Isolation of biocrust cyanobacteria and evaluation of Cu, Pb, and Zn immobilisation potential for soil restoration and sustainable agriculture.

Soil contamination by heavy metals represents an important environmental and public health problem of global concern. Biocrust-forming cyanobacteria offer promise for heavy metal immobilisation in contaminated soils due to their unique characteristics, including their ability to grow in contaminated soils and produce exopolysaccharides (EPS). However, limited research has analysed the representativeness of cyanobacteria in metal-contaminated soils. Additionally, there is a lack of studies examining how cyanobacteria adaptation to specific environments can impact their metal-binding capacity. To address this research gap, we conducted a study analysing the bacterial communities of cyanobacteria-dominated biocrusts in a contaminated area from South Sardinia (Italy). Additionally, by using two distinct approaches, we isolated three Nostoc commune strains from cyanobacteria-dominated biocrust and we also evaluated their potential to immobilise heavy metals. The first isolation method involved acclimatizing biocrust samples in liquid medium while, in the second method, biocrust samples were directly seeded onto agar plates. The microbial community analysis revealed Cyanobacteria, Bacteroidota, Proteobacteria, and Actinobacteria as the predominant groups, with cyanobacteria representing between 13.3 % and 26.0 % of the total community. Despite belonging to the same species, these strains exhibited different growth rates (1.1-2.2 g L-1 of biomass) and capacities for EPS production (400-1786 mg L-1). The three strains demonstrated a notable ability for metal immobilisation, removing up to 88.9 % of Cu, 86.2 % of Pb, and 45.3 % of Zn from liquid medium. Cyanobacteria EPS production showed a strong correlation with the removal of Cu, indicating its role in facilitating metal immobilisation. Furthermore, differences in Pb immobilisation (40-86.2 %) suggest possible environmental adaptation mechanisms of the strains. This study highlights the promising application of N. commune strains for metal immobilisation in soils, offering a potential bioremediation tool to combat the adverse effects of soil contamination and promote environmental sustainability.

Non-rainfall water inputs: A key water source for biocrust carbon fixation.

Links between water and carbon (C) cycles in drylands are strongly regulated by biocrusts. These widespread communities in the intershrub spaces of drylands are able to use non-rainfall water inputs (NRWI) (fog, dewfall and water vapour) to become active and fix carbon dioxide (CO2), converting biocrusts into the main soil C contributors during periods in which vegetation remains inactive. In this study, we first evaluated the influence of biocrust type on NRWI uptake using automated microlysimeters, and second, we performed an outdoor experiment to examine how NRWI affected C exchange (photosynthesis and respiration) in biocrusts. NRWI uptake increased from incipient cyanobacteria to well-developed cyanobacteria and lichen biocrusts. NRWI triggered biocrust activity but with contrasting effects on CO2 fluxes depending on the main NRWI source. Fog mainly stimulated respiration of biocrust-covered soils, reaching net CO2 emissions of 0.68 µmol m-2 s-1, while dew had a greater effect stimulating biocrust photosynthesis and resulted in net CO2 uptake of 0.66 µmol m-2 s-1. These findings demonstrate the key role that NRWI play in biocrust activity and the soil C balance in drylands.

Exopolysaccharide Features Influence Growth Success in Biocrust-forming Cyanobacteria, Moving From Liquid Culture to Sand Microcosms.

Land degradation in drylands is a drawback of the combined action of climate change and human activities. New techniques have been developed to induce artificial biocrusts formation as a tool for restoration of degraded drylands, and among them soils inoculation with cyanobacteria adapted to environmental stress. Improvement of soil properties by cyanobacteria inoculation is largely related to their ability to synthesize exopolysaccharides (EPS). However, cyanobacterial EPS features [amount, molecular weight (MW), composition] can change from one species to another or when grown in different conditions. We investigated the differences in growth and polysaccharidic matrix features among three common biocrust-forming cyanobacteria (Nostoc commune, Scytonema javanicum, and Phormidium ambiguum), when grown in liquid media and on sandy soil microcosms under optimal nutrient and water, in controlled laboratory conditions. We extracted and analyzed the released EPS (RPS) and sheath for the liquid cultures, and the more soluble or loosely-bound (LB) and the more condensed or tightly-bound (TB) soil EPS fractions for the sandy soil microcosms. In liquid culture, P. ambiguum showed the greatest growth and EPS release. In contrast, on the sandy soil, S. javanicum showed the highest growth and highest LB-EPS content. N. commune showed no relevant growth after its inoculation of the sandy soil. A difference was observed in terms of MW distribution, showing that the higher MW of the polymers produced by P. ambiguum and S. javanicum compared to the polymers produced by N. commune, could have had a positive effect on growth for the first two organisms when inoculated on the sandy soil. We also observed how both RPS and sheath fractions reflected in the composition of the soil TB-EPS fraction, indicating the role in soil stabilization of both the released and the cell attached EPS. Our results indicate that the features of the polysaccharidic matrix produced by different cyanobacteria can influence their growth success in soil. These results are of great relevance when selecting suitable candidates for large-scale cyanobacteria applications in soil restoration.

Polyphasic evaluation of key cyanobacteria in biocrusts from the most arid region in Europe.

Cyanobacteria are key microbes in topsoil communities that have important roles in preventing soil erosion, carbon and nitrogen fixation, and influencing soil hydrology. However, little is known regarding the identity and distribution of the microbial components in the photosynthetic assemblages that form a cohesive biological soil crust (biocrust) in drylands of Europe. In this study, we investigated the cyanobacterial species colonizing biocrusts in three representative dryland ecosystems from the most arid region in Europe (SE Spain) that are characterized by different soil conditions. Isolated cyanobacterial cultures were identified by a polyphasic approach, including 16S rRNA gene sequencing, phylogenetic relationship determination, and morphological and ecological habitat assessments. Three well-differentiated groups were identified: heterocystous-cyanobacteria (Nostoc commune, Nostoc calcicola, Tolypothrix distorta and Scytonema hyalinum), which play an important role in N and C cycling in soil; nonheterocystous bundle-forming cyanobacteria (Microcoleus steenstrupii, Trichocoleus desertorum, and Schizothrix cf. calcicola); and narrow filamentous cyanobacteria (Leptolyngbya frigida and Oculatella kazantipica), all of which are essential genera for initial biocrust formation. The results of this study contribute to our understanding of cyanobacterial species composition in biocrusts from important and understudied European habitats, such as the Mediterranean Basin, a hotspot of biodiversity, where these species are keystone pioneer organisms.

Soil Type and Cyanobacteria Species Influence the Macromolecular and Chemical Characteristics of the Polysaccharidic Matrix in Induced Biocrusts.

Inoculation of soils with cyanobacteria is proposed as a sustainable biotechnological technique for restoration of degraded areas in drylands due to the important role that cyanobacteria and their exopolysaccharides (EPS) play in the environment. So far, few studies have analyzed the macromolecular and chemical characteristics of the polysaccharidic matrix in induced cyanobacterial biocrusts and the scarce existing studies have mainly focused on sandy soil textures. However, the characteristics of the cyanobacterial polysaccharidic matrix may greatly depend on soil type. The objective of this study was to examine the macromolecular distribution and monosaccharidic composition of the polysaccharidic matrix induced by inoculation of two cyanobacterial species common in arid environments, Phormidium ambiguum (non N-fixing) and Scytonema javanicum (N-fixing) in different soil types. S. javanicum promoted a higher release in the soil of the more soluble and less condensed EPS fraction (i.e., the loosely bound EPS fraction, LB-EPS), while P. ambiguum showed a higher release of the less soluble and more condensed EPS fraction (i.e., the tightly bound EPS fraction, TB-EPS). LB-EPSs were mainly composed of low MW molecules (< 50 kDa), while TB-EPSs were mainly composed of high MW molecules (1100-2000 kDa). The two EPS fractions showed a complex monosaccharidic composition (from 11 to 12 different types of monosaccharides), with glucose as the most abundant monosaccharide, in particular in the poorer soils characterized by lower organic C contents. In more C-rich soils, high abundances of galactose, mannose, and xylose were also found. Low abundance of uronic acids and hydrophobic monosaccharides, such as fucose and rhamnose, was found in the EPS extracted from the inoculated soils. Our results point to the influence of soil type on the macromolecular distribution and monosaccharide composition of the polysaccharidic matrix in induced biocrusts, which is likely to affect biocrust development and their role in soil structure and nutrient cycling in restored dryland soils.