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Masters’ Seminar - 591

Abiotic Stress Tolerance In Cucurbits

PRABHAT KUMAR SINGH BAC/M/HORT-V/004/2015-16

Department of Horticulture (Veg. & Flori.) Bihar Agricultural University, Sabour, Bhagalpur-

813 210 (Bihar)

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1. CUCURBITS AND THEIR IMPORTANCE

2. ABIOTIC STRESS AND GLOBAL SCENARIO

3. DROUGHT STRESS EFFECT AND TOLERANCE

4. SALINITY STRESS EFFECT AND TOLERANCE

5. THERMAL STRESS EFFECT AND TOLERANCE

6. SECONDARY STRESSES

7. CASE STUDY 1

8. CASE STUDY 2

9. CONCUSION

CONTENTS

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Cucurbits and Their Importance118 genera and 825 species. 14 species widely cultivated in IndiaShares 5.6 % of total vegetable produced in IndiaConsumed in various form-

o Salad (cucumber, gherkins, long melon)o Sweet (ash gourd, pointed gourd)o Pickles (gherkins)o Deserts (melons)

Proteins with abortifacient, ribosome inactivating, immunomodulatory, anti-tumor and anti-AIDS activities have been extracted-o Momordicin (seeds of Momordica charantia)o Trichosanthin (tubers of Trichosanthes kirilowii)o Beta-trichosanthin (tubers of Trichosanthes cucumeroides)o Luffin-a and luffin-b(seeds of Luffa cylindrica) Rai et al. 2008

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Introduction

Change in surrounding environmental factors which adversely affects plant growth and crop productivity.

Abiotic Stress

Stress

Biotic Stress

Due to pathogens, pests, weeds, etc.

Moisture

The Fraction of World Arable Land Subjected to an Abiotic Stress

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Abiotic stress Fraction (%)of arable land

Drought 26

Mineral 20

Freezing 15

No stress 10

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Estimated potential yield losses are-Drought 17 %

Salinity 20 %

High Temperature 40 %

Low Temperature 15 %

Other Stress 8 %

(Ashraf et al., 2008)

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The inadequacy of water availability leading to moisture stress (Sinha,1986)

Plant unable to meet evapotranspiration demand (Blum, 1988)

Moisture stress enhances salinity stress, in saline soil

Results into low water potential due to high solute concentration

Effects Of Drought Stress on Cucurbits

Cucurbits are high water consuming plants have large leaf area,

high water content in fruit

Long-term drought stress causes flower and fruit drop in watermelon (Kaya et al., 2003 )

Affects growth, photosynthesis, nitrogen metabolism, carbohydrate metabolism, and

decrease in nucleic acid and protein (Talbi et al., 2015)

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Drought ToleranceCrop can minimise the loss by-

Drought escape

Dehydration avoidance• Reduced transpiration• Osmotic adjustment By maintaining turgor pressure• Abscisic acid (ABA) Stomata closure

Dehydration tolerance• Enhanced antioxidant enzyme activities• Proline accumulation (Chopra and Paroda, 1986)

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Drought stress tolerance through self-response mechanisms is limited, it can be mediated by exogenous means–Melatonin promotes water-stress tolerance, as it strengthen roots, reduce chlorophyll degradation, increase

antioxidants and ROS-scavenging enzymes in cucumber (Cucumis sativus L.) (Zhang et al., 2012)

Application of beneficial microorganisms such as Arbuscular Mycorrhizal(AM) fungi (Huang et al., 2011)

Grafting a susceptible scion on a resistant rootstock, e.g. grafting watermelon on a commercial rootstock,

PS1313(Cucurbita maxima Duchesne X C. moschata Duchesne)(Schwarz el at., 2010)

Brassinosteroids- Induced antioxidant enzymes such as Superoxide dismutase (SOD), Catalase (CAT) and

Ascorbate peroxidase (APX) associated with drought tolerance (Xia et al., 2009)

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Salinity Stress

Salt affected soils may be grouped into-

• Saline soil

• Sodic soil

Effect of salinity stress on cucurbits-

• Water stress generated by the salts in the soil

• Mineral toxicity stress caused by the salts

• Disturbance in the mineral nutrition of the plant

Marginal yellowing of melon leaves caused by salt toxicity

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Cont..

• Crops with salt injury tend to be more susceptible

to wilting during the hottest hours of the day, even

when moisture level is sufficient

• Majority of cucurbits- moderately sensitive e.g.

muskmelon and cucumber

• Ash gourd and bitter gourd reported to be tolerant

while, squash is moderate tolerantBurning of cucumber leaf margins

due to salt toxicity

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Tolerance To Salt Stress

• Soil reclamation and management practices

• Development of salt tolerant lines by conventional breeding. e.g.

Cucumber lines-CRC-8, CHC-2, G-338, CH-20, and 11411 Sare

Musk melon lines- Calif-525

• Through genetic engineering by using HAL1 gene of Saccharomyces cerevisiae as molecular

tool in watermelon and other crop species (Ellul et al., 2003)

• Grafting as a tool to manage salinity stress e.g.

• Bottle gourd rootstock-grafting promotes photosynthesis by regulating the stomata and non-stomata

performances in leaves of watermelon seedlings under NaCl stress (Yang et al., 2015)

• Watermelon ‘Crimson Tide’ grafted onto rootstock of C. maxima results increase in higher growth under

saline soil (Yeliser and Vygus, 2006)

(Munns et al., 2006)

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Temperature StressHeat stress

Chilling stress

Freezing stress

Effect of heat stress

• Decrease in the photosynthetic rate and increase in respiration

• High temperature and long day condition results more male flowers

• Heat stress of plant will show itself by wilting

• Crops like squashes, melons, cucumber, pumpkins drops their blossom at high temperature

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Tolerance To Heat Stress

• Crop management practices like mulching and the use of shelters protects vegetables from

heat stress

• Stabilization of enzymes and membranes by Heat hardening (50 min at 50◦C)

• Shifts in protein synthesis (e.g. heat shock proteins)

• Osmoregulation ( e.g. Proline and Glycine-betadine:- Protects enzymes and cellular

membrane against heat damage)

• Ethylene biosynthesis inhibitors partly alleviate the effect of heat stress which have positive

effect on leaf water status and stomatal opening (Qin et al., 2007)

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Effect of Chilling and Freezing Stress• Affects seed germination and seedling growth

• Affects water uptake, causing plants to suffer from water stress (Bloom et al., 2004)

• Reduces plant growth and development, causing wilt and necrosis (Ahnet et al., 1999)

• Results in abnormal flower and failure of fruit set

• Freezing may cause membrane disruptions, super cooling of plant water

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Tolerance to Chilling and Freezing Stress

Root-Zone Warming Differently Benefits Mature and Newly Unfolded Leaves of Cucumis sativus L.

Seedlings under chilling Stress in greenhose (Wang et al., 2016)

Brassinosteroids induces tolerance to low temperature by triggering the generation of H2O2 as a

signalling molecule by increased NADPH Oxidase activity, in cucumber leaves (Zhou et al., 2011)

Salicylic acid induces enhancement of cold tolerance through activation of antioxidative capacity

in watermelon (Yang et al., 2008)

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Cont..

Grafting as a tool to manage low temperature stress

e.g.

Figleaf gourd (Cucurbita ficifolia Bouche) and bur cucumber (Sicos angulatus L.) as

rootstocks for cucumber

Grafting of a cucumber scion onto a squash rootstock (Cucurbita moschata Duch)

increase low temperature tolerance

For watermelon, grafting onto Shin-tosa-type (an interspecific squash hybrid,

Cucurbita maxima×C. moschata) rootstocks enhance tolerance (Dietmar et al., 2011)

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Drought

SaltHeat

PLANT CELL

Overproduction of reactive oxygen species (ROS)

Oxidative damage

Response to biotic and abiotic environmental stimuli, cause membrane lipid peroxidation, protein denaturation, and nucleic acid damage

Involved in

Secondary Abiotic Stress

Osmotic stressOxidative stress

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To avoid damage by ROS,

production and detoxification of

ROS is sustained by enzymatic

and non enzymatic antioxidants

Antioxidant enzymes protect

plants against oxidative damage

by scavenging toxic ROS and

restoring redox homeostasis

(Mittler et al., 2004)

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CASE STUDY 1

Regulation of Plant Growth, Photosynthesis,

Antioxidation and Osmosis by an Arbuscular

Mycorrhizal Fungus in Watermelon Seedlings

under Well-Watered and Drought Conditions

Yanling et al., 2016

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For this experiment, drought-sensitive watermelon variety (Y34) and

AM inocula of Glomus versiforme was used

The treatments consisted of

i. Well- watered plants without AM inoculation (WW-NM)

ii. Well- watered plants with AM inoculation (WW+M)

iii. Drought- stressed plants without AM inoculation (DS-NM)

iv. Drought-stressed plants with AM inoculation (DS+M)

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Based on this observation AM Colonization and Plant Growth

The development of the arbuscular mycorrhizal (AM) fungus in watermelon roots (as revealed by trypan blue staining)

Morphological characteristics of the mycorrhizal (+M) and non-mycorrhizal (-NM) watermelon seedlings grown under well-watered(WW) and drought-stressed (DS) conditions

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Leaf Water Status, Chlorophyll Concentration, and Chloroplast Ultrastructure

A-Leaf relative water content (RWC) B-Chlorophyll concentration Mesophyll cells in mycorrhizal (+M) and non-mycorrhizal (-NM) watermelon seedlings grown under WW and DS conditions

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Initial Rubisco Activity, and Photosynthetic and Chlorophyll Fluorescence Parameters

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Total Soluble Sugar and Proline Contents

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CASE STUDY 2

High Throughput Sequencing of Small RNAs in the

Two Cucurbita Germplasm with Different Sodium

Accumulation Patterns Identifies Novel MicroRNAs

Involved in Salt Stress Response

Junjun et al., 2015

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Two Cucurbita germplasm (Inbred lines), namely, N12 (C. maxima Duch.) and N15 (C. moschata

Duch.) with different sodium accumulation pattern were used. Total RNA was isolated from N12

and N15 roots under salt stress or control. Each biological replicate was equally mixed and

designated as-

24hNR- N12 root samples of salt stress

24hR- N12 root samples of control

54hNR- N15 root samples of salt stress

54hR- N15 root samples of control

For each sample, a minimum of 40 mg of total RNA was used for Illumina sequencing from the

Beijing Genomics Institute (BGI, Shenzhen, China)

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Based on the observationN12 and N15 exhibited different Na accumulation patterns

Na concentrations in the leaves, stems and roots (A) and Na distribution (B) in N12 and N15 treated with 100 mM NaCl for 10d

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Identification of conserved miRNAs

Numbers of identical miRNA members in each conserved miRNA family in the four libraries

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Confirmation of predicted miRNAs and target genes by qRTPCR

Expression analysis of miRNAs in N12 by qRT-PCR Expression analysis of miRNAs in N15 by qRTPCR

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From observation it can be inferred that

Two Cucurbita germplasm, namely, N12 (C. maxima Duch.) and N15 (C. moschata

Duch.), exhibited different Na accumulation patterns. N12 accumulated Na in the

shoots, whereas N15 accumulated Na in the roots

• The differential expression of conserved miRNAs and novel miRNAs under salt

stress conditions between the two varieties and their target genes indicated that

novel miRNAs plays important roles in the response of the two Cucurbita

germplasm to salt treatment

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CONCLUSION By use of different crop management practices we can minimise the effect of environmental stress

Grafting of tolerant root stock is gain over slow conventional breeding method practices

Use of exogenous means of beneficial micro-organism like AM fungi, phytohdroxysteriod like

brassinosteroids, use of osmoprotectant, heat shock protein in overcoming the environmental

stress.

Novel microRNA might plays important role in response to salt stress in two Cucurbita germplasm.

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Future Prospects

Through screening of more abiotic stress tolerant genotypes of cucurbits and

hybridising them with high yielding genotypes or transfer of tolerant genes

by back cross breeding method, can overcome environmental stress.

More study is required in the field of novel micro RNA and their targeted

genes in Cucurbits.

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Thank you…..