Development, characterization, and evaluation of a simple polymicrobial colony biofilm model for testing of antimicrobial wound dressings.

Chronic wound infections are generally of polymicrobial nature with aerobic and anaerobic bacteria, as well as fungi frequently observed in them. Wound treatment involves a series of steps, including debridement of the wound, flushing, and often the use of multiple wound dressings many of which are antimicrobial. Yet, many wound dressings are tested versus single species of planktonic microbes, which fails to mirror the real-life presence of biofilms. AIMS: Simple biofilm models are the first step to testing of any antimicrobial and wound dressing; therefore, the aim of this study was to develop and validate a simple polymicrobial colony biofilm wound model comprised of Pseudomonas aeruginosa, Staphylococcus aureus, and Candida albicans on RPMI-1640 agar. The model was then used to evaluate the topical disinfectant chlorohexidine and four commercially available wound dressings using the polymicrobial model. The model used was as a starting point to mimic debridement in clinical care of wounds and the effectiveness of wound dressings evaluated afterwards. METHODS AND RESULTS: Planktonic assessment using AATCC100-2004 demonstrated that all antimicrobial wound dressings reduced the planktonic microbial burden below the limit of detection; however, when challenged with polymicrobial colony biofilms, silver wound dressings showed limited effectiveness (1-2 log CFU reductions). In contrast, a single iodine releasing wound dressing showed potent antibiofilm activity reducing all species CFUs below the limit of detection (>6-10 log) depending on the species. A disrupted biofilm model challenge was performed to represent the debridement of a wound and wound silver-based wound dressings were found to be marginally more effective than in whole colony biofilm challenges while the iodine containing wound dressing reduced microbial recovery below the limit of detection. CONCLUSIONS: In this model, silver dressings were ineffective versus the whole colony biofilms but showed some recovery of activity versus the disrupted colony biofilm. The iodine wound dressing reduced the viability of all species below the level of detection. This suggests that mode of action of wound dressing should be considered for the type of biofilm challenge as should the clinical use, e.g. debridement.

Probing Interkingdom Signaling Molecules via Liquid Extraction Surface Analysis-Mass Spectrometry.

Previously, metabolites diffused or secreted from microbial samples have been analyzed via liquid chromatography-mass spectrometry (LC-MS) approaches following lengthy extraction protocols. Here, we present a model system for growing biofilms on discs before utilizing rapid and direct surface sampling MS, namely, liquid extraction surface analysis, to study the microbial exometabolome. One of the benefits of this approach is its surface-specific nature, enabling mimicking biofilm formation in a way that the study of planktonic liquid cultures cannot imitate. Even though Pseudomonas aeruginosa (P. aeruginosa), Staphylococcus aureus (S. aureus), and Candida albicans (C. albicans) have been studied previously in isolation, very few studies consider the complexity of the interplay between these pathogens, which are commonly combined causative agents of infection. Our model system provides a route to investigate changes in the exometabolome, such as metabolites that become circulatory in the presence of multiple pathogens. Our results agree with previous reports showing that 2-alkyl-4(1H)-quinolone signal molecules produced by P. aeruginosa are important markers of infection and suggest that methods for monitoring levels of 2-heptyl-4-hydroxyquinoline and 2,4-dihydroxyquinoline, as well as pyocyanin, could be beneficial in the determination of causative agents in interkingdom infection including P. aeruginosa. Furthermore, studying changes in exometabolome metabolites between pqs quorum sensing antagonists in treated and nontreated samples suggests suppression of phenazine production by P. aeruginosa. Hence, our model provides a rapid analytical approach to gaining a mechanistic understanding of bacterial signaling.

Growth rate and nutrient limitation as key drivers of extracellular quorum sensing signal molecule accumulation in Pseudomonas aeruginosa.

In Pseudomonas aeruginosa, quorum sensing (QS) depends on an interconnected regulatory hierarchy involving the Las, Rhl and Pqs systems, which are collectively responsible for the co-ordinated synthesis of a diverse repertoire of N-acylhomoserine lactones (AHLs) and 2-alkyl-4-quinolones (AQs). Apparent population density-dependent phenomena such as QS may, however, be due to growth rate and/or nutrient exhaustion in batch culture. Using continuous culture, we show that growth rate and population density independently modulate the accumulation of AHLs and AQs such that the highest concentrations are observed at a slow growth rate and high population density. Carbon source (notably succinate), nutrient limitation (C, N, Fe, Mg) or growth at 25 °C generally reduces AHL and AQ levels, except for P and S limitation, which result in substantially higher concentrations of AQs, particularly AQ N-oxides, despite the lower population densities achieved. Principal component analysis indicates that ~26 % variation is due to nutrient limitation and a further 30 % is due to growth rate. The formation of N-(3-oxododecanoyl)-l-homoserine lactone (3OC12-HSL) turnover products such as the ring opened form and tetramic acid varies with the limiting nutrient limitation and anaerobiosis. Differential ratios of N-butanoyl-homoserine lactone (C4-HSL), 3OC12-HSL and the AQs as a function of growth environment are clearly apparent. Inactivation of QS by mutation of three key genes required for QS signal synthesis (lasI, rhlI and pqsA) substantially increases the concentrations of key substrates from the activated methyl cycle and aromatic amino acid biosynthesis, as well as ATP levels, highlighting the energetic drain that AHL and AQ synthesis and hence QS impose on P. aeruginosa.

Disruption of the Pseudomonas aeruginosa Tat system perturbs PQS-dependent quorum sensing and biofilm maturation through lack of the Rieske cytochrome bc1 sub-unit.

Extracellular DNA (eDNA) is a major constituent of the extracellular matrix of Pseudomonas aeruginosa biofilms and its release is regulated via pseudomonas quinolone signal (PQS) dependent quorum sensing (QS). By screening a P. aeruginosa transposon library to identify factors required for DNA release, mutants with insertions in the twin-arginine translocation (Tat) pathway were identified as exhibiting reduced eDNA release, and defective biofilm architecture with enhanced susceptibility to tobramycin. P. aeruginosa tat mutants showed substantial reductions in pyocyanin, rhamnolipid and membrane vesicle (MV) production consistent with perturbation of PQS-dependent QS as demonstrated by changes in pqsA expression and 2-alkyl-4-quinolone (AQ) production. Provision of exogenous PQS to the tat mutants did not return pqsA, rhlA or phzA1 expression or pyocyanin production to wild type levels. However, transformation of the tat mutants with the AQ-independent pqs effector pqsE restored phzA1 expression and pyocyanin production. Since mutation or inhibition of Tat prevented PQS-driven auto-induction, we sought to identify the Tat substrate(s) responsible. A pqsA::lux fusion was introduced into each of 34 validated P. aeruginosa Tat substrate deletion mutants. Analysis of each mutant for reduced bioluminescence revealed that the primary signalling defect was associated with the Rieske iron-sulfur subunit of the cytochrome bc1 complex. In common with the parent strain, a Rieske mutant exhibited defective PQS signalling, AQ production, rhlA expression and eDNA release that could be restored by genetic complementation. This defect was also phenocopied by deletion of cytB or cytC1. Thus, either lack of the Rieske sub-unit or mutation of cytochrome bc1 genes results in the perturbation of PQS-dependent autoinduction resulting in eDNA deficient biofilms, reduced antibiotic tolerance and compromised virulence factor production.

Evaluation of a high resolution diode array for CyberKnife quality assurance.

PURPOSE: The CyberKnife quality assurance (QA) program relies mainly on the use of radiochromic film (RCF). We aimed at evaluating high-resolution arrays of detectors as an alternative to films for CyberKnife machine QA. METHODS: This study will test the SRS Mapcheck (Sun Nuclear, Melbourne, Florida, USA) diode array and its own software, which allows three tests of the CyberKnife QA program to be performed. The first one is a geometrical accuracy test based on the delivery of two orthogonal beams (Automated Quality Assurance, AQA). Besides comparing the constancy and repeatability of both methods, known errors will be introduced to check their sensitivity. The second checks the constancy of the iris collimator field sizes (Iris QA). Changes in the field sizes will be introduced to study the array sensitivity. The last test checks the correct positioning of the multileaf collimator (MLC). It will be tested introducing known systematic displacements to whole banks and to single leaves. RESULTS: The results of the RCF and diode array were equivalent (maximum differences of 0.18 ± 0.14 mm) for the AQA test, showing the array a higher reproducibility. When known errors were introduced, both methods behaved linearly with similar slopes. Regarding Iris QA, the array measurements are highly linear when changes in the field sizes are introduced. Linear regressions show slopes of 0.96-1.17 with r2 above 0.99 in all field sizes. Diode array seems to detect changes of 0.1 mm. In MLC QA, systematic errors of the whole bank of leaves were not detected by the array, while single leaf errors were detected. CONCLUSIONS: The diode array is sensitive and accurate in the AQA and Iris QA tests, which give us the possibility of substituting RCF with a diode array. QA would be performed faster than using the film procedure, obtaining reliable results. Regarding the MLC QA, the inability to detect systematic displacements make it difficult to confidently use the detector.

The use of a thin guide-wire for urethral definition in prostate SBRT treatments with Cyberknife.

PURPOSE: To study and analyze the effect of the use of a thin guide-wire instead of a Foley catheter for urethral definition in prostate stereotactic body radiation therapy (SBRT) treatments and to compare treatment parameters in both situations. MATERIAL AND METHODS: Thirty-seven prostate SBRT patients were employed in this study. A Foley catheter was employed in nine of them, and a guide-wire was employed in the other 28 patients. For each of the 28 patients in which the guide-wire was employed, a comparison between urethral positions in both situations was performed, allowing for a margin definition of the urethra when a Foley catheter was employed. Displacements of the prostate during treatment were obtained, allowing for an analysis of prostate positions in both situations. Also, different treatment parameters such as the number of treatment interruptions, couch movements performed, and x-rays needed were gathered. RESULTS: Large differences between urethral positions can be found in the anterior-posterior (AP) directions compared to those in the lateral (LAT) direction. Differences are also larger in areas closer to the base of the prostate, where margins applied in the case of using a Foley catheter are 16 mm with a mean displacement of 6 mm in the posterior direction. No differences in the treatment parameters were found during treatment in both situations. The difference found in absolute prostate pitch rotations suggests that the Foley catheter provokes a shift of the prostate position, which does not occur when employing the guide-wire. CONCLUSIONS: Foley catheters shift the urethral position, making them a wrong surrogate of the urethra when no catheters are present. The margins needed to assess uncertainties introduced by the use of a Foley catheter are larger than those usually applied. The use of a Foley catheter did not present any additional difficulty during treatment delivery in terms of images employed or interruptions produced.

Assessment of the Use of Common Juniper (Juniperus communis L.) Foliage following the Cascade Principle.

Juniperus communis L. is a species commonly grown in regions of the Northern Hemisphere, and is a good candidate to be cultivated in marginal lands. Plants coming from a pruning performed in a natural population located in Spain were used to assess the yield and quality of different products obtained following the cascade principle. A total of 1050 kg of foliage biomass were crushed, steam-distilled, and separated into fractions to produce biochar and absorbents for the pet industry using pilot plants. The obtained products were analysed. The essential oil, with a yield of 0.45% dry basis and a qualitative chemical composition similar to that described for the berries in international standards or monographs, showed antioxidant activity with promising CAA results (inhibition of 89% of the cell's oxidation). However, regarding antibacterial and antifungal activities, it only inhibited the growth of microorganisms at the maximum concentration tested, 2.5%. Concerning the hydrolate, it did not show bioactivity. Regarding the biochar, whose yield was 28.79% dry basis, interesting results were obtained for its characterisation as a possible soil improver for agronomic purposes (PFC 3(A)). Finally, promising results were obtained regarding the use of common juniper as absorbent, taking into account the physical characterisation and odour control capacity.

Mushroom-shaped structures formed in Acinetobacter baumannii biofilms grown in a roller bioreactor are associated with quorum sensing-dependent Csu-pilus assembly.

There is currently a need to develop simple biofilm models that facilitate investigation of the architecture/biology of mature bacterial biofilms in a consistent/standardized manner given their environmental and clinical importance and the need for new anti-biofilm interventions. This study introduces a novel biofilm culture system termed the rolling biofilm bioreactor (RBB). This easily operated system allows adherent microbial cells to be repeatedly exposed to air/solid/liquid interfaces optimizing biofilm growth. The RBB was exploited to investigate biofilm formation in Acinetobacter baumannii. High levels of A. baumannii biofilm biomass reproducibly accumulate in the RBB and, importantly, undergo a maturation step to form large mushroom-shaped structures that had not been observed in other models. Based on image analysis of biofilm development and genetic manipulation, we show how N-acylhomoserine lactone-dependent quorum sensing (QS) impacts on biofilm differentiation, composition and antibiotic tolerance. Our results indicate that extracellular DNA (eDNA) is a key matrix component in mature Acinetobacter biofilms as the mushroom-like structures consist of dense cellular masses encased in an eDNA mesh. Moreover, this study reveals the contribution of QS to A. baumannii biofilm differentiation through Csu pilus assembly regulation. Understanding the mechanisms of structural development of mature biofilms helps to identify new biofilm eradication and removal strategies.

Azospirillum baldaniorum Sp245 exploits Pseudomonas fluorescens A506 biofilm to overgrow in dual-species macrocolonies.

Biofilms are essential for plant-associated bacteria to colonize their host. In this work, we analysed the interaction of Azospirillum baldaniorum Sp245 and Pseudomonas fluorescens A506 in mixed macrocolony biofilms. We identified certain culture conditions where A. baldaniorum Sp245 exploits P. fluorescens A506 to boost its growth. Azospirillum growth increased proportionally to the initial number of pseudomonads building the biofilm, which in turn were negatively affected in their growth. Physical contact with P. fluorescens A506 was essential for A. baldaniorum Sp245 growth increase. Biofilm ultrastructure analysis revealed that Pseudomonas produces a thick structure that hosts Azospirillum cells in its interior. Additional experimentation demonstrated that Azospirillum growth boost is compromised when interacting with biofilm-deficient Pseudomonas mutants, and that a low oxygen concentration strongly induce A. baldaniorum Sp245 growth, overriding Pseudomonas stimulation. In this line, we used a microaerophilia reporter strain of A. baldaniorum Sp245 to confirm that dual-species macrocolonies contain a higher number of cells under microaerophilic conditions. Taking all the results into consideration, we propose that A. baldaniorum Sp245 can benefit from P. fluorescens A506 partnership in mixed biofilms by taking advantage of the low oxygen concentration and scaffold made up of Pseudomonas-derived matrix, to expand its growth.

Symbiopectobacterium purcellii, gen. nov., sp. nov., isolated from the leafhopper Empoasca decipiens.

Bacterial endosymbionts are found in multiple arthropod species, where they play crucial roles as nutritional symbionts, defensive symbionts or reproductive parasites. Recent work has highlighted a new clade of heritable microbes within the gammaproteobacteria that enter into both obligate and facultative symbioses, with an obligately required unculturable symbiont recently given the name Candidatus Symbiopectobacterium. In this study, we describe a culturable rod shaped non-flagellated bacterial symbiont from this clade isolated from the leafhopper Empoasca decipiens. The symbiont is related to the transovarially transmitted 'BEV' bacterium that was first isolated from the leafhopper Euscelidius variegatus by Alexander Purcell, and we therefore name the symbiont Symbiopectobacterium purcellii sp. nov., gen. nov. We further report the closed genome sequence for S. purcellii. The genome is atypical for a heritable microbe, being large in size, without profound AT bias and with little evidence of pseudogenization. The genome is predicted to encode Type II, III and VI secretion systems and associated effectors and a non-ribosomal peptide synthase array likely to produce bioactive small molecules. The predicted metabolism is more complete than for other symbionts in the Symbiopectobacterium clade, and the microbe is predicted to synthesize a range of B vitamins. However, Biolog plate results indicate that the metabolism is depauperate compared to the sister clade, represented by Pectobacterium carotovorum. A quorum-sensing pathway related to that of Pectobacterium species (containing an overlapping expI-expR1 pair in opposite directions and a "solo" expR2) is evidenced, and LC-MS/MS analysis reveals the presence of 3-hydroxy-C10-HSL as the sole N-acylhomoserine lactone (AHL) in our strain. This AHL profile is profoundly divergent from that of other Erwinia and Pectobacterium species which produce mostly 3-oxo-C6- and 3-oxo-C8-HSL and could aid group identification. Thus, this microbe denotes one that has lost certain pathways associated with a saprophytic lifestyle but represents an important baseline against which to compare other members of the genus Symbiopectobacterium that show more profound integration into host biology. The type strain of Symbiopectobacterium purcellii gen. nov., sp. nov. is SyEd1T (LMG 32449T=CECT 30436T).

Associations of Mutually Exclusive Categories of Physical Activity and Sedentary Time With Metabolic Syndrome in Older Adults: An Isotemporal Substitution Approach.

The aim of this study was to examine, theoretically, how reallocating time between the intensity of mutually exclusive categories of physical activity and sedentary behavior time is associated with metabolic syndrome. Four hundred and six older adults (61.6% women) from the second wave of the EpiFloripa Aging Cohort Study were included in the study (mean age 71.7 ± 5.9 years). Isotemporal substitution analysis showed a decrease of 35% (odds ratio: 0.65; 95% confidence interval [0.45, 0.96]) in the risk for metabolic syndrome when replacing 30 min/day of sedentary behavior with an equivalent amount of moderate to vigorous physical activity. Furthermore, it has been observed that older adults classified as low sedentary behavior and physically active were 57% less likely to have metabolic syndrome than participants classified as high sedentary and physically inactive (odds ratio: 0.43; 95% confidence interval [0.19, 0.97]). This study highlights the importance of behavioral categories that may emerge concerning the interrelationships of physical activity and health in older adults, having important implications for future health intervention programs.

AhaP, A Quorum Quenching Acylase from Psychrobacter sp. M9-54-1 That Attenuates Pseudomonas aeruginosa and Vibrio coralliilyticus Virulence.

Although Psychrobacter strain M9-54-1 had been previously isolated from the microbiota of holothurians and shown to degrade quorum sensing (QS) signal molecules C6 and C10-homoserine lactone (HSL), little was known about the gene responsible for this activity. In this study, we determined the whole genome sequence of this strain and found that the full 16S rRNA sequence shares 99.78-99.66% identity with Psychrobacter pulmonis CECT 5989T and P. faecalis ISO-46T. M9-54-1, evaluated using the agar well diffusion assay method, showed high quorum quenching (QQ) activity against a wide range of synthetic N-acylhomoserine lactone (AHLs) at 4, 15, and 28 °C. High-performance liquid chromatography-mass-spectrometry (HPLC-MS) confirmed that QQ activity was due to an AHL-acylase. The gene encoding for QQ activity in strain M9-54-1 was identified from its genome sequence whose gene product was named AhaP. Purified AhaP degraded substituted and unsubstituted AHLs from C4- to C14-HSL. Furthermore, heterologous expression of ahaP in the opportunistic pathogen Pseudomonas aeruginosa PAO1 reduced the expression of the QS-controlled gene lecA, encoding for a cytotoxic galactophilic lectin and swarming motility protein. Strain M9-54-1 also reduced brine shrimp mortality caused by Vibrio coralliilyticus VibC-Oc-193, showing potential as a biocontrol agent in aquaculture.

The impaired quorum sensing response of Pseudomonas aeruginosa MexAB-OprM efflux pump overexpressing mutants is not due to non-physiological efflux of 3-oxo-C12-HSL.

Multidrug (MDR) efflux pumps are ancient and conserved molecular machineries with relevant roles in different aspects of the bacterial physiology, besides antibiotic resistance. In the case of the environmental opportunistic pathogen Pseudomonas aeruginosa, it has been shown that overexpression of different efflux pumps is linked to the impairment of the quorum sensing (QS) response. Nevertheless, the causes of such impairment are different for each analysed efflux pump. Herein, we performed an in-depth analysis of the QS-mediated response of a P. aeruginosa antibiotic resistant mutant that overexpresses MexAB-OprM. Although previous work claimed that this efflux pump extrudes the QS signal 3-oxo-C12-HSL, we show otherwise. Our results evidence that the observed attenuation in the QS response when overexpressing this pump is related to an impaired production of alkyl quinolone QS signals, likely prompted by the reduced availability of one of their precursors, the octanoate. Together with previous studies, this indicates that, although the consequences of overexpressing efflux pumps are similar (impaired QS response), the underlying mechanisms are different. This 'apparent redundancy' of MDR efflux systems can be understood as a P. aeruginosa strategy to keep the robustness of the QS regulatory network and modulate its output in response to different signals.

Genome-wide mapping of the RNA targets of the Pseudomonas aeruginosa riboregulatory protein RsmN.

Pseudomonads typically carry multiple non-identical alleles of the post-transcriptional regulator rsmA. In Pseudomonas aeruginosa, RsmN is notable in that its structural rearrangement confers distinct and overlapping functions with RsmA. However, little is known about the specificities of RsmN for its target RNAs and overall impact on the biology of this pathogen. We purified and mapped 503 transcripts directly bound by RsmN in P. aeruginosa. About 200 of the mRNAs identified encode proteins of demonstrated function including some determining acute and chronic virulence traits. For example, RsmN reduces biofilm development both directly and indirectly via multiple pathways, involving control of Pel exopolysaccharide biosynthesis and c-di-GMP levels. The RsmN targets identified are also shared with RsmA, although deletion of rsmN generally results in less pronounced phenotypes than those observed for ΔrsmA or ΔrsmArsmNind mutants, probably as a consequence of different binding affinities. Targets newly identified for the Rsm system include the small non-coding RNA CrcZ involved in carbon catabolite repression, for which differential binding of RsmN and RsmA to specific CrcZ regions is demonstrated. The results presented here provide new insights into the intricacy of riboregulatory networks involving multiple but distinct RsmA homologues.

Gamma Interferon and Interleukin-17A Differentially Influence the Response of Human Macrophages and Neutrophils to Pseudomonas aeruginosa Infection.

Macrophages are important orchestrators of inflammation during bacterial infection, acting as both effector cells and regulators of neutrophil recruitment and life span. Differently activated macrophage populations with distinct inflammatory and microbicidal potentials have been described. Our previous work unveiled a positive and a negative correlation between levels of gamma interferon (IFN-γ) and interleukin-17A (IL-17A), respectively, and lung function in cystic fibrosis, particularly in patients chronically infected with Pseudomonas aeruginosa This study sought to define key parameters in human antibacterial immunity under Th1- and Th17-dominated inflammatory conditions; the final aim was to identify unique characteristics that could be fine-tuned therapeutically to minimize tissue damage while maximizing bacterial clearance. Toward this aim, neutrophils were incorporated into cultures of macrophages treated with IFN-γ or IL-17A and infected with P. aeruginosa The intent of this design was to model (i) initiation of inflammation by infected macrophages and (ii) delayed arrival of neutrophils and their exposure to macrophage-derived cytokines. Under these conditions, IFN-γ decreased bacterial killing and promoted the production of monocyte chemoattractant protein 1 (MCP-1). In contrast, IL-17A promoted bacterial killing but did not affect MCP-1 production. The level of secretion of the pyrogen IL-1ß was significantly lower in the presence of IFN-γ than in the presence of IL-17A and correlated with levels of the IL1B transcript in infected macrophages. These findings support the validity of this model to investigate human antibacterial immunity. Based on these observations, the protective and damaging roles of IFN-γ and IL-17A, respectively, during P. aeruginosa infection could be caused by their contrasting effects on IL-1ß and MCP-1 production.

Associations of daily activities measured by a pattern-recognition activity monitor with overall and abdominal obesity in older people: the IMPACT65+ study.

OBJECTIVES: The aims of the present study were: (i) to analyze the associations of the time spent in daily activities (i.e., lie, recline, passive sit, active sit, stand, walk at slow pace, walk at average pace, walk at brisk pace, and other activities) with body mass index (BMI) and waist circumference (WC); and (ii) to examine how theoretically reallocating time between these daily activities is associated with BMI and WC. METHODS: The sample included 437 older adults (288 women), aged 65 to 92 years, participating in the IMPACT65+ study. The time in daily activities was assessed by the Intelligent Device for Energy Expenditure and Activity (IDEEA). BMI and WC were measured following standardized procedures. Associations of daily activities with BMI and WC were examined using linear regression models adjusting for potential confounders. Isotemporal substitution models were performed to estimate the theoretical effect of replacing one activity with another activity while holding total time constant. RESULTS: The time spent lying and reclining was associated with increased BMI and WC, while the time spent standing, walking at average pace, and walking at brisk pace was associated with decreased BMI and WC. Isotemporal substitution analyses revealed significant hypothetical reductions in BMI and WC when reallocating 15 min from lying or reclining to standing or walking at average pace. Moreover, replacing 15 min from any sedentary activity or light physical activity (except for walking at average pace) with an equal amount of time in walking at brisk pace was associated with lower BMI and WC. CONCLUSIONS: This is the first study examining the activity specific and isotemporal associations of daily behaviors (considering body postures and movements) with overall and abdominal obesity in older people. The results could be used in the development of specific recommendations encouraging an active lifestyle in older people.

Interday Reliability of the IDEEA Activity Monitor for Measuring Movement and Nonmovement Behaviors in Older Adults.

The interday reliability of the Intelligent Device for Energy Expenditure and Activity (IDEEA) has not been studied to date. The study purpose was to examine the interday variability and reliability on two consecutive days collected with the IDEEA, as well as to predict the number of days needed to provide a reliable estimate of several movement (walking and climbing stairs) and nonmovement (lying, reclining, and sitting) behaviors and standing in older adults. The sample included 126 older adults (74 women) who wore the IDEEA for 48 hr. Results showed low variability between the 2 days, and the reliability was from moderate (intraclass coefficient correlation = .34) to high (.80) in most of movement and nonmovement behaviors analyzed. The Bland-Altman plots showed high-moderate agreement between days, and the Spearman-Brown formula estimated that 1.2 and 9.1 days of monitoring with the IDEEA are needed to achieve intraclass coefficient correlations ≥ .70 in older adults for sitting and climbing stairs, respectively.

Dissipation of Three Fungicides and Their Effects on Anthocyanins and Color of Monastrell Red Wines.

The effect of fungicides on fermentation is of paramount importance to control the quality and safety of wines. In this work, the quality (enological parameters, color, phenolic content, antioxidant activity, and fungicide residues) of wines from Monastrell grapes fortified with iprovalicarb, mepanipyrim, and tetraconazole fungicides was evaluated. Along the winemaking process, initial residues of mepanipyrim and tetraconazole were removed in more than 90% while the dissipation of iprovalicarb was around 73%. Significant statistical differences were found in the presence of iprovalicarb and mepanipyrim residues, especially at the highest concentration assayed. For both fungicides, increases in the volatile acidity (between 4 and 8.6 times), the lactic acid content (between 8.6 and 20.5 times), the percentage of polymeric anthocyanins (between 1.3 and 1.7 times), and also a slight increase of the total phenolic index and the total anthocyanin content determined by spectrophotometry were observed. On the contrary, the total monomeric anthocyanins content decreased about 16.3% and 28.6% in the presence of iprovalicarb and mepanipyrim, respectively. These results could be related to a higher development of acetic acid or lactic bacteria in the presence of these fungicides. The color of the final wines was also different in comparison with the control, with a higher yellow component, color intensity, tonality, and hue angle because of pH changes in the medium. Tetraconazole fermentations had a more similar trend to the control wine, probably due to the lower concentration of this fungicide in the grape must at the initial time. No effects on the antioxidant activity was observed for any of the target fungicides. A multivariate statistical analysis was done to view the interrelationships between different variables (color and anthocyanins profile). The obtained model allowed the wines to be separated according to the fungicide treatment applied.

Influence of fungicide residues and in vitro gastrointestinal digestion on total antioxidant capacity and phenolic fraction of Graciano and Tempranillo red wines.

The effect of fenhexamid, mepanipyrim and cyazofamid fungicides on in vitro bioavailability of antioxidant activity and phenolic compounds of Tempranillo and Graciano red wine was studied by simulating the digestive process by dialysis in semipermeable membranes. Determination of antioxidant activity was through reaction with the DPPH • radical and the measurement of phenolic compounds was made with liquid chromatography with diode detector (HPLC-DAD). Fenhexamid, mepanipyrim and cyazofamid reduce the total polyphenol content in both wines. During dialysis there was a large loss of total polyphenols (80-90%) and of antioxidant activity (> 90%). The bioavailability of the phenolic compounds is lower than that for non-treated wines and the highest dialization percentages were found for stilbenes > 50%. While for the remaining phenolic fraction the order is the following hydroxycinnamic derivatives > anthocyanins > flavonols.