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An improved effective microorganism (EM) soil ball-making method for water quality restoration

DOI:10.1007/s11356-015-5617-x
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Gun-Seok Park at AtoGen Co., Ltd.
Gun-Seok Park
  • AtoGen Co., Ltd.
Abdur Rahim Khan at University of California, Davis
Abdur Rahim Khan
Yunyoung Kwak at Kyungpook National University
Yunyoung Kwak
Sung-Jun Hong at Kyungpook National University
Sung-Jun Hong
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Abstract and Figures

Soil balls containing the so-called effective microorganisms (EM) have been applied to improve water quality of small ponds, lakes, and streams worldwide. However, neither the physical conditions facilitating their proper application nor the diversity of microbial community in such soil balls have been investigated. In this study, the application of 0.75 % of hardener to the soil balls exerted almost neutral pH (pH 7.3) which caused up to a fourfold increased hardness of the soil ball. Moreover, the 0.75 % of hardener in the soil ball also improved the water quality due to a significant reduction in dissolved oxygen, total phosphorus, and total nitrogen contents. Metagenomic analysis of the microbial community in the soil ball with 0.75 % hardener was compared with control (traditional soil ball) through next-generation sequencing. The traditional soil ball microbial community comprised 96.1 % bacteria, 2.7 % eukaryota, and 1 % archaea, whereas the soil ball with 0.75 % hardener comprised 71.4 % bacteria, 27.9 % eukaryota, and 0.2 % viruses. Additionally, metagenomic profiles for both traditional and improved soil balls revealed that the various xenobiotic biodegradation, such as those for caprolactam, atrazine, xylene, toluene, styrene, bisphenol, and chlorocyclohexane might be responsible for organic waste cleanup.
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SELECTEDPAPERS FROMTHE 2ND CONTAMINATED LAND, ECOLOGICAL ASSESSMENTAND REMEDIATION(CLEAR 2014) CONFERENCE: ENVIRONMENTALPOLLUTION AND REMEDIATION
An improved effective microorganism (EM) soil ball-making
method for water quality restoration
Gun-Seok Park
1
&Abdur Rahim Khan
1
&Yunyoung Kwak
1
&Sung-Jun Hong
1
&
ByungKwon Jung
1
&Ihsan Ullah
2
&Jong-Guk Kim
3
&Jae-Ho Shin
1
Received: 9 June 2015 /Accepted: 15 October 2015 /Published online: 24 October 2015
#Springer-Verlag Berlin Heidelberg 2015
Abstract Soil balls containing the so-called effective micro-
organisms (EM) have been applied to improve water quality
of small ponds, lakes, and streams worldwide. However, nei-
ther the physical conditions facilitating their proper applica-
tion nor the diversity of microbial community in such soil
balls have been investigated. In this study, the application of
0.75 % of hardener to the soil balls exerted almost neutral pH
(pH 7.3) which caused up to a fourfold increased hardness of
the soil ball. Moreover, the 0.75 % of hardener in the soil ball
also improved the water quality due to a significant reduction
in dissolved oxygen, total phosphorus, and total nitrogen con-
tents. Metagenomic analysis of the microbial community in
the soil ball with 0.75 % hardener was compared with control
(traditional soil ball) through next-generation sequencing. The
traditional soil ball microbial community comprised 96.1 %
bacteria, 2.7 % eukaryota, and 1 % archaea, whereas the soil
ball with 0.75 % hardener comprised 71.4 % bacteria, 27.9 %
eukaryota, and 0.2 % viruses. Additionally, metagenomic pro-
files for both traditional and improved soil balls revealed that
the various xenobiotic biodegradation, such as those for cap-
rolactam, atrazine, xylene, toluene, styrene, bisphenol, and
chlorocyclohexane might be responsible for organic waste
cleanup.
Keywords Biodegradation .Effective microorganisms .
Metagenome .Microbial community .Soil ball .Water quali ty
Introduction
Microbial biotechnology is the most recent approach to waste-
water treatment and is essential for protecting human health
and the environment (Mielczarek et al. 2013). In order to
guarantee the optimal operation using the effective microor-
ganism technology, it is very important to understand the
structure, function, and microbial community dynamics in-
volved in these approaches (Zakaria et al. 2010). Soil balls
have been used as carrier to immobilize effective microorgan-
isms for the remediation of contaminated water environment
(Ekpeghere et al. 2012). The concept of Beffective
microorganisms^(EM) was introduced by Dr. Teruo Higa,
and since then, EMs became an important part of natural
farming (Higa 1998). Those microbes are selected based on
their functions as fixation of atmospheric nitrogen, decompo-
sition of organic wastes and residues, suppression of soil-
borne pathogens, recycling and increased availability of plant
nutrients, solubilization of insoluble nutrient sources, and deg-
radation of toxicants including pesticides (Higa and Parr
1994). EM is co-cultures of naturally occurring beneficial mi-
croorganisms, and it is widely applied as inoculants to soil,
water and plants for the improvement of soil and water quality,
as well as for plant growth and crop yield (Grover et al. 2011;
Javaid 2010). Soil balls with EM can be a good alternative for
stream water treatment in eco-friendly environments, but there
have been no report on their physicochemical properties and
Responsible editor: Robert Duran
Electronic supplementary material The online version of this article
(doi:10.1007/s11356-015-5617-x) contains supplementary material,
which is available to authorized users.
*Jae-Ho Shin
jhshin@knu.ac.kr
1
School of Applied Bioscience, Kyungpook National University,
Daegu 702-701, Republic of Korea
2
Institute of Biotechnology and Genetic Engineering, The University
of Agriculture, Peshawar, Pakistan
3
School of Life Sciences and Biotechnology, Kyungpook National
University, Daegu 702-701, Republic of Korea
Environ Sci Pollut Res (2016) 23:1100–1107
DOI 10.1007/s11356-015-5617-x
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
... A study by Park et al. [31] also sought to improve the quality of water of streams by studying the physical conditions and the diversity of the microorganisms in the water. Park et al. [31] made use of soil balls that contained EM and found that the quality of water was improved by hardener soil balls as it significantly reduced the dissolved oxygen, total nitrogen, and total phosphorus components. ...
... A study by Park et al. [31] also sought to improve the quality of water of streams by studying the physical conditions and the diversity of the microorganisms in the water. Park et al. [31] made use of soil balls that contained EM and found that the quality of water was improved by hardener soil balls as it significantly reduced the dissolved oxygen, total nitrogen, and total phosphorus components. The composition of the hardener soil ball and traditional soil ball microbial community was found to differ with the hardener soil balls comprising 0.2% viruses, 27.9% eukaryote, and 71.4% bacteria while the traditional soil ball consisted of 1% archaea, 2.7% eukaryote, and 96.1% bacteria. ...
... An assessment of pathways that support the xenobiotic degradation was also performed. An examination at the metagenomic profiles for both the hardened soil and the Contaminated harbor sediments Loess balls containing EM Acetic acid, propionic acid, valeric acid [14] Artificial lake Solution COD, TN, TP, TSS, turbidity [47] Synthetic polluted water Solution NH 3 -N, TP, COD [19] Stream water Soil balls containing EM TP, TN, xenobiotic [31] Artificial river water Mudballs containing EM COD, TSS [23] Safwat and Matta Journal of Engineering and Applied Science (2021) 68:48 Page 4 of 12 traditional soils showed that a certain percentage of the organic waste cleanup was due to the xenobiotic biodegradation [31]. Mudballs containing EM were also applied by a study by Nugroho et al. [23] to examine the impact that temperature had on COD and TSS removal. ...
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Effective Microorganisms (EM) Technology for Water Quality Restoration and Potential for Sustainable Water Resources and Management
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Water quality has received considerable attention in allocation processes for maximizing the satisfaction of various sectors. However, pollutant impurities that impede adequate supply of water have a detrimental effect on the quality and harmful for living organisms including aquatic life. For the reduction of water pollution level, various chemical and biological treatments are available but the emergence of an amazing technology of a multiculture of anaerobic and aerobic beneficial microorganisms is presently gaining popularity due to its environmentally friendly nature. This effective microorganism (EM) technology uses naturally occurring microorganisms which are able to purify and revive nature. Applications of EM using the formula known as effective microorganism activated solution (EMAS) have been experimented in several rivers in Malaysia depending on the scale, location, physical and geographical conditions with the principal objective of enhancing and improving the water quality. One of the significant contributions of EM based rehabilitation of polluted and degraded water bodies is to restore aquatic habitats and ecosystems. Existing results of projects via EM technology in solving water quality related problems, and the nationwide campaigns in Malaysia are duly presented. The role of EM-based water restoration approach for sustainability of water resources and prospects of modeling are also discussed. Results clearly demonstrated the effectiveness of this technique for restoration of water quality of degraded/polluted river basin. Valuable lines for further research and acceptance of EM technology for the future are thus suggested as it is believed to be the key to sustained environmental improvement and offers a real opportunity for eco-innovation.
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Quantitative (real-time) PCR
Chapter
Full-text available
  • Jan 2005
Many nucleic acid-based probe and PCR assays have been developed for the detection tracking of specific microbes within the rumen ecosystem [4-6, 8-11, 14, 15]. Conventional PCR assays detect PCR products at the end stage of each PCR reaction, where exponential amplification is no longer being achieved. This approach can result in different end product (amplicon) quantities being generated [3]. In contrast, using quantitative, or real-time PCR, quantification of the amplicon is performed not at the end of the reaction, but rather during exponential amplification, where theoretically each cycle will result in a doubling of product being created w [13]. For real-time PCR, the cycle at which fluorescence is deemed to be detectable above the background during the exponential phase is termed the cycle threshold (Ct). The Ct values obtained are then used for quantitation, which will be discussed later.
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Role of microorganisms in adaptation of agriculture crops to abiotic stresses
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Full-text available
  • May 2011
Increased incidences of abiotic and biotic stresses impacting productivity in principal crops are being witnessed all over the world. Extreme events like prolonged droughts, intense rains and flooding, heat waves and frost damages are likely to further increase in future due to climate change. A wide range of adaptations and mitigation strategies are required to cope with such impacts. Efficient resource management and crop/livestock improvement for evolving better breeds can help to overcome abiotic stresses to some extent. However, such strategies being long drawn and cost intensive, there is a need to develop simple and low cost biological methods for the management of abiotic stress, which can be used on short term basis. Microorganisms could play a significant role in this respect, if we can exploit their unique properties of tolerance to extremities, their ubiquity, genetic diversity, their interaction with crop plants and develop methods for their successful deployment in agriculture production. Besides influencing the physico-chemical properties of rhizospheric soil through production of exopolysaccharides and formation of biofilm, microorganisms can also influence higher plants response to abiotic stresses like drought, chilling injury, salinity, metal toxicity and high temperature, through different mechanisms like induction of osmo-protectants and heat shock proteins etc. in plant cells. Use of these microorganisms per se can alleviate stresses in crop plants thus opening a new and emerging application in agriculture. These microbes also provide excellent models for understanding the stress tolerance, adaptation and response mechanisms that can be subsequently engineered into crop plants to cope with climate change induced stresses. KeywordsAbiotic stress tolerance–Microorganisms–Crop production–Climate change
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Molecular Biology Techniques Used in Wastewater Treatment: An Overview
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Full-text available
Identification of microorganisms by conventional methods requires the isolation of pure cultures followed by laborious characterization experiments. These procedures are therefore inadequate for study of the biodiversity of a natural or engineered ecosystem. A new set of molecular techniques developed during the 1990s revolutionized microbial ecology research. Among these techniques, cloning and the creation of a gene library, denaturant gradient gel electrophoresis (DGGE) and fluorescent in situ hybridization with DNA probes (FISH) stand out. Cloning provides very precise taxonomical information, but is time consuming and requires specialized personnel and so its introduction in wastewater treatment has been slow. DGGE is a rapid and simple method that provides characteristic band patterns for different samples, allowing quick sample profiling, while retaining the possibility of a more thorough genetic analysis by sequencing of particular bands. FISH makes possible to identify microorganisms at any desired taxonomical level, depending on the specificity of the probe used. It is the only quantitative molecular biology technique, although quantification is either complex or tedious and subjective. Combination with a confocal laser-scanning microscope allows the visualization of three-dimensional microbial structures (granules, biofilms). The methods discussed have deepened our understanding of the microbiology of biological wastewater treatment. PCR-based methods (cloning and DGGE) have proved suitable for identifying the microorganisms that form the sludge. Both DGGE and FISH have been extensively employed. FISH is currently being used for elucidation of the composition, quantification and distribution of different bacterial groups in granules and biofilms, as well as their structure and architecture.
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Combined Use of 16S Ribosomal DNA and 16S rRNA to Study the Bacterial Community of Polychlorinated Biphenyl-Polluted Soil
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Full-text available
  • Apr 2001
The bacterial diversity assessed from clone libraries prepared from rRNA (two libraries) and ribosomal DNA (rDNA) (one library) from polychlorinated biphenyl (PCB)-polluted soil has been analyzed. A good correspondence of the community composition found in the two types of library was observed. Nearly 29% of the cloned sequences in the rDNA library were identical to sequences in the rRNA libraries. More than 60% of the total cloned sequence types analyzed were grouped in phylogenetic groups (a clone group with sequence similarity higher than 97% [98% for Burkholderia and Pseudomonas-type clones]) represented in both types of libraries. Some of those phylogenetic groups, mostly represented by a single (or pair) of cloned sequence type(s), were observed in only one of the types of library. An important difference between the libraries was the lack of clones representative of the Actinobacteria in the rDNA library. The PCB-polluted soil exhibited a high bacterial diversity which included representatives of two novel lineages. The apparent abundance of bacteria affiliated to the beta-subclass of the Proteobacteria, and to the genus Burkholderia in particular, was confirmed by fluorescence in situ hybridization analysis. The possible influence on apparent diversity of low template concentrations was assessed by dilution of the RNA template prior to amplification by reverse transcription-PCR. Although differences in the composition of the two rRNA libraries obtained from high and low RNA concentrations were observed, the main components of the bacterial community were represented in both libraries, and therefore their detection was not compromised by the lower concentrations of template used in this study.
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The Microbial Database for Danish wastewater treatment plants with nutrient removal (MiDas-DK) - A tool for understanding activated sludge population dynamics and community stability
Article
  • Jun 2013
Since 2006 more than 50 Danish full-scale wastewater treatment plants with nutrient removal have been investigated in a project called 'The Microbial Database for Danish Activated Sludge Wastewater Treatment Plants with Nutrient Removal (MiDas-DK)'. Comprehensive sets of samples have been collected, analyzed and associated with extensive operational data from the plants. The community composition was analyzed by quantitative fluorescence in situ hybridization (FISH) supported by 16S rRNA amplicon sequencing and deep metagenomics. MiDas-DK has been a powerful tool to study the complex activated sludge ecosystems, and, besides many scientific articles on fundamental issues on mixed communities encompassing nitrifiers, denitrifiers, bacteria involved in P-removal, hydrolysis, fermentation, and foaming, the project has provided results that can be used to optimize the operation of full-scale plants and carry out trouble-shooting. A core microbial community has been defined comprising the majority of microorganisms present in the plants. Time series have been established, providing an overview of temporal variations in the different plants. Interestingly, although most microorganisms were present in all plants, there seemed to be plant-specific factors that controlled the population composition thereby keeping it unique in each plant over time. Statistical analyses of FISH and operational data revealed some correlations, but less than expected. MiDas-DK (www.midasdk.dk) will continue over the next years and we hope the approach can inspire others to make similar projects in other parts of the world to get a more comprehensive understanding of microbial communities in wastewater engineering.
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Phosphorus removal in pilot plant using biofilm filter process from farm wastewater
Article
  • Aug 2006
Various environmental conditions affecting total phosphorus removal from farm wastewater in a biofilm filter, process were investigated using loess balls andChromobacterium LEE-38 at a pilot plant. WhenChromobacterium LEE-38 was used, the removal efficiency of total phosphorous was approximately 10- or 5-fold higher than that ofAcinetobacter CHA-2-14 orAcinetobacter CHA-4-5, respectively. When a loess ball of 11–14 mm manufactured at a 960°C calcining temperature was used, the removal efficiency of total phosphorous was 90.0%. When 70% of the volume fraction was used, the maximum efficiency of total phosphorus removal was 93.1%. Notably, when the initial pH was in the range of 6.0 to 8.0, the maximum removal efficiency of total phosphorus was obtained after 30 days. When the operating temperature was in the range of 30 to 55°C, the maximum removal efficiencies of total phosphorus, 95.6 to 94.6%, were obtained. On the other hand, at operating temperatures below 20°C or above 40°C, the removal efficiency of total phosphorous decreased. Among the various processes, biofilm filter process A gave the highest removal efficiency of 96.4%. Pilot tests of total phosphorus removal using farm wastewater from the biofilm filter process A were carried out for 60 days under optimal condition. WhenAcinetobacters sp. Lee-11 was used, the average removal efficiency in thep-adsorption area was only 32.5%, and the removal efficiencies of chemical oxygen demand (COD) and biological oxygen demand (BOD) were 56.7 and 62.5%, respectively. On the other hand, whenChromobacterium LEE-38 was used, the average removal efficiency was 95.1%, and the removal efficiencies of COD and BOD were 91.3 and 93.2%, respectively.
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Comparison of bacterial community structures in two systems of a sewage treatment plant using PCR-DGGE analysis
Article
The combination of PCR amplification of 16S rRNA genes with denaturing gradient gel electrophoresis (DGGE) analysis was used to reveal the compositions and dynamics of bacterial communities in a sewage treatment plant with two systems, i.e., an anoxic-anaerobic-aerobic system (inverted A20) and an anaerobic-anoxic-aerobic one (conventional A20) over a period from February to July 2009, during which both systems experienced serious sludge bulking problems. The DGGE patterns showed that there were many common bands in both systems, suggesting the high similarity of bacterial communities of the two systems. Meanwhile, the moving window correlation analysis showed that the two systems experienced different microbial community structure changes during the period, which might be related with the different situations of the occurrence and disappearance of sludge bulking, as being reflected by sludge volume index (SVI) values. Major bands of DGGE patterns of sludge samples were further sequenced. Phylogenetic affiliation indicated that the majority of the sequences obtained were affiliated with Actinobacteria, Firmicutes, Bacteroidetes/Chlorobi group and alpha- and beta-Proteobacteria. Two sequences showed high similarities to typical filamentous bacteria Microthrix parvicella and Nostocoida limicola I, indicating that these bacterial species have been involved in the sludge bulking problems.
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