Metagenomic assessment of nitrate-contaminated mine wastewaters and optimization of complete denitrification by indigenous enriched bacteria

Nitrate contamination in water remains to be on the rise globally due to continuous anthropogenic activities, such as mining and farming, which utilize high amounts of ammonium nitrate explosives and chemical-NPK-fertilizers, respectively. This study presents insights into the development of a bioremediation strategy to remove nitrates (NO3−) using consortia enriched from wastewater collected from a diamond mine in Lesotho and a platinum mine in South Africa. A biogeochemical analysis was conducted on the water samples which aided in comparing and elucidating their unique physicochemical parameters. The chemical analysis uncovered that both wastewater samples contained over 120 mg/L of NO3− and over 250 mg/L of sulfates (SO42-), which were both beyond the acceptable limit of the environmental surface water standards of South Africa. The samples were atypical of mine wastewaters as they had low concentrations of dissolved heavy metals and a pH of over 5. A metagenomic analysis applied to study microbial diversities revealed that both samples were dominated by the phyla Proteobacteria and Bacteroidetes, which accounted for over 40% and 15%, respectively. Three consortia were enriched to target denitrifying bacteria using selective media and then subjected to complete denitrification experiments. Denitrification dynamics and denitrifying capacities of the consortia were determined by monitoring dissolved and gaseous nitrogen species over time. Denitrification optimization was carried out by changing environmental conditions, including supplementing the cultures with metal enzyme co-factors (iron and copper) that were observed to promote different stages of denitrification. Copper supplemented at 50 mg/L was observed to be promoting complete denitrification of over 500 mg/L of NO3−, evidenced by the emission of nitrogen gas (N2) that was more than nitrous oxide gas (N2O) emitted as the terminal by-product. Modification and manipulation of growth conditions based on the microbial diversity enriched proved that it is possible to optimize a bioremediation system that can reduce high concentrations of NO3−, while emitting an environmentally-friendly N2 instead of N2O, that is, a greenhouse gas. Data collected and discussed in this research study can be used to model an upscale NO3− bioremediation system aimed to remove nitrogenous and other contaminants without secondary contamination

Whole genome comparison of Thermus sp. NMX2.A1 reveals principal carbon metabolism differences with closest relation Thermus scotoductus SA-01

Genome sequencing of the yellow-pigmented, thermophilic bacterium Thermus sp. NMX2.A1 resulted in a 2.29 Mb draft genome that encodes for 2312 proteins. The genetic relationship between various strains from the genus Thermus was assessed based on phylogenomic analyses using a concatenated set of conserved proteins. The resulting phylogenetic tree illustrated that Thermus sp. NMX2 A.1 clusters together with Thermus scotoductus SA-01, despite being isolated from vastly different geographical locations. The close evolutionary relationship and metabolic parallels between the two strains has previously been recognized; however, neither strain’s genome data were available at that point in time. Genomic comparison of the Thermus sp. NMX2.A1 and T. scotoductus SA-01, as well as other closely related Thermus strains, revealed a high degree of synteny at both the genomic and proteomic level, with processes such as denitrification and natural cell competence appearing to be conserved. However, despite this high level of similarity, analysis revealed a complete, putative Calvin–Benson–Bassham (CBB) cycle in NMX2.A1 that is absent in SA-01. Analysis of horizontally transferred gene islands provide evidence that NMX2 selected these genes due to pressure from its HCO3 - rich environment, which is in stark contrast to that of the deep subsurface isolated SA-01.The National Research Foundation and the Technology Innovation Agency, South Africa.http://www.g3journal.orgam2017Biochemistr

Novel lichen-dominated hypolithic communities in the Namib Desert

The ventral surfaces of translucent rocks from hot desert pavements often harbor hypolithic microbial communities, which are mostly dominated by cyanobacteria. The Namib Desert fog belt supports extensive hypolithic colonization of quartz rocks, which are also colonized by lichens on their dorsal surfaces. Here, we aim to evaluate whether lichens colonize the ventral surface of the rocks (i.e., show hypolithic lifestyle) and compare the bacterial composition of these coastal hypolithic communities with those found inland. Fungal DNA barcoding and fungal and bacterial Illumina metabarcoding were combined with electron microscopy to characterize the composition and spatial structure of hypolithic communities from two (coastal and inland) areas in the Namib Desert. We report, for the first time, the structure and composition of lichen-dominated hypolithic communities found in the coastal zone of the Namib Desert with extensive epilithic lichen cover. Lichen modified areoles with inverted morphology of the genus Stellarangia (three lineages) and Buellia (two lineages) were the main components of these hypolithic communities. Some of these lineages were also found in epilithic habitats. These lichen-dominated hypolithic communities differed in structural organization and bacterial community composition from those found in inland areas. The hypolithic lichen colonization characterized here seems not to be an extension of epilithic or biological soil crust lichen growths but the result of specific sublithic microenvironmental conditions. Moisture derived from fog and dew could be the main driver of this unique colonization.Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. DC acknowledges the financial support of the University of Pretoria and the National Research Foundation of South Africa. AV was supported by the project “CLU-2019–05 – IRNASA/CSIC Unit of Excellence,” funded by the Junta de Castilla y León and co-financed by the European Union (ERDF “Europe drives our growth”). This work was supported by grants CTM2015-64728-C2- 2-R (MINECO/FEDER, EU) and PID2019-105469RB-C22 (MICINN).http://link.springer.com/journal/248am2022BiochemistryGeneticsMicrobiology and Plant Patholog

Endophytic seed mycobiome of six sorghum (Sorghum bicolor) cultivars from commercial seedlots using an Illumina sequencing approach

18 páginas, 6 figuras, 2 tablasPathogen-free seeds are important for the establishment of young seedlings, prevention of health problems upon consumption by animals and livestock, and inadvertent movement of pathogens into and out of countries. However, testing for the presence of fungi, including pathogens from seeds is a time consuming and difficult process. In this study, we characterized the seed fungal microbiome (mycobiome) of six commercial sorghum cultivars from South Africa using a deep amplicon next generation sequencing approach based on the Internal Transcribed Spacer (ITS) region of the ribosomal operon. Sorghum is the fifth most important crop in the world, and widely used by African farmers. We found that the fungi present in each of the seedlots were similar to those reported by cultural studies. By comparing phylotypes of certain key families and genera to phylotypes used in established phylogenies and reputed sequences from public databases, the diagnostic value of the NGS method was also investigated. We showed that a number of molecular operational taxonomic units (MOTUs) could be identified at the species level and established that certain known pathogens are not present in the tested seeds, for instance in the Aspergillus group. Other groups could not be identified, not even to genus level. While acknowledging the shortcomings of using partial ITS data, we demonstrated that deep amplicon sequencing is a valuable diagnostic tool for seed disease control and prevention in some cases.Research funding was provided by the Agricultural Research Council Collaborative Consortium to Broaden the Food Base.Peer reviewe

Characterization of Microbial Diversity of Two Tomato Cultivars through Targeted Next-Generation Sequencing 16S rRNA and ITS Techniques

Even though the nutritional and economic values of Solanum lycopersicum (tomato) are substantially impacted by microbial spoilage, the available data on its microbial community, particularly during spoilage, are limited and have primarily been characterized using conventional culture-dependent methods. This study employed a targeted high-throughput next-generation sequencing method to longitudinally characterize the microbial diversity of two South African tomato cultivars (jam and round) at varied storage intervals (1, 6, and 12 days). Throughout the storage period, the bacterial communities of the two cultivars were more diverse than the fungal communities. The microbial diversity of both bacteria and fungi was greater and comparable between the cultivars on day 1, but becomes distinct as the storage period increases, with round tomatoes being more diverse than jam tomato, though, on day 12, jam tomato develops greater diversity than round tomato. Overall, the most abundant phyla (though Proteobacteria was most dominant) were Proteobacteria, Firmicutes, and Bacteriodota in the bacterial communities, while Ascomycota and Basidiomycota formed most fungal communities with Ascomycota being dominant. At the genus level, Pantoea and Klebsiella (bacteria), Hanseniaspora, Stemphylium, and Alternaria (fungi) were prevalent. Taken together, this study casts light on a broad microbial diversity profile thus, confirms the cultivars’ diversity and abundance differences

A clinically important, plasmid-borne antibiotic resistance gene (β-lactamase TEM-116) present in desert soils

The exhaustive use of antibiotics in humans, animal farming and other agricultural practices has resulted in the frequent appearance of antibiotic resistant bacteria in human-impacted habitats. However, antibiotic resistance in natural (less-impacted) habitats is less understood. Using shotgun metagenomics we analysed soils from relatively low anthropogenic impact sites across the Namib Desert. We report the presence of a clinically significant extended spectrum β-lactamase (TEM-116), on a ColE1-like plasmid also carrying a metal resistance gene (arsC). The co-occurrence of resistance to antimicrobial drugs and metals encoded on a single mobile genetic element increases the probability of dissemination of these resistance determinants and the potential selection of multiple resistance mechanisms. In addition, the presence of a P7 entero-bacteriophage on the same plasmid, may represent a new vehicle for the propagation of TEM-116 in these soil communities. These findings highlight the role of the environment in the One Health initiative.The National Research Foundation (NRF) of South Africahttp://www.elsevier.com/locate/scitotenv2021-06-01hj2020BiochemistryGeneticsMicrobiology and Plant Patholog

Nitrate Water Contamination from Industrial Activities and Complete Denitrification as a Remediation Option

Freshwater is a scarce resource that continues to be at high risk of pollution from anthropogenic activities, requiring remediation in such cases for its continuous use. The agricultural and mining industries extensively use water and nitrogen (N)-dependent products, mainly in fertilizers and explosives, respectively, with their excess accumulating in different water bodies. Although removal of NO3 from water and soil through the application of chemical, physical, and biological methods has been studied globally, these methods seldom yield N2 gas as a desired byproduct for nitrogen cycling. These methods predominantly cause secondary contamination with deposits of chemical waste such as slurry brine, nitrite (NO2), ammonia (NH3), and nitrous oxide (N2O), which are also harmful and fastidious to remove. This review focuses on complete denitrification facilitated by bacteria as a remedial option aimed at producing nitrogen gas as a terminal byproduct. Synergistic interaction of different nitrogen metabolisms from different bacteria is highlighted, with detailed attention to the optimization of their enzymatic activities. A biotechnological approach to mitigating industrial NO3 contamination using indigenous bacteria from wastewater is proposed, holding the prospect of optimizing to the point of complete denitrification. The approach was reviewed and found to be durable, sustainable, cost effective, and environmentally friendly, as opposed to current chemical and physical water remediation technologies

Anaerobic reduction of europium by a Clostridium strain as a strategy for rare earth biorecovery

11 páginas, 7 figurasThe biorecovery of europium (Eu) from primary (mineral deposits) and secondary (mining wastes) resources is of interest due to its remarkable luminescence properties, important for modern technological applications. In this study, we explored the tolerance levels, reduction and intracellular bioaccumulation of Eu by a site-specific bacterium, Clostridium sp. 2611 isolated from Phalaborwa carbonatite complex. Clostridium sp. 2611 was able to grow in minimal medium containing 0.5 mM Eu3+. SEM-EDX analysis confirmed an association between Eu precipitates and the bacterium, while TEM-EDX analysis indicated intracellular accumulation of Eu. According to the HR-XPS analysis, the bacterium was able to reduce Eu3+ to Eu2+ under growth and non-growth conditions. Preliminary protein characterization seems to indicate that a cytoplasmic pyruvate oxidoreductase is responsible for Eu bioreduction. These findings suggest the bioreduction of Eu3+ by Clostridium sp. as a resistance mechanism, can be exploited for the biorecovery of this metal.This research was conducted with financial support from the Technology and Innovation Agency. The financial assistance of the National Research Foundation of South Africa (88833 and 106460) towards this research is hereby acknowledged.Peer reviewe

Baseline Data of the Fungal Phytobiome of Three Sorghum (Sorghum bicolor) Cultivars in South Africa using Targeted Environmental Sequencing

Plant-associated fungi, or the mycobiome, inhabit plant surfaces above ground, reside in plant tissues as endophytes, or are rhizosphere in the narrow zone of soil surrounding plant roots. Studies have characterized mycobiomes of various plant species, but little is known about the sorghum mycobiome, especially in Africa, despite sorghum being one of the most important indigenous and commercial cereals in Africa. In this study, the mycobiome associated with above- and below-ground tissues of three commercial sorghum cultivars, as well as from rhizosphere and surrounding bulk soil samples, were sequenced using targeted sequencing with the Illumina MiSeq platform. Relative abundance differences between fungal communities were found between above-ground and below-ground niches, with most differences mostly in the dominant MOTUs, such as Davidiellaceae sp. (Cladosporium), Didymellaceae sp. 1 (Phoma), Fusarium, Cryptococcus and Mucor. Above-ground communities also appeared to be more diverse than below-ground communities, and plants harboured the most diversity. A considerable number of MOTUs were shared between the cultivars although, especially for NS5511, their abundances often differed. Several of the detected fungal groups include species that are plant pathogens of sorghum, such as Fusarium, and, at low levels, Alternaria and the Ustilaginomycetes. Findings from this study illustrate the usefulness of targeted sequencing of the ITS rDNA gene region (ITS2) to survey and monitor sorghum fungal communities and those from associated soils. This knowledge may provide tools for disease management and crop production and improvement

Plant and Soil Core Mycobiomes in a Two-Year Sorghum–Legume Intercropping System of Underutilized Crops in South Africa

Fungal communities form close beneficial (mutualists) or detrimental (pathogens) associations with their plant hosts. Their diversity and abundance can be affected by agricultural practices which include cropping systems such as rotations and intercropping. Despite the importance of cropping systems in increasing productivity, knowledge of the fungal mycobiome and the core inhabitants for under-utilised cereal and legume crops, particularly over a period, is still limited. The core mycobiomes in plant tissues and bulk soils of a cereal–legume intercrop were characterized over two years using high-throughput sequencing. The intercropping trial consisted of sorghum, Bambara groundnut, cowpea, dry bean, and soybean. A greater number of molecular operational taxonomic units (MOTUs) were found in plant tissues compared to those from the soils and between year one and year two. Principal coordinate analyses revealed that fungal communities for each year were relatively distinct, particularly for the soils. The core mycobiome was dominated by a Davidiellaceae sp. (Cladosporium), Didymellaceae sp. 1 (Phoma), Didymellaceae sp. 2 (Epicoccum), Fusarium sp. 2, Unidentified (Ascomycota), and Cryptococcus MOTUs that were present in all plant tissues and soils of year one and two. Other key MOTUs were only specific to a year, substrate, or crop. Although the mycobiome of sorghum were more distinct than the cores of the legumes, there were still MOTUs dominant across all of the crops. Characterization of this baseline core across two years provides insight into those fungi that are always present in these crops, and that could be utilized in improving crop performance and productivity