A Bioaugmentation Strategy for Promoting the Humification Process during Composting by Microbial Inoculants: A Review

Abstract

Stimulating compost humification is an important method to facilitate carbon sequestration, especially against the background of carbon neutrality. However, the disadvantages of traditional composting, including long humification cycles and high environmental risks, restrict its application. Microbial inoculants markedly increase the humus content of compost, and their performance is considerably influenced by the nature of the material, the microbial species, the inoculation dosages, and the inoculation methods. So far, the effects of microbial inoculants on compost maturity and microbial diversity have been widely studied, whereas an overview of their regulatory role in humus formation is still lacking. This review summarizes the promotional effects of microbial inoculants on humification and related biological mechanisms during composting. Further research on the development of microbial inoculants and the optimization of inoculation methods will promote humification and facilitate the production of high-quality compost.

1. Introduction

The problem of solid organic waste is becoming increasingly serious due to the worldwide expansion of different industries [1]. Until some years ago, landfill was still the most commonly used disposition of solid organic waste in many countries [2]. This not only causes bad odors but also creates greenhouse gas emissions and soil pollution [3]. Composting is a more useful way of converting various solid organic wastes into humus-like stable products by microorganisms [4].

The composting process is mainly a result of microbial metabolism [5]. Different microorganisms have different functions during composting. However, traditional composting is characterized by a long humification cycle, which is related to the poor activity of indigenous microbes. Thus, regulating microorganisms is a feasible way to facilitate nutrient transformation and humus enhancement in the composting process. As a biological regulating method, inoculation is an effective way to improve the composting microenvironment and increase microbial activity, which can accelerate compost maturation [6]. The addition of microbial inoculants is also a main bioaugmentation strategy and can markedly increase the levels of humic substances (HS) during composting [3]. However, the efficiency of this approach is sometimes uncertain because of the differences in inoculation methods and the quantity and types of the microbial inoculant [7, 8, 9, 10, 11, 12]. Therefore, the improvement of the humification process with microbial inoculants during composting is an important research field.

Given that stimulating compost humification is an important method to facilitate carbon sequestration, especially in the context of carbon neutrality [13], studies identifying the driving mechanisms of humus formation and searching for ways to improve the HS amounts during composting have been performed.

Generally, HS formation is the result of polymerization or condensation of humus precursors, such as phenols, amino acids, and reducing sugars, derived from macromolecule degradation and microbial synthesis under the actions of different metabolic pathways (Figure 1) [14, 15]. As one of important components of HS, humic acids (HA) are heterogeneous complexes with various molecular weights and functional groups. A higher HA content indicates a higher degree of humification, facilitating carbon sequestration and soil remediation [16]. Additionally, HA formation might be influenced by various factors, including the precursors, environmental factors, and functional microbial activities [17]. According to a previous study, thermophilic microbes play a key role in humification [18]. However, few reviews have summarized the mechanisms for improving compost humification by bioaugmentation.

Figure 1.

Formation pathway of humus during the aerobic composting process.

The use of microbial inoculants as a bioaugmentation strategy for the green disposal of solid organic waste is widely recommended. Numerous studies reported the considerable promotion of humification by applying microbial inoculants [19]. At the same time, the mechanisms behind these phenomena have attracted increased attention. Nowadays, with the development of novel analytical techniques, we have gained a deeper understanding of how the addition of functional microorganisms affects humification during composting. In this review, the biological mechanisms of microbial inoculants in promoting compost humification are systematically summarized based on recent studies, with the aim to provide scientific guidance for the efficient application of functional microorganisms to promote the conversion of organic components into humus.

2. Development of microbial inoculants used in composting

According to the principles of classical electromagnetism, charged ions move in a closed-loop circular motion within a uniform magnetic field. Herein, we cite a similar concept (“where it comes from, go there”), namely the “golden closed-loop rule.” It implies that microorganisms are to be selected and domesticated to obtain excellent strains for adapting to the native composting environment as well as facilitating the composting process [20, 21]. Accordingly, composting samples are used as the main sources of microbial inoculants. Specially, composting samples taken during the thermophilic period, as a potential microbial mine, have been favored so far [17, 22]. These samples are subjected to a series of microbiological experiments, such as dilution separation, colony purification, molecular identification, high-temperature tolerance tests, inter-strain antagonism tests, and degradative enzyme activity assays [23], to obtain candidates for developing compound microbial inoculants to promote the composting process (Figure 2). In high-temperature tolerance tests, temperatures of 45–55°C are frequently selected to identify thermophile microbes [24, 25]. In terms of degradative enzyme activity, cellulase, amylase, laccase, and FPase were investigated [21]. To improve the ability of the microbial inoculants to degrade the composting matrix, enrichment cultures can be adopted for selecting specialized functional strains, following the sole carbon source method. Suitable carbon sources contain wheat straw [26], coffee husk [27], pectin, or sodium carboxy-methylcellulose [28], but the screening and cultivating of microbes from compost are time consuming and laborious [29]. In this context, the efforts needed to develop microbial inoculants can be reduced via appropriate methods. In addition, since composting needs to be carried out throughout the year, in regions at high latitudes or during winter, the lower ambient temperature limits the composting process [30]. To overcome the adverse effects of low temperatures, it is equally necessary to develop psychrophilic functional strains to initiate composting. Based on previous studies, composting samples derived during a warm period (10°C) are ideal [31, 32, 33]. Considering the simple enzymatic performance and high environmental sensitivity of single strains, composite microbial inoculants need to be constructed to accelerate the biodegradation of organic matter in a synergistic manner [3].

Figure 2.

Illustration of the “golden closed-loop rule” for the development of microbial inoculants used in composting.

According to previous studies, while microbial inoculants facilitate fermentation processes, there are drawbacks of unidirectional nutrient conversion and insignificant compost humification [34]. Above all, the development of microbial inoculants for the directional promotion of compost humification has largely been neglected. However, humification is not only critical for product quality but also facilitates the remediation of polluted soil [4]. In recent years, substantial efforts have been made to develop microbial inoculants for enhancing the humification process in composting. According to various publications, the functional microbial inoculants which could significantly promote the humification process have unique nutritional metabolic properties (Table 1). These functional strains were derived from bacteria, fungi, and actinomycetes and show similar enzymatic features, such as high cellulase and oxidase activities [3, 17, 21, 34, 35, 36, 37, 38]. Additionally, they can also grow at high temperatures (45–60°C) and under adverse environmental conditions. It is therefore expected that these functional microbial inoculants can considerably promote humification during large-scale composting comparison trials. The next step is therefore the development of commercial products using these microbes [39], which represent an innovation in the field of microbial inoculant technology.

Strains	Source	Tolerated temperature (°C)	Cultivation time (d)	Medium	Representative enzymatic activity	Reference
Phanerochaete chrysosporium and Trichoderma longibrachiatum	China General Microbiological Culture Collection Center	_	7	Potato dextrose medium	Ligninase, cellulases, chitinolytic, and pectic enzymes	[3]
Aeromonas caviae, Shinella sp., Rhizobium sp., Corynebacterium pseudotuberculosis, Streptomyces clavuligerus.	Rice straw samples of composting in high temperature period	45	4–5	Gaoshi No.1 medium	Cellulase activity	[17]
Bacillus licheniformis, Aspergillus nidulans, Aspergillus oryzae	Dairy manure and sugarcane leaves compost samples	50	3–5	Luria Bertani medium and potato dextrose medium	Cellulosic enzymatic	[21]
Bacillus halmapalus X-2	Cattle manure compost	55	—	Ammonia-oxidizing bacterial seed medium	—	[34]
Streptomyces sp. and Actinobacteria bacterium	Organic waste compost from different sources	60	5	Gaoshi No.1 medium	Cellulose activity	[35]
A. caviae, Shinella sp., Rhizobium sp., C. pseudotuberculosis, S. clavuligerus.	Rice straw compost samples from the high-temperature period	45	4–5	Gaoshi No.1 medium	Cellulase activity	[36]
Gloeophyllum trabeum	Forest litter	—	7	Potato dextrose medium	Oxidase	[37]
Lactobacillus amylophilus, Geobacillus thermoleovorans, and Bacillus subtilis	China Center of Industrial Culture Collection	—	1–2	De Man Rogosa Sharpe medium and Luria Bertani medium	Cellulase	[38]

Table 1.

Overview of the suitable microbial inoculants for improving compost humification.

“_”: Not given.

3. Inoculation of microbial inoculants: types, amounts, and time

The inherent recalcitrant nature of raw materials hinders the degradation and transformation of organic fractions, leading to a lower composting efficiency [36]. The addition of microbial inoculants can increase microbial activity, accelerate organic matter degradation, release more precursors, and increase the humification degree of compost products [34, 35, 40]. The type, amount, and inoculation time of microbial inoculants [9, 10, 35, 38, 41, 42] have significant effects on the formation of HS during composting. Thus, it is essential to fully explore the correct use of microbial inoculants to improve the HA levels.

The types of microorganisms used for promoting compost humification are bacteria, fungi, and actinomycetes (Table 1). Of these, Bacillus sp [34, 38]., Phanerochaete chrysosporium [3], and Streptomyces sp. [17, 35, 36], can considerably accelerate HS formation. In this regard, actinomycetes have more desirable features than bacteria and fungi, such as thermo-tolerance, adaptability to harsh environments, higher amounts of hydrolytic enzymes that degrade lignocellulose, and a more pronounced response to genetic modification [35, 43]. The formation of HS mainly occurs during the curing period [37], during which actinomycetes are extremely abundant, making them ideal candidates for compost humification bioaugmentation.

The inoculum amount is also a significant factor affecting the humification process during composting [44]. Based on previous findings, inoculation at the level of 4% (in dry weight) is more conducive to the enhancement of HA than that at the level of 2% [9].

Given that various multifunctional microbe populations at different periods drive the composting process [3], it may be justified to inoculate specific microbial inoculants during different periods to reduce competition among microbes [12] and therefore accelerate the humification process. Some studies showed that composting can be considerably promoted by inoculation during the cooling period [35, 36, 37, 38, 39, 40, 41, 42, 43]. The multistage inoculation of composite microbial inoculants during the entire composting process also significantly increased the HA level [10, 38, 42]. Accordingly, the addition of microbial inoculants during the cooling period may be a feasible way for improving humification, especially the inoculation of actinomycetes at the level of 4% or higher.

4. Increasing the compost HA levels by adding microbial inoculants

The increase in the HA level indicates the increase in compost humification. As shown in Table 2, the addition of microbial inoculants can considerably promote compost humification and increases the content of HA. Different microbial inoculants have different effects on the HA content at the same inoculum level. For example, at a level of 1%, the inoculation of protein-, starch-, oil-, and lignocellulose-degrading microbes as well as ammonia-oxidating bacteria resulted in a more significant increase in the HA content, with levels being 216.88% higher compared to those of the control [40]. At an inoculation level of 5%, Bacillus clarkii and B. halmapalus significantly increased the compost HA content by 27.58% compared to the control in the composting of wheat straw and cattle manure [34]. As a lower inoculation amount generally results in a greater potential increase in HAs, the development of specific microbial inoculants is more important than the optimization of inoculation amounts in the improvement of the HA content. In the following section, the promotion of biotic processes by microbial inoculants is discussed.

Composting materials	Main inoculant types	Inoculation time	Dosage	Effects on HA content	Reference
Chicken manure biogas residue, spent mushroom straw, and rice straw	Phanerochaete chrysosporium and Trichoderma longibrachiatum	Day 0	5% (w/w, in fresh weight)	Increase by 23.6%	[3]
Medicinal herbal residues	B. subtilis, Aspergillus niger, Myceliophthora thermophila, Saccharomyces cerevisiae, Streptomyces pratensis, and S. violascens	Days 0, 25, and 55, respectively	0.1% (w/w, in dry weight)	Increase by 12.44%	[10]
Wheat straw and fresh cattle manure	B. clarkii and B. halmapalus	Day 0	5% (v/w, in fresh weight)	Increase by 27.58%	[34]
Pig manure and wheat straw	Gloeophyllum trabeum	Prior to composting	—	Increase by 17.2%	[37]
Biogas residue, sawdust, and food waste	Lactobacillus amylophilus, Geobacillus thermoleovorans, and B. subtilis	Days 0, 2, and 14, respectively	0.5% (w/w, in fresh weight)	Increase by 77.78% in humus index	[38]
Food waste and sawdust	Protein-, starch-, oil-, and lignocellulose-degrading microbes, ammonia-oxidating bacteria	Day 0	1% (w/w, in fresh weight)	Increase by 216.88%	[40]
Corn straw dairy manure	Streptomyces sp. and Actinobacteria	Day 6	2% (w/w, in dry weight)	Increase by 18.75%	[43]
Fresh swine manure and naturally dried corn straw	Acinetobacter pittii, B. subtilis subsp. stercoris, and Bacillus altitudinis	Day 0	1% (v/w, in fresh weight)	Increase by 5.92%	[45]
Fresh cattle manure and dry corn straw	B. subtilis, Bacillus licheniformis, B. cereus, and Streptomyces nogalater	Day 0	5% (v/w, in fresh weight)	Increase by 46.12%	[46]

Table 2.

Overview of the promotion effects of microbial inoculants on compost humification.

“_”: Not given.

5. Bioaugmentation mechanisms and factors related to an increase in compost HA by adding microbial inoculants

The formation of HS is a complex process [21]. Many hypotheses regarding the information of HS include the lignin-protein, phenol-protein, and sugar-amine condensation theories [47]. The core of these theories is that precursors are polymerized to form HS via biotic and abiotic ways [15]. In addition to regulating environmental factors and improving the native microbial community in compost, inoculating exogenous microbial inoculants to increase the humification degree of compost products is the main biological method [34]. Due to the complex interactions among microorganisms, advanced mathematical statistics are needed to accurately reflect the ecological interactions of the humification-associated community in the composting environment. Recent studies revealed the bioaugmentation mechanisms and influencing factors of compost humification, which are summarized in Figure 3.

Figure 3.

Conceptual framework summarizing the bioaugmentation mechanisms for the application of microbial inoculants to composting systems. RS, reducing sugars; AAs, amino acids; PP, polyphenol; TCA, tricarboxylic acid.

The variations in precursor types and concentrations during composting are related to the efficiency of humus synthesis. In addition, the humification process includes the production of precursors and the polymerization of HS, both of which occur sequentially during the whole composting process [17, 36, 47]. Therefore, the deep resolution of bioaugmentation mechanisms about microbial inoculants to composting relies on elucidating the dynamic changes in the precursors, microbial activity, and community structure during different composting periods [48]. Many studies have shown that the inoculation of exogenous microbial inoculants can significantly facilitate the enhancement of functional microbial activity during the warming and thermophilic periods of the composting process, accelerate the degradation of cellulose, proteins, and carbohydrates, and release large amounts of precursors [3, 18, 34, 37, 40, 46]. During these periods, the richness and diversity of the microbial community are markedly increased by the addition of microbial inoculants [21]. Moreover, Firmicutes, involved in the degradation of available organic components, are enriched [45]. The higher relative abundances of lignocellulose-degrading microbes (laccase producing microbes, Chloroflexi, Actinomycetes, fungi, and Luteimonas) go along with a decline in pile temperature during the cooling and maturation periods [3, 37, 38, 43], with the synergistic decomposition of lignin [17, 49]. The effective degradation of lignin can provide more carbon skeletons for humus formation [48]. Additionally, microbial inoculants can also considerably reduce the level of carbon metabolism, which is linked to the tricarboxylic acid cycle [44], and stimulate the microbial assimilation of precursors (polyphenols and quinones) [16, 50]. These processes facilitate HA formation and improve carbon sequestration, resulting in a high-value compost.

Generally, HA and HS formation is affected by environmental factors [46]. Specifically, the suitable C/N ratio and the total organic carbon (TOC) level can increase the richness and activity of microbes, resulting in the accumulation of soluble sugar and amino acids in the early period of composting [36]. The increase in the pH and the decline in the TOC results in the enhanced microbial synthesis of HA at the later period of the composting process [17, 34, 36, 40]. Thus, controlling the environmental factors and functional microbes can accelerate the formation of HAs based on the addition of microbial inoculants.

6. Future perspectives

As described above, the addition of microbial inoculants promotes solid organic waste decomposition, stimulates precursor production, increases the HA content, and alleviates several drawbacks of noninoculated composting systems. However, there are still some uncertainties. We highlight some perspectives for the further exploration of the use of functional microbial inoculants:

1. The instability effect is one of the main restrictions in the promotion of microbial inoculant use. Some investigations have confirmed the positive effects of microbial inoculants on humification only at a suitable inoculation level [44]. Further works will have to focus on the inoculation levels for functional microbes used to promote humus formation and related mechanisms, especially regarding the interactions among microbial inoculants, humification, and compost microbiota during composting.

2. Bioaugmentation using multistage inoculation has been well estimated in small-scale composting systems [10]. However, to guide real production, the effectiveness of multistage inoculation remains to be explored, especially regarding the effects of multistage inoculation on humus formation in large-scale composting systems [51]. Efforts will also be made to identify the carbon and nitrogen cycle functional genes related to HA production pathways, using novel analytical methods.

3. Microbial inoculants are generally cheaper than other compost additives. However, the acquisition of high-efficiency functional strains requires more time. In the near future, in situ selection and omics methods should be used to rapidly develop simple and stable compositive microbial inoculants (including yeast) that can not only adapt to the native compost environment but also degrade novel pollutants.

7. Conclusions

The addition of microbial inoculants to various solid wastes is a potential method for improving the humification process and increasing carbon sequestration. However, further research is needed to gain a deeper understanding of the biotic mechanisms underlying the microbial inoculants’ effect on composting and to explore the relationships among functional microbes, the humification process, and the compost microbiota in large-scale composting systems. A comparison of the influences of different inoculation methods on composting processes and compost quality is also needed. Moreover, the development of multifunctional strains and the optimization of inoculation amounts would further justify the use of microbial inoculants in the compost industry.

Acknowledgments

This work was supported jointly by the research and development project of company’s inner key technologies (Optimization of the sequestration capacity of beneficial microbes on organic fertilizer medium). The authors would like to express their gratitude to MogoEdit (https://www.mogoedit.com/) for the expert linguistic services provided.

Conflict of interest

The authors declare no conflict of interest.