EFFECTS OF TREE SPECIES, STAND AGE AND SOIL
TYPE ON SOIL MICROBIAL BIOMASS AND ITS ACTIVITY
IN A SOUTHERN BOREAL FOREST
J. BAUHUS,1
* D. PAREÂ2,3
and L. COÃTEÂ3
1Department of Forestry, Australian National University, Canberra, ACT 0200, Australia,2UniversiteÂ
du QueÂbec aÁ MontreÂal, DeÂpartment des Sciences Biologiques, Groupe de Recherche en EÂcologie ForestieÁre, C.P. 8888, Succursale Centre-ville, Montreal, Que., Canada H3C 3P8 and3BiodoÃme de
MontreÂal, 4777 avenue Pierre-de-Coubertin, Montreal, Que., Canada H1V 1B3
(Accepted 18 August 1997)
SummaryÐMicrobial C (Cmic) and N (Nmic), the Cmic-to-organic C (Corg) and Nmic-to-total N (Nt)
ratios and the speci®c respiration of microbial biomass were investigated in a southern boreal mixed forest. The forest stands were 50 and 124 years old and consisted of trembling aspen, paper birch and mixed conifers comprising white spruce and balsam ®r. Stands were growing on soils derived either from clay (89% average clay content) or till (46% average clay content) deposits in the clay belt region of northern Quebec. In the forest ¯oors the relative concentrations of microbial C and N and the Cmic
-to-Corgand Nmic-to-Ntratios, regarded as measures of organic matter quality, declined with stand age
whereas the speci®c microbial respiration increased, indicating decreasing C assimilation eciency. In the mineral soils, in contrast, Cmic-to-Corgand Nmic-to-Ntratios increased with stand age. The Cmic
-to-Nmic ratio widened with stand age in both the forest ¯oors and mineral soils, suggesting that the
proportion of fungi had increased. Concentrations of microbial C and N were on average lower in forest ¯oor beneath conifers (Cmic-to-Corg1.9%, Nmic-to-Nt7.5%) than beneath the deciduous species
birch (Cmic-to-Corg2.2%, Nmic-to-Nt8.6%) and aspen (Cmic-to-Corg2.4%, Nmic-to-Nt9.2%). Average
Cmic-to-Nmic ratios were only slightly dierent in the forest ¯oors beneath the dierent tree species
(Cmic-to-Nmic: conifers 8.9, birch 7.2, and aspen 8.3). In both forest ¯oors and mineral soils, average
concentrations of Cmicand Nmicwere generally higher in the clay than in the till soils, but the Cmic
-to-Corgratios were similar in both soil types. The average Nmic-to-Ntratios were lower in till than in clay
soils only beneath conifers. The average speci®c microbial respiration (qCO2=mg CO2-C mg Cmicÿ1dÿ1)
in clay soils (22) was approximately half that in till soils (41). Since the microbial parameters measured were sensitive to the factors stand age, tree species and soil type, they may have the potential to be used as indicators of the in¯uence of forest management on soil organic matter quality. # 1998 Elsevier Science Ltd. All rights reserved
INTRODUCTION
The soil micro¯ora is a small but signi®cant com-ponent in most terrestrial ecosystems. Soil microbial activity contributes to the regulation of soil carbon storage, soil respiration and ecosystem productivity. The role of soil microbial biomass as a relatively labile nutrient pool in the cycling of C, N and P is well established (Marumotoet al., 1982b; Van Veen et al., 1987; Duxbury et al., 1989, Jenkinson and Parry, 1989). Amounts of microbial biomass are in¯uenced by soil texture and soil organic matter (SOM) quality (Wardle, 1992; Ross and Tate, 1993, Bosatta and AÊgren, 1994; Hassink, 1994). The quantity and composition of microbial biomass is sensitive to changes in the soil chemical and physi-cal environment (Wolters and Joergensen, 1991; Wardle, 1992; Bauhus and Khanna, 1994; Beck et al., 1995). In addition, it has been shown that amounts of soil microbial C and N can be
in¯u-enced by forest management (Ohtonen et al., 1992; Bauhus and Barthel, 1995; PietikaÈinen and Fritze, 1995) and increase proportionally with forest pro-ductivity (Myrold et al., 1989). From the above it has been suggested that concentrations of microbial C and N may be regarded as sensitive indicators for changes in the soil ecosystem. Normally the quan-tity and quality of the bulk of SOM changes only slowly and because of the large background value of soil C and the spatial heterogeneity of soils, changes are dicult to measure. Therefore, microbial biomass and, in particular, the ratio of microbial C and N to total organic C and N have been suggested as indicators of the state and modi®-cation of SOM (Anderson and Domsch, 1989; Wolters and Joergensen, 1991; Bosatta and AÊgren, 1994). Soil microbial biomass is usually resource limited and thus microbial C and N concentrations are generally related to amounts of soil C and N (Wardle, 1992). The ratio of microbial C to soil or-ganic carbon has thus been used as an indicator for C availability (Anderson and Domsch, 1986; Insam #1998 Elsevier Science Ltd. All rights reserved Printed in Great Britain 0038-0717/98 $19.00 + 0.00 PII: S0038-0717(97)00213-7
*Author for correspondence.
and Domsch, 1988). Further, the incorporation of N in microbial biomass is aected by the avail-ability of C and N and can be expressed by the pro-portion of microbial N to total soil N, the Nmic
-to-Nt ratio and the Cmic-to-Nmic ratio (Joergensen et
al., 1995).
The amount of information on soil microbial bio-mass in forested ecosystems is limited when com-pared to agricultural systems (Wardle, 1992). This is especially true for the vast area of boreal forests. In this study we wanted to determine how microbial biomass and its activity in the southern boreal for-est is aected by the dominant tree species, soil type and stand age. Our hypotheses were:
(a) The ratio of microbial C to total organic C (Cmic-to-Corg) and the ratio of microbial N to total
N (Nmic-to-Nt) are related to substrate quality. If
this were true, the Cmic-to-Corg and the Nmic-to-Nt
ratios would be expected to be higher in a rich soil substrate than in a poor one, in a forest ¯oor beneath deciduous trees than beneath conifers and they should also decrease with stand age in aggrad-ing forests. The availability of nutrients in the forest ¯oor decreased with the stand age after ®re and was higher in birch and aspen than in coniferous stands (PareÂet al., 1993). Forest ¯oor decomposition rates were related to fungal biomass and were higher in deciduous than in coniferous boreal forests (Flanagan and Van Cleve, 1983). Since the pro-portion of woody litter increases with stand age substrate quality in the forest ¯oor should then decrease (Klinkaet al., 1995).
(b) The speci®c microbial respiration (qCO2=mg
CO2-C mg Cmicÿ1dÿ1) decreases with increasing sub-strate quality. Several studies (e.g. Anderson and Domsch, 1993; Wardle and Ghani, 1995) have shown that speci®c microbial respiration is higher under unfavourable than under favourable con-ditions. Thus speci®c microbial respiration should increase with stand age and should be higher in a poor soil and a coniferous forest ¯oor than in a rich soil and a deciduous forest ¯oor.
MATERIALS AND METHODS
Field sites
The study area was located in the southern boreal forest of Quebec, in the Abitibi region around Lac Duparquet (48830'N, 79820'W). The continental cli-mate is characterized by a mean annual temperature of 0.68C, mean annual precipitation of 823 mm and a frost-free period of 64 days. The study area is part of the northern Clay Belt, where most soils originate from glaciolacustrine clay deposits (Vincent and Hardy, 1977). Forests in the region are mostly composed of trembling aspen (Populus tremuloides) and paper birch (Betula papyrifera) in young successional stages and converge gradually towards dominance of conifers such as balsam ®r
(Abies balsamea), white spruce (Picea glauca) and cedar (Thuja occidentalis) in later successional stages (Bergeron and Dansereau, 1993). The age of forest stands in the area has been determined in dendro-chronological studies (Dansereau and Bergeron, 1993).
Field plots were located in stands originating from ®res in 1944 and 1870. They included 50 and 124 year old stands of aspen, birch and conifers (®r and spruce) on clay and till soils. The patchy occur-rence of conifers in the forest did not allow us to locate plots in pure stands of only one coniferous species. Therefore Abies balsameaandPicea glauca were combined into one conifer group. Thus, although in the following text, plots will be named according to the dominant species or species group, the composition was not always entirely monospeci-®c. Aspen stands in the 124 year old forest were in their second natural rotation (Pare and Bergeron, 1995). Aspen stands disintegrate and rejuvenate at around 70±80 years of age and aspen trees in the 124 year old forest were thus approximately of the same age as in the 50 year old forest. The clay soils were Grey Luvisols with moderate to good drainage and the till soils developed on moraine deposits were classi®ed as Humo-Ferric Podzols (Agriculture Canada Expert Committee on Soil Survey, 1987). Within a randomized design for the factors stand age, tree species and soil type, each treatment was replicated four times amounting to 48 plots of 1010 m in size.
Field sampling
Mineral soil (0±10 cm) and forest ¯oor (OF- and
OH-layer) samples were collected from four
lo-cations at each plot in October 1994. These four samples were combined to form one composite sample for each plot. The forest ¯oor was not sep-arated into OF- and OH-layers, because the layers
were closely attached by a dense ®ne root mat. Mineral soil samples were collected using a 50 mm diameter soil corer. Samples were refrigerated (48C) until further processing in the laboratory. We sampled soils for microbial biomass determinations only once because other studies had shown that amounts of microbial C and N ¯uctuate to only a small extent throughout the year (Patraet al., 1990; Bauhus and Barthel, 1995). We also assumed that potential seasonal ¯uctuations would not disguise dierences between tree species, soil type or stand age. Forest ¯oor samples were sieved (5 mm mesh) to remove coarse woody parts and roots. The till soils were also sieved (2 mm), whereas the clay soils were homogenized by hand because they were too moist for sieving.
Soil analysis
sub-samples, pH was determined in bi-distilled water and in KCl solution; substrate-to-water ratios were 1-to-2 for mineral soil and 1-to-3 for forest ¯oor material. Organic matter and Corg concentrations
were determined by loss on ignition after ashing at 5508C for 2 h; Corg was derived by multiplying the
organic matter concentration by the factor 0.58 (Allen, 1989). Total nitrogen (Nt) was determined
colorimetrically (Tecator FIA Star 5020 Analyzer) following wet digestion in sulphuric acid using mer-cury oxide as a catalyst (Allen, 1989). Bray II-extractable P (McKeague, 1976) was analyzed spec-trophotometrically (Milton Roy, Spectronic 1001plus) at 880 nm. Exchangeable cations were determined in 1M NH4NO3 (Stuanes et al., 1984)
and measured using atomic absorption spectropho-tometry. Particle size distribution of mineral soil samples was determined by the hydrometer method (Sheldrick and Wang, 1993).
Microbial C (Cmic) and N (Nmic) were determined
using the chloroform fumigation±extraction method (Brookeset al., 1985, Vanceet al., 1987). Cmic and
Nmicwere calculated as the dierences in organic C
and total N between fumigated and non-fumigated (control) samples. Two replicates of each sample, 5 g from the organic layer or 10 g mineral soil, were fumigated for 24 h with ethanol-free chloroform at 258C. Subsequently the chloroform was removed by evacuation. Fumigated samples and controls were extracted with 50 ml 0.5M K2SO4 and ®ltered
(Whatman 42); extracts were kept frozen until ana-lyzed. Organic C in K2SO4extracts was determined
by dichromate digestion (Walkley and Black, 1934; Vance et al., 1987) followed by potentiometric (Metrohm, Herisau) end point titration (Raveh and Avnimelech, 1972). Total nitrogen in K2SO4
extracts was determined colorimetrically (Tecator FIA Star 5020 Analyzer) following alkaline persul-fate oxidation (Cabrera and Beare, 1993). The per-sulfate oxidation procedure oxidizes both organic N and ammonium, with N being analyzed as nitrate. Our evaluation of the method using ammonium-nitrate and urea standards showed full recovery of N in K2SO4 extracts. Non-extractable amounts of
microbial C and N were compensated for by using
a correction factor of kC=2.86 (Sparling et al.,
1990) for soils with a high organic C content and kN=1.85 (Brookes et al., 1985; Joergensen and
Mueller, 1996), respectively.
The speci®c respiration rate (Anderson and Domsch, 1990) (qCO2=mg CO2-C mg Cmicÿ1dÿ1) was determined during incubation of fresh soil at room temperature (228C) in sealed tubes after adjusting the water content to 300% in forest ¯oor material and to 40% in mineral soil 3 days prior to measurement. The respiration rate for these samples, measured as CO2 evolution, was
deter-mined by injecting samples of the head space gas of soil samples incubated for 3 h in Vacutainers into a gas chromatograph (Licor 6200). Values for speci®c respiration represent the means of two measure-ments.
Statistics
The eect of the experimental factors, stand age, tree species and soil type on microbial and soil vari-ables was tested by means of ANOVA using the SYSTAT package (Systat Inc., 1992). Homogeneity of variances was examined with Levene's test (Snedecor and Cochran, 1980). In only two cases were the variances between factors not hom-ogenous. In one of the cases, homoscedasticisty was achieved by hyperbolic transformation of the data. In the other case, data were rank transformed since none of the regular transformations yielded hom-ogeneity of variances (Conover and Iman, 1981; Potvin and Ro, 1993). An ANOVA on these ranked data provided similar results for signi®cance and non-signi®cance of factors and interactions as the ANOVA on the non-transformed data. Regression analysis (general linear or non-linear models) was used to assess the in¯uence of soil properties on microbial biomass and the interaction between microbial variables.
RESULTS
Chemical properties of the forest ¯oor and min-eral soil materials are given in Tables 1 and 2,
re-Table 1. Some chemical properties of forest ¯oor material from stands of dierent age, soil type and species composition. Averages for
n= 4 samples, standard deviation is given in parentheses. Values for pH are medians and the range is given in brackets
Age (year) Soil Species Corg (g kgÿ1) Norg (g kgÿ1) pH (in H
2O) Ca (mg gÿ1) K (mg gÿ1) Mg (mg gÿ1) P (mg gÿ1)
50 clay aspen 439 (67) 15.4 (2.2) 5.50 (0.41) 10.72 (1.17) 1.51 (0.06) 0.70 (0.06) 21.3 (1.8) birch 432 (43) 17.7 (1.7) 5.21 (0.26) 7.92 (2.79) 1.32 (0.38) 0.69 (0.14) 21.0 (5.6) conifer 473 (3) 13.3 (2.2) 5.37 (1.00) 8.12 (2.81) 1.38 (0.30) 0.63 (0.07) 21.4 (3.7) till aspen 442 (36) 14.0 (1.6) 4.98 (0.92) 5.03 (1.77) 1.18 (0.28) 0.47 (0.08) 13.0 (4.0) birch 447 (52) 16.2 (1.7) 4.83 (0.87) 5.12 (1.46) 1.31 (0.07) 0.57 (0.15) 16.3 (2.3) conifer 426 (75) 12.6 (3.5) 4.75 (0.35) 4.16 (0.55) 1.03 (0.25) 0.45 (0.10) 14.6 (3.8)
spectively. In the following the results are presented separately for forest ¯oor and mineral soil.
Forest ¯oor
Organic C in the forest ¯oor ranged from 325 to 539 g kgÿ1 and was not aected by stand age, soil
type, or tree species. Forest ¯oor Norg contents,
however, were, on average, signi®cantly lower under conifers (13.2 g kgÿ1) than below a birch (15.9 g kgÿ1) or aspen (15.4 g kgÿ1) canopy. The
average pH was also lower in the forest ¯oor beneath conifers than beneath either deciduous species. However, pH was more strongly in¯uenced by soil type. The forest ¯oor overlaying clay soils (pH 5.4) was less acidic than forest ¯oor over till soils (pH 4.9). Concentrations of base cations and P were higher in the forest ¯oors on clay than on till soils (Table 1).
Microbial carbon
Values for Cmic were between 5570 and
13540mg gÿ1 (Table 3). Cmic contributed 1.33±
3.44% to total organic C in the forest ¯oors. Both Cmic and the Cmic-to-Corg ratio were signi®cantly
aected by stand age, tree species and soil type in the forest ¯oors (Table 5). Cmic, in absolute and
relative terms, was on average higher in young
stands than in old stands. The Cmic-to-Corg ratio
decreased from aspen (2.41%) to birch (2.18%) and to conifers (1.94%) and was higher in forest ¯oors over clay than over till soils.
The best multiple linear regression model for the forest ¯oor data explained only 24% of Cmic
vari-ation, using K and P concentrations as independent variables. Microbial C concentrations were posi-tively related to K beneath aspen, to P beneath birch and to Mg and P beneath conifers (Fig. 1).
Microbial nitrogen
Microbial N ranged between 542 and 1848mg gÿ1
in the forest ¯oors. The contribution of Nmicto Nt
was 4.3±12.4%. Microbial N concentrations and the Nmic-to-Ntratio were signi®cantly in¯uenced by
all experimental factors (Table 5). Microbial N decreased, on average, from young (1385mg gÿ1) to
old (1083mg gÿ1) stands, it was higher in forest
¯oors overlaying clay soils (1366mg gÿ1) than in
for-est ¯oors over till soils (1103mg gÿ1), and lower
beneath conifers (1032mg gÿ1) than beneath
decid-uous species (average 1335mg gÿ1). The interaction
between age and soil for the Nmic-to-Nt ratio was
signi®cant (Table 5), with a decrease in relative Nmic concentrations with stand age occurring only
in forest ¯oor overlaying till soils. In 124 year old Table 2. Some chemical properties and clay contents of the mineral soil from stands of dierent age, soil type and species composition. Averages forn= 4 samples, standard deviation is given in parentheses. Values for pH are medians and the range is given in brackets
Age (year) Soil Species Corg(g kgÿ1) Norg(g kgÿ1) pH (in H2O) Ca (mg gÿ1) K (mg gÿ1) Mg (mg gÿ1) P (mg gÿ1) Clay (%)
50 clay aspen 49.6 (12.9) 1.8 (0.3) 5.40 (0.50) 2623 (1156) 344 (163) 410 (194) 6.3 (1.4) 84.0 (6.6) birch 37.4 (7.8) 1.5 (0.3) 5.01 (0.20) 818 (160) 106 (4) 168 (45) 4.1 (1.7) 90.3 (8.5) conifer 40.3 (6.3) 1.5 (0.4) 5.41 (0.36) 2306 (44) 289 (66) 295 (2) 10.4 (1.4) 100.0 (0.0) till aspen 53.4 (31.0) 1.9 (0.9) 4.74 (0.58) 311 (407) 69 (48) 55 (27) 2.8 (1.5) 54.3 (17.4)
birch 43.1 (5.7) 1.8 (0.1) 4.77 (0.52) 236 (283) 48 (20) 59 (45) 4.4 (1.1) 45.3 (7.2) conifer 37.2 (9.6) 1.4 (1.9) 4.82 (0.23) 130 (148) 43 (11) 33 (20) 5.0 (3.6) 46.9 (5.2) 124 clay aspen 50.5 (9.3) 1.9 (0.2) 5.01 (0.67) 1354 (468) 171 (106) 223 (81) 9.0 (3.2) 86.3 (16.1)
birch 52.0 (5.8) 1.9 (0.3) 4.99 (0.31) 1409 (252) 240 (65) 254 (49) 11.5 (4.1) 87.8 (7.5) conifer 54.0 (21.5) 1.7 (0.4) 4.83 (0.67) 1272 (1365) 169 (103) 176 (136) 6.6 (2.0) 87.8 (15.2) till aspen 19.8 (4.7) 1.1 (0.2) 4.62 (0.53) 208 (22) 39 (5) 44 (2) 5.1 (2.5) 39.4 (8.9)
birch 29.1 (12.4) 1.3 (0.3) 4.93 (0.18) 375 (250) 58 (21) 59 (34) 10.2 (4.5) 46.8 (3.5) conifer 28.3 (5.1) 1.2 (0.1) 4.44 (0.29) 150 (160) 54 (18) 36 (10) 9.8 (4.8) 38.3 (8.1)
Table 3. Microbial biomass C and N, Cmic-to-Nmicratio, Cmic-to-Corgand Nmic-to-Ntratio and the speci®c microbial respiration in forest
¯oor material from stands of dierent age, soil type and species composition. Averages forn= 4 samples, standard deviation is given in parentheses
Age (year) Soil Species Cmic(mg gÿ1) Nmic(mg gÿ1) Cmic-to-Nmic Cmic-to-Corg(%) Nmic-to-Nt(%) qCO2*
50 clay aspen 12344 (975) 1589 (91) 7.8 (0.9) 2.9 (0.53) 10.5 (1.6) 37.9 (4.2) birch 10588 (2251) 1559 (143) 6.8 (1.3) 2.5 (0.53) 8.9 (1.3) 40.4 (3.5) conifer 11347 (2351) 1349 (356) 8.4 (1.3) 2.4 (0.49) 8.7 (0.8) 54.6 (7.8) till aspen 10756 (1753) 1330 (238) 8.0 (0.7) 2.4 (0.24) 9.5 (0.8) 51.3 (12.4)
birch 9285 (606) 1427 (173) 6.5 (0.7) 2.1 (0.15) 8.8 (0.8) 61.1 (11.3) conifer 8873 (2479) 1056 (370) 8.4 (1.8) 2.1 (0.25) 8.3 (0.6) 56.7 (8.3)
124 clay aspen 9939 (574) 1225 (268) 8.1 (1.2) 2.1 (0.22) 9.0 (2.3) 59.9 (8.4) birch 10544 (1220) 1411 (188) 7.5 (1.7) 2.3 (0.36) 9.7 (0.9) 71.3 (11.8) conifer 8366 (996) 1061 (73) 7.9 (1.3) 1.8 (0.26) 8.1 (1.3) 88.5 (18.3) till aspen 10646 (567) 1136 (159) 9.4 (1.1) 2.4 (0.23) 7.8 (1.3) 71.6 (13.5) birch 7830 (962) 1005 (272) 7.8 (2.2) 1.8 (0.09) 6.8 (1.0) 91.9 (13.3) conifer 7254 (1466) 663 (164) 10.9 (1.4) 1.5 (0.20) 4.8 (0.8) 74.3 (12.3)
stands the Nmic-to-Nt ratio was, on average, 6.4
and 8.9% in forest ¯oors over till and clay soils, re-spectively.
The pH, organic N and C explained 31% of the variability of Nmic. Only in the coniferous forest
¯oors, a signi®cant relationship between Nmic and
soil chemical variables was found (Fig. 1), with Nmic positively related to pH, Mg and P
concen-trations.
In the forest ¯oors over the till soils we found a positive correlation between litterfall N concen-trations (Brown, unpublished data) and the Nmic
-to-Ntand the Cmic-to-Corgratios, whereas no
corre-lation between these variables was found in the for-est ¯oors over the clay soils (Fig. 2). The goodness of ®t for the correlation between litterfall N concen-trations and the Cmit-to-Corg ratio in forest ¯oor
over till soils wasR2=0.34.
The microbial C-to-N ratio
Cmic-to-Nmic ratios were highly variable, as
indi-cated by small goodness of ®t values of r2=0.41
and r2=0.68 for linear regressions between Cmic
and Nmic in the forest ¯oors and the mineral soils,
respectively. The Cmic-to-Nmic ratio increased from
young (7.6) to old stands (8.6) in the forest ¯oor. It was higher for conifers (8.9) and aspen (8.3) than for birch (7.1) and it was higher on the till (8.5) than on the clay soils (7.7) (Table 5). However, the ANOVA model explained only 44% of the Cmic
-to-Nmicvariation in the forest ¯oor.
Multiple regression analysis showed that only a small proportion of the variation of the Cmic
-to-Nmic ratio could be explained by the variables
measured. In the forest ¯oor pH, Ntand Corg
con-centrations explained 31% of the variation. The Cmic-to-Nmic ratio in coniferous forest ¯oors was
negatively correlated with pH and exchangeable Ca (Fig. 1).
Speci®c microbial respiration
Speci®c respiration (qCO2) was generally higher
in the forest ¯oors than in the mineral soils (Tables 3 and 4). In the forest ¯oors, it was strongly in¯uenced by the stand age and to a lesser extent by tree species and soil type (Table 5). The qCO2
Fig. 1. Correlation coecients for the relationships between forest ¯oor chemical properties and some mi-crobial parameters for selected tree species. Only
signi®-cant correlations were included (P< 0.05).
Fig. 2. The relationships between litterfall N concen-trations (mg gÿ1) and the N
mic-to-Nt ratio in the forest
¯oor. Signi®cant regressions were found only for till soils.
Table 4. Microbial biomass C and N, Cmic-to-Nmicratio, Cmic-to-Corgand Nmic-to-Ntratio and the speci®c microbial respiration in the
mineral soil from stands of dierent age, soil type and species composition. Averages forn= 4 samples, standard deviation is given in parentheses
Age (year) Soil Species Cmic(mg gÿ1) Nmic(mg gÿ1) Cmic-to-Nmic Cmic-to-Corg(%) Nmic-to-Nt(%) qCO2*
50 clay aspen 588 (207) 85 (18.7) 6.9 (1.0) 1.2 (0.33) 4.8 (0.3) 15.5 (5.8) birch 432 (121) 83 (42.4) 5.2 (1.9) 1.1 (0.12) 5.2 (1.5) 29.7 (10.9) conifer 319 (82) 53 (19.8) 6.0 (1.5) 0.8 (0.14) 3.5 (0.4) 30.5 (7.9) till aspen 444 (243) 78 (42.2) 5.7 (0.9) 0.9 (0.27) 4.1 (1.1) 54.3 (13.0)
birch 587 (58) 84 (18.0) 7.0 (1.0) 1.4 (0.12) 4.8 (1.2) 29.8 (7.7) conifer 279 (71) 38 (7.9) 7.4 (0.7) 0.8 (0.35) 3.1 (1.0) 41.8 (8.7)
124 clay aspen 910 (256) 107 (19.8) 8.5 (1.9) 1.8 (0.28) 5.5 (0.4) 17.0 (7.0) birch 698 (166) 100 (16.1) 7.0 (2.8) 1.3 (0.19) 5.3 (1.6) 17.3 (5.5) conifer 917 (316) 102 (43.3) 9.0 (1.3) 1.8 (0.39) 6.0 (1.3) 23.3 (8.8) till aspen 371 (35) 61 (16.6) 6.1 (1.3) 1.9 (0.33) 5.6 (0.8) 33.3 (8.6) birch 463 (185) 71 (27.3) 6.5 (0.6) 1.6 (0.25) 5.4 (1.1) 41.3 (0.6) conifer 343 (58) 27 (2.7) 12.9 (1.1) 1.2 (0.04) 2.3 (0.3) 36.9 (11.8)
increased for all combinations of soil and species from young (=50.3) to old stands (qCO2=76.3)
(Table 3). Beneath aspen and birch, qCO2 was
higher in the forest ¯oors overlaying till than over the clay soils. It was lower in the aspen forest ¯oor (55.2) than in that of birch (66.2) and conifers (68.5) (Table 3).
The speci®c microbial respiration decreased with increasing concentrations of Cmic [Fig. 3(a)]. A
negative relationship between exchangeable Ca con-centrations andqCO2was found in the forest ¯oors
under birch and aspen (Fig. 1).
Mineral soil
Organic C in the mineral soils ranged between 15 and 94 g kgÿ1, and organic N between 0.9 and
2.9 g kgÿ1. Concentrations of both Corg and Norg
were, on average, signi®cantly (P< 0.01) higher in the clay than in the till soils. However, the dier-ence was more pronounced in old stands (Table 2). The pH and base cation and P concentrations were also higher on average in the clay than in the till soils. Particle size analysis showed that, in Luvisols developed from lacustrine clay deposits, the clay and silt fractions contributed, on average, to 89 and 11% of the mineral soil, respectively, whereas the clay, silt and sand fractions in the Podzols devel-oped on till, comprised 46, 28 and 26% of the min-eral soil, respectively (Table 2).
Microbial carbon
In the mineral soil Cmic ranged from 224 to
1333mg gÿ1 (Table 4). Cmic contributed to 0.53±
2.41% of Corg in the mineral soils. Cmic and the
Cmic-to-Corg ratio in the mineral soils were
in¯u-enced by stand age. However, the trend was inverse compared to the forest ¯oors (Table 4); higher ab-solute and relative Cmic concentrations occurred in
old stands compared to young stands. The Cmic
-to-Corg ratio was 1.05% in 50 year old stands as
com-pared to 1.63% in 124 year old stands. Cmic
signi®-cantly higher in the clay soils than in the till soils. However, soil clay content explained only a small proportion of the variation in Cmic (R2=0.26).
Since the Cmic-to-Corg ratio takes account of the
dierent C content of soils, this dierence between clay and till disappeared when Cmic was expressed
in relative terms (Table 6). The Cmic-to-Corg ratio
was higher beneath deciduous than coniferous trees. The interaction between age and soil for Cmic
showed that Cmic increased with stand age in the
clay but not in the till soils. This was not apparent for the Cmic-to-Corg ratio, because of the increase in
Corg from young to old stands in the clay soils.
Interactions in the Cmic-to-Corg ratio occurred
Table 5. The eect of stand age, dominant tree species and soil type on Cmic, the ratio of microbial C to total soil organic C (Cmic
-to-Corg), Nmic, the ratio of microbial N to total soil N (Nmic-to-Nt), the Cmic-to-Nmicratio and the speci®c microbial respirationqCO2(mg
CO2-C mg Cmicÿ1dÿ1) in the forest ¯oor
F-ratios
Cmic Cmic-to-Corg Nmic Nmic-to-Nt Cmic-to-Nmic qCO2
Age 9.3* 17.0*** 19.5*** 15.7*** 8.1** 56.2***
Species 5.9** 10.2*** 8.9** 7.2** 4.7* 5.6**
Soil 9.0** 7.9* 15.0*** 16.7*** 5.8* 6.9*
Agesoil 0.6 1.6 0.3 7.6* 2.1 0.8
Agespecies 1.0 1.5 0.1 1.2 0.1 0.6
Speciessoil 1.1 1.3 0.5 0.3 1.9 5.3*
Agesoilspecies 1.4 1.9 0.9 1.5 0.2 0.6
Multipler2 0.50 0.60 0.59 0.62 0.44 0.72
*P< 0.05. **P< 0.01. ***P< 0.001.
Fig. 3. The relationship between the concentration of microbial C (mg Cmickgÿ1) and the metabolic quotient
(mg CO2-C mg Cmicÿ1dÿ 1
between age and species and also between species and soil type. Whereas the Cmic-to-Corg ratio was,
on average, higher in clay than in till beneath aspen and conifers, the inverse was found for birch (Table 4). In young stands the Cmic-to-Corg ratio
was highest under birch, whereas it was smallest under birch in old stands. Increasing Cmic-to-Corg
ratios with stand age were found only for aspen and conifers. The variability of the Cmic-to-Corg
ratio was signi®cantly lower in soil under birch than in soil under aspen and conifer.
Multiple linear regressions showed that in the mineral soils 53% of the variation in Cmic could be
explained by the organic C concentration and the clay content (P< 0.05). However, the in¯uence of soil chemical and physical variables on microbial properties diered for dierent tree species groups. Whereas no signi®cant relationship could be found between soil factors and Cmicfor conifers, Cmic was
positively in¯uenced by K in soils under birch and it was related to soil texture in soils under aspen (Fig. 4).
Microbial nitrogen
Values for Nmicin the mineral soils were between
24 to 168mg gÿ1and the contribution of Nmic to Nt
was 2.1±7.5%. Stand age showed no in¯uence on Nmic concentrations (Table 6), but Nmic and the
Nmic-to-Nt ratio were signi®cantly in¯uenced by all
other experimental factors. As found also for Cmic,
the eect of age on the Nmic-to-Ntratio in the
min-eral soils was inverse to the age eect in forest ¯oors, with the Nmic-to-Nt ratio increasing with
stand age in the mineral soils. On the other hand, the eects of species and soil type were the same as in the forest ¯oors. That is, on average, lower ab-solute and relative microbial N concentrations were found beneath conifers (55mg gÿ1) than beneath
deciduous species (83mg gÿ1) and concentrations
were higher in the clay soils (88mg gÿ1) than in the
till soils (60mg gÿ1). In contrast to the Cmic-to-Corg
ratio the Nmic-to-Nt ratio was higher in clay soils
Table 6. The eect of stand age, dominant tree species and soil type on the ratio of microbial C to total soil organic C (Cmic-to-Corg), the
ratio of microbial N to total soil N (Nmic-to-Nt), the Cmic-to-Nmicratio, and the speci®c microbial respirationqCO2(mg CO2-C mg
Cmicÿ1dÿ1) in the mineral soil
F-ratios
Cmic Cmic-to-Corg$ Nmic Nmic-to-Nt Cmic-to-Nmic qCO2
Age 9.3* 51.5*** 0.9 6.7* 17.0*** 4.4*
Species 1.4 5.1* 5.3* 8.5** 8.8** 0.6
Soil 15.9*** 0.2 12.2** 7.5* 0.5 50.8***
Agesoil 14.6** 0.0 6.9* 1.2 0.7 0.0
Agespecies 0.2 5.2* 0.5 0.5 5.7** 1.7
Speciessoil 2.8 3.8* 1.2 3.8* 5.7** 4.2*
Agesoilspecies 0.8 3.5 0.3 4.2* 1.9 8.3**
Multipler2 0.65 0.74 0.52 0.60 0.63 0.74
*P< 0.05. **P< 0.01. ***P< 0.001. $
For comparison with other variables ANOVA results for the Cmic-to-Corgratio are from untransformed data, although variances were
not homogenous for the three tree species groups. However, ANOVA on rank-transformed data gave similar results for signi®cance and non-signi®cance of factors and interactions.
Fig. 4. Correlation coecients for the relationships between mineral soil chemical and physical properties and some microbial parameters for selected tree species. Only
signi®cant correlations were included (P< 0.05).
Fig. 5. The eect of tree species and soil type on speci®c microbial respiration (mg CO2-C mg Cmicÿ1dÿ
1
(0.61%) than in till soils (0.50%). In mineral soils, interactions between species and soil and between all factors were also found for the Nmic-to-Nt ratio
(Table 6). The average Nmic-to-Nt ratios were 5.1,
4.9 and 4.0% for birch, aspen and conifers respect-ively. In the clay soils, there was no dierence between species, but in the till soils the Nmic-to-Nt
ratio was, on average, signi®cantly smaller under conifers (2.7%) than under a deciduous canopy (5.5%). The signi®cant interaction between age and soil type for Nmic (Table 6) was caused by the
increase from an average of 74 to 103mg Nmicgÿ1
soil from young to old stands in the clay soils and a decrease from 67 to 55mg Nmicgÿ1 soil from young
to old stands in the till soils.
Organic N concentrations and clay contents explained 61% of the variability of Nmicin the
min-eral soils. Beneath conifers Nmic was positively
re-lated to exchangeable base cations (Ca, K, and Mg) and also to clay content (Fig. 4). In aspen soils, Nmic also showed a positive relationship to clay
content.
The microbial C-to-N ratio
The Cmic-to-Nmic ratio in mineral soil was
aected by stand age and tree species. It increased from an average of 6.4 in young stands to an aver-age of 8.3 in old stands, and was higher in soil beneath conifers (8.8) than under deciduous species (aspen = 6.8, birch = 6.4). Interactions between species and soil, and between species and age revealed that the Cmic-to-Nmic ratio was higher in
the till than in the clay soils, and was signi®cantly higher in old stands (10.9%) than in young stands (6.7%) in the conifer plots only. The Cmic-to-Nmic
ratio beneath aspen was smaller in the till than in the clay soils (Table 4).
The pH, P and exchangeable K concentrations explained 27% of the variation in the Cmic-to-Nmic
ratio. It was negatively correlated with pH beneath
conifers (Fig. 4). The Cmic-to-Nmic ratio increased
with clay content beneath aspen.
The speci®c respirationIn mineral soils, there was a strong eect of soil type, a small eect of stand age and signi®cant interactions between soil and species, and between all factors. The qCO2 was, on
average, almost twice as high in the till (39.6) as in the clay soils (22.2), and higher in young stands than in old. In the till soils, the qCO2 was highest
under aspen but in the clay soils it was lowest in soil under aspen (Fig. 5).
The speci®c microbial respiration decreased with increasing concentrations of Cmic [Fig. 3(b)]. It was
negatively correlated with Nt, P and exchangeable
Mg concentrations; these factors contributed to 51% of its variation. Soil chemical and physical variables in¯uenced qCO2 beneath dierent species
to dierent degrees (Fig. 6). Although increasing concentrations of exchangeable base cations and clay reducedqCO2in mineral soils under all species,
qCO2 was negatively related (P< 0.05) to soil pH
and P concentrations beneath aspen only.
DISCUSSION
The concentrations of microbial C and N, Cmic
-to-Nmic ratios, Cmic-to-Corg and Nmic-to-Nt ratios
and qCO2 were all within the range of values
reported for boreal forests (Visser and Parkinson, 1989; Martikainen and PalojaÈrvi, 1990; Fritzeet al., 1994; Smolanderet al., 1994; Scheu and Parkinson, 1995).
Anderson and Domsch (1993) showed that soil acidity had a signi®cant in¯uence on the availability of soil C to the micro¯ora. An analysis of many data sets showed that pH had a strong in¯uence on the Cmic-to-Corg ratio (Wardle, 1992). The pH and
base cation concentrations could not explain the variation in the Cmic-to-Corg, Nmic-to-Nt and the
Cmic-to-Nmic ratios for the entire data set in our
study. This result agrees with the ®ndings of Joergensen et al. (1995) and Beck et al. (1995) for large data sets. However, when our data were divided into tree species groups, pH showed a sig-ni®cant in¯uence on the Nmicconcentration and on
the Cmic-to-Nmic ratio in coniferous forest ¯oor and
mineral soil samples. Thus the above relationship may be found only for samples of comparable sub-strate and for data that show a wide range of soil pH extending into strongly acidic values, such as beneath conifers in our study. Joergensen et al. (1995) also suggested the existence of a threshold for the relationship between pH and the Cmic
-to-Nmic ratios, and found little variation above pH
5.0.
The negative relationship betweenqCO2and Cmic
concentrations, which has also been reported by Wolters and Joergensen (1991) and Wardle and Ghani (1995), indicates that microorganisms were
Fig. 6. Correlation coecients for the relationships between mineral soil chemical and physical properties and the speci®c microbial respiration for tree species. Only
more C ecient on substrate supporting high con-centrations of micro¯ora. Wolters and Joergensen (1991) described a negative relationship between soil Ca concentrations and the qCO2, which was
interpreted as metabolic stress of the micro¯ora induced by acidity. In the aspen mineral soils and forest ¯oors, qCO2 was negatively correlated with
both nutrients (K and Ca) and pH, thus supporting the interpretation of Wolters and Joergensen (1991). However, in birch forest ¯oors and mineral soils, qCO2 was negatively correlated only with
concen-trations of Ca and K. Thus high speci®c microbial respiration may be indicative of nutrient stress rather than acid stress. Under most ®eld conditions this will be dicult to distinguish because acid soils have usually a poorer nutrient status than soils with higher pH.
The eect of stand age
We had hypothesized that the Cmic-to-Corg and
Nmic-to-Nt ratios would decrease with stand age,
assuming that substrate quality was higher in young stands than in old stands. Our ®ndings are of inter-est because we observed contrasting trends for mi-crobial variables in the forest ¯oors and the mineral soils. Although no dierence was found between Corg and Norg concentrations in the forest ¯oors of
young and old stands, Cmic-to-Corg and Nmic-to-Nt
ratios decreased from young to old stands, indicat-ing a decline in substrate quality. A decrease in the Cmic-to-Corg ratio in organic horizons with
increas-ing successional stage of vegetation communities has also been observed by Wardle (1993). Neither the decrease of Cmic-to-Corg and Nmic-to-Nt ratios
in the forest ¯oors, nor their increase in the mineral soils, from young to old stands was accompanied by a change in Nt, or exchangeable base cation
con-centrations. However, concentrations of BrayII-extractable P in the mineral soils increased signi®-cantly from an average of 5.5mg gÿ1 in young
stands to 8.7mg gÿ1in old stands. In another study
in the same forests PareÂet al.(1993) demonstrated that the availability of P in the F/H layer over clay soils decreased with stand age, whereas N avail-ability, determined by aerobic laboratory incu-bation, did not change with time. Microbial biomass in the mineral soils of our study may have been P limited in younger stands.
Towards the end of the aggradation period in the mixed boreal forest more litter occurs in the form of woody material (Pare and Bergeron, 1995). If the decrease in microbial concentrations with stand age was caused by a decline in organic matter quality, decomposition and mineralization processes in the forest ¯oor might be less complete in old stands. This could not only lead to the often-observed ac-cumulation of forest ¯oor mass but it could also leave more material that could potentially be lea-ched from the forest ¯oor into the mineral soil.
Qualls et al. (1991) showed that the ¯ux of dis-solved organic matter from the forest ¯oor to the mineral soil can be substantial. At ®elds sites of this study Pare (unpublished data) found that Nt
con-centrations in seepage water below the forest ¯oor overlaying clay stands were twice as high in 124 year old stands as in 50 year old stands. Thus a decrease in organic matter quality in the forest ¯oor may increase the input of soluble organic matter to the mineral soil.
The increases of Cmic-to-Corg and Nmic-to-Nt
ratios with stand age in the mineral soils were ac-companied by a highly signi®cant widening of the Cmic-to-Nmic ratio, indicating a growing importance
of fungi, which have a higher C-to-N ratio than bacteria (Marumoto et al., 1982a), in microbial communities. This trend in Cmic-to-Nmic ratios,
which also occurred in the forest ¯oors, was most prominent in coniferous stands. In the mineral soils, it was also accompanied by a decrease in activity as expressed by the speci®c respiration. Thus the increase in Cmic-to-Corg and Nmic-to-Nt ratios may
be attributable to an increase in microbial eciency or to a larger portion of inactive biomass in the mineral soils of old stands. Nitri®cation in these forests declines sharply from 50 to 124 year old stands (Braiset al., 1995; Pare and Bergeron, 1996) pointing also to a change in substrate and microbial communities.
In the forest ¯oor, qCO2 increased with stand
age. This decrease in microbial C eciency may again be related to a decline in organic matter qual-ity from young to old stands. The decrease in the Cmic-to-Corgratio with stand age in the forest ¯oors
discussed above may be a direct result of the decrease in microbial eciency.
mi-crobial biomass and respiration. If there are equili-bria between microbial C use and Cmic during
suc-cession of plant communities, the forest ¯oors and mineral soils in our study appear to be moving towards distinctly dierent equilibria; for a better understanding of the eect of stand age on mi-crobial biomass and activity, more age classes should have been included. Our hypothesis that stand age would aect substrate quality negatively and thus decrease the Cmic-to-Corg and Nmic-to-Nt
ratio while the speci®c respiration increased was supported only by results for forest ¯oor materials. Our results for the minerals soil suggest the need for a revision of the common belief (which stems from studies of the forest ¯oors) that nutrient avail-ability in boreal forests decreases with stand age.
Eect of soil type
Our common understanding is that clay soils ac-cumulate more organic matter than sandy soils because organic matter is stabilized to a higher degree and less accessible to microbial decompo-sition in clay soils (Van Veen et al., 1987; Oades, 1988). However, most of our knowledge is derived from studies on agricultural soils which are, in con-trast to forest soils, frequently perturbed. The clay soils in our study contained more Corg (53 g kgÿ
1
) and Nt (2.0 g kgÿ1) than the till soils
(Corg=36 g kgÿ1, Nt=1.5 g kgÿ1); the C-to-N ratios
did not dier. Since the turnover of microbial bio-mass is slower in ®ne-textured soils, they contain higher concentrations of Cmicand Nmicthan
coarse-textured soils (Van Veenet al., 1987). The Luvisols in the boreal forest of our study contained on aver-age 55% more Cmic and 48% more Nmic than the
Podzols. Whereas some ®ne-textured soils also have higher Cmic-to-Corg and Nmic-to-Nt ratios than
coarse textured soils (Hassink, 1994; Beck et al., 1995), there is generally no positive relationship between Cmic-to-Corgand soil clay content (Wardle,
1992). This indicates that microbial biomass is pri-marily correlated to the higher C content in clay soils. This was also true in our study. However, the Nmic-to-Nt ratio was higher in the clay than in the
till soils. In ®ne-textured soils the proportion of bacteria, which have a lower C-to-N ratio than fungi, may be higher because they are better pro-tected against desiccation than in coarse-textured soils (Grin, 1981). Smaller microbial C-to-N ratios in ®ne textured soil than in to coarse textured soil were also reported by Hassink (1994).
The absence of a strong relationship between Cmic and clay content has also been reported by
Hassink (1994) and Becket al. (1995). Conditions for microbial biomass can be unfavourable at high soil clay contents, possibly because of insucient drainage and aeration. Aggregate size itself, which clearly in¯uences soil porosity, can also have signi®-cant eects on microbial biomass. Miller and Dick
(1995) showed that Cmic and the Cmic-to-Corg ratio
declined with aggregate size for aggregates smaller than 0.5±1.0 mm.
Soil type had a highly signi®cant eect on the speci®c respiration in the mineral soils, with qCO2
in the till soils being, on average, twice as high as in the clay soil. This observation conforms with the above theory that microbial C turnover is more rapid in coarse-textured soils than in ®ne-textured soils (Van Veen et al., 1987; Verberne et al., 1990). Interactions between species and soil type (Fig. 5) indicated that the dierence in speci®c microbial respiration between clay and till was most pro-nounced in mineral soil beneath aspen. In the aspen stands, small roots were more concentrated at the surface in the till than in the clay soils (Bauhus, unpublished data). In the more nutrient limited soils in till the relative input of C from roots to stimulate N mineralization might be higher than in the clay soils and result in a larger portion of the micro¯ora being metabolically more active (Bradley and Fyles, 1995).
The higher Cmic-to-Corg and Nmic-to-Nt ratios
and a lower Cmic-to-Nmic ratio and speci®c
respir-ation in forest ¯oors overlaying the clay soils com-pared to those in the forest ¯oor over the till soils can be interpreted as the indirect in¯uence of soil nutritional status on forest ¯oor nutrient concen-trations via litter quality. Although litterfall N explained only 48% of the variation in the Nmic
-to-Nt ratio (Fig. 2), this relationship is surprisingly
strong considering that leaf litterfall comprises only a fraction of the total litter input into the forest ¯oor. The input of ®ne root litter is estimated to be high in this boreal forest, where dense root mats are found in the forest ¯oors. The above relation-ship indicates that microbial biomass in the forest ¯oors on the poorer soil substrate may be N lim-ited.
The eect of tree species
despite a high speci®c respiration in birch forest ¯oors might be explained by C input from roots, which was found to be very high under Betula papyrifera(Bradley and Fyles, 1995).
Microbial C utilization in the aspen and birch forest ¯oors was in¯uenced by Ca concentrations. This was also found in a range of soils from European beech forests investigated by Wolters and Joergensen (1991), who described a strong negative linear relationship between qCO2 and soil Ca. The
fact that a similar relationship was not found for the coniferous forest ¯oors, points out that this re-lationship may only be valid for comparable sub-strates.
Higher Cmic-to-Nmicratios in the till mineral soils
under conifer may indicate that fungi were more important than in soil under the deciduous species. However, the dierences in Cmic-to-Nmic ratios in
litter and mineral soil under conifers and under deciduous species was not as profound as we had anticipated. Scheu and Parkinson (1994) found little dierence in the fungal-to-bacterial respiration and biovolume ratios between pine and aspen forest ¯oors. The fungal-to-bacterial ratios of microbial biomass in these forests may be more strongly in¯u-enced by factors such as moisture and temperature than by litter quality.
CONCLUSIONS
The microbial variables measured in our study proved to be sensitive to tree species stand age and soil type. Microbial biomass and its activity may thus be potential indicators for changes in mineral soil and forest ¯oor organic matter quality and be useful for assessing the sustainability of forest man-agement practices such as tree species change. Further studies are needed to investigate the eect of soil aeration and drainage on microbial biomass. If microbial biomass is sensitive to these factors, it will also be sensitive to soil compaction through harvesting operations. Our study showed that it cannot be generally assumed that biological fertility and thus nutrient availability decreases with stand age in boreal forests. This may be true only for the forest ¯oor and biological fertility may actually increase in the mineral soil.
AcknowledgementsÐWe are very thankful to Claire Vasseur for laboratory assistance. The project received funding from the Natural Sciences and Engineering Research Council of Canada and the Quebec Ministry of Natural Resources. JuÈrgen Bauhus received post-doctoral scholarships from the Quebec Ministry of Education and the Foundation of the University of Quebec at Montreal. Thanks to Helen Desmond for English corrections. Two anonymous reviewers provided helpful comments on a previous version of the manuscript.
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