In Morocco, pollution caused by closed mines continues to be a serious threat to the environment, like the generation of acid mine drainage. Mine drainage is produced by environmental and microbial oxidation of sulfur minerals originating from mine wastes. The fundamental role of microbial communities is well known, like implication of Fe-oxidizing and to a lesser extent S-oxidizing microorganism in bioleaching. However, the structure of the microbial communities varies a lot from one site to another, like diversity depends on many factors such as mineralogy, concentration of metals and metalloids or pH, etc. In this study, prokaryotic communities in the pyrrhotite-rich tailings of Kettara mine were characterized using the Illumina sequencing. In-depth phylogenetic analysis revealed a total of 12 phyla of bacteria and 1 phyla of Archaea. The majority of sequences belonged to the phylum of Proteobacteria and Firmicutes with a predominance of Bacillus, Pseudomonas or Corynebacterium genera. Many microbial populations are implicated in the iron, sulfur and arsenic cycles, like Acidiferrobacter, Leptospirillum, or Alicyclobacillus in Fe; Acidiferrobacter and Sulfobacillus in S; and Bacillus or Pseudomonas in As. This is one of the first description of prokaryotic communities in pyrrhotite-rich mine tailings using high-throughput sequencing.

growth promoting microorganisms 2. Acid mine drainage 2.1 Implication of microorganisms in formation and remediation of acid mine drainage 2.1.1 Implication of microorganisms in formation of acid mine drainage Many metals naturally occur as sulfide ores (e.g. Pb in galena, Cu in chalcopyrite, Zn in sphalerite). Acid mine drainage is generated when sulfide minerals (especially pyrite or pyrrhotite) come into contact with oxygen and water (Hallberg 2010). Although AMD occurs naturally, mining operations like excavation and milling accelerate the process by increasing surface area of exposure of sulfide minerals to water, oxygen and microorganisms (Simate and Ndlovu, 2014). AMD is often characterized by pH values below 4 and sometimes as low as-3, as measured at the Richmond mine in California (Nordstrom et al., 2000). Such low pH facilitates the mobilization of metals and metalloids, increasing their dispersion into the water (Humphries et al., 2017). AMD represents one of the most problematic pollution of water in industrial and post-industrial areas throughout the world (Hallberg, 2010). Numerous works have been done to characterize the autochthonous microbial communities (Bacteria, Archaea, Eucaryota) thriving in these extreme environments (

Soil microbiota are vital for successful revegetation, as they play a critical role in nutrient cycles, soil functions, and plant growth and health. A rehabilitation scenario of the abandoned Kettara mine (Morocco) includes covering acidic tailings with alkaline phosphate mine wastes to limit water infiltration and hence acid mine drainage. Revegetation of phosphate wastes is the final step to this rehabilitation plan. However, revegetation is hard on this type of waste in semi-arid areas and only a few plants managed to grow naturally after 5 years on the store-and-release cover. As we know that belowground biodiversity is a key component for aboveground functioning, we sought to know if any structural problem in phosphate waste communities could explain the almost absence of plants. To test this hypothesis, bacterial and archaeal communities present in these wastes were assessed by 16S rRNA metabarcoding. Exploration of taxonomic composition revealed a quite diversified community assigned to 19 Bacterial and two Archaeal phyla, similar to other studies, that do not appear to raise any particular issues of structural problems. The dominant sequences belonged to Proteobacteria, Bacteroidetes, Actinobacteria, and Gemmatimonadetes and to the genera Massilia, Sphingomonas, and Adhaeribacter. LEfSe analysis identified 19 key genera, and metagenomic functional prediction revealed a broader phylogenetic range of taxa than expected, with all identified genera possessing at least one plant growth-promoting trait. Around 47% of the sequences were also related to genera possessing strains that facilitate plant development under biotic and environmental stress conditions, such as drought and heat.

Morocco holds the vast majority of the world’s phosphate reserves, but due to the processes involved in extracting and commercializing these reserves, large quantities of de-structured, nutritionally deficient mine phosphate wastes are produced each year. In a semi-arid climate, these wastes severely hamper plant growth and development leading to huge unvegetated areas. Soil indigenous Plant Growth-Promoting Bacteria (PGPB) play a pivotal role in restauration of these phosphate mining wastes by revegetation, by increasing plants development, soil functioning, and nutrient cycling. The development of a vegetative cover above the degraded phosphate wastes, could stabilize and reintegrate these wastes in the surrounding environment. The current study’s objectives were to isolate, characterize, and identify indigenous bacterial strains, and test their PGP activity in vitro and, for the best-performing strains in planta, in order to assess their potential for acting as biofertilizers. A quantitative test for the synthesis of auxin and the production of siderophores as well as a qualitative test for the solubilization of phosphate were performed on all isolated bacterial strains. The production of hydrogen cyanide (HCN), exopolysaccharides (EPS), and enzymes were also examined. Three bacteria, selected among the best PGPB of this study, were tested in planta to determine whether such indigenous bacteria could aid plant growth in this de-structured and nutrient-poor mining soil. Using 16S rRNA gene sequencing, 41 bacterial strains were isolated and 11 genera were identified: Acinetobacter, Agrococcus, Bacillus, Brevibacterium, Microbacterium, Neobacillus, Paenibacillus, Peribacillus, Pseudarthrobacter, Stenotrophomonas, and Raoultella. Among the three best performing bacteria (related to Bacillus paramycoides, Brevibacterium anseongense, and Stenotrophomonas rhizophila), only Stenotrophomonas rhizophila and Brevibacterium anseongense were able to significantly enhance Lupinus albus L. growth. The best inoculation results were obtained using the strain related to Stenotrophomonas rhizophila, improving the plant’s root dry weight and chlorophyll content. This is also, to our knowledge, the first study to show a PGP activity of Brevibacterium anseongense.

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