T E L I O S P O R E G E R M I N A T I O N A N D C O M P A T I B I L I T Y F A C T O R S I N S O R O S P O R I U M T E X A N U M 1)

by

R U B E N DURAN 2)

(with 2 figs.)

(18.Xli.I967)

Sorosporium texanum was described by ZUNDEL almost 25 years ago (6) from a single known collection made by Mr. HANSEL in Brownsville, Texas on Pennise tum nervosum (NEES) TRIN. (Coll. No. 52794, National Fungus Collection, through communication with J. A. STEVENSON). ZUNDEL apparently never collected this species during his career nor is it known to have been collected or studied b y others since he described K in 1944. That the smut may be rare is suggested by the fact that up until now, S. texanum has been known only from the type specimen on P. nervosum (4,6). In 1966 the smut was collected in the Santa Ri ta Mtns. of Arizona on natal grass, Rhynchety trum roseum (NEEs) STAPF & HUBB., which has not previously been reported as a host for this or any other smut in North America (4). Sorus characters, teliospore morphology, and spore balls of the natal grass smut proved identical to those of the type specimen, thus extending the host range of the fungus to two species in two genera of the tribe Paniceae. Moreover, the collection in Arizona constitutes a new state record for the smut.

Teliospore germination as well as determining the number of alleles which govern compatibility, both reported here for S. texa- hum, are objectives of a continuing s tudy by the author on the smut fungi of North America.

MATERIALS AND METHODS

G e r m i n a t i o n te s t s

Teliospores from the natal grass collection were germinated at room temperature on potato-dextrose agar (PDA) and distilled

x) Scientific Paper No. 3073. Washington Agricultural Experiment Stations, Pull- man, Project 1729. This investigation supported in part by a grant for scientific research from the National Science Foundation.

3) Associate Professor and Associate Plant Pathologist, Washington State Uni- versity, Pullman.

210 R. DU~AN

water. Teliospores in various stages of germinatiort were stained with phloxine to increase cytoplasmic contrast and photographed. Mononucleate condition of sporidia and dikaryotically-paired nu- clei in infection hyphae were demonstrated by the acetic-orcein schedule.

Compatibility tests Three sets of 4 sporidia were isolated by micromanipulation from

three different four-celled promycelia. Each set was isolated from a promycelium bearing only 4 sporidia, one per promycelial cell. Single sporidia were incubated for 36--72 hr at room temperature on hang- ing agar Mocks. Thereafter, the developing sporidial colonies were transferred to potato-malt agar slants (PMA) and kept at 15 ° C to establish cultures for compatibility experiments. Usually 2--3 weeks were required for the colonies to produce sufficient secondary spori- dia for the compatibility experiments. Compatibility of monospori- dial cultures was determined by the Bauct~ test (1) and by observing fusions between sporidia of opposite compatibility with the micro- scope. The 12 sporidial cultures were mated on PDA in all possible combinations for a total of 66 pairings plus 12 unmated controls.

When possible, the position of each sporidium on the promyce- tium was recorded by numbering them in ascending order, begin- ning with the apical cell of the promycelium. Cultures derived from single sporidia were designated accordingly. When arrangement of sporidia on the promycelium could not be determined, cultures were designated A, B, C, D.

]~ESULTS

Germination studies On nutrient media, germination in S. texanum is generally charac-

terized by production of 4-celled promycelia, each cell producing numerous sporidia, or occasionally by promycelia in which the cells fuse directly. Sporidial production and fusion between promycelial cells was observed at times on the same promycelium. When fusions occurred between sporidia or promycelial cells, typical infection hyphae developed within 12--24 hours.

The Ustilago type germination in S. texanum, though somewhat variable, is seemingly no more so than that of other smut fungi re- cently studied (3). The classical 4-celled promycelium is mostly produced on nutrient media, whereas in water, spores tend to germi- nate directly producing typical knee-joints and relatively few spo- ridia. The overall variation is shown in Fig. 1.

Compatibility studies When fresh sporidial cultures of opposite compatibility were

paired, a typical Bauch-type reaction resulted within 12-16 hr. Compatible pairings were identified macroscopically by the cottony

GERMINATION AND COMPATIBILITY OF S, TEXANUM 211

Fig. I. Teliospore germination in Sorosporium texanum on PDA, A--B. Indirect germination. (A) 4-celled promycelium with 1 set of sporidia prior to production of secondary sporidia (basM cell of promycelium obscured by the teliospore). (B) Secondary sporidia budding from the apex and sides of the promycelium (C) Direct

germination; note fusion of apical and subapical cells. × 1350.

aerial mycelium which colonies developed on PDA, a reaction simi- lar to that reported in some races of UstiIago avenae (PERs,) ROSTR. (5). Also, colonies in which compatible sporidia were paired, devel- oped a thin peripheral band of decumbent and dikaryotic infection hyphae. No significant differences in colony size resulted from mat-

2 1 2 R. DURAN

Fig. 2. Sorosporium texanum. Compatibi l i ty between two sets of monosporidial cul- tures, one set (1, 2, 3, and 4) f rom one promycel inm, the second (A, ]3, C, and D)

f rom another . Compatible pair ings indicated by colonies wi th white mycelium.

TABLE I

Sorosporium $exanum. Matings on PDA between ~hree sets of four monosporidial cul- tures, each from different promycelia,

Set 7#1 Set ~2 Set ~3

+I +2 --3 --4 +A +13 --C --D +I --2 +3 --4

-}-1 C I F F I I F F I F I F + 2 C F F I I F F I F I F - - 3 C I F F I I F I F I - - I C F F I I F I F I + A C I F F I F I F + B C F F I F I F --C C I F I F I - - D C F I F I + 1 C F I F - - 2 C F I + 3 C F - - 4 C

C = checks; F = fusions; I = incompat ible pairings.

GERMINATION AND COMPATIBILITY OF S. TEXANUM ~ 1 3

ing compatible or incompatible masses of sporidia. The reaction is shown in Fig. 2.

The results of pairing the 12 monosporidial cultures in all possible ways is summarized in Table I. The pat tern clearly indicates that each set of sporidia is self- as well as cross-fertile with tile others. On the basis of these results, it is hypothesized that compatibility in S. texanum is bipolar. The two sets of sporidia identified as to posi- tion on the promycelia indicate that reduction division of compati- bility alleles may occur in the first or second division of meiosis. Fusions between promycelial cells were always observed to occur between the two basal cells and between the apical cells, suggesting that reduction division occurs frequently during the second division.

S u m m a r y and d i scuss ion

Germination in S. texanum is of the Ustilago type, the teliospores being easily and highly germinable on nutrient agar and water. Sporidia and clamp connections (knee-joints) may be produced on similar or different promycelia, a phenomenon common in other smut fungi (3).

Mating studies indicate that compatibility in S. texanum is con- trolled by one pair of alleles. The BAUCH reaction obtained by mat- ing sporidia of opposite compatibility proved similar to that of Ustilago avenae, but different from that reported recently for Ustilago hilariicola G. W. Flscm (2). In the latter species, signifi- cantly larger colonies developed in mating compatible sporidia than when incompatible sporidia were paired. The larger colonies representing compatible matings were readily identified by wide bands of marginal infection hyphae which were absent in colonies resulting from incompatible pairings. While no significant differ- ences in colony size were noted in S. texanum, even though a thin peripheral band of infection hyphae were present, production of white aerial mycelium occurred only in compatible combinations. I t is interesting that, while the BAUCH test is very useful in studying compatibility relationships in the smut fungi, the test may produce variable reactions ill different species of smut fungi.

References

1. BAU¢~I, R. 1932. Die SexualitStt von Ustilago scorzonereae und Uslilago zea. Phytopa th . Z. 5: 315--321.

2. DURAN, R. & SAFEEULLA, K. ~/L 1965. Teliospore germinat ion and genetic factors controlling compatibi l i ty in Ustilago hilariicola. Mycologia 57: 628--633.

3. DURAN, R. & SAFEEULLA, K. M. 1968. Aspects of telicspore germination in some Nor th American s m u t fungi I. Mycologia. 60: 231--243.

4. FISCHER, G. W. 1953. Manual of the Nor th American s m u t fungi. The Ronald Press Co., New York. 343p.

5. FISCHER, O. \V. & HOLTON, C. S. 1957. ]Biology and control of the s m u t fungi. The Ronald Press Co., New York. 622p.

6. ZUNDEL, CT. L. 1944. Notes on the Ustilaginales of the world. Mycologia 36: 400--412.