Integrating multiple statistical indices to measure the stability of photosynthetic pigment content and composition in Brassica juncea (L.) Czern germplasm under varying environmental conditions.

Understanding the stability of photosynthetic pigments is crucial for developing crop cultivars with high productivity and resilience to the environmental stresses. This study leveraged GGE biplot, WAASB, and MTSI indices to assess the stability of content and composition of photosynthetic pigments in leaves and siliques of 286 Brassica juncea (L.) Czern. genotypes across three environments. The GGE biplot analysis identified NRCQR-9901 as the best genotype in terms of chlorophyll 'a' under conditions of high irradiance and long days (E1). For chlorophyll 'b' and total chlorophyll, NC-533728 performed the best. AJ-2 and NPJ-208 had the maximum total carotenoids levels in leaves. RLC-2 was characterized by maximum values for chlorophyll a, chlorophyll b, and total chlorophyll in the siliques. The low irradiance, short days, and moderate to high temperatures (E2) seemed perfect for the synthesis of photosynthetic pigments. NPJ-182 shows the maximum concentrations of chlorophyll 'a', total chlorophyll, and total carotenoids in leaves. Conversely, IC-597869, RE-389, and IC-597894 exhibited the highest concentrations of chlorophyll 'b' under an environment characterized by low light intensity, shorter daylights, and low temperatures (E3) during flowering and siliqua formation stages. The combined analysis found NPJ-182, NC-533728, CN-105233, RLC-2, CN-101846, JA-96, PBR-357, JM-3, and DTM-34 as top performers with high stability. Comparative transcriptome analysis with two stable and high-performing genotypes (PBR-357 and DTM-34) and two average performers (name the average performers) revealed upregulation of critical photosynthesis-related genes (ELIP1, CAB3.1, ELIP1.5, and LHCB5) in top performers. This study identified promising trait donors for use in breeding programs aimed at improving the mustard crop's photosynthetic efficiency, productivity, and stability.

Quantitative Inheritance of Sclerotinia Stem Rot Resistance in Brassica napus and Relationship to Cotyledon and Leaf Resistances.

Sclerotinia sclerotiorum is a necrotrophic fungus causing devastating stem rot and associated yield losses of canola/rapeseed (Brassica napus) worldwide, including in Australia. Developing host resistance against Sclerotinia stem rot is critical if this disease in canola/rapeseed is to be successfully managed, as cultural or chemical control options provide only partial or sporadic control. Three B. napus breeding populations, C2, C5 and C6, including the parents, F1, F2, BC1P1, and BC2P2, were used in a field study with an objective of exploring the inheritance pattern of disease resistance (based on stem lesion length [SLL]) and the genetic relationships of disease with stem diameter (SD) or days to first flowering (DTF), and to compare these new adult plant stem resistances against S. sclerotiorum with those of seedling (cotyledon and leaf) resistances in earlier studies. Heritability (broad sense) of SLL was 0.57 and 0.73 for population C2 at 3 and 5 weeks postinoculation and 0.21 for population C5 at 5 weeks postinoculation. Additive genetic variance was evident within all 3 populations for DTF but not for SD. Narrow-sense heritability for DTF was 0.48 (C2), 0.42 (C5), and 0.32 (C6). SD, DTF, and SLL were all inherited independently, with no significant genetic covariance between traits in bivariate analysis. Genetic variance for SLL in populations C2 and C5 was entirely nonadditive, and there was significant nonadditive genetic covariance of SLL at 3 and 5 weeks postinoculation. Generation means analysis in population C2 supported the conclusion that complex epistatic interactions controlled SLL. Several C2 and C5 progeny showed high adult plant stem resistance, which may be critical in developing enhanced stem resistance in canola/rapeseed. Although population C6 showed no genetic variation for SLL resistance in this study, it showed significant nonadditive genetic variance at the cotyledon and leaf stages in earlier studies. We conclude that host resistance varies across different plant growth stages, and breeding must be targeted for resistance at each growth stage. In populations C2, C5, and C6, resistance to S. sclerotiorum in stem, leaf, and cotyledon was always controlled by nonadditive effects such as complex epistasis or dominance. Overall, our findings in relation to the quantitative inheritance of Sclerotinia stem rot resistance, together with the new high-level resistances identified, will enable breeders to select/develop genotypes with enhanced resistances to S. sclerotiorum.

Genetic analyses of nitrogen assimilation enzymes in Brassica juncea (L.) Czern & Coss.

Nitrogen (N) is a critical input for plant growth and development. A better understanding of N uptake and utilization is important to develop plant breeding strategies for improving nitrogen use efficiency (NUE). With that objective in mind, we assayed a SNP-genotyped association panel comprising 92 inbred lines for the activities of nitrate reductase (NR), nitrite reductase (NIR), glutamine synthetase (GS) and glutamate synthase (GOGAT). All these enzymes are associated with N assimilation. The experiments were carried out at two levels of N application: no added N (N0) and agrnomically recommened dose (100 kg/ha) of N application (N100). Genome wide association studies (GWAS) helped to identify several marker-trait associations (MTAs), involving chromosomes A01, A06, A08, B02, B04, B05 and B08. These explained high phenotypic variation (up to 32%). Annotation of the genomic region(s) in or around significant SNPs allowed prediction of genes encoding high affinity nitrate transporters, glutamine synthetase 1.3, myb-like transcription factor family protein, bidirectional amino acid transporter 1, auxin signaling F-box 3 and oxidoreductases. This is the first attempt to use GWAS for identification of enzyme QTLs to explain variation for nitrogen assimilation enzymes in Brassica juncea.

Development and molecular-genetic characterization of a stable Brassica allohexaploid.

KEY MESSAGE: We report first-time synthesis of a stable Brassica allohexaploid. It may evolve into a new species and also advance our understanding of pairing regulation and genome evolution in complex allopolyploids. Crop Brassicas include both monogenomic and digenomic species. A trigenomic Brassica (AABBCC) is not known to exist in nature. Past attempts to synthesize a stable allohexaploid were not successful due to aberrant meiosis and very high proportion of aneuploid plants in the selfed progenies. We report the development of a stable allohexaploid Brassica (2n = 54; AABBCC). Genomic in situ hybridization confirmed the complete assemblage of three genomes. Only allohexaploids involving B. rapa cv. R01 (2n = 20; AA) as pollinator with a set of B. carinata (2n = 34; BBCC) were stable. These exhibited a high proportion (0.78-0.94) of pollen mother cells with normal meiosis and an excellent hexaploid ratio (0.80-0.94) in the selfed progenies. Stability of two allohexaploid combinations was demonstrated from H1 to H4 generations at two very diverse locations in India. Graphical genotyping of allohexaploids allowed detection of chromosome fragment exchanges among three genomes. These were much smaller for meiotically stable allohexaploids as compared to unstable ones. The putative hexaploids were morphologically closer to the female donor, B. carinata, for leaf morphology, inflorescence structure and flower shape. The newly formed allohexaploid may also provide unique opportunities to investigate the immediate genetic and genomic consequences of a Brassica allohexaploid with three resident genomes.

Resynthesis of Brassica napus through hybridization between B. juncea and B. carinata.

KEY MESSAGE: First report for the resynthesis of Brassica napus by recombining A and C genome from B. juncea and B. carinata , respectively. Also documents B genome introgressions in resynthesized B. napus. Resynthesis of Brassica napus (AACC) was achieved by hybridizing Brassica juncea (AABB) with Brassica carinata (BBCC). This was facilitated by spontaneous chromosome doubling in the F1 hybrid (ABBC) to yield octaploid (AABBBBCC), elimination of extra B genome chromosomes in the resulting octaploid and in subsequent selfed generations, aided with directed selection for fertile plants having B. napus morphology. Twenty-five plants with varying degrees of resemblance to natural B. napus were identified from 17 A5 progenies and assayed for cytogenetic stability and genetic diversity. Majority of these plants, except six (2n = 38) were hyperploids (2n = 40-56). The six plants with 2n = 38 were designated as derived B. napus types. These showed an expected meiotic configuration of 19II at metaphase-I, with 19-19 distribution at anaphase-I. Genotyping based on A and C genome specific primers confirmed genetic identity of six derived (2n = 38) B. napus plants with natural types whereas genotyping with B genome specific primers indicated introgression of B genome segments. This was also confirmed by genomic in situ hybridization (GISH). Strong signals of B genome probe were detected, proving hitherto unreported genetic exchanges between B and A/C chromosomes. These introgressions possibly occurred en route five generations of selfing. Derived plants yielded fertile hybrids in crosses with natural B. napus var. GSC 6. The selfed derived plants as evaluated in A6 plant to progeny rows were morphologically similar to natural B. napus, and meiotically stable. Agronomic assessment of these progenies revealed variation for key morpho-physiological traits. Of special interest were the progenies with plants having oil content exceeding 47% as against about 39-41% in existing cultivars.

Genetic and molecular analysis of stem rot (Sclerotinia sclerotiorum) resistance in Brassica napus (canola type).

Identifying the molecular and genetic basis of resistance to Sclerotinia stem rot (Sclerotinia sclerotiorum) is critical for developing long-term and cost-effective management of this disease in rapeseed/canola (Brassica napus). Current cultural or chemical management options provide, at best, only partial and/or sporadic control. Towards this, a B. napus breeding population (Mystic x Rainbow), including the parents, F1, F2, BC1P1 and BC1P2, was utilized in a field study to determine the inheritance pattern of Sclerotinia stem rot resistance (based on stem lesion length, SLL). Broad sense heritability was 0.58 for SLL and 0.44 for days to flowering (DTF). There was a significant negative correlation between SLL and stem diameter (SD) (r = -0.39) and between SLL and DTF (r = -0.28), suggesting co-selection of SD and DTF traits, along with SLL, should assist in improving overall resistance. Non-additive genetic variance was evident for SLL, DTF, and SD. In a genome wide association study (GWAS), a significant quantitative trait locus (QTL) was identified for SLL. Several putative candidate marker trait associations (MTA) were located within this QTL region. Overall, this study has provided valuable new understanding of inheritance of resistance to S. sclerotiorum, and has identified QTL, MTAs and transgressive segregants with high-level resistances. Together, these will foster more rapid selection for multiple traits associated with Sclerotinia stem rot resistance, by enabling breeders to make critical choices towards selecting/developing cultivars with enhanced resistance to this devastating pathogen.

Cytogenetic and Molecular Characterization of B-Genome Introgression Lines of Brassica napus L.

Brassica napus introgression lines (ILs), having B-genome segments from B. carinata, were assessed genetically for extent of introgression and phenotypically for siliqua shatter resistance. Introgression lines had 7-9% higher DNA content, were meiotically stable, and had almost normal pollen fertility/seed set. Segment introgressions were confirmed by fluorescent genomic in situ hybridization (fl-GISH), SSR analyses, and SNP studies. Genotyping with 48 B-genome specific SSRs detected substitutions from B3, B4, B6, and B7 chromosomes on 39 of the 69 ILs whereas SNP genotyping detected a total of 23 B-segments (≥3 Mb) from B4, B6, and B7 introgressed into 10 of the 19 (C1, C2, C3, C5, C6, C8, C9, A3, A9, A10) chromosomes in 17 ILs. The size of substitutions varied from 3.0 Mb on chromosome A9 (IL59) to 42.44 Mb on chromosome C2 (IL54), ranging from 7 to 83% of the recipient chromosome. Average siliqua strength in ILs was observed to be higher than that of B. napus parents (2.2-6.0 vs. 1.9-4.0 mJ) while siliqua strength in some of the lines was almost equal to that of the donor parent B. carinata (6.0 vs.7.2 mJ). These ILs, with large chunks of substituted B-genome, can prove to be a useful prebreeding resource for germplasm enhancement in B. napus, especially for siliqua shatter resistance.

Increasing trends of leptospirosis in northern India: a clinico-epidemiological study.

BACKGROUND: Leptospirosis, a zoonosis associated with potentially fatal consequences, has long been a grossly underreported disease in India. There is no accurate estimate of the problem of leptospirosis in non-endemic areas such as north India. METHODS/PRINCIPAL FINDINGS: In order to understand the clinical spectrum and risk factors associated with leptospirosis, we carried out a retrospective study in patients with acute febrile illness in north India over the last 5 years (January 2004 to December 2008). There was increased incidence of leptospirosis (11.7% in 2004 to 20.5% in 2008) as diagnosed by IgM ELISA and microscopic agglutination titer in paired acute and convalescent sera. The disease showed a peak during the rainy season (August and September). We followed up 86 cases of leptospirosis regarding their epidemiological pattern, clinical features, laboratory parameters, complications, therapy, and outcome. Mean age of patients was 32.6 years (2.5 years to 78 years) and males (57%) outnumbered females (43%). Infestation of dwellings with rats (53.7%), working in farm lands (44.2%), and contact with animals (62.1%) were commonly observed epidemiological risk factors. Outdoor workers including farmers (32.6%), labourers (11.6%), para-military personnel (2.3%), and sweepers (1.2%) were commonly affected. Modified Faine's criteria could diagnose 76 cases (88.3%). Renal failure (60.5%), respiratory failure (20.9%), the neuroleptospirosis (11.6%), and disseminated intravascular coagulation (DIC) (11.6%) were the commonest complications. Five patients died, giving a case fatality rate of 5.9%. CONCLUSIONS/SIGNIFICANCE: There has been a rapid rise in the incidence of leptospirosis in north India. Severe complications such as renal failure, respiratory failure, neuroleptospirosis, and DIC are being seen with increasing frequency. Increased awareness among physicians, and early diagnosis and treatment, may reduce mortality due to leptospirosis.