The enterotoxin gene (cpe) of Clostridium perfringens can be chromosomal or plasmid-borne

The location of the cpe gene, encoding the enterotoxin responsible for food poisoning in humans, has been studied in a series of enterotoxigenic Ciostridium perfringens strains by means of pulsed field gel electrophoresis of genomic DNA. The cpe gene was found at the same chromosomal locus in strains associated with food poisoning in humans and was shown to be linked to a repetitive sequence, the Hin dlll repeat, and an open reading frame, ORF3, that may be part of an insertion sequence. In contrast, when the strains originated from domesticated livestock cpe was located on a large episome where it was often close to a copy of the transposable element IS 1151. In these cases, the Hin dlll repeat was not linked to the cpe gene although this was generally preceded by ORF3.     

Biological nitrogen fixation and phosphate solubilization by bacteria isolated from tropical soils

Introduction

In addition to fixing atmospheric nitrogen, some bacterial isolates can also solubilize insoluble phosphates, further contributing to plant growth.

Aims

The objectives of this study were the following: isolate, select, and identify nodulating bacteria in the cowpea that are efficient not only in biological nitrogen fixation (BNF) but also in the solubilization of insoluble inorganic phosphates; identify and quantify the organic acids produced; and establish the relationship between those acids and the solubilizing capacity.

Methods

The bacteria were captured from two soils containing high concentrations of insoluble phosphorus from the cities of Lavras and Patos de Minas, using the cowpea [Vigna unguiculata (L.) Walp.] as bait. We obtained 78 strains, which were characterized according to their cultural attributes in culture medium 79 with the strains UFLA 03-84, INPA 03-11B, and BR3267 (approved by the Ministry of Livestock and Supply Agriculture—MAPA, as inoculants for the cowpea) and Burkholderia cepacia (LMG1222^T), which was used as a positive control for phosphate solubilization. Strains that were selected for their efficiency in both processes were identified by 16S rDNA sequence analysis. We evaluated the symbiotic efficiency (BNF) in a greenhouse and the solubilization efficiency of CaHPO₄, Al(H₂PO₄)₃, and FePO₄.2H₂O in solid and liquid GELP media. Strains that excelled at the solubilization of these phosphate sources were also evaluated for the production of the following organic acids: oxalic, citric, gluconic, lactic, succinic, and propionic.

Results

The presence of Acinetobacter, Bacillus, Firmicutes, Microbacterium, Paenibacillus, and Rhizobium was detected by 16S rDNA sequencing and analysis. Bacterial strains obtained from cowpea nodules varied greatly in the efficiency of their BNF and phosphate solubilization processes, especially in the strains UFLA 03-09, UFLA 03-10, UFLA 03-12, and UFLA 03-13, which were more efficient in both processes. More strains were able to solubilize insoluble inorganic calcium and iron phosphates in liquid medium than in solid medium. The production of organic acids was related to the solubilization of CaHPO₄ and FePO₄.2H₂O for some strains, and the type and concentration of the acid influenced this process.

Conclusions

These are the first results obtained with bacterial isolates from tropical soils in which the production of organic acids was detected and quantified to examine the solubilization of insoluble inorganic phosphates.     

Auxin metabolism and rooting in young and mature clones of Sequoia sempervirens

The capacity of young and mature Sequoia sempervirens clones to produce roots in vitro was studied after wounding and indole-3-butyric acid (IBA) treatments. Rooting was not observed in mature or in young cuttings cultivated for 30 days in medium without IBA. The presence of 25 μ M IBA in the medium resulted in the appearance of roots at the base of the cuttings. More roots appeared and grew faster on cuttings of the young than on the mature clone. This difference in rooting capacity between young and mature cuttings may be related to differences in the hormone levels at the base of the 5 mm long cuttings during the first 4 days of the root inductive period. After HPLC fractionation. IAA. IBA and related compounds, including indole-3-aspartic acid (IAAsp) and IBA-glucose ester (IBA-GE), were determined by MS and MS-MS and their levels measured by ELISA. Another immunoreactive compound was also found and determined to be N,N-dimethyltryptophan (DMT), a compound previously reported to inhibit auxin-enhanced ethylene production. Wounding of the stem without IBA treatment revealed a transient increase in IAA, IAAsp and DMT levels in young cuttings while a dramatic increase in the levels of DMT was observed in mature cuttings. Following IBA treatment. IAA levels increased in both clones, but higher levels were measured in the young than in the mature clone. IBA and IBA-GE were also found but in higher levels in the mature clone. Thus, the difficult-to-root mature clone differs from the young clone in its auxin metabolism.     

Ants impact the composition of the aquatic macroinvertebrate communities of a myrmecophytic tank bromeliad

In an inundated Mexican forest, 89 out of 92 myrmecophytic tank bromeliads (Aechmea bracteata) housed an associated ant colony: 13 sheltered Azteca serica, 43 Dolichoderus bispinosus, and 33 Neoponera villosa. Ant presence has a positive impact on the diversity of the aquatic macroinvertebrate communities (n = 30 bromeliads studied). A Principal Component Analysis (PCA) showed that the presence and the species of ant are not correlated to bromeliad size, quantity of water, number of wells, filtered organic matter or incident radiation. The PCA and a generalized linear model showed that the presence of Azteca serica differed from the presence of the other two ant species or no ants in its effects on the aquatic invertebrate community (more predators). Therefore, both ant presence and species of ant affect the composition of the aquatic macroinvertebrate communities in the tanks of A. bracteata, likely due to ant deposition of feces and other waste in these tanks.     

The long periodicity phase (LPP) controversy part I: The influence of a natural-like ratio of the CER[EOS] analogue [EOS]-br in a CER[NP]/[AP] based stratum corneum modelling system: A neutron diffraction study

This study used neutron diffraction to investigate a ceramide-[NP] C24/[AP] C24 /[EOS]-br C30/cholesterol/lignoceric acid (0.6: 0.3: 0.1: 0.7: 1) based stratum corneum modelling system. By adding specifically deuterated ceramides-[NP]-D₃, [AP]-D₃, and [EOS]-br-D₃, detailed information on the lamellar and the nanostructure of the system was obtained. For the short periodicity phase a natural-like lamellar repeat distance of 5.47?±?0.02?nm was observed, similar to the [NP]/[AP] base system without the [EOS]-br. Unlike in this system the ceramides here were slightly tilted, hinting towards a slightly less natural arrangement. Due to the deuteration it was possible to observe that the long ceramide chains were overlapping in the lamellar mid-plane. This is considered to be an important feature for the natural stratum corneum. Despite the presence of a ceramide [EOS] analogue – able to form a long phase arrangement – no distinct long periodicity phase was formed, despite a slightly higher than natural ω-acyl ceramide ratio of 10?mol%. The deuterated variant of this ceramide determined that the very long ceramide was integrated into the short periodicity phase, spanning multiple layers instead. The – compared to the base system – unchanged repeat distance highlights the stability of this structure. Furthermore, the localisation of the very long ceramide in the short periodicity phase indicates the possibility of a crosslinking effect and thus a multilayer stabilizing role for the ceramide [EOS]. It can be concluded, that additionally to the mere presence of ceramide-[EOS] more complex conditions have to be met in order to form this long phase. This has to be further investigated in the future.     

Robo4 maintains vessel integrity and inhibits angiogenesis by interacting with UNC5B

Robo4 is an endothelial cell-specific member of the Roundabout axon guidance receptor family. To identify Robo4 binding partners, we performed a protein-protein interaction screen with the Robo4 extracellular domain. We find that Robo4 specifically binds to UNC5B, a vascular Netrin receptor, revealing unexpected interactions between two endothelial guidance receptors. We show that Robo4 maintains vessel integrity by activating UNC5B, which inhibits signaling downstream of vascular endothelial growth factor (VEGF). Function-blocking monoclonal antibodies against Robo4 and UNC5B increase angiogenesis and disrupt vessel integrity. Soluble Robo4 protein inhibits VEGF-induced vessel permeability and rescues barrier defects in Robo4(-/-) mice, but not in mice treated with anti-UNC5B. Thus, Robo4-UNC5B signaling maintains vascular integrity by counteracting VEGF signaling in endothelial cells, identifying a novel function of guidance receptor interactions in the vasculature.     

Chemical composition and biological activity of the essential oil of Ballota hispanica (L.) Benth. growing wild in Sicily

Ballota L. comprises several relevant species largely used for their excellent therapeutic properties. Ballota hispanica (L.) Benth. is widely used in herbal medicine, and it is sold in herbalist shops for its sedative and antispasmodic properties. Considering its traditional medicinal use and the lack of scientific studies on the volatile components of this species as well as on its biological activities, in this study the chemical composition of the essential oil from aerial parts of B. hispanica, collected in Sicily, was evaluated by gas chromatography and gas chromatography–mass spectrometry. α-Elemol was the most abundant component of the oil (10.9%), followed by α-ylangene (8.5%), γ-dodecalactone (5.1%), and manoyl oxide (4.8%). A comparison was made of the composition of the different Ballota taxa studied so far showing a peculiar profile of B. hispanica. Futhermore, the antimicrobial and the free radical scavenging activities of the oil were determined.     

Cardiac sodium channel Na(v)1.5 interacts with and is regulated by the protein tyrosine phosphatase PTPH1

In order to identify proteins interacting with the cardiac voltage-gated sodium channel Na(v)1.5, we used the last 66 amino acids of the C-terminus of the channel as bait to screen a human cardiac cDNA library. We identified the protein tyrosine phosphatase PTPH1 as an interacting protein. Pull-down experiments confirmed the interaction, and indicated that it depends on the PDZ-domain binding motif of Na(v)1.5. Co-expression experiments in HEK293 cells showed that PTPH1 shifts the Na(v)1.5 availability relationship toward hyperpolarized potentials, whereas an inactive PTPH1 or the tyrosine kinase Fyn does the opposite. The results of this study suggest that tyrosine phosphorylation destabilizes the inactivated state of Na(v)1.5.     

Identifying and characterizing the most significant β-glucosidase of the novel species Aspergillus saccharolyticus

The newly discovered fungal species Aspergillus saccharolyticus was found to produce a culture broth rich in β-glucosidase activity. In this present work, the main β-glucosidase of A.?saccharolyticus responsible for the efficient hydrolytic activity was identified, isolated, and characterized. Ion exchange chromatography was used to fractionate the culture broth, yielding fractions with high β-glucosidase activity and only 1 visible band on an SDS-PAGE gel. Mass spectrometry analysis of this band gave peptide matches to β-glucosidases from aspergilli. Through a polymerase chain reaction approach using degenerate primers and genome walking, a 2919 bp sequence encoding the 860 amino acid BGL1 polypeptide was determined. BGL1 of A.?saccharolyticus has 91% and 82% identity with BGL1 from Aspergillus aculeatus and BGL1 from Aspergillus niger , respectively, both belonging to Glycoside Hydrolase family 3. Homology modeling studies suggested β-glucosidase activity with preserved retaining mechanism and a wider catalytic pocket compared with other β-glucosidases. The bgl1 gene was heterologously expressed in Trichoderma reesei QM6a, purified, and characterized by enzyme kinetics studies. The enzyme can hydrolyze cellobiose, p-nitrophenyl-β-d-glucoside, and cellodextrins. The enzyme showed good thermostability, was stable at 50?°C, and at 60?°C it had a half-life of approximately 6?h.