Search for Causative Agents of the Sugarbeet Yellow Wilt in Chile 1 C.
URBIl"A-VIDAL AN D
H.
IIIR UM T2 .:1
Received for publicatio1l February 28, J 974
Introduction
Ben nett and co-workers r('po rted the finding s o f th e ir in ves tiga­
tio n of the yellow wilt disease of su ga rbee t in Argentina (4)4 in 1946
and in C hil (' (3) in 1967. Th('y conclud ed that (a) yellow wilt apparen tly
was cau sed by a single viru s, desig nated Chlorogenus jJatagoniensis n. sp.
(or B ela virus 6) ; (b) there were two distinct sets of sy mptom ex pres­
si o n: o ne the "yellowing phase" an d the oth(' r the "wil ting phase" ;
(c) the causativ e agent could be trans mitted by grafting diseased to
hea lthy tissue, by each o f tw o sp ecies of dodder , Cusculcl subinclusa and
C. campestris, a nd by a lea fh o pper ,Pamtanus exitiosus; an d (d) the disease
oc curs in th e fi eJd on sugarbe et, table bee t, fodder bee t, ano Swiss
ch ard in addition to 14 other host plants tested in th e g reen house and
2 t('s ted in the fi eld. Des pite th eir e xte nsiv e studies, many prob lel,ls,
su ch as visua liza tio n of the causative agent ; its fatc in host plants and
in insect vecto rs; isola tion, charac teri za ti o n and id entifi cation of the'
agent; an d effective methods of controlling the disease remained
un solved .
At the time when the studies described above were mad e, th e
existen ce o f a new gro up of pla nt pa th ogens , the mycoplasmalike
organisms (M LO) (7 , 19) had not yet been desc ribed . T he MLO, now
cons ide red to be the eti ologi cal agen ts of a number of ye llow s- typ e
plant di seases, are transmitted by leafh o ppe'r, plan th o ppn , or psyUid
vectors, as well as by gra fting and d od ders (2 5, 33). Th e ivlLO are
se n sitive to tetracycline d er ivat ive s. Base d o n this in fo rm a tion ,
Ehre nfeld (11) de m on strated tha t the ye llow wilt disease of sugarbeet
was supp ressed, but not eliminated, by chlorotetracycline treatments
and suggested that the agent of yellow wilt was not a virus, but a MLO
entity. Subsequently, Urbin a-Vidal and Ehrenfe ld (re ported at the
I st I nternational Co ngress for Bac teri ol ogy, J er usal em , Israel, Sept.
3, 1973) , usin g el ectron mi croscopy thin sectionin g techniques, o b­
'This " 'ork ,,'as s po n sored. in pa n, by the Age ncy fO l' 11llernati on a l Deve lo pm e nt, by resea r c h
grants 'l o s. GB -I 1861 an d GB -2')2 HO to K. :Vlaramoro sc h from th e :'-iational Scien ce Foun dation .
Washin g ton . D.C. ; b\' the C niv e rs il\ of Ch ile, Sant iago , C hil e; a nd b y Boyce Thomp son Institute ,
Yo n kers, ); . Y.
'R es p ecti\'e h: Cel l Bi ologis t, Ln i\'er s it~ ol'Chile, Santidgu, Ch ile; Ce ll Bi o logist. Bo yc e Thomp­
,O il I n st itut e, Yunkers . ); .Y . 107 01
:~ ."ddn'\s for re pr int reque'il: H. Hi nll lli , Bo yce Th ompsoJl I n~l ilUle for Plallt Re sc(-I ]"ch , J08f)
); 01'l h Broa ch ,,1\', YOJ1 kell , :'-i. Y. 107 01
~:\llmbe J) j'n pare!lt! )('st'" refer
(0
li l~ r dlurc ciled.
VOl.. 1R, \:0. '2, O c:-roBL R 1074
I f:l
served microorg an isms in ye ll ow wilt-affected sugarbee t plants,
morphologically rese mbling kn own ML O (25 ,33) . Furth e r stud ies to
visualize the pre sumptive et iologica l age nts of yellow wilt in the
diseased sugarbeet plants we re ca rried out, base d on a working hy­
pothesis that the ye llow wilt disease ma ybe caused by a co m plex of
seve ral agents. Th e pre sent re port d escribes the coex iste nce of at least
tw o possibl e e tiological age nts of the sugarbeet yeil ow wilt disease in
Chil e.
Materials and Methods
Sugarbee t (Beta vulgaris L.) seeds we re seeded directly in a field
of the La Pl atin a Ex pe rim en tal Statio n, Sa n tiago, Chil e, in October
1972. About 50 days later sym pto ms of yell ow wilt began to a ppear.
During the next 9 months, th e a ppare nt in cid e nce of in fect ion in­
creased gradually , finally reaching about 95 % in Se pte mber, ! 973
(Fig. 1). Durin g th3t pe ri od , near ly all of th e affected plants developed
sy mptoms o f the yellowing phase of the disease includin g ve in clearing
a nd yellowin g. In the later stages of disease d evelopme nt during that
period ma ny plants exhibited typical "witches broom" clusters with
d warfing a nd num erous ax il a ry buds (Fig. 2), a nd som e pla nts died.
Sym ptoms of the wilting ph ase developed on a low p erce ntage of the
affected plants, but usually in combination with symptoms of the
yellowing phase.
Portions o f leaves a nd petioles were excised from th e infected
(Figure 2) as well as from norm a l-appea ring pla nts and prefixed with
3% glutarald e hyd e in 0 .05 M sodium cacody late buffe red solution
(p H 7.2) in th e field at th e e nd of Septe mbe r. Specim e ns in the fixativ e
we re processed for electron microscopy examination 5 days later .
A fter rin sin g in the cold bu ffered soluti on with 5% sucrose ove rnight,
they were postfixed with 2% os mium tetro xid e in th e sa me buffe red
solution for 6 hours at 4°C . T he fix ed specim e ns were d e hydrated at
4°C by passing through a graded e thanol series of 50,7 5, 90 ..and 95%
for 15 minu tes ea ch, and at 100% for 60 minute s. Subsequently the
specime ns were passed twice th rough propy le ne oxid e for 60 minutes,
a nd embedd ed in a resin mix ture of Epon 812 and Araldite 506 (27).
U ltrathin sections were cut with diamond knives on a Porter-Blum
MT -2 microtome , and stain ed with 8% urany l magnesi um acetate in
distilled wate r for 3-10 minutes at ro o m te mperatu re, followed by
0.4% le ad ciLra te in 0 . 1 N sodium hyd roxide for 20-60 seco nd s. The
sections were studied with a Sie me ns Elmisko p I electron microscope,
modified to a model lA , at 80KV.
Results
T wo types of possib le et iological agents of th e yellow wilt were
detected in the diseased plants, but were never obse rved in the norm al­
appearing rnate rials. O ne was a mycoplasma like organism (M LO) and
ll -f
J Ol- R,\ .~L OF THE .-\ .
S. S. B. T .
Figure l.-Suga,-beet field in early September at the La Platina"Experi"
mental Station; 11 months old. More than 95 % of the plants showed yellow
wilt symptoms.
Figure 2.-Diseased plants showing advanced stages of the typical
yellowing phase, including witches' broom clusters, dwa rfing, and leaf
necrosis.
the other was a n elongatt"d , fle xuo us viruslike particle associated i~lith
characte ristic inclu sion s.
:V1ycoplasmallhe organism (M LO)
Pleomorphic bod ies wh ose morph o logy resem bl ed that o f micro­
o rga nism s bel ongin g to ~1 yco pl asm ata l es which have been isolated
from animals (l, 13) , a nd M LO wh ich h ave been observed in more than
VOL.
18, ]\io. 2, OCTO[lER 1974
145
50 yellows-type diseased plants (16-18, 25, 33), were detected in the
phloem tissues of th e yellow wilt-affected plants. The MLO were ob­
served either in mature sieve tube elements, devoid of cytoplasm and
filled with sieve tube sap (Fig. 3), or in the cytoplasm of phloem paren­
chyma cells (Fig. 4).
These bodies, bounded by a trilaminar membrane (unit mem­
brane), contained ribosomes and deoxyribonucleic acid (DN A)-like
strands and lacked a nuclear envelope and a cell wall (Figs. 3 to 6). They
varied considerably in size and electron opacity. Wide variations of the
m<:>rphological profiles, such as spherical, ovoidal, amoeboidal, long
ellIptical, horseshoe-shaped, and doughnut-shaped forms, were ob­
served (Figs. 3 to 6). The majority of MLO varied in size from approxi­
mately 100 n m to 800 nm (Figs. 3 and 4). However, some of them, parti­
cularly those in the phloem parenchyma cells, had fine , long cyto­
plasmic protrusions containing electron-lucent cytoplasm (Fig. 6) .
The number of MLO in each host cell varied considerably
(Figs. 3 and 4) . Some sieve tube elements, as well as phloem parenchyma
cells, were tightly packed with numerous MLO, while others contained
a small number of MLO scattered in the elements. In the cytoplasm of
phloem parenchyma cells, "colony-type" accumulations (17) of the
MLO were often seen (Fig. 4). In general, MLO around the sieve plates
were more crowded on one side of the plate than on the other (Fig. 3).
MLO, partially squeezed through the sieve pores, were observed often.
This may indicate that the MLO move from one sieve element to a
neighboring one via the sieve pores.
Morphological configuration indicative of binary fission, budding,
extrusion of the protoplasm and tightly lined bodies, which were
similar to those considered earlier to be undergoing reproduction in
sieve tube elements (16) or in phloem parenchyma cells (17), were also
seen in the phloem of yellow wilt-affected sugarbeet plants.
Elongated viruslike particles
Elongated, flexuous viruslike particles were observed in the cyto­
plasm of mesophyll cells and phloem parenchyma cells of leaves ob­
tained from the diseased plants, but not in the normal-appearing
materials. Morphological characteristics of the particles di ffered from
that of microtubules and phloem protein (P-protein) fibrils , which arc
normal cytoplasmic constituents (Fig. 7). Although measurements of
elongated, flexuous particles in thin sections did not reveal accurately
their length, they appeared to be more than 650 nm in length and
10- 12 11 III in d ia nlf'ter. Tht' partic: les 0 ften appeared in more or less
parallel <llTangelllents, oriented perpendicularly to cell \\'alls (Fig. 7
t () ~I).
116
J OURNAL OF THE
A . S. S. B. T.
Figure 3.- P o rtion of phloem tiss ue fr om a yellow wilt-infected plant.
Pleomorphic mycopl asmalike organisms (MLO), varying in shape and size,
are p rese nt in the sieve tube elements (S). Note some MLO apparently pass­
ing thro ugh the sieve pores (SP). CW: cell wall; P P: Phloem parenc hy ma
cell ; SPL: Sieve plate. X 12,950.
VOl.. I R. :\0. 2.
OClOlllR
1974
147
Figure 4 .-Portion of a phloem parenchyma cell from a yellow wilt­
diseased plant. Note colony type accumulation of intracytoplasmic myco­
plas malike organisms (MLO). CY: Cytoplasm; CW: Cell wall. X 7,000.
1-1 H
.fOU~:\-\l 01 1111.1..
S. S. B.
r.
Figures 5 and 6.-Intracytoplasmic MLO in a phloem parenchyma cell
of a diseased plant. Note the varying morphology and electron opacity of the
MLO (Figure 5, X 48,250), and the long electron lucent extrusions of MLO,
closely associated with filamentous particles (FP) (Figure 6, X 35,000).
VOL. lR , No .2,
O CTOBER
1974
149
Figure 7.-Portion of a mesophyll cell of a yellow wilt-diseased plant,
containing elongated viruslike particles (VLP) associated with a pinwheel
inclusion (PW). Note VLP in more or less parallel arrangement and free
VLP . MT: Microtubules. X 60,500.
150
JO URNA L OF THE
A . S. S. B. T.
Figures 8 and 9.- Portions of mesophyll cells obtained from a yellow
wilt·diseased plant. Note characteristic bundle inclusions (Bi ) (Figure 8,
X 66,000) and laminated aggregates (LA) (Figure 9, X 50,100), oriented
perpendicularly to cell wall (CW ).
\'OL. I
fl,
'.;0. 2 , O c rOllr:l< J 9 74
1-'5 1
The particles were close ly assoc iated \\'ith va ri ous types of cyto­
plasmic inclusio ns, suc h as bundles (Fig. 8), lam inated aggrega tes (Fig .
9), a nd pin wh eel s (F ig. 10). Co mple x inclusio ns co nsistin gofpinwhee l
arms, bund les, a nd/or la mina ted aggrega tes were also seen. The
morph ologica l profiles of these incl usions strikingly resem ble d those
obse rved by others in dis eased plan ts in fecLed with ce r tain types of
el o ngated viruses (2,6 ,8, 10, 14, 15, 20-2 4 ,26,28-32 , 34) .
Single particles , as we ll as inclu sion s, ofte n a ppeared in the
cytop lasm close to the p lasmodesma ta betw ee n p arenchyma ce ll s
(F ig. 11 ), ind icatin g th at they ma y move from ce ll to ce ll th rough the
plasmodesma ta as po in ted o ut rece ntl y by We intraubet al (32) . O ccas­
ionall y the viruslike p articl es were ali gned a long the laminated aggre­
gates (F ig. 12) . At ce rtain angles of sec tioning, very fin e sp ikes were
seen on the surface of the agg regates.
I n so me in stances, the inclusions were obse rved in the p hloe m
par e nch yma ce ll ad jacent to the sieve ele me nts in which MLO were
see n (Fig. 12) . Both MLO and th e in clusion s occasio nall y coex isted
within the same phloem parenchyma cell (Fig. 13).
F igure 10.- Characteristic pinwheel inclusions (PW) in mesophyll
cells of a yellow wilt-diseased plant. Complex inclusions consist of pin­
wheel arms and a laminated aggregate (LA). X 54,600.
.I()L"I{~ .-\l OF TH L .\.
S. S. B. T .
Figure ll.-Elongated viruslike particles (VLP) apparently passing
through the pla smodesmata (PO) of parenchyma cells of a yellow wilt­
diseased plant. CW: Cell wall. X 57,400.
Long fi lamentous jJarticles
Mass es o f lo ng fil a me n to us particles we re frequ e ntl y observed
in th e cytop lasm o f meso ph yll , as well as phloe m pa re nch yma, cells
of bo th ye ll ow wilt-infec ted a nd no rm al-a ppearin g pla nts (Figure 14).
Morph ological feature s, size (ap proxim a tely 1200-1 300 nm in le ngth)
a nd a ppeara nce o f th e particl es in th e host cells we re ve ry simila r to
those o f the pres um ed beet ye ll ows vi r us particles in sugarbeet yellows­
infec ted B eta vulgaris, d esc ribed by Esa u e t al (12) .
T h e fil a mento us particles th at o fte n formed la r ge spindle-sha ped
aggregates are sh own in Fig ures 15 and 16 at high mag nifi ca tion s. The
particles al so appea r in ir regularly alig ned ma sses of va riou s sizes
(fig. 14). I n so me insta nces, the p articles form ed hi ghl y ordered aggre­
gates. A lon gitudi na l section of the ordered aggr egate is show n in
18,
VOL.
~,o.
2,
OCTOBU<
1974
cw
s
@
M,LO
Figure 12.-Portion of phloem t issue from a yello w wilt-infected plant.
Note MLO in the sie ve tube elem ent (S) and elon gated viruslike pa r ticles
(VLP), associated with the laminate d aggre gates (LA ), i n a phloem paren­
chyma cell (PP). C W : Cell wall; SPK: Fine spikes. X 68,2 50.
Figure l3.-Coexistence of m ycoplasmal ike o rga nisms (MLO) and t he
laminated aggregates (LA) in the p h loem p aren chyma cell of a yellow wilt­
infec ted plant. X 4 i ,75l>.
154
JO URNA L OF T H E
A . S. S. B. T.
Figure l4.- Portion of mesophyll cell obtained from a normal-appear­
ing sugarbeet plant. Note a large spindle-shaped aggregate, as well as small
irregular masses, of long filamentous particles (FP). CW: Cell wall.
X 18,750.
VOL. 1H.
'\ 0.
2.
OCTOBER
1974
Figures 15 and 16.-Portion of large spindle aggregate of the long fila­
mentous particles (FP) in the phloem parenchyma cells of a yellow wilt­
diseased plant. Note loosely aligned particles in longitudinal section
(Figure 15, X 49,000) and transverse section (Figure 16, X 49,000 ).
156
J O U H:\AL Of THE
A. S. S. B. T.
in Figure 17 and the transverse section in Figure 18. No enclosin g
membran e was seen arou nd either large or sma ll filamentous masses
(Figures 14, 15, \7, and 18) .
In th e ye ll ow wilt-infected plants, filamentous aggregates were
often observed in phloem parenchyma ce ll s which contained intra­
cytop lasmic MLO (Figure 19). Occasionally the filamentous particles
also were prese nt in host ce ll s, closely associated with t he long cyto­
plasmic protrusions of the MLO (Figure 6) or with sp he rical MLO.
Discussion
Before the ex istence of mycop las malik e organ isms as path ogen ic
agen ts in plants became known, Bennett a nd co-workers (3) me n­
tioned that "the diversity of symptoms of the ye ll ow wilt of sugarbeet
h as led to specu lation as to whet her more than one ca usal agent is
involved." However, they concluded that "results of obser vatio ns and
ex periments strongly support th e co ncept that all the different types
of symptoms observed are the resu lt of th e action of a sin gle agent a nd
that this agent is a virus." They also concluded that the type o f sym p­
toms prod uced was determined largely by environ men tal conditions:
the wilting phase o f the disease appea red und e r co ndition s of high
tempe rature and low humidity, whereas the yell ow in g phase appeared
und e r con d itions of lower temperature and hig her humidity.
T he characteristic sy mpto m expressions of the ye llowing ph ase
described by Be nn e tt et al (3, 4) str ikingl y resemble those of yell ows
ty pe diseases, now kn own to be associa ted with or ca used by MLO
(16-\8, 25, 33). Furth e rmore, successful suppression of the disease
with tetracycl ine trea tments (II) and the present fi ndin gs of MLO in
ye llow wilt-diseased Beta vulgaris plants strongly suppo rt a new co n­
cept that at least symptom exp ressions of the yellowi ng phase are
caused by MLO agents. However, no MLO age nts associated with any
wilting diseases are known to date despite the fact that more than 50
diffe re nt MLO -induced diseases h ave bee n reported. If the MLO a re
indeed the causative age nts of the ye llow in g phase, a comp lex etiology,
most likely MLO and a virus (or viruses), of the suga rbeet yellow wilt
is more acce ptable than the sin gle viral etio logy (3) .
There ex ists a controve rsy as to whether the characteristic in­
clusions, such as pinwheels, bundles, and la min ated aggregates, con­
sist of certain elongated vi r us particles (6 , \ 5, 24, 26, 34) or whether
they are comp lex or ga ni zations of virus particles and/or some sub­
stances of unktlown nature (2, 10,21) . N evertheless , base d on the
in timate association b<."twe en the viruses an d the characteristic in­
dus ions , Edwa rd son (9) proposed that the inclusion s are diagnostic
for infectio n by vir us("s of th e potato virus Y group (5), whose lengths
v:1ry fro m 730 l1r1l to 750 nm. Later similar inclusions in d uced by
shorter (3 1), as well as longer (14), viruses were also reported . It is
VOL.
IS, N o .2 ,
OCT OBER
1974
157
Figures 17 and 18.-- Highly ordered aggregates of the long filam e ntous
particles (FP) in the mesophyll cells obtained from a yellow wilt-infected
plant. Note highly aligned particles in longit udinal section (Figure 17,
X 49,100 ) and transverse section (Figure 18, X 49 , 100),
158
JOURNAL OF THE
A. S. S. B. T.
Figure 19.-Coexistence of the long filamentous particles (FP) and
mycoplasma-like organisms (MLO) in a phloem parenchyma cell obtained
from a yellow wilt-infected plant. MT: Microtubules. X 32,630.
generally accepted that the presence of the ch aracteristic inclusions
indicates the occurrence of infection with certain elon gated viruses.
Thus, the finding of the elongated virus!i ke particles with the charac­
teristic inclusions suggests that the diseased sugarbeet plants examined
may be infected with an inclu sion-inducing viru s. The inclusion­
inducin g elongated viruses are kn own to be ca usative agents of a
number of plant diseases (2, 6, 8-10, 14, 15,19,20,22-24,26,28-31,
34). Although no elongated viruses whi ch induce wi ltin g h ave been
reported, one of the potato virus Y gro up is known to be associated
with the sugarbeet wilting in Brazil (persona l communication: A. S.
Costa, Virus Section, The Agricultural In stitute, C.P. 28,13100 Cam­
pinas, Sao Pau lo). At present, it is un certa in whether elon gated virus­
like particl es arC' responsible in part for ca using symptoms of the
sugarbeet yellow wilt d isease (e.g. in the wilting phase). Fu rther elec­
tron microscopic exam in ation of diseased plants representin g the two
distinct types (i. e. t he yellowing phase and the wiltin g phase) may
provid e more critical information on this problem. Recently the co­
existence of MLO a nd an elongated virus . russet crack virus, asso­
ciated with pinwheel incl usions in sweet potato plants was reported
(20).
VOL. J 8,
:--Jo. 2, OCTOBER 1974
159
The nature of the long filamentous particles observed in both
yellow wilt-infected and appaiently noninfected sugarbeet plants
remains unknown. However, since those particles resembled the beet
yellows virus (12), a nd since one 'or more types of virus yellows usually
is quite prevalent in the sugarbeet crop in Chile, including La Platina
(personal communication: J. O. Gaskill, Beet Sugar Development
Foundation, Fort Collins, Colo.), it seems probable that the long fila­
mentous particles observed in this study resulted from infection by at
least one of the viruses of the sugarbeet virus yellows complex. Since
the effects apparently were mild, as evidenced by the appearance of
plants without yellow wilt symptoms, it is assumed that a mild type of
virus was involved , such as a mild strain of beet yellows virus.
If the above assumption of a yellows-virus involvement is correct
the findings indicate that the yellow wilt-diseased plants examined
were super infected with the yellows virus. The role, if any, of that
virus in the development of symptoms of yellow wilt is not known.
However, it is apparent that controlled conditions of plant growth and
inoculation are needed in future electron microscopy studies of the
causal agent or agents of yellow wilt.
Summary
An electron microscopic search for the causative agents of the
sugarbeet yellow wilt disease in Chile was carried out. Mycoplasmalike
organisms (M LO), as well as elongated viruslike particles associated
with characteristic inclusions (such as pinwheels, bundles, and
laminated aggregates), were observed in the diseased plants, but were
absent from normal-appearing plants. The presence of the MLO
strongly suggests that they are largel y responsible for causing the
yellowing phase of the disease.
The presence of the elongated viruslike particles associated with
characteristic inclusions suggests that the diseased plants were infected
with a virus. Although the elongated particles may be responsible in
part for yellow wilt symptom expression, further studies are needed
to clarify their etiologic role.
These findings suggest the possibility of a complex disease
etiology, presumably with the MLO and a viral agent rather than the
single MLO or viral etiology.
Long filamentous particles, morphologically resembling the beet
yellows virus, were observed in both the yellow wilt diseased and the
normal-appearing plants grown in the same field . This suggests that
the yellow wilt-diseased sugarbeet plants may also have been infected
with a type of beet yellows virus.
JO URN AL
160
OF
THE A. S. S. B. 1'.
Acknowledgmen ts
T he au thors th a nk J. O . Gas kill , Project Consul ta nt, Bee t Sugar
Deve lop me n t Fo und ation , Fo r t Collins , Co lorado, for his valu able
suggestio ns a nd cri tical revicw o f the ma nu sc ri pt. The autho rs also
thank G. L. McNew , Ma naging Directo r, Boyce T h o mp son Insti tute
fo r Plan t Rese arch, Yo nke rs, N.Y., an d the L.S. Age ncy for Interna­
tion al De vel o pm e nt for the ir p articipatio n in ma ki ng the arran ge­
men ts fo r th e se nio r au thor's vi sit to Bo yce Th o mpson In stitute to
ca r ry out this re sea rch.
Tha n ks also to Kazu ko Hirumi of Boyce T ho mp so n Institute for
her valu a bl e ass istance in e lectro n mi cro sc op y, a nd to Rob e rto
E hre nfel d K. of Indu stria Azucarera l\iac io nal S.A., Santiago, Chile ,
fo r co nd uning the field work at La Platina .
Literature Cited
(I) (2 )
(3)
(4) (5) (6) (7 ) (8) (9) (1 0)
(II )
A ~ !)F R SO:-l ,
DOUGLA S, R. 1969. Ul tras tru ctu ral stu dies of m yco plasmas
a n d th e L- p ha se o f ba cte ri a. I n : Lf'ona rd Hay Ric k (ed .), T h f' \ll yco­
plas ma ta le s a nd the L-phase of bacteria, 3 65-402. A pp leto n- Ce nt ur y­
Cro ft s. '\J ew Yo rk .
AR~ o IT , HO\\', \R ll j. a nd KE :-I:-IETH :vi. St-ll TH. 1967 . El ect ro n micro­
sco p y of virus -i n fe cred su n Rower leaves. j. U ltras tr uct. Res. 19 : 173 ­
195 .
BF:\:\r n , C. W ., F. j. HILI S, R. EH Rt.WE LJ) K., J. VALE:-I ZUFLA B., and
C. KLEI ~ K. 196 7 . Ye llow \\il l o f suga r beet. J. A m . Soc. S ugar Bee t
T ec hn o !. 14 :4HO-510.
B t :\:\ETT, C. v\! . a nd CA RLOS M U:\'CK. 1946. Ye llow wil t o f su g ar beet in
\rge nti na . J. Agr. Res . 73: 45-6 4.
BR A~J) E S, J and R . B ERC KS. 1965. Gross morph o logy a nd se r o logy as a
bas is for cl ass ifica tio n of e lon g ated pla llt \' iruse s. A.d\'. Vir us Res .
I 1: 1-24.
C REME R, MAR CARFTHA C. a nd J A.. VA:-.i DlR VEK L'I. 196 4. Lo ca li za t io n
o f vira l a nti ge n in n a rcissus leaves infec te d \\'ith yell o w str ip t' virus ,
dete rmi ne d by m ea n s o f a flu ore scei n co nju ga te d a nti ser um . !'Ie th . J.
Pl a n t Pat hol. 72 :1 05-11 3 .
0 01 , YOJ l , MI CH I \ KI T ERANAKA, K lYOS lI 1 Y ORA, a nd HI DEfC:CII
ASUY AMA. 1967. M yco pl as m a - or PLT gro up-l i ke m inoo r g;] nis m s
fou n d in t he p hl oe m e le m e nts o f p la nts in fected with mulbe rry dwa r f ,
pota to witches' b roo m, a ste r ye llow s, or p aulow n ia witch e s' broo m .
Ann. Ph ), topa th . Soc. j a p a n 33:259-2 66.
ED WAR J)S O:-l , J OH" R. 19 66. Cylindri ca l inclu sion s i n t11f' cytoplas m of
lea f ce lls infected w ith to bacco <:tch viru s . Scie n ce l 53:883-884 .
ED WARDS O:-l , J OHN R. 19 66 . E lectron mi cro sco p y of cyto plas mi c inclu ­
sio n s in ce ll s in fected wit h ro d- sh a p ed viruse s. A m . J. Bo t. 53:359-364.
ED WA RIlS O:-l , j. R. , n. E . PGRCIF ULL, a nd R . G . CH RI STlE. 19 6 8 . Structure
o f cy to pl as mic incl u sions in pla nts i nfected \\'ith ro d- shape d vir uses.
V iro lo g )'34: 25 0-2 63 .
EH RE'IFELD , ROBE RTO K . 1970 . Estu di o pre limi n ar d el efe cto cl e
c1o r ote trac icli na (a ure om icina) so bre e l ca u sa l etio l6gico cle la " m ar­
c hit ez virosa " (yell o w wilt) d e la rem ol ac ha (B eta vu lga ris L.). Agr.
Tec ni ca 30: 4 3 -5 1.
\ ·O L. 18, No.2, OCTOBER 1974
(12)
(13)
(14)
(15)
(16)
( 17)
(18)
(1 9)
(20)
(2 1)
(22)
(23)
(2 4)
(25)
(26)
(27)
(28)
161
ESAu, K. , J. CRONSHAW, and L. L. HOEFERT. 1966. Organization of beet
ye ll ows-virus inclusions in leaf ce lls of B eta . Prot:. Nat!' Acad. Sci.
55 :486-493 .
FREU NDT, E. A . 1969. Ce llul ar morphology and mod e of replication of
thc mycoplasmas. I n: Lco nard Ha yRick (ed.), Th e tvl yco plasmatal es
and th e L-ph as e of bacteria, 28 1-315. Appleton -Cen tur y-Crofts,
Nell" York.
H ARR ISON , B . D . a nd I. ~1. ROIlFRTS. 1971. Pinwh ee ls a nd c r ysta line
stru ctures induced by A trupa mild mosaic viru s, a plant vi ru s with
particles 925 nm. long. J. Ge n . Virol. 10:71-78.
HAY ASH I, T, C. iV[ATSUI , and .\. YAMA GUC HI. 1965 . Elect ron micro­
scopy of intracell ular turnip mosaic virus. Phytopatho lo gy 55:458-1 6 1.
H IRl',fJ , H. ancl K. ivIA RAYlO ROS CH. 1972. :\atural degeneration of
mycoplasmalike bodies in an aster yellow s in fectecl host plant. Phy to­
path. Z. 70:9-26.
HIRUMI , H. and K. MARAMOROSCH. 1973. In tracytop lasmic myco­
plasmalike bodies in phloem parenchyma ce lls of as te r ye llows- infec­
ted l\"icotio.na rustica. Ph )/topal h . Z. 77:7 J- 83.
HI RUMI, H. and K. :VI ARAYIOROSC H. 1973. U lt rast ru ct ure of the aster
yellows agent: Mycoplasma-like bodies in sieve tu be elements of
Nicatiano. ruslica. Ann. N e w York Acad. Sci. 225:201-22 2.
I SH lI E. T-\TSl"jl, Y0.1 1 DOl, KIYOSHI YOR A, and HIDEFUMI ASUYAMA.
1967. Suppressiv e effects of antibioti cs of tetracycline group on
sym pwm clcvelo pment of mulberry dwarf disease. Ann. Phytopath.
Soc. Japan 33:267 -275.
KAHN , ROBERT P ., R. H . LAWSO"l, R. L. MO NROE, an d SUZANNE H EARON.
1972. Sweet potato litt le- lea f (w itc h es' -broom) as sociated with a myco­
plasma-like organ ism. Ph yto pathology 62:903-909.
KAi'l£l, T, Y. H ON!)A , and C. MATSUI. 1969. Intrace llular appearance
of turnip m osa ic a nd bea n yellow mosa ic vir us particl es. Ph yto ­
pathology 59: I 39-14 4.
KI:VI, K\TNG SOO and J. P. FULTON. 1969 . E lect r o n microscopy of poke­
weed leaf cells infected with pokew eed mosa ic virus. Virology 37 :297­
308.
LA~G E :-JBERG, \-VI LLDI G. and H ELE" F. SCHROED ER. 1973. Electron
microsco p y of un stable inclusions induced in m a ize by maize dwarf
mosa ic viru s. Ph ytopathology 63: 1066- 1073.
.
L lE, PITER E. 1965. E lect ro n microscopy of inclusions assoc iated with
w heat stre ak mosaic vir us. J. U Itrastruct. Res. 13: 359-366.
MARA\[OROSC H, KARL, ROBERT R. CRA:-JADOS, an d HIROYL KI HI RU:l-IJ.
1970. Mvcoplas m a diseases of pl a nts and insects. Adv. Virus Res.
16:135-193.
~IATSUI, CHIAKI and AK IRA YA'·[AGUCHI. 1964. Electron microscopy of
host cells infected with tobacco etch virus. I. Fi ne structures of lea f ce ll s
at la ter stages of infection . Virology 22:40-47.
MOLU::-':HAl' ER , HI LIO;\; H . 1964. Plastic e mbe dding mi x ture s for li se
in electron microsco p )' . Stain Techno!. 39: 11/-114 .
Y[CRA:'\T , A. F. an d I. :\I/. ROH£RTS. 197 J. Cylindrical inclu sio ns in
coriand e r leaf ce ll s infected with pars nip mosaic viru s. J. Gen. Virol.
10:65-70.
162
J OU RNAL OF THE A. S. S. B. T.
(29) PURCIF ULL, D. E. andJ. R. EDW ARDSON. 1967. Watermelon mosaic virus:
Tub ul ar inclusion s in p umpkin leaves and aggregates in leaf extracts.
Virology 32 :393-401.
(30) SHEPARD, J. F. 1968. E lec tron microscopy of subtilisin-treated toba cco
etc h virus nuclear and cytop las mic inclu sions. Virology 36:20-29.
(3 1) WFI :-ITR :\ UB , \If. a nd H . W.J. RACETLI. 1970. Distribution of virus like
pa rticl es in leaf cell s of Dianthus barbatus infec ted with carnation vein
mottlt' virus. Virolo gy 40:8("iil-881.
(3 2) WUNTRA UB, M., H . W. J. RA GLTU, and ESTHER L o . 1974 . Po tato v irus
Y particles in plasmodes mata of tobacco leaf ce ll s. J. U lt r as tru ct. Res.
46 131- 148.
(33) WIIITcmIB, R. F. a n d R. E. DAVIS. 1970. MycojJ/asm.a a nd ph ytar bo­
virus es as p la nt pathogens persis tently tra n s mitted by insects. Ann .
Rev. Entomol. 15:403-464.
(34) YAMA(;CCHI, AK IR A, TOSH IO KIKL"\IOTO, an d CHIAKI MATSL:1. 1963.
E lect ron mic roscopy of elon gate d pa rt icles asso ciate d with tu lip
mosa ic. Viro logy 20: 143- 146.