC 16219^T = LMG 26575^T).     

Comparison of PCR fingerprinting techniques for the discrimination of <Emphasis Type="Italic">Salmonella enterica</Emphasis> subsp. <Emphasis Type="Italic">enterica</Emphasis> serovar Weltevreden isolated from indigenous vegetables in Malaysia

Salmonella enterica subsp. enterica (S.) serovar Weltevreden has emerged as a public health problem in many countries. Genomic DNA of S. Weltevreden from indigenous vegetables namely ‘selom’ (Oenanthe stolonifera), ‘pegaga’ (Centella asiatica), ‘kesum’ (Polygonum minus) and ‘kangkong’ (Ipomoea aquatica) were characterized by duplex-polymerase chain reaction (duplex-PCR), multiplex-polymerase chain reaction (multiplex-PCR), random amplified polymorphic DNA (RAPD), enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The results demonstrated that a total of four clusters and three single isolates were generated from ERIC-PCR with primers ERIC-1 and ERIC-2 whereas RAPD with arbitrary primers OPAR2, OPAR17 and OPAR19 discriminated the S. Weltevreden into nine clusters and eight single isolates at a common 65% similarity level with discriminatory index (D) of 0.7443 and 0.9394 respectively. Composite analysis of banding profiles generated from RAPD-PCR and ERIC-PCR showed eight clusters and six single isolates at 65% similarity level with the highest D value that is 0.9508. On the other hand, PCR-RFLP and duplex PCR data exhibited a consistent profile for S. Weltevreden. Multiplex-PCR targeting three different antibiotic resistance genes and a common Salmonella specific gene segment produced two distinguishing profiles among the S. Weltevreden examined. These results demonstrated that the combined analysis of RAPD-PCR and ERIC-PCR is a better tool for characterizing S. Weltevreden than individual methods.     

Comparison of Nitrogen Fixation for North- and South-facing <Emphasis Type="Italic">Robinia pseudoacacia</Emphasis> Stands in Central Korea

The nitrogenase activity, root nodule biomass, and rates of nitrogen (N) fixation were measured in 25-year-old pure north- and south-facing Robinia pseudoacacia stands in an urban forest of Seoul (Kkachisan Mountain) in central Korea. The nitrogenase activity was estimated using an acetylene reduction (AR) assay, which showed an increasing trend during the early growing season, with sustained high rates from June through to September with a decrease thereafter. July had the highest nitrogenase activity rate (micromoles C₂H₄ per gram dry nodule per hour), averaging 95.8 and 115.1 for the north- and south-facing stands, respectively. The maximum root nodule biomass (kilograms per hectare) was 45.7 and 9.1 for the north- and south-facing stands in July, respectively. The AR rate appeared to be strongly correlated to the soil temperature (r ² = 0.68, P < 0.001) and soil pH (r ² = 0.59, P < 0.001) while root nodule biomass was correlated to the soil temperature (r ² = 0.36, P < 0.01) and water content (r ² = 0.35, P < 0.05). The soil temperature showed clear differences between seasons, while there was a significant difference in soil pH, organic matter, total N concentrations, and available phosphorus between the north- and south-facing stands. The N₂ fixation rates during the growing season varied from 0.1 to 37.5 kg N ha⁻¹ month⁻¹ depending on the sampling location and time. The annual N₂ fixation rate (kg N per hectare per year) was 112.3 and 23.2 for the north- and south-facing stands, respectively. The differences in N₂ fixation rate between the two stands were due mainly to the differences in total nodule biomass.     

Inhibitory effects of glycoprotein-120 (G-120) from Ulmus davidiana Nakai on cell growth and activation of matrix metalloproteinases

Excessive breakdown of extracellular matrix by metalloproteinases (MMPs) occurs in many pathological conditions. Consequently, methods for inhibiting MMP activity have therapeutic potential. In this study, we investigated the effect of G-120, a 120 kDa glycoprotein purified from the Oriental herbal plant, Ulmus davidiana Nakai (UDN), on the activity and production of several MMPs by evaluating its growth inhibitory effect on NIH 3T3 cells. Tritium uptake assays showed that proliferation of NIH 3T3 cells was strongly suppressed, and G-120-mediated inhibition of DNA synthesis proved to involve a cytostatic, rather than a cytotoxic, effect, as shown by cytotoxicity and apoptosis assays. More importantly, G-120 strongly reduced the gelatinolytic and collagenase activities of MMP proteins, as well as expression of MMP-2 and MMP-9. Electrophoretic mobility shift assays revealed that it suppressed the DNA binding activity of NF-kappaB. Collectively, our observations show that G-120 strongly inhibits the activation of MMPs and NF-kappaB.     

Simulated Microgravity Induces the Proliferative Inhibition and Morphological Changes in Porcine Granulosa Cells

Astronauts are always faced with serious health problems during prolonged spaceflights. Previous studies have shown that weightlessness significantly affects the physiological function of female astronauts, including a change in reproductive hormones and ovarian cells, such as granulosa and theca cells. However, the effects of microgravity on these cells have not been well characterized, especially in granulosa cells. This study aimed to investigate the effects of simulated microgravity (SMG) on the proliferation and morphology of porcine granulosa cells (pGCs). pGC proliferation from the SMG group was inhibited, demonstrated by the reduced O.D. value and cell density in the WST-1 assay and cell number counting. SMG-induced pGCs exhibited an increased ratio of cells in the G0/G1 phase and a decreased ratio of cells in the S and G2/M phase. Western blot analysis indicated a down-regulation of cyclin D1, cyclin-dependent kinase 4 (cdk4), and cyclin-dependent kinase 6 (cdk6), leading to the prevention of the G1-S transition and inducing the arrest phase. pGCs under the SMG condition showed an increase in nuclear area. This caused a reduction in nuclear shape value in pGCs under the SMG condition. SMG-induced pGCs exhibited different morphologies, including fibroblast-like shape, rhomboid shape, and pebble-like shape. These results revealed that SMG inhibited proliferation and induced morphological changes in pGCs.     

Combined cold, acid, ethanol shocks in Oenococcus oeni: effects on membrane fluidity and cell viability

The effects of combined cold, acid and ethanol on the membrane physical state and on the survival of Oenococcus oeni were investigated. Membrane fluidity was monitored on intact whole O. oeni cells subjected to single and combined cold, acid and ethanol shocks by using fluorescence anisotropy with 1,6-diphenyl-1,3,5-hexatriene (DPH) as a probe. Results showed that cold shocks (14 and 8 degrees C) strongly rigidified plasma membrane but did not affect cell survival. In contrast, ethanol shocks (10-14% v/v) induced instantaneous membrane fluidisation followed by rigidification and resulted in low viability. Acid shocks (pH 4.0 and pH 3.0) exerted a rigidifying effect on membrane without affecting cell viability. Whatever the shock orders, combined cold (14 degrees C) and ethanol (14% v/v) shocks resulted in strong membrane rigidification. Interestingly, O. oeni survived combined cold and ethanol shocks more efficiently than single ethanol shock. Membrane rigidification was induced by ethanol-and-acid (10% v/v - pH 3.5) shock and correlated with total cell death. In contrast, O. oeni recovered its viability when subjected to cold (8 degrees C)-then-ethanol-and-acid shock which strongly rigidified the membrane. Our results suggested a positive short-term effect of combined cold, acid and ethanol shocks on membrane fluidity and viability of O. oeni.