High quality RNA extraction protocol for polyphenolics-rich Cotton tissue.

The extraction of high-quality RNA from cotton (Gossypium spp.) is challenging because of the presence of high polyphenolics, polysaccharides, quinones, and other secondary metabolites. A high-throughput RNA extraction protocol is a prerequisite. This Triton-X-100-based RNA extraction method utilizes Polyvinyl pyrrolidone polymer (PVPP) treatment which efficiently removes phenolics, and the application of Lithium chloride (LiCl) has been found that successfully precipitated the high-quality RNA from cotton tissue. Cytoplasmic male sterility (CMS) is a maternally inherited trait associated with specific mitochondrial genome rearrangements or mutations. The suitability of RNA extracted from Cotton CMS lines was assessed. cDNA was synthesized from RNA and assayed for mitochondrial genes (cox3, nad3, nad9) associated with male sterility. This paper discuss the advantages and limitation of this protocol over existing protocol for RNA extraction for polyphenolics-rich plant tissue.

Verbal autopsy to assess causes of mortality among the economically productive age group in the tribal region of Melghat, central India.

Background & Objectives: Verbal autopsy (VA) is the systematic and retrospective inquiry (from relatives) about the symptoms of an illness prior to death. In tribal India, 67-75 per cent of deaths occur at home with an unknown cause of death (CoD). Hence, the aim of this study was to determine the CoD in the 16-60 yr age group utilizing VA. Methods: A prospective, community based longitudinal study was conducted in 32 tribal villages in the Melghat region of Maharashtra, between 2004 and 2020. Number of deaths and VAs in 16-60 yr age group were collected by village health workers (VHWs) and supervisors, verified by five different persons (internal-external) and cross-checked by three VA interpretation trained physicians. A modified version of WHO VA was used. Cause-specific mortality fractions were calculated. Results: Of the 1011 deaths recorded, mortality in males was significantly higher than females (P<0.001). A total of 763 VAs were conducted which revealed that tuberculosis was the leading CoD, followed by jaundice, heart diseases, diarrhoea, central nervous system infections and suicide. Suicides were significantly more common among males than in females (P=0.046). Significantly, more deaths occurred during the monsoon (P=0.002), especially diarrhoeal deaths (P=0.024). Interpretation & conclusions: The findings of this study suggest that, in Indian tribal areas, infectious diseases are the leading causes of morbidity and one of the major causes of deaths in economically productive age group. Intensified VHW-mediated interventions are required to reduce the premature deaths.

Phase 1 Study of the Anti-HtrA1 Antibody-binding Fragment FHTR2163 in Geographic Atrophy Secondary to Age-related Macular Degeneration.

PURPOSE: FHTR2163 is a novel antigen-binding fragment (Fab) directed against high-temperature requirement protein A1 (HtrA1). HTRA1 inhibition may preserve retinal integrity and slow disease progression in geographic atrophy (GA) secondary to age-related macular degeneration (AMD). This study examined the safety, pharmacokinetics, immunogenicity, and changes in the HTRA1-specific substrate Dickkop-related protein 3 (DKK3) in patients with GA who received FHTR2163. DESIGN: Phase I, open-label, single ascending dose escalation and multiple-dose expansion study. METHODS: Adults aged ≥ 50 years with GA secondary to AMD with best corrected visual acuity ranging between Snellen 20/125 and 20/400 were enrolled. In the first stage, a single intravitreal injection of FHTR2163 was given in 5 dose-escalation cohorts ranging from 1 to 20 mg (n = 3 patients/cohort; n = 15 total patients). The second stage evaluated the maximum tested dose of 20 mg administered every 4 weeks for 3 doses (n = 13 patients). RESULTS: No dose limiting toxicities or ocular serious AEs were reported. The most frequently reported AEs in the study eye were conjunctival hemorrhage (n = 7), conjunctival hyperemia (n = 4), and eye pain (n = 2). No non-ocular or ocular AEs were assessed as drug related. There were no clinically significant changes in ocular exams. A sustained pharmacodynamic effect of anti-HtrA1 was observed in the aqueous humor, as measured by levels of cleaved DKK3. CONCLUSIONS: FHTR2163, a novel Fab directed against HtrA1, was well tolerated with no DLTs or significant ocular AEs. The molecule when injected intravitreally for 3 doses showed a sustained pharmacodynamic effect at the maximum tested dose of 20 mg.

Finding a way for airway: a retrospective study.

BACKGROUND: Maxillofacial trauma is complex kind of injury that requires complex treatment, hence it is difficult in selecting the type of intubation technique depending on trauma. With the advent of various technologies and devices, surgeon and anesthetist should select right method of intubation that will benefit patient. METHODS: In a retrospective study, patients of either sex, admitted in Lata Mangeshkar Hospital under Oral and Maxillofacial Surgery unit for treating maxillofacial trauma operated during year 2018 to year 2019 as elective basis were studied. RESULTS: Of 78 patients, the majority (37, 47.4%) were in the 21-30 age group, followed by the young adults 31-40 years age group (19, 24.3%). Mandible fracture was found to be the most common injury in 35 patients (44.3%) followed by fracture zygoma in 26(33.3%) patients and panfacial in eight patients (10.2%). There was frontal bone fracture in three patients (3.8%). Fiberoptic intubation under sedation was carried out in 34 (43.5%) and submental intubation in 20 (25.6%) and nasal intubation with direct visualization of vocal cords in 14 (17.9%) and blind nasal intubation was performed in eight (10.2%). CONCLUSIONS: The results of this study suggest that the old concept of securing the airway in difficult situation by tracheostomy should be revised.

Neuroendocrine and immunotoxicity of polyaromatic hydrocarbon, chrysene in crustacean post larvae.

Polyaromatic hydrocarbons are a group of chemical pollutants which cause a significant threat to the living organisms in estuaries and marine ecosystems. We report the effect of chrysene, a major PAH pollutant found in Cochin Estuary along the southwest coast of India, on the neuroendocrine and immune gene expression of the post larvae (PL-25) of Penaeus monodon. The PL- 25 of P. monodon were administered with feed coated with increasing concentrations of chrysene (1, 2 and 3 µg/g) for 10 days and the gene expression was studied on 7th, 11th and 15th day. The PL exposed to chrysene showed moulting stress and changes in the levels of moult-inhibiting hormone I (MIH I) indicated by irregular moulting in the experimental tanks. At the molecular level, the higher concentration of chrysene induced two-fold upregulation of neuroendocrine (MIH I) and downregulation of immune (ProPO and crustin) gene on the 7th day of exposure. The expression of MIH I gene reduced on withdrawing the experimental feed (on 11th day), while continued downregulation of ProPO and crustin were observed on the 11th day. The results of the present study indicate that the microgram levels of PAH can impinge the neuroendocrine and immune system of the P. monodon, which may induce morbidity and mortality to the larvae in polluted coastal ecosystems. Therefore, more attention may be given to avoid PAH pollution in the estuaries to maintain a healthy ecosystem and to protect the animals from extinction.

Simultaneous Inhibition of Angiopoietin-2 and Vascular Endothelial Growth Factor-A with Faricimab in Diabetic Macular Edema: BOULEVARD Phase 2 Randomized Trial.

PURPOSE: The phase 2 BOULEVARD trial compared safety and efficacy of faricimab, a novel bispecific antibody targeting angiopoietin-2 and vascular endothelial growth factor-A (VEGF-A), with ranibizumab in patients with diabetic macular edema (DME). DESIGN: The BOULEVARD trial (ClinicalTrials.gov identifier, NCT02699450) was a prospective, randomized, active comparator-controlled, double-masked, multicenter, phase 2 study conducted at 59 sites in the United States. PARTICIPANTS: The trial enrolled patients 18 years of age or older with center-involving DME, best-corrected visual acuity (BCVA) of 73 to 24 Early Treatment Diabetic Retinopathy Study (ETDRS) letters, and central subfield thickness (CST) of 325 µm or more. METHODS: Anti-VEGF treatment-naïve patients were randomized 1:1:1 to intravitreal 6.0 mg faricimab, 1.5 mg faricimab, or 0.3 mg ranibizumab, and patients previously treated with anti-VEGF were randomized 1:1 to 6.0 mg faricimab or 0.3 mg ranibizumab. Patients were dosed monthly for 20 weeks, followed by an observation period up to week 36 to assess durability. MAIN OUTCOME MEASURES: The prespecified primary outcome measure was mean change in BCVA from baseline at week 24 for faricimab versus ranibizumab in treatment-naïve patients. Key secondary and exploratory outcome measures included CST, Diabetic Retinopathy Severity Scale (DRSS) score, and durability as assessed by time to re-treatment. RESULTS: The trial enrolled 229 patients (168 treatment-naïve and 61 previously treated with anti-VEGF). In treatment-naïve patients, 6.0 mg faricimab, 1.5 mg faricimab, and 0.3 mg ranibizumab resulted in mean improvements of 13.9, 11.7, and 10.3 ETDRS letters from baseline, respectively. The 6.0-mg faricimab dose demonstrated a statistically significant gain of 3.6 letters over ranibizumab (P = 0.03). In both patient populations, faricimab resulted in dose-dependent reductions in CST, improvements in DRSS score, and longer time to re-treatment during the observation period compared with ranibizumab. Faricimab showed no new or unexpected safety signals. CONCLUSIONS: The BOULEVARD trial met its primary end point; faricimab demonstrated statistically superior visual acuity gains versus ranibizumab at week 24 in treatment-naïve patients. Central subfield thickness reduction, DRSS score improvement, and extended durability outcomes support the primary outcome. These findings suggest the benefit of simultaneous inhibition of angiopoietin-2 and VEGF-A with faricimab for patients with DME.

Phylogenetic implications and secondary structure analyses of Vigna mungo (L.) Hepper genotypes based on nrDNA ITS2 sequences.

The internal transcribed spacers are highly preferred nuclear markers for the phylogenetic assessment of most eukaryotes, including plants. More recently, ITS2 has shown to possess equivalent phylogenetic significance as the entire ITS region. Vigna mungo L. Hepper is comparatively less explored from the molecular aspects as compared to the other species of the Vigna genus. The study presents the intra-individual characterization of 24 distinct genotypes Vigna mungo L. Hepper, using morphological as well as nrDNA ITS2 sequences and secondary structural data. The morphological characterization has been carried out using nine important agro-morphological traits. The molecular phylogeny of the sequence data, using the maximum parsimony and neighbor joining methods, shows the significant distinction based on the haplotypic variations amongst blackgram genotypes. The ITS2 secondary structures predicted using the homology modeling approach were compatible with the eukaryote-universal ITS2 secondary structure. The sequence-structure phylogeny reconstructed using the profile neighbour joining approach, also showed the presence of haplotypic variations in form of clusters on the phylogenetic tree. Further, the high GC content in the sequence data and highly negative ΔG values of the folded secondary structures ruled out the possibilities of the presence of any pseudogenes in the data set. Our analysis recommends the use of ITS2 sequence and secondary structure data at the intraspecific levels of plant taxonomical classification. Moreover, this study for the very first time reports the combined use morphological, and molecular data (using ITS2 sequence and secondary structural information) for the characterization of plants at the varietal level of taxonomical classification.

Three-dimensional Locking Plate and Conventional Miniplates in the Treatment of Mandibular Anterior Fractures.

CONTEXT: Three-dimensional (3D) locking plates has been designed with the hypothesis that this will overcome the disadvantages of both the systems and also advantages of both systems will be combined for the management of mandibular fractures. AIMS: The purpose of this study was to evaluate the efficacy of 2-mm 3D locking miniplate in the management of anterior mandibular fracture and to compare it with Champy's miniplate. SETTINGS AND DESIGN: A prospective, randomized, clinical trial was carried out in thirty patients who were divided equally in two groups. SUBJECTS AND METHODS: Group I and Group II patients were treated with 2-mm 3D locking plates and 2-mm standard miniplates, respectively. They were evaluated according to the outcomes of the study, that is, working time, wound dehiscence, infection, segmental mobility, postoperative occlusion, need for postoperative intermaxillary fixation (IMF), and radiological evaluation of reduction and fixation. STATISTICAL ANALYSIS USED: Student's t-test and Mann-Whitney test were used to compare the two systems. The data were analyzed using Statistical Package for the Social Science version 14.0. The P value was taken as significant when <0.05 (confidence interval of 95% was taken). RESULTS: The mean duration of procedure for Group I was found to be 49.33 min, whereas for Group II was 59.67 min. There was significantly greater pain on day 1 and at 1 week in Group II patients. 6.7% (n = 1) of both groups showed incidence of infection. Postoperative stability was adequate in most cases except in one patient (n = 1) of 3D locking system, which was revealed as postoperative occlusal disharmony, unsatisfactory radiological reduction of the fracture fragments, and the segmental mobility. There was no incidence of wound dehiscence, tooth damage, and nerve damage in either group. CONCLUSIONS: The result of the study can conclude that there is no major difference between both systems in terms of treatment outcome.

The complete chloroplast genome of Cenchrus ciliaris (Poaceae).

Cenchrus ciliaris is an important pasture resource for arid region and owing to apomixis, it has been very important for development and distribution of cultivars and agro-types. In present study complete chloroplast genome of C. ciliaris was sequenced. The size of chloroplast genome is 138737 bp length with overall GC content 38.6%. It exhibited regular quadripartite structure with 81053 bp of LSC region, 16108 bp of SSC region and 20788 bp of each IR region. A total of 130 genes were identified, including 87 coding genes, 32 tRNAs, 7 ribosomal RNAs and 4 pseudogenes. Phylogenetic analysis was performed using 13 other members representing major subfamilies of Poaceae.

Effectiveness of speech language therapy either alone or with add-on computer-based language therapy software (Malayalam version) for early post stroke aphasia: A feasibility study.

CONTEXT AND AIMS: This study aimed to assess the feasibility of professional based conventional speech language therapy (SLT) either alone (Group A/less intensive) or assisted by novel computer based local language software (Group B/more intensive) for rehabilitation in early post stroke aphasia. SETTINGS AND DESIGN: Comprehensive Stroke Care Center of a tertiary health care institute situated in South India, with the study design being prospective open randomised controlled trial with blinded endpoint evaluation. MATERIAL AND METHODS: This study recruited 24 right handed first ever acute ischemic stroke patients above 15years of age affecting middle cerebral artery territory within 90days of stroke onset with baseline Western Aphasia Battery (WAB) Aphasia Quotient (AQ) score of <93.8 between September 2013 and January 2016.The recruited subjects were block randomised into either Group A/less intensive or Group B/more intensive therapy arms, in order to receive 12 therapy sessions of conventional professional based SLT of 1h each in both groups, with an additional 12h of computer based language therapy in Group B over 4weeks on a thrice weekly basis, with a follow up WAB performed at four and twelve weeks after baseline assessment. The trial was registered with Clinical trials registry India [2016/08/0120121]. STATISTICAL ANALYSIS: All the statistical analysis was carried out with IBM SPSS Statistics for Windows version 21. RESULTS: 20 subjects [14 (70%) Males; Mean age: 52.8years±SD12.04] completed the study (9 in the less intensive and 11 in the more intensive arm). The mean four weeks follow up AQ showed a significant improvement from the baseline in the total group (p value: 0.01). The rate of rise of AQ from the baseline to four weeks follow up (ΔAQ %) showed a significantly greater value for the less intensive treatment group as against the more intensive treatment group [155% (SD: 150; 95% CI: 34-275) versus 52% (SD: 42%; 95% CI: 24-80) respectively: p value: 0.053]. CONCLUSIONS: Even though the more intensive treatment arm incorporating combined professional based SLT and computer software based training fared poorer than the less intensive therapy group, this study nevertheless reinforces the feasibility of SLT in augmenting recovery of early post stroke aphasia.

DNA barcoding and traditional taxonomy: an integrated approach for biodiversity conservation.

Biological diversity is depleting at an alarming rate. Additionally, a vast amount of biodiversity still remains undiscovered. Taxonomy has been serving the purpose of describing, naming, and classifying species for more than 250 years. DNA taxonomy and barcoding have accelerated the rate of this process, thereby providing a tool for conservation practice. DNA barcoding and traditional taxonomy have their own inherent merits and demerits. The synergistic use of both methods, in the form of integrative taxonomy, has the potential to contribute to biodiversity conservation in a pragmatic timeframe and overcome their individual drawbacks. In this review, we discuss the basics of both these methods of biological identification (traditional taxonomy and DNA barcoding), the technical advances in integrative taxonomy, and future trends. We also present a comprehensive compilation of published examples of integrative taxonomy that refer to nine topics within biodiversity conservation. Morphological and molecular species limits were observed to be congruent in ∼41% of the 58 source studies. The majority of the studies highlighted the description of cryptic diversity through the use of molecular data, whereas research areas like endemism, biological invasion, and threatened species were less discussed in the literature.

Genome sequence of Ochrobactrum anthropi strain SUBG007, a plant pathogen and potential xenobiotic compounds degradation bacterium.

Ochrobactrum anthropi SUBG007 was isolated from the fruit of Prunus dulcis in Rajkot (22.30°N, 70.78°E), Gujarat, India. Here we present the 4.37 Mb genome sequence strain SUBG007, which may provide the genetic information for the application in environment pollution degradation and agriculture field. The strain also posses many genes cluster which involved in production of important secondary metabolites. The nucleotide sequence of this genome was deposited into NCBI GenBank under the accession LUAY00000000.

Changes in radical scavenging activity of normal, endoreduplicated and depolyploid root tip cells of Allium cepa.

The plant cell responds to abiotic stress conditions by adjusting its cellular metabolism and various defensive mechanisms. Cellular metabolism involves changes in the cell cycle, in which the cell undergoes repeated rounds of endocycles leading to polyploidization. Defense mechanisms such as role of antioxidants are a key to understand plant adaptation. The present work describes endoreduplication and radical scavenging activity as two different defense mechanisms adapted by plants for their survival under stress condition. The work describes linkage of these two processes with each other under abiotic stress. Endoreduplicated root tip cells of Allium cepa were depolyploidized by exogenous phytohormones. Further, free radical scavenging activity from normal, endoreduplicated and depolyploidized root tips cells was observed to understand the role of phytohormones. Elevated free radical scavenging potential was observed in endoreduplicated cells compared to normal and depolyploidized cells. Based on these results, it was concluded that endoreduplication and antioxidant pathways are linked with each other through phytohormonal activities. The concentration of auxin and cytokinin regulates the activity of ascorbate oxidase enzyme, which in turn maintains the concentration of AsA within the cell. AsA level directs the prolyl-hydroxylation process of cell division proteins in quiescent center cells either toward endoreduplication process or cell division process.

Moult-inhibiting fusion protein augments while polyclonal antisera attenuate moult stages and duration in Penaeus monodon.

Moulting in crustaceans is regulated by moult-inhibiting hormone (MIH) of the CHH family neuropeptides. The inhibitory functions of MIH have pivotal roles in growth and reproduction of Penaeus monodon. In this study, we report the expression of a thioredoxin-fused mature MIH I protein (mf-PmMIH I) of P. monodon in a bacterial system and its use as antigen to raise polyclonal antiserum (anti-mf-PmMIH I). The mature MIH I gene of 231bp, that codes for 77 amino acids, was cloned into the Escherichia coli thioredoxin gene fusion expression system. The translation expression vector construct (mf-PmMIH I+pET32a+) upon induction produced 29.85kDa mature MIH I fusion protein (mf-PmMIH I). The purified fusion protein was used as exogenous MIH I and as antigen to raise polyclonal antisera. When fusion protein (mf-PmMIH I) was injected into D2 and D3 stages of juvenile shrimp, the moult cycle duration was extended significantly to 16.67±1.03 and 14.67±1.03days respectively compared to that of 11.67±1.03days in controls. Moult duration was further reduced to 8.33±0.82days when polyclonal antiserum (anti-mf-PmMIH I - 1:500 dilutions) was injected. Anti-mf-PmMIH I immunolocalized MIH I producing neurosecretory cells in the eyestalk of P. monodon. In short, the present manuscript reports an innovative means of moult regulation in P. monodon with thioredoxin fused MIH I and antisera developed.

Impaired telomerase activity hinders proliferation and in vitro transformation of Penaeus monodon lymphoid cells.

Retaining terminal transferase activity of telomerase, the ribonucleoprotein enzyme which add telomeric repeats on chromosome end is thought to be required to prevent cellular ageing. Additionally, telomerase considered as a marker for cell proliferation and immortalization in eukaryotes. We examined telomerase activity in tissues and lymphoid cell culture of Penaeus monodon. Along with telomerase activity, telomere repeats and an attempt on identification of telomerase reverse transcriptase (PmTERT) were made. Telomeric repeat amplification protocol revealed that telomerase-dependent telomeric lengthening has been taking place in P. monodon and the adult tissues were retaining this capacity throughout their lifespan with the highest activity in ovary, testis and lymphoid organ. However, telomerase activity could not be detected in lymphoid cells in culture. The canonical telomeric repeats added by telomerase of lymphoid tissue extract were identified as TTAGG, but pentameric repeats GGTTA and AGGTT were also added by the telomerase. PmTERT protein sequence (partial) shared 100 % identity with the TERT sequence of Daphnia pulex, 27 % sequence identity with Purple sea urchin and 24-25 % with Zebra fish. Undetectable telomerase activity in lymphoid cell culture supports the hypothesis that the inadequate telomerase activity or gene expression may be a reason that prevents neoplastic transformation and spontaneous immortalization of the cells in vitro. Thus, it is envisaged that telomerase activation in lymphoid cells may surmount cellular ageing for in vitro transformation and cell line establishment.

Whole genome sequences and annotation of Micrococcus luteus SUBG006, a novel phytopathogen of mango.

Actinobaceria, Micrococcus luteus SUBG006 was isolated from infected leaves of Mangifera indica L. vr. Nylon in Rajkot, (22.30°N, 70.78°E), Gujarat, India. The genome size is 3.86 Mb with G + C content of 69.80% and contains 112 rRNA sequences (5S, 16S and 23S). The whole genome sequencing has been deposited in DDBJ/EMBL/GenBank under the accession number JOKP00000000.

Whole genome sequencing of Halomonas sp. SUBG004 isolated from Little Rann of Kutch, a desert of India.

A salt tolerant strain, designated as SUBG004, was isolated from the desert of India, Little Rann of Kutch. The organism is a Gram-negative, facultatively anaerobic and rod shaped bacterium. Chemotaxonomic and phylogenetic properties were consistent with its classification in the genus Halomonas. Here we report the whole genome sequence of Halomonas sp. SUBG004 deposited in DDBJ/EMBL/GenBank under accession number JPEU0100000 which provides insights for salt stress adaptation through betaine synthesis.

High-quality complete genome sequence of Microbacterium sp. SUBG005, a plant pathogen.

Microbacterium sp. SUBG005 is a Gram positive bacterium, isolated from infected leaf of Mangifera indica L. in Rajkot (22.30°N, 70.78°E), Gujarat, India. The genome sequencing of Microbacterium sp. SUBG005 is having type I secretion system genes of pathogenicity as well as heavy metal resistance unique genes. The genome size is 7.01 Mb with G + C content of 64.80% and contains rRNA sequences. Genome sequencing analysis provides information about the microbe role in host-pathogen interaction. The whole genome sequencing has been deposited in DDBJ/EMBL/GenBank under the accession number JNNT00000000.

Genomic analysis of novel phytopathogenic Georgenia sp. strain SUB25.

A Gram positive bacterium, Georgenia sp. SUB25 was isolated from infected leaves of Solanum lycopersicum L. in Rajkot (22.30°N, 70.78°E), Gujarat, India. We sequenced and analyzed Georgenia sp. SUB25 that is novel plant pathogen using next generation sequencing platform and assembly yielded contigs representing a size of 4.84 Mb with 81 tRNAs and 88 rRNAs. The whole genome sequencing has been deposited in DDBJ/EMBL/GenBank under the accession number JNFL00000000. This genome sequence contains Type II secretion system genes, which involved in pathogenicity mechanism that may help to understand plant microbial interaction.

X-ray computed tomography to study rice (Oryza sativa L.) panicle development.

Computational tomography is an important technique for developing digital agricultural models that may help farmers and breeders for increasing crop quality and yield. In the present study an attempt has been made to understand rice seed development within the panicle at different developmental stages using this technique. During the first phase of cell division the Hounsfield Unit (HU) value remained low, increased in the dry matter accumulation phase, and finally reached a maximum at the maturation stage. HU value and seed dry weight showed a linear relationship in the varieties studied. This relationship was confirmed subsequently using seven other varieties. This is therefore an easy, simple, and non-invasive technique which may help breeders to select the best varieties. In addition, it may also help farmers to optimize post-anthesis agronomic practices as well as deciding the crop harvest time for higher grain yield.