9. SEED HEALTH TESTS: SEEDBORNEDISEASES

v.K. AGAR Hi\L & AS!IOK GAUR

Many high yielding varieties have shown susceptibility to dilTerentdieseases

and many of these diseases arc seedborne. The seed primordium or the maturing

seed may be infected either (i) directly from the infected plant through the flower

or fruit stalk and the seed stalk or direcUy from the seed surface, or (ii) infection

from oulside may be inlroduced through stigma or ovary wall or pericarp, and the

Dower or fruit stalk, and later through the seed coat. A pathogen may penetrate

several ofthese parts of the seed and in turn infect them. 'l"he infcslation/contam ina-

lion of the seed may occur during harvesting, threshing and processing. The

pathogen may, thus, be carried with the seeds in three ways.

(i) Admixture: Pathogens are independent of seeds but accom-\pany them. Ergot sclerotia are mixed with healthy seeds

during threshing.

(ii) External: The pathogen may be present on seed surface as

spores, oospores and chlamydospores as in case of kamalbunl of wheat, covered smut of barley, downy mildew ofpearimillet etc.

(iii) Internal: Pathogens establish within the seed with definite

relationship with seed parts.

The seedborne pathogens may result in (i) loss in germination (ii) discoloura-

lionand shrivelling (iii) development ofpJant diseases (iv) distribution of pathogen

to new areas (v) introduction of new strains or physiologic races of the pathogen

along with new germplasm from other countries (vi) toxin production in infected

seedetc. Visualising the importance of seedbome diseases the Central SeedCertification Board has prescribed certification standards for foundation and

certified seed for several diseases Crable 9.1).

The methods as approve by the Central Seed Certification Board as wen as

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suggestive methods for additional diseaseshave been described. The number of

seedsto nl' testedisnasedon the presenceof allem;1one infectedseedperreplication

~ISper ceni fication standards for foundation seedand eenified seedCrable9.2).

I. Kumal Bunt of Wheat and Triticule. Causal Oq,:anism : Nc())'()ssiaindica.

I'r,,('~dllr~

(vii)

. (viii)

: Sodium hydroxide (NaOI!) seed soak method.

(i) Wheat seeds arc soaked in a flask/beaker c(mtaining 500 ml of

0.2 per cent sodium hydroxide solution (2 g NaOHlJ (X)O mlwater for 24 hr at2(1-.~()0c.

(ii) Aller 24 hr the solution is decanted.

(iii) Seeds arc thoroughly washed in tap water.

(iv) Seeds are spread over a blotter paper so that excess water on

the surface of seed is ansorbcd.

(v) Seedsarc examined visua}ly aided with light.

(vi) The seeds exhibiting jet black shiny appearance with hoHow

or without hollownes..'-;are separated.

Such col1ccted seeds .Ire ruptured separately in a drop of water

and observed visually for the release of stream of fungal

spores.

The number of seeds releasing stream of fungal spores arecounted as infected seed.

(ix) Result is reported in percentage.

The Kamal bunt infection can easily be differentiated from black point which.

is mainlycaused byAlternariaalternaw. '1l1esceds'alTected by black pointorblack

tip exhibit dark brown to blackish discolouration which is moslly restricted to

embryo tip. In this case internal tissue of seed is not converted into black powdery

mass. 111cseseeds on rupturing do not relC<lsestrc.lm of fungal spores.

111eabove procedure is applialble to both chemiailly treated as well as

122

untrc:llcdseed lots.

II. Hunt of Paddy. Causal Or~anism : Nem'ossia horrida

"rtJl"edllrc

(vi)

(vii)

(viii)

: Sodium hydroxide (NaUII) seed sank method

(i) Paddy seeds ,Ire soaked in a Ibskibeaker cont,linlng 2:'iO1111of

0,2 percent sodium hydroxIde solulion (2g NaOI1,']()OOml

waler) for 24 hr al 20-.10°('.

(ii) After 24 t1rthe solulion is dcc,mled.

(i ii) Seeds arc thoroughly washed in tap W,IIt.T.

(iv) Seeds arc spread over a hlotter paper so that excess water onthe surface of seed is absorbed,

(v) Seed arc examined visually aided with light.

The seeds exhibiting shiny jet hlack arc separated,

Such shiny jet black seeds arc ruptured separately in a drop of

water hy puncturing and ohserved vbually for the release of

stream 01"fungal spores.

The number of seeds releasing stream of fungal s{X)resarccountl'd as infected seed,

(ix) Resu 11is reported in percentage,

The secds looking only shll1Y Jet hlack have been found to contain buntinfection whereas those with hrown to dull black discolouration do not reveal bunt

infcctum, This is a symptom of hrown spot or brown discolouration of rice.

III. Ergot of I)earlmillet. Cansal organism : Cla~'icepsJusiformis, (=Clm'iceps

microceplzela)

Ergot or sorghum. G1USaIorganism: Sphacelia sol'ghi and C laviccps spp.. and

ergot of triticale. causa I organism: Claviccps pllrpllrea,

123

I'ron-dur<' : Visual ooserv<ttion of dry seeds.

(i) Seeds arc exam ined visually aided with light in dry stat~ for

the presem:e or ergot sc1erot ia. 'Illese sclerotia me purrlish

orown in colour, irregular in shapc, Ilard structurcs and these

may oe two to three times tllgger than healthy seed".

A seed lot o/'sorghum mixed with well developed and broken

sclero! ia is called an ergotted seed jot. Ilowever, a seed lot

mixed with honey dew lumps containing: mycelia] mat with

conidia should no! he called an ergottcd seed lot.

(i i) The sclerotia ;Ire separated and counted.

(iii) Result is reported in percentage (hv number).

IV. Loose Smut of "'heat. Causal organism: Cstilagu trith-';

Pron-dure : Emoryo count method.

l. Soak 20()() seeds in :'i.!)Oper cent sodium hydroxide and 0.02 per cent

trypan olue solution (one liter) for 24 hr at 25<;(f( '.

.., Pass the soaked m.Herialthroligh 10 m(-.h sieve and r~tain the material in

a 20 meshsieve along with showers or tap water.

3. Collect the extracted emoryos in a heaker.

4. Dehydrat(:the embryos in rectified spirit ror 5-] 0 min.

5. 'rake the dehydrated embryos along with charI' etc. in ;ioeaker containing

50 mllactophenol.

6. Add to above beaker 100011 \-vaterand stir it.

7. AJ\ow the materia] to stand for 5 01in to settle the chaff at bottom.

8. Collect the 110ating embryos in another beaker containing 25 011fresh

lactophenol.

124

9. !3oilllll' anon: m:lteriaJ for 2 min.

10. Pour tt1\.'embryos in pctridislle.., and :Irrange in lines along with some

lacwphenol.

11. Observe the embryos under stereobinocular microscope for the presenceofmvcL'lium.

12. Mycelium appe,ir.., ,I'"blue thread like knoll cd structure in the scutdlum

portion of the embryo.

13. Total number of emoryos and infected embryos <Irecounted.

14. Result is reported in percentage.

It is a suggestive procedure for the detection of loose smut of wheat. 1\t themoment there arc no certincat ion standards for I(xlse smut of wheat based on ..,eed

infection.

)otato

Proc~dllr. ; Visual observation of tubers.

Observations:Count infccted tuners based nn S\'lllplnllls described below.

\. I.ale bl ight of Potato: !)In'/rJpluliol'll ifl/c',\{lIm

The tubers exhibit brown to purplish discoloration of the skin, followed ~y a

brownishdry rot which exlcl1lls to about Ie. inch below the surface. Sections cut

through tubers show brown and necrolic tissues,

2. Dry rot of Potato: Fusarium ('(Icru!cum

The surface of infected tubtT is wrinkled and may be sunken, and the rotted

tissues may turn brown, grey or black, Cavities frequently develop in affected

tubers,which may become more or less filled with yellow, pink or red mycelium,

After prolonged storage, a black, blue or violet fungal gro~th develops on the

surface,The margin of the rot is clearly defined. Typical concentric rings appC<lr

onthe tuber surl~K'Cand external mycelium is evident. The tuber dries and hardens.

125

3. Charcoal rot of Potato: il4acrophomina plwseolina

On tubers. lesions appear as somewhat softened and nlackened area..;.The

decayc<.ltissueis dark coloured and usuallyconfined to the outer zone of the tuher.

4. Wet rot of Potato : Sclerotium rolf,,'ii

Lesions develop on the surface of the tuber. 'I 'hey are yellow to tan in colour

and noticeably sunken. I,ater. these lesions become darker in colour. The tubers

decay and the entire tuherhecomes slimy. Sclerotia about the sii'c of mustard seeds

appear in the decayedl issue and in the c,lvities. 'Illey arc white at first and nceome

dark brown with age.

s. (~ommon SG,lh or PoUlin : .<';[,.cpWI11Ycc,~scahies

Two types of symptoms calkd slwllow and deeper pilted seth m~IYaprl~<lr()n

tubers. Shnllowscub consists of u supcrfici::II roughened ;/fe;l. .,ome/imc!> t;uiscd

above. and often slightly hclow (he pl::meof the he<llthyskin. 111elesions consisL~

of corky tissues. The lesions vary in size and shape. sometimes darker than the

healthy skin. Deep pilled scab consists of ]esiolls which are I to more than.1 mm

deep. Ihey may be darker than the shallow lesions. The tissue around the interior

of the pits is corky <l';in shallow scab. Thc..,e lesions may join together so that entiretuber surface is affected.

6. BI::lckscruf or Pot::I!O: Rhizoc/Onia solani

The tuber infection <lppcars hard. superficial. black crust on the skin duc to

formationof sclerotia of the fungus. Tuocrs are often rough. Thesclerotia arcmore

conspicuous when tubers arc wet.

Multiplier Onion

Procedure : Visual observation of onion bulblets.

Obsen'ations : Count infected onion bulblets based on symptoms described below.

1. Basal rot of Onion (Bulblets) :Fusariwll oxysporllm f. cepae

On the base of the bulo whitish moldy growth of the fungus appear on thesurface ofthe decayed portions of the salle. Onsectioning in thebulbs longitudinal-

126

Iy," semiwaterydecay is found to he advancing from the ba<;esof the s(.lIes upward.

Infections occuring about harvest time may continue in transit and storage unt i I

only dry shrivelkd 'mummies' remain.

2. Soft rot of (>nion (Bulhlets) : l~m'inia carotm'orll

One or two outer scales of the onion rots completely. The diseased bulbs can

he detected by gently pressing them, whereupon the watery f1uid is extruded

through the neck. The slimy decay is usually accompanied by a f()ul sulfurous odor.

The scales outside the rooting ones slip off readily in handling.

Sweet Potato

l'nlC~dllr~ : Viswll observation of roots.

Observation.. : Count infected roots based on symptoms described below.

1. Black rot of Sweet Potato: Ceratos(()mclla Ji11lbriata

Dark circular depressed spots of varying size appear on the fleshy roots. The

spots arc grey-hlack when dry and dark greenish-black when moist. In the center

of these spots or in the older portion of the lesions small black fruiting bodies of

the fungus also develop, A shallow. dry decay extends below the surface lesion.

usually not beyond the vascular ring.

2. Scurf of Sweet Potato: Monilochaetes inJilscans

Fleshy roots arc discoloured. "l11elesions may be in the form of spots or may

coveraconsiderable area. The roots shrink during storage due to loss of water.

3. Wilt of Sweet Potato: Fusarium oxysportlm f. batatas

The internal tissue of the roots become disco loured. On the surface of roots

shallow. sunken spots appear which finally shrink.

4. Internal cork of Sweet Potato: A virus disease.

Dark brown to blackish. hard corky spots which vary in size upto 3 cms. across

and 5 cm long appear in the fleshy portion of the root. These corky areas increase

in size and number. the longer the root are held in storage.

127

Table 9.1 Seed standards for foundation and certified seeds based on maximum percent seed infection(Tunwar and Singh, 1988)

Crop Species Disease Causal organism

2 ~

I\J(X)

Pearl millet

Paddy

Sorghum

Wheat

Triticale

Potato

E' b~rgot

Bun!

1, h,rgot

Ear cockle

Kamal bunt

Clal'iceps fusiformis

Neomssia IlOrrida

Spllacelia sorghi

A.nguina tritic'i

Neol'ossia indica

Tundu Coryne/JGcterium michiganl'llesc

pv. trifid andAnguillatritic'i.Neol'Ossia illdicaKamal bunt

Ergot

Late blight

Dry rotCharcoal rot

PIlYlOphthora infes/(/Ils

Fusarium caeru/eum

MacropllOmilla phas('olina

Sclerotium rolj~'iiWet rot

Common scab C Streptomyces scabies

.,.

Certification standards 'i~ (by number)

Foundation seed Certified seed

4 5

0.02

0.0]

0.02

None

O.OS'7c

None

O.OS7<

0.02

\.0

\.0

1.0

None

.-:;.0

(J.O4

0.50

0.04

None

,0.25%

None

0.257<

0,04

1.0

1.0

1.0

None

5.0

2 3 45

Multiplier Onion

Black scllrfd

Total Disease~,c

Basal rot None

None

Rhiwctollia so/alii 5.0

5.0

Fusarium uxY.l'porul11 f. cepac

!:'rll'illia carO!IJ\'uraSoft rot

Brown rot !)seUdIJI11O1I11.1'aerugil/{)s(/ Nonc

Nom:Sweet Potato Storage rot

Black rot

Scurf'

Wilt

Internal cork 5.0 5.0

CcratO.l'lOl11e//afi mhriata

Molli/ocl1aetei: il~rU.l'('alls

Fusarium oxysporul11f.hatatas

None

Non..:

None

I\)CD

hErgot sclerotia seed entirely or partially modified as sclerotia, broken sclerotia or ergolled seed.

5.0

5.0

None

None

None

None

None

None

None

"Even if ~ single tuber infected with common scab is detected on a seed lot, entire seed Jotshall be treated with 0.5% solution of

Agallol-3 or Emisan-6 or Aretan-6 for 30 minutes, before seed lot is declared fit for certification. SeedJotshaving infected tubers

more than the prescribed limitswill not be certified even after treatment.

d(i) A tuber carrying 1O.()%or above scurfed surface wiJJbe consideredas one infected unit.

(ii) Seed lots having black scurf infection more than the prescribed limits couJd be certified after treatmenL<;with approved

chemical/fungicide.

eFor all diseases, the higher di~ease percentage will be considered for the purpose of the specified limiLs of tolerance.

fA root carrying IO.()%or above scurfed surface would be considered as one infected unit.

Table 9.2 Number of seeds to be analysed for.different seed certification standards*.

Percent Seed Infection No. of seeds to be anal ysed/

replication

1. None 5000

50002. 0.02

0.04 2500

lOOO

3.

4. (LOS

0.1 1000

400

5.

6. 0.25

7.

8.

0.5

1.0

200

100

9.

10.

3.0

5.0

:n

20

*The number of seeds to be tested is hased on the presence of atl~lst one infected

seed in the working sample. Each ~Imp'le will have two replicmions. The size of

the working sample has been derived from 4 x n. where n =standard specified.

130