Biology subjects

Boon, N.

Publications and source records attributed to Boon, N..

At least 19 recordsLinked to original sources

bioRxiv2026

Gene Supplementation of MYO7A or activation of Myo7b for treatment of Usher syndrome 1B

Mutations in MYO7A result in the most severe subtype of Usher syndrome, the leading genetic cause of deafblindness. The large size of MYO7A requires dual adeno-associated virus (AAV) vectors for gene transfer or alternative methods to treat retinal defects. Here, we evaluated two treatment approaches: i) Supplementation of the human MYO7A gene via dual mRNA trans-splicing AAVs, and ii) CRISPR/Cas-mediated activation of the related murine Myo7b gene. Upon MYO7A supplementation, the transgenic MYO7A transcript and protein were expressed and correctly localized in retinal pigment epithelial (RPE) and photoreceptors of mice, pigs, and human retinal organoids. In RPE-and photoreceptor-specific Myo7a knockout mice, we could restore MYO7A expression and localization of melanosomes in RPE cells to wild-type levels. Myo7b activation led to partial restoration of melanosome localization, and the localization of MYO7B protein was largely comparable to MYO7A. These findings indicate that both approaches are in principle suitable for the therapy of Usher syndrome.

molecular biology↗

bioRxiv2026

Sympathetic innervation regulates metabolic flexibility of skeletal muscle

The sympathetic nervous system (SNS) is recognized for its role in the physiological regulation of organs, such as heart, vasculature and lungs, and has emerged as a potential player in skeletal muscle metabolic and neuromuscular junction (NMJ) health. However, the mechanism through which SNS signaling influences skeletal muscle function and adaptation to exercise remains unclear. Using molecular, electrophysiological, immunohistochemical, and high-resolution respirometry techniques, we tested the role of sympathetic innervation to skeletal muscle in response to exercise. Our findings reveal that sympathetic denervation disrupts the NMJ, reducing motor and sympathetic receptor expression, with concomitant deficits in skeletal muscle function. Mechanistically, these deficits are linked to diminished CPT1 enzyme activity, which impairs long-chain fatty acid-mediated oxidation in skeletal muscle mitochondria. These findings reveal a key role for sympathetic innervation in maintaining mitochondrial metabolic function and by extension, skeletal muscle performance, offering novel insight into the interplay between the SNS, exercise, and muscle mitochondria.

cell biology↗

bioRxiv2025

An optimized flow cytometry sorting-sequencing workflow reduces storage, sorting and extraction bias in sorted microbial communities

Fluorescence-Activated Cell Sorting (FACS) followed by metataxonomics is commonly used to probe the composition of microbial subpopulations with a functionality of interest. Metataxonomic analysis of sorted low microbial biomass samples is prone to contamination and bias introduced during sorting and DNA extraction. To improve metataxonomic accuracy, the DNA yield obtained by the DNeasy(R) PowerSoil(R) Pro kit was first maximized. FACS-sequencing using the optimized DNA extraction method distorted the composition of sorted microbiota by introducing sheath fluid contaminants, termed the FACSome. Carefully selected controls allowed for FACSome characterization and subsequent in silico decontamination of PacBio and Illumina metataxonomic sequencing data. The optimized extraction and decontamination were validated in soil, water, feces, saliva, gut reactor and mock microbial communities. Metataxonomic profiles of non-selectively sorted and unsorted samples derived from these ecosystems did not perfectly match due to relic DNA depletion during FACS. A nuclease-based digest enhanced the depletion of spiked and naturally present relic DNA, which can interfere with function-driven FACS. Since FACS depleted relic DNA, the preservation of intact cells prior to sorting is essential for activity- or viability-targeted FACS. Intact cells were best preserved during long-term -80{degrees}C storage of fecal samples that were not amended with glycerol-1X Tris-EDTA cryoprotectant.

microbiology↗

bioRxiv2025

Isolation and Characterization of Strains used in Bacterial-Based Strategies for Accelerated Carbonation of Lime Mortars

Over the last century, lime has been quickly replaced by the uptake of Portland cement, mainly due to its faster hardening. Achieving earlier hardening in lime through faster carbonation is thus essential to help overcome one of limes limiting qualities. In this work, we isolated and selected strains suitable for use in lime mortars, and used bacterial suspensions to carbonate lime materials. An isolation campaign from a lime mortar wall returned two alkaliphilic isolates, Shouchella clausii and Shouchella patagoniensis. S. clausii was then further adapted to high pH (> 11) by adaptive laboratory evolution to produce a third strain. All three strains were then followed for a period of 14 days in serum bottles at pH 11 and gas composition of the headspace, intact/damaged cell populations and pH were measured. In parallel, lime mortar samples were incubated in a closed environment with bacterial suspension of the strains. The mortars were then tested at 7 and 14 days with thermogravimetric analysis to study the amount of carbonation through bacterial activity. Overall, S. patagoniensis produced more CO2, close to the estimated maximum CO2 uptake rate of lime, and carbonated the lime mortars faster and to a larger extent than the other strains. Finally, the bacterial suspensions were directly mixed with lime and the carbonation front was followed using analytical techniques. Once again S. patagoniensis led to faster carbonation. Overall, it was shown that bacterial-based strategies for accelerated carbonation of lime are a possibility despite the high alkalinity of the binder. ImportancePortland cement is the dominant binder used in most construction today, but until last century, lime was the ubiquitous construction material. The increase in use of cement has sprung from its higher strength and faster hardening, yet, lime still remain a relevant material, particularly in masonry structures and the built heritage. As such, novel lime materials are necessary to tackle some of the current limitations of lime, such as earlier hardening, which would not only make lime easier to work with but would also limit failure due to environmental conditions. As existing strategies to speed-up lime hardening have had limited uptake due to their reliance on expensive and often toxic chemicals, the need for novel solutions is in place. We show that bacterial-based strategies can help achieve a bio-based solution to go beyond the limitations of current strategies and open up new possibilities for microbial bioengineering in construction materials.

bioengineering↗

bioRxiv2025

Organics Guide Non-Classical Crystallization Of Bacterial Calcite Which Parallels Eukaryotic Biomineralization

Microbially-induced calcium carbonate precipitation is a widespread natural phenomenon with numerous technical applications. Recent advances have shown that bacterial calcium carbonates (BCC) form non-classically via amorphous calcium carbonate (ACC) precursors in the presence of organics, but the role of organics in the formation and nanostructural features of BCCs is not fully understood. Here we show that two bacterial strains produce BCCs with diverse textural and structural features at the macroscale but similar at the micro and nanoscale. We show that bacterial organics guide precipitation of calcite, stabilizing ACC to produce nanogranular crystals and these organics are then trapped within the crystal, rather than being released as previously suggested. These organics are N-rich and create regions of low Z-contrast aligned perpendicular to the c-axis of the crystal, yielding a "Swiss cheese-like" mesostructure. Moreover, it is these occluded organics that lead to the distinctive biosignatures observed in BCC. Finally, we also observe crystalline 2D-films, possibly proteins, templating the oriented crystallization of calcite. These ultrastructural features help to disclose how microbial CaCO3 biomineralization takes place leading to improved technical applications and may provide fingerprints for their identification in nature.

microbiology↗

bioRxiv2025

Impact of flushing procedures on drinking water biostability and invasion susceptibility in distribution systems

Maintaining microbial stability in drinking water distribution systems (DWDS) is a priority for water providers, yet operational events like flushing and the potential invasion of unwanted microorganisms pose significant challenges. This study utilized a pilot-scale DWDS to evaluate the susceptibility of drinking water networks after standard operational flushing procedures (with and without chlorination) as these procedures can alter biostability. Results showed that flushing with chlorination reduced bulk cell counts initially but led to subsequent microbial regrowth, highlighting the limitations of chlorination in long-term biostability. Biofilm cell densities and community composition were similar before and after flushing with and without chlorination. Additionally, the susceptibility of a flushed network against introduced bacterial indicators (Aeromonas media, Pseudomonas putida, and Serratia fonticola) was investigated. The invasion experiments revealed that the species decayed more rapidly in loops flushed with chlorination. This study emphasizes the trade-off between potential regrowth and microbial control, underscoring the need for strategies that enhance microbial resilience without fostering regrowth. These findings are critical for optimizing DWDS management practices to ensure drinking water safety and quality. ImportanceFlushing, with or without chlorination, is a standard procedure used by drinking water providers to address water quality issues such as brown water, as well as following waterworks, or contaminant detection. This process involves discharging large volumes of drinking water, often through fire hydrants. Depending on the contamination, additional disinfectants, such as free chlorine, are dosed during flushing. However, the longer-term effects of these processes on microbial dynamics are unknown. For example, while brown water issues can be solved, the overall biostability of drinking water may be compromised, potentially making the drinking water distribution systems more susceptible to the invasion of unwanted microorganisms. Studying these practices in full-scale distribution networks is challenging. Therefore, in this study, a unique drinking water distribution pilot system that mimics a real network with mature biofilm was used to investigate the changes in both bulk water and biofilm microbiomes following flushing procedures.

microbiology↗

bioRxiv2025

Interindividual differences in caries potential of the salivary microbiome after mouthwash exposure

Prevention and treatment of dental caries is often achieved by fluoride mouthwashes. The most common forms of fluoride are Stannous fluoride (SnF2), amine fluoride (AmF) and sodium fluoride (NaF), each with differing activities against oral bacteria. Additionally, differences in oral microbiome between individuals are linked to differences in response to treatment and the phenotype of the microbiome can provide information on this response. Flow cytometry can successfully track microbial phenotypic heterogeneity and may prove useful for capturing the response to treatment. In this study, we compared the effect of three fluoride-containing mouthwashes, Elmex Anti Caries (AmF/NaF), Listerine Anticaries (NaF) and Meridol (AmF/SnF2), on the salivary microbiome and assessed their anticaries potential. Furthermore, we investigated if the flow cytometric fingerprint could be used to capture the response to treatment. Saliva samples of different donors were treated with the mouthwashes and the intact salivary community composition was assessed using 16S rRNA gene amplicon sequencing. Treated saliva samples were incubated in sucrose-rich medium to assess the organic acid production of the remaining microbiota. Moreover, flow cytometric fingerprints of the intact microbial population were used to track the response to treatment and to train a random forest classifier predicting the most suitable mouthwash for the donor. We found a mouthwash-dependent shift in the salivary microbiome composition and found different anticaries potential of the mouthwashes. Meridol affected more bacterial taxa and had the best overall anticaries potential, followed by Elmex. Finally, the response to treatment could be captured in the flow cytometric fingerprint of the initial intact salivary microbiota and a random forest classifier predicting the most suitable mouthwash for the donor was successfully trained.

microbiology↗

bioRxiv2024

Molybdate application in the early stages of shrimp growth suppresses sulphide formation in a shrimp pond bottom model

Oxygen depletion and sulphide formation, resulting from the accumulation of organic waste, are common challenges in shrimp ponds that could result in complete harvest failure. The stage at which these circumstances occur during the shrimp growth period remains elusive, yet, knowledge of the timing of oxygen depletion and sulphide formation is essential to enable remediating actions. Here, we used an experimental shrimp pond model at different stages in the shrimp growth period to determine when oxygen depletion and sulphide production occur. Microscale depth measurements of oxygen and H2S were determined using microelectrodes to visualize their profiles at different depths of the water-sediment interface and the sediment. We evaluated the potential of different molybdate concentrations at different stages to determine the optimal conditions to suppress H2S formation. Oxygen depletion and sulphide production took place in the middle of the shrimp growth cycle in the simulated model of waste accumulation. The addition of molybdate was only effective in the early stages of the onset of oxygen depletion and H2S formation, and residual molybdate was required to ensure a continuous suppression of sulphide production. However, oxygen depletion could not be prevented and reintroduction of oxygen did not occur when molybdate was added. In conclusion, molybdate appeared to be an effective strategy to suppress H2S formation at the onset of its production in a shrimp pond bottom model.

microbiology↗

bioRxiv2024

Quantifying synthetic bacterial community composition with flow cytometry: efficacy in mock communities and challenges in co-cultures

Determination of bacterial community composition in synthetic communities is critical for understanding microbial systems. The community composition is typically determined through bacterial plating or through PCR-based methods which can be labor-intensive, expensive or prone to bias. Simultaneously, flow cytometry has been suggested as a cheap and fast alternative. However, since the technique captures the phenotypic state of bacterial cells, accurate determination of community composition could be affected when bacteria are co-cultured. We investigated the performance of flow cytometry for quantifying oral synthetic communities and compared it to the performance of strain specific qPCR and 16S rRNA gene amplicon sequencing. Therefore, axenic cultures, mock communities and co-cultures of oral bacteria were prepared. Random forest classifiers trained on flow cytometry data of axenic cultures were used to determine the composition of the synthetic communities, as well as strain specific qPCR and 16S rRNA gene amplicon sequencing. Flow cytometry was shown to have a lower average root mean squared error and outperformed the PCR-based methods in even mock communities (flow cytometry: 0.11 {+/-} 0.04; qPCR: 0.26 {+/-} 0.09; amplicon sequencing: 0.15 {+/-} 0.01). When bacteria were co-cultured, neither flow cytometry, strain specific qPCR and 16S rRNA gene amplicon sequencing resulted in similar community composition. Performance of flow cytometry was decreased compared to mock communities due to changing phenotypes. Finally, discrepancies between flow cytometry and strain specific qPCR were found. These findings highlight the challenges ahead for quantifying community composition in co-cultures by flow cytometry. ImportanceQuantification of bacterial composition in synthetic communities is crucial for understanding and steering microbial interactions. Traditional approaches like plating, strain specific qPCR and amplicon sequencing are often labor-intensive and expensive and limit high-throughput experiments. Recently, flow cytometry has been suggested as a swift and cheap alternative for quantifying communities and has been successfully demonstrated on simple bacterial mock communities. However, since flow cytometry measures the phenotypic state of cells, measurements can be affected by differing phenotypes. Especially changing phenotypes resulting from co-culturing bacteria can have a profound effect on the applicability of the technique in this context. This research illustrates the feasibility and challenges of flow cytometry for the determination of community structure in synthetic mock communities and co-cultures.

microbiology↗

bioRxiv2023

Long-term sulphide mitigation through molybdate at shrimp pond bottoms

Shrimp are commonly cultured in earthen aquaculture ponds where organic-rich uneaten feed and faeces accumulate on and in the sediment to form anaerobic zones. Since the pond water is rich in sulphate, these anaerobic conditions eventually lead to the production of sulphide. Sulphides are toxic and even lethal to the shrimp that live on the pond sediment, but physicochemical and microbial reactions that occur during the accumulation of organic waste and the subsequent formation of sulphide in shrimp pond sediments remain unclear. Molybdate treatment is a promising strategy to inhibit sulphate reduction, thus, preventing sulphide accumulation. We used an experimental shrimp pond model to simulate the organic waste accumulation and sulphide formation during the final 61 days of a full shrimp growth cycle. Sodium molybdate (5 and 25 mg/L Na2MoO4.2H2O) was applied as a preventive strategy to control sulphide production before oxygen depletion. Molybdate addition partially mitigated H2S production in the sediment, and delayed its transfer to the bulk liquid by pushing the higher sulphide concentration zone towards deeper sediment layers. Molybdate treatment at 25 mg/L significantly impacted the overall microbial community composition and treated samples (5 and 25 mg/L molybdate) had about 50% higher relative abundance of sulphate reducing bacteria than the control (no molybdate) treatment. In conclusion, molybdate has the potential to work as mitigation strategy against sulphide accumulation in the sediment during shrimp growth by directly steering the microbial community in a shrimp pond system.

bioengineering↗

bioRxiv2023

Unlocking the Mechanism of Action: A Cost-Effective Flow Cytometry Approach for Accelerating Antimicrobial Drug Development

Antimicrobial resistance is one of the greatest challenges to global health. While the development of new antimicrobials can combat resistance, low profitability is reducing the number of new compounds brought to the market. Elucidating the mechanism of action is crucial for developing new antimicrobials. This process can become expensive as there are no universally applicable pipelines and scientific expertise in different fields is required. One way to determine the mechanism of action is the use of predictive modeling, as antimicrobials can be classified into limited groups.. We demonstrate a cost-effective flow cytometry approach for determining the mechanisms of action of new compounds. Cultures of Actinomyces viscosus and Fusobacterium nucleatum were treated with a range of antimicrobials and measured by flow cytometry. A Gaussian mixture mask was applied over the data to construct a phenotypic fingerprint. The fingerprints were used to train random forest classifiers, and classifiers were used to predict the mechanism of action of cephalothin. Significant statistical differences were found among the 10 different treatment groups. A pairwise comparison between treatment groups showed a statistical difference for 35 out of 45 pairs for Actinomyces viscosus and 32 out of 45 pairs for Fusobacterium nucleatum after 3.5h of treatment. The best performing random forest classifier yielded a Matthews correlation coefficient of 0.92 and the mechanism of action of cephalothin could be successfully predicted. These findings suggest that flow cytometry can be a cheap and fast alternative for determining the mechanism of action of new antimicrobials.

microbiology↗

bioRxiv2023

Unwanted coliforms can hide in mature drinking water biofilms, grown in full-scale distribution networks

Biofilms within drinking water distribution systems serve as a habitat for drinking water microorganisms. However, biofilms can negatively impact drinking water quality by causing water discoloration and deterioration and can be a reservoir for unwanted microorganisms. In this study, we investigated whether indicator organisms for drinking water quality, such as coliforms, can settle in mature drinking water biofilms. Therefore, a biofilm monitor consisting of glass rings was used to grow and sample drinking water biofilms. Two mature drinking water biofilms were characterized by flow cytometry, ATP measurements, confocal laser scanning microscopy and 16S rRNA sequencing. Biofilms developed under treated chlorinated surface water supply exhibited lower cell densities in comparison with biofilms resulting from treated groundwater. Overall, the phenotypic as well as the genotypic characteristics were significantly different between both biofilms. In addition, the response of the biofilm microbiome and possible biofilm detachment after minor water quality changes were investigated. Limited changes in pH and free chlorine addition, to simulate operational changes that are relevant for practice, were evaluated. It was shown that both biofilms remained resilient. Finally, mature biofilms were prone to invasion of the coliform, Serratia fonticola. After spiking low concentrations (i.e. {+/-} 100 cells/100 mL) of the coliform to the corresponding bulk water samples, the coliforms were able to attach and get established within the mature biofilms. These outcomes are emphasizing the need for continued research on biofilm detachment and its implications for water contamination in distribution networks. ImportanceThe revelation that even low concentrations of coliforms can infiltrate into mature drinking water biofilms highlights a potential public health concern. Nowadays, the measurement of coliform bacteria is used as an indicator for fecal contamination and to control the effectiveness of disinfection processes and the cleanliness and integrity of distribution systems. In Flanders (Belgium), 533 out of 18840 measurements exceeded the established norm for the coliform indicator parameter in 2021, however, the source of microbial contamination is mostly unknown. Here, we showed that mature biofilms, are susceptible to invasion of Serratia fonticola. These findings emphasize the importance of understanding and managing biofilms in drinking water distribution systems, not only for their potential to influence water quality, but also for their role in harboring and potentially disseminating pathogens. Further research into biofilm detachment, long-term responses to operational changes, and pathogen persistence within biofilms is crucial to inform strategies for safeguarding drinking water quality.

microbiology↗

bioRxiv2023

mRNA trans-splicing dual AAV vectors for (epi)genome editing and gene therapy

Large genes including several CRISPR-Cas modules, such as gene activators (CRISPRa), require dual adeno-associated viral (AAV) vectors for an efficient in vivo delivery and expression. Current dual AAV vector approaches have important limitations, e.g., low reconstitution efficiency, production of alien proteins, or low flexibility in split site selection. Here, we present a dual AAV vector technology based on reconstitution via mRNA trans-splicing (REVeRT). REVeRT is flexible in split site selection and can efficiently reconstitute different split genes in numerous in vitro models, in human organoids, and in vivo. Furthermore, REVeRT can functionally reconstitute a CRISPRa module targeting genes in various mouse tissues and organs in single or multiplexed approaches upon different routes of administration. Finally, intravitreal supplementation of ABCA4 via REVeRT improves retinal degeneration and function in a mouse model of Stargardt disease. Due to its flexibility and efficiency REVeRT harbors great potential for basic research and clinical applications.

bioengineering↗

bioRxiv2022

Real-Time Flow Cytometry to assess qualitative and quantitative responses of oral pathobionts during exposure to antiseptics

Antiseptics are widely used in oral healthcare to prevent or treat oral diseases, such as gingivitis and periodontitis. However, the incidence of bacteria being tolerant to standard antiseptics has sharply increased over the last few years. This stresses the urgency for surveillance against tolerant organisms, as well as the discovery of novel antimicrobials. Traditionally, susceptibility to antimicrobials is assessed by broth micro-dilution or disc diffusion assays, both of which are time-consuming, labor-intensive and provide limited information on the mode of action of the antimicrobials. The above-mentioned limitations highlight the need for the development of new methods to monitor and further understand antimicrobial susceptibility. Here, we used real-time flow cytometry, combined with membrane permeability staining, as a quick and sensitive technology to study the quantitative and qualitative response of two oral pathobionts to different concentrations of chlorhexidine, cetylpyridinium chloride or triclosan. Apart from the real-time monitoring of cell damage, we further applied a phenotypic fingerprint method to differentiate between the bacterial subpopulations that arose due to treatment. We quantified the pathobiont damage rate of different antiseptics at different concentrations within 15 minutes of exposure and identified the conditions under which the bacteria were most susceptible. Moreover, we detected species-specific and treatment-specific phenotypic subpopulations. This proves that real-time flow cytometry can provide information on the susceptibility of different microorganisms in a short time frame while differentiating between antiseptics and thus could be a valuable tool in the discovery of novel antimicrobial compounds while at the same time deciphering their mode of action.

microbiology↗

bioRxiv2021

Triangulation of microbial fingerprinting in anaerobic digestion reveals consistent fingerprinting profiles

The anaerobic digestion microbiome has been puzzling us since the dawn of molecular methods for mixed microbial community analysis. Monitoring of the anaerobic digestion microbiome can either take place via a non-targeted holistic evaluation of the microbial community through fingerprinting or by targeted monitoring of selected taxa. Here, we compared four different microbial community fingerprinting methods, i.e., amplicon sequencing, metaproteomics, metabolomics and cytomics, in their ability to characterise the full-scale anaerobic digestion microbiome. Cytometric fingerprinting through cytomics reflects a, for anaerobic digestion, novel, single cell-based approach of direct microbial community fingerprinting by flow cytometry. Three different digester types, i.e., sludge digesters, digesters treating agro-industrial waste and dry anaerobic digesters, each reflected different operational parameters. The -diversity analysis yielded inconsistent results, especially for richness, across the different methods. In contrast, {beta}-diversity analysis resulted in comparable profiles, even when translated into phyla or functions, with clear separation of the three digester types. In-depth analysis of each methods features i.e., operational taxonomic units, metaproteins, metabolites, and cytometric traits, yielded certain similar features, yet, also some clear differences between the different methods, which was related to the complexity of the anaerobic digestion process. In conclusion, cytometric fingerprinting through flow cytometry is a reliable, fast method for holistic monitoring of the anaerobic digestion microbiome, and the complementary identification of key features through other methods could give rise to a direct interpretation of anaerobic digestion process performance.

bioengineering↗

bioRxiv2021

Predicting the presence and abundance of bacterial taxa in environmental communities through flow cytometric fingerprinting

Microbiome management research and applications rely on temporally-resolved measurements of community composition. Current technologies to assess community composition either make use of cultivation or sequencing of genomic material, which can become time consuming and/or laborious in case high-throughput measurements are required. Here, using data from a shrimp hatchery as an economically relevant case study, we combined 16S rRNA gene amplicon sequencing and flow cytometry data to develop a computational workflow that allows the prediction of taxon abundances based on flow cytometry measurements. The first stage of our pipeline consists of a classifier to predict the presence or absence of the taxon of interest, with yields an average accuracy of 88.13{+/-}4.78 % across the top 50 OTUs of our dataset. In the second stage, this classifier was combined with a regression model to predict the relative abundances of the taxon of interest, which yields an average R2 of 0.35{+/-}0.24 across the top 50 OTUs of our dataset. Application of the models on flow cytometry time series data showed that the generated models can predict the temporal dynamics of a large fraction of the investigated taxa. Using cell-sorting we validated that the model correctly associates taxa to regions in the cytometric fingerprint where they are detected using 16S rRNA gene amplicon sequencing. Finally, we applied the approach of our pipeline on two other datasets of microbial ecosystems. This pipeline represents an addition to the expanding toolbox for flow cytometry-based monitoring of bacterial communities and complements the current plating- and marker gene-based methods. ImportanceMonitoring of microbial community composition is crucial for both microbiome management research and applications. Existing technologies, such as plating and amplicon sequencing, can become laborious and expensive when high-throughput measurements are required. Over the recent years, flow cytometry-based measurements of community diversity have been shown to correlate well to those derived from 16S rRNA gene amplicon sequencing in several aquatic ecosystems, suggesting there is a link between the taxonomic community composition and phenotypic properties as derived through flow cytometry. Here, we further integrated 16S rRNA gene amplicon sequencing and flow cytometry survey data in order to construct models that enable the prediction of both the presence and the abundance of individual bacterial taxa in mixed communities using flow cytometric fingerprinting. The developed pipeline holds great potential to be integrated in routine monitoring schemes and early warning systems for biotechnological applications.

microbiology↗

bioRxiv2021

Selective metal extraction by biologically produced siderophores during bioleaching from low-grade primary and secondary mineral resources

Siderophores are a class of biogenic macromolecules that have high affinities for metals in the environment, thus could be exploited for alternate sustainable metal recovery technologies. Here, we assess the role of siderophores in the extraction and complexation of metals from an iron oxide-rich metallurgical processing residue and a low-grade primary Ni ore. Evaluation of the biological siderophore bioproduction by three pseudomonads, P. fluorescens, P. azotoformans and P. putida identified that P. putida could generate the highest siderophore yield, which was characterized as a hydroxamate and catecholate mixed-type pyoverdine PyoPpC-3B. Key physiochemical parameters involved in raw siderophore mediated metal extraction were identified using a fractional factorial design of experiments (DOE) and subsequently employed in purified PyoPpC-3B leaching experiments. Further targeted experiments with hydroxamate and catecholate functional analogues of PyoPpC-3B confirmed their marked ability to competitively or selectively leach and chelate hard metal ions, including Al(OH)4-, Mn2+ and Zn2+. Interestingly, complexation of Mn and Zn ions exceeded the natural affinity of pyoverdine for Fe3+, thus despite the low metal recoveries from the materials tested in this study, this work provides important new insights in siderophore-metal interactions.

biochemistry↗

bioRxiv2020

Rearing water microbiomes in white leg shrimp (Litopenaeus vannamei) larviculture assemble stochastically and are influenced by the microbiomes of live feed products

The development of effective management strategies to reduce the occurrence of diseases in aquaculture is hampered by the limited knowledge on the microbial ecology of these systems. In this study, the dynamics and dominant community assembly processes in the rearing water of Litopenaeus vannamei larviculture tanks were determined. Additionally, the contribution of peripheral microbiomes, such as those of live and dry feeds, to the rearing water microbiome were quantified. The community assembly in the hatchery rearing water over time was dominated by stochasticity, which explains the observed heterogeneity between replicate cultivations. The community undergoes two shifts that match with the dynamics of the algal abundances in the rearing water. Source tracking analysis revealed that 37% of all bacteria in the hatchery rearing water were either introduced by the live or dry feeds, or during water exchanges. The contribution of the microbiome from the algae was the largest, followed by that of the Artemia, the exchange water and the dry feeds. Our findings provide fundamental knowledge on the assembly processes and dynamics of rearing water microbiomes and illustrate the crucial role of these peripheral microbiomes in maintaining health-promoting rearing water microbiomes. Originality-Significance StatementMost studies on rearing water microbiomes are characterized by sampling resolutions of multiple days and by few replicate cultivations. Through an 18-day sampling campaign in a Litopenaeus vannamei hatchery where five replicate cultivations were studied at a sampling resolution of one day, we studied the microbiome dynamics in this system. We show that the community assembly is dominated by stochasticity, which explains the heterogeneity between replicate cultivations. The dynamics of the algal community in the rearing water induced shifts in community composition at two differerent timepoints. Finally, we quantified the contribution of live and dry feed microbiomes to the rearing water community for the first time. We found that the contribution of each source was dependent on its taxonomic composition, the bacterial load caused by the addition of this source and the timing of the introduction. These new insights will aid in the further development of effective microbiome management to reduce the frequency and magnitude of bacterial diseases.

microbiology↗