Telomeric expression sites are highly conserved in trypanosoma brucei

Subtelomeric regions are often under-represented in genome sequences of eukaryotes. One of the best known examples of the use of telomere proximity for adaptive purposes are the bloodstream expression sites (BESs) of the African trypanosome Trypanosoma brucei. To enhance our understanding of BES structure and function in host adaptation and immune evasion, the BES repertoire from the Lister 427 strain of T. brucei were independently tagged and sequenced. BESs are polymorphic in size and structure but reveal a surprisingly conserved architecture in the context of extensive recombination. Very small BESs do exist and many functioning BESs do not contain the full complement of expression site associated genes (ESAGs). The consequences of duplicated or missing ESAGs, including ESAG9, a newly named ESAG12, and additional variant surface glycoprotein genes (VSGs) were evaluated by functional assays after BESs were tagged with a drug-resistance gene. Phylogenetic analysis of constituent ESAG families suggests that BESs are sequence mosaics and that extensive recombination has shaped the evolution of the BES repertoire. This work opens important perspectives in understanding the molecular mechanisms of antigenic variation, a widely used strategy for immune evasion in pathogens, and telomere biology

Get Organised: The 'Do's' Preceding Successful Field Research

There is no shortage in the political science literature on field research regarding issues of research design, methodology, and data evaluation. Yet, the practical and organisational intricacies that precede successful fieldwork are frequently overlooked. This lack of methodical advice may be due to the impression that field research is highly contextual, and so case-specific that general guidelines, which apply to all field research endeavours alike, are inconceivable. While we acknowledge the organisational complexity of field research, we disagree with the notion that the preparatory dimension of fieldwork is by necessity unique for every undertaking. Rather, recommendations for common challenges that occur during the preparation and organisation phase of a field trip can be identified and formulated. Consequently, we present and discuss ten organisational ?do's? preceding successful field research. Current graduate students and future field researchers will regard these ten pointers as useful hints in the organisation of their own endeavour. While the list is by no means exhaustive, the ten recommendations will lower the organisational entry costs of aspiring field researchers, and enable them to hit the ground running when arriving in the field

Acquisition of naturally occurring antibody responses to recombinant protein domains of Plasmodium falciparum erythrocyte membrane protein 1

Background: Antibodies targeting variant antigens expressed on the surface of Plasmodium falciparum infected erythrocytes have been associated with protection from clinical malaria. The precise target for these antibodies is unknown. The best characterized and most likely target is the erythrocyte surface-expressed variant protein family Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). Methods: Using recombinant proteins corresponding to five domains of the expressed A4 var gene, A4 PfEMP1, the naturally occurring antibody response was assessed, by ELISA, to each domain in serum samples obtained from individuals resident in two communities of differing malaria transmission intensity on the Kenyan coast. Using flow cytometry, the correlation in individual responses to each domain with responses to intact A4-infected erythrocytes expressing A4 PfEMP1 on their surface as well as responses to two alternative parasite clones and one clinical isolate was assessed. Results: Marked variability in the prevalence of responses between each domain and between each transmission area was observed, as wasa strong correlation between age and reactivity with some but not all domains. Individual responses to each domain varied strikingly, with some individuals showing reactivity to all domains and others with no reactivity to any, this was apparent at all age groups. Evidence for possible cross-reactivity in responses to the domain DBL4γ was found. Conclusion: Individuals acquire antibodies to surface expressed domains of a highly variant protein. The finding of potential cross-reactivity in responses to one of these domains is an important initial finding in the consideration of potential vaccine targets

Antigenic Variation in Plasmodium falciparum Malaria Involves a Highly Structured Switching Pattern

Many pathogenic bacteria, fungi, and protozoa achieve chronic infection through an immune evasion strategy known as antigenic variation. In the human malaria parasite Plasmodium falciparum, this involves transcriptional switching among members of the var gene family, causing parasites with different antigenic and phenotypic characteristics to appear at different times within a population. Here we use a genome-wide approach to explore this process in vitro within a set of cloned parasite populations. Our analyses reveal a non-random, highly structured switch pathway where an initially dominant transcript switches via a set of switch-intermediates either to a new dominant transcript, or back to the original. We show that this specific pathway can arise through an evolutionary conflict in which the pathogen has to optimise between safeguarding its limited antigenic repertoire and remaining capable of establishing infections in non-naïve individuals. Our results thus demonstrate a crucial role for structured switching during the early phases of infections and provide a unifying theory of antigenic variation in P. falciparum malaria as a balanced process of parasite-intrinsic switching and immune-mediated selection

Are the pneumococcal polysaccharide vaccines effective? Meta-analysis of the prospective trials

The objective was to review the evidence of effectiveness of the polyvalent polysaccharide pneumococcal vaccine from prospective properly randomised controlled trials comparing pneumococcal vaccines with placebo in subjects who are immunocompetent and those likely to have an impaired immune system. Databases searched included the Cochrane Library, (issue 2, 2000), MEDLINE (1966-August 2000), PubMed (to August 2000) and EMBASE ( to August 2000). Reference lists of reports and reviews were also searched. To be included in the analysis, a study had to have been a prospective randomised comparison of a polysaccharide pneumococcal vaccine (any valency) and to have a placebo or no treatment comparison group. Papers had to report important clinical outcomes, such as rates of pneumonia, pneumococcal pneumonia, lower respiratory tract infections, pneumonia deaths or bacteraemia. Serological outcomes were not sought. Thirteen randomised comparisons with over 45,000 subjects were identified in an extensive literature review. Eight studies had a quality score of 3 or more on a scale of 1 to 5. In three comparisons with 21,152 immunocompetent subjects (South African gold miners, New Guinea highlanders) pneumococcal vaccination was effective in reducing the incidence of all-cause pneumonia (relative risk 0.56, 95% confidence interval 0.47 to 0.66), pneumococcal pneumonia (0.16; 0.11 to 0.23), pneumonia deaths (0.70; 0.50 to 0.96) and bacteraemia (0.18; 0.09 to 0.34). In ten comparisons in over 24,000 people who were elderly or likely to have impaired immune systems, pneumococcal vaccination was without effect for any outcome. Present guidelines recommend pneumococcal vaccination for "high-risk" groups. There is no evidence from randomised trials that this is of any benefit

Left dorsolateral prefrontal cortex supports context-dependent prioritisation of off-task thought

When environments lack compelling goals, humans often let their minds wander to thoughts with greater personal relevance; however, we currently do not understand how this context-dependent prioritisation process operates. Dorsolateral prefrontal cortex (DLPFC) maintains goal representations in a context-dependent manner. Here, we show this region is involved in prioritising off-task thought in an analogous way. In a whole brain analysis we established that neural activity in DLPFC is high both when ‘on-task’ under demanding conditions and ‘off-task’ in a non-demanding task. Furthermore, individuals who increase off-task thought when external demands decrease, show lower correlation between neural signals linked to external tasks and lateral regions of the DMN within DLPFC, as well as less cortical grey matter in regions sensitive to these external task relevant signals. We conclude humans prioritise daydreaming when environmental demands decrease by aligning cognition with their personal goals using DLPFC

Interaction between gemcitabine and topotecan in human non-small-cell lung cancer cells: effects on cell survival, cell cycle and pharmacogenetic profile

The pyrimidine analogue gemcitabine is an established effective agent in the treatment of non-small-cell lung cancer (NSCLC). The present study investigates whether gemcitabine would be synergistic with the topoisomerase I inhibitor topotecan against the NSCLC A549 and Calu-6 cells. Cells were treated with gemcitabine and topotecan for 1 h and the type of drug interaction was assessed using the combination index (CI). Cell cycle alterations were analysed by flow cytometry, while apoptosis was examined by the occurrence of DNA internucleosomal fragmentation, nuclear condensation and caspase-3 activation. Moreover, the possible involvement of the PI3K-Akt signalling pathway was investigated by the measurement of Akt phosphorylation. Finally, quantitative, real-time PCR (QRT-PCR) was used to study modulation of the gemcitabine-activating enzyme deoxycytidine kinase (dCK) and the cellular target enzyme ribonucleotide reductase (RR). In results, it was found that simultaneous and sequential topotecan → gemcitabine treatments were synergistic, while the reverse sequence was antagonistic in both cell lines. DNA fragmentation, nuclear condensation and enhanced caspase-3 activity demonstrated that the drug combination markedly increased apoptosis in comparison with either single agent, while cell cycle analysis showed that topotecan increased cells in S phase. Furthermore, topotecan treatment significantly decreased the amount of the activated form of Akt, and enhanced the expression of dCK (+155.0 and +115.3% in A549 and Calu-6 cells, respectively), potentially facilitating gemcitabine activity. In conclusion, these results indicate that the combination of gemcitabine and topotecan displays schedule-dependent activity in vitro against NSCLC cells. The gemcitabine → topotecan sequence is antagonistic while drug synergism is obtained with the simultaneous and the sequential topotecan → gemcitabine combinations, which are associated with induction of decreased Akt phosphorylation and increased dCK expression

Evolution of the Multi-Domain Structures of Virulence Genes in the Human Malaria Parasite, Plasmodium falciparum

The var gene family of Plasmodium falciparum encodes the immunodominant variant surface antigens PfEMP1. These highly polymorphic proteins are important virulence factors that mediate cytoadhesion to a variety of host tissues, causing sequestration of parasitized red blood cells in vital organs, including the brain or placenta. Acquisition of variant-specific antibodies correlates with protection against severe malarial infections; however, understanding the relationship between gene expression and infection outcome is complicated by the modular genetic architectures of var genes that encode varying numbers of antigenic domains with differential binding specificities. By analyzing the domain architectures of fully sequenced var gene repertoires we reveal a significant, non-random association between the number of domains comprising a var gene and their sequence conservation. As such, var genes can be grouped into those that are short and diverse and genes that are long and conserved, suggesting gene length as an important characteristic in the classification of var genes. We then use an evolutionary framework to demonstrate how the same evolutionary forces acting on the level of an individual gene may have also shaped the parasite's gene repertoire. The observed associations between sequence conservation, gene architecture and repertoire structure can thus be explained by a trade-off between optimizing within-host fitness and minimizing between-host immune selection pressure. Our results demonstrate how simple evolutionary mechanisms can explain var gene structuring on multiple levels and have important implications for understanding the multifaceted epidemiology of P. falciparum malaria

Inhibition of macrophage migration inhibitory factor decreases proliferation and cytokine expression in bladder cancer cells

BACKGROUND: The importance of various inflammatory cytokines in maintaining tumor cell growth and viability is well established. Increased expression of the proinflammatory cytokine macrophage migration inhibitory factor (MIF) has previously been associated with various types of adenocarcinoma. METHODS: MIF IHC was used to localize MIF in human bladder tissue. ELISA and Western blot analysis determined the synthesis and secretion of MIF by human bladder transitional cell carcinoma cells. The effects of MIF inhibitors (high molecular weight hyaluronate (HA), anti-MIF antibody or MIF anti-sense) on cell growth and cytokine expression were analyzed. RESULTS: Human bladder cancer cells (HT-1376) secrete detectable amounts of MIF protein. Treatment with HA, anti-MIF antibody and MIF anti-sense reduced HT-1376 cell proliferation, MIF protein secretion, MIF gene expression and secreted inflammatory cytokines. Our evidence suggests MIF interacts with the invariant chain, CD74 and the major cell surface receptor for HA, CD44. CONCLUSIONS: This study is the first to report MIF expression in the human bladder and these findings support a role for MIF in tumor cell proliferation. Since MIF participates in the inflammatory response and bladder cancer is associated with chronic inflammatory conditions, these new findings suggest that neutralizing bladder tumor MIF may serve as a novel therapeutic treatment for bladder carcinoma