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Effect of collembolan grazing upon nitrogen and cation leaching from decomposing leaf litter
0038-0717/8’;060601-05$03.00/O Copyright 0 1982 Pergamon Press Ltd
Soil Biol. Bioche~~~.Vol. 14, pp. 601 to 605, 1982 Printed in Great Britain. All rights reserved
EFFECT OF COLLEMBOLAN GRAZING UPON NITROGEN AND CATION LEACHING FROM DECOMPOSING LEAF LITTER P. INESON.M. A. LEONARDand J. M. ANDERSON Department of Biological Sciences, Wolfson
Ecology Laboratory, Exeter EX4 4PS. Devon, U.K.
(Accepted 20 March
University
of Exeter.
1982)
Summary-Fragmented oak litter was incubated in the laboratory for 4 months with and without collembola. The effects of the animals upon fungal standing crop and leaching of inorganic nitrogen and cations was monitored over this period. The results showed that the fungal standing crop was higher in the presence of small numbers of animals than in litter lacking animals, yet at higher grazing intensities the fungal standing crop fell markedly. Significant increases in the leaching of ammonium, nitrate and calcium occurred as a consequence of animal grazing. but potassium and sodium losses from the litter were unaffected.
INTRODUCTION Despite the small contribution that saprotrophic microarthropods make towards total carbon mineralization in forest and woodland soils (Edwards et a/., 1981) they have an important role in affecting the quantity, community composition and activity of the soil microflora (Hanlon and Anderson. 1979; Parkinson et al., 1979). Studies suggest that protozoa, nematodes and annelids are important in soil N mineralization processes (Anderson et al., 1981) yet there are few data concerning the part played by microarthropods in nutrient transformations in soils. despite much speculation (Harding and Stuttard, 1974; Seastedt and Crossley, 1980; Anderson et al., 1981). Fungal mycelium in soil can contain a major proportion of nitrogen, potassium and other cations in the soil pool (Cromack et al.. 1975) and thus the sensitivity of fungal hyphae to microarthropod grazing may well have considerable significance for nutrient mobilization (Hanlon and Anderson, 1979; Parkinson et al., 1979). In the experiment described here a microcosm system was used to follow changes in collembolan numbers, fungal standing crop and cation and ammonium release over several months in grazed and ungrazed litter. The aim was to assess the functional significance of collembolan grazing in nutrient release from decomposing litter. MATERIALS AND Preparation
METHODS
and watering of microcosms
Oak (Quercus robur L.) leaves were collected shortly after leaf fall from the litter layer of a mature oak woodland, air dried at 20°C and brushed free of faecal material and debris. Midribs were removed and the remaining laminae cut into small fragments of 0.5-l.Ocm. The fragments were mixed well and aliquots, 1.5 g, placed in microcosm chambers (Anderson and Ineson, 1982). 601 S.BB 146-F
The litter in each microcosm was rehydrated by the addition of 100 ml of distilled water for 24 h, which served to leach out soluble tannins and readily-metabolizable materials. The litter was drained and inoculated with a coarsely sieved suspension of macerated fresh leaf litter and then incubated in a cooled cabinet at 15°C. Microcosms were placed according to a randomized block design and were leached weekly with 60 ml of distilled water, leachates being retained for chemical analysis. Prior to animal addition three microcosms were sampled to determine litter dry weight and nutrient content. Culture, addition and enumeration
of collembola
Folsomia candida Will. were maintained in the laboratory on a diet of dried baker’s yeast and housed in plastic boxes containing a substrate of plaster of Paris and activated charcoal (Goto. 1960). Individuals were selected for experimentation by passing the culture through a 0.3 mm pore size sieve, those being retained were added, 20 per microcosm, to half of the microcosms. The additions were not made until 6 weeks after initial leaching, permitting time for the development of a decomposer flora. No further animal additions were made and the population was allowed to develop freely. Six microcosms were destructively sampled, three per treatment and at 2 week intervals, in order to enumerate collembola and fungal standing crop. Collembolan numbers were assessed after leaching by flooding the microcosms with distilled water and then gently stirring the litter. The collembola readily float to the surface and are easily counted. Determination of jiingal standing crop Litter derived from the sampled microcosms was examined for fungal standing crop using a modification of the method of Hering and Nicholson (1964). Litter was left overnight in bleaching solution, washed in distilled water, stained with phenolic aniline blue
and subsequently debyd~ted in alcohols. The bleached fragments were cleared in methyl salicylate so that hyphal lengths could be enumerated using phase contrast microscopy (Frankland, 1974) and the intersection method of Olson (1950). Ten fields per fragment and 20 fragments per microcosm were counted, at a magnification of x 400 and, by changing the depth of focus, the standing crop of fungi was partitioned into that present on the leaf surface and that within the leaf matrix. Hyphal lengths were converted to fungal standing crop using the factors given by Frankland rf al. (1978). Chemicul analysis qf leadmtrs Shortly after collection. leachates were analysed for pH using a Pye Unicam pH meter fitted with a combination electrode. Samples were stored at 5’C and analysed for K, Na and Ca using a Pye Unicam SP9 atomic absorption spectrophotometer. K and Na were determined using flame emission, Ca by atomic absorption using a nitrous oxide-acetylene flame. Ammonium-N and nitrate-N were measured on a Bemas continuous flow autoanalyser (Burkard Scientific. U.K.) using the alkaline phenate-nitroprusside and sulphanilamide-copper hydrazine methods, respectively (Allen ef al., 1974). Samples were normally analysed for all elements within 1 week of collection: any sample not being analysed within that time was stored at - 15°C and then thawed immediately before analysis. RESULTS
C/ranges in F. candida
population
Figure 1 shows changes in the number of live collembola recovered from the grazed microcosms during the course of the experiment. The collembola bred freely in the experimenta chambers and the numbers increased rapidly after week 6 from 20 individuals to a maximum of 300 individuals at week 10. The numbers then showed a significant decrease to around 240 per chamber by the end of the experiment at week 12. No animals were found in the control microcosms on any sampling occasion. Some mortality was observed amongst the initial 20 collembola added to the microcosms; possibly because of the inclusion of old individuals from the stock culture.
Observations of the gut contents of recovered collembola indicated that all size classes of itldividuals were actively grazing fungal hyphae.
Practically all of the fungal colonization of the leaf litter was external. with very little hyphae being seen within the leaf tissue. For this reason no distinction is drawn between internal and external colonization. and Fig. I shows total fungal standing crop for both the grazed and the ungrazed litter. The fungal standing crop increased substantially in both series over the first 4 weeks. with the grazed microcosm having a 53 per cent greater standing crop than the ungrazed system at this time. Subsequently the standing crop in the ungrazed microcosms remained contant. whilst that in the grazed series dropped markedly. By the end of the experiment the amount of fungal tissue in the grazed microcosms was only one third that present in the ungrazed ones.
Data for the N, Ca and Na release from the microcosms are given in Figs 2 and 3. Both NH,i and NO; release were significantly enhanced by the grazing of collembola. yet in absolute terms the enhancement of NO; release was very small. In contrast, the increase in NHf-N release became very marked towards the end of the study and by week 12 was over twice that of the controls. The enhancement of NH,‘-N release resulting from grazing only became apparent 8 weeks after animal addition and can be related to the rapid increase in collembolan population observed at that time (Fig. 1). Of the cations analysed only Ca showed any significant leaching response to grazing (Fig. 3), with Na and K losses remaining the same in treatments with and without animals. Similarly. the pH of leachate remained unaffected by the collembola populations.
In the ungrazed microcosms fungal development followed the classical growth curve exhibiting lag, exponential and stationary phases. In contrast, changes in fungal standing crop in the grazed microcosms were more complicated. showing a stimulation
Time,
Tame,
weeks
Fig. 1. Fungal standing crop on leaf litter with (e) and without (0) the effect of grazing. and changes in the collembola populations (A). Values shown are means + t SE.
weeks
Fig. 2. The effect of collembola feeding activities on mobilization of nitrogen as ammonium (0) and nitrate (A). Open symbols represent cultures without animals. Values shown are means f 1 SE.
Collembolan grazing and mineralization processes 200
r
Ttme,
weeks
Fig. 3. The effect of collembola feeding activities on mobilization of calcium (0) and sodium (A). Open symbols represent cultures without animals. Values shown are means + 1 SE.
of fungal growth at low grazing intensities (Fig. 1) similar to the results of Hanlon and Anderson (1979). This phenomenon may be caused by effects of grazing on the fungi, gut secretions or the remobilization of nutrients, but the relative contributions of these processes have not been quantified. At high grazing intensities, resulting from a marked increase in collembola numbers after 6 weeks (Fig. l), a reduction in fungal standing crop occurred. Parkinson et al. (1979) and Hanlon and Anderson (1979) also observed that grazing rates may exceed the production of fungal hyphae but Leonard (in Anderson and Ineson, 1983) has shown that the bafance of these processes is determined by the physical structure of the substrate and the available nutrient supply. Parkinson et al. (1979) showed that such grazing may be species selective, in that certain members of the fungal flora of litters are heavily grazed whereas other, less palatable species, remain unaffected or may even be stimulated at high animal levels. Although such specific grazing effects may be occurring here, the net effect was a reduction of total fungal standing crop by nearly 70 per cent. Examination of grazed leaf litter and gut contents for basidiomycete fungi suggested that animal grazing was selective. Basidiomycete hyphae were less severely grazed than non-basidiomycete hyphae, despite the fact that both groups were growing on the leaf surface and appeared equaliy accessible to animal attack. In the period between 6 and 8 weeks the number of collembola increased, fungal standing crop decreased, and increases in NH: and Ca leaching were observed as a consequence. Thus, the grazing of fungal hyphae or leaf material or both by the collembola resulted in increased nutrient loss from the litter. Nitrogen is readily immobilized by fungi in decomposing litter (Ausmus et al., 1976) and it appears that collembolan grazing inhibits this process. Thus, NH:-N normally released during leaf decomposition and subsequently used by soil fungi, is leached from the system when the fungal biomass is reduced. This hypothesis is supported by results from microcosm studies where stimulation of the microflora by glucose addition greatly reduced leaching losses of N (B&&h et al., 1978).
603
An alternative explanation for the observed increase in NH: leaching is that the N is derived from animal excretion or excretory products. Collembola lack Malphigian tubules and carry out storage excretion of N, accumulating uric acid in special urate cells in the fat bodies (Chapman, 1972). It is therefore unlikely that the NH: observed here was derived from excretory sources. However, ~llembola moult frequently and the exuviae produced rapidly disappear in litter culture, either via decomposition or ingestion by the animal (Poole, 1952). The amount of inorganic N released by these routes has not been measured. Whatever the mechanism, collembolan grazing of leaf litter increases mineralization of N, and this N is in a form suitable for uptake by primary producers. The increases in NO; release from the decomposing litter were very small in absolute terms, yet represented a doubling of inorganic N in this form. Nitrate may have been derived heterotrophically from grazed fungi, or alternatively, from autotrophic nitrification stimulated by increased NH: availability. In contrast to our results, Visser et al. (1981) failed to detect any change in NO; leaching from decomposing aspen poplar leaves exposed to grazing by Onychiurus &tent&, however their study terminated after only 10 days and similar conclusions could have been drawn from our work after the same period. The effect of collembolan grazing on cation release was far less pronounced than that for NH:. On only one occasion did grazing significantly affect cation release; Ca2+ leaching at week 8 was increased by 13 per cent over control values. Although this trend remained for Ca2+ throughout the remainder of the study, week 8 was the only time that it differed significantly from control. KC and Na+ leaching remained unaffected by the animals, but much of the initial concentrations of these highly mobile ions will have been leached during the preparatory microcosm incubation. Ca’+, unlike K” and Na+, is a structurally important ion in higher plants and therefore less subject to leaching. Table 1 summarizes the results of the experiment and provides estimates of the nutrient pools in the fungi and collembola. The data emphasize the relatively low order of nutrients immobilized in the organisms in comparison to those of the litter. Leaching losses of N, Ca, K and Na represent, respectively, 1.0, 8.5, 28.9 and 20.8 per cent of the litter content in the ungrazed system. Approximately 30 per cent of the N released from the fungal tissue by grazing is accounted for by increases in the collembolan population but only 5 of the remaining 70 per cent was seen as leachate loss. Collembola grazing above optimal intensity causes an increase in bacterial biomass (Hanlon and Anderson, 1979) and the transfer of the remaining 65 per cent (24mg) of mobilized fungal N to this pool is within the scale of the observed effects in experimental systems. We do not, however, have direct evidence for the transfer of N between fungal and bacterial biomass in these experiments. The low concentration of cations in insect tissue resulted in a small collembola pool of Ca, K and Na and the same was true for the fungal pool, with the exception of Ca. The process of Ca sequestration by fungal mycelium has been described by Cromack et
Table 1. Initial litter nutrient content and changes in the location 01 nutrients with and without collcmholan grazing (mg g-’ dry wt leaf tissue) Componenl Initial litter content Fungal standing crop* ungrazed grared dit&rence
Nitrogen
Calcium
I I .I0
13.10
0.38
1.43
0.56
0.66
0.72 0.44
0.0’7 0.01 0.01
0.02
0.19 0.37
0.00 0.1 1 0.1 I
0.00
0.00 0.00 0.00
0.00 0.01 0.01
0.12
1.11 0.19 0.08
Collembola standing crop? ungtazed grazed difference Leaching losses ungrazed grazed difference
0.14 0.02
0.00
0.00
Sodium Potassium
O.il 0.10 -0.01
0.01 0.01
0.30 u.33 0.03
* Values for iiutriellt content of litter fungi from Swift et (II. (19X0). t Values for nutrient cantent of collembola from Allen ct tll. (1974). Collembolan standing crops derived from regression equations of Petersen (1975).
01.(1975). Since there were no significant changes in K and Na leaching as a consequence of grazing, these ions do not appear to transfer directly from fungal standing crop to leachate. However, the limited amount of these ions present in the fungal pool (Table 1) means that such transfers would be analytically difficult to detect. Although the experiments described here were carried out in the laboratory, the collembola densities recorded in the microcosms were similar to those determined for natural ~pulations (Usher, 1969. 1976: Grkgoire-Wibo, 1979). We conclude, therefore. that aggregations of collembola and other microarthropods will enhance Ca and N mineralization processes through short term effects on the litter microflora. Nutrients immobilized in animal biomass are released by more long-term, spatio-temporally unpredictable mortality processes. The effects of microarthropods on nutrient fluxes, particularly N mineralization, appear to be small in comparison with those of litterfeeding macroarthropods (Anderson and Ineson. 1983) but could significantly influence nutrient availability in acid forest soils where large immobilized nutrient pools are located in the accumulated organic matter.
Acknorvledyements-We thank Sally Huish for skilful technical assistance and the Royal Society for provision of analytical equipment. This work was supported by the Natural Environment Research Council.
REFERENCES S., GKIMSHAW M., PARKINSONJ. and QUARMBY C. (1974) Chen~ical Analysis of Ecological Materiats. Black-
ALLEN
well, Oxford. AN~ERXI~ J. M. and INESONP. (1982) A soil microcosm system and its application to measurements of respir-
ation and nutrient leaching. Soil Bio(og) & Einchen~ibrr> 14.415-416. ANDERSOUJ. M. and INESONP. (1983) interactions between soil arthropods and microbial populations in carbon, nitrogen and mineral nutrient fluxes from decomposing leaf litter. In Nirrogen as tin Ecological Facrur (J. A. Lee, S. McNeil1 and I. H. Rorison. Eds). Blackwell. Oxford (in press). AUDERSON R. V., COLEMA% D, C. and COLE C. V. (1981) Effects of saprotrophic grazing on net mineralization. In Terrestrial hriirogen Cycles. Processes, tegies and ~~~affage~t~e~t.~ impacts (F.
Roswall. 201&236. AUSM~S B.
Ecosystems
Stra-
E. Clark and T. Eds). ~e~~~~j~u~ Btrliefins (S~~)e~~~o/?~) 33,
S., EDWARDS N. T. and WITI;AMP M. (1976) Microb~di immobilization of carbon. nitrogen. phosphorus and potassium: implications for forest ecosystem processes. In The Role of’ Terrestrial and Aquatic Oryurb isms in Decomposition Processes. (J. M. Anderson and A. Macfadyen. Eds), pp. 397-416. Blackwell. Oxford. I%&TH E.. LOIIM U.. LUNDGRENB.. RWSWALL T.. SBIXRsr~RijMB., S~HLENIUSB. and WIKL!NA. (1978) The effect of nitrogen and carbon supply on the development of soil organism populations and pine seedlings: a microcosm experiment. Oikos 31. 153-163. CHAPMAWR. F. (19721 Tlrr Insects. English Universities Press, London. CROMACEK. JR, TODD R. L. and MOE;KC. D. (1975) Patterns of basidiomycete nutrient accumu~dtion in conifer and deciduous forest litter. Soi! Bj~~~o~~ & ~i~e~t~/~ti.~r~~ 7, 265-768. EDWARDSN. T.. SHUGAR~H. H. JR, MCLAUGHLIN S. B., HARRIS W. F. and REICHLED. E. (1981) Carbon metabolism in terrestrial ecosystems. in ~~nff~z~eProperties of Forest Ecosystems (D. E. Reichle. Ed.), .oo. . 499-536. Cambridge University Press. GOTO H. F. (1960) Simnle techniaues for the rearine of ~oIlembola and a note on the use of fungistatic substance in the cultures. ~F:torn~~ogjsts ~ont~~~~ ~u~u=i~e 96. 138-140. GR&GOIRE-WIBO C. (1979) Cycle phCnologique de Foisomia ~uudriocl~laza en for&. &males de ta Societe ropzle zoologique de ~e~gi~ae 109, 43-65. HANLONR. D. G. and AND~SON J. M. (1979) The effect of
Collembolan grazing and mineralization processes Collembola grazing on microbial activity in decomposition of leaf litter. Oecologia (Berlin) 38, 93-99. HARIXNG D. J. L. and STUTTARDR. A. (1974) Microarthropods. In BioEogy of Plant Litter Decomposition. (C. H. Dickinson and G. J. F. Pugh, Eds), Vol. 2, pp. 4X9-532. Academic Press, London. %3RING T. F. and NICHGL%?NP. B. (1964) A clearing technique for the examination of fur@ in plant tissue. Nature 201, 942-943. FRANKLAND 3. C. t1974) fmportance of phase contrast microscopy for estimation of mycelial biomass in leaf litter. Soil Biology & Biochemistry 7, 408410. FRANKLANDJ. C.. LINDLEYD. K. and SWIFT M. J. (1978) A comparison of two methods for the estimation of myceha1 biomass in leaf litter. Soil Biology & Biochemistry 10, 323-333. OLSONF. C. W. (1950) Quantitative estimates of filamentous algae. Tr~~~~ci~o~sof tlze American ~~~rascQp~~a1 society
59, 272-279.
PAKKKNSOND., Vrssm of
Pedobiologia
of various Collembola
species.
15, 222-243.
POOLET. B. (1952) Studies on the food of callembola in a Douglas fir plantation. Proceedings of the Zoological Society ofLondon 132, 71-82. SEASTEDTT. R. and CROSSLEYD. A. JR (1980) Effects of microarthropods on the seasonal dynamics of nutrients in forest litter. Srrii 3~~~~9~d 3jff&~~~~~srr~ 12, 337342. SU’IFTM. J.. HEAL 0. W. and ANDERSONJ. M. IL98Oi Decomposition ‘in Terrestrial Ecosystems. Studies in Ecology, Vol. 5. Blackwell, Oxford. Usnra M. B. (1969) Some properties of the aggregations in soil arthropods: Collembola. Journal of Animal Ecoloqt _. 38, 607-622. USHERM. B. (1976) Aggregation responses of soil arthropods in relation to the soil environment. In The Role of’ Terrestrial and Aquatic Organisms in Decomposition Pro”cesses (J. M. Anderson and A. Macfadyen, Ed+. pp.
6 1-94. BlackweIt, Oxford. S. and WHITTAKER
J_
B,
(1979)
colIembo~an grazing on fungal cotonization of leaf litter. Soii Biofog_r & R~~~~ern~stryIt, 529-535. PETERSENH. (1975) Estimation of dry weight, fresh weight, Effects
and calorific content
605
VISSER S., WHETAKER J. 33.
and PARKIXSON D. (1981) Effects of colkembolan grazing on nutrient release and respiration of a leaf litter inhabiting fungus. Soil Bioirtgy & Biochemistry 13, 215-218.
Soil Biol. Bioche~~~.Vol. 14, pp. 601 to 605, 1982 Printed in Great Britain. All rights reserved
EFFECT OF COLLEMBOLAN GRAZING UPON NITROGEN AND CATION LEACHING FROM DECOMPOSING LEAF LITTER P. INESON.M. A. LEONARDand J. M. ANDERSON Department of Biological Sciences, Wolfson
Ecology Laboratory, Exeter EX4 4PS. Devon, U.K.
(Accepted 20 March
University
of Exeter.
1982)
Summary-Fragmented oak litter was incubated in the laboratory for 4 months with and without collembola. The effects of the animals upon fungal standing crop and leaching of inorganic nitrogen and cations was monitored over this period. The results showed that the fungal standing crop was higher in the presence of small numbers of animals than in litter lacking animals, yet at higher grazing intensities the fungal standing crop fell markedly. Significant increases in the leaching of ammonium, nitrate and calcium occurred as a consequence of animal grazing. but potassium and sodium losses from the litter were unaffected.
INTRODUCTION Despite the small contribution that saprotrophic microarthropods make towards total carbon mineralization in forest and woodland soils (Edwards et a/., 1981) they have an important role in affecting the quantity, community composition and activity of the soil microflora (Hanlon and Anderson. 1979; Parkinson et al., 1979). Studies suggest that protozoa, nematodes and annelids are important in soil N mineralization processes (Anderson et al., 1981) yet there are few data concerning the part played by microarthropods in nutrient transformations in soils. despite much speculation (Harding and Stuttard, 1974; Seastedt and Crossley, 1980; Anderson et al., 1981). Fungal mycelium in soil can contain a major proportion of nitrogen, potassium and other cations in the soil pool (Cromack et al.. 1975) and thus the sensitivity of fungal hyphae to microarthropod grazing may well have considerable significance for nutrient mobilization (Hanlon and Anderson, 1979; Parkinson et al., 1979). In the experiment described here a microcosm system was used to follow changes in collembolan numbers, fungal standing crop and cation and ammonium release over several months in grazed and ungrazed litter. The aim was to assess the functional significance of collembolan grazing in nutrient release from decomposing litter. MATERIALS AND Preparation
METHODS
and watering of microcosms
Oak (Quercus robur L.) leaves were collected shortly after leaf fall from the litter layer of a mature oak woodland, air dried at 20°C and brushed free of faecal material and debris. Midribs were removed and the remaining laminae cut into small fragments of 0.5-l.Ocm. The fragments were mixed well and aliquots, 1.5 g, placed in microcosm chambers (Anderson and Ineson, 1982). 601 S.BB 146-F
The litter in each microcosm was rehydrated by the addition of 100 ml of distilled water for 24 h, which served to leach out soluble tannins and readily-metabolizable materials. The litter was drained and inoculated with a coarsely sieved suspension of macerated fresh leaf litter and then incubated in a cooled cabinet at 15°C. Microcosms were placed according to a randomized block design and were leached weekly with 60 ml of distilled water, leachates being retained for chemical analysis. Prior to animal addition three microcosms were sampled to determine litter dry weight and nutrient content. Culture, addition and enumeration
of collembola
Folsomia candida Will. were maintained in the laboratory on a diet of dried baker’s yeast and housed in plastic boxes containing a substrate of plaster of Paris and activated charcoal (Goto. 1960). Individuals were selected for experimentation by passing the culture through a 0.3 mm pore size sieve, those being retained were added, 20 per microcosm, to half of the microcosms. The additions were not made until 6 weeks after initial leaching, permitting time for the development of a decomposer flora. No further animal additions were made and the population was allowed to develop freely. Six microcosms were destructively sampled, three per treatment and at 2 week intervals, in order to enumerate collembola and fungal standing crop. Collembolan numbers were assessed after leaching by flooding the microcosms with distilled water and then gently stirring the litter. The collembola readily float to the surface and are easily counted. Determination of jiingal standing crop Litter derived from the sampled microcosms was examined for fungal standing crop using a modification of the method of Hering and Nicholson (1964). Litter was left overnight in bleaching solution, washed in distilled water, stained with phenolic aniline blue
and subsequently debyd~ted in alcohols. The bleached fragments were cleared in methyl salicylate so that hyphal lengths could be enumerated using phase contrast microscopy (Frankland, 1974) and the intersection method of Olson (1950). Ten fields per fragment and 20 fragments per microcosm were counted, at a magnification of x 400 and, by changing the depth of focus, the standing crop of fungi was partitioned into that present on the leaf surface and that within the leaf matrix. Hyphal lengths were converted to fungal standing crop using the factors given by Frankland rf al. (1978). Chemicul analysis qf leadmtrs Shortly after collection. leachates were analysed for pH using a Pye Unicam pH meter fitted with a combination electrode. Samples were stored at 5’C and analysed for K, Na and Ca using a Pye Unicam SP9 atomic absorption spectrophotometer. K and Na were determined using flame emission, Ca by atomic absorption using a nitrous oxide-acetylene flame. Ammonium-N and nitrate-N were measured on a Bemas continuous flow autoanalyser (Burkard Scientific. U.K.) using the alkaline phenate-nitroprusside and sulphanilamide-copper hydrazine methods, respectively (Allen ef al., 1974). Samples were normally analysed for all elements within 1 week of collection: any sample not being analysed within that time was stored at - 15°C and then thawed immediately before analysis. RESULTS
C/ranges in F. candida
population
Figure 1 shows changes in the number of live collembola recovered from the grazed microcosms during the course of the experiment. The collembola bred freely in the experimenta chambers and the numbers increased rapidly after week 6 from 20 individuals to a maximum of 300 individuals at week 10. The numbers then showed a significant decrease to around 240 per chamber by the end of the experiment at week 12. No animals were found in the control microcosms on any sampling occasion. Some mortality was observed amongst the initial 20 collembola added to the microcosms; possibly because of the inclusion of old individuals from the stock culture.
Observations of the gut contents of recovered collembola indicated that all size classes of itldividuals were actively grazing fungal hyphae.
Practically all of the fungal colonization of the leaf litter was external. with very little hyphae being seen within the leaf tissue. For this reason no distinction is drawn between internal and external colonization. and Fig. I shows total fungal standing crop for both the grazed and the ungrazed litter. The fungal standing crop increased substantially in both series over the first 4 weeks. with the grazed microcosm having a 53 per cent greater standing crop than the ungrazed system at this time. Subsequently the standing crop in the ungrazed microcosms remained contant. whilst that in the grazed series dropped markedly. By the end of the experiment the amount of fungal tissue in the grazed microcosms was only one third that present in the ungrazed ones.
Data for the N, Ca and Na release from the microcosms are given in Figs 2 and 3. Both NH,i and NO; release were significantly enhanced by the grazing of collembola. yet in absolute terms the enhancement of NO; release was very small. In contrast, the increase in NHf-N release became very marked towards the end of the study and by week 12 was over twice that of the controls. The enhancement of NH,‘-N release resulting from grazing only became apparent 8 weeks after animal addition and can be related to the rapid increase in collembolan population observed at that time (Fig. 1). Of the cations analysed only Ca showed any significant leaching response to grazing (Fig. 3), with Na and K losses remaining the same in treatments with and without animals. Similarly. the pH of leachate remained unaffected by the collembola populations.
In the ungrazed microcosms fungal development followed the classical growth curve exhibiting lag, exponential and stationary phases. In contrast, changes in fungal standing crop in the grazed microcosms were more complicated. showing a stimulation
Time,
Tame,
weeks
Fig. 1. Fungal standing crop on leaf litter with (e) and without (0) the effect of grazing. and changes in the collembola populations (A). Values shown are means + t SE.
weeks
Fig. 2. The effect of collembola feeding activities on mobilization of nitrogen as ammonium (0) and nitrate (A). Open symbols represent cultures without animals. Values shown are means f 1 SE.
Collembolan grazing and mineralization processes 200
r
Ttme,
weeks
Fig. 3. The effect of collembola feeding activities on mobilization of calcium (0) and sodium (A). Open symbols represent cultures without animals. Values shown are means + 1 SE.
of fungal growth at low grazing intensities (Fig. 1) similar to the results of Hanlon and Anderson (1979). This phenomenon may be caused by effects of grazing on the fungi, gut secretions or the remobilization of nutrients, but the relative contributions of these processes have not been quantified. At high grazing intensities, resulting from a marked increase in collembola numbers after 6 weeks (Fig. l), a reduction in fungal standing crop occurred. Parkinson et al. (1979) and Hanlon and Anderson (1979) also observed that grazing rates may exceed the production of fungal hyphae but Leonard (in Anderson and Ineson, 1983) has shown that the bafance of these processes is determined by the physical structure of the substrate and the available nutrient supply. Parkinson et al. (1979) showed that such grazing may be species selective, in that certain members of the fungal flora of litters are heavily grazed whereas other, less palatable species, remain unaffected or may even be stimulated at high animal levels. Although such specific grazing effects may be occurring here, the net effect was a reduction of total fungal standing crop by nearly 70 per cent. Examination of grazed leaf litter and gut contents for basidiomycete fungi suggested that animal grazing was selective. Basidiomycete hyphae were less severely grazed than non-basidiomycete hyphae, despite the fact that both groups were growing on the leaf surface and appeared equaliy accessible to animal attack. In the period between 6 and 8 weeks the number of collembola increased, fungal standing crop decreased, and increases in NH: and Ca leaching were observed as a consequence. Thus, the grazing of fungal hyphae or leaf material or both by the collembola resulted in increased nutrient loss from the litter. Nitrogen is readily immobilized by fungi in decomposing litter (Ausmus et al., 1976) and it appears that collembolan grazing inhibits this process. Thus, NH:-N normally released during leaf decomposition and subsequently used by soil fungi, is leached from the system when the fungal biomass is reduced. This hypothesis is supported by results from microcosm studies where stimulation of the microflora by glucose addition greatly reduced leaching losses of N (B&&h et al., 1978).
603
An alternative explanation for the observed increase in NH: leaching is that the N is derived from animal excretion or excretory products. Collembola lack Malphigian tubules and carry out storage excretion of N, accumulating uric acid in special urate cells in the fat bodies (Chapman, 1972). It is therefore unlikely that the NH: observed here was derived from excretory sources. However, ~llembola moult frequently and the exuviae produced rapidly disappear in litter culture, either via decomposition or ingestion by the animal (Poole, 1952). The amount of inorganic N released by these routes has not been measured. Whatever the mechanism, collembolan grazing of leaf litter increases mineralization of N, and this N is in a form suitable for uptake by primary producers. The increases in NO; release from the decomposing litter were very small in absolute terms, yet represented a doubling of inorganic N in this form. Nitrate may have been derived heterotrophically from grazed fungi, or alternatively, from autotrophic nitrification stimulated by increased NH: availability. In contrast to our results, Visser et al. (1981) failed to detect any change in NO; leaching from decomposing aspen poplar leaves exposed to grazing by Onychiurus &tent&, however their study terminated after only 10 days and similar conclusions could have been drawn from our work after the same period. The effect of collembolan grazing on cation release was far less pronounced than that for NH:. On only one occasion did grazing significantly affect cation release; Ca2+ leaching at week 8 was increased by 13 per cent over control values. Although this trend remained for Ca2+ throughout the remainder of the study, week 8 was the only time that it differed significantly from control. KC and Na+ leaching remained unaffected by the animals, but much of the initial concentrations of these highly mobile ions will have been leached during the preparatory microcosm incubation. Ca’+, unlike K” and Na+, is a structurally important ion in higher plants and therefore less subject to leaching. Table 1 summarizes the results of the experiment and provides estimates of the nutrient pools in the fungi and collembola. The data emphasize the relatively low order of nutrients immobilized in the organisms in comparison to those of the litter. Leaching losses of N, Ca, K and Na represent, respectively, 1.0, 8.5, 28.9 and 20.8 per cent of the litter content in the ungrazed system. Approximately 30 per cent of the N released from the fungal tissue by grazing is accounted for by increases in the collembolan population but only 5 of the remaining 70 per cent was seen as leachate loss. Collembola grazing above optimal intensity causes an increase in bacterial biomass (Hanlon and Anderson, 1979) and the transfer of the remaining 65 per cent (24mg) of mobilized fungal N to this pool is within the scale of the observed effects in experimental systems. We do not, however, have direct evidence for the transfer of N between fungal and bacterial biomass in these experiments. The low concentration of cations in insect tissue resulted in a small collembola pool of Ca, K and Na and the same was true for the fungal pool, with the exception of Ca. The process of Ca sequestration by fungal mycelium has been described by Cromack et
Table 1. Initial litter nutrient content and changes in the location 01 nutrients with and without collcmholan grazing (mg g-’ dry wt leaf tissue) Componenl Initial litter content Fungal standing crop* ungrazed grared dit&rence
Nitrogen
Calcium
I I .I0
13.10
0.38
1.43
0.56
0.66
0.72 0.44
0.0’7 0.01 0.01
0.02
0.19 0.37
0.00 0.1 1 0.1 I
0.00
0.00 0.00 0.00
0.00 0.01 0.01
0.12
1.11 0.19 0.08
Collembola standing crop? ungtazed grazed difference Leaching losses ungrazed grazed difference
0.14 0.02
0.00
0.00
Sodium Potassium
O.il 0.10 -0.01
0.01 0.01
0.30 u.33 0.03
* Values for iiutriellt content of litter fungi from Swift et (II. (19X0). t Values for nutrient cantent of collembola from Allen ct tll. (1974). Collembolan standing crops derived from regression equations of Petersen (1975).
01.(1975). Since there were no significant changes in K and Na leaching as a consequence of grazing, these ions do not appear to transfer directly from fungal standing crop to leachate. However, the limited amount of these ions present in the fungal pool (Table 1) means that such transfers would be analytically difficult to detect. Although the experiments described here were carried out in the laboratory, the collembola densities recorded in the microcosms were similar to those determined for natural ~pulations (Usher, 1969. 1976: Grkgoire-Wibo, 1979). We conclude, therefore. that aggregations of collembola and other microarthropods will enhance Ca and N mineralization processes through short term effects on the litter microflora. Nutrients immobilized in animal biomass are released by more long-term, spatio-temporally unpredictable mortality processes. The effects of microarthropods on nutrient fluxes, particularly N mineralization, appear to be small in comparison with those of litterfeeding macroarthropods (Anderson and Ineson. 1983) but could significantly influence nutrient availability in acid forest soils where large immobilized nutrient pools are located in the accumulated organic matter.
Acknorvledyements-We thank Sally Huish for skilful technical assistance and the Royal Society for provision of analytical equipment. This work was supported by the Natural Environment Research Council.
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