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Error: It's not possible to reach RSS file... BMC Cell Biology - Latest Articles
The latest research articles published by BMC Cell Biology
- ERAD and protein import defects in a sec61 mutant lacking ER-lumenal loop 7
Background: The Sec61 channel mediates protein translocation across the endoplasmic reticulum (ER) membrane during secretory protein biogenesis, and likely also during export of misfolded proteins for ER-associated degradation (ERAD). The mechanisms of channel opening for the different modes of translocation are not understood so far, but the position of the large ER-lumenal loop 7 of Sec61p suggests a decisive role. Results: We show here that the Y345H mutation in L7 which causes diabetes in the mouse displays no ER import defects in yeast, but a delay in misfolded protein export. A complete deletion of L7 in Sec61p resulted in viable, cold- and tunicamycin-hypersensitive yeast cells with strong defects in posttranslational protein import of soluble proteins into the ER, and in ERAD of soluble substrates. Membrane protein ERAD was only moderately slower in sec61[increment]L7 than in wildtype cells. Although Sec61[increment]L7 channels were unstable in detergent, co-translational protein integration into the ER membrane, proteasome binding to Sec61[increment]L7 channels, and formation of hetero-heptameric Sec complexes were not affected. Conclusions: We conclude that L7 of Sec61p is required for initiation of posttranslational soluble protein import into and misfolded soluble protein export from the ER, suggesting a key role for L7 in transverse gating of the Sec61 channel.
- Primary porcine proximal tubular cells as an alternative to human primary renal cells in vitro: an initial characterization
Background: A good in vitro model should approximate an in vivo-like behavior as closely as possible in order to reflect most likely the in vivo situation. Regarding renal physiology of different species, humans are more closely related to pigs than to rodents, therefore primary porcine kidney cells (PKC) and their subsequent cell strain could be a valid alternative to primary human cells for renal in vitro toxicology. For this PKC must display inherent characteristics (e.g. structural organization) and functions (e.g. transepithelial transport) as observed under in vivo conditions within the respective part of the kidney.FindingsWe carried out a comprehensive characterization of PKC and their subsequent cell strain, including morphology and growth as well as transporter expression and functionality. The data presented here demonstrate that PKC express various transporters including pMrp1 (abcc1), pMrp2 (abcc2), pOat1 (slc22a6) and pOat3 (slc22a8), whereas pMdr1 (abcb1) and pOatp1a2 (slco1a2) mRNA could not be detected in either the PKCs or in the porcine cortical tissue. Functionality of the transporters was demonstrated by determining the specific PAH transport kinetics. Conclusions: On the basis of the presented results it can be concluded that PKC and to some extent their subsequent cell strain represent a valuable model for in vitro toxicology, which might be used as an alternative to human primary cells.
- Differential marker expression by cultures rich in mesenchymal stem cells
Background: Mesenchymal stem cells have properties that make them amenable to therapeutic use. However, the acceptance of mesenchymal stem cells in clinical practice requires standardized techniques for their specific isolation. To date, there are no conclusive marker (s) for the exclusive isolation of mesenchymal stem cells. Our aim was to identify markers differentially expressed between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. We compared and contrasted the phenotype of tissue cultures in which mesenchymal stem cells are rich and rare. By initially assessing mesenchymal stem cell differentiation, we established that bone marrow and breast adipose cultures are rich in mesenchymal stem cells while, in our hands, foreskin fibroblast and olfactory tissue cultures contain rare mesenchymal stem cells. In particular, olfactory tissue cells represent non-stem cell mesenchymal cells. Subsequently, the phenotype of the tissue cultures were thoroughly assessed using immuno-fluorescence, flow-cytometry, proteomics, antibody arrays and qPCR. Results: Our analysis revealed that all tissue cultures, regardless of differentiation potential, demonstrated remarkably similar phenotypes. Importantly, it was also observed that common mesenchymal stem cell markers, and fibroblast-associated markers, do not discriminate between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. Examination and comparison of the phenotypes of mesenchymal stem cell and non-stem cell mesenchymal cell cultures revealed three differentially expressed markers -- CD24, CD108 and CD40. Conclusion: We indicate the importance of establishing differential marker expression between mesenchymal stem cells and non-stem cell mesenchymal cells in order to determine stem cell specific markers.
- Suppression of anoikis in human intestinal epithelial cells: differentiation state-selective roles of ¿2ß1, ¿3ß1, ¿5ß1, and ¿6ß4 integrins
Background: Regulation of anoikis in human intestinal epithelial cells (IECs) implicates differentiation state-specific mechanisms. Human IECs express distinct repertoires of integrins according to their state of differentiation. Therefore, we investigated whether α2β1, α3β1, α5β1, and α6β4 integrins perform differentiation state-specific roles in the suppression of IEC anoikis. Results: Human (HIEC, Caco-2/15) IECs were exposed to specific antibodies that block the binding activity of integrin subunits (α2, α3, α5, α6, β1 or β4) to verify whether or not their inhibition induced anoikis. The knockdown of α6 was also performed by shRNA. Additionally, apoptosis/anoikis was induced by pharmacological inhibition of Fak (PF573228) or Src (PP2). Anoikis/apoptosis was assayed by DNA laddering, ISEL, and/or caspase activity (CASP-8, -9, or -3). Activation levels of Fak and Src, as well as functional Fak-Src interactions, were also assessed. We report herein that differentiated IECs exhibit a greater sensitivity to anoikis than undifferentiated ones. This involves an earlier onset of anoikis when kept in suspension, as well as significantly greater contributions from β1 and β4 integrins in the suppression of anoikis in differentiated cells, and functional distinctions between β1 and β4 integrins in engaging both Fak and Src, or Src only, respectively. Likewise, Fak performs significantly greater contributions in the suppression of anoikis in differentiated cells. Additionally, we show that α2β1 and α5β1 suppress anoikis in undifferentiated cells, whereas α3β1 does so in differentiated ones. Furthermore, we provide evidence that α6β4 contributes to the suppression of anoikis in a primarily α6 subunit-dependent manner in undifferentiated cells, whereas this same integrin in differentiated cells performs significantly greater contributions in anoikis suppression than its undifferentiated state-counterpart, in addition to doing so through a dependence on both of its subunits. Conclusions: Our findings indicate that the suppression of human IEC anoikis implicates differentiation state-selective repertoires of integrins, which in turn results into distinctions in anoikis regulation, and sensitivity, between undifferentiated and differentiated IECs. These data further the functional understanding of the concept that the suppression of anoikis is subjected to cell differentiation state-selective mechanisms.
- BORIS/CTCFL is an RNA-binding protein that associates with polysomes
Background: BORIS (CTCFL), a paralogue of the multifunctional and ubiquitously expressed transcription factor CTCF, is best known for its role in transcriptional regulation. In the nucleus, BORIS is particularly enriched in the nucleolus, a crucial compartment for ribosomal RNA and RNA metabolism. However, little is known about cytoplasmic BORIS, which represents the major pool of BORIS protein. Results: We show, firstly, that BORIS has a putative nuclear export signal in the C-terminal domain. Furthermore, BORIS associates with mRNA in both neural stem cells and young neurons. The majority of the BORIS-associated transcripts are different in the two cell types. Finally, by using polysome profiling we show that BORIS is associated with actively translating ribosomes. Conclusion: We have demonstrated the RNA binding properties of cellular BORIS and its association with actively translating ribosomes. We suggest that BORIS is involved in gene expression at both the transcriptional and post-transcriptional levels.
- THOC5, a member of the mRNA export complex, contributes to processing of a subset of wingless/integrated (Wnt) target mRNAs and integrity of the gut epithelial barrier
Background: THO (Suppressors of the transcriptional defects of hpr1 delta by overexpression) complex 5 (THOC5), an mRNA export protein, is involved in the expression of only 1% of all genes. Using an interferon inducible knockout mouse system, we have previously shown that THOC5 is an essential element in the maintenance of hematopoietic stem cells and cytokine-mediated hematopoiesis in adult mice. Here we interrogate THOC5 function in cell differentiation beyond the hematopoietic system and study pathological changes caused by THOC5 deficiency. Results: To examine whether THOC5 plays a role in general differentiation processes, we generated tamoxifen inducible THOC5 knockout mice. We show here that the depletion of THOC5 impaired not only hematopoietic differentiation, but also differentiation and self renewal of the gut epithelium. Depletion of the THOC5 gene did not cause pathological alterations in liver or kidney.We further show that THOC5 is indispensable for processing of mRNAs induced by Wnt (wingless/integrated) signaling which play key roles in epithelial cell differentiation/proliferation. A subset of Wnt target mRNAs, SRY-box containing gene 9 (Sox9), and achaete-scute complex homolog 2 (Ascl2), but not Fibronectin 1 (Fn1), were down-regulated in THOC5 knockout intestinal cells. The down-regulated Wnt target mRNAs were able to bind to THOC5. Furthermore, pathological alterations in the gastrointestinal tract induced translocation of intestinal bacteria and caused sepsis in mice. The bacteria translocation may cause Toll-like receptor activation. We identified one of the Toll-like receptor inducible genes, prostaglandin-endoperoxidase synthase 2 (Ptgs2 or COX2) transcript as THOC5 target mRNA. Conclusion: THOC5 is indispensable for processing of only a subset of mRNAs, but plays a key role in processing of mRNAs inducible by Wnt signals. Furthermore, THOC5 is dispensable for general mRNA export in terminally differentiated organs, indicating that multiple mRNA export pathways exist. These data imply that THOC5 may be a useful tool for studying intestinal stem cells, for modifying the differentiation processes and for cancer therapy.
- A mouse protein that localizes to acrosome and sperm tail is regulated by Y-chromosome
Background: Acrosomal proteins play crucial roles in the physiology of fertilization. Identification of proteins localizing to the acrosome is fundamental to the understanding of its contribution to fertilization. Novel proteins are still being reported from acrosome. In order to capture yet unreported proteins localizing to acrosome in particular and sperm in general, 2D-PAGE and mass spectrometry analysis of mouse sperm proteins was done. Results: One of the protein spots identified in the above study was reported in the NCBI database as a hypothetical protein from Riken cDNA 1700026L06 that localizes to chromosome number 2. Immunofluorescence studies using the antibody raised in rabbit against the recombinant protein showed that it localized to mouse acrosome and sperm tail. Based on the localization of this protein, it has been named mouse acrosome and sperm tail protein (MAST, [Q7TPM5 (http://www.ncbi.nlm.nih.gov/protein/Q7TPM5)]). This protein shows 96% identity to the rat spermatid specific protein RSB66. Western blotting showed that MAST is expressed testis-specifically. Co-immunoprecipitation studies using the MAST antibody identified two calcium-binding proteins, caldendrin and calreticulin as interacting partners of MAST. Caldendrin and calreticulin genes localize to mouse chromosomes 5 and 8 respectively. In a Yq-deletion mutant mouse, that is subfertile and has a deletion of 2/3rd of the long arm of the Y chromosome, MAST failed to localize to the acrosome. Western blot analysis however, revealed equal expression of MAST in the testes of wild type and mutant mice. The acrosomal calcium-binding proteins present in the MAST IP-complex were upregulated in sperms of Yq-del mice. Conclusions: We have identified a mouse acrosomal protein, MAST, that is expressed testis specifically. MAST does not contain any known motifs for protein interactions; yet it complexes with calcium-binding proteins localizing to the acrosome. The misexpression of all the proteins identified in a complex in the Yq-del mice invokes the hypothesis of a putative pathway regulated by the Y chromosome. The role of Y chromosome in the regulation of this complex is however not clear from the current study.
- The role of c-Src in integrin (¿6ß4) dependent translational control
Background: Integrin α6β4 contributes to cancer progression by stimulating transcription as well as translation of cancer related genes. Our previous study demonstrated that α6β4 stimulates translation initiation of survival factors such as VEGF by activating mTOR pathway. However, the immediate early signaling events that link α6β4 to mTOR activation needs to be defined. Results: In the current studies, we demonstrated that c-Src is an immediate early signaling molecule that acts upstream of α6β4 dependent mTOR activation and subsequent translation of VEGF in MDA-MB-435/β4 and MDA-MB-231 cancer cells. m7GTP-Sepharose–binding assay revealed that Src activity is required to form eIF4F complex which is necessary for Cap-dependent translation in α6β4 expressing human cancer cells. Conclusions: Overall, our studies suggest that integrin β4 and c-Src activation is important early signaling events to lead mTOR activation and cap-dependent translation of VEGF.
- Functional and differential proteomic analyses to identify platelet derived factors affecting ex vivo expansion of mesenchymal stromal cells
Background: Multilineage differentiation, immunomodulation and secretion of trophic factors render mesenchymal stromal cells (MSC) highly attractive for clinical application. Human platelet derivatives such as pooled human platelet lysate (pHPL) and thrombin-activated platelet releasate in plasma (tPRP) have been introduced as alternatives to fetal bovine serum (FBS) to achieve GMP-compliance. However, whereas both pHPL and tPRP support similar proliferation kinetics of lipoaspirate-derived MSC (LA-MSC), only pHPL significantly accelerates bone marrow-derived MSC (BM-MSC) expansion. To identify functionally bioactive factors affecting ex vivo MSC expansion, a differential proteomic approach was performed and identified candidate proteins were evaluated within a bioassay. Results: Two dimensional difference gel electrophoresis (2D-DIGE), MALDI-TOF analyses and complementary Western blotting revealed 20 differential protein species. 14 candidate proteins occured at higher concentrations in pHPL compared to tPRP and 6 at higher concentrations in tPRP. The candidate proteins fibrinogen and apolipoprotein A1 differentially affected LA- and BM-MSC proliferation.In a second set of experiments, reference cytokines known to foster proliferation in FBS were tested for their effects in the human supplements. Interestingly although these cytokines promoted proliferation in FBS, they failed to do so when added to the humanized system. Conclusions: The differential proteomic approach identified novel platelet derived factors differentially acting on human MSC proliferation. Complementary testing of reference cytokines revealed a lack of stimulation in the human supplements compared to FBS. The data describe a new coherent approach to combine proteomic technologies with functional testing to develop novel, humanized, GMP-compliant conditions for MSC expansion.
- New insights in osteogenic differentiation revealed by mass spectrometric assessment of phosphorylated substrates in murine skin mesenchymal cells
Background: Bone fractures and loss represent significant costs for the public health system and often affect the patients quality of life, therefore, understanding the molecular basis for bone regeneration is essential. Cytokines, such as IL-6, IL-10 and TNFα, secreted by inflammatory cells at the lesion site, at the very beginning of the repair process, act as chemotactic factors for mesenchymal stem cells, which proliferate and differentiate into osteoblasts through the autocrine and paracrine action of bone morphogenetic proteins (BMPs), mainly BMP-2. Although it is known that BMP-2 binds to ActRI/BMPR and activates the SMAD 1/5/8 downstream effectors, little is known about the intracellular mechanisms participating in osteoblastic differentiation. We assessed differences in the phosphorylation status of different cellular proteins upon BMP-2 osteogenic induction of isolated murine skin mesenchymal stem cells using Triplex Stable Isotope Dimethyl Labeling coupled with LC/MS. Results: From 150 μg of starting material, 2,264 proteins were identified and quantified at five different time points, 235 of which are differentially phosphorylated. Kinase motif analysis showed that several substrates display phosphorylation sites for Casein Kinase, p38, CDK and JNK. Gene ontology analysis showed an increase in biological processes related with signaling and differentiation at early time points after BMP2 induction. Moreover, proteins involved in cytoskeleton rearrangement, Wnt and Ras pathways were found to be differentially phosphorylated during all timepoints studied. Conclusions: Taken together, these data, allow new insights on the intracellular substrates which are phosphorylated early on during differentiation to BMP2-driven osteoblastic differentiation of skin-derived mesenchymal stem cells.
Genome Biology - Latest Articles
The latest research articles published by Genome Biology
- Learning to swim in a sea of genomic data
A report on the 63rd American Society of Human Genetics (ASHG) meeting held in Boston, USA, 22–26 October 2013.
- Transcriptome and proteome quantification of a tumor model provides novel insights into post-transcriptional gene regulation
Background: Genome-wide transcriptome analyses have allowed for systems-level insights into gene regulatory networks. Due to the limited depth of quantitative proteomics, however, our understanding of post-transcriptional gene regulation and its effects on protein complex stoichiometry are lagging behind. Results: Here, we employ deep sequencing and iTRAQ technology to determine transcript and protein expression changes of a Drosophila brain tumor model at near genome-wide resolution. In total, we quantify more than 6,200 tissue-specific proteins, corresponding to about 70% of all transcribed protein-coding genes. Using our integrated data set, we demonstrate that post-transcriptional gene regulation varies considerably with biological function and is surprisingly high for genes regulating transcription. We combine our quantitative data with protein-protein interaction data and show that post-transcriptional mechanisms significantly enhance co-regulation of protein complex subunits beyond transcriptional co-regulation. Interestingly, our results suggest that only about 11% of the annotated Drosophila protein complexes are co-regulated in the brain. Finally, we refine the composition of some of these core protein complexes by analyzing the co-regulation of potential subunits. Conclusions: Our comprehensive transcriptome and proteome data provide a valuable resource for quantitative biology and offer novel insights into understanding post-transcriptional gene regulation in a tumor model.
- Epidermal stem cells ride the circadian wave
An intriguing study shows that, in epidermal progenitor cells, circadian genes are expressed in successive waves that modulate responses to differentiation signals.
- Characterizing 5-methylcytosine in the mammalian epitranscriptome
The post-transcriptional modification 5-methylcytosine (m5C) occurs in a wide range of coding and non-coding RNAs. We describe transcriptome-wide approaches to capture the global m5C RNA methylome. We also discuss the potential functions of m5C in RNA and compare them to 6-methyladenosine modifications.
- Chromatin signatures at transcriptional start sites separate two equally populated yet distinct classes of intergenic long noncoding RNAs
Background: Mammalian transcriptomes contain thousands of long noncoding RNAs (lncRNAs). Some lncRNAs originate from intragenic enhancers which, when active, behave as alternative promoters producing transcripts that are processed using the canonical signals of their host gene. We have followed up this observation by analyzing intergenic lncRNAs to determine the extent to which they might also originate from intergenic enhancers. Results: We integrated high-resolution maps of transcriptional initiation and transcription to annotate a conservative set of intergenic lncRNAs expressed in mouse erythroblasts. We subclassified intergenic lncRNAs according to chromatin status at transcriptional initiation regions, defined by relative levels of histone H3K4 mono- and trimethylation. These transcripts are almost evenly divided between those arising from enhancer-associated (elncRNA) or promoter-associated (plncRNA) elements. These two classes of 5[prime] capped and polyadenylated RNA transcripts are indistinguishable with regard to their length, number of exons or transcriptional orientation relative to their closest neighbouring gene. Nevertheless, elncRNAs are more tissue-restricted, less highly expressed and less well conserved during evolution. Of considerable interest, we found that expression of elncRNAs, but not plncRNAs, is associated with enhanced expression of neighboring protein-coding genes during erythropoiesis. Conclusions: We have determined globally the sites of initiation of intergenic lncRNAs in erythroid cells, allowing us to distinguish two similarly abundant classes of transcripts. Different correlations between the levels of elncRNAs, plncRNAs and expression of neighboring genes suggest that functional lncRNAs from the two classes may play contrasting roles in regulating the transcript abundance of local or distal loci.
- Single-cell genomics reveal low recombination frequencies in freshwater bacteria of the SAR11 clade
Background: The SAR11 group of Alphaproteobacteria is highly abundant in the oceans. It contains a recently diverged freshwater clade, which offers the opportunity to compare adaptations to salt- and freshwaters in a monophyletic bacterial group. However, there are no cultivated members of the freshwater SAR11 group and no genomes have been sequenced yet. Results: We isolated ten single SAR11 cells from three freshwater lakes and sequenced and assembled their genomes. A phylogeny based on 57 proteins indicates that the cells are organized into distinct microclusters. We show that the freshwater genomes have evolved primarily by the accumulation of nucleotide substitutions and that they have among the lowest ratio of recombination to mutation estimated for bacteria. In contrast, members of the marine SAR11 clade have one of the highest ratios. Additional metagenome reads from six lakes confirm low recombination frequencies for the genome overall and reveal lake-specific variations in microcluster abundances. We identify hypervariable regions with gene contents broadly similar to those in the hypervariable regions of the marine isolates, containing genes putatively coding for cell surface molecules. Conclusions: We conclude that recombination rates differ dramatically in phylogenetic sister groups of the SAR11 clade adapted to freshwater and marine ecosystems. The results suggest that the transition from marine to freshwater systems has purged diversity and resulted in reduced opportunities for recombination with divergent members of the clade. The low recombination frequencies of the LD12 clade resemble the low genetic divergence of host-restricted pathogens that have recently shifted to a new host.
- Rumors of the death of consumer genomics are greatly exaggerated
David Mittelman and Razib Khan argue that DTC genetics, such as 23andme, has an enduring role to play in our society
- Characterization of chromosomal architecture in Arabidopsis by chromosome conformation capture
Background: The packaging of long chromatin fibres in the nucleus poses a major challenge, as it must fulfil both physical and functional requirements. Until recently, insight into the chromosomal architecture of plants was mainly provided by cytogenetic studies. Complementary to these analyses, chromosome conformation capture technologies promise to refine and improve our view on chromosomal architecture and to provide a more generalized description of nuclear organization. Results: Employing circular chromosome conformation capture, this study describes chromosomal architecture in Arabidopsis nuclei from a genome-wide perspective. Surprisingly, the linear organisation of chromosomes is reflected in the genome-wide interactome. In addition, we study the interplay of the interactome and epigenetic marks and report that the heterochromatic knob on the short arm of chromosome 4 (hk4s) maintains a pericentromere-like interaction profile and interactome despite its euchromatic surrounding. Conclusion: Despite the extreme condensation that is necessary to pack the chromosomes into the nucleus, the Arabidopsis genome appears to be packed in a predictive manner, according to the following criteria: heterochromatin and euchromatin represent two distinct interactomes; interactions between chromosomes correlate with the linear position on the chromosome arm; and distal chromosome regions have a higher potential to interact with other chromosomes.
- The non-coding genome: a universe in expansion for fine-tuning the coding world
A report on the EMBO/EMBL Symposium on The Non-Coding Genome, held in Heidelberg, Germany, 9-12 October, 2013.
- Genomic analysis reveals key aspects of prokaryotic symbiosis in the phototrophic consortium 'Chlorochromatium aggregatum'
Background: 'Chlorochromatium aggregatum' is a phototrophic consortium, a symbiosis that may represent the highest degree of mutual interdependence between two unrelated bacteria not associated with a eukaryotic host. 'Chlorochromatium aggregatum' is a motile, barrel-shaped aggregate formed from a single cell of "Candidatus Symbiobacter mobilis," a polarly flagellated, non-pigmented, heterotrophic bacterium, which is surrounded by approximately 15 epibiont cells of Chlorobium chlorochromatii, a non-motile photolithoautotrophic green sulfur bacterium. Results: We analyzed the complete genome sequences of both organisms to understand the basis for this symbiosis. Chl. chlorochromatii has acquired relatively few symbiosis-specific genes; most acquired genes are predicted to modify the cell wall or function in cell-cell adhesion. In striking contrast, "Ca. S. mobilis" appears to have undergone massive gene loss, is probably no longer capable of independent growth, and thus may only reproduce when consortia divide. A detailed model for the energetic and metabolic bases of the dependency of "Ca. S. mobilis" on Chl. chlorochromatii is described. Conclusions: Genomic analyses suggest that three types of interactions lead to a highly sophisticated relationship between these two organisms. Firstly, extensive metabolic exchange, involving carbon, nitrogen, and sulfur sources as well as vitamins, occurs from the epibiont to the central bacterium. Secondly, "Ca. S. mobilis" can sense and move towards light and sulfide, resources that only directly benefit the epibiont. Thirdly, electron cycling mechanisms, particularly those mediated by quinones and potentially involving shared protonmotive force, could provide an important basis for energy exchange in this and other symbiotic relationships.
Malaria Journal - Latest Articles
The latest research articles published by Malaria Journal
- Serum lipids and lipoproteins in malaria - a systematic review and meta-analysis
Background: Serum lipid profile changes have been observed during malaria infection. The underlying biological mechanisms remain unclear. The aim of this paper is to provide an overview on those serum lipid profile changes, and to discuss possible underlying biological mechanisms and the role of lipids in malaria pathogenesis. Methods: A systematic review and meta-analysis to determine lipid profile changes during malaria was conducted, following PRISMA guidelines. Without language restrictions, Medline/PubMed, Embase, Cochrane Central Register of Controlled Trials, Web of Science, LILACS, Biosis Previews and the African Index Medicus were searched for studies published up to 11 July, 2013, that measured serum lipid parameters in malaria patients. Also, major trial registries were searched. Mean differences in lipid profile parameters were combined in fixed and random effects meta-analysis, with a separate analysis for different groups of controls (healthy, other febrile illnesses or very low parasitaemia). These parameters were also compared between severe malaria and uncomplicated malaria. Funnel plots were used to test for publication bias. Results: Of 2,518 studies reviewed, 42 met the criteria for inclusion in the qualitative analysis, and of these, 15 reported the necessary data for inclusion in the meta-analysis for cholesterol; nine for high-density lipoprotein (HDL), eight for low-density lipoprotein (LDL), and nine for triglycerides, respectively. Total cholesterol, HDL and LDL concentrations were lower in malaria and other febrile diseases compared to healthy controls. The decline was more pronounced and statistically significant during malaria compared to other febrile diseases. These results were consistent across included studies. Triglycerides were raised compared to healthy controls, but not statistically significant when compared to symptomatic controls. Conclusions: This meta-analysis suggests that the observed lipid profile changes are characteristic for malaria. Although a definite link with the pathogenesis of malaria cannot yet be demonstrated, plausible hypotheses of biological mechanisms involving host lipid alterations and the pathogenesis of malaria exist. An increased research effort to elucidate the precise pathways is warranted, since this could lead to better understanding of malaria pathophysiology and consequently to novel treatment approaches.
- Changes in drug sensitivity and anti-malarial drug resistance mutations over time among Plasmodium falciparum parasites in Senegal
Background: Malaria treatment efforts are hindered by the rapid emergence and spread of drug resistant parasites. Simple assays to monitor parasite drug response in direct patient samples (ex vivo) can detect drug resistance before it becomes clinically apparent, and can inform changes in treatment policy to prevent the spread of resistance. Methods: Parasite drug responses to amodiaquine, artemisinin, chloroquine and mefloquine were tested in approximately 400 Plasmodium falciparum malaria infections in Thies, Senegal between 2008 and 2011 using a DAPI-based ex vivo drug resistance assay. Drug resistance-associated mutations were also genotyped in pfcrt and pfmdr1. Results: Parasite drug responses changed between 2008 and 2011, as parasites became less sensitive to amodiaquine, artemisinin and chloroquine over time. The prevalence of known resistance-associated mutations also changed over time. Decreased amodiaquine sensitivity was associated with sustained, highly prevalent mutations in pfcrt, and one mutation in pfmdr1 -- Y184F -- was associated with decreased parasite sensitivity to artemisinin. Conclusions: Directly measuring ex vivo parasite drug response and resistance mutation genotyping over time are useful tools for monitoring parasite drug responses in field samples. Furthermore, these data suggest that the use of amodiaquine and artemisinin derivatives in combination therapies is selecting for increased drug tolerance within this population.
- Mosquito transmission, growth phenotypes and the virulence of malaria parasites
Background: A series of elegant experiments was recently published which demonstrated that transmission of malaria parasites through mosquitoes elicited an attenuated growth phenotype, whereby infections grew more slowly and reached peak parasitaemia at least five-fold lower than parasites which had not been mosquito transmitted. To assess the implications of these results it is essential to understand whether the attenuated infection phenotype is a general phenomenon across parasites genotypes and conditions. Methods: Using previously published data, the impact of mosquito transmission on parasite growth rates and virulence of six Plasmodium chabaudi lines was analysed. Results: The effect of mosquito transmission varied among strains, but did not lead to pronounced or consistent reductions in parasite growth rate. Conclusions: Mosquito-induced attenuated growth phenotype is sensitive to experimental conditions.
- A case report of transfusion-transmitted Plasmodium malariae from an asymptomatic non-immune traveller
Background: The incidence of transfusion-transmitted malaria is very low in non-endemic countries due to strict donor selection. The optimal strategy to mitigate the risk of transfusion-transmitted malaria in non-endemic countries without unnecessary exclusion of blood donations is, however, still debated and asymptomatic carriers of Plasmodium species may still be qualified to donate blood for transfusion purposes.Case description: In April 2011, a 59-year-old Dutch woman with spiking fevers for four days was diagnosed with a Plasmodium malariae infection. The patient had never been abroad, but nine weeks before, she had received red blood cell transfusion for anaemia. The presumptive diagnosis of transfusion-transmitted quartan malaria was made and subsequently confirmed by retrospective PCR analysis of donor blood samples. The donor was a 36-year-old Dutch male who started donating blood in May 2006. His travel history outside Europe included a trip to Kenya, Tanzania and Zanzibar in 2005, to Thailand in 2006 and to Costa Rica in 2007. He only used malaria prophylaxis during his travel to Africa. The donor did not show any abnormalities upon physical examination in 2011, while laboratory examination demonstrated a thrombocytopenia of 126 x 109/L as the sole abnormal finding since 2007. Thick blood smear analysis and the Plasmodium PCR confirmed an ongoing subclinical P. malariae infection. Chloroquine therapy was started, after which the infection cleared and thrombocyte count normalized. Fourteen other recipients who received red blood cells from the involved donor were traced. None of them developed malaria symptoms.DiscussionThis case demonstrates that P. malariae infections in non-immune travellers may occur without symptoms and persist subclinically for years. In addition, this case shows that these infections pose a threat to transfusion safety when subclinically infected persons donate blood after their return in a non-endemic malaria region.Since thrombocytopenia was the only abnormality associated with the subclinical malaria infection in the donor, this case illustrates that an unexplained low platelet count after a visit to malaria-endemic countries may be an indicator for asymptomatic malaria even when caused by non-falciparum Plasmodium species.
- Evaluation of Plasmodium falciparum gametocyte detection in different patient material
Background: For future eradication strategies of malaria it is important to control the transmission of gametocytes from humans to the anopheline vector which causes the spread of the disease. Sensitive, non-invasive methods to detect gametocytes under field conditions can play a role in monitoring transmission potential. Methods: Microscopically Plasmodium falciparum-positive patients from Jimma, Ethiopia donated finger-prick blood, venous blood, saliva, oral mucosa and urine samples that were spotted on filter paper or swabs. All samples were taken and stored under equal, standardized conditions. RNA was extracted from the filter paper and detected by real-time QT-NASBA. Pfs16-mRNA and Pfs25-mRNA were measured with a time to positivity to detect gametocyte specific mRNA in different gametocyte stages. They were compared to 18S-rRNA, which is expressed in all parasite stages. Results were quantified via a known dilution series of artificial RNA copies. Results: Ninety-six samples of 16 uncomplicated malaria patients were investigated. 10 (66.7%) of the slides showed gametocyte densities between 0.3-2.9 gametocytes/mul. For all RNA-targets, molecular detection in blood samples was most sensitive; finger-prick sampling required significantly smaller amounts of blood than venous blood collection. Detection of asexual 18S-rRNA in saliva and urine showed sensitivities of 80 and 67%, respectively. Non-invasive methods to count gametocytes proved insensitive. Pfs16-mRNA was detectable in 20% of urine samples, sensitivities for other materials were lower. Pfs25-mRNA was not detectable in any sample. Conclusions: The sensitivity of non-invasively collected material such as urine, saliva or mucosa seems unsuitable for the detection of gametocyte-specific mRNA. Sensitivity in asymptomatic carriers might be generally even lower. Finger-prick testing revealed the highest absolute count of RNA copies per muL, especially for Pfs25-mRNA copies. The method proved to be the most effective and should preferably be applied in future transmission control and eradication plans. A rapid test for gametocyte targets would simplify efforts.
- The changing burden of malaria and association with vector control interventions in Zambia using district-level surveillance data, 2006--2011
Background: Malaria control was strengthened in Zambia over the past decade. The two primary interventions for vector control are indoor residual spraying (IRS) and long-lasting insecticide-treated nets (LLINs). Using passive malaria surveillance data collected from 2006 to 2011 through the Zambian District Health Information System, the associations between increased coverage with LLINs and IRS and the burden of malaria in Zambia was evaluated. Methods: National passive malaria surveillance data from 2006 to 2011 were analysed. A district-level, random-effects model with Poisson regression was used to explore the association between malaria cases and coverage with LLINs and IRS. Malaria cases and LLINs and IRS coverage were mapped to visualize spatiotemporal variation in malaria for each year. Results: From 2006--2011, 24.6 million LLINs were distributed and 6.4 million houses were sprayed with insecticide. Coverage with LLINs was not uniformly distributed over the study period and IRS was targeted to central and southern districts where malaria transmission was low. LLIN coverage was associated with a reduction in malaria cases, although an increase in the number of malaria cases was reported in some districts over the study period. A high burden of malaria persisted in north-eastern Zambia, whereas a reduction in the number of reported malaria cases was observed in western and southern Zambia. Conclusion: Enhanced and targeted interventions in north-eastern Zambia where the burden of malaria remains high, as well as efforts to sustain low malaria transmission in the south-west, will be necessary for Zambia to achieve the national goal of being malaria free by 2030.
- Qualitative study of presumptive treatment of childhood malaria in third tier tertiary hospitals in southeast Nigeria: a focus group and in-depth study
Background: Presumptive treatment of childhood-malaria (PTCM) is common in Nigeria. Delayed laboratory result is blamed, with little attention on patients’ and providers’ roles. This study aimed to determine patient, provider and laboratory attributes that sustain PTCM in Nigeria. Methods: Data collection was from focus-group discussions for parents/guardians, and in-depth interviews involving providers and laboratory scientists in two tertiary hospitals. Results: All parents/guardians agreed to a malaria test. Majority accepted to come back later for full treatment, provided that some treatment was commenced. Majority affirmed that their interests are on their children’s improvement.The providers practice presumptive treatment of childhood malaria, for the following reasons: (1) malaria is endemic and should be suspected and treated; (2) microscopy takes two days to be available and parents want immediate treatment for their children, thus delay may lead to self-medication; (3) relying on results for decision to treat creates an impression of incompetence; (4) rapid diagnostic test kits (RDTs) are not available in the consulting rooms and there is doubt about their reliability; (5) patients have already wasted time before being reviewed, so wasting more time on investigation is not advisable; (6) withhold of malaria treatment may be feasible in suspected uncomplicated malaria, but if severe, then anti-malarial treatment has to start immediately.Interviews of laboratory scientists showed that (1) malaria microscopy test cannot be urgent; it is done in batches and takes 24 hours to be ready; (2) a request of malaria test with other investigations on the same form, contributes to the delay; (3) RDTs are unavailable in the facilities. Conclusions: Provision of RDTs is the only feasible means to treatment of confirmed malaria at the time healthcare providers review a patient on day zero. In facilities that depend on microscopy; a common practice in resource poor countries, healthcare providers can depend on parental willingness to return later for full medication, to commence adjunctive care with antipyretics and multivitamins for uncomplicated malaria. In complicated malaria, supportive care - intravenous fluids, blood transfusion, oxygen therapy - can be commenced while awaiting the inclusion of anti-malarial drugs when the diagnosis of malaria is confirmed.
- Haemoglobin levels for population from Gambo, a rural area of Ethiopia, and their association with anaemia and malaria
Background: Knowledge of appropriate reference intervals is critical not only to provide optimal clinical care, but also to enrol populations in medical research. The aim of this study was to generate normal ranges of laboratory values for haemoglobin among healthy Ethiopian adults and children and to determine if anaemia is a possible indicator of malaria in women and children in this area of Ethiopia. Methods: This study was carried out from January to May 2010. The reference sample population with malaria-negative consisted of 454 individuals, divided women, men and children. The malaria-infected sample population consisted of 117 individuals. The reference ranges were based on the guidelines from the Clinical and Laboratory Standards Institute. Haemoglobin concentration was determined by Hemo-Control EKF Diagnostic Analyser on whole blood. Testing for malaria-positive and negative infection was done by microscopy and by PCR. Results: The lower limits for adult haemoglobin range obtained from this population were slightly higher than those derived from other African populations, but were equal to those established by other studies in Ethiopia and the World Health Organization (WHO). Regarding children, the minimum values were lower than those obtained from different African populations and those established by WHO. The malaria-negative group had anaemia in 35.6% of cases and in the malaria-positive group in 70.9%. There was a stronger, statistically significant association between anaemia and malaria-positive samples than between anaemia and malaria-negative samples in women and both groups of children. Conclusions: The results from this study are a contribution in the definition of the haemoglobin parameters in African populations, which could be taken as standards for interpretation of laboratory results. The haemoglobin indices in adults from Gambo tended to be higher than other African populations and in children were lower than other studies in Africa. The results also suggest that anaemia is not useful as a supportive diagnostic criterion to monitor and evaluate malaria in women and children from Ethiopia, because a 29.1% of malaria cases will be not detected, because of not having anaemia.
- Reticulocyte-prone malaria parasites predominantly invade CD71hi immature cells: implications for the development of an in vitro culture for Plasmodium vivax
Background: The lack of a continuous in vitro culture system for blood stages of malarial parasites with a unique tropism for reticulocytes, such as Plasmodium vivax and the Plasmodium yoelii 17X reticulocyte-prone strain, hinders research in these organisms. The maturation of reticulocytes into erythrocytes is a complex process involving the selective removal of membrane proteins such as the transferrin receptor, CD71. In order to advance in the characterization of infected cells during experimental infections of BALB/c mice with P. yoelii 17X, CD71 expression in erythroid cells (TER119+) was assessed and in vitro culture of P. yoelii 17X was attempted by adding reticulocytes highly expressing CD71. Methods: BALB/c mice were infected with P. yoelii 17X-GFP transgenic parasites and erythroid cells (TER119+) were analysed in blood, spleen and bone marrow cells. TER119, CD71 and GFP expression was assessed at different points post-infection by flow cytometry. Moreover, in vitro culture of P. yoelli 17X was attempted by adding red blood cells (RBCs) from mice with a pyruvate kinase deficiency, which contain high percentages of CD71hi cells in peripheral blood as compared to healthy animals. Results: A predominance of erythroid cells lacking expression of CD71 (CD71-) was observed in peripheral blood and spleen in normal and infected animals up to ten days post-infection (pi). At ten days pi, however, a dramatic temporal switch to erythroid cells highly expressing CD71 (CD71hi) was observed in the spleen and at day 15 pi in peripheral blood of the infected cells. A distribution of erythroid cells expressing differently CD71 was noticed in the bone marrow. Yet, similar to peripheral blood and spleen, a predominance of CD71hi cells was observed at 15 days pi. Remarkably, CD71hi cells were the cells predominantly infected in these organs as well as in peripheral blood. Attempts were thus made to culture in vitro the P. yoelli 17X strain by adding RBCs from pyruvate kinase-deficient mice containing high percentages of CD71hi cells in peripheral blood. Conclusions: The parasite preference for immature cells that are rare in normal peripheral blood could have important implications for the development of an in vitro culture system for P. vivax.
- Malaria case management in Papua New Guinea following the introduction of a revised treatment protocol
Background: This paper reports on the availability of diagnostic tools and recommended anti-malarials in the 12-month period immediately following the implementation of a new national malaria treatment protocol (NMTP) in Papua New Guinea (PNG). Health worker adherence to the new NMTP is also examined and comparisons made with previously reported pre-implementation findings. Methods: A countrywide cross-sectional survey in randomly selected primary health care facilities (n = 88). Data were collected via passive observation of the clinical case management of fever or suspected malaria patients and via an interviewer administered questionnaire completed with the officer in charge of each participating health care facility. Results: Malaria rapid diagnostic tests (RDTs) and the new first-line anti-malarial medication, artemether-lumefantrine (AL), were available in 53.4% and 51.1% of surveyed heath facilities, respectively. However, they were more widely available in the larger health centres as compared to the smaller aid-posts (90.2% vs. 21.3% and 87.8% vs. 19.2%, respectively). Overall, 68.3% of observed fever cases (n = 445) were tested for malaria by RDT and 39% prescribed an anti-malarial, inclusive of 98.2% of RDT positive patients and 19.8% of RDT negative cases. The availability and use of malaria RDTs was greater in the current survey as compared to pre-implementation of the new NMTP (8.9% vs. 53.4% & 16.2% vs. 68.3%, respectively) as was the availability of AL (0% vs. 51.1%). The percentage of fever patients prescribed anti-malarials decreased substantially post implementation of the new NMTP (96.4% vs. 39.0%). Conclusions: PNG has achieved high coverage of malaria RDTs and AL at the health centre level, but these resources have yet to reach the majority of aid-posts. Malaria case management practice has substantially changed in the 12-month period immediately following the new NMTP, although full protocol adherence was rarely observed.
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Comprehensive Transcriptome Profiling in Tomato Reveals a Role for Glycosyltransferase in Mi-Mediated Nematode Resistance Plant Physiology 144:1079-1092 (2007)
Jennifer E. Schaff, Dahlia M. Nielsen, Chris P. Smith, Elizabeth H. Scholl and David McK. Bird
Phylogenetic Analysis of Pasteuria penetrans by Use of Multiple Genetic Loci Journal of Bacteriology, August 2005, p. 5700-5708, Vol. 187, No. 16
Lauren Charles, Ignazio Carbone, Keith G. Davies, David Bird, Mark Burke, Brian R. Kerry, and Charles H. Opperman
Root-knot nematodes and bacterial Nod factors elicit common signal transduction events in Lotus japonicus PNAS | February 22, 2005 | vol. 102 | no. 8 | 3147-3152
Ravisha R. Weerasinghe, David McK. Bird and Nina S. Allen
Virulence Genes in Heterodera glycines: Allele Frequencies and Ror Gene Groups Among Field Isolates and Inbred Lines (pdf file)
Phytopathology | February 2005 | Volume 95, Number 2
K. Dong, K. R. Barker, and C. H. Opperman. Pages 186-191.
Lotus japonicus: A New Model to Study Root-Parasitic Nematodes
Dasharath Prasad Lohar and David McK. Bird
Plant and Cell Physiology, 2003, Vol. 44, No. 11 1176-1184
Horizontally transferred genes in plant-parasitic nematodes: a high-throughput genomic approach
Elizabeth H Scholl, Jeffrey L Thorne, James P McCarter, and David Mck Bird
Genome Biology 2003, Vol. 4, Issue 6
Cytokinins play opposite roles in lateral root formation, and nematode and Rhizobial symbioses
Lohar, Dasharath Prasad, Schaff, Jennifer E., Laskey, James G., Kieber, Joseph J., Bilyeu, Kristin D., Bird, David McK.
The Plant Journal 38 (2), 203-214. (2004)
Model systems in agriculture: Lessons from worms.
Bird, David McK.
Annals of Applied Biology 146 (2), 147-154. (2005)
Resolving tylenchid evolutionary relationships through multiple gene analysis derived from EST data
Elizabeth H. Scholl and David McK. Bird
In Press, Corrected Proof, Available online 3 May 2005
Plant Parasitic Nematodes: Habitats, Hormones, and Horizontally-Acquired Genes
David McKenzie Bird, Hinanit Koltai
Journal of Plant Growth Regulation, Volume 19, Issue 2, Jun 2000, Pages 183 - 194
Alkahest NuclearBLAST : a user-friendly BLAST management and analysis system
Diener SE, Houfek TD, Kalat SE, Windham DE, Burke M, Opperman C, Dean RA
BMC Bioinformatics 2005, 6:147 (15 June 2005)
Wormbase
Nematode.Net
DNA Analysis
Entrez
Blaxter Lab
C. Elegans WWW server
Agriculture Network Information Center
Genome Research Laboratory
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