Ecosystem engineering in space and time

The ecosystem engineering concept focuses on how organisms physically change the abiotic environment and how this feeds back to the biota. While the concept was formally introduced a little more than 10 years ago, the underpinning of the concept can be traced back to more than a century to the early work of Darwin. The formal application of the idea is yielding new insights into the role of species in ecosystems and many other areas of basic and applied ecology. Here we focus on how temporal, spatial and organizational scales usefully inform the roles played by ecosystem engineers and their incorporation into broader ecological contexts. Two particular, distinguishing features of ecosystem engineers are that they affect the physical space in which other species live and their direct effects can last longer than the lifetime of the organism – engineering can in essence outlive the engineer. Together, these factors identify critical considerations that need to be included in models, experimental and observational work. The ecosystem engineering concept holds particular promise in the area of ecological applications, where influence over abiotic variables and their consequent effects on biotic communities may facilitate ecological restoration and counterbalance anthropogenic influences.     

Increased expression of OsPT1, a high-affinity phosphate transporter, enhances phosphate acquisition in rice

Most high-affinity phosphate transporter genes (OsPTs) in rice were highly induced in roots when phosphate was depleted. OsPT1, however, was highly expressed in primary roots and leaves regardless of external phosphate concentrations. This finding was confirmed histochemically using transgenic rice plants that express the GUS reporter gene under the control of the OsPT1 promoter, which exhibited high GUS activity even in the phosphate sufficient condition. Furthermore, transgenic rice plants overexpressing the OsPT1 gene accumulated almost twice as much phosphate in the shoots as did wild-type plants. As a result, transgenic plants had more tillers than did wild-type plants, which is a typical physiological indicator for phosphate status in rice.     

The in vitro anti-platelet,antioxidant and cellular immunity activity of <Emphasis Type="Italic">Phellinus gilvus</Emphasis> fractional extracts

The in vitro anti-platelet and antioxidant activities of various solvent extracts from Phellinus gilvus (PG), and the effects of hot water extract from PG (PGW) on murine cellular immunity were investigated. Chloroform extract (CE), methanol extract (ME) and butanol extract (BE) from PG could significantly inhibit platelet aggregation induced by thrombin. Ethyl acetate extract (EAE), BE, ME from PG had significant 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity compared with the control, and the EAE showed the highest effect with IC₅₀ values of 13.34 μg/ml, which is higher than that of ascorbic acid (40 μg/ml). In addition, EAE displayed the inhibition of xanthine oxidase (XO) activity with IC₅₀ value of 2.45 μg/ml. As to the cellular immunity activity, PGW could enhance both the lipopolysaccharide (LPS)-induced B lymphocyte proliferation and concanavalin A (Con A)-induced T lymphocyte proliferation in vitro. The phagocytosis of both peritoneal macrophages and RAW264.7 macrophage cells were also increased by the addition of PGW. Moreover, PGW was found to inhibit the nitric oxide (NO) production of RAW264.7 macrophages induced by LPS in a concentration-dependant manner.     

Improved cryopreservation of bovine preimplantation embryos cultured in chemically defined medium

The aim of this study was to examine the effects of modifications to a standard slow freezing protocol on the viability of in vitro produced bovine embryos. Bovine oocytes were matured, fertilized with frozen-thawed semen, and presumptive zygotes cultured in defined two-step culture media. The standard freezing medium was 1.5M ethylene glycol (EG), 0.1M sucrose, 10% fetal bovine serum (FBS) in Dulbecco's phosphate buffered saline (D-PBS). A preliminary trial showed that in vitro produced embryos cryopreserved in this medium had a survival rate of 74.6% at 24h and 53.5% at 48 h post-thaw. Experiment 1 studied the effects of omitting the sucrose supplement or replacing it with 0.1M xylose. In Experiment 2, the effects of partial (0%, 25% or 50%) or total (100%) replacement of sodium chloride with choline chloride in the cryopreservation medium were examined (the medium with 100% replacement was designated CJ1). The effects of replacing the 10% FBS with 0.4% BSA or 0.4% lipid-rich BSA (Albumax I) in CJ1 was studied in Experiment 3. In Experiment 4, pregnancy/calving rates following the post-thaw transfer of in vitro produced embryos cryopreserved in the standard freezing medium were compared with those of in vitro and in vivo produced embryos cryopreserved in the improved medium (Albumax I in CJ1). Supplementation of the cryopreservation medium with 0.1M sucrose resulted in higher post-thaw survival rates at 24 h (71.3% versus 53.5 and 51.7%; P<0.05), 48 h (51.1% versus 45.3 and 40.2%), and 72 h (34.0% versus 24.4 and 23.0%) than 0.1M xylose or no supplement, respectively, in Experiment 1. Experiment 2 showed that embryos cryopreserved in the standard medium had poorer survival rates at 24 h (72.8% versus 86.5%; P<0.05), 48 h (53.1% versus 66.3%) or 72 h (28.4% versus 44.9%) than those frozen in CJ1. The post-thaw survival rate of embryos frozen in medium supplemented with Albumax I was better than that for the FBS or BSA supplements at 24h (92.0% versus 90.7 and 87.3%), 48 h (87.3% versus 76.9 and 70.9%; P<0.05), and 72 h (70.4% versus 49.1 and 46 4%; P<0.05; Experiment 3). In Experiment 4, in vitro produced embryos cryopreserved in CJ1 medium supplemented with Albumax I resulted in higher pregnancy rates at Day 35 (31.9% versus 22.9%) and Day 60 (24.1% versus 14.3%) of gestation, and calving rates (22.6% versus 10.0%; P<0.05) than similar embryos frozen in the standard medium. However, in vivo produced embryos cryopreserved in Albumax I in CJ1 resulted in higher pregnancy rates at Day 35 (50.7%; P<0.05) and Day 60 (45.1%; P<0.05) of gestation, and calving rate (43.7%; P<0.05). It was concluded that modification of the freezing medium by addition of lipid-rich BSA and replacing sodium chloride with choline chloride improves the post-thaw survival of in vitro produced embryos, and their viability post-transfer.     

Short-term Administration of Water-soluble Silicon Improves Mineral Density of the Femur and Tibia in Ovariectomized Rats

Silicon is important for the proper growth and development of bone and connective tissues. This study was designed to investigate if water-soluble silicon could be used for the treatment of postmenopausal osteoporosis. Silicon (Si 20 mg/kg body weight/day) was administrated orally to 17-week-old ovariectomized (OVX) rats for 4 weeks. Silicon did not alter weight gain in OVX rats. Silicon supplementation significantly increased the bone mineral density of the femur (p < 0.05, vs. OVX control group) and tibia in OVX rats (p < 0.05, vs. OVX control group). Serum alkaline phosphatase and osteocalcin, two bone formation biomarkers tested, were not significantly altered, but urinary calcium and phosphorous excretion tended to decrease with silicon supplementation. OVX rats with silicon supplementation showed a relatively higher serum CTx compared to the nonsupplemented OVX group (p < 0.01, vs. OVX control group). According to these results, short-term soluble silicon supplementation improved bone mineral density in OVX-induced osteoporosis.     

Hepatitis C virus (HCV) genotyping by annealing reverse transcription-PCR products with genotype-specific capture probes

The genotype of the hepatitis C virus (HCV) strain infecting a given patient is an important predictive factor for the clinical outcome of chronic liver disease and its response to anti-viral therapeutic agents. We herein sought to develop a new easy, sensitive and accurate HCV genotyping method using annealing genotype-specific capture probes (AGSCP) in an automation-friendly 96-well plate format. The validation of our new AGSCP was performed using the Standard HCV Genotype Panel. We then used both our AGSCP and the commercially available INNO-LiPA assay to analyze the HCV genotypes from 111 Korean patients. Discordant results were analyzed by direct sequencing. AGSCP successfully genotyped the standard panel. The genotypes of 111 patient samples were also obtained successfully by AGSCP and INNO-LiPA. We observed a high concordance rate (93 matched samples, 83.8%) between the two assays. Sequencing analysis of the 18 discordant results revealed that the AGSCP had correctly identified 12 samples, whereas the INNO-LiPA had correctly identified only 6. These results collectively indicate that AGSCP assay is a convenient and sensitive method for large-scale genotyping, and it may be a promising tool for the determination of HCV and other genotypes in clinical settings.     

15N NMR relaxation studies of Y14F mutant of ketosteroid isomerase: the influence of mutation on backbone mobility

The backbone dynamics of Y14F mutant of Delta(5)-3-ketosteroid isomerase (KSI) from Comamonas testosteroni has been studied in free enzyme and its complex with a steroid analogue, 19-nortestosterone hemisuccinate (19-NTHS), by (15)N NMR relaxation measurements. Model-free analysis of the relaxation data showed that the single-point mutation induced a substantial decrease in the order parameters (S(2)) in free Y14F KSI, indicating that the backbone structures of Y14F KSI became significantly mobile by mutation, while the chemical shift analysis indicated that the structural perturbations of Y14F KSI were more profound than those of wild-type (WT) KSI upon 19-NTHS binding. In the 19-NTHS complexed Y14F KSI, however, the key active site residues including Tyr14, Asp38 and Asp99 or the regions around them remained flexible with significantly reduced S(2) values, whereas the S(2) values for many of the residues in Y14F KSI became even greater than those of WT KSI upon 19-NTHS binding. The results thus suggest that the hydrogen bond network in the active site might be disrupted by the Y14F mutation, resulting in a loss of the direct interactions between the catalytic residues and 19-NTHS.     

Enhancement of centelloside production from cultured plants of Centella asiatica by combination of thidiazuron and methyl jasmonate

In order to produce centellosides from whole plant cultures of Centella asiatica (L.) Urban, we evaluated the synergistic effects of thidiazuron (TDZ) and methyl jasmonate (MJ) on whole plant growth and centelloside production. After 4 weeks of treatment with 0.025 mg/L of TDZ coupled with 0.1 mM MJ, the production of madecassoside and asiaticoside from whole plant cultures was estimated to be 2.40- and 2.44-fold, respectively, above that of MJ elicitation alone. When whole plants were treated with a growth regulator and an elicitor, the growth of whole plants, as compared to the controls, did not differ. Additionally, total phytosyterol content in the leaves of whole plants co-treated with MJ and TDZ was 1.08-fold greater than those of MJ alone. These results demonstrate that combined treatments not only stimulate the accumulation of centellosides in the leaves but also inhibit the reduction of phytosterol levels caused by MJ elicitation.     

Functional characterization of the interaction between bacterial adhesin multivalent adhesion molecule 7 (MAM7) protein and its host cell ligands

The ability of a pathogen to rapidly form a stable interaction with the host cell surface is key to its success. Bacterial pathogens use a repertoire of virulence factors, but their efficient use relies on close contact between the host and the pathogen. We have recently identified a constitutively expressed MAM7 (multivalent adhesion molecule 7), which is widely distributed in gram-negative pathogens and enables them to establish initial contact with the host cell. Here, we describe the dissection of the MAM7 interaction with the host cell surface into two distinct binding events, involving the host protein fibronectin and the membrane phospholipid phosphatidic acid. We analyzed which domains within MAM7 and fibronectin are necessary for complex formation. We further studied phosphatidic acid binding by MAM7 using site-directed mutagenesis and liposome association assays and demonstrated that a specific distribution of basic charge on MAM7 is required for high affinity binding. Finally, we showed that fibronectin and phosphatidic acid binding to MAM7 are not mutually exclusive and that the three molecules likely assemble into a tripartite complex on the host cell surface.