Spore dormancy in vesicular-arbuscular mycorrhizal fungi

Spore dormancy in vesicular-arbuscular mycorrhizal fungi

[ 37 ] Trans. Br. mycol. Soc. 81 (1), 37-45 (1983) Printed in Great Britain SPORE DORMANCY IN VESICULAR-ARBUSCULAR MYCORRHIZAL FUNGI By I. C. TOMMER...

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[ 37 ] Trans. Br. mycol. Soc. 81 (1), 37-45 (1983)

Printed in Great Britain

SPORE DORMANCY IN VESICULAR-ARBUSCULAR MYCORRHIZAL FUNGI By I. C. TOMMERUP Department of Soil Science and Plant Nutrition, University of Western Australia, Nedlands, W.A. 6009, Australia Spores of four species of VA mycorrhizal fungi were unable to germinate when first formed. The change in their capacity to germinate was examined by using populations of newly-formed spores with a narrow age range. Percentage germination and length of hyphae produced by the spores when incubated in soil or on nutrient agar media was measured during storage at constant temperatures between 5 and 37°C in both wet (-0'15 MPa) and dry (-300 MPa) soil. The results supported the hypothesis that spores of these species were innately dormant when first formed and suggest that changes occurred with close synchrony. The dormancy period in wet soil was approximately 6 weeks for Glomus caledonium and G. monosporum, and 12 weeks for Gigaspora calospora. This period was significantly reduced to 1 week for the Glomus spp. and 6 weeks for Gigaspora calospora in dry soil. For Acaulospora laevis the dormancy period was 6 months under all conditions. Plant roots had no effect on the change to quiescence of spores of any species, however newly-quiescent spores were effective propagules. Spores are important propagules for many vesicular-arbuscular (VA) mycorrhizal fungi but their value as inoculum at a particular time depends on their capacity to germinate and infect plants. The problem of considerable (and unexplained) variability in germination between populations of spores has been mentioned by many authors (Gerdemann & Trappe, 1974; Godfrey, 1957; Daniels & Graham, 1976; Hepper & Smith, 1976; Mosse, 1959, 1970; Sward, Hallam & Holland, 1978) and has also been encountered with organisms isolated from soils under either agricultural crops or virgin vegetation in Western Australia. Several explanations for the failure of spores of VA mycorrhizal fungi to germinate have been suggested, excluding that of innate dormancy. Dormancy of spores, sporocarps or sclerotia has been recorded for fungi from all major taxonomic groups. Many of them are biotrophs (Turian & Hohl, 1981; Weber & Hess, 1976). The present study was undertaken to determine whether dormancy is a factor contributing to the inability of spores of VA mycorrhizal fungi to germinate. In addition, a procedure is described for routinely producing spore populations that have a consistently high capacity to germinate with close synchrony. In this paper, a dormant spore is one that fails to germinate although it is exposed to physical and chemical conditions that will support germination and hyphal growth of apparently identical, but non-dormant, spores of the same species. The

germination of a dormant spore is prevented by structural and, or, physiological characteristics of the spore. A quiescent spore fails to germinate unless the above physical and chemical conditions are fulfilled, and germination of a quiescent spore is prevented only because the environment is unsuitable. Germination is defined as the production of a germ-tube up to 5 Jlm in length. Further elongation is described as hyphal growth. MATERIALS AND METHODS

Production of spores Isolates of Glomus caledonium (Nicol. & Gerd.)

Trappe & Gerdemann, Gigasporacalospora (Nicol. & Gerd.) Gerdemann & Trappe, Acaulospora laevis Gerdemann & Trappe, and Glomus monosporum Gerdemann & Trappe (isolate MGM of Abbott & Robson, 1979) were those used previously (Tommerup & Kidby, 1979, 1980). Cultures of each fungus were grown in association with subterranean clover (Trifolium subterraneum L. cv. Seaton Park) in closed, 17 em diam pots containing 3 kg of Lancelin sand (Hill, Robson & Loneragan, 1978) which had been steamed for 1 h. Nutrients, except phosphorus and nitrogen, were applied at rates adequate for maximum growth. Phosphorus (as KH 2PO 4) was added at a rate sufficient to give 4050 % of the maximum growth of tops of uninoculated plants. The nutrients were applied in solution to the surface of dry soil in each pot and dried before mixing (Hill et al., 1978). After pregermin-

I. C. Tommerup ating for 24 h, seeds of uniform size and radical length were inoculated with Rhizobium trifolii strain TAl and planted. The pots were watered to field capacity (-0'03 MPa) two days prior to planting and maintained at that matric potential by frequent weighing and watering. The soil matric potential was monitored during each experiment using the filter paper method (Hamblin, 1981 ). Pots were incubated at 20°C in temperaturecontrolled tanks. The cultures were established with spore-free, fresh mycorrhizas using the following schedule, in which the primary source of inoculum was spores isolated from pot cultures using the sieving, centrifugation and flotation procedure (T ommerup & Kidby, 1979). Fifty spores from the 100 to 250 11m fraction were placed at 3-4 em depth, with two pregerminated clover seeds above them at 1 em. After 4 weeks the seedlings were removed, the roots chopped to approximately 1 em lengths and layered in new pots at 3 em depth, and the pots seeded. The plants were grown for 3 weeks and the procedure repeated twice more before the inoculum was used to initiate infection in experimental pots. At each transfer three sub-samples of root inoculum were cleared, stained and examined (Phillips & Hayman, 1970). No new spores were found and the mature spores used as primary inoculum were not observed after the second transfer. Approximately 1 g of heavily infected root inoculum was placed at 3 cm depth in each pot which was to produce the spores, and four pairs of pregerminated clover seeds were planted and thinned 2 weeks after emergence to one seedling per site . To determine the uniformity of sporulation, the production and development of spores was monitored by weekly core samplings of two replicate pots (not subsequently included in the final harvests ) from 3 weeks after planting. Spores were separated from the core samples by sieving, flotation and centrifuging as described above. Root fragments with adhering extramatrical hyphae were collected from the coarser mesh grids and examined for spores before being cleared and stained (T ommerup , 1982). The onset of sporulation, i.e, the initial appearance of small, immature spores « 30 11m diam) and/or mother cells, occurred at 8 weeks for Acaulospora laevis and Gigaspora calospora, and at 4 weeks for Glomus caledonium and G. monosporum, but large numbers of immature spores were not found until 8 weeks. Pots were harvested at 11 weeks when the morphological development of most of the spores greater than roo sm diam appeared to be complete. At each harvest the tops of the clover plants were removed. Then the soil containing spores and

infected roots of each replicate pot was finely chopped and thoroughly mixed. Germination test Quiescent spores from populations which had shown high ( > 85 % ) germination for between 5 and 7 months were separated from soil as described above and incubated at 22° between membrane filters in soil wet to a matric potential equivalent to -0'15 MPa (T ommeru p & Kidby, 1979). Their germination and hyphal lengths were measured (at x 50 to X 1000) at invervals of 2 to 3 days. This system was also used to germinate newly formed spores which were stored at 25° in either wet or dry soil as described later under spore storage. They were sampled at harvest (0 days) and subsequently at 3 days and 1 week for Glomus caledonium and G. monosporum at 1,3,6 and 12 weeks for Gigaspora calospora, and 1, 3, 6, 12 and 26 weeks for Acaulospora laevis. The effect on germination of flotation in 50 % (wIv) aqueous sucrose during the separation procedure, compared to flotation in water, was examined using both the old quiescent and the newly-formed spores of the four species. For the water separation, the soil fractions retained on the 100 11m sieves were suspended in water and examined in a nematode counting dish (H aym an & Stovold, 1979). The spores were removed by suction into a micropipette, transferred to membrane filters and their germination tested in the same way as with spores from the sucrose phase. RESUL TS

The definition of a dormant spore presumes a knowledge of the environment in which spores of the four species of VA mycorrhizal fung i germinated. During the germination test, since the lapse of time or the exposure to some of the environmental changes could have resulted in the spores being able to germinate and grow, the rate of germination and hyphal growth of non-dormant spores in each system was defined. Germination test

The time course of germination of old quiescent spores incubated between membrane filters in soil is shown (Fig. 1) for each species. There was no difference in the rate of germination, maximum germination, rate of hyphal elongation or maximum hyphal length between spores separated from soil by water only and those separated by sedimentation in water and flotation in aqueous sucrose. The rates of germination and hyphal elongation

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of newly formed spores of each species (T able 1) were the same as those of old quiescent spores (Fig. 1), showing that populations of newly quiescent spores had near synchronous germination. At all sampling times prior to those indicated on Table 1 no spores germinated, showing that the separation methods did not affect the time at which spores became quiescent . Both separation methods differentiated between dormant and quiescent spores. Since the flotation in sucrose for a total of 2 minutes did not affect the change to quiescence in dormant spores or

the developmental stages used to measure germination, the method has been used for all other results reported. It had the advantage of being a much more rapid procedure and a slightly more effective recovery method than the water system (T ornmerup, unpubl.). The influence of several physicochemical treatments on the subsequent capacity of newly formed spores to germinate was examined in a series of time-course experiments to determine whether they affected dormant spores , the change to quie scence or the behaviour of quiescent spores.

I. C. Tommerup



Table 1 . Time taken for maximum germination and hyphal elongation of newly-quiescent spores offour VA mycorrhizal fungi, when incubated in soil after separation from wet or dry soil by flotation in sucrose

Sampling time (weeks)

Time of maximum % germination (days)'

Time at which hyphae of all spore s reached maximum length"

Glomus caledonium

Dry soil 1 Wet soil 6

12'2 11 ,8

14 '8 15'4

G. monosporum

Dry soil 1 Wet soil 6

12'9 13 '4

16'5 16 '0

Gigaspora calospora

Dry soil 6 Wet soil 12

14'6 16'3

21 '2 21'5

A caulospora laevis

Dry soil 26 Wet soil 26

16'3 15'7

17'4 17'1

a b Values are means of 12 replicates (4 replicates of each of 3 batches) of 20 spores. No standard errors were greater than 1'0.

Storage Spores were stored at two moisture regimes and four temperatures for periods up to 42 weeks. For each fungus half the soil containing spores was dried from a moisture content equivalent to -0'03 MPa at harvest to -0'15 MPa at 24 hand maintained at that. The other half was dried at 20° to - 300 MPa by 7 days after harvest. At day 7, pairs of replicate 2 kg samples of both wet and dry soil were transferred to each of the storage temperatures, 5°, 10°,25° and 37°, in S kg vacuum sealed jars in darkness. During storage the matric potentials and water contents of the wet and the dry soils were monitored using both the filter paper (H amblin, 1981) and gravimetric methods. Desorption curves for soil water were obtained from direct suction plant, direct pressure plate and pressure membrane plate apparatus. The germination of spores from 4 replicates of each treatment was tested at harvest (0 days ) and subsequently at 1, 3, 6, 12, 26 and 42 weeks. Incubation periods were 2 weeks for Glomus caledonium and G. monosporum, and 3 weeks for Acaulospora laevis and Gigaspora calospora. All newly-formed spores were dormant at the time of harvest (Fig. 2). At the subsequent sampling times their capacity to germinate varied according to the species, the soil water matric potential and the storage temperature. When spores germinated, the hyphal lengths of each species did not differ (P > 0 '1) from the maximum lengths in Fig . 1, showing that the physiological changes involved in attaining a quiescent state were complete. No spores of any species stored at 37° in wet soil germinated, indicating that this temperature is lethal to hydrated, dormant cells. The pattern of behaviour of spores of Glomus

caledonium and G. monosporum was similar (Fig. 2). After one week's storage in drying soil, a high proportion of spores germinated, whereas none germinated in wet soil until after 3 weeks of storage, The capacity of spores from wet soil to germinate at 6 weeks was not changed by the storage temperature except for G. monosporum between SO and 2SO (P = o-oy). Only at 5° was there an increase in germination between 6 and 12 weeks (P < o-oy). Unlike the Glomus spp., drying soil did not immediately induce a quiescent state in spores of Gigaspora calospora. Germination of spores from dry soil occurred first at 6 weeks (Fig. 2), and storage of spores in wet soil at 2So delayed the capacity to germinate until after six weeks. Very few spores of G. calospora from the dry soil at 37° germinated and the higher temperature did not alter the storage time at which germination could be induced, indicating that 37° is close to the maximum for survival during the change from dormancy to quiescence. The lag phase during which no spores of Acaulospora laevis germinated was at least 12 weeks for all treatments . Soil matric potential did not affect the rate of change to quiescence at 10° or 2So (Fig. 2). In dry soil at So, the rate was the same but in wet soil at So, it was slower at 26 weeks (P < 0 '01) and 42 weeks (P ~ 0 '01 ). Compared with the other treatments, few spores from dry soil at 37° germinated (P ~ 0 '01) indicating this temperature was lethal to most dormant spores. Host plant roots

Spores were stored in wet soil ( - 0' 1S MPa) at 20° for 0, 3, 6, 9, 12, 18 and 26 weeks before being separated. Then they were placed at 4 em depth in

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Fig. 2. Percentage germination of spores in soil at 22° as a function of the period of storage in wet or dry soil at constant temperatures between 5 and 37°" Values are means with standard errors of four replicates of 60-150 spores. soil maintained at 20° and - 0 '15 MPa, with two subterranean clover seeds planted at 1 cm above each of the 10 replicate groups of 25 spores, After 4 week's growth, seedlings and soil were removed in 2 cm diam core wh ich was sliced at 2 em intervals below the seed . The roots were floated free of the soil, cleared and stained. They were examined for the pres ence of germinated spores, and pre- and post-penetration infection structures. The soil suspension was also stained and examined for

germinated spores. In the control treatments spores were separated from soil, then incu bated for 4 weeks in pots without plants. Germination of newl y-formed spores was not influenced by roo ts of host plant seedlings, At no planting time was there a difference (P < 0 '1 ) between percentage germination of spores from the two treatments . The percentage germination, hyphal length and t ime at which germination was first found for spores of Glomus caledonium and

42

I. C. Tomm erup

G. monosporum incubated directly in soil with or without roots did not differ from those reported above for spores stored in wet soil at 10° and 25° and germinated between membrane filters (F ig. 2, Table 1) . Early post penetration infection occurred in seedlings planted at six weeks. Fewer than 5 % of the spores of Gigaspora calospora germinated at 9 weeks and there were no infection structures. By 12 weeks, the percentage germination and hyphal growth was the same as in Fig. 2 and roots were infected. Approximately 16 % of spores of Acaulospora laeuis germinated at 18 weeks but the hyphae were different from those produced at 26 weeks. They were unbranched, narrow, very short (less than 100 pm), and stained less intensely than those from the later harvest, and they did not form any pre- or post-penetration structures. Germination at the 26 week planting time was the same as shown above (Fig. 2) and the plants were infected. These results also showed that compared to incubation directly in soil, incubation between membrane filters in soil had no effect on dormant spores or the germination of quiescent spores. Soil temperature Spores ofGlomus caledonium and Acaulospora laeois were separated from soil maintained at - 0'15 MPa and 20° at harvest (0 days) and after 3 , 6, 9,12,18 and 26 weeks, and then incubated between membranes in soil at 10°, 15° and 22° for the same periods as in the spore storage experiment. These temperatures were selected to cover the range recorded during the growing season in the upper 3-10 em of the soil profile in the natural habitat of these fungi. The incubation temperatures did not modify the onset of quiescence. For each species, the lapsed storage time at which germination was first found, the percentage germination and hyphallengths, did not differ from those of spores sampled at the same storage times in the preceding experiments, except for A. laevis incubated at 10°. Germination at this temperature was zero at 18 weeks and 10 % at 26 weeks . The temperature is close to the minimum for spore germination in A . laevis (T ommerup, unpubl.). Agar media Spores were separated from soil stored at -0'15 MPa and 20° after storage for 0 days (harvest) and 3, 6, 9, 12, 18 and 26 weeks. Their surfaces were treated for 20 min in 0'42 % chlorine (5 % w [v chloramine-T) at 30° (Tommerup & Kidby, 1980) and incubated on (i) 1 % Difco agar

in deionized water; (ii) 0 '02 % yeast extract (D ifco) and 1 % agar in deionized water and (iii) 1 % agar in a soil extract (modification of Sivasithamparam & Parker, 1981 ). Spore germination and hyphal lengths were measured after 2 weeks incubation for the Glomus spp . and 3 weeks for A caulospora laevis and Gigaspora calospora. There were five replicates each having 30 spores. Selection of the harvest time was based on previous information (T omm erup & Kidby, 1980; Figs 1-3 and Table 1 this paper). In the control treatments, immediately after spores were separated, they were incubated in the soil germination test system. On all the media, germination was first found for spores of both Glomus spp, that had been stored for 6 weeks and for Gigaspora calospora and Acaulospora laeois after 12 and 18 weeks respectively (T able 2 ). At these times spore populations of each of the first three species had attained both maximum percenrage germination and hyphal growth on each of the media within the incubation time chosen. However, fewer than 10 % of the spores of A. laevis had germinated at 18 weeks. The hyphae formed were less than 100 pm long and had the different morphology described above, indicating that the spores had commenced but not completed the change to a quiescent state. At 26 weeks, maximum germination occurred. For each species, the storage period before germination was first observed was the same on agar (T able 2) as in soils (F ig. 2, Table 1). These findings suggested that neither the mild oxidising treatment nor the nutrient status or physical properties of the agar substrates were environmental factors that affected the rate of change to a quiescent state. The time at which the change from dormancy to quiescence was completed in a high percentage of the spores of each species can be determined by taking into account storage periods, the total time of incubation periods, the lapsed time prior to the onset ofgermination ofnewly quiescent spores, and the time at which hyphae attain maximum lengths. This information from all the experiments is summarized in Table 3. The actual time limits during which spores become quiescent may have been narrower than the values given, especially in the case of Acaulospora laeuis in dry soil for which no intervening measurements were made. The change to quiescence occurred in a few days for the two Glomus species in dry soil, over narrow periods in wet soil and also over narrow periods for the other species under most conditions despite great differences in the dormancy periods. These findings indicated that the change occurred with near synchrony in each species and therefore that the stage of physiological development was simi-

Spore dormancy in V A fungi Table

2.

43

Effect of storage time on the capacity of newly-formed spores of four V A mycorrhizal fungi to germinate on agar media Yeast Agar

Water Agar Storage time (weeks)

Soil Agar

Germination"

Hyphal length"

Germination

Hyphal length

Germination

Hyphal length

("!o)

(rom)

("!o)

(rom)

("!o)

(rom)

0 458±33 474±21 468±27

0 73±4 85±3 88±4

0 469±24 43 1±29 486±33

0 34 1±33 394±28 382±31

0 68±5 76±3 75±4

0 362± 17 374±23 379±30

0 0 67±4 4 18±32 476± 19 75±5 Acaulospora laevis 0 0 12±4 6±3 86±5 489±24

0 71 ±3 73±4

0 473±21 481±24

0 0 10±5 7±2 89±3 438±43 Values are means with standard errors of 5 replicates of 30 spores.

0 21±3 528±24

0-3 6 9 12

0 69±4 82±4 87±3

0 402±28 431±23 462±35

0-3 6 9 12

0 64±5 72±4 74±3

0 386±29 352±19 376±3 2

00-9 12 18

0 64±6 72±3

0 421±26 462±31

0-12 18 26

0 4±2 79±4

Glomus caledonium 0 73±3 84±4 86±4 G. monosporum 0 65±4 74±3 76±4 Gigaspora calospora

a, b

Table 3. Time limits between which spores offour V A mycorrhizal fungi became quiescent when they had been stored in soil between 10 and 25 0 Time after harvest (weeks) Spores from dry soil

Spores from wet soil

0'5-1 0'5-1 5'5-7'0 13'5-26'0

4'5-6'0 4'5-6'0 11'5-1 3'0 21'5-26'0

Glomus caledonium G, monosporum

Gigaspora calospora Acaulosporalaevis

Table 4. Frequency distribution of spore sizes in four V A mycorrhizal fungi Spore width* (um)

Glomus caledonium G. monosporum

100-129 4 11

56 0

Gigaspora calospora

Acaulospora laevis

* Width (diameter) of

100

130-159 56 27 39 3

160-189 32 38 5 18

190-210 5 18 0 48

220-250 0 6 0 31

spores, measured perpendicular to the attachment hyphae.

lady synchronized. Developmental synchrony appeared to be related to the age rather than the size of spores. At harvest, spores of each species were at a similar stage of anatomical development and mostly alike in age but spore size was less uniform (Table 4), a feature that was not reflected in the change to quiescence.

DISCUSSION

Populations of spores of all four species having a high capacity to germinate with near synchrony were produced. Results indicate that there were two major contributing factors; the uniformity of spore age and innate dormancy in newly formed spores.

44

I. C. Tommerup

All spores of each species were innately dormant when first formed. They did not germinate under a range of physical and chemical soil conditions, characteristic of the natural environment of the cultures, which were conducive to the germination of quiescent spores. Furthermore the dormant spores failed to germinate on a range of laboratory media which supported the germination and hyphal growth of quiescent spores (Tommerup & Kidby, 1980 ; unpubl.). The change from dormancy to quiescence occurred with near synchrony in the populations used and this probably reflects the near uniformity of the physiological characteristics associated with dormancy. The change to quiescence was not influenced by any of the conditions used to separate or germinate the spores, but by the environment under which spores were stored following development. The length of the dormancy period varied considerably. Dormancy in wet soil was probably related to spore age for both species of Glomus and for the Gigaspora, but drying the spores in soil caused a rapid change to quiescence, thereby diminishing the significance of spore age. However, for Acaulospora laevis, spores of which apparently have a long period ofinnate dormancy, none of the storage conditions shortened the period. Dormancy was not relieved by the presence of growing plants and the capacity of quiescent spores to germinate is also not influenced by growing host roots (Tommerup, unpubl.). Similar findings have been reported for other species (Daniels & Trappe,

history or innate physiological differences (Daniels & Duff, 1978 ). Spore germination of several cultures of Glomus mosseae were improved by storage (Daniels & Graham, 1976; Hepper & Smith, 1976; Mosse, 1959 (yellow vacuolate )) and found to be high in old cultures of G. caledonium (Hepper, 1979). These results, together with those reported in this paper, indicate that the dormancy period among several species of Glomus in moist soil may be similar. Other cultures of the same two Glomus spp. isolated from geographically and edaphically diverse sites had identical dormancy periods to those reported here (T ommerup, unpubl.). Short dormancy phases would prevent the spores germinating immediately after development, thereby contributing to survival of inoculum particularly in moist field soils. The failure of quiescent spores to germinate in moist soils is due to fungistasis (Tommerup, in prep.). A long dormant period such as that of Acaulospora laevis would have advantages for survival in several diverse environments. They include those having regular but limited growing seasons interspersed with extreme conditions that are unfavourable for plant growth or those where the growing seasons occur at irregular intervals. Dormancy would protect the spores against an early false break in the dry season in climates such as those of the semiarid Mediterranean regions .

I thank Susan Grahame and Teresa Gigengack for technical assistance and J. P. Beilby and. The inability of dormant spores to infect plants D. K. Kidby for useful discussions. This work was may be one factor contributing to the lack of a supported by Post-Doctoral Research Fellowship relationship between spore numbers and the level from the University of Western Australia and the ofinfeetion (Hayman & Stovold, 1979; Kianmehr, Rural Credits Development Fund of the Reserve 1981). The occurrence of spore dormancy in Bank of Australia. species of VA mycorrhizal fungi and the length of the dormancy period are characteristics that need REFERENCES to be considered when inoculum includes spores. L. K. & ROBSON, A. D. (1979). A quantitative They are features that could affect the infectivity ABBOTT, study of the spores and anatomy of mycorrhizas formed and effectiveness of inoculum and therefore by a species of Glomus with reference to its taxonomy. influence the choice of inoculum type for field Australian Journal of Botany 1.7, 363-375. application (Abbott & Robson, 1981). ABBOTT, L. L. & ROBSON, A. D. (1981). Infectivity and effectiveness of vesicular arbuscular mycorrhizal fungi: Some of the low or erratic germination reported effect of inoculum type . Australian Journal of Agriculfor . spores of A. laevis (Mosse, 1970) or other tural Research 31., 631-639 . species (Godfrey, 1957; Hepper & Smith, 197 6; Mosse, 1959; Daniels & Graham, 1976) may be DANIELS, B. A. & DUFF, D. M. (1978). Variation in germination and spore morphology among four isolates attributable to innate dormancy especially in of Glomus mosseae. Mycologia 70, 1261-1267. populations having spores of variable ages, but DANIELS, B. A. & GRAHAM, S. O. (1976). Effects ofnutriincluding a proportion of newly formed spores. tion and soil extracts on germination of Glomus mosseae The causes suggested by other workers are spores. Mycologia 68, 108-116. consistent with this idea and they include the DANIELS, B. A. & TRAPPE, J. M. (1980). Factors affecting spore germination of the vesicular-arbuscular maturity and condition of sporocarps (Godfrey, mycorrhizal fungus, Glomus epigaeus. Mycologia 71., 1957); the time of year at which spores were col457-471. lected (Mosse , 1959; Sward et al., 1978); cultural

1980).

Spore dormancy in VA fungi GERDEMANN, J. W. & TRAPPE, J. M . (1974). The Endogonaceae in the Pacific North-West. Mycological Memoirs 5, 1-786. GODFREY, R. M. (1957). Studies on British species of Endogene. III. Germination of spores. Transactions of the British Mycological Society 40,203-210. HAMBLIN, A. P. (1981). Filter-paper method of routine measurement of field water potential. Journal of Hydrology 53, 355-3 60. HAYMAN, D . S. & STOVOLD, G. E. (1979). Spore populations and infectivity of vesicular arbuscular mycorrhizal fungi in New South Wales. Australian Journal of Botany 27, 227-233 . HEPPER, C. M . (1979). Germination and growth of Glomus caledonius spores: the effects of inhibitors and nutrients. Soil Biology and Biochemistry 11,269-277. HEPPER, C. M. & SMITH, G . A. (1976). Observations on the germination of Endogene spores . Transactions of the British Mycological Society 66, 189-194. HILL, J., ROBSON, A. D. & LONERAGAN, J. F. (1978). The effects of copper and nitrogen supply on the retranslocation of copper in four cultivars of wheat. Australian Journal of Agricultural Research :29, 925"""939. KIANMEHR, H. (1981). Vesicular-arbuscular mycorrhizal spore population and infectivity of saffron (Crocus sativus) in Iran. New Phytologist 88, 79-82. MOSSE, B. (1959). The regular germination of resting spores and some observations on the growth requirements of an Endogone sp. causing vesicular-arbuscular mycorrhiza. Transactions of the British Mycological Society 42, 273-286.

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MOSSE, B. (1970). Honey-coloured, sessile Endogone spores 1. Life history . Archiv fur Mikrobiologie 70, 167-175. PHILLIPS, J. M. & HAYMAN, D . S. ( 1970) . Improved procedures for clearing roots and staining parasitic vesicular arbu scular mycorrhizal fungi for rapid assessment of infection . Transactions of the Br itish Mycological Society 55, 158-161. SIVASITHAMPARAM, K. & PARKER, C. A. (1981). The physiology and nutrition of Gaeumannomyces graminis in culture. In Biology and Control of Take All (ed. P. J. Shipton&M. J. C. Asher ),pp.125-150. London: Academic Press. SWARD, R. J., HALLAM, N . D. & HOLLAND, A. A. (1978). Endogone spores in a heathland area of south-eastern Australia. Australian Journal of Botany :26, 29-43 . TOMMERUP, 1. C. (1982). Airstream fractionation of vesicular-arbuscular mycorrhizal fungi: concentration and enumeration of propagules, Applied and Environmental Microbiology 44, 533-539. TOMMERUP,1. C. & KIDBY, D. K. (1979). Preservation of spores of vesicular-arbuscular endophytes by L-drying. Applied and Environmental Microbiology 37, 831-835. TOMMERUP, 1. C. & KIDBY, D. K. (1980). Production of aseptic spores of vesicular-arbuscular endophytes and their viability after chemical and physical stress. Applied and Env ironmental Microbiology 39,1111-1119. TURIAN, G. & HOHL, H. R. (eds.) (1981). The Fungal Spore : Morphogenetic Controls. London: Academic Press . WEBER, D . J. & HESS, W. M. (1976). The Fungal Spore : Form and Funct ion. Wiley-Interscience.

(Received for publication 8 July 1982)