Multi-locus sequence typing and in vitro antimicrobial resistance of equine Streptococcus equi subspecies zooepidemicus strains.

Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is one of the most important pathogens frequently associated with the main causes of equine infertility. In this study, we surveyed 22 strains of S. zooepidemicus collected during 2021 from cervico-uterine swabs of mares with endometritis. The genetic variability of the isolated strains was studied by multi-locus sequence typing (MLST) from whole-genome sequencing (WGS) data. The average length of reconstructed genomes was 2,088,286 bp (95% CI: 2,061,569 bp-2,114,967 bp), which was expected for S. zooepidemicus genomes. The assembled genomes were assigned to sequence types (STs) using the S. zooepidemicus scheme targeting seven loci (arcC, nrdE, proS, spi, tdk, tpi, yqiL) available in PubMLST database. MLST revealed a wide variability of STs with two (9.1%) novel STs identified in this study, precisely ST521 with two isolates and ST522 with one isolate. Furthermore, 4/22 (18.2%) isolates were assigned to ST92, 3/22 (13.6%) to ST205, 2/22 (9.1%) to ST475, and one strain (4.5%) for each of the following STs: ST10, ST30, ST39, ST49, ST101, ST132, ST147, ST314, ST369, ST467. Isolates were also tested for antimicrobial resistance using Kirby-Bauer disk diffusion method. Resistance to amoxicillin-clavulanate, ampicillin, amikacin, gentamicin, streptomycin, enrofloxacin, sulfamethoxazole-trimethoprim, tetracycline, oxytetracycline represented the most common resistance profile (13/22, 59.1%). No correlation between specific ST and antimicrobial resistance profile was found. Our study provides a comprehensive insight into the epidemiology, ST diversity and antimicrobial resistance profile of S. zooepidemicus strains, isolated in Italy, causing subfertility problems in mares.

Waste management beyond the COVID-19 pandemic: Bibliometric and text mining analyses.

The outbreak of the COVID-19 pandemic has significantly increased the demand for personal protective equipment, in particular face masks, thus leading to a huge amount of healthcare waste generated worldwide. Consequently, such an unprecedented amount of newly emerged waste has posed significant challenges to practitioners, policy-makers, and municipal authorities involved in waste management (WM) systems. This research aims at mapping the COVID-19-related scientific production to date in the field of WM. In this vein, the performance indicators of the target literature were analyzed and discussed through conducting a bibliometric analysis. The conceptual structure of COVID-19-related WM research, including seven main research themes, were uncovered and visualized through a text mining analysis as follows: (1) household and food waste, (2) personnel safety and training for waste handling, (3) sustainability and circular economy, (4) personal protective equipment and plastic waste, (5) healthcare waste management practices, (6) wastewater management, and (7) COVID-19 transmission through infectious waste. Finally, a research agenda for WM practices and activities in the post-COVID-19 era was proposed, focusing on the following three identified research gaps: (i) developing a systemic framework to properly manage the pandemic crisis implications for WM practices as a whole, following a systems thinking approach, (ii) building a circular economy model encompassing all activities from the design stage to the implementation stage, and (iii) proposing incentives to effectively involve informal sectors and local capacity in decentralizing municipal waste management, with a specific focus on developing and less-developed countries.

Diagnosis of SARS-CoV-2 during the Pandemic by Multiplex RT-rPCR hCoV Test: Future Perspectives.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has rapidly become a significant threat to public health. However, among the Coronaviridae family members, there are other viruses that can also cause infections in humans. Among these, severe acute respiratory syndrome (SARS-CoV) and Middle East respiratory syndrome (MERS-CoV) have posed significant threats to human health in the past. Other human pathogenic coronaviruses have been identified, and they are known to cause respiratory diseases with manifestations ranging from mild to severe. In this study, we evaluated the performance of a multiplex RT-rPCR specific to seven human pathogenic coronaviruses in mainly detecting SARS-CoV-2 directly from nasopharyngeal swabs obtained from suspected COVID-19 infected patients, while simultaneously detecting different human pathogenic coronaviruses in case these were also present. We tested 1195 clinical samples suspected of COVID-19 infection. The assay identified that 69% of the samples tested positive for SARS-CoV-2 (1195), which was confirmed using another SARS-CoV-2 RT-PCR kit available in our laboratory. None of these clinical samples were positive for SARS-CoV, MERS-CoV or HCoV. This means that during the endemic phase of COVID-19, infection with other human pathogenic coronaviruses, even the common cold coronavirus (HCoV), is very uncommon. Our study also confirmed that the multiplex RT-rPCR is a sensitive assay for detecting SARS-CoV-2 regardless of differences among the variants. This multiplex RT-rPCR is also time- and cost-saving and very easy to apply in the diagnostic laboratory due to its simple procedure and its stability in storage after preparation. These features make the assay a valuable approach in screening procedures for the rapid detection of SARS-CoV-2 and other human pathogenic coronaviruses that could affect public health.

An inclusive trend study of techno-economic analysis of biofuel supply chains.

Biofuels have gained much attention as a potentially sustainable alternative to fossil fuels to tackle climate change and energy scarcity. Hence, the increasing global interest in contributing to the biofuel supply chain (BSC), from biomass feedstock to biofuel production, has led to a huge amount of scientific production in recent years. In this vein, techno-economic analysis (TEA) of biofuel production to estimate total costs and revenues is highly important for transitioning towards a bioeconomy. This research aims to provide a comprehensive image of the body of knowledge in TEA evolution within the BSC domain. To this end, a systematic science mapping analysis, supported by a bibliometric analysis, is carried out on 1104 articles from 1986 to 2021. As a result, performance indicators of the scientific production within the target literature are presented to explain how this literature has evolved. Besides, thematic trends and conceptual structures of TEA of biofuel production are discovered. The results show that (i) biofuel production and consumption need promotion through tax measures and price subsidies, (ii) the development of cost-competitive algal biofuels has faced many challenges over recent years, and (iii) TEA of algal biofuels to identify commercial improvements and increase the economic feasibility is still lacking, which calls for more in-depth investigations. Consequently, current challenges and future perspectives of TEA in the BSC domain are rendered. The provided insights enable researchers and decision-makers involved in BSCs to (i) capture the most influential contributors to the field and (ii) identify major research hotspots and potential directions for further development.

Monkeypox Virus Infections in Southern Italy: Is There a Risk for Community Spread?

The ongoing outbreak of the Monkeypox virus (MPXV) is characterized by sustained human-to-human transmission, particularly among men who have sex with men (MSM). The aim of the study was to describe the characteristics of the MPXV infection identified in Southern Italy. Clinical samples for each suspected case identified from 1 June to 1 August 2022 were tested for MPXV, and whole-genome sequencing (WGS) was performed on two strains. Ten cases were identified: eight were young adult males, including six MSMs, and two were female. Nine subjects reported recent sexual exposure. One female subject without sexual exposure only reported attendance at a social gathering. Overall, 7 of 10 skin lesion samples had a high viral load of MPXV DNA, and 6/9 whole blood samples and 6/8 nasopharyngeal swab samples also tested positive. The analyzed sequences belonged to Clade 3, lineage B.1, and B.1.5, respectively. Despite this recent multinational outbreak of MPXV cases having revealed a high proportion of cases occurring among MSM, the identification of cases among heterosexual subjects and in a female subject without sexual risk factors should raise awareness among clinicians about the possible spread of MPXV in the general population.

Toxigenic Genes, Pathogenic Potential and Antimicrobial Resistance of Bacillus cereus Group Isolated from Ice Cream and Characterized by Whole Genome Sequencing.

Bacillus cereus is isolated from a variety of foods where it may cause food spoilage and/or food poisoning due to its toxigenic and pathogenic nature. In this study, we identified members of B. cereus groups in 65% of the ice cream samples analyzed, which were characterized based on multi locus variable number tandem repeats analysis (MLVA) and whole genome sequencing (WGS). The MLVA revealed that 36 strains showed different allelic profiles. Analyses of WGS data enabled the identification of three members of the B. cereus group: B. cereus sensu stricto, B. mosaicus and B. thuringiensis. Based on the multi locus sequence typing (MLST) scheme, the strains were classified in 27 sequence types (STs), including ST26 that causes food poisoning. Toxin genes' detection revealed the presence of the genes encoding nonhemolytic enterotoxin (NHE), hemolysin BL (HBL), cytotoxin K (cytK) and cereulide (ces) in 100%, 44%, 42% and 8% of the strains, respectively. The identification of eleven antimicrobial resistance (AMR) genes predicted the resistance to five different antimicrobials, and the resistance to beta-lactam antibiotics was confirmed with a phenotypic antimicrobial test. Taken together, the results showed that the B. cereus strains isolated from ice cream were a potential hazard for consumer safety.

An Outbreak of Human Systemic Anthrax, including One Case of Anthrax Meningitis, Occurred in Calabria Region (Italy): A Description of a Successful One Health Approach.

In this report, three cases of human cutaneous anthrax are described, one complicated by meningitis, and all were linked to a single infected bullock. A 41-year-old male truck driver, along with two male slaughterhouse workers, 45 and 42, were hospitalized for necrotic lesions of the arm associated with edema of the limb and high fever. All three patients were involved in transporting a bullock to the slaughterhouse. Microbiological examination on the prescapular lymph node and a piece of muscle from the bullock carcass showed the presence of Bacillus anthracis. The three patients underwent a biopsy of the affected tissues, and all samples tested positive for B. anthracis DNA using PCR. Furthermore, the truck driver also complained of an intense headache, and a CSF sampling was performed, showing him positive for B. anthracis by PCR, confirming the presumptive diagnosis of meningitis. Fast diagnosis and appropriate treatment are crucial for the management of human anthrax. Cooperation between human and veterinary medicine proved successful in diagnosing and resolving three human anthrax cases, confirming the reliability of the One Health approach for the surveillance of zoonoses.

Emerging Mutations Potentially Related to SARS-CoV-2 Immune Escape: The Case of a Long-Term Patient.

SARS-CoV-2 isolates from long-term COVID-19 patients play a significant role in understanding the mechanisms of infection and virus persistence. This study describes a SARS-CoV-2 isolate from a 53-year-old woman from Apulia (Italy), who was COVID-19 positive for approximately four months. In this paper we aimed to investigate any potential correlation between genetic mutations and clinical features of this case of infection. The viral isolate was assigned to lineage B.1.177.51 through whole-genome sequencing (WGS) and harbored a novel set of mutations on the Spike protein (V143D, del144/145 and E484K); furthermore, seroneutralization assays showed impaired response of the surveyed strain to BNT162b2 (Comirnaty) Pfizer/BioNTech vaccine-induced (average reduction of 70%) and convalescent sera (average reduction of 19.04%), when compared to VOC P.1. This study highlights the importance of genomic surveillance for the management of the COVID-19 pandemic, the relevance of monitoring of emerging SARS-CoV-2 mutations in all lineages, and the necessity of testing the response of emerging variants to available therapies and vaccines.

Three pillars of sustainability in the wake of COVID-19: A systematic review and future research agenda for sustainable development.

The COVID-19 pandemic has immensely impacted the economic, social, and environmental pillars of sustainability in human lives. Due to the scholars' increasing interest in responding to the urgent call for action against the pandemic, the literature of sustainability research considering COVID-19 consequences is very fragmented. Therefore, a comprehensive review of the COVID-19 implications for sustainability practices is still lacking. This research aims to analyze the effects of COVID-19 on the triple bottom line (TBL) of sustainability to support the future sustainable development agenda. To achieve that, the following research questions are addressed by conducting a systematic literature review: (i) what is the current status of research on the TBL of sustainability considering COVID-19 implications? (ii) how does COVID-19 affect the TBL of sustainability? and (iii) what are the potential research gaps and future research avenues for sustainable development post COVID-19? The results manifest the major implications of the COVID-19 outbreak for the triple sustainability pillars and the sustainable development agenda from the economic, social, and environmental points of view. The key findings provide inclusive insights for governments, authorities, practitioners, and policy-makers to alleviate the pandemic's negative impacts on sustainable development and to realize the sustainability transition opportunities post COVID-19. Finally, five research directions for sustainable development corresponding to the United Nations' sustainable development goals (SDGs) post COVID-19 are provided, as follows: (1) sustainability action plan considering COVID-19 implications: refining sustainability goals and targets and developing measurement framework; (2) making the most of sustainability transition opportunities in the wake of COVID-19: focus on SDG 12 and SDG 9; (3) innovative solutions for economic resilience towards sustainability post COVID-19: focus on SDG 1, SDG 8, and SDG 17; (4) in-depth analysis of the COVID-19 long-term effects on social sustainability: focus on SDG 4, SDG 5, and SDG 10; and (5) expanding quantitative research to harmonize the COVID-19-related sustainability research.

Energy efficiency and biological interactions define the core microbiome of deep oligotrophic groundwater.

While oligotrophic deep groundwaters host active microbes attuned to the low-end of the bioenergetics spectrum, the ecological constraints on microbial niches in these ecosystems and their consequences for microbiome convergence are unknown. Here, we provide a genome-resolved, integrated omics analysis comparing archaeal and bacterial communities in disconnected fracture fluids of the Fennoscandian Shield in Europe. Leveraging a dataset that combines metagenomes, single cell genomes, and metatranscriptomes, we show that groundwaters flowing in similar lithologies offer fixed niches that are occupied by a common core microbiome. Functional expression analysis highlights that these deep groundwater ecosystems foster diverse, yet cooperative communities adapted to this setting. We suggest that these communities stimulate cooperation by expression of functions related to ecological traits, such as aggregate or biofilm formation, while alleviating the burden on microorganisms producing compounds or functions that provide a collective benefit by facilitating reciprocal promiscuous metabolic partnerships with other members of the community. We hypothesize that an episodic lifestyle enabled by reversible bacteriostatic functions ensures the subsistence of the oligotrophic deep groundwater microbiome.

Salmonella enterica Subsp. houtenae Associated with an Abscess in Young Roe Deer (Capreolus capreolus).

BACKGROUND: S. enterica subsp. houtenae has been rarely documented, and very limited genomic information is available. This report describes a rare case of primary extraintestinal salmonellosis in a young roe deer, associated with Salmonella enterica subsp. houtenae. Methods: A traditional cultural-based analysis was carried out from the contents of a neck abscess; biochemical identification and PCR assay were performed to isolate and identify the pathogen. Through whole-genome sequencing (WGS), multilocus sequence typing (MLST), core genome MLST (cgMLST), and the Salmonella pathogenicity islands (SPIs) survey, resistome and virulome genes were investigated to gain insight into the virulence and antimicrobial resistance of S. houtenae. RESULTS: Biochemical identification and PCR confirmed the presence of Salmonella spp. in the swelling. The WGS analysis identified Salmonella enterica subspecies houtenae serovar 43:z4,z23:- and ST 958. The virulence study predicted a multidrug resistance pattern with resistance shown against aminoglycosides, tetracycline, beta-lactamase, fluoroquinolones, fosfomycin, nitroimidazole, aminocoumarin, and peptide. Fifty-three antibiotic-resistant genes were identified. No plasmids were detected. CONCLUSION: This study demonstrates the importance of continuous surveillance of pathogenic salmonellae. Biomolecular analyses combined with epidemiological data can provide important information about poorly described Salmonella strains and can help to improve animal welfare.

Recovery agenda for sustainable development post COVID-19 at the country level: developing a fuzzy action priority surface.

As a response to the urgent call for recovery actions against the COVID-19 crisis, this research aims to identify action priority areas post COVID-19 toward achieving the targets of the sustainable development goals (SDGs) within the 2030 Agenda for Sustainable Development launched by the United Nations (UN). This paper applies a mixed-method approach to map the post-COVID-19 SDGs targets on a fuzzy action priority surface at the country level in Iran, as a developing country, by taking the following four main steps: (1) using a modified Delphi method to make a list of the SDGs targets influenced by COVID-19; (2) using the best-worst method, as a multi-criteria decision-making tool, to weight the COVID-19 effects on the SDGs targets achievement; also (3) to weight the impact of the SDGs targets on the sustainable development implementation; and finally (4) designing a fuzzy inference system to calculate the action priority scores of the SDGs targets. As a result, reduction of poor people proportion by half (SDG 1.2), development-oriented policies for supporting creativity and job creation (SDG 8.3), end the pandemics and other epidemics (SDG 3.3), reduction of deaths and economic loss caused by disasters (SDG 11.5), and financial support for small-scale enterprises (SDG 9.3) were identified as the highest priorities for action, respectively, in the recovery agenda for sustainable development post COVID-19. The provided fuzzy action priority surface supports the UN's SDGs achievement and implementing the 2030 Agenda for Sustainable Development in Iran. It also serves as a guideline to help the government, stakeholders, and policy-makers better analyze the long-term effects of the pandemic on the SDGs and their associated targets and mitigate its adverse economic, social, and environmental consequences.

Genomic Surveillance of Circulating SARS-CoV-2 in South East Italy: A One-Year Retrospective Genetic Study.

In order to provide insights into the evolutionary and epidemiological viral dynamics during the current COVID-19 pandemic in South Eastern Italy, a total of 298 genomes of SARS-CoV-2 strains collected in the Apulia and Basilicata regions, between March 2020 and January 2021, were sequenced. The genomic analysis performed on the draft genomes allowed us to assign the genetic clades and lineages of belonging to each sample and provide an overview of the main circulating viral variants. Our data showed the spread in Apulia and Basilicata of SARS-CoV-2 variants which have emerged during the second wave of infections and are being currently monitored worldwide for their increased transmission rate and their possible impact on vaccines and therapies. These results emphasize the importance of genome sequencing for the epidemiological surveillance of the new SARS-CoV-2 variants' spread.

The Fennoscandian Shield deep terrestrial virosphere suggests slow motion 'boom and burst' cycles.

The deep biosphere contains members from all three domains of life along with viruses. Here we investigate the deep terrestrial virosphere by sequencing community nucleic acids from three groundwaters of contrasting chemistries, origins, and ages. These viromes constitute a highly unique community compared to other environmental viromes and sequenced viral isolates. Viral host prediction suggests that many of the viruses are associated with Firmicutes and Patescibacteria, a superphylum lacking previously described active viruses. RNA transcript-based activity implies viral predation in the shallower marine water-fed groundwater, while the deeper and more oligotrophic waters appear to be in 'metabolic standby'. Viral encoded antibiotic production and resistance systems suggest competition and antagonistic interactions. The data demonstrate a viral community with a wide range of predicted hosts that mediates nutrient recycling to support a higher microbial turnover than previously anticipated. This suggests the presence of 'kill-the-winner' oscillations creating slow motion 'boom and burst' cycles.

The Plant Health Monitoring System of the EDEN ISS Space Greenhouse in Antarctica During the 2018 Experiment Phase.

The EDEN ISS project has the objective to test key technologies and processes for higher plant cultivation with a focus on their application to long duration spaceflight. A mobile plant production facility was designed and constructed by an international consortium and deployed to the German Antarctic Neumayer Station III. Future astronaut crews, even if well-trained and provided with detailed procedures, cannot be expected to have the competencies needed to deal with all situations that will arise during a mission. Future space crews, as they are today, will be supported by expert backrooms on the ground. For future space-based greenhouses, monitoring the crops and the plant growth system increases system reliability and decreases the crew time required to maintain them. The EDEN ISS greenhouse incorporates a Plant Health Monitoring System to provide remote support for plant status assessment and early detection of plant stress or disease. The EDEN ISS greenhouse has the capability to automatically capture and distribute images from its suite of 32 high-definition color cameras. Collected images are transferred over a satellite link to the EDEN ISS Mission Control Center in Bremen and to project participants worldwide. Upon reception, automatic processing software analyzes the images for anomalies, evaluates crop performance, and predicts the days remaining until harvest of each crop tray. If anomalies or sub-optimal performance is detected, the image analysis system generates automatic warnings to the agronomist team who then discuss, communicate, or implement countermeasure options. A select number of Dual Wavelength Spectral Imagers have also been integrated into the facility for plant health monitoring to detect potential plant stress before it can be seen on the images taken by the high-definition color cameras. These imagers and processing approaches are derived from traditional space-based imaging techniques but permit new discoveries to be made in a facility like the EDEN ISS greenhouse in which, essentially, every photon of input and output can be controlled and studied. This paper presents a description of the EDEN ISS Plant Health Monitoring System, basic image analyses, and a summary of the results from the initial year of Antarctic operations.

Statistical Analysis of Community RNA Transcripts between Organic Carbon and Geogas-Fed Continental Deep Biosphere Groundwaters.

Life in water-filled bedrock fissures in the continental deep biosphere is broadly constrained by energy and nutrient availability. Although these communities are alive, robust studies comparing active populations and metabolic processes across deep aquifers are lacking. This study analyzed three oligotrophic Fennoscandian Shield groundwaters, two "modern marine" waters that are replenished with organic carbon from the Baltic Sea and are likely less than 20 years old (171.3 and 415.4 m below sea level) and an extremely oligotrophic "thoroughly mixed" water (448.8 m below sea level) of unknown age that is composed of very old saline and marine waters. Cells were captured either using a sampling device that rapidly fixed RNA under in situ conditions or by filtering flowing groundwater over an extended period before fixation. Comparison of metatranscriptomes between the methods showed statistically similar transcript profiles for the respective water types, and they were analyzed as biological replicates. Study of the small subunit (SSU) rRNA confirmed active populations from all three domains of life, with many potentially novel unclassified populations present. Statistically supported differences between communities included heterotrophic sulfate-reducing bacteria in the modern marine water at 171.3 m below sea level that has a higher organic carbon content than do largely autotrophic populations in the H2- and CO2-fed thoroughly mixed water. While this modern marine water had signatures of methanogenesis, syntrophic populations were predominantly in the thoroughly mixed water. The study provides a first statistical evaluation of differences in the active microbial communities in groundwaters differentially fed by organic carbon or "geogases."IMPORTANCE Despite being separated from the photosynthesis-driven surface by both distance and time, the deep biosphere is an important driver for the earth's carbon and energy cycles. However, due to the difficulties in gaining access and low cell numbers, robust statistical omics studies have not been carried out, and this limits the conclusions that can be drawn. This study benchmarks the use of two separate sampling systems and demonstrates that they provide statistically similar RNA transcript profiles, importantly validating several previously published studies. The generated data are analyzed to identify statistically valid differences in active microbial community members and metabolic processes. The results highlight contrasting taxa and growth strategies in the modern marine waters that are influenced by recent infiltration of Baltic Sea water versus the hydrogen- and carbon dioxide-fed, extremely oligotrophic, thoroughly mixed water.

VEGF, Microvessel Density, and CD44 as Inflammation Markers in Peri-implant Healthy Mucosa, Peri-implant Mucositis, and Peri-implantitis: Impact of Age, Smoking, PPD, and Obesity.

Several biologic processes affect the supporting peri-implant tissue leading to implant failure and complications, mainly referred to inflammation that is still poorly investigated in the peri-implant soft tissues. Our aim was to investigate in peri-implant healthy mucosa, peri-implant mucositis, and peri-implantitis the expression of some angiogenesis markers highly associated with inflammation, and evaluate its relationships with age, smoking, peri-implant pocket depth (PPD), and body max index (BMI). Moreover, we wanted to study the impact of these clinical parameters in the disease pathogenesis. Forty-eight total patients were recruited. Sixteen had at least one successfully osteointegrated dental implant (group A) and 32 had at least one osseointegrated implant in need of a peri-implant treatment for inflammatory/infectiveous reasons: precisely 16 for mucositis (group B) and 16 for peri-implantitis (group C). VEGF, CD34, and CD44 immunohistochemical expression was evaluated in the interproximal biopsies of marginal peri-implant tissue and correlated with the clinical parameters. A significant difference between groups in mean PPD was found, while the distribution by age, gender, smoking, and BMI resulted similar. Group C had significantly higher levels of VEGF, CD34, and CD44 expression compared to the other groups. VEGF, CD34, CD44, and peri-implant pocket depth were all positively correlated. Our study revealed that peri-implantitis is a condition characterized by unique and distinctive features. Our results supported that PPD has a great impact on the peri-implantitis and it is closely related to the inflammation marker expression. The identification of specific biomarkers might help in choosing distinct treatment approaches for target individuals.

A Novel Inorganic Sulfur Compound Metabolizing Ferroplasma-Like Population Is Suggested to Mediate Extracellular Electron Transfer.

Mining and processing of metal sulfide ores produces waters containing metals and inorganic sulfur compounds such as tetrathionate and thiosulfate. If released untreated, these sulfur compounds can be oxidized to generate highly acidic wastewaters [termed 'acid mine drainage (AMD)'] that cause severe environmental pollution. One potential method to remediate mining wastewaters is the maturing biotechnology of 'microbial fuel cells' that offers the sustainable removal of acid generating inorganic sulfur compounds alongside producing an electrical current. Microbial fuel cells exploit the ability of bacterial cells to transfer electrons to a mineral as the terminal electron acceptor during anaerobic respiration by replacing the mineral with a solid anode. In consequence, by substituting natural minerals with electrodes, microbial fuel cells also provide an excellent platform to understand environmental microbe-mineral interactions that are fundamental to element cycling. Previously, tetrathionate degradation coupled to the generation of an electrical current has been demonstrated and here we report a metagenomic and metatranscriptomic analysis of the microbial community. Reconstruction of inorganic sulfur compound metabolism suggested the substrate tetrathionate was metabolized by the Ferroplasma-like and Acidithiobacillus-like populations via multiple pathways. Characterized Ferroplasma species do not utilize inorganic sulfur compounds, suggesting a novel Ferroplasma-like population had been selected. Oxidation of intermediate sulfide, sulfur, thiosulfate, and adenylyl-sulfate released electrons and the extracellular electron transfer to the anode was suggested to be dominated by candidate soluble electron shuttles produced by the Ferroplasma-like population. However, as the soluble electron shuttle compounds also have alternative functions within the cell, it cannot be ruled out that acidophiles use novel, uncharacterized mechanisms to mediate extracellular electron transfer. Several populations within the community were suggested to metabolize intermediate inorganic sulfur compounds by multiple pathways, which highlights the potential for mutualistic or symbiotic relationships. This study provided the genetic base for acidophilic microbial fuel cells utilized for the remediation of inorganic sulfur compounds from AMD.

Metatranscriptomes Reveal That All Three Domains of Life Are Active but Are Dominated by Bacteria in the Fennoscandian Crystalline Granitic Continental Deep Biosphere.

The continental subsurface is suggested to contain a significant part of the earth's total biomass. However, due to the difficulty of sampling, the deep subsurface is still one of the least understood ecosystems. Therefore, microorganisms inhabiting this environment might profoundly influence the global nutrient and energy cycles. In this study, in situ fixed RNA transcripts from two deep continental groundwaters from the Äspö Hard Rock Laboratory (a Baltic Sea-influenced water with a residence time of <20 years, defined as "modern marine," and an "old saline" groundwater with a residence time of thousands of years) were subjected to metatranscriptome sequencing. Although small subunit (SSU) rRNA gene and mRNA transcripts aligned to all three domains of life, supporting activity within these community subsets, the data also suggested that the groundwaters were dominated by bacteria. Many of the SSU rRNA transcripts grouped within newly described candidate phyla or could not be mapped to known branches on the tree of life, suggesting that a large portion of the active biota in the deep biosphere remains unexplored. Despite the extremely oligotrophic conditions, mRNA transcripts revealed a diverse range of metabolic strategies that were carried out by multiple taxa in the modern marine water that is fed by organic carbon from the surface. In contrast, the carbon dioxide- and hydrogen-fed old saline water with a residence time of thousands of years predominantly showed the potential to carry out translation. This suggested these cells were active, but waiting until an energy source episodically becomes available.IMPORTANCE A newly designed sampling apparatus was used to fix RNA under in situ conditions in the deep continental biosphere and benchmarks a strategy for deep biosphere metatranscriptomic sequencing. This apparatus enabled the identification of active community members and the processes they carry out in this extremely oligotrophic environment. This work presents for the first time evidence of eukaryotic, archaeal, and bacterial activity in two deep subsurface crystalline rock groundwaters from the Äspö Hard Rock Laboratory with different depths and geochemical characteristics. The findings highlight differences between organic carbon-fed shallow communities and carbon dioxide- and hydrogen-fed old saline waters. In addition, the data reveal a large portion of uncharacterized microorganisms, as well as the important role of candidate phyla in the deep biosphere, but also the disparity in microbial diversity when using standard microbial 16S rRNA gene amplification versus the large unknown portion of the community identified with unbiased metatranscriptomes.

Microbial Community and Metabolic Activity in Thiocyanate Degrading Low Temperature Microbial Fuel Cells.

Thiocyanate is a toxic compound produced by the mining and metallurgy industries that needs to be remediated prior to its release into the environment. If the industry is situated at high altitudes or near the poles, economic factors require a low temperature treatment process. Microbial fuel cells are a developing technology that have the benefits of both removing such toxic compounds while recovering electrical energy. In this study, simultaneous thiocyanate degradation and electrical current generation was demonstrated and it was suggested that extracellular electron transfer to the anode occurred. Investigation of the microbial community by 16S rRNA metatranscriptome reads supported that the anode attached and planktonic anolyte consortia were dominated by a Thiobacillus-like population. Metatranscriptomic sequencing also suggested thiocyanate degradation primarily occurred via the 'cyanate' degradation pathway. The generated sulfide was metabolized via sulfite and ultimately to sulfate mediated by reverse dissimilatory sulfite reductase, APS reductase, and sulfate adenylyltransferase and the released electrons were potentially transferred to the anode via soluble electron shuttles. Finally, the ammonium from thiocyanate degradation was assimilated to glutamate as nitrogen source and carbon dioxide was fixed as carbon source. This study is one of the first to demonstrate a low temperature inorganic sulfur utilizing microbial fuel cell and the first to provide evidence for pathways of thiocyanate degradation coupled to electron transfer.