Pyramiding of qDTY1.1 and qDTY3.1 into rice mega-variety Samba Mahsuri-Sub1: physiological performance under water deficit conditions.

Water deficit is a crucial factor causing huge loss to rice productivity. The present study aimed to develop a multiple stress tolerant genotype by pyramiding drought tolerant yield QTLs qDTY1.1 and qDTY3.1 into high yielding rice variety Samba Mahsuri Sub-1(SMS) through marker assisted pyramiding. To achieve this six introgression lines of SMS carrying qDTY1.1 (SAB) were crossed with DRR-50, an Essentially Derived Variety of SMS carrying qDTY3.1. The SAB lines are taller than SMS due to tight linkage between qDTY1.1 and wild type SD-1. Therefore, F2 generation of crosses were screened for recombinants between SD-1 and qDTY1.1. Phenotyping of 1530 F2 plants representing three F2 populations from 35 F1 hybrids, identified 305 dwarf plants. Three dwarf F2 plants along with three others carrying qDTY1.1 and qDTY3.1 were forwarded to F3 generation. From the six F3 (SABD) lines fourteen pyramided progenies were selected and forward to F4 generation. The six SABD F3 lines SABD-7, SABD-8, SABD-9, SABD-76, SABD-79 and SABD-80 along with parents were evaluated under moisture stress (MS) for various physiological parameters. Chlorophyll and relative water content were more, while canopy temperature and malonaldehyde (MDA) content were lesser in SABD lines compared to parents indicating tolerance under MS. Variance due to genotypes was highly significant for all the yield related traits except test weight. Based on seed morphology, agronomic characters and physiological parameters six superior lines SABD-9-3, SABD-9-2, SABD-9-6, SABD-9-7, SABD-76-2 and SABD-76-6 performing better under MS were identified, which could be released after multi-location evaluation. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01387-5.

Outcomes of Ahmed glaucoma valve implantation and risk factors for graft failure in eyes with Descemet's stripping endothelial keratoplasty.

PURPOSE: The aim of this study was to assess the efficacy of Ahmed glaucoma valve (AGV) in eyes with Descemet's stripping endothelial keratoplasty (DSEK) and glaucoma and evaluate the risk factors for failure. METHODS: Retrospective interventional study included 27 eyes (27 subjects) that underwent AGV implantation following DSEK. The main outcome measure was AGV success, defined as IOP between 6 and 21 mmHg with or without topical antiglaucoma medications (AGMs). Secondary outcome measure was graft survival. Graft failure was defined as corneal edema for ≥ 1 month or irreversible graft opacity requiring intervention. RESULTS: Eighteen eyes (66.7%) had anterior chamber tube and 9 eyes (33.3%) had sulcus tube placement. Median follow-up was 1.2 years (interquartile range, IQR, 0.5-2.5 years). Following AGV implantation, the median IOP decreased significantly (p < 0.0001), median number of AGMs reduced significantly (p < 0.001) and median visual acuity was maintained (p = 0.76). Cumulative success probability of AGV was 75.8 ± 10.6% at 1 year, and 75.8 ± 10.6% at 2 years. For 24 clear grafts before AGV, estimates of success post-AGV were 100% at 1 year, 77.8 ± 10% at 2 years and 51.9 ± 16.4% at 3 years. Postoperative tube intervention was a borderline significant risk factor for graft failure (p = 0.05) with hazards ratio of 7.2; however, tube location was not associated with failure. CONCLUSIONS: AGV can be considered an effective surgical option for IOP control in eyes with DSEK and glaucoma. However, high risk of long-term graft failure in these eyes needs to be considered specially those eyes needing tube intervention.

In silico characterization, and expression analysis of rice golden 2-like (OsGLK) members in response to low phosphorous.

The availability of phosphorus (P) affects productivity of rice. Under acidic soil conditions (pH < 5.5), P is rapidly immobilized in the soil. Several transcription factors play an important role in low Pi tolerance response, including MYB family members but their role in acidic soil is yet unknown. In this study, genome wide identification and characterization of golden 2-like (GLK) members belonging to GARP superfamily from rice (OsGLK) led to identification of 46 members distributed over 12 chromosomes. We assigned gene nomenclature, analyzed gene structure and identified mutant orthologs and phenotypes in maize and rice, respectively. On the basis of biological functions three categories viz., (a) two-component response regulator (five members), (b) putative transcription factor (21 members) and (c) phosphate starvation response (8 members) were identified. Phylogenetic analysis revealed a total of nine subgroups with MYB homeodomain-like and MYB CC-type domains conserved across members. Expression profiling of OsGLKs in response to 24 and 48 h of low Pi in four contrasting rice genotypes, revealed significantly higher expression of OsGLK10, OsGLK15, OsGLK22 and OsGLK30 in tolerant genotypes as compared to susceptible genotypes, suggesting their role in Pi starvation tolerance. Meta analyses and cis-regulatory elements (CREs) profiling of OsGLK showed diverse expression pattern in various tissues and organs and also modulation in response to various abiotic and biotic stresses. Our results highlight the versatile role of this diverse and complex GLK family, in particular to abiotic stress. These genes will form the basis of future studies on low Pi tolerance in acidic soils.

Marker-trait association for low-light intensity tolerance in rice genotypes from Eastern India.

Light intensity is a crucial environmental factor that affects photosynthesis and ultimately, grain yield in rice. However, no gene or marker directly associated with improved performance under low-light intensity under field conditions has been identified till date. With an aim of identifying genes and markers associated with improved performance (measured in terms of better yields) under low-light intensity, an integrated field screening, in silico and wet lab validation analysis was performed. Field-based screening of a diverse set of 110 genotypes led to the identification of two physiological and three morphological parameters critical for low-light tolerance in rice. In silico analysis using information available in public databases led to the identification of a set of 90 potential candidate genes which were narrowed to thirteen genic targets for possible marker-trait association. Marker-trait association on the panel of 48 diverse rice genotypes varying in their response to low-light intensity led to the identification of six markers [HvSSR02-44 (biological yield), HvSSR02-52 (spikelet fertility), HvSSR02-54 (grain yield), HvSSR06-56 (spikelet fertility), HvSSR06-69 (spikelet fertility; biological yield), HvSSR09-45 (spikelet fertility)] lying on chromosomes 2, 6 and 9 showing significant association (R2 > 0.1) for traits like grain yield/plant, biological yield and spikelet fertility under low light. Eight rice genes [including member of BBX (B-box) family] lying within 10 kb distance of these identified markers already reported for their role in response to stress or change in plant architecture in rice were also identified. The eight rice genotypes, five traits, eight genes and six markers identified in the current study will help in devising strategies to increase yield under low light intensity and pave way for future application in marker-assisted breeding.

In silico characterisation of novel rice transcripts differentially expressed in phosphorus dificient conditions suggests a role of these transcripts in multiple abiotic stresses.

Phosphorus deficiency adversely affects crop productivity. The mechanism of tolerance in plants is not well understood. The current study successfully annotated a set of highly significant (Log2 RPKM ≥3) nine novel sequences up-regulated in P deficient condition identified from a low P tolerant rice genotype. Sequence annotation identified two transcripts (Os01g37260 and Os02g11060) carrying known domains, F-box and WD, respectively. Multiple Expectation maximization for Motif Elicitation (MEME) revealed presence of conserved domains like D[LP][HY][CL]D[CM][DT]C[AP][DQ][IQ]C, [EH][DN]HN[HS] [ER][FY][EP]I[HN]H which might play a role in phosphorus deficiency tolerance. Analysis of the upstream regions indicated presence of stress responsive elements like E Box, ABRE, and MYBCORE suggesting regulation of the novel transcripts by DNA binding. Protein localization prediction tool suggests that these novel proteins might be targeted to nucleus, chloroplast and cell wall. Transcripts Os02g03640 and Os02g10250 revealed potential target sites for microRNA binding suggesting role of novel miRNAs in low phosphorus response. Our analysis suggests that an F-box protein, Os01g37260 (OSFBx14) might be a promising candidate gene playing a role in multiple abiotic stresses including P deficiency.

Genetic analysis of purple pigmentation in rice seed and vegetative parts - implications on developing high-yielding purple rice (Oryza sativa L.).

Pigmentation in rice grains is an important quality parameter. Purple-coloured rice (Oryza sativa L.) indicates the presence of high anthocyanin with benefits of antioxidant properties. However, the genetic mechanism of grain colour is not fully understood. Therefore, the study focused on understanding pigmentation in grain pericarp and vegetative parts, and its relationship with blast resistance and enhanced grain yield. Three local cultivars from the northeastern region (NER) of India - Chakhao Poireiton (purple), Mang Meikri (light brown), and Kala Joha (white) - along with high-yielding varieties (HYVs) Shasharang (light brown) and Sahbhagi dhan (white) were used to develop biparental populations. The findings suggested that pigmentation in vegetative tissue was governed by the inter-allelic interaction of several genes. Haplotype analysis revealed that Kala3 complemented Kala4 in enhancing purple pigmentation and that Kala4 is not the only gene responsible for purple colour as evident by the presence of a desired allele for markers RID3 and RID4 (Kala4 locus) in Chakhao Poireiton and Kala Joha irrespective of their pericarp colour, implying the involvement of some other additional, unidentified genes/loci. RID3 and RID4 together with RM15191 (Kala3 locus) could be employed as a reliable marker set for marker-assisted selection (MAS). Pericarp colour was strongly correlated with colour in different vegetative parts, but showed a negative correlation with grain yield. Pb1, reported to be associated with panicle blast resistance, contributed to leaf blast resistance. Transgressive segregants for improved pigmentation and high yield were identified. The selection of lines exhibiting coloured pericarp, high anthocyanin content, aroma, blast resistance, and increased yield compared to their respective HYV parents will be valuable resources in the rice breeding programme.

Outcomes of Ahmed Glaucoma Valve Implantation in Eyes with Pediatric Keratoplasty.

PURPOSE: This study aimed to assess the efficacy and safety of Ahmed glaucoma valve (AGV) (New World Medical Inc.) implantation in eyes with pediatric keratoplasty and glaucoma. DESIGN: Retrospective, noncomparative, interventional case series. PARTICIPANTS: Twenty-eight eyes of 25 children who underwent AGV after penetrating keratoplasty (PK) in 22 eyes and Descemet's stripping endothelial keratoplasty (DSEK) in 6 eyes were included. METHODS: Children (age ≤ 16 years) treated with AGV implant after keratoplasty from 2008 to 2019 with documented 6 months follow-up operated by a single surgeon were included. MAIN OUTCOME MEASURES: The primary outcome measure was AGV survival (intraocular pressure [IOP] between 6 and 21 mmHg with or without topical anti-glaucoma medications [AGMs]), and the secondary outcome measure was corneal graft survival. RESULTS: The mean age (± standard deviation) of patients at AGV surgery was 4.8 ± 0.7 years. The mean follow-up after AGV was 3.4 ± 0.5 years. The mean IOP decreased significantly from 31.5 ± 2.0 mmHg to 21.1 ± 2.3 mmHg at 6 months postsurgery (P < 0.001). The mean number of AGM decreased significantly (P < 0.001), and there was no change in mean visual acuity (P > 0.04 with Bonferroni correction). The cumulative survival probability of AGV was 92.9% ± 4.9% at 1 year and 81.3% ± 11.7% at 5 years. The AGV success was comparable between PK and DSEK eyes (P = 0.73). The cumulative probability of graft survival was 96.2% ± 3.8% at 1 year and 77.8% ± 9.1% at 5 years. The graft survival was also comparable between PK and DSEK eyes (P = 0.18). One eye needed tube trimming; none had tube or implant exposure. The major complication noted was corneal graft infection in 8 eyes (28.5%). CONCLUSIONS: In this cohort, the long-term AGV success and graft survival outcomes are satisfactory considering that AGV was performed in complex eyes with keratoplasty.

Playing with colours: genetics and regulatory mechanisms for anthocyanin pathway in cereals.

Cereals form the most important source of energy in our food. Currently, demand for coloured food grains is significantly increasing globally because of their antioxidant properties and enhanced nutritional value. Coloured grains of major and minor cereals are due to accumulation of secondary metabolites like carotenoids and flavonoids such as anthocyanin, proanthocyanin, phlobaphenes in pericarp, aleurone, lemma, testa or seed coat of grains. Differential accumulation of colour in grains is regulated by several regulatory proteins and enzymes involved in flavonoid and caroteniod biosynthesis. MYB and bHLH gene family members are the major regulators of these pathways. Genes for colour across various cereals have been extensively studied; however, only a few functional and allele-specific markers to be utilized directly in breeding programmes are reported so far. In this review, while briefly discussing the well studied and explored carotenoid pathway, we focus on a much more complex anthocyanin pathway that is found across cereals. The genes and their orthologs that are responsible for encoding key regulators of anthocyanin biosynthesis are discussed. This review also focuses on the genetic factors that influence colour change in different cereal crops, and the available/reported markers that can be used in breeding programs for utilizing this pathway for enhancing food and nutritional security.

A 1.84-Mb region on rice chromosome 2 carrying SPL4, SPL5 and MLO8 genes is associated with higher yield under phosphorus-deficient acidic soil.

Phosphorus (P) deficiency is one of the major limiting factors for rice productivity with only one locus (PSTOL1) available for field based application. A biparental mapping population (F6) derived from two P deficiency tolerant genotypes (Sahbhagi Dhan (SD) (PSTOL1+) and Chakhao Poreiton (CP) (PSTOL1-)), in which, transcriptome data generated from our lab had previously shown existence of diverse mechanisms was used to identify novel regions for better yield under lowland acidic soils. Phenotyping at F4, F5 and F6 generations revealed significant correlation between traits like tiller number at 30 days (TN 30), tiller number at 60 days (TN 60), filled grains (FG), percent spikelet fertility (SF%), panicle number (PN) and grain yield per panicle (GYPP) and also association with better yield/performance under low P acidic soil conditions. Through selected genotyping on a set of forty superior and inferior lines using SSR, candidate gene-based and SNP polymorphic markers, 5 genomic regions associated with various yield-related traits were identified. Marker trait association studies revealed 13 markers significantly associated with yield attributing traits and PUE under lowland acidic field conditions. Chi-square and regression analyses of markers run on the entire population identified seven and six markers for SF% and GYPP, respectively, and two for biological yield with positive allele derived from SD which constitute a novel 1.847-Mb region on chromosome 2 flanked by two markers RM12550 and PR9-2. Expression analysis of 7 candidate genes lying within this region across SD, CP and two low P susceptible rice genotypes has revealed that expression of four genes including SPL4, SPL5, ACA9 and MLO8 is significantly upregulated only in SD under low P conditions. In CP, there is low expression of MLO8 under low P conditions, whereas SPL4, SPL5 and Os02g08120 are downregulated. In the case of the two susceptible genotypes, there is no expression of Os02g08120 either in optimum or limiting conditions. Sequence data across a panel of 3024 rice genotypes also suggests that there is polymorphism for these differentially expressed genes. The genes and underlying markers identified on chromosome 2 will be key to imparting tolerance to low P in diverse genetic backgrounds and for marker-assisted selection for higher yield under lowland acidic conditions.

Root transcriptome reveals efficient cell signaling and energy conservation key to aluminum toxicity tolerance in acidic soil adapted rice genotype.

Aluminium (Al) toxicity is the single most important contributing factor constraining crop productivity in acidic soils. Hydroponics based screening of three rice genotypes, a tolerant (ARR09, AR), a susceptible (IR 1552, IR) and an acid soil adapted landrace (Theruvii, TH) revealed that AR accumulates less Al and shows minimum decrease in shoot and root biomass under Al toxicity conditions when compared with IR. Transcriptome data generated on roots (grown in presence or absence of Al) led to identification of ~1500 transcripts per genotype with percentage annotation ranging from 21.94% (AR) to 29.94% (TH). A total of 511, 804 and 912 DEGs were identified in genotypes AR, IR and TH, respectively. IR showed upregulation of transcripts involved in exergonic processes. AR appears to conserve energy by downregulating key genes of glycolysis pathway and maintaining transcript levels of key exergonic step enzymes under Al stress. The tolerance in AR appears to be as a result of novel mechanism as none of the reported Al toxicity genes or QTLs overlap with significant DEGs. Components of signal transduction and regulatory machinery like transcripts encoding zinc finger protein, calcieurin binding protein and cell wall associated transcripts are among the highly upregulated DEGs in AR, suggesting increased and better signal transduction in response to Al stress in tolerant rice. Sequencing of NRAT1 and glycine-rich protein A3 revealed distinct haplotype for indica type AR. The newly identified components of Al tolerance will help in designing molecular breeding tools to enhance rice productivity in acidic soils.

Comparative functional analysis of three abiotic stress-inducible promoters in transgenic rice.

To identify minimal effective promoters for driving abiotic stress-inducible transgene expression in rice, we selected promoter elements of three stress-responsive genes, viz. rab16A coding for dehydrin, OsABA2 coding for zeaxanthin epoxidase, and a gene coding for a hypothetical protein (HP1) based on the presence of ABA-, salt- and drought-responsive cis-acting elements. These were translationally fused to the gusA reporter gene and introduced into rice to study their effect on heterologous gene expression. The OsABA2 promoter was found to be the most effective and desirable promoter among the three in terms of driving a low constitutive transgene expression under normal conditions and high induction in response to ABA, salt and drought stress, the highest being a 12-fold induction in response to ABA. The rab16A and HP1 promoters resulted in high levels of constitutive expression. While induction of GUS activity was generally two- to threefold for all the treatments in roots for both the promoters, induction in leaves was generally insignificant, the exceptions being rab16A in response to continuous salt stress and HP1 in response to water deficit. It was also observed that the three promoters, in general, resulted in lower constitutive expression, but higher induction in roots as compared to leaves.

Root transcriptomes of two acidic soil adapted Indica rice genotypes suggest diverse and complex mechanism of low phosphorus tolerance.

Low phosphorus (P) tolerance in rice is a biologically and agronomically important character. Low P tolerant Indica-type rice genotypes, Sahbhagi Dhan (SD) and Chakhao Poreiton (CP), are adapted to acidic soils and show variable response to low P levels. Using RNAseq approach, transcriptome data was generated from roots of SD and CP after 15 days of low P treatment to understand differences and similarities at molecular level. In response to low P, number of genes up-regulated (1318) was more when compared with down-regulated genes (761). Eight hundred twenty-one genes found to be significantly regulated between SD and CP in response to low P. De novo assembly using plant database led to further identification of 1535 novel transcripts. Functional annotation of significantly expressed genes suggests two distinct methods of low P tolerance. While root system architecture in SD works through serine-threonine kinase PSTOL1, suberin-mediated cell wall modification seems to be key in CP. The transcription data indicated that CP relies more on releasing its internally bound Pi and coping with low P levels by transcriptional and translational modifications and using dehydration response-based signals. Role of P transporters seems to be vital in response to low P in CP while sugar- and auxin-mediated pathway seems to be preferred in SD. At least six small RNA clusters overlap with transcripts highly expressed under low P, suggesting role of RNA super clusters in nutrient response in plants. These results help us to understand and thereby devise better strategy to enhance low P tolerance in Indica-type rice.

Understanding Fe2+ toxicity and P deficiency tolerance in rice for enhancing productivity under acidic soils.

Plants experience low phosphorus (P) and high iron (Fe) levels in acidic lowland soils that lead to reduced crop productivity. A better understanding of the relationship between these two stresses at molecular and physiological level will lead to development of suitable strategies to increase crop productivity in such poor soils. Tolerance for most abiotic stresses including P deficiency and Fe toxicity is a quantitative trait in rice. Recent studies in the areas of physiology, genetics, and overall metabolic pathways in response to P deficiency of rice plants have improved our understanding of low P tolerance. Phosphorous uptake and P use efficiency are the two key traits for improving P deficiency tolerance. In the case of Fe toxicity tolerance, QTLs have been reported but the identity and role played by underlying genes is just emerging. Details pertaining to Fe deficiency tolerance in rice are well worked out including genes involved in Fe sensing and uptake. But, how rice copes with Fe toxicity is not clearly understood. This review focuses on the progress made in understanding these key environmental stresses. Finally, an opinion on the key genes which can be targeted for this stress is provided.

Marker-free transgenic (MFT) near-isogenic introgression lines (NIILs) of 'golden' indica rice (cv. IR64) with accumulation of provitamin A in the endosperm tissue.

We have developed near-isogenic introgression lines (NIILs) of an elite indica rice cultivar (IR64) with the genes for beta-carotene biosynthesis from dihaploid (DH) derivatives of golden japonica rice (cv. T309). A careful analysis of the DH lines indicated the integration of the genes of interest [phytoene synthase (psy) and phytoene desaturase (crtI)] and the selectable marker gene (hygromycin phosphotransferase, hph) in two unlinked loci. During subsequent crossing, progenies could be obtained carrying only the locus with psy and crtI, which was segregated independently from the locus containing the hph gene during meiotic segregation. The NIILs (BC(2)F(2)) showed maximum similarity with the recurrent parent cultivar IR64. Further, progenies of two NIILs were devoid of any fragments beyond the left or right border, including the chloramphenicol acetyltransferase (cat) antibiotic resistance gene of the transformation vector. Spectrophotometric readings showed the accumulation of up to 1.06 microg total carotenoids, including beta-carotene, in 1 g of the endosperm. The accumulation of beta-carotene was also evident from the clearly visible yellow colour of the polished seeds.

Seedling stage low temperature response in tolerant and susceptible rice genotypes suggests role of relative water content and members of OsSNAC gene family.

Low temperature (LT) severely affects rice growth and grain yield. Recently, we reported contrasting genotypes including ARR 09 and Takyer for seedling stage long duration low temperature response. Here we show that susceptible rice genotypes show an increase in lipid peroxide levels and decrease in relative water content (RWC) to a higher extent in comparison to tolerant genotypes in response to 3 h LT. Stress induced NAC family members (OsNAC1, OsNAC2, OsNAC3, and OsNAC5) showed a higher transcript accumulation in tolerant genotypes than in sensitive genotypes after LT treatment suggesting stress tolerance might be due to higher expression of stress-responsive transcription factors. Furthermore, ARR 09 can be used as an important genetic resource to better understand LT tolerance mechanism.

Allele mining across DREB1A and DREB1B in diverse rice genotypes suggest a highly conserved pathway inducible by low temperature.

Low temperature stress is one of the major limiting factors affecting rice productivity in higher altitudes. DREB1A and DREB1B, are two transcription factors that have been reported to play key regulatory role in low temperature tolerance. In order to understand whether natural genetic variation in these two loci leads to cold tolerance or susceptibility, OsDREB1A and OsDREB1B were targeted across several rice genotypes showing differential response to low temperature. Expression data suggests induction of gene expression in shoots in response to low temperature in both tolerant and susceptible genotypes. Upon sequence analysis of 20 rice genotypes, eight nucleotide changes were identified including two in the coding region and six in the 5'UTR. None of the discovered novel variations lie in the conserved region of the genes under study, thereby causing little or no changes in putative function of the corresponding proteins. In silico analysis using a diverse set of 400 O. sativa revealed much lower nucleotide diversity estimates across two DREB loci and one other gene (MYB2) involved in DREB pathway than those observed for other rice genes. None of the changes showed association with seedling stage cold tolerance, suggesting that nucleotide changes in DREB loci are unlikely to contribute to low temperature tolerance. So far, data concerning the physiological role and regulation of DREB1 in different genetic background are very limited; it is to be expected that they will be studied extensively in the near future.