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Background: We estimate the prevalence of drug injection, the variables associated with having ever injected and the proportion of ever injectors whose first drug injection was for having sex; we describe the first drug injection episode, analyze the drugs most frequently injected and estimate the prevalence of risky injecting behaviors. Methods: The participants were 3387 MSM without a previous HIV diagnosis attending four HIV/STI diagnosis services in Madrid and Barcelona. Lifetime prevalence and prevalence ratios (PRs) by different factors were calculated using Poisson regression models with robust variance. We compared the characteristics of first drug injection episode, lifetime injection and risky injecting behaviors of those whose first injection was for sex (FIS) with those whose was not (non-FIS). Results: Lifetime prevalence of injection was 2.1% (CI 1.7–2.7). In the multivariate analysis, it was strongly associated with having been penetrated by more than five men in the last 12 months (aPR = 10.4; CI 2.5–43.4) and having met most of their partners at private parties (aPR = 7.5; CI 4.5–12.3), and less strongly with other factors. Of those who had ever injected drugs, 81.9% injected for sex the first time they injected drugs (FIS). At first injection, FIS participants had a mean age of 31 years, 62.7% used mephedrone and 32.2% methamphetamine on that occasion. Of this FIS group 39.0% had ever shared drugs or equipment and 82.6% had always shared for sex. Some 30.8% of non-FIS reported having also injected drugs for sex later on. Conclusions: Only two out of a hundred had ever injected, most to have sex and with frequent drug or injecting equipment sharing. Injecting for sex is the most common first episode of drug injection and is the most efficient risky behavior for the transmission of HIV, hepatitis B or C and other blood-borne infections. MSM participating in private parties should be considered a priority group for prevention policies. [ABSTRACT FROM AUTHOR]

Background: Toll-like receptors (TLRs) are transmembrane pattern-recognition receptors of the innate immune system recognizing diverse pathogen-derived and tissue damage-related ligands. It has been suggested that TLR signaling contributes to the pathogenesis of age-related, neurodegenerative diseases, including Alzheimer's disease (AD). AD is associated to oligomers of the amyloid β peptide (Aβo) that cause intracellular Ca2+ dishomeostasis and neuron cell death in rat hippocampal neurons. Here we assessed the interplay between inflammation and Aβo in long-term cultures of rat hippocampal neurons, an in vitro model of neuron aging and/or senescence.Methods: Ca2+ imaging and immunofluorescence against annexin V and TLR4 were applied in short- and long-term cultures of rat hippocampal neurons to test the effects of TLR4-agonist LPS and Aβo on cytosolic [Ca2+] and on apoptosis as well as on expression of TLR4.Results: LPS increases cytosolic [Ca2+] and promotes apoptosis in rat hippocampal neurons in long-term culture considered aged and/or senescent neurons, but not in short-term cultured neurons considered young neurons. TLR4 antagonist CAY10614 prevents both effects. TLR4 expression in rat hippocampal neurons is significantly larger in aged hippocampal cultures. Treatment of aged hippocampal cultures with Aβo increases TLR4 expression and enhances LPS-induced Ca2+ responses and neuron cell death.Conclusions: Aging and amyloid β oligomers, the neurotoxin involved in Alzheimer's disease, enhance TLR4 expression as well as LPS-induced Ca2+ responses and neuron cell death in rat hippocampal neurons aged in vitro. [ABSTRACT FROM AUTHOR]

Background: The increasing availability of DNA markers provides new metrics of inbreeding based on single nucleotide polymorphisms (SNPs), i.e. molecular inbreeding or the proportion of runs of homozygosity (ROH), as alternatives to traditional pedigree-based inbreeding coefficients. However, none of these metrics incorporate the length of ROH as an indicator of recent inbreeding. Novel inbreeding coefficients that incorporate length of ROH as a random variable with an associated density are investigated. Methods: New inbreeding metrics based on the distribution of the length of ROH are proposed: (1) the Kolmolgorov– Smirnov test, (2) a function of the quantiles of the cumulative distribution function of an individual versus the population, and (3) fitting of an exponential distribution to ROH lengths (mean, variance, and the probability of drawing at random a ROH larger than a given threshold). The new inbreeding and pedigree-based metrics were compared using 217 sows of an Iberian line that belong to three groups: C1 (conservation), C2 (conservation derived from C1), and S (selected and derived from C1), with complete pedigrees and genotyped for 35,023 SNPs. Results: Correlations between pedigree-based and the new genomic inbreeding coefficients ranged from 0.22 to 0.72 but most ranged from 0.60 to 0.70. The correlation between quantile chromosomal inbreeding coefficients (using molecular information of just one chromosome at the time) and chromosomal length was 0.84 (SE = 0.14), supporting the hypothesis that these coefficients incorporate information on ROH length as an indication of recent inbreeding. Kolmogorov–Smirnov and exponential chromosomal inbreeding coefficients were also correlated with chromosomal length (0.57). Chromosome 1 had the largest quantile ROH inbreeding coefficient (largest ROH sizes), whereas chromosome 10 had the lowest (shortest ROH sizes). Selection for lean growth increased ROH-based inbreeding coefficients for group S when compared to unselected groups C1 and C2. At the chromosomal level, this comparison showed that the level of autozygosity and the length of ROH for most of the autosomes increased in the selection line. Conclusions: Quantile and exponential probability inbreeding coefficients using ROH length as a random variable provide additional information about recent inbreeding compared to existing inbreeding coefficients such as molecular, pedigree-based or total ROH content inbreeding coefficients. [ABSTRACT FROM AUTHOR]

Background Hepatitis E virus (HEV) has been reported in the human population and pigs are a recognized reservoir for HEV and a possible source of HEV transmission to humans. Spray-dried porcine plasma (SDPP) is an ingredient commonly used in feed for pigs around the world. Even though processing conditions used to produce SDPP should be adequate to inactivate HEV, it was of interest to analyze commercial SDPP samples for presence of genome and antibodies (AB) against HEV and to retrospectively analyze serum samples collected from pigs used in past experiments that had been fed diets containing either 0% or 8% SDPP to detect potential transmission of HEV as determined by seroconversion. Results Eighty-five commercial SDPP samples were analyzed by ELISA and 100% of them contained AB against HEV, while 22.4% (11 of 49 samples analyzed) were positive for HEV RNA. Frozen sera samples (n = 140) collected from 70 pigs used in past experiments that had been fed diets containing either 0% or 8% commercial SDPP was analyzed by ELISA for AB against HEV. Age of pigs at sera sampling ranged from 3 to 15 weeks and feeding duration of diets ranged from approximately 4 to 9 weeks. One lot of SDPP used in one experiment was analyzed and confirmed to contain HEV RNA. Regardless of the diet fed, some sera samples collected at the beginning of an experiment contained AB titer against HEV. These sera samples were collected from weaned pigs prior to feeding of the experimental diets and the HEV titer was probably from maternal origin. However, by the end of the experiments, HEV titer was not detected or had declined by more than 50% of the initial titer concentration. Conclusions To our knowledge, this is the first study reporting presence of HEV AB titer and RNA in SDPP. Retrospective analysis of serum collected from pigs fed diets with SDPP revealed no indication of seroconversion to HEV. The results indicate that feeding SDPP in diets for pigs does not represent a risk of transmitting HEV, even though HEV genome may be detected in SDPP. [ABSTRACT FROM AUTHOR]

Background The two main genetic types in Iberian pig production show important phenotypic differences in growth, fattening and tissue composition since early developmental stages. The objective of this work was the evaluation of muscle transcriptome profile in piglets of both genetic types, in order to identify genes, pathways and regulatory factors responsible for their phenotypic differences. Contemporary families coming from pure Iberian pigs (IB) or from crossing with Duroc boars (DUxIB) were generated. Piglets (14 from each genetic type) were slaughtered at weaning (28 days) and longissimus dorsi was sampled for composition and gene expression studies. RNA was obtained and hybridized to Affymetrix Porcine Genechip expression arrays. Results Loin muscle chemical composition showed significant differences between genetic types in intramuscular fat content (6.1% vs. 4.3% in IB and DUxIB animals, respectively, P = 0.009) and in saturated (P = 0.019) and monounsaturated fatty acid proportions (P = 0.044). The statistical analysis of gene expression data allowed the identification of 256 differentially expressed (DE) genes between genetic types (FDR < 0.10), 102 upregulated in IB and 154 upregulated in DUxIB. Transcript differences were validated for a subset of DE genes by qPCR. We observed alteration in biological functions related to extracellular matrix function and organization, cellular adhesion, muscle growth, lipid metabolism and proteolysis. Candidate genes with known effects on muscle growth were found among the DE genes upregulated in DUxIB. Genes related to lipid metabolism and proteolysis were found among those upregulated in IB. Regulatory factors (RF) potentially involved in the expression differences were identified by calculating the regulatory impact factors. Twenty-nine RF were found, some of them with known relationship with tissue development (MSTN, SIX4, IRX3), adipogenesis (CEBPD, PPARGC1B), or extracellular matrix processes (MAX, MXI1). Correlation among the expression of these RF and DE genes show relevant differences between genetic types. Conclusion These results provide valuable information about genetic mechanisms determining the phenotypic differences on growth and meat quality between the genetic types studied, mainly related to the development and function of the extracellular matrix and also to some metabolic processes as proteolysis and lipid metabolism. Transcription factors and regulatory mechanisms are proposed for these altered biological functions. [ABSTRACT FROM AUTHOR]

Background: Prolificacy is the most important trait influencing the reproductive efficiency of pig production systems. The low heritability and sex-limited expression of prolificacy have hindered to some extent the improvement of this trait through artificial selection. Moreover, the relative contributions of additive, dominant and epistatic QTL to the genetic variance of pig prolificacy remain to be defined. In this work, we have undertaken this issue by performing one-dimensional and bi-dimensional genome scans for number of piglets born alive (NBA) and total number of piglets born (TNB) in a three generation Iberian by Meishan F2 intercross. Results: The one-dimensional genome scan for NBA and TNB revealed the existence of two genome-wide highly significant QTL located on SSC13 (P < 0.001) and SSC17 (P < 0.01) with effects on both traits. This relative paucity of significant results contrasted very strongly with the wide array of highly significant epistatic QTL that emerged in the bi-dimensional genome-wide scan analysis. As much as 18 epistatic QTL were found for NBA (four at P < 0.01 and five at P < 0.05) and TNB (three at P < 0.01 and six at P < 0.05), respectively. These epistatic QTL were distributed in multiple genomic regions, which covered 13 of the 18 pig autosomes, and they had small individual effects that ranged between 3 to 4% of the phenotypic variance. Different patterns of interactions (a × a, a × d, d × a and d × d) were found amongst the epistatic QTL pairs identified in the current work. Conclusions: The complex inheritance of prolificacy traits in pigs has been evidenced by identifying multiple additive (SSC13 and SSC17), dominant and epistatic QTL in an Iberian × Meishan F2 intercross. Our results demonstrate that a significant fraction of the phenotypic variance of swine prolificacy traits can be attributed to first-order gene-by-gene interactions emphasizing that the phenotypic effects of alleles might be strongly modulated by the genetic background where they segregate. [ABSTRACT FROM AUTHOR]

Background: Graham Little - Piccardi - Lassueur (GLPL) syndrome is a rare dermatosis characterized by scarring alopecia, loss of pubic and axillary hair, and progressive development of variously located follicular papules. We report a first case ever of an autoimmune response in a patient suffering from GLPL syndrome. Methods: Immunofluorescence and immunoblot analysis were used in a variety of cell culturesincluding human, monkey, hamster, mouse and bovine cells to analyze the presence of autoantibodies in a GLPL patient. Results: The autoimmune serum showed a pattern of centromere and spindle microtubule staining resembling that of the chromosomal passenger protein complex. By using a complex of proteins expressed in baculovirus, immunoblot analysis demonstrated that the INCENP protein is a major autoantigen in this patient with GLPL syndrome. Conclusion: An autoimmune response in GLPL syndrome is reported against the INCENP centromere protein. The occasional development of autoimmunity in GLPL patients could serve as a test in continuing efforts to detect this disease and for a more directed therapy based on the autoantigen response. [ABSTRACT FROM AUTHOR]

An abstract of the article "Prevalence and changing distribution of HTLV-1 and HTLV-2 infections in Spain," by Ana Treviño and colleagues is presented.

An abstract of the article "Seroprevalence of HTLV infection in Spain among immigrant pregnant women," by Rafael Benito, Estrella Caballero and colleagues, is presented.

Background: Toll-like receptors (TLRs) are transmembrane pattern-recognition receptors of the innate immune system recognizing diverse pathogen-derived and tissue damage-related ligands. It has been suggested that TLR signaling contributes to the pathogenesis of age-related, neurodegenerative diseases, including Alzheimer's disease (AD). AD is associated to oligomers of the amyloid β peptide (Aβo) that cause intracellular Ca 2+ dishomeostasis and neuron cell death in rat hippocampal neurons. Here we assessed the interplay between inflammation and Aβo in long-term cultures of rat hippocampal neurons, an in vitro model of neuron aging and/or senescence., Methods: Ca 2+ imaging and immunofluorescence against annexin V and TLR4 were applied in short- and long-term cultures of rat hippocampal neurons to test the effects of TLR4-agonist LPS and Aβo on cytosolic [Ca 2+ ] and on apoptosis as well as on expression of TLR4., Results: LPS increases cytosolic [Ca 2+ ] and promotes apoptosis in rat hippocampal neurons in long-term culture considered aged and/or senescent neurons, but not in short-term cultured neurons considered young neurons. TLR4 antagonist CAY10614 prevents both effects. TLR4 expression in rat hippocampal neurons is significantly larger in aged hippocampal cultures. Treatment of aged hippocampal cultures with Aβo increases TLR4 expression and enhances LPS-induced Ca 2+ responses and neuron cell death., Conclusions: Aging and amyloid β oligomers, the neurotoxin involved in Alzheimer's disease, enhance TLR4 expression as well as LPS-induced Ca 2+ responses and neuron cell death in rat hippocampal neurons aged in vitro.

Background: Conventional vaccination programs using a single injection of a combined vaccine against porcine circovirus type 2 (PCV2) and Mycoplasma hyopneumoniae (MHYO) can promote a strong immune response that reduces feed intake for 24 to 48 h post injection. Often such vaccines are given around the time of weaning during a critical stress period in which feed intake is already compromised. Spray dried porcine plasma (SDPP) is a protein source used in starter diets that increases post-weaning feed intake of pigs. The objectives of this study were to determine the effects of a conventional vaccination program along with feeding SDPP in a starter diet on antibody development and wean to finish performance of pigs., Results: Pigs fed the starter diet with SDPP had improved body weight, average daily weight gain and average daily feed intake during the initial 14 d after weaning along with improved feed efficiency during the initial 7 d after weaning and these responses were independent of vaccination. Vaccination at 3 d after weaning had no significant effect on performance during the initial 14 d after weaning. Cumulative mortality was reduced for pigs fed the starter diet with SDPP, while vaccinated pigs had reduced mortality from d 48 to 145. Both vaccinated pigs and those fed the starter diet with SDPP had heavier carcass weight. One pig per pen was challenged with PCV2 at d 63. A higher percentage of vaccinated pigs were sero-positive for antibodies against PCV2 and MHYO at d 35, 63 and 78. Antibody values against PCV2 were higher for vaccinated pigs at d 35 and 63, but lower at d 146. Percentage of positive samples for PCV2 genome in serum was reduced for vaccinated pigs at d 117 and 146. Antibody values against MHYO were increased for vaccinated pigs at d 35, 63 and 78., Conclusions: Vaccination supported a long term antibody response against PCV2 and a moderate but weaker antibody response against MHYO for early finishing pigs challenged with PCV2. Using SDPP in the starter diet along with vaccination supported the best long-term beneficial effects on survival to market and carcass weight.

Background: Thymidine analogue resistance mutations (TAMs) selected under treatment with nucleoside analogues generate two distinct genotypic profiles in the HIV-1 reverse transcriptase (RT): (i) TAM1: M41L, L210W and T215Y, and (ii) TAM2: D67N, K70R and K219E/Q, and sometimes T215F. Secondary mutations, including thumb subdomain polymorphisms (e.g. R284K) have been identified in association with TAMs. We have identified mutational clusters associated with virological failure during salvage therapy with tenofovir/emtricitabine-based regimens. In this context, we have studied the role of R284K as a secondary mutation associated with mutations of the TAM1 complex., Results: The cross-sectional study carried out with > 200 HIV-1 genotypes showed that virological failure to tenofovir/emtricitabine was strongly associated with the presence of M184V (P < 10-10) and TAMs (P < 10-3), while K65R was relatively uncommon in previously-treated patients failing antiretroviral therapy. Clusters of mutations were identified, and among them, the TAM1 complex showed the highest correlation coefficients. Covariation of TAM1 mutations and V118I, V179I, M184V and R284K was observed. Virological studies showed that the combination of R284K with TAM1 mutations confers a fitness advantage in the presence of zidovudine or tenofovir. Studies with recombinant HIV-1 RTs showed that when associated with TAM1 mutations, R284K had a minimal impact on zidovudine or tenofovir inhibition, and in their ability to excise the inhibitors from blocked DNA primers. However, the mutant RT M41L/L210W/T215Y/R284K showed an increased catalytic rate for nucleotide incorporation and a higher RNase H activity in comparison with WT and mutant M41L/L210W/T215Y RTs. These effects were consistent with its enhanced chain-terminated primer rescue on DNA/DNA template-primers, but not on RNA/DNA complexes, and can explain the higher fitness of HIV-1 having TAM1/R284K mutations., Conclusions: Our study shows the association of R284K and TAM1 mutations in individuals failing therapy with tenofovir/emtricitabine, and unveils a novel mechanism by which secondary mutations are selected in the context of drug-resistance mutations.

Background: Type 2 diabetes mellitus and depression are highly prevalent diseases that are associated with an increased risk of cardiovascular disease and mortality. There is evidence about a bidirectional association between depressive symptoms and type 2 diabetes mellitus. However, prognostic implications of the joint effects of these two diseases on cardiovascular morbidity and mortality are not well-known., Method/design: A three-year, observational, prospective, cohort study, carried out in Primary Health Care Centres in Madrid (Spain). The project aims to analyze the effect of depression on cardiovascular events, all-cause and cardiovascular mortality in patients with type 2 diabetes mellitus, and to estimate a clinical predictive model of depression in these patients.The number of patients required is 3255, all them with type 2 diabetes mellitus, older than 18 years, who regularly visit their Primary Health Care Centres and agree to participate. They are chosen by simple random sampling from the list of patients with type 2 diabetes mellitus of each general practitioner.The main outcome measures are all-cause and cardiovascular mortality and cardiovascular morbidity; and exposure variable is the major depressive disorder.There will be a comparison between depressed and not depressed patients in all-cause mortality, cardiovascular mortality, coronary artery disease and stroke using the Chi-squared test. Logistic regression with random effects will be used to adjust for prognostic factors. Confounding factors that might alter the effect recorded will be taken into account in this analysis. To assess the effect of depression on the mortality, a survival analysis will be used comparing the two groups using the log-rank test. The control of potential confounding variables will be performed by the construction of a Cox regression model., Discussion: Our study's main contribution is to evaluate the increase in the risk of cardiovascular morbidity and mortality, in depressed Spanish adults with type 2 diabetes mellitus attended in Primary Health Care Setting. It would also be useful to identify subgroups of patients for which the interventions could be more beneficial.

Background: Although most HTLV infections in Spain have been found in native intravenous drug users carrying HTLV-2, the large immigration flows from Latin America and Sub-Saharan Africa in recent years may have changed the prevalence and distribution of HTLV-1 and HTLV-2 infections, and hypothetically open the opportunity for introducing HTLV-3 or HTLV-4 in Spain. To assess the current seroprevalence of HTLV infection in Spain a national multicenter, cross-sectional, study was conducted in June 2009., Results: A total of 6,460 consecutive outpatients attending 16 hospitals were examined. Overall, 12% were immigrants, and their main origin was Latin America (4.9%), Africa (3.6%) and other European countries (2.8%). Nine individuals were seroreactive for HTLV antibodies (overall prevalence, 0.14%). Evidence of HTLV-1 infection was confirmed by Western blot in 4 subjects (prevalence 0.06%) while HTLV-2 infection was found in 5 (prevalence 0.08%). Infection with HTLV types 1, 2, 3 and 4 was discarded by Western blot and specific PCR assays in another two specimens initially reactive in the enzyme immunoassay. All but one HTLV-1 cases were Latin-Americans while all persons with HTLV-2 infection were native Spaniards., Conclusions: The overall prevalence of HTLV infections in Spain remains low, with no evidence of HTLV-3 or HTLV-4 infections so far.

Background: Computerized Clinical Records, which are incorporated in primary health care practice, have great potential for research. In order to use this information, data quality and reliability must be assessed to prevent compromising the validity of the results.The aim of this study is to validate the diagnosis of hypertension and diabetes mellitus in the computerized clinical records of primary health care, taking the diagnosis criteria established in the most prominently used clinical guidelines as the gold standard against which what measure the sensitivity, specificity, and determine the predictive values.The gold standard for diabetes mellitus was the diagnostic criteria established in 2003 American Diabetes Association Consensus Statement for diabetic subjects. The gold standard for hypertension was the diagnostic criteria established in the Joint National Committee published in 2003., Methods: A cross-sectional multicentre validation study of diabetes mellitus and hypertension diagnoses in computerized clinical records of primary health care was carried out. Diagnostic criteria from the most prominently clinical practice guidelines were considered for standard reference.Sensitivity, specificity, positive and negative predictive values, and global agreement (with kappa index), were calculated. Results were shown overall and stratified by sex and age groups., Results: The agreement for diabetes mellitus with the reference standard as determined by the guideline was almost perfect (κ=0.990), with a sensitivity of 99.53%, a specificity of 99.49%, a positive predictive value of 91.23% and a negative predictive value of 99.98%.Hypertension diagnosis showed substantial agreement with the reference standard as determined by the guideline (κ=0.778), the sensitivity was 85.22%, the specificity 96.95%, the positive predictive value 85.24%, and the negative predictive value was 96.95%. Sensitivity results were worse in patients who also had diabetes and in those aged 70 years or over., Conclusions: Our results substantiate the validity of using diagnoses of diabetes and hypertension found within the computerized clinical records for epidemiologic studies.

Introduction: Systemic lupus erythematosus (SLE) is characterized by B cell hyper-activation and auto-reactivity resulting in pathogenic auto-antibody generation. The phenotypic analysis of blood B cell subsets can be used to understand these alterations., Methods: The combined detection of CD19, CD27 and IgD (or IgM) by flow cytometry (FC) analysis delineates five well-defined blood B cell-subsets: naive, switched (S) memory, double negative (DN) memory and CD27 IgD IgM (non-switched memory) B lymphocytes, and plasma cells (PCs). This phenotypic study was performed in 69 consecutive SLE patients and 31 healthy controls., Results: SLE patients exhibited several abnormalities in the distribution of these B cell subsets, including elevated levels of DN memory B cells and PCs, and decreased CD27 IgD IgM B cells. Active SLE patients also showed decreased presence of S memory B cells and increased proportions of naive B lymphocytes. Nevertheless, when the patients in remission who did not require treatment were studied separately, the only remaining abnormality was a reduction of the CD27 IgD IgM B cell-subset detectable in most of these patients. The level of reduction of CD27 IgD IgM B cells was associated with elevated values of serum SLE auto-antibodies. Further analysis of this latter B cell-subset specifically showed increased expression of CD80, CD86, CD95, 9G4 idiotype and functional CXCR3 and CXCR4., Conclusions: The presence of a reduced blood CD27 IgD IgM B cell-subset, exhibiting an activated state and enriched for auto-reactivity, is a consistent B cell abnormality in SLE. These findings suggest that CD27 IgD IgM B lymphocytes play a role in the pathogenesis of this disease.

One QTL affecting backfat thickness (BF), intramuscular fat content (IMF) and eye muscle area (MA) was previously localized on porcine chromosome 6 in an F2 cross between Iberian and Landrace pigs. This work was done to study the effect of two positional candidate genes on these traits: H-FABP and LEPR genes. The QTL mapping analysis was repeated with a regression method using genotypes for seven microsatellites and two PCR-RFLPs in the H-FABP and LEPR genes. H-FABP and LEPR genes were located at 85.4 and 107 cM respectively, by linkage analysis. The effects of the candidate gene polymorphisms were analyzed in two ways. When an animal model was fitted, both genes showed significant effects on fatness traits, the H-FABP polymorphism showed significant effects on IMF and MA, and the LEPR polymorphism on BF and IMF. But when the candidate gene effect was included in a QTL regression analysis these associations were not observed, suggesting that they must not be the causal mutations responsible for the effects found. Differences in the results of both analyses showed the inadequacy of the animal model approach for the evaluation of positional candidate genes in populations with linkage disequilibrium, when the probabilities of the parental origin of the QTL alleles are not included in the model.