Jéliro Castaño Z: ... - .. _-~---
Rice (!l~:;e~d_ ~;"ª_tJ~ L_) is thc most widely grOi-m cereal crap of the VI01'ld,
the avprag<? product'í on accounti 119 to about 2',0 mi 11 i ons tons ilnnUi) l1y
(98, 99). Aboul half of the production is grown in the tropics (66 )
*
populatioll (104), Since rice is net ~ropa9atcd vegetativ~ly the pe¡'pctuation
of the trop depends on the sexua' seed production. lhe importance of rice
,in world foad production and the significance of seed transmission of
several important diseases of this ho~t is, tl1erefore, of concern of nnn
in his unending quest for more and better food. Heavy losses of rice are
due to I'arasitic diseases of which the principal are bli1st (Pyricul_ilLi_!!.
!lryzé\~), bro\'ln leaf spot ol' seedling blight (Cochliobolus miyabean~~;
conidial stage Hell11inthosporiul11 oryzae), leaf scald (metasphceri!!. !!.lE!,sc~r~,,-;
conidial stage Rhynchosporium Qryzae), bacterial leaf blight (Xanthong..nas_
oryzae) and !'ice leaf or white tip nematode (Aphelenchoides be~yiJ. All
are seed borne. Fungi cause the largest nUl11ber of plant diseases and
these mi crool'gan i sms occurs more common ly in seeds than do bacteri a or
nematodes. So far,no virus disease is knoVl to be seed-borne. The
Il\Ycofl ora Il\ay cons i st of saprophyti c fungi aod/or pathogeni e fun9i. The
saprophytes are non-host specific species and may be found on seeds of
various plants, whereas each pathogenic fungal species in generally confined
to a limited host-range. 80th I'lay be borne superficiill1y attached to the
* Postdoctoral Fellow, Rice 1983 . t ~ . " (\
f
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outcr secd surface in cracks, 01' insidC' the seed eo,lt, but the putl10gens
may illso be prescnt witllin the cotyledons and other tissucs of the
embryo (73). Considcring just rice and wheat, tha ccreals uf whicll most
uf the hilrvest qoes far human foad, the lasses are obout 30 mil1ian metric
tons. This rcpresents enough food for keep 150 millian people alive for
ayear, The most important part of this picture, howcver, is that these
losscs ilre more severe in thosc countries which can lcast afford them;
the poorer and undcr developed or emerging countries of the world.
Although the losses are 5% on a Vlorld Vlide basis in India, and in parts )
of Africa and South America tile losses may reach 30% of the harvested
crop. In a11 of these losses fungi playa major and irl1portant role,
although this was 110t fully realized until recently.
The damilge caused by fungi va ri es from spots i nvi si b 1 e to the naked eye
to complete rot of the endosperm. Further damage may occur during
storage. Generally, the same fungi cause heavy spotting or discoloration
of the hulls. Kernel spotting increases in damp or warm, rainy weather
and in rice that matures late in the season. The presenee of spotted
or sta i ned kernel s reduces the grade of .ri ce. Kernel s tila t are s potted
severely and therefore chalky break into pieees in the milling proeess;
thus; kernel spot reduces yield of head rice (5).
Today that agriculture has reached a very high level of technification
it is imperative to obtain a good qualite of seed. Seeds are both •
vietims and cfficient vehieles of diseases. Seed tral1smission of
pa thogcns i s no\.¡ recogni zed as the method for excc 11 enee by wh i eh p 1 ant
pathogens (a) are introdueed il1to new areas, (b) survive periads when
f
1
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thc Ilo~t is laekillg, (e) are selceted ilnd disseminated as host speeHie
strail15, and (ct) are distributcd through the plant population as foei of
infection (8). Sccds that harbor pathogcnic fungi are important to
a(Jriculturc beeausc: (al infeeted seeds may not gernrinate and the resulting
decrcascd sccdling stand may lead to a reduetion in yield; (b) infeeled
sccds Ciln provide inoculum that, under suitable conditions may initiate
an cpiphytotic that can lower yields; (e) as it was mentioned above,
infected sceds can introduce pathogens into areas that are free of the
patllogens; (d) infected seeds, even though they are treated with a chemical
may stil1 harbor ellough viable pathogens to resuH in any of the aboye
situations and, (e) infection of seed by pathogenic lIlieroorganisrlS before
harvest may cause a reduction in seed quality and yield.
AIl idea oY tile importan ce of microorganisms on seeds can be aehieved by
lookill9 at the number of genera which have been reported frolll rice seeds.
Padwiek (105) lIlentiOlled about 17 genera which inhabit seeds; the United
Sta tes Department of Agriculture (152) listed about 24 genera; Riehardson
(117, 118) prepared a list of sorne 40 genera comprising the most important
disease agents af this cereal. More recently, 1 made an extensive
literature review of seed borne pathogens of rice and 1 accounted more
than 50 genera of mieroarganisms that inhabit rice seeds (Table 1). From
this table it can be observed that more than 100 species of fungi an a
redueed nUlIlber of bacteria and nematodes are assoeiated with tile rice
grains.
The fungi may be divided into two major groups: one group consists of
-.
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TACLA 1. LISTA PARCII\L DE t1IC1UlORGI\r¡IS~10S ASOCIADOS CON LA SG11 LL~ DE I\RROZ
---_._-------....,--~--_ ... _- ORGAN 1 S 1·',0 S
Ac1'ocy1 i ndd um oryzilc, syn. Saroel adi um oryzae
A He rrw r i a 1 on gis s i mil A. tenllis, syn. A. alternata Ascochyta aryzae Salansia oryzae, syn. Ephe1is oryzae
Botrytis sp.
eochl iabolus miyabeanus, syns. Kelminthasporium oryzae, D,-cchs 1 el-a aryzae, Ophi obo 1 us miyabeanus, SeptosphHcria rostrata
Cochliobolus lunatus, syn. Curvularia lunata .
C. spicifcr
Chaetomium spp.
Cladosporium cladosporioides
el. herbarum
D. tetramera
Epicoccum neglcctum
E. ol'yzue
F. dimerulll
F. equiscti
F. heterosporulll
F. lateritium
F. longipes
F. nivale
F. oxysporum
F. poae
F. semitectulll
F. solani
Helicoceras nymphaearum
H, oryzae
Hendlersonia oryzae
Macrophoma spp.
Cuadro 1. (Continuación)
ORGi\tllS¡·lQS
~1agnJportlic sal vinii, syns, Lcptosphaeria sa 1 vi lIi i, He 1 mi thospori um sygl1loi dceurn SclerotiullI oryzae
Helanornllla glumarunI
Monascus purpureus
Ophiobolus oryzinus
Oospori1 oryzetorum
Phaeotrichoconis crotalariae
Phoma indianensis
Ph. glolllerata
Ph. jolyana
Ph. sorghina, syn. Ph. glumicola, Phyllosticta glumicola
Pithornyces chartarum
Pleosphaerulina oryzae
REFERENCIAS
10,36,89,101,137
105
114
19,107
Septoria oryzde
Sphaerul ina oryzae, syn. Cercospora oryzae
Stachybotrys spp.
Trematosphaerella oryzae, syn. Phaeosphaería oryzae
Tríchoderma viride
Trichoconis caudata
Trichothecium sp.
Ulocladium sp.
Bacterias
X. kresek
REFERENCIAS
So
10
59
152
23,27,79,103
25,40,135,152,154
114
10,11,135
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harvest. The othcr group contains the storage fun:¡i \/hich usually are
Sa¡lfophyLcs and dcvelops aflcr harvest. Gcnerally, most of the seed
tl't1nsrllittcd j1ilthogens are fungi. Some classes ar genera are frequent in
seed othcrs occur only occasionally)othcrs do occur but cannot be )
revealcd·by conventional testing procedurcs.and the extent to Vlhich fungi }
occur in sceds depr!nd s on thei r capabi 1 i ty to survi ve uncler the extreme
dry condition of secd as a carrier (99). The fungí have the abi1ity to
invade the seed prior tO,during, or fo11o\"ling germination, either killing
th~ yOllng secclling outright, causing root and cro\"ln rots, or becoming
established on the cotyledons and froln there to the above ground-poJ'tions
of tile plant later on. Fung·¡ may be present as mycel ium (e.g. Rhi zoctonJa
sp.) spores (e.g. Helminthospol'ium Q!J'z.ae) or fruiting structures (e.g.
Phol~ 9Ji1marum). In addition, there are a great many soil inhabiting
plant pathogens which are able to attack the young seed or seedling. Sucil
fungi are species of Sclerotium, Pythium, Achly~,Fusarium, Helrninthosporiucl
and Rhizoctonia alllong others.
FIELD FUr'iGI
Field fungi invade the seed either while the plant is still groVling or
after it has been cut befare threshing. According to Christenscn (32),
"it is comlnon to isolate field fungí from close to lOO;i tlf freshly harvested
surface disinfected Kernels of sound rice of the highest grade". In sto red
rice, field fungi die gradually or rapidly, depending on moisture content
and temperature and on the competition of other fungi especially the
storage fungi. One of the major environlllental fa.ctors which determines
'.
•
•
tIJe JI10isture rcquircments of the organismo All plant products are
hY9l'oscopic. All hygl'oscopic matcrials will tend to come to a steady
moisture contcnt condition in equilibrium with the surrounding atl~10sphere;
eithcr gaining 01' 10Sin9 water to do so. This steady moisture content i5
called the equilibrium moist0re content (EMe), which depends on both
relative humidíty (RIl) and temperature. Any changes in either or both
of these factors will change the EMe of the material. lIJe relationship
betwcen the EMe and Rfl Is direct being inverse with respect to temperature.
for the starchy cereal s such as rice,the field fungi require betvleen 20-25%
of moisture contento These moisture contents correspond to the EI1C of
cereal s at about 90% RH. Once the moisture content drops to about 14%
the organism die rapidly. Although it was stated that these.organisms
usua11y do not cause storage losses, they can occasionally cause such
losses. If the seeds or grain is harvested at high molsture contents
(above 20-25%) and storage at hlgh RIJ, these fungi can continue to groN
and cause damage. The main reason that storage losses due to field fungi
seldom occurs Is due to the utilization of seed certification programs,
which require seeds to be dried to moisture content closer to those in
equilibrium al relative humidity between ?O-90%.Christensen and Lopez
(34) observed that the percentage of surface disinfected grains of
Bluebonnet 50 yielding field fungi decreases with increasing moisture
content and with 10ng¿r time and the fjeld fungi die in close to 100% . )
of the Kernels before germination percentage began to decrease. Death of
field fungi in barley and l~heat seeds stored at appl'oximatcly the same
•
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Tlle dCllla(JC ciJused by ficld fungi varies from srnall brovm color spots to
complete staincd of thc glumcs. Removing those, the disturb may affects
thc endospcrm and even the embryo; thus reducing germination severily
(Figul'e J). TiJe effect of field fungi in rice secd gcnnination i5 evident
(4,IQ19,22,24,94,131,136,156).
The field fungi muy live for years in grain sto)'ed at 10'11 mQisture content
(30), their survival 1ik8 that of the embryos of the seeds themselves,
being favored by low moisture content and 10\'1 tempel'atul'e. Under the
usual cOl1dition of storage of food and feed grains, for instance, Fusal'ium
may die within a 'few months; Helminthosporiurn may not survive much longe!';
and I\lternaria may not be recoverable from more than a sl~all percentage
of secd storage for él year (41,71,148).
STOf1AGE FUNGI
Pioneering work of the role of fungi in- the deterioration of storage
grains was not begun until the 1930's. It continued through the 1940's,
and full realization of their importance was not achieved until 1950'5.
Even today, many of the grain merchandasing business do not realizo that
fungi playa decisivo role in their operations, profits and losses. The
storage fungi are genera lly saprophytes or facultati ve parasites. These
fungi do 110t invade grai ns to any appreci ab 1 e degree or extent befo)'e
harvest (149,150). This is now well establ ished from tests with m3ny
different lots of cereal graios, over a long periad of time,and with
samples from many different sources, Storage fungi invade ilnd grow in
;
.. ..
•
Thesc> ¡Ir'l: rnoisture contents in cquil'ibrium with rclat'ivc humidities of
about 70-90~. In thoso environmrnls storagc fungi are among the most
abundant alld succcssful of a11 living things. A1most incvidently thcy
contarni na te· a 11 secds. The organi srn5 compri si ng thi 5 group are about
t\'lclve spccies of I\sl)['.r:D_ill.LJs,scveral species of PenicilJltJm anel less
frcqucnlly a few oth"r genera of fungi such as Rhi.zopus _' l~u..c.2!: and
~bsid~ (33,114,124,143). Species of Penicillium are encountered at
times, u5ua11y in lots of grains stared at low ternperatures and with
moistun~ content aboye 16%. Each of the spccies of Aspc!:.gillus has its
o\,m rathel' sharply defined lovler limit of moisture cantent (or relative
humidity) below which it \'Ii11 110t grow, and competition with associated
species may estab1ish a less sharp1y defined upper limito Thus,
determination of tlle number and kinds of fungi in a given lot of seed
at times indieates the moisture eontent and temperature at \-Ihieh the
grain has been stored, and sometimes even the approximate 1en9ht of time
it has beell stored (33).Although severa1 factors are involved in the
se1ection and succession of microorganisms, the most important by far
i5 the moisture eontent of the seed. Temperature plays a role in this
se1ection and suecession but, sinee temperature requeriments of thcsc
or9ani5ms are not nearly as restl'Ícted in range as are their moisture
requirclllcnts, the cffect of telllperature is lIsua11y on1y important at the
very 10\'1 tcmperatul'es (Near freezing) or the very high ternperatures
(lO-7S"C). Huch rice is harvested \'Iith a moisture content aboye 16%
(123) and, like other seeds, is subjected to postharvest invasion by
storagc fungi. Thi5 invasion m~y result in decreased gcrminability
(34,35,49), discoloration of thc grains (123) even in th~ absencc of
;
•
. e
- 14·-
food illld, p¡'oduction of toxins that constitutcs a health haZill'd for
anilnals ilnd hUl\1ill1S Uil). I\ccording to Chl'istensen and Lopcz (34) at
moisture contctlts aboye about 1t,.5;; invasion of rough rice by storage
fungí and decrcase in germinalion percentage of the seedi wcre proportional
to increasiQ9 moisturc contcnt a"d the increasing lcl1gth of storage. In
rice, tile nlost obviolls economic 1055 associated I~ith fungal deterioratioll
during storJge is 1055 of grade in white milled rice (121,122). Rice
may be dovm~jI'adcd for: (a) damaged Kernels, (b) heat-damaged Kerne1s,
(e) change in general color, and (d) musty or moldy odor .. All the othe¡' , faetors are or may be caused by the activities of fungi d~ring storage
(124). Losses in storage are a direct result of the activitics of various
mucro and mic¡'oorganisms made possible by manis failure to understand and
fo110lv sound storage principles and practices.
Storage fungi are post harvest invaders. ~lany cammon species can inyade
and g¡'OW in seed with little or not free water. Christensen (32) found
that many of the common species invaue stored grain with ITIoisture contents
between 13-18% in equilibrium with re1ative humidities of 70-85%. However,
sorne species can invade and compete Ivith field fungi, growing ¡'apidly and
causing 10ss in quality in seed with ITIoisture contents aboye 18%. Since
rice is usually harvested vlith a moisture content between 18-22%, drying .
or conditíoning until the moisture ís be10w 18% must be done promptly .
Propagative units of coanon species of storage fungi are found on the
surface of almost every Kernel of rice after harvest (124). I\lthough
the fungal flora of rice has been studied in the Unitcd States and abroad,
a complete listing of all fungí that ha ve bcen observed associated with
storcd rice has not been assembled.
:
,
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- 15 ..
Til(! s torage fUII9 i are genera lly saprophytes al' faculta t i ve paras i tes and
are main1y spccies of the Asp~i.E.t~ and Pc>nic:2.l.:U.um groups (124). The
genl1S ~l.E-ºEiL1.Ll~2. comprise s species of~ .. yersicolJ!.l.:.,!l. ~dm'ii" !l. repens,
1\ .11.<11'(\ s it.is.~.' !l. rc_s}r.Lc tus, ~.r.!1~."'-r:, ~. .anls te 1 od o.I11.L ~. ca n5.!.i dll:~,
1\ • .f..1avus" [l.. !!.¡~lopb.ibll..'.'.., [l.. niDer, .f2. glaucus, [l.. ochraceu~ and .!!,.cheval.1.eri,
(14,8.l,12~,I~G). !l.glaucl1s and !l. flavu~ groups, however, appear to
constitute the major funga1 species that invade stored rice (124).
A1though insects direct1y damage grain through penetration and consumption
of Kerne1s, they a1so influence the activities of fungi (29). Sel'ving as
carriers and vectol'S of propagative units they a1so increase avai1ab1e
moisture and ri se the temperature, thus fostering and stimu1ating increased .
fungal activity.Aspergi11us f1avus, ~. repens and !l. restrictus, for
i ns tance, ha ve be en found associ a ted vlith the i ncreased mol s ture content
in rice containing \.¡eevils (Sitophilus oryza) (62).
Species al' Penicillium such as ~. J2uberu1um are encounted at times,
usua1ly in lots of seed stored at 10w temperature and with moisture content
above 16% (33,117).
Preventive methods in seed fields.
1. Selection of seed production areas. Seed should be produced in areas
where the pathogen popu1ations is unable to establish or maintain
itself. The physica1 environment-especially 101'1 re1ative humidity­
marked1y restricts incidence of many diseases and prevents infection
of the secd produced.
- 16 -
2. Cultul'il_l_J'2::1stiees. Secd pl¡¡nted in seed fields silould be free of
pathoucns; such scrd can be obtained: (a) frorn areas free of pathogcns
or where it is unable to infect and (b) by secd treatmcnt to erildicatc
tilc patbogcns. The life cycle of a plant pathogenic organism can be
viewcd as con5isting of four basic phases, survival, transmission,
infection, and diseasc developrnent. Methods of control11n9 secd
diseases have th8 objetive of interfiring ,lith the disease cycle in a
way that will reduce subsequent disease development. Seed treatment
affeets tho cycle at the survival and transmission phases by reducing
seed-borne or soil borne inoculurn. Seed h~alth testing can be used
when enough is kno'o'In about the discase cycle to establ'ish tolerance
levels for seed borne pathogens. Sorne of the chernicals listed in
Toble 2 have bren reported to be effeetive in reducing the arnount of
primary inoculum when they are spray at the reproductive stage.
Fungicide application to the growing crop usually affects the infection
and disease developrnent phases. Cultural practices, such as host
resistance, erop rotation, destruction of residues, or alteration of
the harvest date, can affect any of the four phases of the cyele.
Researeh is needed to find better methods of control. Sorne promising
results hav~ been reported. It has been demostrated, for instance,
that increasing spacing between rice plants reduce the incidence of
seed borne infection causes by Trichoconis ~dwickii and; decreasing the
rate of nitrogen fertilization will reduce th~ incidence of both T .
Eadwickii and Tilletio barclayana (3,139,140). Additionally, field
experim~nts have sl10wcd that early heading decreases the severity of
l. barclayiID..'!.. (140)and early or late planting reduce the incidencl? of
Balansia 2.I'yzae (92).
- 17 -
3. ?o"i.~!_~.~lri9_i.l.1..J_~.-,~~ctiQ.l.1... This m~thod of controlis useful in rejecting
seed froro flelds witl, high incidenee of secd transmitted pathogens.
Ho,.¡cvcr, appilfcnt absence of a diseilse may not insure freedom from the
pathogcns in tho seod bccause (a) the pathogen may be- present without
producine¡ sYlllptorns in thc field and (b) pathogen free seed may be
contarninatcd by pathogens during threshing, cleaning, or other handling
operatiol1s.
Preventive and Control Methods for Harvested Seed.
1. Sced Indexing. The detection of seed borne pathogens is best done after
seed has been cleaned and packaged. The best method is to plant the
seed in soi 1, as thi s approximates aetua 1 condi ti ons. The soi 1 used
shauld be: (a) non-treated field soil, to determine the actual field
hazard; (b) tl'eated at 1006C for 30 minutes \vith steam to indicate the
maxilllulII potential transrnission; (e) treated at 60·C for 30 minutes with
aerated steam to give sorne indieatian of the extent to which soil
antagonists will inhibit transmission (9).
Speci a 1 di rect mc;thods ha ve been devi s~d for mi cl'oseopi C exami nat i on of
secd on blotter or agar tests .. These methods are described at the end
of this ftrticle. The methods are subject to great variability in
comparative tests in different labaratories, and must take inta account
spccial requircrnents of different microorganisms.
,
..
- 18 -
2. Srcd trratmcnt~. Much seed should receive a protcction either as oil --_._-----------_. bascd dusts, wcttable powdcr slurrics or as a direct application of
conC(~ntl'ated solution of llctive ingredients.One purpose of seed
trcatment by the use of chernical protectants is to destroy seed surface
borne microorganisJTls that cause secdling blights and seed decay anel to
provido a protcctive zone around the seed, through which soil borne
microorganisms cilnnot penetrates, thus protecting the developing
seedling against attack until it is established on a well devcloped
root system, and has acquin,d the capability of outgrow·ing attacks frorn
·soil borne pathogens. Hence, the principle of seed treatrnent is mainly
control of prilllary inocululll in the infection court, although chemical
seed treatment may also protect 'against infection by organisrns in the
soil. The inorganic and organic rnercurials are exarnples of seed
treating materials used to eradicate certain pathogens from the surface
of seeds. Organic fungicides, including rnercurials, are used as seed
protcctants to coat the seed and provide protection against damping off
and seed-rottihg organisms in the soil.
Control or prevention of seed rnicroorganisrns depends upon both pre and
post harvest rneasures, including those taken during harvesting, threshing
and drying. There exists in the rnarket several chemicals vlhich are
reported to be useful to control the most frequent seed-borne pathogcns
of rice (Table 2). Treated seed, should be stored in a cool dl'y place
and JTloisture content of the seed maintained below the critical level
which will induce germination. A1though seed treated \11th orthocide,
• •
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t • ' " l , ~ l ~ . 1 ~ ~ ) v ~ j J l l t
IÜ H
L .i'j)U
h ~ ' 1 ~ q l r ' I ~ ~ I ' ' ; ' " I ~ ; :
'''''''llr''J 'II.J~ql~
'11, -,'1""'1 'd~
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- 20 -
treated seed should not be stored for long pcriods of time.
Seed born~ fungí. Ilellllit~~o~riul!1. oryzae anel Eyricularia oryzae, tl'lO of ----,._---~-----"'-- -~ -
the 1ll0st devastating pathogens of rice can be control1ed.by a wide range
of chclllicals. fungicides su~h as vitavax-maneb, vitavax-orthoc-ide,
dyrene and ceresan Illily retain thcir effectiveness against both fungi on
rice seeds after G lIlonths of storage (157). Ca re should be taken, hov/eve,',
with organic fungicides such as arasan 75, brestan 60, beam and hinosan
wllich are classified as intermediate toxic compounds, or ceresan which i5
classified as highly toxic cOlllpound (lable 2). Although other chemicals
have 1Il0derate to 101'1 toxicity, cllemically treated seed should NEVER be
used for focd, for livestock 01' food for animals 01' humag consumption even
though it may have be en storcd far months ol' yeal's.
Although a lot is know about seed treatment, we sti 11 do not have good
chemical seed treatlllents for certain plant pathogens. lhis Is particularly
t rue si nce the a 1ky] organi c me I'curi e éompounds ha ve been banned from use.
According to Rangaswdllli and Ramalingam (111) the seed treatment with culture
filtrates of flacillus mycoides increased germination of naturally infected
seed \~i th He 1 mi nthospori U\l1 oryzae, res tri cted the spread of the fungus in
the seedl i ngs, and i mproved the growth of these. Thi s type of bi 01 09i ca 1
control offers an exciting field of research and as long as new seed problcllls
come out this king of control are g01ng to have a rewarded practical
application .
}_0,:_d __ .bQC!~('_Jl~ct('rÍjlc' Ilactcria are oftcn prescnt as dry bilcter-ial ooze
cíther on thc 01 umcs or in the mi cropyl c. Seco trea tment reduces i nfect i on
by Xilnthoi'lnllilS oryzilc, ill1tibiotics being more effective than fungicides --'".~----------. '.'--- --~.--
(82). Secd trcatl,lcnt I'lith ilntibiotics such as aurcofungin and a91'il1\ycin
and, fungicidcs sucil as cercsan and orthocide give good control of f..oryz~.~
(110) and 'f. S!!YJ.:~ improvino germination and seedling growth (10,82).
Tllere are, howcvcr, other efficient and less dangerou$ methods to control
this bacteria. t:auffman and Reddy (70), for instance, reported that
bacterioplloges may playa good role in reducing the bacterial population
in gCflllinating secd. ]i.. oryzae may be also controlled by Ü'eatments uscd
to break dormancy. I\ccording to Subramoney and Abraham (133), soaking
thc secd in dilute nitrie acid for 16-24 hours washed artd sun-dried, then
soaking ¡¡gain in water and germinated is obtained a good control of the
bactel'i a. 1 t has been reported that Xantholnonaas oryzi ea 1 a can be
controlled efficiently by soaking the seed for 12 hours in a mixtul'e of
agrimyci 11 and ceresan ¡¡nd 1 ater hot ,.¡ater treatmcnt at 52°C fot' 30 mi I1UtCS
(125). Ily irradiating the seed, packing it into polyethylene bags and
stored under suitilble environmental conditions it is possible to obtain
a good control of Pseudomonas spp.(64).
Secd borne nematodes. The leaf or white tip nematode Aphelenchoides besse2'l
can ramain in the rice Kemcls as long as 2 years (5). The pl'esenee of
more than 200 nematodes per 100 seeds of rice warranty contl'ol mcasures
(51). Some nematicides have be en repol'ted to control this nematode efficiently.
Chemi ca 1 s such as ~1ethyl bromi de, neguvon aod sys tox among others have shü\·m
to reduce thc population of~. besseyi significantly (37,~l). I\ceording to
Templeton ~~. (142) seed treatment with benlate erildicilte the nCllliltodc
;
. .
•
•
from infcstcd seeds. A1though the applicatioll of nematicídes is ve/'y
expcnsivl', i t has been rcported that the soi1 trcatmcnt \Vith '-'asudin
givcs satisfactory results in controlling the stem nematode Ditylcellcl/_lJS_
.3. Therll·otl.~cl'i)py. Heat treatment is effective against all classcs of
sccd Dome pathogens, but usually is used ugainst pathogens so
Sit"Uiltcd as to be protected fl'om chemicals. There are thrce typcs of
such treatment: hot water, aerated steam, and hot dry airo Hater has
twice the thermal capacity of saturated aerated steam, and five times
that of dry airo Temperature or time of treatment therefore increases
from 49-51 oC for 30 minutes for water, through 54-60 d C for 30 minutes
for aerated steam, to a range of 54°C for 5 hours, to 95-100·C for
12 hours for dry ait,. Since temperature and time are more or less
mutually compensating, it is desirable to standarize on a 30 minute
period (9). Hot Nater treatment has long been used and is very effective
in controlling seed borne pathogcs of rice. Suryanarayan ~ 2.l (136),
for instance, reported that the treatment of seed infected by Trichoconis
padl.¡ickiiwith hot \.¡ater at 52°C for 10 minutes increases the
germinatian significantly. Although the increase of germination in
many cuses is not very high, many of the seeds may have lost their
viability as a result of infection befare the treatment. Excellent
control of Galansia oryzae has been a1so obtained with hot vJater at
54°C for 10 minutes ~nd by solar treatrnent (92,93). According to Sinha
and Nene (130), soaking the seed in water at room tcmperature for 12 . .
hours, then at 53°C for 30 minutes can eradicate Xal~JlOrnonas 9.!..l'zae
.'
- 23 -
minutes givcs 900<1 control of !\phelenchoidC's b~scyi (38). Follo,ling any
thcrtnothcrilpy trc,atmcnt the secd shou1d be trcated \;itb a mild fungicide.
Sincc, seecl gCfminatiol1 is sOlncwhat retilrdcd and Vleakcnccl, this pl'otectiol1
frolll sDi 1 mi crool'gilrli SlnS i s benefi ci al. , .
CONTROL OF STORAGE PATHOGENS
Tbere are three basic direct methods utilized to control losses by storage
pathoCjcns; al drying to a moisture content safe for storage, b) ael'ation to
mainté\ i n 10Vl and uni form temperatures and thereby prevent moi sture tré\ns fefS
and, e) refrigeration.
Q!J'in_~. The proeess of drying is somewhat complicated and invo1ves more
than just drying to a moisture content necessary to meet grade standards
and for protection fl'om storage losses. The methods used must a1so maintain
quality such as gennination, mínimum breakage, etc. Sometimes this becomes
difficult because of the economics of drying. The 1011ger it takes to dl'y
a 'load, the more it costs. Thus rapid drying becomes desirable from an
economic point of víew. Rapid drying, however, requíl'es hígh tempe¡'ature
(60°C) and this tcmperature results in low seed germínation and chemical
a lterat ion. 1 f the gra in i s to be used for seed i t mus t germi na te and a
high telllperature decreases germination resuHing in poorer quality and
price for use as seed. If the graio i5 to be used as feed the importance •
of high temperature drying i5 lessened.
If the moistul'e content is maintained at a lIniform1y 10\"1 level, rice can
be stored for several years \'Iith 1 ittle detcrioration, cven \'lh(,11 otiler
factors are lInfavorable (124).
- 24 -
r\er~~tio_f!.. lile IIID:;t 91'llcl'a1 ~Iay tD maintain low and uniform moisture
content, as I'¡cll as unifonn tcmperutul'cS, is acriltion, Thc ma'in fUllctions
of acration ül'C to a) reduce thc tcmpcrature of storcd grain to about
5-10~C, and 1») Illdintuin il uniform tcrnpcrature, VJhich in turn pl-events
moi s ture mi gl'at ion.
AC1'ation is 1",Her and clleaper than other ways of maintaining tempel-atur~
ond moiSturc. It also has the advantage in that not on1y docs it prevcnt
moisture migration and I'¡ater from being absorbed into the 91'oin, but it
often resu1ts in additional reduction of initia1 moisture content, somc-
times as much as 1%. This reduction becomes important for two reasons.
First, most of the water removed is lost from the outer l~yers of the
seed, wlli1e tlle inner 1ayers may stay at a high 1evel. Thus, \1hi1e the
i/lner parts of tlle seed may have moisture levels conducive to fungal growth
or infection, the outer layers may be too dry to allow init'ial infection
to occur. Secondly, such a moisture condition in the seed allows one
good protection from fungi under a favorable economic condition since
one does not llave to reduce tlle moisture content of the entire seed be10w
tile grade requi rement.
According to Christensen (35) temperatures below 25°C are preferable, since
low temperotllrcs lIsually decrease the activity of most storage fungi.
Ilowcver, sonle fungi grow well in the range of 5-10°C. r1any Penicillilll\l
species grow at low tel\lperatures in the presence of sufficient moisture.
Ile also observed more Kernels invoded by species of AspergillllS gli\uCUS ,
group in rice adjusted to an 18.5 percent moisture content and stored
at 12°C tilan in similar rice stored at 27°C. The relation, however, \,as
I
- 25 .-
reversed for spccies Di the !l. ilayus anel 12. candidus groups. Fanse and
Chl'istcnscn (49) stored rice \'Iith moistllre contents of 12,14 ancl 16 Jler
cent ilt 5"C for 465 days withollt a reduction in germinability, althol1gh
thcy. obscrvcd an incrc¡¡se in tile ¡'ate of invasion by storage fungí in rice
~lith 16 Jlor cent of moisture.
Refrigcrntion. This process is usually one of storing grains at o·e with ----'--_._ .. _- a moistl1ro content of 20 per cent or more. Cooling to too 101' a temperature
results in large difforences betweeJ1 grain temperature an air temperatul'e,
leading to moisture migl'ation. Hilen this Decurs, organisms sucll as
Penieil1iu!il, yeasts and bacteria, which can grow at low temperatures
provided that 1I10isture content is about 20-25 per cent,can invade and
cause slow deterioration. 80th Penicillium and bacteria can grow at or
belO\~ froezing provided that moisture content is about 22 per cent. Then.
if the temperature rises, sucll as when the grain is removed for transport,
rapid colonization can occur, resulting in incipient deterioration within
24 hours. So in essence, while refrigeration gets rid of the Aspergilli
and other fungi, \'le may often simply be seleeting a new group of organisms
as the cause of.storage losses. Much more information needs to be obtained
about l'efrigeration with high moisture content befo re it becomes establ ishcd
as a predominant form of storage and control of microbial deterioration.
Fungicid0s. In general, the fungicides have little utilization in the
control of storagc fungi. First, at present, they are genorally ineffective
and secondly, governmcntal regulations prevent many potential control
fungicidcs from bcing used in grain destincd for consumption. More
.-
bp.comin~ mor-e prC'valcnt. One intcrested in the possibi1ity of chelllicill
control of storilgc fungí should consult both a specialist in seed [lathology
and a reccnt pl'sticide manual for recommendations.
,
LI\r\Oi\.~TORY 11ETIIOllS FOR DETECTION OF SEEDGORNE MICROOHGANISI1S
A. Ayar Jl.l_i~t..c'._t('~;~. TI'IO sets of foul'-hundl'ed seeds per each cultival'
are sccded on petri dishes containing 2% agar. Four-hundred seeds
are sceded \,¡ith gIUlIlp.s,the other ones are seeded without them. Scc>ds
are pn'trcated by soaking 5-10 minutes in a 1% hypochlorite solution.
The solution is poured off and the seeds rinse with sterile water
prior lo planting onto the agar surface. Ten seeds are placed per
plateo The seeds are well distributed over the plate and incubated . at 20-25% for a per'iod of 4-8 days (Sheppard, 1979). Readings of
seed genllinatíon anó number of seeds from vlhich bacteria or fungí
01' both ha ve grown after 4 and 8 days are recorded. The different
types of bacteria are transferred to nutrient agar plus calcium
carbonate tubes and the fungi to potato-dextrose-agar (PDA) plus
lactic acid and V-8 vegetable juice agar for later identification and
pathogenicity test. The isolates are storeged in tubes containing
sterilized distilled water.
B. ~4-D blotter test. The blotter test is actually a modification of a
germination test as the seeds are placed on I.he surface of moistened
filter papel' discs. The blotters are saturated with sterile water and
the excess water is poured off. As in the agar plate test,two sets
of 400 seeds with and withou~ glumes are tested. Seeds wíth glumes
are sown onto tile blotter with a pair of forceps. As in the agar
;
,
tcst thc grovlth of SQcdlings often makes ObSCI'viltion of fungi
difficu1t and dctermination of percentage infection impossiblc, a
0.2% solution of 2,4-0ichlorophenoxy ilcetic-acid (2,4-0) to prevent
growth of seed1ing5 i5 uscd (Neegaard and Saad, 1973; Sheppard 1979)
Tlle petri dishcs are wrapped in papcr in order to reduce cvaporation
of the 2,4-0 in the incubator. They are incubatcd at 20-25"C with
a 12 hour dily/night NUV light cycle fur a period of 4-8 days.
BIOflSS!\Y OF FUNGICIDES ON TREATED SEED
Test J!..S@.r::.. A suspension of a given pathogen washed from the surface
of PDA (or other medial petri di5h with sterile distilled water is
diluted to approximately 150,000 spores/m1. This suspension is
mixed with either POA or V-8 vegetab1e juice agar in the ratio of
1:4, giving a final spore concentration of 37,500 spores/ml of seeded
aga\'. Twenty m1 of the seeded agar are poured into each p1ate. Five
p1ates with four seeds are used for each chemica1 and each standard.
Stand a rd CUI've. A standard curve i s prepared to l'e 1 a te tile di ameter
of tlle zone of inhibition to the concentration of chemical presento
Five 10ts of seeds are treated at five different concentrations, i.e.
0.50, lOO, and 1000 ppm. Tilese seeds are p1anted out on test agar,
and tile diameter o, the resulting zone of inhibition is measured. The
average zone for each treatment is plotted on tho linear scale of
scmi10g papar against the concentration of treatment (on tlle 10g sca1e).
Dy entet'ing thc curve with the zone of inhibition, th~ concentration
of "equivalellL effectivcness" fo!' each chell1ical can be dctcnnined.
,
•
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:tC: . .sJ:_i!l5L~.t!!'~L~.. Tile seeds to be tcsted are pressed into the sced
it(¡ilr to the botton of tile plate so the enti t'e surface of the seed
wil1 be in contact with the agar. Tile p1ates are incubated for 40-48
hours at 20-25°C. The diameter of inhibition lone around each seed
is lIleasurecl anel the average determined. This diameter is converted to
equivalcnt concentration af seed treatment chemical by reference to
the standard curve (Mitchell, 1967).
Dama(Lc as~~nt. A lat of seeds is used to study the eHect of tile
given patnogens on seed germination and disease transmission. The
secds are artificially infested in the follüwing manner: Twenty ml of
sterile dlsti1led water are placed on each retri di~h culture and the
myceliunl and sporcs from the agar are separated. The resulting
suspcnsions are combined and enougil water is added to bring the
conecntration at approximately 150,000 spores/ml. Two-hundred seeds
are dipped into it and the they are placed to dry on a piece of filte\'
paper at room temperature.
Olle-haH of ea eh 10t of infested seedsis treated by placing
approximately 20 at a time in a small beaker with a small amount of
thc best chemical from the bioassay test. The other half of the
seed will serve as untreated contro15. Another 10t of seeds is
reserved that is neither infested non treated to serve as germination
contro1s. Also several seeds that have not been infested are treatcd.
These seeds will serve to show any rhytotoxic effects the chemical
a10ne may have on the secds or seedlings.
j/r;,~~7\:-~!·--1-
- 30 -
$I11G11 plilstic )lDtS ilre fi11cd with soil or silnd autoclilved for 1 hou)' <lt
a presSure of lS lbs/se¡. in. The secds are plantcd (five of the sai1le
treiltment pe)' pot) in the gl'eenhollse in labeled pots and watcred as
nec'ued. Every otiler dily observatiol1s a)'e made. The number of seeds tilat
germinate and Lhe number of seedlings showing disease symptoms are
recorded (Stevens, 1967).
1. Anon. J962. Anntlal Report 1961-1962. Research and specialist services, /1inistr'y of J\grictllttlre, Northern lligo¡Oía. 53 pp.
2. Aganlal, V.K. and O.V. SínrJh. 1974. Relative percelltage incidence of secd borne fU~Ji associated with different varieties of rice seeds. Seed Rcsearch 2: 23-25
3. and S.C. Modgal. 1975. ¡nfluence of different doses of nitrogen and -'--Sf)ilcing on the seed borne infections of rice. Indian Phytopath.
28 (l): 33-40
4. Aguiero, V.M.; O.R. Exconde and r.J .. Quintana. 1966. Seedborne fungi of rice and their response to secd treatmcnt. Philipp. Agri. 49 (10) 871-873
5. Atkins, J.G. and M.A. Marchetti. 1979. Rice diseases. United Sta tes Department of Agriculture. Farmers' Bulletin No.2120. 19p.
6. Aulakh, K.S. 1966. Rice, a new host of Curvularia verruculosa. Plant Ois. Reptr. 50: 314-316
7-. Aulakh, IC.S., S.B. ~jathulo E. P. Neergaard. 1974. Comparison of secd­ llome infection of Drechslera oryzae as recorded on blotter and in soil. Seed Sci(~nce and Technology 2(3): 385-391
8. Baker, K.F. 1972. Secd Pathology, In seed biology, Vol. 11 Germination control, metabolism, and pathology, (Ed.) T.L Kozl1o\'lsk"i. Academíc Press, New York pp.317-346
9.
10.
Baker, K.F. 1979. Seed Pathology - Concepts and methods of control. Journal of seed technology 4(2): 57-67
Bedi,P.S.and D.S.Dhaliwal 1970a. Studies on the seed microflora of rice variety Taichung native - 1 \~ith special reference to its pathological significance. Jnl Res., Ludhiana 7(4): 600-605 .
11. __ ~ 19l0b. spermatoshere of rice variety native 1 from different states of India. Indian Phytopath. 23 (4): 708-710
12. Benoit, M.A. and S.8. Mathur. 1970. ldentification of species of Curvularia on rice secd. Proc.lnt. Seed Test. Ass. Vol.35(1): 99-119.
1.3. Bernaux, P. 1966. Principal fungi recorded on rice in Camargue. Annl. [piphytics 17: 43-52 .
14. Boller, R.A. and II.li.sch\'oecler. 1974. Influence.of temperature production of aflatoxin in rice by J\sp(~rgillus parasiticus. 64: 283-286.
on Phytopathology
:l'
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15. Bradbury, J.I'. 1971. Nom0nclaLlIre of the bacterial leilf streak pathogcn f 'A 1 t J Syst. !Jilct .. 21(1):72. o n e". n' . .
16. Btlgnicoul,t, r. 1955. Deux espcces nouvelles & Hell1linthosporiul1lisDl~s de SClIl8nCeS de I'iz. Rev. Gen. Got., G2: 238-243
17. Casti\rlO, J. 1974. Iden.tificociÓn de ncmátodos fitopar{¡sitos del arroz (OI'YZil sativil L.) en arcas del Estado de: 1-lorel05, ¡·léxico. Universidad I\utonomil Chapincjo, Colegio de Postgraduados, Rama de Fitapatologia, Chapin90, ¡·léxico. 1Gp.
18. Caro'lis, C. Oc. 1972. t'lyrothecium venucaria (Alb. and Sch\'l.) Ditm.ex Fr. su I'iso in Italia. Riso 21(3): 253-258.
19. Centro Internacional de Agricultura Tropical. 1982. Rice Grain Oiscoloration. Annual Report Rice Programo CIAT, Cali, Colombia.
20. Chakravarti, B.P. and M. Rangarajan. 1967. A virulent strain of Xanthomonas orylae isolatcd frolil rice seeds in Indía. Phytopathology 57 (7): 688-690'
21. 1968. Studies on bacterial blight of rice: association of Xantholllonas --oryzac with secds and varietal reactions.Oryza 5(1): 20-25
22. Ctlilntarasnit, A. 1971. Identification and role of pycnidíal fungí associated \vith rice seed. 'Thailand J. r\gric. Sci. 4(1): 17-28.
23. Cha ttopadhyaj, 5.8. and C. Dasgupta. 1959. f\elminthosporium rostratum Drechs.on !'ice in India. Plant Dis. Reptr. 43(12): 1241-1244.
24. Chceran, 1\. and J.S. Rao 1966. Effect of seed treatment on the germination of ri ce seeds i nfected by Tri choco))i s padvli cky Ganguly, Agri c. Res. J. Kerala, 4(1): 57-59.
25. Cheerwick, W.J. 1954. Studies on seed-borne microflora and the effect of seed tt'eatment of rice. Malay. Agríc.J.37: 169-172.
26. Chevaugcon, J. and A. Revise. 1957, discases of rice in West Africa. 4: 143-151
The ~ater regime and parasitic Journal Agrie. Trop. Bot. Appl.
27. Chidambaram, P., 5.8. Mathur and P. Neergaard. 1974. sced borne Drechslera species. Handbook on secd Series 2B (3). ISTA. Nonlay. pp.165-207.
Identification of health testing.
28. Chien, C.C.) C.L. Chu and J.M. Liu. 1973. , Studies 011 seed disinfection of rice. Journal of Taiwan Agtic. Res. 22(4) :263-269.
.,
•
- 33 .-
31. Chl'istensc,n, C./1. and P. Lilll~o 1963. /~oisture contents of harel red wintel' ilS dl'tr,rmi neo by 11Il,tcrs ¡¡nd by oven dryi n9, ilnd i nfl uencc of sllIilll di ffcrr,"Cl'<, .¡ n n¡(J"i s Lun: content upon subsqucnt dctcri Ol'at i on of the grilin stOI'.J<]c. Cc¡,c,al chcmistry 40: 120-13/.
32. , 196';. Funrji in cereal grains ilnd their products. In mycotoxina in foodstuffs. G.N. llagan (Editor). Prcss: Cambridge, Hass. 9p.
33, and 11.11. Killlffll1illl. 1965. Deterioratíon of stored grains by fungi. ---i\liñüJl R<.:vic:¡'/ of Phytol'Jtholo9Y 3: 69-84.
34. and L,r, Lopez 19G5, Relation of moisture content and length of - stordgi' to Chill1r!esi 11 til0 mi crofl ora and ge¡"mi nati 011 percentage of
rough rice. Phytopat:hology 55: 953-956.
3~;. ,1969. Influence of moistlwe content, tenlperature, and time of ---storagc tipon invasion of rough rice by storage fungi. Phytopathology
59: 145-Fl8.
36. Corbctta, G. 1953. Researches and experiments 011 diseases of rice (Oryzil sutiva L.) VIl¡. The rice disease caused by sclcrotium ol"yzae catt. (= Helminthosporiul11 sigllloideulll cavo = Leptosphaeria salvínii ciltt.). Phytopath. Z. 20 (3): 260-296.
37. Cralley, E./'1. and R.G. FI'cnch. 1952. Studíes on the control of v/hite tip of rice. Phytopatholo9Y 42: 6
38. _----,~' 1956. A new control measure for whíte tipo Arkansas Fm. Res. 5(4):5
39. De Mello, R.E. and De Sonja, O.M. 1962. nos Arrojíils do Estado de Sao Paulo.
Ocorrencias de doencas e pragas Biologico 28:33-45
40. De /'le110, R.E. and A.F. Cintra. 1968. Condicoes fítossanítarías , de sell1entes de arroz no Estado de Sao Paulo. Bilogico, 34(8): 175-178
41. Del Prado, r.A. and C.M. Christensen, 1952. Grain storage studies 12, The funglls flora of stored ríce seed. Cereal chemistry 29:456-462
42. Devi, L.R. and M.R. Menon. 1970. Effect of treatments on viabílity of paddy seeds. 54~55
fungieidal pre-storage Agríe. Res. J. Kerala 8(1):
4.3. Días de Toledo, A.C., Amaral, R.E., D.t1. SOl/za and H.V. de Arruda. 1971. FlIngicidas sucedaneos dos mercuriais para tratalllento de sementes de arroz.[liologico .37(11): 300-302
,
,
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- 31) ,
45. Dunsmore, J.R. 1970. InvcsU9dtions on the vilrieti<::s, pests and diseascs of llplilnd rice (hin [lildí) in Sarill'/ak, Millilsya. Int. Rice COIDIIl. Nev/sl. 19(1): 1''1-35
46. Eamchit, S.iJ/ld S.H.Ou. 1970. baclerid! bliyht of rice 33-45.
Some studies on the transmission of through seed. Phillip. Agric. 54 (1-2):
47. [suruoso, o.r., C.O. Komolafe ane! ¡'1.0. Aluko. 1975. Seed bOrne func]i of rice (oryza sativJ) in Nigeria. Seed Sciencc afld Technology 3(3/4): 661-666.
48. Faan, II.C. and S.C. Wu.1965. Fielel control of bacterial leaf streak (Xanthornonas oryzicola) of rice in K\'.Jangtung. Acta Phytophylac. Sin. 4(1):1-6.
49. Fanse, H.A. alld C.M. Christensen. 1970. rough ri ce in commel'ci al storage ami 60: 228-231.
Invas ion by storage fungi in in the 1 abora tory. Phytopatho 1 ogy
50. Fazli, S.F.and H.\'.Schroeder. 1966. Kernel infection of Bluebonnet 50 rice by Helminthospol'ium oryzae. Phytopathology 56:507
51. FCilkin, S.D. (Editor). 1971. Pest control in rice: Rice leaf or Ivhite tip nematode. PANS Manual 3: 99-100.
52. Ferrer, A., P. WillialD y R. Moises 1980. Control de hongos patogenos transmitidos por semilla en arroz. Ciencia Agropecuaria (3): 113-117
53. Govindu, H.C. 1969. Occurrenee of Ephelis on rice variety IR8 and cotton grass in India. Plant Ois. Reptr. 53(5) :360.
54. Hasbioka, Y. 1970. Rice diseases in tlle world - IV. Phycomycetes (Funga 1 di seases - 1 J. V. Smuts Riso 18(4): 279-290.
Disecases due to (Fungal diseases-2)
55. Ilashioka, Y. 1971. Rice diseases in the world.- VIII. Diseases due to l1ypocrealcs', Ascomyeetes (Fungal diseases - 5) Riso 20(3): 235-258
56. Haware, M.P. and N.O. Sharma. 1973. A new glum~ blotch of rice (Oryza sativa). Plant Disease Reporter 57(5): 436-437.
57. Ilesseltine, e.IL, O.L. Shotl,ell, J.J.Elis and R.O.Stubblefield. 19G6. Aflatoxin formation by Aspergi11us flavus. Bateriol. Rev. 30:795-805
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59 .
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Huang, Y.T. and C.M. Yu. 1973. thc non-sublllcr'Cj('r! sccldl inq by sccd treo tlllcnts. 1'1 ant
Studies on sccdling discases of rice in bed. L Control of secdling blilJilt of rice PraL Bull, TaÍYlan, 15(4) 139-146
60. HlI¡¡n~J, CS., F.r. Cupi'rtino und N.t·L Martinclli. 1977. Incidence of whiLe-tip nematodc, ~.pl\Clenchoides besseyi in stored rice secd frolll CI,nLI'al vlcst llrazil. Pf\NS 23(1): 65-67.
• 61. Hung, Y.C. ilnd C.C. Chicn. 1971. Reevaluation of mercul'ic fungicides for rice secd treaLlilcnt. J. Tai\'lan I\grie. Res. 20(1):61-66.
62. ¡'¡yun, J.S. 1963. Dev(C'lopmcnt of st.orage fungi in polished rice infestcd with rice Vlcevil, Sitophilus ol'yzae L. Seoul Univ. J. Ser. B. 13(8): 71-86
63. Ibrilhim, I.A. and S.A. Farag. 1965. 1\ study of some fungi isolated from grains of Egyptian rice varictics. Alex. J. Agl'ic. Res.13(2): 427-437
64. Iizuka, H. and H. Ito. 1968. Effect of gamma-irradiation on the microflora ol' rice. Cereal chernistl'y 45(5): 503-511.
65. Imilm Fazli, S.F. and H.H. Schroeder. 1966. EHect of Kernel infection of rice by Helminthosporium oryzae 011 yield and quality. Phytopathology 56: 1103-1105.
66. Jackson, E.A. 1966. Tropical rice: The quest for high yield. Agr. Rev. 4:21-26.
67. Jain, B.L. 1962. T\;o new species of Curvularia. Trans. Brit. IQycol. Soco 45(4):539-544.
68. Kang, C.S., P.Neel'gaard and S.B. ttlathur. 1972. Seed health tcsting of rice. VI. Detection of seed bOfne fungi on blotters under different incubation conditions of light and temperature. Proccedings of the International seed ~esting Association 37(3): 731-740.
69. Kang, C.S. and CK. Kim. 1972. Studies on the fungi associated \'Iith ear blight of rice. Korean J. of plant Protection 1l(2): 101-107.
70. Kauffman, K.E. and A.P. Reddy. 1975. Seed transmission studies on Xantliolllonas oryzae in rice, Phytopathology 65(':';) :663-666
71. Kothcimcr, J.Il. and C.t1. Christensen. 1961. Microflora of barley Kernels. Hallerstein Lab. Commun., 24: 21-27.
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73.
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Kuli~, M.M. Jnd J.F. Scll0rn. 1977. Proccdures for thc routinc of SC(,U hnrlH' jJatllofjenic fllllgi in seed tcsting laboratory. of sccd T('(:llnol09Y 2: 29-39.
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