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The ecology of microfungi on decaying stems of Urtica dioica
[ 249 ] Trans. Br. my col. Soc. 51 (2), 249-259 ( 1968) Printed in Great Britain
THE ECOLOGY OF MICROFUNGI ON DECAYING STEMS OF URTICA DIOICA By A. S. YADAV
Department
of Botany, Science College, Patna, India AND
Department
M. F. MADELIN
ofBotany, The University, Bristol
The succession and distribution of microfungi associated with stems of Urtica
dioica L. decaying above ground have been studied. The percentage frequency of occurrence of species of fungi sporulating on the stems has been determined, and the intensity ofsporulation ofsome ofthese fungi and the percentage ofstem area covered by them have also been estimated. Certain fungus species show definite patterns of distribution on the stems. A primary microflora, consisting of species visible on the stems in the year ofth eir growth, and a secondary microflora, comprizing species appearing in the winter and following summer, may be recognized . A quiescent period, in which very few new species appeared, intervened. All the species discovered by isolation from the decayingstems had also been isolated previously from Heracleum sphondylium L. (Yad av, 1966). However Acremoniella atra (Corda) Sacco which was common on H. sphondylium was never isolated from U. dioica, Colonization of the moribund shoots started from the top. Alternaria tenuis Nees. ex Pers., Cladosporium herbarum Link ex Fr., Epicoccum nigrum Link, and Botrytis cinerea Pers, ex Fr. first invaded the fading leaves and adjoining segments. Lower segments later died and soon supported sporulation of these same fungi as well as of Phoma acuta Fuckel and Cladosporium sphaerospermum Penzig. A . tenuis and C. herbarum were rare on the lower segments.
This account of the ecology of microfungi on stems of Urtica dioica L. contributes to our knowledge of the decay of dicotyledonous herbaceous remains. One of the authors has already reported on the decay of stems of Heracleum sphondylium L. (Yadav, 1966). Like the latter plant, U. dioica has erect, slightly woody stems which do not rapidly decay beyond recognition; it is widespread and abundant in the British Isles and harbours a large population of microfungi during the decay of its shoots. MATERIALS AND METHODS
U. dioica is a herb with a perennial, creeping, much-branched root-stock from which in the spring arise erect, hispid, ribbed shoots, 0'84:>'9 or even 1'2 m high, with many rather short internodes. It flowers in summer and autumn, and then dies back, the lower internodes remaining green longest. The general methods of study were the same as those used by Yadav (1966). Stems which had recently died back and which were in the first stages of decay were gathered in October 1957 from fields at Swanley (N.G. Ref. TG 515670) and Eynsford (N.G. Ref. TQ531626) in Kent and placed upon the ground at Chelsea Physic Gardens, London, S.W. 3. Each
25°
Transactions British Mycological Society
month, three to five stems were removed at random from th e pile, cut into three equal segments, labelled A-C in ascending order, and examined as described below. Because of the rather large number of small internodes in these stems, it was not practicable to study quantitatively each internode individually. These collections were supplemented at irregular intervals by others taken from the original field sites at Swanley and Eynsford, as well as one at St Albans, Herts. (N .G. Ref. TL 132042). When gathering the stems, care was taken to ensure that they included the first internode exposed above soil level. The total number of U. dioica stems sampled from all four sources between No vember 1957 and October 1958 was eighty-eight. After collection, stem segments were incubated individually at high humidity for 2 days to facilitate identification and mapping of the sporulating fungal colonies. Then for each segment records were made of the following: (I) Total surface area of segment. (2) Names of all species of microfungi present. (3) Total area covered by each of those species which experience had shown to be important (Table 3). (4) Intensity of production of reproductive structures by certain important species. For pyrenomycetes, this was expressed as the mean number of ascocarps per field of a binocular dissecting microscope (78'5 mm") determined in the way described for pycnidia by Yadav (1966) (see this reference also for relevant methods for hyphomycetes). Although the principal site from which samples were drawn was artificially created, a number of observations indicated that the microflora on stems in this situation was substantially the same as that in natural habitats. First, the fungus microfloras of stems from the four different sampling sites were qualitatively alike in respect of all their principal components. The sixteen species which occurred on six or more of the eighty-eight stems studied quantitatively were found at all four sites. The microflora on stems from the artificial pile at Chelsea resembled that from the three natural sites in that, of the total of forty-four species observed on Urtica stems, thirty-nine were recorded at Chelsea. Conversely, only eight of these species at Chelsea were not collected also from natural sites elsewhere. Secondly, the rnicroflora in the year in which the principal quantitative studies were made closely resembled that in a second year of observations on stems from natural sites. Of the forty-four species seen in the first year, thirty-two were seen also at field sites in the second year. The twelve not seen were species which had been rare in the first season. Although the foregoing evidence indicates only a qualitative similarity between the microfloras of the artificial and natural sites, the succession and development of the geographically separated microfloras were not noticed to be significantly different. However, such differences as might exist would admittedly not be revealed by the quantitative data presented below which are based on a sampling procedure which combined the samples drawn from different locations.
Microfungi on Urtica. A. S. Yadav and M. F. Madelin
25 1
Table I. Percentage frequencies at various periods recorded for diiferent species offungi on decaying stems of U rtica dioica which had grown in 1957 On whole stems (
'I
Nov, 57- Nov, 57- May 58Oct. 58 Apr, 58 Oct, 58
Alternaria tenuis N ees ex Pers. Botryosphaeria sp. Botrytis carnea Schum. B, cinerea Pers. ex Fr. Calloria fusarioides (Berk,) Fr, Camposporium peliucidum (Grove) Hughes Circinotrichum sp. Cladosporium herbarum Link ex Fr. C. sphaerospermum Penzig Coniothyrium conoideum Sacc, Cryptodidymosphaeria conoidea (Niessl) Rehm Cylindrocolla urticae (Pers. ex Fr.) Bon, Dasyscypha sulphurea (Pers. ex Fr.) Massee Dendryphium comosum Wallr. D, griseum Berk, & Br. D, laxum Berk, & n-. Dictyosporium toruloides (Corda) Gueg. Dinemasporium hispidulum Sacc, Epicoccum nigrum Link Fusarium sambucinum Fuckel Fusarium sp. Graphium cuneiferum (Berk, & Br.) Mason & Ellis Helotium herbarum (Pers, ex Fr.) Fr. Leptosphaeriaacuia (Hoffm. ex Fr.) Karst, L. doliolum (Pers. ex Fr,) deNot. Oospora sp, Ophiobolus erythrosporus (Riess) Wint. 0, immersus Trail Paecilomyces betae (Delacr.) Cornford Periconia byssoides Pers, ex Corda P. cookei Mason & Ellis Phaeoisaria cornui (Bainier) Mason Phialea cyathoidea (Bull, ex Fr.) Gill, Phoma acuta Fuckel P, berkeleyi Sacc, P, herbarum West, P, urticae Sch, & Sacc, Pleurophragmium simplex (Berk, & Br,) Hughes Pyrenopeziza urticicola (Phil!.) Boud, Torula herbarum Link ex Fr. Trichocladium opacum (Corda) Hughes Trichothecium sp, Tricladium sp. Volutella ciliata Fr.
41 4'2
r-r
0
3'4 6'8
8'3 0 0
27 0 "6 ,,6 9'4 1,6
0 4' 46 8'3 8'3
"6 35 1,6 0 0
3'
,,'
r-r I"
37 '4 2'3 2'3 6,8
25
5'7
0
4'5
3'4 '00 i -r ,6 2'3 26
8'3 13 0 42 0 0 100 4'2 13 8'3 8'3
8'0 22 2'3
0 46 0
3'4 5'7 r-r
27
,,'
r-r
r-r 3'4
0 0 4'2
I"
3'4 I" I"
3'4 8'0 -r
i
r-r
0 5'7 8'0 r-r
,,'
0 0 3' '0 0 0 r-r
7,8
3'4
3'4
0
'3 2'3
6'3 7,8 1,6 3" 1,6
8'3 0 0
0 2'3 4'5 0
3'4
0 8'3 4'2 0 0
r-r
,.,
r-r
5'7
4'5 8'0 2'3 2'3
,6
C 27 0 0 3'4
0
3' ,,6 4'7 3"
0 25
B 3'4
0
4'2 8'3 0 0
2'3 26
A 0
2'3
23 3'4 3'4 2'3
,,'
On individual segments, Nov, '957-0ct, '958
r-r
II
3'5
,,'
9'0 0 3'4
i -t
0
r-r
2'3
4'5 3'4
I"
0
I"
0
0
r-r
i
-r
i-r
r-r
2'3 5'7 0
3" 27
2'3 21
1'1 4'5
0 2'3
19 ,,6 6'3
'4
4'5
2'3 0
"6 3'1 ,,6 20 1,6 4'7 '00 0 '7 0 33
r-r
2'3
,., 3'5 ,,' 9'7 I"
3'4 77 r-r
8'0 2'3 24
0
i
-r
2'3
,,'
2'3
"I
r-r
0 8'5 0
0 II
5'7
0 0 56 0 g'o 0 6,8
6,8 2'3 0
4'5 0 0 0
,,'
65 0 8'0 r-r
'3 3'1
8'0 20 2'3
"6 "6 3"
r-r
0
0 3'4
r-r
r-r
0
0
II
Transactions British Mycological Society
25 2
RESULTS
Succession and distribution rif species riffungi sporulating on decaying stems Frequency of occurrence Eighty-eight stems were gathered between November 1957 and October 1958 from all four sites. In Table I are shown the frequencies with which the forty-four fungal species seen occurred on the stems throughout the whole year and within the two component half-year periods. The table also shows the frequencies of occurrence throughout the year on the three segments separately. To facilitate description, the named categories of frequencies previously Table
2.
Species
riffungi recorded each month on stems rif Urtica dioica,
November 195 7-0ctober 1958 N. D. J. F. M. A. Alternaria tenuis + + + + Botrytis cinerea + + Phoma acuta + + + + + + Cladosporium herbarum + + + + Periconia cookei + + Epicoccum nigrum + + Coniotbyrium conoideum + Ophiobolus immersus + Phoma berkeleyi + Cladosporium sphaerospermum + + + + Paecilomy ces betae + + Dinemasporium hispidulum + + Phoma herbarum + + + Cryptodidymosphaeria conoidea + Dendryphium griseum + + Leptosphaeria acuta + + + Torula herbarum + + Leptosphaeria doliolum + Pleurophragmium simplex + + Dendryphium comosum + Botryosphaeria sp. + Cylindrocolla urticae + + + + Fusarium sambucinum + Volutella ciliata + Phoma urticae + Calloriafusarioides Fusarium sp. Phialea cyathoidea Ophiobolus erythrosporus Graphium cuneiferum Pyrenopeziza urticicola Periconiabyssoides Phaeoisaria cornui Circinotrichum sp. Trichothecium sp. Trichocladium opacum Dendryphium laxum Camposporium pellucidum Tricladium sp . Dasyscypha sulphurea
Dictyosporium toruloides Helotium hetbarum Botrytis carnea Oospora sp.
M. J.
J. + + + + + + + + + + + +
A. S. O.
+ + + + + + + + + + + +
+ +
+ + + + +
+ + + + + + + + + + + + + + + + + + + + + + +
+
+
+ + + + +
+ + + + + + + +
+ + + + + + + + + +
+ + + + +
+ + + + + + +
+
Microfungi on Urtica. A. S. Yadav and M. F. Madelin
253
defined (Yadav, 1966) are here employed. The microflora on the stems in the year after death was as follows: Very common: Phoma acuta Fuckel (100%). Occasional: Cladosporium herbarum Link ex Fr. (37 %), Alternaria tenuis Nees ex Pers. (31 'Yo), Periconia cookei Mason & Ellis (27 'Yo), Leptosphaeria acuta (Hoffm. ex Fr.) Karst (26%), Pleurophragmium simplex (Berk. & Br.) Hughes (26%), Dendryphium comosum Wallr. (23 %) and Torula herbarum Link ex Fr. (22 %). The remaining thirty-six species recorded were rare. It is noteworthy that no species fell in the categories of frequent (41-60 %) or common (61-80%). During the winter period from November 1957 to April 1958, in which twenty-four stems were studied, the principal components of the microflora were: Very common: P. acuta (100%). Frequent: Cladosporium sphaerospermum Penzig (46 %), T. herbarum (46 %), P. cookei (42 %), A. tenuis (41 'Yo), and C. herbarum (41 %). Occasional: Cylindrocolla urticae (Pers. ex Fr.) Bon. (25 'Yo), and L. acuta (25 %). . In the summer period from May 1958 until October 1958, during which sixty-four stems were examined, the microflora showed principally quantitative changes in its major components: Very common: P. acuta (100%). Occasional: C. herbarum (35 %), P. simplex (33 %), D. comosum (31 %), A. tenuis (27 %), and L. acuta (27 %). D. comosum and P. simplex, both rare during the winter period (4'2 and 8'3 % respectively) now became occasional (31 and 33 %), while C. urticae which was formerly occasional (25 %) was absent. Its disappearance was accompanied by the appearance of its apothecial state, Calloria fusariodes (Berk.) Fr. Alternaria tenuis, C. herbarum, C. sphaerospermum, P. cookei and T. herbarum all declined in frequency during the year, but P. acuta and its perithecial state, L. acuta (Grove, 1935), remained more or less uniformly frequent. The data in Table 1 on the frequency ofoccurrence ofsporulating species on different segments show that they fall into one or other of three different categories ofdistribution oftheir reproductive structures: (i) more or less uniformly distributed; (ii) more frequent on upper segments; (iii) more frequent on lower segments. Of those species which were at least 10% frequent on one segment of the stem, C, sphaerospermum, D. comosum and P. cookei were more or less uniformly distributed; A. tenuis and C. herbarum were much more frequent on upper segments; and L. acuta, Leptosphaeria doliolum (Pers. ex Fr.) de Not., P. simplex and T. herbarum were much more frequent on lower ones. P. acuta increased from frequent above to common lower down the stem.
Monthly occurrence Table 2 presents the records made each month between November 1957 and October 1958 of the species sporulating on decaying stems taken from
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the four sites. It shows that six species were already sporulating on the stems in November of the year in which the stems grew, and that fourteen more species were evident in December. From January to April, only five new species appeared. However, from May onwards, the microflora changed rapidly, nineteen species being seen to sporulate then for the first time.
Extent and intensity of occurrence of reproductive structures of important members of the primary microflora Because of their widespread occurrence and high abundance on at least some parts of the stems, certain species (listed in Table 3) were more closely investigated. The presence of these species on the stems is expressed in terms of their extent and, in certain instances, the intensity with which they produced reproductive structures. Table 3. Frequency ofoccurrence ofsix important members of theprimary microflora and the percentage mean area covered by them on those segments (A-C) of Urtica dioica on which they were present No. of segments with fungus (maximum of 30)
Percentage mean area covered
,------A-------
,----'-----.
A
Torula herbarum Leptosphaeria acuta Phoma acuta Cladosporium sphaerospermum C. herbarum Alternaria tenuis
7 8 28 8 1 0
B
C
A
B
C
4 2 27 7 3
0 0 20 6 10 6
36 33 86 53 1·6 0
14 56 81 89 75 12
0 0
1
95 72 45 55
To determine their mean extents, measurements were made of the areas over which each species produced reproductive structures more or less continuously on each segment of thirty stems selected at random from the eighty-eight employed in this study. Sixteen stems were drawn from the winter half of the year and fourteen from the summer half. The percentage mean area covered was calculated as follows: total area covered by the species on all segments A (B or C) -- - - - - total surface area of those segments A (B or C) bearing the species -
X 100
.
The result in Table 3 shows that T. herbarum which sporulated more frequently at the base of the stem (see Table I) was also more extensive there, while A. tenuis and C. herbarum which were rarest below were similarly least extensive below. P. acuta covered a large proportion of the available area on all parts of the stem. Of the four species which cover appreciable areas of the top segment (Table 3), P. acuta is on average the most extensive but, as Table 4 shows, on individual segments anyone of the four species may be the most extensive. The intensities with which reproductive structures were produced were determined for five of the six species whose extent had been measured. T. herbarum was omitted because the way it forms spores makes it virtually
Microfungi on Urtica. A. S. Yadav and M. F. Made/in
255
impossible to assess by the direct visual methods employed. The data for the three segments in ascending order are P. acuta 47, 59, and 79, and L. acuta 17, 4, and 0 fructifications per 78'5 mm"; Cladosporium sphaerospermum 5, 4, and 5, C. herbarum 4, 6, and 6, and A. tenuis 0, 5, and 6 tufts of conidiophores per 0'102 mm-. Thus while P. acuta was more frequent on the lower parts of the stems, its pycnidia were produced more densely on the upper parts. Its perithecial state, L. acuta was similarly more frequent on the lowest part of the stem, but by contrast was also more dense there. A. tenuis and C. herbarum sporulated more intensely on segment C, on which both occurred more frequently than elsewhere. C. sphaerospermum, which was more or less uniformly frequent on the three segments (Table I), was also uniformly dense. Table 4. The areas (mm2 ) occupied byfungi on the top segments of ten representative individual stems qf U rtica dioica Stem
Area of segment C
Area occupied
, P. acuta
I
1200
1200
2
3975 3000 30 0 0 340 0 3600 340 0 3600 1600 35 0 0
3975 3000 3000 0 36 0 0 900 0 1600 1000
3 4
5 6 7
8 9 10
, A. tenuis C. herbarum C. sphaerospermum 1200 0 0 0 0 0 2000 0 30 0 35 0 0
0
0 0 0 1000 1000 1000 340 0 3600 0
3975 15 0 0 0 150 0 0 150 0 0 300 15 0 0
0
Table 5. Species isolated from stems, at different stages of decay, grown in 1958 and 1957 Date of isolation
, Oct.
Jan.
Mar. July
Aug.
1958
1958 1959 1959 1958 stems
Alternariatenuis Fusarium sp. Mucor hiemalis Weymer Mucorsp. Aureobasidium pullulans (de Bary) Arn. Pythium intermedium de Bary Rhizopus arrhizus Fischer Trichoderma koningii Oudem.
~
+ +
+
+ + + + +
+ + + +
Oct.
1958
1958
195 8
1957
,stems
I
+ + + + +
, Sept.
+ + + +
+ + + + + +
+ + + +
+
Survey of species in different internodes by isolation Isolations were made from thoroughly washed fragments of seven stems representing different stages of decay. The methods of sampling and isolating were those employed by Yadav (1966). The data are presented (Table 5) in a sequence representing stems dead for increasingly long periods, even though the chronological sequence of isolation was different. The results show principally that: (a) Six fungi were detected which had not been seen on the stems;
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Mucor hiemalis Weymer, Mucor sp., Aureobasidium pullulans (de Bary) Arn., Pythium intermedium de Bary, Rhizopus arrhizus Fischer, and Trichoderma koningii Oudem. (b) Though absent in October, M. hiemalis was consistently present after January. (c) A. tenuis and Fusarium sp. were present throughout the year. The number of occasions out of a possible maximum of seven on which each species was isolated from the stem as a whole and from individual segments is presented in Table 6. It is notable that A. tenuis was isolated from two out of seven basal segments even though its conidiophores had never been seen in this position. Mucor hiemalis and Fusarium sp. were widespread despite the absence or rarity of their reproductive structures. These results for these three moulds are very similar to those for these same species on Heracleum sphondylium (Yadav, 1966). Table 6. The number of occasions that different species were detected by isolation in artificial culture from decaying stems and segments of stems of Urtica dioica (maximum of seven occasions)
.
Segments
Alternaria tenuis Fusarium sp. * Mucor kiemalis Mucor sp. Aureobasidium pullulans Pythium intermedium Rhizopus arrhizus
Trichoderma koningii
Stems
A
B
5
2
2
7 6
5 5
7
2 1
2 1
4
4
1 1
0
1
C
4 6
5
6
1 1 2 0 0
0 1 1 0 [
* This isolate was identical with the Fusarium sp. isolated from Heracleum sphondylium (Yadav, 1966). Fungi observed on moribund and very recently dead stems Shoots of U. dioica were gathered from the three field sites between July and September 1959, in order to observe which were the first species of fungi to sporulate on them. InJuly the shoots were all alive and green and showed no sign of fungal attack. Although some leaves which were formed early in the season and were low upon the stems did become moribund during the growing season, they normally were shed from the stems by a clean abscission, and were replaced by leaves arising from their axillary buds. The adjoining parts of the stem remained healthy. Fungi were first seen sporulating on shoots gathered in August. These had begun to die back. Their upper segments and leaves attached thereto were first colonized by A. tenuis and C. herbarum, The leaves of the middle and lower segments of some of the stems showed dead areas which contained a sterile mycelium. By September, A. tenuis and C. herbarum, together with Epicoccum nigrum Link, had extended downwards to the leaves and even to the stems of the middle (B) segments. C. herbarum and E. nigrum appeared on the lowest segments of the shoots by September, but A. tenuis never appeared there. Meanwhile, Botrytis cinerea Pers. ex Fr. appeared in the microflora of the upper shoot segments.
Microfungi on Urtica. A. S. Yadav and M . F . Made/in
257
This sequence of appearances of these fungi was reflected in the distribution of sporulating species on ten shoots which had died back as far as the base of the stems, and which were collected in August and September 19S8 (Table 7). These bore the four pioneer species already mentioned whose successive appearances were basipetal, together with C. sphaerospermum which was more or less uniformly frequent; and P. acuta which was most frequent on the lower parts of the stem and which appeared to be extending gradually upwards. Table 7. Number of occurrences of species offungi present on ten recently, but entirely, dead stems collected during August and September 1958 (maximum of ten occurrences) Segments A
Alternaria tenuis Botrytis cinerea Cladosporium herbarum C. sphaerospermum Epicoccum nigrum Phoma acuta
A
B
0 0
2 2
I
6 7 5 6
5 I
8
C 9
4 6 6 6 3
DISCUSSION
A picture of the succession and distribution of reproductive structures of microfungi on decaying stems of U. dioica may be constructed from the data presented above. The first shoots to die back began to do so in August, the leaves dying before the stems. On most stems a wave of colonization by A. tenuis and/or C. herbarum proceeded basipetally, usually closely followed by E. nigrum and B. cinerea. This initial phase of colonization of the dying shoots was very similar to that of the culms of Dactylis glomerata L., where C. herbarum, A. tenuis and Epicoccum purpurascens (= E. nigrum) were dominant members of the flora during the decline of vegetative activity in August (Webster, 1956). C. herbarum, which Webster found to be a vigorous primary saprophyte of moribund tissues, progressed up the stems of Dactylis as the leaves unfolded, colonizing them as they became senescent. In U. dioica, senescence and the subsequent appearance of C. herbarum proceeded basipetally. Unlike the pioneer species with which it was associated on Urtica dioica, A. tenuis never sporulated on the lower part of the stem, even though its mycelium was detected there in some stems by isolation. In September C. sphaerospermum and P. acuta appeared. Perhaps because the stem was by then completely moribund, these species did not display a basipetal sequence of appearances. Until December there were only six species visible on the stems but it has been shown that other mycelia were in fact present; namely, Fusarium sp. and Aureobasidium pullulans, Of these pioneer colonizers, P. acuta was seen on every stem examined, being by far the most frequent species. In December fourteen new species appeared. During the 6 months from November to April inclusive the microflora was composed principally of Phoma acuta (100 % frequency), C. sphaerospermum and T . herbarum (46 % ), Periconia cookei (42 % ), A. tenuis and C. herbarum (41 %), 17
Myc.5 1
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Cylindrocolla urticae and L. acuta (both 25 %). Nineteen more species oflow individual frequency appeared for the first time between May and October inclusive. It may be significant that, of the species which pioneered the colonization of dying shoots, those whose spread was basipetal (A. tenuis, B. cinerea, C. herbarum and E. nigrum) were all dry-spored. So too was C. sphaerospermum which established itself more or less uniformly over the stem. However, P. acuta, which appeared on the lower parts of the stems when the latter had died back to their bases, and then appeared to spread upwards, was slimy-spored. The microflora recorded throughout the year was readily divisible into primary and secondary parts, like that on Heracleum, on the basis ofwhether or not the species sporulated on the stems in the year in which the latter had grown. The two microfloras were separated in time by a quiescent period in which reproductive structures of very few new species appeared (only five species between January and April). The precise boundary between the appearances of the reproductive structures of members of the primary and secondary microfloras is somewhat arbitrary, yet it nevertheless serves to divide the population of colonizing species into what appear to be two distinct biological group s. The primary microflora species sporulated rapidly on the moribund or newly dead shoots. They commonly persisted throughout the winter and the following season, and indeed in some instances attained their highest development in the second summer period (for example Phoma herbarum, Leptosphaeria doliolum, Pleurophragmium simplex and Dendryphium comosum). The secondary microflora was composed principally of species which sporulated rather late in the course of decay, usually from May onwards, although it is possible, even probable, that they invaded the stems appreciably earlier. As with H. sphondylium (Yadav, 1966) and Agropyron repens (Hudson & Webster, 1958), so too with Urtica dioica it was found that there were more species of fungus present than were seen to sporulate on the stems. Six microfungi were detected only by isolation. Of these, five were species which had been seen on, or isolated from, stems of H. sphondylium also. It was only by isolation that four phycomycetes were detected on the stems. It is interesting that these could not be isolated in the earliest phase of decay, i.e. in October, but could after January. Hudson & Webster (1958) drew attention to the fact that though mucoraceous fungi, penicilli and aspergilli are frequently recorded from the soil, they were very infrequently recorded from stems of Agropyron repens, even very old ones which had lain on the soil for several months. They concluded that during this phase of decay these organisms were relatively unimportant. In the present authors' studies of stems of H. sphondylium and of U. dioica, no species of Penicillium or Aspergillus were observed or isolated. In respect of these fungi, we concur with Hudson and Webster's conclusion. However, mucoraceous fungi have been isolated from thoroughly washed pieces of stems of both of the dicotyledonous herbaceous species studied, which suggests that these fungi play some part in their decomposition, although the magnitudes of their roles are quite unknown. Because they are species presumably incap-
Microfungi on Urtica. A. S. Yadav and M. F. Madelin
259
able of cellulose decomposition, their roles may be secondary to those of cellulolytic species. A feature of the microflora of U. dioica stems which contrasts with that of H. sphondylium (Yadav, 1966) was the variability of the microfloras on different individual nettle stems. Whereas on H. sphondylium there were nine different species which sporulated on over 50 % of the stems sampled in the period November to April, there was only one which did so on U. dioica, viz. P. acuta (100 %). There were in fact only six fungi on nettle stems that occurred with more than 40 % frequency. Several of the early colonizers showed definite patterns of distribution of their reproductive structures on the stems. Thus P. acuta was rather more frequent on the lower parts of the stems, and was approximately uniformly extensive on all the segments on which it occurred, but it fructified more intensively above. However, its ascigerous stage, L. acuta, was not only more frequent, but also more extensive (see Table 3) and more densely fructifying in the lower parts of the stem. In all three measurements of presence it ranked much lower than P. acuta. Also more frequent and extensive below was Torula herbarum, This species was never seen on the top third of the stems, and only very rarely and restrictedly on the middle one. Leptosphaeria doliolum and Pleurophragmium simplex were also commonest below, though recorded on all three segments of the stems. A. tenuis sporulated only on the mid and upper segments of the stem, but was more frequent, extensive and intensive at the top. C. herbarum was similar except that it did sporulate, albeit rarely and sparsely, on the lowest segment. In contrast with these species, C. sphaerospermum was essentially uniformly frequent and intensive on all three segments, though rather more extensive on the middle one. Some experimental studies which bear upon the differential distributions of some of these fungi will be reported separately. We gratefully acknowledge the facilities provided in the herbarium of the Commonwealth Mycological Institute for comparing specimens, and the help received from Mr E. W. Mason, Dr G. C. Ainsworth, Dr M. B. Ellis, Dr C. Booth, Mr J.J. Elphick, and Dr A. C. Hayward. We wish also to thank Dr J. Gremmen and Dr R. W. G. Dennis for identifying certain discomycetes. REFERENCES
E. (1935). British stem and leaffungi (Coelomycetes), vol. 1,488 pp. Cambridge University Press. HUDSON, H. J. & WEBSTER, J. (1958). Succession of Fungi on decaying stems of Agropyron repens. Trans. Br. mycol, Soc. 41, 165-177. WEBSTER, J. (1956). Succession of Fungi on decaying cocksfoot culms. I. ]. Ecol. 44, 517544· YADAV, A. S. (1966). The ecology of microfungi on decaying stems of Heracleum sphondylium. Trans. Br. mycol. Soc. 49, 471-485. GROVE, W.
(Accepted for publication 7 March 1967)
17-2
THE ECOLOGY OF MICROFUNGI ON DECAYING STEMS OF URTICA DIOICA By A. S. YADAV
Department
of Botany, Science College, Patna, India AND
Department
M. F. MADELIN
ofBotany, The University, Bristol
The succession and distribution of microfungi associated with stems of Urtica
dioica L. decaying above ground have been studied. The percentage frequency of occurrence of species of fungi sporulating on the stems has been determined, and the intensity ofsporulation ofsome ofthese fungi and the percentage ofstem area covered by them have also been estimated. Certain fungus species show definite patterns of distribution on the stems. A primary microflora, consisting of species visible on the stems in the year ofth eir growth, and a secondary microflora, comprizing species appearing in the winter and following summer, may be recognized . A quiescent period, in which very few new species appeared, intervened. All the species discovered by isolation from the decayingstems had also been isolated previously from Heracleum sphondylium L. (Yad av, 1966). However Acremoniella atra (Corda) Sacco which was common on H. sphondylium was never isolated from U. dioica, Colonization of the moribund shoots started from the top. Alternaria tenuis Nees. ex Pers., Cladosporium herbarum Link ex Fr., Epicoccum nigrum Link, and Botrytis cinerea Pers, ex Fr. first invaded the fading leaves and adjoining segments. Lower segments later died and soon supported sporulation of these same fungi as well as of Phoma acuta Fuckel and Cladosporium sphaerospermum Penzig. A . tenuis and C. herbarum were rare on the lower segments.
This account of the ecology of microfungi on stems of Urtica dioica L. contributes to our knowledge of the decay of dicotyledonous herbaceous remains. One of the authors has already reported on the decay of stems of Heracleum sphondylium L. (Yadav, 1966). Like the latter plant, U. dioica has erect, slightly woody stems which do not rapidly decay beyond recognition; it is widespread and abundant in the British Isles and harbours a large population of microfungi during the decay of its shoots. MATERIALS AND METHODS
U. dioica is a herb with a perennial, creeping, much-branched root-stock from which in the spring arise erect, hispid, ribbed shoots, 0'84:>'9 or even 1'2 m high, with many rather short internodes. It flowers in summer and autumn, and then dies back, the lower internodes remaining green longest. The general methods of study were the same as those used by Yadav (1966). Stems which had recently died back and which were in the first stages of decay were gathered in October 1957 from fields at Swanley (N.G. Ref. TG 515670) and Eynsford (N.G. Ref. TQ531626) in Kent and placed upon the ground at Chelsea Physic Gardens, London, S.W. 3. Each
25°
Transactions British Mycological Society
month, three to five stems were removed at random from th e pile, cut into three equal segments, labelled A-C in ascending order, and examined as described below. Because of the rather large number of small internodes in these stems, it was not practicable to study quantitatively each internode individually. These collections were supplemented at irregular intervals by others taken from the original field sites at Swanley and Eynsford, as well as one at St Albans, Herts. (N .G. Ref. TL 132042). When gathering the stems, care was taken to ensure that they included the first internode exposed above soil level. The total number of U. dioica stems sampled from all four sources between No vember 1957 and October 1958 was eighty-eight. After collection, stem segments were incubated individually at high humidity for 2 days to facilitate identification and mapping of the sporulating fungal colonies. Then for each segment records were made of the following: (I) Total surface area of segment. (2) Names of all species of microfungi present. (3) Total area covered by each of those species which experience had shown to be important (Table 3). (4) Intensity of production of reproductive structures by certain important species. For pyrenomycetes, this was expressed as the mean number of ascocarps per field of a binocular dissecting microscope (78'5 mm") determined in the way described for pycnidia by Yadav (1966) (see this reference also for relevant methods for hyphomycetes). Although the principal site from which samples were drawn was artificially created, a number of observations indicated that the microflora on stems in this situation was substantially the same as that in natural habitats. First, the fungus microfloras of stems from the four different sampling sites were qualitatively alike in respect of all their principal components. The sixteen species which occurred on six or more of the eighty-eight stems studied quantitatively were found at all four sites. The microflora on stems from the artificial pile at Chelsea resembled that from the three natural sites in that, of the total of forty-four species observed on Urtica stems, thirty-nine were recorded at Chelsea. Conversely, only eight of these species at Chelsea were not collected also from natural sites elsewhere. Secondly, the rnicroflora in the year in which the principal quantitative studies were made closely resembled that in a second year of observations on stems from natural sites. Of the forty-four species seen in the first year, thirty-two were seen also at field sites in the second year. The twelve not seen were species which had been rare in the first season. Although the foregoing evidence indicates only a qualitative similarity between the microfloras of the artificial and natural sites, the succession and development of the geographically separated microfloras were not noticed to be significantly different. However, such differences as might exist would admittedly not be revealed by the quantitative data presented below which are based on a sampling procedure which combined the samples drawn from different locations.
Microfungi on Urtica. A. S. Yadav and M. F. Madelin
25 1
Table I. Percentage frequencies at various periods recorded for diiferent species offungi on decaying stems of U rtica dioica which had grown in 1957 On whole stems (
'I
Nov, 57- Nov, 57- May 58Oct. 58 Apr, 58 Oct, 58
Alternaria tenuis N ees ex Pers. Botryosphaeria sp. Botrytis carnea Schum. B, cinerea Pers. ex Fr. Calloria fusarioides (Berk,) Fr, Camposporium peliucidum (Grove) Hughes Circinotrichum sp. Cladosporium herbarum Link ex Fr. C. sphaerospermum Penzig Coniothyrium conoideum Sacc, Cryptodidymosphaeria conoidea (Niessl) Rehm Cylindrocolla urticae (Pers. ex Fr.) Bon, Dasyscypha sulphurea (Pers. ex Fr.) Massee Dendryphium comosum Wallr. D, griseum Berk, & Br. D, laxum Berk, & n-. Dictyosporium toruloides (Corda) Gueg. Dinemasporium hispidulum Sacc, Epicoccum nigrum Link Fusarium sambucinum Fuckel Fusarium sp. Graphium cuneiferum (Berk, & Br.) Mason & Ellis Helotium herbarum (Pers, ex Fr.) Fr. Leptosphaeriaacuia (Hoffm. ex Fr.) Karst, L. doliolum (Pers. ex Fr,) deNot. Oospora sp, Ophiobolus erythrosporus (Riess) Wint. 0, immersus Trail Paecilomyces betae (Delacr.) Cornford Periconia byssoides Pers, ex Corda P. cookei Mason & Ellis Phaeoisaria cornui (Bainier) Mason Phialea cyathoidea (Bull, ex Fr.) Gill, Phoma acuta Fuckel P, berkeleyi Sacc, P, herbarum West, P, urticae Sch, & Sacc, Pleurophragmium simplex (Berk, & Br,) Hughes Pyrenopeziza urticicola (Phil!.) Boud, Torula herbarum Link ex Fr. Trichocladium opacum (Corda) Hughes Trichothecium sp, Tricladium sp. Volutella ciliata Fr.
41 4'2
r-r
0
3'4 6'8
8'3 0 0
27 0 "6 ,,6 9'4 1,6
0 4' 46 8'3 8'3
"6 35 1,6 0 0
3'
,,'
r-r I"
37 '4 2'3 2'3 6,8
25
5'7
0
4'5
3'4 '00 i -r ,6 2'3 26
8'3 13 0 42 0 0 100 4'2 13 8'3 8'3
8'0 22 2'3
0 46 0
3'4 5'7 r-r
27
,,'
r-r
r-r 3'4
0 0 4'2
I"
3'4 I" I"
3'4 8'0 -r
i
r-r
0 5'7 8'0 r-r
,,'
0 0 3' '0 0 0 r-r
7,8
3'4
3'4
0
'3 2'3
6'3 7,8 1,6 3" 1,6
8'3 0 0
0 2'3 4'5 0
3'4
0 8'3 4'2 0 0
r-r
,.,
r-r
5'7
4'5 8'0 2'3 2'3
,6
C 27 0 0 3'4
0
3' ,,6 4'7 3"
0 25
B 3'4
0
4'2 8'3 0 0
2'3 26
A 0
2'3
23 3'4 3'4 2'3
,,'
On individual segments, Nov, '957-0ct, '958
r-r
II
3'5
,,'
9'0 0 3'4
i -t
0
r-r
2'3
4'5 3'4
I"
0
I"
0
0
r-r
i
-r
i-r
r-r
2'3 5'7 0
3" 27
2'3 21
1'1 4'5
0 2'3
19 ,,6 6'3
'4
4'5
2'3 0
"6 3'1 ,,6 20 1,6 4'7 '00 0 '7 0 33
r-r
2'3
,., 3'5 ,,' 9'7 I"
3'4 77 r-r
8'0 2'3 24
0
i
-r
2'3
,,'
2'3
"I
r-r
0 8'5 0
0 II
5'7
0 0 56 0 g'o 0 6,8
6,8 2'3 0
4'5 0 0 0
,,'
65 0 8'0 r-r
'3 3'1
8'0 20 2'3
"6 "6 3"
r-r
0
0 3'4
r-r
r-r
0
0
II
Transactions British Mycological Society
25 2
RESULTS
Succession and distribution rif species riffungi sporulating on decaying stems Frequency of occurrence Eighty-eight stems were gathered between November 1957 and October 1958 from all four sites. In Table I are shown the frequencies with which the forty-four fungal species seen occurred on the stems throughout the whole year and within the two component half-year periods. The table also shows the frequencies of occurrence throughout the year on the three segments separately. To facilitate description, the named categories of frequencies previously Table
2.
Species
riffungi recorded each month on stems rif Urtica dioica,
November 195 7-0ctober 1958 N. D. J. F. M. A. Alternaria tenuis + + + + Botrytis cinerea + + Phoma acuta + + + + + + Cladosporium herbarum + + + + Periconia cookei + + Epicoccum nigrum + + Coniotbyrium conoideum + Ophiobolus immersus + Phoma berkeleyi + Cladosporium sphaerospermum + + + + Paecilomy ces betae + + Dinemasporium hispidulum + + Phoma herbarum + + + Cryptodidymosphaeria conoidea + Dendryphium griseum + + Leptosphaeria acuta + + + Torula herbarum + + Leptosphaeria doliolum + Pleurophragmium simplex + + Dendryphium comosum + Botryosphaeria sp. + Cylindrocolla urticae + + + + Fusarium sambucinum + Volutella ciliata + Phoma urticae + Calloriafusarioides Fusarium sp. Phialea cyathoidea Ophiobolus erythrosporus Graphium cuneiferum Pyrenopeziza urticicola Periconiabyssoides Phaeoisaria cornui Circinotrichum sp. Trichothecium sp. Trichocladium opacum Dendryphium laxum Camposporium pellucidum Tricladium sp . Dasyscypha sulphurea
Dictyosporium toruloides Helotium hetbarum Botrytis carnea Oospora sp.
M. J.
J. + + + + + + + + + + + +
A. S. O.
+ + + + + + + + + + + +
+ +
+ + + + +
+ + + + + + + + + + + + + + + + + + + + + + +
+
+
+ + + + +
+ + + + + + + +
+ + + + + + + + + +
+ + + + +
+ + + + + + +
+
Microfungi on Urtica. A. S. Yadav and M. F. Madelin
253
defined (Yadav, 1966) are here employed. The microflora on the stems in the year after death was as follows: Very common: Phoma acuta Fuckel (100%). Occasional: Cladosporium herbarum Link ex Fr. (37 %), Alternaria tenuis Nees ex Pers. (31 'Yo), Periconia cookei Mason & Ellis (27 'Yo), Leptosphaeria acuta (Hoffm. ex Fr.) Karst (26%), Pleurophragmium simplex (Berk. & Br.) Hughes (26%), Dendryphium comosum Wallr. (23 %) and Torula herbarum Link ex Fr. (22 %). The remaining thirty-six species recorded were rare. It is noteworthy that no species fell in the categories of frequent (41-60 %) or common (61-80%). During the winter period from November 1957 to April 1958, in which twenty-four stems were studied, the principal components of the microflora were: Very common: P. acuta (100%). Frequent: Cladosporium sphaerospermum Penzig (46 %), T. herbarum (46 %), P. cookei (42 %), A. tenuis (41 'Yo), and C. herbarum (41 %). Occasional: Cylindrocolla urticae (Pers. ex Fr.) Bon. (25 'Yo), and L. acuta (25 %). . In the summer period from May 1958 until October 1958, during which sixty-four stems were examined, the microflora showed principally quantitative changes in its major components: Very common: P. acuta (100%). Occasional: C. herbarum (35 %), P. simplex (33 %), D. comosum (31 %), A. tenuis (27 %), and L. acuta (27 %). D. comosum and P. simplex, both rare during the winter period (4'2 and 8'3 % respectively) now became occasional (31 and 33 %), while C. urticae which was formerly occasional (25 %) was absent. Its disappearance was accompanied by the appearance of its apothecial state, Calloria fusariodes (Berk.) Fr. Alternaria tenuis, C. herbarum, C. sphaerospermum, P. cookei and T. herbarum all declined in frequency during the year, but P. acuta and its perithecial state, L. acuta (Grove, 1935), remained more or less uniformly frequent. The data in Table 1 on the frequency ofoccurrence ofsporulating species on different segments show that they fall into one or other of three different categories ofdistribution oftheir reproductive structures: (i) more or less uniformly distributed; (ii) more frequent on upper segments; (iii) more frequent on lower segments. Of those species which were at least 10% frequent on one segment of the stem, C, sphaerospermum, D. comosum and P. cookei were more or less uniformly distributed; A. tenuis and C. herbarum were much more frequent on upper segments; and L. acuta, Leptosphaeria doliolum (Pers. ex Fr.) de Not., P. simplex and T. herbarum were much more frequent on lower ones. P. acuta increased from frequent above to common lower down the stem.
Monthly occurrence Table 2 presents the records made each month between November 1957 and October 1958 of the species sporulating on decaying stems taken from
254
Transactions British Mycological Society
the four sites. It shows that six species were already sporulating on the stems in November of the year in which the stems grew, and that fourteen more species were evident in December. From January to April, only five new species appeared. However, from May onwards, the microflora changed rapidly, nineteen species being seen to sporulate then for the first time.
Extent and intensity of occurrence of reproductive structures of important members of the primary microflora Because of their widespread occurrence and high abundance on at least some parts of the stems, certain species (listed in Table 3) were more closely investigated. The presence of these species on the stems is expressed in terms of their extent and, in certain instances, the intensity with which they produced reproductive structures. Table 3. Frequency ofoccurrence ofsix important members of theprimary microflora and the percentage mean area covered by them on those segments (A-C) of Urtica dioica on which they were present No. of segments with fungus (maximum of 30)
Percentage mean area covered
,------A-------
,----'-----.
A
Torula herbarum Leptosphaeria acuta Phoma acuta Cladosporium sphaerospermum C. herbarum Alternaria tenuis
7 8 28 8 1 0
B
C
A
B
C
4 2 27 7 3
0 0 20 6 10 6
36 33 86 53 1·6 0
14 56 81 89 75 12
0 0
1
95 72 45 55
To determine their mean extents, measurements were made of the areas over which each species produced reproductive structures more or less continuously on each segment of thirty stems selected at random from the eighty-eight employed in this study. Sixteen stems were drawn from the winter half of the year and fourteen from the summer half. The percentage mean area covered was calculated as follows: total area covered by the species on all segments A (B or C) -- - - - - total surface area of those segments A (B or C) bearing the species -
X 100
.
The result in Table 3 shows that T. herbarum which sporulated more frequently at the base of the stem (see Table I) was also more extensive there, while A. tenuis and C. herbarum which were rarest below were similarly least extensive below. P. acuta covered a large proportion of the available area on all parts of the stem. Of the four species which cover appreciable areas of the top segment (Table 3), P. acuta is on average the most extensive but, as Table 4 shows, on individual segments anyone of the four species may be the most extensive. The intensities with which reproductive structures were produced were determined for five of the six species whose extent had been measured. T. herbarum was omitted because the way it forms spores makes it virtually
Microfungi on Urtica. A. S. Yadav and M. F. Made/in
255
impossible to assess by the direct visual methods employed. The data for the three segments in ascending order are P. acuta 47, 59, and 79, and L. acuta 17, 4, and 0 fructifications per 78'5 mm"; Cladosporium sphaerospermum 5, 4, and 5, C. herbarum 4, 6, and 6, and A. tenuis 0, 5, and 6 tufts of conidiophores per 0'102 mm-. Thus while P. acuta was more frequent on the lower parts of the stems, its pycnidia were produced more densely on the upper parts. Its perithecial state, L. acuta was similarly more frequent on the lowest part of the stem, but by contrast was also more dense there. A. tenuis and C. herbarum sporulated more intensely on segment C, on which both occurred more frequently than elsewhere. C. sphaerospermum, which was more or less uniformly frequent on the three segments (Table I), was also uniformly dense. Table 4. The areas (mm2 ) occupied byfungi on the top segments of ten representative individual stems qf U rtica dioica Stem
Area of segment C
Area occupied
, P. acuta
I
1200
1200
2
3975 3000 30 0 0 340 0 3600 340 0 3600 1600 35 0 0
3975 3000 3000 0 36 0 0 900 0 1600 1000
3 4
5 6 7
8 9 10
, A. tenuis C. herbarum C. sphaerospermum 1200 0 0 0 0 0 2000 0 30 0 35 0 0
0
0 0 0 1000 1000 1000 340 0 3600 0
3975 15 0 0 0 150 0 0 150 0 0 300 15 0 0
0
Table 5. Species isolated from stems, at different stages of decay, grown in 1958 and 1957 Date of isolation
, Oct.
Jan.
Mar. July
Aug.
1958
1958 1959 1959 1958 stems
Alternariatenuis Fusarium sp. Mucor hiemalis Weymer Mucorsp. Aureobasidium pullulans (de Bary) Arn. Pythium intermedium de Bary Rhizopus arrhizus Fischer Trichoderma koningii Oudem.
~
+ +
+
+ + + + +
+ + + +
Oct.
1958
1958
195 8
1957
,stems
I
+ + + + +
, Sept.
+ + + +
+ + + + + +
+ + + +
+
Survey of species in different internodes by isolation Isolations were made from thoroughly washed fragments of seven stems representing different stages of decay. The methods of sampling and isolating were those employed by Yadav (1966). The data are presented (Table 5) in a sequence representing stems dead for increasingly long periods, even though the chronological sequence of isolation was different. The results show principally that: (a) Six fungi were detected which had not been seen on the stems;
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Mucor hiemalis Weymer, Mucor sp., Aureobasidium pullulans (de Bary) Arn., Pythium intermedium de Bary, Rhizopus arrhizus Fischer, and Trichoderma koningii Oudem. (b) Though absent in October, M. hiemalis was consistently present after January. (c) A. tenuis and Fusarium sp. were present throughout the year. The number of occasions out of a possible maximum of seven on which each species was isolated from the stem as a whole and from individual segments is presented in Table 6. It is notable that A. tenuis was isolated from two out of seven basal segments even though its conidiophores had never been seen in this position. Mucor hiemalis and Fusarium sp. were widespread despite the absence or rarity of their reproductive structures. These results for these three moulds are very similar to those for these same species on Heracleum sphondylium (Yadav, 1966). Table 6. The number of occasions that different species were detected by isolation in artificial culture from decaying stems and segments of stems of Urtica dioica (maximum of seven occasions)
.
Segments
Alternaria tenuis Fusarium sp. * Mucor kiemalis Mucor sp. Aureobasidium pullulans Pythium intermedium Rhizopus arrhizus
Trichoderma koningii
Stems
A
B
5
2
2
7 6
5 5
7
2 1
2 1
4
4
1 1
0
1
C
4 6
5
6
1 1 2 0 0
0 1 1 0 [
* This isolate was identical with the Fusarium sp. isolated from Heracleum sphondylium (Yadav, 1966). Fungi observed on moribund and very recently dead stems Shoots of U. dioica were gathered from the three field sites between July and September 1959, in order to observe which were the first species of fungi to sporulate on them. InJuly the shoots were all alive and green and showed no sign of fungal attack. Although some leaves which were formed early in the season and were low upon the stems did become moribund during the growing season, they normally were shed from the stems by a clean abscission, and were replaced by leaves arising from their axillary buds. The adjoining parts of the stem remained healthy. Fungi were first seen sporulating on shoots gathered in August. These had begun to die back. Their upper segments and leaves attached thereto were first colonized by A. tenuis and C. herbarum, The leaves of the middle and lower segments of some of the stems showed dead areas which contained a sterile mycelium. By September, A. tenuis and C. herbarum, together with Epicoccum nigrum Link, had extended downwards to the leaves and even to the stems of the middle (B) segments. C. herbarum and E. nigrum appeared on the lowest segments of the shoots by September, but A. tenuis never appeared there. Meanwhile, Botrytis cinerea Pers. ex Fr. appeared in the microflora of the upper shoot segments.
Microfungi on Urtica. A. S. Yadav and M . F . Made/in
257
This sequence of appearances of these fungi was reflected in the distribution of sporulating species on ten shoots which had died back as far as the base of the stems, and which were collected in August and September 19S8 (Table 7). These bore the four pioneer species already mentioned whose successive appearances were basipetal, together with C. sphaerospermum which was more or less uniformly frequent; and P. acuta which was most frequent on the lower parts of the stem and which appeared to be extending gradually upwards. Table 7. Number of occurrences of species offungi present on ten recently, but entirely, dead stems collected during August and September 1958 (maximum of ten occurrences) Segments A
Alternaria tenuis Botrytis cinerea Cladosporium herbarum C. sphaerospermum Epicoccum nigrum Phoma acuta
A
B
0 0
2 2
I
6 7 5 6
5 I
8
C 9
4 6 6 6 3
DISCUSSION
A picture of the succession and distribution of reproductive structures of microfungi on decaying stems of U. dioica may be constructed from the data presented above. The first shoots to die back began to do so in August, the leaves dying before the stems. On most stems a wave of colonization by A. tenuis and/or C. herbarum proceeded basipetally, usually closely followed by E. nigrum and B. cinerea. This initial phase of colonization of the dying shoots was very similar to that of the culms of Dactylis glomerata L., where C. herbarum, A. tenuis and Epicoccum purpurascens (= E. nigrum) were dominant members of the flora during the decline of vegetative activity in August (Webster, 1956). C. herbarum, which Webster found to be a vigorous primary saprophyte of moribund tissues, progressed up the stems of Dactylis as the leaves unfolded, colonizing them as they became senescent. In U. dioica, senescence and the subsequent appearance of C. herbarum proceeded basipetally. Unlike the pioneer species with which it was associated on Urtica dioica, A. tenuis never sporulated on the lower part of the stem, even though its mycelium was detected there in some stems by isolation. In September C. sphaerospermum and P. acuta appeared. Perhaps because the stem was by then completely moribund, these species did not display a basipetal sequence of appearances. Until December there were only six species visible on the stems but it has been shown that other mycelia were in fact present; namely, Fusarium sp. and Aureobasidium pullulans, Of these pioneer colonizers, P. acuta was seen on every stem examined, being by far the most frequent species. In December fourteen new species appeared. During the 6 months from November to April inclusive the microflora was composed principally of Phoma acuta (100 % frequency), C. sphaerospermum and T . herbarum (46 % ), Periconia cookei (42 % ), A. tenuis and C. herbarum (41 %), 17
Myc.5 1
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Cylindrocolla urticae and L. acuta (both 25 %). Nineteen more species oflow individual frequency appeared for the first time between May and October inclusive. It may be significant that, of the species which pioneered the colonization of dying shoots, those whose spread was basipetal (A. tenuis, B. cinerea, C. herbarum and E. nigrum) were all dry-spored. So too was C. sphaerospermum which established itself more or less uniformly over the stem. However, P. acuta, which appeared on the lower parts of the stems when the latter had died back to their bases, and then appeared to spread upwards, was slimy-spored. The microflora recorded throughout the year was readily divisible into primary and secondary parts, like that on Heracleum, on the basis ofwhether or not the species sporulated on the stems in the year in which the latter had grown. The two microfloras were separated in time by a quiescent period in which reproductive structures of very few new species appeared (only five species between January and April). The precise boundary between the appearances of the reproductive structures of members of the primary and secondary microfloras is somewhat arbitrary, yet it nevertheless serves to divide the population of colonizing species into what appear to be two distinct biological group s. The primary microflora species sporulated rapidly on the moribund or newly dead shoots. They commonly persisted throughout the winter and the following season, and indeed in some instances attained their highest development in the second summer period (for example Phoma herbarum, Leptosphaeria doliolum, Pleurophragmium simplex and Dendryphium comosum). The secondary microflora was composed principally of species which sporulated rather late in the course of decay, usually from May onwards, although it is possible, even probable, that they invaded the stems appreciably earlier. As with H. sphondylium (Yadav, 1966) and Agropyron repens (Hudson & Webster, 1958), so too with Urtica dioica it was found that there were more species of fungus present than were seen to sporulate on the stems. Six microfungi were detected only by isolation. Of these, five were species which had been seen on, or isolated from, stems of H. sphondylium also. It was only by isolation that four phycomycetes were detected on the stems. It is interesting that these could not be isolated in the earliest phase of decay, i.e. in October, but could after January. Hudson & Webster (1958) drew attention to the fact that though mucoraceous fungi, penicilli and aspergilli are frequently recorded from the soil, they were very infrequently recorded from stems of Agropyron repens, even very old ones which had lain on the soil for several months. They concluded that during this phase of decay these organisms were relatively unimportant. In the present authors' studies of stems of H. sphondylium and of U. dioica, no species of Penicillium or Aspergillus were observed or isolated. In respect of these fungi, we concur with Hudson and Webster's conclusion. However, mucoraceous fungi have been isolated from thoroughly washed pieces of stems of both of the dicotyledonous herbaceous species studied, which suggests that these fungi play some part in their decomposition, although the magnitudes of their roles are quite unknown. Because they are species presumably incap-
Microfungi on Urtica. A. S. Yadav and M. F. Madelin
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able of cellulose decomposition, their roles may be secondary to those of cellulolytic species. A feature of the microflora of U. dioica stems which contrasts with that of H. sphondylium (Yadav, 1966) was the variability of the microfloras on different individual nettle stems. Whereas on H. sphondylium there were nine different species which sporulated on over 50 % of the stems sampled in the period November to April, there was only one which did so on U. dioica, viz. P. acuta (100 %). There were in fact only six fungi on nettle stems that occurred with more than 40 % frequency. Several of the early colonizers showed definite patterns of distribution of their reproductive structures on the stems. Thus P. acuta was rather more frequent on the lower parts of the stems, and was approximately uniformly extensive on all the segments on which it occurred, but it fructified more intensively above. However, its ascigerous stage, L. acuta, was not only more frequent, but also more extensive (see Table 3) and more densely fructifying in the lower parts of the stem. In all three measurements of presence it ranked much lower than P. acuta. Also more frequent and extensive below was Torula herbarum, This species was never seen on the top third of the stems, and only very rarely and restrictedly on the middle one. Leptosphaeria doliolum and Pleurophragmium simplex were also commonest below, though recorded on all three segments of the stems. A. tenuis sporulated only on the mid and upper segments of the stem, but was more frequent, extensive and intensive at the top. C. herbarum was similar except that it did sporulate, albeit rarely and sparsely, on the lowest segment. In contrast with these species, C. sphaerospermum was essentially uniformly frequent and intensive on all three segments, though rather more extensive on the middle one. Some experimental studies which bear upon the differential distributions of some of these fungi will be reported separately. We gratefully acknowledge the facilities provided in the herbarium of the Commonwealth Mycological Institute for comparing specimens, and the help received from Mr E. W. Mason, Dr G. C. Ainsworth, Dr M. B. Ellis, Dr C. Booth, Mr J.J. Elphick, and Dr A. C. Hayward. We wish also to thank Dr J. Gremmen and Dr R. W. G. Dennis for identifying certain discomycetes. REFERENCES
E. (1935). British stem and leaffungi (Coelomycetes), vol. 1,488 pp. Cambridge University Press. HUDSON, H. J. & WEBSTER, J. (1958). Succession of Fungi on decaying stems of Agropyron repens. Trans. Br. mycol, Soc. 41, 165-177. WEBSTER, J. (1956). Succession of Fungi on decaying cocksfoot culms. I. ]. Ecol. 44, 517544· YADAV, A. S. (1966). The ecology of microfungi on decaying stems of Heracleum sphondylium. Trans. Br. mycol. Soc. 49, 471-485. GROVE, W.
(Accepted for publication 7 March 1967)
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