Microbial Nursery Production of High-Quality Biological Soil Crust Biomass for Restoration of Degraded Dryland Soils

Abstract

Biological soil crusts (biocrusts) are slow-growing, phototroph-based microbial assemblages that develop on the topsoils of drylands. Biocrusts help maintain soil fertility and reduce erosion. Because their loss through human activities has negative ecological and environmental health consequences, biocrust restoration is of interest. Active soil inoculation with biocrust microorganisms can be an important tool in this endeavor. We present a culture-independent, two-step process to grow multispecies biocrusts in open greenhouse nursery facilities, based on the inoculation of local soils with local biocrust remnants and incubation under seminatural conditions that maintain the essence of the habitat but lessen its harshness. In each of four U.S. Southwest sites, we tested and deployed combinations of factors that maximized growth (gauged as chlorophyll a content) while minimizing microbial community shifts (assessed by 16S rRNA sequencing and bioinformatics), particularly for crust-forming cyanobacteria. Generally, doubling the frequency of natural wetting events, a 60% reduction in sunlight, and inoculation by slurry were optimal. Nutrient addition effects were site specific. In 4 months, our approach yielded crusts of high inoculum quality reared on local soil exposed to locally matched climates, acclimated to desiccation, and containing communities minimally shifted in composition from local ones. Our inoculum contained abundant crust-forming cyanobacteria and no significant numbers of allochthonous phototrophs, and it was sufficient to treat ca. 6,000 m² of degraded dryland soils at 1 to 5% of the typical crust biomass concentration, having started from a natural crust remnant as small as 6 to 30 cm² IMPORTANCE: Soil surface crusts can protect dryland soils from erosion, but they are often negatively impacted by human activities. Their degradation causes a loss of fertility, increased production of fugitive dust and intensity of dust storms with associated traffic problems, and provokes general public health hazards. Our results constitute an advance in the quest to actively restore biological soil covers by providing a means to obtain high-quality inoculum within a reasonable time (a few months), thereby allowing land managers to recover essential, but damaged, ecosystem services in a sustainable, self-perpetuating way as provided by biocrust communities.

Keywords: 16S rRNA; biological soil crusts; cyanobacteria; degraded soils; drylands; erosion control; soil microbiome; soil restoration.

FIG 1

Boxplots for the final phototrophic biomass (as aerial chl a content) obtained after greenhouse incubation of native soils from 4 sites (each panel shows a site) inoculated with natural biocrusts from their respective site under 18 different treatments. Boxes denote lower and upper quartiles (with median values depicted as black solid lines), and whiskers denote lower and upper extremes (n = 3). Blue lines indicate the chl a content of field biocrust samples used as inoculum (INOC), and red lines indicate initial chl a content in the inoculated soils (INIT) (color solid lines indicate mean, and color dashed lines indicate standard deviations of n = 3).

FIG 2

Endpoint bacterial community composition by phylum for each of the treatments in the fractional factorial experiments. Each panel corresponds to a different site. Data are averages of three independent determinations (biological replicates). Also included are the community composition determined for the biocrust samples used as inoculum (INOC; n = 3, technical replicates).

FIG 3

Endpoint cyanobacterial community composition by major clades for each of the treatments in the fractional factorial experiments. Each panel corresponds to a different site. Data are averages of three independent determinations (biological replicates). Also included are the community composition determined for the biocrust samples used as inoculum (INOC; n = 3, technical replicates).

FIG 4

Boxplots showing aerial chl a contents at the end of the greenhouse incubation period for the second set of experiments. Boxes denote lower and upper quartiles (with median values depicted as black solid lines), and whiskers denote lower and upper extremes (n = 5). Blue lines indicate the chl a content of field biocrust samples used as inoculum (INOC), and red lines indicate the initial chl a content in the inoculated soils (INIT) (color solid lines indicate mean, and color dashed lines indicate standard deviations of n = 5) (FB, Fort Bliss; J, Jornada; N, Nosecone; BB, Burr Buttercup).

FIG 5

Endpoint bacterial community composition by phylum at the end of the greenhouse incubation period for the second set of experiments that were optimized according to previous results versus that obtained for the field biocrusts used as inoculum. Each panel corresponds to one site. Data are averages from five technical replicates (INOC, inoculum biocrust samples) or five biological replicates (cultivated biocrusts) (FB, Fort Bliss; J, Jornada; N, Nosecone; BB, Burr Buttercup).

FIG 6

Endpoint cyanobacterial community composition by clades at the end of the greenhouse incubation period for the second set of experiments that were optimized according to previous results versus that obtained for the field biocrusts used as inoculum. Each panel corresponds to one site. Data are averages from five technical replicates (INOC, inoculum biocrust samples) or five biological replicates (cultivated biocrusts) (FB, Fort Bliss; J, Jornada; N, Nosecone; BB, Burr Buttercup).

FIG 7

General aspect of the microbial nursery facilities. Initial fractional factorial experiment (first phase experiments) at the Arizona State University (ASU) greenhouse (A), with a detailed view of Fort Bliss (B) and Jornada (C) soil incubations; plastic containers are 15 by 15 cm, and greenhouse benches are 2.74 by 0.91 m. Large-scale incubations (second phase experiments) in the Northern Arizona University (NAU) nursery (D), with top views of the final biocrusts produced for Nosecone (E) and Burr Buttercup (F) samples; plastic containers are 86 by 14 cm.