Detection of Mycobacterium tuberculosis (TB) in vitro and in situ using an electronic nose in combination with a neural network system

The use of volatile production patterns produced by Mycobacterium tuberculosis and associated bacterial infections from sputum samples were examined in vitro and in situ using an electronic nose based on a 14 sensor conducting polymer array. In vitro, it was possible to successfully discriminate between M. tuberculosis (TB) and control media, and between M. tuberculosis and M. avium, M. scrofulaceum and Pseudomonas aeruginosa cultures in the stationary phase after 5-6h incubation at 37 degrees C based on 35 samples. Using neural network (NN) analysis and cross-validation it was possible to successfully identify 100% of the TB cultures from others. A second in vitro study with 61 samples all four groups were successfully discriminated with 14 of 15 unknowns within each of the four groups successfully identified using cross-validation and discriminant function analysis. Subsequently, lipase enzymes were added to 46 sputum samples directly obtained from patients and the head space analysed. Parallel measurements of bacterial contamination were also carried out for confirmation using agar media. NN analysis was carried out using some of the samples as a training set. Based on the NN and genetic algorithms of up to 10 generations it was possible to successfully cross-validate 9 of 10 unknown samples. PCA was able to discriminate between TB infection alone, the controls, M. avium, P. aeruginosa and a mixed infection. These findings will have significant implications for the development of rapid qualitative systems for screening of patient samples and clinical diagnosis of tuberculosis.     

Molecular probes for the cannabinoid receptors

Cannabinoids produce most of their biochemical and pharmacological effects by interacting with CB1 and CB2 cannabinoid receptors, both of which are G-protein coupled membrane-bound functional proteins. CB1 is found in the central nervous system and in a variety of other organs including heart, vascular endothelium, uterus, vas deferens, testis and small intestine. Conversely, the CB2 receptor appears to be associated exclusively with the immune system and is found in the periphery of the spleen and other cells associated with immunochemical functions. Although both CB1 and CB2 have been cloned and the primary sequences are known, their three dimensional structures and the amino acid residues at the active site, critical for ligand recognition, binding and activation have not been characterized. In the absence of any X-ray crystallographic and NMR data, information on the structural requirements for ligand-receptor interactions is obtained with the help of suitably designed molecular probes. These ligands either interact with the receptor in a reversible fashion (reversible probes) or, alternatively, attach at or near the receptor active site with the formation of a covalent bond (irreversible probes). Subsequently, information related to ligand binding and receptor activation is further amplified with the help of receptor mutants and computer modeling. This review focuses on molecular probes related to the classical and non-classical cannabinoids that have been reported since the discovery of the first cannabinoid receptor over a decade ago.     

Mutations in KIF11 cause autosomal-dominant microcephaly variably associated with congenital lymphedema and chorioretinopathy

We have identified KIF11 mutations in individuals with syndromic autosomal-dominant microcephaly associated with lymphedema and/or chorioretinopathy. Initial whole-exome sequencing revealed heterozygous KIF11 mutations in three individuals with a combination of microcephaly and lymphedema from a microcephaly-lymphedema-chorioretinal-dysplasia cohort. Subsequent Sanger sequencing of KIF11 in a further 15 unrelated microcephalic probands with lymphedema and/or chorioretinopathy identified additional heterozygous mutations in 12 of them. KIF11 encodes EG5, a homotetramer kinesin motor. The variety of mutations we have found (two nonsense, two splice site, four missense, and six indels causing frameshifts) are all predicted to have an impact on protein function. EG5 has previously been shown to play a role in spindle assembly and function, and these findings highlight the critical role of proteins necessary for spindle formation in CNS development. Moreover, identification of KIF11 mutations in patients with chorioretinopathy and lymphedema suggests that EG5 is involved in the development and maintenance of retinal and lymphatic structures.     

Work stress, obesity and the risk of type 2 diabetes: gender-specific bidirectional effect in the Whitehall II study

Psychosocial work stress has been linked to higher risk of type 2 diabetes (T2DM), with the effect being consistently higher among women than men. Also, work stress has been linked to prospective weight gain among obese men but weight loss among lean men. Here, we aimed to examine the interaction between work stress and obesity in relation to T2DM risk in a gender‐specific manner. We studied 5,568 white middle‐aged men and women in the Whitehall II study, who were free from diabetes at analysis baseline (1993). After 1993, diabetes was ascertained at six consecutive phases by an oral glucose tolerance test supplemented by self‐reports. Cox regression analysis was used to assess the association between job strain (high job demands/low job control) and 18‐year incident T2DM stratifying by BMI (BMI <30 kg/m² vs. BMI ≥30 kg/m²). Overall, work stress was associated with incident T2DM among women (hazard ratio (HR) 1.41: 95% confidence intervals: 1.02; 1.95) but not among men (HR 0.87: 95% confidence interval 0.69; 1.11) (P_INTERACTION = 0.017). Among men, work stress was associated with a lower risk of T2DM in nonobese (HR 0.70: 0.53; 0.93) but not in obese individuals (P_INTERACTION = 0.17). Among women, work stress was associated with higher risk of T2DM in the obese (HR 2.01: 1.06; 3.92) but not in the nonobese (P_INTERACTION = 0.005). Gender and body weight status play a critical role in determining the direction of the association between psychosocial stress and T2DM. The potential effect‐modifying role of gender and obesity should not be ignored by future studies looking at stress‐disease associations.     

Effects of a selective cannabinoid CB2 agonist and antagonist on intravenous nicotine self administration and reinstatement of nicotine seeking

Over the last decade there have been significant advances in the discovery and understanding of the cannabinoid system along with the development of pharmacologic tools that modulate its function. Characterization of the crosstalk between nicotine addiction and the cannabinoid system may have significant implications on our understanding of the neurobiological mechanisms underlying nicotine dependence. Two types of cannabinoid receptors (CB1 and CB2) have been identified. CB1 receptors are expressed in the brain and modulate drug taking and drug seeking for various drugs of abuse, including nicotine. CB2 receptors have been recently identified in the brain and have been proposed to play a functional role in mental disorders and drug addiction. Our objective was to explore the role of CB2 receptors on intravenous nicotine self administration under two schedules of reinforcement (fixed and progressive ratio) and on nicotine seeking induced by nicotine priming or by nicotine associated cues. For this, we evaluated the effects of various doses of the selective CB2 antagonist AM630 (1.25 to 5 mg/kg) and CB2 agonist AM1241 (1 to 10 mg/kg) on these behavioral responses in rats. Different groups of male Long Evans rats were trained to lever press for nicotine at a unit dose of 30 μg/kg/infusion. Subsequently, animals were randomized using a Latin-square design and injected with either AM1241 or AM630 using a counterbalanced within subject design. Administration of the CB2 ligands did not affect either nicotine-taking nicotine-seeking behavior. Our results do not support the involvement of CB2 receptors in nicotine-taking or nicotine-seeking behavior.     

The structure of Mediterranean rocky reef ecosystems across environmental and human gradients, and conservation implications

Historical exploitation of the Mediterranean Sea and the absence of rigorous baselines makes it difficult to evaluate the current health of the marine ecosystems and the efficacy of conservation actions at the ecosystem level. Here we establish the first current baseline and gradient of ecosystem structure of nearshore rocky reefs at the Mediterranean scale. We conducted underwater surveys in 14 marine protected areas and 18 open access sites across the Mediterranean, and across a 31-fold range of fish biomass (from 3.8 to 118 g m(-2)). Our data showed remarkable variation in the structure of rocky reef ecosystems. Multivariate analysis showed three alternative community states: (1) large fish biomass and reefs dominated by non-canopy algae, (2) lower fish biomass but abundant native algal canopies and suspension feeders, and (3) low fish biomass and extensive barrens, with areas covered by turf algae. Our results suggest that the healthiest shallow rocky reef ecosystems in the Mediterranean have both large fish and algal biomass. Protection level and primary production were the only variables significantly correlated to community biomass structure. Fish biomass was significantly larger in well-enforced no-take marine reserves, but there were no significant differences between multi-use marine protected areas (which allow some fishing) and open access areas at the regional scale. The gradients reported here represent a trajectory of degradation that can be used to assess the health of any similar habitat in the Mediterranean, and to evaluate the efficacy of marine protected areas.     

Dose-dependent effects of thyroid hormone on post-ischemic cardiac performance: potential involvement of Akt and ERK signalings

The present study explored the effects of thyroid hormone (TH) treatment on post-ischemic cardiac function and potential implicated mechanisms. Acute myocardial infarction (AMI) was induced in mice by coronary artery ligation while sham-operated animals served as controls. This procedure resulted in a marked depression of cardiac function and significant reduction in TH levels in plasma. TH was given at a dose aiming to normalize T3 levels in plasma [AMI-TH (A)] and also at higher doses. The group of animals treated with the highest dose of TH, which displayed significantly increased mortality rate was included in the study [AMI-TH (B)]. In AMI-TH (A) mice, TH significantly improved left ventricular (LV) ejection fraction (EF%), [27.9% (1.4) in AMI versus 38.0 (3.1) in AMI-TH (A), P < 0.05], and favorably remodeled LV chamber while α-MHC was the dominant isoform expressed. In AMI-TH (B) mice, TH treatment resulted in increased mortality as compared to untreated mice (73% vs 47%, P < 0.05), while the favorable effect of TH was not evident in the survived animals. At the molecular level, TH, at the replacement dose, modestly increased p-Akt levels in the myocardium without any change in p-ERK levels. On the contrary, TH at the higher dose resulted in further increase in p-Akt along with an increase in p-ERK levels. In conclusion, TH appears to have a dose-dependent bimodal effect on post-ischemic cardiac performance and this effect may, at least in part, be mediated by a distinct pattern of activation of Akt and ERK signaling.     

The turbulent life of Sirevirus retrotransposons and the evolution of the maize genome: more than ten thousand elements tell the story

Sireviruses are one of the three genera of Copia long terminal repeat (LTR) retrotransposons, exclusive to and highly abundant in plants, and with a unique, among retrotransposons, genome structure. Yet, perhaps due to the few references to the Sirevirus origin of some families, compounded by the difficulty in correctly assigning retrotransposon families into genera, Sireviruses have hardly featured in recent research. As a result, analysis at this key level of classification and details of their colonization and impact on plant genomes are currently lacking. Recently, however, it became possible to accurately assign elements from diverse families to this genus in one step, based on highly conserved sequence motifs. Hence, Sirevirus dynamics in the relatively obese maize genome can now be comprehensively studied. Overall, we identified >10 600 intact and approximately 28 000 degenerate Sirevirus elements from a plethora of families, some brought into the genus for the first time. Sireviruses make up approximately 90% of the Copia population and it is the only genus that has successfully infiltrated the genome, possibly by experiencing intense amplification during the last 600 000 years, while being constantly recycled by host mechanisms. They accumulate in chromosome-distal gene-rich areas, where they insert in between gene islands, mainly in preferred zones within their own genomes. Sirevirus LTRs are heavily methylated, while there is evidence for a palindromic consensus target sequence. This work brings Sireviruses in the spotlight, elucidating their lifestyle and history, and suggesting their crucial role in the current genomic make-up of maize, and possibly other plant hosts.     

Morphological differences in human zygotes and embryos cultured in different media

Summary Purpose: To compare the effects of four culture media on the quality of human zygotes and embryos. Methods: Prospective study analyzing 2289 human embryos cultivated simultaneously in two different culture media: HTF, the default medium, with either Universal IVF, Global or IVF-30 as the secondary media. The sibling oocytes by each patient were randomly divided between the two culture media following intracytoplasmic sperm injection (ICSI). On day 1 the pronuclear stage of zygotes were evaluated and on day 2 embryos were evaluated according to the number of cells, percentage of fragmentation and number of nuclei. Z-test and odds ratios were used in the statistical analysis. Results: There was a higher percentage (55.2%) of class A1 + A2 zygotes with IVF-30 compared with HTF, Global or Universal IVF media (49.1%, 44.7% and 44.2%, respectively). The percentage of Top embryos was significantly higher with Global (40.2%) compared with HTF (21.3%), IVF-30 (25.0%) or Universal IVF media (11.2%). Conclusions: Global medium produced more Top embryos evaluated on day 2 of development.     

Changes in the conformation of 5S rRNA cause alterations in principal functions of the ribosomal nanomachine

5S rRNA is an integral component of the large ribosomal subunit in virtually all living organisms. Polyamine binding to 5S rRNA was investigated by cross-linking of N¹-azidobenzamidino (ABA)-spermine to naked 5S rRNA or 50S ribosomal subunits and whole ribosomes from Escherichia coli cells. ABA-spermine cross-linking sites were kinetically measured and their positions in 5S rRNA were localized by primer extension analysis. Helices III and V, and loops A, C, D and E in naked 5S rRNA were found to be preferred polyamine binding sites. When 50S ribosomal subunits or poly(U)-programmed 70S ribosomes bearing tRNA^Phe at the E-site and AcPhe-tRNA at the P-site were targeted, the susceptibility of 5S rRNA to ABA-spermine was greatly reduced. Regardless of 5S rRNA assembly status, binding of spermine induced significant changes in the 5S rRNA conformation; loop A adopted an apparent ‘loosening’ of its structure, while loops C, D, E and helices III and V achieved a more compact folding. Poly(U)-programmed 70S ribosomes possessing 5S rRNA cross-linked with spermine were more efficient than control ribosomes in tRNA binding, peptidyl transferase activity and translocation. Our results support the notion that 5S rRNA serves as a signal transducer between regions of 23S rRNA responsible for principal ribosomal functions.