A genome-scale integrated approach aids in genetic dissection of complex flowering time trait in chickpea

A combinatorial approach of candidate gene-based association analysis and genome-wide association study (GWAS) integrated with QTL mapping, differential gene expression profiling and molecular haplotyping was deployed in the present study for quantitative dissection of complex flowering time trait in chickpea. Candidate gene-based association mapping in a flowering time association panel (92 diverse desi and kabuli accessions) was performed by employing the genotyping information of 5724 SNPs discovered from 82 known flowering chickpea gene orthologs of Arabidopsis and legumes as well as 832 gene-encoding transcripts that are differentially expressed during flower development in chickpea. GWAS using both genome-wide GBS- and candidate gene-based genotyping data of 30,129 SNPs in a structured population of 92 sequenced accessions (with 200–250 kb LD decay) detected eight maximum effect genomic SNP loci (genes) associated (34 % combined PVE) with flowering time. Six flowering time-associated major genomic loci harbouring five robust QTLs mapped on a high-resolution intra-specific genetic linkage map were validated (11.6–27.3 % PVE at 5.4–11.7 LOD) further by traditional QTL mapping. The flower-specific expression, including differential up- and down-regulation (>three folds) of eight flowering time-associated genes (including six genes validated by QTL mapping) especially in early flowering than late flowering contrasting chickpea accessions/mapping individuals during flower development was evident. The gene haplotype-based LD mapping discovered diverse novel natural allelic variants and haplotypes in eight genes with high trait association potential (41 % combined PVE) for flowering time differentiation in cultivated and wild chickpea. Taken together, eight potential known/candidate flowering time-regulating genes [efl1 (early flowering 1), FLD (Flowering locus D), GI (GIGANTEA), Myb (Myeloblastosis), SFH3 (SEC14-like 3), bZIP (basic-leucine zipper), bHLH (basic helix-loop-helix) and SBP (SQUAMOSA promoter binding protein)], including novel markers, QTLs, alleles and haplotypes delineated by aforesaid genome-wide integrated approach have potential for marker-assisted genetic improvement and unravelling the domestication pattern of flowering time in chickpea

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Development and Integration of Genome-Wide Polymorphic Microsatellite Markers onto a Reference Linkage Map for Constructing a High-Density Genetic Map of Chickpea

<div><p>The identification of informative <i>in silico</i> polymorphic genomic and genic microsatellite markers by comparing the genome and transcriptome sequences of crop genotypes is a rapid, cost-effective and non-laborious approach for large-scale marker validation and genotyping applications, including construction of high-density genetic maps. We designed 1494 markers, including 1016 genomic and 478 transcript-derived microsatellite markers showing <i>in-silico</i> fragment length polymorphism between two parental genotypes (<i>Cicer arietinum</i> ICC4958 and <i>C</i>. <i>reticulatum</i> PI489777) of an inter-specific reference mapping population. High amplification efficiency (87%), experimental validation success rate (81%) and polymorphic potential (55%) of these microsatellite markers suggest their effective use in various applications of chickpea genetics and breeding. Intra-specific polymorphic potential (48%) detected by microsatellite markers in 22 <i>desi</i> and <i>kabuli</i> chickpea genotypes was lower than inter-specific polymorphic potential (59%). An advanced, high-density, integrated and inter-specific chickpea genetic map (ICC4958 x PI489777) having 1697 map positions spanning 1061.16 cM with an average inter-marker distance of 0.625 cM was constructed by assigning 634 novel informative transcript-derived and genomic microsatellite markers on eight linkage groups (LGs) of our prior documented, 1063 marker-based genetic map. The constructed genome map identified 88, including four major (7–23 cM) longest high-resolution genomic regions on LGs 3, 5 and 8, where the maximum number of novel genomic and genic microsatellite markers were specifically clustered within 1 cM genetic distance. It was for the first time in chickpea that <i>in silico</i> FLP analysis at genome-wide level was carried out and such a large number of microsatellite markers were identified, experimentally validated and further used in genetic mapping. To best of our knowledge, in the presently constructed genetic map, we mapped highest number of new sequence-based robust microsatellite markers (634) which is an advancement over the previously documented (~300 markers) inter-specific genetic maps. This advanced high-density map will serve as a foundation for large-scale marker validation and genotyping applications, including identification and targeted mapping of trait-specific genes/QTLs (quantitative trait loci) with sub-optimal use of resources and labour in chickpea.</p></div

Markers mapped on the eight LGs of an integrated and inter-specific genetic map of chickpea.

<p>^aMarkers mapped on the eight LGs of a previously reported inter-specific genetic map (ICC4958 x PI489777) constructed by Gaur et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125583#pone.0125583.ref026" target="_blank">26</a>].</p><p>Markers mapped on the eight LGs of an integrated and inter-specific genetic map of chickpea.</p

An advanced inter-specific, high-resolution and integrated genetic map (ICC4958 x PI489777) of chickpea constructed by assigning 634 novel genomic and transcript-derived microsatellite markers on eight LGs of a previously reported similar 1063 marker-based genetic map (Gaur et al. [26]).

<p>The genetic distance (cM) and identity of the marker loci integrated are indicated on the left and right side of eight LGs, respectively. The earlier reported markers are considered as anchor markers to define eight LGs. The LGs are specified with <i>Arabic</i> numerals on the upper-side corresponding with the genetic map as reported by Gaur et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125583#pone.0125583.ref026" target="_blank">26</a>]. The newly integrated genomic and transcript-derived microsatellite markers in this study are highlighted with red colour. Markers designated as CaGMS represent <i>Cicer arietinum</i> genomic microsatellite markers, whereas CaTMS represent genic <i>C</i>. <i>arietinum</i> transcript-derived microsatellite markers.</p

Segregation pattern of transcript-derived (A) (CaTMS651) and genomic (B) (CaGMS16) microsatellite markers in a representative set of mapping individuals of a RIL population derived from the inter-specific cross between ICC4958 and PI489777 along with parental genotypes.

<p>The amplified microsatellite marker alleles are resolved using agarose gel-based assay and fluorescent dye-labeled automated fragment analyzer. The fragment sizes (bp) of the amplified parental polymorphic alleles are indicated. M: 50 bp DNA ladder size standard. *indicates the heterozygous alleles amplified by microsatellite markers which are further confirmed through automated fragment analysis (C). The identities of two markers with their detailed information are provided in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125583#pone.0125583.s002" target="_blank">S2 Table</a>.</p

Summary of genomic and transcript-derived in <i>silico</i> polymorphic microsatellite markers used in the present study.

<p>Summary of genomic and transcript-derived in <i>silico</i> polymorphic microsatellite markers used in the present study.</p

Markers mapped on the eight LGs of an integrated and inter-specific genetic map of chickpea.

<p>^aMarkers mapped on the eight LGs of a previously reported inter-specific genetic map (ICC4958 x PI489777) constructed by Gaur et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125583#pone.0125583.ref026" target="_blank">26</a>].</p><p>Markers mapped on the eight LGs of an integrated and inter-specific genetic map of chickpea.</p

Validation of a representative set of novel transcript-derived (I) and genomic (II) microsatellite markers showing <i>in silico</i> FLP between ICC4958 (1) and PI489777 (2) using the gel-based assay (I and II) and fluorescent dye labeled automated fragment analyzer (III and IV).

<p>The fragment sizes (bp) of the amplified polymorphic alleles are indicated. The identities of markers (A: CaTMS616, B: CaTMS654, C: CaTMS715, D: CaTMS716, E: CaTMS561, F: CaTMS577, G: CaTMS783, H: CaTMS651, I: CaGMS1, J: CaGMS3, K: CaGMS13, L: CaGMS16, M: CaGMS18, N: CaGMS19, O: CaGMS24, P: CaGMS23, Q: CaGMS20, R: CaGMS41, S: CaGMS43 and T: CaGMS45) with their detailed information are provided in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125583#pone.0125583.s002" target="_blank">S2 Table</a>. The primers CaTMS606 and CaGMS40 were used for automated fragment analysis (III and IV). M: 50 bp DNA ladder size standard.</p