Development of loop mediated isothermal amplification (LAMP): A new tool for rapid diagnosis of cotton leaf curl viral disease.

Cotton leaf curl disease (CLCuD) ranks top among all endemic diseases transmitted by whitefly (Bemisia tabaci) affecting cotton (Gossypium hirsutum) causing severe economic losses to the cotton growers in the Indian subcontinent. For its effective management, robust tools for detection are a prerequisite and it is important to diagnose the virus titre in early stage of infection in plants as well as in the disease transmitting vector. Considering the limitations in current PCR-based techniques we have standardised rapid and sensitive Loop Mediated Isothermal Amplification (LAMP) protocol for the diagnosis of cotton leaf curl virus (CLCuV) in cotton leaves and in its transmitting vector whitefly. Perhaps, this is the first report of use of LAMP tool for rapid diagnosis of CLCuV in cotton and its transmitting vector the whitefly. Further, the colorimetric detection for diagnostic simplicity of amplified LAMP product by using different dyes lead to enhanced applicability of this technique in the field of disease diagnostics. The merit of present study is that the diagnostic failure of PCR and LAMP due to low virus titre in the infected leaf has been circumvented through the combination of rolling circle amplification (RCA) with LAMP. Thus RCA-LAMP can be an option for ultra-sensitive detection of samples with low virus titre. The potential applications of this advanced diagnostic tool in laboratory research on diagnosis of CLCuV, an important viral pathogen of cotton have been discussed.

Genome-wide identification and expression analysis of the plant specific LIM genes in Gossypium arboreum under phytohormone, salt and pathogen stress.

Asiatic cotton (Gossypium arboreum) cultivated as 'desi cotton' in India, is renowned for its climate resilience and robustness against biotic and abiotic stresses. The genome of G. arboreum is therefore, considered as a valued reserve of information for discovering novel genes or gene functions for trait improvements in the present context of cotton cultivation world-wide. In the present study, we carried out genome-wide analysis of LIM gene family in desi cotton and identified twenty LIM domain proteins (GaLIMs) which include sixteen animals CRP-like GaLIMs and four plant specific GaLIMs with presence (GaDA1) or absence (GaDAR) of UIM (Ubiquitin Interacting Motifs). Among the sixteen CRP-like GaLIMs, eleven had two conventional LIM domains while, five had single LIM domain which was not reported in LIM gene family of the plant species studied, except in Brassica rapa. Phylogenetic analysis of these twenty GaLIM proteins in comparison with LIMs of Arabidopsis, chickpea and poplar categorized them into distinct αLIM1, ßLIM1, γLIM2, δLIM2 groups in CRP-like LIMs, and GaDA1 and GaDAR in plant specific LIMs group. Domain analysis had revealed consensus [(C-X2-C-X17-H-X2-C)-X2-(C-X2-C-X17-C-X2-H)] and [(C-X2-C-X17-H-X2-C)-X2-(C-X4-C-X15-C-X2-H)] being conserved as first and/or second LIM domains of animal CRP-like GaLIMs, respectively. Interestingly, single LIM domain containing GaLIM15 was found to contain unique consensus with longer inter-zinc-motif spacer but shorter second zinc finger motif. All twenty GaLIMs showed variable spatio-temporal expression patterns and accordingly further categorized into distinct groups of αLIM1, ßLIM1, γLIM2 δLIM2 and plant specific LIM (DA1/DAR). For the first time, response of GaDA1/DAR under the influence of biotic and abiotic stresses were studied in cotton, involving treatments with phytohormones (Jasmonic acid and Abscisic acid), salt (NaCl) and wilt causing pathogen (Fusarium oxysporum). Expressions patterns of GaDA1/DAR showed variable response and identified GaDA2 as a probable candidate gene for stress tolerance in G. arboreum.

Degree day-based model predicts pink bollworm phenology across geographical locations of subtropics and semi-arid tropics of India.

There is a global concern about the effects of climate change driven shifts in species phenology on crop pests. Using geographically and temporally extensive data set of moth trap catches and temperatures across the cotton growing states of India, we predicted the phenology of cotton pink bollworm Pectinophora gossypiella (Saunders). Our approach was centered on growing degree days (GDD), a measure of thermal accumulation that provides a mechanistic link between climate change and species' phenology. The phenology change was predicted by calculating absolute error associated with DD and ordinal date, an alternative predictor of phenology, for peak moth abundance. Our results show that GDD outperformed the ordinal dates in predicting peak moth abundance in 6 out of 10 selected locations. Using established thresholds of 13.0/34.0 °C, mean DD accumulated between the consecutive moth peaks across different years were estimated at 504.05 ± 4.84. Seven generations were determined for pink bollworm in a cropping season, the length of which varied between 35 and 73 days in response to temperature. Pink bollworm population reached its peak during third generation which can be the target for management actions. The study provides essential information for developing pink bollworm management strategies under climate change.

Evidence for population expansion of Cotton pink bollworm Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae) in India.

Pink bollworm, Pectinophora gossypiella (Saunders) infestation on Bt cotton is a major concern to cotton production in India. The genetic diversity and phylogeographic structure of the insect in light of PBW resistance needs to be revisited. The objective of this study was to identify different haplotypes of pink bollworm and their distribution in India. To achieve this we studied the population structure in 44 cotton growing districts of India. The partial mitochondrial COI sequence analyses of 214 pink bollworm populations collected from 44 geographical locations representing 9 cotton growing states of India were analysed. Genetic diversity analysis exhibited presence of 27 haplotypes, among them Pg_H1 and Pg_H2 were the most common and were present in 143 and 32 populations, respectively. Distributions of pairwise differences obtained with partial COI gene data from the overall Indian populations are unimodal, suggesting population expansion in India. Significant neutrality test on the basis of Tajima' D and Fu's Fs presented a star-shaped haplotype network together with multiple haplotypes. The unimodal mismatch distribution, rejection of neutrality test with significant negative values supported the theory of demographic expansion in cotton pink bollworm populations in India. Genetic data not only provides us with a perspective of population genetics, but also that the two populations of pink bollworm, those occurring early in the season are genetically close to the late season populations with respect to their partial CO1 region. Resistance to Cry toxins does not seem to have had an impact on this region of the mt DNA in populations of pink bollworm.

Morpho-physiological and molecular characterization of drought tolerance traits in Gossypium hirsutum genotypes under drought stress.

Drought stress is one of the major abiotic stresses affecting lint yield and fibre quality in cotton. With increase in population, degrading natural resources and frequent drought occurrences, development of high yielding, drought tolerant cotton cultivars is critical for sustainable cotton production across countries. Six Gossypium hirsutum genotypes identified for drought tolerance, wider adaptability and better fibre quality traits were characterized for various morpho-physiological and biochemical characters and their molecular basis was investigated under drought stress. Under drought conditions, genotypes revealed statistically significant differences for all the morpho-physiological and biochemical traits. The interaction (genotype × treatment) effects were highly significant for root length, excised leaf water loss and cell membrane thermostability indicating differential interaction of genotypes under control and stress conditions. Correlation studies revealed that under drought stress, relative water content had significant positive correlation with root length and root-to-shoot ratio while it had significant negative correlation with excised leaf water loss, epicuticular wax, proline, potassium and total soluble sugar content. Analysis of expression of fourteen drought stress related genes under water stress indicated that both ABA dependent and ABA independent mechanisms of drought tolerance might be operating differentially in the studied genotypes. IC325280 and LRA5166 exhibited ABA mediated expression of stress responsive genes and traits. Molecular basis of drought tolerance in IC357406, Suraj, IC259637 and CNH 28I genotypes could be attributed to ABA independent pathway. Based on physiological phenotyping, the genotypes IC325280 and IC357406 were identified to possess better root traits and LRA5166 was found to have enhanced cellular level tolerance. Variety Suraj exhibited good osmotic adjustment and better root traits to withstand water stress. The identified drought component trait(s) in specific genotypes would pave way for their pyramiding through marker assisted cotton breeding.

Clinical Profile and Treatment Outcome of Drug Resistant Tuberculosis Patients of Western Maharashtra, India.

BACKGROUND: Drug-resistant tuberculosis (DR-TB) imposes formidable burden on national health systems. OBJECTIVES: This study aims to evaluate clinical profile and treatment outcome of drug resistant tuberculosis patients at tertiary care centre using a standardised treatment regimen (STR). METHODS: Retrospective analysis of 146 patients with DR-TB from Solapur district who were treated with STR from period of September 2012 to December 2014 was done. Statistical analysis of treatment outcome data was done to know predictors of treatment success of DR-TB. RESULTS: Out of total 146 bacteriologically proven cases of DR-TB, 95 were males and 51 females. 41% of patients were residents of Solapur city and the rest from different parts of Solapur district. Out of the 146 patients, 130 (89%) patients achieved sputum culture conversion within three months. Treatment outcome of these patients was as followstreatment success in 84 (58%), 20(14%) died, 28(19%) defaulted and failure in 14(9%) patients. Three predictors were identified for successful treatment outcome of DR-TB that include urban residence, patients with chest x-ray findings of moderately advanced disease and patients whose DR-TB status diagnosed by genexpert technology. CONCLUSION: In resource-poor settings, well-designed STR under national programme provides satisfactory results.