- Email: [email protected]
Belowground growth of Spartina alterniflora Loisel.: habit, functional biomass and non-structural carbohydrates
Estuarine, Coastal and Shelf Science 0 9 8 0 x2, 579-587
Belowground Growth of Spartina alterniflora Loisel.: Habit, Functional Biomass and Non-structural Carbohydrates
Diane C. Livingstone and David G.
Patriqttin
Biology Department, Dalhousie University, Halifax, Nova Scotia, B3H 4fit, Canada Received 25 October I979 and in revised form 24 August I98o
Keywords: salt marshes; vegetation; growth rates; age determination; annual cycles; seasonal variations; carbohydrates; Nova Scotia coast Observations on the growth habit of a thatch island Sparthla alterniflora colony in. Nova Scotia indicated that an individual shoot chtmp lives for 2 years and then dies. There were three distinct phases of growth during the year: rapid leaf and root growth in June and July, vigorous rhizome growth and production of ovenvintering tillers concurrent with reproductive development in August and September, and finally, a period of no obvious change from October until the following May. There was a distinct autumn accumulation of non-structural carbohydrates in underground parts, with sugars accounting for 31% of the dry weight in October and starch 4%. Ratios of functional belowground biomass to end of season aboveground biomass were roughly constant for stands more than i year old; the ratio of annual net production of belowground parts to end of season aboveground biomass was estimated as x'75.
Introduction Although the aboveground productivity of saltmarsh angiosperms has been extensively studied (Turner, I976; Jefferies, x977), relatively little is known of the belowground growth habits and production of most species. Recent interest in sulfur metabolism of salt marshes has emphasized the high magnitude of belowground production (Howarth, I979) and has indicated as well, that much of this production may become available aboveground through diffusion of reduced hydrogen acceptors to the sediment surface (Fenchel & Riedl, x97o). There is no generally acceptable technique for estimating belowground production of perennial species ~Iilner & Hughes, x968; Newbould, 1967). Valiela et al. 0976) estimated belowground production of ,S'. alterniflora from the observed increases of dead materials during the growing season. It was assumed that these increases represented turnover of belowground biomass within one season. For fertilized and unfertilized low marsh plots, their estimates of belowground production were in the range of 33x5 to 5687 g dry matter m -2. GaUagher & Plumley (x979) estimated belowground production from the differences between maximum and minimum amounts of total macro-organic matter within one growing season; their estimates varied between 2x8 and 2ioo g dry matter m -2 year -1. There are several reports of aboveground/belowground biomass ratios (Smalley, i959; Seneca, x974; 579 0272-7714/8t/o5o579+to :$02.o0/0
~) t98t Academic Press Inc. (London) Ltd.
580
D. C. Livingstone r D. G. Patriquin
Broome et aL, x975) but particularly for older stands, it is not clear what this biomass represented. Observations on the growth habit of S. alterniflora reported in this study provided a means of separating functional from non-functional belowground biomass in young stands. We examined aboveground]belowground biomass ratios and estimated net belowground production for stands of increasing plant density and age on a recently colonized sandbar in a Nova Scotian sahmarsh. We also investigated the relationship of non-structural carbo9 hydrate (NSC) content to plant growth during a growing season. In Georgia, where year round growth of S. alterniflora occurs, ~IcIntire & Dunstan (i976) found that S. alterniflora retained low levels of NSC throughout the year. They raised the question of whether this represented a genetically fixed characteristic of incorporating most production directly into structural components, or whether in northern regions (such as Nova Scotia) storage of NSC may occur. Materials and methods
The S. alterniflora stand was located on a recently established (within Io years) sandbar at Conrad Beach, on the Atlantic coast near Halifax, Nova Scotia. This stand was actively expanding along its edges, and was similar in general aspect to tile 'thatch islands' described by Redfield (I972). For studies of growth habit and biomass, samples were taken, as described in the text, and later washed with distilled water, dried in a forced air oven at 80 ~ and. weighed. For determination of non-structural carbohydrates, sods were removed from the marsh to a depth of about 3~ cm and plants washed free of soil. Only belowground tissue directly attached to live foliage was collected. Plants were divided it/to three sections: roots and rhizomes, leaves and culms and reproductive parts. Data for reproductive parts are reported in Livingstone (x978). Approximately 5o-Ioo g of each type were sliced into sections a few centimeters in length, mixed, and 5 or xo g snbsamples removed. One Io g sample of each tissue type was dried at 80 ~ and then ground to 60 mesh in a Wiley Mill. Subsamplcs of this matelial were used for the determination of starch content (Hassid & Neufeld, x964; Dubois et al., x956). Another ,o g sample of each tissue type was extracted three times in ioo ml portions of hot 8o% ethanol. Following centrifugation, supernatants were combined and evaporated to dryness in a rotary evaporator at 4 ~ ~ Residues were resuspended in Io ml deionized water and an aliquot of the resuspended extract separated into acidic, basic and neutral fractions by ion-exchange chromatography (Atkins & Canvin, I97x ). Total carbohydrates in the neutral fraction were measured by the total carbon assay of Umbreit et aL (i97z) and expressed as equivalents of glucose (laequiv. glucose (g dry wt)-l), by comparison of absorbance values to a glucose standard curve. Carbohydrates in the neutral fraction were separated by thin-layer chromatography on cellulose MN 300 (25o ~tm thickness) plates in water: ethyl acetate: pyridine (25 : Ioo : 35) (Raadsveld & Klomp, x97 Q. Spots were identified against standards using p-anisidine hydrochloride spray (Pridham, x956). Results and discussion
Growth habit The growth habit of S. alterniflora was examined by carefully digging up connected 'shoot dumps' (Caldwell, x957), beginning with the apical region of recently produced long
Belozcground grorcth of Spartina aherrfiflora
3
581
!
2
o
/ ..
Foliooe in foil of 197,5
Ttller bud in foil of 1975
P
,.__/
Long chizome with filler bud, foil 1975
Slems ond roofs ossociofed with 197.5 folloqe Old deod folioge, produced 1 9 7 4 e~
J
5temso roofs ond long rhizornes Of 1 9 7 4
t t I
Slems, roofs, fohoqe (deod} o n d long rhizomes of 1973
Figure x. Structure of connected shoot clumps in the autumn of x975. rhizomes. The structure of successive shoot clumps examined in the fall is illustrated in Figure x. The youngest clump (o) consisted of a long rhizome which terminated at this time in a single tiller bud or small (less than I I cm length) ovcrwintering shoot. The next clump (x) consisted of several mature shoots (length greater than x8 cm), of which the oldest one or two were usually flowering shoots, and a number of tiller buds or overwintering shoots, and a large mass of roots. Clump (z) included mature shoots as in (x) but few or no buds or overwintering tillers and no recently produced long rhizomes. Basal portions of old dead shoots from the previous year were present in type (z) clumps. Clump (3) consisted of dead shoots only, or occasionally a single living shoot was present. The pattern of occurrence of dead shoots in these clumps suggested that the successive clumps (o), (i), (z), (3) were, in the autumn, respectively o, z, z, and 3 years of age.
582
D. C. Livingstone C.~ D. G. Patriquln
Observations of clump development at 2-week intervals from May to October and on three occasions between October and ~'Iay illustrated three more or less distinct seasonal phases of belowground growth: (i) vigorous root growth at bases of new tillers in June and July, (ii) vigorous rhizome growth and production of overavintering shoots in August and September, and (iii) a period of no obvious change during the winter (Plate x). These periods corresponded respectively to vegetative growth, reproductive development, and senescence of aboveground vegetation (Figure 2).
e ~
~
8
I/'\
p-
J
50(
Ioo
o
V - * / t May
I June
t July
I Aug
I Sept
[al 0r
Jan
l,,l " Apr
Month
Figure 2. Seasonal changes in seawater and sediment temperatures and above-
ground biomass of S. alterniflora; (r seawater, (u) sediment at 4cm depth, data are means of two readings taken at randomly selected sites at ~2.ooh (Atlantic Standard Time); (O) standing crop, data are means of all live biomass on three t/16 m t quadrats_+S.E.M.; ( , ) maximum plant height on three x/i6 m 2 quadrats, data are means 4-s.e..~t. Valiela et al. (x976) suggested that a large proportion of roots and rhizomes produced during one growing season die during the same season and that roots do not overwinter. We observed some decomposition of old stems above the point of attachment of new tillers, and of portions of long rhizomes in type (2) clumps. However, we could find no evidence of substantial decomposition of new roots (current year; solid lines in Figure I) during the growing season, or of old roots (produced the previous year; broken lines in Figure 1) in type (2) clumps. These latter, old roots remain attached and turgid (Plate x, 2 June) and new roots were not observed emerging from old nodes. Carbon-x 4 translocation studies indicated that one year old roots retained vital activity (Patriquin & McClung, x978). When x-year old roots were incubated with 2,3,5-triphenyltetrazolium clfloride (Patfiquin, i978 ), tetrazolium reduction was observed in the stele but to a limited extent or not at all in the cortex. New roots (current year) exhibited intense tetrazolium reduction in both the stele and cortex. Stands at various positions between the edge and the center of the thatch island colony exhibited decreasing numbers of shoots, buds and long rhizomes per clump with increasing
Belozcground growth of Spartina aherniflora
583
TABLE 1. Numbers of shoots, tiller buds and long rhizomes per clump in stands of differing densities and age. Observations conducted September-October Clump description and no. clumps examined
No. mature* shoots per clump
No. tiller buds per clump
One isolated type (t) clump (t year old)
4-8 b
16
13 type (1) clumps from middle of 3-year old stand
1-3
22 type (1) clumps from 4-6-year old stand
x-3
No. long rhizomes per clump
5+6 ~
3"4
z'z3
1"14
*Non-overwintering shoot, but not necessarily flowering. Generally only the oldest one or two shoots in a clump flowered. bRange for many such clumps. CFive long rhizomes with a total of six branches from these.
stand density and age (Table x). In mature stands, production of new clumps appears to roughly balance loss of old clumps. Essentially, then, a shoot clump produces foliage and roots for two years, and then dies. Our observations for S. alterniflora are somewhat similar to those reported by Caldwell (x957) for S. townsendii, except that S. toz~tsendii clumps generally, but not always, die after one season. As for S. townsendii, concentric alternating bands of dead and living clumps can be observed in isolated, young S. alterniflora stands. In older stands these bands become obscured but alternating patches of living and dead clumps can still be distinguished. '.-. 1,500
r i
I
yl
._~ IOOC
g *~ 50C x3 I
I-
I Moy
I June
I July
I Aug
I Sept
Iii Ocl
I Jon
Month
Figure 3. Seasonal changes in ethanol-soluble carbohydrate content of ,S'. alterniflora during I977/1978; (@) roots and rhizomes, (0) leaves and culms, data are means of two or three samples -I-S.E.M. and expressed as Itequiv. glucose (g dry weight tissue) -t, wet weights were converted to dry weights using dry/wet weight factors determined monthly during the study period.
584
D.C. Livlngstone ~ D. G. Patrlquln
Non-structural carbohydrates Total sugar concentrations in belowground and aboveground tissues exhibited a decline during the period of vigorous root growth and rapid leaf grox~th, increased to previous levels during the period of rhizome grox~lh and then changed little over the winter (Figure 3). Chromatography of "ethanol-soluble sugars revealed only glucose, fructose and sucrose. Starch concentrations in belowground tissues exhibited a somewhat different pattern of change, with declining concentrations over the winter (Figure 4). At all times the sugars appeared to be the predominant non-structural carbohydrates (NSC). The high NSC content of S. alterniflora in Nova Scotia (9-31%) in comparison to Georgia (4-io%, A[cIntire & Durtstan, i976 ) and the distinct seasonal pattern in Nova Scotia suggest that, near this northern extreme of S. alterniflora, NSC are of importance as storage compounds. As has been suggested for alpine plants (Mooney & Billings, x96o), these carbohydrate reserves probably support rapid early season growth (Figure 2). High sugar content may also function in frost hardiness (Bidwell, i974) of both the overwintering tillers and underground parts. In January, i978, ice was observed to a depth of about io cm in the marsh sediments.
I "o
200
o
T~
3
100
g
I
Moy
I
June
I
July
I
Aug
I
Sep!
,,
Oct
I
t=l
Jon Apt
Month
Figure 4. Seasonal changes in tissue starch content; (Q) roots and rhizomes, ( , ) leaves and culms, data are means of two or three samples 4-s.e..~t., starch content expressed as pequiv, glucose (g dry weight tissue) - t .
Functional belowground biomass In newly established stands, it is relatively easy to separate type (3) clumps (non-functional) from types (o), (I) and (2) clumps, which include the functional biomass. The ratio of functional belowground biomass to aboveground biomass in the autumn was estimated for six i/i6 m * samples from areas of differing shoot densities. For each sample, a trench was dug around the specified area to a depth below the root zone, and sediment was then washed away from roots and rhizomes by flushing with seawater. In this manner, the entire biomass was removed in an intact condition. In such a sample, belowground biomass of type (3) clumps was recognized as that attached to dead shoots only. For stands above a density of about 2o shoots per 1/16 m ~ or for stands including more than i year of growth, the ratio of functional belowground biomass to aboveground biomass was nearly constant (Table 2).
Belozcground grozvth of Spartina altcrniflora
.]. ,-~ ~
585
2,~ P-. ~ P-. &
0
~'__..I. r
~
tq el el el r
8 r', " ~
0
'9. ;,,b,.b ;,,~ ~
0
0
~
:.
g'~
e~ e-.
0 ~o
~
0
de~ E "6
-~-~
e.,
~
u"~O 0 ~ o
.s
C ~ 0 0 00
r
~
~d
8
el t#3
o=
2 .+'=B 0 " ~
Z ~0
d
. 0
"~
"~' "~ x "~
0 "~. e., ~
~2~g .~
m
586
D. C. Livlngstone'~ D. G. Patriquln
The belowground-aboveground biomass ratios are of similar magnitude to those observed for young stands in more southerly regions (Smalley, i959; Seneca, 1974; Broome et al., 1975). In the oldest stands (3 to 5 in Table 2), roots accounted for 32 to 39% of the belowground biomass. Ratios of net belowground production to end-of-season aboveground biomass (equal to about 83% of aboveground production, Hatcher & Mann, I975) were estimated by dividing functional belowground biomass by shoot biomass produced over two years (Table 2). Biomass for shoots produced during the previous growing season was estimated by multiplying the number of these shoots (as indicated by dead shoot bases) by present end-ofseason average shoot blomass. As for the above, these ratios were nearly constant for stands including more than I year of growth. Such values give a reasonable indication of actual production of belowground biomass only if there is not substantial decomposition of this belowground biomass after one winter. For one stand, it was possible to separate the belowground biomass produced during the current year from that produced during the previous year (solid vs. broken lines in clump types (o), (i), (2), Figure 1). The ratio of belowground biomass (current year only) to aboveground biomass for this stand, 1.75, was only 11% greater than the ratio estimated for this stand on the basis of 2 years of accumulation of belowground biomass (stand 4, Table 2). This supports the qualitative observations above Of limited decomposition of belowground parts during their second year. The ratios of belowground production to aboveground production (as indicated by the end of season biomass) are much lower than values (8-12) estimated for S. alterniflora in Massachusetts (Valiela et aL, 1976). The absolute values of belowground production estimated by Gallagher & Plumley (1979) are considerably lower than those of Valiela et al. (1976), and are similar to our values. Our method of estimating belowground production is essentially a direct one in that it is based on measurements of living, functional biomass, but the values so obtained represent only net seasonal production. The techniques of Gallagher & Plumley (1979) and Valiela et aL (1976) are indirect ones in that increments in living materials are inferred from changes in the total belowground macro-organlc matter, or in total dead belowground biomass. It is not clear precisely what these changes, which were sometimes irregular, represent. Our observations do not concur with Vallela et al. (I976) who suggested (i) that roots do not overwinter, (ii) that there is a rapid turnover of belowground biomass during the growing season, and (iii) that root production constitutes a minor fraction (3-11%) of total belowground production. It does appear necessary, however, to postulate higher belowground production than suggested by our values in order to account for observed rates of sulfate reduction (Howarth, x979). Perhaps this sulfate reduction is dependent on components that do undergo rapid turnover, such as root exudates (Martin, I977) and root cap cells (Clowes & Woolston, 1978). Techniques to discriminate these other components of belowground production are required.
Acknowledgements This work was supported by an Operating Grant from the Natural Sciences and Engineering Research Council of Canada (NSERC). The first author was supported by a NSERC Postgraduate Scholarship.
References Atkins, C. A. & Canvin, D. T. I97x Photosynthesis and CO2 evolution by leaf discs: gas exchange, extraction, and ion-exchange fractionation of t~C-labeled photosynthetic products. Canadian ffournal of Botany 49, xz25-1234.
Belowground growth of Spartina alterniflora
587
Bidwell, R. G. S. x974 Plant Physiology, MacMillan Publishing Co., Inc., New York. 643pp. Broome, S. W., SVoodhouse, W. W., Jr. & Seneca, E. D. t975 The relationship of mineral nutrients to growth of Spartina alterniflora in North Carolina: II. The effects of N, P, and Fe fertilizers. Soll Science Society of America Proceedings39(2), 3ox-3o7 . Caldwell, P. x957 The spatial development of Spartina colonies growing without competition. Annals of Botany ~I~ 2ox-zx5. Clowes, F. A. L. & Woolston, R. E. x978 Sloughing of root cap cells. Annals of Botany 4z, 83-89. Dubois, M., Gilles, K. A., Hamilton, J. K., Rebers, P. A. & Smith, F. x956 Colorimetric method for determination of sugars and related substances. Analytical Chemistry z8(3), 35o-356. Fenehel, T. M. & Riedl, R. J. x97o The sulfide system: a new biotic community underneath the oxidized layer of marine sand bottoms. International ffournal on Life in Oceans attd Coastal Waters 7(3), 255-268. Gallagher, J. L. & Plumley, F. G. x979 Underground biomass profiles and productivity in Atlantic coastal marshes. Americanffournal of Botany 66(2), I56-16I. Hassid, "~V.Z. & Neufeld, E. F. x964 Quantitative determination of starch in plant tissues. Methods in Carbohydrate Chemistry 4~ 33-36. Hatcher, B. G. & Mann, K. H. x975 Above-ground production of marsh cordgrass (Spartlna alterniflora) near the northern end of its range,ffournal of the Fisheries Research Board of Canada 32, 83-87. Howarth, R. W. x979 Pyrite: its rapid formation in a salt marsh and its importance in ecosystem metabolism. Science 2o3~ 49-5 x. Jefferies, R. L. I977 The vegetation of salt marshes at some coastal sites in Arctic North America. ffournal of Ecology 65~ 661-672. Livingstone, D. C. 1978 Growth and nitrogen accumulation in Sparthta alterniflora Loisel. in relation to certain environmental and physiological factors. Unpublished 1VI.Sc. Thesis, Dalhousie University, Halifax, Nova Scotia. l~iartin, J. K. x977 Factors influencing loss of organic-carbon from wheat roots. Soil Biology and Biochemistry 9(x), x--7. IMeIntire, G. L. & Dunstan, SV. M. x976 Non-structural carbohydrates in Spartina alterniflora Loisel. Botanica 3larina xg~ 93-96. l~iilner, C. & Hughes, R. E. x968 3letlwdr for the Measurement of the Primary Production of Grassland. IBP Handbook 6, Blackwell. l~iooney, H. A. & Billings, W. D. x96o The annual carbohydrate cycle of alpine plants as related to growth. American ffournal of Botany 47~ 594-598. Newbould, P. J. x967 Methods for Estimating the Primary Production of Forests. IBP Handbook No. 2. Blackwell. Patriquin, D. G. x978 Nitrogen fixation (acetylene reduction) associated with cord grass, Spartina aIterniflora Loisel. In Environmental Role of Nitrogen-fixing Blue-green Algae and Asymbiotic Bacteria (Granhall, U., ed.). Ecological Bulletin (Stockholm) 26~ so-a7. Patriquin, D. G. & McCIung, (2. R. x978 Nitrogen accretion, and the nature and possible significance of Nz-fixatlon (acetylene reduction) in a Nova Scotia Spartina alterniflora stand. ~Iarine Biology 47~ zz7-z4z. Pridham, J. B. x956 Determination of sugars on paper chromatograms with p-anisidine hydrochloride. Analytical Chemistry ~8, x967-x968. Raadsveld, C. W. & Klomp, H. x97x Thin-layer chromatographic analysis of sugar mixtures, ffournal of Chromatography 57J 99-xo6. Redfield, A. (2. x97z Development of a New England salt marsh. Ecological ~Xonographs 42j sol-z37. Seneca, E. D. x974 Germination and seedling response of Atlantle and Gulf Coasts populations of Spartlna alterniflora. American ffournal of Botany 6x(9), 947-956. Smalley, A. E. x959 The growth cycle of Spartlna and its relation to the insect populations in the marsh. Proceedings of the Salt Marsh Conference, Marine Institute of the University of Georgia. pp. 96-xoo. Turner, R. E. x976 Geographic variations in salt marsh maerophyte production: a review. Contributions in Marine Science 2o, 47-68. Umbreit, W. ~,V.,Burris, R. H. & Stauffer, J. F. x97z Manometric and Biochemical Techniques. Burgess Publishing Co., Minneapolis. 387pp. Valiela, I., Teal, J. iVI. & Persson, N. Y. 1976 Production and dynamics of experimentally enriched salt marsh vegetation: belmvground blomass. Limnology and Oceanography ~x(2), 245-252.
2 June
) ~--~
.
.~.'J
1'~
O§ k
9
I
9
, .
~ P
!
f 14 August
9.
/
"'
f
( Plate z. P/ant specimens taken at the beginning (2 June) and middle ([4 August) of the growing season. 2 June specimen includes a I ,-b type clump which had produced foliage during the previous )'ear (OT, old tiller) and a long rhizome terminating in a short overwlntering tiller (o type clump in the previous fall, now o-b). 14 August specimen is a new ( o + ) clump, and illt*strates the vigorous root growth that occurs in June and July. Several tillers are present, of which the oldest is flowering. Some new overwintering tillers (indicated by arrow) and long rhizomes (R) have recently bccn produced. Scale=8 cm.
[Faci~ p. 58~
Belowground Growth of Spartina alterniflora Loisel.: Habit, Functional Biomass and Non-structural Carbohydrates
Diane C. Livingstone and David G.
Patriqttin
Biology Department, Dalhousie University, Halifax, Nova Scotia, B3H 4fit, Canada Received 25 October I979 and in revised form 24 August I98o
Keywords: salt marshes; vegetation; growth rates; age determination; annual cycles; seasonal variations; carbohydrates; Nova Scotia coast Observations on the growth habit of a thatch island Sparthla alterniflora colony in. Nova Scotia indicated that an individual shoot chtmp lives for 2 years and then dies. There were three distinct phases of growth during the year: rapid leaf and root growth in June and July, vigorous rhizome growth and production of ovenvintering tillers concurrent with reproductive development in August and September, and finally, a period of no obvious change from October until the following May. There was a distinct autumn accumulation of non-structural carbohydrates in underground parts, with sugars accounting for 31% of the dry weight in October and starch 4%. Ratios of functional belowground biomass to end of season aboveground biomass were roughly constant for stands more than i year old; the ratio of annual net production of belowground parts to end of season aboveground biomass was estimated as x'75.
Introduction Although the aboveground productivity of saltmarsh angiosperms has been extensively studied (Turner, I976; Jefferies, x977), relatively little is known of the belowground growth habits and production of most species. Recent interest in sulfur metabolism of salt marshes has emphasized the high magnitude of belowground production (Howarth, I979) and has indicated as well, that much of this production may become available aboveground through diffusion of reduced hydrogen acceptors to the sediment surface (Fenchel & Riedl, x97o). There is no generally acceptable technique for estimating belowground production of perennial species ~Iilner & Hughes, x968; Newbould, 1967). Valiela et al. 0976) estimated belowground production of ,S'. alterniflora from the observed increases of dead materials during the growing season. It was assumed that these increases represented turnover of belowground biomass within one season. For fertilized and unfertilized low marsh plots, their estimates of belowground production were in the range of 33x5 to 5687 g dry matter m -2. GaUagher & Plumley (x979) estimated belowground production from the differences between maximum and minimum amounts of total macro-organic matter within one growing season; their estimates varied between 2x8 and 2ioo g dry matter m -2 year -1. There are several reports of aboveground/belowground biomass ratios (Smalley, i959; Seneca, x974; 579 0272-7714/8t/o5o579+to :$02.o0/0
~) t98t Academic Press Inc. (London) Ltd.
580
D. C. Livingstone r D. G. Patriquin
Broome et aL, x975) but particularly for older stands, it is not clear what this biomass represented. Observations on the growth habit of S. alterniflora reported in this study provided a means of separating functional from non-functional belowground biomass in young stands. We examined aboveground]belowground biomass ratios and estimated net belowground production for stands of increasing plant density and age on a recently colonized sandbar in a Nova Scotian sahmarsh. We also investigated the relationship of non-structural carbo9 hydrate (NSC) content to plant growth during a growing season. In Georgia, where year round growth of S. alterniflora occurs, ~IcIntire & Dunstan (i976) found that S. alterniflora retained low levels of NSC throughout the year. They raised the question of whether this represented a genetically fixed characteristic of incorporating most production directly into structural components, or whether in northern regions (such as Nova Scotia) storage of NSC may occur. Materials and methods
The S. alterniflora stand was located on a recently established (within Io years) sandbar at Conrad Beach, on the Atlantic coast near Halifax, Nova Scotia. This stand was actively expanding along its edges, and was similar in general aspect to tile 'thatch islands' described by Redfield (I972). For studies of growth habit and biomass, samples were taken, as described in the text, and later washed with distilled water, dried in a forced air oven at 80 ~ and. weighed. For determination of non-structural carbohydrates, sods were removed from the marsh to a depth of about 3~ cm and plants washed free of soil. Only belowground tissue directly attached to live foliage was collected. Plants were divided it/to three sections: roots and rhizomes, leaves and culms and reproductive parts. Data for reproductive parts are reported in Livingstone (x978). Approximately 5o-Ioo g of each type were sliced into sections a few centimeters in length, mixed, and 5 or xo g snbsamples removed. One Io g sample of each tissue type was dried at 80 ~ and then ground to 60 mesh in a Wiley Mill. Subsamplcs of this matelial were used for the determination of starch content (Hassid & Neufeld, x964; Dubois et al., x956). Another ,o g sample of each tissue type was extracted three times in ioo ml portions of hot 8o% ethanol. Following centrifugation, supernatants were combined and evaporated to dryness in a rotary evaporator at 4 ~ ~ Residues were resuspended in Io ml deionized water and an aliquot of the resuspended extract separated into acidic, basic and neutral fractions by ion-exchange chromatography (Atkins & Canvin, I97x ). Total carbohydrates in the neutral fraction were measured by the total carbon assay of Umbreit et aL (i97z) and expressed as equivalents of glucose (laequiv. glucose (g dry wt)-l), by comparison of absorbance values to a glucose standard curve. Carbohydrates in the neutral fraction were separated by thin-layer chromatography on cellulose MN 300 (25o ~tm thickness) plates in water: ethyl acetate: pyridine (25 : Ioo : 35) (Raadsveld & Klomp, x97 Q. Spots were identified against standards using p-anisidine hydrochloride spray (Pridham, x956). Results and discussion
Growth habit The growth habit of S. alterniflora was examined by carefully digging up connected 'shoot dumps' (Caldwell, x957), beginning with the apical region of recently produced long
Belozcground grorcth of Spartina aherrfiflora
3
581
!
2
o
/ ..
Foliooe in foil of 197,5
Ttller bud in foil of 1975
P
,.__/
Long chizome with filler bud, foil 1975
Slems ond roofs ossociofed with 197.5 folloqe Old deod folioge, produced 1 9 7 4 e~
J
5temso roofs ond long rhizornes Of 1 9 7 4
t t I
Slems, roofs, fohoqe (deod} o n d long rhizomes of 1973
Figure x. Structure of connected shoot clumps in the autumn of x975. rhizomes. The structure of successive shoot clumps examined in the fall is illustrated in Figure x. The youngest clump (o) consisted of a long rhizome which terminated at this time in a single tiller bud or small (less than I I cm length) ovcrwintering shoot. The next clump (x) consisted of several mature shoots (length greater than x8 cm), of which the oldest one or two were usually flowering shoots, and a number of tiller buds or overwintering shoots, and a large mass of roots. Clump (z) included mature shoots as in (x) but few or no buds or overwintering tillers and no recently produced long rhizomes. Basal portions of old dead shoots from the previous year were present in type (z) clumps. Clump (3) consisted of dead shoots only, or occasionally a single living shoot was present. The pattern of occurrence of dead shoots in these clumps suggested that the successive clumps (o), (i), (z), (3) were, in the autumn, respectively o, z, z, and 3 years of age.
582
D. C. Livingstone C.~ D. G. Patriquln
Observations of clump development at 2-week intervals from May to October and on three occasions between October and ~'Iay illustrated three more or less distinct seasonal phases of belowground growth: (i) vigorous root growth at bases of new tillers in June and July, (ii) vigorous rhizome growth and production of overavintering shoots in August and September, and (iii) a period of no obvious change during the winter (Plate x). These periods corresponded respectively to vegetative growth, reproductive development, and senescence of aboveground vegetation (Figure 2).
e ~
~
8
I/'\
p-
J
50(
Ioo
o
V - * / t May
I June
t July
I Aug
I Sept
[al 0r
Jan
l,,l " Apr
Month
Figure 2. Seasonal changes in seawater and sediment temperatures and above-
ground biomass of S. alterniflora; (r seawater, (u) sediment at 4cm depth, data are means of two readings taken at randomly selected sites at ~2.ooh (Atlantic Standard Time); (O) standing crop, data are means of all live biomass on three t/16 m t quadrats_+S.E.M.; ( , ) maximum plant height on three x/i6 m 2 quadrats, data are means 4-s.e..~t. Valiela et al. (x976) suggested that a large proportion of roots and rhizomes produced during one growing season die during the same season and that roots do not overwinter. We observed some decomposition of old stems above the point of attachment of new tillers, and of portions of long rhizomes in type (2) clumps. However, we could find no evidence of substantial decomposition of new roots (current year; solid lines in Figure I) during the growing season, or of old roots (produced the previous year; broken lines in Figure 1) in type (2) clumps. These latter, old roots remain attached and turgid (Plate x, 2 June) and new roots were not observed emerging from old nodes. Carbon-x 4 translocation studies indicated that one year old roots retained vital activity (Patriquin & McClung, x978). When x-year old roots were incubated with 2,3,5-triphenyltetrazolium clfloride (Patfiquin, i978 ), tetrazolium reduction was observed in the stele but to a limited extent or not at all in the cortex. New roots (current year) exhibited intense tetrazolium reduction in both the stele and cortex. Stands at various positions between the edge and the center of the thatch island colony exhibited decreasing numbers of shoots, buds and long rhizomes per clump with increasing
Belozcground growth of Spartina aherniflora
583
TABLE 1. Numbers of shoots, tiller buds and long rhizomes per clump in stands of differing densities and age. Observations conducted September-October Clump description and no. clumps examined
No. mature* shoots per clump
No. tiller buds per clump
One isolated type (t) clump (t year old)
4-8 b
16
13 type (1) clumps from middle of 3-year old stand
1-3
22 type (1) clumps from 4-6-year old stand
x-3
No. long rhizomes per clump
5+6 ~
3"4
z'z3
1"14
*Non-overwintering shoot, but not necessarily flowering. Generally only the oldest one or two shoots in a clump flowered. bRange for many such clumps. CFive long rhizomes with a total of six branches from these.
stand density and age (Table x). In mature stands, production of new clumps appears to roughly balance loss of old clumps. Essentially, then, a shoot clump produces foliage and roots for two years, and then dies. Our observations for S. alterniflora are somewhat similar to those reported by Caldwell (x957) for S. townsendii, except that S. toz~tsendii clumps generally, but not always, die after one season. As for S. townsendii, concentric alternating bands of dead and living clumps can be observed in isolated, young S. alterniflora stands. In older stands these bands become obscured but alternating patches of living and dead clumps can still be distinguished. '.-. 1,500
r i
I
yl
._~ IOOC
g *~ 50C x3 I
I-
I Moy
I June
I July
I Aug
I Sept
Iii Ocl
I Jon
Month
Figure 3. Seasonal changes in ethanol-soluble carbohydrate content of ,S'. alterniflora during I977/1978; (@) roots and rhizomes, (0) leaves and culms, data are means of two or three samples -I-S.E.M. and expressed as Itequiv. glucose (g dry weight tissue) -t, wet weights were converted to dry weights using dry/wet weight factors determined monthly during the study period.
584
D.C. Livlngstone ~ D. G. Patrlquln
Non-structural carbohydrates Total sugar concentrations in belowground and aboveground tissues exhibited a decline during the period of vigorous root growth and rapid leaf grox~th, increased to previous levels during the period of rhizome grox~lh and then changed little over the winter (Figure 3). Chromatography of "ethanol-soluble sugars revealed only glucose, fructose and sucrose. Starch concentrations in belowground tissues exhibited a somewhat different pattern of change, with declining concentrations over the winter (Figure 4). At all times the sugars appeared to be the predominant non-structural carbohydrates (NSC). The high NSC content of S. alterniflora in Nova Scotia (9-31%) in comparison to Georgia (4-io%, A[cIntire & Durtstan, i976 ) and the distinct seasonal pattern in Nova Scotia suggest that, near this northern extreme of S. alterniflora, NSC are of importance as storage compounds. As has been suggested for alpine plants (Mooney & Billings, x96o), these carbohydrate reserves probably support rapid early season growth (Figure 2). High sugar content may also function in frost hardiness (Bidwell, i974) of both the overwintering tillers and underground parts. In January, i978, ice was observed to a depth of about io cm in the marsh sediments.
I "o
200
o
T~
3
100
g
I
Moy
I
June
I
July
I
Aug
I
Sep!
,,
Oct
I
t=l
Jon Apt
Month
Figure 4. Seasonal changes in tissue starch content; (Q) roots and rhizomes, ( , ) leaves and culms, data are means of two or three samples 4-s.e..~t., starch content expressed as pequiv, glucose (g dry weight tissue) - t .
Functional belowground biomass In newly established stands, it is relatively easy to separate type (3) clumps (non-functional) from types (o), (I) and (2) clumps, which include the functional biomass. The ratio of functional belowground biomass to aboveground biomass in the autumn was estimated for six i/i6 m * samples from areas of differing shoot densities. For each sample, a trench was dug around the specified area to a depth below the root zone, and sediment was then washed away from roots and rhizomes by flushing with seawater. In this manner, the entire biomass was removed in an intact condition. In such a sample, belowground biomass of type (3) clumps was recognized as that attached to dead shoots only. For stands above a density of about 2o shoots per 1/16 m ~ or for stands including more than i year of growth, the ratio of functional belowground biomass to aboveground biomass was nearly constant (Table 2).
Belozcground grozvth of Spartina altcrniflora
.]. ,-~ ~
585
2,~ P-. ~ P-. &
0
~'__..I. r
~
tq el el el r
8 r', " ~
0
'9. ;,,b,.b ;,,~ ~
0
0
~
:.
g'~
e~ e-.
0 ~o
~
0
de~ E "6
-~-~
e.,
~
u"~O 0 ~ o
.s
C ~ 0 0 00
r
~
~d
8
el t#3
o=
2 .+'=B 0 " ~
Z ~0
d
. 0
"~
"~' "~ x "~
0 "~. e., ~
~2~g .~
m
586
D. C. Livlngstone'~ D. G. Patriquln
The belowground-aboveground biomass ratios are of similar magnitude to those observed for young stands in more southerly regions (Smalley, i959; Seneca, 1974; Broome et al., 1975). In the oldest stands (3 to 5 in Table 2), roots accounted for 32 to 39% of the belowground biomass. Ratios of net belowground production to end-of-season aboveground biomass (equal to about 83% of aboveground production, Hatcher & Mann, I975) were estimated by dividing functional belowground biomass by shoot biomass produced over two years (Table 2). Biomass for shoots produced during the previous growing season was estimated by multiplying the number of these shoots (as indicated by dead shoot bases) by present end-ofseason average shoot blomass. As for the above, these ratios were nearly constant for stands including more than I year of growth. Such values give a reasonable indication of actual production of belowground biomass only if there is not substantial decomposition of this belowground biomass after one winter. For one stand, it was possible to separate the belowground biomass produced during the current year from that produced during the previous year (solid vs. broken lines in clump types (o), (i), (2), Figure 1). The ratio of belowground biomass (current year only) to aboveground biomass for this stand, 1.75, was only 11% greater than the ratio estimated for this stand on the basis of 2 years of accumulation of belowground biomass (stand 4, Table 2). This supports the qualitative observations above Of limited decomposition of belowground parts during their second year. The ratios of belowground production to aboveground production (as indicated by the end of season biomass) are much lower than values (8-12) estimated for S. alterniflora in Massachusetts (Valiela et aL, 1976). The absolute values of belowground production estimated by Gallagher & Plumley (1979) are considerably lower than those of Valiela et al. (1976), and are similar to our values. Our method of estimating belowground production is essentially a direct one in that it is based on measurements of living, functional biomass, but the values so obtained represent only net seasonal production. The techniques of Gallagher & Plumley (1979) and Valiela et aL (1976) are indirect ones in that increments in living materials are inferred from changes in the total belowground macro-organlc matter, or in total dead belowground biomass. It is not clear precisely what these changes, which were sometimes irregular, represent. Our observations do not concur with Vallela et al. (I976) who suggested (i) that roots do not overwinter, (ii) that there is a rapid turnover of belowground biomass during the growing season, and (iii) that root production constitutes a minor fraction (3-11%) of total belowground production. It does appear necessary, however, to postulate higher belowground production than suggested by our values in order to account for observed rates of sulfate reduction (Howarth, x979). Perhaps this sulfate reduction is dependent on components that do undergo rapid turnover, such as root exudates (Martin, I977) and root cap cells (Clowes & Woolston, 1978). Techniques to discriminate these other components of belowground production are required.
Acknowledgements This work was supported by an Operating Grant from the Natural Sciences and Engineering Research Council of Canada (NSERC). The first author was supported by a NSERC Postgraduate Scholarship.
References Atkins, C. A. & Canvin, D. T. I97x Photosynthesis and CO2 evolution by leaf discs: gas exchange, extraction, and ion-exchange fractionation of t~C-labeled photosynthetic products. Canadian ffournal of Botany 49, xz25-1234.
Belowground growth of Spartina alterniflora
587
Bidwell, R. G. S. x974 Plant Physiology, MacMillan Publishing Co., Inc., New York. 643pp. Broome, S. W., SVoodhouse, W. W., Jr. & Seneca, E. D. t975 The relationship of mineral nutrients to growth of Spartina alterniflora in North Carolina: II. The effects of N, P, and Fe fertilizers. Soll Science Society of America Proceedings39(2), 3ox-3o7 . Caldwell, P. x957 The spatial development of Spartina colonies growing without competition. Annals of Botany ~I~ 2ox-zx5. Clowes, F. A. L. & Woolston, R. E. x978 Sloughing of root cap cells. Annals of Botany 4z, 83-89. Dubois, M., Gilles, K. A., Hamilton, J. K., Rebers, P. A. & Smith, F. x956 Colorimetric method for determination of sugars and related substances. Analytical Chemistry z8(3), 35o-356. Fenehel, T. M. & Riedl, R. J. x97o The sulfide system: a new biotic community underneath the oxidized layer of marine sand bottoms. International ffournal on Life in Oceans attd Coastal Waters 7(3), 255-268. Gallagher, J. L. & Plumley, F. G. x979 Underground biomass profiles and productivity in Atlantic coastal marshes. Americanffournal of Botany 66(2), I56-16I. Hassid, "~V.Z. & Neufeld, E. F. x964 Quantitative determination of starch in plant tissues. Methods in Carbohydrate Chemistry 4~ 33-36. Hatcher, B. G. & Mann, K. H. x975 Above-ground production of marsh cordgrass (Spartlna alterniflora) near the northern end of its range,ffournal of the Fisheries Research Board of Canada 32, 83-87. Howarth, R. W. x979 Pyrite: its rapid formation in a salt marsh and its importance in ecosystem metabolism. Science 2o3~ 49-5 x. Jefferies, R. L. I977 The vegetation of salt marshes at some coastal sites in Arctic North America. ffournal of Ecology 65~ 661-672. Livingstone, D. C. 1978 Growth and nitrogen accumulation in Sparthta alterniflora Loisel. in relation to certain environmental and physiological factors. Unpublished 1VI.Sc. Thesis, Dalhousie University, Halifax, Nova Scotia. l~iartin, J. K. x977 Factors influencing loss of organic-carbon from wheat roots. Soil Biology and Biochemistry 9(x), x--7. IMeIntire, G. L. & Dunstan, SV. M. x976 Non-structural carbohydrates in Spartina alterniflora Loisel. Botanica 3larina xg~ 93-96. l~iilner, C. & Hughes, R. E. x968 3letlwdr for the Measurement of the Primary Production of Grassland. IBP Handbook 6, Blackwell. l~iooney, H. A. & Billings, W. D. x96o The annual carbohydrate cycle of alpine plants as related to growth. American ffournal of Botany 47~ 594-598. Newbould, P. J. x967 Methods for Estimating the Primary Production of Forests. IBP Handbook No. 2. Blackwell. Patriquin, D. G. x978 Nitrogen fixation (acetylene reduction) associated with cord grass, Spartina aIterniflora Loisel. In Environmental Role of Nitrogen-fixing Blue-green Algae and Asymbiotic Bacteria (Granhall, U., ed.). Ecological Bulletin (Stockholm) 26~ so-a7. Patriquin, D. G. & McCIung, (2. R. x978 Nitrogen accretion, and the nature and possible significance of Nz-fixatlon (acetylene reduction) in a Nova Scotia Spartina alterniflora stand. ~Iarine Biology 47~ zz7-z4z. Pridham, J. B. x956 Determination of sugars on paper chromatograms with p-anisidine hydrochloride. Analytical Chemistry ~8, x967-x968. Raadsveld, C. W. & Klomp, H. x97x Thin-layer chromatographic analysis of sugar mixtures, ffournal of Chromatography 57J 99-xo6. Redfield, A. (2. x97z Development of a New England salt marsh. Ecological ~Xonographs 42j sol-z37. Seneca, E. D. x974 Germination and seedling response of Atlantle and Gulf Coasts populations of Spartlna alterniflora. American ffournal of Botany 6x(9), 947-956. Smalley, A. E. x959 The growth cycle of Spartlna and its relation to the insect populations in the marsh. Proceedings of the Salt Marsh Conference, Marine Institute of the University of Georgia. pp. 96-xoo. Turner, R. E. x976 Geographic variations in salt marsh maerophyte production: a review. Contributions in Marine Science 2o, 47-68. Umbreit, W. ~,V.,Burris, R. H. & Stauffer, J. F. x97z Manometric and Biochemical Techniques. Burgess Publishing Co., Minneapolis. 387pp. Valiela, I., Teal, J. iVI. & Persson, N. Y. 1976 Production and dynamics of experimentally enriched salt marsh vegetation: belmvground blomass. Limnology and Oceanography ~x(2), 245-252.
2 June
) ~--~
.
.~.'J
1'~
O§ k
9
I
9
, .
~ P
!
f 14 August
9.
/
"'
f
( Plate z. P/ant specimens taken at the beginning (2 June) and middle ([4 August) of the growing season. 2 June specimen includes a I ,-b type clump which had produced foliage during the previous )'ear (OT, old tiller) and a long rhizome terminating in a short overwlntering tiller (o type clump in the previous fall, now o-b). 14 August specimen is a new ( o + ) clump, and illt*strates the vigorous root growth that occurs in June and July. Several tillers are present, of which the oldest is flowering. Some new overwintering tillers (indicated by arrow) and long rhizomes (R) have recently bccn produced. Scale=8 cm.
[Faci~ p. 58~