Development of a sequence-based in silico OspA typing method for Borrelia burgdorferi sensu lato.

Lyme disease (LD), caused by spirochete bacteria of the genus Borrelia burgdorferi sensu lato, remains the most common vector-borne disease in the northern hemisphere. Borrelia outer surface protein A (OspA) is an integral surface protein expressed during the tick cycle, and a validated vaccine target. There are at least 20 recognized Borrelia genospecies, that vary in OspA serotype. This study presents a new in silico sequence-based method for OspA typing using next-generation sequence data. Using a compiled database of over 400 Borrelia genomes encompassing the 4 most common disease-causing genospecies, we characterized OspA diversity in a manner that can accommodate existing and new OspA types and then defined boundaries for classification and assignment of OspA types based on the sequence similarity. To accommodate potential novel OspA types, we have developed a new nomenclature: OspA in silico type (IST). Beyond the ISTs that corresponded to existing OspA serotypes 1-8, we identified nine additional ISTs that cover new OspA variants in B. bavariensis (IST9-10), B. garinii (IST11-12), and other Borrelia genospecies (IST13-17). The IST typing scheme and associated OspA variants are available as part of the PubMLST Borrelia spp. database. Compared to traditional OspA serotyping methods, this new computational pipeline provides a more comprehensive and broadly applicable approach for characterization of OspA type and Borrelia genospecies to support vaccine development.

PfaSTer: a machine learning-powered serotype caller for Streptococcus pneumoniae genomes.

Streptococcus pneumoniae (pneumococcus) is a leading cause of morbidity and mortality worldwide. Although multi-valent pneumococcal vaccines have curbed the incidence of disease, their introduction has resulted in shifted serotype distributions that must be monitored. Whole genome sequence (WGS) data provide a powerful surveillance tool for tracking isolate serotypes, which can be determined from nucleotide sequence of the capsular polysaccharide biosynthetic operon (cps). Although software exists to predict serotypes from WGS data, most are constrained by requiring high-coverage next-generation sequencing reads. This can present a challenge in respect of accessibility and data sharing. Here we present PfaSTer, a machine learning-based method to identify 65 prevalent serotypes from assembled S. pneumoniae genome sequences. PfaSTer combines dimensionality reduction from k-mer analysis with a Random Forest classifier for rapid serotype prediction. By leveraging the model's built-in statistical framework, PfaSTer determines confidence in its predictions without the need for coverage-based assessments. We then demonstrate the robustness of this method, returning >97 % concordance when compared to biochemical results and other in silico serotyping tools. PfaSTer is open source and available at: https://github.com/pfizer-opensource/pfaster.

Effects of digestate-encapsulated biochar on plant growth, soil microbiome and nitrogen leaching.

The increasing amount of food waste and the excessive use of mineral fertilizers have caused detrimental impacts on soil, water, and air quality. Though digestate derived from food waste has been reported to partially replace fertilizer, its efficiency requires further improvement. In this study, the effects of digestate-encapsulated biochar were comprehensively investigated based on growth of an ornamental plant, soil characteristics, nutrient leaching and soil microbiome. Results showed that except for biochar, the tested fertilizers and soil additives, i.e., digestate, compost, commercial fertilizer, digestate-encapsulated biochar had positive effects on plants. Especially, the digestate-encapsulated biochar had the best effectiveness as evidenced by 9-25% increase in chlorophyll content index, fresh weight, leaf area and blossom frequency. For the effects of fertilizers or soil additives on soil characteristics and nutrient retention, the digestate-encapsulated biochar leached least N-nutrients (<8%), while the compost, digestate and mineral fertilizer leached up to 25% N-nutrients. All the treatments had minimal effects on the soil properties of pH and electrical conductivity. According to the microbial analysis, the digestate-encapsulated biochar has the comparable role with compost in improving the soil immune system against pathogen infection. The metagenomics coupling with qPCR analysis suggested that digestate-encapsulated biochar boosted the nitrification process and inhibited the denitrification process. This study provides an extensive understanding into the impacts of the digestate-encapsulated biochar on an ornamental plant and offers practical implications for the choice of sustainable fertilizers or soil additives and food-waste digestate management.

Genetic Surveillance of SARS-CoV-2 Mpro Reveals High Sequence and Structural Conservation Prior to the Introduction of Protease Inhibitor Paxlovid.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to represent a global health emergency as a highly transmissible, airborne virus. An important coronaviral drug target for treatment of COVID-19 is the conserved main protease (Mpro). Nirmatrelvir is a potent Mpro inhibitor and the antiviral component of Paxlovid. The significant viral sequencing effort during the ongoing COVID-19 pandemic represented a unique opportunity to assess potential nirmatrelvir escape mutations from emerging variants of SARS-CoV-2. To establish the baseline mutational landscape of Mpro prior to the introduction of Mpro inhibitors, Mpro sequences and its cleavage junction regions were retrieved from ~4,892,000 high-quality SARS-CoV-2 genomes in the open-access Global Initiative on Sharing Avian Influenza Data (GISAID) database. Any mutations identified from comparison to the reference sequence (Wuhan-Hu-1) were catalogued and analyzed. Mutations at sites key to nirmatrelvir binding and protease functionality (e.g., dimerization sites) were still rare. Structural comparison of Mpro also showed conservation of key nirmatrelvir contact residues across the extended Coronaviridae family (α-, ß-, and γ-coronaviruses). Additionally, we showed that over time, the SARS-CoV-2 Mpro enzyme remained under purifying selection and was highly conserved relative to the spike protein. Now, with the emergency use authorization (EUA) of Paxlovid and its expected widespread use across the globe, it is essential to continue large-scale genomic surveillance of SARS-CoV-2 Mpro evolution. This study establishes a robust analysis framework for monitoring emergent mutations in millions of virus isolates, with the goal of identifying potential resistance to present and future SARS-CoV-2 antivirals. IMPORTANCE The recent authorization of oral severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antivirals, such as Paxlovid, has ushered in a new era of the COVID-19 pandemic. The emergence of new variants, as well as the selective pressure imposed by antiviral drugs themselves, raises concern for potential escape mutations in key drug binding motifs. To determine the potential emergence of antiviral resistance in globally circulating isolates and its implications for the clinical response to the COVID-19 pandemic, sequencing of SARS-CoV-2 viral isolates before, during, and after the introduction of new antiviral treatments is critical. The infrastructure built herein for active genetic surveillance of Mpro evolution and emergent mutations will play an important role in assessing potential antiviral resistance as the pandemic progresses and Mpro inhibitors are introduced. We anticipate our framework to be the starting point in a larger effort for global monitoring of the SARS-CoV-2 Mpro mutational landscape.

Bioaugmentation of Methanosarcina thermophila grown on biochar particles during semi-continuous thermophilic food waste anaerobic digestion under two different bioaugmentation regimes.

This study presents the effect of bioaugmentation of thermophilic anaerobic digestion of food waste with Methanosarcina thermophila grown on a wood-derived biochar. Two different supplementation regimes were tested, namely a single bioaugmentation (SBABC) in which 10% v/v of the microbes grown on biochar (1 g/L) is added at setup of the reactors, versus a routine bioaugmentation (RBABC) wherein the same amount of supplements were added over 10 feeding cycles. The optimally performing 'R' and 'S' reactors had increased methane yields by 37% and 32% over their respective controls while reactors SBABC 2 and 3 produced 21.89% and 56.09% higher average methane yield than RBABC 2 and 3, respectively. It appears that a single dose bioaugmentation is advantageous for improving AD as analysed in terms of average methane yield and VFA production. This study provides the basis for understanding how biochar and bioaugmentation can be used for engineering sustainable pilot-scale AD processes.

Methanosarcina thermophila bioaugmentation and its synergy with biochar growth support particles versus polypropylene microplastics in thermophilic food waste anaerobic digestion.

Both biochar supplementation as well as bioaugmentation have been shown in literature to improve the methane yield of anaerobic digestion. In this study, the combination of both are evaluated by growing Methanosarcina thermophila on biochar support particles prior to augmentation of thermophilic food waste anaerobic digestion. Biochar stand alone, bioaugmentation solely, a combination of both added separately or grown together, and utilizing polypropylene (PP) microplastics as growth support instead were all tested when starting up a thermophilic process from mesophilic inoculum. Methanosarcina thermophila and biochar supplementation displayed synergy, with 5% M. thermophila on 1 g/L biochar presenting a 32% increase in specific methane yield over the control. Double the bioaugmentation dosage/concentration was also trialled with a thermophilic inoculum, and 10% M. thermophila grown on 2 g/L biochar displayed the best results with a 20% increase specific methane yield from its control standard.

Enhancing microbial lipids yield for biodiesel production by oleaginous yeast Lipomyces starkeyi fermentation: A review.

The enhanced production of microbial lipids suitable for manufacturing biodiesel from oleaginous yeast Lipomyces starkeyi is critically reviewed. Recent advances in several aspects involving the biosynthetic pathways of lipids, current conversion efficiencies using various carbon sources, intensification strategies for improving lipid yield and productivity in L. starkeyi fermentation, and lipid extraction approaches are analyzed from about 100 papers for the past decade. Key findings on strategies are summarized, including (1) optimization of parameters, (2) cascading two-stage systems, (3) metabolic engineering strategies, (4) mutagenesis followed by selection, and (5) co-cultivation of yeast and algae. The current technical limitations are analyzed. Research suggestions like examination of more gene targets via metabolic engineering are proposed. This is the first comprehensive review on the latest technical advances in strategies from the perspective of process and metabolic engineering to further increase the lipid yield and productivity from L. starkeyi fermentation.

Food-waste anaerobic digestate as a fertilizer: The agronomic properties of untreated digestate and biochar-filtered digestate residue.

Anaerobic digestion produces large quantities of digestate as a by-product, which can potentially be applied as an organic fertilizer, but untreated anaerobic digestate (AD) may contain phytotoxins and the large volume of AD makes transportation and storage difficult. This study explored two relatively inexpensive processing methods to improve the agronomic performance of AD as a fertilizer via vegetable cultivation experiments. We first investigated the effect of dilution on AD's performance using four leafy vegetables (Chinese spinach, water spinach, Chinese cabbage and lettuce). The optimal concentrations of the AD were 20-40% (v/v in 250 mL applications per single-plant pot) for all four vegetables based on shoot fresh weight and comparable to the control treatment using commercial fertilizer. AD application also introduced Synergistetes bacteria into the growing medium, but the overall bacterial diversity and composition were similar to those of the control treatment. Considering the nutrient separation in the liquid and solid fractions of AD and the need to reduce the volume, we then experimented with the recovery of nutrients from both the liquid and solid fractions by filtering AD using two types of wood-based biochar (100 g biochar: 1 L AD) before applying the AD-biochar residues as side dressing at 1% (w/w). Both types of biochar achieved yields comparable to the treatment using a commercial fertilizer for the three vegetables tested (kale, lettuce and rocket salad). Our results show that dilution and biochar filtration can improve the agronomic performance of AD, making it a sustainable substitute for commercial fertilizer.

Timing of biochar dosage for anaerobic digestion treating municipal leachate: Altered conversion pathways of volatile fatty acids.

In this study, the anaerobic digestion (AD) applications of early & late biochar dosage were compared for municipal leachate treatment, with the objective of studying the flexible use of biochar as a mitigation measure for biomethane recovery. In two experimental phases, biochar was favourable for the immediate promotion of AD performances, as revealed by Gompertz's model of reduced lag phases, higher biomethane generation rates, and increased biomethane yields. Irrespective of late biochar dosage, it could still retrieve 89% of the ultimate biomethane potential. Comparing the residual VFAs (volatile fatty acids) compositions, it was found that the fraction of long-chain VFAs accounted for 81% of total VFAs in reactor set of early biochar dosage, while it was only 38% in the reactor of late one. Parallel evidence suggested that the schedule of biochar dosage not only could affect methanogenic responses but also the VFAs conversion pathways.

Biochar utilisation in the anaerobic digestion of food waste for the creation of a circular economy via biogas upgrading and digestate treatment.

A wood waste-derived biochar was applied to food-waste anaerobic digestion to evaluate the feasibility of its utilisation to create a circular economy. This biochar was first purposed for the upgrading of the biogas from the said anaerobic digestion, before treating and recovering the nutrients in the solid fraction of the digestate, which was finally employed as a biofertilizer for the organic cultivation of three green leafy vegetables: kale, lettuce and rocket salad. Whilst the amount of CO2 the biochar could absorb from the biogas was low (11.17 mg g-1), it could potentially be increased by modifying through physical and chemical methods. Virgin as well as CO2-laden biochar were able to remove around 31% of chemical oxygen demand, 8% of the ammonia and almost 90% of the total suspended solids from the digestate wastewater, which was better than a dewatering process via centrifugation but worse than the industry standard of a polytetrafluoroethylene membrane bioreactor. Nutrients were recovered in the solid fraction of the digestate residue filtered by the biochar, and utilised as a biofertilizer that performed similarly to a commercial complete fertilizer in terms of aerial fresh weight growth for all three vegetables cultivated. Contingent on the optimal upgrading of biogas, the concept of a circular economy based on biochar and anaerobic digestion appears to be feasible.

Influence of wet oxidation pretreatment with hydrogen peroxide and addition of clarified manure on anaerobic digestion of oil palm empty fruit bunches.

Food and energy requirements are increasing globally, and the challenge is to meet these demands in a sustainable manner. Oil palm has a relatively high productivity, but produces the lignocellulosic residue of empty fruit bunches (OPEFB). In this study, wet oxidation pretreatment is utilized to overcome the recalcitrance of OPEFB during semi-continuous anaerobic digestion (AD) with between 19.7 and 52.7% improvement over the control, and near total cellulose and hemicellulose content could be degraded. Clarified manure, the water phase of cattle and dairy manure after filtration, is further tested for its effect on methane production by providing necessary micronutrients and vitamins. An increase of 49% was found after addition of clarified manure to OPEFB compared to without this addition.

Closing the food waste loop: Food waste anaerobic digestate as fertilizer for the cultivation of the leafy vegetable, xiao bai cai (Brassica rapa).

The increasing world population necessitates the production of larger amounts of food in a safe and environmentally sustainable manner, while concomitantly managing an increasing amount of food waste similarly. These needs can theoretically be met by the recycling of the nutrients in food waste via anaerobic digestion, which also produces renewable energy. This hypothesis is proven by the growing of a commonly consumed leafy vegetable, xiao bai cai (Brassica rapa), by the addition of food waste anaerobic digestate in place of commercial fertilizer. Different concentrations of the digestate were tested, as well as different heat treatments to simulate hygienization, and the results for most part (aerial fresh weight, dry weight, chlorophyll content) are not significantly different from growth utilizing commercial inorganic 15:15:15 NPK fertilizer. Microbial analysis of the growth media was also carried out to explicate digestate effects and to show that some common foodborne disease pathogens were not detected.

The Safety of Continuing Therapeutic Anticoagulation During Inferior Vena Cava Filter Retrieval: A 6-Year Retrospective Review from a Tertiary Centre.

PURPOSE: Assess the safety of inferior vena cava (IVC) filter retrieval in patients taking anticoagulation, compared to a non-anticoagulated cohort. MATERIALS AND METHODS: Single-centre retrospective analysis of patients who underwent IVC filter retrieval between January 2012 and February 2018. Information about patient demographics, anticoagulation, tilt, major and minor complications was collected. Major complications were defined as: IVC injury from the filter retrieval, retained fragment of filter, filter fracture and filter embolisation. Minor complications were defined as: neck haematoma and puncture site infection. RESULTS: Total of 357 patients (age 18-95, Male: 231) underwent IVC filter retrieval, comprising of Cook Celect Platinum, Cook Celect, and ALN-branded filters. Of these 182 patients were on anticoagulation and 175 patients were not on anticoagulation, based on the indication for the filter (thrombosis or prophylaxis) and at the discretion of the referring unit who were managing the anticoagulation. IVC filter retrieval was technically successful in 349 patients. Five major complications (1.4% of retrievals) were recorded and no minor complications (0% of retrievals). In the anticoagulation cohort, there were two major complications (1.1% of retrievals) both related to IVC injury. In the non-anticoagulated cohort, there were three major complications (1.7% of retrievals) relating to filter embolisation, IVC injury, and filter fracture. CONCLUSIONS: IVC filter retrieval is a safe procedure with a low complication rate. Being on anticoagulation does not increase the risk of a major complication or change the management of major complication compared with a non-anticoagulated cohort. IVC filter retrieval is safe to perform in patients currently taking prophylactic or therapeutic anticoagulation based on our cohort. LEVEL OF EVIDENCE: Level 3, retrospective cohort study.

Layers of Cryptic Genetic Variation Underlie a Yeast Complex Trait.

Cryptic genetic variation may be an important contributor to heritable traits, but its extent and regulation are not fully understood. Here, we investigate the cryptic genetic variation underlying a Saccharomyces cerevisiae colony phenotype that is typically suppressed in a cross of the laboratory strain BY4716 (BY) and a derivative of the clinical isolate 322134S (3S). To do this, we comprehensively dissect the trait's genetic basis in the BYx3S cross in the presence of three different genetic perturbations that enable its expression. This allows us to detect and compare the specific loci that interact with each perturbation to produce the trait. In total, we identify 21 loci, all but one of which interact with just a subset of the perturbations. Beyond impacting which loci contribute to the trait, the genetic perturbations also alter the extent of additivity, epistasis, and genotype-environment interaction among the detected loci. Additionally, we show that the single locus interacting with all three perturbations corresponds to the coding region of the cell surface gene FLO11 While nearly all of the other remaining loci influence FLO11 transcription in cis or trans, the perturbations tend to interact with loci in different pathways and subpathways. Our work shows how layers of cryptic genetic variation can influence complex traits. Here, these layers mainly represent different regulatory inputs into the transcription of a single key gene.

CD90 Identifies Adventitial Mesenchymal Progenitor Cells in Adult Human Medium- and Large-Sized Arteries.

Mesenchymal stem cells (MSCs) reportedly exist in a vascular niche occupying the outer adventitial layer. However, these cells have not been well characterized in vivo in medium- and large-sized arteries in humans, and their potential pathological role is unknown. To address this, healthy and diseased arterial tissues were obtained as surplus surgical specimens and freshly processed. We identified that CD90 marks a rare adventitial population that co-expresses MSC markers including PDGFRα, CD44, CD73, and CD105. However, unlike CD90, these additional markers were widely expressed by other cells. Human adventitial CD90+ cells fulfilled standard MSC criteria, including plastic adherence, spindle morphology, passage ability, colony formation, and differentiation into adipocytes, osteoblasts, and chondrocytes. Phenotypic and transcriptomic profiling, as well as adoptive transfer experiments, revealed a potential role in vascular disease pathogenesis, with the transcriptomic disease signature of these cells being represented in an aortic regulatory gene network that is operative in atherosclerosis.

Environmental impact comparison of four options to treat the cellulosic fraction of municipal solid waste (CF-MSW) in green megacities.

Megacities are characterized by a high urban population density, and timely solid waste management is crucial for the wellbeing of its citizens. In this study, the cellulosic fraction of municipal solid waste is the target for management in Singapore (which has one of the highest population densities in the world) using four pertinent options - incineration, anaerobic digestion, gasification and composting. The energy sustainability and environmental impacts in the form of greenhouse gas emissions are considered for each scenario together with the status quo to achieve the most favorable recommendation, with a sensitivity analysis in the case that no bio-fertilizers are allowed for use locally. In terms of GWP and energy profile for grass waste, AD is the most environmentally friendly option (2 kilo tonnes of CO2-eq and 23.3 GWh respectively), while for leaf waste, gasification is the best, with AD a close second.

Acclimatization of a mixed-animal manure inoculum to the anaerobic digestion of Axonopus compressus reveals the putative importance of Mesotoga infera and Methanosaeta concilii as elucidated by DGGE and Illumina MiSeq.

In this study, a multifarious microbial mix from different sources is acclimatized over a period of three months to digesting cowgrass, and the changes in the community structure are examined with both a traditional denaturing gradient gel electrophoresis method as well as a next generation sequencing MiSeq method. It is shown that the much more in depth analysis by Illumina gives more information about the relative abundance and thus putative importance of the role of various microbes, in particular the bacterium Mesotoga infera and the archaeon Methanosaeta concilii.

Genetic suppression: Extending our knowledge from lab experiments to natural populations.

Many mutations have deleterious phenotypic effects that can be alleviated by suppressor mutations elsewhere in the genome. High-throughput approaches have facilitated the large-scale identification of these suppressors and have helped shed light on core functional mechanisms that give rise to suppression. Following reports that suppression occurs naturally within species, it is important to determine how our understanding of this phenomenon based on lab experiments extends to genetically diverse natural populations. Although suppression is typically mediated by individual genetic changes in lab experiments, recent studies have shown that suppression in natural populations can involve combinations of genetic variants. This difference in complexity suggests that sets of variants can exhibit similar functional effects to individual suppressors found in lab experiments. In this review, we discuss how characterizing the way in which these variants jointly lead to suppression could provide important insights into the genotype-phenotype map that are relevant to evolution and health.