A comprehensive evaluation of SAM, the SAM R-package and a simple modification to improve its performance

<p>Abstract</p> <p>Background</p> <p>The Significance Analysis of Microarrays (SAM) is a popular method for detecting significantly expressed genes and controlling the false discovery rate (FDR). Recently, it has been reported in the literature that the FDR is not well controlled by SAM. Due to the vast application of SAM in microarray data analysis, it is of great importance to have an extensive evaluation of SAM and its associated R-package (sam2.20).</p> <p>Results</p> <p>Our study has identified several discrepancies between SAM and sam2.20. One major difference is that SAM and sam2.20 use different methods for estimating FDR. Such discrepancies may cause confusion among the researchers who are using SAM or are developing the SAM-like methods. We have also shown that SAM provides no meaningful estimates of FDR and this problem has been corrected in sam2.20 by using a different formula for estimating FDR. However, we have found that, even with the improvement sam2.20 has made over SAM, sam2.20 may still produce erroneous and even conflicting results under certain situations. Using an example, we show that the problem of sam2.20 is caused by its use of asymmetric cutoffs which are due to the large variability of null scores at both ends of the order statistics. An obvious approach without the complication of the order statistics is the conventional symmetric cutoff method. For this reason, we have carried out extensive simulations to compare the performance of sam2.20 and the symmetric cutoff method. Finally, a simple modification is proposed to improve the FDR estimation of sam2.20 and the symmetric cutoff method.</p> <p>Conclusion</p> <p>Our study shows that the most serious drawback of SAM is its poor estimation of FDR. Although this drawback has been corrected in sam2.20, the control of FDR by sam2.20 is still not satisfactory. The comparison between sam2.20 and the symmetric cutoff method reveals that the relative performance of sam2.20 to the symmetric cutff method depends on the ratio of induced to repressed genes in a microarray data, and is also affected by the ratio of DE to EE genes and the distributions of induced and repressed genes. Numerical simulations show that the symmetric cutoff method has the biggest advantage over sam2.20 when there are equal number of induced and repressed genes (i.e., the ratio of induced to repressed genes is 1). As the ratio of induced to repressed genes moves away from 1, the advantage of the symmetric cutoff method to sam2.20 is gradually diminishing until eventually sam2.20 becomes significantly better than the symmetric cutoff method when the differentially expressed (DE) genes are either all induced or all repressed genes. Simulation results also show that our proposed simple modification provides improved control of FDR for both sam2.20 and the symmetric cutoff method.</p

Parvovirus B19 infection and severe anaemia in Kenyan children: a retrospective case control study

Background: During acute Human parvovirus B19 (B19) infection a transient reduction in blood haemoglobin concentration is induced, due to a 5-7 day cessation of red cell production. This can precipitate severe anaemia in subjects with a range of pre-existing conditions. Of the disease markers that occur during B19 infection, high IgM levels occur closest in time to the maximum reduction in haemoglobin concentration. Previous studies of the contribution of B19 to severe anaemia among young children in Africa have yielded varied results. This retrospective case/control study seeks to ascertain the proportion of severe anaemia cases precipitated by B19 among young children admitted to a Kenyan district hospital.Methods: Archival blood samples from 264 children under 6 years with severe anaemia admitted to a Kenyan District Hospital, between 1999 and 2004, and 264 matched controls, were tested for B19 IgM by Enzyme Immunosorbent Assay and 198 of these pairs were tested for B19 DNA by PCR. 536 samples were also tested for the presence of B19 IgG.Results: 7 (2.7%) cases and 0 (0%) controls had high B19 IgM levels (Optical Density > 5 x cut-off value) (McNemar's exact test p = 0.01563), indicating a significant association with severe anaemia. The majority of strongly IgM positive cases occurred in 2003.10/264 (3.7%) cases compared to 5/264 (1.9%) controls tested positive for B19 IgM. This difference was not statistically significant, odds ratio (OR) = 2.00 (CI95 [0.62, 6.06], McNemar's exact test p = 0.3018. There was no significant difference between cases and controls in the B19 IgG (35 (14.8%) vs 32 (13.6%)), OR = 1.103 (CI95 [0.66, 1.89], McNemar's exact test, p = 0.7982), or the detection of the B19 DNA (6 (3.0%) vs 5 (2.5%)), OR = 1.2 (CI95 [0.33, 4.01], McNemar's exact test p = 1).Conclusions: High B19 IgM levels were significantly associated with severe anaemia, being found only among the cases. This suggests that 7/264 (2.7%) of cases of severe anaemia in the population of children admitted to KDH were precipitated by B19. While this is a relatively small proportion, this has to be evaluated in the light of the IgG data that shows that less than 15% of children in the study were exposed to B19, a figure much lower than reported in other tropical areas

The need for an integrated approach for chronic disease research and care in Africa.

With the changing distribution of infectious diseases, and an increase in the burden of non-communicable diseases, low- and middle-income countries, including those in Africa, will need to expand their health care capacities to effectively respond to these epidemiological transitions. The interrelated risk factors for chronic infectious and non-communicable diseases and the need for long-term disease management, argue for combined strategies to understand their underlying causes and to design strategies for effective prevention and long-term care. Through multidisciplinary research and implementation partnerships, we advocate an integrated approach for research and healthcare for chronic diseases in Africa

Photoswitchable diacylglycerols enable optical control of protein kinase C.

Increased levels of the second messenger lipid diacylglycerol (DAG) induce downstream signaling events including the translocation of C1-domain-containing proteins toward the plasma membrane. Here, we introduce three light-sensitive DAGs, termed PhoDAGs, which feature a photoswitchable acyl chain. The PhoDAGs are inactive in the dark and promote the translocation of proteins that feature C1 domains toward the plasma membrane upon a flash of UV-A light. This effect is quickly reversed after the termination of photostimulation or by irradiation with blue light, permitting the generation of oscillation patterns. Both protein kinase C and Munc13 can thus be put under optical control. PhoDAGs control vesicle release in excitable cells, such as mouse pancreatic islets and hippocampal neurons, and modulate synaptic transmission in Caenorhabditis elegans. As such, the PhoDAGs afford an unprecedented degree of spatiotemporal control and are broadly applicable tools to study DAG signaling

Application of Equilibrium Models of Solution Hybridization to Microarray Design and Analysis

Background: The probe percent bound value, calculated using multi-state equilibrium models of solution hybridization, is shown to be useful in understanding the hybridization behavior of microarray probes having 50 nucleotides, with and without mismatches. These longer oligonucleotides are in widespread use on microarrays, but there are few controlled studies of their interactions with mismatched targets compared to 25-mer based platforms. Principal Findings: 50-mer oligonucleotides with centrally placed single, double and triple mismatches were spotted on an array. Over a range of target concentrations it was possible to discriminate binding to perfect matches and mismatches, and the type of mismatch could be predicted accurately in the concentration midrange (100 pM to 200 pM) using solution hybridization modeling methods. These results have implications for microarray design, optimization and analysis methods. Conclusions: Our results highlight the importance of incorporating biophysical factors in both the design and the analysis of microarrays. Use of the probe ‘‘percent bound’ ’ value predicted by equilibrium models of hybridization is confirmed to be important for predicting and interpreting the behavior of long oligonucleotide arrays, as has been shown for shor

The Regulation of Sulfur Metabolism in Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) has evolved into a highly successful human pathogen. It deftly subverts the bactericidal mechanisms of alveolar macrophages, ultimately inducing granuloma formation and establishing long-term residence in the host. These hallmarks of Mtb infection are facilitated by the metabolic adaptation of the pathogen to its surrounding environment and the biosynthesis of molecules that mediate its interactions with host immune cells. The sulfate assimilation pathway of Mtb produces a number of sulfur-containing metabolites with important contributions to pathogenesis and survival. This pathway is regulated by diverse environmental cues and regulatory proteins that mediate sulfur transactions in the cell. Here, we discuss the transcriptional and biochemical mechanisms of sulfur metabolism regulation in Mtb and potential small molecule regulators of the sulfate assimilation pathway that are collectively poised to aid this intracellular pathogen in its expert manipulation of the host. From this global analysis, we have identified a subset of sulfur-metabolizing enzymes that are sensitive to multiple regulatory cues and may be strong candidates for therapeutic intervention

Expression profiling identifies genes involved in neoplastic transformation of serous ovarian cancer

Background: The malignant potential of serous ovarian tumors, the most common ovarian tumor subtype, varies from benign to low malignant potential (LMP) tumors to frankly invasive cancers. Given the uncertainty about the relationship between these different forms, we compared their patterns of gene expression. Methods: Expression profiling was carried out on samples of 7 benign, 7 LMP and 28 invasive (moderate and poorly differentiated) serous tumors and four whole normal ovaries using oligonucleotide microarrays representing over 21,000 genes. Results: We identified 311 transcripts that distinguished invasive from benign tumors, and 20 transcripts that were significantly differentially expressed between invasive and LMP tumors at p < 0.01 (with multiple testing correction). Five genes that were differentially expressed between invasive and either benign or normal tissues were validated by real time PCR in an independent panel of 46 serous tumors (4 benign, 7 LMP, 35 invasive). Overexpression of SLPI and WNT7A and down-regulation of C6orf31, PDGFRA and GLTSCR2 were measured in invasive and LMP compared with benign and normal tissues. Over-expression of WNT7A in an ovarian cancer cell line led to increased migration and invasive capacity. Conclusion: These results highlight several genes that may play an important role across the spectrum of serous ovarian tumorigenesis