Association of paratuberculosis sero-status with milk production and somatic cell counts across 5 lactations, using multilevel mixed models, in dairy cows.

The aim of this work was to investigate associations between individual cow Mycobacterium avium ssp. paratuberculosis (MAP) seropositivity, 305-d corrected milk production, and somatic cell count during 5 lactations lifespan in Portuguese dairy herds using multilevel mixed models. We used MAP serum ELISA (Idexx MAP Ac, Idexx Laboratories Inc., Westbrook, ME) results (n = 23,960) from all the 20,221 adult cows present in 329 farms and corresponding 47,586 lactation records from the National Dairy Improvement Association. Cows and farms were classified as positive or negative. Multilevel mixed models were used to investigate the association of cow MAP status with variation in milk production and somatic cell count. Cow MAP status, farm status, and lactation number were considered as independent variables. A quadratic function of lactation number was used to mimic the effect of lactation order on milk production. The models considered 3 levels: measurement occasion (level 1) within cow (level 2) and cow within farm (level 3). Four final models were produced, including all herds and cows, to address the effect of farm status (models 1 and 2) or the effect of cow status (models 3 and 4) on the outcome variables. Our results show that MAP status affects milk production. Losses are detectable from third lactation onward. During the first 5 lactations, positive cows accumulated an average loss of 1,284.8 kg of milk when compared with the negative cows. We also observed that somatic cell counts were higher in positive cows and a positive interaction occurs between cow status and lactation number, suggesting a positive association between MAP infection and increased somatic cell counts. Our results are in line with previous studies, suggesting a possible positive relation between cow milk production and susceptibility to MAP infection.

The interactions between municipal socioeconomic status and age on hip fracture risk.

SUMMARY: Age modifies the effect of area-level socioeconomic status (SES) in the risk of fragility hip fractures (HF). For older individuals, the risk of HF increases as SES increases. For younger individuals, risk of HF increases as SES decreases. Our study may help decision-makers to better direct the implementation of political decisions. INTRODUCTION: The effect of socioeconomic status (SES) on hip fracture (HF) incidence remains unclear. The objective of this study is to evaluate the association between HF incidence and municipality-level SES as well as interactions between age and SES. METHODS: From the Portuguese Hospital Discharge Database, we selected hospitalizations (2000-2010) of patients aged 50+, with HF diagnosis (codes 820.x, ICD9-CM), caused by traumas of low/moderate energy, excluding bone cancer cases and readmissions for aftercare. Municipalities were classified according to SES (deprived to affluent) using 2001 Census data. A spatial Bayesian hierarchical regression model (controlling for data heterogeneity and spatial autocorrelation), using the Poisson distribution, was used to quantify the relative risk (RR) of HF, 95% credible interval (95%CrI), and analyze the interaction between age and SES after adjusting for rural conditions. RESULTS: There were 96,905 HF, 77.3% of which were on women who, on average, were older than men (mean age 81.2±8.5 vs 78.2±10.1 years) at admission (p<0.001). In women, there was a lower risk associated with better SES: RR=0.83 (95%CrI 0.65-1.00) for affluent versus deprived. There was an inverse association between SES and HF incidence rate in the youngest and a direct association in the oldest, for both sexes, but significant only between deprived and affluent in older ages (≥75 years). CONCLUSIONS: Interaction between SES and age may be due to inequalities in lifestyles, access to health systems, and preventive actions. These results may help decision-makers to better understand the epidemiology of hip fractures and to better direct the available funding.

Epicardial adipose tissue and coronary artery calcification in psoriasis patients.

BACKGROUND: Psoriasis is a chronic, immune-mediated disease associated with several cardio-metabolic comorbidities, accelerated atherosclerosis and cardiovascular disease (CVD). Other causes beyond systemic inflammation and traditional cardiovascular risk factors (CVRF) may be implicated in the increased risk of CVD observed in these patients. Epicardial adipose tissue (EAT), a type of visceral adipose tissue surrounding the heart and coronary vessels has been implicated in the development of coronary artery disease, by endocrine mechanisms, but particularly by local inflammation. OBJECTIVE: To compare EAT volumes in psoriasis patients and controls using multidetector computed tomography (MDCT) and to analyse if eventual differences were independent from abdominal visceral adiposity; to determine, within psoriasis patients, its relation with subclinical atherosclerosis and other markers of cardiometabolic risk. METHODS: One hundred patients with severe psoriasis, without CVD underwent MDCT, with EAT and abdominal visceral fat (AVF) assessment and coronary artery calcification (CAC) quantification and were compared with 202 control patients. RESULTS: EAT volume was increased in psoriasis patients compared to control subjects, independently from age, sex and AVF, being, on average, 15.2 ± 4.41 mL higher (95% CI: 6.5-26.0, P = 0.001) than in controls. Moreover, psoriasis patients had a statistically significant higher risk of having subclinical atherosclerosis (OR 2.52, 95% CI: 1.23-5.16) than controls, after adjusting for traditional CVRF. Within psoriasis patients EAT volume was associated with subclinical atherosclerosis, independently of age, sex, psoriasis duration, classical CVRF and AVF. CONCLUSION: This study showed that psoriasis was associated with increased EAT volume independently of visceral abdominal fat and with subclinical atherosclerosis. Within psoriasis patients EAT volume was independently associated with CAC. EAT may be another important contributor to the higher cardiovascular risk observed in psoriasis.

Antigen-triggered interferon-γ and interleukin-10 pattern in cured mucosal leishmaniasis patients is shaped during the active phase of disease.

An exacerbated type 1 response to leishmanial antigens is the basis of tissue destruction observed in mucosal leishmaniasis (ML). After therapy, a persistent production of high levels of inflammatory cytokines can confer a poor prognosis. Herein we investigated whether the clinical conditions defined during the active phase of ML affect the magnitude of long-term anti-Leishmania immune response. Twenty clinically cured ML cases were studied. Peripheral blood mononuclear cells (PBMC) were cultured with L. braziliensis antigens (Lb-Ag), Toxoplasma gondii antigens (Tg-Ag), concanavalin-A (Con-A) or medium alone, and the lymphocyte proliferative response and cytokine secretion were quantified. Medical records were reviewed for Montenegro skin test (MST) during diagnosis, duration of ML disease or time elapsed after clinical cure. The duration of disease was correlated positively with MST (r = 0·61). Lb-Ag induced interferon (IFN)-γ was correlated positively with duration of illness (r = 0·69) as well as the frequency of secreting cells [enzyme-linked immunospot (ELISPOT)] assay. No association was observed for Tg-Ag or Con-A. Disease duration was correlated negatively with interleukin (IL)-10 production (r = -0·76). Moreover, a negative correlation between length of time after clinical cure and TNF levels (r = -0·94) or the IFN-γ : IL-10 ratio (r = -0·89) were also seen. We suggest that the magnitude of the IFN-γ inflammatory response triggered by ML can be driven by the time of leishmanial antigens exposition during the active phase of the disease. This pattern could persist even long-term after cure. However, despite IFN-γ levels, the decrease of the TNF and IFN-γ : IL-10 ratio reflects the control of proinflammatory responses achieved by cure of ML, possibly preventing disease relapses.

Strategies to work with HLA data in human populations for histocompatibility, clinical transplantation, epidemiology and population genetics: HLA-NET methodological recommendations.

HLA-NET (a European COST Action) aims at networking researchers working in bone marrow transplantation, epidemiology and population genetics to improve the molecular characterization of the HLA genetic diversity of human populations, with an expected strong impact on both public health and fundamental research. Such improvements involve finding consensual strategies to characterize human populations and samples and report HLA molecular typings and ambiguities; proposing user-friendly access to databases and computer tools and defining minimal requirements related to ethical aspects. The overall outcome is the provision of population genetic characterizations and comparisons in a standard way by all interested laboratories. This article reports the recommendations of four working groups (WG1-4) of the HLA-NET network at the mid-term of its activities. WG1 (Population definitions and sampling strategies for population genetics' analyses) recommends avoiding outdated racial classifications and population names (e.g. 'Caucasian') and using instead geographic and/or cultural (e.g. linguistic) criteria to describe human populations (e.g. 'pan-European'). A standard 'HLA-NET POPULATION DATA QUESTIONNAIRE' has been finalized and is available for the whole HLA community. WG2 (HLA typing standards for population genetics analyses) recommends retaining maximal information when reporting HLA typing results. Rather than using the National Marrow Donor Program coding system, all ambiguities should be provided by listing all allele pairs required to explain each genotype, according to the formats proposed in 'HLA-NET GUIDELINES FOR REPORTING HLA TYPINGS'. The group also suggests taking into account a preliminary list of alleles defined by polymorphisms outside the peptide-binding sites that may affect population genetic statistics because of significant frequencies. WG3 (Bioinformatic strategies for HLA population data storage and analysis) recommends the use of programs capable of dealing with ambiguous data, such as the 'gene[rate]' computer tools to estimate frequencies, test for Hardy-Weinberg equilibrium and selective neutrality on data containing any number and kind of ambiguities. WG4 (Ethical issues) proposes to adopt thorough general principles for any HLA population study to ensure that it conforms to (inter)national legislation or recommendations/guidelines. All HLA-NET guidelines and tools are available through its website http://hla-net.eu.

SNP identification and polymorphism analysis in exon 2 of the horse myostatin gene.

The myostatin gene (MSTN) belongs to the TGF-ß superfamily of secreted growth and differentiation factors and is responsible for embryonic and adult skeletal muscle development. In this study, exon 2 of the MSTN gene, which encodes part of the TGF-ß pro-peptide, was sequenced in 332 horses of 20 different breeds and compared with the horse MSTN gene sequence deposited in GenBank. The sequences obtained revealed the presence of 11 haplotypes represented by 10 variable nucleotide mutations, eight of them corresponding to amino acid sequence changes. This gene shows a high variability when compared with other genes. This might be an indication that some breeds have the same ancestry but different pressures of selection.

A role for endothelin receptor type A in migraine without aura susceptibility? A study in Portuguese patients.

BACKGROUND AND PURPOSE: Migraine is a common neurological disabling disorder, and anomalies of vascular function have been implied in its pathophysiology. Several findings point to a possible role of the endothelin receptor type A (EDNRA) in migraine. We aim to assess the involvement of endothelin receptor type A (EDNRA) in migraine susceptibility in a sample of Portuguese migraineurs. METHODS: Three tagging SNPs (rs702757, rs5333 and rs5335) were analysed in 188 cases - 111 without aura (MO) and 77 with aura (MA) - and 287 controls. A multivariable logistic regression was performed, including the three SNPs, adjusted for gender. Allelic and haplotypic frequencies were compared between cases and controls. Significant or promising results were confirmed by a multifactor dimensionality reduction analysis (MDR). RESULTS: We found a nominal association for the rs702757 T-allele [odds ratio (OR) = 1.44, 95% confidence intervals (CI): 1.05-1.99] and for the TT-genotype (OR = 2.34, 95% CI: 1.12-4.90) for MO, that do not remain significant after multiple test correction. A trend towards an increased risk for MA regarding the C-allele of rs5333 was also found. However, an additional MDR analysis was performed, and highly significant results were found for the two SNPs. The T-C-G haplotype (rs702757-rs5333-rs5335) was found to be significantly overrepresented in the MO subgroup, even after permutation was performed. CONCLUSIONS: Our results show additional findings for a role of EDNRA as a susceptibility factor for MO, although we cannot exclude the involvement of this gene in MA susceptibility in our population. Our study also emphasizes the need for replication of association findings in different populations.

HFE gene polymorphisms and severity in Portuguese patients with multiple sclerosis.

BACKGROUND: High iron concentrations have been reported in oligodendrocytes, myelin and macrophages in multiple sclerosis (MS) lesions. It has been proposed that HFE gene polymorphisms could have a role in MS. METHODS: The C282Y and H63D HFE variants frequencies were determined in 373 patients with MS and compared with a normal population. RESULTS: No significant association was found between HFE polymorphisms and disease susceptibility. An analysis of the association of genotypes with disease severity was performed, and the C282Y allele was more frequent in the aggressive group. CONCLUSIONS: Patients carrying the C282Y variant seem to have a worse prognosis.

Correlates of physical activity in Portuguese adolescents from 10 to 18 years.

This study examined the association between demographic [age, sex, socioeconomic status (SES)] and socio-cultural [father, mother, sibling physical activity (PA); peers and physical education teacher influences] correlates and low, moderate and high levels of PA among Portuguese adolescents aged 10-18 years. A total of 3352 males and females attending basic and secondary schools, their parents and siblings were sampled across four regions of Portugal. PA was assessed with a psychometrically validated questionnaire. Multinomial logistic regression was used. Age was positively related with moderate and high PA. Boys and adolescents of high SES were more likely to participate in moderate and high PA. Adolescents were more likely to participate in high PA when theirs mother and sibling(s) also participated. Peers had a positive influence on participation in moderate and high PA, while physical education teachers did not have an influence. The results indicated that demographic and socio-cultural correlates--in particular age, sex, SES, mother and sibling PA, and peer influence--were significantly associated with adolescent PA. These results also suggested that interventions should focus on girls and low SES adolescents who face higher risk of inactivity.

Genetic diversity in the Maremmano horse and its relationship with other European horse breeds.

The Maremmano is an Italian warmblood horse breed from central Italy. We characterized the genetic diversity and the degree of admixture in Maremmano in comparison to 14 other European horse breeds using 30 microsatellites. Between-breed diversity explained about 9 per cent of the total genetic diversity. Cluster analysis, genetic distances and genetic differentiation coefficients showed a close relationship of Maremmano with Hanoverian and Lusitano in accordance with breed history.

A 3-year longitudinal analysis of changes in fitness, physical activity, fatness and screen time.

AIM: To analyse whether changes in physical activity index (PAI), screen time (ST: television, computer) and body mass index (BMI) made a contribution to longitudinal changes in fitness of children and adolescents. Additionally, we analysed the interaction between baseline fitness level and changes in fitness. METHODS: This is a 3-year longitudinal study of 345 high school students aged 11-19 years. Students performed curl-ups, push-ups and 20-m shuttle run tests from Fitnessgram. PA and ST were evaluated using a standard questionnaire. Standardized scores of fitness tests were summed. Changes over time were calculated as Delta(1) (2007 minus 2006), Delta(2) (2008 minus 2007) and Delta(3) (2008 minus 2006). RESULTS: Changes in PAI were positively and independently associated with changes in fitness in Delta(1), Delta(2) and Delta(3). Changes in BMI were negatively associated with changes in fitness in Delta(3). Participants highly fit at baseline were those who showed positive changes in PAI over Delta(3), decreased changes in ST and had the lowest increase in BMI over 3 years compared with those low-fit at baseline. CONCLUSIONS: Changes in BMI were associated with changes in fitness over 3 years. However, changes in PAI were the best predictor for changes in fitness in each year and over the 3 years of evaluation in youth.

A 3-year longitudinal analysis of changes in Body Mass Index.

The aim of this study was to analyse whether Physical Activity Index (PAI), Physical Fitness, Screen Time (watching TV and computer use), Socio-economic Status and Commuting to School made a significant contribution to longitudinal changes in Body Mass Index (BMI) in youth. This longitudinal study was carried out over a period of 3 years with 345 students (147 boys) who were between 11 and 16 years old at the beginning of the study. Students were invited to perform tests from FITNESSGRAM Battery for Curl-Ups, Push-Ups, Back-Saver Sit and Reach, and 20 m Shuttle-Run (CRF). Fitness tests were categorized in "Healthy Zone" (HZ) and "Under Healthy Zone" (UHZ), PAI in "less active" and "active"; Socio-economic Status, in low, middle and high education level, and Commuting in active and passive. BMI was corrected for age and gender meaning that we subtracted the age-and-sex-specific cut points for overweight. Corrected body mass index was used as dependent variable in a Linear Mixed Model. The main result was the strong positive and independent association of individuals with CRF performances UHZ with corrected body mass index. In conclusion, the results of this longitudinal study showed markedly an important relationship of lower fitness levels with the risk of being overweight/obese, in particular CRF and abdominal strength.

New insights on the genetic basis of Portuguese grapevine and on grapevine domestication.

As the ancestor of cultivated grape, Vitis vinifera subsp. sylvestris represents a unique, invaluable genetic resource for the improvement of cultivated grapevines. Recently, five populations of wild grapevines were identified in Portugal. Sixty vines were characterized with 11 nuclear SSR markers and further compared with 70 genotypes of Portuguese Vitis vinifera subsp. sativa. The obtained data demonstrate moderate genetic differentiation between wild grapevine populations and moderate to high genetic differentiation between wild and cultivated grapevines. However, the identification of high degrees of similarity between wild and cultivated grapes (up to 87%) and a putative parent-progeny relationship between wild and cultivated grapes with 17 additional SSR markers is indicative of gene flow between local wild grapevine populations and Portuguese domesticated vines. Also, the ancestry of some Azorean cultivars was ascertained. The obtained data further support the hypothesis of several domestication centres, with Portugal, Spain, and Italy playing a particular role after the last glaciation, giving rise to many of the Western European cultivars.

Beta blockers for peripheral arterial disease.

OBJECTIVES: To quantify the potential harm of beta blockers in patients with peripheral arterial disease. MATERIALS AND METHODS: All randomised controlled trials (RCTs) comparing beta blockers with placebo for the outcomes of claudication and maximal walking distance and time, calf blood flow, vascular resistance and skin temperature were searched using the Cochrane Controlled Trials Register, PubMed and CINAHL. Trials comparing different types of beta blockers were excluded. RESULTS: Six RCTs fulfilling the above criteria, with a total of 119 patients, were included. The beta blockers studied were atenolol, propranolol, pindolol and metoprolol. None of the trials showed a statistically significant worsening effect of beta blockers on the outcomes measured. There were no reports of any adverse events with the beta blockers studied. CONCLUSIONS: Currently, there is no evidence to suggest that beta blockers adversely affect walking distance in people with intermittent claudication. Beta blockers should be used with caution if clinically indicated, especially in patients with critical ischaemia where acute lowering of blood pressure is contraindicated.

New insights of HLA class I association to Behçet's disease in Portuguese patients.

Human leukocyte antigen (HLA)-B*51 is a well-known genetic factor associated with Behçet's disease (BD). To analyse the influence of HLA-B*51 and other HLA class I alleles in BD susceptibility in a Portuguese population and its association with disease severity, we studied 78 BD patients and 208 healthy controls. The patients were classified into two severity groups as described by Gul et al. As expected, a higher frequency of HLA-B*51 was found. The frequency of HLA-Cw*16 alleles was significantly higher in patients. Regarding severity, HLA-B*27 frequency was higher in the severe group compared with controls and with the mild group. Thus, HLA-B*51 and HLA-Cw*16 seem to confer susceptibility to BD in this patients. HLA-B*27 may be important as a prognostic factor.

Chromosomal G-dark bands determine the spatial organization of centromeric heterochromatin in the nucleus.

Gene expression can be silenced by proximity to heterochromatin blocks containing centromeric alpha-satellite DNA. This has been shown experimentally through cis-acting chromosome rearrangements resulting in linear genomic proximity, or through trans-acting changes resulting in intranuclear spatial proximity. Although it has long been been established that centromeres are nonrandomly distributed during interphase, little is known of what determines the three-dimensional organization of these silencing domains in the nucleus. Here, we propose a model that predicts the intranuclear positioning of centromeric heterochromatin for each individual chromosome. With the use of fluorescence in situ hybridization and confocal microscopy, we show that the distribution of centromeric alpha-satellite DNA in human lymphoid cells synchronized at G(0)/G(1) is unique for most individual chromosomes. Regression analysis reveals a tight correlation between nuclear distribution of centromeric alpha-satellite DNA and the presence of G-dark bands in the corresponding chromosome. Centromeres surrounded by G-dark bands are preferentially located at the nuclear periphery, whereas centromeres of chromosomes with a lower content of G-dark bands tend to be localized at the nucleolus. Consistent with the model, a t(11; 14) translocation that removes G-dark bands from chromosome 11 causes a repositioning of the centromere, which becomes less frequently localized at the nuclear periphery and more frequently associated with the nucleolus. The data suggest that "chromosomal environment" plays a key role in the intranuclear organization of centromeric heterochromatin. Our model further predicts that facultative heterochromatinization of distinct genomic regions may contribute to cell-type specific patterns of centromere localization.

A pilot study on the effect of topical negative pressure on quality of life.

OBJECTIVE: To discover the impact of topical negative pressure (TNP) on quality of life. METHOD: An exploratory prospective cohort study was conducted on 26 patients undergoing TNP. The Cardiff Wound Impact Schedule (CWIS), a wound-specific tool, was used to investigate quality-of-life scores before therapy and four weeks after therapy or at wound closure. Wound dimensions were measured at both assessments, and the values for the CWIS domains (physical symptoms, social functioning, well-being and overall quality of life) were investigated using parametric and non-parametric tests. RESULTS: The mean duration of TNP therapy was 3.3 +/- 1.7 weeks. Topical negative pressure therapy helped to achieve complete wound closure in 14 patients (54%), and there was a mean reduction in wound surface area from 52.2 cm2 (range 4-150) to 26.8 cm2 (0-120). While there was no significant change in quality of life in patients whose wounds healed (1 +/- 11.9), the physical-functioning domain improved in obese patients (20 +/- 21, p < 0.05) and worsened in ambulatory patients (-3 +/- 13, p < 0.05). The portableTNP system had no significant impact on quality of life (-3 +/- 16), while the global quality-of-life score worsened with surgical intervention (-0.5 +/- 2, p < 0.05). CONCLUSION: Although TNP aids wound closure in patients with complex wounds, in selected cases their quality of life can worsen. This is the first exploratory cohort study of its kind, and has identified an urgent need to validate the use of patient-based outcome measures in TNP therapy. Such data can be useful in allocating resources and justifying funding in wound care.

Expression of ubiquitin and proteasome in motorneurons and astrocytes of spinal cords from patients with amyotrophic lateral sclerosis.

Proteasome, ubiquitin, GFAP and neurofilament were evaluated in motorneurons and astrocytes of spinal cords of ALS and control cases. ALS neurons exhibited ubiquitin positive inclusions and areas of strong immunoreaction for proteasome. Areas of proteasome stain were observed close to neurofilament positive proximal process enlargement. The percentage of neurons strongly immunoreacted, for proteasome was higher in ALS cases than in controls. Many astrocytes were positive for ubiquitin and proteasome. These results suggest that the ubiquitin-proteasome pathway is involved in the ALS pathogenesis and agree with the view that ALS is a disorder of protein aggregation that affects neurons and nonneuronal cells.