To evaluate the interobserver agreement for the Neoplastic Spine Instability Score (SINS) among spine surgeons with or without experience in vertebral metastasis treatment and physicians in other specialties.
Case descriptions were produced based on the medical records of 40 patients with vertebral metastases. The descriptions were then published online. Physicians were invited to evaluate the descriptions by answering questions according to the Neoplastic Spine Instability Score (SINS). The agreement among physicians was calculated using the kappa coefficient.
Seventeen physicians agreed to participate: three highly experienced spine surgeons, seven less-experienced spine surgeons, three surgeons of other specialties, and four general practitioners (n = 17). The agreement for the final SINS score among all participants was fair, and it varied according to the SINS component. The agreement was substantial for the spine location only. The agreement was higher among experienced surgeons. The agreement was nearly perfect for spinal location among the spine surgeons who were highly experienced in vertebral metastases.
This study demonstrates that the experience of the evaluator has an impact on SINS scale classification. The interobserver agreement was only fair among physicians who were not spine surgeons and among spine surgeons who were not experienced in the treatment of vertebral metastases, which may limit the use of the SINS scale for the screening of unstable lesions by less-experienced evaluators.
Spine; Health Services Research; Models, Statistical; Observer Variation
A leaching experiment, where liquid manure spiked with Salmonella enterica serovar Typhimurium (Tet+) DSM554 was applied to soil surfaces, was conducted on intact soil monoliths (60 cm in diameter and 100 cm long). A total of 6.5 × 1010 CFU was applied to each column. We found that Salmonella serovar Typhimurium could be transported to a 1-m depth in loamy soil at concentrations reaching 1.3 × 105 CFU/ml of leachate. The test strain was found in concentrations ranging from 300 to 1.35 cells/ml in loamy soil throughout the 27 days of the experiment, while concentrations below 20 cells/ml were sporadically detected in the leachates from sandy monoliths. Real-time PCR targeting invA DNA showed a clear correspondence between the total and culturable numbers of cells in the leachate, indicating that most cells leached were viable. On day 28, distribution of Salmonella serovar Typhimurium at five depths in the four monoliths was determined. The highest recovery rate, ranging from 1.5% to 3.8% of the total applied inoculum, was found in the top 0.2 m.
A bacterial community from Danish agricultural soil was enriched with linuron [N-(3,4-dichlorophenyl)-N′-methoxy-N′-methylurea] as the sole carbon and nitrogen source. The community mineralized [ring-U-14C]linuron completely to 14CO2 and 14C-biomass. Denaturing gradient gel electrophoresis analysis and cultivation revealed that a Variovorax sp. was responsible for the mineralization activity.
Stem cell factor (SCF), a key regulator of hematopoiesis, potently synergizes with a number of hematopoietic growth factors. However, little is known about growth factors capable of inhibiting the actions of SCF. TNF-alpha has been shown to act as a bidirectional regulator of myeloid cell proliferation and differentiation. This study was designed to examine interactions between TNF-alpha and SCF. Here, we demonstrate that TNF-alpha potently and directly inhibits SCF-stimulated proliferation of CD34+ hematopoietic progenitor cells. Furthermore, TNF-alpha blocked all colony formation stimulated by SCF in combination with granulocyte colony-stimulating factor (CSF) or CSF-1. The synergistic effect of SCF observed in combination with GM-CSF or IL-3 was also inhibited by TNF-alpha, resulting in colony numbers similar to those obtained in the absence of SCF. These effects of TNF-alpha were mediated through the p55 TNF receptor, whereas little or no inhibition was signaled through the p75 TNF receptor. Finally, TNF-alpha downregulated c-kit cell-surface expression on CD34+ bone marrow cells, and this was predominantly a p55 TNF receptor-mediated event as well.
Ethanol reassimilation in Pichia stipitis CBS 6054 was studied by using continuous cultures, and the oxidation of [1-13C]ethanol was monitored by in vivo and in vitro 13C nuclear magnetic resonance spectroscopy. Acetate was formed when ethanol was reassimilated. The ATP/ADP ratio and the carbon dioxide production decreased, whereas the malate dehydrogenase activity increased, in ethanol-reassimilating cells. The results are discussed in terms of the low ethanol tolerance in P. stipitis compared with that in Saccharomyces cerevisiae (S. W. Brown, S. G. Oliver, D. E. F. Harrison, and R. C. Righelato, Eur. J. Appl. Microbiol. Biotechnol. 11:151-155, 1981).
Recent research suggests that sleep disturbance, fatigue, and depressed mood form a symptom cluster in patients treated with chemotherapy. To date, however, no studies have examined lagged relationships among these symptoms during chemotherapy, a time when symptom variability is high. The aim of the current study was to examine lagged changes among daily symptoms during platinum-based chemotherapy.
Participants were 78 women with gynecologic cancer (mean age 63, SD=11; 91% Caucasian; 97% non-Hispanic). Sleep disturbance was assessed via wrist actigraphy, while fatigue and depressed mood were assessed via daily diary in the week after participants’ first chemotherapy infusion. Latent change score models (LCS) were used to examine lagged relationships between symptom pairs.
High levels of sleep disturbance (i.e., minutes awake at night) were associated with earlier subsequent peaks in fatigue, while high levels of fatigue were associated with higher subsequent levels of depressed mood.
These findings suggest that sleep disturbance, fatigue, and depressed mood occur in a cascade pattern during chemotherapy, in which increases in sleep disturbance contribute to fatigue, which in turn contributes to depressed mood. Interventions targeting symptoms early in the cascade, such as sleep disturbance, may provide benefits across multiple downstream symptoms.
neoplasms; gynecologic neoplasms; sleep; fatigue; depression
Candida albicans is a major fungal pathogen of humans. This yeast is carried by many individuals as a harmless commensal, but when immune defences are perturbed it causes mucosal infections (thrush). Additionally, when the immune system becomes severely compromised, C. albicans often causes life-threatening systemic infections. A battery of virulence factors and fitness attributes promote the pathogenicity of C. albicans. Fitness attributes include robust responses to local environmental stresses, the inactivation of which attenuates virulence. Stress signalling pathways in C. albicans include evolutionarily conserved modules. However, there has been rewiring of some stress regulatory circuitry such that the roles of a number of regulators in C. albicans have diverged relative to the benign model yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe. This reflects the specific evolution of C. albicans as an opportunistic pathogen obligately associated with warm-blooded animals, compared with other yeasts that are found across diverse environmental niches. Our understanding of C. albicans stress signalling is based primarily on the in vitro responses of glucose-grown cells to individual stresses. However, in vivo this pathogen occupies complex and dynamic host niches characterised by alternative carbon sources and simultaneous exposure to combinations of stresses (rather than individual stresses). It has become apparent that changes in carbon source strongly influence stress resistance, and that some combinatorial stresses exert non-additive effects upon C. albicans. These effects, which are relevant to fungus–host interactions during disease progression, are mediated by multiple mechanisms that include signalling and chemical crosstalk, stress pathway interference and a biological transistor.
Candida albicans; Fungal pathogenicity; Heat shock; Oxidative stress; Nitrosative stress; Osmotic stress; Cationic stress; Stress adaptation; Carbon metabolism
There is increasing evidence that global climate change will alter the spatiotemporal occurrences and abundances of many species at continental scales. This will have implications for efficient conservation of biodiversity. We investigate if the general public in Denmark are willing to pay for the preservation of birds potentially immigrating and establishing breeding populations due to climate change to the same extent that they are for native species populations currently breeding in Denmark, but potentially emigrating due to climate change. We find that Danish citizens are willing to pay much more for the conservation of birds currently native to Denmark, than for bird species moving into the country – even when they are informed about the potential range shifts associated with climate change. The only exception is when immigrating species populations are under pressure at European level. Furthermore, people believing climate change to be man-made and people more knowledgeable about birds tended to have higher WTP for conservation of native species, relative to other people, whereas their preferences for conserving immigrant species generally resembled those of other people. Conservation investments rely heavily on public funding and hence on public support. Our results suggest that cross-country coordination of conservation efforts under climate change will be challenging in terms of achieving an appropriate balance between cost-effectiveness in adaptation and the concerns of a general public who seem mostly worried about protecting currently-native species.
To investigate the use of manual vacuum aspiration in postabortion care in Malawi between 2008–2012.
A retrospective cross-sectional study was done at the referral hospital Queen Elisabeth Central Hospital, and the two district hospitals of Chiradzulu and Mangochi. The data were collected simultaneously at the three sites from Feb-March 2013. All records available for women admitted to the gynaecological ward from 2008-2012 were reviewed. Women who had undergone surgical uterine evacuation after incomplete abortion were included and the use of manual vacuum aspiration versus sharp curettage was analysed.
Altogether, 5121 women were included. One third (34.2%) of first trimester abortions were treated with manual vacuum aspiration, while all others were treated with sharp curettage. There were significant differences between the hospitals and between years. Overall there was an increase in the use of manual vacuum aspiration from 2008 (19.7%) to 2009 (31.0%), with a rapid decline after 2010 (28.5%) ending at only 4.9% in 2012. Conversely there was an increase in use of sharp curettage in all hospitals from 2010 to 2012.
Use of manual vacuum aspiration as part of the postabortion care in Malawi is rather low, and decreased from 2010 to 2012, while the use of sharp curettage became more frequent. This is in contrast with current international guidelines.
Twenty-four individuals with transtibial amputation were recruited to a randomized, crossover design study to examine stride-to-stride fluctuations of lower limb joint flexion/extension time series using the largest Lyapunov exponent (λ). Each individual wore a “more appropriate” and a “less appropriate” prosthesis design based on the subject's previous functional classification for a three week adaptation period. Results showed decreased λ for the sound ankle compared to the prosthetic ankle (F1,23 = 13.897, p = 0.001) and a decreased λ for the “more appropriate” prosthesis (F1,23 = 4.849, p = 0.038). There was also a significant effect for the time point in the adaptation period (F2,46 = 3.164, p = 0.050). Through the adaptation period, a freezing and subsequent freeing of dynamic degrees of freedom was seen as the λ at the ankle decreased at the midpoint of the adaptation period compared to the initial prosthesis fitting (p = 0.032), but then increased at the end compared to the midpoint (p = 0.042). No differences were seen between the initial fitting and the end of the adaptation for λ (p = 0.577). It is concluded that the λ may be a feasible clinical tool for measuring prosthesis functionality and adaptation to a new prosthesis is a process through which the motor control develops mastery of redundant degrees of freedom present in the system.
A highly enantioselective addition of phenyl carbamate to meso-epoxides has been developed to efficiently generate protected trans-1,2-amino alcohols. This transformation is promoted by an oligomeric (salen)Co–OTf catalyst and has been used to prepare two useful 2-aminocycloalkanol hydrochlorides in enantiopure formon a multigram scale from commercially-available starting materials.
The two North Atlantic eel species, the European and the American eel, represent an ideal system in which to study parallel selection patterns due to their sister species status and the presence of ongoing gene flow. A panel of 80 coding-gene SNPs previously analyzed in American eel was used to genotype European eel individuals (glass eels) from 8 sampling locations across the species distribution. We tested for single-generation signatures of spatially varying selection in European eel by searching for elevated genetic differentiation using FST-based outlier tests and by testing for significant associations between allele frequencies and environmental variables.
We found signatures of possible selection at a total of 11 coding-gene SNPs. Candidate genes for local selection constituted mainly genes with a major role in metabolism as well as defense genes. Contrary to what has been found for American eel, only 2 SNPs in our study correlated with differences in temperature, which suggests that other explanatory variables may play a role. None of the genes found to be associated with explanatory variables in European eel showed any correlations with environmental factors in the previous study in American eel.
The different signatures of selection between species could be due to distinct selective pressures associated with the much longer larval migration for European eel relative to American eel. The lack of parallel selection in North Atlantic eels could also be due to most phenotypic traits being polygenic, thus reducing the likelihood of selection acting on the same genes in both species.
Adaptation; European eel; Genetic-by-environment associations; Parallel selection; Single nucleotide polymorphisms
The opportunistic fungal pathogen Candida glabrata is a frequent cause of candidiasis, causing infections ranging from superficial to life-threatening disseminated disease. The inherent tolerance of C. glabrata to azole drugs makes this pathogen a serious clinical threat. To identify novel genes implicated in antifungal drug tolerance, we have constructed a large-scale C. glabrata deletion library consisting of 619 unique, individually bar-coded mutant strains, each lacking one specific gene, all together representing almost 12% of the genome. Functional analysis of this library in a series of phenotypic and fitness assays identified numerous genes required for growth of C. glabrata under normal or specific stress conditions, as well as a number of novel genes involved in tolerance to clinically important antifungal drugs such as azoles and echinocandins. We identified 38 deletion strains displaying strongly increased susceptibility to caspofungin, 28 of which encoding proteins that have not previously been linked to echinocandin tolerance. Our results demonstrate the potential of the C. glabrata mutant collection as a valuable resource in functional genomics studies of this important fungal pathogen of humans, and to facilitate the identification of putative novel antifungal drug target and virulence genes.
Clinical infections by the yeast-like pathogen Candida glabrata have been ever-increasing over the past years. Importantly, C. glabrata is one of the most prevalent causes of drug-refractory fungal infections in humans. We have generated a novel large-scale collection encompassing 619 bar-coded C. glabrata mutants, each lacking a single gene. Extensive profiling of phenotypes reveals a number of novel genes implicated in tolerance to antifungal drugs that interfere with proper cell wall function, as well as genes affecting fitness of C. glabrata both during normal growth and under environmental stress. This fungal deletion collection will be a valuable resource for the community to study mechanisms of virulence and antifungal drug tolerance in C. glabrata, which is particularly relevant in view of the increasing prevalence of infections caused by this important human fungal pathogen.
Ensuring adherence to treatment and retention is important in clinical trials, particularly in remote areas and minority groups. We describe a novel approach to improve adherence, retention and clinical review rates of Indigenous children.
This descriptive study was nested within a placebo-controlled, randomised trial (RCT) on weekly azithromycin (or placebo) for 3-weeks. Indigenous children aged ≤24-months hospitalised with acute bronchiolitis were recruited from two tertiary hospitals in northern Australia (Darwin and Townsville). Using mobile phones embedded within a culturally-sensitive approach and framework, we report our strategies used and results obtained. Our main outcome measure was rates of adherence to medications, retention in the RCT and self-presentation (with child) to clinic for a clinical review on day-21.
Of 301 eligible children, 76 (21%) families declined participation and 39 (13%) did not have access to a mobile phone. 186 Indigenous children were randomised and received dose one under supervision in hospital. Subsequently, 182 (99%) children received dose two (day-7), 169 (93%) dose three (day-14) and 180 (97%) attended their clinical review (day-21). A median of 2 calls (IQR 1–3) were needed to verify adherence. Importantly, over 97% of children remained in the RCT until their clinical endpoint at day-21.
In our setting, the use of mobile phones within an Indigenous-appropriate framework has been an effective strategy to support a clinical trial involving Australian Indigenous children in urban and remote Australia. Further research is required to explore other applications of this approach, including the impact on clinical outcomes.
ACTRN12608000150347 (RCT component).
Mobile phones; SMS; Adherence; Randomised controlled trial; ALRTI; Bronchiolitis; Indigenous
Intervertebral disc degeneration is accompanied by elevated levels of inflammatory cytokines that have been implicated in disease etiology and matrix degradation. While the effects of inflammatory stimulation on disc cell metabolism have been well-studied, their effects on cell biophysical properties have not been investigated. The hypothesis of this study is that inflammatory stimulation alters the biomechanical properties of isolated disc cells and volume responses to step osmotic loading. Cells from the nucleus pulposus (NP) of bovine discs were isolated and treated with either lipopolysaccharide (LPS), an inflammatory ligand, or with the recombinant cytokine TNF-α for 24 hours. We measured cellular volume regulation responses to osmotic loading either immediately after stimulation or after a 1 week recovery period from the inflammatory stimuli. Cells from each group were tested under step osmotic loading and the transient volume-response was captured via time-lapse microscopy. Volume-responses were analyzed using mixture theory framework to investigate two biomechanical properties of the cell, the intracellular water content and the hydraulic permeability. Intracellular water content did not vary between treatment groups, but hydraulic permeability increased significantly with inflammatory treatment. In the 1 week recovery group, hydraulic permeability remained elevated relative to the untreated recovery control. Cell radius was also significantly increased both after 24 hours of treatment and after 1 week recovery. A significant linear correlation was observed between hydraulic permeability and cell radius in untreated cells at 24 hours and at 1-week recovery, though not in the inflammatory stimulated groups at either time point. This loss of correlation between cell size and hydraulic permeability suggests that regulation of volume change is disrupted irreversibly due to inflammatory stimulation. Inflammatory treated cells exhibited altered F-actin cytoskeleton expression relative to untreated cells. We also found a significant decrease in the expression of aquaporin-1, the predominant water channel in disc NP cells, with inflammatory stimulation. To our knowledge, this is the first study providing evidence that inflammatory stimulation directly alters the mechanobiology of NP cells. The cellular biophysical changes observed in this study are coincident with documented changes in the extracellular matrix induced by inflammation, and may be important in disease etiology.
Motivation: Large-scale phenotyping projects such as the Sanger Mouse
Genetics project are ongoing efforts to help identify the influences of genes and their
modification on phenotypes. Gene–phenotype relations are crucial to the improvement
of our understanding of human heritable diseases as well as the development of drugs.
However, given that there are ∼20 000 genes in higher vertebrate genomes
and the experimental verification of gene–phenotype relations requires a lot of
resources, methods are needed that determine good candidates for testing.
Results: In this study, we applied an association rule mining approach to
the identification of promising secondary phenotype candidates. The predictions rely on a
large gene–phenotype annotation set that is used to find occurrence patterns of
phenotypes. Applying an association rule mining approach, we could identify 1967 secondary
phenotype hypotheses that cover 244 genes and 136 phenotypes. Using two automated and one
manual evaluation strategies, we demonstrate that the secondary phenotype candidates
possess biological relevance to the genes they are predicted for. From the results we
conclude that the predicted secondary phenotypes constitute good candidates to be
experimentally tested and confirmed.
Availability: The secondary phenotype candidates can be browsed through at
firstname.lastname@example.org or email@example.com
Supplementary data are available at Bioinformatics
Lack of consensus regarding how to identify cancer patients with significant fatigue has hampered research regarding cancer-related fatigue (CRF).
Specific criteria were used to identify CRF cases in women with stage 0-II breast cancer (BC group, n = 304). Women completed assessments before adjuvant therapy (baseline), end of adjuvant therapy (Post-Tx), and 6 and 42 months after end of adjuvant therapy (6 and 42 Month Post-Tx). At each, women completed a clinical interview and questionnaires assessing physical and mental health. A healthy control (HC) group with no history of BC (n = 337) completed 2 similar assessments 36 months apart.
Off-treatment CRF prevalence was 9% and 13% at the 6 and 42 Month Post-Tx assessments, respectively. Thus, 15% of the sample evidenced off-treatment CRF with 7% evidencing delayed onset CRF. CRF at the 6 Month Post-Tx assessment was associated only with CRF at baseline (OR = 3.2) and Post-Tx assessments (OR = 3.9). CRF at the 42 Month Post-Tx assessment was associated with CRF at the Post-Tx assessment (OR = 6.1), obesity at baseline, and several baseline measures of coping in response to fatigue. Off-treatment CRF cases differed markedly from CRF noncases and healthy controls on a spectrum of health status indices (mean effect size >1.0 SD).
Results document the prevalence of off-treatment and delayed onset CRF, suggest the utility of a cognitive-behavioral model of CRF, and support NCCN guidelines recommending monitoring fatigue across the cancer trajectory.
fatigue; survivorship; assessment; outcomes; late effects
Spectrins and plakins are important communicators linking cytoskeletal components to each other and to cellular junctions. Microtubule-actin cross-linking factor 1 (MACF1) belongs to the spectraplakin family and is involved in control of microtubule dynamics. Complete knock out of MACF1 in mice is associated with developmental retardation and embryonic lethality. Here we present a family with a novel neuromuscular condition. Genetic analyses show a heterozygous duplication resulting in reduced MACF1 gene product. The functional consequence is affected motility observed as periodic hypotonia, lax muscles and diminished motor skills, with heterogeneous presentation among the affected family members. To corroborate these findings we used RNA interference to knock down the VAB-10 locus containing the MACF1 homologue in C. elegans, and we could show that this also causes movement disturbances. These findings suggest that changes in the MACF1 gene is implicated in this neuromuscular condition, which is an important observation since MACF1 has not previously been associated with any human disease and thus presents a key to understanding the essential nature of this gene.
To estimate the overall and age-specific associations between obesity
and extremity musculoskeletal injuries and pain in children.
This cross-sectional study used information from electronic medical
records of 913 178 patients aged 2–19 years enrolled in an
integrated health plan in the period 2007–2009. Children were
classified as underweight, normal weight, overweight, or
moderately/extremely obese and, using multivariable logistic regression
methods, the associations between weight class and diagnosis of upper or
lower extremity fractures, sprains, dislocations and pain were
Overweight (OR 1.18, 95% CI 1.15 to 1.20), moderately obese
(OR 1.24, 95% CI 1.20 to 1.27) and extremely obese (OR 1.34,
95% CI 1.30 to 1.39) children had statistically significantly higher
odds of lower extremity injuries/pain compared to normal weight, adjusted
for sex, age, race/ethnicity and insurance status. Age-stratified analyses
yielded similar results. No consistent association was observed between body
mass index and injuries/pain of the upper extremities.
Greater body mass index is associated with increased odds of lower
extremity injuries and pain issues. Because the benefits of physical
activity may still outweigh the risk of injury, attention should be paid to
injury prevention strategies for these children at greater risk for lower
Pregnancy associated breast cancers (PABC) generally present at advanced stages and have a poor prognosis. The reasons are unclear but we hypothesized that the continuous high levels of estrogens and progesterone were involved. We have now carried out a detailed analysis of PABC compared to tumors of age-matched non-pregnant (Non-PABC) women.
Malignant epithelia and tumor-associated stroma of PABC and Non-PABC were isolated by laser capture microdissection and gene expression profiled. Additionally, normal breast epithelia and stroma adjacent to the two tumor types were analyzed. Lastly, subsets of previously identified E- and P-regulated genes were defined in all tissues.
We find that PABC signatures cluster with established breast cancer subtypes. Major hormone-regulated genes whose expression correlated with epithelia of PABC dealt with regulation of cell proliferation, metabolism and tumor aggressiveness, including genes used to predict tumor recurrence. Compared to normal epithelia, a significant number of genes associated with cell cycle processes were enriched in PABC, many of which are hormone regulated. Thus compared to normal epithelia, many of the genes that were differentially expressed in epithelia of PABC were distinct from those differentially expressed in Non-PABC. With regard to the tumor microenvironment, immune-related genes were enriched in tumor-associated stroma of PABC. Compared to normal stroma, PABC-associated stroma overexpressed immune response genes, while genes involved in angiogenesis and extracellular matrix deposition were more commonly downregulated. This suggests that the heightened aggressiveness of PABC may involve a predisposition to metastasis through extracellular matrix degradation, plus angiogenesis independence. Moreover, genes encoding cell proliferative factors, signaling, immunomodulators and cell death, were hormone regulated in stroma.
In sum, these analyses demonstrate complex patterns of enrichment and hormonal regulation of genes in PABC and suggest that it may have a distinct biological nature.
breast cancer; pregnancy; expression profiling; estrogen; stroma vs. epithelium
During August 2010–December 2012, we conducted a study of patients in Ghana who had Buruli ulcer, caused by Mycobacterium ulcerans, and found that 23% were co-infected with Mansonella perstans nematodes; 13% of controls also had M. perstans infection. M. perstans co-infection should be considered in the diagnosis and treatment of Buruli ulcer.
Mycobacterium ulcerans; Buruli ulcer; Mansonella perstans; co-infection; bacteria; Ghana; parasites; nematodes; filariae
Phytophthora infestans, causing late blight in potato, remains one of the most devastating pathogens in potato production and late blight resistance is a top priority in potato breeding. The introduction of multiple resistance (R) genes with different spectra from crossable species into potato varieties is required. Cisgenesis is a promising approach that introduces native genes from the crops own gene pool using GM technology, thereby retaining favourable characteristics of established varieties.
We pursued a cisgenesis approach to introduce two broad spectrum potato late blight R genes, Rpi-sto1 and Rpi-vnt1.1 from the crossable species Solanum stoloniferum and Solanum venturii, respectively, into three different potato varieties. First, single R gene-containing transgenic plants were produced for all varieties to be used as references for the resistance levels and spectra to be expected in the respective genetic backgrounds. Next, a construct containing both cisgenic late blight R genes (Rpi-vnt1.1 and Rpi-sto1), but lacking the bacterial kanamycin resistance selection marker (NPTII) was transformed to the three selected potato varieties using Agrobacterium-mediated transformation. Gene transfer events were selected by PCR among regenerated shoots. Through further analyses involving morphological evaluations in the greenhouse, responsiveness to Avr genes and late blight resistance in detached leaf assays, the selection was narrowed down to eight independent events. These cisgenic events were selected because they showed broad spectrum late blight resistance due to the activity of both introduced R genes. The marker-free transformation was compared to kanamycin resistance assisted transformation in terms of T-DNA and vector backbone integration frequency. Also, differences in regeneration time and genotype dependency were evaluated.
We developed a marker-free transformation pipeline to select potato plants functionally expressing a stack of late blight R genes. Marker-free transformation is less genotype dependent and less prone to vector backbone integration as compared to marker-assisted transformation. Thereby, this study provides an important tool for the successful deployment of R genes in agriculture and contributes to the production of potentially durable late blight resistant potatoes.
Potato; Late blight; Resistance gene; Cisgenesis; Marker-free transformation
Breast self-examinations (BSE) and skin self-examinations (SSE) represent cost-effective and time-efficient approaches to cancer detections. Given their utility, it is important to determine who is likely to perform these behaviors regularly and why. Because BSE and SSE require close examination of one's body, women who are less satisfied and less comfortable with their bodies may perform these behaviors less often. This study sought to determine if a relationship exists between body image and BSE and SSE behaviors and intentions. Ninety-three women completed measures assessing body image, past performance of and future intentions to perform BSE and SSE. Results indicated that body image was related to past performance of SSE. Having greater satisfaction with overall appearance and evaluating oneself as more attractive were related to having performed SSE more frequently in the past year. Future research should further examine this relationship utilizing longitudinal designs and more diverse populations.
Studies have reported that heparin may be unsuitable as an anticoagulant in human plasma samples when quantifying cytokines using multiplex bead array assays. For mouse samples, multiplex assays have been validated for serum and EDTA-plasma, but it remains to be elucidated whether heparin influences the quantification of cytokines, and if so – to what extent. Furthermore, laboratory mice are often anesthetized for blood sampling, which causes acute stress that may influence circulating cytokine concentrations and thus bias experimental results. The objectives of the present study were to identify whether specific cytokine concentrations varied between heparin-plasma, serum, and EDTA-plasma, and whether short isoflurane anesthesia would influence the concentrations of these cytokines in the circulation. Twenty-three acute phase and pro-inflammatory cytokines were quantified in matched serum, EDTA-plasma, and heparin-plasma samples from anesthetized and unanesthetized male NMRI mice using a multiplex assay. In addition, samples from unanesthetized mice were spiked with three levels of heparin.
The concentrations of five out of 23 cytokines were significantly different between sample types, but only one cytokine (IL-17A) differed between heparin-plasma and serum. When further spiking the heparin-plasma with increasing concentrations of heparin, there was a significant effect on 11 cytokines, where the cytokine recovery could be correlated to the heparin concentration for ten of these cytokines. Anesthesia resulted in lower concentrations of G-CSF, but had no significant impact on the concentrations of the other 22 cytokines.
In mice, heparin seems like a suitable anticoagulant for obtaining plasma for multiplex assays for the cytokines IL-1α, IL-1β, IL-2, IL-6, IL-9, IL-12p40, IL-12p70, IL-13, G-CSF, GM-CSF, IFN-γ, KC, MCP-1, MIP-1α, MIP-1β, RANTES and TNFα, but an effect of heparin in high concentrations should be considered for the cytokines IL-9, IL-12p40, IL-12p70, KC, MCP-1, MIP-1β and RANTES. Short isoflurane anesthesia had significant impact on G-CSF, but none of the other cytokines.
Multiplex assay; Anticoagulant; Isoflurane; Stress; Laboratory mice