Bioresource Technology 289 (2019) 121653

Contents lists available at ScienceDirect

Bioresource Technology
journal homepage: www.elsevier.com/locate/biortech

Microbial inoculation influences bacterial community succession and T

physicochemical characteristics during pig manure composting with corn
straw
⁎
Changning Lia,b, Haiyun Lia,b, Tuo Yaoa,b, , Ming Sua,b, Fu Rana,b, Bing Hana,b, Jianhong Lia,b,
Xiaojun Lana,b, Yincui Zhanga,b, Xiaomei Yanga,b, Shuangbao Guna,c
a
College of Grassland Science, Gansu Agricultural University, Lanzhou 730070, Gansu, China
b
Key Laboratory of Grassland Ecosystem, Gansu Agricultural University, Lanzhou 730070, Gansu, China
c
College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, Gansu, China

G R A P H I C A L A B S T R A C T

Effects of compost water extract on germination of Medicago sativa seed at different periods between C and T. Among them 4, 12, 24 and 32 represent compost
samples on the 4th, 12th, 24th and 32th day in composting.

A R T I C LE I N FO A B S T R A C T

Keywords: This study determined the physicochemical changes and bacterial community succession in the pig manure
Microbial inoculation composting process with microbial inoculant. Microbial inoculant could prolong the thermophilic stage by
Pig manure 2 days and increased the germination index (GI). Analysis with 16S rDNA showed that the Chao1 and Shannon
Corn straw indices increased at the thermophilic stage in the treatment (T), while those of the control (C) decreased.
Microbial succession
Microbial inoculant increased the relative abundance of Flavobacterium and Solibacillus in 4–12 and 12–24 days,
Physicochemical
respectively. Acinetobacter was reduced at 4–12 days. The key physicochemical factors affecting microbial suc-
cessions were revealed by canonical correspondence analysis (CCA) and correlation analysis. Linear discriminant
analysis (LDA) effect size (LEfse) analysis showed that there were 78 biomarkers, while in piles T and C, there
were 35 and 43 biomarkers, respectively. These results indicated the addition of microbial inoculant improved
the maturity and fertility, as well as significantly regulating the microbial community structure.

1. Introduction
With the improvement of the living standards of residents, the de-
mand for animal protein has been increasing and has boosted the rapid
development of pig production in China (Wang et al., 2018). In 2015,
the number of breeding pigs was estimated to be 0.48 billion in China
(Fan et al., 2018; Yang et al., 2019). Fresh manure production is esti-
mated at 0.49 billion tons with the calculated parameter of 5.1 kg d−1
provided by the American Society of Agricultural Engineers (ASAE).
However, most pig farms are lacking complete facilities to treat manure
and urine, which not only can pollute the soil but also can enable many
parasites and bacteria to be transmitted through the air, spreading

⁎
Corresponding author at: College of Grassland Science, Gansu Agricultural University, Lanzhou 730070, Gansu, China.
E-mail address: yaotuo@gsau.edu.cn (T. Yao).

https://doi.org/10.1016/j.biortech.2019.121653
Received 31 March 2019; Received in revised form 11 June 2019; Accepted 12 June 2019
Available online 13 June 2019
0960-8524/ © 2019 Elsevier Ltd. All rights reserved.
C. Li, et al.
Table 1
Some basic characteristics of physical–chemical of the raw materials.
Moisture (%) OM (%)
Pig manure 46.20 ± 0.31 42.65 ± 0.19
Corn straw 8.75 ± 0.14 80.74 ± 0.12
diseases among people and animals and posing an unnecessary threat to
the life and health of livestock and poultry (Sanchez et al., 2017; Zhu
et al., 2013). In 2017, the government promulgated the policy of the
action plan of livestock and poultry manure utilization (2017–2020) to
accelerate the resource utilization of livestock and poultry manure (Ma
et al., 2018), which is necessary to determine a way to address the
pollution of livestock and poultry wastes.
Composting is an effective means of recycling, minimizing the harm
and reducing the quantity of agricultural solid waste. Specifically,
composting is the process of mineralizing and humifying of agricultural
solid waste through the physiological metabolism of microorganisms
(Zorpas, 2014). Microorganisms encounter difficulty in breeding in
composting waste because pig manure has a low carbon-to-nitrogen (C/
N) ratio (Guo et al., 2012). The planting area of corn is large and widely
distributed in China. Furthermore, corn straw, which has a high C/N
ratio and contains abundant nutrients, such as nitrogen, phosphorus,
potassium and organic matter, cellulose, hemicellulose and other non-
starch macromolecules, is a prime renewable biomass resource and a
good organic bulking agent (Janczak et al., 2017). Gansu is in north-
western China and has a dry climate and low temperature. Usually,
when corn straw was used as fertilizer compost, the use of indigenous
microbial content was too low to decompose sufficiently rapidly
(Huang et al., 2017). Therefore, a large quantity of corn straw was
burned, which not only caused the waste of straw and the loss of nu-
trient elements but also represented one of the main reasons for autumn
atmospheric haze. It is urgently important to solve the problem of re-
source utilization of straw (Wei et al., 2018a,b). Consequently, the
combination of pig manure and corn straw in the composting process at
a certain proportion not only can adjust the C/N ratio to one that is
suitable for microbial metabolism but also can adjust the porosity of the
pile, the final product of which can be used in edible fungus cultivation,
protected horticultural culture and as a soil regulator with irreplaceable
advantages to those of chemical fertilizers (Zhang and Sun, 2014).
Hence, the application of corn straw has many benefits in pig manure
composting, not only reducing the pollution associated with pig manure
and increasing the utilization of corn straw but also promoting co-
operation between livestock-based and crop-based farming.
However, the number of indigenous microorganisms in natural
compost fermentation is insufficient, which contributes to many de-
fects, including poor decomposition ability, prolonged fermentation
period and reduced nutrient content in compost products (Wei et al.,
2016). To accelerate the process of high-temperature composting and to
improve the quality of products, researchers have focused on the ap-
plication of additives to composting. Common additives include natural
adsorbents, such as peat and zeolite (Wang et al., 2016; Wu et al., 2017;
Mao et al., 2018), straw rich in calcium magnesium and other metal
salts (Dias et al., 2010; Yuan et al., 2016; Chen et al., 2017), and or-
ganic carbon and exogenous microbial inoculants (Chen et al., 2010; Li
et al., 2012; Wang et al., 2016).
Microbial inoculants have become a heavily researched subject due
to their ability to accelerate the degradation of agricultural wastes and
their advantages of low cost, absence of secondary pollution and ease of
operation (Huang et al., 2017). In this study, two pig manure com-
postings were tested in which compound microbial inoculants were
added to the treatment. According to the change of temperature in the
composting process, we divide composting processes into four phases,
the rising temperature stage, the thermophilic stage, the cooling phase
and the maturity stage (Zhao et al., 2016), and we evaluate compost
Bioresource Technology 289 (2019) 121653
TN (%) C/N TK (%) TP (%)
2.03 ± 0.08 12.18 ± 1.02 0.56 ± 0.12 0.21 ± 0.03
0.81 ± 0.26 57.81 ± 1.62 1.56 ± 0.04 0.27 ± 0.18
maturity and fertilizer efficiency through physicochemical parameters.
The changes in microbial community structure in four phases of pig
manure composting were determined using 16S rRNA technology. In
addition, the relationship between microbial community structure and
physicochemical parameters in the compost process was studied
through canonical correspondence analysis (CCA) (Zhou et al., 2019a).
The biomarker bacteria were explored through linear discriminant
analysis (LDA) effect size (LEfSe) during the compost process. De-
termining the changes of physicochemical parameters and bacterial
community in the compost process can not only help to elucidate the
composting mechanism of pig manure but can also provide theoretical
support for screening functional strains and accelerating the com-
posting fermentation process.
2. Materials and methods
2.1. Compost materials and preparations
Fresh pig manure was collected from 2400 breeding pigs on farms in
Zheng Da in Heishi town, Lanzhou, China. Corn stalk was purchased
from a local farmer and cut into 3–5-cm-long pieces. Microbial in-
oculation was provided by the grassland microbiology laboratory at
Gansu Agricultural University. The inoculant of composting in this
study was a kind of compound bacterium agent (a combination of
Acinetobacter pittii, Bacillus subtilis sub sp. Stercoris and Bacillus altitu-
dinis) that was screened from high temperature compost of pig manure
and then cultivated by cellulose medium domestication. Some basic
physicochemical characteristics of the raw materials are shown in
Table 1.
2.2. Compost process and compost samples
The experiment was carried out at Zheng Da in Heishi town,
Lanzhou, China, from August of 2018 to September of 2018 for a total
of 35 days. Pig manure and corn stalk were mixed at a ratio of 6:1 of
wet weight, and the moisture content was adjusted to approximately
60%. The C/N ratio was approximately 30:1, and the initial compost
mixtures density of 0.5 kg/L. Two groups of composting experiments
were carried out. The first group, without the addition of compound
microbial inoculant, was regarded as the control (C) for comparison
purposes; the second group, with the addition of the compound mi-
crobial inoculant at 1.0% (inoculant volume/wet weight of composting
sample), was regarded as the treatment (T), and the concentration of
the liquid inoculant suspended in liquid medium was 1.0 × 109 CFU
mL−1. The mixture of corn stalk and pig manure was carried out by
composting 3.00 m × 1.50 m × 1.35 m (length × width × height).
When the compost temperature rose above 50 °C, to provide aeration,
the compost piles were turned manually; the mixture was turned twice
a week in the first 2 weeks and once a week thereafter. Composting
samples from the top, middle, and bottom layers were collected from
each compost pile, and the five-point sampling method was employed
at days 0, 4, 12, 24, 28 and 32 of the experiment. Approximately 500 g
of sample was taken from each compost pile at each sampling time
(Meng et al., 2017). The fresh samples were mixed gently and divided
into three parts. The first part after natural air drying in the shade was
used for the determination of physiochemical properties and then
passed through a 60-mesh sieve stored in a cotton bag. The second part
was stored at −80 °C for the determination of 16S rDNA, while the
2
C. Li, et al.
third part was stored at −4 °C until being used for the pH, electrical
conductivity (EC) and germination index (GI) determination. Com-
posting was divided into four stages. The first stage was the rising
temperature stage (0–4 days), the second was the thermophilic stage
(4–12 days), the third was the cooling stage (12–24 days), and the
fourth was the maturity stage (24–32 days). The temperature of com-
posts and ambient air was measured twice per day (09:00 and 15:00),
and then recorded. A stainless steel compost thermometer approxi-
mately 1.50 m in length (KT, BJ8A, Shanghai, China) was employed to
measure the temperature of compost at 9: 00 and 15:00 every day (Sun
et al., 2016), the upper layer of the compost sample (0–40 cm), the
middle layer (40–80 cm), and the lower layer (80–120 cm), and the
average temperature was measured twice at each position as the
average temperature at that point. The average temperature at five
locations of the same depth was taken as the temperature of the com-
post, and the average temperature of the upper, middle and lower
layers of the compost was taken as the temperature of the composting
body. At the same time, the environmental temperature of the day was
recorded to calculate the average value as the environmental tem-
perature of the day. The pH and EC values were measured at a ratio of
1:5 (wet weight of composting sample/water volume) after shaking
equilibration for approximately 30 min using a pH meter (thunder
magnetic PHS-3C type pH meter, Shanghai, China) and a conductivity
meter (DDS-307A type conductivity meter, Shanghai, China) according
to Mao et al. (2018). Total nitrogen (TN) was determined according to
the methods described by Barrington et al. (2002). The sample was
decomposed by HClO4-H2SO4, and the total phosphorus (TP) con-
centration was estimated by the colorimetry method (UV2800A ultra-
violet spectrophotometer, UNIC Inc., China). Additionally, the total
potassium (TK) was measured by the flame emission technique with an
FP6431 flame photometer (YD LTD. Shanghai, China). The air-dried
compost samples were ignited at 550 °C in a muffle furnace for 24 h to
detect the content of organic matter (OM) and total organic carbon
(TOC) calculated from the formula TOC = OM/1.724. GI is a method to
determine the toxicity to plants and the maturity of compost samples in
different periods. The compost sample and distilled water were pre-
pared at a 1:10 (W/V) ratio to obtain water extract, and the GI assay
was determined using alfalfa seeds according to Cui et al. (2017).
seedgermiation × rootlengthoftreatment(mm)
GI(%) = × 100
seedgermiation × rootlengthofcontrol(mm)
2.3. DNA extraction and high-throughput 16S rDNA pyrosequencing
For extraction of total genome DNA from the compost samples,
samples were extracted using MOBIO DNeasy Power Soil Kit, following
the manufacturer’s instructions. 16S rRNA genes of distinct regions
(16S V4) were amplified used specific primer 515F (5′-GTGCCAGCM
GCCGCGGTAAT-3′) and 806R (5′-GGACTACHVGGGTWTCTAA-3′) with
the barcode. All PCR reactions were carried out in 30 μL reactions with
15 μL of Phusion® High-Fidelity PCR Master Mix (New England
Biolabs); 0.2 μM of forward and reverse primers, and about 10 ng
templates DNA. Thermal cycling consisted of initial denaturation at
98 °C for 1 min, followed by 30 cycles of denaturation at 98 °C for 10 s,
annealing at 50 °C for 30 s, elongation at 72 °C for 30 s. and finally 72 °C
for 5 min. Mix same volume of 1 × loading buffer with PCR products
and operate electrophoresis on 2% agarose gel for detection. PCR pro-
ducts were mixed in equidensity ratios, then, mixture PCR products
were purified with GeneJETTM Gel Extraction Kit (Thermo Scientific).
Sequencing libraries were generated using Ion Plus Fragment Library
Kit 48 rxns (Thermo Scientific) following manufacturer's re-
commendations. The library quality was assessed on the Qubit@ 2.0
Fluorometer (Thermo Scientific). At last, the library was sequenced on
an Ion S5TM XL platform and 400 bp single-end reads were generated.
3
Bioresource Technology 289 (2019) 121653
2.4. Data analysis
All compost samples were repeated four times. The temperature,
pH, EC, GI, TOC, TN, TP, TK, and C/N data were analysed by SPSS 19.0
(SPSS for Windows, USA). The significance test was performed by using
one-way ANOVA (P < 0.05) in the different periods of the group,
student’s test for the same period between C and T.
Based on the IonS5TM XL sequencing platform, the sequences were
clustered into OTUs (Operational Taxonomic Units) with 97% identity,
a total of 1827 OTUs were obtained, and then the OTU sequence and
the Silva132 database species were annotated. In this study, the
Observed species, Chao1, Shannon, and Good-coverage indices were
calculated with QIIME (Version 1.7.0) and displayed with R software
(Version 2.15.3). Multiple variations of correlation between selected
influenced factors (EC, temperature, pH, TN, TOC, TP, TK and C/N) and
bacterial community composition (Top 10 genes) were analysed by CCA
using the vegan package in R software. The bacteria community were
analysed by the linear discriminant analysis (LDA) effect size (LEfSe)
method. First, the non-parametric factorial Kruskal-Wallis (KW) sum-
rank test was used to detect species with significant differences in
abundance between different groups. Then, groups of Wilcoxon were
used. The rank sum test was used to judge the differences between
groups. Finally, linear discriminant analysis (LDA) was used to achieve
dimensionality reduction and assess the magnitude of the difference
(i.e., biomarkers) (Segata et al., 2011).
To perform cluster analysis of functional relative abundance ac-
cording to the functional notes and abundance information of the
samples in the database (SILVA SSU Ref NR), the clustering was carried
out from the functional difference level in R software by the Tax4Fun
package (Segata et al., 2011). Spearman correlation analysis was per-
formed between the environmental factors and the abundance of bac-
terial community composition (Top 30 genes) in R software by the
Corr.test function of the psych package and the pheatmap function in
the pheatmap package (Segata et al., 2011).
3. Results and discussion
3.1. Changes in pile temperature
Changes in temperature during the composting process reflect mi-
crobial community changes in the compost, and it is also important for
pig manure composting to be harmless and stable (Ma et al., 2018). In
this study, the ambient temperature was 23 °C on the first day (Fig. 1a),
but the temperature of compost increased sharply, with T reaching
50 °C within only 2 days and reaching a peak of 67.3 °C, while C
reached 50 °C at 4 days and reached a peak of 64.7 °C; also, the tem-
perature of T was higher than 60 °C for 10 days, while C was higher
than 60 °C for 8 days. This result can be explained as follows: the ad-
dition of microbial inoculation can accelerate the temperature increase
and prolong the thermophilic phase, the high temperature produced
during the composting process can not only kill the pathogenic bacteria,
weed seeds, eggs and so on but can also determine the microbial
community that plays a dominant role in the composting process. This
was supported by the research of Wu et al. (2017) and Zhong et al.
(2018). In addition, the thermophilic phase (temperature greater than
50 °C) of T and C lasted for 25 days and 23 days, respectively. The
temperature fluctuation of T is greater than that of C during the ther-
mophilic phase, indicating that the addition of microbial inoculation in
composting can accelerate the degradation of organic matter and con-
tribute considerable heat from microorganisms, thereby promotinge
large temperature fluctuations (Li et al., 2015). As the reaction pro-
gressed, the temperature of the pile began to decrease (the temperature
fell below 50 °C) and gradually approached the ambient temperature on
the 28th day; the temperature of T decreased faster than that of C in the
cooling stage and maturity stage. This result indicated that the addition
of microbial inoculation can not only accelerate the degradation rate of
C. Li, et al.
Fig. 1. Changes of temperature (°C) (a), pH (b), EC (c), GI (d) in C and T during the composting period. Among them, EC and GI are abbreviations for Electrical
conductivity and Germination index, respectively.
organic matter and harmless standards in the pile but can also shorten
the composting period (Nakasaki et al., 2013).
3.2. Changes in physicochemical properties during composting
3.2.1. pH
The pH values reached 8.6 and 8.3 on the 4th day for T and C,
respectively (Fig. 1b). The pH value showed the most direct evidence of
microbial activity during composting. The appropriate pH range was
6.0 to 9.0. Otherwise, if the pH is too high or too low, the normal
composting reaction would be affected (Zhao et al., 2016). The pH
values of C and T were 8.8 and 9.0 on the 12th day, respectively. This
was mainly due to in the initial stage of composting, there is more
organic matter available in the pile, the microbial metabolism is fast,
and the small molecular organic acids were volatilized with the increase
of composting temperature (Chen et al., 2017). Meanwhile, ammonia
produced by microorganism decomposition of nitrogen-containing or-
ganic compounds causes the value of pH to begin to rise in compost
(Mao et al., 2018). The final pH values were 8.8 and 8.7 in T and C
(P > 0.05), which were mainly caused by the decomposition of or-
ganic matter by microorganisms in the later composting period and the
production of large amounts of organic acids, such as acetic acid and
butyric acid (Wei et al., 2016).
3.2.2. Ec
EC was used to indicate the total concentration of water-soluble salt.
The EC value of T and C were 3.1 ms cm-1 and 3.5 ms cm−1, respec-
tively, at 4th day. This result attributed to the addition of microbial
inoculation, which could accelerate the ammonia volatilization and the
transformations NH4+ and NO3–, as recommended by Zhang et al.
4
Bioresource Technology 289 (2019) 121653
(2017). The value of EC increased gradually at 24th day during com-
posting for both T and C. This increase responsible for the rapid de-
gradation of organic matter in the pile, resulting in a large number of
small molecules, including various anions and cation, such as HCO3–,
Mg2+, Na+, K+, and Ca2+, which increased the value of EC (Zhang and
Sun, 2014). However, the value of EC in T was higher than that in C for
day 12, which might be attributed to the active metabolism of micro-
organisms and thermophilic bacteria in T at the early period of com-
posting, which contributed to the decomposition of abundant organic
matter in straw, resulting in more nutrient ions than C (Garcia-Gomez
et al., 2005). After 24th day, the EC of T and C was 3.8 and 3.7, re-
spectively, which remained largely unchanged and tended to be stable.
3.2.3. Germination index
Using a typical seed germination index (GI) to estimate compost
maturity by evaluating the phytotoxicity of a plant is regarded as ac-
ceptable by researchers; GI greater than 80% indicates that the compost
is not toxic to plant growth, which is one of the basic requirements for
composting to maturity and achieving non-toxicity (Zhang et al., 2018).
In the 4th day (Fig. 1d), GI values were 35.5% and 36.5% in C and T
(P > 0.05), respectively, indicating that the plant toxicity of compost
was higher in both C and T during the initial stage of composting. With
the composting process for both the C and T compost, the GI value was
gradually increased, and the GI was more than 80% in the 24th day;
meanwhile, the GI of T was greater than Zhou et al. (2019a) reported a
similar phenomenon that compost with corn straw and pig manure can
cause the content of the toxic substances in the compost to be gradually
transferred or disappeared. Our results also indicated that the addition
of microbial inoculation could promote the growth of alfalfa seeds.
C. Li, et al.
Fig.2. The character of TOC (a), TN (b), TP (c), TK (d) and C/N (e) during composting. The difference significance in the group was expressed in lowercase letters a,
b, c and d. (P < 0.05). The difference between groups was expressed with *, ns indicate no difference significance.
3.2.4. ToC, TN, TP, TK, C/N
The TOC content decreased constantly during the composting pro-
cess at 4–12 days (Fig. 2a). The content of TOC in C did not change
significantly, while the TOC content decreased sharply in T with a
significant difference, and the TOC reduction was observed from 46.7%
to 42.5%. This phenomenon indicates that the addition of microbial
inoculation could accelerate the reduction of TOC in composting. This
effect responsible for the fact that active thermophilic microorganisms
decompose organic carbon resource into CO2 and H2O in the com-
posting process, resulting in the decomposition of a large amount of
TOC in compost (Huang et al., 2017). Chefetz et al. (1996) reported that
composting is a metabolic process in which microorganisms participate,
and TOC can provide an organic carbon source for microorganisms.
Correlation analysis also indicated (Fig. 6b) that such microbes as Pu-
sillimonas, Taibaiella, Hydrogenispora, Lysobacter, Cellvibrio, Paenibacillus
and Idiomarina at the genus level have a significant negative correlation
with TOC content.
The content of TN were 2.1% and 1.8% in C and T in the 4th day
(P < 0.05), respectively (Fig. 2b).This result indicated that the addi-
tion of microbial inoculation could not effectively reduced nitrogen loss
in 4th day, Zhao et al. (2016) explained the fact that microbial
5
Bioresource Technology 289 (2019) 121653
inoculation could accelerate temperature increases and lead to TN vo-
latilization in the form of ammonia at the rising temperature stage. The
TN content showed an upward trend in 12th day compared with 4th
day in T, while C remains unchanged. This might be because nitrogen-
containing organic compound was accelerated degradation by micro-
bial inoculation (Zhang, 2016). As the fermentation continued, the
content of TN was 2.7% in both C and T, and the results showed that
microbial inoculation could not effectively increase the TN content of
compost.
The range of TP was 1.8–2.3% and 1.2–2.6% in C and T, respec-
tively, in the composting process (Fig. 2c), which showed that the ad-
dition of microbial inoculation can effectively improve the TP to pro-
mote the nutrient content of the product (Wei et al., 2018a,b). This
finding suggests that Thermobifida, Thermopolyspora, Caldicoprobacter,
Limnochorda, Luteimonas, Oceanobacillus, Vulgatibacter, Lysobacter,
Iamia, Thermomonospora and Microbispora at the genus level have a
significant positive correlation with TP (Fig. 6b).
TK has been an increasing trend, and the change in TK was
1.2–1.8% and 1.3–1.9% in C and T throughout the composting process
(Fig. 2d). The results of Khan et al. (2014) indicated that TK increased
continuously with increasing fermentation, and the absolute contents
C. Li, et al.
would not change but would only transform among different forms.
Through correlation analysis (Fig. 6b), it was found that Iamia, Ther-
momonospora, Microbispora, Limnochorda, Caldicoprobacter, Thermo-
polyspora, and Thermobifida were positively correlated with TK.
Through fermentation of microorganisms, the volume and weight of
compost were reduced with the decomposition and conversion of vo-
latile organic compounds (Gaind, 2014; Sanchez et al., 2017). The
content of TK in C and T was not significantat at the end of composting,
which is observed because TK in compost is relatively stable with
continuous fermentation, and the total dry matter of the pile decreases
(Zhou et al., 2019a); therefore, the relative content of TK in the pile
increases.
The suitable C/N is an indispensable nutritional condition to pro-
vide the optimal metabolic efficiency of microorganisms in the compost
(Ma et al., 2018). The addition of microbial inoculation increased C/N
in pile T compared with C at 4th day (Fig. 2e). Which observed because
the addition microbial inoculation contributed a high metabolic rate
and the volatilization of nitrogen; thus the content of C/N in pile T was
higher than that in Meng et al. (2017) reported that the degradation
rate of TOC in the early composting period is smaller than that of ni-
trogen loss and volatilization, which leads to the increased trend of C/
N. The final C/N decreased 13.3 and 15.3 in C and T, respectively. C/N
is an important parameter to evaluate the maturity and stability of
compost, theoretically, the final C/N was approximately 15 after
composting when the pile was fully matured (Guo et al., 2012). Cor-
relation analysis found that the microorganisms negatively correlated
with C/N were mainly Microbispora, Thermomonospora, Iamia, Vulgati-
bacter, Limnochorda, Caldicoprobacter, Thermopolyspora, and Thermo-
bifida at the genus level (Fig. 6b).
3.3. Changes in bacterial communities and diversity with microbial
inoculants of the microbial community
3.3.1. Diversity and richness of the bacterial community
During the 4–12 days of composting, the Chao1 index in C showed a
decreasing trend, while the Chao1 index in T presented an increasing
trend (Fig. 3a). The Chao1 index was used to indicate the bacterial
community richness, and the larger value indicated higher community
richness (Liu et al., 2017). The Shannon index also showed the same
trend in C and T with Chao1 (Fig. 3b). The Shannon index was chosen
to reflect the degree of bacterial community diversity, and the greater
Shannon index indicated the higher diversity of the bacterial commu-
nity (Liu et al., 2017). Both the Chao1 and Shannon indices indicated
that microbial inoculation can promote the large-scale reproduction of
community in the thermophilic stage (4–12 days) of compost. Sun et al.
(2016) found that a microbial inoculant not only enhanced the richness
and diversity of bacteria but also accelerated the decomposition rate.
After 24 days, the Shannon index was larger in C compared with T. This
finding help to explain why the addition of microbial inoculation pro-
moted the rapid increase in temperature, and the abundant thermo-
philic bacteria reduced the contents of organic matter and other nu-
trients available in pile T, indicating that the diversity of the bacterial
community in the thermophilic period was more notable after the ad-
dition of microbial inoculant to compost (Xi et al., 2012).
A good coverage index was commonly used to describe the depth of
sequencing, which presented results that accurately reflect the depth of
sequencing samples (Li et al., 2015). The good coverage index of all
samples was more than 0.99 during composting (Fig. 3c), which in-
dicated the reasonableness of sequencing data (Mao et al., 2018). The
observed species index reflects the number of bacterial communities
that was intuitively observed (Fig. 3d); on the 4th day, for observed
species, C was 700, and T was 350. Compared with C, T was decreased
by 50.0%, which is observed because at the beginning of composting,
adding microbial inoculant could accelerate the rapid increase of
temperature in compost T and kill a large number of pathogenic bac-
teria (Zhang et al., 2013; Zhou et al., 2019a). Intolerant bacteria died in
6
Bioresource Technology 289 (2019) 121653
large numbers; therefore, the number of observed species in compost T
was significantly lower than in compost C.
3.3.2. Changes in the bacterial community structure
The bacteria at the dominant phylum level were Proteobacteria,
Actinobacteria, Firmicutes and Bacteroidetes (Fig. 4a); however, the
relative abundances of each sample were clearly different. The relative
abundances of Proteobacteria, Actinobacteria, Firmicutes and Bacter-
oidetes were 11.7–51.1%, 0.9–31.1%, 11.4–46.7% and 4.5–27.8%, re-
spectively, in compost C, and they were 10.8–74.5%, 2.1–49.0%,
18.2–59.8% and 2.7–30.5%, respectively, in compost T. These phylum-
level bacteria are widely used in the treatment of agricultural waste
(Wang et al., 2018; Sanchez et al., 2017).
At the genus level (Fig. 4b), the relative abundances of Acinetobacter
(which belongs to the Proteobacteria phylum) were 2.2–24.1% and
48.7–22.2% in C and T, respectively, at 4–12 days. It was found that
Acinetobacter increased in C and decreased in T. The results showed that
the content of Acinetobacter in the compost could be significantly re-
duced by adding microbial inoculation. Acinetobacter at the genus level
of many bacteria not only plays an important role in the rapid de-
gradation of straw in compost but also relates to the metabolism of
carbon and nitrogen (Awasthi et al., 2017). Additionally, Acinetobacter
decomposes a large amount of organic matter to produce organic acid
due to the high temperature of the compost (4–12 days), the organic
acid was volatilized in the form of gas, resulting in an increase in the pH
value of the compost (Wei et al., 2016). This effect was also indicated
by correlation analysis, which showed a positive correlation with TOC
(Fig. 5b). Studies have shown that inoculation of Actinobacteria at dif-
ferent stages of composting can accelerate the maturity and stability of
compost products (Zhao et al., 2016).
The dominant genus populations of Flavobacterium (belonging to
Bacteroidetes phylum) in C and T were 17.4–22.7% and 2.5–21.7%,
respectively, at 4–12 days. It was found that microbial inoculation
could make Flavobacterium propagate in larger quantities compared
with C. The number of Flavobacterium gradually increased with the
composting process; it was not killed during the thermophilic stage of
composting and can inhabit thermophilic and extreme environments
(Jurado et al., 2014). Through correlation analysis, it was found that
the temperature (P < 0.05) and C/N (P < 0.01) were positively cor-
related (Fig. 5b). In addition, Flavobacterium can accelerate the de-
gradation of cellulose, hemicellulose and lignin, which are difficult to
degrade in compost (Mao et al., 2018).
The dominant genus populations of Solibacillus (belonged to
Firmicutes phylum) in C and T were 7.9–1.3% and 1.4–10.3%, re-
spectively, at 12–24 days. It was found that microbial inoculation could
increase the relative abundance of Solibacillus at 12–24 days. The pos-
sible reason is that a large amount of TOC produced in compost T
provides an abundant carbon source for Solibacillus metabolism, and
correlation analysis showed that Solibacillus and TOC were positively
correlated (P < 0.05) (Fig. 5b). The dominant genera of Lysinibacillus
(belonging to Firmicutes) in C and T were 0.7–1.0% and 17.6–1.1%,
respectively, at 24–32 days. This evidence supports that after the con-
tinuous effect of high temperature at thermophilic stage, microbial in-
oculation could cause the relative abundance of Lysinibacillus to exhibit
a decreasing trend. This finding is observed because the sharp decrease
in Lysinibacillus content contributes to the decrease in temperature
during the cooling stage (24–32 days). Zhao et al. (2016) reported Ly-
sinibacillus can not only withstand high temperature but also help to
increase the EC in composting, Additionally, the amount of organic
matter that can be easily absorbed by Lysinibacillus during composting,
usually with a positive correlation between Lysinibacillus and nitrogen
C/N (Liu et al., 2018). Correlation analysis also support this evidence
that Lysinibacillus are positively correlated with temperature and C/N
(P < 0.05) and are negatively correlation with EC (P < 0.01),
(Fig. 5b).
Both C and T had the highest abundance of Thermopolyspora
C. Li, et al.
Fig. 3. Alpha diversity of C and T, Chao 1 (a), Shannon (b), Good coverage (c), Observed species (d) in all composting stages.
(belonging to the Actinobacteria phylum) on the 32nd day (maturity
stage), which was 6.2% and 11.0%, respectively. Compared with C, the
addition of microbial inoculation can promote the increase relative
abundance of Thermopolyspora in compost raw materials at the genus
level. The higher relative abundance of Thermopolyspora indicates that
the toxicity of compost mature products is lower than that of (Zhou
et al., 2019a). This abundance can also promote the dissolution of in-
soluble phosphorus and potassium in compost materials (Sun et al.,
2016), correlation analysis showed that Thermopolyspora and TK, TP
were positively correlated (P < 0.01) and negatively correlated with
temperature (P < 0.05) (Fig. 5b). In conclusion, the change in micro-
bial communities is related to the microhabitat in which it was located
(Huang et al., 2017); at the same time, this change also provided the
basis for directional screening of microbes at the genus level to accel-
erate the composting process.
3.3.3. LEfSe analysis
LEfSe analysis was used to identify bacterial communities or species
that differed significantly in each sample (Zhou et al., 2019b). To fur-
ther analyse the bacterial community structure, LEfSe analysis showed
that there were 78 biomarkers (Fig. 5a, 5b) with LDA scores > 4. There
were 11 taxa in C4, the most important contribution to C4 was Flavo-
bacterium_sp_M1_I (at the species level), which can accelerate the com-
post degradation rate and transformation of organic phosphorus (Wei
et al., 2018a,b). In addition, there are Sphingobacterium_jejuense and
7
Bioresource Technology 289 (2019) 121653
paenibacillus_camelliae at the species level. Of the 10 taxa in T4, the
largest contribution to T4 was Pseudomonadales (at the order level),
which is the most important in disease suppression and promotion plant
growth (Mehta et al., 2014; Zhao et al., 2016; Fan et al., 2018). 4 taxa
were observed in C12, 7 taxa in T12, 8 taxa in C24, 12 taxa in T24, and
the most important contributions to C12, T12, C24 and T24 were Fla-
vobacteriales (at the order level), Bacteroidetes (at the phylum level),
Cellvibrionales (at the order level) and Firmicutes (at the phylum level).
It was found that the addition of microbial inoculation could increase
the number of biomarkers. Which Firmicutes were thought to play a
major role in lignocellulose degradation (Jurado et al., 2014). 12 taxa
in C32 and 14 taxa in T32, the most important contributions to C32
were Limnochordales (at the order level), Limnochordales (at the fa-
mily level) and Limnochordales (at the class level), while T32 was
Actinobacteria (at the phylum level). Actinobacteria could form spores
in severe environments, such as high temperature and low water con-
tent; therefore, it was one of the most important microbial communities
in the composting process (Zhao et al., 2016). These findings indicated
significant differences in the distribution of microbes between C and T
in different stages, which could reflect the effect of microbial inoculant
on the composting process to some extent.
3.3.4. Cluster analysis of microbial metabolism functions
Compost samples can be grouped into three categories. C4, C12, T12
gather in one category (Fig. 6b), and it was found that the abundance of
C. Li, et al. Bioresource Technology 289 (2019) 121653

Fig. 4. The relative abundance of bacteria at the phylum level (a), at the genus level (b) among all samples. The bar chart shows the top 10 bacterial community with
the largest relative abundance.

functional genes was higher than that related to metabolism and human likely observed because the thermophilic stage of compost was pro-
diseases in the thermophilic stage (4–12 days). Toledo et al. (2017) longed by adding microbial agents (Fig. 1a), causing a large number of
reported that a higher abundance of metabolism was helpful to de- pathogenic bacteria to be killed. Wu et al. (2017) reported reaching the
compose the degradation of cellulose and hemicellulose of compost. A compost sanitation requirements when the temperature more than
higher abundance of human diseases indicated a large number of pa- 7 days during the thermophilic stage of composting.
thogenic bacteria. In all kinds of infectious diseases, bacterial diseases
are the most common, such as dysentery in livestock faces, which often
3.4. Correlations between composting environment factors and bacterial
occurs around the world (Zhou et al., 2019b). T4, C24, C32, T24 come
communities at the top 10 genera
together in the same category, and it was found that the content of
cellular processes, genetic information processing and organismal sys-
The results showed that eight environmental factors (TN, TOC, T,
tems were higher, and the content of human diseases gene in C32 was
TP, TK, EC, pH, C/N) explained the first axis by 41.9% by CCA (Fig. 6a).
lower in compost. The results showed that the abundance of functional
Environmental factors determine the succession of microorganisms in
genes related to human diseases was low at the maturity stage in
the process of composting (Zhao et al., 2016; Huang et al., 2017). The
compost C.
results showed that temperature was positively correlated with the first
T32 is grouped into one category alone. It was found that the
axis (r = 0.999) followed by C/N (r = 0.918) and TOC (r = 0.551)
abundance of human disease-related functional genes could be reduced
(Table 2). This finding indicated that at the 4th day to the 12th day, the
by adding microbial inoculant at the maturity period. This finding is
compost was rich in TOC and high in C/N content, and with the

8
C. Li, et al. Bioresource Technology 289 (2019) 121653

Fig. 5. The evolutionary branch diagram of differential bacterial community or species (a). Indicator microbial in groups with LDA scores higher than 4 (b). The
circle of size from inside to outside of evolutionary branching graphs in a group indicated the importance of abundance from phylum to genus or species, the same
color as a group indicated that it is important in this group. The LDA value (influence value of linear discriminant analysis) distribution histogram shows bacterial
community or species (Biomarker) with LDA score greater than 4. All the bacterial community or species displayed have statistical difference among all groups. The
letters k, p, c, o, f, g and s are stand for the abbreviation of kingdom, phylum, class, order, family, genus and species. The bacterial community or species with
significant abundance difference in different groups were shown, and the length of histogram represented the influence of Biomarkers.

increased temperature, which promotes the positive correlation of
bacteria at the genus level, Acinetobacter, Solibacillus, Pseudomonas, and
Flavobacterium were observed (Fig. 5a). Wei et al. (2016) noted that the
abundance of heat-resistant bacteria increased during the thermophilic
phase, and a large amount of TOC served as a nutrient for bacterial
community growth.
A negative correlation was found in TN (r = −0.918), pH
(r = −0.369), EC (r = −0.549), TP (r = −0.656) with the first axis.
This negative correlation is observed because such bacteria as
Lysinibacillus, Bacillus, Thermobifida, Limnochorda, Thermopolyspora, and
Caldicoprobacter, which cannot withstand high temperature, were killed
or rendered dormant, while the abundance of the heat-resistant bacteria
increased. Meng et al. (2017) reported that during this period, the de-
creased pH values resulted from the organic acids produced by heat-
resistant microbial community metabolism decreased in the compost,
and part of TN was volatilized in the form of ammonia.
The composting period with positive correlation with the second
axis were C24, T24, C32 and T32 from bottom to top. A negative cor-
relation was found in TP (r = −0.683), pH (r = −0.610), and T
(r = −0.019). Zhang et al. (2013) reported that the organic matter was
decomposed completely, the temperature gradually decreased, the TN
volatilization decreased, and the change in pH tended to be stable
during the maturity stage. This effect might be responsible for a large
number of positive correlations of microbial communities, such as Ly-
sinibacillus, bacillus, Thermobifida, Thermopolyspora, Caldicoprobacter
and Limnochorda, on the 24th day to the 32 day, resulting in an increase
in the content of EC at the end of composting. This effect was also
supported by CCA, which indicated that the compost environmental
factors positively correlated with the second axis, not only for EC
(r = 0.647) but also for TK (r = 0.375) and TN (r = 0.035). The T24
microorganism was affected more by EC, TK, TN, TOC, and C/N than by
C24, mainly by Lysinibacillus and Bacillus. The addition of bacterial
inoculation to pig manure compost promoted the mineralization of
TOC, increased the porosity of the substrate and stimulated the mi-
crobial activity (Zhang and Sun, 2015).
At the end of composting, the microbial communities of C and T
were dominated by Thermopolyspora, Caldicoprobacter and Limnochorda.
4. Conclusions
This study used 16S rDNA technology to demonstrate that em-
ploying microbial inoculant can effectively increase the content of TP
and reduce the abundance of human disease-related functional genes,
and there was no plant toxicity at the end of composting. In addition,
microbial inoculation did not increase pH, EC and TN when the com-
posting was finished; however, Chao1 and Shannon indices increased at

9
C. Li, et al. Bioresource Technology 289 (2019) 121653

Fig. 6. CCA analysis between composting environment factors and bacterial communities (a). Spearman correlation analysis between composting environment
factors and bacterial communities (b). Cluster analysis between microbial metabolism functions and each sample (c). In CCA analysis the environmental factor is
represented by arrow, and its length represents the correlation degree between a certain environmental factor and community distribution. The angle between the
arrow line and the sorting axis represents the correlation between a certain environmental factor and the sorting axis. The smaller angle represents the higher the
correlation is, on the contrary, the lower the correlation is. In Spearman correlation analysis the intermediate heat map corresponding value is the Spearman
correlation coefficient r, r > 0 is the positive correlation, r < 0 is negative correlation, * indicates the significance test P value < 0.05, ** indicates the significance
test P value < 0.01.

Table 2 Acknowledgements
The result of significance test of environmental factors.
Environmental Correlation coefficient Coefficient of Significance
All the authors gratefully acknowledge the Utilization of pig and
factors between environmental determination test (Pr > r) poultry wastes of Gansu province (No.GARS-ZJ-6). The Selection and
factors and sorting axis (R2) degradation mechanism of Pesticide degrading bacteria in Agricultural
and Animal Husbandry production system of Gansu Agricultural
Axis 1 Axis 2
University (No.GAU-XKJS-2018-007), and National Natural Science
TN −0.9994 0.0354 0.8238 0.0005*** Foundation of China (No.31660688).
TOC 0.5512 0.8344 0.5798 0.0005***
T 0.9998 −0.0194 0.8118 0.0005*** Appendix A. Supplementary data
TP −0.7301 −0.6833 0.8032 0.0005***
TK −0.9272 0.3746 0.6587 0.0005***
EC −0.7626 0.6468 0.5198 0.0010*** Supplementary data to this article can be found online at https://
pH −0.7926 −0.6097 0.2144 0.0840 doi.org/10.1016/j.biortech.2019.121653.
C/N 0.9179 0.3967 0.6985 0.0005***
References

the thermophilic stage (4–12 days). Microbial inoculation increased the
number of biomarkers with LDA scores great that 4 at the rising tem-
perature stage, thermophilic stage, cooling phase and maturity stage.
Awasthi, M.K., Zhang, Z., Wang, Q., Shen, F., Li, R., Li, D., Ren, X., Wang, M., Chen, H.,
Zhao, J., 2017. New insight with the effects of biochar amendment on bacterial di-
versity as indicators of biomarkers support the thermophilic phase during sewage
sludge composting. Bioresour. Technol. 238, 589–601.
Barrington, S., Choiniere, D., Trigui, M., Knight, W., 2002. Effect of carbon source on
compost nitrogen and carbon losses. Bioresour. Technol. 83, 189–194.

10
C. Li, et al.
Chefetz, B., Hatcher, P.G., Hadar, Y., Chen, Y., 1996. Chemical and biological char-
acterization of organic matter during composting of municipal solid waste. J.
Environ. Qual. 25, 776–785.
Chen, W., Liao, X., Wu, Y., Liang, J., Mi, J., Huang, J., Zhang, H., Wu, Y., Qiao, Z., Li, X.,
Wang, Y., 2017. Effects of different types of biochar on methane and ammonia mi-
tigation during layer manure composting. Waste Manage. 61, 506–515.
Chen, Y., Huang, X., Han, Z., Huang, X., Hu, B., Shi, D., Wu, W., 2010. Effects of bamboo
charcoal and bamboo vinegar on nitrogen conservation and heavy metals immobility
during pig manure composting. Chemosphere 78, 1177–1181.
Cui, H., Zhao, Y., Chen, Y., Zhang, X., Wang, X., Lu, Q., Jia, L., Wei, Z., 2017. Assessment
of phytotoxicity grade during composting based on EEM/PARAFAC combined with
projection pursuit regression. J. Hazard. Mater. 326, 10–17.
Dias, B., Silva, C., Higashikawa, F., Roig, A., Sanchez-Monedero, M., 2010. Use of biochar
as bulking agent for the composting of poultry manure: effect on organic matter
degradation and humification. Bioresour. Technol. 101, 1239–1246.
Gaind, S., 2014. Effect of fungal consortium and animal manure amendments on phos-
phorus fractions of paddy-straw compost. Int. Biodeterior. Biodegrad. 94, 90–97.
Garcia-Gomez, A., Bernal, M., Roig, A., 2005. Organic matter fractions involved in-
degradation and humification processes during composting. Compost. Sci. Util. 13,
127–135.
Fan, H., Liao, J., Abass, O., Liu, L., Huang, X., Wei, L., Li, J., Xie, W., Liu, C., 2018. Effects
of compost characteristics on nutrient retention and simultaneous pollutant im-
mobilization and degradation during co-composting process. Bioresour. Technol.
275, 61–69.
Guo, R., Li, G., Jiang, T., Schuchardt, F., Chen, T., Zhao, Y., Shen, Y., 2012. Effect of
aeration rate, C/N ratio and moisture content on the stability and maturity of com-
post. Bioresour. Technol. 112, 171–178.
Huang, C., Zeng, G., Huang, D., Lai, C., Xu, P., Zhang, C., Cheng, M., Wan, J., Hu, L.,
Zhang, Y., 2017. Effect of Phanerochaete chrysosporium inoculation on bacterial
community and metal stabilization in lead-contaminated agricultural waste com-
posting. Bioresour. Technol. 243, 294–303.
Janczak, D., Malinska, K., Czekała, W., Caceres, R., Lewicki, A., Dach, J., 2017. Biochar to
reduce ammonia emissions in gaseous and liquid phase during composting of poultry
manure with wheat straw. Waste Manag. 66, 36–45.
Jurado, M., Lopez, M., Suarez-Estrella, F., Vargas-Garcia, M., Lopez-Gonzalez, J., Moreno,
J., 2014. Exploiting composting biodiversity: Study of the persistent and biotechno-
logically relevant microorganisms from lignocellulose-based composting. Bioresour.
Technol. 162, 283–293.
Khan, N., Clark, I., Sanchez-Monedero, M.A., Shea, S., Meier, S., Bolan, N., 2014. Maturity
indices in co-composting of chicken manure and sawdust with biochar. Bioresour.
Technol. 168, 245–251.
Li, R., Wang, J., Zhang, Z., Shen, F., Zhang, G., Qin, R., Li, X., Xiao, R., 2012. Nutrient
transformations during composting of pig manure with bentonite. Bioresour.
Technol. 121, 362–368.
Liu, N., Hou, T., Yin, H., Han, L., Huang, G., 2018. Effects of amoxicillin on nitrogen
transformation and bacterial community succession during aerobic composting. J.
Hazard. Mater. 362, 258–265.
Liu, N., Zhou, J., Han, L., Ma, S., Sun, X., Huang, G., 2017. Role and multi-scale char-
acterization of bamboo biochar during poultry manure aerobic composting.
Bioresour. Technol. 241, 190–199.
Li, R., Wang, Q., Zhang, Z., Zhang, G., Li, Z., Wang, L., Zheng, J., 2015. Nutrient trans-
formation during aerobic composting of pig manure with biochar prepared at dif-
ferent temperatures. Environ. Technol. 36, 815–826.
Ma, S., Fang, C., Sun, X., Han, L., He, X., Huang, G., 2018. Bacterial community suc-
cession during pig manure and wheat straw aerobic composting covered with a semi-
permeable membrane under slight positive pressure. Bioresour. Technol. 259,
221–227.
Mao, H., Lv, Z., Sun, H., Li, R.H., Zhai, B., Wang, Z., Awasthi, M., Wang, Q., Zhou, L.,
2018. Improvement of biochar and bacterial powder addition on gaseous emission
and bacterial community in pig manure compost. Bioresour. Technol. 258, 195–202.
Mehta, C., Palni, U., Franke-Whittle, I., Sharma, A., 2014. Compost: Its role, mechanism
and impact on reducing soil-borne plant diseases. Waste Manage. 34, 607–622.
Meng, X., Liu, B., Xi, C., Luo, X., Yuan, X., Wang, X., Zhu, W., Wang, H., Cui, Z., 2017.
Effect of pig manure on the chemical composition and microbial diversity during co-
composting with spent mushroom substrate and rice husks. Bioresour. Technol. 251,
22–30.
Nakasaki, Araya, S., Mimoto, H., 2013. Inoculation of Pichiakudriavzevii RB1 degrades
the organic acids present in raw compost material and accelerates composting.
Bioresour. Technol. 144, 521–528.
Sanchez, O., Ospina, D., Montoya, S., 2017. Compost supplementation with nutrients and
microorganisms in composting process. Waste Manage. 69, 136–153.
Segata, N., Izard, J., Waldron, L., Gevers, D., Miropolsky, L., Garrett, W., Huttenhower, C.,
11
Bioresource Technology 289 (2019) 121653
2011. Metagenomic biomarker discovery and explanation. Genome boil. 12 (6), R60.
Sun, D., Lan, Y., Xu, E., Meng, J., Chen, W., 2016. Biochar as a novel niche for culturing
microbial communities in composting. Waste Manage. 54, 93–100.
Toledo, M., Gutierrez, M.C., Siles, J.A., Garcia-Olmo, J., Martin, M.A., 2017.
Chemometric analysis and NIR spectroscopy to evaluate odorous impact during the
composting of different raw materials. J. Cleaner Prod. 167, 154–162.
Wang, Q., Awasthi, M., Ren, X., Zhao, J., Li, R., Wang, Z., Wang, M., Chen, H., Zhang, Z.,
2018. Combining biochar, zeolite and wood vinegar for composting of pig manure:
The effect on greenhouse gas emission and nitrogen conservation. Waste Manage. 74,
221–230.
Wang, Q., Wang, Z., Awasthi, M., Jiang, Y., Li, R., Ren, X., Zhao, J., Shen, F., Wang, M.,
Zhang, Z., 2016. Evaluation of medical stone amendment for the reduction of ni-
trogen loss and bioavailability of heavy metals during pig manure composting.
Bioresour. Technol. 220, 297–304.
Wei, Y., Wu, D., Wei, D., Zhao, Y., Wu, J., Xie, X., Zhang, R., Wei, Z., 2018a. Improved
lignocellulose-degrading performance during straw composting from diverse sources
with actinomycetes inoculation by regulating the key enzyme activities. Bioresour.
Technol. 271, 66–74.
Wei, Y., Zhao, Y., Lu, Q., Cao, Z., Wei, Z., 2018b. Organophosphorus – degrading bacterial
community during composting from different sources and their roles in phosphorus
transformation. Bioresour. Technol. 264, 277–284.
Wei, Y., Zhao, Y., Wang, H., Lu, Q., Cao, Z., Cui, H., Zhu, L., Wei, Z., 2016. An optimized
regulating method for composting phosphorus fractions transformation based on
biochar addition and phosphate-solubilizing bacteria inoculation. Bioresour. Technol.
221, 139–146.
Wu, J., He, S., Liang, Y., Li, G., Li, S., Chen, S., Nadeem, F., Hu, J., 2017. Effect of
phosphate additive on the nitrogen transformation during pig manure composting.
Environ. Sci. Pollut. Res. Int. 24, 17760–17768.
Xi, B., He, X., Wei, Z., Jiang, Y., Li, M., Li, D., Li, Y., Dang, Q., 2012. Effect of inoculation
methods on the composting efficiency of municipal solid wastes. Chemosphere 88,
744–750.
Yang, F., Li, Y., Han, Y., Qian, W., Li, G., Luo, W., 2019. Performance of mature compost
to control gaseous emissions in kitchen waste composting. Sci. Total Environ. 675,
262–269.
Yuan, J., Chadwick, D., Zhang, D., Li, G., Chen, S., Luo, W., Du, L., He, S., Peng, S., 2016.
Effects of aeration rate on maturity and gaseous emissions during sewage sludge
composting. Waste Manag. 56, 403–410.
Zhu, Y., Johnson, T.A., Su, J., Qiao, M., Guo, G., Stedtfeld, R.D., Hashsham, S.A., Tiedje,
J.M., 2013. Diverse and abundant antibiotic resistance genes in Chinese swine farms.
PNAS 110, 3435–3440.
Zhang, D., Luo, W., Li, Y., Wang, G., Li, G., 2018. Performance of co-composting sewage
sludge and organic fraction of municipal solid waste at different proportions.
Bioresour. Technol. 250, 853–859.
Zhang, L., Sun, X., 2015. Effects of earthworm casts and zeolite on the two-stage com-
posting of green waste. Waste Manag. 39, 119–129.
Zhang, L., Sun, X., Tian, Y., Gong, X., 2013. Effects of brown sugar and calcium super-
phosphate on the secondary fermentation of green waste. Bioresour. Technol. 131,
68–75.
Zhang, P., 2016. Effects of straw incorporation on the soil nutrient contents, enzyme
activities, and crop yield in a semiarid region of China. Soil Tillage Research. 160,
65–72.
Zhao, Y., Lu, Q., Wei, Y., Cui, H., Zhang, X., Wang, X., Shan, S., Wei, Z., 2016. Effect of
actinobacteria agent inoculation methods on cellulose degradation during com-
posting based on redundancy analysis. Bioresour. Technol. 219, 196–203.
Zhong, X., Ma, S., Wang, S., Wang, T., Sun, Z., Tang, Y., Deng, Y., Kida, K., 2018.
Acomparative study of composting the solid fraction of dairy manure with or without
bulking material: Performance and microbial community dynamics. Bioresour.
Technol. 247, 443–452.
Zhou, G., Xu, X., Qiu, X., Zhang, J., 2019a. Biochar influences the succession of microbial
communities and the metabolicfunctions during rice straw composting with pig
manure. Bioresour. Technol. 272, 10–18.
Zorpas, A., 2014. Recycle and reuse of natural zeolites from composting process: a 7-year
project. Desalinat. Water Treat. 52, 109–117.
Zhang, D., Luo, W., Yuan, J., Li, G., Luo, Y., 2017. Effects of woody peat and superpho-
sphate on compost maturity and gaseous emissions during pig manure composting.
Waste Manag. 68, 56–63.
Zhou, H., Zhang, D., Jiang, Z., Sun, P., Xiao, H., Yu, W., Chen, J., 2019b. Changes in the
soil microbial communities of alpine steppe at Qinghai-Tibetan Plateau under dif-
ferent degradation levels. Sci. Total. Environ. 651, 2281–2291.
Zhang, L., Sun, X., 2014. Changes in physical, chemical, and microbiological properties
during the two-stage co-composting of green waste with spent mushroom compost
and biochar. Bioresour. Technol. 171, 274–284.