INVESTIGATIONS ON SUGARCANE MOSAIC VIRUS IN PUNJAB AND NWFP:

CHARACTERIZATION AND IDENTIFICATION OF RESISTANCE SOURCES

 PRESENTATION BY
MUHAMMAD ASAD FAROOQ

SUPERVISORY COMMITTEE

SUPERVISOR DR. ABID RIAZ

CO-SUPERVISOR DR. TAHIRA YASMIN
MEMBER DR. S. M. MUGHAL
MEMBER HUMAYUN JAVED
INTRODUCTION
Sugarcane -- an important cash crop of
the world
-- accounts for over two third
of world sugar production
In Pakistan -- sugarcane is a high value
cash crop.
-- mainly grown for production
of essential item for
industries like sugar, chipboard,
and paper etc.
Share in -- Value added of agriculture: 3.6 %
-- GDP: 0.8 % (Anonymous, 2010).
World sugarcane yield is limited by various diseases
caused by -- fungi, bacteria, virus and phytoplasma.
A significant share of yield losses is attributed to:
VIRUSES and PHYTOPLASMA
The most important among them is:
Sugarcane Mosaic Virus (SCMV)
Belongs to family POTYVIRIDAE
Occurs -- in almost all the sugarcane growing
countries, except Mauritius and Guyana
(Viswanathan and Mohanraj, 2001).
 OBJECTIVES
• To conduct quantitative surveys of Punjab and NWFP
to study occurrence, prevalence and distribution of
SCMV.
• To identify weed hosts of SCMV.
• Identification and characterization of SCMV through:
– Symptomatology
– Differential Hosts
– Serology (DAS-ELISA)
• Screening of sugarcane germplasm against SCMV
under inoculated conditions to identify/find sources of
resistance.
MATERIALS AND METHODS
The study was conducted in the Department of
Plant Pathology, PMAS-Arid Agriculture
University, Rawalpindi in collaboration with Crop
Disease Research Program and Sugar Crops
Research Program of National Agricultural
Research Center, Islamabad during the year
2008-09.
FIELD SURVEYS AND COLLECTION OF SAMPLES
Punjab NWFP
Mandi Bahauddin Mardan
Sargodha Peshawar
Faisalabad Charsadda
Jhang Dargai
Toba Tek Singh
Okara
Rahim Yar Khan
Bahawalpur
Muzaffargarh
Layyah
Data were collected regarding:
– Percent Disease Incidence
– Relative Occurrence and
– Varietal Reaction
Percent Disease Incidence:

An area of 10-X-10 m was selected --- all plants were counted

No. of infected plants

% D.I. = --------------------------------------- x 100
Total No. of plants observed

Relative Occurrence (%):

Samples -- Whole leaves with characteristic symptoms of SCMV -- from the top
of the plants

Kept in sterilized polythene bags -- Placed in a refrigerator at 4 0C until

processed.

Tested to confirm the presence of SCMV through DAS-ELISA

The relative occurrence was determined by applying following formula;

Number of samples positive for a Virus

Relative Occurrence (%) = ------------------------------------------------------ x 100
Total number of samples tested
 MAINTENANCE OF VIRUS ISOLATES
• ELISA positive samples were stored out

• Virus from samples having maximum

concentrations were multiplied on virus
susceptible sugarcane varieties-- through
mechanical inoculation – besides glass house
conditions for further studies.
INDEXING OF SAMPLES THROUGH ELISA

• Samples -- both provinces -- tested

through DAS-ELISA – standard buffers
and protocol.

• The samples were replicated twice.

• The optical density -- measured at 405 nm

-- ELISA plate reader -- with continuous
movement absorbance mode.
 IDENTIFICATION OF WEED HOSTS
Weeds -- from sugarcane fields -- carrying SCMV.

Record of local/vernacular and botanicals names was

maintained.

All samples were kept in sterilized polythene bags --

placed in a refrigerator at 4 0C until processed.

Samples were then tested to confirm the presence of

SCMV through DAS-ELISA.
 DIFFERENTIAL HOST STUDIES
Differential hosts tested: wheat, barley, maize, millet and
sorghum.
Seeds were obtained from Crop Sciences Institute,
NARC, Islamabad.
Test plants were grown in 4 clay pots (9”dia) -- filled with
a mixture of field soil, sand and FYM (1:1:1) -- under
glasshouse conditions -- in an insect free environment.
Plants in 3 clay pots -- 3 to 4 leaves stage -- were
mechanically inoculated -- a mixture of representative
isolates -- Punjab, NWFP and Islamabad.
The plants were observed for symptom appearance
weekly – 25-30 days after inoculation the young growing
leaves were collected -- tested through ELISA to confirm
the presence or absence of SCMV.
SCREENING OF SUGARCANE GERMPLASM
Germplasm lines of diverse sources being maintained at NARC.
Hot water treatment at 55 oC for 10 minutes.

These lines were mechanically inoculated with SCMV at 3-4

leaves stage.
The plants were observed for symptom appearance weekly.
25-30 days after inoculation -- young growing leaves were
collected -- tested through ELISA to confirm the presence or
absence of SCMV.
The lines thus tested were grouped as resistant -- moderately
resistant -- moderately susceptible -- susceptible on the basis of
disease rating scale of Piper et al., (1996).
 SCALE USED FOR RATING SEVERITY OF SCMV ON SEEDLING

Rating Description of symptoms

0 Healthy, no virus visible.

1 Very mild symptoms on one or more leaves.

2 Mild symptoms on one or more leaves.

3 Moderate symptoms on one or more leaves.

4 Moderate symptoms (as in 2 or 3) but more widespread than 3.

5 Severe symptoms, widespread on plant. Especially, to all leaves on a tiller showing

symptoms.

6 Severe symptoms, as in 5, but in addition either noticeable stunting or small to

moderate amount of necrosis.

7 Very severe symptoms, severe stunting, obvious and significant amount of necrosis.

Where,
0 = - = Resistant (R)
1-2 = + = Moderately Resistant (MR)
3-4 = ++ = Moderately Susceptible (MS)
5-7 = +++ = Susceptible (S)
RESULTS AND DISCUSSION
Disease Incidence (percent) of SCMV in different districts of Punjab on
the basis of symptom observation

S. No. Name of District Mean D.I. (%) of two locations

of each district
1. Mandi Bahauddin 41.89

2. Sargodha 70.46

3. Toba Tek Singh 51.12

4. Jhang 32.74

5. Faisalabad 15.95

6. Okara 75.53

7. Bahawalpur 35.87

8. Rahim Yar Khan 45.75

9. Muzaffargarh 34.34

10. Layyah 33.81

Disease Incidence (percent) of SCMV in different districts of NWFP on the
basis of symptom observation

S. No. Name of District D.I. (percent) Mean of two locations

of each district

1. Peshawar 43.16

2. Mardan 53.03

3. Dargai 52.71

4. Charsadda 52.13
Relative occurrence (%) of random and non-randomly collected samples from Punjab

% R.O. (Random) % R.O. (Non Random)

70

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Relative occurrence (%) of random and non-randomly collected samples from NWFP

% R.O. (Random) % R.O. (Non Random)

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 Response of different cultivars to SCMV in different districts of Punjab
S. No. Name of District Locations Name of Variety Location vise D. I. (%)

1. Mandi Bahauddin Field 1 HSF-240 14.11

Field 2 Kala India 69.68
2. Sargodha Field 1 Unknown 90.26
Field 2 CPF-238 50.67
3. Toba Tek Singh Field 1 CPF-237 21.50
Field 2 China 80.75
4. Jhang Field 1 SPSG-79 35.94
Field 2 NSG-555 29.55

5. Faisalabad Field 1 HSF-242 19.01

Field 2 HSF-240 12.90
6. Okara Field 1 CP-90 71.93
Field 2 Mixture 79.14
7. Bahawalpur Field 1 CPF-234 35.95
Field 2 CPF-234 35.79
8. Rahim Yar Khan Field 1 CPF-234 44.64
Field 2 CPF-234 46.86
9. Muzaffargarh Field 1 HSF-242 29.53
Field 2 Desi + Mixture 39.15
10. Layyah Field 1 CPF-238 46.22
Field 2 CPF-234 21.40
 Response of different cultivars to SCMV in different districts of NWFP

S. No. Name of District Locations Name of Variety Location vise D. I. (%)

1. Peshawar Field 1 CP-77/400 41.71

Field 2 Sanober 44.60

2. Mardan Field 1 CP-77/400 62.44

Field 2 CP-77/400 43.62

3. Dargai Field 1 CP-77/400 52.94

Field 2 CP-77/400 52.48

4. Charsadda Field 1 CP-77/400 60.76

Field 2 CP-77/400 43.49

 Different weeds identified as natural hosts of SCMV
S. No. Local Names Scientific Names of Weeds OD405 Value Reaction against
SCMV
1. Baroo Sorghum halepense Pers. 0.439 Positive
2. Bathoo Chenopodium album L. 0.129 Negative
3. Bhakra Tribulus terristris L. 0.119 Negative
4. Bhoin Cyperus iria L. 0.121 Negative
5. Damb Grass Polypogon monspeliensis Desf. 0.395 Positive
6. Deela Scirpus maritimum L. 0.207 Negative
7. Ghoin Cyprus difformis L. 0.112 Negative
8. It-sit Trianthema portulacastrum L. 0.126 Negative
9. Kara Bara Digitaria adscendens L. 0.108 Negative
10. Khabbal Cynodon dactylon (L.) Pres. 0.328 Positive
11. Lelhi Convolvulus arvensis L. 0.119 Negative
12. Loomar Grass Setaria pumila (Poir.) Roem & Schult. 0.304 Positive
13. Lumb Grass Leptocloa panacea (Retz.) Ohwi. 0.459 Positive

14. Madhana Dactyloctenium aegyptium (L.) Willd. 0.349 Positive

15. Moorak Cyperus rotundus L. 0.127 Negative

16. Naroo Paspalum distichum L. 0.411 Positive
17. Swanki Echinochloa crusgalli (L.) Beauv 0.512 Positive
 Reaction of different host species on inoculation with SCMV

Host plant Cultivars Symptom Observations DAS-ELISA

(SCMV)
Wheat Chakwal-97 No visible symptoms -

NARC No. 1542 No visible symptoms -

Barani-73 No visible symptoms -
Sorghum Js-62 No visible symptoms -
Millet MB-87 Light yellowing & mosaic +

Super-1 Light mosaic +

Acc. No. 8808 Light yellowing & mosaic +

Oats PD2LV65 No visible symptoms +

Maize EU-1097 Mosaic +
Rakaposhi Mosaic (7-8 leaf stage) +
Islamabad Gold Mosaic +
Islamabad white Mosaic +
Sweet Corn Mosaic and light yellowing +

Barley (Soorab-96) No visible symptoms -

 Resistance potential of commercial sugarcane varieties against SCMV

Reaction against SCMV Name of Varieties

Resistant (22) HSF-240, BF-129, CP-48-103, CPM-13, CO-1321,

SPF-213, M-93, CP-43-33, M-92, CPF-237, NIA-98,
SPF-234, Thatta-10, BF-162, COL-54, LRK-2001,
BL-4, HSF-242, L-113, CP-51-21, L-118 and CP-44-
107

Moderately Resistant (04) COJ-84, SPSG-26, PR- 1000 and L-116

Moderately Susceptible (06) CP-72-2086, Triton, CP-77-400, IM-61, L-357 and

L-62-96

0 = - = Resistant (R)
1-2 = + = Moderately Resistant (MR)
3-4 = ++ = Moderately Susceptible (MS)
5-7 = +++ = Susceptible (S)
 Resistance potential of sugarcane germplasm lines against SCMV

Reaction against SCMV Name of Germplasm Lines

Resistant (38) TCP-88-3480, CO-436, THATTA-8, PR-66-1240, SPHS-8, PR-68-123,

HSTH-18, CP-88-1165, CPTH-1, HSTH-10, HSTH-9, TCP-85-1432, TCP-
69-1059, COJ-76, COJ-64, CO-312, CPTH-19, CP-81-1238, RB-72-454,
CP-88-1540, L-89-152, CPHS-3, SPHS-2, HS-12, RB-82-5336, CPTH-20,
CP-50-28, CP-88-1573, CP-70-321, HOSG-449, S-00-SPSG-1607, S-01-
HOSG-795, S-02-HOSG-155, HOCP-92-631, N-60, Malakand-7, SPF-213
and CP-84-1198

Moderately Resistant (10) RB-78-5148, GT-1, NCO-310, L-89-113, TCP-81-3067, S-98-CSSG-668,

S-03-CPSG-433, S-03-HOSG-1275, S-03-HOSG-945 and S-97-US-183

Moderately Susceptible (15)
Susceptible (13)
CPF-150, AEC-81-89, CP-76-331, CPF-222, GT-11, CO-285, TCP-83-
3211, COJ-79, CP-90-951, COJ-78, TCP-83-3210, HOSG-104, HS-4, S-
98-SP-729 and CP-85-1382
CP-73-1030, CPTH-16, CPTH-3, TCP-86-3368, TCP-81-10, CPM-76-611,
MEX-57-473, SPSG-93, HOCP-91-552, HOCP-91-559, CP-89-2376, CP-
85-1491 and CP-89-1945

0 = - = Resistant (R)
1-2 = + = Moderately Resistant (MR)
3-4 = ++ = Moderately Susceptible (MS)
5-7 = +++ = Susceptible (S)
CONCLUSION
• The sugarcane fields of major cane growing districts of Punjab and NWFP
were found overwhelmingly infected with SCMV.

• SCMV was prevalent in all sugarcane-growing districts of Punjab and

NWFP with the maximum disease incidence in Okara and Mardan
respectively.

• The relative occurrence of SCMV was found highest in Muzaffargarh district

of Punjab, while it was highest Peshawar district of NWFP.

• Eight weed species were recognized as natural hosts of SCMV and were
among the dominant weed flora of both provinces.

• Common maize and millet cultivars may also harbor SCMV in natural
conditions.

• Disease may spread rapidly when vector population, particularly of Corn

leaf aphid (Rhopalosiphum maidis) increases in the maize fields
surrounding the fields of sugarcane, a commonly observed situation in
Punjab and NWFP.

• Out of 32 commercial sugarcane cultivars evaluated for their resistance

against SCMV, four were found moderately resistant and six were found
moderately susceptible to SCMV.
• Cultivation of improved commercial varieties with known potential against
SCMV is therefore recommended so that, unaddressed losses due to this
disease may be minimized.

• Ten sugarcane advance lines were found moderately resistant, fifteen lines
were found moderately susceptible and only thirteen lines were found
susceptible to SCMV.

• Preferred parameters are quantitative i.e. cane weight, yield, germination,

vigor & sugar contents, rather than qualitative i.e. resistance against pests or
diseases, drought tolerance, frost tolerance etc.

• This results in failure of varieties due to low level of resistance against

diseases especially of viruses. Therefore the lines identified as completely or
at least partially resistant to SCMV must be included in yield trials as so that
resistant varieties with good yield potential may be put forward to the farmers

• Week crop stand due to unavailability of required inputs well on time, poor
phytosanitary measures and meager check on insect pests worsen the
situation collectively favor the virus to establish and become inhabitant in
potential areas as poor crop stand, higher weed infestation and large vector
population play a role that is more favorable for SCMV to become so
widespread in nature.
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