Unravelling the metabolomic diversity of pigmented and non-pigmented traditional rice from Tamil Nadu, India.

Rice metabolomics is widely used for biomarker research in the fields of pharmacology. As a consequence, characterization of the variations of the pigmented and non-pigmented traditional rice varieties of Tamil Nadu is crucial. These varieties possess fatty acids, sugars, terpenoids, plant sterols, phenols, carotenoids and other compounds that plays a major role in achieving sustainable development goal 2 (SDG 2). Gas-chromatography coupled with mass spectrometry was used to profile complete untargeted metabolomics of Kullkar (red colour) and Milagu Samba (white colour) for the first time and a total of 168 metabolites were identified. The metabolite profiles were subjected to data mining processes, including principal component analysis (PCA), Orthogonal Partial Least Square Discrimination Analysis (OPLS-DA) and Heat map analysis. OPLS-DA identified 144 differential metabolites between the 2 rice groups, variable importance in projection (VIP) ≥ 1 and fold change (FC) ≥ 2 or FC ≤ 0.5. Volcano plot (64 down regulated, 80 up regulated) was used to illustrate the differential metabolites. OPLS-DA predictive model showed good fit (R2X = 0.687) and predictability (Q2 = 0.977). The pathway enrichment analysis revealed the presence of three distinct pathways that were enriched. These findings serve as a foundation for further investigation into the function and nutritional significance of both pigmented and non-pigmented rice grains thereby can achieve the SDG 2.

Identification, validation and quantification of thymoquinone in conjunction with assessment of bioactive possessions and GC-MS profiling of pharmaceutically valuable crop Nigella (Nigella sativa L.) varieties.

Background: Plants have been pivotal in traditional and modern medicine globally, with historical evidence supporting their therapeutic applications. Nigella (Nigella sativa L.) is an annual herbaceous plant of the Ranunculaceae family and is cultivated in the Middle East, Eastern Europe, and Western and Central Asia. The medicinal use of plants dates back thousands of years, documented in ancient writings from various civilizations. Alkaloids, phenolics, saponins, flavonoids, terpenoids, anthraquinones, and tannins found in plants exhibit antioxidant, immunomodulatory, anti-inflammatory, anticancer, antibacterial, and antidiabetic activities. Methodology: This study specifically examines the pharmacological potential of Nigella sativa L., emphasizing thymoquinone-a compound with diverse nutraceutical benefits. The extraction, characterization, and quantification of thymoquinone, alongside other physicochemical parameters, were carried out using ethanol through Soxhlet extraction procedures on five nigella varieties. HPLC analysis was performed to determine the maximum accumulation of thymoquinone in the released variety of the plant and the chemical composition of the seed oil isolated from Nigella sativa L., varieties utilized in the study was determined through GC-MS analysis. Results: The research revealed that the Ajmer nigella-20 variety stands out, exhibiting elevated levels of thymoquinone (0.20 ± 0.07%), antioxidants (76.18 ± 1.78%), and substantial quantities of total phenols (31.85 ± 0.97 mg GAEg-1 seed) and flavonoids (8.150 ± 0.360 mg QE 100 g-1 seed) compared to other varieties. The GC-MS profiling showed the presence of 11 major compounds in the studied varieties, with p-cymene, longifolene, and myristic acid identified as the major chemical compounds present in the oil. Conclusion: The observed variations among Nigella varieties indicate the Ajmer nigella-20 variety as particularly promising for thymoquinone and bioactive compound extraction. This study underscores Nigella's potential as a source of pharmacologically active compounds, highlighting the need for further exploration in therapeutic applications.

Characterization of terminal flowering cowpea (Vigna unguiculata (L.) Walp.) mutants obtained by induced mutagenesis digs out the loss-of-function of phosphatidylethanolamine-binding protein.

Cowpea (Vigna unguiculata (L.) Walp) is one of the major food legume crops grown extensively in arid and semi-arid regions of the world. The determinate habit of cowpea has many advantages over the indeterminate and is well adapted to modern farming systems. Mutation breeding is an active research area to develop the determinate habit of cowpea. The present study aimed to develop new determinate habit mutants with terminal flowering (TFL) in locally well-adapted genetic backgrounds. Consequently, the seeds of popular cowpea cv P152 were irradiated with doses of gamma rays (200, 250, and, 300 Gy), and the M1 populations were grown. The M2 populations were produced from the M1 progenies and selected determinate mutants (TFLCM-1 and TFLCM-2) from the M2 generation (200 Gy) were forwarded up to the M5 generation to characterize the mutants and simultaneously they were crossed with P152 to develop a MutMap population. In the M5 generation, determinate mutants (80-81 days) were characterized by evaluating the TFL growth habit, longer peduncles (30.75-31.45 cm), erect pods (160°- 200°), number of pods per cluster (4-5 nos.), and early maturity. Further, sequencing analysis of the VuTFL1 gene in the determinate mutants and MutMap population revealed a single nucleotide transversion (A-T at 1196 bp) in the fourth exon and asparagine (N) to tyrosine (Y) amino acid change at the 143rd position of phosphatidylethanolamine-binding protein (PEBP). Notably, the loss of function PEPB with a higher confidence level modification of anti-parallel beta-sheets and destabilization of the protein secondary structure was observed in the mutant lines. Quantitative real-time PCR (qRT-PCR) analysis showed that the VuTFL1 gene was downregulated at the flowering stage in TFL mutants. Collectively, the insights garnered from this study affirm the effectiveness of induced mutation in modifying the plant's ideotype. The TFL mutants developed during this investigation have the potential to serve as a valuable resource for fostering determinate traits in future cowpea breeding programs and pave the way for mechanical harvesting.

Pilot-scale genome-wide association mapping in diverse sorghum germplasms identified novel genetic loci linked to major agronomic, root and stomatal traits.

This genome-wide association studies (GWAS) used a subset of 96 diverse sorghum accessions, constructed from a large collection of 219 accessions for mining novel genetic loci linked to major agronomic, root morphological and physiological traits. The subset yielded 43,452 high quality single nucleotide polymorphic (SNP) markers exhibiting high allelic diversity. Population stratification showed distinct separation between caudatum and durra races. Linkage disequilibrium (LD) decay was rapidly declining with increasing physical distance across all chromosomes. The initial 50% LD decay was ~ 5 Kb and background level was within ~ 80 Kb. This study detected 42 significant quantitative trait nucleotide (QTNs) for different traits evaluated using FarmCPU, SUPER and 3VmrMLM which were in proximity with candidate genes related and were co-localized in already reported quantitative trait loci (QTL) and phenotypic variance (R2) of these QTNs ranged from 3 to 20%. Haplotype validation of the candidate genes from this study resulted nine genes showing significant phenotypic difference between different haplotypes. Three novel candidate genes associated with agronomic traits were validated including Sobic.001G499000, a potassium channel tetramerization domain protein for plant height, Sobic.010G186600, a nucleoporin-related gene for dry biomass, and Sobic.002G022600 encoding AP2-like ethylene-responsive transcription factor for plant yield. Several other candidate genes were validated and associated with different root and physiological traits including Sobic.005G104100, peroxidase 13-related gene with root length, Sobic.010G043300, homologous to Traes_5BL_8D494D60C, encoding inhibitor of apoptosis with iWUE, and Sobic.010G125500, encoding zinc finger, C3HC4 type domain with Abaxial stomatal density. In this study, 3VmrMLM was more powerful than FarmCPU and SUPER for detecting QTNs and having more breeding value indicating its reliable output for validation. This study justified that the constructed subset of diverse sorghums can be used as a panel for mapping other key traits to accelerate molecular breeding in sorghum.

Plant Metabolomics: Current Initiatives and Future Prospects.

Plant metabolomics is a rapidly advancing field of plant sciences and systems biology. It involves comprehensive analyses of small molecules (metabolites) in plant tissues and cells. These metabolites include a wide range of compounds, such as sugars, amino acids, organic acids, secondary metabolites (e.g., alkaloids and flavonoids), lipids, and more. Metabolomics allows an understanding of the functional roles of specific metabolites in plants' physiology, development, and responses to biotic and abiotic stresses. It can lead to the identification of metabolites linked with specific traits or functions. Plant metabolic networks and pathways can be better understood with the help of metabolomics. Researchers can determine how plants react to environmental cues or genetic modifications by examining how metabolite profiles change under various crop stages. Metabolomics plays a major role in crop improvement and biotechnology. Integrating metabolomics data with other omics data (genomics, transcriptomics, and proteomics) provides a more comprehensive perspective of plant biology. This systems biology approach enables researchers to understand the complex interactions within organisms.

Exploring bixin from Bixa orellana L. seeds: quantification and in silico insights into its anti-cancer potential.

Bixin, the key pigment of Bixa orellana L., is an apo-carotenoid found in the seed arils. The present study aimed to quantitatively determine the bixin content of seeds and explore its anti-cancer activity through in silico studies. The bixin content from the seeds of the local genotype, TNMTP8, quantified by RP-HPLC was 4.58 mg per gram. The prediction of pharmacological activity suggested that bixin may serve as a BRAF, MMP9, TNF expression inhibitors, and TP53 expression enhancer. According to molecular docking analysis, bixin interacted with eight different skin cancer targets and had the lowest binding energy compared to the standard drug, 5-fluorouracil. The binding score between bixin and the targets ranged from -4.7 to -8.7 kcal/mol. The targets BRAF and SIRT3 interacted well with bixin, with binding energies as low as -8.3 and -8.7 kcal/mol, respectively. Hence, the dynamic behavior of these two docked complexes throughout a 500 ns trajectory run was investigated further. The Root Mean Square Deviation (RMSD), Root Mean Square Fluctuation (RMSF) values, and total contacts as a function of time recorded during scrutiny suggest that both complexes were stable. This was validated by post-molecular dynamics analysis using Molecular Mechanics Generalized Born Surface Area (MM-GBSA). Principal component analysis (PCA) was used to analyze the significant differences in motion exhibited by BRAF-Bixin and SIRT3-Bixin. The results showed that bixin is a promising source for potential treatment interventions in skin cancer therapies.Communicated by Ramaswamy H. Sarma.

Molecular characterization and SNP identification using genotyping-by-sequencing in high-yielding mutants of proso millet.

Proso millet (Panicummiliaceum L.) is a short-duration C4 crop that is drought tolerant and nutritionally rich and can grow well in marginal lands. Though the crop has many climate-resilient traits like tolerance to drought and heat, its yield is lower than that of common cereals like rice, wheat, and maize. Being an underutilized crop, the molecular resources in the crop are limited. The main aim of the present study was to develop and characterize contrasting mutants for yield and generate functional genomic information for the trait in proso millet. Gamma irradiation-induced mutant population was screened to identify high-yielding mutants, which were evaluated up to M4 generation. One mutant with a dense panicle and high yield (ATL_hy) and one with a lax panicle and low yield (ATL_ly) along with the wild type were sequenced using the genotyping-by-sequencing approach. The variants detected as single nucleotide polymorphisms (SNPs) and insertions-deletions (InDels) were annotated against the reference genome of proso millet. Bioinformatic analyses using the National Center for Biotechnology Information (NCBI) and UniProt databases were performed to elucidate genetic information related to the SNP variations. A total of 25,901, 30,335, and 31,488 SNPs, respectively, were detected in the wild type, ATL_hy mutants, and ATL_ly mutants. The total number of functional SNPs identified in high-yielding and low-yielding mutants was 84 and 171, respectively. Two functional SNPs in the high-yielding mutant (ATL_hy) and one in the low-yielding mutant (ATL_ly) corresponded to the gene coding for "E3 ubiquitin-protein ligase UPL7". Pathway mapping of the functional SNPs identified that two SNPs in ATL_ly were involved in the starch biosynthetic pathway coding for the starch synthase enzyme. This information can be further used in identifying genes responsible for various metabolic processes in proso millet and in designing useful genetic markers.

"Are we barking up the wrong tree? Too much emphasis on Cutibacterium acnes and ignoring other pathogens"- a study based on next-generation sequencing of normal and diseased discs.

BACKGROUND: The majority of literature on bacterial flora in the disc stands disadvantaged in utilizing traditional culture methods and targeting a single bacterium, Cutibacterium acnes. PURPOSE: Our objective was to document the diversity in the bacterial flora between normal and degenerated discs for shortlisting potential pathogens using next-generation genomic tools. STUDY DESIGN: Experimental case-control study. METHODS: Researchers employed 16S metagenome sequencing to profile bacterial diversity in magnetic resonance imaging normal healthy discs from brain-dead organ voluntary donors (n=20) and 40 degenerated disc samples harvested during surgery (Modic [MC]=20 and non-Modic [NMC]=20). The V3-V4 region was amplified using universal bacterial primers 341F and 806R, and the libraries were sequenced using Illumina NovoSeq 6000 platform. Statistical significance was set at bacteria with a minimum of 100 operational taxonomic unit (OTU) and present in at least 70% of the samples. The quality check-filtered reads were processed using the QIIME-2 pipeline. The OTU clustering and taxonomic classification were carried out for the merged reads using the Greengenes/SILVA reference database. Validation was done by identification of bacterial metabolites in samples using the liquid chromatography-mass spectrometry approach. RESULTS: Abundant bacteria differing widely in diversity, as evidenced by Alfa and Beta diversity analysis, were present in all control and degenerative samples. The number of bacterial genera was 27 (14-gram-positive: 13-gram-negative) in the control group, 23 (10-gram-positive: 11-gram-negative) in the Modic group, and 16 (11-gram-positive: 5-gram-negative) in the non-Modic group. In the Modic group, gram-negative bacteria OTUs were found to be predominant (more than 50% of the total bacteria identified), whereas in control and non-Modic groups the OTUs of gram-positive bacteria were predominant. Species-level analysis revealed an abundance of opportunistic gram-negative pathogens like Pseudomonas aeruginosa, Sphingomonos paucibacillus, and Ochrobactrum quorumnocens in the discs with Modic changes, more than in non-Modic discs. The presence of bacterial metabolites and quorum-sensing molecules like N-decanoyl-L-homoserine lactone, 6-hydroxynicotinic acid, 2-aminoacetophenone, 4-hydroxy-3-polyprenylbenzoate, PE (16:1(9Z)/18:0) and phthalic acid validated the colonization and cell-cell communication of bacteria in disc ruling out contamination theory. Cutibacterium acnes was not the predominant bacteria in any of the three groups of discs and in fact was in the 16th position in the order of abundance in the control discs (0.72%), seventh position in the Modic discs (1.41%), and 12th position (0.53%) in the non-Modic discs. CONCLUSION: This study identified a predominance of gram-negative bacteria in degenerated discs and highlights that Cutibacterium acnes may not be the only degeneration-causing bacteria. This may be attributed to the environment, diet, and lifestyle habits of the sample population. Though the study does not reveal the exact pathogen, it may pave the way for future studies on the subject. CLINICAL SIGNIFICANCE: These findings invite further investigation into causal relationships of bacterial profile with disc degeneration phenotypes as well as phenotype-driven clinical treatment protocols.

Altering Stomatal Density for Manipulating Transpiration and Photosynthetic Traits in Rice through CRISPR/Cas9 Mutagenesis.

Stomata regulates conductance, transpiration and photosynthetic traits in plants. Increased stomatal density may contribute to enhanced water loss and thereby help improve the transpirational cooling process and mitigate the high temperature-induced yield losses. However, genetic manipulation of stomatal traits through conventional breeding still remains a challenge due to problems involved in phenotyping and the lack of suitable genetic materials. Recent advances in functional genomics in rice identified major effect genes determining stomatal traits, including its number and size. Widespread applications of CRISPR/Cas9 in creating targeted mutations paved the way for fine tuning the stomatal traits for enhancing climate resilience in crops. In the current study, attempts were made to create novel alleles of OsEPF1 (Epidermal Patterning Factor), a negative regulator of stomatal frequency/density in a popular rice variety, ASD 16, using the CRISPR/Cas9 approach. Evaluation of 17 T0 progenies identified varying mutations (seven multiallelic, seven biallelic and three monoallelic mutations). T0 mutant lines showed a 3.7-44.3% increase in the stomatal density, and all the mutations were successfully inherited into the T1 generation. Evaluation of T1 progenies through sequencing identified three homozygous mutants for one bp insertion. Overall, T1 plants showed 54-95% increased stomatal density. The homozygous T1 lines (# E1-1-4, # E1-1-9 and # E1-1-11) showed significant increase in the stomatal conductance (60-65%), photosynthetic rate (14-31%) and the transpiration rate (58-62%) compared to the nontransgenic ASD 16. Results demonstrated that the genetic alterations in OsEPF1 altered the stomatal density, stomatal conductance and photosynthetic efficiency in rice. Further experiments are needed to associate this technology with canopy cooling and high temperature tolerance.

Targeting bone cancer with 4-Allylbenzene-1,2-diol purified from Piper betle L.: an in silico and cytotoxicity scrutiny.

Piper betle L., a well-known medicinal plant with rich source of bioactive compounds, is widely used in several therapeutics. The present study was performed to scrutinize the anti-cancer potential of compounds P. betle petiole by means of in silico studies, purification of 4-Allylbenzene-1,2-diol from petioles and assessing its cytotoxicity on bone cancer metastasis. Subsequent to SwissADME screening, 4-Allylbenzene-1,2-diol and Alpha terpineol were chosen for molecular docking together with eighteen approved drugs against fifteen important bone cancer targets accompanied with molecular dynamics simulation studies. 4-Allylbenzene-1,2-diol was found to be multi-targeting, interacted effectively with all targets, particularly exhibited good stability with MMP9 and MMP2 during molecular dynamics simulations and Molecular Mechanics- Generalized Born and Surface Area (MM-GBSA) analysis using Schrodinger. Later, the compound was isolated, purified and the cytotoxicity studies on MG63 bone cancer cell lines confirmed the cytotoxicity nature (75.98% at 100 µg/ml concentration). The results demonstrated the compound as a matrix metalloproteinase inhibitor, and therefore 4-Allylbenzene-1,2-diol may possibly be prescribed in targeted therapy for alleviating the bone cancer metastasis upon further wet lab experimental validations.Communicated by Ramaswamy H. Sarma.

Marker Assisted Introgression of Resistance Genes and Phenotypic Evaluation Enabled Identification of Durable and Broad-Spectrum Blast Resistance in Elite Rice Cultivar, CO 51.

Across the globe, rice cultivation is seriously affected by blast disease, caused by Magnaporthe oryzae. This disease has caused heavy yield loss to farmers over the past few years. In this background, the most affordable and eco-friendly strategy is to introgress blast-resistant genes from donors into elite rice cultivars. However, it is not only challenging to evolve such resistance lines using conventional breeding approaches, but also a time-consuming process. Therefore, the marker-assisted introduction of resistance genes has been proposed as a rapid strategy to develop durable and broad-spectrum resistance in rice cultivars. The current study highlights the successful introgression of a blast resistance gene, i.e., Pi9, into CO 51, an elite rice cultivar which already has another resistance gene named Pi54. The presence of two blast resistance genes in the advanced backcross breeding materials (BC2F2:3) was confirmed in this study through a foreground selection method using functional markers such as NBS4 and Pi54MAS. The selected positive introgressed lines were further genotyped for background selection with 55 SSR markers that are specific to CO 51. Consequently, both Pi9 as well as Pi54 pyramided lines, with 82.7% to 88.1% of the recurrent parent genome recovery, were identified and the selected lines were evaluated under hotspot. The analysis outcomes found that both the lines possessed a high level of resistance against blast disease during the seedling stage itself. In addition to this, it was also noticed that the advanced breeding rice lines that carry Pi9 + Pi54 were effective in nature and exhibited a higher degree of resistance against blast disease compared to the lines that were introgressed with a single blast resistance gene. Thus, the current study demonstrates a rapid and a successful introgression and pyramiding of two blast resistance genes, with the help of markers, into a susceptible yet high-yielding elite rice cultivar within a short period of time. Those gene pyramided rice lines can be employed as donors to introgress the blast-resistant genes in other popular susceptible cultivars.

Comparative Metabolomic Profiling of Horse Gram (Macrotyloma uniflorum (Lam.) Verdc.) Genotypes for Horse Gram Yellow Mosaic Virus Resistance.

Horse gram (Macrotyloma uniflorum (Lam.) Verdc.) is an under-utilized legume grown in India. It is a good source of protein, carbohydrates, dietary fiber, minerals, and vitamins. We screened 252 horse gram germplasm accessions for horse gram yellow mosaic virus resistance using the percent disease index and scaling techniques. The percentage values of highly resistant, moderately resistant, moderately susceptible, susceptible, and highly susceptible were 0.34, 13.89, 38.89, 46.43, and 0.34, respectively. Repetitive trials confirmed the host-plant resistance levels, and yield loss was assessed. The present disease index ranged from 1.2 to 72.0 and 1.2 to 73.0 during the kharif and rabi seasons of 2018, respectively. The maximum percent yield loss was noticed in the HS (75.0 -89.4), while HR possessed the minimum (1.2-2.0). The methanolic leaf extracts of highly resistant and highly susceptible genotypes with essential controls were subjected to gas chromatography-mass spectrometry analysis. Differential accumulation of metabolites was noticed, and a total of 81 metabolites representing 26 functional groups were identified. Both highly resistant and susceptible genotypes harbored eight unique classes, while ten biomolecules were common. The hierarchical cluster analysis indicated a distinct metabolite profile. Fold change in the common metabolites revealed an enhanced accumulation of sugars, alkanes, and carboxylic acids in the highly resistant genotype. The principal component analysis plots explained 93.7% of the variation. The metabolite profile showed a significant accumulation of three anti-viral (octadecanoic acid, diphenyl sulfone, and 2-Aminooxazole), one insecticidal (9,10-Secocholesta-5,7,10(19)-triene-3,24,25-triol), one antifeedant (cucurbitacin B), and six metabolites with unknown biological function in the highly resistant genotype.

Identification of Newer Stable Genetic Sources for High Grain Number per Panicle and Understanding the Gene Action for Important Panicle Traits in Rice.

Rice is an important food crop extensively cultivated worldwide, and rice's grain yield should be improved to meet future food demand. Grain number per panicle is the main trait that determines the grain yield in rice, and other panicle-related traits influence the grain number. To study the genetic diversity, 50 diverse Indian-origin germplasm were evaluated for grain number per panicle and other panicle traits for two consecutive seasons (Rabi 2019 and Kharif 2020). The maximum genotypic and phenotypic coefficient of variation was obtained for the number of spikelets and filled grains per panicle. The genotypes were grouped into eight clusters with Mahalanobis' D2 analysis and six groups using Principal component analysis. Based on, per se, performance for grain number per panicle and genetic distances, six parents were selected and subjected to full diallel mating. The genotypes CB12132, IET 28749, and BPT 5204 were the best general combiners for the number of filled grains per panicle and other panicle branching traits, viz., the number of primary and secondary branches per panicle. The hybrid BPT 5204 × CB 12132 identified as the best specific combination for most of the studied panicle traits. The additive gene effects were high for the number of filled grains per panicle, the number of primary branches, and secondary branches, whereas non-additive gene action was high for the number of productive tillers and grain yield per plant. The information obtained from this study will be useful in rice breeding programs to improve panicle traits, especially the grain number, which would result in higher grain yield.

Bacteria in human lumbar discs - subclinical infection or contamination? Metabolomic evidence for colonization, multiplication, and cell-cell cross-talk of bacteria.

BACKGROUND CONTEXT: The accumulating evidence associating sub-clinical infection with disc degeneration (DD) and the controversy of contamination versus infection mandates a further understanding of the microbial activity in the disc and host-microbiome interaction. PURPOSE: To utilize a novel approach of metabolomics to probe the presence of bacterial metabolites involved in colonization, survival, and replication in human lumbar intervertebral discs (LIVD). STUDY DESIGN: An observational case-control study. PATIENT SAMPLE: Nucleus pulposus from the LIVD of three brain-dead voluntary organ donors (MRI normal and classified as controls) and of three patients undergoing surgery for disc degeneration (DD) (cases) were utilized. METHODS: Untargeted metabolite profiling was carried out in six discs (3-controls and 3-cases) after extraction using methanol: acetonitrile: water (2:2:1) solvent system and acquired through HPLC-MS/MS platform using C18 reversed-phase column. From the total IVD metabolome, microbial metabolites were filtered by mapping against HMDB, ChEBI, SigMol, Siderophore database, ecdmb database, and PaMet databases. The biological functions of the metabolites were then studied by MSEA pipeline from Metaboanalyst, and the enrichment ratio, p-value, and Variably Importance Projection scores of the metabolites were calculated. Degeneration responsive changes in the abundance of the microbial metabolites were calculated based on the peak intensities between the control and cases. RESULTS: Mass spectrometry identified a total of 17601 and 15003 metabolites, respectively, in the control and degenerated discs. Preliminary mapping of the above metabolites against HMDB indicated the multiple sources, and of these, 64 metabolites were of microbial origin, accounting for 1.6% of the total IVD metabolome. Principle Component Analysis and Orthogonal Partial Least Square-Discriminant Analysis (OPLS-DA) showed distinct clustered patterns between control and disc degene`ration, indicating a strong variation in concentration, peak, and spectral values of the 64 metabolites between controls and cases. After the exclusion of metabolites that were also associated with humans, drugs, and food, 39 metabolites specific to bacteria were isolated. Nine were primary metabolites related to bacterial growth and survival, and the remaining 30 were secondary metabolites related to different environmental stress response activities. The three significant pathways (p<.001) which were predominant in the bacterial metabolites were autoinducer-2 biosynthesis, peptidoglycan biosynthesis, and chorismate pathway. In addition, a significant fold change of >1.0 was found for nine metabolites which included (S)-14-Methyilhexadecanoic acid related to P. acnes, 9-OxoODE, and 13-OxoODE related to gut flora, vibriobactin - a siderophore, tuberculosinol and iso-tuberculosinol, virulence factors of M. tuberculosis. There was also upregulation of Autoinducer- 2, an important "Quorum sensing molecule" involved in bacterial cross-talk. CONCLUSION: We identified several bacterial-specific metabolites participating in bacterial growth, survival, and cross-talk pathways. These were found in both groups but up-regulated in degenerated discs. The presence of Quorum sensing molecules and cell-cell interactions provides firm proof of colonization and growth. These findings indicate that the bacterial presence may not be mere contamination but could be colonization with a possible role in infection-mediated inflammation in DD. CLINICAL SIGNIFICANCE: Proof of subclinical infection as an initiator of DD and documentation of exact germ and drug sensitivity will change the way millions of patients with non-specific low back pain (NSLBP) are treated across the world.

Whole genome sequencing of ASD 16 and ADT 43 to identify predominant grain size and starch associated alleles in rice.

BACKGROUND: The rice cultivars ASD 16 and ADT 43 are the most popular high-yielding Indica rice cultivars in southern India. Despite their popularity very little is known about their genetic basis due to lack of studies on the complete genome. In the current study, efforts were made to identify alleles and SNP markers that differentiate the two contrasting rice genotypes, ASD 16 and ADT 43 for grain shape and starch content. METHODS AND RESULTS: The complete genome of bold grain ASD 16 and slender grain ADT 43 were sequenced via Illumina's paired-end sequencing and the reads obtained were mapped to the Oryza sativa Indica Group cultivar 93-11 reference genome. The grain size of rice is controlled by Quantitative Trait Loci (QTL) that has a robust effect on grain yield and quality. To gain insight into genes that controlling grain size and starch content, an in-silico analysis was performed by taking into account of 72 grain elongation and starch biosynthesis genes. The identified alleles were further validated in the whole genome sequencing data of 32 bold grain and 25 slender grain varieties that were retrieved from the 3 K rice genome project. CONCLUSION: An "A to G" polymorphism leading to SER 74 PRO was identified at the CDS position 220 of the An-1 gene, encoding bHLH domain-containing protein that regulates awn formation and increase in grain length. The non-synonymous substitutions such as A545C variant leading PHE 182 CYS in ADP Glucose Pyrophosphorylase large subunit IV (AGPL4) and C3094G variant leading to VAL 1032 LEU in Starch synthase IIIb (OsSSIIIb) were also identified in the starch biosynthesis genes. These identified allelic variants may contribute to the crop improvement programs in rice.

Comparative metagenomic analysis of human intervertebral disc nucleus pulposus and cartilaginous end plates.

Study design: The diversity of microflora inhabiting endplate (EP) and nucleus pulposus (NP) tissues of human intervertebral disc (IVD) was profiled through NGS-supported 16S rRNA amplicon sequencing. Sixteen EP and their corresponding NP were excised from the brain-dead voluntary organ donors with no clinical history of low back pain, and 12 herniated and 8 degenerated NP tissues isolated from the patients undergoing spinal surgery were subjected to study the alteration in the microbial diversity. Objectives: To understand in normal IVD, whether the colonization of bacteria to the NP is through the EP in discs with intact annulus fibrosus. To identify significantly differing microbial population(s) between normal and diseased IVD (NP). Background of the study: There is increasing evidence for subclinical infection by fastidious low, growing bacteria to be a cause of disc degeneration. Although the presence of bacteria in NP has been reported well in literature, the source of bacteria is not clearly proved as the disc is avascular in healthy condition. Documentation of similar bacterial populations in the EP and NP may add proof that bacterial inoculation of NP occurs via the EP. Materials and methods: Sixteen EP and their corresponding NP excised from brain-dead voluntary organ donors with no history of back pain and 20 diseased discs collected from patients undergoing microdiscectomy/fusion surgery were used for profiling microbiome through 16S rRNA amplicon sequencing using primers specific for V1-V9 hypervariable regions. Changes in bacterial diversity and abundance were analysed to identify the key microbial populations in normal IVD NP and EP tissues and those significantly altered in diseased IVD (NP). Results: NP and EP shared a similar spectrum of microbiome but with varying abundance. The five dominant phyla identified were Proteobacteria, Firmicutes, Actinobacteria, OD1, and Bacteroidetes. Proteobacteria was found to be the most abundant phyla in both NP (62%) and EP (53%) of the normal IVD. This was followed by Firmicutes (16%), Actinobacteriota (11%), OD1 (Parcubacteria) (7.6%), and Bacteroidetes (2%) in NP and Firmicutes (23.4%), OD1 (Parcubacteria) (17.6%), Actinobacteriota (2.8%), and Bacteroidetes (2.6%) in EP, respectively. Under diseased conditions, Proteobacteria (68%) was dominant when compared with other phyla. However, there was no significant difference in the abundance of Proteobacteria between the normal and diseased discs. Interestingly, the other dominant phyla such as Firmicutes (Normal-NP: 16.2%; Diseased-NP: 4.02%) and Actinobacteria (Normal-NP: 11%; Diseased-NP: 0.99%) showed a significant reduction in degenerated discs. To understand the key microbial populations that are significantly altered during disease, correlation analysis was performed among the three phyla, which revealed a negative correlation in the ratio of Actinobacteria + Firmicutes vs. Proteobacteria (p = 0.001) in DD. Conclusion: Results of our study clearly demonstrated a similar bacterial diversity but with varying abundance between the EP and NP, suggesting the existence of the endplate-nucleus pulposus axis in the normal IVD microbiome. Further, our results have indicated that the changes in the abundance of Actinobacteria + Firmicutes vs. Proteobacteria during DDD need further investigation.

Complete Genome Sequence Analysis of Bacillus subtilis Bbv57, a Promising Biocontrol Agent against Phytopathogens.

Plant growth-promoting rhizobacteria (PGPR) are a group of root-associated beneficial bacteria emerging as one of the powerful agents in sustainable plant disease management. Among the PGPR, Bacillus sp. has become a popular biocontrol agent for controlling pests and the diseases of several crops of agricultural and horticultural importance. Understanding the molecular basis of the plant growth-promoting and biocontrol abilities of Bacillus spp. will allow us to develop multifunctional microbial consortia for sustainable agriculture. In our study, we attempted to unravel the genome complexity of the potential biocontrol agent Bacillus subtilis Bbv57 (isolated from the betelvine's rhizosphere), available at TNAU, Coimbatore. A WGS analysis generated 26 million reads, and a de novo assembly resulted in the generation of 4,302,465 bp genome of Bacillus subtilis Bbv57 containing 4363 coding sequences (CDS), of which 4281 were functionally annotated. An analysis of 16S rRNA revealed its 100% identity to Bacillus subtilis IAM 12118. A detailed data analysis identified the presence of >100 CAZymes and nine gene clusters involved in the production of secondary metabolites that exhibited antimicrobial properties. Further, Bbv57 was found to harbor 282 unique genes in comparison with 19 other Bacillus strains, requiring further exploration.

Comparative RNA-Seq analysis unravels molecular mechanisms regulating therapeutic properties in the grains of traditional rice Kavuni.

Developing rice varieties with enhanced levels of functional bioactives is an important intervention for achieving food and nutritional security in Asia where rice is the staple food and Type II diabetes incidences are higher. The present study was aimed at dissecting out the molecular events underlying the accumulation of bio active compounds in pigmented traditional rice Kavuni. Comparative transcriptome profiling in the developing grains of Kavuni and a white rice variety ASD 16 generated 37.7 and 29.8 million reads respectively. Statistical analysis identified a total of 9177 exhibiting significant differential expression (DEGs) between the grains of Kavuni and ASD 16. Pathway mapping of DEGs revealed the preferential up-regulation of genes involved in the biosynthesis of amylose and dietary fibres in Kavuni accounting for its low glycemic index (GI). Transcripts involved in the biosynthesis of carotenoids, flavonoids, anthocyanins, phenolic acids and phenylpropanoids were also found to be up-regulated in the grains of Kavuni. This study identified up-regulation of key transcripts involved in the accumulation of phenolic acids having potential for inhibiting major hydrolytic enzymes α-amylase and α-glucosidase and thus accounting for the slow digestibility leading to low GI. Overall, this study has identified molecular targets for the genetic manipulation of anti-diabetic and anti-oxidant traits in rice.

Are Modic changes 'Primary infective endplatitis'?-insights from multimodal imaging of non-specific low back pain patients and development of a radiological 'Endplate infection probability score'.

PURPOSE: To probe the pathophysiological basis of Modic change (MC) by multimodal imaging rather than by MRI alone. METHODS: Nineteen radiological signs found in mild infections and traumatic endplate fractures were identified by MRI and CT, and by elimination, three signs unique to infection and trauma were distilled. By ranking the Z score, radiological 'Endplate Infection Probability Score' (EIPS) was developed. The score's ability to differentiate infection and traumatic endplate changes (EPC) was validated in a fresh set of 15 patients each, with documented infection and trauma. The EIPS, ESR, CRP, and Numeric Pain Rating Scale (NRS) were then compared between 115 patients with and 80 patients without MC. RESULTS: The EIPS had a confidence of 66.4%, 83% and, 100% for scores of 4, 5 and, 6, respectively, for end plate changes suggesting infection. The mean EIPS was 4.85 ± 1.94 in patients with Modic changes compared to - 0.66 ± 0.49 in patients without Modic changes (p < 0.001). Seventy-eight (67.64%) patients with MC had a score of 6, indicating high infection possibility. There was a difference in the NRS (p < 0.01), ESR (p = 0.05), CRP (p < 0.01), and type of pain (p < 0.01) between patients with and without MC. CONCLUSION: Multimodal imaging showed many radiological signs not easily seen in MRI alone and thus missed in Modic classification. There were distinct radiological differences between EPCs of trauma and infection which allowed the development of an EIPS. The scores showed that 67.64% of our study patients with Modic changes had EPCs resembling infection rather than trauma suggesting the possibility of an infective aetiology and allowing us to propose an alternate theory of 'Primary Endplatitis'.

Walking through crossroads-rice responses to heat and biotic stress interactions.

Rice, the most important source of calories for humans is prone to severe yield loss due to changing climate including heat stress. Additionally, rice encounters biotic stresses in conjunction with heat stress, which exacerbates the adverse effects, and exponentially increase such losses. Several investigations have identified biotic and heat stress-related quantitative trait loci (QTLs) that may contribute to improved tolerance to these stresses. However, a significant knowledge gap exists in identifying the genomic regions imparting tolerance against combined biotic and heat stress. Hereby, we are presenting a conceptual meta-analysis identifying genomic regions that may be promising candidates for enhancing combined biotic and heat stress tolerance in rice. Fourteen common genomic regions were identified along chromosomes 1, 2, 3, 4, 6, 10 and 12, which harbored 1265 genes related to heat stress and defense responses in rice. Further, the meta expression analysis revealed 24 differentially expressed genes (DEGs) involved in calcium-mediated stress signaling including transcription factors Myb, bHLH, ROS signaling, molecular chaperones HSP110 and pathogenesis related proteins. Additionally, we also proposed a hypothetical model based on GO and MapMan analysis representing the pathways intersecting heat and biotic stresses. These DEGs can be potential candidate genes for improving tolerance to combined biotic and heat stress in rice. We present a framework highlighting plausible connecting links (QTLs/genes) between rice response to heat stress and different biotic factors associated with yield, that can be extended to other crops.