Publications search

Background Staphylococcus aureus is a major human pathogen that can colonize healthy people mainly in the anterior nares. The aim of the present study was to evaluate S. aureus nasal colonization over time among Portuguese nursing students, including methicillin-resistant S. aureus (MRSA). Methods and findings In this longitudinal cohort study, we collected 280 nasal swabs from nursing students at 14 time points over four years of schooling (2012-2016). The isolates were characterized by pulsed-field gel electrophoresis (PFGE), spa typing, multilocus sequence typing (MLST), and SCCmec typing for MRSA. Among 47 students, 20 (43%) carried methicillin-susceptible S. aureus (MSSA) at admission, but none was colonized with MRSA. A total of 19 students (40%) became colonized after exposure during the nursing training, out of which five carried MRSA. Overall, 39 students (83%) had S. aureus detected at least once during the study period. Among the 97 MSSA isolates, most (65%) belonged to four clones: PFGE A-ST30 (21%), B-ST72 (20%), C-ST508 (13%), and D-ST398 (11%). Three of the five MRSA carriers were colonized with the predominant clone circulating in Portuguese hospitals (ST22-IVh) and two with ST3162-II. Colonization of nursing students was highly dynamic with continuous appearance of strains with distinct PFGE types in the same individual. Conclusions A considerable proportion of students became colonized by S. aureus, including MRSA, during the nursing education, evidencing this population represents an important reservoir of S. aureus. Therefore, education on infection control measures in nursing schools is of major importance.

S531 of Escherichia coli RNA polymerase (RNAP) beta subunit is a part of RNA binding domain in transcription complex. While highly conserved, S531 is not involved in interactions within the transcription complex as suggested by X-ray analysis. To understand the basis for S531 conservation we performed systematic mutagenesis of this residue. We find that the most of the mutations significantly decreased initiation-to-elongation transition by RNAP. Surprisingly, some changes enhanced the production of full-size transcripts by suppressing abortive loss of short RNAs. S531-R increased transcript retention by establishing a salt bridge with RNA, thereby explaining the R substitution at the equivalent position in extremophilic organisms, in which short RNAs retention is likely to be an issue. Generally, the substitutions had the same effect on bacterial doubling time when measured at 20 degrees. Raising growth temperature to 37 degrees ablated the positive influence of some mutations on the growth rate in contrast to their in vitro action, reflecting secondary effects of cellular environment on transcription and complex involvement of 531 locus in the cell biology. The properties of generated RNAP variants revealed an RNA/protein interaction network that is crucial for transcription, thereby explaining the details of initiation-to-elongation transition on atomic level.

Polycomb group proteins (PcG) are transcriptional repressors that control cell identity and development. In mammals, five different CBX proteins associate with the core Polycomb repressive complex 1 (PRC1). In mouse embryonic stem cells (ESCs), CBX6 and CBX7 are the most highly expressed CBX family members. CBX7 has been recently characterized, but little is known regarding the function of CBX6. Here, we show that CBX6 is essential for ESC identity. Its depletion destabilizes the pluripotency network and triggers differentiation. Mechanistically, we find that CBX6 is physically and functionally associated to both canonical PRC1 (cPRC1) and non-canonical PRC1 (ncPRC1) complexes. Notably, in contrast to CBX7, CBX6 is recruited to chromatin independently of H3K27me3. Taken together, our findings reveal that CBX6 is an essential component of ESC biology that contributes to the structural and functional complexity of the PRC1 complex.

Calcium phosphate-based mineralo-organic particles form spontaneously in the body and may represent precursors of ectopic calcification. We have shown earlier that these particles induce activation of caspase-1 and secretion of IL-1 beta by macrophages. However, whether the particles may produce other effects on immune cells is unclear. Here, we show that these particles induce the release of neutrophil extracellular traps (NETs) in a size-dependent manner by human neutrophils. Intracellular production of reactive oxygen species is required for particle-induced NET release by neutrophils. NETs contain the high-mobility group protein B1 (HMGB1), a DNA-binding protein capable of inducing secretion of TNF-alpha by a monocyte/macrophage cell line and primary macrophages. HMGB1 functions as a ligand of Toll-like receptors 2 and 4 on macrophages, leading to activation of the MyD88 pathway and TNF-alpha production. Furthermore, HMGB1 is critical to activate the particle-induced pro-inflammatory cascade in the peritoneum of mice. These results indicate that mineral particles promote pro-inflammatory responses by engaging neutrophils and macrophages via signaling of danger signals through NETs.

Intestinal intraepithelial lymphocytes (IELs) are located at the critical interface between the intestinal lumen, which is chronically exposed to food and microbes, and the core of the body. Using high-resolution microscopy techniques and intersectional genetic tools, we investigated the nature of IEL responses to luminal microbes. We observed that TCRgd IELs exhibit unique microbiota-dependent location and movement patterns in the epithelial compartment. This behavioral pattern quickly changes upon exposure to different enteric pathogens, resulting in increased interepithelial cell (EC) scanning, expression of antimicrobial genes, and glycolysis. Both dynamic and metabolic changes to gamma delta IEL depend on pathogen sensing by ECs. Direct modulation of glycolysis is sufficient to change gd IEL behavior and susceptibility to early pathogen invasion. Our results uncover a coordinated EC-IEL response to enteric infections that modulates lymphocyte energy utilization and dynamics and supports maintenance of the intestinal epithelial barrier.

Voltage-gated Na+ channels (Na-v) modulate neuronal excitability, but the roles of the various Na-v subtypes in specific neuronal functions such as synaptic transmission are unclear. We investigated expression of the three major brain Na-v subtypes (Na(v)1.1, Na(v)1.2, Na(v)1.6) in area CA1 and dentate gyrus of rat hippocampus. Using light and electron microscopy, we found labeling for all three Na-v subtypes on dendrites, dendritic spines, and axon terminals, but the proportion of pre- and post-synaptic labeling for each subtype varied within and between subregions of CA1 and dentate gyrus. In the central hilus (CH) of the dentate gyrus, Na(v)1.1 immunoreactivity was selectively expressed in presynaptic profiles, while Na(v)1.2 and Na(v)1.6 were expressed both pre- and post-synaptically. In contrast, in the stratum radiatum (SR) of CA1, Na(v)1.1, Na(v)1.2, and Na(v)1.6 were selectively expressed in postsynaptic profiles. We next compared differences in Na-v subtype expression between CH and SR axon terminals and between CH and SR dendrites and spines. Na(v)1.1 and Na(v)1.2 immunoreactivity was preferentially localized to CH axon terminals compared to SR, and in SR dendrites and spines compared to CH. No differences in Na(v)1.6 immunoreactivity were found between axon terminals of CH and SR or between dendrites and spines of CH and SR. All Na-v subtypes in both CH and SR were preferentially associated with asymmetric synapses rather than symmetric synapses. These findings indicate selective presynaptic and postsynaptic Na-v expression in glutamatergic synapses of CH and SR supporting neurotransmitter release and synaptic plasticity.

In psoriasis, an IL-17-mediated inflammatory skin disease, skin lesions resolve with therapy, but often recur in the same locations when therapy is discontinued. We propose that residual T cell populations in resolved psoriatic lesions represent the pathogenic T cells of origin in this disease. Utilizing high-throughput screening (HTS) of the T cell receptor (TCR) and immunostaining, we found that clinically resolved psoriatic lesions contained oligoclonal populations of T cells that produced IL-17A in both resolved and active psoriatic lesions. Putative pathogenic clones preferentially utilized particular V beta and V alpha subfamilies. We identified 15 TCR beta and 4 TCR alpha antigen receptor sequences shared between psoriasis patients and not observed in healthy controls or other inflammatory skin conditions. To address the relative roles of alpha beta versus gamma delta T cells in psoriasis, we carried out TCR/delta HTS. These studies demonstrated that the majority of T cells in psoriasis and healthy skin are alpha beta T cells. gamma delta T cells made up 1% of T cells in active psoriasis, less than 1% in resolved psoriatic lesions, and less than 2% in healthy skin. All of the 70 most frequent putative pathogenic T cell clones were alpha beta T cells. In summary, IL-17-producing alpha beta T cell clones with psoriasis-specific antigen receptors exist in clinically resolved psoriatic skin lesions. These cells likely represent the disease-initiating pathogenic T cells in psoriasis, suggesting that lasting control of this disease will require suppression of these resident T cell populations.

Psoriasis is a common immune-mediated disease that affects 2%-4% of individuals in North America and Europe. In the past decade, advances in research have led to an improved understanding of immune pathways involved in the pathogenesis of psoriasis and has spurred the development of targeted therapeutics. Recently, three psoriasis autoantigens have been described: cathelicidin (LL37), a disintegrin and metalloprotease domain containing thrombospondin type 1 motif-like 5 (ADAMTSL5), and lipid antigens generated by phospholipase A2 (PLA2) group IVD (PLA2G4D). It is important to establish the expression, regulation and therapeutic modulation of these psoriasis autoantigens. In this study, we performed immunohistochemistry and two-colour immunofluorescence on non-lesional and lesional psoriasis skin to characterize ADAMTSL5 and LL37, and their co-expression with T cells, dendritic cells, neutrophils and macrophages, which are the main immune cells that drive this disease. Our results showed that ADAMTSL5 and LL37 are significantly (P<.05) increased in lesional skin and are co-expressed by many dendritic cells, macrophages and some T cells in the dermis. Gene expression analysis showed significant (P<.05) upregulation of LL37 in lesional skin and significant downregulation following treatment with etanercept. ADAMTSL5 and LL37 are also significantly decreased by IL-17 or TNF- blockade, suggesting feed-forward induction of psoriasis autoantigens by disease-related cytokines.

Background: Oil palm is the most productive oil crop and the efficiency of pollination has a direct impact on the yield of oil. Pollination by wind can occur but maximal pollination is mediated by the weevil E. kamerunicus. These weevils complete their life cycle by feeding on male flowers. Attraction of weevils to oil palm flowers is due to the emission of methylchavicol by both male and female flowers. In search for male flowers, the weevils visit female flowers by accident due to methylchavicol fragrance and deposit pollen. Given the importance of methylchavicol emission on pollination, we performed comparative transcriptome analysis of oil palm flowers and leaves to identify candidate genes involved in methylchavicol production in flowers. Results: RNA sequencing (RNA-Seq) of male open flowers, female open flowers and leaves was performed using Illumina HiSeq 2000 platform. Analysis of the transcriptome data revealed that the transcripts of methylchavicol biosynthesis genes were strongly up-regulated whereas transcripts encoding genes involved in lignin production such as, caffeic acid O-methyltransferase (COMT) and Ferulate-5-hydroxylase (F5H) were found to be suppressed in oil palm flowers. Among the transcripts encoding transcription factors, an EAR-motif-containing R2R3-MYB transcription factor (EgMYB4) was found to be enriched in oil palm flowers. We determined that EgMYB4 can suppress the expression of a monolignol pathway gene, EgCOMT, in vivo by binding to the AC elements present in the promoter region. EgMYB4 was further functionally characterized in sweet basil which also produces phenylpropenes like oil palm. Transgenic sweet basil plants showed significant reduction in lignin content but produced more phenylpropenes. Conclusions: Our results suggest that EgMYB4 possibly restrains lignin biosynthesis in oil palm flowers thus allowing enhanced carbon flux into the phenylpropene pathway. This study augments our understanding of the diverse roles that EAR-motif-containing MYBs play to fine tune the metabolic flux along the various branches of core phenylpropanoid pathway. This will aid in metabolic engineering of plant aromatic compounds.