<Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> Cystic Fibrosis isolates of similar RAPD genotype exhibit diversity in biofilm forming ability <Emphasis Type="Italic">in vitro</Emphasis>

Background  

Pseudomonas aeruginosa is considered to grow in a biofilm in cystic fibrosis (CF) chronic lung infections. Bacterial cell motility is one of the main factors that have been connected with P. aeruginosa adherence to both biotic and abiotic surfaces. In this investigation, we employed molecular and microscopic methods to determine the presence or absence of motility in P. aeruginosa CF isolates, and statistically correlated this with their biofilm forming ability in vitro.     

Optimization and SAR for dual ErbB-1/ErbB-2 tyrosine kinase inhibition in the 6-furanylquinazoline series

Synthetic modifications on a 6-furanylquinazoline scaffold to optimize the dual ErbB-1/ErbB-2 tyrosine kinase inhibition afforded consistent SAR whereby a 4-(3-fluorobenzyloxy)-3-haloanilino provided the best enzyme potency and cellular selectivity. Changes made to the 6-furanyl group had little impact on the enzyme activity, but appeared to dramatically affect the cellular efficacy. The discovery of lapatinib emerged from this work.     

Culturable Bacterial Flora Associated with the Dinoflagellate Green Noctiluca miliaris During Active and Declining Bloom Phases in the Northern Arabian Sea

A massive algal bloom of the dinoflagellate Noctiluca miliaris (green) was located in the Northern Arabian Sea by IRS-P4-2 (OCM-II) for microbiological studies, during two consecutive cruises of February-March 2009. Culturable bacterial load during bloom were ～2–3-fold higher in comparison to non-bloom waters and ranged from 3.20?×?10⁵ to 6.84?×?10⁵?cfu?ml^?1. An analysis of the dominant heterotrophs associated with Noctiluca bloom resulted in phylogenetic and a detailed metabolic characterization of 70 bacterial isolates from an overlapping active and declining bloom phase location near north-central Arabian Sea. The active phase flora was dominated by Gram-positive forms (70.59 %), a majority of which belonged to Bacillus (35.29 %) of Firmicutes. As the bloom declined, Gram-negative forms (61.11 %) emerged dominant, and these belonged to a diverse γ-proteobacterial population consisting of Shewanella (16.67 %) and equal fractions of a Cobetia–Pseudomonas-Psychrobacter–Halomonas population (36.11 %). A Unifrac-based principal coordinate analysis of partial 16S rDNA sequences showed significant differences among the active and declining phase flora and also with reported endocytic flora of Noctiluca (red). A nonparametric multidimensional scaling (NMDS) of antibiogram helped differentiation among closely related strains. The organic matter synthesized by N. miliaris appears to be quickly utilized and remineralized as seen from the high efficiency of isolates to metabolize various complex and simple C/N substrates such as carbohydrates, proteins/amino acids, lipids, sulfide production from organic matter, and solubilize phosphates. The ability of a large fraction of these strains (50–41.67 %) to further aerobically denitrify indicates their potential for nitrogen removal from these high-organic microniches of the Noctiluca bloom in the Arabian Sea, also known for high denitrification activity. The results indicate that culturable euphotic bacterial associates of Noctiluca are likely to play a critical role in the biogeochemical ramifications of these unique seasonally emerging tropical open-water blooms of the Northern Arabian Sea.     

A Failure Reveals Success

Although environmental education and education for sustainable development have become well‐established areas of scholarship and practice, there has not been a similar development focused on “industrial ecology education.” A review of the historical context and guiding philosophies for each of these areas finds many similarities, as well as key differences. Environmental education traces its modern roots to the idealism of the 1960s and 1970s. It has focused mostly on improving environmental conditions. Education for sustainable development arose along with international concerns about social justice. It has emphasized general education as well as education about sustainability as necessary to ensure human prosperity. Industrial ecology, in its contemporary form, evolved as an applied approach to address environmental concerns and to meet sustainability goals. It has developed into a diverse, multifaceted approach to address the complexity inherent in industrial society. Education focused on industrial ecology remains decentralized, with core principles and tools being integrated into existing disciplinary programs as well as development of industrial‐ecology–specific curricula. These efforts have not coalesced into a formalized, industrial ecology education. Rather than reflecting a shortcoming, this potentially offers a more robust method for applying industrial ecology principles and tools widely.     

A simple and rapid microwave-assisted hematoxylin and eosin staining method using 1,1,1 trichloroethane as a dewaxing and a clearing agent

The use and practicability of microwave-assisted staining procedures in routine histopathology has been well established for more than 17 years. In the study reported here, we aimed to examine an alternative approach that would shorten the duration of dewaxing and clearing steps of hematoxylin and eosin (H & E) staining of paraffin sections by using a microwave oven. Although xylene is one of the most popular dewaxing and clearing agents, its flammability restricts its use in a microwave oven; thus we preferred 1,1,1 trichloroethane, which is not flammable, as the dewaxing and clearing agent in the present study. In Group I and Group II (control groups), intestine was processed with xylene and 1,1,1 trichloroethane, respectively. The sections were then stained with H & E according to the conventional staining protocol at room temperature and subdivided into two groups according to the duration of dewaxing and clearing in xylene. In Groups III and IV (experimental groups) similar tissues were processed with xylene and 1,1,1 trichloroethane, respectively; however, sections from these groups were divided into four subgroups to study the period required for dewaxing and clearing in 1,1,1 trichloroethane, then stained with H & E in the microwave oven at 360 W for 30 sec. Our conventional H & E staining procedure, which includes dewaxing, staining and clearing of sections, requires approximately 90 min, while our method using 1,1,1 trichloroethane and microwave heating required only 2 min. Our alternative method for H & E staining not only reduced the procedure time significantly, but also yielded staining quality equal or superior to those stained the conventional way. Our results suggest that 1,1,1 trichloroethane can be used effectively and safely as a dewaxing and clearing agent for H & E staining in a microwave oven.     

Multivalent poly(ethylene glycol)-containing conjugates for in vivo antibody suppression

Poly(ethylene glycol) (PEG) was incorporated into multivalent conjugates of the N-terminal domain of beta(2)GPI (domain 1). PEG was incorporated to reduce the rate of elimination of the conjugates from plasma and to putatively improve their efficacy as toleragens for the suppression of anti-beta(2)GPI antibodies and the treatment of antiphospholipid syndrome (APS). Three structurally distinct types of multivalent platforms were constructed by incorporating PEG into the platform structures in different ways. The amount of PEG incorporated ranged from about 5000 g per mole to about 30000 g per mole. The platforms were functionalized with either four or eight aminooxy groups. The conjugates were prepared by forming oxime linkages between the aminooxy groups and N-terminally glyoxylated domain 1 polypeptide. The plasma half-life of each conjugate, labeled with (125)I, was measured in both mice and rats. The half-lives of the conjugates ranged from less than 10 min to about 1 h in mice, and from less than 3 h to about 19 h in rats. The ability of five tetravalent conjugates to suppress anti-domain 1 antibodies in immunized rats was also measured. Incorporation of PEG in the conjugates significantly reduced the doses required for suppression, and the amount of reduction correlated with the amount of PEG incorporated.     

Purification and characterization of Mycobacterium tuberculosis KatG, KatG(S315T), and Mycobacterium bovis KatG(R463L)

Isoniazid, a first-line antibiotic used for the treatment of tuberculosis, is a prodrug that requires activation by the Mycobacterium tuberculosis enzyme KatG. The KatG(S315T) mutation causes isoniazid resistance while the KatG(R463L) variation is thought to be a polymorphism. Much of the work to date focused on isoniazid activation by KatG has utilized recombinant enzyme overexpressed in Escherichia coli. In this work, native KatG and KatG(S315T) were purified from M. tuberculosis, and KatG(R463L) was purified from Mycobacterium bovis. The native molecular weight, enzymatic activity, optical, resonance Raman, and EPR spectra, K(D) for isoniazid binding, and isoniazid oxidation rates were measured and compared for each native enzyme. Further, the properties of the native enzymes were compared and contrasted with those reported for recombinant KatG, KatG(S315T), and KatG(R463L) in order to assess the ability of the recombinant enzymes to act as good models for the native enzymes.     

The effect of salt extraction on the structure of transcriptionally active genes; evidence for a DNAseI-sensitive structure which could be dependent on chromatin structure at levels higher than the 30 nm fibre.

The procedure developed by Lawson and Cole (Biochemistry, 1979, 18 2161-2166) for removing lysine-rich histones from nuclei at low pH also quantitatively extracts proteins HMG14 and 17. The effect of this low pH extraction on the DNAseI-sensitive structures of active genes in avian red blood cells has been investigated. No major perturbation of a developmentally regulated DNAseI hypersensitive site in the beta-globin domain and at the 5' end of the alpha D gene was seen. The overall DNAseI-sensitive conformation of the beta A-globin gene (relative to the ovalbumin gene) is minimally affected by pH3 salt extraction, but there is some loss of sensitivity of the alpha D gene. Removal of HMG proteins at neutral pH had no effect on the sensitivity of active genes in erythroid or fibroblast nuclei. These results, together with those carried out on DNAseI sensitivity and HMG binding to monomer nucleosomes, indicate that there is a major structural feature of active genes responsible for DNAseI-sensitivity which is independent of HMG proteins or nucleosome core particle structure but may be dependent on higher order chromatin structures.