A broadly neutralizing human monoclonal antibody that recognizes a conserved, novel epitope on the globular head of the influenza H1N1 virus hemagglutinin

The conserved influenza virus hemagglutinin (HA) stem domain elicits cross-reactive antibodies, but epitopes in the globular head typically elicit strain-specific responses because of the hypervariability of this region. We isolated human monoclonal antibody 5J8, which neutralized a broad spectrum of 20th century H1N1 viruses and the 2009 pandemic H1N1 virus. Fine mapping of the interaction unexpectedly revealed a novel epitope between the receptor-binding pocket and the Ca₂ antigenic site on HA. This antibody exposes a new mechanism underlying broad immunity against H1N1 influenza viruses and identifies a conserved epitope that might be incorporated into engineered H1 virus vaccines.     

Complement resistance of Borrelia burgdorferi correlates with the expression of BbCRASP-1, a novel linear plasmid-encoded surface protein that interacts with human factor H and FHL-1 and is unrelated to Erp proteins

The etiologic agent of Lyme disease, Borrelia burgdorferi, is capable of circumventing the immune defense of a variety of potential vertebrate hosts. Previous work has shown that interaction of host-derived complement regulators, factor H and factor H-like protein 1 (FHL-1), with up to five complement regulator-acquiring surface proteins (CRASPs) expressed by resistant B. burgdorferi sensu lato isolates conferred complement resistance. In addition expression of CRASP-1 is directly correlated with complement resistance of Borrelia species. This work describes the functional characterization of BbCRASP-1 as the dominant factor H and FHL-1-binding protein of B. burgdorferi. The corresponding gene, zs7.a68, is located on the linear plasmid lp54 and is different from factor H-binding Erp proteins that are encoded by genes localized on circular plasmids (cp32). Deletion mutants of BbCRASP-1 were generated, and a high affinity binding site for factor H and FHL-1 was mapped to the C terminus of BbCRASP-1. Similarly, the predominant binding site of factor H and FHL-1 was localized to the short consensus repeat 7. Factor H and FHL-1 maintain their cofactor activity for factor I-mediated C3b inactivation when bound to BbCRASP-1, and factor H is up to 6-fold more efficient in mediating C3b conversion than FHL-1. In conclusion, BbCRASP-1 (i). binds the host complement regulators factor H and FHL-1 with high affinity, (ii). is the key molecule of the complement resistance of spirochetes, and (iii). is distinct from the Erp protein family. Thus, BbCRASP-1 most likely contributes to persistence of B. burgdorferi and to pathogenesis of Lyme disease.     

CULTIVATION‐INDEPENDENT SPECIES IDENTIFICATION OF DINOBRYON SPECIES (CHRYSOPHYCEAE) BY MEANS OF MULTIPLEX SINGLE‐CELL PCR1

With the discovery of a high molecular diversity of protists, a discrepancy between morphological and molecular species richness estimates became apparent. Solving the current concerns requires a comparative analysis of different sequences combined with morphological analyses of single cells originating from preserved field samples. We refined a single‐cell PCR (SC‐PCR) protocol for analyzing cells from field samples preserved with Lugol’s iodine solution. We linked microscopic screening with multiplex PCR targeting the SSU rDNA, internal transcribed spacer 1 (ITS1), 5.8S rDNA, internal transcribed spacer 2 (ITS2), and the mitochondrial cytochrome oxidase 1 (CO1) in a single PCR reaction. Using this method, we investigated the intraspecific molecular variation in Dinobryon populations originating from two lakes in the Salzkammergut area of Austria. All investigated genetic markers showed two separated clusters within the investigated populations of Dinobryon divergens O. E. Imhof, indicating a reproductive isolation of the two coexisting populations. Based on these findings, we describe a lineage, which is morphologically similar to D. divergens but, based on the molecular data, is reproductively isolated.     

Factors shaping community patterns of protists and bacteria on a European scale

Factors shaping community patterns of microorganisms are controversially discussed. Physical and chemical factors certainly limit the survival of individual taxa and maintenance of diversity. In recent years, a contribution of geographic distance and dispersal barriers to distribution patterns of protists and bacteria has been demonstrated. Organismic interactions such as competition, predation and mutualism further modify community structure and maintenance of distinct taxa. Here, we address the relative importance of these different factors in shaping protists and bacterial communities on a European scale using high-throughput sequencing data obtained from lentic freshwater ecosystems. We show that community patterns of protists are similar to those of bacteria. Our results indicate that cross-domain organismic factors are important variables with a higher influence on protists as compared with bacteria. Abiotic physical and chemical factors also contributed significantly to community patterns. The contribution of these latter factors was higher for bacteria, which may reflect a stronger biogeochemical coupling. The contribution of geographical distance was similar for both microbial groups.     

Unbiased Approach for Virus Detection in Skin Lesions

To assess presence of virus DNA in skin lesions, swab samples from 82 squamous cell carcinomas of the skin (SCCs), 60 actinic keratoses (AKs), paraffin-embedded biopsies from 28 SCCs and 72 kerathoacanthomas (KAs) and fresh-frozen biopsies from 92 KAs, 85 SCCs and 92 AKs were analyzed by high throughput sequencing (HTS) using 454 or Ion Torrent technology. We found total of 4,284 viral reads, out of which 4,168 were Human Papillomavirus (HPV)-related, belonging to 15 known (HPV8, HPV12, HPV20, HPV36, HPV38, HPV45, HPV57, HPV59, HPV104, HPV105, HPV107, HPV109, HPV124, HPV138, HPV147), four previously described putative (HPV 915 F 06 007 FD1, FA73, FA101, SE42) and two putatively new HPV types (SE46, SE47). SE42 was cloned, sequenced, designated as HPV155 and found to have 76% similarity to the most closely related known HPV type. In conclusion, an unbiased approach for viral DNA detection in skin tumors has found that, although some new putative HPVs were found, known HPV types constituted most of the viral DNA.     

Association between Body Mass Index,Asymmetric Dimethylarginine and Risk of Cardiovascular Events and Mortality in Norwegian Patients with Suspected Stable Angina Pectoris

Background

Asymmetric dimethylarginine (ADMA) is associated with increased risk of atherosclerotic cardiovascular disease and mortality through inhibition of nitrogen oxide (NO) synthesis. As positive correlations between serum concentrations of NO and body mass index (BMI) have been observed, we aimed to explore whether the potential associations between plasma ADMA levels and the risk of acute myocardial infarction (AMI) and mortality were modified by BMI.

Methods

Multivariable Cox proportional hazard models were used to estimate the hazard ratios (HR) for AMI, cardiovascular death and all-cause mortality according to baseline plasma ADMA levels in 4122 patients with suspected stable angina pectoris. Analyses were subsequently repeated in patients with BMI below (low BMI) or above (high BMI) median.

Results

A total of 2982 patients (72%) were men. Median (range) age, plasma ADMA level and BMI were 62 (21–88) years, 0.54 (0.10–1.25) μmol/L and 26.3 (18.5–54.3) kg/m², respectively. During a mean (standard deviation) follow-up time of 4.7 (1.4) years, 337 (8%) patients suffered from an AMI, 300 (7%) died, whereof 165 (55%) due to cardiovascular disease. Each 0.1 μmol/L increment in plasma ADMA level was associated with an increased risk of AMI (HR (95% CI) 1.21 (1.08, 1.35) and cardiovascular death 1.30 (1.13, 1.49) in participants with low BMI only. Interactions were significant for AMI (p = 0.04) and CV death (p = 0.03). BMI did not modify the association between plasma ADMA levels and all-cause mortality.

Conclusion

Plasma ADMA levels were associated with risk of AMI and cardiovascular death among patients with low BMI only.     

Early-Life Stress Affects Stress-Related Prefrontal Dopamine Activity in Healthy Adults,but Not in Individuals with Psychotic Disorder

Early life stress may have a lasting impact on the developmental programming of the dopamine (DA) system implicated in psychosis. Early adversity could promote resilience by calibrating the prefrontal stress-regulatory dopaminergic neurotransmission to improve the individual’s fit with the predicted stressful environment. Aberrant reactivity to such match between proximal and distal environments may, however, enhance psychosis disease risk. We explored the combined effects of childhood adversity and adult stress by exposing 12 unmedicated individuals with a diagnosis of non-affective psychotic disorder (NAPD) and 12 healthy controls (HC) to psychosocial stress during an [¹⁸F]fallypride positron emission tomography. Childhood trauma divided into early (ages 0–11 years) and late (12–18 years) was assessed retrospectively using a questionnaire. A significant group x childhood trauma interaction on the spatial extent of stress-related [¹⁸F]fallypride displacement was observed in the mPFC for early (b = -8.45, t(1,23) = -3.35, p = .004) and late childhood trauma (b = -7.86, t(1,23) = -2.48, p = .023). In healthy individuals, the spatial extent of mPFC DA activity under acute psychosocial stress was positively associated with the severity of early (b = 7.23, t(11) = 3.06, p = .016) as well as late childhood trauma (b = -7.86, t(1,23) = -2.48, p = .023). Additionally, a trend-level main effect of early childhood trauma on subjective stress response emerged within this group (b = -.7, t(11) = -2, p = .07), where higher early trauma correlated with lower subjective stress response to the task. In the NAPD group, childhood trauma was not associated with the spatial extent of the tracer displacement in mPFC (b = -1.22, t(11) = -0.67), nor was there a main effect of trauma on the subjective perception of stress within this group (b = .004, t(11) = .01, p = .99). These findings reveal a potential mechanism of neuroadaptation of prefrontal DA transmission to early life stress and suggest its role in resilience and vulnerability to psychosis.     

Antagonism of microRNA-122 in mice by systemically administered LNA-antimiR leads to up-regulation of a large set of predicted target mRNAs in the liver

MicroRNA-122 (miR-122) is an abundant liver-specific miRNA, implicated in fatty acid and cholesterol metabolism as well as hepatitis C viral replication. Here, we report that a systemically administered 16-nt, unconjugated LNA (locked nucleic acid)-antimiR oligonucleotide complementary to the 5′ end of miR-122 leads to specific, dose-dependent silencing of miR-122 and shows no hepatotoxicity in mice. Antagonism of miR-122 is due to formation of stable heteroduplexes between the LNA-antimiR and miR-122 as detected by northern analysis. Fluorescence in situ hybridization demonstrated uptake of the LNA-antimiR in mouse liver cells, which was accompanied by markedly reduced hybridization signals for mature miR-122 in treated mice. Functional antagonism of miR-122 was inferred from a low cholesterol phenotype and de-repression within 24 h of 199 liver mRNAs showing significant enrichment for miR-122 seed matches in their 3′ UTRs. Expression profiling extended to 3 weeks after the last LNA-antimiR dose revealed that most of the changes in liver gene expression were normalized to saline control levels coinciding with normalized miR-122 and plasma cholesterol levels. Combined, these data suggest that miRNA antagonists comprised of LNA are valuable tools for identifying miRNA targets in vivo and for studying the biological role of miRNAs and miRNA-associated gene-regulatory networks in a physiological context.     

Improving the phenotype predictions of a yeast genome‐scale metabolic model by incorporating enzymatic constraints

Genome‐scale metabolic models (GEMs) are widely used to calculate metabolic phenotypes. They rely on defining a set of constraints, the most common of which is that the production of metabolites and/or growth are limited by the carbon source uptake rate. However, enzyme abundances and kinetics, which act as limitations on metabolic fluxes, are not taken into account. Here, we present GECKO, a method that enhances a GEM to account for enzymes as part of reactions, thereby ensuring that each metabolic flux does not exceed its maximum capacity, equal to the product of the enzyme's abundance and turnover number. We applied GECKO to a Saccharomyces cerevisiae GEM and demonstrated that the new model could correctly describe phenotypes that the previous model could not, particularly under high enzymatic pressure conditions, such as yeast growing on different carbon sources in excess, coping with stress, or overexpressing a specific pathway. GECKO also allows to directly integrate quantitative proteomics data; by doing so, we significantly reduced flux variability of the model, in over 60% of metabolic reactions. Additionally, the model gives insight into the distribution of enzyme usage between and within metabolic pathways. The developed method and model are expected to increase the use of model‐based design in metabolic engineering.