Reverse Genetics in Ecological Research

By precisely manipulating the expression of individual genetic elements thought to be important for ecological performance, reverse genetics has the potential to revolutionize plant ecology. However, untested concerns about possible side-effects of the transformation technique, caused by Agrobacterium infection and tissue culture, on plant performance have stymied research by requiring onerous sample sizes. We compare 5 independently transformed Nicotiana attenuata lines harboring empty vector control (EVC) T-DNA lacking silencing information with isogenic wild types (WT), and measured a battery of ecologically relevant traits, known to be important in plant-herbivore interactions: phytohormones, secondary metabolites, growth and fitness parameters under stringent competitive conditions, and transcriptional regulation with microarrays. As a positive control, we included a line silenced in trypsin proteinase inhibitor gene (TPI) expression, a potent anti-herbivore defense known to exact fitness costs in its expression, in the analysis. The experiment was conducted twice, with 10 and 20 biological replicates per genotype. For all parameters, we detected no difference between any EVC and WT lines, but could readily detect a fitness benefit of silencing TPI production. A statistical power analyses revealed that the minimum sample sizes required for detecting significant fitness differences between EVC and WT was 2–3 orders of magnitude larger than the 10 replicates required to detect a fitness effect of TPI silencing. We conclude that possible side-effects of transformation are far too low to obfuscate the study of ecologically relevant phenotypes

More frequent, more costly? Health economic modelling aspects of monitoring glaucoma patients in England

BACKGROUND: Chronic open angle glaucoma (COAG) is an age-related eye disease causing irreversible loss of visual field (VF). Health service delivery for COAG is challenging given the large number of diagnosed patients requiring lifelong periodic monitoring by hospital eye services. Yet frequent examination better determines disease worsening and speed of VF loss under treatment. We examine the cost-effectiveness of increasing frequency of VF examinations during follow-up using a health economic model. METHODS: Two different VF monitoring schemes defined as current practice (annual VF testing) and proposed practice (three VF tests per year in the first 2 years after diagnosis) were examined. A purpose written health economic Markov model is used to test the hypothesis that cost effectiveness improves by implementing proposed practice on groups of patients stratified by age and severity of COAG. Further, a new component of the model, estimating costs of visual impairment, was added. Results were derived from a simulated cohort of 10000 patients with quality-adjusted life years (QALYs) and incremental cost-effectiveness ratios (ICERs) used as main outcome measures. RESULTS: An ICER of £21,392 per QALY was derived for proposed practice improving to a value of £11,382 once savings for prevented visual impairment was added to the model. Proposed practice was more cost-effective in younger patients. Proposed practice for patients with advanced disease at diagnosis generated ICERs > £60,000 per QALY; these cases would likely be on the most intensive treatment pathway making clinical information on speed of VF loss redundant. Sensitivity analysis indicated results to be robust in relation to hypothetical willingness to pay threshold identified by national guidelines, although greatest uncertainty was allied to estimates of implementation and visual impairment costs. CONCLUSION: Increasing VF monitoring at the earliest stages of follow-up for COAG appears to be cost-effective depending on reasonable assumptions about implementation costs. Our health economic model highlights benefits of stratifying patients to more or less monitoring based on age and stage of disease at diagnosis; a prospective study is needed to prove these findings. Further, this works highlights gaps in knowledge about long term costs of visual impairment

Micropropagation and conservation of selected endangered anticancer medicinal plants from the Western Ghats of India

Globally, cancer is a constant battle which severely affects the human population. The major limitations of the anticancer drugs are the deleterious side effects on the quality of life. Plants play a vital role in curing many diseases with minimal or no side effects. Phytocompounds derived from various medicinal plants serve as the best source of drugs to treat cancer. The global demand for phytomedicines is mostly reached by the medicinal herbs from the tropical nations of the world even though many plant species are threatened with extinction. India is one of the mega diverse countries of the world due to its ecological habitats, latitudinal variation, and diverse climatic range. Western Ghats of India is one of the most important depositories of endemic herbs. It is found along the stretch of south western part of India and constitutes rain forest with more than 4000 diverse medicinal plant species. In recent times, many of these therapeutically valued herbs have become endangered and are being included under the red-listed plant category in this region. Due to a sharp rise in the demand for plant-based products, this rich collection is diminishing at an alarming rate that eventually triggered dangerous to biodiversity. Thus, conservation of the endangered medicinal plants has become a matter of importance. The conservation by using only in situ approaches may not be sufficient enough to safeguard such a huge bio-resource of endangered medicinal plants. Hence, the use of biotechnological methods would be vital to complement the ex vitro protection programs and help to reestablish endangered plant species. In this backdrop, the key tools of biotechnology that could assist plant conservation were developed in terms of in vitro regeneration, seed banking, DNA storage, pollen storage, germplasm storage, gene bank (field gene banking), tissue bank, and cryopreservation. In this chapter, an attempt has been made to critically review major endangered medicinal plants that possess anticancer compounds and their conservation aspects by integrating various biotechnological tool

Effective refractive error coverage in adults aged 50 years and older: estimates from population-based surveys in 61 countries

Background: In 2021, WHO Member States endorsed a global target of a 40-percentage-point increase in effective refractive error coverage (eREC; with a 6/12 visual acuity threshold) by 2030. This study models global and regional estimates of eREC as a baseline for the WHO initiative. Methods: The Vision Loss Expert Group analysed data from 565 448 participants of 169 population-based eye surveys conducted since 2000 to calculate eREC (met need/[met need + undermet need + unmet need]). A binary logistic regression model was used to estimate eREC by Global Burden of Disease (GBD) Study super region among adults aged 50 years and older. Findings: In 2021, distance eREC was 79·1% (95% CI 72·4–85·0) in the high-income super region; 62·1% (54·7–68·8) in north Africa and Middle East; 49·5% (45·0–54·0) in central Europe, eastern Europe, and central Asia; 40·0% (31·7–48·2) in southeast Asia, east Asia, and Oceania; 34·5% (29·4–40·0) in Latin America and the Caribbean; 9·0% (6·5–12·0) in south Asia; and 5·7% (3·1–9·0) in sub-Saharan Africa. eREC was higher in men and reduced with increasing age. Global distance eREC increased from 2000 to 2021 by 19·0%. Global near vision eREC for 2021 was 20·5% (95% CI 17·8–24·4). Interpretation: Over the past 20 years, distance eREC has increased in each super region yet the WHO target will require substantial improvements in quantity and quality of refractive services in particular for near vision impairment. Funding: WHO, Sightsavers, The Fred Hollows Foundation, Fondation Thea, Brien Holden Vision Institute, Lions Clubs International Foundation

FLOW CYTOMETRIC AND KARYOLOGICAL ANALYSIS OF POLYSOMATY AND POLYPLOIDIZATION DURING CALLUS FORMATION FROM LEAF SEGMENTS OF VARIOUS POTATO GENOTYPES

Flow cytometry and karyological analysis were used to study polysomaty and polyploidization during the first 15 days of callus formation in leaf segments from shoot cultures and greenhouse-grown plants of various lines and genotypes of Solanum tuberosum and S. phureja. The greenhouse-grown plants showed a higher degree of polysomaty (77% and 49% of polyploidized nuclei) than the shoot cultures (< 3%). During the in vitro culture period, polyploidization occurred through endoreduplication. Segments of the five shoot cultures showed up to 87%, 53%, 59%, 45% and 56% polyploidization, respectively; the DNA content of corresponding interphase nuclei amounted to 8C, 16C, 16C, 16C and 8C, and the chromosome numbers to 96. Segments from the two greenhouse-grown genotypes showed up to 87% and 84% polyploidization; the DNA content amounted to 32C and 16C, and the chromosome numbers to 192 and 96. The number of reduplication cycles was species-dependent; the degree of polyploidization was dependent on the initial ploidy level of the genotypes. Cell proliferation did not take place at a constant rate. The maximum frequencies of metaphases (52–171 per segment) occurred after 1 week of culture and were correlated with the ploidy level of the genotypes. Cells were triggered to mitosis rather than to endoreduplication. Cell cycles with normal monochromosomes could be shorter than 1 day, and those with diplochromosomes lasted at least 1 day. Polysomaty, degree of polyploidization and abnormal nuclear processes are discussed in relation to the origin of genetic instability early in cultur