RET mutational spectrum in Hirschsprung disease: evaluation of 601 Chinese patients

Rare (RVs) and common variants of the RET gene contribute to Hirschsprung disease (HSCR; congenital aganglionosis). While RET common variants are strongly associated with the commonest manifestation of the disease (males; short-segment aganglionosis; sporadic), rare coding sequence (CDS) variants are more frequently found in the lesser common and more severe forms of the disease (females; long/total colonic aganglionosis; familial).Here we present the screening for RVs in the RET CDS and intron/exon boundaries of 601 Chinese HSCR patients, the largest number of patients ever reported. We identified 61 different heterozygous RVs (50 novel) distributed among 100 patients (16.64%). Those include 14 silent, 29 missense, 5 nonsense, 4 frame-shifts, and one in-frame amino-acid deletion in the CDS, two splice-site deletions, 4 nucleotide substitutions and a 22-bp deletion in the intron/exon boundaries and 1 single-nucleotide substitution in the 5' untranslated region. Exonic variants were mainly clustered in RET the extracellular domain. RET RVs were more frequent among patients with the most severe phenotype (24% vs. 15% in short-HSCR). Phasing RVs with the RET HSCR-associated haplotype suggests that RVs do not underlie the undisputable association of RET common variants with HSCR. None of the variants were found in 250 Chinese controls.     

Accurate genome relative abundance estimation based on shotgun metagenomic reads

Accurate estimation of microbial community composition based on metagenomic sequencing data is fundamental for subsequent metagenomics analysis. Prevalent estimation methods are mainly based on directly summarizing alignment results or its variants; often result in biased and/or unstable estimates. We have developed a unified probabilistic framework (named GRAMMy) by explicitly modeling read assignment ambiguities, genome size biases and read distributions along the genomes. Maximum likelihood method is employed to compute Genome Relative Abundance of microbial communities using the Mixture Model theory (GRAMMy). GRAMMy has been demonstrated to give estimates that are accurate and robust across both simulated and real read benchmark datasets. We applied GRAMMy to a collection of 34 metagenomic read sets from four metagenomics projects and identified 99 frequent species (minimally 0.5% abundant in at least 50% of the data-sets) in the human gut samples. Our results show substantial improvements over previous studies, such as adjusting the over-estimated abundance for Bacteroides species for human gut samples, by providing a new reference-based strategy for metagenomic sample comparisons. GRAMMy can be used flexibly with many read assignment tools (mapping, alignment or composition-based) even with low-sensitivity mapping results from huge short-read datasets. It will be increasingly useful as an accurate and robust tool for abundance estimation with the growing size of read sets and the expanding database of reference genomes.     

Altered connectivity pattern of hubs in default-mode network with Alzheimer's disease: an Granger causality modeling approach

Background

Evidences from normal subjects suggest that the default-mode network (DMN) has posterior cingulate cortex (PCC), medial prefrontal cortex (MPFC) and inferior parietal cortex (IPC) as its hubs; meanwhile, these DMN nodes are often found to be abnormally recruited in Alzheimer''s disease (AD) patients. The issues on how these hubs interact to each other, with the rest nodes of the DMN and the altered pattern of hubs with respect to AD, are still on going discussion for eventual final clarification.

Principal Findings

To address these issues, we investigated the causal influences between any pair of nodes within the DMN using Granger causality analysis and graph-theoretic methods on resting-state fMRI data of 12 young subjects, 16 old normal controls and 15 AD patients respectively. We found that: (1) PCC/MPFC/IPC, especially the PCC, showed the widest and distinctive causal effects on the DMN dynamics in young group; (2) the pattern of DMN hubs was abnormal in AD patients compared to old control: MPFC and IPC had obvious causal interaction disruption with other nodes; the PCC showed outstanding performance for it was the only region having causal relation with all other nodes significantly; (3) the altered relation between hubs and other DMN nodes held potential as a noninvasive biomarker of AD.

Conclusions

Our study, to the best of our knowledge, is the first to support the hub configuration of the DMN from the perspective of causal relationship, and reveal abnormal pattern of the DMN hubs in AD. Findings from young subjects provide additional evidence for the role of PCC/MPFC/IPC acting as hubs in the DMN. Compared to old control, MPFC and IPC lost their roles as hubs owing to the obvious causal interaction disruption, and PCC was preserved as the only hub showing significant causal relations with all other nodes.     

A novel genotype of GB virus C: its identification and predominance among injecting drug users in Yunnan, China

GB virus C (GBV-C) is prevalent globally and particularly among individuals at risk of parental exposures. Based on genetic diversity, this virus is now classified into six genotypes and many subtypes with distinct geographical distribution. In this study, 120 Injecting Drug Users (IDUs) were recruited from Yunnan province, China. Among them, 43 (35.8%) were positive for GBV-C RNA, 70 (58.3%) and 103 (85.8%) sero-positive for HIV-1 and HCV respectively. This revealed 18.3% of IDUs having GBV-C/HIV/HCV triple infection, which is significantly higher than 7.5% of GBV-C/HIV-1 and 10% of GBV-C/HCV dual infection rates (P<0.05). Based on 5'UTR sequences, the identified 43 viral isolates can be classified into three phylogenetic groups: one (2.3%) and two (4.7%) belonged to genotype 3 and 4, respectively, and the remaining 40 (93%) formed a new group with 97% of bootstrap support. This new GBV-C group was further confirmed by characterizing the E2 region and full-length genome sequences. Analysis of 187 nt 5'UTR sequence showed three previous reported isolates from Southeast Asia were re-classified into this new group. It implies they have the same origin with strains from Yunnan. Although we provisionally assigned this new group as GBV-C genotype 7, a simpler five groups of GBV-C nomenclature is recommended. Genotype 4, 6 and the newly designated genotype 7 could be reclassified as one group, which may represent a single GBV-C genotype. The classification of the other four groups was corresponding to that of previous reported genotype 1, 2, 3 and 5. Furthermore, the diversity of amino acid sequence in the E2 region was analyzed. The inhibitory effect of GBV-C genotype 7 on HIV-1 cell entry could be deduced. Since GBV-C may have a beneficial effect on AIDS disease progression and interact with HCV during co-infection, this finding may raise interests in future studies on this virus that was previously thought to be a "non-pathogenic virus".     

The nucleosome (histone-DNA complex) is the TLR9-specific immunostimulatory component of Plasmodium falciparum that activates DCs

The systemic clinical symptoms of Plasmodium falciparum infection such as fever and chills correspond to the proinflammatory cytokines produced in response to the parasite components released during the synchronized rupture of schizonts. We recently demonstrated that, among the schizont-released products, merozoites are the predominant components that activate dendritic cells (DCs) by TLR9-specific recognition to induce the maturation of cells and to produce proinflammatory cytokines. We also demonstrated that DNA is the active constituent and that formation of a DNA-protein complex is essential for the entry of parasite DNA into cells for recognition by TLR9. However, the nature of endogenous protein-DNA complex in the parasite is not known. In this study, we show that parasite nucleosome constitute the major protein-DNA complex involved in the activation of DCs by parasite nuclear material. The parasite components were fractionated into the nuclear and non-nuclear materials. The nuclear material was further fractionated into chromatin and the proteins loosely bound to chromatin. Polynucleosomes and oligonucleosomes were prepared from the chromatin. These were tested for their ability to activate DCs obtained by the FLT3 ligand differentiation of bone marrow cells from the wild type, and TLR2(-/-), TLR9(-/-) and MyD88(-/-) mice. DCs stimulated with the nuclear material and polynucleosomes as well as mono- and oligonucleosomes efficiently induced the production of proinflammatory cytokines in a TLR9-dependent manner, demonstrating that nucleosomes (histone-DNA complex) represent the major TLR9-specific DC-immunostimulatory component of the malaria parasite nuclear material. Thus, our data provide a significant insight into the activation of DCs by malaria parasites and have important implications for malaria vaccine development.     

Conspecific crop-weed introgression influences evolution of weedy rice (Oryza sativa f. spontanea) across a geographical range

Background

Introgression plays an important role in evolution of plant species via its influences on genetic diversity and differentiation. Outcrossing determines the level of introgression but little is known about the relationships of outcrossing rates, genetic diversity, and differentiation particularly in a weedy taxon that coexists with its conspecific crop.

Methodology/Principal Findings

Eleven weedy rice (Oryza sativa f. spontanea) populations from China were analyzed using microsatellite (SSR) fingerprints to study outcrossing rate and its relationship with genetic variability and differentiation. To estimate outcrossing, six highly polymorphic SSR loci were used to analyze >5500 progeny from 216 weedy rice families, applying a mixed mating model; to estimate genetic diversity and differentiation, 22 SSR loci were analyzed based on 301 weedy individuals. Additionally, four weed-crop shared SSR loci were used to estimate the influence of introgression from rice cultivars on weedy rice differentiation. Outcrossing rates varied significantly (0.4∼11.7%) among weedy rice populations showing relatively high overall Nei''s genetic diversity (0.635). The observed heterozygosity was significantly correlated with outcrossing rates among populations (r² = 0.783; P<0.001) although no obvious correlation between outcrossing rates and genetic diversity parameters was observed. Allelic introgression from rice cultivars to their coexisting weedy rice was detected. Weedy rice populations demonstrated considerable genetic differentiation that was correlated with their spatial distribution (r² = 0.734; P<0.001), and possibly also influenced by the introgression from rice cultivars.

Conclusions/Significance

Outcrossing rates can significantly affect heterozygosity of populations, which may shape the evolutionary potential of weedy rice. Introgression from the conspecific crop rice can influence the genetic differentiation and possibly evolution of its coexisting weedy rice populations.     

Reproducibility and relative validity of a food frequency questionnaire developed for female adolescents in Suihua, North China

Background

This study aims to evaluate the reproducibility and validity of a food frequency questionnaire (FFQ) developed for female adolescents in the Suihua area of North China. The FFQ was evaluated against the average of 24-hour dietary recalls (24-HRs).

Methodology/Principal Findings

A total of 168 female adolescents aged 12 to 18 completed nine three consecutive 24-HRs (one three consecutive 24 HRs per month) and two FFQs over nine months. The reproducibility of the FFQ was estimated using intraclass correlation coefficients (ICCs), and its relative validity was assessed by comparing it with the 24-HRs. The mean values of the 24-HRs were lower than those of the FFQs, except for protein (in FFQ1) and iron (in FFQ2). The ICCs for all nutrients and food groups in FFQ1 and FFQ2 were moderately correlated (0.4–0.8). However, all the ICCs decreased after adjusting for energy. The weighted κ statistic showed moderate agreement (0.40–0.6) for all nutrients and food groups, except for niacin and calcium, which showed poor agreement (0.35). The relative validity results indicate that the crude Spearman''s correlation coefficients of FFQ1 and the 24-HRs ranged from 0.41 (for Vitamin C) to 0.65 (for fruit). The coefficients of each nutrient and food group in FFQ2 and the 24-HRs were higher than those in FFQ1 and the 24-HRs, indicating good correlation. Although all energy-adjusted Spearman''s correlation coefficients were lower than the crude coefficients, de-attenuation to correct for intra-individual variability improved the correlation coefficients. The weighted κ coefficients of nutrients and food groups ranged from 0.32 for beans to 0.52 for riboflavin in FFQ1 and the 24-HRs, and 0.32 for Vitamin C to 0.54 for riboflavin in FFQ2 and the 24-HRs.

Conclusion

The FFQ developed for female adolescents in the Suihua area is a reliable and valid instrument for ranking individuals within this study.