Efficacy and Safety of Ayurveda interventions in the management of conjunctivitis: A systematic review and meta-analysis.

BACKGROUND: Conjunctivitis is the inflammation of the conjunctiva. Although data on clinical efficacy and safety of various ayurvedic treatments in conjunctivitis is published, systematic review is not done. This systematic review and meta-analysis aims to evaluate the efficacy and safety of ayurvedic treatments in conjunctivitis. METHODS: A literature search of the Cochrane Library (Cochrane central register of controlled trials: issue 6 of 12, June 2018), Pub Med, AYUSH research portal (Govt. of India), DHARA portal, Google scholar and online clinical trials registers was done. Randomized controlled trials (RCTs), quasi-randomized controlled trials (QRCTs), controlled clinical trials (CCTs) and multiple arms clinical trials were identified in which Ayurveda treatments with any dose, type, schedule, drug, dosage form, and advised Pathayapathya (lifestyle changes) were selected. RESULTS: We identified 13 eligible RCTs, five CCTs and two multiple arms clinical trials which includes a total of 816 participants. Meta analysis of data from five trials showed that ayurvedic treatments benefitted more compared with non-ayurveda interventions in symptoms like itching (SMD = -0.98, 95% CI (-1.30,-0.65) p < 0.00001, I2 = 38%), pain (SMD = -0.57, 95% CI (-0.87, -0.29, P = 0.0001, I2 = 0%), ropy discharge (SMD = -1.02, 95% CI(-1.45, -0.59), P < 0.00001, I2 = 0%), conjunctival congestion (SMD = -0.67, 95% CI (-0.91, -0.43), p < 0.00001, I2 = 0%), foreign body sensation (SMD = -0.68, 95% CI(-1.06, -0.29), p = 0.0006, I2 = 46%, Fig. 8) and lid heaviness (SMD = -0.66, 95% CI(- 0.98, -0.33), p < 0.0001, I2 = 0%). CONCLUSIONS: Although some findings confirm the benefit of ayurveda as opposed to non ayurveda for the treatment of conjunctivitis, since the studies have high risk of bias and are of lower quality, the findings could not be generalized. There is a need for high quality studies in ayurveda in this regard. PROSPERO REGISTRATION: CRD42019129436.

Genetic transformation of tropical maize (Zea mays L.) inbred line with a phytase gene from Aspergillus niger.

A full-length cDNA of phyA gene of Aspergillus niger, encoding phytase enzyme, was cloned and expressed in E. coli BL21 cells and assayed for its activity. The phyA cDNA consisted of 1404 bp, which encoded 467 amino acid residues. The phytase activity of purified phytase was 826.33 U/mL. The phyA gene under the control of endosperm-specific promoters was transformed into an Indian maize inbred line, UMI29, using particle bombardment-mediated transformation method to generate transgenic maize plants over-expressing phytase in seeds. PCR and GUS analyses demonstrated the presence of transgenes in T0 transgenic plants and their stable inheritance in the T1 progenies. Three transgenic events expressing detectable level of A. niger phytase were characterized by western blot analysis. Phytase activity of 463.158 U/kg of seed was observed in one of the events, JB-UMI29-Z17/2. The phytase activity of transgenic maize seeds was 5.5- to 7-fold higher than the wild-type UMI29 seeds and, consequently, the seeds had 0.6- to 5-fold higher inorganic phosphorus content.

SNPs in ERCC1, ERCC2, and XRCC1 genes of the DNA repair pathway and risk of male infertility in the Asian populations: association study, meta-analysis, and trial sequential analysis.

PURPOSE: We investigated if substitutions in the ERCC1, ERCC2, and XRCC1 genes of the DNA repair pathway correlate with non-obstructive azoospermia and male infertility. METHODS: A total of 548 azoospermic infertile males and 410 fertile controls were genotyped for XRCC1 399A > G, 280G > A, and ERCC1 C > A 3' UTR and 541 azoospermic infertile males and 416 fertile controls were genotyped for ERCC2 751A > C using iPLEX Gold Assay. Meta-analyses were performed on XRCC1 399A > G (1022 cases and 1004 controls), ERCC1 C > A 3' UTR (879 cases and 1059 controls), and ERCC2 751A > C (914 cases and 850 controls) polymorphisms to quantitatively estimate the significance of the association between these polymorphisms and the risk of infertility. RESULTS: Statistically significant association between ERCC2 751A > C SNP and male infertility was found using the codominant model (p = 0.03). Results of meta-analysis suggested a lack of correlation with male infertility risk, which could be due to pooling of studies from different ethnic populations. Due to limited the number of studies, a stratified analysis for different ethnic groups could not be performed. CONCLUSION (S): In conclusion, AA genotype of 751A > C SNP in ERCC2 correlated with a higher risk of male infertility and may contribute to an increased risk of azoospermia and male infertility in Indian men.

NR5A1 mutations are not associated with male infertility in Indian men.

NR5A1 or steroidogenic factor 1 (SF1) is an autosomal gene, which encodes a protein that is a member of nuclear receptor family. NR5A1 regulates the transcription of numerous genes that are expressed in hypothalamic-pituitary-gonadal axis and adrenal cortex which in turn, coordinate the gonadal development, steroidogenesis and sex differentiation. Several mutations in NR5A1 have been reported to cause gonadal dysgenesis with adrenal insufficiency in individuals with 46,XY karyotype. However, studies in the past few years have shown that NR5A1 mutations can also contribute to primary ovarian insufficiency and impaired spermatogenesis. As there is no genetic study on NR5A1 in Indian infertile men, we have sequenced the entire coding region (exons 2-7) of NR5A1 in 502 infertile men of which, 414 were non-obstructive azoospermic and 88 severe oligozoospermic, along with 427 ethnically matched fertile controls. Interestingly, none of the mutations reported to be associated with male infertility were found in our study, except one polymorphism, rs1110061. However, it was not significantly different between infertile and fertile groups (p = .76). In addition, we have identified six intronic variants; but none of them was significantly associated with male infertility.

Docking and molecular dynamics simulation study of EGFR1 with EGF-like peptides to understand molecular interactions.

Epidermal growth factor receptors (EGFRs) are oncogenes, which regulate the expression of genes in various pathways, allowing cells to grow and divide. EGF-like growth factors bind to the extracellular region of EGFR causing EGFR dimerization (homo/hetero) and activation of the intrinsic protein kinase activity of the EGFR. The binding potentials of different growth factors vary from nanomolar to micromolar, because of changes in conformational state of EGFR1 bound to different growth factors. Our aim is to predict the key amino acid residues that are vital to activation of EGFR1, stimulating subsequent conformational changes in the extracellular region and its dimerization. Protein-peptide docking was performed using HADDOCK and molecular dynamics simulations of complexes were done using Gromacs. Dynamic domain movement studies were performed to understand the conformational changes in the domains of EGFR1. We predicted epidermal growth factor, transforming growth factor-α, heparin-binding epidermal growth factor, and betacellulin would show better interactions than epiregulin and neuregulin with EGFR1. The study identifies the altered behavior of crucial EGFR1 residues Cys305, Gly307, Arg310, and Val312, which is suggestive of their probable role in dimerization of EGFRs and activation of the tyrosine kinase domain, which is associated with cancer.

Development of leaffolder resistant transgenic rice expressing cry2AX1 gene driven by green tissue-specific rbcS promoter.

The insecticidal cry genes of Bacillus thuringiensis (Bt) have been successfully used for development of insect resistant transgenic rice plants. In this study, a novel cry2AX1 gene consisting a sequence of cry2Aa and cry2Ac gene driven by rice rbcS promoter was introduced into a rice cultivar, ASD16. Among 27 putative rice transformants, 20 plants were found to be positive for cry2AX1 gene. The expression of Cry2AX1 protein in transgenic rice plants ranged from 5.95 to 122.40 ng/g of fresh leaf tissue. Stable integration of the transgene was confirmed in putative transformants of rice by Southern blot hybridization analysis. Insect bioassay on T0 transgenic rice plants against rice leaffolder (Cnaphalocrosis medinalis) recorded larval mortality up to 83.33%. Stable inheritance and expression of cry2AX1 gene in T1 progenies was demonstrated using Southern and ELISA. The detached leaf bit bioassay with selected T1 plants showed 83.33-90.00% mortality against C. medinalis. The whole plant bioassay for T1 plants with rice leaffolder showed significant level of resistance even at a lower level of Cry2AX1 expression varying from 131 to 158 ng/g fresh leaf tissue during tillering stage.

Transgenic rice plants expressing synthetic cry2AX1 gene exhibits resistance to rice leaffolder (Cnaphalocrosis medinalis).

Bacillus thuringiensis is a major source of insecticidal genes imparting insect resistance in transgenic plants. Level of expression of transgenes in transgenic plants is important to achieve desirable level of resistance against target insects. In order to achieve desirable level of expression, rice chloroplast transit peptide sequence was fused with synthetic cry2AX1 gene to target its protein in chloroplasts. Sixteen PCR positive lines of rice were generated by Agrobacterium mediated transformation using immature embryos. Southern blot hybridization analysis of T0 transgenic plants confirmed the integration of cry2AX1 gene in two to five locations of rice genome and ELISA demonstrated its expression. Concentration of Cry2AX1 in transgenic rice events ranged 5.0-120 ng/g of fresh leaf tissue. Insect bioassay of T0 transgenic rice plants against neonate larvae of rice leaffolder showed larval mortality ranging between 20 and 80 % in comparison to control plant. Stable inheritance and expression of cry2AX1 gene was demonstrated in T1 progenies through Southern and ELISA. In T1 progenies, the highest concentration of Cry2AX1 and mortality of rice leaffolder larvae were recorded as 150 ng/g of fresh leaf tissue and 80 %, respectively. The Cry2AX1 expression even at a very low concentration (120-150 ng/g) in transgenic rice plants was found effective against rice leaffolder larvae.

In silico screening for novel inhibitors of DNA polymerase III alpha subunit of Mycobacterium tuberculosis (MtbDnaE2, H37Rv).

Tuberculosis, a pandemic disease is caused by Mycobacterium tuberculosis (Mtb). DNA polymerase III encoded by DnaE2 of Mtb is specifically required for its survival in vivo, and hence can be considered to be a potential drug target. Amino acid sequence analysis of the MtbDnaE2 and its human counterpart does not show any significant similarity. Therefore, a 3D model of the MtbDnaE2 was generated using Modeller 9v10 with the template structure of E. Coli DNA polymerase III alpha subunit (2HNH_A). The generated models were validated using a number of programmes such as RAMPAGE/PROCHECK, VERIFY_3D, and ProSA. MtbDnaE2 has few conserved residues and four conserved domains similar to that present in DNA polymerase III of E. coli. In silico screening was performed with bioactive anti-tuberculosis compounds and 6-AU (a known inhibitor of DNA polymerase III of Bacillus subtilis) and its analogues against the modeled MtbDnaE2 structure. Docking was performed using GOLD v5.2 software which resulted in the identification of top ten compounds with high GOLD fitness scores and binding affinity (X-Score). To further evaluate the efficacy of these compounds, in silico ADMET analysis was performed using MedChem Designer v3. Given their high binding affinity to the targeted MtbDnaE2, which is essential for DNA replication in the Mtb and good ADMET properties, these compounds are promising candidates for further evaluation and development as anti-tubercular agents.

A maize α-zein promoter drives an endosperm-specific expression of transgene in rice.

An alpha-zein promoter isolated from maize containing P-box, E motif sequence TGTAAAGT, opaque-2 box and TATA box was studied for its tissue-specific expression in rice. A 1,098 bp promoter region of alpha-zein gene, fused to the upstream of gusA reporter gene was used for transforming rice immature embryos (ASD 16 or IR 64) via the particle bombardment-mediated method. PCR analysis of putative transformants demonstrated the presence of transgenes (the zein promoter, gusA and hpt). Nineteen out of 37 and two out of five events generated from ASD 16 and IR 64 were found to be GUS-positive. A histological staining analysis performed on sections of mature T1 seeds revealed that the GUS expression was limited to the endosperm and not to the pericarp or the endothelial region. GUS expression was observed only in the following seed development stages : milky (14-15 DAF), soft dough (17-18 DAF), hard dough (20-23 DAF), and mature stages (28-30 DAF) of zein-gusA transformed (T0) plants. On the contrary a constitutive expression of GUS was evident in CaMV35S-gusA plants. PCR and Southern blotting analyses on T1 plants demonstrated a stable integration and inheritance of transgene in the subsequent T1 generation. GUS assay on T2 seeds revealed that the expression of gusA gene driven by alpha-zein promoter was stable and tissue-specific over two generations. Results suggest that this alpha-zein promoter could serve as an alternative promoter to drive endosperm-specific expression of transgenes in rice and other cereal transformation experiments.

Potential use of Pseudomonas koreensis AGB-1 in association with Miscanthus sinensis to remediate heavy metal(loid)-contaminated mining site soil.

Endophytic bacteria have the potential to promote plant growth and heavy metal(loid) (HM) removal from contaminated soil. Pseudomonas koreensis AGB-1, isolated from roots of Miscanthus sinensis growing in mine-tailing soil, exhibited high tolerance to HMs and plant growth promoting traits. Transmission electron microscope (TEM) analysis revealed that AGB-1 sequestered HMs extracellularly and their accumulation was visible as dark metal complexes on bacterial surfaces and outside of the cells. DNA sequencing of HM resistance marker genes indicated high homology to the appropriate regions of the arsB, ACR3(1), aoxB, and bmtA determinants. Inoculating mining site soil with AGB-1 increased M. sinensis biomass by 54%, chlorophyll by 27%, and protein content by 28%. High superoxide dismutase and catalase activities, and the lower malondialdehyde content of plants growing in AGB-1-inoculated soil indicate reduced oxidative stress. Metal(loid) concentrations in roots and shoots of plants grown in inoculated soil were higher than those of the controls in pot trials with mine tailing soil. Results suggest that AGB-1 can be used in association with M. sinensis to promote phytostabilization and remediation of HM-contaminated sites.

Insight into the effect of inhibitor resistant S130G mutant on physico-chemical properties of SHV type beta-lactamase: a molecular dynamics study.

Bacterial resistance is a serious threat to human health. The production of ß-lactamase, which inactivates ß-lactams is most common cause of resistance to the ß-lactam antibiotics. The Class A enzymes are most frequently encountered among the four ß-lactamases in the clinic isolates. Mutations in class A ß-lactamases play a crucial role in substrate and inhibitor specificity. SHV and TEM type are known to be most common class A ß-lactamases. In the present study, we have analyzed the effect of inhibitor resistant S130G point mutation of SHV type Class-A ß-lactamase using molecular dynamics and other in silico approaches. Our study involved the use of different in silico methods to investigate the affect of S130G point mutation on the major physico-chemical properties of SHV type class A ß-lactamase. We have used molecular dynamics approach to compare the dynamic behaviour of native and S130G mutant form of SHV ß-lactamase by analyzing different properties like root mean square deviation (RMSD), H-bond, Radius of gyration (Rg) and RMS fluctuation of mutation. The results clearly suggest notable loss in the stability of S130G mutant that may further lead to decrease in substrate specificity of SHV. Molecular docking further indicates that S130G mutation decreases the binding affinity of all the three inhibitors in clinical practice.

Transcriptome analysis of salinity responsiveness in contrasting genotypes of finger millet (Eleusine coracana L.) through RNA-sequencing.

Finger millet (Eleusine coracana L.) is a hardy cereal known for its superior level of tolerance against drought, salinity, diseases and its nutritional properties. In this study, attempts were made to unravel the physiological and molecular basis of salinity tolerance in two contrasting finger millet genotypes viz., CO 12 and Trichy 1. Physiological studies revealed that the tolerant genotype Trichy 1 had lower Na(+) to K(+) ratio in leaves and shoots, higher growth rate (osmotic tolerance) and ability to accumulate higher amount of total soluble sugar in leaves under salinity stress. We sequenced the salinity responsive leaf transcriptome of contrasting finger millet genotypes using IonProton platform and generated 27.91 million reads. Mapping and annotation of finger millet transcripts against rice gene models led to the identification of salinity responsive genes and genotype specific responses. Several functional groups of genes like transporters, transcription factors, genes involved in cell signaling, osmotic homeostasis and biosynthesis of compatible solutes were found to be highly up-regulated in the tolerant Trichy 1. Salinity stress inhibited photosynthetic capacity and photosynthesis related genes in the susceptible genotype CO 12. Several genes involved in cell growth and differentiation were found to be up-regulated in both the genotypes but more specifically in tolerant genotype. Genes involved in flavonoid biosynthesis were found to be down-regulated specifically in the salinity tolerant Trichy 1. This study provides a genome-wide transcriptional analysis of two finger millet genotypes differing in their level of salinity tolerance during a gradually progressing salinity stress under greenhouse conditions.

Differential regulation of defense-related gene expression in response to red rot pathogen Colletotrichum falcatum infection in sugarcane.

Red rot is a serious disease of sugarcane caused by the fungus Colletotrichum falcatum imposing a considerable economic loss annually in all sugarcane-producing countries. In this study, we analyzed the early resistance response of sugarcane to red rot fungus by comparing the differences between control and inoculated stalk tissues. Differential display reverse transcription polymerase chain reaction (DD-RT-PCR) was employed to identify altered expression of genes in disease-resistant cv Co 93009, in response to pathogen infection. DD-RT-PCR identified 300 differentially expressed transcripts of which 112 were selected for further analysis. Cloning and sequence analysis of the isolated cDNA fragments resulted in functional categorization of these clones into five categories, of which the defense/stress/signaling group was the largest, with clones homologous to genes known to be actively involved in various pathogenesis-related functions in plant species. This group showed overexpression of several transcripts related to ethylene-mediated and jasmonic acid pathway of plant defense mechanisms. Of the 112 expressed sequence tags, validation of expression was carried out for five important genes whose role in plant defense mechanisms is well established. This is the first report of Colletotrichum-mediated gene regulation in sugarcane which has provided a set of candidate genes for detailed molecular dissection of signaling and defense responses in tropical sugarcane during the onset of red rot resistance.

In-silico designing of a potent analogue against HIV-1 Nef protein and protease by predicting its interaction network with host cell proteins.

BACKGROUND: HIV-1 has numerous proteins encoded within its genome, which acquaints it with the required arsenal to establish a favorable host cell environment suitable for viral replication and pathogenesis. Among these proteins, one protein that is indispensable and ambiguous is the Nef protein. AIM: Interaction of Nef protein with different host-cell protein was predicted and subsequently the down regulation of cluster of differentiation 4 (CD4) was targeted through designing of inhibitors of Nef protein for either preventing or if not at least delaying pathogenesis. MATERIALS AND METHODS: The interaction network of Nef protein with host-cell proteins were predicted by PIMRider. Analogue of Lopinavir were prepared and evaluated considering all factors affecting the drug stability and toxicity. Finally Docking simulation were performed using an Auto-Dock Tool 4.0. RESULTS: In the interaction network of Nef protein with different host-cell proteins it was found out that 22 host cell proteins are involved in the interaction and execution of different types of functions in host cell but these functions are altered with the interaction with the Nef protein. After extensive and controlled in silico analysis it has been observed that the analogue LOPI1 binds to Nef protein (2NEF) at CD4 interacting site residues giving minimum binding energy of -7.68 Kcal/mole, low Ki value of 2.34 µM, maximum number of hydrogen bonds (8), good absorption, distribution, metabolism and excretion properties, and less toxicity in comparison with the standard Lopinavir against HIV1 protease (1HPV). CONCLUSION: The newly designed analogue (LOPI1) is showing significant in silico interaction with Nef protein and protease and can be taken forward as a potent drug lead, which may finally emerge out to be even better than the standard Lopinavir.

Landslide hazard assessment: recent trends and techniques.

Landslide hazard assessment is an important step towards landslide hazard and risk management. There are several methods of Landslide Hazard Zonation (LHZ) viz. heuristic, semi quantitative, quantitative, probabilistic and multi-criteria decision making process. However, no one method is accepted universally for effective assessment of landslide hazards. In recent years, several attempts have been made to apply different methods of LHZ and to compare results in order to find the best suited model. This paper presents the review of researches on landslide hazard mapping published in recent years. The advanced multivariate techniques are proved to be effective in spatial prediction of landslides with high degree of accuracy. Physical process based models also perform well in LHZ mapping even in the areas with poor database. Multi-criteria decision making approach also play significant role in determining relative importance of landslide causative factors in slope instability process. Remote Sensing and Geographical Information System (GIS) are powerful tools to assess landslide hazards and are being used extensively in landslide researches since last decade. Aerial photographs and high resolution satellite data are useful in detection, mapping and monitoring landslide processes. GIS based LHZ models helps not only to map and monitor landslides but also to predict future slope failures. The advancements in Geo-spatial technologies have opened the doors for detailed and accurate assessment of landslide hazards.

Novel mutations in calcium/calmodulin-dependent protein kinase IV (CAMK4) gene in infertile men.

Calcium/calmodulin-dependent protein kinase IV (CAMK4) is a multifunctional serine/threonine protein kinase, which plays an important role in the spermatogenesis by phosphorylating protamines. It has been shown to be involved in the regulation of human sperm motility. Moreover, the Camk4 knockout mice were infertile because of severely reduced sperm count and morphological abnormalities. As no study is available on the association of this gene with male infertility, we analysed all the exons of CAMK4 gene in ethnically matched 283 infertile and 268 fertile Indian men. We identified twenty nucleotide substitutions, of which twelve were novel. Of these novel variants, eight were exclusively detected in infertile men. Moreover, two infertile men-specific mutations were non-synonymous replacing amino acids at the highly conserved region. In silico analysis predicted both of these mutations as 'deleterious'. In addition to nucleotide substitutions, we identified five novel insertion-deletion mutations; of these, g.150264_66delGCG was exclusively found in two oligoasthenoteratozoospermic men. In silico analysis of infertile men exclusive mutations predicted that they can alter/diminish the potential binding sites of splicing factors, which may affect the mRNA splicing and protein translation. Our study suggests that the mutations in CAMK4 may lead to abnormal semen parameters.

Portulaca oleracea L. extract ameliorates the cisplatin-induced toxicity in chick embryonic liver.

Cisplatin, a cytotoxic agent used in treating cancer, at high doses induces hepatotoxicity. In this study, we investigated the protective role of aqueous extract of aerial parts of Portulaca oleracea L. (Po) against cisplatin-induced hepatotoxicity in chick embryonic liver. A group of 12 day old chick embryos, acclimatized to laboratory conditions were treated with a single dose of cisplatin (100 microg), while another group received Po extract at different doses (1 and 3 mg) 6 h prior to cisplatin treatment. The biochemical parameters were estimated after 24 and 72 h of incubation. A dose-dependent increase in biochemical parameters, such as alanine transaminase, aspartate transaminase, alkaline phosphatase, lactate dehydrogenase, malondialdehyde levels and a decrease in antioxidant enzymes levels like superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione-s-transferase and reduced glutathione were observed in cisplatin-treated animals, indicating a definite damage to the liver tissue. Pre-treatment with Po extract was found to provide significant protection against cisplatin-induced hepatotoxicity, as evident by the recovered levels of the altered changes in the measured biochemical parameters.

Embryo protective effect of pomegranate (Punica granatum L.) fruit extract in adriamycin-induced oxidative stress.

The possible protective role of pomegranate (Punica granatum L.) fruit extract which has shown antioxidant capacity higher than that of red wine and green tea was evaluated against adriamycin-induced oxidative stress in chick embryos. Adriamycin (ADR), an anthracycline broad spectrum of chemotherapeutic drug is used for the treatment of variety of cancers; however, its prolonged use is limited by an irreversible, dose-dependant and progressive cardiomyopathy, hepatotoxicity and general toxicity to other organs in human beings, due to oxidative stress. The morphological changes (malformation of different organs), changes in body weight, volume of amniotic fluid (AF) and biochemical parameters of AF were studied after 24 and 48 h of incubation by comparing ADR alone and pomegranate fruit extract pretreated groups with their respective controls of 12 days old chick embryos. ADR alone at a dose of 70 microg/egg showed a significant dose versus time- dependent reduction in body weight, volume of AF. A dose-related increase in embryo gross morphological deformities and significant changes in the levels of biochemical parameters in AF were observed in ADR-treated group. These changes were significantly ameliorated to normal by pre-administration of pomegranate fruit extract at a dose of 200 microg/egg. Thus, the present study demonstrated the embryo protective nature of pomegranate fruit extract against ADR-induced oxidative stress.