Structural analysis of the sheath of a sheathed bacterium, Leptothrix cholodnii

Leptothrix cholodnii is an aerobic sheath-forming bacterium often found in oligotrophic and metal-rich aquatic environments. The sheath of this bacterium was isolated by selectively lysing the cells. Glycine and cysteine were the major amino acids of the sheath. The sheath was readily dissolved in hydrazine, and a polysaccharide substituted with cysteine was recovered from the solution. Galactosamine, glucosamine and galacturonic acid were detected in the hydrazinolysate by gas liquid chromatography analysis. FAB-MS analysis of the hydrazinolysate suggested a sugar sequence of HexN-GalA-HexN-HexN. Methylation linkage analysis revealed the presence of 4-linked GalA, 3-linked HexN and 4-linked HexN. The sulfhydryl groups of the sheath were used for labeling with the fluorogenic reagent, 4-(aminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole (ABD-F). The labeled sheath (ABD-sheath) was partially hydrolyzed and three fluorescent fragments were purified by HPLC. One of them was identified as ABD-cysteine. The second one was found to be the ABD-cysteine tetramer. Another fragment was indicated to be a pentasaccharide substituted with ABD-cysteine by nuclear magnetic resonance (NMR) analysis. It can be assumed that the polysaccharide and peptide moieties of the sheath are connected by a cysteine residue. NMR analysis of the hydrazinolysate revealed that the polysaccharide moiety of the sheath was constructed from a pentasaccharide repeating unit containing 2-amino-2-deoxygalacturonic acid (GalNA), as shown below. -->4)-alpha-GalNA-(1-->4)-alpha-D-GalN(p)-(1-->4)-alpha-D-GalA(p)-(1-->4)-beta-D-GlcN(p)-(1-->3)-beta-D-GalN(p)-(1-->.     

Use of microelectrodes to investigate the effects of 2-chlorophenol on microbial activities in biofilms

In order to assess the applicability of using microelectrodes as a tool for inhibition tests, temporal and spatial inhibitory effects of 2-chlorophenol (2-CP) on O(2) respiration and nitrification activities in municipal wastewater biofilms were investigated using microelectrodes for O(2) and NH(4)(+). The time-course microelectrode measurements demonstrated that 2-CP inhibited O(2) respiration and nitrification activities within 6-18 min. The microbial activities were inhibited only in the upper 400 microm of the biofilms by 2-CP, and the bacteria present in the deeper parts of the biofilms were still active, probably due to limited penetration of 2-CP. These results could reasonably explain the difference in inhibitory ratios of the O(2) respiration and nitrification activities in the biofilms. O(2) respiration activity was incompletely inhibited, which was attributed to the presence of O(2) respiration activities in the deeper parts of the biofilm. In contrast, nitrification activity was significantly inhibited because ammonia-oxidizing bacteria were present in the upper parts of the biofilm. These results indicate that the microelectrodes with a very quick response time and a high spatial resolution are useful tools to study temporal and spatial inhibitory effects of inhibitors on in situ microbial activities in biofilms.     

Synthesis and characterization of a novel reagent containing dansyl group, which specifically alkylates sulfhydryl group: an example of application for protein chemistry

We have synthesized a novel reagent containing dansyl group, iodoacethyl dansylcadaverine (IADC), which specifically alkylates sulfhydryl groups. The carboxyl group of iodoacetic acid was activated with dicyclohexylcarbodiimide and was condensed with amino group of dansylcadaverine. Purity and chemical structure of IADC was confirmed with mass spectrometry (MS) and NMR. IADC alkylated GSH but not GSSG, which was confirmed by MS. The reactivity of IADC with proteins was also investigated with Western blotting using anti-dansyl antibody. IADC reacted only with sulfhydryl-containing proteins. The specificity of the interaction of IADC with sulfhydryl groups in proteins was confirmed by adding excessive amount of a well-known sulfhydryl-specific reagent, 5, 5'-dithiobis(2-nitrobenzoic acid), which led to a complete inhibition. To show the usefulness of IADC, the cysteines in glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from chicken muscle were modified with this reagent, and GAPDH was then digested by lysyl endopeptidase. The peptides generated from digestion of IADC-incorporated GAPDH were applied to an anti-dansyl immunoaffinity column. The peptide fragments bound and eluted from the column were separated by HPLC, and the amino acid sequence of each peptide was analyzed, and peptide was identified as the one containing a Cys residue(s). These data showed that IADC is a useful reagent to specifically identify the positions of a Cys residue(s) in proteins.     

Study of biological communities subject to imperfect detection: bias and precision of community <Emphasis Type="Italic">N</Emphasis>-mixture abundance models in small-sample situations

Community N-mixture abundance models for replicated counts provide a powerful and novel framework for drawing inferences related to species abundance within communities subject to imperfect detection. To assess the performance of these models, and to compare them to related community occupancy models in situations with marginal information, we used simulation to examine the effects of mean abundance \((\bar{\lambda }\): 0.1, 0.5, 1, 5), detection probability \((\bar{p}\): 0.1, 0.2, 0.5), and number of sampling sites (n _site : 10, 20, 40) and visits (n _visit : 2, 3, 4) on the bias and precision of species-level parameters (mean abundance and covariate effect) and a community-level parameter (species richness). Bias and imprecision of estimates decreased when any of the four variables \((\bar{\lambda }\), \(\bar{p}\), n _site , n _visit ) increased. Detection probability \(\bar{p}\) was most important for the estimates of mean abundance, while \(\bar{\lambda }\) was most influential for covariate effect and species richness estimates. For all parameters, increasing n _site was more beneficial than increasing n _visit . Minimal conditions for obtaining adequate performance of community abundance models were n _site  ≥ 20, \(\bar{p}\) ≥ 0.2, and \(\bar{\lambda }\) ≥ 0.5. At lower abundance, the performance of community abundance and community occupancy models as species richness estimators were comparable. We then used additive partitioning analysis to reveal that raw species counts can overestimate β diversity both of species richness and the Shannon index, while community abundance models yielded better estimates. Community N-mixture abundance models thus have great potential for use with community ecology or conservation applications provided that replicated counts are available.     

Role of mitochondrial phosphate carrier in metabolism-secretion coupling in rat insulinoma cell line INS-1

In pancreatic β-cells, glucose-induced mitochondrial ATP production plays an important role in insulin secretion. The mitochondrial phosphate carrier PiC is a member of the SLC25 (solute carrier family 25) family and transports Pi from the cytosol into the mitochondrial matrix. Since intramitochondrial Pi is an essential substrate for mitochondrial ATP production by complex V (ATP synthase) and affects the activity of the respiratory chain, Pi transport via PiC may be a rate-limiting step for ATP production. We evaluated the role of PiC in metabolism-secretion coupling in pancreatic β-cells using INS-1 cells manipulated to reduce PiC expression by siRNA (small interfering RNA). Consequent reduction of the PiC protein level decreased glucose (10 mM)-stimulated insulin secretion, the ATP:ADP ratio in the presence of 10 mM glucose and elevation of intracellular calcium concentration in response to 10 mM glucose without affecting the mitochondrial membrane potential (Δψm) in INS-1 cells. In experiments using the mitochondrial fraction of INS-1 cells in the presence of 1 mM succinate, PiC down-regulation decreased ATP production at various Pi concentrations ranging from 0.001 to 10 mM, but did not affect Δψm at 3 mM Pi. In conclusion, the Pi supply to mitochondria via PiC plays a critical role in ATP production and metabolism-secretion coupling in INS-1 cells.     

Role of Nrf2 in host defense against influenza virus in cigarette smoke-exposed mice

Influenza virus is a common respiratory tract viral infection. Although influenza can be fatal in patients with chronic pulmonary diseases such as chronic obstructive pulmonary disease, its pathogenesis is not fully understood. The Nrf2-mediated antioxidant system is essential to protect the lungs from oxidative injury and inflammation. In the present study, we investigated the role of Nrf2 in protection against influenza virus-induced pulmonary inflammation after cigarette smoke exposure with both in vitro and in vivo approaches. For in vitro analyses, peritoneal macrophages isolated from wild-type and Nrf2-deficient mice were treated with poly(I:C) and/or cigarette smoke extract. For in vivo analysis, these mice were infected with influenza A virus with or without exposure to cigarette smoke. In Nrf2-deficient macrophages, NF-κB activation and the induction of its target inflammatory genes were enhanced after costimulation with cigarette smoke extract and poly(I:C) compared with wild-type macrophages. The induction of antioxidant genes was observed for the lungs of wild-type mice but not those of Nrf2-deficient mice after cigarette smoke exposure. Cigarette smoke-exposed Nrf2-deficient mice showed higher rates of mortality than did wild-type mice after influenza virus infection, with enhanced peribronchial inflammation, lung permeability damage, and mucus hypersecretion. Lung oxidant levels and NF-κB-mediated inflammatory gene expression in the lungs were also enhanced in Nrf2-deficient mice. Our data indicate that the antioxidant pathway controlled by Nrf2 is pivotal for protection against the development of influenza virus-induced pulmonary inflammation and injury under oxidative conditions.     

Indigenous utilization of termite mounds and their sustainability in a rice growing village of the central plain of Laos

Background

This study was conducted to identify medicinal plants and spices used for medicine by the community of Beni-Sueif, Upper Egypt.

Methods

Ethnobotanical data from local people was collected using direct interviews and a semi-structured questionnaire.

Results

Forty-eight plant species belonging to twenty-seven families and forty-seven genera were encountered during the study. Their botanical and vernacular names, plant parts used and medicinal uses are given. Results of the study were analyzed using two quantitative tools. The factor informant consensus indicated the agreement in the use of plants and the fidelity level indicated the ratio between the number of informants who independently suggested the use of a species for the same major purpose and the total number of informants who mentioned the plant for any use. The results of the factor informant consensus showed that the cardiovascular category has the greatest agreement, followed by the immunological, gastrointestinal and respiratory categories. The most important species according to their fidelity are: Hibiscus sabdariffa L. for the cardiovascular category; Trigonella foenum-graecum L. for the immunological category; Mentha piperita L. for the gastrointestinal category and Pimpinella anisum L. for the respiratory category.

Conclusions

Medicinal plants are still used for treatment in Beni-Sueif community despite the availability of prescribed medications. Documentation of this ethnomedicinal knowledge is important. Evaluation of pharmacological activity for the promising medicinal plants is suggested.     

Effects of Adding Alkaline Material on the Heavy Metal Chemical Fractions in Soil under Flooded and Non-Flooded Conditions

We investigated the effect of adding an alkaline material (containing calcium carbonate and gypsum) on the immobilization of heavy metals (Cd, Cu, Pb, and Zn) in a paddy soil slightly contaminated with Cd and Zn under flooded and non-flooded conditions in the laboratory. Adding the alkaline material increased the soil pH and significantly decreased the exchangeable fraction of all of the metals, especially for Cd (>75% decrease) and Zn (ca. 90% decrease), under both flooded and non-flooded conditions. Drying the flooded soil samples increased the ratio of exchangeable fraction to the total fraction, particularly for Cd. The exchangeable fraction ratio was lower in the dried, previously flooded samples that contained the alkaline material than in the samples that did not contain the alkaline material, indicating that adding the alkaline material would be an effective way of immobilizing heavy metals during the oxidation of anoxic soils. These results show that the alkaline material can be used to immobilize heavy metals under both anoxic and oxic conditions, and that the effects of flooding and amending a paddy soil with alkaline material on the chemical forms will be different between heavy metals.