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PubMed2005

Molecular characterization of Agrotis segetum nucleopolyhedrovirus from Poland.

The turnip moth, Agrotis segetum (Lepidoptera, Noctuidae), is an important pest insect in Europe, Asia, and Africa. We have genetically characterized and classified a nucleopolyhedrovirus isolated from A. segetum larvae in Poland (AgseNPV-P). The restriction pattern of AgseNPV-P was distinct from an isolate from England/France (AgseNPV-UK and AgseNPV-F). Sequence analysis of three conserved baculovirus genes, polyhedrin, lef-8 and pif-2, revealed that AgseNPV-P differs substantially from the already described NPVs isolated from A. segetum and possibly represents a new NPV species. Phylogenetic analysis placed AgseNPV-P among group II NPVs and showed the closest relationship to Agrotis ipsilon (Agip) NPV and Spodoptera exigua (Se) MNPV.

Animals↗
PubMed1992

A discussion of the U.S. EPA methodology for determining Water Quality Standards (WQS).

Based on material published by the U.S. Environmental Protection Agency (U.S. EPA) in the Federal Register for 19 November 1991, many state environmental agencies have proposed and/or adopted revisions to their State Water Quality Standards (WQS) for organic and inorganic chemicals in fresh and marine waters (see, for example, State of Connecticut, Department of Environmental Protection, Bureau of Water Management, (1992), memorandum to Interested Parties concerning the Water Quality Standards Hearing Report). Generally, many states simply republish the U.S. EPA's proposed Water Quality Criteria (WQC) as the State's proposed WQS. Many of the state WQS and federal WQC values--especially those for organic compounds regulated as human or animal carcinogens--are much more stringent than the values now in effect because the U.S. EPA's new methodology (i) for estimating exposure point concentrations, exposure doses, carcinogenic potency, and incremental lifetime cancer risk and (ii) for setting the target acceptable risk combine a series of conservative assumptions into an equally conservative set of results. In the Federal Register proposal, the U.S. EPA failed to honor its standard risk assessment methodology in that (i) it failed to perform a quantitative or even qualitative uncertainty analysis and (ii) it failed to analyze the overall degree of conservatism in the results. The U.S. EPA suggested that the analysis is suitably conservative for the average exposed adult, but it failed to consider various phenomena that make the proposed WQC far more conservative than acknowledged or intended. To focus on a central problem of manageable size, this article dissects the method by which the U.S. EPA calculates proposed WQC for organic chemicals regulated as human or animal carcinogens. Because the results for most such chemicals are driven by the pathway for the human ingestion of fish which have bioconcentrated the chemicals from the water column (as opposed to the pathway for direct ingestion of water by humans), this article focuses exclusively on the fish-to-human pathway. These considerations form the basis of general quality assurance criteria and standards.

Animals↗
PubMed2001

Understanding clonal diversity patterns through allozyme polymorphism in an endangered and geographically restricted Australian shrub, Zieria baeuerlenii and its implications for conservation.

An electrophoretic genetic analysis utilising starch gel electrophoresis was employed to assess clonality in endangered Zieria baeuerlenii populations distributed over an area of less than a kilometre square in Nowra (NSW). Eleven enzymes systems encoded by 19 loci (41 alleles) when assayed to estimate levels of genetic diversity within and among populations, revealed moderate levels of genetic diversity. Despite finding seven loci being fixed at a particular locus, 20 unique multilocus genotypes/clones restricted to a particular population were detected within a sample of 179 ramets collected from throughout the range of the species. The probability of finding any clone produced by sexual reproduction is <0.0025 and is further supported by the fact that reproduction appears to be exclusively by vegetative spread and there is virtually no pollen viability and seed production. The lack of seed set in this area may be due to additional factors inhibiting sexual reproduction. Overall, such genetic studies play a crucial role in devising conservation and management strategies for rare and endangered taxa.

Journal Article↗
PubMed1992

An analysis of prognostic factors in response to conservative treatment of postmastectomy lymphedema.

Postmastectomy lymphedema of the upper limb is usually managed conservatively. Patients are often advised to use elastic bandages or sleeves during the day: elastic compression is, in fact, generally considered the mainstay of lymphedema therapy by authors who advocate the addition of cyclical instrumental or physical treatments, such as pneumatic compression or manual lymphatic drainage. The results of elastic compression, however, have never been extensively analyzed. In the present study, we describe the patterns of response during six months of use of a standard elastic sleeve in a series of 120 patients with postmastectomy lymphedema. Univariate and multivariate analyses were performed to identify possible prognostic factors for a better or worse response. Overall, limb measurements were reduced by 14.7 percent, with weight gain after surgical treatment resulting as the only significant independent negative factor: patients who did not increase weight after mastectomy achieved nearly a 25 percent reduction of lymphedema. These results should be considered during the selection of patients for protocols of lymphedema treatment and the analysis of results.

Aged↗
PubMed2005

[Role of surgery in spinal degenerative disease. Analysis of systematic reviews on surgical and conservative treatments from an evidence-based approach].

INTRODUCTION: The lifetime prevalence of invalidating back pain in general population caused by Spinal Degenerative Disease (SDD) is about 70-80%. Global costs related to this disease are enormous (1-2% gross domestic product). From an Evidence-based point of view, there is a striking discrepancy between the use of many available surgical techniques (especially for spinal fusion) and the lack of scientific support. METHODS: The authors carefully reviewed all published metaanalysis on SDD therapies up to December 2003. Treatment recommendations were classified according to levels of evidence (strong, moderate, mild or lack of evidence) for both surgical and conservative measures. RESULTS: Forty-four metaanalysis were selected (nine on lumbar surgery, three on cervical surgery and thirty-two on other therapies). Relating surgery, there is strong evidence favouring early laminectomy in cauda equina syndrome secondary to lumbar disc herniation; discectomy or microdiscectomy are superior to chemo-nucleolysis in lumbar prolapse and spondylosis; and fusion surgery (probably noninstrumented) in adult isthmic spondylolysthesis or degenerative spondylolysthesis with spinal stenosis. In cervical spondylosis and radiculomyelopathy, discectomy seems as effective as discectomy plus fusion, which does not seem to be better than untreated SDD beyond 24 months. Preoperative antibiotics seem to prevent infection in spinal surgery. No benefit of surgery is demonstrated in discogenic pain. None of conservative therapies are supported by strong evidence. Antidepressants improve pain perception but do not influence the functional status. DISCUSSION: Although lumbar instrumented surgery has nearly doubled over two decades and the annual growth is about 20%, clinical results do not seem to have improved, not even global fusion rates. The increasing use of fusion surgery for cases other than spinal deformities, spondylolysthesis or spinal stenosis plus lysthesis may be related to multiple technical and clinical-epidemiological factors where huge financial and commercial interests must be considered. It is crucial to differentiate subsets of patients prone to benefit from surgery. It is discussed whether randomized trials incorporating sham operations are ethically justifiable, because of the lack of sound evidence for many spinal procedures. The efficacy of most conservative treatments is mild or moderate (mainly transient) and they should be probably used in combination. CONCLUSIONS. There is no strong evidence favouring most of surgical procedures for SDD from an evidence-based approach. It seems necessary that scientific organizations studying SDD create clinical guidelines relating its multidisciplinary and integral management, recognizing that, up to now, few interventions positively modify in the long-term the natural history of the disease.

Diskectomy↗
PubMed2005

Comparative analysis of HIV-1 Tat variants.

HIV-1 Tat protein is a crucial element for viral replication; therefore, its inhibition might be exploited against the AIDS infection. To gain insights on the natural variability of this protein, we present a comparative investigation on the relationship between the primary sequences and the experimentally available three-dimensional structures from the HIV-1 Tat variants Z2, BRU, and MAL. Our computational tools include sequence conservation algorithms, structural analysis, electrostatic modeling, and molecular dynamics (MD) simulations. We find that two regions located between residues 10-18 and 41-52 display the highest primary sequence conservation, while the most conserved region among the available structures corresponds approximately to the segment between positions approximately 44 and 50. Furthermore, in spite of their large structural divergence, Tat variants share a common mode for long-range intramolecular interactions. Finally, the flexibility of the Z2, BRU, and MAL variants, as emerging from multinanosecond MD simulations, is rather similar. Based on this work, we conclude that the turnlike region between amino acids 44 and 50 is structurally most conserved, emerging as an important motif for pharmaceutical targeting aimed toward inhibiting Tat action.

Algorithms↗
PubMed1997

Evolutionary conserved cathepsin E substrate specificity as defined by N-terminal and C-terminal sequencing of peptide pools.

The substrate specificity of the non-lysosomal aspartic protease cathepsin E from three different species has been studied using the method of automated N-terminal sequencing and a newly developed method for C-terminal sequencing of peptides and peptide pools. The combination of N-terminal and C-terminal sequencing of peptide pools is a fast and easy method to identify and compare the substrate specificity of endopeptidases. Our analysis shows a conserved hydrolytic specificity between human, mouse and bovine cathepsin E, with only small differences in fine specificity. Furthermore, our results confirm and extend the rules governing the interactions of the substrate with the amino acid (aa) side chains of the various pockets within the enzyme's active cleft. We found that the positions flanking the scissile peptide bond P1-P1' are occupied exclusively by hydrophobic aa with both aliphatic or aromatic side chains; Val and Ile, however, are not allowed in the S1 binding site. The S2 and S2' subsites accept hydrophilic aa. Additional requirements concerning the S3' to S5' subsites were also revealed. Finally, the sequences of single peptides generated by cathepsin E from the three different species can be easily aligned to the determined cleavage motif, showing the reliability of our pool sequencing methods.

Amino Acid Sequence↗
PubMed2006

The Rhizobium etli sigma70 (SigA) factor recognizes a lax consensus promoter.

A collection of Rhizobium etli promoters was isolated from a genomic DNA library constructed in the promoter-trap vector pBBMCS53, by their ability to drive the expression of a gusA reporter gene. Thirty-seven clones were selected, and their transcriptional start-sites were determined. The upstream sequence of these 37 start-sites, and the sequences of seven previously identified promoters were compared. On the basis of sequence conservation and mutational analysis, a consensus sequence CTTGACN16-23TATNNT was obtained. In this consensus sequence, nine on of twelve bases are identical to the canonical Escherichia coli sigma70 promoter, however the R.etli promoters only contain 6.4 conserved bases on average. We show that the R.etli sigma factor SigA recognizes all R.etli promoters studied in this work, and that E.coli RpoD is incapable of recognizing them. The comparison of the predicted structure of SigA with the known structure of RpoD indicated that regions 2.4 and 4.2, responsible for promoter recognition, are different only by a single amino acid, whereas the region 1 of SigA contains 72 extra residues, suggesting that the differences contained in this region could be related to the lax promoter recognition of SigA.

Bacteria↗
PubMed2004

Comparison of the effectiveness of the conservative treatment of the periodontal pockets with or without the use of laser biostimulation.

The use of laser therapy as the agent reinforcing conventional treatment of the periodontal diseases becomes more and more common. In the physiotherapy of the periodontal diseases the biostimulating, laser is eagerly used because of its action which accelerates the healing of wounds and also because of its antioedematous, anti-inflammatory and analgesic action. The aim of work was the evaluation of the influence of laser biostimulation on the change of the periodontological pockets depth after the routine conservative periodontological treatment with additional use of laser biostimulation and without it for two groups of pockets: above and below 5 mm. In six patients having periodontitis 613 sites were submitted to the statistic analysis (290 treated conservatively only, including 251 with the depth 2-5 mm and 39 above 5 mm as well as 323 with the use of laser therapy including 297 shallow pockets and 26 deep ones). The initial values of API, SBI, PPD and their changes in the course of the treatment were registered. During each control appointment the patients subjectively estimated periodontal pain occurrence. In both studied groups statistically essential decrease of the evaluated parameters was obtained. Reinforcing the conventional treatment with laser biostimulation shortens its duration and leads to the elimination of pain faster than with the use of conservative treatment only. The changes of the PPD index among the successive examinations were statistically essentially higher in the therapy with the use of laser, especially in relation to deep pockets.

Combined Modality Therapy↗
PubMed2003

Characterization of the last subunit of the Arabidopsis COP9 signalosome: implications for the overall structure and origin of the complex.

The COP9 signalosome (CSN) is an evolutionarily conserved protein complex that resembles the lid subcomplex of proteasomes. Through its ability to regulate specific proteasome-mediated protein degradation events, CSN controls multiple aspects of development. Here, we report the cloning and characterization of AtCSN2, the last uncharacterized CSN subunit from Arabidopsis. We show that the AtCSN2 gene corresponds to the previously identified FUS12 locus and that AtCSN2 copurifies with CSN, confirming that AtCSN2 is an integral component of CSN. AtCSN2 is not only able to interact with the SCF(TIR1) subunit AtCUL1, which is partially responsible for the regulatory interaction between CSN and SCF(TIR1), but also interacts with AtCUL3, suggesting that CSN is able to regulate the activity of other cullin-based E3 ligases through conserved interactions. Phylogenetic analysis indicated that the duplication and subsequent divergence events that led to the genes that encode CSN and lid subunits occurred before the divergence of unicellular and multicellular eukaryotic organisms and that the CSN subunits were more conserved than the lid subunits during evolution. Comparative analyses of the subunit interaction of CSN revealed a set of conserved subunit contacts and resulted in a model of CSN subunit topology, some aspects of which were substantiated by in vivo cross-link tests.

Amino Acid Sequence↗
PubMed1997

[The conservative and surgical therapy of traumatic humeral shaft fractures].

Sixty-three patients with humeral shaft fractures were evaluated clinically and radiographically 18 months after injury; 27 patients were treated surgically (group A) and 36 patients conservatively (group B). Analysis of the results according to a score by Kwasny revealed 6.2 points in group A and 2.2 points in group B (P < 0.0001; F = 46.9). The results of these two comparable groups suggest that conservative treatment of humeral shaft fractures is superior regarding mobility of the shoulder and elbow, strength, the incidence of neurological complications, pain, subjective rating and cosmesis. There were no differences on roentgenograms between the two groups (P = 0.48).

Adolescent↗
PubMed2003

Twelve new MotA-dependent middle promoters of bacteriophage T4: consensus sequence revised.

Bacteriophage T4 middle-mode transcription requires Escherichia coli RNA polymerase, phage-encoded transcriptional activator MotA and co-activator AsiA that form a complex at a middle promoter DNA. T4 middle promoters have been defined by a consensus sequence deduced from the list of 14 middle promoters identified in earlier studies. To date, 33 middle promoters have been mapped on the T4 genome. Of these, 12 contain differences even at the highly conserved positions of the consensus sequence. In the T4 prereplicative gene cluster between genes e and rpbA, we have identified 12 new middle promoters, most of which contain differences from the consensus sequence deduced previously. Analysis of base conservation in the different sequence positions of new middle promoters, as well as those identified previously, revealed some new features of middle T4 promoters. We propose to define these promoters by a MotA box (a/t)(a/t)(a/t)TGCTTtA centred at the position -30, the sequence TAtaAT centred at -10 relative to the transcriptional start site, and the spacer region of 12(+/-1) base-pairs between them.

Bacteriophage T4↗
PubMed1996

Internal initiation of translation of hepatitis C virus RNA: the ribosome entry site is at the authentic initiation codon.

Hepatitis C viral (HCV) RNA includes an internal ribosome entry segment (IRES) that extends some 30 nt into the coding region and promotes internal initiation of translation at the authentic initiation codon at nt 342. The 5'-boundary of this IRES was mapped by in vitro translation and transfection assays and was found to lie between nt 42 and 71. Within these IRES boundaries there are, in most HCV strains, three AUG triplets upstream of the authentic initiation site. Although the first, 5'-proximal, of these is absolutely conserved, a mutational analysis showed that it is not a functional initiation codon. In particular, the G residue could be substituted provided compensatory mutations were made to maintain base pairing. The other two upstream AUGs are not absolutely conserved, and mutation of the third (5'-distal) had little effect on IRES activity. When an additional AUG codon was introduced by single-site mutation just upstream of the authentic initiation codon, it was found to be used when most of the IRES had been deleted to generate an RNA translated by the scanning ribosome mechanism, but was not used in the background of the full-length IRES when internal initiation is operative. These results argue that the IRES promotes direct ribosome entry immediately at, or indeed very close to, the authentic initiation codon, and that the upstream AUGs do not serve as ribosome entry sites.

Base Sequence↗
PubMed1996

Conservation of aconitase residues revealed by multiple sequence analysis. Implications for structure/function relationships.

Aconitases have recently regained much attention, because one member of this family, iron regulatory protein-1 (IRP-1), has been found to play a dual role as a cytoplasmic aconitase and a regulatory RNA-binding protein. This finding has highlighted a novel role for Fe-S clusters as post-translational regulatory switches. We have aligned 28 members of the Fe-S isomerase family, identified highly conserved amino acid residues, and integrated this information with data on the crystallographic structure of mammalian mitochondrial aconitase. We propose structural and/or functional roles for the previously unrecognized conserved residues. Our findings illustrate the value of detailed protein sequence analysis when high-resolution crystallographic data are already available.

Aconitate Hydratase↗
PubMed1995

Should the audiometric database analysis method (draft ANSI S12.13-1991) for evaluating the effectiveness of hearing conservation programs be accepted as a US national standard?.

The value of audiometric database analysis (ADBA) procedures for evaluating the effectiveness of hearing conservation programs (HCPs) was studied in a population of 82,195 workers. We rated the HCP using four ADBA procedures. Results showed that the HCP ratings ran the gamut from "unacceptable" to "acceptable" from year to year and from procedure to procedure. A mere 7 percent of the total population was eligible for ADBA analysis. This drastically reduced sample size precludes subgroup analyses of even a large cohort and likely could not be applied to the HCPs of most small- and medium-sized businesses. The low levels of agreement observed (weighted kappa = 15%-25%) between the ADBA procedures reflect poor validity. Generalizability of these results would be inappropriate when so few nonrandomly selected individuals contribute to the evaluation. In view of these problems, acceptance of the ADBA method as a national standard may not be warranted.

Audiology↗
PubMed2005

Dimerization and enzymatic activity of fungal 17beta-hydroxysteroid dehydrogenase from the short-chain dehydrogenase/reductase superfamily.

BACKGROUND: 17beta-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17beta-HSDcl) is a member of the short-chain dehydrogenase/reductase (SDR) superfamily. SDR proteins usually function as dimers or tetramers and 17beta-HSDcl is also a homodimer under native conditions. RESULTS: We have investigated here which secondary structure elements are involved in the dimerization of 17beta-HSDcl and examined the importance of dimerization for the enzyme activity. Sequence similarity with trihydroxynaphthalene reductase from Magnaporthe grisea indicated that Arg129 and His111 from the alphaE-helices interact with the Asp121, Glu117 and Asp187 residues from the alphaE and alphaF-helices of the neighbouring subunit. The Arg129Asp and His111Leu mutations both rendered 17beta-HSDcl monomeric, while the mutant 17beta-HSDcl-His111Ala was dimeric. Circular dichroism spectroscopy analysis confirmed the conservation of the secondary structure in both monomers. The three mutant proteins all bound coenzyme, as shown by fluorescence quenching in the presence of NADP+, but both monomers showed no enzymatic activity. CONCLUSION: We have shown by site-directed mutagenesis and structure/function analysis that 17beta-HSDcl dimerization involves the alphaE and alphaF helices of both subunits. Neighbouring subunits are connected through hydrophobic interactions, H-bonds and salt bridges involving amino acid residues His111 and Arg129. Since the substitutions of these two amino acid residues lead to inactive monomers with conserved secondary structure, we suggest dimerization is a prerequisite for catalysis. A detailed understanding of this dimerization could lead to the development of compounds that will specifically prevent dimerization, thereby serving as a new type of inhibitor.

17-Hydroxysteroid Dehydrogenases↗
PubMed2006

VISTA family of computational tools for comparative analysis of DNA sequences and whole genomes.

Comparative analysis of DNA sequences is becoming one of the major methods for discovery of functionally important genomic intervals. Presented here the VISTA family of computational tools was built to help researchers in this undertaking. These tools allow the researcher to align DNA sequences, quickly visualize conservation levels between them, identify highly conserved regions, and analyze sequences of interest through one of the following approaches: . Browse precomputed whole-genome alignments of vertebrates and other groups of organisms. . Submit sequences to Genome VISTA to align them to whole genomes. . Submit two or more sequences to mVISTA to align them with each other (a variety of alignment programs with several distinct capabilities are made available).. Submit sequences to Regulatory VISTA (rVISTA) to perform transcription factor binding site predictions based on conservation within sequence alignments.Use stand-alone alignment and visualization programs to run comparative sequence analysis locally All VISTA tools use standard algorithms for visualization and conservation analysis to make comparison of results from different programs more straightforward. The web page http://genome.lbl.gov/vista/ serves as a portal for access to all VISTA tools. Our support group can be reached by email at vista@lbl.gov.

Algorithms↗
PubMed2005

Characterization of LST-R533: uncovering a novel repetitive element in Leishmania.

We have previously isolated and sequenced a novel repetitive element, now named LST-R533, which is present in four different regions of one extremity of Leishmania major chromosome 20. The repeats are polymorphic in size, ranging from 367 to 533 bp and contain an internal 81 bp sequence with highly conserved segments (14-81 bp long) dispersed throughout the parasite's genome. These sequences were not found in coding regions of any predicted gene in L. major Friedlin genome, but are part of untranslated regions of some Leishmania transcripts. Analysis of the 81 bp sequence revealed significant degrees of identity with retrotransposons described in several other organisms. The presence of the sequence in other species from genus Leishmania was determined by Southern hybridisation and DNA sequencing. This analysis indicated the conservation of the 81-nucleotide element in all the Leishmania species evaluated. No sequences corresponding to LST-R533 or the 81 bp element were found on either Trypanosoma brucei or Trypanosoma cruzi databanks.

Animals↗