bluuebunny/crossref_metadata_2025 huggingface dataset:
Multiple lineage specific expansions within the guanylyl cyclase gene family
Authors: Fitzpatrick David A , O'Halloran Damien M , Burnell Ann M
Published: April 2006

Background

Guanylyl cyclases (GCs) are responsible for the production of the secondary messenger cyclic guanosine monophosphate, which plays important roles in a variety of physiological responses such as vision, olfaction, muscle contraction, homeostatic regulation, cardiovascular and nervous function. There are two types of GCs in animals, soluble (sGCs) which are found ubiquitously in cell cytoplasm, and receptor (rGC) forms which span cell membranes. The complete genomes of several vertebrate and invertebrate species are now available. These data provide a platform to investigate the evolution of GCs across a diverse range of animal phyla.

Results

In this analysis we located GC genes from a broad spectrum of vertebrate and invertebrate animals and reconstructed molecular phylogenies for both sGC and rGC proteins. The most notable features of the resulting phylogenies are the number of lineage specific rGC and sGC expansions that have occurred during metazoan evolution. Among these expansions is a large nematode specific rGC clade comprising 21 genes in C. elegans alone; a vertebrate specific expansion in the natriuretic receptors GC-A and GC-B; a vertebrate specific expansion in the guanylyl GC-C receptors, an echinoderm specific expansion in the sperm rGC genes and a nematode specific sGC clade. Our phylogenetic reconstruction also shows the existence of a basal group of nitric oxide (NO) insensitive insect and nematode sGCs which are regulated by O2. This suggests that the primordial eukaryotes probably utilized sGC as an O2 sensor, with the ligand specificity of sGC later switching to NO which provides a very effective local cell-to-cell signalling system. Phylogenetic analysis of the sGC and bacterial heme nitric oxide/oxygen binding protein domain supports the hypothesis that this domain originated from a cyanobacterial source.

Conclusion

The most salient feature of our phylogenies is the number of lineage specific expansions, which have occurred within the GC gene family during metazoan evolution. Our phylogenetic analyses reveal that the rGC and sGC multi-domain proteins evolved early in eumetazoan evolution. Subsequent gene duplications, tissue specific expression patterns and lineage specific expansions resulted in the evolution of new networks of interaction and new biological functions associated with the maintenance of organismal complexity and homeostasis.
Population genetic estimation of the loss of genetic diversity during horizontal transmission of HIV-1
Authors: Edwards Charles TT , Holmes Edward C , Wilson Daniel J , Viscidi Raphael P , Abrams Elaine J , Phillips Rodney E , Drummond Alexei J
Published: April 2006

Background

Genetic diversity of the human immunodeficiency virus type 1 (HIV-1) population within an individual is lost during transmission to a new host. The demography of transmission is an important determinant of evolutionary dynamics, particularly the relative impact of natural selection and genetic drift immediately following HIV-1 infection. Despite this, the magnitude of this population bottleneck is unclear.

Results

We use coalescent methods to quantify the bottleneck in a single case of homosexual transmission and find that over 99% of theenvandgagdiversity present in the donor is lost. This was consistent with the diversity present at seroconversion in nine other horizontally infected individuals. Furthermore, we estimated viral diversity at birth in 27 infants infected through vertical transmission and found there to be no difference between the two modes of transmission.

Conclusion

Assuming the bottleneck at transmission is selectively neutral, such a severe reduction in genetic diversity has important implications for adaptation in HIV-1, since beneficial mutations have a reduced chance of transmission.
Assessment of methods for amino acid matrix selection and their use on empirical data shows that ad hoc assumptions for choice of matrix are not justified
Authors: Keane Thomas M , Creevey Christopher J , Pentony Melissa M , Naughton Thomas J , Mclnerney James O
Published: April 2006

Background

In recent years, model based approaches such as maximum likelihood have become the methods of choice for constructing phylogenies. A number of authors have shown the importance of using adequate substitution models in order to produce accurate phylogenies. In the past, many empirical models of amino acid substitution have been derived using a variety of different methods and protein datasets. These matrices are normally used as surrogates, rather than deriving the maximum likelihood model from the dataset being examined. With few exceptions, selection between alternative matrices has been carried out in an ad hoc manner.

Results

We start by highlighting the potential dangers of arbitrarily choosing protein models by demonstrating an empirical example where a single alignment can produce two topologically different and strongly supported phylogenies using two different arbitrarily-chosen amino acid substitution models. We demonstrate that in simple simulations, statistical methods of model selection are indeed robust and likely to be useful for protein model selection. We have investigated patterns of amino acid substitution among homologous sequences from the three Domains of life and our results show that no single amino acid matrix is optimal for any of the datasets. Perhaps most interestingly, we demonstrate that for two large datasets derived from the proteobacteria and archaea, one of the most favored models in both datasets is a model that was originally derived from retroviral Pol proteins.

Conclusion

This demonstrates that choosing protein models based on their source or method of construction may not be appropriate.
Spontaneous rescue from cystic fibrosis in a mouse model
Authors: Charizopoulou Nikoletta , Wilke Martina , Dorsch Martina , Bot Alice , Jorna Huub , Jansen Silke , Stanke Frauke , Hedrich Hans J , de Jonge Hugo R , Tümmler Burkhard
Published: April 2006

Background

From the original Cftr

TgH(neoim)Hgu
mutant mouse model with a divergent genetic background (129P2, C57BL/6, MF1) we have generated two inbred Cftr

TgH(neoim)Hgu
mutant strains named CF/1-Cftr

TgH(neoim)Hgu
and CF/3-Cftr

TgH(neoim)Hgu
, which are fertile and show normal growth and lifespan. Initial genome wide scan analysis with microsatellite markers indicated that the two inbred strains differed on the genetic level. In order to further investigate whether these genetic differences have an impact on the disease phenotype of cystic fibrosis we characterised the phenotype of the two inbred strains.

Results

Reduced amounts, compared to wild type control animals, of correctly spliced Cftr mRNA were detected in the nasal epithelia, lungs and the intestine of both inbred Cftr

TgH(neoim)Hgu
strains, with higher residual amount observed for CF/1-Cftr

TgH(neoim)Hgu
than CF/3-Cftr

TgH(neoim)Hgu
for every investigated tissue. Accordingly the amounts of wild type Cftr protein in the intestine were 9% for CF/1-Cftr

TgH(neoim)Hgu
and 4% for CF/3-Cftr

TgH(neoim)Hgu
. Unlike the apparent strain and/or tissue specific regulation of Cftr mRNA splicing, short circuit current measurements in the respiratory and intestinal epithelium revealed that both strains have ameliorated the basic defect of cystic fibrosis with a presentation of a normal electrophysiology in both tissues.

Conclusion

Unlike the outbred Cftr

TgH(neoim)Hgu
insertional mouse model, which displayed the electrophysiological defect in the gastrointestinal and respiratory tracts characteristic of cystic fibrosis, both inbred Cftr

TgH(neoim)Hgu
strains have ameliorated the electrophysiological defect. On the basis of these findings both CF/1-Cftr

TgH(neoim)Hgu
and CF/3-Cftr

TgH(neoim)Hgu
offer an excellent model whereby determination of the minimal levels of protein required for the restoration of the basic defect of cystic fibrosis can be studied, along with the modulating factors which may affect this outcome.
Comparison of the latest commercial short and long oligonucleotide microarray technologies
Authors: de Reyniès Aurélien , Geromin Daniela , Cayuela Jean-Michel , Petel Fabien , Dessen Philippe , Sigaux François , Rickman David S
Published: April 2006

Background

We compared the relative precision and accuracy of expression measurements obtained from three different state-of-the-art commercial short and long-oligonucleotide microarray platforms (Affymetrix GeneChip™, GE Healthcare CodeLink™ and Agilent Technologies). The design of the comparison was chosen to judge each platform in the context of a multi-project program.

Results

All wet-lab experiments and raw data acquisitions were performed independently by each commercial platform. Intra-platform reproducibility was assessed using measurements from all available targets. Inter-platform comparisons of relative signal intensities were based on a common and non-redundant set of roughly 3,400 targets chosen for their unique correspondence toward a single transcript. Despite many examples of strong similarities we found several areas of discrepancy between the different platforms.

Conclusion

We found a higher level of reproducibility from one-color based microarrays (Affymetrix and CodeLink) compared to the two-color arrays from Agilent. Overall, Affymetrix data had a slightly higher level of concordance with sample-matched real-time quantitative reverse-transcriptase polymerase chain reaction (QRT-PCR) data particularly for detecting small changes in gene expression levels.
Linkage disequilibrium interval mapping of quantitative trait loci
Authors: Boitard Simon , Abdallah Jihad , de Rochambeau Hubert , Cierco-Ayrolles Christine , Mangin Brigitte
Published: April 2006

Background

For many years gene mapping studies have been performed through linkage analyses based on pedigree data. Recently, linkage disequilibrium methods based on unrelated individuals have been advocated as powerful tools to refine estimates of gene location. Many strategies have been proposed to deal with simply inherited disease traits. However, locating quantitative trait loci is statistically more challenging and considerable research is needed to provide robust and computationally efficient methods.

Results

Under a three-locus Wright-Fisher model, we derived approximate expressions for the expected haplotype frequencies in a population. We considered haplotypes comprising one trait locus and two flanking markers. Using these theoretical expressions, we built a likelihood-maximization method, called HAPim, for estimating the location of a quantitative trait locus. For each postulated position, the method only requires information from the two flanking markers. Over a wide range of simulation scenarios it was found to be more accurate than a two-marker composite likelihood method. It also performed as well as identity by descent methods, whilst being valuable in a wider range of populations.

Conclusion

Our method makes efficient use of marker information, and can be valuable for fine mapping purposes. Its performance is increased if multiallelic markers are available. Several improvements can be developed to account for more complex evolution scenarios or provide robust confidence intervals for the location estimates.
Developmental disturbances in early life stage mortality (M74) of Baltic salmon fry as studied by changes in gene expression
Authors: Vuori Kristiina AM , Koskinen Heikki , Krasnov Aleksei , Koivumäki Paula , Afanasyev Sergey , Vuorinen Pekka J , Nikinmaa Mikko
Published: April 2006

Background

We have studied alterations of gene expression associated with naturally-occurring early life stage mortality (M74) in Baltic salmon using a cDNA microarray and real time PCR. M74-affected fry have several typical neurological, cardiovascular and pathological symptoms. They are also characterized by low thiamine content and show signs of oxidative stress.

Results

Affected fry can be divided into three major groups with early, intermediate or late onset of mortality. If mortality starts during the first third of the yolk-sac stage, virtually all the responses are compatible with stress, which rapidly leads to the common terminal responses. If death occurs during the second third of the yolk sac stage, the terminal stage is preceded by a decrease in globin gene expression, which leads to internal hypoxia when the animals grow and shift from skin- to gill-breathing. Fry will eventually proceed to the terminal responses. The group developing M74 most slowly appears to compensate for reduced oxygen delivery by downregulation of metabolism, and hence some fry can escape death.

Conclusion

Our study is the first demonstration of diverse transcriptional responses to a naturally-occurring developmental disturbance. Since many of the genes differentially expressed in M74-fry are evolutionarily conserved, the M74 of Baltic salmon can serve as a model for developmental disturbances and environmental stress responses in vertebrates in general.
Trypanosoma cruzi mitochondrial maxicircles display species- and strain-specific variation and a conserved element in the non-coding region
Authors: Westenberger Scott J , Cerqueira Gustavo C , El-Sayed Najib M , Zingales Bianca , Campbell David A , Sturm Nancy R
Published: April 2006

Background

The mitochondrial DNA of kinetoplastid flagellates is distinctive in the eukaryotic world due to its massive size, complex form and large sequence content. Comprised of catenated maxicircles that contain rRNA and protein-coding genes and thousands of heterogeneous minicircles encoding small guide RNAs, the kinetoplast network has evolved along with an extreme form of mRNA processing in the form of uridine insertion and deletion RNA editing. Many maxicircle-encoded mRNAs cannot be translated without this post-transcriptional sequence modification.

Results

We present the complete sequence and annotation of the Trypanosoma cruzi maxicircles for the CL Brener and Esmeraldo strains. Gene order is syntenic with Trypanosoma brucei and Leishmania tarentolae maxicircles. The non-coding components have strain-specific repetitive regions and a variable region that is unique for each strain with the exception of a conserved sequence element that may serve as an origin of replication, but shows no sequence identity with L. tarentolae or T. brucei. Alternative assemblies of the variable region demonstrate intra-strain heterogeneity of the maxicircle population. The extent of mRNA editing required for particular genes approximates that seen in T. brucei. Extensively edited genes were more divergent among the genera than non-edited and rRNA genes. Esmeraldo contains a unique 236-bp deletion that removes the 5'-ends of ND4 and CR4 and the intergenic region. Esmeraldo shows additional insertions and deletions outside of areas edited in other species in ND5, MURF1, and MURF2, while CL Brener has a distinct insertion in MURF2.

Conclusion

The CL Brener and Esmeraldo maxicircles represent two of three previously defined maxicircle clades and promise utility as taxonomic markers. Restoration of the disrupted reading frames might be accomplished by strain-specific RNA editing. Elements in the non-coding region may be important for replication, transcription, and anchoring of the maxicircle within the kinetoplast network.
DNA deformability changes of single base pair mutants within CDE binding sites in S. Cerevisiae centromere DNA correlate with measured chromosomal loss rates and CDE binding site symmetries
Authors: Hennemuth Brad , Marx Kenneth A
Published: April 2006

Background

The centromeres in yeast (S. cerevisiae) are organized by short DNA sequences (125 bp) on each chromosome consisting of 2 conserved elements: CDEI and CDEIII spaced by a CDEII region. CDEI and CDEIII are critical sequence specific protein binding sites necessary for correct centromere formation and following assembly with proteins, are positioned near each other on a specialized nucleosome. Hegemann et al. BioEssays 1993, 15: 451–460 reported single base DNA mutants within the critical CDEI and CDEIII binding sites on the centromere of chromosome 6 and quantitated centromere loss of function, which they measured as loss rates for the different chromosome 6 mutants during cell division. Olson et al. Proc Natl Acad Sci USA 1998, 95: 11163–11168 reported the use of protein-DNA crystallography data to produce a DNA dinucleotide protein deformability energetic scale (PD-scale) that describes local DNA deformability by sequence specific binding proteins. We have used the PD-scale to investigate the DNA sequence dependence of the yeast chromosome 6 mutants' loss rate data. Each single base mutant changes 2 PD-scale values at that changed base position relative to the wild type. In this study, we have utilized these mutants to demonstrate a correlation between the change in DNA deformability of the CDEI and CDEIII core sites and the overall experimentally measured chromosome loss rates of the chromosome 6 mutants.

Results

In the CDE I and CDEIII core binding regions an increase in the magnitude of change in deformability of chromosome 6 single base mutants with respect to the wild type correlates to an increase in the measured chromosome loss rate. These correlations were found to be significant relative to 105 Monte Carlo randomizations of the dinucleotide PD-scale applied to the same calculation. A net loss of deformability also tends to increase the loss rate. Binding site position specific, 4 data-point correlations were also created using the wild type sequence and the 3 associated alternate base mutants at each binding site position. These position specific slope magnitudes, or sensitivities, correlated with and reflected the underlying position symmetry of the DNA binding sequences.

Conclusion

These results suggest the utility of correlating quantitative aspects of sequence specific protein-DNA complex single base mutants with changes in the easily calculated PD-deformability scale of the individual DNA sequence mutants. Using this PD approach, it may be possible in the future to understand the magnitude of biological or energetic functional effects of specific DNA sequence mutants within DNA-protein complexes in terms of their effect on DNA deformability.
A potential role of alternative splicing in the regulation of the transcriptional activity of human GLI2 in gonadal tissues
Authors: Speek Mart , Njunkova Olga , Pata Illar , Valdre Eola , Kogerman Priit
Published: April 2006

Background

Mammalian Gli proteins are important transcription factors involved in the regulation of Sonic hedgehog signal transduction pathway. Association of Gli2 with mammalian development and human disease led us to study the structure and expression of the human GLI2.

Results

We show that the region encoding GLI2 repressor domain is subject to alternative splicing in the gonadal tissues and different cell lines. Two major alternatively spliced forms of GLI2 mRNA arise from skipping exon 3 (GLI2 Δ3) or exons 4 and 5 (GLI2 Δ4–5). Both forms contain premature translational stop codons in the GLI2 open reading frame (ORF) starting from exon 2. Translation of GLI2 Δ3 and GLI2 Δ4–5 in vitro, initiated from downstream AUG codons, produced N-terminally truncated proteins. In Gli-dependent transactivation assay, expression of GLI2 Δ3 induced activation of the reporter gene similar to that of the full-length construct (GLI2 fl) containing complete ORF. However, expression of the GLI2 Δ4–5 resulted in about 10-fold increase in activation, suggesting that deletion of the major part of repressor domain was responsible for the enhanced activation of GLI2 protein.

Conclusion

Our data suggest that in addition to proteolytic processing, alternative splicing may be another important regulatory mechanism for the modulation of repressor and activator properties of GLI2 protein.
Behavioral effects of ketamine and toxic interactions with psychostimulants
Authors: Hayase Tamaki , Yamamoto Yoshiko , Yamamoto Keiichi
Published: April 2006

Background

The anesthetic drug ketamine (KT) has been reported to be an abused drug and fatal cases have been observed in polydrug users. In the present study, considering the possibility of KT-enhanced toxic effects of other drugs, and KT-induced promotion of an overdose without making the subject aware of the danger due to the attenuation of several painful subjective symptoms, the intraperitoneal (i.p.) KT-induced alterations in behaviors and toxic interactions with popular co-abused drugs, the psychostimulants cocaine (COC) and methamphetamine (MA), were examined in ICR mice.

Results

A single dose of KT caused hyperlocomotion in a low (30 mg/kg, i.p.) dose group, and hypolocomotion followed by hyperlocomotion in a high (100 mg/kg, i.p.) dose group. However, no behavioral alterations derived from enhanced stress-related depression or anxiety were observed in the forced swimming or the elevated plus-maze test. A single non-fatal dose of COC (30 mg/kg, i.p.) or MA (4 mg/kg, i.p.) caused hyperlocomotion, stress-related depression in swimming behaviors in the forced swimming test, and anxiety-related behavioral changes (preference for closed arms) in the elevated plus-maze test. For the COC (30 mg/kg) or MA (4 mg/kg) groups of mice simultaneously co-treated with KT, the psychostimulant-induced hyperlocomotion was suppressed by the high dose KT, and the psychostimulant-induced behavioral alterations in the above tests were reversed by both low and high doses of KT. For the toxic dose COC (70 mg/kg, i.p.)- or MA (15 mg/kg, i.p.)-only group, mortality and severe seizures were observed in some animals. In the toxic dose psychostimulant-KT groups, KT attenuated the severity of seizures dose-dependently. Nevertheless, the mortality rate was significantly increased by co-treatment with the high dose KT.

Conclusion

Our results demonstrated that, in spite of the absence of stress-related depressive and anxiety-related behavioral alterations following a single dose of KT treatment, and in spite of the KT-induced anticonvulsant effects and attenuation of stress- and anxiety-related behaviors caused by COC or MA, the lethal effects of these psychostimulants were increased by KT.
Cyclooxygenase expression is not required for release of arachidonic acid from cells by some nonsteroidal anti-inflammatory drugs and cancer preventive agents
Authors: Levine Lawrence
Published: April 2006

Background

Nonsteroidal anti-inflammatory drugs (NSAIDs) have been shown to be effective in inhibiting colorectal cancer. Cyclooxygenase activity is thought to mediate, in part, this cancer preventive effect. From observations made when cells that express cyclooxygenase activity were treated with NSAIDs and known cancer preventive agents, I have postulated that arachidonic acid (AA) release is associated with cancer prevention. In this study, the effects of NSAIDs on two cells that do not express cycloxygenase activity are detailed.

Results

NSAIDs and several cancer preventive agents release AA from human colon cancer cells (the HCT-15 cell line). The concentrations of NSAIDs required to release significant amounts of AA from the HCT-15 cells are greater than those required to inhibit the lactacystin plus 12-0-tetradecanoyl-13-acetate stimulated cyclooxygenase activity of rat liver cells. NSAIDs, tamoxifen and simvastatin were found to hemolyze erythrocyte cells which also do not express cyclooxygenase activity

Conclusion

The data demonstrate that AA release is independent of cyclooxygenase activity and together with hemolysis suggest that intercalation of the plasma membrane by some NSAIDs and cancer preventive agents, e.g. tamoxifen, mediates this release. A mechanism by which many of these drugs affect several diverse biologic properties including deesterification of membrane phospholipids by phospholipases to release AA is presented.
Reviewer agreement trends from four years of electronic submissions of conference abstract
Authors: Rowe Brian H , Strome Trevor L , Spooner Carol , Blitz Sandra , Grafstein Eric , Worster Andrew
Published: April 2006

Background

The purpose of this study was to determine the inter-rater agreement between reviewers on the quality of abstract submissions to an annual national scientific meeting (Canadian Association of Emergency Physicians; CAEP) to identify factors associated with low agreement.

Methods

All abstracts were submitted using an on-line system and assessed by three volunteer CAEP reviewers blinded to the abstracts' source. Reviewers used an on-line form specific for each type of study design to score abstracts based on nine criteria, each contributing from two to six points toward the total (maximum 24). The final score was determined to be the mean of the three reviewers' scores using Intraclass Correlation Coefficient (ICC).

Results

495 Abstracts were received electronically during the four-year period, 2001 – 2004, increasing from 94 abstracts in 2001 to 165 in 2004. The mean score for submitted abstracts over the four years was 14.4 (95% CI: 14.1–14.6). While there was no significant difference between mean total scores over the four years (p = 0.23), the ICC increased from fair (0.36; 95% CI: 0.24–0.49) to moderate (0.59; 95% CI: 0.50–0.68). Reviewers agreed less on individual criteria than on the total score in general, and less on subjective than objective criteria.

Conclusion

The correlation between reviewers' total scores suggests general recognition of "high quality" and "low quality" abstracts. Criteria based on the presence/absence of objective methodological parameters (i.e., blinding in a controlled clinical trial) resulted in higher inter-rater agreement than the more subjective and opinion-based criteria. In future abstract competitions, defining criteria more objectively so that reviewers can base their responses on empirical evidence may lead to increased consistency of scoring and, presumably, increased fairness to submitters.
"When patients have cancer, they stop seeing me" – the role of the general practitioner in early follow-up of patients with cancer – a qualitative study
Authors: Anvik Tor , Holtedahl Knut A , Mikalsen Hege
Published: April 2006

Background

The role of the general practitioner (GP) in cancer follow-up is poorly defined. We wanted to describe and analyse the role of the GP during initial follow-up of patients with recently treated cancer, from the perspective of patients, their relatives and their GPs.

Methods

One focus group interview with six GPs from the city of Bodø and individual interviews with 17 GPs from the city of Tromsø in North Norway. Text analysis of the transcribed interviews and of free text comments in two questionnaires from 91 patients with cancer diagnosed between October 1999 and September 2000 and their relatives from Tromsø.

Results

The role of the GP in follow-up of patients with recently treated cancer is discussed under five main headings: patient involvement, treating the cancer and treating the patient, time and accessibility, limits to competence, and the GP and the hospital should work together.

Conclusion

The GP has a place in the follow-up of many patients with cancer, also in the initial phase after treatment. Patients trust their GP to provide competent care, especially when they have more complex health care needs on top of their cancer. GPs agree to take a more prominent role for cancer patients, provided there is good access to specialist advice. Plans for follow-up of individual patients could in many cases improve care and cooperation. Such plans could be made preferably before discharge from in-patient care by a team consisting of the patient, a carer, a hospital specialist and a general practitioner. Patients and GPs call on hospital doctors to initiate such collaboration.
Blood test ordering for unexplained complaints in general practice: the VAMPIRE randomised clinical trial protocol. [ISRCTN55755886]
Authors: van Bokhoven Marloes A , Koch Hèlen , van der Weijden Trudy , Grol Richard PTM , Bindels Patrick JE , Dinant Geert-Jan
Published: April 2006

Background

General practitioners (GPs) frequently order blood tests when they see patients presenting with unexplained complaints. Due to the low prevalence of serious pathology in general practice, the risk of false-positive test results is relatively high. This may result in unnecessary further testing, leading to unfavourable effects such as patient anxiety, high costs, somatisation and morbidity. A policy of watchful waiting is expected to lower both the number of patients to be tested and the risk of false-positive test results, without missing serious pathology. However, many general practitioners experience barriers when trying to postpone blood testing by watchful waiting. The objectives of this study are (1) to determine the accuracy of blood tests in patients presenting with unexplained complaints in terms of detecting pathology, (2) to determine the accuracy of a watchful waiting strategy and (3) to determine the effects of a quality improvement strategy to promote the postponement of blood test ordering by GPs for patients with unexplained complaints.

Design

General practices are randomised over three groups. Group 1 is instructed to order blood tests immediately, group 2 to apply a watchful waiting policy and group 3 also to postpone testing, but supported by our quality improvement strategy. The trial consists of two sub-studies: a diagnostic study at patient level (group 1 versus groups 2 and 3) and a quality improvement study at GP level (group 2 versus group 3). The diagnostic strategy to be used involves of both customary and innovative tests. The quality improvement strategy consists of two small-group meetings and a practice outreach visit. Patient follow-up ends at 12 months after the initial consultation. Primary outcome measures are the accuracy and added value of blood tests for detecting pathology, the effect of a 4-week postponement of test ordering on the blood test characteristics and the quantity of tests ordered. Secondary outcome measures are the course of complaints, quality of life, satisfaction with care, anxiety of patients and practitioners, determinants of physicians' behaviour, health care utilisation and costs.

Discussion

The innovative aspect of this trial is that it combines a clinical-epidemiological study and a quality of care study.
A record review of reported musculoskeletal pain in an Ontario long term care facility
Authors: D'Astolfo Connie J , Humphreys B Kim
Published: April 2006

Background

Musculoskeletal (MSK) pain is one of the leading causes of chronic health problems in people over 65 years of age. Studies suggest that a high prevalence of older adults suffer from MSK pain (65% to 80%) and back pain (36% to 40%). The objectives of this study were:

1. To investigate the period prevalence of MSK pain and associated subgroups in residents of a long-term care (LTC) facility.

2. To describe clinical features associated with back pain in this population.

3. To identify associations between variables such as age, gender, cognitive status, ambulatory status, analgesic use, osteoporosis and osteoarthritis with back pain in a long-term care facility.

Methods

A retrospective chart review was conducted using a purposive sampling approach of residents' clinical charts from a LTC home in Toronto, Canada. All medical records for LTC residents from January 2003 until March 2005 were eligible for review. However, facility admissions of less than 6 months were excluded from the study to allow for an adequate time period for patient medical assessments and pain reporting/charting to have been completed. Clinical data was abstracted on a standardized form. Variables were chosen based on the literature and their suggested association with back pain and analyzed via multivariate logistic regression.

Results

140 (56%) charts were selected and reviewed. Sixty-nine percent of the selected residents were female with an average age of 83.7 years (51–101). Residents in the sample had a period pain prevalence of 64% (n = 89) with a 40% prevalence (n = 55) of MSK pain. Of those with a charted report of pain, 6% (n = 5) had head pain, 2% (n = 2) neck pain, 21% (n = 19) back pain, 33% (n = 29) extremity pain and 38% (n = 34) had non-descriptive/unidentified pain complaint. A multivariate logistic regression analysis revealed that osteoporosis was the only significant association with back pain from the variables studied (P = 0.001).

Conclusion

Residents with back pain represent 13.6% (n = 19) of the sample population studied. This is as frequent as other serious conditions commonly found in LTC. Of the variables studied, only osteoporosis and the self-report of back pain were found to be associated. The back pain resident in this facility can typically be described as female, osteoporotic, with mild to moderate dementia, an independent or assisted walker having low levels of depression. Further research using other sites is needed to determine the overall prevalence of this condition and its impact on quality of life issues. The results of this study should inform future research in this area.
Antibiotic susceptibility of Atopobium vaginae
Authors: De Backer Ellen , Verhelst Rita , Verstraelen Hans , Claeys Geert , Verschraegen Gerda , Temmerman Marleen , Vaneechoutte Mario
Published: April 2006

Background

Previous studies have indicated that a recently described anaerobic bacterium, Atopobium vaginae is associated with bacterial vaginosis (BV). Thus far the four isolates of this fastidious micro-organism were found to be highly resistant to metronidazole and susceptible for clindamycin, two antibiotics preferred for the treatment of BV.

Methods

Nine strains of Atopobium vaginae, four strains of Gardnerella vaginalis, two strains of Lactobacillus iners and one strain each of Bifidobacterium breve, B. longum, L. crispatus, L. gasseri and L. jensenii were tested against 15 antimicrobial agents using the Etest.

Results

All nine strains of A. vaginae were highly resistant to nalidixic acid and colistin while being inhibited by low concentrations of clindamycin (range: < 0.016 μg/ml), rifampicin (< 0.002 μg/ml), azithromycin (< 0.016 – 0.32 μg/ml), penicillin (0.008 – 0.25 μg/ml), ampicillin (< 0.016 – 0.94 μg/ml), ciprofloxacin (0.023 – 0.25 μg/ml) and linezolid (0.016 – 0.125 μg/ml). We found a variable susceptibility for metronidazole, ranging from 2 to more than 256 μg/ml. The four G. vaginalis strains were also susceptible for clindamycin (< 0.016 – 0.047 μg/ml) and three strains were susceptible to less than 1 μg/ml of metronidazole. All lactobacilli were resistant to metronidazole (> 256 μg/ml) but susceptible to clindamycin (0.023 – 0.125 μg/ml).

Conclusion

Clindamycin has higher activity against G. vaginalis and A. vaginae than metronidazole, but not all A. vaginae isolates are metronidazole resistant, as seemed to be a straightforward conclusion from previous studies on a more limited number of strains.
Neutropenia induced in outbred mice by a simplified low-dose cyclophosphamide regimen: characterization and applicability to diverse experimental models of infectious diseases
Authors: Zuluaga Andres F , Salazar Beatriz E , Rodriguez Carlos A , Zapata Ana X , Agudelo Maria , Vesga Omar
Published: April 2006

Background

For its low cost and ease of handling, the mouse remains the preferred experimental animal for preclinical tests. To avoid the interaction of the animal immune system, in vivo antibiotic pharmacodynamic studies often employ cyclophosphamide (CPM) to induce neutropenia. Although high doses (350–450 mg/kg) are still used and their effects on mouse leukocytes have been described, a lower dose (250 mg/kg) is widely preferred today, but the characteristics and applicability of this approach in outbred mice have not been determined.

Methods

Fifteen female ICR mice were injected intraperitoneally with 150 and 100 mg/kg of CPM on days 1 and 4, respectively. Blood samples (~160 μL) were drawn from the retro-orbital sinus of each mouse on days 1, 4, 5, 6, 7 and 11. Leukocytes were counted manually and the number of granulocytes was based on microscopic examination of Wright-stained smears. The impact of neutropenia induced by this method was then determined with a variety of pathogens in three different murine models of human infections: pneumonia (Klebsiella pneumoniae, Streptococcus pneumoniae, Staphylococcus aureus), meningoencephalitis (S. pneumoniae), and the thigh model (S. aureus, Escherichia coli, Bacteroides fragilis).

Results

The basal count of leukocytes was within the normal range for outbred mice. On day 4, there was an 84% reduction in total white blood cells, and by day 5 the leukopenia reached its nadir (370 ± 84 cells/mm3). Profound neutropenia (≤10 neutrophils/mm3) was demonstrated at day 4 and persisted through days 5 and 6. Lymphocytes and monocytes had a 92% and 96% decline between days 1 and 5, respectively. Leukocytes recovered completely by day 11. Mice immunosupressed under this protocol displayed clinical and microbiological patterns of progressive and lethal infectious diseases after inoculation in different organs with diverse human pathogens.

Conclusion

A CPM total dose of 250 mg/kg is sufficient to induce profound and sustained neutropenia (<10 neutrophils/mm3) at least during 3 days in outbred mice, is simpler than previously described methods, and allows successful induction of infection in a variety of experimental models.
Occurrence of invasive pneumococcal disease and number of excess cases due to influenza
Authors: Grabowska Katarzyna , Högberg Liselotte , Penttinen Pasi , Svensson Åke , Ekdahl Karl
Published: April 2006

Background

Influenza is characterized by seasonal outbreaks, often with a high rate of morbidity and mortality. It is also known to be a cause of significant amount secondary bacterial infections. Streptococcus pneumoniae is the main pathogen causing secondary bacterial pneumonia after influenza and subsequently, influenza could participate in acquiring Invasive Pneumococcal Disease (IPD).

Methods

In this study, we aim to investigate the relation between influenza and IPD by estimating the yearly excess of IPD cases due to influenza. For this purpose, we use influenza periods as an indicator for influenza activity as a risk factor in subsequent analysis. The statistical modeling has been made in two modes. First, we constructed two negative binomial regression models. For each model, we estimated the contribution of influenza in the models, and calculated number of excess number of IPD cases. Also, for each model, we investigated several lag time periods between influenza and IPD. Secondly, we constructed an "influenza free" baseline, and calculated differences in IPD data (observed cases) and baseline (expected cases), in order to estimate a yearly additional number of IPD cases due to influenza. Both modes were calculated using zero to four weeks lag time.

Results

The analysis shows a yearly increase of 72–118 IPD cases due to influenza, which corresponds to 6–10% per year or 12–20% per influenza season. Also, a lag time of one to three weeks appears to be of significant importance in the relation between IPD and influenza.

Conclusion

This epidemiological study confirms the association between influenza and IPD. Furthermore, negative binomial regression models can be used to calculate number of excess cases of IPD, related to influenza.
Deep neck infection complicating lymphadenitis caused by Streptococcus intermediusin an immunocompetent child
Authors: Rigante Donato , Spanu Teresa , Nanni Lorenzo , Tornesello Assunta , Sanguinetti Maurizio , D'Inzeo Tiziana , Stabile Achille , Fadda Giovanni
Published: April 2006

Background

Streptococcus intermediusbelongs to theStreptococcus anginosusgroup. It is part of the normal flora of the human mouth, but it can be etiologically associated with deep-site infections.

Case presentation

We present a case of deep neck infection complicatingStreptococcus intermediuslymphadenitis, which developed in an immunocompetent 14-year-old boy with a history of recent dental work. The infection was ultimately eradicated by a combined medical and surgical approach.

Conclusion

Our report suggests that combined medical and surgical therapy is essential for the complete resolution of deep infections caused byStreptococcus intermedius. Molecular biological techniques can be useful in guiding the diagnostic investigation and providing insight into the possibility of occult abscesses, which are particularly common withStreptococcus intermediusinfections.
Evaluation of real-time PCR for diagnosis of Bordetella pertussis infection
Authors: Knorr Laina , Fox Julie D , Tilley Peter AG , Ahmed-Bentley Jasmine
Published: April 2006

Background

Nucleic acid amplification of the IS481 region by PCR is more sensitive than culture for detection and diagnosis of Bordetella pertussis but the assay has known cross-reactivity for Bordetella holmesii and its use as a routine diagnostic assay has not been widely evaluated.

Methods

The objectives of this study were: 1) to assess the diagnostic utility of real-time IS481 PCR by comparison of results with culture and direct fluorescent antigen (DFA) testing for B. pertussis, 2) to employ a PCR assay designed against a different insertion sequence (IS1001) to assess the incidence of B. holmesii in symptomatic individuals and 3) to design and evaluate a new PCR-based assay which could be used for B. pertussis confirmation. A total of 808 nasopharyngeal specimens were included in the study the majority of which were submitted in charcoal transport medium (88%) with the rest submitted in Regan-Lowe medium.

Results

Concordant results for PCR, DFA and culture were obtained for 21 B. pertussis positive and 729 B. pertussis negative specimens. DFA was prone to false positive and negative reactions when compared with both PCR and culture. The IS481 PCR identified 28 positive results for specimens that were DFA and culture negative. A novel real-time PCR targeting the B. pertussis toxin promoter was found to be specific and useful for confirming the majority of IS481 positive specimens as B. pertussis. B. holmesii was not detected in any of the submitted samples.

Conclusion

The potential pick up of B. holmesii by the IS481 PCR had minimal diagnostic relevance in the Alberta population during the time period of our study. The IS481 PCR assay is now used in our laboratory routinely for front-line screening of samples for B. pertussis with associated enhancement in diagnostic sensitivity compared with DFA and culture. Retrospectively, patients' samples are batched and tested by the IS1001 MB and TPR assays for research purposes and to ensure there is no change in B. holmesii incidence in the population.
Cerebrospinal fluid HIV-1 RNA, intrathecal immunoactivation, and drug concentrations after treatment with a combination of saquinavir, nelfinavir, and two nucleoside analogues: the M61022 study
Authors: Yilmaz Aylin , Fuchs Dietmar , Hagberg Lars , Nillroth Ulrika , Ståhle Lars , Svensson Jan-Olof , Gisslén Magnus
Published: April 2006

Background

The way various antiretroviral drugs and drug combinations affect HIV-1 infection in the central nervous system is still largely unknown. The aim of this study was to determine the cerebrospinal fluid (CSF) steady-state concentrations of saquinavir and nelfinavir in relation to plasma concentrations, and to study their effect in combination with two nucleoside reverse transcriptase inhibitors (NRTIs) on CSF viral loads, intrathecal immunoactivation, and blood-brain barrier integrity.

Methods

Paired CSF and plasma samples from 8 antiretroviral-naïve HIV-1 infected patients starting combination therapy with saquinavir, nelfinavir, and two nucleoside analogues were collected prior to treatment, and again after approximately 12 and 48 weeks of antiretroviral therapy. Additional plasma samples were taken at weeks 2, 4, 8, 24, and 36. The concentrations of protease inhibitors were analysed, as were levels of HIV-1 RNA, CD4+ T-cell count, β2-microglobulin, neopterin, albumin ratio, IgG index, and monocytic cell count.

Results

None of the patients in the study presented with HIV-1 RNA < 50 copies/mL in CSF or plasma prior to treatment, compared to 5/7 at the end of the study. Signs of cell-mediated intrathecal immunoactivation, measured by neopterin and β2-microglobulin, decreased significantly in both CSF and serum, although only 1/7 reached normal CSF neopterin levels after 48 weeks of treatment. There was no significant reduction of albumin ratio, IgG index or CSF monocytic cell count. Saquinavir median (range) concentrations were < 2.5 (< 2.5–96.0) nM unbound in plasma, and < 2.5 (< 2.5–9.0) nM total in CSF. Nelfinavir median (range) concentrations were 10.0 (< 2.0–31.0) nM unbound in plasma, and < 2.0 (< 2.0–23.0) nM total in CSF. Saquinavir and nelfinavir were detectable in 7/15 and 9/15 CSF samples, respectively.

Conclusion

Saquinavir and nelfinavir, in combination with two NRTIs, decrease the CSF viral load and, to a lesser extent, intrathecal immunoactivation. We found reasonably high CSF concentrations of nelfinavir, but suboptimal concentrations of saquinavir.
Arthritis in adults with community-acquired bacterial meningitis: a prospective cohort study
Authors: Weisfelt Martijn , van de Beek Diederik , Spanjaard Lodewijk , de Gans Jan
Published: April 2006

Background

Although the coexistence of bacterial meningitis and arthritis has been noted in several studies, it remains unclear how often both conditions occur simultaneously.

Methods

We evaluated the presence of arthritis in a prospective nationwide cohort of 696 episodes of community-acquired bacterial meningitis, confirmed by culture of cerebrospinal fluid, which occurred in patients aged >16 years. The diagnosis of arthritis was based upon the judgment of the treating physician. To identify differences between groups Fisher exact statistics and the Mann-Whitney U test were used.

Results

Arthritis was recorded in 48 of 696 (7%) episodes of community-acquired bacterial meningitis in adults. Joint-fluid aspirations were performed in 23 of 48 patients (48%) and joint-fluid cultures yielded bacteria in 6 of 23 patients (26%). Arthritis occurred most frequently in patients with meningococcal meningitis (12%). Of the 48 patients with bacterial meningitis and coexisting arthritis, four died (8%) and 10 (23%) had residual joint symptoms.

Conclusion

Arthritis is a common manifestation in patients with community-acquired bacterial meningitis. Functional outcome of arthritis in bacterial meningitis is generally good because meningococcal arthritis is usually immune-mediated, and pneumococcal arthritis is generally less deforming than staphylococcal arthritis. Nevertheless, additional therapeutic measures should be considered if clinical course is complicated by arthritis. In patients with infectious arthritis prolonged antibiotic therapy is mandatory.