bluuebunny/crossref_metadata_2025 huggingface dataset:
Increased Use of Pneumococcal Vaccination in a Medicine Clinic
Following Initiation of a Quality Assessment Monitor
Authors: Tobacman Joanne K.
Published: September 2009

Obectives:

To determine the use of pneumococcal vaccination in a general medicine
ambulatory clinic and the effect of a quality assessment monitor on
use.

Design:

A prospective 8-month (October 1990 through May 1991) survey and 8-month
(October 1989 through May 1990) retrospective survey of use of
pneumococcal vaccination in the clinic.

Setting:

A general medicine clinic where patients are seen for their first
internal medicine clinic visit at The University of Iowa Hospitals and
Clinics, Iowa City, Iowa.

Patients:

One thousand sixty adult patients who presented to the clinic during the
study interval.

Interventions:

The use of the form constituted an intervention.

Results:

Use of pneumococcal vaccination was increased more than 5 times over the
baseline level following implementation of the quality assessment
monitor.

Conclusions:

Underuse of pneumococcal vaccination occurred in this general medicine
clinic, and the introduction of a quality assessment monitor constituted
an intervention that significantly increased the use of pneumococcal
vaccination.
Clustered Bacteremias in a Hemodialysis Unit Cross-Contamination
of Blood Tubing From Ultrafiltrate Waste
Authors: Longfield Robert N. , Wortham William G. , Fletcher Linda L. , Nauscheutz William F.
Published: September 2009

Objectve:

To determine the cause of clustered bacteremias occurring among chronic
hemodialysis patients.

Design:

A retrospective investigation of clinical and laboratory records with
direct observation of dialysis facilities and technique. Bacterial blood
isolates were identified and compared with environmental isolates.

Setting:

The 1 1-station chronic hemodialysis unit that serves approximately 50
patients in a 450-bed military hospital.

Patients:

Hemodialysis unit patients with aerobic gram-negative bacillus or
Enterococcus casseliflavus blood isolates between
April 1988 and February 1990.

Results:

The recovery and species identification of the unique isolate, E
casseliflavus, from 2 index cases of bacteremia in
February 1990 helped identify the cluster and demonstrated its protracted
course. Dialysis blood tubing was contaminated with ultrafiltrate waste
during dialyzer setup.

Intervention:

Bacteremias were controlled by halting the practice of attaching the
venous tubing directly to a waste container while priming the membrane,
by emphasizing glove changes and handwashing after contact with
ultrafiltrate waste and by daily decontamination of ultrafiltrate waste
bags.

Conclusions:

We recommend that other hemodialysis units institute these
interventions.
Continuous Quality Improvement in a Hospital System: Implications for Hospital Epidemiology
Authors: Decker Michael D. , Scheckler William E.
Published: September 2009

The purpose of this report is to describe the “Continuous Quality Improvement” (CQI) paradigm as adopted by one of the 30 largest hospital systems in the United States and to explore the implications for hospital epidemiology and infection control. Hospital epidemiology has its roots in the application of epidemiologic tools and principles to the problems of nosocomial infections. Key steps in the development of hospital epidemiology came from physicians in Great Britain and the United States who were part of the public health systems of those countries. In the United States, physicians trained in infectious diseases as a subspecialty occupy the position of hospital epidemiologist in most university, Veterans Affairs, and larger community teaching hospitals. Some of these individuals argue that hospital epidemiologists should continue to focus principally on infection control. Others are just as convinced that the premises and knowledge of epidemiology honed by experiences in infection control are very well suited to many other problems facing hospitals in the 1990s.
Needlestick Prevention Devices: A Pointed Discussion
Authors: Crow Sue , Wugofski Lee
Published: September 2009

Institutions and employees in the healthcare industry recently have been flooded with a variety of new preventive devices, with the promise of reducing the risk of needlestick accidents. With the known risk of occupational human immunodeficiency virus (HIV) infection and the greater risk of morbidity and mortality of occupational hepatitis B virus (HBV) infection, it is hard not to get swept away by the tsunami of “safer devices.” The ultimate question of course, is whether these new or improved products actually contribute to the reduction of accidents and diminish the transmission of these diseases. With the demand for safer working conditions in the context of the HIV epidemic and in times of shrinking budgets, the safest and most cost-effective devices must be determined. Although the Occupational Safety and Health Administrations (OSHA) final ruling on occupational exposure to bloodborne pathogens may not directly address the issue of needlestick prevention devices, institutions should be prepared to justify their practices and selection of equipment, given its emphasis on engineering and work practices that states “Engineering and work practice controls shall be used to eliminate or minimize employee exposure.”
Nosocomial Human Parovirus B19 Infection: Lack of Transmission From a Chronically Infected Patient to Hospital Staff
Authors: Koziol Deloris E. , Kurtzrnan Gary , Ayub Jamshed , Young Neal S. , Henderson David K.
Published: September 2009

Objective:

To assess the potential for nosocomial spread of parvovirus B19 from a chronically infected patient.

Design:

Employees exposed to the index case and control (unexposed) employees were evaluated by baseline and follow up parvovirus B19 serologies and hematologic assessments, and completed baseline and follow up epidemiologic questionnaires.

Setting:

A chronically infected patient was hospitalized on a hematology ward in a research referral hospital for 3.5 weeks prior to a diagnosis of parvovirus B19 infection and the institution of isolation precautions.

Methods:

Sera were screened for parvovirus B19 DNA (dot blot analysis), and IgG and IgM anti-B19 antibodies (capture immunoassay). Hematologic assessment included CBC, differential, and reticulocyte count.

Results:

The index case had parvovirus B19 DNA at approximately lo6 genome copies per ml of serum, elevated IgM and low levels of IgG B19 antibodies. Of the 21 exposed staff, 11 (52%) had IgG B19 antibodies and were immune; of the 8 unexposed staff, 6 (75%) had IgG B19 antibodies. No employees developed IgM B19 antibodies, B19 DNA, hematologic abnormalities, or clinical symptoms.

Conclusions:

In contrast to reports of documented nosocomial transmission of B19 parvovirus from patients in transient aplastic crisis, nosocomial transmission did not occur-even in the absence of isolation precautions-presumably from the lower level of B19 viremia in our chronically infected (rather than acutely infected) patient.
SHEA's Initiative for Confronting the Cost-Quality Conundrum
Authors: Massanari R. Michael , Simmons Bryan ,   The Society for Hospital Epidemiology of America
Published: September 2009

The US healthcare system is being buffeted by winds of change. Once viewed as a model of excellence, economic and social disequilibrium have uncovered a multitude of frailties within the system. Spiraling healthcare costs in an otherwise stagnant economy have given rise to skepticism regarding the motivations of healthcare providers. What are the issues that confront the healthcare system? What new demands do these issues place on the delivery system? How should an organization such as SHEA position itself to meet these new demands?

Three major issues will confront the healthcare system during the next decade. First, 10% to 14% of the US population have limited or no access to healthcare. Steps must be taken through the political process to assure that every person living in the United States has access to a reasonable standard of medical care. Second, despite implicit rationing, the costs of healthcare continue to consume an ever-increasing proportion of society's resources. In response, a potpourri of strategies have been implemented to contain increasing costs. Third, as society learns more about processes and outcomes of medical care, concern for quality intensifies. Efforts to constrain costs are juxtaposed with demands to assure excellence and quality in medical care. During the decade of the 90s the critical questions will be how much medical care should be provided to whom and at what level of quality? If society insists that medical care should be provided to all who live in the United States but refuses to increase present resources, difficult decisions will have to be made regarding how to reallocate those resources and how to minimize the impact of reallocation on quality of care.
Azithromycin and Clarithromycin Overview and Comparison With Erythromycin
Authors: Scheld W. Michael , Whitman Marc S. , Tunkel Allan R.
Published: September 2009

Macrolides currently account for 10% to 15% of the worldwide oral antibiotic market.' Erythromycin, the first macrolide antibiotic, was discovered in 1952 from a strain of Streptomyces erythreus obtained from soil samples in the Phillipines. Originally, erythromycin was marketed as an alternative to penicillin because of its activity against gram-positive organisms such as staphylococci, pneumococci, and streptococci. Subsequently, its clinical use broadened to include species of Mycoplasma, Legionella, Campylobacter, and Chlamydia. Although several other macrolides have been marketed in countries other than the United States, they have failed to achieve erythromycin's widespread use. Unfortunately, erythromycin suffers from several drawbacks, including gastrointestinal side effects, a short serum elimination half-life, and only borderline in vitro activity against common gram-negative respiratory pathogens such as Haemophilus influenzae.
Nonincreased Risk of Nosocomial Infection During a 22-Day Housekeeping Personnel Strike in a Tertiary Hospital
Authors: Delgado-Rodríquez Miguel , Ramos-Cuadra Amalia , Bueno-Cavanillas Aurora , Jimenez-Romano Elena , Guillen-Solvas Jose , Galvez-Vargas Ramon
Published: September 2009

Background:

From April 2-23, 1992, the housekeeping staff of the University of Granada Hospital was on strike. Measures were implemented to minimize the effects of the strike on patients' health and especially to diminish the risk of hospital infection.

Objective:

To assess the risk of nosocomial infection during the housekeeping personnel strike.

Setting:

An 800-bed, tertiary care hospital.

Methods:

A case-cohort approach was used. One hundred forty-eight infected patients (with 184 hospital infections) were detected prospectively from March 1, 1992, to May 31, 1992. A sample of 459 of the base population (patients admitted during the same period) was selected. Information on relevant risk factors for hospital infection was abstracted from patients' clinical charts after hospital discharge. Crude odds ratios and adjusted (by proportional hazards model) relative risks (RRs) for the strike period were estimated.

Results:

Risk of nosocomial infection did not increase during the strike period (multiple-risk factor adjusted RR= 0.99, 0.96 to 1.01/day of strike). Similar results were observed for major sites of infection (especially surgical wound) and major areas of the hospital (including gynecology, surgery, and intensive care).

Conclusion:

We concluded that there was no increase in the risk of nosocomial infection during the housekeeping strike.
Selecting Optimal Statistical Tools
Authors: Birnbaum David
Published: September 2009

The first consideration in analyzing any data is to consider the architecture of study design and quality of the data. Numerous study designs exist, and they vary in their susceptibility to distorting biases. Maclure identifies 32 distinct combinations of taxonomic dimensions by ranking study designs on axes regarding (in decreasing order of importance) randomization, aggregation, parameter timing, selection/allocation, blinded-ness, and proximity. Maclure uses the term aggregation in favor of the often-used term ecologic because groupings may be based on nonspatial units (eg, occupation, family membership, birth periods, etc.), rather than places per se. Parameter timing refers to whether exposure or outcome is measured first or simultaneously; proximity refers to the relative time of measurements, ranging from historic to concurrent. His final figure, which summarizes a composition of six preliminary figures of taxonomic dimension, is reproduced here (Figure 1). Validity is strongest as one moves to the right of center in his summary figure. This taxonomy indicates that blinded randomized trials provide the strongest designs, nonrandomized aggregational studies suffer the greatest susceptibility to bias, and nonrandomized person-based studies rest between.
Molecular Markers for Differentiation of Multiresistant Klebsiella Pneumoniae Isolates in a Pediatric Hospital
Authors: Lhopital Sylvie , Bonacorsi Stephane , Meis Daniele , Brahimi Naima , Mathy Stephanie , Navarro Jean , Aigrain Yves , Bingen Edouard
Published: September 2009

Objective:

To study the spread of extended spectrum ß-lactamase-producing, but aminoglycoside-susceptible, Klebsiella pneumoniae strains in our hospital over an 8-month period, by using two genotypic markers.

Design:

Ribotyping (using two endonucleases) and randomly amplified polymorphic DNA analysis (RAPD; using two different 10-mer primers) were applied to the epidemiological typing of clinical K pneumoniae isolates from stools, ileal fluid, or urine of hospitalized children.

Setting and Patients:

The surgical intensive care ward (S1: 9 patients, 17 isolates), surgical unit (S2: 2 patients, 2 isolates), and gastroenterology ward (GE: 1 patient, 1 isolate) of the Robert Debre Hospital of Paris, France.

Results:

Ribotyping of the 20 clinical isolates, the type strain of the species, and two epidemiologically unrelated isolates with EcoRI and HindlII revealed 6 and 5 different patterns, respectively. Six ribotypes were identified by using these two enzymes. RAPD generated 6 distinct patterns, in complete agreement with ribotyping. Our geno-typic results showed that 11 patients from wards Sl, S2, and GE harbored genotypically related strains, suggesting nosocomial transmission and cross-colonization between and within the three wards.

Conclusions:

Ribotyping and RAPD appear to be reliable methods for distinguishing K pneumoniae strains. The spread of one strain of K pneumoniae in different units of our hospital was demonstrated by both methods. However, RAPD has the advantage of simplicity and rapidity conferred by polymerase chain reaction.
The Epidemiology of Fecal Carriage of Vancomycin-Resistant Enterococci
Authors: Lai Kwan Kew , Fontecchio Sally A , Kelley Anita L , Melvin Zita S. , Baker Stephen
Published: September 2009

An outbreak of vancomycin-resistant enterococci (VRE) began at the University of Massachusetts Medical Center in May 1993. As of September 1995, we had a total of 253 patients infected or colonized with VRE, with consequent increasing demand for private rooms. We analyzed results of surveillance cultures for VRE of 49 patients known to be colonized or infected with VRE. Of these, 34 (70%) were classified as persistent carriers, defined as patients with at least three consecutively positive cultures from any site taken over at least a 2-week period. The length of carriage varied from 19 to 303 days (median, 41 days); 11 patients were converters, defined as patients with three consecutive negative cultures from all previously colonized sites taken over a 3-week period. These patients were free of VRE for 39 to 421 days (median, 142 days). Four were recolonizers after they were documented to be clear of VRE for 33 to 106 days. VRE carriage tends to be prolonged, and hospitalization of patients with VRE will require continued isolation and contact precautions for control of transmission.
Rate of Seasonal Spread of Respiratory Syncytial Virus in a Pediatric Hospital
Authors: McLaws Mary-Louise , Murphy Cathryn , Taylor Peter , Ferson Mark , Gold Julian , Donnellan Robyn , Dalton Dianne
Published: September 2009

The rate of nosocomial respiratory syncytial virus infection was measured in a large pediatric hospital using an incidence density method. The at-risk days for nosocomial RSV were summed during a defined winter period in which there were 54 admissions with community-acquired RSV infection giving a rate of 2.9 cases per 1,000 at-risk days (95% confidence interval, 0.3-5.4 per 1,000).
Retrieve-ensembl-seq: user-friendly and large-scale retrieval of single or multi-genome sequences from Ensembl
Authors: Sand Olivier , Thomas-Chollier Morgane , van Helden Jacques
Published: September 2009

Summary: The preparation of an appropriate sequence dataset is the starting point of all genomic analyses. We present retrieve-ensembl-seq, an application that considerably eases the retrieval of sequences from the Ensembl database, via our user-friendly web site or web services. The user provides Ensembl identifiers or gene names, and the program returns corresponding upstream, downstream, intronic, exonic, UTR or whole gene sequences. retrieve-ensembl-seq also offers a multiple organism mode to retrieve sequences from homologous genes at any taxonomical level. And we introduce various original filters such as the masking of coding fragments and the avoidance of sequence redundancy for genes with multiple transcripts. retrieve-ensembl-seq is included in the software suite regulatory sequence analysis tools (RSAT), allowing instant submission of retrieved sequences to further analysis tools.

Availability:  retrieve-ensembl-seq is integrated in the RSAT suite: http://rsat.ulb.ac.be/rsat. Web site: http://rsat.ulb.ac.be/rsat/retrieve-ensembl-seq_form.cgi. Web services: http://rsat.ulb.ac.be/rsat/web_services/RSATWS.wsdl. Stand-alone distribution: freely available under an academic licence to download from the RSAT web site. The complete manual, a convenient tutorial and demos are available from the RSAT website. Additional help can be found on the RSAT public forum.

Contact:  oly@bigre.ulb.ac.be
Comparative analysis and unification of domain–domain interaction networks
Authors: Björkholm Patrik , Sonnhammer Erik L. L.
Published: September 2009

Motivation: Certain protein domains are known to preferentially interact with other domains. Several approaches have been proposed to predict domain–domain interactions, and over nine datasets are available. Our aim is to analyse the coverage and quality of the existing resources, as well as the extent of their overlap. With this knowledge, we have the opportunity to merge individual domain interaction networks to construct a comprehensive and reliable database.

Results: In this article we introduce a new approach towards comparing domain–domain interaction networks. This approach is used to compare nine predicted domain and protein interaction networks. The networks were used to generate a database of unified domain interactions, UniDomInt. Each interaction in the dataset is scored according to the benchmarked reliability of the sources. The performance of UniDomInt is an improvement compared to the underlying source networks and to another composite resource, Domine.

Availability:  http://sonnhammer.sbc.su.se/download/UniDomInt/

Contact:  Erik.Sonnhammer@sbc.su.se
A Review of a Historical Summit on Integrative Medicine
Authors: Ullman Dana
Published: September 2009

The US Institute of Medical sponsors a “Summit on Integrative Medicine and the Health of the Public” on February 25–27, 2009. A prestigious body of speakers and attendees created a dynamic conference in which the content and discussions provided vital information for transforming the US health care system. Topics included: patient‐centered care, the scientific basis of integrative medicine, health care financing reform and value‐driven care, and mind‐body relationships and health.
Age and growth of longtail tuna (Thunnus tonggol) in tropical and temperate waters of the central Indo-Pacific
Authors: Griffiths Shane P. , Fry Gary C. , Manson Fiona J. , Lou Dong C.
Published: September 2009

Griffiths, S. P., Fry, G. C., Manson, F. J., and Lou, D. C. 2010. Age and growth of longtail tuna (Thunnus tonggol) in tropical and temperate waters of the central Indo-Pacific. – ICES Journal of Marine Science, 67: 125–134. Age and growth of longtail tuna (Thunnus tonggol) were assessed by examination of annual growth increments in sectioned sagittal otoliths from 461 fish (238–1250 mm fork length, LF) sampled from tropical and temperate waters in the central Indo-Pacific between February 2003 and April 2005. Edge and microincrement analyses (presumed daily increments) suggest that longtail tuna deposit a single annual growth increment mainly between August and October. Age was, therefore, estimated for all fish by counting assumed annual growth increments. Ages ranged from 154 d to 18.7 years, with most fish being 3–9 years. Five growth models were fitted to length-at-age data, all of which indicated that the species is relatively slow-growing and long-lived. Recaptures of two tagged fish at liberty for 6.2 and 10.5 years support this notion. A bias-corrected form of Akaike's Information Criterion determined that the Schnute–Richards model provided the best fit to length-at-age data, with model parameter estimates (sexes combined) of L∞ = 135.4 cm LF, K = 22.3 year−1, t0 = 0.120 years, δ = 150.0, v = 0.019, and γ = 2.7 × 10−8. There was no significant difference in growth between sexes. The results suggest that longtail tuna grow more slowly and live longer than other tuna species of similar size. Coupled with their restricted neritic distribution, longtail tuna may be vulnerable to overexploitation by fisheries, and caution needs to be exercised in managing the species until more reliable biological and catch data are collected to assess the status of the population.
Presenilins, Notch dose control the fate of pancreatic endocrine progenitors during a narrow developmental window
Authors: Cras-Méneur Corentin , Li Lin , Kopan Raphael , Permutt M. Alan
Published: September 2009

Canonical Notch signaling is thought to control the endocrine/exocrine decision in early pancreatic progenitors. Later, RBP-Jκ interacts with Ptf1a and E12 to promote acinar differentiation. To examine the involvement of Notch signaling in selecting specific endocrine lineages, we deregulated this pathway by targeted deletion ofpresenilin1andpresenilin2, the catalytic core of γ-secretase, in Ngn3- or Pax6-expressing endocrine progenitors. Surprisingly, whereas Pax6+progenitors were irreversibly committed to the endocrine fate, we discovered that Ngn3+progenitors were bipotential in vivo and in vitro. Whenpresenilinamounts are limiting, Ngn3+progenitors default to an acinar fate; subsequently, they expand rapidly to form the bulk of the exocrine pancreas. γ-Secretase inhibitors confirmed that enzymatic activity was required to block acinar fate selection by Ngn3 progenitors. Genetic interactions identified Notch2 as the substrate, and suggest that γ-secretase and Notch2 act in a noncanonical titration mechanism to sequester RBP-Jκ away from Ptf1a, thus securing selection of the endocrine fate by Ngn3 progenitors. These results revise the current view of pancreatic cell fate hierarchy, establish that Ngn3 is not in itself sufficient to commit cells to the endocrine fate in the presence of Ptf1a, reveal a noncanonical action for Notch2 protein in endocrine cell fate selection, and demonstrate that acquisition of an endocrine fate by Ngn3+progenitors is γ-secretase-dependent until Pax6 expression begins.
MicroRNAs miR-143 and miR-145 modulate cytoskeletal dynamics and responsiveness of smooth muscle cells to injury
Authors: Xin Mei , Small Eric M. , Sutherland Lillian B. , Qi Xiaoxia , McAnally John , Plato Craig F. , Richardson James A. , Bassel-Duby Rhonda , Olson Eric N.
Published: September 2009

Vascular injury triggers dedifferentiation and cytoskeletal remodeling of smooth muscle cells (SMCs), culminating in vessel occlusion. Serum response factor (SRF) and its coactivator, myocardin, play a central role in the control of smooth muscle phenotypes by regulating the expression of cytoskeletal genes. We show that SRF and myocardin regulate a cardiovascular-specific microRNA (miRNA) cluster encoding miR-143 and miR-145. To assess the functions of these miRNAs in vivo, we systematically deleted them singly and in combination in mice. Mice lacking both miR-143 and miR-145 are viable and do not display overt abnormalities in smooth muscle differentiation, although they show a significant reduction in blood pressure due to reduced vascular tone. Remarkably, however, neointima formation in response to vascular injury is profoundly impeded in mice lacking these miRNAs, due to disarray of actin stress fibers and diminished migratory activity of SMCs. These abnormalities reflect the regulation of a cadre of modulators of SRF activity and actin dynamics by miR-143 and miR-145. Thus, miR-143 and miR-145 act as integral components of the regulatory network whereby SRF controls cytoskeletal remodeling and phenotypic switching of SMCs during vascular disease.
The Hsp70 homolog Ssb is essential for glucose sensing via the SNF1 kinase network
Authors: von Plehwe Ulrike , Berndt Uta , Conz Charlotte , Chiabudini Marco , Fitzke Edith , Sickmann Albert , Petersen Astrid , Pfeifer Dietmar , Rospert Sabine
Published: September 2009

Yeast senses the availability of external energy sources via multiple interconnected signaling networks. One of the central components is SNF1, the homolog of mammalian AMP-activated protein kinase, which in yeast is essential for the expression of glucose-repressed genes. When glucose is available hyperphosphorylated SNF1 is rendered inactive by the type 1 protein phosphatase Glc7. Dephosphorylation requires Reg1, which physically targets Glc7 to SNF1. Here we show that the chaperone Ssb is required to keep SNF1 in the nonphosphorylated state in the presence of glucose. Using a proteome approach we found that the Δssb1Δssb2 strain displays alterations in protein expression and suffers from phenotypic characteristics reminiscent of glucose repression mutants. Microarray analysis revealed a correlation between deregulation on the protein and on the transcript level. Supporting studies uncovered that SSB1 was an effective multicopy suppressor of severe growth defects caused by the Δreg1 mutation. Suppression of Δreg1 by high levels of Ssb was coupled to a reduction of Snf1 hyperphosphorylation back to the wild-type phosphorylation level. The data are consistent with a model in which Ssb is crucial for efficient regulation within the SNF1 signaling network, thereby allowing an appropriate response to changing glucose levels.
Cephalochordates (Amphioxus or Lancelets): A Model for Understanding the Evolution of Chordate Characters: Figure 1.
Authors: Yu J.K. Sky , Holland Linda Z.
Published: September 2009

INTRODUCTION

Cephalochordates, commonly called amphioxus or lancelets, are marine invertebrate chordates. They were traditionally held as the closest living relatives of vertebrates and have long occupied a key phylogenetic position in our discussions on the origin of vertebrates. However, recent molecular phylogenetic analyses have placed tunicates as the sister taxon of vertebrates and cephalochordates as the most basal chordate group. Therefore, studies on cephalochordates can also provide important insights for understanding the conserved patterning mechanisms for all chordates. Cephalochordates and vertebrates share several key chordate features, namely, the dorsal nerve cord, notochord, segmented somites, and pharyngeal gill slits. However, vertebrates have ~25% more genes than amphioxus and several features not present in cephalochordates, including a more elaborate head and forebrain, migratory neural crest, and neurogenic placodes. During the last two decades, expression patterns of important developmental genes have been used extensively to infer homologies between cephalochordate and vertebrate embryos. These studies have answered some long-standing questions concerning the evolution of vertebrates from their invertebrate ancestors and have also generated interesting hypotheses for further investigations. With the completion of a cephalochordate genome project in 2008, comparison between cephalochordate and vertebrate genomes has allowed us to postulate a picture for the genome of the last common ancestor of all chordates. Such comparisons are also providing important biological insights into the functional biology of cephalochordates and the evolutionary origin of developmental mechanisms that led to the emergence of the vertebrate body plan.
The Western Clawed Frog (Xenopus tropicalis): An Emerging Vertebrate Model for Developmental Genetics and Environmental Toxicology
Authors: Showell Chris , Conlon Frank L.
Published: September 2009

INTRODUCTION

Xenopus tropicalis, also known as Silurana tropicalis or the Western clawed frog, is a small, wholly aquatic frog that is found in the countries that lie along the west coast of equatorial Africa from Gabon to Sierra Leone. It is a diploid relative of Xenopus laevis, a frog that has found many uses in laboratories throughout the world. X. tropicalis closely resembles its relative in anatomy throughout its life cycle. It shares its advantages as a model organism for studying many aspects of vertebrate biology, particularly the genetic, biochemical, and environmental factors that influence vertebrate development from embryonic stages through adulthood. It also shares much of its biology with more distantly related species, including mammals, but unlike mammalian model organisms, routine manipulation of X. tropicalis in the laboratory can produce thousands of embryos for experimental analysis. X. tropicalis is also finding uses as an important test species for assessing the impact of environmental toxins and disease on amphibians, which are in decline in many areas of the world due to waterborne pollutants and infectious agents such as the chytrid fungus.
In Vivo Imaging of Mammalian Cells: Cell Engineering and Viability
Authors: Swedlow Jason R. , Andrews Paul D. , Platani Melpomeni
Published: September 2009

INTRODUCTION

There are a number of instances in which the use of live cell imaging has provided critical insight into cellular and tissue function. It has therefore become a requisite analytical tool for use in cell biology, neurobiology, and developmental biology, as well as a routine method practiced in many biomedical research laboratories. The technical requirements for performing live cell imaging include a capacity to engineer appropriate cells for the imaging experiment and access to a digital image acquisition system. The availability of fluorescent protein (FP) technology allows the molecular specificity of fluorescent markers to be used in a genetically encoded manner. In this article, we describe techniques for the generation and maintenance of fluorescently labeled cells and their use in live cell imaging experiments.
In Vivo Dimethyl Sulfate (DMS) Footprinting via Ligation-Mediated Polymerase Chain Reaction (LM-PCR): Figure 1.
Authors: Carey Michael F. , Peterson Craig L. , Smale Stephen T.
Published: September 2009

INTRODUCTION

Ligation-mediated polymerase chain reaction (LM-PCR) is a powerful technique for detecting DNA strand breaks within a complex sample because of its extreme sensitivity and specificity. This general method has proven to be invaluable for several purposes, in particular, in analysis of the properties of an endogenous DNA locus. LM-PCR can be coupled to a number of DNA modification and cleavage reagents. Coupling to dimethyl sulfate (DMS) modification, described here, results in assays that are analogous to in vitro DNase I footprinting or methylation protection. In this method, DMS is added directly to intact cells. After quenching the modification reaction, genomic DNA is purified, and cleavage adjacent to modified bases is induced by piperidine treatment. LM-PCR is then performed on the cleaved genomic DNA.
Chromatin Immunoprecipitation (ChIP)
Authors: Carey Michael F. , Peterson Craig L. , Smale Stephen T.
Published: September 2009

INTRODUCTION

Chromatin immunoprecipitation (ChIP) is an invaluable method for studying interactions between specific proteins or modified forms of proteins and a genomic DNA region. ChIP can be used to determine whether a transcription factor interacts with a candidate target gene and is used with equal frequency to monitor the presence of histones with post-translational modifications at specific genomic locations. In early ChIP studies, UV light from a transilluminator was used to cross-link proteins to DNA irreversibly. The cross-linked chromatin was then either sonicated or cleaved with restriction enzymes to generate smaller DNA fragments, followed by immunoprecipitation with the desired antibodies. The precipitated protein-DNA adducts were then purified, treated with a protease, and analyzed by dot blot or Southern blot using a radiolabeled probe derived from the cloned DNA fragment of interest. The use of formaldehyde as a reversible protein-DNA and protein-protein cross-linking agent for ChIP and the use of polymerase chain reaction (PCR) to detect precipitated DNA fragments were later added as components of the modern ChIP procedure. The protocol below represents a standard ChIP procedure for use in mammalian cells. Cross-linking is performed by adding formaldehyde to growing cells, and chromatin is prepared, sheared by sonication, and precleared to reduce nonspecific immunoprecipitation. Immunoprecipitation is performed with a specific antibody. After elution of the protein-DNA complexes from protein A- or protein G-agarose resin, the samples are heated to reverse the covalent cross-links. The DNA fragments are purified and analyzed by PCR or real-time PCR.
Position-Based Clustering of Microarray Expression Data
Authors: Faulk Christopher , Kim Joomyeong
Published: September 2009

INTRODUCTION

Microarray expression analysis has traditionally focused only on genes with the highest expression level changes, ignoring the majority of genes with lower fold changes. To address this problem, we provide a simple method that can derive additional useful biological information from the rest of the genes included on an array. Chromosomal position information combined with expression data, either raw or in expression-level changes, gives a regional overview of expression from an array. This protocol is tailored to Affymetrix data, but can be used for other types of microarray results. The procedure is illustrated by the reanalysis of a data set, GSE5230, previously deposited in the Gene Expression Omnibus (GEO). Using this procedure, we identified several classes of chromosomal regions where expression levels were affected in concert. Linking expression data to chromosomal position also allowed us to identify several genomic regions displaying low but steady fold change, which were missed by traditional approaches. Overall, this method is useful in detecting regional regulatory changes. It should allow for greater use of the large quantity of previously overlooked microarray data.
Induction of Spawning in the Starlet Sea Anemone Nematostella vectensis, In Vitro Fertilization of Gametes, and Dejellying of Zygotes
Authors: Genikhovich Grigory , Technau Ulrich
Published: September 2009

INTRODUCTION

This protocol describes a reliable method for inducing spawning in the starlet sea anemone Nematostella vectensis under laboratory conditions. Spawning occurs at ~13±3 h after the onset of the inductive cues, which are a particular combination of light and temperature. This method has been used successfully on a daily basis with rotating cultures, yielding thousands of embryos every day. Each given female or male can be induced once per week. For long-term health, an induction once every 10-14 d is less exhausting to the animals. The protocol also describes the dejellying of the zygotes for further manipulation of the embryos.
Anti-acetylated Tubulin Antibody Staining and Phalloidin Staining in the Starlet Sea Anemone Nematostella vectensis
Authors: Genikhovich Grigory , Technau Ulrich
Published: September 2009

INTRODUCTION

This protocol describes anti-acetylated tubulin antibody staining in the starlet sea anemone Nematostella vectensis. The antibody against acetylated tubulin mainly recognizes cilia in N. vectensis. In planula larvae, it not only stains the cilia covering the ectoderm, but also is a good marker for the apical tuft at the aboral pole. Variations of this protocol can be used for staining with other antibodies (Marlow et al. 2009). However, not all antigens are preserved by fixation in paraformaldehyde; thus, alternative fixation methods are also described. In addition, an optional step of phalloidin staining of fibrillar actin is included.
Amphioxus (Branchiostoma floridae) Spawning and Embryo Collection
Authors: Yu J.K. Sky , Holland Linda Z.
Published: September 2009

INTRODUCTION

This protocol describes how to obtain and collect embryos from the amphioxus Branchiostoma floridae. Currently, it is only possible to obtain fertilizable gametes from this species on the days in summer when ripe animals would normally spawn in the field once about every 2 wk. Sexes are separate, and in the natural environment, mature oocytes undergo meiotic maturation and arrest at the second meiotic metaphase in the early afternoon, while sperm gain the ability to become motile upon exposure to seawater. In the evening, animals with mature gametes emerge from the sand to spawn ~30 min after sunset, that is, ~9:00 p.m. Animals collected in the afternoon of the day of spawning can be brought into the laboratory and prevented from spawning by the normal room light in the laboratory, allowing the investigator to obtain freshly spawned eggs and sperm throughout the night either by placing adults in the dark or by applying a mild electric shock (50 V DC). Unfortunately, toward dawn of the next day, the quality of gametes begins to drop, and the animals stop spawning. Therefore, investigators need to work during the night to fertilize eggs and perform experiments on early embryos.
Amphioxus Whole-Mount In Situ Hybridization
Authors: Yu J.K. Sky , Holland Linda Z.
Published: September 2009

INTRODUCTION

The in situ hybridization (ISH) technique is one of the basic methods of developmental biology. It allows the distributions of specific transcripts to be detected in fixed cells or embryos during development. This protocol describes ISH of digoxigenin (DIG)-labeled antisense RNA probes to whole-mount amphioxus embryos and larvae. Embryos are fixed and permeabilized before being hybridized with the DIG-labeled antisense RNA probes. Following hybridization, transcripts are detected immunohistochemically with an anti-DIG antibody conjugated to alkaline phosphatase. The gene expression patterns are visualized in the embryos with colorimeric substrates of alkaline phosphatase. Labeled embryos are photographed as whole mounts, after which they can be embedded in Spurr’s resin and sectioned if desired. To facilitate the handling of small amphioxus embryos during whole-mount ISH, we developed a method of solution exchange using homemade mesh-bottom baskets and multiwell plates. This method speeds up time-consuming aspiration and decanting of buffers and avoids accidental loss of precious embryos during these steps. Because ISH conditions are the same for most of the probes tested, this design allows high-throughput analysis of gene expression during embryogenesis to be carried out.
Natural Mating and Tadpole Husbandry in the Western Clawed Frog Xenopus tropicalis
Authors: Showell Chris , Conlon Frank L.
Published: September 2009

INTRODUCTION

The routine generation and successful husbandry of tadpoles is essential for investigators using the Western clawed frog Xenopus tropicalis for developmental and genetic studies. We describe a method to induce natural mating by injecting a hormone into sexually mature X. tropicalis, and to raise the offspring through the embryonic, tadpole, and metamorphosis stages. X. tropicalis develop at a faster rate than Xenopus laevis, reaching metamorphosis in as little as 4 wk. Moreover, natural mating produces large numbers of offspring and is therefore particularly useful for mutation screening and genetic mapping. The method includes a technique for the anesthesia of frogs, which makes them easier to handle during injection of the hormone, reducing the likelihood of injury to the animals.
RNA Extraction from Arabidopsis for Northern Blots and Reverse Transcriptase-PCR
Authors: Ahn Ji H.
Published: September 2009

INTRODUCTION

Analysis of the cellular distribution of gene products (RNA and protein) can provide important clues to the function of genes in vivo. RNA expression can be monitored either after extraction from plants or tissues or in situ. The former procedure is easier to perform and more quantitative, whereas the latter provides much more detailed resolution of spatial and temporal expression patterns but is often more technically challenging. This protocol describes a method for RNA extraction from Arabidopsis thaliana. Frozen plant tissue is homogenized using a hand drill fitted with a plastic micropestle instead of using a conventional mortar and pestle, which can be inefficient and physically demanding. This homogenization method allows samples to be processed quickly (up to 12 samples/hour). Typical yields are 20-40 μg of total RNA from 100 mg of Arabidopsis seedlings. The RNA extracted using this method is suitable for reverse transcriptase-polymerase chain reaction (RT-PCR) and Northern hybridization.
Semiquantitative Analysis of Arabidopsis RNA by Reverse Transcription Followed by Noncompetitive PCR
Authors: Ahn Ji H.
Published: September 2009

INTRODUCTION

This article describes the analysis of Arabidopsis RNA by reverse transcriptase-polymerase chain reaction (RT-PCR). RNA is first reverse-transcribed into cDNA, which is then amplified by PCR. This method is quick and easy. Although RT-PCR can be performed quantitatively, it is often technically challenging to obtain highly accurate results. We recommend the use of an RT-PCR kit; the procedure given here is adapted from a commercial kit protocol. In the PCR step, it is advisable to use oligonucleotide primers that span at least one intron. In this way, amplification of any contaminating genomic DNA is readily detected, because the product derived from genomic DNA will be larger than the cDNA product. Expression levels are determined by comparing against a gene that is expressed fairly uniformly, such as the ubiquitin gene UBQ10.
Semiquantitative Analysis of Arabidopsis RNA by Reverse Transcription Followed by PCR Using Mimics
Authors: Kardailsky Igor
Published: September 2009

INTRODUCTION

This article describes the analysis and quantification of Arabidopsis RNA by reverse transcriptase-polymerase chain reaction (RT-PCR). RNA is first reverse-transcribed into cDNA, which is then amplified by competitive PCR using a mimic. Multiple PCRs are performed using a constant amount of unknown cDNA and varying amounts of a mimic; that is, a template of known concentration that is close in size and GC content to the quantified target and that contains the same annealing sites for primers. Mimics are usually designed to produce PCR products ~10%-20% smaller or larger than the endogenous target. A mimic can be used as a DNA standard (as in this protocol), to be included after the reverse transcription reaction, or, even better, as an RNA standard. In the latter case, an RNA molecule must be generated from a vector designed for in vitro transcription, such as pGEM or pBluescript series vectors. The procedure described here is based on commercial kit protocols.
The role of lockups in takeover contests
Authors: Che Yeon‐Koo , Lewis Tracy R.
Published: September 2009

We examine breakup fees and stock lockups as devices for prospective target firms to encourage bidder participation in takeover contests. Unless bidding costs for the first bidder are too high, breakup fees provide for the socially desirable degree of competition and ensure the efficient allocation of the target to the highest‐valued buyer in a takeover auction. In contrast, stock lockups permit the target firm to subsidize entry of a new bidder at the expense of an incumbent bidder. Stock lockups induce too much competition when offered to a second bidder and too little competition when offered to a first bidder. Despite their socially wasteful properties, target management would favor stock lockups, as they induce takeover competition at least cost to the target.
Alignment, clade robustness and fungal phylogenetics— Crepidotaceae and sister families revisited
Authors: Petersen Gitte , Knudsen Henning , Seberg Ole
Published: September 2009

The circumscription and delimitation of the agaric family Crepidotaceae and related families in the Agaricales have been analysed using nLSU sequence data and direct optimization. The influence of alignment parameters on clade robustness has been explored, and many clades are found to have a low robustness. Nevertheless, based on clades with a high or moderately high robustness to increased gap costs, a few taxonomic changes are suggested: Crepidotaceae has been amended to include Crepidotus, Episphaeria, Simocybe, Pleuroflammula and Inocybe, the latter genus adding the homoplasious character mycorrhization to the family, Pellidiscus is included in Crepidotus, a position supported by micromorphology, two other genera formerly included in the family, Chromocyphella and Phaeosolenia, are excluded, and a new family, Chromocyphellaceae fam. nov., is erected for them based on fruit body shape, microcharacters, and sequences. Monophyly of Tubariaceae as recently circumscribed was not recovered, but tentatively we accept the genera Tubaria and Phaeomarasmius as members of the family.

© The Willi Hennig Society 2009.
Program note: Cladescan, a program for automated phylogenetic sensitivity analysis
Authors: Sanders Jon G.
Published: September 2009

Examination of trees for the presence of particular nodes is a fundamental aspect of systematics, and is the basis of phylogenetic sensitivity analysis, but becomes unwieldy when performed manually for complex nodes or over large numbers of trees. The program Cladescan is presented here as a stand‐alone application to facilitate the detection of nodes in such situations. Cladescan includes features useful for phylogenetic sensitivity analysis, such as automatic generation of “Navajo rug” sensitivity plots. In addition, researchers may find it useful for general comparisons among large data sets.

© The Willi Hennig Society 2009.