The inversion of multiresponse partial least squares models, a useful tool to improve analytical methods in the framework of analytical quality by design.

Analytical Quality by Design (AQbD) is the adaptation of Quality by Design (QbD) when it is applied to the development of an analytical method. The main idea is to develop the analytical method in such a way that the desired quality of the Critical Quality Attributes (CQAs), stated via the analytical target profile (ATP), is maintained while allowing some variation in the Control Method Parameters (CMPs). The paper presents a general procedure for selecting factor levels in the CMPs to achieve the desired responses, characterized by the CQAs, when liquid chromatographic methods are to be used for the simultaneous determination of several analytes. In such a case, the CMPs are usually the composition of the ternary mobile phase, its flow rate, column temperature, etc., while typical CQAs refer to the quality of the chromatograms in terms of the resolution between each pair of consecutive peaks, initial and final chromatographic time, etc. The analytical target profile in turn defines the desired characteristics for the CQAs, the reason for the whole approach. The procedure consists of four steps. The first is to construct a D-optimal combined design (mixture-process design) to select the domain and levels of the CMPs. The second step is to fit a PLS2 model to predict the analytical responses expressed in the ATP (the good characteristics of the chromatogram) as a function of the CMPs. The third step is the inversion of the PLS2 model to obtain the conditions necessary to obtain the preset ATP in the corresponding CQAs. The inversion is performed computationally in order to estimate the Pareto front of these responses, namely, a set of experimental conditions to perform the chromatographic determination for which the desired critical quality attributes are met. The fourth final step is to obtain the Method Operable Design Region (MODR), that is, the region where the CMPs can vary while maintaining the quality of the CQAs. The procedure has been applied to some cases involving different analytes, all of which are regulated by the European Union due to their toxicity to human health, namely five bisphenols and ten polycyclic aromatic hydrocarbons.

Short-term impact of noise, other air pollutants and meteorological factors on emergency hospital mental health admissions in the Madrid region.

BACKGROUND: A number of environmental factors, such as air pollution, noise in urbanised settings and meteorological-type variables, may give rise to important effects on human health. In recent years, many studies have confirmed the relation between various mental disorders and these factors, with a possible impact on the increase in emergency hospital admissions due to these causes. The aim of this study was to analyse the impact of a range of environmental factors on daily emergency hospital admissions due to mental disorders in the Madrid Autonomous Region (MAR), across the period 2013-2018. METHODOLOGY: Longitudinal ecological time series study analysed by Generalised Linear Models with Poisson regression, with the dependent variable being daily Emergency Hospital Mental Health Admissions (EHMHA) in the MAR, and the independent variable being mean daily concentrations of chemical pollutants, noise levels and meteorological variables. RESULTS: EHMHA were related statistically significantly in the short term with diurnal noise levels. Relative risks (RRs) for total admissions due to mental disorders and self-inflicted injuries, in the case of diurnal noise was RR: 1.008 95%CI (1.003 1.013). Admissions attributable to diurnal noise account for 5.5% of total admissions across the study period. There was no association between hospital admissions and chemical air pollution. CONCLUSION: Noise is a variable that shows a statistically significant short-term association with EHMHA across all age groups in the MAR region. The results of this study may serve as a basis for drawing up public health guidelines and plans, which regard these variables as risk factors for mental disorders, especially in the case of noise, since this fundamentally depends on anthropogenic activities in highly urbanised areas with high levels of traffic density.

Gender differences in adaptation to heat in Spain (1983-2018).

In Spain the average temperature has increased by 1.7 °C since pre-industrial times. There has been an increase in heat waves both in terms of frequency and intensity, with a clear impact in terms of population health. The effect of heat waves on daily mortality presents important territorial differences. Gender also affects these impacts, as a determinant that conditions social inequalities in health. There is evidence that women may be more susceptible to extreme heat than men, although there are relatively few studies that analyze differences in the vulnerability and adaptation to heat by sex. This could be related to physiological causes. On the other hand, one of the indicators used to measure vulnerability to heat in a population and its adaptation is the minimum mortality temperature (MMT) and its temporal evolution. The aim of this study was to analyze the values of MMT in men and women and its temporal evolution during the 1983-2018 period in Spain's provinces. An ecological, longitudinal retrospective study was carried out of time series data, based on maximum daily temperature and daily mortality data corresponding to the study period. Using cubic and quadratic fits between daily mortality rates and the temperature, the minimum values of these functions were determined, which allowed for determining MMT values. Furthermore, we used an improved methodology that provided for the estimation of missing MMT values when polynomial fits were inexistent. This analysis was carried out for each year. Later, based on the annual values of MMT, a linear fit was carried out to determine the rate of evolution of MMT for men and for women at the province level. Average MMT for all of Spain's provinces was 29.4 °C in the case of men and 28.7 °C in the case of women. The MMT for men was greater than that of women in 86 percent of the total provinces analyzed, which indicates greater vulnerability among women. In terms of the rate of variation in MMT during the period analyzed, that of men was 0.39 °C/decade, compared to 0.53 °C/decade for women, indicating greater adaptation to heat among women, compared to men. The differences found between men and women were statistically significant. At the province level, the results show great heterogeneity. Studies carried out at the local level are needed to provide knowledge about those factors that can explain these differences at the province level, and to allow for incorporating a gender perspective in the implementation of measures for adaptation to high temperatures.

Method operable design region obtained with a partial least squares model inversion in the determination of ten polycyclic aromatic hydrocarbons by liquid chromatography with fluorescence detection.

A chromatographic method with the Analytical Quality by Design (AQbD) methodology is developed for the simultaneous determination by HPLC-FLD of ten PAHs (naphthalene, phenanthrene, anthracene, fluoranthene, pyrene, chrysene, benzo[a]anthracene, perylene, benzo[b]fluoranthene, and benzo[a]pyrene), widely spread in the environment. The construction of the Method Operable Design Region (MODR) is conducted, for the first time, via the inversion of a multiresponse Partial Least Squares (PLS2) model, which is needed to maintain the correlations among the Critical Method Parameters (CMP), among the Critical Quality Attributes (CQA), and the covariance between one another. The five CMP considered were the composition of the mobile phase (water, methanol, acetonitrile), flow rate, and column temperature. The eight CQA were linked to resolution between peaks recorded in the same emission wavelength (greater than 1.4) and the total time (less than 15 minutes). By systematic use of experimental design and parallel coordinates plots to explore the Pareto optimal front obtained with the PLS2 model inversion, the computed MODR is formed by convex combinations of eight specific settings of Critical Method Parameters that have a mobile phase with percentages of water between 37 and 38 %, of methanol from 13 and 22 %, and of acetonitrile between 41 and 49 %, together with a flow rate between 1.47 and 1.50 mL min-1, and column temperature between 41.9 and 44.0 °C in their adequate combinations. All the chromatographic peaks are well resolved, with total time varying between 12.96 and 15.66 min inside the estimated MODR and the analytical method is accurate with CCß between 0.9 and 7.0 µg L-1 with probability of both false positive and false negative equal to 0.05.

A new approach based on inversion of a partial least squares model searching for a preset analytical target profile. Application to the determination of five bisphenols by liquid chromatography with diode array detector.

The paper shows a procedure for selecting the control method parameters (factors) to obtain a preset 'analytical target profile' when a liquid chromatographic technique is going to be carried out for the simultaneous determination of five bisphenols (bisphenol-A, bisphenol-S, bisphenol-F, bisphenol-Z and bisphenol-AF), some of them regulated by the European Union. The procedure has three steps. The first consists of building a D-optimal combined design (mixture-process design) for the control method parameters, which are the composition of the ternary mobile phase and its flow rate. The second step is to fit a PLS2 model to predict six analytical responses (namely, the resolution between each pair of consecutive peaks, and the initial and final chromatographic time) as a function of the control method parameters. The third final step is the inversion of the PLS2 model to obtain the conditions needed for attaining a preset analytical target profile. The computational inversion of the PLS2 prediction model looking for the Pareto front of these six responses provides a set of experimental conditions to conduct the chromatographic determination, specifically 22% of water, mixed with 58% methanol and 20% of acetonitrile, keeping the flow rate at 0.66 mL min-1. These conditions give a chromatogram with retention times of 2.180, 2.452, 2.764, 3.249 and 3.775 min for BPS, BPF, BPA, BPAF and BPZ, respectively, and excellent resolution among all the chromatographic peaks. Finally, the analytical method is validated under the selected experimental conditions, in terms of trueness and precision. In addition, the detection capability for the five bisphenols were: 596, 334, 424, 458 and 1156 µg L-1, with probabilities of false positive and of false negative equal to 0.05.

Ghrelin treatment leads to dendritic spine remodeling in hippocampal neurons and increases the expression of specific BDNF-mRNA species.

Ghrelin (Gr) is an orexigenic peptide that acts via its specific receptor, GHSR-1a distributed throughout the brain, being mainly enriched in pituitary, cortex and hippocampus (Hp) modulating a variety of brain functions. Behavioral, electrophysiological and biochemical evidence indicated that Gr modulates the excitability and the synaptic plasticity in Hp. The present experiments were designed in order to extend the knowledge about the Gr effect upon structural synaptic plasticity since morphological and quantitative changes in spine density after Gr administration were analyzed "in vitro" and "in vivo". The results show that Gr administered to hippocampal cultures or stereotactically injected in vivo to Thy-1 mice increases the density of dendritic spines (DS) being the mushroom type highly increased in secondary and tertiary extensions. Spines classified as thin type were increased particularly in primary extensions. Furthermore, we show that Gr enhances selectively the expression of BDNF-mRNA species.

Cytoplasmic interaction of the tumour suppressor protein hSNF5 with dynamin-2 controls endocytosis.

Human SNF5 (hSNF5; INI1, SMARCB1 or BAF47) is a component of the human SWI/SNF chromatin remodelling complex and a tumour suppressor mutated in rhabdoid tumours. It also associates with the integrase of the human immunodeficiency virus (HIV)-1. We show by fluorescence loss in photobleaching that hSNF5 is constantly shuttling between the nucleus and the cytoplasm, raising the question of what the role of hSNF5 is in the cytoplasm. Here, we demonstrate that hSNF5 directly interacts with the GTPase dynamin-2 (DNM2) in the cytoplasm. DNM2 is a large GTPase involved in endocytosis and vesicle dynamics, which has been related to HIV-1 internalization. We show that hSNF5 colocalizes with DNM2 in endocytic vesicles. Depletion of hSNF5, but not of other components of the SWI/SNF complex, destabilizes DNM2 and impairs DNM2-dependent endocytosis. Furthermore, we show that hSNF5 inhibits assembly-stimulated DNM2 GTPase activity but not basal GTPase activity in vitro. Altogether, these results indicate that hSNF5 affects both the stability and the activity of DNM2, uncovering an unexpected role of hSNF5 in modulating endocytosis, and open new perspectives in understanding the role of hSNF5 in tumour genesis.

[Delay in the acquisition of sucking-swallowing-breathing in the preterm; efects of early stimulation]. / Retraso de la adquisición de la succión-deglución-respiración en el pretérmino; efectos de una estimulación precoz.

INTRODUCTION: Premature baby's oral feeding is not possible until the reflex of sucking-swallowing-breathing adquisition. Its delay extends hospital stay and increases the incidence of oral motor disorders in early childhood. AIMS: To analyze the transition from enteral to oral nutrition, the comorbidity associated with its delay and the impact of an early suction stimulation in a cohort of premature babies. PATIENTS AND METHODS: Retrospective checking of 95 infants less than 32 gestation weeks (GW) admitted to a neonatal ICU in the last 4 years. It was revised the gestational age, anthropometric at birth and discharge, comorbidity, duration of mechanical ventilation, oxygen requirements, time of beginning and end of enteral/oral nutrition, beginning of Kangaroo method and the suction stimulation and the daily weight gain average. RESULTS: Suction stimulation began between weeks 29 and 40 GW (average and median 32 GW). Oral nutrition was initiated between 31-40 GW (average and median 33 GW) and completed between 33-44 GW (average and median 35 GW). Oral nutrition was delayed in patients who required longer mechanical ventilation and oxygen therapy. There was a positive correlation between the beginning of suction stimulation and the time of acquisition of a complete oral nutrition (84% Spearman correlation test) and length of hospital stay (80% Spearman correlation test). CONCLUSIONS: [corrected] Early suction stimulation in a preterm patient seems to facilitate full oral nutrition at an early stage and it is associated with a hospital stay decrease and the improvement in the daily weight gain average.

On the construction of experimental designs for a given task by jointly optimizing several quality criteria: Pareto-optimal experimental designs.

Experimental designs for a given task should be selected on the base of the problem being solved and of some criteria that measure their quality. There are several such criteria because there are several aspects to be taken into account when making a choice. The most used criteria are probably the so-called alphabetical optimality criteria (for example, the A-, E-, and D-criteria related to the joint estimation of the coefficients, or the I- and G-criteria related to the prediction variance). Selecting a proper design to solve a problem implies finding a balance among these several criteria that measure the performance of the design in different aspects. Technically this is a problem of multi-criteria optimization, which can be tackled from different views. The approach presented here addresses the problem in its real vector nature, so that ad hoc experimental designs are generated with an algorithm based on evolutionary algorithms to find the Pareto-optimal front. There is not theoretical limit to the number of criteria that can be studied and, contrary to other approaches, no just one experimental design is computed but a set of experimental designs all of them with the property of being Pareto-optimal in the criteria needed by the user. Besides, the use of an evolutionary algorithm makes it possible to search in both continuous and discrete domains and avoid the need of having a set of candidate points, usual in exchange algorithms.

Improving the visualization of the Pareto-optimal front for the multi-response optimization of chromatographic determinations.

The paper shows tools to visualize and more easily interpret the effect that some experimental factors may exert on analytical responses of interest when optimization of several responses is needed. It is based on an adaptation of the parallel coordinate plot, a tool for graphical representation of points in multidimensional spaces that, theoretically and contrary to the usual Cartesian plots, does not have limits in the dimension of the points being depicted. The joint use of the Pareto-optimal solutions and their visualization allows a deeper knowledge about the problem at hand as well as the wise selection of the conditions of experimental factors for achieving specific goals about the responses. Although the methodology is for a general use, the procedure, its interpretation and usefulness is shown with several analytical cases in chromatography. The first one refers to the experimental conditions to obtain simultaneously the maximum allowable area for both the peak of the malachite green and its metabolite leucomalachite green in fish by liquid chromatography with tandem mass spectrometry detection (LC-MS/MS). The second one is about the simultaneous determination of steroid hormones estrone and 17-α-ethinylestradiol by gas chromatography-mass spectrometry (GC/MS). In the last case, the chromatographic separation by GC/MS of the diastereoisomers, α- and ß-estradiol is needed taking into account that these hormones have the same mass fragments.

Two class-modelling techniques that give families of class-models and their relation with the structure of the data.

In class-modelling problems, which are again becoming increasingly important, there are two parameters to value the quality of the class-model built for a category, namely sensitivity and specificity. Using them as criteria, in this paper, two different approaches to class-modelling problems are presented, approaches that differ from other usual methods in the fact that they provide not just one class-model per category but a set of different class-models that accounts for the possible pairs of sensitivity-specificity values attainable for a given data set. One of the proposals is partial least squares class-modelling (PLS-CM) that, by the joint use of PLS with binary responses and the posterior statistical modelling of the distribution of the computed responses, permits the estimation of the risks related to the decision of assigning a sample into a class, and thus, the values of sensitivity and specificity. The other proposed method, Pareto-optimal front in class-modelling, is an analytical approach posed in a multi-response optimization framework, the one that corresponds to trying to simultaneously maximise the sensitivity and specificity of a class-model. Additionally, the whole family of computed class-models is validated in prediction by using cross-validation, showing the stability of both methods for prediction. The case-studies show the complementariness of both approaches and, in particular, that the joint use of both techniques allows the user to detect possible structures in the data set especially inadequate for PLS. The results, i.e. the whole set of sensitivity-specificity values achievable for a given problem, are graphically represented to improve its study and make it easy to make a decision about the model.

Tutorial on evaluation of type I and type II errors in chemical analyses: from the analytical detection to authentication of products and process control.

Uncertainty is inherent in all experimental determinations. Nevertheless, these measurements are used to make decisions including the performance of the own measurement systems. The link between the decision and the true implicit system that generates the data (measurement system, production process, category of samples, etc.) is a representation of this uncertainty as a probability distribution. This representation leads to the probabilistic formalization of the possibility of making errors. In the context of regulations established by official agencies, it is important to use these statistical decision methods in some cases because the own norm makes them mandatory and, in general, because this is the way of reasonably evaluating whether a working hypothesis is rejected on the basis of the experimental data. The aim of the present tutorial is to introduce some ideas and basic methods for the critical analysis of experimental data. With this goal, the basic elements of the Neyman-Pearson theory of hypothesis testing are formally introduced in connection with the common problems in chemical analysis and, if this is the case, their relation to the norms of regulatory agencies. The notion of decision with 'enough quality' is modelled when explicitly considering: (1) the null, H(0), and alternative, H(1), hypotheses. (2) The significance level of the test, which is the probability, alpha, of rejecting H(0) when it is true, and the power of the test, 1-beta, beta being the probability of accepting H(0) when it is false. (3) The difference between H(0) and H(1) that has to be detected with experimental data. (4) The needed sample size. These four concepts should be explicitly defined for each problem and, under the usual assumption of normal distribution of the data, the mathematical relations among these concepts are shown, which allow the analyst to design a decision rule with pre-set values of alpha and beta. To illustrate the unifying character of this inferential methodology, several situations are exposed along the tutorial: the design of a hypothesis test to decide on the performance characteristics of analytical methods, the capability of detection of both quantitative and qualitative analytical methods (including its generalization to the case of multivariate and/or multiway signals), the analytical sensitivity with multivariate signals, the class-modelling and the process control.

Identification and quantification of ciprofloxacin in urine through excitation-emission fluorescence and three-way PARAFAC calibration.

Due to the second-order advantage, calibration models based on parallel factor analysis (PARAFAC) decomposition of three-way data are becoming important in routine analysis. This work studies the possibility of fitting PARAFAC models with excitation-emission fluorescence data for the determination of ciprofloxacin in human urine. The finally chosen PARAFAC decomposition is built with calibration samples spiked with ciprofloxacin, and with other series of urine samples that were also spiked. One of the series of samples has also another drug because the patient was taking mesalazine. The mesalazine is a fluorescent substance that interferes with the ciprofloxacin. Finally, the procedure is applied to samples of a patient who was being treated with ciprofloxacin. The trueness has been established by the regression "predicted concentration versus added concentration". The recovery factor is 88.3% for ciprofloxacin in urine, and the mean of the absolute value of the relative errors is 4.2% for 46 test samples. The multivariate sensitivity of the fit calibration model is evaluated by a regression between the loadings of PARAFAC linked to ciprofloxacin versus the true concentration in spiked samples. The multivariate capability of discrimination is near 8 microg L(-1) when the probabilities of false non-compliance and false compliance are fixed at 5%.

Pareto-optimal front as a tool to study the behaviour of experimental factors in multi-response analytical procedures.

This work presents a methodology to analyse the behaviour of an analytical procedure, above all when optimization of the procedure is needed. The methodology starts by the design of an experiment suitable to fit response surfaces to some analytical responses of interest in the problem being studied. Then, a pareto-optimal front is estimated that accounts for the optimal possibly trading-off solutions among the responses. The analysis of the behaviour of the optimal values of the response surfaces and the experimental conditions that provide these values allows going deeply into the analytical procedure.

Effect of neuropeptide-EI on the binding of [3H]SCH 23390 to the dopamine D1 receptor in rat striatal membranes.

We have previously demonstrated that neuropeptide-EI, at high doses, stimulates the production of cAMP, in caudate putamen, through the activation of adenylate cyclase coupled to specific D1 receptors. The aim of the present work was to find evidences for a probable interaction between this neuropeptide and the dopamine D1 receptor in the mammalian central nervous system. The present data show that neuropeptide-EI, at high concentrations, affected both the maximum binding and the apparent affinity of [n-methyl-3H] (R)-(+)-8 chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepin-7-ol hemimaleate to the dopamine D1 receptor in a concentration-dependent manner.

Correlation of increased grooming behavior and motor activity with alterations in nigrostriatal and mesolimbic catecholamines after alpha-melanotropin and neuropeptide glutamine-isoleucine injection in the rat ventral tegmental area.

1. We wished to further study the behavioral effects of alpha-melanotropin (alpha-MSH), melanin-concentrating hormone (MCH), and neuropeptide glutamine-isoleucine (NEI). 2. To this effect we administered alpha-MSH, MCH, and NEI in the ventral tegmental area of the rat, a structure where these neuropeptides are highly concentrated. To further elucidate the biochemical mechanisms of the behavioral effect of these neuropeptides, we determined the degree of grooming behavior and the levels of catecholamines. after neuropeptide administration. 3. We preselected those animals responding to the central injection of alpha-MSH with excessive grooming behavior. We administered the neuropeptides at the dose of 1 microg/0.5 microL, in each side of the ventral tegmental area, bilaterally. We studied grooming behavior, locomotor activity, and total behavior scores, 30 and 65 min after administration of the peptides. 4. Three groups of animals were decapitated immediately after the injection of the neuropeptides, and 30 or 65 min after injection. We measured dopamine (DA), noradrenaline (NA), and the dopac/dopamine ratio (DOPAC/DA) to determine steady state levels of catecholamines and an indirect measure of DA release and metabolism, respectively. 5. Injections of alpha-MSH produced significant elevations in grooming behavior, locomotor activity, and total behavior scores, both 30 and 65 min after peptide administration. This was correlated with significant decreases in DA content, increases in DOPAC content, and increases in the DOPAC/DA ratio. In the caudate putamen, changes in catecholamines occurred both at 30 and 65 min after injection. In the nucleus accumbens, changes were present at 65 min after injection. Conversely, there were no alterations in NA content, either in the caudate putamen or in the nucleus accumbens, at any time after the injection. 6. Injections of NEI resulted in significant elevations in grooming behavior, locomotor activity, and total behavior scores, both 30 and 65 min after peptide administration. This was correlated with increased DOPAC/DA ratio in the nucleus caudatus but not in the nucleus accumbens. Conversely, NEI produced increased NA concentrations in the nucleus accumbens, but not in the nucleus caudatus. 7. Injections of MCH did not produce significant changes in behavior or significant changes in nucleus caudatus or nucleus accumbens catecholamines. 8. Our results indicate (a) There is a correlation with alterations in behavior as induced for the neuropeptides injected here, and changes in extrapyramidal catecholamines. (b) There is a correlation between alterations in behavior and increases in DOPAC/DA ratio in the nucleus caudatus. (c) There is a correlation between alterations in behavior and alterations in catecholamines in the nucleus accumbens. In the nucleus accumbens, DOPAC/DA ratio is changed after alpha-MSH, and NA ratio is changed after NEI injection. (d) Absence of alterations in extrapyramidal catecholamines, and in particular in catecholamines in the nucleus accumbens, correlates with absence of behavioral alterations after neuropeptide administration to the ventral tegmental area. 9. In conclusion, the behavioral effect of exogenous administration of neuropeptides in the ventral tegmental area is peptide-specific, and is probably associated with alterations in catecholamine metabolism and release in the nucleus caudatus and the nucleus accumbens. Both alpha-MSH and NEI seem to stimulate the nigrostriatal DA system. While alpha-MSH appears to stimulate the mesolimbic DA system as well, NEI may exert its actions not through the DA, but through the NA mesolimbic system. The precise contribution of DA and NA, and the relative role of the nucleus caudatus and nucleus accumbens in these behaviors remain to be elucidated.

Evidence that alpha-MSH induced grooming is not primarily mediated by any of the cloned melanocortin receptors.

It is well established that melanocortic peptides, such as melanocyte-stimulating hormone (MSH) and adrenocorticotropin, induce grooming behavior. The MC3 and MC4 receptors are the MC receptors which are most abundantly expressed in the brain. gamma-MSH, a peptide with preference to the MC3 receptor, however, does not induce grooming. Recent studies have shown that MC4 receptor antagonists are very effective in inhibiting alpha-MSH induced grooming. These data have indicated that grooming behavior in rodents may be mediated by the MC4 receptor. In this study we investigated if the recently developed MC1 receptor selective agonist MS05 was able to induce grooming in comparison with alpha-MSH. The results show that MS05 is effective in inducing grooming after either intracerebroventricular or ventral tegmental area administration in rats. Central administration of either MS05 or alpha-MSH besides grooming also induced stretching, yawning, rearing and locomotion. The results indicate that the earlier hypothesis that the MC4 receptor is the main mediator of grooming behavior has to be modified. Moreover, as this behaviour does not pharmacologically correlate to the profile of any of the five cloned MC receptors, we suggest that alpha-MSH induced grooming may not primarily be mediated by any of these receptors.

Symbiont survival and host-symbiont disequilibria under differential vertical transmission.

Interspecific genetic interactions in host-symbiont systems raise intriguing coevolutionary questions and may influence the effectiveness of public health and management policies. Here we present an analytical and numerical investigation of the effects of host genetic heterogeneity in the rate of vertical transmission of a symbiont. We consider the baseline case with a monomorphic symbiont and a single diallelic locus in its diploid host, where vertical transmission is the sole force. Our analysis introduces interspecific disequilibria to quantify nonrandom associations between host genotypes and alleles and symbiont presence/absence. The transient and equilibrium behavior is examined in simulations with randomly generated initial conditions and transmission parameters. Compared to the case where vertical transmission rates are uniform across host genotypes, differential transmission (i) increases average symbiont survival from 50% to almost 60%, (ii) dramatically reduces the minimum average transmission rate for symbiont survival from 0.5 to 0.008, and (iii) readily creates permanent host-symbiont disequilibria de novo, whereas uniform transmission can neither create nor maintain such associations. On average, heterozygotes are slightly more likely to carry and maintain the symbiont in the population and are more randomly associated with the symbiont. Results show that simple evolutionary forces can create substantial nonrandom associations between two species.

Attenuation and immunogenicity of Deltacya Deltacrp derivatives of Salmonella choleraesuis in pigs.

Six different isogenic Deltacya Deltacrp derivatives of a strain of Salmonella choleraesuis var. kunzendorf-chi3246 virulent for pigs were constructed by transposon-mediated deletion mutagenesis. These strains were evaluated for virulence and ability to elicit a protective immune response in young weaned pigs after oral administration and were compared to a commercially available vaccine which lacks the 50-kb virulence plasmid (vpl(-)). These derivatives were Deltacya Deltacrp vpl(+), Deltacya Deltacrp vpl(-), Deltacya Delta(crp-cdt) vpl(+), Deltacya Delta(crp-cdt) vpl(-), Deltacya Deltacrp pmi-3834 vpl(+), and Deltacya Delta(crp-cdt) pmi-3834. In experiments to evaluate safety, no significant adverse effects of any of the vaccine constructs were observed, except that two of the strains which carried the virulence plasmid (vpl(+)) caused a small, short-term elevation in maximum temperature compared to pretreatment temperature values. Orally immunized animals, except for those vaccinated with the Deltacya Deltacrp pmi-3834 vpl(+) strain or SC-54, developed significant serum antibody responses 21 days postvaccination as measured by enzyme-linked immunosorbent assay. No cell-mediated immune responses to heat-killed S. choleraesuis were noted at the same time point as measured with heat-killed bacteria as antigen in a lymphocyte proliferation assay. In an oral challenge exposure model with a highly virulent heterologous strain of S. choleraesuis, the Deltacya Deltacrp strains with deletions in pmi were not protective. As measured by morbidity scores, the responses to challenge of the pigs vaccinated with the other four Deltacya Deltacrp derivatives were significantly better than those of the nonvaccinated, challenged group. With the exception of temperature elevation and slight differences in diarrhea scores postchallenge, none of these strains differed significantly from each other in the other clinical parameters analyzed. While the commercial vaccine was protective by most of the parameters measured, it was not fully protective against challenge with virulent S. choleraesuis as judged by diarrhea scores and temperature elevation. Collectively, these data demonstrate that Deltacya Deltacrp derivatives, with or without the virulence plasmid but not with deletions in the pmi gene, are candidates for vaccines for protection against salmonellosis in pigs.

Effects and interactions between alpha-MSH and MCH/NEI upon striatal cAMP levels.

It is known that alpha-MSH augments cAMP levels in rat brain slices containing accumbens and caudate-putamen nuclei. In this study we examined: a) the effect of other neuropeptides: MCH and NEI, on this cyclic nucleotide; b) if the effects of alpha-MSH on cAMP production can be modulated by addition of MCH or NEI to the incubation medium. Both MCH and NEI (3.6 microM) increased the production of cAMP, whereas at doses of 0.6 microM exerted no effects. When alpha-MSH 0.6 microM was added with NEI or MCH (0.6 microM), only MCH blocked the increase in the cAMP induced by alpha-MSH. Neither MCH nor NEI at the highest dose used (3.6 microM) had any additive effect on AMPc when added together with alpha-MSH. We conclude that, at a high concentration, (MCH/NEI)-like peptides can use the intracellular signal transduction linked to cyclic nucleotides in the CNS.