Adirianto_2023_IOP_Conf._Ser.__Earth_Environ._Sci._1263_012047.pdf

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Effect of biochar in soil on microbial diversity: a
meta-analysis
To cite this article: B Adirianto and T Bachtiar 2023 IOP Conf. Ser.: Earth Environ. Sci. 1263 012047
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6th International Symposium on Sustainable Urban Development 2023
IOP Conf. Series: Earth and Environmental Science 1263 (2023) 012047
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doi:10.1088/1755-1315/1263/1/012047
1
Effect of biochar in soil on microbial diversity: a meta-analysis
B Adirianto1,* and T Bachtiar2
1
Bogor Agricultural Development Polytechnic, Bogor, Indonesia
2
National Research and Innovation Agency of Indonesia, Jakarta, Indonesia
*bayuadirianto@polbangtan-bogor.ac.id
Abstract. The diversity, structure, and behavior of soil microbes communities,
which are crucial to the breakdown of organic matter, cycling of nutrients, and
general health of the soil, can be impacted by biochar. This study uses a meta-
analysis approach to examine how biochar affects soil microbial diversity, and it
anticipates that the results will take the form of a summary of the information that
has already been published in journals. This study presents a meta-analysis of 24
articles published between 2018 and 2023 that reported biochar's effect on soil
microbial diversity and richness. Alpha diversity indexes such as Shannon, Simpson
(Diversity index), Chao1, and ACE (Richness Index) were measured as parameters,
as well as the operational taxonomic units (OTUs) count. The levels of biochar
dosage varied from 0 to 50% w/w. Simpson (0.546), the OTUs (0.473), Chao1
(0.227), Shannon (0.125), and ACE (0.056) had the most significant effect sizes for
the biochar (Hedges'd), with the majority of the values impact sizes being on the
right. According to aggregate-driven tree analysis, the type of biochar, application
rate, use of the soil, and length of the experiment all play a significant role in how
biochar affects soil microbial diversity. In conclusion, adding biochar requires
considering biochar application rates and type to improve microbes' diversity.
1. Introduction
Advanced modification techniques can affect the characteristics of biochar, which typically depend
on the type of feedstock and pyrolysis temperature [1]. For improving soil quality for plant growth,
biochar is frequently used as a supplement [2,3], reduce environmental pollution [4], and increase
carbon sequestration [5]. However, there is still room for improvement in our systematic
comprehension of how adding biochar affects the biomass and diversity of soil microbes. It can
selectively enhance the growth of specific microbial taxa, potentially supporting beneficial

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doi:10.1088/1755-1315/1263/1/012047
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microbes involved in nutrient cycling or disease suppression [6]. Biochar can provide physical and
chemical support for the soil microbes community by regulating soil aggregation, soil
characteristics, nutrients, and enzyme activity [7].
In contrast, the changes in the pore structure of soil modified with biochar can affect nutrient
cycling [8]. The surface area of biochar is significant [9] and can absorb and retain nutrients,
preventing leaching and making them available for microbial uptake [10]. Biochar application can
increase the availability of nutrients for soil microbes, promoting their growth and activity.
Furthermore, biochar has the potential to buffer soil pH, reducing fluctuations that can affect the
number and movement of the microbes community [11,12]. Biochar can interact with soil organic
matter, affecting its decomposition rate and changing the carbon cycle dynamics in soil [13].
Biochar's ability to increase the soil's water-holding capacity [14] prevents faster moisture loss.
The effect of biochar on soil microbes can vary depending on factors such as biochar feedstock,
pyrolysis conditions, application dose, soil type, and climate. Since soil microbes composition is
one of the parameters of soil quality, using biochar is worth considering.
We hypothesize that applying biochar with various properties and using it in different soil
conditions causes major changes in the soil microbial population. The 16S rRNA gene sequencing
has become a standard method to assess microbial diversity. This culture-independent DNA-based
study resulted in an illustrative large-scale data set microbial composition of a particular niche [15].
The quantity and design of the species present and the diversity of the bacteria all contribute to
defining characteristics of the bacterial community in a given niche. To compare the bacterial
diversity of the sampled microorganisms, various bioinformatics methods have been created [16–
18]. Tools such as the ShannonWeaver and Simpson indexes can describe the population's diversity
in a sample [15]. Microbial diversity was assessed using the Shannon and Simpson indices, whereas
microbial richness was evaluated using the Ace and Chao1 indices [19]. Few research have looked
at how biochar affects the variety of soil microbes. Based on this, the researchers aimed to analyze
the effect of biochar in soil on microbial diversity (OTUs, Shannon, Simpson, Chao1, and ACE)
through the meta-analysis method. We operated a meta-analysis to identify the impact of varied
control situations (the term of the experiment and biochar application dose), soil attributes (initial
soil pH), and pyrolysis terms (feedstock and pyrolysis temperature).
2. Methods
2.1 Paper classification
Researchers collect information from Scopus (https://www.sciencedirect.com). The researcher
investigates several keywords, such as "biochar", "community", "microbes", "richness", and
"diversity". Researchers only cite fulfilled articles for which all facts are accessible for analysis.
Literature was collected from the period 2018 to 2023. Inclusion criteria for an article are as
follows: (1) the articles were published in English; (2) the investigation was conducted based on
the application of biochar; (3) the biochar addition reported rate; and (4) the microbes diversity
index (OTUs, Shannon, Simpson, Chao1 and ACE) to be compared from the control group with
added to biochar; (5) study included the mean, standard error (SE) or standard deviation (SD), and
any stated or quantified treatment, at least three independent replicates. After evaluating the
abstract and full text, 24 articles (describing 106 experiments) met the inclusion criteria (Table-1).

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doi:10.1088/1755-1315/1263/1/012047
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When published studies report multiple trials, each entity is coded separately. All experiments were
carried out directly.
2.2. Data categorization and treatment.
For the sub-group meta-analysis, biochar is categorized based on the raw material: rice straw, wheat
straw, bamboo, pine chips, wood chips, corn stalk, weed stalk, canola stalk, tree branch, and rice
husk. Biochar rates were grouped into <2%, 2-4%, 5-10%, and >10%. Soil acidity is categorized
into three different soil pH (H2O) levels into: very acid (<4.5), acid (4.5-5.5), slightly acid (5.5-
6.5), neutral (6.6-7.5), slightly alkaline (7.6-8.5), alkaline (>8.5). Trial duration (time since biochar
was applied) was categorized into <3 Months, 3-6 months, 6-12 months, and >12 months. Pyrolysis
temperatures of biochar are categorized into four groups (≤ 400 °C, 401–500 °C, 501–600 °C and
> 600 °C). The microbial diversity was categorized into bacteria and fungi, and the habitat was
categorized into agricultural soil, polluted soil, wetland constructed, and compost.
2.3. Data computation and numerical analyses.
For each standardized pairwise comparison (control and biochar treatment), Hedges standard mean
differences [20,21] were counted between the control and biochar application groups to verify the
metric effect size. Using the OpenMee program [22], A 95% bootstrap confidence interval (CI)
and average effect sizes for each grouping were calculated. The 95% confidence interval did not
overlap with 0 for the effect magnitude to be considered significant.
3. Results and Discussions
3.1. The overall effect of biochar on soil microbial diversity
Based on a meta-analysis of the effect of biochar in soil on microbial diversity, the "overall effect
size" value is in the range of 0.125 to 0.546, where all of the "effect size" value is on the right.
Nevertheless, the effects of biochar on soil microbes diversity were not significant at Shannon,
Chao1, and ACE (Figure 1). Although the effect size value of all diversity indexes is on the right,
only OTUs and Simpson show a significant effect size value. OTUs showed effect size values of
0.473 (P= 0.032). Meanwhile, Simpson showed effect values of 0.546 (P<0.001). Based on this,
only the OTUs and Simpson indices were continued to test moderating variables.
Table 1. Experiments considered in the meta-analysis of the impact of applying biochar on the
diversity index of soil microbes.
Study
no.
Reference Biochar Type Pyrolisis
Temperature
(C)
Biochar rate
(%) w/w
Duration of
experiment
1 [12] Rice straw 500 0; 0.5; 1; 2; and
5
3 months
2 [23] Corn stalk; grass, wood chip 450; 800 0 and 5 3 months
3 [4] Wheat straw 500 0; 1; 2; and 4 2 weeks
4 [24] Rice straw 500 0 and 10 1.5 month
5 [25] Leaves and wood chips 300 0 and 4 3 months
6 [26] Bamboo 500 0 and 50 24 months
7 [27] Tree branches 550 0 and 4 4 months

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Study
no.
Reference Biochar Type Pyrolisis
Temperature
(C)
Biochar rate
(%) w/w
Duration of
experiment
9 [29] Cornstalk 600 0 and 2 1 month
10 [30] Cornstalk 500 0; 0.01; and
0,04
3 months
11 [31] Pine chip 700 0; 2.5; and 5 3 months
12 [32] Wood chip 500 0 and 2 24 months
13 [33] Sewage sludge 700 0 and 5 2 months
14 [34] Rice straw 500 0 and 10 1 month
15 [35] Weed stalk 500 0; 1; 3; and 4 2 months
16 [36] Cornstalk 450 0 and 1.25 3 months
17 [37] Wheat straw 500 and 700 0 and 0.4 1.5 months
18 [38] Bamboo 600 0; 1; and 3 5 months
19 [8] Rice straw and Canola stalk 350 and 550 0 and 1 15 months
20 [39] Rice husk 500 0 and 3 1 month
21 [40] Cornstalk 550 0 and 10 0; 0.5; 1; 2; 2.5
months
22 [41] Rice husk 600 0.05; 0.1; 0.25 2 weeks
23 [42] Cornstalk 400 0 and 1 7 months
Figure 1. The total effect of biochar on microbes diversity: Operational Taxonomic Unit (OTUs), Shannon, Simpson,
Chao1 and Ace. If the effect size's 95% bootstrap confidence interval (CI) did not include 0, it was deemed statistically
significant.
Table 2. Summary of the effect size Hedges'd of soil microbes diversity index.
Response
parameter
n Effect size Lower bound Upper bound Standard
Error
P-value
OTUs 504 0.473 0.04 0.907 0.221 0.032*
Shannon 762 0.125 -0.195 0.446 0.164 0.444
Simpson 504 0.546 0.237 0.854 0.157 < 0.001*
Chao1 599 0.227 -0.111 0.566 0.173 0.188
ACE 268 0.056 -0.384 0.496 0.224 0.803
* changes considered significant when the 95% confidence interval for the effect magnitude does not include zero.
The wheat straw biochar under the OTUs index showed an effect size value of 2.616 (P< 0.001),
while the Simpson effect size showed an effect size value of 1.004 (P <0.001). The cornstalk only
showed a significant effect under the Simpson index with a value of 2.92 (P< 0.001). The pyrolysis

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temperature of 401-500o
C and > 600o
C significantly affected soil microbes diversity based on
OTUs with effect size 1.144 (P<0.001). At the same time, the Simpson index showed pyrolysis
temperature at 401-500o
C (effect size=0.58, P=0.009) and 501-600 C (effect size=0.85, P=0.015)
significantly affected soil microbes diversity. However, biochar's effects on soil microbes diversity
were not significant at biochar pyrolysis temperature <400o
C (Figure 2). The effect size of OTUs
(0.966) was more effective in <2% biochar application rates (P= 0.042) than in soil with other
biochar rates, while Simpson had the highest responses to 5-10% biochar rates (effect size=1.185,
P<0.001). On average, soil microbial diversity-based OTUs and Simpson showed consistently
significant effects to biochar addition at <3 Month duration of the experiment. Based on the
duration of the experiment, OTUs showed an effect size value of 1.184 (P= 0.002). Meanwhile,
Simpson showed an effect size value of 0.934 (P<0.001).
The biochar treatment on fungal diversity was significantly favorable under the OTUs index
(effect size value=1.249, P= 0.006). At the same time, the Simpson index showed that biochar
application had a significant effect on bacteria (effect size value=0.502, P=0.006) and fungal
diversity (effect size value=0.683, P=0.032). Soil pH from different ranges affects soil microbial
diversity under biochar application (Figure 3). Soil with neutral pH significantly affected soil
microbial diversity based on OTUs (effect size 0.865, P= 0.011) and Simpson (effect size=0.87, P=
0.003). The Simpson index showed that biochar was significantly positive on microbial diversity
when applied to the polluted soil (effect size=1.321, P<0.001).
3.2. Impacts of moderating variables on soil microbial diversity
(A)OTUs (B) Simpson
Feedstock
Application
Rate

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Effect size (Hedges'd) Effect size (Hedges'd)
Figure 2. Effect size Hedges'd for the soil microbial diversity (A) OTUs; and (B) Simpson; of soil under different
biochar characteristics (feedstock of biochar, biochar pyrolysis temperature, rate of biochar, and duration of the
experiment). If the effect size's 95% bootstrap confidence interval (CI) did not include 0, it was deemed statistically
significant.
(A)OTUs (B) Simpson
Figure 3. Effect size Hedges'd for the soil microbial diversity (A) OTUs; and (B) Simpson; of soil under different
conditions (microbial group, soil pH, and soil use type). If the effect size's 95% bootstrap confidence interval (CI) did
not include 0, it was deemed statistically significant.
3.3. Discussions
The diversity of soil microbial changed according to the biochar feedstock, biochar pyrolysis
temperature, biochar application rates, the duration of the experiment (Figure 2), microbes group,
soil pH, and soil use type (Figure 3). However, the Cohen Benchmark provides rough estimates
with a mean effect size d > 0.8, indicating a large effect, 0.2 < d < 0.8 a moderate effect, and 0 < d
< 0.2 small effects [44,45]. The result showed that the Simpson index has a larger mean effect size
than the OTUs.
3.3.1 Result of biochar on soil microbes heterogeneity under different biochar characteristics
(feedstock of biochar, biochar temperature, biochar rate, and experiment duration)
Soil microbial diversity is an essential evidence of soil properties and significantly influences soil
health [46]. The biochar amendment undoubtedly improves the physicochemical properties such
as pH, especially in acid soils [47]. Due to its characteristics or by the absorption of nutrients with
surface functional groups, such as carboxylic groups, biochar supplies nutrition to soil bacteria.
The C/N ratio of some biochar fragments suggests that they can be utilized as a carbon source by
Microbes
Soil pH
Duration

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soil bacteria. The properties of biochar are critical in affecting soil microbes' diversity (Table 1)
because they provide a unique microenvironment that shapes microorganisms and can transform
soil microbes' community structure (Fig. S1). Wheat straw biochar is thought to be physically able
to have good pores as a place for microbes and protect from environmental stress. It is suspected
that wheat straw biochar has high stability in soil due to its high carbon availability, complex
aromatic structure, and inherent chemical inertia.
The biochar produced at 501-6000
C can increase the diversity of soil microbes. This follows
research by Li et al. [48] that biochar with pyrolysis of 600-700 °C can increase the total microbes
significantly and bacterial diversity. Biochar produced at higher temperatures (500 ◦C to 700 ◦C)
has a more specific surface area that might be capable of facilitating microbial life. Biochar's
specific surface area and total pore volume rise as pyrolysis temperature rises [37]. This meta-
analysis showed that a 5-10% biochar application rate gave the highest effect size value (Figure 2)
on microbial diversity. Xu et al. [49] also showed that in Pb spiked mud clay, 5% biochar treatment
enhanced microbial respiration, microbial biomass C, and soil C utilizing efficiency. Biochar with
more than 10% is thought to make the environment for growing soil microbes richer in oxygen due
to increased soil porosity. Li et al. [48] concluded that biochar tends to impair microbial variety
when applied at a high pace because it can severely disturb the microenvironment for microbial
growth. In addition, the residual organic carbon in biochar stimulates microbial growth at low
levels but inhibits it at high rates [50]. Microbes can use volatile carbon provided by fresh biochar
but aromatic carbon by aging biochar, and variations in the use of carbon resources inferred from
biochar may cause noticeable changes in the pattern of the soil microbial population [51–53].
Biochar accelerates microbial biomass while enhancing the fast growth of some microbes and
altering the community composition of soil microbes, although diversity is decreased [54].
3.3.2. Effect of biochar on soil microbes heterogeneity below varied conditions (type of soil
microbes, soil pH, and soil use).
Different pore sizes significantly affect microbial community structures, and the abundance of
microbes changes at the pore scale. Smaller holes are crucial for biochar's water-holding capacity
and nutrient adsorption, while bigger pores enable soil aeration, water storage, and internal
transportation [8]. Because fungal hyphae adhere to soil's solid particles, the fungi community is
typically more stable and less mobile [55]. Meantime, bacteria and fungi prefer various carbon
sources and nutrient materials. Some toxic elements are available to some bacteria, even harmful
to others [37]. Our study findings demonstrated that soil pH is essential in establishing microbial
community structure. When added to neutral soils, biochar triggers many actions. First, soil pH
changes the available nutrient content in the soil, which may affect specific microbial communities
or even support other microbial life. Secondly, the acidic or alkaline nature itself can make soil
microbes resistant to their environment. Along with pH shifts brought on by biochar, specific
reaction methods between fungi and bacteria are also present [56]. Simpson's index shows that
soils with polluted conditions significantly differed in soil microbial diversity when treated with
biochar. In addition, hazardous substances being present in the soil allows leaving environment
microbes resistant to toxic components, thereby reducing microbial diversity.

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4. Conclusions
In conclusion, based on the OTUs and Simpson indices, our results show that soil microbial
diversity increased significantly after soil was treated with biochar. Microbial diversity increases
more with biochar produced under pyrolysis temperature of 501-600 o
C, which comes from wheat
straw or cornstalk. In addition, this research revealed that a biochar application rate of 5-10% and
a study duration of <3 Months result in higher soil microbes heterogeneity. We come to the
conclusion that applying biochar increases the variety of fungi more significantly than bacteria. We
also came to the conclusion that neutral soil pH, with polluted soil being more susceptible to
biochar application, dramatically influences the response of microbial diversity to biochar. This
research showed that biochar can improve polluted soil by stimulating the growth of soil microbes.
This study will also help apply biochar as a soil amendment, which is critical for restoring the
microecological environment and health of the soil, sustainable land use management, and soil
quality improvement.
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