Efficiency of Electronic Nose in Detecting the Microbial Spoilage of Fresh Sardines (Sardinella longiceps).

The assessment of microbial spoilage in fresh fish is a major concern for the fish industry. This study aimed to evaluate the efficiency and reliability of an electronic nose (E-nose) to detect microbial spoilage of fresh sardines (Sardinella longiceps) by comparing its measurements with Total Bacterial Count (TBC), Hydrogen Sulfide (H2S) producing bacterial count and Trimethylamine Oxide (TMAO) reducing bacterial count after variable storage conditions. The samples were stored at 0 °C (0, 2, 4, 6, and 8 days) and 25 °C (0, 3, 6, and 9 h), while day 0 was used as a control. The E-nose measurements were analyzed by Principal Component Analysis (PCA), Linear Discriminant Analysis (LDA) and Artificial Neural Network (ANN). Microbial counts increased significantly and simultaneously with the changes in E-nose measurements during storage. The LDA and ANN showed a good classification of E-nose data for different storage times at two storage temperatures (0 °C and 25 °C) compared to PCA. It is expected as PCA is based on linear relationships between the factors, while ANN is based on non-linear relationships. Correlation coefficients between E-nose and TBC, TMAO-reducing bacterial and H2S-producing bacterial counts at 0 °C were 0.919, 0.960 and 0.915, respectively, whereas at 25 °C, the correlation coefficients were 0.859, 0.945 and 0.849, respectively. These positive correlations qualify the E-nose as an efficient and reliable device for detecting microbial spoilage of fish during storage.

Characterization of Streptomyces Cell Surface by the Microbial Adhesion to Solvents Method.

The cell surface physicochemical properties of Streptomyces should influencing the dispersal and adsorption of spores and hyphae in soil and should conditioning there interactions with organic or metal substances in the bioremediation of contaminated environment. These properties are concerning surface hydrophobicity, electron donor/acceptor, and charge surface. To date, only hydrophobicity of Streptomyces was studied by contact angle measurements and microbial adhesion to hydrocarbons (MATH). In this work, we studied the electron donor/acceptor character of the Streptomyces cell surface in two ionic strength 10-3 M and 10-1 M of KNO3. Thus, to facilitate the characterisation of the surfaces of microbial cells, we used a simple, rapid, and quantitative technique, the microbial adhesion method to solvents (MATS), which is based on the comparison of the affinity of microbial cells for a monopolar solvent with a polar solvent. The monopolar solvent can be acid (electron acceptor) or basic (electron donor), but both solvents should have a surface tension similar to that of the Kifshitz van der Waals components. At the significant ionic strength of the biological medium, the electron donor character is well expressed for all 14 Streptomyces strains with very significant differences among them ranging from 0% to 72.92%. When the cells were placed in a solution with a higher ionic strength, we were able to classify the donor character results into three categories. The first category is that the weak donor character of strains A53 and A58 became more expressed at 10-1 M KNO3 concentration. The second category is that three strains A30, A60, and A63 expressed a weaker character in a higher ionic strength. For the other strains, no expression of the donor trait was obtained at higher ionic strength. In a suspension with a concentration of 10-3 KNO3, only two strains expressed an electron acceptor character. This character is very important for strains A49, A57, A58, A60, A63, and A65 at 10-1M KNO3. This work has shown that these properties vary greatly depending on the Streptomyces strain. It is important to consider the change in physicochemical properties of surface cells with ionic strength when using Streptomyces in different bioprocesses.

Current Understanding on Adhesion and Biofilm Development in Actinobacteria.

Biofilm formation and microbial adhesion are two related and complex phenomena. These phenomena are known to play an important role in microbial life and various functions with positive and negative aspects. Actinobacteria have wide distribution in aquatic and terrestrial ecosystems. This phylum is very large and diverse and contains two important genera Streptomyces and Mycobacteria. The genus Streptomyces is the most biotechnologically important, while the genus Mycobacteria contains the pathogenic species of Mycobacteriaceae. According to the literature, the majority of studies carried out on actinomycetes are focused on the detection of new molecules. Despite the well-known diversity and metabolic activities, less attention has been paid to this phylum. Research on adhesion and biofilm formation is not well developed. In the present review, an attempt has been made to review the literature available on the different aspects on biofilm formation and adhesion of Actinobacteria. We focus especially on the genus Streptomyces. Furthermore, a brief overview about the molecules and structures involved in the adhesion phenomenon in the most relevant genus is summarized. We mention the mechanisms of quorum sensing and quorum quenching because of their direct association with biofilm formation.

Clinical laboratory services for primary healthcare centers in urban cities: a pilot ACO model of ten primary healthcare centers.

BACKGROUND: Primary healthcare centers (PHC) ensure that patients receive comprehensive care from promotion and prevention to treatment, rehabilitation, and palliative care in a familiar environment. It is designed to provide first-contact, continuous, comprehensive, and coordinated patient care that will help achieve equity in the specialty healthcare system. The healthcare in Saudi Arabia is undergoing transformation to Accountable Care Organizations (ACO) model. In order for the Kingdom of Saudi Arabia (KSA) to achieve its transformational goals in healthcare, the improvement of PHCs' quality and utilization is crucial. An integral part of this service is the laboratory services. METHODS: This paper presents a pilot model for the laboratory services of PHC's in urban cities. The method was based on the FOCUS-PDCA quality improvement method focusing on the pre-analytical phase of the laboratory testing as well as the Saudi Central Board for Accreditation of Healthcare Institutes (CBAHI) gap analysis and readiness within the ten piloted primary healthcare centers. RESULTS: The Gap analysis, revealed in-consistency in the practice, lead to lower the quality of the service, which was seen in the low performance of the chosen key performance indicators (KPI's) (high rejection rates, lower turn-around times (TAT) for test results) and also in the competency of the staff. Following executing the interventions, and by using some of the ACO Laboratory strategies; the KPI rates were improved, and our results exceeded the targets that we have set to reach during the first year. Also introducing the electronic connectivity improved the TAT KPI and made many of the processes leaner. CONCLUSIONS: Our results revealed that the centralization of PHC's laboratory service to an accredited reference laboratory and implementing the national accreditation standards improved the testing process and lowered the cost, for the mass majority of the routine laboratory testing. Moreover, the model shed the light on how crucial the pre-analytical phase for laboratory quality improvement process, its effect on cost reduction, and the importance of staff competency and utilization.

Novel Lipid Species for Detecting and Predicting Atrial Fibrillation in Patients With Type 2 Diabetes.

The incidence of atrial fibrillation (AF) is higher in patients with diabetes. The goal of this study was to assess if the addition of plasma lipids to traditional risk factors could improve the ability to detect and predict future AF in patients with type 2 diabetes. Logistic regression models were used to identify lipids associated with AF or future AF from plasma lipids (n = 316) measured from participants in the ADVANCE trial (n = 3,772). To gain mechanistic insight, follow-up lipid analysis was undertaken in a mouse model that has an insulin-resistant heart and is susceptible to AF. Sphingolipids, cholesteryl esters, and phospholipids were associated with AF prevalence, whereas two monosialodihexosylganglioside (GM3) ganglioside species were associated with future AF. For AF detection and prediction, addition of six and three lipids, respectively, to a base model (n = 12 conventional risk factors) increased the C-statistics (detection: from 0.661 to 0.725; prediction: from 0.674 to 0.715) and categorical net reclassification indices. The GM3(d18:1/24:1) level was lower in patients in whom AF developed, improved the C-statistic for the prediction of future AF, and was lower in the plasma of the mouse model susceptible to AF. This study demonstrates that plasma lipids have the potential to improve the detection and prediction of AF in patients with diabetes.

Changes in plasma lipids predict pravastatin efficacy in secondary prevention.

BACKGROUNDStatins have pleiotropic effects on lipid metabolism. The relationship between these effects and future cardiovascular events is unknown. We characterized the changes in lipids upon pravastatin treatment and defined the relationship with risk reduction for future cardiovascular events.METHODSPlasma lipids (n = 342) were measured in baseline and 1-year follow-up samples from a Long-Term Intervention with Pravastatin in Ischaemic Disease (LIPID) study subcohort (n = 4991). The associations of changes in lipids with treatment and cardiovascular outcomes were investigated using linear and Cox regression. The effect of treatment on future cardiovascular outcomes was examined by the relative risk reduction (RRR).RESULTSPravastatin treatment was associated with changes in 206 lipids. Species containing arachidonic acid were positively associated while phosphatidylinositol species were negatively associated with pravastatin treatment. The RRR from pravastatin treatment for cardiovascular events decreased from 23.5% to 16.6% after adjustment for clinical risk factors and change in LDL-cholesterol (LDL-C) and to 3.0% after further adjustment for the change in the lipid ratio PI(36:2)/PC(38:4). Change in PI(36:2)/PC(38:4) mediated 58% of the treatment effect. Stratification of patients into quartiles of change in PI(36:2)/PC(38:4) indicated no benefit of pravastatin in the fourth quartile.CONCLUSIONThe change in PI(36:2)/PC(38:4) predicted benefit from pravastatin, independent of change in LDL-C, demonstrating its potential as a biomarker for monitoring the clinical benefit of statin treatment in secondary prevention.TRIAL REGISTRATIONAustralian New Zealand Clinical Trials Registry identifier ACTRN12616000535471.FUNDINGBristol-Myers Squibb; NHMRC grants 211086, 358395, and 1029754; NHMRC program grant 1149987; NHMRC fellowship 108026; and the Operational Infrastructure Support Program of the Victorian government of Australia.

Large-scale plasma lipidomic profiling identifies lipids that predict cardiovascular events in secondary prevention.

BACKGROUND: Plasma lipidomic measures may enable improved prediction of cardiovascular outcomes in secondary prevention. The aim of this study is to determine the association of plasma lipidomic measurements with cardiovascular events and assess their potential to predict such events. METHODS: Plasma lipids (n = 342) were measured in a retrospective subcohort (n = 5,991) of the LIPID study. Proportional hazards regression was used to identify lipids associated with future cardiovascular events (nonfatal myocardial infarction, nonfatal stroke, or cardiovascular death) and cardiovascular death. Multivariable models adding lipid species to traditional risk factors were created using lipid ranking established from the Akaike information criterion within a 5-fold cross-validation framework. The results were tested on a diabetic case cohort from the ADVANCE study (n = 3,779). RESULTS: Specific ceramide species, sphingolipids, phospholipids, and neutral lipids containing omega-6 fatty acids or odd-chain fatty acids were associated with future cardiovascular events (106 species) and cardiovascular death (139 species). The addition of 7 lipid species to a base model (11 conventional risk factors) resulted in an increase in the C-statistics from 0.629 (95% CI, 0.628-0.630) to 0.654 (95% CI, 0.653-0.656) for prediction of cardiovascular events and from 0.673 (95% CI, 0.671-0.675) to 0.727 (95% CI, 0.725-0.728) for prediction of cardiovascular death. Categorical net reclassification improvements for cardiovascular events and cardiovascular death were 0.083 (95% CI, 0.081-0.086) and 0.166 (95% CI, 0.162-0.170), respectively. Evaluation on the ADVANCE case cohort demonstrated significant improvement on the base models. CONCLUSIONS: The improvement in the prediction of cardiovascular outcomes, above conventional risk factors, demonstrates the potential of plasma lipidomic profiles as biomarkers for cardiovascular risk stratification in secondary prevention. FUNDING: Bristol-Myers Squibb, the National Health and Medical Research Council of Australia (grants 211086, 358395, and 1029754), and the Operational Infrastructure Support Program of the Victorian government of Australia.

Weight Loss and Exercise Alter the High-Density Lipoprotein Lipidome and Improve High-Density Lipoprotein Functionality in Metabolic Syndrome.

OBJECTIVE: High-density lipoprotein (HDL) lipid composition and function may better reflect cardiovascular risk than HDL cholesterol concentration. This study characterized the relationships between HDL composition, metabolism, and function in metabolic syndrome (MetS) patients and how changes in composition after weight loss (WL) and exercise treatments are related to function. APPROACH AND RESULTS: Plasma samples from MetS patients (n=95) and healthy individuals (n=40) were used in this study. Subsets of the MetS group underwent 12 weeks of no treatment (n=17), WL (n=19), or WL plus exercise (WLEX; n=17). HDL was isolated using density-gradient ultracentrifugation. The HDL lipidome was analyzed by mass spectrometry, and particle size determined by nuclear magnetic resonance. Cholesteryl ester transfer protein activity and ex vivo HDL cholesterol efflux capacity (CEC) were assessed. The HDL lipidome in the MetS patients was substantially different from that in healthy individuals, mean particle size was smaller, and CEC was lower. Several HDL phospholipid and sphingolipid species were associated with HDL diameter and CEC. The HDL lipidome and particle size were modified toward the healthy individuals after WL and WLEX treatments, with greater effects observed in the latter group. Cholesteryl ester transfer protein activity was reduced after WL and WLEX, and CEC was improved after WLEX. CONCLUSIONS: WLEX treatment in MetS patients normalizes the HDL lipidome and particle size profile and enhances CEC. HDL lipids associated with diminished CEC may represent novel biomarkers for early prediction of HDL dysfunction and disease risk and may represent potential therapeutic targets for future HDL therapies. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique identifier: NCT00163943.

Plasma Lipidomic Profiles Improve on Traditional Risk Factors for the Prediction of Cardiovascular Events in Type 2 Diabetes Mellitus.

BACKGROUND: Clinical lipid measurements do not show the full complexity of the altered lipid metabolism associated with diabetes mellitus or cardiovascular disease. Lipidomics enables the assessment of hundreds of lipid species as potential markers for disease risk. METHODS: Plasma lipid species (310) were measured by a targeted lipidomic analysis with liquid chromatography electrospray ionization-tandem mass spectrometry on a case-cohort (n=3779) subset from the ADVANCE trial (Action in Diabetes and Vascular Disease: Preterax and Diamicron-MR Controlled Evaluation). The case-cohort was 61% male with a mean age of 67 years. All participants had type 2 diabetes mellitus with ≥1 additional cardiovascular risk factors, and 35% had a history of macrovascular disease. Weighted Cox regression was used to identify lipid species associated with future cardiovascular events (nonfatal myocardial infarction, nonfatal stroke, and cardiovascular death) and cardiovascular death during a 5-year follow-up period. Multivariable models combining traditional risk factors with lipid species were optimized with the Akaike information criteria. C statistics and NRIs were calculated within a 5-fold cross-validation framework. RESULTS: Sphingolipids, phospholipids (including lyso- and ether- species), cholesteryl esters, and glycerolipids were associated with future cardiovascular events and cardiovascular death. The addition of 7 lipid species to a base model (14 traditional risk factors and medications) to predict cardiovascular events increased the C statistic from 0.680 (95% confidence interval [CI], 0.678-0.682) to 0.700 (95% CI, 0.698-0.702; P<0.0001) with a corresponding continuous NRI of 0.227 (95% CI, 0.219-0.235). The prediction of cardiovascular death was improved with the incorporation of 4 lipid species into the base model, showing an increase in the C statistic from 0.740 (95% CI, 0.738-0.742) to 0.760 (95% CI, 0.757-0.762; P<0.0001) and a continuous net reclassification index of 0.328 (95% CI, 0.317-0.339). The results were validated in a subcohort with type 2 diabetes mellitus (n=511) from the LIPID trial (Long-Term Intervention With Pravastatin in Ischemic Disease). CONCLUSIONS: The improvement in the prediction of cardiovascular events, above traditional risk factors, demonstrates the potential of plasma lipid species as biomarkers for cardiovascular risk stratification in diabetes mellitus. CLINICAL TRIAL REGISTRATION: URL: https://clinicaltrials.gov. Unique identifier: NCT00145925.

An Efficient Single Phase Method for the Extraction of Plasma Lipids.

Lipidomic approaches are now widely used to investigate the relationship between lipid metabolism, health and disease. Large-scale lipidomics studies typically aim to quantify hundreds to thousands of lipid molecular species in a large number of samples. Consequently, high throughput methodology that can efficiently extract a wide range of lipids from biological samples is required. Current methods often rely on extraction in chloroform:methanol with or without two phase partitioning or other solvents, which are often incompatible with liquid chromatography electrospray ionization-tandem mass spectrometry (LC ESI-MS/MS). Here, we present a fast, simple extraction method that is suitable for high throughput LC ESI-MS/MS. Plasma (10 µL) was mixed with 100 µL 1-butanol:methanol (1:1 v/v) containing internal standards resulting in efficient extraction of all major lipid classes (including sterols, glycerolipids, glycerophospholipids and sphingolipids). Lipids were quantified using positive-ion mode LC ESI-MS/MS. The method showed high recovery (>90%) and reproducibility (%CV < 20%). It showed a strong correlation of all lipid measures with an established chloroform:methanol extraction method (R2 = 0.976). This method uses non-halogenated solvents, requires no drying or reconstitution steps and is suitable for large-scale LC ESI-MS/MS-based lipidomic analyses in research and clinical laboratories.

Factors affecting variability in distribution of tacrolimus in liver transplant recipients.

AIMS: Therapeutic drug monitoring (TDM) of tacrolimus is complicated by conflicting data on the correlation between tacrolimus trough blood concentrations and the incidence of rejection. The aim of this cross-sectional study was to investigate the blood distribution and protein binding of tacrolimus in liver transplant recipients to explore better predictors of clinical outcome. METHODS: Blood and plasma distribution of 3H-dihydro-tacrolimus was investigated in 40 liver transplant recipients using Ficoll Paque and density gradient ultracentrifugation, respectively, and equilibrium dialysis to investigate plasma protein binding. RESULTS: In blood tacrolimus was mainly associated with the erythrocyte fraction (83.2%, range 74.6-94.9%), followed by diluted plasma (16.1%, range 4.5-24.9%), and lymphocyte fraction (0.61%, range: 0.11-1.53%). In plasma, lipoprotein deficient serum fraction (54.2%, range 38.5-68.2%) was the main reservoir of tacrolimus. The unbound fraction of tacrolimus was found to be 0.47 +/- 0.18% (range 0.07-0.89%). The percentage of tacrolimus associated with the lymphocytes (0.8 +/- 0.4 vs 0.3 +/- 0.1%, P = 0.012) and estimated unbound concentration (0.42 +/- 0.21 ng l-1vs 0.24 +/- 0.08 ng l-1, P < 0.001) of tacrolimus were significantly different in stable transplant recipients and those experiencing rejection. Haematocrit and red blood cell count significantly influenced the percentage of tacrolimus associated with erythrocytes. The fraction unbound of tacrolimus was correlated with alpha1-acid glycoprotein and high density lipoprotein cholesterol concentrations. CONCLUSIONS: Tacrolimus unbound concentration was observed to be lower in liver transplant recipients experiencing rejection and further study is required to evaluate its utility in the TDM of tacrolimus.

Coral bleaching and mortality on artificial and natural reefs in Maldives in 1998, sea surface temperature anomalies and initial recovery.

The bleaching and subsequent mortality of branching and massive corals on artificial and natural reefs in the central atolls of Maldives in 1998 are examined with respect to sea surface temperature (SST) anomalies. SST normally peaks in April-May in Maldives. The UK Meteorological Office's Global sea-Ice and SST data set version 2.3 b shows that in 1998 monthly mean SST was 1.2-4 S.D. above the 1950-1999 average during the warmest months (March-June), with the greatest anomaly in May of +2.1 degrees C. Bleaching was first reported in mid-April and was severe from late April to mid-May with some recovery evident by late-May. At least 98% of branching corals (Acroporidae, Pocilloporidae) on artificial structures deployed on a reef flat in 1990 died whereas the majority of massive corals (Poritidae, Faviidae, Agariciidae) survived the bleaching. The pre-bleaching coral community on the artificial reefs in 1994 was 95% branching corals and 5% massives (n = 1589); the post-bleaching community was 3% branching corals and 97% massives (n = 248). Significant reductions in live coral cover were seen at all natural reefs surveyed in the central atolls, with average live coral cover decreasing from about 42% to 2%, a 20-fold reduction from pre-bleaching levels. A survey of recruitment of juvenile corals to the artificial structures 10 months after the bleaching event showed that 67% of recruits (> or = 0.5 cm diameter) were acroporids and pocilloporids and 33% were from massive families (n = 202) compared to 94% and 6%, respectively, in 1990-1994 (n = 3136). Similar post-bleaching dominance of recruitment by branching corals was seen on nearby natural reef (78% acroporids and pocilloporids; 22% massives). A linear regression of April mean monthly SST against year was highly significant (p < 0.001) and suggests a rise of 0.16 degree C per decade. If this trend continues, by 2030 mean April SST in the central atolls will normally exceed the anomaly level at which corals appear there are susceptible to mass bleaching.

Validation of methods to study the distribution and protein binding of tacrolimus in human blood.

INTRODUCTION: Tacrolimus is a macrolide immunosuppressant that has a narrow therapeutic index, displays considerable variability in response, and has the potential for serious drug interactions. Therapeutic drug monitoring and dose individualisation for tacrolimus is complicated but essential. Few studies have investigated the blood distribution and protein binding of tacrolimus and the results of these studies are conflicting. The aim of the present study is to establish and validate methods to investigate the distribution of tacrolimus in human blood. To conduct these studies at clinically relevant concentrations the use of 3H-dihydro-tacrolimus instead of tacrolimus was investigated. METHODS: The use of radiolabelled tacrolimus was validated by conducting studies with a mixture of both labelled and unlabelled drug where tacrolimus was analysed by LC-MS/MS. The in vitro distribution of tacrolimus and 3H-dihydro-tacrolimus was investigated in blood collected from healthy subjects using Ficoll-Paque reagent and density gradient ultracentrifugation, respectively. The unbound fraction of tacrolimus in plasma was studied using equilibrium dialysis conducted at 37 degrees C. RESULTS: In blood, tacrolimus was found to be mainly associated with erythrocytes (85.3+/-1.5%), followed by diluted plasma proteins (14.3+/-1.5%) and lymphocytes (0.46+/-0.10%). In plasma, tacrolimus was found to mainly be associated with the soluble protein fraction (61.2+/-2.5%), high-density lipoproteins (HDL, 28.1+/-5.4%), low-density lipoproteins (LDL, 7.8+/-1.6%), and very low-density lipoproteins (VLDL, 1.4+/-0.3%). The unbound fraction of tacrolimus was found to be only 1.2+/-0.12%. Statistical comparison indicated that there was no significant difference in the blood distribution and plasma protein binding of 3H-dihydro-tacrolimus when compared with tacrolimus. DISCUSSION: These results have important implications for therapeutic drug monitoring of tacrolimus and subsequent studies of tacrolimus distribution in transplant recipients.

Effects of subinhibitory concentrations of nitroxoline on the surface properties of Escherichia coli.

Nitroxoline (5-nitro-8-quinolinol; NIQ) at subinhibitory concentrations (sub-MIC) decreased the adherence of uropathogenic Escherichia coli to catheter surface and significantly enhanced cell surface hydrophobicity. The surface hydrophobicity increased in the presence of sub-MIC of NIQ and also in an excess of Mg2+. The effect of NIQ on the cell surface was not related to the bacteriostatic effect of this agent. The increase in nitrogen and decrease in phosphate content in the cell surface was found in the presence of NIQ. NIQ did not inhibit the expression of fimbriae.

Extractive separation of trivalent lanthanide metals with a combination of Di(2-ethylhexyl)phosphoric acid and 1,10-phenanthroline.

The equilibrium extraction behavior of a series of trivalent lanthanide ions (Ln(3+)) using a chloroform-Kerosine solution containing Di(2-ethylhexyl)phosphoric acid, combined with an adductant, 1,10-phenanthroline monohydrate (phen), was studied. The enhancement of the extraction by addition of such a neutral adductant is explained in terms of the extraction of the quaternary complex, M(HX(2))(3)(phen)(2), in addition to the neutral complex, M(HX(2))(3), into the organic phase. The stoichiometry, extraction constants and separation factors of these systems were determined. The extraction constants of these systems partially follow the order of the atomic numbers. The synergistic extraction constants increased in the other Gd > Er > Ho > Eu > Ce > La > Pr and the highest separation factor was observed for ErHo (2.09). pH (1 2 ) values were also obtained. In this synergistic extraction system, both the extraction equilibrium constants and the separation factors were found to be greater than those of commercial extractants.