OPERATIONAL MANUAL FOR BIOCRUST RESTORATION IN DRYLANDS Akasha Faist, Colin Tucker, Sasha C. Reed, Anita Antoninka, Matt Bowker, Nichole Barger, Kara Dohrenwend, Natalie Day, Sue Bellagamba, Jayne Belnap, Michael Duniway, Stephen Fick, Ana Giraldo-Silva, Corey Nelson, Julie Bethany, Sergio Velasco-Ayuso, Ferran Garcia-Pichel Abstract The purpose of this manual is to synthesize current information about biological soil crust restoration for resource managers making decisions on the ground. Biological soil crusts (biocrusts) are communities of photosynthetic organisms (mosses, lichens, cyanobacteria, and/or microalgae) with accompanying populations of bacteria, archaea and fungi, that occupy the upper few millimeters of dryland or other sparsely vegetated soils across the globe. These communities play several fundamental roles in the functioning of ecosystems, being critical for soil stabilization and erosion prevention. Biocrusts may also affect vascular plant success, help dictate soil hydrology, and can be the dominant force for sustained fertility in some dryland soils. This manual is based on the experience gathered through work conducted as part of a Wildlife Conservation Society (WCS) project focused on elucidating the role of climate adaptation in biocrust restoration practice on the Colorado Plateau, as well as a Strategic Environmental Research and Development Program (SERDP) project assessing best practices for biocrust restoration carried out in the Chihuahuan and Great Basin deserts. The manual starts by introducing biocrusts, why biocrusts matter for ecological function of dryland ecosystems, and discussing the need for restoration of this important component of drylands in the face of anthropogenic pressures. The manual also explicitly addresses new options for biocrust restoration, including information about climate-adapted biocrust restoration, farm-grown biocrust inoculum sources, opportunities to acquire biocrust sources through salvage, and mechanizing the harvesting of biocrust. Methods for monitoring biocrust growth, development, and organismal composition are also provided. Finally, the operational manual ends with practical findings and potential areas that could benefit from further investigation as we progress in our understanding of best biocrust restoration practices for a changing world. Our understanding of biocrust restoration is rapidly improving, and this manual is expected to be a “living document” with improvements and updates made as our knowledge advances. Acknowledgements We would like to thank the Wildlife Conservation Society Climate Adaptation Fund, which was established with funds provided by the Doris Duke Charitable Foundation, for their support of the restoration work that informed this manual. We are also grateful to the Strategic Environmental Research and Development Program (SERDP) for their support of research included in the manual, and to the U.S. Geological Survey Ecosystems Mission Area. Finally, we thank the resource managers who continue to help facilitate the research and biocrust collections from the Canyonlands Research Center (Kristen Redd and Matt Redd), on the Jornada Experimental Range (John Anderson), Fort Bliss (John Kipp), and the Utah Test and Training Range (Russ Lawrence, Mike Shane, and Jace Taylor). We are also indebted to the large number of biological science technicians from the U.S. Geological Survey, Northern Arizona University, Arizona State University, and the University of Colorado, Boulder who were critically important in implementing the field and laboratory work that was the foundation for this manual. Cover photos: (upper left) Kristina Young, (all others) Anita Antoninka. OPERATIONAL MANUAL FOR BIOCRUST RESTORATION IN DRYLANDS Akasha Faist1, Colin Tucker2,3, Sasha C. Reed3, Anita Antoninka4, Matt Bowker4, Nichole Barger5, Kara Dohrenwend6, Natalie Day3,7, Sue Bellagamba8, Jayne Belnap3, Michael Duniway3, Stephen Fick3,5, Ana Giraldo-Silva9, Corey Nelson9, Julie Bethany9, Ferran Garcia-Pichel9, Sergio Velasco-Ayuso9 1 Department of Animal and Range Sciences, New Mexico State University, Las Cruces, NM 2 U.S. Forest Service, Northern Research Station, Houghton, MI 3 U.S. Geological Survey, Southwest Biological Science Center, Moab, UT 4 School of Forestry, Northern Arizona University, Flagstaff, AZ 5 Department of Ecology and Evolutionary Biology, University of Colorado, Boulder, CO 6 Mayberry Native Plant Propagation Center, Rim to Rim Restoration, Moab, UT 7 U.S. Geological Survey, Colorado Water Science Center, Grand Junction, CO 8 The Nature Conservancy-Utah, Moab, UT 9 School of Life Sciences and Center for Fundamental and Applied Microbiomics, Arizona State University, Tempe, AZ TABLE OF CONTENTS SECTION 1: BIOCRUSTS AND RESTORATION 1 Introduction: What is a biocrust and why should we restore it? ................ 1 SECTION 2: INOCULUM SOURCES, HARVESTING, PREPARATION, AND STORAGE 2.1 Introduction to inoculum sources, harvesting, preparation, and storage . 3 2.2 Propagule collection and growth . 4 2.2.1 Field collection . 4 2.2.2 Salvaged biocrusts . 4 2.2.3 Greenhouse-grown biocrusts ..................................... 5 2.2.4 Lab-grown biocrusts. 8 2.2.5 Outdoor biocrust production .................................... 11 2.3 Climate adapted biocrusts ........................................... 12 2.4 Preparation after growth/collection .................................... 13 2.4.1 Hardening . 13 2.4.2 Sieving size considerations and instructions ........................ 14 2.4.3 Storage . 14 CLIMATE-ADAPTED BIOCRUST RESTORATION INFOGRAPHIC ........... 15 SECTION 3: INOCULUM ADDITION 3.1 Introduction to inoculum additions. 17 3.2 Ways of adding inoculum ............................................ 17 3.3 Amounts of inoculum to add. 17 3.4 Season of inoculation. 18 SECTION 4: HABITAT PREPARATION, MODIFICATION, AND SOIL STABILITY 4.1 Introduction to habitat preparation, modification, and soil stability .......... 19 4.2 Habitat modifications. 19 4.2.1 Surface roughening ............................................ 19 4.2.2 Shading . 19 4.2.3 Irrigation and Water addition .................................... 19 4.2.4 Other habitat considerations. 20 4.3 Soil stability considerations ........................................... 20 4.3.1 Polyacrylamides ............................................... 20 4.3.2 Straw borders and checkerboards ................................ 20 4.3.3 Psyllium and other plant-based stabilizers . 21 SECTION 5: MONITORING METHODS AND PROTOCOLS 5.1 Introduction to monitoring methods and protocols ....................... 22 5.2 Types of monitoring methods . 22 5.2.1 Biocrust cover . 22 5.2.2 Chlorophyll a. 22 5.2.3 16S rRNA gene abundance . 23 5.2.4 Microbial community composition: 16S rRNA community diversity ..... 23 5.2.5 Soil aggregate stability ......................................... 23 5.2.6 Soil tensile strength and compaction . 24 5.2.7 Repeat photography ........................................... 25 5.2.8 Level of development .......................................... 25 5.2.9 Soil nutrients . 25 SECTION 6: MAJOR FINDINGS 6.1 Major findings, challenges, and opportunities ........................... 27 References. 28 SECTION 1: BIOCRUSTS AND RESTORATION 1. Introduction: What is biocrust and why (Belnap 2006) by increasing their access to water, or should we restore it? decrease infiltration by increasing runoff. In addition to the potentially beneficial role of biocrusts in native Biological soil crusts (biocrusts) are a community vascular plant growth and establishment, there is of lichens, mosses, cyanobacteria, and other evidence that the presence of lichen biocrusts inhibits nonvascular photosynthetic organisms and associated the establishment of grasses including the exotic, decomposers living on soil surfaces worldwide invasive annual grass, Bromus tectorum (Leines et al. (see Garcia-Pichel 2003 for a primer; Belnap et al. 2007). Inhibition of annual grasses by well-developed 2001, Weber et al. 2016 for monographs). Wherever biocrusts has also been shown in the Great Basin soils have direct access to the sun, biocrusts have and Australia (Crisp 1975, Larsen 1995, Howell 1998). the potential to exist. Indeed, biocrusts occur on A recent metaanalysis of biocrust effects on plants all continents and across contrasting biomes (for shows that relationships are context-dependent, but example, polar, temperate, arid), and comprise a support the assertion that intact biocrust promotes majority of surface cover in many arid and polar native over exotic plant growth (Havrilla et al. 2019). systems (Belnap et al. 2016; Torres-Cruz et al. 2018). A recent global estimate suggested that biocrusts cover Biocrusts play a central role in stabilizing dryland soils. about 12% of the Earth’s terrestrial surface (Rodriguez- A matrix of filamentous, motile cyanobacteria (Garcia- Caballero et al. 2018), and these photosynthetic soil Pichel and Wojciechowski 2009) forms throughout communities play foundational roles in the ecosystems the uppermost soil layers, providing most of the where they occur (Belnap et al. 2016). cohesion (Belnap and Gardner 1993). Lichens and mosses physically shield soils with their aboveground Biocrusts are especially influential in numerous thalli and with anchoring structures that penetrate the aspects of dryland ecosystems, such as those in soil matrix (Sanders 1994). The resultant dense aerial the western United States. They help set a system’s and subterranean network reduces wind (Belnap