Plastics are widely used in the global economy, and each year, at least 350 to 400 million tons are being produced. Due to poor recycling and low circular use, millions of tons accumulate annually in terrestrial or marine environments. Today it has become clear that plastic causes adverse effects in all ecosystems and that microplastics are of particular concern to our health. Therefore, recent microbial research has addressed the question of if and to what extent microorganisms can degrade plastics in the environment. This review summarizes current knowledge on microbial plastic degradation. Enzymes available act mainly on the high-molecular-weight polymers of polyethylene terephthalate (PET) and ester-based polyurethane (PUR). Unfortunately, the best PUR- and PET-active enzymes and microorganisms known still have moderate turnover rates. While many reports describing microbial communities degrading chemical additives have been published, no enzymes acting on the high-molecular-weight polymers polystyrene, polyamide, polyvinylchloride, polypropylene, ether-based polyurethane, and polyethylene are known. Together, these polymers comprise more than 80% of annual plastic production. Thus, further research is needed to significantly increase the diversity of enzymes and microorganisms acting on these polymers. This can be achieved by tapping into the global metagenomes of noncultivated microorganisms and dark matter proteins. Only then can novel biocatalysts and organisms be delivered that allow rapid degradation, recycling, or value-added use of the vast majority of most human-made polymers.

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The cohort of the ammonia-oxidizing archaea (AOA) of the phylum Thaumarchaeota is a diverse, widespread and functionally important group of microorganisms in many ecosystems. However, our understanding of their biology is still very rudimentary in part because all available genome sequences of this phylum are from members of the Nitrosopumilus cluster. Here we report on the complete genome sequence of Candidatus Nitrososphaera gargensis obtained from an enrichment culture, representing a different evolutionary lineage of AOA frequently found in high numbers in many terrestrial environments. With its 2.83 Mb the genome is much larger than that of other AOA. The presence of a high number of (active) IS elements/transposases, genomic islands, gene duplications and a complete CRISPR/Cas defence system testifies to its dynamic evolution consistent with low degree of synteny with other thaumarchaeal genomes. As expected, the repertoire of conserved enzymes proposed to be required for archaeal ammonia oxidation is encoded by N. gargensis, but it can also use urea and possibly cyanate as alternative ammonia sources. Furthermore, its carbon metabolism is more flexible at the central pyruvate switch point, encompasses the ability to take up small organic compounds and might even include an oxidative pentose phosphate pathway. Furthermore, we show that thaumarchaeota produce cofactor F420 as well as polyhydroxyalkanoates. Lateral gene transfer from bacteria and euryarchaeota has contributed to the metabolic versatility of N. gargensis. This organisms is well adapted to its niche in a heavy metal-containing thermal spring by encoding a multitude of heavy metal resistance genes, chaperones and mannosylglycerate as compatible solute and has the genetic ability to respond to environmental changes by signal transduction via a large number of two-component systems, by chemotaxis and flagella-mediated motility and possibly even by gas vacuole formation. These findings extend our understanding of thaumarchaeal evolution and physiology and offer many testable hypotheses for future experimental research on these nitrifiers.

Esterases receive special attention because of their wide distribution in biological systems and environments and their importance for physiology and chemical synthesis. The prediction of esterases' substrate promiscuity level from sequence data and the molecular reasons why certain such enzymes are more promiscuous than others remain to be elucidated. This limits the surveillance of the sequence space for esterases potentially leading to new versatile biocatalysts and new insights into their role in cellular function. Here, we performed an extensive analysis of the substrate spectra of 145 phylogenetically and environmentally diverse microbial esterases, when tested with 96 diverse esters. We determined the primary factors shaping their substrate range by analyzing substrate range patterns in combination with structural analysis and protein-ligand simulations. We found a structural parameter that helps rank (classify) the promiscuity level of esterases from sequence data at 94% accuracy. This parameter, the active site effective volume, exemplifies the topology of the catalytic environment by measuring the active site cavity volume corrected by the relative solvent accessible surface area (SASA) of the catalytic triad. Sequences encoding esterases with active site effective volumes (cavity volume/SASA) above a threshold show greater substrate spectra, which can be further extended in combination with phylogenetic data. This measure provides also a valuable tool for interrogating substrates capable of being converted. This measure, found to be transferred to phosphatases of the haloalkanoic acid dehalogenase superfamily and possibly other enzymatic systems, represents a powerful tool for low-cost bioprospecting for esterases with broad substrate ranges, in large scale sequence data sets.

Polyethylene terephthalate (PET) is one of the most important synthetic polymers used today. Unfortunately, the polymers accumulate in nature and to date no highly active enzymes are known that can degrade it at high velocity. Enzymes involved in PET degradation are mainly α- and β-hydrolases, like cutinases and related enzymes (EC 3.1.1). Currently, only a small number of such enzymes are well characterized. In this work, a search algorithm was developed that identified 504 possible PET hydrolase candidate genes from various databases. A further global search that comprised more than 16 Gb of sequence information within 108 marine and 25 terrestrial metagenomes obtained from the Integrated Microbial Genome (IMG) database detected 349 putative PET hydrolases. Heterologous expression of four such candidate enzymes verified the function of these enzymes and confirmed the usefulness of the developed search algorithm. In this way, two novel and thermostable enzymes with high potential for downstream application were partially characterized. Clustering of 504 novel enzyme candidates based on amino acid similarities indicated that PET hydrolases mainly occur in the phyla of Actinobacteria, Proteobacteria, and Bacteroidetes. Within the Proteobacteria, the Betaproteobacteria, Deltaproteobacteria, and Gammaproteobacteria were the main hosts. Remarkably enough, in the marine environment, bacteria affiliated with the phylum Bacteroidetes appear to be the main hosts of PET hydrolase genes, rather than Actinobacteria or Proteobacteria, as observed for the terrestrial metagenomes. Our data further imply that PET hydrolases are truly rare enzymes. The highest occurrence of 1.5 hits/Mb was observed in sequences from a sample site containing crude oil.IMPORTANCE Polyethylene terephthalate (PET) accumulates in our environment without significant microbial conversion. Although a few PET hydrolases are already known, it is still unknown how frequently they appear and with which main bacterial phyla they are affiliated. In this study, deep sequence mining of protein databases and metagenomes demonstrated that PET hydrolases indeed occur at very low frequencies in the environment. Furthermore, it was possible to link them to phyla that were previously not known to harbor such enzymes. This work contributes novel knowledge on the phylogenetic relationships, the recent evolution, and the global distribution of PET hydrolases. Finally, we describe the biochemical traits of four novel PET hydrolases.

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Viruses represent the most abundant life forms on the planet. Recent experimental and computational improvements have led to a dramatic increase in the number of viral genome sequences identified primarily from metagenomic samples. As a result of the expanding catalog of metagenomic viral sequences, there exists a need for a comprehensive computational platform integrating all these sequences with associated metadata and analytical tools. Here we present IMG/VR (https://img.jgi.doe.gov/vr/), the largest publicly available database of 3908 isolate reference DNA viruses with 264 413 computationally identified viral contigs from >6000 ecologically diverse metagenomic samples. Approximately half of the viral contigs are grouped into genetically distinct quasi-species clusters. Microbial hosts are predicted for 20 000 viral sequences, revealing nine microbial phyla previously unreported to be infected by viruses. Viral sequences can be queried using a variety of associated metadata, including habitat type and geographic location of the samples, or taxonomic classification according to hallmark viral genes. IMG/VR has a user-friendly interface that allows users to interrogate all integrated data and interact by comparing with external sequences, thus serving as an essential resource in the viral genomics community.

Metagenomics as a new field of research has been developed over the past decade to elucidate the genomes of the non-cultured microbes with the goal to better understand global microbial ecology on the one side, and on the other side it has been driven by the increasing biotechnological demands for novel enzymes and biomolecules. Since it is well accepted that the majority of all microbes has not yet been cultured, the not-yet-cultivated microbes represent a shear unlimited and intriguing resource for the development of novel genes, enzymes and chemical compounds for use in biotechnology. However, with respect to biotechnology, metagenomics faces now two major challenges. Firstly, it has to identify truly novel biocatalysts to fulfil the needs of industrial processes and green chemistry. Secondly, the already available genes and enzymes need to be implemented in production processes to further prove the value of metagenome-derived sequences.