1WELCOME

BIG EVENTS HAPPEN IN

SMALL WORLD

ROLE OF NANOTECHNOLOGY FOR CROP PROTECTION IN HORTICULTURE

Speaker

Girija Kumari, Ch.

ID No. 10468

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Seminar Incharge

Dr. T. M. Rao

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Nanotechnology/Nanoscience

Nanotechnology

The design, characterization, production and application of

structures, devices and systems by controlling shape and size

at the nanoscale

British Standards Institution (BSI 2005)

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Nano scale Nano scale

The term ‘nanotechnology’ is based on the prefix ‘nano’- Greek word

meaning ‘dwarf’

Word ‘nano’ means 10 ⁹ or one billionth part of a metreˉ

1 nanometre= one billionth (10 ⁹) of metreˉ

Size range between 1 and 100 nm

The term ‘nanotechnology’ is based on the prefix ‘nano’- Greek word

meaning ‘dwarf’

Word ‘nano’ means 10 ⁹ or one billionth part of a metreˉ

1 nanometre= one billionth (10 ⁹) of metreˉ

Size range between 1 and 100 nm

Nanoscale

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Richard Feynman, Physicist

“ The father of nanotechnology”

Richard Feynman, Physicist

“ The father of nanotechnology”

“There’s Plenty of Room at the Bottom”

- at American Physical Society meeting at the California Institute of Technology on Dec-29, 1959.

“There’s Plenty of Room at the Bottom”

- at American Physical Society meeting at the California Institute of Technology on Dec-29, 1959.

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Richard Feynman“Adapability to manipulate, control, assemble, produce and

manufacture things at atomic precision”

Norio Taniguchi, Professor

- coined the term “Nanotechnology” (1974)

Norio Taniguchi, Professor

- coined the term “Nanotechnology” (1974)

“Nano-technology’’ - Processing, separation, consolidation and

deformation of materials by one atom or by one molecule.

“Nano-technology’’ - Processing, separation, consolidation and

deformation of materials by one atom or by one molecule.

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Nanotechnology received its greatest momentum with the invention of the scanning tunneling microscope (STM)

It was invented by Gerd K. Binning and Heinrich Rohrer in 1985

What STM does?

It allows imaging solid surfaces with atomic scale resolution. It operates based on tunneling current, which starts to flow when a sharp tip is mounted on a piezoelectric scanner approaches a conducting surface at a distance of about 1 nm. This scanning is recorded and displayed as an image of the surface can be resolved an displayed using STM

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Scanning Tunneling Microscopy

Time Line of Nanotechnology

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~ 2000 Years Ago

Sulfide nanocrystals used by Greeks and Romans to dye hair

~ 1000 Years Ago

Gold nanoparticles of different sizes used to produce different colors in stained glass windows

1959 “There is plenty of room at the bottom” by R. Feynman1974 “Nanotechnology” - Taniguchi uses the term nanotechnology for the first

time

1981 IBM develops Scanning Tunneling Microscope1985 “Buckyball” - Scientists at Rice University and University of Sussex

discover C60

1986 • “Engines of Creation” - First book on nanotechnology by K. Eric Drexler.

• Atomic Force Microscope invented by Binnig, Quate and Gerbe

1989 IBM logo made with individual atoms1991 Carbon nanotube discovered by S. Iijima 1999 “Nanomedicine” – 1st nanomedicine book by R. Freitas 2000 “National Nanotechnology Initiative” launched

(British Standards Institution, 2005)

Physics

BiologyMaterial Science

Medicine Engineering

Bionanoscience

Chemistry

Bionanoscience / Technology

Exploitation of biomaterials, devices or methodologies on the Nanoscale Exploitation of biomaterials, devices or methodologies on the Nanoscale

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Nanoparticles

Nanoparticles are particles between 1 and 100 nanometers in

size

 In nanotechnology a particle is defined as a small object that behaves as a whole unit with respect to its transport and properties 

Arranged or assembled into ordered layers, or mine layers

Possess distinct physical, biological and chemical properties associated with their atomic strength

(Bhattacharyya et al., 2010)

Nanoparticles are particles between 1 and 100 nanometers in

size

 In nanotechnology a particle is defined as a small object that behaves as a whole unit with respect to its transport and properties 

Arranged or assembled into ordered layers, or mine layers

Possess distinct physical, biological and chemical properties associated with their atomic strength

(Bhattacharyya et al., 2010)

Nanocarbon

FullereneTubes ConesCarbon blackHornsRodsFoamsNanodiamonds

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Carbon nanotubes

Iijima in 1991

Appeared to be made up of a perfect network of hexagonal graphite rolled up to form a tube

Exhibit unusual photochemical, electronic , thermal and mechanical properties

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Nanoparticles in disease and pest management (Crop Protection)

Biopolymer nanoparticles eg. Chitosan

Metallic nanoparticles eg. Silver nanoparticles

Silica nanoparticles

Copper nanoparticles

Zinc nanoparticles

Nanocomposites eg. Chitosan Silver NP

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ChitosanHas various applications in biology due to its biodegradable and nontoxic properties

chitosan and chitosan nanoparticles are found to be more effective against plant pathogens like Fusarium solani

The chitosan therefore could be formulated and applied as a natural antifungal agent in nanoparticles form to enhance its antifungal activity (Ing et al., 2012)

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Mode of action of chitosan against fungi can be explained by following mechanisms:

The positive charge of chitosan interacts with negatively charged phospholipid components of fungi membrane, which in turn alter cell permeability of plasma membrane and causes the leakage of cellular contents, which consequently leads to death of the cell (García-Rincón et al., 2010)

Chitosan chelates with metal ions, which has been implicated as a possible mode of antimicrobial action (Rabea et al., 2003). On binding to trace elements, it interrupts normal growth of fungi by making the essential nutrients unavailable for its development (Roller and Covill, 1999).

It is suggested that chitosan could penetrate fungal cell wall and bind to its DNA and inhibit the synthesis of mRNA and, in turn, affect the production of essential proteins and enzymes (Sudarshan et al. , 1992; Kong et al., 2010)

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Metallic nanoparticles

Metallic nanoparticles ( Ag, Zn, Cu, Si) possess unique chemical and

physical properties, small size, huge surface to volume ratio, structural

stability and strong affinity to their targets (Kumar et al., 2010)

These can be used as new antimicrobial agents and an alternative to

synthetic fungicide to delay or inhibit the growth of many pathogens

species because of its multiple mode of inhibition

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Silver nanoparticles

Silver has been used as an antimicrobial agent since ancient civilizations; it has been used extensively due to its broadspectrum and multiple modes of antimicrobial activity (Wei et al., 2009)

Silver exhibits higher toxicity to microorganism and lower toxicity to mammalian cells

The application of silver nanoparticles as antimicrobial agents is because of its economical production and multiple modes of inhibitory action to microorganisms (Clement and Jarrett, 1994)

Its specific antimicrobial mechanisms are still unclear

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Ag acts as plant-growth stimulator and reduces unwanted microorganisms in soils and hydroponics systems (Sharma et al., 2012)

Silver in ionic or nanoparticle forms has a high antimicrobial activity and is therefore widely used for various sterilization purposes (Park et al. , 2006)

Some studies found that inhibition of fungal pathogens with silver nanoparticles is concentration dependent and also on type of silver nanoparticles used

Cont........

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Silica nanoparticles

Silicon (Si) increases disease resistance and stress resistance in plants (Brecht et al., 2004)

It also stimulates the physiological activity and growth of plants (Carver et al., 1998)

Torney et al. (2007) used honeycomb mesoporous silica nanoparticle (MSN) system with 3nm pores to deliver DNA and chemicals into plant cells and intact leaves

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Copper nanoparticles

Copper-based fungicides produce highly reactive hydroxyl radicals which can damage lipids, proteins, DNA, and other biomolecules

It plays an important role in disease prevention and treatment of large variety of plants (Borkow and Gabbay, 2005)

Because of its bio-compatibility, these nanohydrogels are included as a new generation of copper-based bio-pesticides and it could also be developed into an efficient delivery system for copper based fungicides for plant protection (Brunel et al., 2013)

Complexation of copper with chitosan nanogel was shown to have strong synergistic effect between chitos an and copper in inhibiting the growth of phytopathogenic fungus Fusarium graminearum

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Zinc nanoparticles

Zinc oxide nanoparticles (ZnO NPs) could be used as an effective fungicide in agricultural and food safety applications

Mechanism of action of zinc nitrate derived nano-ZnO on important fungal pathogen Aspergillus fumigatus showed hydroxyl and superoxide radicals mediated fungal cellwall deformity and death due to high energy transfer (Prasun Patra and Goswami,2012)

ZnO nanoparticles can cause deformation of fungal hyphae and prevent the conidiophores and conidial development which ultimately leads to the death of fungal hyphae

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Nanoparticles for pest management

Globally insect pests cause a huge crop loss of 14% and plant pathogens cause an estimated loss up to 13% with a value of US $2,000 billion per year (Pimentel,2009)

Nano pesticide formulations increase the solubility of poorly soluble active ingredient and helps in releasing the active ingredient slowly

Nanoparticles are loaded with pesticides and released slowly based on environmental trigger (Lauterwasser, 2005)

Rotenone, a water-insoluble botanical insecticide used to control aphids, thrips , acari from decades , however its effective utilization has limited due to its poor water solubility, stability, degradation and isomerization when exposed to sunlight

Nanosilica showed 100% mortality against insect pests whereas nanosulfur inhibited the sporulation and growth of fungi (Goswami et al., 2010)

(Royal Society and Royal Academy of Engineering, 2004)25

METHODS OF NANOPARTICLE PRODUCTION

(Royal Society and Royal Academy of Engineering, 2004)26

TOP DOWN APPROACH

(Royal Society and Royal Academy of Engineering, 2004)27

BOTTOM UP APPORAOCH

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1 m

1 m

1 m

Each edge is 1 m Each edge is 0.1 m, but there are 1000

cube

Volume (in cubic mts ) = 1m x 1m x 1m =1m³

Surface area (in sq. m) = (1m x 1m ) x 6 sides

= 6 sq. m

Volume = (0.1 x 0.1 x 0.1) x 1000 cubes = 1m³

Surface area = (0.1 x 0.1) x 6 sides x 1000 = 60 sq.m

Mode of action of nano particles

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Dendrimers

Quantumdots

Nanosensors

FullerenesCarbon Nanotubes

Nano Chips

C60 Cadmium selinade

3D

macromolecules

Sequence nanoscale

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Tools of Nanotechnology

C60

Applications In Agriculture

31 (Mahendra et al., 2012)

32Applications of nanotechnology in crop protection and plant nutrition

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CASE STUDY : 1

Antifungal effectiveness of nanosilver colloid against rose

powdery mildew in greenhousesKim et al., 2008

Solid State Phenomena Vol. 135 , pp 15-18

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IntroductionPowdery mildew, caused by Sphaerotheca pannosa var. rosae, is one of the most common and widespread fungal diseases of greenhouse and outdoor roses which reduces flower production and causes weakening of the plants

It mainly appear first on the under surface of young leaves in early summer and the infection spreads to stems, shoots and buds

Silver have long been known to have strong antimicrobial activity and AgNP because of their specific surface area have high activity than their bulk siver metal

Nanosilver colloid that is a well-dispersed and stabilized silver nanoparticle solution will be more adhesive on bacteria and fungus and so have enhanced antibacterial activity

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Objective of the study

To examine the effectiveness of nanosilver colloid as new fungicide against rose powdery mildew in greenhouses

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Material and Methods

31.5 g of 99.8 % AgNO3, was dissolved in 3.7 l distilled water and 40g PVP as stabilizer was added

1g of 98.0 % NaBH4, as reducing agent was dissolved in distilled water of 0.2 l and this solution was slowly dropped in silver ion/PVP solution under sonication

After adding 28.5 g quaternary ammonium chloride (Cluster Instruments Co., 80 %) as another stabilizer was dissolved and vigorously stirred for 1 h

The particle size of nanosilver and UV-visible spectrum of nanosilver colloidal solutions was characterized by Transmission electron microscopy (TEM) and UV spectrometer, respectively

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The antifungal effects of nanosilver solution carried out at a commercial greenhouse (Sung-Ju Farm), located at SungJu (Gyeongsangbuk-Do, Korea), an important area for cut rose production

Rose plants, belonging to the 'Suncity' cultivar, were grown according to the cultural practices normally adopted by local growers

The nanosilver solution of 500 kg with concentration of 10 ppm was sprayed at large area of 3306 m2 infected by the rose powdery mildew

Cont....

The antifungal effects wereobserved by an optical microscope

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Results and discussion

TEM images of the nanosilver

in colloidal solution of 1000 ppm

Average size was 1.5 nm with size

distribution of 1-5 nm

In addition the images showed that

nanosilver particles were densely and well

dispersed in the colloidal solution

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Absorption spectra of a nanosilver colloidal solution afterdilution; (a) only stabilizers (b) 1 ppm, (c) 5 ppm.

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Photographs of rose effected with powdery mildew

before treatment immediately after treatment 2 days after

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Photographs of leaves with powdery mildew

Before treatment a week after treatment

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Before treatment

Optical microscope (80 magnification) images of powdery mildew on leaf of rose

week after treatment

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Conclusion of the study

The double-capsulized nanosilver showed high dispersity and stability

The photographic results showed that the effects of nanosilver colloidal solution against rose powdery mildew was very high and durable for a week

In addition, the nanosilver did not have phyto-toxicity on the plants cell of leaves, stem and buds of rose plants

As a result, well dispersive and stabilized nanosilver could be recommended as new fungicide for powdery mildew

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DNA-tagged nano gold: A New Tool For The Control Of The Spodoptera

litura Fab. (Chakravarthy et al., 2012)

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CASE STUDY : 2

African Journal of Biotechnology Vol. 11(38), pp. 9295-9301, 10 May, 2012

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Introduction

Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) is an extremely serious pest, the larvae of which can defoliate many economically important crops cutting across over 40 families

Polyphagous, voracious feeder and very prolific pest

The use of insecticides in agricultural fields leads to an ecological imbalance in nature and thus in some countries including India, several insecticides have been banned (Yadav, 2010)

A very recent approach to the control of insect pests is the use of DNA-tagged nano particles

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To examine the potential of DNA-tagged nano particles for the control of Spodoptera litura

Objective of the study

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Material and Methods

A solution of HAuCl4 dissolved in 20 ml of distilled water was used as the solvent for the preparation of gold (Au) nanoparticle

The solution was continuously stirred in a bath at 110°C for an hour and then quickly treated with C6H5Na3O7

The Au nanoparticle solution was then further reacted with an aqueous solution of calf-thymus DNA-sodium salt to obtain DNA tagged Au nanoparticles

To determine the virulence/lethal concentration of DNA-tagged nano particle solution, serial dilutions of the nano particle solution ranging from 200, 300, 400 to 500 ppm were prepared

10 μl of the suspension was dispensed on the semi-synthetic chickpea (Cicer arietinum) based diet filled into 5 ml glass vials

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Second instar larvae of S. litura of uniform age and size were released onto the diet 20 min after surface treatment with DNA-tagged gold nanoparticles, at all four concentrations viz., 200, 300, 400 and 500 ppm

A control diet was maintained where chickpea based semi-synthetic diet was applied and used without DNA-tagged gold nanoparticles

Observations of larval settlement on the diet were taken from first day onwards

Larval mortality was recorded from 3rd till 10th day at 24 h interval. Each treatment was replicated thrice

Cont.....

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Results and discussion

As the concentration and days after treatment increased, the larval

mortality of 2nd instar S. litura larvae also increased. The maximum

mortality of 30.0 (33.2), 57.5 (49.6) and 75.0 (60.5) was obtained at 500

ppm on 3rd, 4th and 5th day, respectively

There were statistically significant differences between the treatments

and the days after treatment required for 50% mortality of the larvae

At the highest concentration (500 ppm) of the DNA-tagged gold nano

particle, feeding was reduced, larvae turned sluggish and were unable

to orientate towards the source

Treatment (PPM) Percentage of larval mortality days after treatment

3rd 4th 5th

200 10.0 (16.0)b 27.5 (31.4)c 35.0 (36.0)b

300 22.5 (28.2)a 42.5(40.7)b 62.5(52.3)a

400 25.0(29.7)a 55.0 (47.9)ab 72.5 (58.6)a

500 30.0 (33.2)a 57.5 (49.6) a 75.0(60.6)a

Control 0.0 (0.6)c 0.0 (0.6)d 0.0 (0.6)c

SEM ± 2.92 2.87 3.05

CD at 5 % 8.81 8.66 9.20

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Effect Of Different Concentrations Of DNA-tagged With Nano Particle On 2nd Instar S. litura, Three,

Fourth And Fifth Days After Treatment

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Effect of different concentrations of DNA-tagged gold nanoparticles on 2nd instar S. litura larvae

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2nd instar S. litura larva at three days after treatment

2nd instar S. litura larva four days after treatment

The larvae ceased active movement, the skin and entire body became stiff and hard and oozing of the body content (lysis)

The body became swollen, pulpy and fragile. It attained almost a ‘C’- shape and body turned dark brown

2nd instar S. litura larva at four days after treatment

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2nd instar S. litura larva at five days after treatment

2nd instar S. litura larva six days after treatment

The larvae showed premature molting in and half of the body became discoloured (fleshy white) and the other half turned brown

The larvae attained pupal shape, all the internal contents oozed out, and eventually death occurred

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2nd instar S. litura larva seven days after treatment

Control

The dead larvae turned black

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Conclusion of the study

This study clearly demonstrates that DNA-tagged gold nanoparticle has a devastating effect on the larval tissue of S. Litura and would therefore be a useful component of an integrated pest management strategy

Metal nano-particles could be a better alternative to synthetic insecticides, in addition to being a toxicant that inhibits biological and physiological systems of insects

This experiments clearly established that the DNA tagged gold nano particle should be tested at concentrations higher than 500 ppm to determine the effective dose resulting in 50% larval mortality

Subsequent to the laboratory tests on S. litura, field tests on a small scale need to be initiated

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Other Applications Of Nanotechnology

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According to Mariya Khodakovskaya, a plant biologist, and Alexandru Biris, a nanotechnologist founded a technique used carbon nanotubes to encourage fast and early germination of tomato plants

Tomato seeds were planted, some with a growth medium containing carbon nanotubes, and some without nanotubes in the growth medium

It took only three days for more than 30% of the nanotube tomato seeds to begin sprouting. In that time, none of the non-treated seeds had even germinated

In fact, it took 12 days for 32% of the tomato seeds without nanotube help to germinate

After four weeks, the researchers noticed that the tomato plants that had been treated with carbon nanotubes had two times the biomass and two times the height of their non-treated counterparts. The current theory is that the nanotubes penetrate the seed coat of the tomato seeds, allowing water to more rapidly penetrate the seeds and boost their development

http://phys.org/news174066714.html#jcp

Other Applications of nanotechnology (carbon nanotubes)

Nanotechnology for seed germination

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Interestingly, the root systems were similar in all of the plants, so the nanotubes did not change the way the roots established themselves

Another issue is that the nanotubes seem to be causing abnormally long internodes, and that might affect the ultimate outcome regarding the viability of mature plantsThe mechanism of ionic interactions with the CNT surface (Miskovic 2008) implies that redox type changes of the nutrient ion of a given oxidation state might take place with the MWCNTs in the medium.

There is a potential for the utilization of CNTs for optimizing water transport in arid-zone agriculture and Horticulture and of improving crop biomass yields

http://phys.org/news174066714.html#jcp

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Nanoparticles in post-harvest disease management

Chitosan, a deacetylated derivative of chitin, is found to be very effective

in reducing postharvest decay of fruit and vegetables (Liu et al. , 2007)

Chitosan/nanosilica hybrid film, extended shelf life, reduced browning

index, retarded weight loss and inhibited the increase of malondialdehyde

amount and polyphenoloxidase activity in fresh longan fruit (Shi et al.

2013)

Pulsing of nano silver (with 2.5 nm diameters) on cut gerbera (Gerbera

jamesonii) cv. Ruikou flowers for 24 h with 5 mg/L nano solution extended

vase life and inhibited the bacteria growth in vase solution for initial 2 days

when observed in vitro under microscope (Liu et al. 2009)

Biological toxicity Environmental toxicity

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Nanotoxicity

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Conclusion

Nanoparticles are biodegradable and target specific, so they can be

successfully employed in production of nanocapsules for delivery of

fungicides, pesticides, fertilizers, and other agrochemicals

Nanotechnology will revolutionize agriculture including crop protection

in the near future

Over the next two decades, the green and golden revolutions would be

accelerated by means of nanotechnology

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Future prospects

More studies are needed to explore the mode of action of NPs, their

interaction with biomolecules, and their impact on the regulation of gene

expressions in plants

Research on nanoparticles with respect to crop protection should be

geared towards introduction of faster and ecofriendly nanoformulations in

future

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Thank you