Genomic characterization of ZIP genes in pigeonpea (CcZIP) and their expression analysis among the genotypes with contrasting host response to pod borer.

Zinc (Zn) is a vital micronutrient from the perspective of biofortification and biotic stress endurance in pigeonpea. The ZIP transporters with domain (Pfam: PF02535) regulate uptake and transport of metal ions, including Zn, in consonance with plant metal homeostasis. Genome-wide analysis in pigeonpea identified 19 non-redundant members of ZIP family (CcZIP) that were analyzed for gene structure, conserved motifs and homology besides other structural and biochemical parameters. Intra-specific as well as the inter-specific phylogenetic relationships of these 19 CcZIPs were elucidated by comparison with ZIP proteins of Arabidopsis thaliana, Medicago truncatula, Phaseolus vulgaris and Glycine max. In addition to gene structure, the cis-regulatory elements (CREs) in the promoter region were also identified. It revealed several stress responsive CREs that might be regulatory for differential expression of CcZIP proteins. Expression analysis showed that both CcZIP3 and CcZIP15, having zinc deficiency responsive element, up-regulated in the reproductive leaf tissues and down-regulated in matured green pods of the pod borer resistant genotypes with higher zinc content. Alternately, the expression of CcZIP6 and CcZIP13 was higher in matured green pods than reproductive leaves of the resistant genotypes. These findings on differential expression indicate the possible role of these CcZIPs on the mobilization of Zn from leaves to pods, phloem loading and unloading, and higher accumulation of seed zinc in pod borer resistant genotypes used in this study. Further functional characterization of CcZIP genes could shed light on their role in bio-fortification and genetic improvement to inhibit the pod borer herbivory in pigeonpea. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01111-1.

Comparative transcriptome analysis of Gossypium hirsutum L. in response to sap sucking insects: aphid and whitefly.

BACKGROUND: Cotton (Gossypium hirsutum L.) is a major fiber crop that is grown worldwide; it faces extensive damage from sap-sucking insects, including aphids and whiteflies. Genome-wide transcriptome analysis was performed to understand the molecular details of interaction between Gossypium hirsutum L. and sap-sucking pests, namely Aphis gossypii (Aphid) and Bemisia tabacci (Whiteflies). Roche's GS-Titanium was used to sequence transcriptomes of cotton infested with aphids and whiteflies for 2 h and 24 h. RESULTS: A total of 100935 contigs were produced with an average length of 529 bp after an assembly in all five selected conditions. The Blastn of the non-redundant (nr) cotton EST database resulted in the identification of 580 novel contigs in the cotton plant. It should be noted that in spite of minimal physical damage caused by the sap-sucking insects, they can change the gene expression of plants in 2 h of infestation; further change in gene expression due to whiteflies is quicker than due to aphids. The impact of the whitefly 24 h after infestation was more or less similar to that of the aphid 2 h after infestation. Aphids and whiteflies affect many genes that are regulated by various phytohormones and in response to microbial infection, indicating the involvement of complex crosstalk between these pathways. The KOBAS analysis of differentially regulated transcripts in response to aphids and whiteflies indicated that both the insects induce the metabolism of amino acids biosynthesis specially in case of whiteflies infestation at later phase. Further we also observed that expression of transcript related to photosynthesis specially carbon fixation were significantly influenced by infestation of Aphids and Whiteflies. CONCLUSIONS: A comparison of different transcriptomes leads to the identification of differentially and temporally regulated transcripts in response to infestation by aphids and whiteflies. Most of these differentially expressed contigs were related to genes involved in biotic, abiotic stresses and enzymatic activities related to hydrolases, transferases, and kinases. The expression of some marker genes such as the overexpressors of cationic peroxidase 3, lipoxygenase I, TGA2, and non-specific lipase, which are involved in phytohormonal-mediated plant resistance development, was suppressed after infestation by aphids and whiteflies, indicating that insects suppressed plant resistance in order to facilitate their infestation. We also concluded that cotton shares several pathways such as phagosomes, RNA transport, and amino acid metabolism with Arabidopsis in response to the infestation by aphids and whiteflies.

Comparative transcriptomic analysis of roots of contrasting Gossypium herbaceum genotypes revealing adaptation to drought.

BACKGROUND: Root length and its architecture govern the adaptability of plants to various stress conditions, including drought stress. Genetic variations in root growth, length, and architecture are genotypes dependent. In this study, we compared the drought-induced transcriptome of four genotypes of Gossypium herbaceum that differed in their drought tolerance adaptability. Three different methodologies, namely, microarray, pyrosequencing, and qRT-PCR, were used for transcriptome analysis and validation. RESULTS: The variations in root length and growth were found among four genotypes of G.herbaceum when exposed to mannitol-induced osmotic stress. Under osmotic stress, the drought tolerant genotypes Vagad and GujCot-21 showed a longer root length than did by drought sensitive RAHS-14 and RAHS-IPS-187. Further, the gene expression patterns in the root tissue of all genotypes were analyzed. We obtained a total of 794 differentially expressed genes by microarray and 104928 high-quality reads representing 53195 unigenes from the root transcriptome. The Vagad and GujCot-21 respond to water stress by inducing various genes and pathways such as response to stresses, response to water deprivation, and flavonoid pathways. Some key regulatory genes involved in abiotic stress such as AP2 EREBP, MYB, WRKY, ERF, ERD9, and LEA were highly expressed in Vagad and GujCot-21. The genes RHD3, NAP1, LBD, and transcription factor WRKY75, known for root development under various stress conditions, were expressed specifically in Vagad and GujCot-21. The genes related to peroxidases, transporters, cell wall-modifying enzymes, and compatible solutes (amino acids, amino sugars, betaine, sugars, or sugar alcohols) were also highly expressed in Vagad and Gujcot-21. CONCLUSION: Our analysis highlights changes in the expression pattern of genes and depicts a small but highly specific set of drought responsive genes induced in response to drought stress. Some of these genes were very likely to be involved in drought stress signaling and adaptation, such as transmembrane nitrate transporter, alcohol dehydrogenase, pyruvate decarboxylase, sucrose synthase, and LEA. These results might serve as the basis for an in-depth genomics study of Gossypium herbaceum, including a comparative transcriptome analysis and the selection of genes for root traits and drought tolerance.

Multimodal Analysis of Dysphonia in Smokers: A Two Year Comprehensive Study.

To study the specific effects of smoking on laryngeal mucosa and voice using video-stroboscopy, subjective and objective voice analysis. This cross-sectional and observational study was carried out in the Department of Otorhinolaryngology, M.G.M. Medical College & M.Y. Hospital, Indore on 103 subjects with dysphonia (49 smokers and 54 non-smokers), between October 2018 to October 2020. After detailed history and general examination, all the cases underwent video-stroboscopic analysis and voice analysis by simple parameters-Maximum Phonation Time (M.P.T.) & s/z score. The cases answered the Voice Related Quality Of Life questionnaire (V.R.Q.O.L.). The observations between smokers and non-smokers were compared. Statistically significant (P < 0.05) associations were observed between smoking and various vocal fold pathological characteristics. There was statistically significant association of smoking with overall possibility of benign vocal fold lesions (P = 0.0129). Significant association was noted of smoking with specific lesions like leukoplakia (P = 0.02), erythema (P = 0.0161) and Reinke's edema (P = 0.322). The mean M.P.T. was 9.5 s in smokers and 11 s in non-smokers. We observed statistically significant relationship between smoking and abnormal s/z ratio (P = 0.0165). This study shows the observable effects of smoking on vocal fold micro-anatomy and voice. Dysphonia was primary complaint in both groups, but all observations were worse in smokers, indicating harmful effects of smoking.

Title-Optical Forceps: The Real Boon for Surgeon, Residents and Patients with Foreign Body Aspiration.

Management of aspirated foreign bodies in children becomes very challenging in absence of most appropriate instruments. Rigid bronchoscopy has evolved in the long course of history. The advent of Hopkins rod telescope and optical forceps has enhanced the efficacy as well as the safety of removing tracheobronchial foreign bodies. Total 36 children of age less than 16 year with confirmed diagnosis of airway F.B. were included in the study. Two types of forceps were used: (1)Optical forceps combined with Hopkins rod lens telescope[n = 25] (2)Standard forceps [n = 11]. Most common age group was 1-5 year [n = 26] with male preponderance [n = 20]. Most common site was right bronchus in 13 cases (RMB-11, RBI-1, RLLB-1) followed by Left main bronchus in 11 cases, trachea in 10 cases, bilateral bronchus and sub-glottis each in 1 case. Vegetative F.B. were found in majority of cases [n = 26]. Mean grasping attempt, Mean bronchoscopic insertion and Mean time taken from insertion of forceps to removal of F.B. were less in optical forceps as compared to standard forceps with significant p-value. Optical forceps have advantage of high resolution & magnified view of airway, spring action in handle and better tactile sensation. These helps in correctly identifying the type, size and site of tracheobronchial foreign body. Precise grasping attempts with optical forceps reduces the chances of complications. Optical forceps have given the new dimension to the Rigid Bronchoscopy and proved to be the real boon for surgeons, residents and patients with F.B. aspiration.

Whitefly and aphid inducible promoters of Arabidopsis thaliana L.

Lack of regulated expression and tissue specificity are the major drawbacks of plant and virus-derived constitutive promoters. A precise tissue or site-specific expression, facilitate regulated expression of proteins at the targeted time and site. Publically available microarray data on whitefly and aphid infested Arabidopsis thaliana L. was used to identify whitefly and aphid-inducible genes. The qRT-PCR further validated the inducible behaviour of these genes under artificial infestation. Promoter sequences of genes were retrieved from the Arabidopsis Information Resources database with their corresponding 5'UTR and cloned from the A. thaliana genome. Promoter reporter transcriptional fusions were developed with the beta-glucuronidase (GUS) gusA gene in a binary expression vector to validate the inducible behaviour of these promoters in eight independent transgenic Nicotiana tabaccum lines. Histochemical analysis of the reporter gene in T2 transgenic tobacco lines confirmed promoter driven expression at the sites of aphid and whitefly infestation. The qRT-PCR and GUS expression analysis of transgenic lines revealed that abscisic acid largely influenced the expression of both aphid and whitefly inducible promoters. Further, whitefly-specific promoter respond to salicylic acid and jasmonic acid (JA), whereas aphid-specific promoters to JA and 1-aminocyclopropane carboxylic acid. The response of promoters to phytohormones correlated to the presence of corresponding conserved cis-regulatory elements.

Acute stroke secondary to internal carotid artery pseudoaneurysm: MRI findings and treatment with endovascular coiling.

Internal carotid artery pseudoaneurysms are uncommon. We report a young man with history of trivial trauma who presented with middle cerebral artery stroke. Head CT scan revealed a right parapharyngeal mass. MRI confirmed the diagnosis of internal carotid pseudoaneurysm. Cerebral angiogram showed dissection of internal carotid artery with a large pseudoaneurysm and almost complete occlusion of vessel distal to aneurysm. Vessel was obliterated by fibercoiling and gluing by endovascular technique to diminish risk of further embolization. MRI characteristics of carotid pseudoaneurysms and role of endovascular treatment are discussed.

Enhanced Host-Parasite Resistance Based on Down-Regulation of Phelipanche aegyptiaca Target Genes Is Likely by Mobile Small RNA.

RNA silencing refers to diverse mechanisms that control gene expression at transcriptional and post-transcriptional levels which can also be used in parasitic pathogens of plants that Broomrapes (Orobanche/Phelipanche spp.) are holoparasitic plants that subsist on the roots of a variety of agricultural crops and cause severe negative effects on the yield and yield quality of those crops. Effective methods for controlling parasitic weeds are scarce, with only a few known cases of genetic resistance. In the current study, we suggest an improved strategy for the control of parasitic weeds based on trans-specific gene-silencing of three parasite genes at once. We used two strategies to express dsRNA containing selected sequences of three Phelipanche aegyptiaca genes PaACS, PaM6PR, and PaPrx1 (pma): transient expression using Tobacco rattle virus (TRV:pma) as a virus-induced gene-silencing vector and stable expression in transgenic tomato Solanum lycopersicum (Mill.) plants harboring a hairpin construct (pBINPLUS35:pma). siRNA-mediated transgene-silencing (20-24 nt) was detected in the host plants. Our results demonstrate that the quantities of PaACS and PaM6PR transcripts from P. aegyptiaca tubercles grown on transgenic tomato or on TRV-infected Nicotiana benthamiana plants were significantly reduced. However, only partial reductions in the quantity of PaPrx1 transcripts were observed in the parasite tubercles grown on tomato and on N. benthamiana plants. Concomitant with the suppression of the target genes, there were significant decreases in the number and weight of the parasite tubercles that grew on the host plants, in both the transient and the stable experimental systems. The results of the work carried out using both strategies point to the movement of mobile exogenous siRNA from the host to the parasite, leading to the impaired expression of essential parasite target genes.

Clinical drug-drug interaction assessment of ivacaftor as a potential inhibitor of cytochrome P450 and P-glycoprotein.

Ivacaftor is approved in the USA for the treatment of cystic fibrosis (CF) in patients with a G551D-CFTR mutation or one of eight other CFTR mutations. A series of in vitro experiments conducted early in the development of ivacaftor indicated ivacaftor and metabolites may have the potential to inhibit cytochrome P450 (CYP) 2C8, CYP2C9, CYP3A, and CYP2D6, as well as P-glycoprotein (P-gp). Based on these results, a series of clinical drug-drug interaction (DDI) studies were conducted to evaluate the effect of ivacaftor on sensitive substrates of CYP2C8 (rosiglitazone), CYP3A (midazolam), CYP2D6 (desipramine), and P-gp (digoxin). In addition, a DDI study was conducted to evaluate the effect of ivacaftor on a combined oral contraceptive, as this is considered an important comedication in CF patients. The results indicate ivacaftor is a weak inhibitor of CYP3A and P-gp, but has no effect on CYP2C8 or CYP2D6. Ivacaftor caused non-clinically significant increases in ethinyl estradiol and norethisterone exposure. Based on these results, caution and appropriate monitoring are recommended when concomitant substrates of CYP2C9, CYP3A and/or P-gp are used during treatment with ivacaftor, particularly drugs with a narrow therapeutic index, such as warfarin.

Role of fluorodeoxyglucose positron emission tomography in the diagnosis of neurosarcoidosis.

A 45-year-old man developed seizures and myelopathy. MRI showed bitemporal and cervical spinal cord hyperintense lesions on T2-weighted and FLAIR images that contrast-enhanced. Initial evaluation for sarcoidosis was negative, including serum angiotensin converting enzyme (ACE) and chest X-ray. Whole body fluorodeoxyglucose positron emission tomography (FDG-PET) revealed multiple hypermetabolic hilar and mediastinal foci and spinal cord hypermetabolism at the site of MRI abnormality. Temporal lobe MRI lesions were hypometabolic. Mediastinal lymph node biopsy was consistent with sarcoidosis. The brain, spinal cord, and chest metabolic abnormalities together with the clinical presentation were interpreted as being most consistent with sarcoidosis. FDG-PET helped target the site of biopsy that subsequently confirmed the diagnosis histologically. In patients with perplexing neurologic presentations, whole body FDG-PET can help secure a timely and minimally invasive diagnosis of neurosarcoidosis.

Diffusion-weighted magnetic resonance imaging in superior sagittal sinus thrombosis.

BACKGROUND AND PURPOSE: Diffusion-weighted imaging (DWI) has shown high sensitivity in the diagnosis of acute arterial strokes. The pathophysiology of cerebral venous thrombosis with associated venous stroke appears to differ from that of arterial strokes. The purpose of this study was to describe DWI findings in venous strokes. METHODS: The authors reviewed 3 adults with superior sagittal sinus thrombosis who underwent DWI and magnetic resonance imaging within 24 hours of symptom onset. DWI was obtained at 1.5 T using the multishot echo planar technique (TR = 8000, TE = 97, field of view = 30 x 19 cm, slice thickness = 6.0 mm, interslice gap = 0.5 mm, matrix 128 x 128, NEX = 1). The diffusion gradients were applied in 3 orthogonal directions with 3 increasing b values (0-1000 s/mm2) to create average (trace) DWI images. Apparent diffusion coefficient (ADC) values were calculated on a pixel-by-pixel basis and displayed as ADC maps. RESULTS: DWI showed hyperintensities in patients 1 and 2 and hypointensity in patient 3 in corresponding to parenchymal lesions on conventional images. As compared to the homologous uninvolved location in the contralateral hemisphere, ADC values were decreased (0.53 x 10(-3) mm2/s [patient 1] and 0.68 x 10(-3) mm2/s [patient 2]) and increased (1.1 x 10(-3) mm2/s [patient 3]). The ADC ratio of the lesion in the involved to uninvolved side was 88% (patient 1), 81% (patient 2), and 120% (patient 3). CONCLUSION: Acute cerebral venous strokes may contain cytotoxic edema and/or vasogenic edema on DWI scans. DWI may be helpful in diagnosing cerebral venous thrombosis in cases with cryptic presentations.

Gene silencing of CCD7 and CCD8 in Phelipanche aegyptiaca by tobacco rattle virus system retarded the parasite development on the host.

Strigolactones are phytohormones that stimulate seed germination of parasitic plants including Phelipanche aegyptiaca. Strigolactones are derived from carotenoids via a pathway involving the carotenoid cleavage dioxygenases CCD7 and CCD8. We report here identification of PaCCD7 and PaCCD8 orthologous genes from P. aegyptiaca. Expression analysis of PaCCD7 and PaCCD8 genes showed significant variation in their transcript levels in seeds and tubercles of P. aegyptiaca at different developmental stages. These two parasitic PaCCD7 and PaCCD8 genes were silenced in P. aegyptiaca using a trans-silencing approach in Nicotiana benthamiana. The transient knock-down of PaCCD7 and PaCCD8 inhibited tubercle development and the infestation process in host plants. Our results suggest an important role of the strigolactone associated genes (PaCCD7 and PaCCD8) in the parasite life cycle.

Cucumber Mosaic Virus as a carotenoid inhibitor reducing Phelipanche aegyptiaca infection in tobacco plants.

Cucumber Mosaic Virus (CMV) is a highly infectious cucumovirus, which infects more than 800 plant species and causes major diseases in greenhouse and field crops worldwide. Parasitic weeds such as Phelipanche aegyptiaca are a major constraint to the production of many crops in the world and the parasite's lifestyle makes control extremely difficult. The parasite seeds can germinate after conditioning and perceiving strigolactones secreted by the host roots. Strigolactones are rhizosphere signaling molecules in plants that are biosynthesized through carotenoid cleavage. In the present study we investigated the possibility of reducing ß-carotene and then strigolactone production in the host roots by blocking carotenoid biosynthesis using CMV-infected tobacco. It was found that CMV downregulated the enzyme phytoene desaturase(PDS) and reduced significantly both carotenoid production and Phelipanche infection in tobacco host roots infected with both CMV and P. aegyptiaca. Based on our results (decrease of ß-carotene and repression of PDS transcripts in tobacco roots), we hypothesized that the reduction of Phelipanche tubercles and shoots occurred due to an effect of CMV on secondary metabolite stimulators such as strigolacetones. Our study indicated that mass production of the host roots was not affected by CMV; however, most inflorescences of Phelipanche grown on CMV-infected tobacco developed abnormally (deformed shoots and short nodes). Carotenoid biosynthesis inhibitors such as CMV can be used to reduce the production of strigolactones, which will lead to decreased Phelipanche attachment. Interestingly, attenuated CMV strains may provide a safe means for enhancing crop resistance against parasitic weeds in a future plan.

Formation of norisoprenoid flavor compounds in carrot (Daucus carota L.) roots: characterization of a cyclic-specific carotenoid cleavage dioxygenase 1 gene.

Carotenoids are isoprenoid pigments that upon oxidative cleavage lead to the production of norisoprenoids that have profound effect on flavor and aromas of agricultural products. The biosynthetic pathway to norisoprenoids in carrots (Daucus carota L.) is still largely unknown. We found the volatile norisoprenoids farnesylacetone, α-ionone, and ß-ionone accumulated in Nairobi, Rothild, and Purple Haze cultivars but not in Yellowstone and Creme de Lite in a pattern reflecting their carotenoid content. A cDNA encoding a protein with carotenoid cleavage dioxygenase activity, DcCCD1, was identified in carrot and was overexpressed in Escherichia coli strains previously engineered to produce different carotenoids. The recombinant DcCCD1 enzyme cleaves cyclic carotenes to generate α- and ß-ionone. No cleavage products were found when DcCCD1 was co-expressed in E. coli strains accumulating non-cyclic carotenoids, such as phytoene or lycopene. Our results suggest a role for DcCCD1 in carrot flavor biosynthesis.

Purification and characterization of a lectin with high hemagglutination property isolated from Allium altaicum.

A lectin was purified from the leaves of Allium altaicum and corresponding gene was cloned. The lectin namely Allium altaicum agglutinin (AAA) was ~24 kDa homodimeric protein and similar to a typical garlic leaf lectin. It was synthesized as 177 amino acid residues pre-proprotein, which consisted of 28 and 43 amino acid long N and C-terminal signal peptides, respectively. The plant expressed this protein more in scapes and flowers in comparison to the bulbs and leaves. Hemagglutination activity (with rabbit erythrocytes) was 1,428 fold higher as compared to Allium sativum leaf agglutinin (ASAL) although, the insecticidal activity against cotton aphid (Aphis gossypii) was relatively low. Glycan array revealed that AAA had higher affinity towards GlcAb1-3Galb as compared to ASAL. Homology analysis showed 57-94% similarity with other Allium lectins. The mature protein was expressed in E. coli as a fusion with SUMO peptide in soluble and biologically active form. Recombinant protein retained high hemagglutination activity.