Arbuscular mycorrhizal fungal (AMF) inoculants are sustainable biological materials that can provide several benefits to plants, especially in disturbed agroecosystems and in the context of phytomanagement interventions.
1. Introduction
Mitigating the environmental impacts of intensive agriculture, such as greenhouse gas emission, eutrophication, the pollution of surface and underground water, global soil loss to salinity and compaction, loss of microbial diversity, etc., calls for the integration of multiple sustainable management approaches including the diversification of crop rotations, intercropping, integrated farm management, conservation, precision agriculture, and the ‘4R’ framework of nutrient management (meaning applying the right type and quantity of nutrient in right place and at the right time). Another important strategy that can help to reduce the environmental footprint of agriculture, restore soil health, and protect biodiversity is the introduction of beneficial microorganisms into agroecosystems
[1][2].
Arbuscular mycorrhizal fungi (AMFs) are essential soil microbial communities that form obligate symbiosis with 80% of terrestrial plants
[3]. AMF promotes plant growth by facilitating nutrient acquisition through the extraradical mycelia that spread from the host’s roots into surrounding soils
[4][5]. By increasing plant access to nutrients, AMF application can offset phosphorus fertilizer demand by ~50%
[6]. A growing number of studies also indicate that the application of AMF can reduce nitrogen losses in the form of nitrous oxide emission and nitrate leaching
[7][8][9][10]. AMF can also stimulate the production of phytohormones and secondary metabolites, that can improve plant productivity
[11], crop quality
[12][13], and build resilience against environmental stress such as salinity
[14], drought
[15][16], heat, and pathogens
[17][18]. Thus, intervention with commercial AMF is tenable in soils with high P-fixing potential, anthropized sites that exhibit low diversity and richness of native AMF species, and arid and semi-arid areas
[19][20][21][22][23][24].
The AMF inoculant market is burgeoning and gaining wide recognition, although commercial products are majorly based on a few AMF strains belonging to the
Glomeraceae genera:
Rhizophagus,
Funneliformis, and
Claroideoglomus strains
[2]. AMF inoculants consisting of several AMF strains can produce additive or synergistic effects on target crops
[25], however increasing the diversity of AMF in inoculants may fail to produce additional benefits if constituent strains are redundant i.e., performing similar functions
[26]. Similarly, the concerted application of AMF with other bioinoculants such as growth-promoting rhizobacteria (PGPR) or nitrogen-fixing bacteria, and organic substances such as humic acid can produce greater plant response
[27][28]; however, only a few commercial products are based on such complex formulation
[2].
Despite the growing demand for biofertilizers, results from commercial AMF inoculants have been largely context-dependent, especially under field conditions, contrary to the common laboratory successes
[29][30][31][32]. The inconsistent narrations from the research community regarding the reliability of AMF inoculation as a valid agricultural management technique
[33] are generating low consumer confidence that is hindering large-scale adoption of the technology. Although, quantifiable or marketable gains (such as yield enhancement, nutrient replacement, watering costs, seedling growth, and nursery raising) are the mostly sought benefits of AMF, agricultural benefits of AMF extend beyond immediate monetary gains. The non-marketable benefits of AMF include the improvement of food and fiber quality; plant stress mitigation as well as ecosystem services, such as soil erosion control; the prevention of nutrient losses (nitrous oxide emission and N leaching); carbon sequestration; and landscape recreation
[34][35]. Notwithstanding, to win growers’ confidence and market sustainability, investment in inoculant technology must pay off either in the short- or long-term.
Field success of commercial inoculants relies on the ability of the introduced strains to establish and persist at the target site for a desired or specified period. However, AMF establishment is complex and are often determined by the inherent features of the target agroecosystem. Unfortunately, most microbial inoculants are selected based on their expressed functional traits in a greenhouse, without proper consideration of the ecologically relevant traits that determine establishment and persistence under natural conditions
[36]. While many cases of inoculation failures could be attributed to poor product quality stemming from the lack of regulatory or quality control frameworks that mandate best practices, resulting in a market flooded by substandard products
[29][32][37], there is also lack of understanding of ecology and of the mode of action of inoculants
[38]. Duell et al.
[29] recently demonstrated that inoculants could decouple native plant symbiosis with indigenous strains in a natural ecosystem already containing a large diversity of AMF community, without conferring additional benefits. Thus, inoculation in such ecosystems with high mycorrhizal potential may prove unproductive.
Therefore, understanding the factors that influence the establishment and persistence of AMF inoculants will inform decisions, as well as the management practices necessary, to ensure inoculation success
[39]. Tracing the survival and persistence of the introduced strains will help identify which AMF strains adapt well to the local conditions and induce the observed plant response following long-term persistence and will help determine whether another inoculation exercise is necessary. Moreover, monitoring the downstream impacts of the introduced inoculant
[40] is necessary to prevent the intentional propagation of detrimental or ineffective strains and the elimination of keystone taxa
[41].
2. Establishment and Persistence of Introduced AMF in Field
2.1. Establishment
The influence of AMF on plants and microbial communities relies both on the successful colonization of plant roots and on the maintenance of live propagules that can form mycorrhizal association for a given period after their application
[42]. However, the establishment success of introduced AMF in the field is context-specific, varying according to the type and composition of the inoculant, as well as the physical and biotic conditions of the target field
[31]. The establishment of AMF is largely successful regardless of soil type, nutrient concentration, or the composition of the indigenous AMF communities (
Table 1). In other contexts, AMF may fail to establish at the target site due various factors. For example,
Rhizophagus irregularis (DAOM197198), often regarded as a generalist AMF with a cosmopolitan distribution, did not influence grapevine growth in a five-year trail
[43][44]. Renault et al., 2020
[45] also observed no difference in root colonization between control and non-inoculated plots of corn soybean, or in wheat treated with same isolate of
R. irregularis. In composite trials, where AMF inoculants were applied in different fields, establishment was successful at one site and failed at the other
[46][47].
AMF Name, Isolate (Brand, Manufacturer) |
Location of Trial |
Soil Type |
Climate |
Crop |
Persistence Tracking Period |
Change in Abundance of Introduced AMF over Time |
References |
Funneliformis mossaeae BEG12 and AZ225C, Rhizophagus irregularis BEG141 |
Manciano (Grosseto) Italy |
Haplic Calcisol or Inceptisol |
Humid Mediterranean |
Medicago sativa L. |
2 years |
F. mosseae AZ225C was most persistent, followed by BEG12 and BEG141. Abundance of local F. mosseae cluster was reduced, while the abundance of native R. irregularis increased. |
[45] |
Rhizophagus Irregularis, DAOM 197198 (Myke Pro, Premier Tech) |
Canada (Swift Current) |
Brown Chernozem |
Semi-arid |
Pea-wheat |
2 years, 5 months |
Inoculant decreased from 27% in first season to 15% in years two and three. |
[41] |
Canada (Outlook) |
Dark Brown Chernozem |
Semi-arid |
|
2 years, 5 months |
Introduced AMF decreased from 17% in year one to 4% in years two and three. |
Canada (Scott) |
Chernozem |
Sub-humid |
|
3 months |
Relative abundance of introduced AMF was 33% but detection failed in subsequent years. |
Canada (Melfort) |
Black Chernozem |
Sub-humid |
|
1 year, 3 months |
AMF detected in year one with 10% abundance and 4% in year two but declined completely in year three. |
Rhizophagus irregulare, DAOM 197198 (Myke Pro GR, Premier Tech) |
Swift Current, Saskatchewan |
_ |
_ |
Lentil Lens culinaris, Linum usitatissimum (Flax var. Bethune) rotation inoculation year 1 only |
_ |
Inoculation did not affect abundance of target AMF in roots. |
[46] |
Beaverlodge, Alberta |
_ |
_ |
Pisum sativum, Linum usitatissimum (Flax var. Bethune) |
_ |
Inoculation did not affect abundance of target AMF in roots. |
Melfort, Saskatchewan |
_ |
_ |
|
_ |
Abundance of introduced isolate was less in inoculated site than control |
STR Saskatchewan |
_ |
_ |
|
2 years |
Abundance of introduced isolate was higher in inoculated plots in year one, but did not differ in year two. |
Rhizophagus Irregularis IR27 |
Senegal |
Tropical ferruginous |
Semi-arid |
Jujube (Ziziphus mauritiana lam., Tasset and Gola) |
1 years, 5 months |
Abundance of R. irregularis was low (15% after 18 months). |
[47] |
Funneliformis Mosseae AZ225C and IMA1 |
Pisa, Italy |
Sandy loam |
Mediterranea climate |
M. sativa |
2 years |
2 years, relative proportion of F. mosseae decreased in favor of native species, abundance of isolate AZ225C dropped from 100% to 16.3%, while isolate IMA1 survived only three months. |
[48] |
Glomus sp., G. intraradices and a mixture of both |
Vicente Banes, Molina de Segura, Southeastern Spain) |
Typic Torriorthent (silty clay) |
Semi-arid Mediteeranean climate |
O. europaea |
1 year, 2 months |
Abundance varied by AMF species: Glomus sp.: 20%, G. intraradices: 48.2%, and mixed (G. intraradices: 14%, Glomus sp.: 39.7%). |
[49] |
G. intraradices IMA6 and F. mosseae AZ 225C, and mixture of both |
Italy |
|
Mediterranean climate |
Artichoke (Cynara cardunculus L. var. scolymus F.) |
3 months |
Inoculation increased the abundance of Glomus OTUs in inoculated plants compared to control. |
[12] |
Gigaspora margarita CK (cerakinkong, Central glass Co., Tokyo, Japan) |
Mizunashi River, Mt Fugendake, Nagasaki Prefecture, Japan |
Reforested soil |
|
Eragrosis curvula (weeping love grass) and Miscanthus sinensis (Japanese Pampas grass) |
4 years |
G. margariata isolate CK was detected in rhizosphere and root of E. curvula. |
[50] |
Funneliformis (syn. Glomus) mosseae BEG12, 167 G. intraradices BEG 141 (IBG), G. eutenicatum BEG 168, (Endol, Biorize) |
Daxing, Hebei Province, China |
- |
|
Sweet potato, (Ipomoea batatas L.) |
3 months |
Contaminating AMF, G eutenicatum had longer persistence than G. mosseae, the establishment of which was not successful in year 2; similarly, low amounts of F. mosseae BEG 167 were detected compared to contaminating G. eutenicatum. |
[51] |
R. irregularis GEG140, F. mosseae BEG95, C. claroideum BEG96, (Symbiom) |
Coal mine spoil bank, Mekur, North Bohemia, Czech Republic |
_ |
_ |
Phalaris arundinacea |
3 years |
Introduced AMF persisted and co-existed with native strains. |
[52] |
Surprisingly, there seems to be greater discrepancies among commercial and laboratory inoculants tested in the same field. In 2017, Berruti et al.
[48] reported that a commercial inoculant failed to establish in the field, while other studies obtained inconsistent results, where some commercial inoculants survived better under greenhouse conditions
[30][31][32]. A recent global evaluation of commercial AMF inoculants in greenhouse and field conditions demonstrated that only 4 out of 28 AMF inoculants were successfully established in a greenhouse experiment, and only one successfully influenced plant performance in the field, whereas the inoculum obtained from the laboratory established successfully in both ecosystems
[32].
2.2. Persistence
Most studies indicate that AMF could survive for a long period after the first inoculation. More specifically, introduced AMF could survive for relatively few months after inoculation to several years (
Table 1); however, no study has monitored the persistence of AMF in the field beyond four years, according to the information available while gathering data for this study. The survival limit, as well as the abundance of the introduce species, is majorly affected by the type of AMF as well as the biotic and abiotic conditions of the target sites. For example, out of the two non-native
Funneliformis mosseae isolates—AZ2256C and IMA1, which originated from the USA and the UK, respectively—only the former was detected two years after inoculation, but in lower proportions compared to the native
R. irregularis and
R. intraradices, despite early establishment success and the stimulatory effects of both strains on the roots of
Medicago sativa [49]. In a recent study, Pellegrino et al.
[50] also reported that although the AMF strains
F. mossaeae BEG12 and AZ225C were detected in alfalfa roots two years post-treatment, the AZ225C strains had longer persistence.
Does persisting inoculant consistently improve plant growth, as in the initial stage of its introduction? Results from a 2012 study by Pellegrino et al.
[51] suggest that persistent AMF could sustain yields two years after inoculation. A recent study by Pellegrino et al.
[50] also indicated that AMF could sustain positive effects on the host crop several months after inoculation, whereby both
F. mossaeae BEG12 and AZ225C enhanced alfalfa yield, nutrients, and fatty acid content. Similar findings were reported by Thioye et al.
[52] where the growth-promoting effects of R. irregularis IR27 on Ziziphus mauritiana Lam continued 18 months after planting. However, Farmer et al.
[53] and Alguacil et al.
[54] could not correlate the persistence of inoculants with plant growth, even though inoculation increased plant growth in both studies.
Moreover, most studies reported a decline in the abundance of the persistent strains over time due to competition and suppression by native communities. For example,
R. irregularis MUCL 41833 DNA was detected at a concentration 100 to 1000 times lower than native
R. irregularis strains in three potato cultivars grown in the field in Belgium
[55]. In a recent trial, the levels of the AMF abundance of inoculates introduced in four Canadian agricultural fields varied between 3 and 27 months
[41].
Rhizophagus irregularis IR27 represented 11 to 15% of the
Rhizophagus genus in
Ziziphus mauritiana roots after 13 and 18 months
[56]. A low abundance of introduced AMF compared to native strains indicates that exotic AMF can co-exist with native strains without posing a negative threat to local biodiversity. Lastly, the results obtained by Kokkoris et al., 2019
[57] also demonstrated that the establishment and persistence of AMF was site-dependent and not related to crop management.