Significance of the prime factors regulating arsenic toxicity and associated health risk: a hypothesis-based investigation in a critically exposed population of West Bengal, India.

The suffering from arsenic toxicity is a long-standing concern in Asian countries. The role of the key factors (arsenic intake, age and sex) regulating arsenic toxicity is aimed to evaluate for a severely exposed population from Murshidabad district, West Bengal. Mean arsenic concentrations in drinking water supplied through tube well, Sajaldhara treatment plant and pipeline were observed as 208, 27 and 54 µg/l, respectively. Urinary arsenic concentration had been observed as < 3-42.1, < 3-56.2 and < 3-80 µg/l in children, teenagers and adults, respectively. Mean concentrations of hair and nail arsenic were found to be 0.84 and 2.38 mg/kg; 3.07 and 6.18 mg/kg; and 4.41 and 9.07 mg/kg, respectively, for the studied age-groups. Water arsenic was found to be associated with hair and nail (r = 0.57 and 0.60), higher than urine (r = 0.37). Arsenic deposition in biomarkers appeared to be dependent on age; however, it is independent of sex. Principal component analysis showed a direct relationship between dietary intake of arsenic and chronic biomarkers. Nail was proved as the most fitted biomarker of arsenic toxicity by Dunn's post hoc test. Monte Carlo sensitivity analysis and cluster analysis showed that the most significant factor regulating health risk is 'concentration of arsenic' than 'exposure duration', 'body weight' and 'intake rate'. The contribution of arsenic concentration towards calculated health risk was highest in teenagers (45.5-61.2%), followed by adults (47.8-49%) and children (21-27.6%). Regular and sufficient access to arsenic-safe drinking water is an immediate need for the affected population.

Nematode Genome Announcement: A Draft Genome of Seed Gall Nematode, Anguina tritici.

Anguina tritici is the first plant-parasitic nematode described in literature, dating back to the year 1743. It is responsible for causing earcockle (seed gall) and tundu diseases in wheat and rye. Notably, this nematode has been observed to survive in an anhydrobiotic state for up to 32 years within wheat seed galls. These exceptional characteristics have inspired the sequencing of the A. tritici genome. In this study, we present the initial draft genome of A. tritici, obtained using the Illumina MiSeq platform with coverage of 60-fold. The genome is estimated to have a size of 164 Mb and comprises 39,965 protein-coding genes, exhibiting a GC content of 39.1%. The availability of this genome data will serve as a foundation for future functional biological investigations, particularly for genes whose functions remain unknown to this day.

Structural and functional analysis of CCT family genes in pigeonpea.

BACKGROUND: Pigeonpea (Cajanus cajan L.) is a photoperiod-sensitive short-day plant. Understanding the flowering-related genes is critical to developing photoperiod insensitive cultivars. METHODS: The CCT family genes were identified using 'CCT DOMAIN PROTEIN' as a keyword and localized on the chromosomes using the BLAST search option available at the LIS database. The centromeric positions were identified through BLAST search using the centromeric repeat sequence of C. cajan as a query against the chromosome-wise FASTA files downloaded from the NCBI database. The CCT family genes were classified based on additional domains and/or CCT domains. The orthologous and phylogenetic relationships were inferred using the OrthoFinder and MEGA 10.1 software, respectively. The CCT family genes' expression level in photoperiod-sensitive and insensitive genotypes was compared using RNA-seq data and qRT-PCR analysis. RESULTS: We identified 33 CCT family genes in C. cajan distributed on ten chromosomes and nine genomic scaffolds. They were classified into CMF-type, COL-type, PRR-type, and GTCC- type. The CCT family genes of legumes exhibited an extensive orthologous relationship. Glycine max showed the maximum similarity of CCT family genes with C. cajan. The expression analysis of CCT family genes using photoperiod insensitive (ICP20338) and photoperiod sensitive (MAL3) genotypes of C. cajan demonstrated that CcCCT4 and CcCCT23 are the active CONSTANS in ICP20338. In contrast, only CcCCT23 is active in MAL3. CONCLUSION: The CCT family genes in C. cajan vary considerably in structure and domain types. They are maximally similar to soybean's CCT family genes. The differential photoperiod response of pigeonpea genotypes, ICP20338 and MAL3, is possibly due to the difference in the number and types of active CONSTANS in them.

Rice seed (IR64) priming with potassium humate for improvement of seed germination, seedling growth and antioxidant defense system under arsenic stress.

The intolerable levels of arsenic (As) in groundwater and its application in rice cultivation are continuously affecting the rice production in Ganga-Meghna-Brahmaputra (GMB) plain. The reduced germination and plant growth rates under excessive As stress ultimately lead to lower yield. To mitigate this concerning issue, the present study was carried out to evaluate the potential of K-humate priming on seed germination and plant growth under As stress. Seeds were primed with 100 mg/l K-humate for 12 h prior to germination. The germination percentages in unprimed seeds were 65 ±â€¯5.0% and 58.3 ±â€¯7.6% under stress level of 50 µM AsV and 50 µM AsIII, respectively. However, germination percentage in K-humate primed seeds were 75 ±â€¯5.0% and 68.3 ±â€¯2.9% under AsV and AsIII stress, respectively. The vigour index I (VG I) and vigour index II (VG II) recorded on 12 DAS (days after seeding) were also increased by 1.47 and 1.51 fold, respectively with K-humate supplementation under As stress. Detrimental effects of AsIII on seed germination, seedling growth and other physiological parameters were more suppressive than AsV. Application of K-humate not only improved seed germination, seedling growth and nutrient uptake but also decreased the oxidative stress markers and antioxidant activities by minimizing As uptake and translocation in the seedlings.

Health effect and risk assessment of the populations exposed to different arsenic levels in drinking water and foodstuffs from four villages in arsenic endemic Gaighata block, West Bengal, India.

Health exposure and perception of risk assessment have been evaluated on the populations exposed to different arsenic levels in drinking water (615, 301, 48, 20 µg/l), rice grain (792, 487, 588, 569 µg/kg) and vegetables (283, 187, 238, 300 µg/kg) from four villages in arsenic endemic Gaighata block, West Bengal. Dietary arsenic intake rates for the studied populations from extremely highly, highly, moderately, and mild arsenic-exposed areas were 56.03, 28.73, 11.30, and 9.13 µg/kg bw/day, respectively. Acute and chronic effects of arsenic toxicity were observed in ascending order from mild to extremely highly exposed populations. Statistical interpretation using 'ANOVA' proves a significant relationship between drinking water and biomarkers, whereas "two-tailed paired t test" justifies that the consumption of arsenic-contaminated dietary intakes is the considerable pathway of health risk exposure. According to the risk thermometer (SAMOE), drinking water belongs to risk class 5 (extremely highly and highly exposed area) and 4 (moderately and mild exposed area) category, whereas rice grain and vegetables belong to risk class 5 and 4, respectively, for all the differently exposed populations. The carcinogenic (ILCR) and non-carcinogenic risks (HQ) through dietary intakes for adults were much higher than the recommended threshold level, compared to the children. Supplementation of arsenic-safe drinking water and nutritional food is strictly recommended to overcome the severe arsenic crisis.

Arsenic toxicity in livestock growing in arsenic endemic and control sites of West Bengal: risk for human and environment.

The present study aims to estimate geochemical arsenic toxicity in the domestic livestock and possible risk for human and environment caused by them. Daily dietary arsenic intake of an exposed adult cow or bull is nearly 4.56 times higher than control populace and about 3.65 times higher than exposed goats. Arsenic toxicity is well exhibited in all the biomarkers through different statistical interpretations. Arsenic bioconcentration is faster through water compared to paddy straw and mostly manifested in faeces and tail hair in cattle. Cow dung and tail hair are the most pronounced pathways of arsenic biotransformation into environment. A considerable amount of arsenic has been observed in animal proteins such as cow milk, boiled egg yolk, albumen, liver and meat from the exposed livestock. Cow milk arsenic is mostly accumulated in casein (83%) due to the presence of phosphoserine units. SAMOE-risk thermometer, calculated for the most regularly consumed foodstuffs in the area, shows the human health risk in a distinct order: drinking water > rice grain > cow milk > chicken > egg > mutton ranging from class 5 to 1. USEPA health risk assessment model reveals more risk in adults than in children, subsisting severe cancer risk from the foodstuffs where the edible animal proteins cannot be ignored. Therefore, the domestic livestock should be urgently treated with surface water, while provision of both arsenic-free drinking water and nutritional supplements is mandatory for the affected human population to overcome the severe arsenic crisis situation.

Non-coding RNAs having strong positive interaction with mRNAs reveal their regulatory nature during flowering in a wild relative of pigeonpea (Cajanus scarabaeoides).

In higher plants, flower development is a result of crosstalk between many factors like photoperiod, vernalization, hormone concentration, epigenetic modification etc. and is also regulated by non-coding RNAs (ncRNAs). In the present study, we are reporting the involvement of long non-coding RNAs (lncRNAs) and miRNAs during the process of flower development in Cajanus scarabaeoides, an important wild relative of pigeonpea. The transcriptome of floral and leaf tissues revealed a total of 1672 lncRNAs and 57 miRNAs being expressed during flower development. Prediction analysis of identified lncRNAs showed that 1593 lncRNAs were targeting 3420 mRNAs and among these, 98 were transcription factors (TFs) belonging to 48 groups. All the identified 57 miRNAs were novel, suggesting their genera specificity. Prediction of the secondary structure of lncRNAs and miRNAs followed by interaction analysis revealed that 199 lncRNAs could interact with 47 miRNAs where miRNAs were acting in the root of interaction. Gene Ontology of the ncRNAs and their targets showed the potential role of lncRNAs and miRNAs in the flower development of C. scarabaeoides. Among the identified interactions, 17 lncRNAs were endogenous target mimics (eTMs) for miRNAs that target flowering-related transcription factors. Expression analysis of identified transcripts revealed that higher expression of Csa-lncRNA_1231 in the bud sequesters Csa-miRNA-156b by indirectly mimicking the miRNA and leading to increased expression of flower-specific SQUAMOSA promoter-binding protein-like (SPL-12) TF indicating their potential role in flower development. The present study will help in understanding the molecular regulatory mechanism governing the induction of flowering in C. scarabaeoides.

An extensive review of arsenic dynamics and its distribution in soil-aqueous-rice plant systems in south and Southeast Asia with bibliographic and meta-data analysis.

Millions of people worldwide are affected by arsenic (As) contamination, particularly in South and Southeast Asian countries, where large-scale dependence on the usage of As-contaminated groundwater in drinking and irrigation is a familiar practice. Rice (Oryza sativa) cultivation is commonly done in South and Southeast Asian countries as a preferable crop which takes up more As than any other cereals. The present article has performed a scientific meta-data analysis and extensive bibliometric analysis to demonstrate the research trend in global rice As contamination scenario in the timeframe of 1980-2023. This study identified that China contributes most with the maximum number of publications followed by India, USA, UK and Bangladesh. The two words 'arsenic' and 'rice' have been identified as the most dominant keywords used by the authors, found through co-occurrence cluster analysis with author keyword association study. The comprehensive perceptive attained about the factors affecting As load in plant tissue and the nature of the micro-environment augment the contamination of rice cultivars in the region. This extensive review analyses soil parameters through meta-data regression assessment that influence and control As dynamics in soil with its further loading into rice grains and presents that As content and OM are inversely related and slightly correlated to the pH increment of the soil. Additionally, irrigation and water management practices have been found as a potential modulator of soil As concentration and bioavailability, presented through a linear fit with 95% confidence interval method.

Appraisal of treated drinking water quality from arsenic removal units in West Bengal, India: Approach on safety, efficiency, sustainability, future health risk and socioeconomics.

The present study depicts the true failed scenario of the arsenic (As) removal units (ARU) in West Bengal by evaluating their treated water quality. Annual As removal efficiency of the 12 studied ARUs range between 35.2% and 82.6%. A comprehensive physico-chemical parameters and trace elements analysis find almost 25% and 16.7% of treated drinking water samples with poor water quality index (WQI) and high heavy metal evaluation index (HEI), respectively. The pond-based water treatment plant maintains the production of continuous As-safe water with a range between 60.2% and 66.7% due to its high Fe/As ratio. It's a discontent concluding the treated drinking water of the groundwater based-ARUs were observed with sufficient As mediated cancer risk (3 ×10-3). The non-cancer risk (HQ) of As is safe for the surface water treatment plant (0.38), whereas it is threatening for the groundwater based-ARUs (7.44). However, the drinking water samples are safe in view of HQ from the other trace elements like Hg, Al, Cd, Cr, Pb, F- and NO3-. Small scale ARU could be a feasible mitigation strategy in reducing the As menace in the long run if the plants are maintained correctly. Nevertheless, surface treated water is the most sustainable solution as withdrawal of groundwater for drinking purpose is not a viable practice.

Genomic survey of high-throughput RNA-Seq data implicates involvement of long intergenic non-coding RNAs (lincRNAs) in cytoplasmic male-sterility and fertility restoration in pigeon pea.

BACKGROUND: Long-intergenic non-coding RNAs (lincRNAs) originate from intergenic regions and have no coding potential. LincRNAs have emerged as key players in the regulation of various biological processes in plant development. Cytoplasmic male-sterility (CMS) in association with restorer-of-fertility (Rf) systems makes it a highly reliable tool for exploring heterosis for producing commercial hybrid seeds. To date, there have been no reports of lincRNAs during pollen development in CMS and fertility restorer lines in pigeon pea. OBJECTIVE: Identification of lincRNAs in the floral buds of cytoplasmic male-sterile (AKCMS11) and fertility restorer (AKPR303) pigeon pea lines. METHODS: We employed a computational approach to identify lincRNAs in the floral buds of cytoplasmic male-sterile (AKCMS11) and fertility restorer (AKPR303) pigeon pea lines using RNA-Seq data. RESULTS: We predicted a total of 2145 potential lincRNAs of which 966 were observed to be differentially expressed between the sterile and fertile pollen. We identified, 927 cis-regulated and 383 trans-regulated target genes of the lincRNAs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the target genes revealed that these genes were specifically enriched in pathways like pollen and pollen tube development, oxidative phosphorylation, etc. We detected 23 lincRNAs that were co-expressed with 17 pollen-related genes with known functions. Fifty-nine lincRNAs were predicted to be endogenous target mimics (eTMs) for 25 miRNAs, and found to be associated with pollen development. The, lincRNA regulatory networks revealed that different lincRNA-miRNA-mRNA networks might be associated with CMS and fertility restoration. CONCLUSION: Thus, this study provides valuable information by highlighting the functions of lincRNAs as regulators during pollen development in pigeon pea and utilization in hybrid seed production.

Investigating spatial distribution of fluoride in groundwater with respect to hydro-geochemical characteristics and associated probabilistic health risk in Baruipur block of West Bengal, India.

Fluoride (F-) enrichment in groundwater of the lower Gangetic plain in West Bengal, India is a major concern. Fluoride contamination and its toxicity were reported earlier in this region; however, limited evidence was available on the precise site of contamination, hydro-geochemical attributions of F- mobilization and probabilistic health risk caused by fluoridated groundwater. The present study addresses the research gap by exploring the spatial distribution and physico-chemical parameters of fluoridated groundwater along with the depth-wise sedimental distribution of F-. Approximately, 10 % of the groundwater samples (n = 824) exhibited high F- ≥ 1.5 mg/l from 5, out of 19 gram-panchayats and Baruipur municipality area and the maximum F- was observed in Dhapdhapi-II gram-panchayat with 43.7 % of samples showed ≥1.5 mg/l (n = 167). The distribution patterns of cations and anions in fluoridated groundwater were Na+ > Ca2+ > Mg2+ > Fe > K+ and Cl- > HCO3- > SO42- > CO32- > NO3- > F-. Different statistical models like Piper and Gibbs diagram, Chloro Alkaline plot, Saturation index were applied to better understand the hydro-geochemical characteristics for F- leaching in groundwater. Fluoridated groundwater is of Na-Cl type which implies strong saline character. The intermediate zone between evaporation and rock dominance area controls F- mobilization along with ion-exchange process occurring between groundwater and host silicate mineral. Furthermore, saturation index proves geogenic activities related to groundwater F- mobilization. All cations present in sediment samples are closely interlinked with F- in the depth range of 0-18.3 m. Mineralogical analyses revealed that muscovite is the most responsible mineral for F- mobilization. The probabilistic health risk assessment disclosed severe health hazard in the order of infants > adults > children > teenagers through F- tainted groundwater. At P95 percentile dose, all the studied age groups showed THQ >1 from Dhapdhapi-II gram-panchayat. Supply of F- safe drinking water is required through reliable water supply strategies in the studied area.

Different levels of arsenic exposure through cooked rice and its associated benefit-risk assessment from rural and urban populations of West Bengal, India: a probabilistic approach with sensitivity analysis.

Rice arsenic (As) contamination and its consumption poses a significant health threat to humans. The present study focuses on the contribution of arsenic, micronutrients, and associated benefit-risk assessment through cooked rice from rural (exposed and control) and urban (apparently control) populations. The mean decreased percentages of As from uncooked to cooked rice for exposed (Gaighata), apparently control (Kolkata), and control (Pingla) areas are 73.8, 78.5, and 61.3%, respectively. The margin of exposure through cooked rice (MoEcooked rice) < 1 signifies the existence of health risk for all the studied exposed and control age groups. The respective contributions of iAs (inorganic arsenic) in uncooked and cooked rice are nearly 96.6, 94.7, and 100% and 92.2, 90.2, and 94.2% from exposed, apparently control, and control areas. LCR analysis for the exposed, apparently control, and control populations (adult male: 2.1 × 10-3, 2.8 × 10-4, 4.7 × 10-4; adult female: 1.9 × 10-3, 2.1 × 10-4, 4.4 × 10-4; and children: 5.8 × 10-4, 4.9 × 10-5, 1.1 × 10-4) through cooked rice is higher than the recommended value, i.e., 1 × 10-6, respectively, whereas HQ > 1 has been observed for all age groups from the exposed area and adult male group from the control area. Adults and children from rural area showed that ingestion rate (IR) and concentration are the respective influencing factors towards cooked rice As, whereas IR is solely responsible for all age groups from urban area. A vital suggestion is to reduce the IR of cooked rice for control population to avoid the As-induced health risks. The average intake (µg/day) of micronutrients is in the order of Zn > Se for all the studied populations and Se intake is lower for the exposed population (53.9) compared to the apparently control (140) and control (208) populations. Benefit-risk assessment supported that the Se-rich values in cooked rice are effective in avoiding the toxic effect and potential risk from the associated metal (As).

Rice grain arsenic and nutritional content during post harvesting to cooking: A review on arsenic bioavailability and bioaccessibility in humans.

Rice (Oryza sativa L.) is considered as the staple food for 50% of the world's population. Humans are exposed to arsenic (As) through rice consumption, which is a global health issue that requires attention. The present review reflects the scenario of rice grown in As endemic regions of Asia that has a significant portion of inorganic As (iAs) compared to other rice grown areas around the world. Post-harvesting, pre-cooking, and cooking procedures in South and South-East Asian countries employ As-contaminated groundwater. Polishing of brown rice and parboiling, washing and cooking with As-safe water can reduce As concentration and nutrient level in cooked rice. However, in rural parts of South-east Asia, rice is usually cooked using As-contaminated groundwater and consumption of this As enriched rice and water may cause a significant health exposure in humans. Bioaccessibility and bioavailability of As can be determined using in-vitro and in-vivo techniques that can be utilized as a tool to assess As exposure in humans. Arsenic in cooked rice may be reduced by using newly developed cooking procedures such as Kateh cooking, steam percolating, and the parboiled and absorbed (PBA) method. For individuals living in rural regions, using rainwater or treated surface water for drinking and cooking is also an alternative. Although this study examined the processes involved in the post-harvesting, pre-cooking, and cooking stages, there are still significant research gaps in this area that must be addressed in near future.

Effect of sulfate application on inhibition of arsenic bioaccumulation in rice (Oryza sativa L.) with consequent health risk assessment of cooked rice arsenic on human: A pot to plate study.

Arsenic (As) in rice is posing a serious threat worldwide and consumption of As contaminated rice by human is causing health risks. A pot experiment with different levels of sulfate dosage (0, 20, 40, 60 and 80 mg/kg) was set up in this study to explore the influence of sulfate fertilizer on rice plant growth, yield, and As accumulation in rice grain. Apart from As bioaccumulation in rice grains, the As fraction of cooked rice was quantified, and the health risks associated with cooked rice consumption were also investigated. The sulfate application significantly (p ≤ 0.05) enhanced the chlorophyll, tiller number, grains per panicle, grain and biomass yield under As stressed condition. The sulfate application also reduced the oxidative stress and antioxidant activity in rice plants. Sulfate fertigation improved the accumulation of total sulfur (S) and reduced the uptake and translocation of As in rice plants. Arsenic concentration in rice grain was reduced by 50.1% in S80 treatment (80 mg of sulfate/kg of soil) as compared to S0 set. The reduction percentage of As in cooked parboiled and sunned rice with correspond to raw rice ranged from 55.9 to 74% and 40.3-60.7%, respectively. However, the sulfate application and cooking of parboiled rice reduced the potential non-cancer and cancer risk as compared to sunned rice. The S80 treatment and cooking of parboiled rice reduce the As exposure for both children and adults by 51% as compared to cooked sunned rice under S80 treatment and this trend was similar for all treatments. Therefore, sulfate application in soil can be recommended to produce safer rice grains and subsequent cooking of parboiled rice grain with low-As contaminated water need to be done to avoid any potential health risk in As endemic areas.

Application of potassium humate to reduce arsenic bioavailability and toxicity in rice plants (Oryza sativa L.) during its course of germination and seedling growth.

Arsenic (As), a metalloid is a class I carcinogen and is a major problem in various parts of the world. Food crops are severely affected due to As poisoning and suffer from low germination, yield and disfiguration of morphological and anatomical traits. To attenuate such adverse effects and tone down As uptake by plants, the present study attempts to explore the role of K-humate (KH) in alleviation of As toxicity in rice. KH was administered in the growth media containing 800 ppb As (III) at varying doses to observe the stress alleviating capacity of the amendment. Five treatments were investigated, viz: (a) 800 ppb As (control), (b) 800 ppb As + 25 ppm KH, (c) 800 ppb As + 50 ppm KH, (d) 800 ppb As + 75 ppm KH and (e) 800 ppb As + 100 ppm KH. The results of the amendment administration were noted at 14 days after seeding (DAS). Application of KH significantly improved germination percentage, vigour indices and chlorophyll content by reducing the oxidative stress, antioxidant and antioxidant enzyme activities under As stress. In vivo detection of reactive oxygen species (ROS) using DCF-2DA fluorescent dye and scanning electron microscope (SEM) study of root further depicted that KH application effectively reduced ROS formation and improved root anatomical structure under As stress, respectively. Gradually increasing concentrations of KH was capable of decreasing the bioavailability of As to the rice plants, thus minimizing toxic effect of the metalloid.

Single trait versus principal component based association analysis for flowering related traits in pigeonpea.

Pigeonpea, a tropical photosensitive crop, harbors significant diversity for days to flowering, but little is known about the genes that govern these differences. Our goal in the current study was to use genome wide association strategy to discover the loci that regulate days to flowering in pigeonpea. A single trait as well as a principal component based association study was conducted on a diverse collection of 142 pigeonpea lines for days to first and fifty percent of flowering over 3 years, besides plant height and number of seeds per pod. The analysis used seven association mapping models (GLM, MLM, MLMM, CMLM, EMLM, FarmCPU and SUPER) and further comparison revealed that FarmCPU is more robust in controlling both false positives and negatives as it incorporates multiple markers as covariates to eliminate confounding between testing marker and kinship. Cumulatively, a set of 22 SNPs were found to be associated with either days to first flowering (DOF), days to fifty percent flowering (DFF) or both, of which 15 were unique to trait based, 4 to PC based GWAS while 3 were shared by both. Because PC1 represents DOF, DFF and plant height (PH), four SNPs found associated to PC1 can be inferred as pleiotropic. A window of ± 2 kb of associated SNPs was aligned with available transcriptome data generated for transition from vegetative to reproductive phase in pigeonpea. Annotation analysis of these regions revealed presence of genes which might be involved in floral induction like Cytochrome p450 like Tata box binding protein, Auxin response factors, Pin like genes, F box protein, U box domain protein, chromatin remodelling complex protein, RNA methyltransferase. In summary, it appears that auxin responsive genes could be involved in regulating DOF and DFF as majority of the associated loci contained genes which are component of auxin signaling pathways in their vicinity. Overall, our findings indicates that the use of principal component analysis in GWAS is statistically more robust in terms of identifying genes and FarmCPU is a better choice compared to the other aforementioned models in dealing with both false positive and negative associations and thus can be used for traits with complex inheritance.

Isolation and expression analysis of partial sequences of heavy metal transporters from Brassica juncea by coupling high throughput cloning with a molecular fingerprinting technique.

Heavy metal transporters play a key role in regulating metal accumulation and transport in plants. These are important candidate genes to study in metal tolerant and accumulator plants for their potential use in environmental clean up. We coupled a degenerate primer-based RT-PCR approach with a molecular fingerprinting technique based on amplified rDNA restriction analysis (ARDRA) to identify novel ESTs corresponding to heavy metal transporters from metal accumulator Brassica juncea. We utilized this technique to clone several family members of natural resistance-associated macrophage proteins (NRAMP) and yellow stripe-like proteins (YSL) in a high throughput manner to distinguish between closely related isoforms and/or allelic variants from the allopolyploid B. juncea. Partial clones of 23 Brassica juncea NRAMPs and 27 YSLs were obtained with similarity to known Arabidopsis thaliana and Noccaea (Thlaspi) caerulescens NRAMP and YSL genes. The cloned transporters showed Brassica-specific changes in domains, which can have important functional consequences. Semi-quantitative RT-PCR-based expression analysis of chosen members indicated that even closely related isoforms/allelic variants of BjNRAMP and BjYSL have distinct tissue-specific and metal-dependent expressions which might be essential for adaptive fitness and heavy metal tolerance. Consistent to this, BjYSL6.1 and BjYSL5.8 were found to show elevated expressions specifically in cadmium-treated shoots and lead-treated roots of B. juncea, respectively.

A Behavioral Screen for Heat-Induced Seizures in Mouse Models of Epilepsy.

Transgenic mouse models have proved to be powerful tools in studying various aspects of human neurological disorders, including epilepsy. The SCN1A-associated genetic epilepsies comprise a wide spectrum of seizure disorders with incomplete penetrance and clinical variability. SCN1A mutations can result in a large variety of seizure phenotype ranging from simple, self-limited fever-associated febrile seizures (FS), moderate-level genetic epilepsy with febrile seizures plus (GEFS+) to more severe Dravet Syndrome (DS). Although FS are commonly seen in children below 6-7 years of age who do not have genetic epilepsy, FS in GEFS+ patients continue to occur into adulthood. Traditionally, experimental FS have been induced in mice by exposing the animal to a stream of dry air or heating lamps, and the rate of change in body temperature is often not well controlled. Here, we describe a custom-built heating chamber, with a plexiglass front, that is fitted with a digital temperature controller and a heater-equipped electric fan, which can send heated forced air into the test arena in a temperature-controlled manner. The body temperature of a mouse placed in the chamber, monitored through a rectal probe, can be increased to 40-42 °C in a reproducible manner by increasing the temperature inside the chamber. Continual visual monitoring of the animals during the heating period demonstrates induction of heat-induced seizures in mice carrying an FS mutation at a body temperature that does not elicit behavioral seizures in wild-type litter mates. Animals can be easily removed from the chamber and placed on a cooling pad to rapidly return body temperature to normal. This method provides for a simple, rapid, and reproducible screening protocol for the occurrence of heat-induced seizures in epilepsy mouse models.

Interneuron Dysfunction in a New Mouse Model of SCN1A GEFS.

Advances in genome sequencing have identified over 1300 mutations in the SCN1A sodium channel gene that result in genetic epilepsies. However, it still remains unclear how most individual mutations within SCN1A result in seizures. A previous study has shown that the K1270T (KT) mutation, linked to genetic epilepsy with febrile seizure plus (GEFS+) in humans, causes heat-induced seizure activity associated with a temperature-dependent decrease in GABAergic neuron excitability in a Drosophila knock-in model. To examine the behavioral and cellular effects of this mutation in mammals, we introduced the equivalent KT mutation into the mouse (Mus musculus) Scn1a (Scn1aKT) gene using CRISPR/Cas9 and generated mutant lines in two widely used genetic backgrounds: C57BL/6NJ and 129X1/SvJ. In both backgrounds, mice homozygous for the KT mutation had spontaneous seizures and died by postnatal day (P)23. There was no difference in mortality of heterozygous KT mice compared with wild-type littermates up to six months old. Heterozygous mutants exhibited heat-induced seizures at ∼42°C, a temperature that did not induce seizures in wild-type littermates. In acute hippocampal slices at permissive temperatures, current-clamp recordings revealed a significantly depolarized shift in action potential threshold and reduced action potential amplitude in parvalbumin (PV)-expressing inhibitory CA1 interneurons in Scn1aKT/+ mice. There was no change in the firing properties of excitatory CA1 pyramidal neurons. These results suggest that a constitutive decrease in inhibitory interneuron excitability contributes to the seizure phenotype in the mouse model.

Identification and characterization of MADS box gene family in pigeonpea for their role during floral transition.

MADS box genes are class of transcription factors involved in various physiological and developmental processes in plants. To understand their role in floral transition-related pathways, a genome-wide identification was done in Cajanus cajan, identifying 102 members which were classified into two different groups based on their gene structure. The status of all these genes was further analyzed in three wild species i.e. C. scarabaeoides, C. platycarpus and C. cajanifolius which revealed absence of 31-34 MADS box genes in them hinting towards their role in domestication and evolution. We could locate only a single copy of both FLOWERING LOCUS C (FLC) and SHORT VEGETATIVE PHASE (SVP) genes, while three paralogs of SUPPRESSOR OF ACTIVATION OF CONSTANS 1 (SOC1) were found in C. cajan genome. One of those SOC1 paralogs i.e. CcMADS1.5 was found to be missing in all three wild relatives, also forming separate clade in phylogeny. This SOC1 gene was also lacking the characteristic MADS box domain in it. Expression profiling of major MADS box genes involved in flowering was done in different tissues viz shoot apical meristem, vegetative leaf, reproductive meristem, and reproductive bud. Gene-based time tree of FLC and SOC1 gene dictates their divergence from Arabidopsis before 71 and 23 million year ago (mya), respectively. This study provides valuable insights into the functional characteristics, expression pattern, and evolution of MADS box proteins in grain legumes with emphasis on C. cajan, which may help in further characterizing these genes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-020-02605-7.