Crop losses due to plant diseases and pests are a problem worldwide, with average yield losses ranging from 17 to 30% in the five major food crops (i.e., wheat, rice, maize, potato, and soybean) (1
). Preventative measures are the most effect way to protect a crop against soilborne pathogens. However, some of these measures (e.g., the use of chemical soil fumigants) are banned or restricted due to environmental hazards. Creating disease-suppressive soils can be an effective strategy against multiple diseases (3
), and the addition of organic materials to the soil is one of the methods found to induce such suppressiveness (5
). In addition, organic amendments provide essential nutritional elements and can improve other soil quality parameters. As a result of the European Waste Framework Directive (Council Directive 2008/98/EC), more organic-rich waste products are promoted for agricultural use, which helps contribute to a circular economy (7
). Nowadays, a wide range of organic amendments with the potential to support plant health and improve nutrient use efficiency can be applied in agriculture (9
Compost is the most commonly used organic amendment used to create enhanced soil suppressiveness against soilborne diseases (10–12
), but it yields variable results, and the effects are case specific (5
). Chitin, a by-product from the seafood industry (crab, lobster, and shrimp shells), has been used since the 1970s to reduce damage by plant-parasitic nematodes (13–15
) and fungal pathogens (15–18
). Other organic waste products have been applied to control plant parasitic nematodes, such as oil cakes derived from several plant species, cellulosic waste, sugarcane bagasse, bone and horn meal, sewage sludge, manure, and various crop residues (19
). Similarly, several plant- and animal-derived materials have been tested to reduce the detrimental effect of soilborne plant diseases (12
). Although disease reduction has been demonstrated with several types of organic waste products, the effects are often inconsistent, and the level of control depends on the product, type of soil, plant-pathogen combination, and cultivation system (5
). Moreover, disease suppression via organic amendments operates through various mechanisms (6
). However, most studies to date have focused on the organic amendment suppression mechanisms related to chemical structure and decomposition rate, rather than on effects mediated through the soil microbial community.
To better predict the effect of organic amendments on soil suppressiveness, the physicochemical properties of the organic products, as well as their impact on soil properties such as their microbial community, are being studied. For example, organic products have been characterized in terms of their chemical composition, pH, C/N ratio, and decomposition rate (9
). Other studies have assessed chemical, physical, and/or biological soil properties after being amended with organic products to determine possible links to disease suppression (24
). The use of organic amendments can affect soil microbial communities (26–28
), and several recent studies have demonstrated correlations between soil microbial community composition and soil suppressiveness against plant pathogens (18
). Therefore, integration of organic amendments and beneficial soil microorganisms might be the key for controlling soilborne pathogens (6
The development of a tailor-made pipeline to select the best organic amendment and/or microbial consortium treatments to increase suppressiveness against a specific pathogen is of great interest to farmers. To achieve this, further insight into the mechanisms behind pathogen suppression in soil is needed. In order to better understand the mechanisms behind the stimulation of soil suppressiveness via the addition of organic products and which specific factors can be traced as disease suppressiveness markers, the following questions must be answered. (i) Which types of organic soil amendments have the potential to stimulate disease suppressiveness? (ii) Can the stimulation of disease suppression by organic amendments be ascribed to enrichment of specific microbial groups? (iii) Does enhanced disease suppression correlate with biological and physicochemical soil properties?
To address these questions, 10 organic products with different characteristics (i.e., variation in C/N ratio and decomposition rate) were selected to assess their potential to enhance disease suppression in soil. After two different soils from arable fields were amended with the products, pot experiments were performed to assess soil suppressiveness against the fungal pathogen Rhizoctonia solani. In addition, several soil characteristics were analyzed, as well as bacterial and fungal community composition and how microorganisms interact within these communities, to better understand the effect of the organic amendments on creating disease-suppressive soils.
We found that chitin- and keratin-rich soil amendments play an important role in R. solani disease suppression. Furthermore, we showed that in soil treated with chitin- and keratin-rich amendments, the microbial community composition was very different from those of the nonsuppressive soils. Specifically, chitin- and keratin-rich amended soils were enriched in Oxalobacteraceae and Mortierelleceae species abundance. In those treatments, soil microbial communities related to disease suppression were positively correlated with zinc and copper, although the concentrations of these elements did not vary significantly across treatments. On the other hand, calcium and selenium concentrations were higher in the suppressive treatments. Based on these results, it might be possible to use specific microbial taxonomic groups and/or available calcium and selenium compounds as indicators for R. solani suppression in soils or as soil supplements to boost R. solani disease suppression.
We aimed to find explanatory factors for soil suppressiveness by using organic products with a wide range of properties and effects on disease suppression. A positive effect of keratin- and chitin-rich soil amendments on R. solani
disease suppression was demonstrated in the current experiments, as well as in previous research (21
). Suppressiveness was not induced by the nitrogen in these products, since the addition of mineral nitrogen [Ca(NO3
] alone did not increase soil disease suppressiveness. Furthermore, the possibility that nitrogen in the organic products may be converted to other forms (e.g., NH3
) that are toxic to some microbial species (30
) was discarded.
We found that the microbial communities in all amended soils were different, with these differences becoming more apparent when treatments were grouped by amendment type (i.e., plant, manure, keratin, manure-chitin, seed). However, in some instances these differences in soil microbial community composition correlated with disease suppressiveness, and at other times they did not. It has been shown that chitin-enriched compost improves soil suppressive capacity against R. solani
), where chitin- and keratin-degrading fungi (i.e., Trichoderma
) are more abundant and act as active antagonists of R. solani
. Furthermore, keratin-based products have shown plant growth promotion via increased activity of keratinolytic bacteria, which can act as antagonists to plant pathogens (32
). The keratin- and chitin-rich amendments in this study were enriched in chitinolytic/keratinolytic fungi such as Cutaneotrichosporum
(Onygenales), and Mucor
(Mucorales). Although chitin and keratin are very different molecules (a glucosamine polymer and a protein, respectively), their degradation can be performed by the same microorganisms, as keratinolytic fungi are able to produce other hydrolytic enzymes such as chitinases (34
). It can be hypothesized that the presence of high levels of keratin (as well as chitin) promotes the presence of both keratinolytic and chitinolytic fungi. The most abundant group found in the keratin-amended soils is Mortierellales. It has been reported that Mortierella
, and Mucor
species have a chitinolytic activity (35
). Moreover, Mortierella
spp. have been reported for their capacity to degrade other toxic and recalcitrant compounds, to accumulate heavy metals such as zinc, and to induce the production of plant hormones such as indole-3-acetic acid (IAA) and abscisic acid (ABA), as well as antibiotics (36–38
Some soil bacteria are able to degrade both chitin and keratin (35
), and some of them were more abundant in keratin- and chitin-rich amended soils in the present study. Bacteria such as Flavobacterium
), and Rhizobium
), which were abundant in chitin-rich samples, have been shown to have pathogen (e.g., R. solani
) suppression effects (40
) were the dominant species in chitin-rich amended soils, and they were less abundant in soils treated with plant-derived amendments, which has also been observed in other studies (42
). These species are an important component of the rhizosphere microbiota and they can thrive on the seed coat, radicle, and roots of plants, as well as on the hyphae of the pathogen Pythium
, hence reducing its pathogenicity. Furthermore, Massilia
is a well-known chitinolytic bacterium that is able to degrade keratin (43
). Because of its keratin degradation property, it is likely that it can thrive in keratin-rich environments and is able to reduce the growth of fungal pathogens as well.
In the present work, fungal and bacterial communities showed strong correlations. In R. solani
-suppressive amended soils (keratin- and chitin-rich), there were clear interactions between fungal and bacterial soil communities. The abundance of Mortierella
spp. negatively correlated with several bacterial taxa, among them an uncultured Alcaligenaceae
taxon. Some bacteria belonging to this taxon (e.g., Alcaligenes
spp.) are keratinase-producing bacteria (45
One remarkable observation is that microbial diversity (inverse Simpson and Shannon indices) was lower in all amended soils and that the disease-suppressive treatments were among the soils with the lowest diversity indices, together with the seed-based amendment treatment (Terrafit-Biofum). Other microbial ecology studies support the importance of having a highly diverse microbial community to help create a better ecosystem and resilience (46
). In the present study, we measured the short-term response (after 3 weeks) of the physicochemical soil properties and found that they were drastically changed by the amended products. This presumably allowed more opportunistic, fast-growing microbes to proliferate, as they are more adapted to carbon-enriched environments. It has been shown that disturbance reduces the microbial Shannon diversity index, as well as microbial community functions in agricultural environments (47
). A highly diverse environment (in this case, the soil before treatment) is essential to provide the soil system with the plasticity to better respond against pathogens and/or other inputs. Future research is needed to focus on how long these changes in the microbial communities will persist.
Keratin- and chitin-rich amended soils had very distinct bacterial and fungal microbial communities compared to the other treatments. Furthermore, these microbial communities were linked to specific soil chemical properties (zinc and copper). Another factor, potential mineralizable nitrogen (PMN), which usually positively correlates with total microbial biomass (48
), positively correlated with R. solani
suppression. However, PMN was not associated with the microbial community composition in amended, suppressive soils. On the other hand, total fungal biomass was higher only in the chitin-rich amended soils that were colonized by A. bisporus
. Selenium compounds are accumulated in A. bisporus
hyphae and fruiting bodies (49
), which explains why selenium concentrations were highly positively correlated with the microbial communities in the chitin-rich treatment. Although zinc levels were not significantly different between treatments, zinc was positively correlated with microbial community structure in keratin-rich treatments. Therefore, it is suspected that positive correlations between zinc and disease suppression are the result of the ability of zinc to enhance specific microbial groups that reduce disease. This supports findings that zinc-enriched products have been shown to have a fungicidal effect on several soilborne pathogens (51
). Microbial keratin decomposition can be compared with that of other recalcitrant polymers such as cellulose and chitin. A specialized family of metalloenzymes that cleave cellulose and chitin, lytic polysaccharide monooxygenases (LPMOs), are produced by keratin-degrading fungi as well (e.g., Trichoderma
]) and contain a metal binding site that can oxidize cellulose, hemicellulose, and chitin (54
). We hypothesize that the observed higher zinc levels in keratin treatments might be due to higher production of specialized LPMOs containing a zinc binding site by keratinolytic microorganisms.
Bacterial taxa that have been correlated with R. solani
disease suppression in former studies (29
), such as Massilia
spp., and Cutaneotrichosporon
spp., could be used together with zinc and copper as markers for disease suppression. In addition, chitin-rich amendments have been more thoroughly studied, and their effects on soil quality and abiotic/biotic plant stress tolerance observed here are supported by past research (35
). Keratin-based amendments can likewise boost the plant-growth-promoting effect of symbiotic microorganisms such as Bacillus subtilis
Despite the advances in knowledge about using soil amendments to enhance disease suppression presented in this study, still there are open questions. For example, is disease suppression due to enhancement of specific microbial functions? How long do changes to the soil microbial community composition and soil chemical properties last? Metatranscriptomic analyses could help to gain insight into the molecular functions behind the effects of amendments on R. solani disease spread by, for example, looking into the production and function of specialized enzymes that degrade chitin and keratin (e.g., LPMOs). Finally, the data presented here were collected at one time point. However, to better understand the duration of the effects of treatments on disease suppression, a time line study is needed, in which microbial populations and functions are tracked together with levels of disease suppression across multiple time points.
We showed that soil amendments rich in keratin and chitin are able to reduce R. solani disease in sugar beet. Furthermore, the microbial community is distinct in soils treated with different types of amendments (plant, seed, manure, keratin, and manure-chitin) and nonamended soils. Available forms of zinc and copper were related to soil microbial communities and disease suppression. We demonstrated that soil microorganisms likely play an important role in disease suppression in soils. In the case of arable soils, it is possible to steer the soil microbial community by applying various organic amendments that can boost specific biological and chemical soil properties that increase plant and soil resilience and/or disease suppression.