The copper-resistant bacterium ACU isolated from the rhizosphere of Eichhornia crassipes (Mart.) increased the endurance of Potamogeton crispus L. to copper toxicity

Aims: This study aimed to develop endurance to copper stress in Potamogeton crispus L. by inoculation with the anti-copper strain ACU – a novel Enterobacteriaceae bacterium isolated from the rhizosphere of Eichhornia crassipes with high copper-removal ability. Methods and Results: A spherical copper-resistant bacterium, namely ACU, was isolated from the rhizosphere of E. crassipes. It was demonstrated to have substantial copper-removing capability, even at copper concentrations as high as 69 mg l⁻¹. The 16S rRNA gene sequence of ACU suggested it to be a novel Enterobacteriaceae bacterium most closely related to Providencia sp. With increasing copper concentrations, the growth rate of ACU gradually decreased with a delay in the logarithmic growth phase. ACU demonstrated high copper-removal ability at the lag phase when cultivated in media with high copper concentrations. A 48-kDa extracellular copper-binding protein was detected in ACU. When P. crispus was inoculated with ACU, the growth ability of P. crispus significantly improved at all the tested copper concentrations, and the lethal time for 10 mg l⁻¹ was delayed. Further study revealed that while ACU cells were rarely detected in the culture solution, they were associated with the surface of P. crispus. These findings indicated that ACU grew by anchoring itself on the surface of P. crispus and could increase the ability of P. crispus to resist copper toxicity. Conclusion: To the best of our knowledge, the Enterobacteriaceae bacterium ACU is a novel nonpathogenic bacterium with high copper-removing ability from water. Significance and Impact of the Study: This study demonstrated that the Enterobacteriaceae bacterium ACU has potential applicability for use in copper removal and in the protection of aquatic plants in copper-polluted water.     

Sequence analysis of a cryptic plasmid from Flavobacterium sp. KP1, a psychrophilic bacterium

A cryptic plasmid found at high copy number was isolated from Flavobacterium sp. KP1, a psychrophilic Gram-negative bacterium, cloned, and sequenced. The sequence will appear in the DDBJ/EMBL/GenBank databases under the accession number AB007196. The pFL1 plasmid is 2311 nucleotides in length with 32.7% GC content, and shows a distinctive nucleotide sequence without homology to other plasmids of similar length. The plasmid contains two open reading frames of significant length, ORFI and ORFII. ORFI encodes a protein similar to the replication proteins found in Gram-negative bacterial plasmids, Bacteroides fragilis plasmid pBI143 and Zymomonas mobilis plasmid pZM2. The putative translation product of ORFII shows homologies with plasmid recombination proteins found mainly in Gram-positive bacterial plasmids such as Staphylococcus aureus plasmid pT181.     

Occurrence of Pasteuria-like Organisms on Selected Plant-Pamsitic Nematodes of Pineapple in the Hawaiian Islands

Soils from 320 sites representing diverse undisturbed habitats from five Hawaiian Islands were assessed for occurrence of Pasteuria-like organisms. Mean annual rainfall at sites ranged from 125-350 cm, elevation from 69-2,286 m, and annual mean temperature from 12-24 C. Seven different natural communities were represented: wet lowland, mesic lowland, wet montane, mesk montane, dry montane, mesic subalpine, and dry alpine. Pasteuria spp. in a soil sample was detected by baiting with infective stages of Helicotylenchus dihystera, Meloidogyne javanica, Pratylenchus brachyurus, and Rotylenchulus reniformis, followed by cultivation of the nematodes on pineapple plants for 10-11 months. All nematode baits except R. reniformis were readily recovered from the soil samples. A sample was considered Pasteuria-positive if at least 5 % of the nematode specimens showed endospore attachment. Thirteen percent of all samples were positive for Pasteuria-like organisms. The frequencies of association between Pasteuria spp. and Meloidogyne, Helicotylenchus, or Pratylenchus species were 52%, 24%, and 24%, respectively. Positive samples were more prevalent on the older islands of Kauai and Oahu (75%), in lowland communities (61%), and in areas with introduced vegetation (60%). More than 27% of the positive samples were associated with plant species in a few selected families that included Meliaceae and Myrtaceae. Occurrence of Pasteuria spp. seemed to be positively associated with mean annual rainfall or temperature, but negatively associated with elevation.     

Expansion of the geographic distribution of a novel lineage of epsilon-Proteobacteria to a hydrothermal vent site on the Southern East Pacific Rise

The diversity associated with a microbial mat sample collected from a deep-sea hydrothermal vent on the Southern East Pacific Rise was determined using a molecular phylogenetic approach based on the comparison of sequences from the small subunit ribosomal RNA gene (16S rDNA). The DNA was extracted from the sample and the 16S rDNA was amplified by PCR. Sixteen different phylotypes were identified by restriction fragment length polymorphism analysis; four phylotypes were later identified as putative chimeras. Analysis of the 16S rDNA sequences placed all the phylotypes within the Proteobacteria. The majority of the sequences (98%) were most closely related to a new clade of epsilon-Proteobacteria that were initially identified from an in situ growth chamber deployed on a deep-sea hydrothermal vent on the Mid-Atlantic Ridge in 1995. The similarity between phylotypes identified from Atlantic and Pacific deep-sea hydrothermal vent sites indicates that this new clade of Proteobacteria may be endemic to and widely distributed among deep-sea hydrothermal vents.     

Biochemical and morphological alterations in lungs induced by experimental inhibition of fibrinolytic activity

The fibrinolytic system is known to play an important role in the protection of lung architecture and function. This study investigated the effects on lungs of inhibiting the fibrinolytic system using tranexamic acid (TXA). Thirty cats were used, 15 experimental and 15 control. TXA was administered intravenously to the experimental animals for 3 h at 200 mg/kg (acute) and 7 days at 100 mg/kg (chronic). Blood samples were obtained from the carotid artery. The acute dose cats were sacrificed at 3 h and 24 h and the chronic dose cats at 8 days. Samples of inflated and fixed lung were examined morphologically and their collagen contents were determined. Fibrinolytic activity in blood samples was determined by fibrinogen degradation products levels, fibrin plate lytic area diameter, and the euglobulin lysis time. Hyperemia, lung interstitial oedema, haemorrhaging, inflammatory cell infiltration, pneumocyte type II cell proliferation, thrombosis and emphysema-related changes, characterized by enlargement of air spaces accompanied by destruction of alveolar walls, were observed in experimental cats group. None of these alterations except hyperemia and lung interstitial oedema were observed in two control animals. Electron microscopy results revealed oedema fluid in the interstitium, proliferation of pneumocyte type II cells, thickening of the alveolar septa and presence of marked amounts of collagen. Vacuoles were seen in the capillary endothelial cells. Elastic tissue was observed as elastic masses and partly disrupted, although elastic fibers were not prominent in all parts of the interstitium. Collagen content in the chronic dose experimental group was significantly higher than in all control and acute dose experimental groups. The inhibition of fibrinolytic system appears to have caused the emphysematous alterations, alveolar wall destruction and collagen accumulation possibly by causing microthromboses leading to mechanical blockage-ischemic changes, or by causing secondary fibrinolysis as a result of fibrin degradation products affecting local plasminogen activators and proteases. An injury-repair process also appears to have occurred.     

Structural Revision of Sulfated Polysaccharide B-1 Isolated from a Marine <Emphasis Type="Italic">Pseudomonas</Emphasis> Species and Its Cytotoxic Activity Against Human Cancer Cell Lines

Abstract The structure of a sulfated polysaccharide (B-1) isolated and purified from the culture filtrate of marine Pseudomonas sp. WAK-1 was revised to have a repeating unit as follows: -2)-β-D-Galp(4SO₄)(1-4)[β-D-Glcp(1-6)]-β-D-Galp(3SO₄)(1-. B-1 was evaluated for anticancer activity using a human cancer cell line panel coupled with a drug sensitivity database. The average B-1 concentration required for 50% growth inhibition against the panel of 39 cell lines was 63.2 μg/ml. Among the cancer cell lines tested, high sensitivities to B-1 were observed in central nervous system cancer and lung cancer cell lines. The COMPARE analysis revealed that the differential growth inhibition pattern of B-1 had no significant correlation with those of more than 200 standard compounds, most of which were anticancer drugs and different types of inhibitors. This lack of similarities in the cytotoxic patterns appears to reflect previously unrecognized biological properties of B-1. It was revealed that B-1 induced apoptosis in U937 cells, as shown by cell morphology and internucleosomal DNA fragmentation.     

Structure of an acidic O-specific polysaccharide from marine bacterium Shewanella fidelis KMM 3582T containing Nepsilon-[(S)-1-carboxyethyl]-Nalpha-(D-galacturonoyl)-L-lysine

The O-specific polysaccharide was isolated by mild acid degradation of the lipopolysaccharide of the marine bacterium Shewanella fidelis type strain KMM 3582T and studied by sugar analysis along with 1H and 13C NMR spectroscopy including one-dimensional NOE in difference mode and two-dimensional experiments. The polysaccharide was found to consist of linear tetrasaccharide repeating units containing Nepsilon-[(S)-1-carboxyethyl]-Nalpha-(D-galacturonoyl)-L-lysine and having the following structure: [See text.] The amide of D-galacturonic acid with Nepsilon-[(S)-1-carboxyethyl]-L-lysine ('alaninolysine', 2S,8S-AlaLys) was found for the first time in nature as a component of the O-specific polysaccharide of Providencia rustigianii O14 (Carbohydr. Res. 2003, 338, 1009-1016).     

Structural basis for broad substrate specificity of earthworm fibrinolytic enzyme component A

Earthworm fibrinolytic enzyme component A (EFE-a) possesses an S1 pocket, which is typical for an elastase-like enzyme, but it can still hydrolyze varieties of substrates, and it exhibits wide substrate specificity. Former structure studies suggested that the four-residue insertion after Val(217) might endow EFE-a with this specificity. Based on the native crystal structure at a resolution of 2.3A, we improved the native crystal structure to 1.8A and determined its complex structure with the inhibitor Meo-Suc-Ala-Ala-Pro-Val-CMK at a resolution of 1.9A. The final structures show that: (1) EFE-a possesses multisubstrate-binding sites interacting with the substrates; (2) significant conformation adjustment takes place at two loops binding to the N-terminal of the substrates, which may enhance the interaction between the enzyme and the substrates. These characteristics make the substrate-specificity of EFE-a less dependent on the property of its S1-pocket and may endow the enzyme with the ability to hydrolyze chymotrypsin-specific substrates and even trypsin-specific substrates.