Molecular cloning and characterization of FHL2, a novel LIM domain protein preferentially expressed in human heart

A full-length cDNA clone encoding a novel LIM-only protein was isolated and sequenced from a human fetal heart cDNA library. This full-length clone consists of 1416 base pairs and has a predicted open reading frame (ORF) encoding 279 amino acids. The ORF of this polypeptide codes for the human heart-specific

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our and a

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alf

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IM-only protein

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(FHL2). It possesses an extra zinc finger that is a half LIM domain and four repeats of LIM domain. When the human FHL2 cDNA probe was used to hybridize with poly-A RNA of various human tissues, a very strong signal could be seen in heart tissues, and only moderately low signals could be detected in placenta, skeletal muscle and ovary. Virtually no signal could be detected in brain, lung, liver, kidney, pancreas, spleen, thymus, prostate, testis, small intestine, colon or peripheral blood leukocyte. FHL2 was mapped to chromosome 2q12–q13 by fluorescent in-situ hybridization (FISH).     

Chemotaxonomy of Trichoderma spp. Using mass spectrometry-based metabolite profiling

In this study, seven Trichoderma species (33 strains) were classified using secondary metabolite profile-based chemotaxonomy. Secondary metabolites were analyzed by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS) and multivariate statistical methods. T. longibrachiatum and T. virens were independently clustered based on both internal transcribed spacer (ITS) sequence and secondary metabolite analyses. T. harzianum formed three subclusters in the ITS-based phylogenetic tree and two subclusters in the metabolitebased dendrogram. In contrast, T. koningii and T. atroviride strains were mixed in one cluster in the phylogenetic tree, whereas T. koningii was grouped in a different subcluster from T. atroviride and T. hamatum in the chemotaxonomic tree. Partial least-squares discriminant analysis (PLS-DA) was applied to determine which metabolites were responsible for the clustering patterns observed for the different Trichoderma strains. The metabolites were hetelidic acid, sorbicillinol, trichodermanone C, giocladic acid, bisorbicillinol, and three unidentified compounds in the comparison of T. virens and T. longibrachiatum; harzianic acid, demethylharzianic acid, homoharzianic acid, and three unidentified compounds in T. harzianum I and II; and koninginin B, E, and D, and six unidentified compounds in T. koningii and T. atroviride. The results of this study demonstrate that secondary metabolite profiling-based chemotaxonomy has distinct advantages relative to ITSbased classification, since it identified new Trichoderma clusters that were not found using the latter approach.     

Screening and identification of antimicrobial compounds from Streptomyces bottropensis suppressing rice bacterial blight

Xanthomonas oryzae pv. oryzae (Xoo) is the most devastating pathogen to Oryza sativa and has been shown to cause bacterial blight. Two bioactive compounds showing antimicrobial activities against Xoo strain KACC 10331 were isolated from a Streptomyces bottropensis strain. The ethyl acetate extract was fractionated on a Sephadex LH-20 column, and then purified by preparative HPLC. The purified compounds were identified as bottromycin A2 and dunaimycin D3S by HR/MS and 1H NMR analyses. The MIC value against Xoo and the lowest concentration still capable of suppressing rice bacterial blight were 2 microgram/ml and 16 microgram/ml for bottromycin A2, and 64 microgram/ml and 0.06 microgram/ml for dunaimycin D3S, respectively. These two compounds were shown to exert different bioactivities in vitro and in rice leaf explants.     

Culture condition-dependent metabolite profiling of Aspergillus fumigatus with antifungal activity

Three sections of Aspergillus (five species, 21 strains) were classified according to culture medium-dependent and time-dependent secondary metabolite profile-based chemotaxonomy. Secondary metabolites were analysed by liquid chromatography–electrospray ionisation tandem mass spectrometry (LC–ESI-MS–MS) and multivariate statistical methods. From the Aspergillus sections that were cultured on malt extract agar (MEA) and Czapek yeast extract agar (CYA) for 7, 12, and 16 d, Aspergillus sections Fumigati (A. fumigatus), Nigri (A. niger), and Flavi (A. flavus, A. oryzae, and A. sojae) clustered separately on the basis of the results of the secondary metabolite analyses at 16 d regardless of culture medium. Based on orthogonal projection to latent structures discriminant analysis by partial least squares discriminant analysis (PLS-DA), we identified the secondary metabolites that helped differentiate sections between A. fumigatus and Aspergillus section Flavi to be gliotoxin G, fumigatin oxide, fumigatin, pseurotin A or D, fumiquinazoline D, fumagillin, helvolic acid, 1,2-dihydrohelvolic acid, and 5,8-dihydroxy-9,12-octadecadienoic acid (5,8-diHODE). Among these compounds, fumagillin, helvolic acid, and 1,2-dihydrohelvolic acid of A. fumigatus showed antifungal activities against Malassezia furfur, which is lipophilic yeast that causes epidermal skin disorders.     

Histopathological and parasitological study of the gastrointestinal tract of dogs naturally infected with Leishmania infantum

Background

A nationwide survey on the microbial etiology of cases of subclinical mastitis in dairy cows was carried out on dairy farms in Sweden. The aim was to investigate the microbial panorama and the occurrence of antimicrobial resistance. Moreover, differences between newly infected cows and chronically infected cows were investigated.

Methods

In total, 583 quarter milk samples were collected from 583 dairy cows at 226 dairy farms from February 2008 to February 2009. The quarter milk samples were bacteriological investigated and scored using the California Mastitis Test. Staphylococci were tested for betalactamase production and presence of resistance was evaluated in all specific udder pathogens. Differences between newly infected cows and chronically infected cows were statistically investigated using logistic regression analysis.

Results

The most common isolates of 590 bacteriological diagnoses were Staphylococcus (S) aureus (19%) and coagulase-negative staphylococci (CNS; 16%) followed by Streptococcus (Str) dysgalactiae (9%), Str. uberis (8%), Escherichia (E.) coli (2.9%), and Streptococcus spp. (1.9%). Samples with no growth or contamination constituted 22% and 18% of the diagnoses, respectively. The distribution of the most commonly isolated bacteria considering only bacteriological positive samples were: S. aureus - 31%, CNS - 27%, Str. dysgalactiae - 15%, Str. uberis - 14%, E. coli - 4.8%, and Streptococcus spp. - 3.1%. There was an increased risk of finding S. aureus, Str. uberis or Str. dysgalactiae in milk samples from chronically infected cows compared to findings in milk samples from newly infected cows. Four percent of the S. aureus isolates and 35% of the CNS isolates were resistant to penicillin G. Overall, resistance to other antimicrobials than penicillin G was uncommon.

Conclusions

Staphylococcus aureus and CNS were the most frequently isolated pathogens and resistance to antimicrobials was rare.     

Generating neuron-like cells from BM-derived mesenchymal stromal cells in vitro

BACKGROUND: The multipotency of stromal cells has been studied extensively. It has been reported that mesenchymal stromal cells (MSC) are capable of differentiating into cells of multilineage. Different methods and reagents have been used to induce the differentiation of MSC. We investigated the efficacy of different growth factors in inducing MSC differentiation into neurons. METHODS: MSC from human BM were isolated and cultured in media supplemented with 10% FBS. These cells were identified and later induced to differentiate into neuron-like cells using different neurotrophic factors. Three different growth factors were used, either alone or in combination: brain-derived neurotrophic factor, epidermal growth factor and neural growth factor. RESULTS: After 10 days of culture, MSC showed neuron-like morphologic changes. Immunostaining showed that these cells expressed markers for neurons (growth-associated protein-43, neuron-specific nuclear protein and neurofilament 200 kDa) and expression of these markers suggested the transition of immature stages to more mature stages of neuron-like cells. DISCUSSION: Our results show that BM-derived MSC can differentiate not only into target cells of mesodermal origin but also neuron-like cells of ectodermal origin. The findings show that a combination of growth factors is more effective in inducing MSC into neuron-like cells.     

Genome sequence of the leaf-colonizing Bacterium Bacillus sp. strain 5B6, isolated from a cherry tree

Plant growth-promoting bacteria colonize various habitats, including the phyllosphere. Here, we present the high-quality draft genome sequence of Bacillus sp. strain 5B6, which was isolated from the leaf of a cherry tree. The 3.9-Mb genome uncovers its potential for understanding the nature of leaf colonization as well as antibiosis against plant pathogens.     

Comparison of Glial Activation in the Hippocampal CA1 Region Between The Young and Adult Gerbils After Transient Cerebral Ischemia

It has been reported that young animals are less vulnerable to brain ischemia. In the present study, we compared gliosis in the hippocampal CA1 region of the young gerbil with those in the adult gerbil induced by 5?min of transient cerebral ischemia by immunohistochemistry and western blot for glial cells. We used male gerbils of postnatal month 1 (PM 1) as the young and PM 6 as the adult. Neuronal death in CA1 pyramidal neurons in the adult gerbil occurred at 4?days posti-schemia; the neuronal death in the young gerbil occurred at 7?days post-ischemia. The findings of glial changes in the young gerbil after ischemic damage were distinctively different from those in the adult gerbil. Glial fibrillary acidic protein-immunoreactive astrocytes, ionized calcium-binding adapter molecule (Iba-1), and isolectin B4-immunoreactive microglia in the ischemic CA1 region were activated much later in the young gerbil than in the adult gerbil. In brief, very less gliosis occurred in the hippocampal CA1 region of the young gerbil than in the adult gerbil after transient cerebral ischemia.     

Suppression of androgen receptor signaling in prostate cancer cells by an inhibitory receptor variant

There is increasing evidence that sensitization of the androgen receptor (AR) signaling pathway contributes to the failure of androgen ablation therapy for prostate cancer, and that direct targeting of the AR may be a useful therapeutic approach. To better understand how AR function could be abrogated in prostate cancer cells, we have developed a series of putative dominant-negative variants of the human AR, containing deletions or mutations in activation functions AF-1, AF-5, and/or AF-2. One construct, AR inhibitor (ARi)-410, containing a deletion of AF-1 and part of AF-5 of the AR, had no intrinsic transactivation activity but inhibited wild-type AR (wtAR) in a ligand-dependent manner by at least 95% when transfected at a 4:1 molar ratio. ARi-410 was an equally potent inhibitor of gain-of-function AR variants. Ectopic expression of ARi-410 inhibited the proliferation of AR-positive LNCaP cells, but not AR-negative PC-3 cells. Whereas ARi-410 also marginally inhibited progesterone receptor activity, this was far less pronounced than the effect on AR (50% vs. 95% maximal inhibition, respectively), and there was no inhibition of either vitamin D or estrogen receptor activity. In the presence of ligand, ARi-410 interacted with wtAR, and both receptors translocated into the nucleus. Whereas the amino-carboxy terminal interaction was not necessary for optimal dominant-negative activity, disruption of dimerization through the ligand binding domain reduced the efficacy of ARi-410. In addition, although inhibition of AR function by ARi-410 was not dependent on DNA binding, the DNA binding domain was required for dominant-negative activity. Taken together, our results suggest that interaction between ARi-410 and the endogenous AR in prostate cancer cells, potentially through the DNA binding and ligand binding domains, results in a functionally significant reduction in AR signaling and AR-dependent cell growth.     

Genome Sequences of Escherichia coli B strains REL606 and BL21(DE3)

Escherichia coli K-12 and B have been the subjects of classical experiments from which much of our understanding of molecular genetics has emerged. We present here complete genome sequences of two E. coli B strains, REL606, used in a long-term evolution experiment, and BL21(DE3), widely used to express recombinant proteins. The two genomes differ in length by 72,304 bp and have 426 single base pair differences, a seemingly large difference for laboratory strains having a common ancestor within the last 67 years. Transpositions by IS1 and IS150 have occurred in both lineages. Integration of the DE3 prophage in BL21(DE3) apparently displaced a defective prophage in the λ attachment site of B. As might have been anticipated from the many genetic and biochemical experiments comparing B and K-12 over the years, the B genomes are similar in size and organization to the genome of E. coli K-12 MG1655 and have > 99% sequence identity over ∼ 92% of their genomes. E. coli B and K-12 differ considerably in distribution of IS elements and in location and composition of larger mobile elements. An unexpected difference is the absence of a large cluster of flagella genes in B, due to a 41 kbp IS1-mediated deletion. Gene clusters that specify the LPS core, O antigen, and restriction enzymes differ substantially, presumably because of horizontal transfer. Comparative analysis of 32 independently isolated E. coli and Shigella genomes, both commensals and pathogenic strains, identifies a minimal set of genes in common plus many strain-specific genes that constitute a large E. coli pan-genome.