J. Elem. s. 365–375 DOI: 10.5601/jelem.2011.16.3.

02
365

SOIL FUNGAL COMMUNITIES SHAPED

UNDER THE INFLUENCE OF ORGANIC
FERTILIZATION

Bo¿ena Cwalina-Ambroziak1, Jadwiga Wierzbowska2

1Chair of Phytopathology and Entomology
2Chair of Agricultural Chemistry and Environmental Protection
University of Warmia and Mazury in Olsztyn

Abstract

The results presented in this paper have been obtained during a three-year experi-
ment (from the second cycle of four-year trials set up in 2004) established in Ba³cyny. The
following crops were grown in succession: potato, fodder barley, winter oilseed rape and
winter wheat. The fertilization treatments included: mineral NPK, FYM and organic fertili-
zation with composted and dried sewage sludge and composted municipal waste. Treatments
without soil enrichment or with mineral NPK fertilization were taken for comparison. Far-
myard manure and organic fertilizers were applied in two different ways: as a single dose
of 10 t ha–1 before planting the potato or in two doses, each 5 t d.m. ha–1, before planting
the potato and before sowing the oilseed rape. The effect of the applied fertilizers on as-
semblages of soil fungi was tested. The fungi were cultured on Martin medium.
The fertilizers were determined to have affected the populations of soil fungi. Yeast-
like fungi prevailed among the isolated colonies (60-85% of the total isolates). The richest
fungal community was obtained from the soil amended with dried and pelleted sewage
sludge or municipal green waste compost. However, suppressed growth of pathogenic fun-
gi represented by the species Aureobasidium pullulans, Botrytis cinerea, of the genus Fu-
sarium and Sclerotinia sclerotiorum was observed in the soil amended with natural fertili-
zers and composts. The growth of pathogens was most strongly inhibited in the soil
enriched with a double dose of FYM, composted urban green waste or with dried and pel-
leted sewage sludge in either of the two application variants. The pathogen-antagonistic
fungi, represented by three species of Gliocladium and four species of the genera Paecilo-
myces and Trichoderma, were more often isolated from the soil fertilized with FYM or
organic fertilizers than the unfertilized or NPK-nourished soil. The soil enriched with Dano
compost applied in a single rate of 10 t d.m. ha–1 was most abundantly populated by these
fungi. Among the remaining saprotrophs, fungi of the genus Penicillium and the orders
Mucorales and Sporotrichum olivaceum were isolated.

Key words : organic fertility, NPK fertility, soil pathogens, antagonists.

dr hab. Bo¿ena Cwalina-Ambroziak prof. UWM, Chair of Phytopathology and Entomology,

University of Warmia and Mazury, Prawocheñskiego 17, 10-720 Olsztyn, Poland, e-mail:
bambr@[Link]
366

ZBIOROWISKO GRZYBÓW GLEBOWYCH UKSZTA£TOWANE POD WP£YWEM

NAWO¯ENIA ORGANICZNEGO

Abstrakt

Pezentowane w pracy wyniki pochodz¹ z 3-letniego doœwiadczenia (z 2. cyklu 4-letnich

doœwiadczeñ za³o¿onych w 2004 r.) za³o¿onego w Ba³cynach. Uprawiano po sobie nastêpu-
j¹ce gatunki roœlin: ziemniak, jêczmieñ paszowy, rzepak ozimy i pszenicê ozim¹. Zastoso-
wano nawo¿enie mineralne NPK, naturalne obornikiem i organiczne: kompostowanymi i su-
szonymi osadami œciekowymi oraz kompostowanymi odpadami komunalnymi. Kombinacje
bez nawo¿enia i z nawo¿eniem mineralnym NPK uwzglêdniono jako porównawcze. Obor-
nik i nawozy organiczne aplikowano w dwojaki sposób: jednorazowo w dawce 10 t ha–1
przed sadzeniem ziemniaka lub dwukrotnie w dawce po 5 t s.m. ha–1 przed sadzeniem
ziemniaka i przed siewem rzepaku. W badaniach okreœlono wp³yw aplikowanych nawozów
na sk³ad zbiorowiska grzybów glebowych. Hodowlê grzybów prowadzono na pod³o¿u
Martina.
Stwierdzono wp³yw zastosowanych nawozów na populacjê grzybów glebowych. Naj-
czêœciej izolowano z gleby grzyby dro¿d¿opodobne (60-85% ogó³u izolatów). Najbogatsze zbio-
rowisko grzybów otrzymano z gleby u¿yŸnionej osadem suszonym i granulowanym oraz
kompostem z odpadów zieleni miejskiej. Jednak w glebie z zastosowanym nawo¿eniem na-
turalnym i kompostami stwierdzono ograniczenie liczebnoœci grzybów patogenicznych re-
prezentowanych przez gatunki: Aureobasidium pullulans, Botrytis cinerea, rodzaju Fusa-
rium, Sclerotinia sclerotiorum. Najbardziej zahamowany rozwój patogenów zanotowano
w glebie z wniesionym dwukrotnie obornikiem, kompostem z odpadów zieleni miejskiej oraz
osadem suszonym i granulowanym zastosowanymi w obydwu wariantach. Grzyby antago-
nistyczne wzglêdem patogenów, reprezentowane przez 3 gatunki Gliocladium, 4 gatunki
rodzaju Paecilomyces i Trichoderma, czêœciej ni¿ z gleby bez nawo¿enia i z nawo¿eniem
NPK izolowano z gleby nawo¿onej obornikiem i organicznie, z najliczniej zasiedlon¹ gleb¹
u¿yŸnion¹ Dano w dawce 10 t s.m. ha –1. Wœród pozosta³ych saprotrofów wyodrêbniono
grzyby rodzaju Penicillium, rzêdu Mucorales i Sporotrichum olivaceum.

S³owa kluczowe : nawo¿enie organiczne, nawo¿enie NPK, patogeny glebowe, antagoniœci.

INTRODUCTION

Organic substances are added to soil in order to improve its quality by

changing the physical (aggregation of soil particles) and microbiological prop-
erties (increased biomass content, microbial diversity and enzymatic activi-
ty) (DISSANAYAKE, HOY 1999, PUGET et al. 2000, CESPEDES LEON et al. 2006). Soil
improvement is achieved, inter alies, by adding more carbon to soil (SIKORA,
STOTT 1996, van BRUGGEN, SEMENOV 2000). In a study reported by KUNDU and
NANDI (1985), a broader C:N ratio stimulated the development of bacteria
while inhibiting that of fungi. This effect depended on the concentration of
chemical and biological components in organic matter. Natural fertilizers
applied in adequate doses, apart from large amounts of organic matter, en-
rich soil with some macro- and micronutrients. Certain characteristic micro-
bial communities develop in soil under the influence of these fertilizers. For
the phytosanitary safety, the share of fungi demonstrating antibiotic and
 367

parasitic activity is important. The fungi whose presence in soil is particu-

larly desirable are the ones with enzymatic properties belonging to the gen-
era Gliocldium and Trichoderma, which can inhibit the development of path-
ogens (not only the soil ones) by activating the following mechanisms:
antibiosis, competition and hyperparasitism (CHERNIN, CHET 2002). Organic fer-
tilizers (SARAIVA et al. 2004), composted plant waste (MAZZOLA, MULLINIX 2005),
municipal waste and sewage sludge (LEWIS et al. 1992, TRANKNER 1992) as
well as paper-mill residue (COOPERBAND 2001, STONE et al. 2003) provide effec-
tive plant protection against such pathogens as Botrytis cinerea, Pythium
spp., Phytophthora spp., Rhizoctonia solani, Sclerotinia sclerotiorum.
This paper contains an analysis of the structure of soil fungal assem-
blages shaped under the influence of mineral, FYM or organic fertilization.

MATERIAL AND METHODS

The experiment was carried out in 2008-2010 (the second cycle of a four-
year experiment started in 2004) on experimental plots in Ba³cyny, on grey-
brown podzolic soil originating from light silty loam, complex 4 class III (the
Polish Norms, BN-78/9180-11). Prior to the experiment, the concentration of
P (38.2 mg kg –1), K (105.8 mg kg –1) and Mg (48.3 mg kg –1) in the soil as
well as the soil pH = 5.04 were determined. The following crops were grown
in succession: potato, fodder barley, winter oilseed rape and winter wheat.
The applied fertilization consisted of natural fertilizer: mixed farmyard ma-
nure or organic fertilizers: sewage sludge composted with straw (municipal
sewage sludge from the WTP in Dzia³dowo, mixed with cereal straw in a 1 :
0.5 ratio and composted in heaps), dried and pelleted sludge (sludge from
the WTP in I³awa), sludge composted with no added substances (sludge from
the WTP in Ostróda), municipal waste compost (Dano compost from unsort-
ed municipal waste in Suwa³ki), and urban green waste compost from
Suwa³ki. The chemical composition of the organic fertilizers is presented in
Table 1. The treatments without fertilization or with the NPK mineral ferti-
lization were taken for comparison. The farmyard manure and organic ferti-
lizers were applied in two ways: in a single dose of 10 t d.m. ha–1 before
planting the potato or as two doses, each of 5 t d.m. ha–1 before planting
the potato and before sowing the oilseed rape. For the treatments with the
organic fertilizers, depending on their N-total content, nitrogen was balanced
to 150 kg ha–1 (under potato) and to 120 kg ha–1 (under oilseed rape). Spring
barley and winter wheat received only mineral fertilization.
For determination of the quantitative and qualitative composition of soil
fungal assemblages, in the early August each year, soil samples were col-
lected from the depth of 10 cm from all the plots (at three sites) which
made up a given experimental treatment. In the laboratory, the soil sam-
368

ples were shaken and portions of 10 g each were weighed out to place in
250 ml flasks, in which they were shaken for 20 minutes in 90 ml of sterile
water, thus obtaining 10–1 dilution. The suspension was further diluted (from
10–2 to 10–4). Fungi were cultured on Martin medium at 22-23oC. Fungal
colonies grown after 5-day incubation were re-calculated per 1 gram of dry
matter of soil (DHINGRA, SINCLAIR 1995). Next, the fungi were inoculated onto
agar slabs for further species identification. The results of the in vitro tests,
carried out in a completely randomized design, were processed statistically
using analysis of variance (Statistica® 9.0 2010). The means were compared
with Duncan’s test at the significance level of 0.01.
Table 1
Chemical composition of organic fert
C org. N P K Mg
Fertilizer C:N
(g kg–1 d. m.)
FYM 364.00 20.00 3.72 20.80 4.80 18.2
Sludge composted with straw 276.00 20.56 23.15 9.81 6.11 13.4
Dried and pelleted sludge 386.00 35.15 24.48 3.56 5.65 11.0
Sludge composted with no added
410.00 50.45 31.82 3.18 7.27 8.1
substances
Municipal waste compost (Dano) 151.00 12.57 6.92 7.83 5.46 12.0
Urban green waste compost 59.40 5.00 1.88 3.25 4.25 11.9

RESULTS AND DISCUSSION

The applied natural and organic fertilization differentiated counts of the

soil fungal populations; in total, 6,192 colonies of fungi were obtained, in
which yeast-like fungi made up 76.3%. Filamentous fungi were represented
by 50 species. The phytopathological tests have demonstrated that most of
the colony-forming units were obtained from the soil amended with urban
waste compost and dried and pelleted sludge (Figure 1). The literature con-
tains contradictory reports on the influence of soil organic amendments on
assemblages of soil fungi. GÓRSKA and STÊPIEÑ (2007) as well as PRATT (2008)
claim that organic fertilization does not differentiate the abundance of fila-
mentous fungal populations. AWAD and FAWN (2004) formulate contradictory
conclusions as they suggest that increasing rates of sewage sludge stimulate
the development of soil microorganisms and increase their diversity (WEY-
MAN-KACZMARKOWA et al. 2002). As researchers claim, the amount of organic
carbon added to soil plays an important role; to a lesser degree, added ni-
trogen affects microbial communities.
 369

Fig. 1 Number of colony-forming units of fungi in the soil

In the present study, the dominant organisms in the fungal assemblages

were yeast-like fungi, which made up from 63 to 85% of total communities
(in the treatment with municipal waste compost 10 t d. m. ha–1 and urban
waste compost in both application variants, respectively) – Figures 2a,c,d.
Fifty species and non-sporulating colonies were identified among the fila-
mentous fungi The pathogenic fungi, important for the phytosanitary soil
condition, were represented by the following species: Aureobasidium pullu-
lans, Botrytis cinerea, four species of the genus Fusarium and Sclerotinia
sclerotiorum, most frequently colonizing the unfertilized and NPK-fertilized
soil: 4.6 and 3.6%, respectively, of the total number of isolates (Figures 3a,
2b). The smallest population of the above fungi was cultured from the soil
fertilized twice with FYM and urban green waste compost, under spring
barley (Figures 3d, 2d). A single application of these fertilizers stimulated
the development of potentially pathogenic fungi. However, this effect was
accompanied by a growing number of antagonistic fungi, whose presence in
soil is desirable (3 species of the genus Gliocladium, 5 – Paecilomyces and 4
– Trichoderma). Based on their experiments, SERRA-WITTLING et al. (1996)
concluded that natural fertilizers and composts inhibited the development of
Fusarium spp. fungi as well as those of Pythium and Phytophthora, all wide-
370

Fig. 2. Fungi isolated from soil fertilized with composted industrial waste (%)

spread in soil. TSROR LAKHIM et al. (2001) as well as LAZAROVITZ et al. (2008)
report that the infection of potato by Rhizoctonia solani and Streptomyces
scabies is less severe in soil fertilized with cattle manure. Other researchers
(MILLS et al. 2002) point to less severe infection of solanaceous plants by
P. capsici, Alternaria solani and Septoria lycopersici in soil amended with com-
posted plant waste. CATXARRERA et al. (2002) observed weaker symptoms of Fusar-
ium wilt of tomato plants when the soil had been enriched with the fungus
Trichoderma asperellum and compost. BARAKAT and AL.-MASRI (2009) found out
a similar effect for the soil enriched with T. harzianum and sheep manure.
 371

Fig. 3. Fungi isolated from unfertilized, NPK and FYM fertilized soil (%)

Our comparison of the analysed composted sewage sludge has demon-

strated that the most favourable soil conditions, in respect of the plants’
health, prevailed in the soil amended with dried and pelleted sludge, applied
in either of the two systems: 0.6% of the pathogens and 3% of the antago-
nists (Figure 4c,d). However, the highest number of antagonists colonized
the soil enriched with the compost made from unsorted municipal waste,
372

Fig. 4. Fungi isolated from soil fertilized with composted sewage sludge (%)
 373

applied in a single dose under fodder barley (Figure 2a). For the phytosani-
tary safety of crops, it is desirable to maintain dynamic changes of fungal
populations in the soil. Organic fertilizers added to soil create favourable
conditions for the development of antagonistic fungi (HOITIN, BOEHM 1999,
PANDEY et al. 2006), especially the ones belonging to the genus (BULLOCK et
al. 2002). Such fungi produce antibiotic and parasitic effects on pathogenic
organisms, which means that they can be employed for the biological con-
trol of plant pathogens (PAPAVIZAS 1985, NELSON et al. 1983, ZAK et al. 2003).
Soil cropped with Brassicaceae has been demonstrated to contain a growing
population of the antagonists Streptomyces spp., which produce secondary
metabolites inhibiting the development of plant pathogens, e.g. Rhizoctonia
solani (MAZZOLA, MULLINIX 2005).
Among the saprotrophic fungi, the species of the order Mucorales pro-
duced the most numerous populations in the soil amended with the munici-
pal waste compost added in a dose of 10 t d.m. ha–1 (6.4%, Figure 2a). In
the treatments fertilized with composted sewage sludge and FYM
(Figures 3c,d, 4a-f), the highest counts of saprotrophs were isolated from the
soil amended with a double application of the fertilizers, except sewage
sludge composted with no added substances. Species of the genus Penicil-
lium had a similar share in the total number of fungi as saptrotrophs, and
their development was promoted by the soil environment amended with
a single dose of FYM or dried and pelleted sludge (Figures 3c, 4c). The sapro-
trophs of the genus Penicillium, analogously to those of the genus Aspergil-
lus, as well as the pathogens belonging to Fusarium spp. are the most com-
mon components of fungal assemblages dwelling in soil enriched with organic
fertilizers (SARAIVA et al. 2004).
Recapitulating, the positive influence of farmyard manure as well as
sewage sludge or composted municipal waste applied as soil amending sub-
stances on the structure of soil fungal assemblages is noteworthy. These
fertilizers have reduced the development of potential plant pathogens but
stimulated that of antagonistic fungi.

CONCLUSIONS

In brief, the experiment has demonstrated a suppressed growth of popu-

lations of pathogenic fungi in soil amended with farmyard manure or organ-
ic fertilizers. The highest count of fungal colonies was obtained from the
soil enriched with dried and pelleted sludge. At the same time, the soil from
this treatment presented the most desirable phytosanitary conditions, i.e.
a small count of pathogens but a high number of antagonistic fungi.
374

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