Biosensors in plant pathogen detection is one of the recent advancements in plant pathology for the detection of plant pathogens

1. BIOSENSORS IN PLANT
PATHOGEN DETECTION
S. HARISHINI
REG. NO: 207090
II-M.SC AGRICULTUR
DEPT. OF PLANT PATHOLOGY,
ANNAMALAI UNIVERSITY.
1

2. BIOSENSORS
✓ An analytical device used
for the detection of a
chemical substance, that
combines a biological
component with a
physiochemical detector.
✓ Father of Biosensors –
Leland C. Clark.
2
(Khater et al. 2017)

3. COMPONENTS OF A BIOSENSOR
BIOLOGICAL
COMPONENTS
• Polysaccharides,
• Microorganisms,
• Nucleic acid,
• Tissue,
• Enzyme,
• Antibody.
PHYSICAL
COMPONENTS
• Transducers,
• A/D Converter,
• Amplifier,
• Display
3
(Ali et al. 2017)

4. WORKING PRINCIPLE
4

5. 5

6. Singh et al. 2013)
6

7. BIO-ELEMENT AND SENSOR ELEMENT
COUPLING MECHANISMS
A. Covalent
fabrication,
B. Matrix
immobilization,
C. Membrane
encapsulation,
D. Physical
adsorption
fabrication.
7

8. (Ali et al. 2017)
8

9. TYPES OF TRANSDUCERS
9
(Rani et al. 2019)

10. TYPES OF BIOSENSORS
Electrochemical Biosensors,
Optical Biosensors,
Phage-based Biosensors,
Graphene-based Biosensors,
E-Nose based Biosensors,
Magnetic recognition Biosensors,
Mobile Biosensing,
Food system based (Post-harvest),
Agricultural Biosensors,
Nanotechnology based
Biosensors.
10

11. 1. ELECTROCHEMICAL BIOSENSORS
▪ Two core components – Molecular recognition layer, an
electrochemical transducer.
▪ Detection of target pathogens under different conditions – Air,
water and on seeds.
▪ Different platforms – Greenhouses, in-field and in postharvest
storage vessels.
▪ Advantageous biological sensing components – Plant’s antibody
and DNA.
(Dyussembayev et al. 2021)
11

12. CLASSIFICATION:
Electrochemical
biosensors
A. Antibody
based
Label-free EIS
QCM
Microfluidic
ECEIA
B. DNA based
Voltammetric
detection
Label free
impedimetric
method 12

13. ANTIBODY BASED
BIOSENSORS
• High sensitivity and
specificity.
• Detection of target antigens
at low concentration.
• High affinity level.
• Fundamental principle –
Biomolecular interaction
between analyte and
antibody.
DNA-BASED BIOSENSORS
• Hybridization between
target DNA sequence and
DNA probe sequence.
• Use of Nano structured
materials:
✓Gold
✓Cadmium sulfide
✓Silver.
• On-site environmental
monitoring
13
(Prittesh et al. 2021)

14. 14
(Fang and Ramasamy 2015)

15. A. ANTIBODY BASED BIOSENSORS
1. LABEL FREE ELECTROCHEMICAL IMPEDANCE
SPECTROSCOPY (EIS) BASED DETECTION
✓Self-assembled monolayers – Immobilization method.
✓Achievement of better Antigen-Antibody interaction efficiency.
✓Most reported substrate – Thiol SAMs formation on gold electrodes.
Examples:
1. Detection of Plum pox virus (PPV) on gold electrodes.
2. Detection of Prunus necrotic ringspot virus (PNRSV) using carbon
electrodes. 15
(Sambasivam et al. 2021)

16. EIS FOR PNRSV
16
(Khater et al. 2017)

17. 2. LABEL-FREE QUARTZ CRYSTAL MICROBALANCE (QCM)-
BASED APPROACHES
✓Oscillation frequency based.
✓Measurement of very small mass changes.
✓Alternative bioreceptor – DNA probe.
✓New biorecognition elements – Aptamers.
✓Identification of orchid viruses.
✓Piezoelectric label-free immunosensors based.
Examples:
1. QCM immunosensor based on SAMs – Maize chlorotic mottle virus
2. Higher selectivity against Wheat streak mosaic virus.
17

18. SUPPORTIVE ARTCLE
18

19. 3. MICROFLUIDIC IMMUNOSENSOR
✓Amperometric detection.
✓Three microfabricated gold electrodes used.
✓Introduction of solutions in the sensor – A syringe pump
system.
✓Detection of Absorbance – Bio Rad Benchmark microplate
reader.
✓Electrode – Glass combination.
Example:
Early detection of Xanthomonas arabicola in walnut samples.
19
(Lie et al. 2006)

20. 4. ENZYMATIC LABEL BASED TECHNOLOGY
(INDIRECT ELISA)
✓Electrochemical enzyme-linked immunoassay (ECEIA).
✓Amperometric and Voltammetric techniques.
✓Enzyme labels – Horseradish peroxidase (HRP), Alkaline
phosphate (AP).
✓Sensitivity – 10 times higher than that of standard ELISA.
✓Measurement of current – Linear sweep voltammetry using a
hanging mercury electrode.
(Khater et al. 2017)
20

21. Examples:
1. Detection of Cucumber mosaic virus
▪ Substrates – o-aminophenol and o-phenylenediamine.
2. Detection of Tobacco mosaic virus, Potato virus Y, Southern bean mosaic
virus, Tomato aspermy virus, Turnip mosaic virus.
3. PSS antigen:
✓Detection of panicle blight,
✓Cercospora leaf spot on rice,
✓ Black spot of crucifer.
21

22. B. DNA-BASED BIOSENSORS
22

23. 1. DNA HYBRIDIZATION VOLTAMMETRIC DETECTION
✓Amperometric and voltametric, impedimetric detection.
✓Detection of pathogen related DNA with complementary target
immobilized on the electrode monitored by OSWV.
✓Hybridization, Redox indicator – Methylene Blue.
Example:
1. Voltammetric detection of Sugarcane white leaf disease on Glassy
electode modified with chitosan.
2. High selectivity for identification of Trichoderma harzianum against
other Trichoderma sp.
(Abdhurehman and Jaffer 2020)
23

24. 2. LABEL – FREE IMPEDIMETRIC METHOD
▪ Quantitative detection of CTV by Label less
impedimetric biosensor.
▪ Thiolated DNA probe immobilized and optimized for
DNA hybridization.
▪ Method of high potential.
▪ Gold nano particles distributed on carbon electrodes.
24

25. LABEL FREE HYBRIDIZATION FOR
PLUM POX DETECTION
(Khater et al. 2017)
25

26. 26

27. 2. OPTICAL BIOSENSORS
Measurement of interaction by:
❑Light source,
❑An optical transmission medium,
❑An immobilized biorecognition element and,
❑A signal detection system.
27

28. CLASSIFICATION:
Optical biosensors
A. Antibody
based
Lateral flow
immunoassay
SPR
Microsphere
immunoassay
B. DNA based
Bridging
Flocculation
AuNP based
Electrochemilu
minescence 28

29. A. ANTIBODY BASED BIOSENSORS
1. LATERAL FLOW IMMUNOASSAYS (LFIAs)
▪ Paper analytical device.
▪ Main LF formats – Sandwich and Lateral flow
Immunoassays.
▪ Labelling – AuNPs, Magnetic nanoparticles,
Fluorescent nanoparticles, Enzymes.
▪ First LFIA – TMV Detection.
(Tothill 2001)
29

30. 30

31. 2. SURFACE PLASMON RESONANCE
31
❑ Label-free,
❑ Real-time,
❑ Highly accurate,
❑ Detection of orchid viruses – CMV, ORSV

32. 3. MICROSPHERE IMMUNOASSAY
➢Application of Fluorescent loaded magnetic microsphere and fluorophore
antibodies for detection.
➢Sensitivity and the ability to detect multiple pathogens in single assay.
➢Limitation – Complexity of assay, need of fluorescent readers.
32

33. B. DNA BASED BIOSENSORS
1. BRIDGING FLOCCULATION
▪ Reversible adsorption to differentiate long and short
DNA polymers.
▪ Main applications is the visual detection of:
✓ Pseudomonas syringae,
✓Fusarium oxysporum and Botrytis cinerea.
▪ Detection in very early stage.
(Sambasivam et al. 2021)
33

34. 34

35. 2. LATERAL FLOW BASED ON DNA HYBRIDIZATION FOR
BBTV
35

36. LATERAL FLOW
SYSTEMS FOR THE
DETECTION OF
PATHOGENS:
A – Phytophthora sp.
B – Erwinia amylovora
C – Potato virus y
(Khater et al. 2017)
36

37. 37

38. 38

39. A – MEDIATED ELECTRON
TRANSFER.
B – DIRECT ELECTRON TRANSFER 39
3. ENZYMATIC ELECTROCHEMICAL
BIOSENSORS

40. 4. PHAGE – BASED BIOSENSORS
▪ Interaction between phage and the targeted bacterial component.
▪ Very sensitive, cost effective approach.
▪ Stable at high temperature.
Example:
Phage-based Magnoelastic biosensors for the detection of S.
typhimurium on the surface of tomato and spinach leaves.
40

41. (Fang and Ramasamy 2015)
41

42. 5. GRAPHENE BASED BIOSENSORS
42
(Bahamonde et al. 2018)

43. SUPPORTIVE ARTICLE
43

44. 6. E-NOSE BIOSENSORS
▪ Detection – Changes in VOC
▪ Disease diagnosis in wheat and
pear plants.
▪ High stability and Easy
processing of data.
▪ Soil-borne pathogen detection.
(Abdallahi et al. 2020 )
44

45. SUPPORTIVE ARTICLE
45

46. 7. NMR BASED BIOSENSORS
▪ Magnetic recognition
transduction.
▪ Target bacteria – Antibody
based SPIONS.
▪ Superparamagnetics discs for
bacteria biosensing.
(Mclamore et al. 2021)
46

47. 8. MOBILE BIOSENSING
✓ Portable optical
biosensors,
✓ Portable electrochemical
biosensors.
✓ Portable NMR biosensors.
47
(Mclamore et al. 2021)

48. 8. FOOD SYSTEM BASED BIOSENSORS
▪ Optical-SPR
biosensors.
▪ Post harvest pathogen
detection.
✓ Botrytis sp.
✓ Aspergillus,
✓ Colletotrichum.
48
(Jazib et al. 2017)

49. 9. AGRICULTURAL BIOSENSORS
❑Plant wearable biosensors,
❑Multiplexing sensors,
❑NMR and MRI,
❑Smart farming,
❑UAVs for near field communication
(Mclamore et al. 2021)
49

50. 10. NANOTECHNOLOGY BASED
BIOSENSORS
1. FSNP– Detection of
Xanthomonas axonopodis
pv. vesicatoria.
2. CuO – Detection of
Aspergillus niger.
3. AgNPs – Soil borne
pathogens.
50
(Adetunji et al. 2018)

51. IMPORTANCE OF NANOPARTICLES IN
BIOSENSORS
• High and targeted surface area,
• Particle size and charge,
• Core and surface properties,
• Size and Flexibilities,
• Multivalency and controlled synthesis.
(Mudasir et al. 2021)
51

52. 52

53. (Fang and Ramasamy 2015)
53

54. PROPERTIES OF A GOOD BIOSENSOR
▪ Highly specific for the analyte,
▪ Response should be linear over a broad spectrum range,
▪ Small, compatible, rapid and accurate, sterilizable,
▪ Low cost and easy to use,
▪ Assay costs should be lower than the conventional tests,
▪ Assay should be fast, reliable and repeatable.
54

55. 1.
Linearity
2.
Sensitivity
3.
Selectivity
4.
Response
time
55
CHARACTERISTICS

56. APPLICATIONS OF BIOSENSORS
56

57. CHALLENGES AND FUTURE
PERSPECTIVES
• Need of careful validation within a particular pathosystem.
• Pathogen detection and quantification should be used in
conjugation with other multiple factors of farming systems.
• Cultural cropping and climatic factors should be considered.
• Futureproofing of IDM strategies.
• Validated biosensors improve the accuracy of epidemiological
models.
(Dyssembayev et al 2021)
57

58. SUPPORTIVE
ARTICLES
58

59. 59

60. 60

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63. 63

64. CONCLUSION
64

65. 65