Caryologia International Journal of Cytology, Cytosystematics and Cytogenetics ISSN: 0008-7114 (Print) 2165-5391 (Online) Journal homepage: https://www.tandfonline.com/loi/tcar20 Cytology and Teliospore Development of Entyloma Thirumalachari R.A. Singh & M.S. Pavgi To cite this article: R.A. Singh & M.S. Pavgi (1975) Cytology and Teliospore Development of EntylomaThirumalachari, Caryologia, 28:1, 15-28, DOI: 10.1080/00087114.1975.10796592 To link to this article: https://doi.org/10.1080/00087114.1975.10796592 Published online: 30 Jan 2014. Submit your article to this journal Article views: 73 Citing articles: 2 View citing articles Full Terms & Conditions of access and use can be found at https://www.tandfonline.com/action/journalinformation?journalcode=tcar20
CYTOLOGY AND TELIOSPORE DEVELOPMENT OF ENTYLOMA THIRUMALACHARI R.A. SINGH and M.S. PAVGI Faculty of Agriculture, Banaras Hindu University, Varanasi, India INTRODUCTION Received: 19 1 11 July 1973 Entyloma thirumalachari PA VGI and SINGH, paraslttc on the composite Blumea oxydonta DC. produces creamy-yellow, circular to irregular, galllike raised sori on both leaf surfaces with leaf crinkles. The sori are nonerumpent containing dense spore masses aggregated in the mesophyllar interspaces. The teliospores are globose to globoid, light brown, smooth, thickwalled and ensheathed in a thick hyaline firm layer (PAVGI and SINGH 1967). Cytology of relatively few Entyloma species viz. Entyloma calendulae (Oud.) DeBary, E. dahliae Sydow, E. eleocharidis PAVGI and SINGH, E. microsporum (Unger) Schroter, E. nj'mphaeae (Cunn.) Setchell and E. scirpicola Thirumalachar and Dickson has been studied (DAs 1949; FISCHER and HoLTON 1957; KAISER 1936; PAVGI and SINGH 1970; THIRUMALACHAR and DICKSON 1949). The development of parasitic mycelium and teliospores in the hosts of Entyloma species has been studied by only few workers (DAs 1949; KAISER 1936; LUTMAN 1910; PAVGI and SINGH 1970; RACIBORSKI 1897; THIRUMA.LACHAR and DICKSON 1949). The teliospores of E. thirumalachari germinate readily, without indication of dormancy, at room temperature (26-30 (), providing excellent material for studying the cytological development in detail. MATERIALS AND METHODS Mature teliospores from air-dried sori on the leaves of Blumea oxydonta DC. were germinated on slide mounts at 26-30 C by the method described by THIRU MALACHAR et al. (1950, 1953). The germinating teliospores were killed and fixed in situ at different stages of development in modified Carnoy's fluid (95% ethyl alcohol 30 ml, chloroform 10 ml, glacial acetic acid 10 ml), downgraded to water through an alcohol series, treated in Taka Diastase ( 1 tablet Taka Diastase, Parke [ Caryologia, Vol. 28, n. 1, 197.5
16 SINGH and PAVGI Davis (India) Ltd., Bombay in 50 ml dist. water) soln., washed in tap water and mordanted in 0.5% ferric ammonium sulphate 'AnalaR' (iron alum) soln. (10 min.), washed in tap water (5 min.), rinsed in dist. water, upgraded to 70% alcohol through an alcohol series and stained in Belling's 0.5% aceto-carmin (JoHANSEN 1940). Germinating teliospores in another set were killed and fixed in Randolph's modified Navashin soln. (12-14 hrs) and stained with Heidenhain's hematoxylin for complementary observations (JoHANSEN 1940). Developmental cytology of the fungus in host tissues was studied by killing and fixing sori on infected leaves of Blumea oxydonta DC. at progressive stages of development in Randolph's modified Navashin soln. (24 hrs) and embedded in paraffin (JoHANSEN 1940). Serial sections were microtomed (8-10 IJ.), stained with Heidenhain's iron hematoxylin and counterstained with orange G. OBSERVATIONS A) Teliospore germination and meiotic nuclear division. A mature teliospore contains a deeply stainable diploid fusion nucleus measuring 3-3.51J. in diam. (Fig. 1 ). During germination, the teliospore imbibes water, exospore swells, becomes turgid and the protoplasm becomes densely granular. The endospore enlarges and pushes out through a germ pore and sheath into a stout promycelium (Fig. 2). The teliospores of the other Entyloma species, except E. calendulae, studied cytologically have germinated by rupturing the exospore. Synchronously the fusion nucleus gradually enlarges, moves toward the germ pore and squeezes out along with the cytoplasm into the promycelium. The nucleus becomes oval-elongate, while passing through the narrow germ pore becoming spherical again in the promycelium and usually moves towards the center (Figs. 3, 4, 7); but nuclei were also commonly found located near the tip or base of the promycelium entering the interphase. The diploid fusion nucleus continues to enlarge Plate I. Meiotic nuclear division and nuclear behaviour during teliospore germination and sporidia! development in Entyloma thirumalachari. Fig. 1. - Teliospore with a diploid fusion nucleus. - Figs. 2, 3. - Teliospore germination and nuclear migration into the promycelium. - Fig. 4. - Diploid nucleus in interphase. - Fig. 5. - Diplotene stage of prophase. - Figs. 6, 7. - Stages in metaphase I showing chromosome migration towards the spindle plate. - Fig. 8. - Precocious movement of chromosomes on the equatorial plate. - Fig. 9. - Early anaphase I. - Fig. 10. - Anaphase I. - Fig. 11. - Two daughter nuclei. - Fig. 12. - Metaphase II. - Figs. 13-16. - Differentiation of terminal sporidia and migration of a haploid nucleus in each. - Figs. 17-19. - Conjugation between compatible sporidia in situ and formation of dikaryotic secondary sporidia. - Figs. 20, 21. - Secondary sporidia with and without lateral branches. - Figs. 22-24. - Formation of infection hyphae, nuclear multiplication and their migration. - Figs. 25, 26. - Haploid, uninucleate and dikaryotic sporidia budding in artificial culture. (Scale A: Figs. 1 to 12; B: Figs. 13 to 26).
CYTOLOGY AND TELIOSPORE DEVELOPMENT 17
18 SINGH and PAVGI (4-4.51J.), merges into early prophase and the nucleolus becomes more conspicuous in the chromatin reticulum surrounded by a well-defined nuclear membrane (Fig. 4 ). A typical leptotene or any stage was not discerned in any of the preparations as possibly it had merged with late interphase in the continuum. Zygotene and pachytene are of very short duration and are only partially distinguished in the sequence. The chromosomes gradually shorten, becoming more prominent showing intertwining loops or chiasmata occasionally or along most of their length. Nucleolus remains faintly stainable but the nuclear wall disappears (Fig. 5). The 6 short bivalents become darkstained and irregularly oriented in different planes. The nucleolus disappears as the chromosomes elongate slightly and gradually move toward the equatorial region (early metaphase) to become arranged on the equatorial plate representing metaphase in the sequence. The metaphase spindle is usually oriented parallel to the long axis of the promycelium (Figs. 6, 7 ). The chromosomes are rarely seen along the spindle due to precocious. movement from the equatorial plate (Fig. 8). The chromatids of each bivalent chromosome split and moved to the poles representing early anaphase (Fig. 9), and anaphase (Fig. 10). The movement of chromosomes was often asynchronous as in the higher plants. The chromosomes of each daughter nucleus aggregate into a dense mass as they lose their identity (telophase). The 2 daughter nuclei remain close to each other as a deep staining network of chromosomes (Fig. 11) and undergo subsequent division without any intervening rest period. Interphase of the 2 nuclei probably coincides with the early prophase II. The chromosomes again become distinct and start thickening indicating late prophase or early metaphase II. The chromosomes became irregularly arranged over the equatorial region and the 2 spindles appear similarly oriented. The chromatids from each chromosome move towards their respective poles (Fig. 12) and become aggregated into individual daughter nuclei (telophase II). The 4 haploid nuclei gradually move towards the promycelial tip (Fig. 13 ). B) Development and cytology of sporidia. Synchronously with the nuclear divisions in the promycelium, 4 to 8 small papillate projections differentiate at the tip of the promycelium, which gradually develop into a whorl of sporidia (Figs. 13-16). The 4 haploid nuclei undergo a further mitotic division as necessary. One nucleus moves from the promycelium into each of the fully developed sporidia to become located approximately in the middle (Figs. 15, 16 ). Sometimes 2 nuclei pass into a few of the sporidia in the whorl, while the remaining sporidia remain uninucleate. Formation of a septum at the base of a sporidium after nuclear migration has been reported in some species (DAs 1949; THIRUMALACHAR
CYTOLOGY AND TELIOSPORE DEVELOPMENT 19 and DICKSON 1949), but it was not observed in E. thirumalachari. Occasionally nuclear migration into the sporidia is delayed until after mitotic division of the nuclei in the promycelium increasing the number to 8. One nucleus migrates into each sporidium (Fig. 16) and the supernumerary nuclei remain in the promycelium (Fig. 17). Such multinucleate apical cells beneath the whorl of sporidia have been described in some species of Doassansia Cornu, Entyloma De Bary and Tuburcinia Fries in the Tilletiaceae (THIRU MALACHAR and DICKSON 1949). A mature sporidium sends out a narrow fusion tube generally from the tip, occasionally from the middle, which arches over to approach a compatible sporidium dissolving the walls at the point of contact to form a fusion bridge (Fig. 17 ). The nucleus along with the cytoplasm from the approaching sporidium migrates into the other (Fig. 18 ). Sometimes 2 compatible sporidia send out conjugation tubes which are attracted towards each other and fuse into a bridge. The nucleus from each sporidium migrates into the bridge. Fusion of 2 primary sporidia in situ through a conjugation tube has been reported in all Entyloma species studied. PARA VICINI ( 1917) reported fusion of detached sporidia in E. calendulae, which was not confirmed by KAISER (1936). One to 3 secondary sporidia develop directly from the fusion bridge or are borne on small hyphae produced on the bridge (Fig. 19 ). Morphologically 2 types of secondary sporidia are frequently observed; one an acicular type with a pointed tip and truncate base (Fig. 21) and the other morphologically similar to the first, but with 1 to 2 lateral branches (Fig. 20). The Entyloma species have been reported to produce fusoid to acicular secondary sporidia except in E. magnusii (Ule) Woronin, which produces secondary sporidia with lateral branches like those in E. thirumalachari (FISCHER and HoLTON 1957). The 2 compatible nuclei associated in the fusion bridge enter into the secondary sporidium (Fig. 19) or divide conjugately forming 2 to several pairs of dikaryons, which enter several secondary sporidia as paired nuclei. When only a single secondary sporidium is produced with a dikaryon entering into it, the other pair(s) remains in the primary sporidium awaiting successive formation of secondary sporidia. Mature secondary sporidia are detached, intermingled and netted with lateral processes. They are dikaryotic, the 2 nuclei remaining a short distance away from each other (Fig. 21 ). Occasionally secondary sporidia with 3 nuclei were observed. Binucleate secondary sporidia are regularly cut off after fusion in Entyloma species, but development of uninucleate secondary sporidia at the apex of conjugating sporidia was noted in E. scirpicola (THIRUMALACHAR and DICKSON 1949). Odd numbers of sporidia in a whorl which do not fuse, cut off uninucleate secondary sporidia in E. dahliae, E. calendulae and E. scirpicola (JosHI 1960;
20 SINGH and PAVGI KAISER 1936; THIRUMALACHAR and DICKSON 1949), but were not observed in E. thirumalachari. Even an unfused sporidium gives rise to a dikaryotic secondary sporidium or directly to an infection hypha, which later is furnished with several pairs of nuclei. This phenomenon was also observed by KAISER (19 36) in E. calendulae. Production of dikaryotic secondary sporidia from an unconjugated sporidium may be due to migration of an additional compatible nucleus from the promycelium, which was also illustrated for E. calendulae (KAISER 1936). Haploid primary sporidia detach and germinate occasionally in E. microsporum (DAs 1949), but the phenomenon has not been observed in the present fungus. Terminal budding of haploid dikaryotic, secondary sporidia also occurs in artificial culture after fusion and consummation between 2 compatible sporidia in the latter case (SINGH 1968; SINGH and PAVGI 1973) (Figs. 25, 26). The 2 nuclei in a secondary sporidium divide conjugately to form 4 nuclei (Fig. 20 ). Nuclear division in and elongation of a secot:tdary sporidium from either end and/or from the tip of a lateral branch occur simultaneously. Normally a pair of nuclei proceeds towards each elevating tip of the sporidium (Figs. 22, 23 ), but unidirectional migration of all the dikaryotic pairs is not uncommon (Fig. 24 ). The acute tips of terminal and lateral branches become blunt (obtuse) at the time of elongation (Figs. 23, 24). The hyphal branches become irregularly wavy in thickness after short growth intervals (Fig. 23 ). The infection hypha remains aseptate containing several pairs of nuclei by successive conjugate divisions of the dikaryons, which may infect a susceptible host under suitable environmental conditions later. C) Aberrations in nuclear behaviour. Anomalous but interesting nuclear behaviour (movement) was observed during teliospore germination. Differentiation and development of sporidia relative to nuclear condition may be broadly grouped into 4 patterns: Plate II. Aberrations in teliospore germination, sporidia) development and nuclear behaviour in Entyloma thirumalachari. Fig. 27. - Two sporidia remaining anucleate. - Figs. 28, 29. - Few sporidia remaining anucleate due to migration of more than one nucleus into some and subsequent formation of infection hyphae. - Figs. 31, 32. - Bifurcation of promycelium bearing sporidia on either or both branches. - Figs. 30, 33-37. - Suppression of sporidia) fusion and formation of secondary sporidia or infection hyphae directly with diviations in nuclear behaviour. - Fig. 38. - Bifurcation of promycelium, increasing the sporidia) number; 2 infection hyphae are produced from one fusion bridge. - Fig. 39. - Conjugation occurring in presence of 2 or more nuclei in each sporidium. - Fig. 40. - Nuclei dividing several times in promycelium and migrating irregularly leaving one sporidium anucleate. - Fig. 41. - Two conjugation bridges between 3 sporidia.
CYTOLOGY AND TELIOSPORE DEVELOPMENT 21
22 SINGH and PAVGI a) Some of the primary sporidia in a whorl remain anucleate either due to lesser number of nuclei than sporidia (Fig. 27) or due to migration of more than one nucleus into a few sporidia (Figs. 28, 29). Such sporidia either directly produce infection hyphae or secondary sporidia without conjugation and the cytoplasm is gradually withdrawn from the anucleate sporidia to provide for the developing infection hyphae (Figs. 28, 29). Such an irregular movement of nuclei has been seen in E. calendulae, parasitic on another host genus of the Compositae (KAISER 1936). b) Bifurcation of promycelium bearing terminal sporidia on either or both branches occurs with no significant increase in sporidia! numbers (Figs. 31, 32). Infrequently one of the branches remains abortive and all the nuclei migrate in the sporidia! whorl of the other branch. Almost two thirds of the sporidia conjugated in pairs with their compatible partners (Fig. 31 ). Sometimes both the branches terminate in 2 symmetrical whorls of sporidia, wherein an equal number of nuclei migrate into each branch irrespective of the sporidia! number borne in the whorl (Fig. 32). c) The most frequent nuclear anomalies observed during germination show that the fusion of primary sporidia in situ has been suppressed with sporidia directly producing either infection hyphae or secondary sporidia with other minor diviations in the nuclear behaviour (Figs. 30, 33 to 37, 40). All the 4 nuclei formed, after meiotic division, continue to divide mitotically and whether 2 compatible nuclei pass directly into the sporidia or one nucleus divides after migration in a sporidium is uncertain; but the sporidium produces dikaryotic infection hyphae or secondary sporidia without cellular fusion. Some nuclei lag behind, possibly later entering the anucleate sporidia or may remain in the promycelium (Figs. 30, 33 to 37). A physiological barrier possibly impedes their movement into the sporidia. Occasionally, a fusion bridge is initiated between 2 compatible sporidia by an unknown physiological stimulus and plasmogamy is consummated even in the absence of a nucleus in one, while the nucleus of the other sporidium divides and functions as a dikaryon (Fig. 34 ). Rarely a few nuclei pass into sporidia and the others remain in the promycelium, which later divide several times forming multinucleate sporidia and promycelia (Fig. 40). d) A most interesting nuclear behaviour was observed, when fusion of 2 compatible sporidia occurred in situ (Figs. 38, 39, 41). The sporidia} number was increased by bifurcation of the promycelium, 2 infection threads were formed on the fusion bridge and the several nuclei divided simultaneously. Sometimes the nuclei divided and moved irregularly without following a
CYTOLOGY AND TELIOSPORE DEVELOPMENT 23 definite pattern and conjugation occurred between compatible sporidia even in the presence of more than 2 nuclei in both sporidia (Fig. 39). Rarely 2 fusion bridges were observed conjugating 3 sporidia. It was thought that 2 nuclei migrated into one sporidium (the third nucleus remaining stationary), while the fourth nucleus moved into the infection thread through an arching bridge from the other sporidium (Fig. 41 ). D) Development of teliospore. The dikaryotic infection hyphae enter through stomata on the under surface of the leaf (Fig. 42). The 2 compatible nuclei remain in close proximity in the vigorously growing tip of the infection hypha. Early stages of infection showed several hyphae collected in the substomatal chamber, later ramifying intercellularly in the mesophyll. Young hyphae are hyaline, sparsely septate, irregularly branched, dikoryotic and 1.5 to 2.5!J. in diam. (Fig. 43 ), similar to those observed in E. microsporum, E. nymphaeae and E. scirpicola (DAS 1949; LUTMAN 1910; THIRUMALACHAR and DICKSON 1949). Development of large, intercellular, binucleate haustoria has been observed only in E. nymphaeae (LUTMAN 1910; RACIBORSKI 1897). Clamp connections were observed proximal to the hyphal tips, where a small tubular branch bent downward and the nuclei underwent a conjugate mitotic division. One nucleus became oriented obliquely toward a daughter nucleus in the hook, while the other nucleus became spindled along the axis of the hypha. Later a daughter nucleus from each of the nuclei became separated by formation of 2 septa (Fig. 45). Clamp connections were also formed on intercalary cells resulting in the formation of small, dikaryotic lateral sporogenous cells or at times on short, few-celled branches (Figs. 44, 45). Clamp connec\ \ons have been observed in only a few species of the smut fungi. SEYFERT ( 1927) reported an intensive study on conjugate nuclear division and the formation of clamp connections in the smuts. He investigated a dozen species representing the genera Entyloma, Tilletia Tulasne, Tuburcinia, Urocystis Rabenhorst and Ustilago (Pers.) Roussek finding clamp connections in all. Clamp connections had previously been reported in E. calendulae, E. chrysoplenii (BERKELEY and BROOME) Schroter and E. ranunculi (Bonorden) Schroter (SEYFERT 1927). The mycelium became closely septate and irregular in thickness with several interwoven hyphal strands occupying the intercellular spaces (Figs. 46, 47). The terminal and lateral sporogenous cells formed by the clamp connections enlarged, became dense in cytoplasm and the nuclei appeared more prominent due to deep staining (Fig. 48 ). The host cells were pushed apart by repeated formation of clamps and new cells occupying the intercellular spaces (Figs. 47 to 51). Sometimes a young enlarged sporogenous cell budded
24 SINGH and PAVGI a new sporogenous cell and the nuclei underwent conjugate division. The daughter nuclei passed into the newly formed cell, which developed into a teliospore. Three to 4 or more teliospores formed in this manner became enveloped in a common sheath. The terminal and lateral sporogenous cells enlarge and become 8-10 1J. in diam. (Figs. 48 to 50) followed by the formation of a thin initial spore wall, which gradually becomes prominent and thick (Fig. 51) with further enlargement of the spore cell. The peripheral portion of the cytoplasm not included in the spore wall becomes the sheath layer around the developing teliospore. The density of the cytoplasm increases and 2 nuclei lying side by side fuse, consummating karyogamy and forming a diploid nucleus of the mature teliospore (Figs. 49, 50). Normally karyogamy occurs after the formation of the spore wall, but occasionally earlier. Teliospore development has been studied in detail by LUTMAN ( 1910) and RACIBORSKI (1897) in E. nymphaeae, in which it occurs in the same manner as observed for E. thirumalachari. The appearance of a large vacuole pushing th~ diploid nucleus to one side in the mature teliospores of E. nymphaeae was not observed in E. thirumalachari. The mycelium continues to ramify in the healthy tissues at the peripheral region, developing into a diffuse sorus by continued formation of fresh teliospores. Development of teliospores in the sorus is centrifugal in the epicentre of the infection site. In a young sorus, only a few young teliospores are seen initially in the mesophyll; soon they develop in large numbers and aggregate in a maturing sorus (Figs. 50, 52). The fungus does not invade the vascular tissues (Fig. 52). E) Histopathology. Serial sections through a sorus indicate no difference in reaction to infection between the palisade and spongy parenchyma tissues. The number and size of the chloroplasts is diminished with the development of a sorus and the intercellular spaces widen as the mesophyllar (spongy) cells are pushed upwards and sideways by the developing teliospores. Similar pathological Plate III. Development of mycelium and teliospores of Entyloma thirttmalachari in the leaves of Blumea oxydonta. Fig. 42. - Infection hypha entering through a stoma. - Figs. 43-45. - Intercellular myce lium, dikaryotic nuclear condition and formation of clamp connections on terminal and intercalary cells. - Figs. 46, 47. - Profuse development of mycelium giving prosenchymatous appearance. - Fig. 48. - Enlargement of sporogenous cells with prominent dikaryotic nuclei in each. - Figs. 49, 50. - Teliospores in different developmental stages showing demarcation of spore wall and outer sheath, dikaryons and their fusion. - Fig. 51. - Mature teliospores in intercellular spaces of mesophyll. - Fig. 52. - Distribution pattern of teliospores in the mesophyll. (Scale A: Figs. 42 to 51; B: Fig. 52).
CYTOLOGY AND TELIOSPORE DEVELOPMENT 25
26 SINGH and PAVGI changes have been reported earlier for other Entyloma species (DAs 1949; JosHI 1960 ). Aggregation of teliospores in the intercellular spaces also contributes to the formation of galls by pushing the mesophyllar cells apart (Fig. 52). DAs (1949) reported hypertrophy of parenchymatous cells antecedent to gall formation in E. micros porum, with an increase in the number of parenc-hymatous cell layers contributing to the gall formation on the leaves of Blum a species. The infection induces hypertrophy of cells rather than hyperpla!'y in E. thirumalachari. DISCUSSION Blumea oxydonta, the host of Entyloma thirumalachari is a rainy season (July to November), terrestrial plant thriving on uncultivated, waste lands, bunds of cultivated fields or shady places, being exposed to various ecological conditions.md often remaining in standing water for few days during heavy rains. Teliospores from the previous season, embedded in the tissues of the host plant Jebris remain in the soil and are released from the disintegrating tissues with the advent of monsoon rains to germinate and serve as the primary inoculum. Normally, the teliospores germinate by a promycelium bearing a whorl of sporidia. The diploid nucleus migrates into the promycelium, divides meiotically and mitotically to increase the nuclear number to 8. A single haploid nucleus passes into each sporidium of the apical whorl. Compatible sporidia conjugate in situ and produce dikaryotic secondary sporidia. The needleshaped secondary sporidia equipped with lateral process( es) may be an adaptation by the fungus to become wind-borne, inciting local infection on the leaves and petioles. Budding of dikaryotic secondary sporidia has not been observed in this species as in E. eleocharidis PAVGI and SINGH (PAVGI and SINGH 1970; SINGH 1968). The site of nuclear division during teliospore germination differs from species to species. Both the nuclear divisions in E. microsporum occur in the teliospore (DAs 1949 ), while the first division occurs in the teliospore and subsequent 2 successive divisions in the promycelium of E. scirpicola (THIRU MALACHAR and DICKSON 1949 ). In the remaining species studied including E. thirumalachari under study, they have occurred in the promycelium. Chromosome complements and their behaviour during nuclear division for the Entyloma species previously studied cytologically have not been critically observed. WANG ( 1934) reported it as the diploid number for E. dahliae and the same number was reported for E. microsporum (DAs 1949). He reported the first division as equatorial and the second division as reductional making the haploid chromosome number 2. Six was found to be the haploid number of E. thirumalachari with the first division observed to be reductional followed
CYTOLOGY AND TELIOSPORE DEVELOPMENT 27 by an equatorial one. The chromosomes are distinct during metaphase I. A chromosome complement of 4 has been reported for most of the Ustilaginales except the pioneer contribution by HARPER (1898), in which he reported 8 to 10 chromosomes in Ustilago scabiosae (Sowerby) Winter (FISCHER and HOLTON 1957). Several aberrant types of germination were frequently observed. Migration of more than one nucleus into only few sporidia increases the elongation potentiality of the infection hypha, which may be helpful to the fungus in locating nearby susceptible host leaves to incite infection of plants submerged in water. These aberrant germination types may be an adaptation by the fungus for better survival under varied ecological conditions. Further studies are required to correlate different aberrations with the factors conditioning their production. The diplobiontic type of nuclear rhythm possessed by the fungus is typical of the family Tilletiaceae (GA.uMANN and WYND 1952), which is more rigid in nuclear behaviour than observed in other species of Entyloma. Consequentely, lesser chance prevails for widespread occurrence of this smut in nature. This weakness of the pathogen has been partially compensated for its potentiality to incite secondary host infection (SINGH and PAVGI 1968). The cytological studies of the fungus indicate close similarity in life cycle with other Entyloma species parasitic on members of the fam. Compositae e.g. E. calendulae and E. dahliae, than the species parasitic on members of other host families. REFERENCES DAs M. C., 1949. - Morphology and cytology of Entyloma microsporum (Unger) Schroet. and Urocystis anemones (Pers.) Winter on Ranunculus repens L. Indian Phytopath., 2: 108-127. FISCHER G. W. and HoLTON C. S., 1957. - Biology and control of the smut fungi. The Ronald Press Co., New York, 622 p. GXUMANN E. A. and WYND F. L., 1952. - The fungi. Hafner Publishing Co., New York, 420 p. HARPER R. A., 1898. - Nuclear phenomenon in certain stages in the development of the smuts. Trans. Wise. Acad. Arts and Letters, 5: 475-498. JoHANSEN D. A., 1940. - Plant microtechnique. McGraw Hill Book Co., Inc., New York, 523 p. JosHI N. C., 1960. - Studies in Ustilaginales 3. Morphology, cytology and spore germination of Entyloma dahliae on Dahlia variabilis Des/. Indian Phytopath., 13: 6-11. KAISER W., 1936. - Zur Biologie und Entwicklungsgeschichte einiger Entyloma Arten. Angew. Bot., 18: 81-131. LuTMAN B. F., 1910. - Some contributions to the life history and cytology of the smuts. Trans. Wise. Acad. Sci. Arts and Letters, 16: 1191-1244. OLIVE L. S., 1965. - Nuclear behavior during meiosis. In: G. C. Ainsworth and A. Sussman (Ed.). The Fungi. Academic Press Inc., New York, Vol. 1: 143-161.
28 SINGH and PAVGI PARAVICINI E., 1917. - Untersuchungen iiber das Verbalten der Zellkerne bei der Fortpflan zung der Brandpilze. Ann. Mycol., 15: 57-96. PAVGI M. S. and SINGH R. A., 1967. - An interesting Ent} loma species from India. Can. ]. Botany, 45: 939-943. -, 1970. - Teliospore germination, cytology and development of Entyloma eleocharidis. Cytologia, 35: 391-401. RACIBORSKI M., 1897. - Ramphospora nymphaeae. Flora, 83: 75. SEYFERT R., 1927. - Ober Scbnallenbildung in Paarkernmyzel der Brandpilze. Zeitscher. f. Bot., 19: 577-601. SINGH R. A., 1968. - Studies 011 some diseases of rice and smuts of forage plants from Uttar Pradesh. Ph. D. Thesis, Banaras Hindu University, India, 184 p. SINGH R. A. and PAVGI M. S., 1968. - Secondary host infection by an Entyloma species. Curr. Sci., 37: 26. -, 1973. - Artificial culture of Entyloma thirumalachari. Sci. Cult., 39: 456-458. THIRUMALACHAR M. J. and DICKSON ]. G., 1949. - The cytology and life cycle of a CJ pericolous Entyloma species. Am. J. Botany, 36: 404-408. THIRUMALACHAR M. ]. and NARASIMHAN M. ]., 1953. - Notes on some mycological methods. Mycologia, 45: 461-468. THIRUMALACHAR M. J. and PAVGI M. S., 1950. - Notes on some spore germination and mounting techniques. Indian Phytopath., 3: 177-178. WANG D. T., 1934. - Contribution a l'itude des Ustilaginees (Cytologie du parasite et pathologie de la cellule bote). Le Botaniste, 26: 540-670. SUMMARY Cytology and development of Entyloma thirumalachari Pavgi and Singh, parasltlc on Blumea oxydonta DC. have been described. During teliospore germination, the diploid nucleus migrates into and divides in the promycelium. The first division is reductional followed by an equational one. Some of the meiotic divisional stages were observed and the haploid chromosome complement determined to be n=6. Several aberrations in teliospore germination and nuclear behavior were observed and their repercussions discussed. Sporidia! development and their conjugation, parasitic mycelium and formation of damp connections, and development of teliospores in the host tissue have been studied. The fungus possesses a diplobiontic rhythm of life cycle, which is well adapted for its effecth e dissemination and survival under the ecological conditions to which the host is subjected.