ELOVL6 Genetic Variation Is Related to Insulin Sensitivity: A New Candidate Gene in Energy Metabolism

BACKGROUND: The elongase of long chain fatty acids family 6 (ELOVL6) is an enzyme that specifically catalyzes the elongation of saturated and monounsaturated fatty acids with 12, 14 and 16 carbons. ELOVL6 is expressed in lipogenic tissues and it is regulated by sterol regulatory element binding protein 1 (SREBP-1). OBJECTIVE: We investigated whether ELOVL6 genetic variation is associated with insulin sensitivity in a population from southern Spain. DESIGN: We undertook a prospective, population-based study collecting phenotypic, metabolic, nutritional and genetic information. Measurements were made of weight and height and the body mass index (BMI) was calculated. Insulin resistance was measured by homeostasis model assessment. The type of dietary fat was assessed from samples of cooking oil taken from the participants' kitchens and analyzed by gas chromatography. Five SNPs of the ELOVL6 gene were analyzed by SNPlex. RESULTS: Carriers of the minor alleles of the SNPs rs9997926 and rs6824447 had a lower risk of having high HOMA_IR, whereas carriers of the minor allele rs17041272 had a higher risk of being insulin resistant. An interaction was detected between the rs6824447 polymorphism and the intake of oil in relation with insulin resistance, such that carriers of this minor allele who consumed sunflower oil had lower HOMA_IR than those who did not have this allele (P = 0.001). CONCLUSIONS: Genetic variations in the ELOVL6 gene were associated with insulin sensitivity in this population-based study.

Mutant prevention concentrations of fluoroquinolones for Enterobacteriaceae expressing the plasmid-carried quinolone resistance determinant qnrA1.

The influence of qnrA1 on the development of quinolone resistance in Enterobacteriaceae was evaluated by using the mutant prevention concentration parameter. The expression of qnrA1 considerably increased the mutant prevention concentration compared to strains without this gene. In the presence of qnrA1, mutations in gyrA and parC genes were easily selected to produce high levels of quinolone resistance.

In vitro activity of fosfomycin against extended-spectrum-beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae: comparison of susceptibility testing procedures

The agar dilution, broth microdilution, and disk diffusion methods were compared to determine the in vitro susceptibility of 428 extended-spectrum-beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae to fosfomycin. Fosfomycin showed very high activity against all ESBL-producing strains. Excellent agreement between the three susceptibility methods was found for E. coli, whereas marked discrepancies were observed for K. pneumoniae.

Surgical menopause increases salt sensitivity of blood pressure

Salt sensitivity of blood pressure is associated with an elevated risk of developing hypertension (HTN) and is an independent risk factor for cardiovascular disease. The prevalence of HTN increases after menopause. The aim of this study was to investigate prospectively whether the loss of ovarian hormones increases the occurrence of salt sensitivity among healthy premenopausal women. We enrolled 40 normotensive, nondiabetic women (age 47.2+/-3.5), undergoing hysterectomy-oophorectomy for nonneoplastic processes and not on hormone replacement, to determine the effect of changes in sodium intake on blood pressure the day before and subsequently 4 months after surgical menopause. Salt loading was achieved using a 2-L normal saline infusion and salt depletion produced by 40 mg of intravenous furosemide. A decrease >10 mm Hg in systolic blood pressure between salt loading and salt depletion was used to define salt sensitivity. Before and after menopause, salt-sensitive women exhibited higher waist/hip and waist/thigh ratios (P<0.01). Although all of the women remained normotensive, the prevalence of salt sensitivity was significantly higher after surgical menopause (21 women; 52.5%) than before (9 women; 22.5%; P=0.01), because 12 (38.7%) salt-resistant women developed salt sensitivity after menopause. In summary, we demonstrated that the prevalence of salt sensitivity doubled as early as 4 months after surgical menopause, without an associated increase in blood pressure. Epidemiological studies indicate that development of HTN may not occur until 5 to 10 years after menopause. The loss of ovarian hormones may unmask a population of women prone to salt sensitivity who, with aging, would be at higher risk for the subsequent development of HTN and cardiovascular disease.

Loss of ovarian hormones increases salt-sensitivity in normotensive women

Salt sensitivity (SS) is associated with an elevated risk of developing hypertension(HTN) and is an independent risk factor for cardiovascular (CV) morbidity and mortality. Cross-sectional studies have suggested that postmenopausal women are more salt sensitive than premenopausal women. The purpose of the present study was to investigate prospectively the prevalence of SS among healthy premenopausal women and determine whether the loss of ovarian hormones increases SS.

Impact of dietary betaine and conjugated linoleic acid on insulin sensitivity, protein and fat metabolism of obese pigs.

To determine possible mechanisms of action that might explain the nutrient partitioning effect of betaine and conjugated linoleic acid (CLA) in Iberian pigs and to address potential adverse effects, twenty gilts were restrictively fed from 20 to 50 kg BW Control, 0.5% betaine, 1% CLA or 0.5% betaine + 1% CLA diets. Serum hormones and metabolites profile were determined at 30 kg BW and an oral glucose test was performed before slaughter. Pigs were slaughtered at 50 kg BW and livers were obtained for chemical and histological analysis. Decreased serum urea in pigs fed betaine and betaine + CLA diets (11%; P = 0.0001) indicated a more efficient N utilization. The increase in serum triacylglycerol (58% and 28%, respectively; P = 0.0098) indicated that CLA and betaine + CLA could have reduced adipose tissue triacylglycerol synthesis from preformed fatty acids. Serum glucose, low-density lipoprotein (LDL) cholesterol and non-esterified fatty acids were unaffected. CLA and betaine + CLA altered serum lipids profile, although liver of pigs fed CLA diet presented no histopathological changes and triglyceride content was not different from Control pigs. Compared with controls, serum growth hormone decreased (20% to 23%; P = 0.0209) for all treatments. Although serum insulin increased in CLA, and especially in betaine + CLA pigs (28% and 83%; P = 0.0001), indices of insulin resistance were unaffected. In conclusion, CLA, and especially betaine + CLA, induced changes in biochemical parameters and hormones that may partially explain a nutrient partitioning effect in young pigs. Nevertheless, they exhibited weak, although detrimental, effects on blood lipids. Moreover, although livers were chemically and histologically normal, pigs fed CLA diet challenged with a glucose load had higher serum glucose than controls.

Trends in yearly prevalence of third-generation cephalosporin and fluoroquinolone resistant Enterobacteriaceae infections and antimicrobial use in Spanish hospitals, Spain, 1999 to 2010.

Escherichia coli, Klebsiella pneumoniae, and Enterobacter spp. are a major cause of infections in hospitalised patients. The aim of our study was to evaluate rates and trends of resistance to third-generation cephalosporins and fluoroquinolones in infected patients, the trends in use for these antimicrobials, and to assess the potential correlation between both trends. The database of national point prevalence study series of infections and antimicrobial use among patients hospitalised in Spain over the period from 1999 to 2010 was analysed. On average 265 hospitals and 60,000 patients were surveyed per year yielding a total of 19,801 E. coli, 3,004 K. pneumoniae and 3,205 Enterobacter isolates. During the twelve years period, we observed significant increases for the use of fluoroquinolones (5.8%-10.2%, p<0.001), but not for third-generation cephalosporins (6.4%-5.9%, p=NS). Resistance to third-generation cephalosporins increased significantly for E. coli (5%-15%, p<0.01) and for K. pneumoniae infections (4%-21%, p<0.01) but not for Enterobacter spp. (24%). Resistance to fluoroquinolones increased significantly for E. coli (16%30%, p<0.01), for K. pneumoniae (5%-22%, p<0.01), and for Enterobacter spp. (6%-15%, p<0.01). We found strong correlations between the rate of fluoroquinolone use and the resistance to fluoroquinolones, third-generation cephalosporins, or co-resistance to both, for E. coli (R=0.97, p<0.01, R=0.94, p<0.01, and R=0.96, p<0.01, respectively), and for K. pneumoniae (R=0.92, p<0.01, R=0.91, p<0.01, and R=0.92, p<0.01, respectively). No correlation could be found between the use of third-generation cephalosporins and resistance to any of the latter antimicrobials. No significant correlations could be found for Enterobacter spp.. Knowledge of the trends in antimicrobial resistance and use of antimicrobials in the hospitalised population at the national level can help to develop prevention strategies.

Virulence profiles of bacteremic extended-spectrum β-lactamase-producing Escherichia coli: association with epidemiological and clinical features.

There is scarce data about the importance of phylogroups and virulence factors (VF) in bloodstream infections (BSI) caused by extended-spectrum β-lactamase-producing Escherichia coli (ESBLEC). A prospective multicenter Spanish cohort including 191 cases of BSI due to ESBLEC was studied. Phylogroups and 25 VF genes were investigated by PCR. ESBLEC were classified into clusters according to their virulence profiles. The association of phylogropus, VF, and clusters with epidemiological features were studied using multivariate analysis. Overall, 57.6%, 26.7%, and 15.7% of isolates belonged to A/B1, D and B2 phylogroups, respectively. By multivariate analysis (adjusted OR [95% CI]), virulence cluster C2 was independently associated with urinary tract source (5.05 [0.96-25.48]); cluster C4 with sources other than urinary of biliary tract (2.89 [1.05-7.93]), and cluster C5 with BSI in non-predisposed patients (2.80 [0.99-7.93]). Isolates producing CTX-M-9 group ESBLs and from phylogroup D predominated among cluster C2 and C5, while CTX-M-1 group of ESBL and phylogroup B2 predominantes among C4 isolates. These results suggest that host factors and previous antimicrobial use were more important than phylogroup or specific VF in the occurrence of BSI due to ESBLEC. However, some associations between virulence clusters and some specific epidemiological features were found.

Inoculum effect on the efficacies of amoxicillin-clavulanate, piperacillin-tazobactam, and imipenem against extended-spectrum β-lactamase (ESBL)-producing and non-ESBL-producing Escherichia coli in an experimental murine sepsis model.

Escherichia coli is commonly involved in infections with a heavy bacterial burden. Piperacillin-tazobactam and carbapenems are among the recommended empirical treatments for health care-associated complicated intra-abdominal infections. In contrast to amoxicillin-clavulanate, both have reduced in vitro activity in the presence of high concentrations of extended-spectrum β-lactamase (ESBL)-producing and non-ESBL-producing E. coli bacteria. Our goal was to compare the efficacy of these antimicrobials against different concentrations of two clinical E. coli strains, one an ESBL-producer and the other a non-ESBL-producer, in a murine sepsis model. An experimental sepsis model {~5.5 log10 CFU/g [low inoculum concentration (LI)] or ~7.5 log(10) CFU/g [high inoculum concentration (HI)]} using E. coli strains ATCC 25922 (non-ESBL producer) and Ec1062 (CTX-M-14 producer), which are susceptible to the three antimicrobials, was used. Amoxicillin-clavulanate (50/12.5 mg/kg given intramuscularly [i.m.]), piperacillin-tazobactam (25/3.125 mg/kg given intraperitoneally [i.p.]), and imipenem (30 mg/kg i.m.) were used. Piperacillin-tazobactam and imipenem reduced spleen ATCC 25922 strain concentrations (-2.53 and -2.14 log10 CFU/g [P < 0.05, respectively]) in the HI versus LI groups, while amoxicillin-clavulanate maintained its efficacy (-1.01 log10 CFU/g [no statistically significant difference]). Regarding the Ec1062 strain, the antimicrobials showed lower efficacy in the HI than in the LI groups: -0.73, -1.89, and -1.62 log10 CFU/g (P < 0.05, for piperacillin-tazobactam, imipenem, and amoxicillin-clavulanate, respectively, although imipenem and amoxicillin-clavulanate were more efficacious than piperacillin-tazobactam). An adapted imipenem treatment (based on the time for which the serum drug concentration remained above the MIC obtained with a HI of the ATCC 25922 strain) improved its efficacy to -1.67 log10 CFU/g (P < 0.05). These results suggest that amoxicillin-clavulanate could be an alternative to imipenem treatment of infections caused by ESBL- and non-ESBL-producing E. coli strains in patients with therapeutic failure with piperacillin-tazobactam.

Contribution of efflux pumps, porins, and β-lactamases to multidrug resistance in clinical isolates of Acinetobacter baumannii.

We investigated the mechanisms of resistance to carbapenems, aminoglycosides, glycylcyclines, tetracyclines, and quinolones in 90 multiresistant clinical strains of Acinetobacter baumannii isolated from two genetically unrelated A. baumannii clones: clone PFGE-ROC-1 (53 strains producing the OXA-58 β-lactamase enzyme and 18 strains with the OXA-24 β-lactamase) and clone PFGE-HUI-1 (19 strains susceptible to carbapenems). We used real-time reverse transcriptase PCR to correlate antimicrobial resistance (MICs) with expression of genes encoding chromosomal β-lactamases (AmpC and OXA-51), porins (OmpA, CarO, Omp33, Dcap-like, OprB, Omp25, OprC, OprD, and OmpW), and proteins integral to six efflux systems (AdeABC, AdeIJK, AdeFGH, CraA, AbeM, and AmvA). Overexpression of the AdeABC system (level of expression relative to that by A. baumannii ATCC 17978, 30- to 45-fold) was significantly associated with resistance to tigecycline, minocycline, and gentamicin and other biological functions. However, hyperexpression of the AdeIJK efflux pump (level of expression relative to that by A. baumannii ATCC 17978, 8- to 10-fold) was significantly associated only with resistance to tigecycline and minocycline (to which the TetB efflux system also contributed). TetB and TetA(39) efflux pumps were detected in clinical strains and were associated with resistance to tetracyclines and doxycycline. The absence of the AdeABC system and the lack of expression of other mechanisms suggest that tigecycline-resistant strains of the PFGE-HUI-1 clone may be associated with a novel resistance-nodulation-cell efflux pump (decreased MICs in the presence of the inhibitor Phe-Arg β-naphthylamide dihydrochloride) and the TetA(39) system.

Prospective multicenter study of the impact of carbapenem resistance on mortality in Pseudomonas aeruginosa bloodstream infections.

The impact of antimicrobial resistance on clinical outcomes is the subject of ongoing investigations, although uncertainty remains about its contribution to mortality. We investigated the impact of carbapenem resistance on mortality in Pseudomonas aeruginosa bacteremia in a prospective multicenter (10 teaching hospitals) observational study of patients with monomicrobial bacteremia followed up for 30 days after the onset of bacteremia. The adjusted influence of carbapenem resistance on mortality was studied by using Cox regression analysis. Of 632 episodes, 487 (77%) were caused by carbapenem-susceptible P. aeruginosa (CSPA) isolates, and 145 (23%) were caused by carbapenem-resistant P. aeruginosa (CRPA) isolates. The median incidence density of nosocomial CRPA bacteremia was 2.3 episodes per 100,000 patient-days (95% confidence interval [CI], 1.9 to 2.8). The regression demonstrated a time-dependent effect of carbapenem resistance on mortality as well as a significant interaction with the Charlson index: the deleterious effect of carbapenem resistance on mortality decreased with higher Charlson index scores. The impact of resistance on mortality was statistically significant only from the fifth day after the onset of the bacteremia, reaching its peak values at day 30 (adjusted hazard ratio for a Charlson score of 0 at day 30, 9.9 [95% CI, 3.3 to 29.4]; adjusted hazard ratio for a Charlson score of 5 at day 30, 2.6 [95% CI, 0.8 to 8]). This study clarifies the relationship between carbapenem resistance and mortality in patients with P. aeruginosa bacteremia. Although resistance was associated with a higher risk of mortality, the study suggested that this deleterious effect may not be as great during the first days of the bacteremia or in the presence of comorbidities.

Impact of inadequate empirical therapy on the mortality of patients with bloodstream infections: a propensity score-based analysis.

The impact of the adequacy of empirical therapy on outcome for patients with bloodstream infections (BSI) is key for determining whether adequate empirical coverage should be prioritized over other, more conservative approaches. Recent systematic reviews outlined the need for new studies in the field, using improved methodologies. We assessed the impact of inadequate empirical treatment on the mortality of patients with BSI in the present-day context, incorporating recent methodological recommendations. A prospective multicenter cohort including all BSI episodes in adult patients was performed in 15 hospitals in Andalucía, Spain, over a 2-month period in 2006 to 2007. The main outcome variables were 14- and 30-day mortality. Adjusted analyses were performed by multivariate analysis and propensity score-based matching. Eight hundred one episodes were included. Inadequate empirical therapy was administered in 199 (24.8%) episodes; mortality at days 14 and 30 was 18.55% and 22.6%, respectively. After controlling for age, Charlson index, Pitt score, neutropenia, source, etiology, and presentation with severe sepsis or shock, inadequate empirical treatment was associated with increased mortality at days 14 and 30 (odds ratios [ORs], 2.12 and 1.56; 95% confidence intervals [95% CI], 1.34 to 3.34 and 1.01 to 2.40, respectively). The adjusted ORs after a propensity score-based matched analysis were 3.03 and 1.70 (95% CI, 1.60 to 5.74 and 0.98 to 2.98, respectively). In conclusion, inadequate empirical therapy is independently associated with increased mortality in patients with BSI. Programs to improve the quality of empirical therapy in patients with suspicion of BSI and optimization of definitive therapy should be implemented.

UV-fixed-thick-blotch preparation improves sensitivity of auramine staining.

We describe here a very simple modification of the auramine staining procedure based on preparation of a UV-fixed thick blotch which allowed us to reach an overall sensitivity of 0.82 (592 acid-fast bacillus [AFB]-positive specimens/722 initial respiratory specimens with positive mycobacterial culture) and sensitivities of 0.93 (526 AFB-positive specimens/564 culture-positive specimens) for Mycobacterium tuberculosis complex and 0.42 (66 AFB-positive specimens/158 culture-positive specimens) for nontuberculous mycobacteria.

Analysis of genes encoding penicillin-binding proteins in clinical isolates of Acinetobacter baumannii.

There is limited information on the role of penicillin-binding proteins (PBPs) in the resistance of Acinetobacter baumannii to β-lactams. This study presents an analysis of the allelic variations of PBP genes in A. baumannii isolates. Twenty-six A. baumannii clinical isolates (susceptible or resistant to carbapenems) from three teaching hospitals in Spain were included. The antimicrobial susceptibility profile, clonal pattern, and genomic species identification were also evaluated. Based on the six complete genomes of A. baumannii, the PBP genes were identified, and primers were designed for each gene. The nucleotide sequences of the genes identified that encode PBPs and the corresponding amino acid sequences were compared with those of ATCC 17978. Seven PBP genes and one monofunctional transglycosylase (MGT) gene were identified in the six genomes, encoding (i) four high-molecular-mass proteins (two of class A, PBP1a [ponA] and PBP1b [mrcB], and two of class B, PBP2 [pbpA or mrdA] and PBP3 [ftsI]), (ii) three low-molecular-mass proteins (two of type 5, PBP5/6 [dacC] and PBP6b [dacD], and one of type 7 (PBP7/8 [pbpG]), and (iii) a monofunctional enzyme (MtgA [mtgA]). Hot spot mutation regions were observed, although most of the allelic changes found translated into silent mutations. The amino acid consensus sequences corresponding to the PBP genes in the genomes and the clinical isolates were highly conserved. The changes found in amino acid sequences were associated with concrete clonal patterns but were not directly related to susceptibility or resistance to β-lactams. An insertion sequence disrupting the gene encoding PBP6b was identified in an endemic carbapenem-resistant clone in one of the participant hospitals.

Klebsiella pneumoniae strains producing extended-spectrum beta-lactamases in Spain: microbiological and clinical features.

Extended-spectrum β-lactamases (ESBL) of the CTX-M, SHV, and TEM families were recognized in 76 (67%), 31 (27%), and 6 (5%) isolates, respectively, among 162 ESBL-producing Klebsiella pneumoniae (ESBL-Kp) strains obtained in a multicenter study in Spain. Predisposing factors for ESBL-Kp acquisition included invasive procedures, mechanical ventilation, and previous antimicrobial use.