16 A.H.C. van Bruggen, A.M. Semenov Applied Soil Ecology 15 2000 13–24
Fig. 2. Ratio of CFUs of copiotrophic to oligotrophic bacteria 1 day before, 1 day after and 1, 2, 3, 5, and 7 weeks after incorporation
of a vetchoats cover crop ‘Cover crop’ or the same amount of vetchoats cover crop foliage ‘Fallow+debris’ into soil, or after
leaving the soil unamended ‘Unamended’.
This general concept was verified in a series of exper- iments in which the effects of various stress factors
temperature, mineral salts, drying-rewetting, and combinations of these factors on cellulase activity
and fungal communities were investigated for peat bogs Nizovtseva and Semenov, 1995; Semenov and
Nizovtseva, 1995. Maximal fungal diversity occurred when cellulase activity was minimal, namely under
oligotrophic conditions without addition of mineral elements or drying-rewetting stress. Any of the ap-
plied stress factors resulted in increased cellulase activity and a succession in the micromycete commu-
nity leading to an initial decrease in fungal diversity followed by an increase.
In other studies, metabolic diversity in terms of evenness as well as richness decreased after tillage
which results in an increase in available carbon compounds from necromass Lupwayi et al., 1998.
Similarly, scarification of forest soil resulted in a temporary lower metabolic diversity than in control
plots Staddon et al., 1997. Such a decline in di- versity after temporary eutrophication will likely be
followed by an increase in diversity as the soil returns to an oligotrophic state. The same may hold for other
disturbances, for example introduction of a denitrify- ing Pseudomonas fluorescens into a small-pore soil
fraction resulted in a temporary decline in microbial diversity White et al., 1994. From all these examples
it appears that consideration of microbial succession resulting from various stress factors will likely pro-
vide the means to determine the health status of a soil.
4. Indicators for soil health
Many authors have attempted to develop soil health indicators by measuring various soil characteristics
and relating these to different management practices, productivity, environmental quality, or plant disease
severity Workneh et al., 1993; Pankhurst et al., 1995; Grunwald, 1997; Staben et al., 1997. Doran et al.
1996 presented a list of properties affecting soil ecological functions and quality, for example soil
bulk density, water infiltration and holding capacity, total organic C and N, electrical conductivity, pH,
plant-available nutrients, and measures of microbial biomass and activity. Although these properties may
be useful as indicators for soil quality, they are not necessarily associated with soil health and the main-
tenance of essential soil ecological functions. The general approach to measure as many variables as pos-
sible and relate them to different uses natural versus agricultural soil or soil management practices such
as conventional versus alternative practices in terms of tillage, plant nutrition, or pest control has not resulted
in indicators that are consistently correlated with soil health Table 1 Pankhurst et al., 1995; Staben et al.,
1997. One of the reasons for the inconsistencies may be the sensitivity of many of these measurements to
the time of sampling in relation to significant man- agement or environmental events tillage, irrigation,
residue incorporation, fertilization, rainfall, etc..
A different approach has been to search for in- dicator organisms associated with healthy or dete-
riorated soil. Keystone species or higher taxonomic groups may function as indicators for soil health since
Table 1 Variables associated with severity of corky root Pyrenochaeta
lycopersici on tomato
a
Corky root field studies in 1989 and 1990 Variables selected
in both years Variables selected in
one of the years N mineralization potential
Organic C Tissue N
Soil N Soil NO
3
Clay Soil NH
4
Microbial activity pH
Aggregate stability
a
Determined by stepwise discriminant analysis on physical, chemical and microbial variables measured on soil and plant tissues
from organic and conventional farming systems from a field study by Workneh et al. 1993.
A.H.C. van Bruggen, A.M. Semenov Applied Soil Ecology 15 2000 13–24 17
these control the interactions between other species within an ecosystem. For example, certain springtail
species were identified as keystone species in oak for- est soils due to their ability to promote detoxification
of phenolic compounds and enhance microbial activity and nutrient cycling Lee, 1994. Rhizobium species
have also been considered as keystone species, and Rhizobium populations have been proposed as indi-
cators of soil quality Visser and Parkinson, 1992. Organisms that are sensitive to anthropogenic influ-
ences and thrive only within a narrow range of environ- mental conditions could also act as indicator species.
For example, Rhizobium and nitrifying bacteria could be used as indicators in this respect because of their
high sensitivity to agrochemical Domsch et al., 1983. Members of the soil fauna such as earthworms, nema-
todes, collembola or predatory mites have also been suggested as potential indicators for soil health Lee,
1994; Pankhurst et al., 1995; Lau et al., 1997, while soil algae, bacteria-feeding or predacious nematodes,
or basidiomycete fungi could function as indicators of industrial pollution of soil Shtina, 1981; Won-
dratschek and Roeder, 1993; Korthals et al., 1996; Lau et al., 1997. However, searching for indicator species
does not constitute a systematic approach either.
Instead of a random search for indicators of soil health, we need a unified concept of soil health, and
base our search for indicators on this concept. First of all, we have to realize that biological indicators for
soil health can be divided into general and specific indicators. General indicators are universal, while the
usefulness of specific indicators depends on the geo- graphic zone, climate, soil type, and soil history. The
soil characteristics mentioned in the previous para- graph mostly fall under the category of specific indi-
cators, so that the interpretation of the values for these characteristics depends on the location and climatolog-
ical conditions. Universal indicators may include bio- diversity Parr et al., 1992, stability and self-recovery
from stress. However, microbial diversity determined by phospholipid analysis and diversity of bacteria and
fungi cultured on rich media did not differ among soils in conventional or low-input agroecosystems Buyer
and Kaufman, 1996. On the other hand, Moore et al. 1993 showed that the time needed for a soil to re-
turn to steady state after perturbation indicative of resilience declined exponentially with an increase in
primary productivity. Primary productivity in soil is determined by the return of plant-derived organic mat-
ter to the soil which may be higher in natural than in agricultural ecosystems, and higher in biologically
than chemically based farming systems. Considering a healthy soil as a stable ecosystem with tolerance or
resilience to stress, we propose that the responses of a soil sample to a variety of stresses may be better indi-
cators of soil health than soil characteristics measured without imposing stress. Similar ideas were proposed
by Kozhevin 1989 and Crowley 1997. However, these have not generally been adopted to measure soil
health.
We suggest to apply stress factors such as dry- ing and rewetting, mechanical disturbance, ex-
cess nutrients or pollutants, or inundation with a micro-organism to soil samples differing in man-
agement history, and then monitor the amplitude of changes in microbial populations and the time needed
to return to dynamic steady-state conditions as ob- served before application of stress. An important con-
sideration with this approach is the frequency of the measurements, since trends in microbial populations
could be missed or misinterpreted if data are collected too infrequently, especially if wave-like fluctuations
result from a perturbation Semenov et al., 1999. This was also shown for nematode successions Ettema
and Bongers, 1993.
Alternatively, thresholds of increasing levels of stress could be determined until the soil ecosystem
would not rebound within a reasonable time in terms of several weeks. Effects of increasing concentra-
tions of heavy metals or hydrocarbons on soil fun- gus communities were studied by Guzev and Levin
1995, but only for a few days after the treatments. They distinguished different response zones, namely
homeostasis, stress, resistance and repression zones, corresponding to particular concentration ranges of
the pollutants. The concentration ranges for these zones could also be used to compare soil health of
different soils.
The response of the microbial community to stresses could be measured with a large variety of microbio-
logical techniques. The most common traditional mi- crobial measurements such as microbial activity and
biomass by respiration or fumigation–extraction or metabolic profiles do not include important micro-
bial groups such as oligotrophs Hu and van Bruggen, 1998. Thus, these methods have severe limitations as
18 A.H.C. van Bruggen, A.M. Semenov Applied Soil Ecology 15 2000 13–24
indicators for the response to stress, since the transi- tion from eutrophic to oligotrophic conditions likely
is an important characteristic for soil health. We suggest to test the following methods to track the
bacterial succession after application of a stress fac- tor: 1 the ratio of CFUs to total microscopic counts
Fig. 1, coined the index of microbial succession stage Zvyagintsev et al., 1984; Kozhevin, 1989, 2 the ra-
tio of copiotrophic to oligotrophic CFU Fig. 2 van Bruggen and Semenov, 1999, or 3 the ratio of res-
piration to microbial biomass Visser and Parkinson, 1992 which all increase in ecosystems after pertur-
bation and decrease with soil maturity. These indica- tors, and in particular, the second indicator reflect the
nutrient stress tolerance of the species present in soil. Interestingly, measures of nutrient stress tolerance of
the plant species in various communities were posi- tively correlated with resistance to frost and drought
stress and negatively correlated with resilience after fire MacGillivray et al., 1995.
It is somewhat more difficult to track fungal suc- cession in soil. CFUs obtained from dilution plating
are more indicative of fungal sporulation than of hy- phal growth. It is therefore preferable to use a direct
soil plating technique. Since many fungi have both oligotrophic and copiotrophic capacities, it is not very
useful to distinguish trophic groups on media differ- ing in nutrient contents. However, it is easier to iden-
tify fungi than bacteria using traditional classification techniques. Characteristic ‘sugar fungi’ such as the
Mucorales and Moniliales occur early in succession, while cellulolytic and ligninolytic fungi occur typi-
cally later in succession.
The above-mentioned techniques are quite insen- sitive to subtle changes in microbial communities.
Moreover, only a small proportion of micro-organisms can be isolated from soil. Techniques assessing mi-
crobial composition by measuring phospholipid fatty acid PLFA profiles and various DNA hybridization
or DNARNA fingerprinting techniques are more sen- sitive and culture-independent Torsvik et al., 1996;
Bossio et al., 1998. For example, microbial composi- tion as assessed by PLFA profiles was more affected
by specific farming operations disturbances than by farming system organic, integrated, or conventional,
but the time to return to the initial composition re- silience was not compared for the different farming
systems Bossio et al., 1998. Besides monitoring changes in microbial succession after a perturba-
tion, PLFA and DNA- or RNA-based techniques also give the opportunity to estimate microbial diversity
Torsvik et al., 1996. These techniques can then be used to compare recovery of the microbial community
from stress for soils varying in microbial diversity Crowley, 1997.
The problem with these techniques is that the extraction efficiencies of phospholipids and DNA de-
pend on soil type and microbial community Zhou et al., 1996, and that they give little if any information
about changes in functional capabilities. Moreover, fatty acids or DNA sequences characteristic for olig-
otrophs have not been identified yet. For soil health assessment, the number and identity of phylogenetic,
physiological and trophic groups is probably more important than biodiversity as such. Hence, charac-
teristic PLFA peaks and function-specific DNA or mRNA probes or primers are necessary to track spe-
cific microbial populations after a disturbance Liu et al., 1997.
Alternatively, substrate utilization tests such as pro- vided by ‘Biolog’ could be used to trace shifts in
metabolic activities by microbial communities Bossio and Scow, 1995; Grunwald, 1997. Differences in sub-
strate utilization patterns were detected after carbon enrichment Bossio and Scow, 1995; Grunwald, 1997
and soil moisture treatment Bossio and Scow, 1995. However, segments of the microbial community are
excluded on these substrate utilization plates, in partic- ular, slower growing organisms such as gram-positive
bacteria and oligotrophs Wuensche and Babel, 1996; Verschuere et al., 1997. Thus, all biological methods
listed above have some deficiencies. It is therefore im- portant to use more than one method to monitor mi-
crobial succession in soil.
In addition to these potential indicators, inoculum levels of root pathogens and subsequent disease were
also considered as potential bioindicators of soil health Visser and Parkinson, 1992; Pankhurst et al., 1995,
but this idea was rejected by Hornby and Bateman 1997 for a variety of reasons, in particular, the de-
pendence of root pathogens on cropping history.
5. Soil health and disease suppression
