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Microbial Community Structure During
Composting With and Without Mass
Inocula
Spela Velikonja Boltaa, Rok Mihelica, Franc Lobnika & Domen Lestana
a Center for Soil and Environmental Science, Agronomy Department,
Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Published online: 23 Jul 2013.
To cite this article: Spela Velikonja Bolta, Rok Mihelic, Franc Lobnik & Domen Lestan (2003) Microbial
Community Structure During Composting With and Without Mass Inocula, Compost Science &
Utilization, 11:1, 6-15, DOI: 10.1080/1065657X.2003.10702104
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Compost Science & Utilization, (2003), Vol. 11, No. 1, 6-15
Microbial Community Structure During Composting
With and Without Mass Inocula
Spela Velikonja Bolta, Rok Mihelic, Franc Lobnik and Domen Lestan
Center for Soil and Environmental Science, Agronomy Department,
Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Inoculation of a mixture of household organics and shredded wood with inocula from
the active phase of composting enhanced mineralization of organic matter and yield-
ed a biologically stabilized product with a more favorable C/N ratio than in a non-
inoculated treatment. Analysis of phospholipid fatty acids was used to determine to-
tal viable microbial biomass and relative amounts of bacteria, Gram positive and
Gram negative bacteria, actinomycetes, fungi, thermophiles and nonthermophiles in
compost. The initial viable microbial biomass was 6-fold higher in inoculated than in
noninoculated compost. The thermophilic phase occurred immediately after inocu-
lation and the total viable biomass afterwards followed the course of temperature.
The ratio of viable (sum of phospholipid fatty acids) to dead (sum of diglyceride fat-
ty acids) microbial biomass during this period was generally higher at lower com-
posting temperatures in both inoculated and noninoculated compost. During com-
posting, fluctuations in total viable microbial biomass and in the relative amounts of
indicator PLFAs of all microbial groups except Gram negative bacteria was more in-
tensive in noninoculated compost.
Introduction
Bacteria and fungi are the main organisms responsible for the decomposition of or-
ganic wastes during composting processes (Zibilske 1998). One of the predominant is-
sues is the management of the materials by addition of microorganisms capable of
speeding up the composting process. Some studies deal with the isolation of compost
microorganisms and use of pure cultures as inoculants (Strom 1985b, Janzen et al. 1995,
Maplestone et al. 1991). The hunt for a magic inoculum appears futile, as it is unlikely
that any particular organism could govern the whole composting process (Hermann
and Shan 1997). It is also recognized, that a better understanding of microbial commu-
nity structure and dynamics during composting is needed in order to improve and an-
alyze the effect of different management alternatives.
Recently new techniques, such as carbon source utilization studies (Atkinson et
al. 1997, Carpenter-Boggs et al. 1998, Minna Laine et al. 1997) (Biolog plates), bacterial
and fungal DNA fingerprints (Dees and Ghiorse 2001), and phospholipid fatty acids
analysis (PLFA) have been introduced to describe functional and physiological di-
versity of microbial populations during composting (Klamer and Bååth 1998, Car-
penter-Boggs et al. 1998). The PLFA method is based on the fact that different groups
of microorganisms differ in their phospholipid fatty acid compositions (Frostegard
1996, Vestal and White 1989, Zelles 1999). Although few PLFAs can be considered to
be absolute signature substances for a single species, it is possible to get indications
of major groups of microorganisms such as fungi, actinomycetes, Gram positive,
Gram negative bacteria, thermophilic and nonthermophilic bacteria (Klamer and
Bååth 1998, Carpenter-Boggs et al. 1998). Total PLFAs were suggested as a measure
of the total viable microbial biomass (VMB) by Zelles et al. (1995), and were shown
to correlate well with the biomass measurements using the fumigation extraction
technique during the period after peak temperature in a municipal solid waste com-
6
Compost Science & Utilization
Winter 2003

Microbial Community Structure During Composting With and Without Mass Inocula
post (Hellmann et al. 1997). Phospholipids are components of viable biomass. After
the death of microbial cells, the cellular phospholipids are quickly hydrolyzed to
diglycerides (White and Tucker 1969). The ratio between phospholipids and diglyc-
erides therefore indicates the ratio of living to dead microbial biomass in natural me-
dia (Guckert et al. 1985).
In this study we determined the influence of inoculation with a consortium of mi-
croorganisms from the active phase of composting on both the structure of microbial
populations and the composting process. The changes in viable biomass of microbial
groups and the structure of communities were investigated by analyzing the PLFAs
during 198 days of the composting process.
Materials and Methods
Experimental Set Up
The experiment was conducted in Ljubljana (Slovenia, Europe) from December
1999 to June 2000. The starting compost (C) consisted of shredded wood (length ap-
prox.1 - 5 cm) and organic household waste in the volume ratio of 2:1. The starting C/N
ratio of the compost was 23.7:1. Inoculated compost (IC) was prepared by bulk inocu-
lation with 14 vol. % of 1 week old compost in its active phase, prepared from the same
material and the same way as a starting compost ( C). The C/N ratio of IC was 25.5:1.
The initial moisture content of IC and C was 50.7 % and 49.7 %, respectively. The com-
posts were placed in 810 L (surface area 90 x 90 cm) insulated wooden containers with
the height of 100 cm. Composting containers were equipped with a system for forced
3
-1
aeration (axial blower, Δp = 200 Pa, flow rate = 1m min ) and a set of temperature
probes. Three groups of three probes were vertically and equidistantly aligned around
the center axis of the container, from the bottom to the top, and used for continuous
temperature measurement. The daily average of temperature readings of the top three
probes closest to the sampling point were reported as the composting temperature. The
set point temperature of the compost was 55°C. The forced aeration was switched on
after temperature readings of at least 3 probes exceeded the set temperature. Below the
set point temperature aeration was controlled by a timer (2 min on 18 min off), in order
to supply enough air. Containers were topped with a metal cover with an opening for
exhaust air. On days 8 and 32 composts were mixed and 78 L of water was added to the
compost to obtain approximately 55% of moisture. Composting containers were kept
outdoors where daily mean air temperatures were low (–10° and 5°C) during the active
phase (first 32 days) of composting. Later on, the outdoor temperatures were higher.
Sampling of the Compost
Compost was sampled on days 0, 1, 2, 4, 6, 8, 12, 18, 27, 32, 89 and 198. Twelve sub-
samples of approximately 250 g of compost were taken from various points 15 cm be-
low the top of compost. Two representative samples were collected by combining six
subsamples. The dry weight and moisture of compost were determined after 24 hour
drying in an oven at 105°C (ÖNORM S 2023, 1993). For the PLFA and DGFA extrac-
tions, performed immediately after sampling , the fresh representative samples were
sieved and 3 g of size fraction below 4.4 mm was used to ensure better homogeneity
of the sample. This fraction was taken instead of the whole compost sample because
some particles in compost mixture alone weighed more than the required 3 g needed
for PLFA and DGFA extraction. By sieving, we believe that the sample for PLFA analy-
sis was more homogeneous.
Compost Science & Utilization
Winter 2003 7

Spela Velikonja Bolta, Rok Mihelic, Franc Lobnik and Domen Lestan
Analyses of Compost
The total organic matter, N content, pH value, phosphorus and potassium content
of compost were analyzed in accord with Austrian standard ÖNORM S 2023 from a
representative sample in two replicates. The organic matter content was obtained as a
loss on ignition (550°C) and multiplied by factor 0,58 to obtain organic matter C – TOC
(ÖNORM S 2023 1993).
PLFA Analysis
The modified procedures of Frostegard et al. (1991) and Guckert et al. (1985) were
used for the lipid extraction, fractionation and mild alkaline methanolysis. Briefly, 3 g
of sieved compost was weighed in a centrifuge tube. One-phase mixture of methanol,
chloroform and citric buffer, (2:1:0.8 v/v/v), pH 4.0 was added. After 2 minutes in ul-
-1
trasonic bath and 2 hours of shaking on horizontal shaker (120 strokes min ) the mix-
ture was centrifuged for 25 min. at 14000 g, the supernatant was poured into a sepa-
ration funnel and water and chloroform were added. Two phases were formed and
allowed to separate overnight. The lower lipid containing organic phase was removed
into a glass tube and evaporated to dryness at 40°C under the gentle stream of nitro-
gen. At this point, dried extracts were kept at –20°C until further analysis. Extracts
were redissolved 3-fold in 100 and 2-fold in 50 µL of chloroform and separated on the
preconditioned SPE-Si column. Lipids of different polarity were eluded with a series
of organic solvents with increasing polarity. Diglycerides were eluded with chloro-
form, glycolipids with acetone and phospholipids with methanol. Diglyceride and
phospholipid fractions were evaporated under a gentle stream of nitrogen and stored
dry at –20°C until alkaline methanolysis. For this process extracts were redissolved in
1 ml of methanol and 1 ml of 0.2 M KOH in methanol. After 30 min of heating at 60°C
the samples were cooled to room temperature and the pH was adjusted to 6.0 with 10%
acetic acid in methanol. Methyl esters were extracted with hexane. Hexane was evap-
orated under a stream of nitrogen and the final extract was prepared by addition of
200 µL of hexane with methyl nonadecanoate as an internal standard. The final extracts
of methyl esters of phospholipid and diglyceride fatty acids were analyzed with Var-
ian Saturn 3 GC/MS, using Supelco SPB-5 column (30m x 0.2mm x 0.25mm) and the
-1
temperature program: 150°C, 4 min, 4° min to 250°C, 250°C, 5 min. Helium flow was
-1
0.7 mL min , temperature of injector 200°C and ion trap temperature 150°C.
Frequent sampling time at the beginning of the composting and labware con-
straints permitted the concurrent PLFA analysis of at most two sample replicates. The
results of PLFA analysis are therefore presented as the mean values of two represen-
tative samples. The average difference in PLFAs values between data pairs obtained
from the two representative samples was 28%, and was within the range expected due
to the variability of PLFA analysis method. The variability of the PLFA analysis
method was determined at the end of composting on a series of four replicates; vari-
ability coefficient was 26.9 % for total PLFAs, and between 22 % (16:0) and 36 % (17:1)
for individual PLFAs.
Fatty Acid Nomenclature
Fatty acids were designated using the following scheme: A:BꢀC, A being a total
number of carbon atoms, B a number of double bonds and C the position of double
bonds. The prefixes i and a designate iso and anteiso branching of alkylic terminal of
8
Compost Science & Utilization
Winter 2003

Microbial Community Structure During Composting With and Without Mass Inocula
molecule, 10Me indicates branching on the tenth carbon from carbonyl end of the mol-
ecule. When the position of branching is not known, it is designated as br. Cis and trans
configurations of double bonds are indicated by cis and trans, respectively (Frostegard
and Bååth 1996).
Viable and Dead Microbial Biomass and Structure of Microbial Groups
Viable microbial biomass (VMB) was determined by summing all fatty acid con-
tents in the sample (Zelles et al. 1995, White et al. 1985) and expressed as pmol of total
-
PLFAs in g 1 (dry weight) of compost. The ratio between viable and dead biomass was
calculated by dividing total phospholipids (PLFA) by total diglycerides (DGFA)
(Ringelberg et al. 1997).
The sum of the following fatty acids was used to represent bacteria: i15:0, a15:0,
15:0, i16:0. 16:1ꢀ9c, i17:0, a17:0, 17:0 and cy17 (Frostegard and Bååth 1996). Gram pos-
itive bacteria were represented by i15:0, a15:0, i17:0 and a17:0 (Buyer et al. 1999) and
Gram negative bacteria by cy17:0 and cy19:0 (Klamer and Bååth 1998). Fungi were rep-
resented by 18:2ꢀ6 (Vestal and White 1989, Frostegard 1996) and actinomycetes by
10Me 18:0 (Klamer and Bååth 1998). Fatty acids used to represent thermophiles were
all branched (Carpenter-Boggs et al. 1998). For nonthermophiles anteiso-branched and
unsaturated fatty acids (Carpenter-Boggs et al. 1998) were used.
The structure of microbial groups in compost was presented in relative shares of
microbial groups, determined as mol % of PLFAs indicative for particular microbial
groups against total PLFAs. One must bear in mind that the development of different
groups of microorganisms during composting inferred from the changes in the PLFA
pattern does not give absolute amounts of biomass of different groups, since conver-
sion factors from the microorganism groups to actual biomass are lacking.
Results
The temperature evolution during the composting of C and IC showed that the
temperature increase in IC was much faster (Figure 1). A peak temperature of 70°C was
achieved after 2 days and the thermophilic (temperatures above 50°C) lasted until day
8 of composting. After 8 days the temperature sharply decreased due to the mixing and
watering of the compost. Afterwards the temperature rose again and the thermophilic
phase continued until day 17. The tem-
perature in C reached maximum of 69°C
on day 6 of composting. The blower
cooled the compost below 50°C within
the same day. After compost was mixed
and watered on day 8 the temperature of
C first decreased to the ambient temper-
ature and than increased but only to
o
about 35 C. After day 11 the temperature
in C steadily decreased, most likely due
to the low outside temperatures and low-
er available organic matter (Table 1). The
temperature in IC and C rose again to-
wards the end of composting, apparent-
ly indicating secondary microbial
growth (Figure 6).
Figure 1: Average daily temperature in IC (ꢀ) and C (□)
and outdoor temperature (ᮡ) during the composting ex-
periment measured closest to the sampling point. The ar-
rows indicate the time when compost was mixed and
watered.
Compost Science & Utilization
Winter 2003 9

Spela Velikonja Bolta, Rok Mihelic, Franc Lobnik and Domen Lestan
Table 1 summarizes some
TABLE 1.
Moisture content, pH, P and K content, organic matter
content, total N content, and C/N ratio during
composting of IC and C
pertinent parameters of the
composting process. Organic
matter breakdown was much
higher in IC than in C, as dur-
ing 198 days of composting
37.8% of organic matter was
mineralized in IC compared
to only 9.6% in C. N was well
preserved in both treatments,
resulting in a significantly
lower final C/N ratio. C/N
ratio (16:1) in IC indicates a bi-
ologically stabilized end-
product whereas higher C/N
Day 1 Day 32 Day 198
Day 1 Day 32 Day 198
IC
IC
IC
C
C
C
Moisture (%)
pH
45.2
6.1
3.5
59.2
7.3
4.6
2.3
-
44.3
7.5
47.2
6.2
54.3
7.2
4.1
2.8
-
43.0
7.4
P (mg/g)
K (mg/g)
8.2
3.2
5.7
2.5
10.2
51,0
1.85
16:1
2.6
9.8
Organic matter (%) 62.6
57.1
1.40
23.6:1
54.6
1.60
19.8:1
N
(%)
1.41
1.39
-
1.31
-
total
C/N ratio
25.7:1
ratio (19.8:1) in C could lead to immobilization of N if such compost was applied to the
soil (Larson and Schuman 1977). Composts from both treatments are relatively rich in
P but poor in K compared to the average farmyard manure. Due to the mineralization
of organic matter the concentrations of both elements increased during composting.
The initial moisture level of compost was rather low. Additional watering on day 8 of
composting raised the moisture content close to 60%, which is assumed to be optimal
for microbial activity (Keener et al. 1993). pH of IC and C increased during the com-
posting and was almost identical in both treatments.
In total 31, different PLFAs were detected during the composting process and 26 of
these were identified. Figure 2 presents GC-MS chromatogram of PLFAs in IC on day 6
of composting during the thermophilic phase. The marker PLFAs (sum of fatty acids that
represented total bacteria, Gram positive bacteria, Gram negative bacteria, fungi, actin-
omycetes, thermophiles and non.thermophiles) were from 71.6-93.1 % of total PLFAs.
VMB in the compost inoculum was 3-fold and 16-fold higher than initial VMB in IC
and C (Table 2). The ratio of viable to dead microbial biomass in compost inoculum was
very favorable (1,04 for C, and 4,03 for IC) indicating rapid turnover of the dead micro-
bial biomass and high micro-
bial activity. High VMB and
microbial activity of compost
inoculum are a plausible ex-
planation for 6-fold higher ini-
tial VMB in IC than in C (Table
2) and consequently for rapid
temperature increase (Figure
1) and intensive organic mat-
ter breakdown in IC (Table 1)
compared to C. The relative
share of thermophilic microor-
ganisms in compost inoculum
was low. The initial structure
of C, compost inoculum and
IC microbial communities are
Figure 2: GC-MS chromatogram of PLFA in thermophilic phase of IC on
the day 6 of composting. Peaks represent methyl esters of the following
fatty acids: 1- 12:0, 2- i14:0, 3-14:0, 4- i15:0, 5- a15:0, 6- 15:0, 7- i16:0, 8-
16:1?9c, 9- 16:0, 10- 16:1, 11- i17:0, 12- i17:0, 13- cy 17:0, 14- 17:0, 15- 18:2 ?
6, 16- 18:1 ? 9c, 17- 18:1 ? 9t, 18- 18:0, 19- 10Me 18:0, 20 – cy19:0, 21- 19:0
(internal standard), 22- 20:0, 23- 22:0.
presented in Table 2 as relative
shares of particular microbial
groups.
10 Compost Science & Utilization
Winter 2003

Microbial Community Structure During Composting With and Without Mass Inocula
TABLE 2.
-1
The initial concentrations of indicative PLFAs (pmol g (dry weight)) and relative shares of
-1
microbial groups (mol %) in C, compost inoculum (I) and IC. VMB (pmol total PLFAa g (dry
weight) compost), relative ratio of compost fungi and bacteria, and ratio of viable and dead
microbial biomass in samples of compost (PLFA/DGFA), are also shown. PLFAs in IC and C
were determined 1 hour after inoculation and at beginning of the composting.
C
I
IC
(pmol g
C
I
IC
(mol%)
-1
-1
-1
(pmol g
)
(pmol g
)
)
(mol%)
(mol%)
Bacteria
25661
15404
1101
6599
707
300796
135186
37438
67436
15049
111186
689736
1186367
0.57
70171
37908
7106
34.0
20.4
1.5
25.4
11.4
3.16
5.68
1.27
9.37
58.14
16.0
8.6
Gram positive bacteria
Gram negative bacteria
Fungi
1.6
63473
4129
8.7
14.4
0.9
Actinomycetes
Thermophiles
0.9
21940
36102
75590
0.26
59235
29.0
47.8
13.5
64.2
Nonthermophiles
Total PLFAs
282104
439466
0.90
Ratio Fungi/Bacteria
Ratio PLFA/DGFA
2.07
29.72
2.59
The total mol% exceeds 100% because some fatty acids are common inicators for different microbial groups.
After inoculation the VMB in IC in-
creased until the peak temperature on
day 2 and then decreased steadily to-
wards the end of the active phase (Figure
3). In C an increase in VMB occurred at
the beginning of composting and com-
pensated for the initial difference in VMB
in IC (Table 2). When compost tempera-
o
ture rose above 50 C, VMB in C de-
creased. The highest VMB in C was de-
termined after compost was mixed and
watered, indicating that perhaps part of
readily biodegradable biomass remained
not consumed in an early phase of the
Figure 3: Viable microbial biomass during in IC (ꢀ) and
C (□) during composting. The arrows indicate the time
when compost was mixed and watered.
composting process. Towards the end of composting the difference in VMB between
IC and C was smaller. One would expect that mixing and watering of compost fol-
lowed by a temperature decrease would be a shock to thermophilic microorganisms.
However, as shown in Figure 3 and 6 low temperatures influenced neither VMB nor
the ratio between viable and dead microbial biomass. Afterwards the ratio between vi-
able and dead microbial biomass was higher in both treatments.
Figure 4 presents relative shares of bacteria, Gram positive and Gram negative
bacteria, actinomycetes, fungi, thermophilic and nonthermophilic microorganisms
in IC and C during composting. The relative shares of particular microbial groups
changed in IC more smoothly and steadily (except for Gram negative bacteria) than
in C, where more intensive fluctuations of bacteria, actinomycetes, Gram positive
bacteria, and thermophiles were detected during the course of composting. Fluctua-
tions in C coincided with compost mixing and watering and might have been in part
enhanced by readily biodegradable biomass in C, that was apparently still available
at that time. Generally, in both IC and C the relative share of fungi decreased while
shares of actinomycetes and Gram negative bacteria increased during the course of
Compost Science & Utilization
Winter 2003 11

Spela Velikonja Bolta, Rok Mihelic, Franc Lobnik and Domen Lestan
Figure 4: Relative shares of bacteria (A), Gram positive
bacteria (B), Gram negative bacteria (C), actinomycetes
(D), fungi (E), thermophilic (F) and nonthermophilic mi-
croorganisms (G) in IC (ꢀ) and C (□) during compost-
ing. The arrows indicate the time when compost was
mixed and watered.
composting. The relative shares of bacteria, Gram positive bacteria, and ther-
mophiles increased during the active phase of composting, while the share of non-
thermophiles remained relatively constant.
The ratio between fungal and bacterial PLFAs (F/B ratio) at the beginning of com-
posting was 4-fold higher in IC compared to C (Figure 5). Afterwards the ratio steadi-
ly decreased in IC. As compost contained a lot of material rich in cellulose (shredded
wood and bark as a bulking agent), the high initial F/B ratio in IC may have contributed
to effectiveness of the composting process (Figure 1, Table 1). In C the initial increase of
the F/B ratio was observed on days 1 and 2 followed by a decrease. Towards the end
of composting no significant changes in F/B ratio were observed either in IC or C.
Figure 6 shows the ratio between living and dead microbial biomass in composts
determined as PLFA/DGFA ratio. The ratio in general decreased with the increasing
compost temperature in both IC and C, with the exception of C on day 2, when an in-
creased ratio was determined despite higher compost temperature. The share of viable
microbial biomass was therefore high in the mesophilic phase in both treatments. In
o
IC the highest ratio was observed on day 32 and was coincident with low (15 C) com-
post temperature. Mixing and watering on day 8 shifted the ratio in both treatments
towards living microbial biomass.
We must emphasize the problems that we encountered with the analyses of diglyc-
erides. At the beginning of composting, we obtained chromatograms with a lot of sat-
urated peaks which were not resolved, presumably because of high amounts of diglyc-
12 Compost Science & Utilization
Winter 2003

Microbial Community Structure During Composting With and Without Mass Inocula
Figure 6: Ratio between viable (total PLFA) and dead mi-
Figure 5: Ratio of fungal to bacterial biomass in IC (ꢀ)
crobial biomass (diglycerides) in IC (ꢀ) and C (□) dur-
and C (□) during composting. The arrows indicate the
ing composting. The arrows indicate the time when com-
time when compost was mixed and watered.
post was mixed and watered.
erides present in organic household waste. These problems were solved by excluding
saturated peaks from chromatograms. Towards the end of the experiment, the satu-
rated peaks disappeared and the chromatograms became well resolved.
Discussion
Mass inocula of consortium of microorganisms from the active phase of compost-
ing substantially enhanced the composting of mixture of organic household waste and
shredded wood. Mineralization of organic matter was much more efficient in IC than
in C, yielding biologically stabilized product with appropriate properties (Table 1). The
course of VMB (Figures 1,3) in IC (rapid increase with temperature and afterwards
steadily decrease) was consistent with results reported earlier (Klamer and Bååth 1998,
Hellmann et al. 1997). Fast increase of VMB in IC could be explained by the high
amount of active microbial biomass present in the inoculum (Table 2). Presumably, in-
oculum microorganisms, adapted to composting conditions rapidly out competed mi-
croorganisms initially present in the fresh compost with a lot of available nutrients
(Table 1). It also seems that inoculation enhanced the robustness of compost microbial
populations. In IC less fluctuations in temperature, VMB, and relative shares of indi-
vidual microbial groups were observed than in C.
The analysis of structure of microbial groups during composting revealed that the
relative share of bacteria in both IC and C were highest in the thermophilic phase (Fig-
ure 4A). At this point VMB was very high in IC (Figure 3), but ratio of living to dead
microbial biomass was rather low (Figure 6), presumably because of high tempera-
tures. The high share of bacterial biomass was associated with the high share of Gram
positive bacteria (Figure 4B), as judged by the increase in total branched fatty acids,
which include iso- and anteiso- branched fatty acids, common in the genus Bacillus, a
genus well known to be dominant in composts at high temperatures (Strom 1985a,
Strom 1985b, Beffa et al. 1996). The relative share of Gram positive bacteria in micro-
bial consortia in IC and C was higher above 50°C as observed also by Klamer and Bååth
(1998). The relative share of Gram negative bacteria decreased in the thermophilic
phase of composting (Figure 4C). Overall, the relative shares of Gram positive and
Gram negative bacteria in IC and C were lower than those reported by Klammer and
Bååth (1998) who composted straw material and pig slurry.
The increase of actinomycetes (Figure 4D) in later phases of composting is in ac-
cord with findings of Klamer and Bååth (1998). Hellmann et al. (1997), who compost-
Compost Science & Utilization
Winter 2003 13

Spela Velikonja Bolta, Rok Mihelic, Franc Lobnik and Domen Lestan
ed municipal solid waste, however, reported high relative shares of actinomycetes in
thermophilic phase, which was not the case in our study. The relative share of fungi in
IC and C (Figure 4E) did not decrease significantly at the temperatures above 50°C
which is contrary to the findings of Klamer and Bååth (1998).
The relative shares of thermophiles (Figure 4F) followed the course expected re-
garding the temperatures in IC and C. Temperatures around 70°C do not seem to have
been lethal for thermophiles, as a relatively high share (approx. 30%) of thermophiles
survived the entire thermophilic phase. The changes of relative shares of nonther-
mophilic microbial populations during the composting were rather small in IC and C
(Figure 4G). This is in agreement with the observation of Carpenter-Boggs et al. (1998).
However, these changes were not related to compost temperatures in the way that one
would expect. In IC for example, maximum share of nonthermophiles was achieved on
day 2 and was coincident with the peak temperature. High shares of nonthermophiles
were accompanying high temperatures also in C, quite contrary to expectations that
their share should decrease at temperatures above 50°C. Our observation may therefore
indicate that the biomarker PLFAs for nonthermophiles (sum of anteiso-branched and
unsaturated fatty acids) are not correctly defined in available literature (Carpenter-Bog-
gs et al. 1998). It is possible that nonthermophiles present such a diverse assembly of mi-
croorganisms that it would be difficult to find PLFAs indicative for them as a group.
As explained in Materials and methods section, the relative shares of particular mi-
crobial groups shown in Figure 4 must be interpreted with caution, although the main
dynamics of the different groups are well-supported by data obtained with traditional
isolation techniques (Beffa et al. 1997b, Fermor et al. 1979, Hayes 1968). Some conclusions
on the share of fungi and bacteria in IC and C can, however be drawn from the data on
PLFA based F/B (Fungal/Bacterial biomass) ratios found in other substrates. Frostegard
and Bååth (1996) found that the F/B ratio varied from 0.02-0.04 in different agricultural
soils low in organic matter to 0.3-0.5 in forest soils. The latter are believed to be domi-
nated by fungal biomass. In our study the F/B ratios at the end of composting were 0.15
and 0.12 for IC and C (Figure 5), respectively. Therefore it appears that fungi were not
the predominant compost group, as was reported by Klamer and Bååth (1998). Visually
we observed brown fungal growth in IC, which were not identified.
The PLFA/DGFA ratio was found to correlate well with the course of temperature
in IC and C (Figure 6). At highest compost temperatures in the thermophilic phase the
ratio of viable to dead microbial biomass was low, as expected. When the temperatures
decreased, the share of viable and active microorganisms increased, presumably be-
cause microorganisms that survived the thermophilic phase feed on the riches of dead
microbial biomass.
Acknowledgements
The research work was supported by the Slovenian Ministry of Science and Tech-
nology, grant No. L4-14347.
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