Protein structure, amino acid composition and sequence determine proteome vulnerability to oxidation-induced damage.

Oxidative stress alters cell viability, from microorganism irradiation sensitivity to human aging and neurodegeneration. Deleterious effects of protein carbonylation by reactive oxygen species (ROS) make understanding molecular properties determining ROS susceptibility essential. The radiation-resistant bacterium Deinococcus radiodurans accumulates less carbonylation than sensitive organisms, making it a key model for deciphering properties governing oxidative stress resistance. We integrated shotgun redox proteomics, structural systems biology, and machine learning to resolve properties determining protein damage by γ-irradiation in Escherichia coli and D. radiodurans at multiple scales. Local accessibility, charge, and lysine enrichment accurately predict ROS susceptibility. Lysine, methionine, and cysteine usage also contribute to ROS resistance of the D. radiodurans proteome. Our model predicts proteome maintenance machinery, and proteins protecting against ROS are more resistant in D. radiodurans. Our findings substantiate that protein-intrinsic protection impacts oxidative stress resistance, identifying causal molecular properties.

Diel Cycle Proteomics: Illuminating Molecular Dynamics in Purple Bacteria for Optimized Biotechnological Applications.

Circadian rhythms, characterized by approximately 24 h cycles, play a pivotal role in enabling various organisms to synchronize their biological activities with daily variations. While ubiquitous in Eukaryotes, circadian clocks remain exclusively characterized in Cyanobacteria among Prokaryotes. These rhythms are regulated by a core oscillator, which is controlled by a cluster of three genes: kaiA, kaiB, and kaiC. Interestingly, recent studies revealed rhythmic activities, potentially tied to a circadian clock, in other Prokaryotes, including purple bacteria such as Rhodospirillum rubrum, known for its applications in fuel and plastic bioproduction. However, the pivotal question of how light and dark cycles influence protein dynamics and the expression of putative circadian clock genes remains unexplored in purple non-sulfur bacteria. Unraveling the regulation of these molecular clocks holds the key to unlocking optimal conditions for harnessing the biotechnological potential of R. rubrum. Understanding how its proteome responds to different light regimes-whether under continuous light or alternating light and dark cycles-could pave the way for precisely fine-tuning bioproduction processes. Here, we report for the first time the expressed proteome of R. rubrum grown under continuous light versus light and dark cycle conditions using a shotgun proteomic analysis. In addition, we measured the impact of light regimes on the expression of four putative circadian clock genes (kaiB1, kaiB2, kaiC1, kaiC2) at the transcriptional and translational levels using RT-qPCR and targeted proteomic (MRM-MS), respectively. The data revealed significant effects of light conditions on the overall differential regulation of the proteome, particularly during the early growth stages. Notably, several proteins were found to be differentially regulated during the light or dark period, thus impacting crucial biological processes such as energy conversion pathways and the general stress response. Furthermore, our study unveiled distinct regulation of the four kai genes at both the mRNA and protein levels in response to varying light conditions. Deciphering the impact of the diel cycle on purple bacteria not only enhances our understanding of their ecology but also holds promise for optimizing their applications in biotechnology, providing valuable insights into the origin and evolution of prokaryotic clock mechanisms.

Towards the discovery of novel molecular clocks in Prokaryotes.

Diel cycle is of enormous biological importance as it imposes daily oscillation in environmental conditions, which temporally structures most ecosystems. Organisms developed biological time-keeping mechanisms - circadian clocks - that provide a significant fitness advantage over competitors by optimising the synchronisation of their biological activities. While circadian clocks are ubiquitous in Eukaryotes, they are so far only characterised in Cyanobacteria within Prokaryotes. However, growing evidence suggests that circadian clocks are widespread in the bacterial and archaeal domains. As Prokaryotes are at the heart of crucial environmental processes and are essential to human health, unravelling their time-keeping systems provides numerous applications in medical research, environmental sciences, and biotechnology. In this review, we elaborate on how novel circadian clocks in Prokaryotes offer research and development perspectives. We compare and contrast the different circadian systems in Cyanobacteria and discuss about their evolution and taxonomic distribution. We necessarily provide an updated phylogenetic analysis of bacterial and archaeal species that harbour homologs of the main cyanobacterial clock components. Finally, we elaborate on new potential clock-controlled microorganisms that represent opportunities of ecological and industrial relevance in prokaryotic groups such as anoxygenic photosynthetic bacteria, methanogenic archaea, methanotrophs or sulphate-reducing bacteria.

Survival of human enteric and respiratory viruses on plastics in soil, freshwater, and marine environments.

The public health significance of plastics and microplastics in different environmental matrices has mainly focused on the toxicological effects of human ingestion. But these pollutants can also harbour pathogenic bacteria as the surfaces of plastics in the environment quickly become colonised by microbial biofilm. This novel microbial habitat has been termed the 'plastisphere' and could facilitate the survival and dissemination of important bacterial and fungal pathogens. Importantly, however, the role of plastic pollution as a secondary pathway for the transmission of human pathogenic viruses has never been addressed. Due to the high prevalence of both enteric and respiratory viruses in the population and in the environment, there is significant potential for human viruses to become associated with the plastisphere. In this review we critically evaluate current knowledge on the interaction of human enteric and respiratory viruses with plastic surfaces and identify the main environmental conditions and plastic characteristics that could affect virus survival and persistence in the environment. Our hypothesis is that the plastisphere can enhance the adhesion, survival and dissemination of human pathogenic viruses and potentially lead to more effective transfer and transmission of viral diseases within the environment. We identify key research questions needed to more fully assess the potential human health risks associated with viruses on plastic surfaces. These include understanding, (1) the mechanisms of viral attachment to either naked or biofilm-colonised plastic (2) how the structural characteristics of viruses (e.g., enveloped, or non-enveloped), affect their persistence in the plastisphere, (3) whether the plastisphere offers protection and increases the persistence of infectious viruses in soil, freshwater, and marine environments.

A Closer Look at Plastic Colonisation: Prokaryotic Dynamics in Established versus Newly Synthesised Marine Plastispheres and their Planktonic State.

The taxonomy of marine plastisphere communities has been extensively studied, demonstrating the ubiquity of hydrocarbonoclastic bacteria of potential biotechnological significance. However, prokaryotic functioning on plastic surfaces has received limited attention, and the question of whether these microorganisms are active and expressing specific molecular mechanisms underpinning plastisphere colonisation remains to be addressed. The aim of this study was to investigate the plastic colonisation process, to identify the active taxa involved in biofilm formation and the mechanisms used to initiate colonisation. To achieve this, a marine plastisphere characterised by active hydrocarbonoclastic genera was used as the inoculum for a short-term microcosm experiment using virgin low-density polyethylene as the sole carbon source. Following incubation for 1 and 2 weeks (representing early and late colonisation, respectively), a taxonomic and comparative metaproteomic approach revealed a significant shift in plastisphere diversity and composition, yet highlighted stability in the predominance of active Proteobacteria spanning 16 genera, including Marinomonas, Pseudomonas, and Pseudoalteromonas. Relative quantification of 1,762 proteins shared between the initial plastisphere inoculum, the microcosm plastisphere and the planktonic cells in the surrounding artificial seawater, provided insights into the differential regulation of proteins associated with plastisphere formation. This included the upregulation of proteins mediating cellular attachment in the plastisphere, for example flagellin expressed by Marinomonas, Cobetia, Pseudoalteromonas, and Pseudomonas, and curli expressed by Cobetia. In addition to the differential regulation of energy metabolism in Marinomonas, Psychrobacter, Pseudomonas and Cobetia within the plastisphere relative to the surrounding seawater. Further, we identified the upregulation of amino acid metabolism and transport, including glutamine hydrolysis to glutamate in Marinomonas and unclassified Halomonadaceae, potentially coupled to ammonia availability and oxidative stress experienced within the plastisphere. Our study provides novel insights into the dynamics of plastisphere formation and function, highlighting potential targets for regulating plastisphere growth to enhance plastic bioremediation processes.

Novel functional insights into the microbiome inhabiting marine plastic debris: critical considerations to counteract the challenges of thin biofilms using multi-omics and comparative metaproteomics.

BACKGROUND: Microbial functioning on marine plastic surfaces has been poorly documented, especially within cold climates where temperature likely impacts microbial activity and the presence of hydrocarbonoclastic microorganisms. To date, only two studies have used metaproteomics to unravel microbial genotype-phenotype linkages in the marine 'plastisphere', and these have revealed the dominance of photosynthetic microorganisms within warm climates. Advancing the functional representation of the marine plastisphere is vital for the development of specific databases cataloging the functional diversity of the associated microorganisms and their peptide and protein sequences, to fuel biotechnological discoveries. Here, we provide a comprehensive assessment for plastisphere metaproteomics, using multi-omics and data mining on thin plastic biofilms to provide unique insights into plastisphere metabolism. Our robust experimental design assessed DNA/protein co-extraction and cell lysis strategies, proteomics workflows, and diverse protein search databases, to resolve the active plastisphere taxa and their expressed functions from an understudied cold environment. RESULTS: For the first time, we demonstrate the predominance and activity of hydrocarbonoclastic genera (Psychrobacter, Flavobacterium, Pseudomonas) within a primarily heterotrophic plastisphere. Correspondingly, oxidative phosphorylation, the citrate cycle, and carbohydrate metabolism were the dominant pathways expressed. Quorum sensing and toxin-associated proteins of Streptomyces were indicative of inter-community interactions. Stress response proteins expressed by Psychrobacter, Planococcus, and Pseudoalteromonas and proteins mediating xenobiotics degradation in Psychrobacter and Pseudoalteromonas suggested phenotypic adaptations to the toxic chemical microenvironment of the plastisphere. Interestingly, a targeted search strategy identified plastic biodegradation enzymes, including polyamidase, hydrolase, and depolymerase, expressed by rare taxa. The expression of virulence factors and mechanisms of antimicrobial resistance suggested pathogenic genera were active, despite representing a minor component of the plastisphere community. CONCLUSION: Our study addresses a critical gap in understanding the functioning of the marine plastisphere, contributing new insights into the function and ecology of an emerging and important microbial niche. Our comprehensive multi-omics and comparative metaproteomics experimental design enhances biological interpretations to provide new perspectives on microorganisms of potential biotechnological significance beyond biodegradation and to improve the assessment of the risks associated with microorganisms colonizing marine plastic pollution. Video Abstract.

Microplastics contamination in aquaculture-rich regions: A case study in Gresik, East Java, Indonesia.

The widespread use of plastic has resulted in the accumulation of plastic waste across a range of sizes, notably including microplastics (MPs). The introduction of MPs into aquatic ecosystems can lead to the contamination of organisms, mainly fish. This study reports for the first time a quantitative and qualitative analysis conducted on the abundance of MPs encountered in water and sediment of milkfish aquaculture ponds in Gresik, East Java, Indonesia. Water and sediment samples were collected at three stations between February to April 2021. The abundance of MPs was analyzed through the application of one-way ANOVA tests and Pearson's correlation analysis. The results identified four types of MPs: fragments, fibers, films, and pellets. The highest abundance of MPs in both water (10.40 particle/L) and sediment samples (1.15 particle/g) was observed in March. The predominant MPs size in the water samples is 100-500 µm, while it is below 100 µm in the sediment. The color of the MPs varied across eight colors: black, purple, red, blue, yellow, pink, green, and transparent. The identification of MPs polymers was found to be polypropylene (PP), Polyurethane (PU), Polycarbonate (PC), Polyethylene terephthalate (PETE), High-density polyethylene (HDPE), and low-density polyethylene (LDPE). The presence of MPs in the water column and sediments was correlated with human activities around the ponds. Hence, the abundance of MPs is a source of pollution that has the potential to damage the nutritional quality of farmed milkfish. This study provides important information for the local governments to develop waste management policies for a cleaner environment and improved human health.

Diversifying the concept of model organisms in the age of -omics.

In today's post-genomic era, it is crucial to rethink the concept of model organisms. While a few historically well-established organisms, e.g. laboratory rodents, have enabled significant scientific breakthroughs, there is now a pressing need for broader inclusion. Indeed, new organisms and models, from complex microbial communities to holobionts, are essential to fully grasp the complexity of biological principles across the breadth of biodiversity. By fostering collaboration between biology, advanced molecular science and omics communities, we can collectively adopt new models, unraveling their molecular functioning, and uncovering fundamental mechanisms. This concerted effort will undoubtedly enhance human health, environmental quality, and biodiversity conservation.

From rivers to marine environments: A constantly evolving microbial community within the plastisphere.

Plastics accumulate in the environment and the Mediterranean Sea is one of the most polluted sea in the world. The plastic surface is rapidly colonized by microorganisms, forming the plastisphere. Our unique sampling supplied 107 plastic pieces from 22 geographical sites from four aquatic ecosystems (river, estuary, harbor and inshore) in the south of France in order to better understand the parameters which influence biofilm composition. In parallel, 48 enrichment cultures were performed to investigate the presence of plastic degrading-bacteria in the plastisphere. In this context, we showed that the most important drivers of microbial community structure were the sampling site followed by the polymer chemical composition. The study of pathogenic genus distribution highlighted that only 11% of our plastic samples contained higher proportions of Vibrio compared to the natural environment. Finally, results of the enrichment cultures showed a selection of hydrocarbon-degrading microorganisms suggesting their potential role in the plastic degradation.

Binding, recovery, and infectiousness of enveloped and non-enveloped viruses associated with plastic pollution in surface water.

Microplastics in wastewater and surface water rapidly become colonised by microbial biofilm. Such 'plastisphere' communities are hypothesised to persist longer and be disseminated further in the environment and may act as a vector for human pathogens, particularly as microplastics entering wastewater treatment plants are exposed to high concentrations of pathogenic bacteria. However, the potential for human viral pathogens to become associated with the plastisphere has never before been quantified. Here, we have used rotavirus (RV) SA11 (a non-enveloped enteric virus) and the enveloped bacteriophage Phi6 as model viruses to quantify binding and recovery from biofilm-colonised microplastic pellets in three different water treatments (filtered and non-filtered surface water, and surface water with added nutrients). Viruses associated with biofilm-colonised pellets were more stable compared to those remaining in the water. While infectious particles and genome copies of RV remained stable over the 48 h sampling period, Phi6 stability was highly impacted, with a reduction ranging from 2.18 to 3.94 log10. Virus particles were protected against inactivation factors when associated with the biofilm on microplastic surfaces, and when there was a high concentration of particulate matter in the liquid phase. Although our results suggest that the presence of an envelope may limit virus interaction with the plastisphere, the ability to recover both enveloped and non-enveloped infectious viruses from colonised microplastic pellets highlights an additional potential public health risk of surface waters becoming contaminated with microplastics, and subsequent human exposure to microplastics in the environment.

Priorities to inform research on marine plastic pollution in Southeast Asia.

Southeast Asia is considered to have some of the highest levels of marine plastic pollution in the world. It is therefore vitally important to increase our understanding of the impacts and risks of plastic pollution to marine ecosystems and the essential services they provide to support the development of mitigation measures in the region. An interdisciplinary, international network of experts (Australia, Indonesia, Ireland, Malaysia, the Philippines, Singapore, Thailand, the United Kingdom, and Vietnam) set a research agenda for marine plastic pollution in the region, synthesizing current knowledge and highlighting areas for further research in Southeast Asia. Using an inductive method, 21 research questions emerged under five non-predefined key themes, grouping them according to which: (1) characterise marine plastic pollution in Southeast Asia; (2) explore its movement and fate across the region; (3) describe the biological and chemical modifications marine plastic pollution undergoes; (4) detail its environmental, social, and economic impacts; and, finally, (5) target regional policies and possible solutions. Questions relating to these research priority areas highlight the importance of better understanding the fate of marine plastic pollution, its degradation, and the impacts and risks it can generate across communities and different ecosystem services. Knowledge of these aspects will help support actions which currently suffer from transboundary problems, lack of responsibility, and inaction to tackle the issue from its point source in the region. Being profoundly affected by marine plastic pollution, Southeast Asian countries provide an opportunity to test the effectiveness of innovative and socially inclusive changes in marine plastic governance, as well as both high and low-tech solutions, which can offer insights and actionable models to the rest of the world.

Shedding light on the bacterial resistance to toxic UV filters: a comparative genomic study.

UV filters are toxic to marine bacteria that dominate the marine biomass. Ecotoxicology often studies the organism response but rarely integrates the toxicity mechanisms at the molecular level. In this study, in silico comparative genomics between UV filters sensitive and resistant bacteria were conducted in order to unravel the genes responsible for a resistance phenotype. The genomes of two environmentally relevant Bacteroidetes and three Firmicutes species were compared through pairwise comparison. Larger genomes were carried by bacteria exhibiting a resistant phenotype, favoring their ability to adapt to environmental stresses. While the antitoxin and CRISPR systems were the only distinctive features in resistant Bacteroidetes, Firmicutes displayed multiple unique genes that could support the difference between sensitive and resistant phenotypes. Several genes involved in ROS response, vitamin biosynthesis, xenobiotic degradation, multidrug resistance, and lipophilic compound permeability were shown to be exclusive to resistant species. Our investigation contributes to a better understanding of UV filters resistance phenotypes, by identifying pivotal genes involved in key pathways.

Unraveling the molecular effects of oxybenzone on the proteome of an environmentally relevant marine bacterium.

The use of Benzophenone-3 (BP3), also known as oxybenzone, a common UV filter, is a growing environmental concern in regard to its toxicity on aquatic organisms. Our previous work stressed that BP3 is toxic to Epibacterium mobile, an environmentally relevant marine α-proteobacterium. In this study, we implemented a label-free quantitative proteomics workflow to decipher the effects of BP3 on the E. mobile proteome. Furthermore, the effect of DMSO, one of the most common solvents used to vehicle low concentrations of lipophilic chemicals, was assessed to emphasize the importance of limiting solvent concentration in ecotoxicological studies. Data-independent analysis proteomics highlighted that BP3 induced changes in the regulation of 56 proteins involved in xenobiotic export, detoxification, oxidative stress response, motility, and fatty acid, iron and amino acid metabolisms. Our results also outlined that the use of DMSO at 0.046% caused regulation changes in proteins related to transport, iron uptake and metabolism, and housekeeping functions, underlining the need to reduce the concentration of solvents in ecotoxicological studies.

Diel Protein Regulation of Marine Picoplanktonic Communities Assessed by Metaproteomics.

The diel cycle is of enormous biological importance in that it imposes temporal structure on ecosystem productivity. In the world's oceans, microorganisms form complex communities that carry out about half of photosynthesis and the bulk of life-sustaining nutrient cycling. How the functioning of microbial communities is impacted by day and night periods in surface seawater remains to be elucidated. In this study, we compared the day and night metaproteomes of the free-living and the particle-attached bacterial fractions from picoplanktonic communities sampled from the northwest Mediterranean Sea surface. Our results showed similar taxonomic distribution of free-living and particle-attached bacterial populations, with Alphaproteobacteria, Gammaproteobacteria and Cyanobacteria being the most active members. Comparison of the day and night metaproteomes revealed that free-living and particle-attached bacteria were more active during the day and the night, respectively. Interestingly, protein diel variations were observed in the photoautotroph Synechococcales and in (photo)-heterotrophic bacteria such as Flavobacteriales, Pelagibacterales and Rhodobacterales. Moreover, our data demonstrated that diel cycle impacts light-dependent processes such as photosynthesis and UV-stress response in Synechococcales and Rhodobacterales, respectively, while the protein regulation from the ubiquitous Pelagibacterales remained stable over time. This study unravels, for the first time, the diel variation in the protein expression of major free-living and particle-attached microbial players at the sea surface, totaling an analysis of eight metaproteomes.

Toxicity of UV filters on marine bacteria: Combined effects with damaging solar radiation.

Organic UV filters are of emerging concern due to their occurrence and persistence in coastal ecosystems. Because marine bacteria are crucial in the major biogeochemical cycles, there is an urgent need to understand to what extent these microorganisms are affected by those chemicals. This study deciphers the impact of five common sunscreen UV filters on twenty-seven marine bacteria, combining both photobiology and toxicity analysis on environmentally relevant species. Seven bacteria were sensitive to different organic UV filters at 1000 µg L-1, including octinoxate and oxybenzone. This is the first report demonstrating inhibition of bacterial growth from 100 µg L-1. None of the UV filters showed any toxicity at 1000 µg L-1 on stationary phase cells, demonstrating that physiological state was found to be a key parameter in the bacterial response to UV-filters. Indeed, non-growing bacteria were resistant to UV filters whereas growing cells exhibited UV filter dependent sensitivity. Octinoxate was the most toxic chemical at 1000 µg L-1 on growing cells. Interestingly, photobiology experiments revealed that the toxicity of octinoxate and homosalate decreased after light exposure while the other compounds were not affected. In terms of environmental risk characterization, our results revealed that the increasing use of sun blockers could have detrimental impacts on bacterioplanktonic communities in coastal areas. Our findings contribute to a better understanding of the impact of the most common UV filters on bacterial species and corroborate the importance to consider environmental parameters such as solar radiation in ecotoxicology studies.

mPies: a novel metaproteomics tool for the creation of relevant protein databases and automatized protein annotation.

Metaproteomics allows to decipher the structure and functionality of microbial communities. Despite its rapid development, crucial steps such as the creation of standardized protein search databases and reliable protein annotation remain challenging. To overcome those critical steps, we developed a new program named mPies (metaProteomics in environmental sciences). mPies allows the creation of protein databases derived from assembled or unassembled metagenomes, and/or public repositories based on taxon IDs, gene or protein names. For the first time, mPies facilitates the automatization of reliable taxonomic and functional consensus annotations at the protein group level, minimizing the well-known protein inference issue, which is commonly encountered in metaproteomics. mPies' workflow is highly customizable with regards to input data, workflow steps, and parameter adjustment. mPies is implemented in Python 3/Snakemake and freely available on GitHub: https://github.com/johanneswerner/mPies/. REVIEWER: This article was reviewed by Dr. Wilson Wen Bin Goh.

Deciphering the Functioning of Microbial Communities: Shedding Light on the Critical Steps in Metaproteomics.

Unraveling the complex structure and functioning of microbial communities is essential to accurately predict the impact of perturbations and/or environmental changes. From all molecular tools available today to resolve the dynamics of microbial communities, metaproteomics stands out, allowing the establishment of phenotype-genotype linkages. Despite its rapid development, this technology has faced many technical challenges that still hamper its potential power. How to maximize the number of protein identification, improve quality of protein annotation, and provide reliable ecological interpretation are questions of immediate urgency. In our study, we used a robust metaproteomic workflow combining two protein fractionation approaches (gel-based versus gel-free) and four protein search databases derived from the same metagenome to analyze the same seawater sample. The resulting eight metaproteomes provided different outcomes in terms of (i) total protein numbers, (ii) taxonomic structures, and (iii) protein functions. The characterization and/or representativeness of numerous proteins from ecologically relevant taxa such as Pelagibacterales, Rhodobacterales, and Synechococcales, as well as crucial environmental processes, such as nutrient uptake, nitrogen assimilation, light harvesting, and oxidative stress response, were found to be particularly affected by the methodology. Our results provide clear evidences that the use of different protein search databases significantly alters the biological conclusions in both gel-free and gel-based approaches. Our findings emphasize the importance of diversifying the experimental workflow for a comprehensive metaproteomic study.

Proteogenomic Analysis of Epibacterium Mobile BBCC367, a Relevant Marine Bacterium Isolated From the South Pacific Ocean.

Epibacterium mobile BBCC367 is a marine bacterium that is common in coastal areas. It belongs to the Roseobacter clade, a widespread group in pelagic marine ecosystems. Species of the Roseobacter clade are regularly used as models to understand the evolution and physiological adaptability of generalist bacteria. E. mobile BBCC367 comprises two chromosomes and two plasmids. We used gel-free shotgun proteomics to assess its protein expression under 16 different conditions, including stress factors such as elevated temperature, nutrient limitation, high metal concentration, and UVB exposure. Comparison of the different conditions allowed us not only to retrieve almost 70% of the predicted proteins, but also to define three main protein assemblages: 584 essential core proteins, 2,144 facultative accessory proteins and 355 specific unique proteins. While the core proteome mainly exhibited proteins involved in essential functions to sustain life such as DNA, amino acids, carbohydrates, cofactors, vitamins and lipids metabolisms, the accessory and unique proteomes revealed a more specific adaptation with the expression of stress-related proteins, such as DNA repair proteins (accessory proteome), transcription regulators and a significant predominance of transporters (unique proteome). Our study provides insights into how E. mobile BBCC367 adapts to environmental changes and copes with diverse stresses.