Use of lignocellulosic waste materials in the passive treatment of highly alkaline wastewater contaminated with sulfates and metals - From a laboratory study to pilot scale.

Passive wastewater treatment systems are an alternative to costly and ineffective chemical wastewater treatment methods. Lignocellulosic waste materials (LWM) are often used in passive wastewater treatment systems as a cheap and accessible source of nutrients. LWM, such as spent mushroom compost and woodchips, have been implemented for the successful management of mildly alkaline effluents, which constitute a large fraction of industrial wastewater. The objective of the study was to provide an extensive study of the parameters in four types of commonly used LWM (raw and composted sawdust, spent mushroom compost and woodchips), which can be used in the planning of a passive wastewater treatment plant. LWM were shown to remove up to 90% Zn2+ and Pb2+ from a model solution and neutralize wastewater. Moreover, the LWM were inhabited by a physiologically diverse microbial consortium containing sulfate-reducing and cellulolytic microbes, which can influence the treatment process. Another purpose of this study was to construct a pilot wastewater treatment plant based on the use of LWM and gravel and to present its ability to effectively treat extremely alkaline flotation wastewater (pH = 12) originating from a lead and zinc mine located in Montenegro. The treated wastewater had a unique, but challenging chemical composition for passive treatment, as it was heavily contaminated with sulfates (∼1200 mg/L) and lead (∼1 g/L). The removal within the developed installation reached a rate of 66%, while the treated effluent, after initial neutralization, was maintained at a pH of approximately 7. Lead and zinc concentrations after treatment were also kept at levels required by Montenegrin law for wastewater disposal.

Recent Advances in Cardiovascular Diseases Research Using Animal Models and PET Radioisotope Tracers.

Cardiovascular diseases (CVD) is a collective term describing a range of conditions that affect the heart and blood vessels. Due to the varied nature of the disorders, distinguishing between their causes and monitoring their progress is crucial for finding an effective treatment. Molecular imaging enables non-invasive visualisation and quantification of biological pathways, even at the molecular and subcellular levels, what is essential for understanding the causes and development of CVD. Positron emission tomography imaging is so far recognized as the best method for in vivo studies of the CVD related phenomena. The imaging is based on the use of radioisotope-labelled markers, which have been successfully used in both pre-clinical research and clinical studies. Current research on CVD with the use of such radioconjugates constantly increases our knowledge and understanding of the causes, and brings us closer to effective monitoring and treatment. This review outlines recent advances in the use of the so-far available radioisotope markers in the research on cardiovascular diseases in rodent models, points out the problems and provides a perspective for future applications of PET imaging in CVD studies.

Diversity and Horizontal Transfer of Antarctic Pseudomonas spp. Plasmids.

Pseudomonas spp. are widely distributed in various environments around the world. They are also common in the Antarctic regions. To date, almost 200 plasmids of Pseudomonas spp. have been sequenced, but only 12 of them were isolated from psychrotolerant strains. In this study, 15 novel plasmids of cold-active Pseudomonas spp. originating from the King George Island (Antarctica) were characterized using a combined, structural and functional approach, including thorough genomic analyses, functional analyses of selected genetic modules, and identification of active transposable elements localized within the plasmids and comparative genomics. The analyses performed in this study increased the understanding of the horizontal transfer of plasmids found within Pseudomonas populations inhabiting Antarctic soils. It was shown that the majority of the studied plasmids are narrow-host-range replicons, whose transfer across taxonomic boundaries may be limited. Moreover, structural and functional analyses enabled identification and characterization of various accessory genetic modules, including genes encoding major pilin protein (PilA), that enhance biofilm formation, as well as active transposable elements. Furthermore, comparative genomic analyses revealed that the studied plasmids of Antarctic Pseudomonas spp. are unique, as they are highly dissimilar to the other known plasmids of Pseudomonas spp.

Genomic Analysis of Shewanella sp. O23S-The Natural Host of the pSheB Plasmid Carrying Genes for Arsenic Resistance and Dissimilatory Reduction.

Shewanella sp. O23S is a dissimilatory arsenate reducing bacterial strain involved in arsenic transformations within the abandoned gold mine in Zloty Stok (SW Poland). Previous physiological studies revealed that O23S may not only release arsenic from minerals, but also facilitate its immobilization through co-precipitation with reduced sulfur species. Given these uncommon, complementary characteristics and the application potential of the strain in arsenic-removal technologies, its genome (~5.3 Mbp), consisting of a single chromosome, two large plasmids (pSheA and pSheB) and three small plasmid-like phages (pSheC-E) was sequenced and annotated. Genes encoding putative proteins involved in heavy metal transformations, antibiotic resistance and other phenotypic traits were identified. An in-depth comparative analysis of arsenic respiration (arr) and resistance (ars) genes and their genetic context was also performed, revealing that pSheB carries the only copy of the arr genes, and a complete ars operon. The plasmid pSheB is therefore a unique natural vector of these genes, providing the host cells arsenic respiration and resistance abilities. The functionality of the identified genes was determined based on the results of the previous and additional physiological studies, including: the assessment of heavy metal and antibiotic resistance under various conditions, adhesion-biofilm formation assay and BiologTM metabolic preferences test. This combined genetic and physiological approach shed a new light on the capabilities of O23S and their molecular basis, and helped to confirm the biosafety of the strain in relation to its application in bioremediation technologies.

Genomic and Biotechnological Characterization of the Heavy-Metal Resistant, Arsenic-Oxidizing Bacterium Ensifer sp. M14.

Ensifer (Sinorhizobium) sp. M14 is an efficient arsenic-oxidizing bacterium (AOB) that displays high resistance to numerous metals and various stressors. Here, we report the draft genome sequence and genome-guided characterization of Ensifer sp. M14, and we describe a pilot-scale installation applying the M14 strain for remediation of arsenic-contaminated waters. The M14 genome contains 6874 protein coding sequences, including hundreds not found in related strains. Nearly all unique genes that are associated with metal resistance and arsenic oxidation are localized within the pSinA and pSinB megaplasmids. Comparative genomics revealed that multiple copies of high-affinity phosphate transport systems are common in AOBs, possibly as an As-resistance mechanism. Genome and antibiotic sensitivity analyses further suggested that the use of Ensifer sp. M14 in biotechnology does not pose serious biosafety risks. Therefore, a novel two-stage installation for remediation of arsenic-contaminated waters was developed. It consists of a microbiological module, where M14 oxidizes As(III) to As(V) ion, followed by an adsorption module for As(V) removal using granulated bog iron ores. During a 40-day pilot-scale test in an abandoned gold mine in Zloty Stok (Poland), water leaving the microbiological module generally contained trace amounts of As(III), and dramatic decreases in total arsenic concentrations were observed after passage through the adsorption module. These results demonstrate the usefulness of Ensifer sp. M14 in arsenic removal performed in environmental settings.

Granulated Bog Iron Ores as Sorbents in Passive (Bio)Remediation Systems for Arsenic Removal.

The main element of PbRS (passive (bio)remediation systems) are sorbents, which act as natural filters retaining heavy metals and carriers of microorganisms involved in water treatment. Thus, the effectiveness of PbRS is determined by the quality of the (ad)sorbents, which should be stable under various environmental conditions, have a wide range of applications and be non-toxic to (micro)organisms used in these systems. Our previous studies showed that bog iron ores (BIOs) meet these requirements. However, further investigation of the physical and chemical parameters of BIOs under environmental conditions is required before their large-scale application in PbRS. The aim of this study was (i) to investigate the ability of granulated BIOs (gBIOs) to remove arsenic from various types of contaminated waters, and (ii) to estimate the application potential of gBIOs in technologies dedicated to water treatment. These studies were conducted on synthetic solutions of arsenic and environmental samples of arsenic contaminated water using a set of adsorption columns filled with gBIOs. The experiments performed in a static system revealed that gBIOs are appropriate arsenic and zinc adsorbent. Dynamic adsorption studies confirmed these results and showed, that the actual sorption efficiency of gBIOs depends on the adsorbate concentration and is directly proportional to them. Desorption analysis showed that As-loaded gBIOs are characterized by high chemical stability and they may be reused for the (ad)sorption of other elements, i.e., zinc. It was also shown that gBIOs may be used for remediation of both highly oxygenated waters and groundwater or settling ponds, where the oxygen level is low, as both forms of inorganic arsenic (arsenate and arsenite) were effectively removed. Arsenic concentration after treatment was <100 µg/L, which is below the limit for industrial water.

Analysis of the Genome and Mobilome of a Dissimilatory Arsenate Reducing Aeromonas sp. O23A Reveals Multiple Mechanisms for Heavy Metal Resistance and Metabolism.

Aeromonas spp. are among the most ubiquitous microorganisms, as they have been isolated from different environmental niches including waters, soil, as well as wounds and digestive tracts of poikilothermic animals and humans. Although much attention has been paid to the pathogenicity of Aeromonads, the role of these bacteria in environmentally important processes, such as transformation of heavy metals, remains to be discovered. Therefore, the aim of this study was a detailed genomic characterization of Aeromonas sp. O23A, the first representative of this genus capable of dissimilatory arsenate reduction. The strain was isolated from microbial mats from the Zloty Stok mine (SW Poland), an environment strongly contaminated with arsenic. Previous physiological studies indicated that O23A may be involved in both mobilization and immobilization of this metalloid in the environment. To discover the molecular basis of the mechanisms behind the observed abilities, the genome of O23A (∼5.0 Mbp) was sequenced and annotated, and genes for arsenic respiration, heavy metal resistance (hmr) and other phenotypic traits, including siderophore production, were identified. The functionality of the indicated gene modules was assessed in a series of minimal inhibitory concentration analyses for various metals and metalloids, as well as mineral dissolution experiments. Interestingly, comparative analyses revealed that O23A is related to a fish pathogen Aeromonas salmonicida subsp. salmonicida A449 which, however, does not carry genes for arsenic respiration. This indicates that the dissimilatory arsenate reduction ability may have been lost during genome reduction in pathogenic strains, or acquired through horizontal gene transfer. Therefore, particular emphasis was placed upon the mobilome of O23A, consisting of four plasmids, a phage, and numerous transposable elements, which may play a role in the dissemination of hmr and arsenic metabolism genes in the environment. The obtained results indicate that Aeromonas sp. O23A is well-adapted to the extreme environmental conditions occurring in the Zloty Stok mine. The analysis of genome encoded traits allowed for a better understanding of the mechanisms of adaptation of the strain, also with respect to its presumable role in colonization and remediation of arsenic-contaminated waters, which may never have been discovered based on physiological analyses alone.

The role of dissimilatory arsenate reducing bacteria in the biogeochemical cycle of arsenic based on the physiological and functional analysis of Aeromonas sp. O23A.

Dissimilatory arsenate reducing bacteria (DARB) are known to contribute to the mobilization of arsenic and other elements from minerals. Despite this, metabolic capabilities of only a few DARB strains have been thoroughly investigated so far, and the influence of these bacteria on the bioavailability of arsenic in the environment is still a topic for discussion. In this study, Aeromonas sp. O23A, one of the dominant DARB strains found in the Zloty Stok mine (SW Poland), was subjected to a detailed physiological and functional analysis aimed to identify the actual environmental impact of this strain. Physiological analyses revealed that O23A is a facultative anaerobe, capable of utilizing arsenate as a respiratory substrate and acetate, citrate and lactate as electron donors. Arsenate reduction was observed within the first 24h of culturing. The strain shows high resistance to arsenic and several other heavy metals (i.a. Cu, Fe, Ni and Zn) as well tolerance to a broad range of physico-chemical conditions. Metabolic preferences of O23A were thoroughly investigated using Biolog™ MicroArray assay. The strain was found to produce hydroxamate siderophores, potentially involved in the mobilization of iron and co-occurring heavy metals from minerals. On the other hand, O23A showed high adherence abilities, and its involvement in biofilm formation may lead to the entrapment of dissolved arsenic species and other toxic ions. The results shed light on the importance of arsenic respiration and resistance in the overall metabolism of Aeromonas sp. O23A and confirmed its key role in the biogeochemical cycle of arsenic, also in the context of self-purification of heavy-metal-contaminated waters.

Shewanella sp. O23S as a Driving Agent of a System Utilizing Dissimilatory Arsenate-Reducing Bacteria Responsible for Self-Cleaning of Water Contaminated with Arsenic.

The purpose of this study was a detailed characterization of Shewanella sp. O23S, a strain involved in arsenic transformation in ancient gold mine waters contaminated with arsenic and other heavy metals. Physiological analysis of Shewanella sp. O23S showed that it is a facultative anaerobe, capable of growth using arsenate, thiosulfate, nitrate, iron or manganite as a terminal electron acceptor, and lactate or citrate as an electron donor. The strain can grow under anaerobic conditions and utilize arsenate in the respiratory process in a broad range of temperatures (10-37 °C), pH (4-8), salinity (0%-2%), and the presence of heavy metals (Cd, Co, Cr, Cu, Mn, Mo, Se, V and Zn). Under reductive conditions this strain can simultaneously use arsenate and thiosulfate as electron acceptors and produce yellow arsenic (III) sulfide (As2S3) precipitate. Simulation of As-removal from water containing arsenate (2.5 mM) and thiosulfate (5 mM) showed 82.5% efficiency after 21 days of incubation at room temperature. Based on the obtained results, we have proposed a model of a microbially mediated system for self-cleaning of mine waters contaminated with arsenic, in which Shewanella sp. O23S is the main driving agent.

Plasmid diversity in arctic strains of Psychrobacter spp.

Six strains of Psychrobacter spp. isolated from guano of little auks collected on Spitsbergen island (Arctic) carried nine plasmids that were fully sequenced. These replicons (ranging in size from 2917 to 14924 bp) contained either repA (ColE2-type) or repB (iteron-type) replication systems of a relatively narrow host range, limited to Psychrobacter spp. All but one of the plasmids carried predicted mobilization for conjugal transfer systems, encoding relaxases of the MOBQ, MOBV or MOBP families. The plasmids also contained diverse additional genetic load, including a type II restriction-modification system and a gene encoding a putative subunit C of alkyl hydroperoxide reductase (AhpC)-an antioxidant enzyme and major scavenger of reactive oxygen species. Detailed comparative sequence analyses, extended to all plasmids identified so far in psychrophilic bacteria, distinguished groups of the most ubiquitous replicons, which play a key role in horizontal gene transfer in cold environments.