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Ecological studies on the community structure of arbuscular mycorrhizal fungi (AMF) are generally restricted to the main rooting zone (10–30 cm soil depth, e.g. Stutz & Morton, 1995, Douds et al., 1995, Guadarrama & Alvarez-Sanchez, 1999; Bever et al., 2001) regardless of whether they were based on spore morphotyping (e.g. Blaszkowski, 1994; Kurle & Pfleger, 1996; Franke-Snyder et al., 2001) or on molecular techniques allowing identification of AMF species directly in the roots (Helgason et al., 1998, 2002; Husband et al., 2002; Redecker, 2002; Vandenkoornhuyse et al., 2002; Jansa et al., 2003; Johnson et al., 2003). Only a few studies included the subsoil. With increasing soil depth, a decrease was found in the percentage of roots colonized by AMF (Jakobsen & Nielsen, 1983; Rillig & Field, 2003), in the number of infective propagules (An et al., 1990), in the amount of extraradical AMF hyphae (Kabir et al., 1998, already in 15–25 cm soil depth when compared to the upper 15 cm), in AMF spore (Jakobsen & Nielsen, 1983; Zajicek et al., 1986; Thompson, 1991) and in species numbers (Zajicek et al., 1986). The latter authors detected only two AMF species deeper than 40 cm and only one species deeper than 60 cm in a prairie grassland of the Great Plains. Nothing has been reported about the vertical distribution of AMF species in soils under extensive compared with intensive agricultural management. This is of interest because soil management methods and agronomic practices may affect the AMF community structure positively or negatively, to the benefit or disadvantage of crop yields and land productivity (van der Heijden et al., 1998; Mäder et al., 2002).
In the present study, we investigate the abundance and diversity of AMF spores at different soil depths, considering three agricultural land use systems widely differing in management intensity and prevailing in the Upper Rhine Valley: extensively managed, permanent grasslands, a vineyard managed at intermediate intensity, and intensively managed, continuously mono-cropped maize fields. We have previously analysed the abundance and diversity of AMF spores in topsoils of both the grasslands and the maize fields, using samples from the same field sites, and this previous study has clearly demonstrated that agricultural intensification, as practiced in temperate Central Europe, negatively affects AMF abundance and diversity in the topsoils (Oehl et al., 2003a). We now show that deeper soil layers differ considerably in the abundance and diversity of AMF species, as analysed by morphotyping of spores, and that in intensively managed agroecosystems, these deep soil layers may represent a hidden source of additional AMF diversity. This insight is particularly relevant for current attempts to restore degraded soils that – due to detrimental agricultural or other landuse practices – have been impoverished with respect to AMF diversity (An et al., 1990; Cuenca et al., 1998a,b).
The present study is based on spore counts and on identification of the AMF by spore morphology. Since these AMF spores accumulated over a certain time (weeks to months), our study provides an integrative holistic, but somewhat static picture of the AMF community. Clearly, spore populations do not directly reflect the AMF community that is actually colonizing the plant roots (Clapp et al., 1995). In fact, some AMF may not be detected at all because they are not sporulating or are only doing so occasionally. In the future, it may be possible to obtain a more dynamic and more complete picture of the AMF community actually present and active in the roots, using molecular identification tools. However, this approach is currently limited due to the considerable costs involved, the lack of adequate primers to cover the whole range of AMF, and the difficulties to assign sequences to taxonomic units. For the time being, therefore, morphological identification of AMF spores remains the most economical, the fastest, and, indeed, the only feasible way to assess AMF communities in studies on soil samples taken from the field (Douds & Millner, 1999; Oehl et al., 2003a).
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Previously, we have reported that intensification of land use (Oehl et al., 2003a) and conventional, as opposed to organic, farming practices (Oehl et al., 2004) cause a reduction in AMF spore abundance and AMF species diversity in the agroecosystems of Central Europe. According to common practice, we had analysed only the topsoil (0–10 cm depth) in these studies. In the present work, we investigated the soil over a large depth range. As expected, the topsoil layers from 0 to 20 cm depth contained by far the largest AMF spore abundances, a feature that was most expressed in the grasslands. More remarkably, however, the AMF community composition changed towards deeper soil layers and a surprisingly high species richness was observed even in the deepest soil layers examined (50–70 cm). Scutellospora species (S. calospora and S. castanea) in particular were found to occur more abundantly with increasing soil depth, in relative and sometimes even in absolute terms (Table 2). Scutellospora castanea was detected only below 20 cm soil depth. Thus, at least with respect to spore formation, the Scutellospora species appear to be specialized for deeper layers of the Loess soils in the agro-climatic region under study. The increase of Scutellospora spore abundance towards deeper soil layers was more pronounced in the intensively managed maize fields (particularly at site R) when compared with the less intensively managed vineyard and the extensive grasslands (Table 2). This observation agrees well with the recent finding in a long-term field trial – using molecular tools – that maize roots from plowed and chiseled plots were colonized by Scutellospora to a much lower extent than roots from no-till, less intensively managed plots (Jansa et al., 2003). In another long-term field trial comparing different farming systems (Mäder et al., 2002), the occurrence of S. calospora and S. pellucida spores (Oehl et al., 2004) was found to be negatively correlated with the soil contents of available phosphorus (Oehl et al., 2002). These findings suggest two possible reasons for the stimulation of development of S. calospora and S. castanea in deeper soil layers (Table 2), namely the reduced mechanical soil disturbance or the decreased supply of available phosphorus (Table 1).
In the present study, where sampling was carried out in autumn, a much lower percentage of spores could be positively identified than in a previous study where sampling took place in springtime at the same field sites, except in the vineyard (Oehl et al., 2003a). Only 15–50% of all spores isolated, or 30–70% of the spores mounted on slides could be assigned to a species or at least to a species group, compared with 30–60% of all isolated spores, or 60–85% of the spores mounted on slides in the previous study (Oehl et al., 2003a). Most likely, this is due to the problem that in autumn, a major proportion of the spores is still immature and cannot be identified. Nevertheless, a very similar AMF community composition was obtained for both the grasslands and the maize fields in these studies. Only for a few species, spore abundance significantly changed from springtime to autumn; Scutellospora calospora spores, for instance, were more abundant in spring than in autumn; in trap cultures under natural ambient light and temperature conditions, this species exhibited a burst of sporulation between October and December (Oehl et al., 2003a, 2004), which could explain our observation. The increased spore abundance of S. calospora towards the deep soil layers and the characteristic of late-seasonal sporulation of this species might be related since in the deep soil layers the soil temperature is more constant and, during autumn until early winter, generally higher than in the topsoil, which is exposed to the frequently low temperature during late fall in Central Europe.
Several species found frequently in the extensive grasslands were not found in the intensively managed maize fields, neither in the topsoils nor in the deeper soil layers (Tables 2 and 3). This finding is in agreement with the observation that the majority of these AMF (G. sp. strains BR8 and BR9, G. microcarpum, G. rubiforme and G. sinuosum) were strongly decreasing in abundance with increasing soil depth or were not detected at all in the subsoil layers of the grasslands (Table 2). It appears that these AMF, at least in Central Europe, preferentially inhabit undisturbed topsoil rich in organic matter as occurring in grasslands. Another possibility is that they might need specific plant hosts. Accordingly, most of them were not recovered in the trap cultures (Table 3) containing a substrate devoid of organic matter (Table 1). It is not clear why, in previous studies (Oehl et al., 2003a, 2004), some of these AMF species sporulated abundantly.
Glomus aggregatum, G. geosporum, G. constrictum, G. fasciculatum, G. diaphanum, G. tunicatum, G. caledonium and G. mosseae are commonly found even in intensively managed arable lands (Land & Schönbeck, 1991; Blaszkowski, 1993; Kurle & Pfleger, 1996; Franke-Snyder et al., 2001; Oehl et al., 2003a). These species are sometimes called ‘typical AMF of arable lands’ or AMF ‘generalists’ (Oehl et al., 2003a) or even AMF ‘weed’ species (JPW Young, pers. comm.). According to our study they appear to belong to different groups with respect to their vertical distribution in the soils (Groups A–D, Table 2); some of them (G. mosseae and G. caledonium) differed in this respect depending on the agroecosystem. We assume that even these AMF ‘generalists’ might fulfil different ecological functions.
AMF spore abundance and species richness in general decreased with increasing soil depth (Figs 1 and 2). The decreases were much more pronounced in the extensive grasslands and in the vineyard than in the intensively managed maize fields. In fact, in one instance (maize field R), the highest number of species (Fig. 2) and the highest diversity (Fig. 3) was found in the sample corresponding to a soil depth of 20–35 cm, just below ploughing depth. This finding suggests that several AMF species apparently eradicated from the intensively managed maize field (top)soils (Oehl et al., 2003a) may have found a refuge or at least a preferred habitat below ploughing depth and, thus, are not completely lost through the agricultural practices. These species were, for example Glomus invermaium, Pacispora dominikii, Acaulospora paulinae, Entrophospora infrequens and Scutellospora castanea (Table 2). In general, our finding corroborates observations in other studies indicating that intensification of tillage practices (Jansa et al., 2002) and high-input conventional farming, compared with low-input organic farming (Oehl et al., 2004), negatively affect AMF abundance and diversity, especially with respect to AMF species not belonging to the genus Glomus.
The possibility of the survival of sensitive AMF species in the subsoil under adverse conditions, caused by intensive farming practices deleterious to AMF diversity, has important implications. An AMF ‘gene bank’, may persist in the subsoil, facilitating agro-ecological restoration when switching from a high-input to a low-input farming system that has to rely more on internal biotic and abiotic resources.