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A series of spectacular scientific discoveries and technological advances in the second half of the 20th century have provided the basis for the ongoing genome editing revolution. The elucidation of structural and functional features of DNA and RNA was followed by pioneering studies on genome editing: Molecular biotechnology was born. Since then, four decades followed during which progress of scientific insights and technological methods continued at an overwhelming pace. Fundamental insights into microbial host-virus interactions led to the development of tools for genome editing using restriction enzymes or the revolutionary CRISPR-Cas technology. In this review, we provide a historical overview of milestones that led to the genome editing revolution and speculate about future trends in biotechnology.

Chlamydiae is a bacterial phylum composed of obligate animal and protist endosymbionts. However, other members of the Planctomycetes–Verrucomicrobia–Chlamydiae superphylum are primarily free living. How Chlamydiae transitioned to an endosymbiotic lifestyle is still largely unresolved. Here we reconstructed Planctomycetes–Verrucomicrobia–Chlamydiae species relationships and modelled superphylum genome evolution. Gene content reconstruction from 11,996 gene families suggests a motile and facultatively anaerobic last common Chlamydiae ancestor that had already gained characteristic endosymbiont genes. Counter to expectations for genome streamlining in strict endosymbionts, we detected substantial gene gain within Chlamydiae. We found that divergence in energy metabolism and aerobiosis observed in extant lineages emerged later during chlamydial evolution. In particular, metabolic and aerobic genes characteristic of the more metabolically versatile protist-infecting chlamydiae were gained, such as respiratory chain complexes. Our results show that metabolic complexity can increase during endosymbiont evolution, adding an additional perspective for understanding symbiont evolutionary trajectories across the tree of life.

There is a need for standardized in vitro models emulating the functionalities of the human intestinal tract to study human intestinal health without the use of laboratory animals. The Caco-2 cell line is a well-accepted and highly characterized intestinal barrier model, which has been intensively used to study intestinal (drug) transport, host–microbe interactions and chemical or drug toxicity. This cell line has been cultured in different in vitro models, ranging from simple static to complex dynamic microfluidic models. We aimed to investigate the effect of these different in vitro experimental variables on gene expression. To this end, we systematically collected and extracted data from studies in which transcriptome analyses were performed on Caco-2 cells grown on permeable membranes. A collection of 13 studies comprising 100 samples revealed a weak association of experimental variables with overall as well as individual gene expression. This can be explained by the large heterogeneity in cell culture practice, or the lack of adequate reporting thereof, as suggested by our systematic analysis of experimental parameters not included in the main analysis. Given the rapidly increasing use of in vitro cell culture models, including more advanced (micro) fluidic models, our analysis reinforces the need for improved, standardized reporting protocols. Additionally, our systematic analysis serves as a template for future comparative studies on in vitro transcriptome and other experimental data.

Les archées représentent l’un des trois domaines du vivant, au côté des bactéries et des eucaryotes. Très tôt après leur découverte, en 1977, l’auteur a entrepris l’étude de ces microorganismes. Il décrit la découverte dans son équipe à Orsay de nouvelles familles d’enzymes impliquées dans la modification des super-hélices d’ADN. L’une de ces enzymes, la reverse gyrase, est essentielle à la vie à très haute température, une autre, l’ADN topoisomérase VI « chez les archées », a permis d’identifier une protéine clef impliquée dans la diversification de notre matériel génétique. Archaea represent one of the three domains of life, alongside bacteria and eukaryotes. Very soon after their discovery, in 1977, the author undertook the study of these microorganisms. He describes the discovery in his team at Orsay of new families of enzymes involved in the modification of DNA superhelices. One of these enzymes, reverse gyrase, is essential for life at very high temperatures, another, DNA topoisomerase VI "in the archaea", has identified a key protein involved in the diversification of our genetic material

Mayaro virus (MAYV) is a mosquito-transmitted alphavirus that causes often debilitating rheumatic disease in tropical Central and South America. There are currently no licensed vaccines or antiviral drugs available for MAYV disease. Here, we generated Mayaro virus-like particles (VLPs) using the scalable baculovirus-insect cell expression system. High-level secretion of MAYV VLPs in the culture fluid of Sf9 insect cells was achieved, and particles with a diameter of 64 to 70 nm were obtained after purification. We characterize a C57BL/6J adult wild-type mouse model of MAYV infection and disease and used this model to compare the immunogenicity of VLPs from insect cells with that of VLPs produced in mammalian cells. Mice received two intramuscular immunizations with 1 μg of nonadjuvanted MAYV VLPs. Potent neutralizing antibody responses were generated against the vaccine strain, BeH407, with comparable activity seen against a contemporary 2018 isolate from Brazil (BR-18), whereas neutralizing activity against chikungunya virus was marginal. Sequencing of BR-18 illustrated that this virus segregates with genotype D isolates, whereas MAYV BeH407 belongs to genotype L. The mammalian cell-derived VLPs induced higher mean neutralizing antibody titers than those produced in insect cells. Both VLP vaccines completely protected adult wild-type mice against viremia, myositis, tendonitis, and joint inflammation after MAYV challenge. IMPORTANCE Mayaro virus (MAYV) is associated with acute rheumatic disease that can be debilitating and can evolve into months of chronic arthralgia. MAYV is believed to have the potential to emerge as a tropical public health threat, especially if it develops the ability to be efficiently transmitted by urban mosquito vectors, such as Aedes aegypti and/or Aedes albopictus. Here, we describe a scalable virus-like particle vaccine against MAYV that induced neutralizing antibodies against a historical and a contemporary isolate of MAYV and protected mice against infection and disease, providing a potential new intervention for MAYV epidemic preparedness.

Fan et al. challenge the finding that mitochondria represent a sister group to Alphaproteobacteria1, arguing that support for thisposition can be explained by unreliable site removal methods andoutgroup attraction. The authors chose to reduce compositionalheterogeneity by replacing AT-rich Rickettsiales and mitochondriawith GC-rich alternatives and attempted to attenuate long branchattraction (LBA) effects by removing all fast-evolving lineages butone. The study suggests that mitochondria form a sister clade toRickettsiales, within the ‘Alpha-II’ clade. Here we show that the association with Alpha-II is an artefact caused by a problematic taxonand that residual support for Rickettsiales-sister can be explained byconvergent evolution of the selected mitochondria and Rickettsialestowards high %GC. We further argue that site removal methods arein fact reliable and that outgroup attraction is unlikely to explain theAlphaproteobacteria-sister tree

BackgroundInfants are frequently exposed to antibiotics (AB) in the first week of life for suspected bacterial infections. Little is known about the effect of AB on the developing intestinal microbiota. Therefore, we studied intestinal microbiota development with and without AB exposure in the first week of life in term born infants.MethodsWe analysed the faecal microbiota from birth until 2.5 years of age by 16S rRNA gene amplicon sequencing in a cohort with 56 term born infants, exposed to AB in the first week of life (AB+) (AB for 2–3 days (AB2, n=20), AB for 7 days (AB7, n=36)), compared with 126 healthy controls (AB-). The effects of AB and duration were examined in relation to delivery and feeding mode.ResultsAB+ was associated with significantly increased relative abundance ofEnterobacteriaceaeat 3 weeks and 1 year and a decrease ofBifidobacteriaceae, from 1 week until 3 months of age only in vaginally delivered, but not in C-section born infants. Similar deviations were noted in AB7, but not in AB2. After AB, breastfed infants had lower relative abundance of potentially pathogenicEnterobacteriaceaecompared with formula fed infants and recovered 2 weeks faster towards controls.ConclusionsAB exposure in the first week of life alters faecal microbiota development with deviations in the relative abundance of individual taxa until 1 year of age. These alterations can have long-term health consequences, which emphasises the need for future studies aiming at restoring intestinal microbiota after AB administration.

Multiple models have been developed in the field to simulate growth and product accumulation of microalgal cultures. These models heavily depend on the accurate estimation of growth parameters. In this paper growth parameters are presented for three industrially relevant microalgae species: Nannochloropsis sp., Neochloris oleoabundans, and Picochlorum sp. (BPE23). Dedicated growth experiments were done in photobioreactors to determine the maximal biomass yield on light and maintenance rate, while oxygen evolution experiments were performed to estimate the maximal specific growth rate. Picochlorum sp. exhibited the highest specific growth rate of 4.98 ± 0.24 day−1 and the lowest specific maintenance rate of 0.079 day−1, whereas N. oleoabundans showed the highest biomass yield on light of 1.78 gx·molph−1. The measured growth parameters were used in a simple kinetic growth model for verification. When simulating growth under light conditions as found at Bonaire (12 °N, 68° W), Picochlorum sp. displayed the highest areal biomass productivity of 32.2 g.m−2·day−1 and photosynthetic efficiency of 2.8%. The presented growth parameters show to be accurate compared to experimental data and can be used for model calibration by scientists and industrial communities in the field.

Abstract In biodesulfurization (BD) at haloalkaline and dO2-limited conditions, sulfide-oxidizing bacteria (SOB) effectively convert sulfide into elemental sulfur that can be used in agriculture as a fertilizer and fungicide. Here we show which bacteria are present in this biotechnological process. 16S rRNA gene amplicon sequencing of biomass from ten reactors sampled in 2018 indicated the presence of 444 bacterial Amplicon Sequence Variants (ASVs). A core microbiome represented by 30 ASVs was found in all ten reactors, with Thioalkalivibrio sulfidiphilus as the most dominant species. The majority of these ASVs are phylogenetically related to bacteria previously identified in haloalkaline BD processes and in natural haloalkaline ecosystems. The source and composition of the feed gas had a great impact on the microbial community composition followed by alkalinity, sulfate, and thiosulfate concentrations. The halophilic SOB of the genus Guyparkeria (formerly known as Halothiobacillus) and heterotrophic SOB of the genus Halomonas were identified as potential indicator organisms of sulfate and thiosulfate accumulation in the BD process. Key points • Biodesulfurization (BD) reactors share a core microbiome • The source and composition of the feed gas affects the microbial composition in the BD reactors • Guyparkeria and Halomonas indicate high concentrations of sulfate and thiosulfate in the BD process

Billions of years of Darwinian evolution has led to the emergence of highly sophisticated and diverse life forms on Earth. Inspired by natural evolution, similar principles have been adopted in laboratory evolution for the fast optimization of genes and proteins for specific applications. In this review, we highlight state-of-the-art laboratory evolution strategies for protein engineering, with a special emphasis on in vitro strategies. We further describe how recent progress in microfluidic technology has allowed the generation and manipulation of artificial compartments for high-throughput laboratory evolution experiments. Expectations for the future are high: we foresee a revolution on-a-chip.

Rural household income growth may have two counteracting effects on energy-related carbon emissions, by stimulating total energy consumption and by changing the energy structure towards low-carbon energy sources. This study aims to provide more insight into this relationship for the case of rural central China, a region where coal-fired electricity dominates modern energy consumption. Based on survey data collected in 2019, energy-related carbon emissions (ECE) and energy-related carbon intensity (ECI) per unit of energy are calculated. A mediation effect model is employed to distinguish between energy structure changes and total electricity consumption as mediating factors in the income growth – carbon emission relationship. We argue that whether biomass fuels are treated as either carbon-intensive or as carbon neutral has major consequences for the analysis. When biomass fuel is treated as carbon-intensive, income-related changes in energy structure and electricity consumption have opposite effects on ECE, but both are conducive to the reduction of ECI. When biomass fuel is treated as carbon-neutral, the positive effect of growth in coal-fired electricity consumption on ECE is much larger than the negative effect of the change in energy structure. Based on these findings, we present several recommendations for balancing two main targets of China's rural revitalization vitalization policy, i.e. rural household income growth and the transition towards low-carbon energy sources in China.

Rosa (Rosaceae) is an important ornamental and medicinal plant genus worldwide, with several species being cultivated in China. Members of Sporocadaceae (pestalotioid fungi) are globally distributed and include endophytes, saprobes but also plant pathogens, infecting a broad range of host plants on which they can cause important plant diseases. Although several Sporocadaceae species were recorded to inhabit Rosa spp., the taxa occurring on Rosa remain largely unresolved. In this study, a total of 295 diseased samples were collected from branches, fruits, leaves and spines of eight Rosa species (R. chinensis, R. helenae , R. laevigata, R. multiflora, R. omeiensis, R. rugosa, R. spinosissima and R. xanthina) in Gansu, Henan, Hunan, Qinghai, Shaanxi Provinces and the Ningxia Autonomous Region of China. Subsequently 126 strains were obtained and identified based on comparisons of DNA sequence data. Based on these results 15 species residing in six genera of Sporocadaceae were delineated, including four known species (Pestalotiopsis chamaeropis, Pes. rhodomyrtus, Sporocadus sorbi and Spo. trimorphus) and 11 new species described here as Monochaetia rosarum, Neopestalotiopsis concentrica, N. subepidermalis, Pestalotiopsis tumida, Seimatosporium centrale, Seim. gracile, Seim. nonappendiculatum, Seim. parvum, Seiridium rosae, Sporocadus brevis, and Spo. spiniger. This study also represents the first report of Pes. chamaeropis, Pes. rhodomyrtus and Spo. sorbi on Rosa. The overall data revealed that Pestalotiopsis was the most prevalent genus, followed by Seimatosporium, while Pes. chamaeropis and Pes. rhodomyrtus were the two most prevalent species. Analysis of Sporocadaceae abundance on Rosa species and plant organs revealed that spines of R. chinensis had the highest species diversity.

Summary Eubacterium maltosivorans YIT is a human intestinal isolate capable of acetogenic, propionogenic and butyrogenic growth. Its 4.3-Mb genome sequence contains coding sequences for 4227 proteins, including 41 different methyltransferases. Comparative proteomics of strain YIT showed the WoodLjungdahl pathway proteins to be actively produced during homoacetogenic growth on H2 and CO2 while butyrogenic growth on a mixture of lactate and acetate significantly upregulated the production of proteins encoded by the recently identified lctABCDEF cluster and accessory proteins. Growth on H2 and CO2 unexpectedly induced the production of two related trimethylamine methyltransferases. Moreover, a set of 16 different trimethylamine methyltransferases together with proteins for bacterial microcompartments were produced during growth and deamination of the quaternary amines, betaine, carnitine and choline. Growth of strain YIT on 1,2-propanediol generated propionate with propanol and induced the formation of bacterial microcompartments that were also prominently visible in betaine-grown cells. The present study demonstrates that E. maltosivorans is highly versatile in converting low-energy fermentation end-products in the human gut into butyrate and propionate whilst being capable of preventing the formation of the undesired trimethylamine by converting betaine and other quaternary amines in bacterial microcompartments into acetate and butyrate., This work was supported by the SIAM Gravitation Grant 024.002.002 of the Netherlands Organization for Scientific Research., info:eu-repo/semantics/publishedVersion

The class 2 type V CRISPR effector Cas12 is thought to have evolved from the IS200/IS605 superfamily of transposon-associated TnpB proteins1. Recent studies have identified TnpB proteins as miniature RNA-guided DNA endonucleases2,3. TnpB associates with a single, long RNA (ωRNA) and cleaves double-stranded DNA targets complementary to the ωRNA guide. However, the RNA-guided DNA cleavage mechanism of TnpB and its evolutionary relationship with Cas12 enzymes remain unknown. Here we report the cryo-electron microscopy (cryo-EM) structure of Deinococcus radiodurans ISDra2 TnpB in complex with its cognate ωRNA and target DNA. In the structure, the ωRNA adopts an unexpected architecture and forms a pseudoknot, which is conserved among all guide RNAs of Cas12 enzymes. Furthermore, the structure, along with our functional analysis, reveals how the compact TnpB recognizes the ωRNA and cleaves target DNA complementary to the guide. A structural comparison of TnpB with Cas12 enzymes suggests that CRISPR–Cas12 effectors acquired an ability to recognize the protospacer-adjacent motif-distal end of the guide RNA–target DNA heteroduplex, by either asymmetric dimer formation or diverse REC2 insertions, enabling engagement in CRISPR–Cas adaptive immunity. Collectively, our findings provide mechanistic insights into TnpB function and advance our understanding of the evolution from transposon-encoded TnpB proteins to CRISPR–Cas12 effectors.

The present study systematically evaluated the potential of Candida subhashii, Fusarium solani and their consortium for the abatement of n-hexane, trichloroethylene (TCE), toluene and α-pinene in biofilters (BFs) and biotrickling filters (BTFs). Three 3.2 L BFs packed with polyurethane foam and operated at a gas residence time of 77 s with an air mixture of hydrophobic volatile organic compounds (VOCs) were inoculated with C. subhashii, F. solani and a combination of thereof. The systems were also operated under a BTF configuration with a liquid recirculating rate of 2.5 L h-1. Steady state elimination capacities (ECs) of total VOCs of 17.4 ± 0.7 g m−3 h−1 for C. subhashii, 21.2 ± 0.8 g m−3 h−1 for F. solani and 24.4 ± 1.4 g m−3 h−1 for their consortium were recorded in BFs, which increased up to 27.2 ± 1.6 g m−3 h−1, 29.2 ± 1.9 g m−3 h−1, 37.7 ± 3.3 g m−3 h−1 in BTFs. BTFs supported a superior biodegradation performance compared to BF, regardless of the VOCs. Moreover, a more effective VOC biodegradation was observed when C. subhashii and F. solani were grown as a consortium. The microbial analysis conducted revealed that the fungi initially introduced in each BF represented the dominant species by the end of the experiment, with C. subhashii gradually overcoming F. solani in the system inoculated with the fungal consortium.

Background Dilute ethanol streams generated during fermentation of biomass or syngas can be used as feedstocks for the production of higher value products. In this study, we describe a novel synthetic microbial co-culture that can effectively upgrade dilute ethanol streams to odd-chain carboxylic acids (OCCAs), specifically valerate and heptanoate. The co-culture consists of two strict anaerobic microorganisms: Anaerotignum neopropionicum, a propionigenic bacterium that ferments ethanol, and Clostridium kluyveri, well-known for its chain-elongating metabolism. In this co-culture, A. neopropionicum grows on ethanol and CO2 producing propionate and acetate, which are then utilised by C. kluyveri for chain elongation with ethanol as the electron donor. Results A co-culture of A. neopropionicum and C. kluyveri was established in serum bottles with 50 mM ethanol, leading to the production of valerate (5.4 ± 0.1 mM) as main product of ethanol-driven chain elongation. In a continuous bioreactor supplied with 3.1 g ethanol L−1 d−1, the co-culture exhibited high ethanol conversion (96.6%) and produced 25% (mol/mol) valerate, with a steady-state concentration of 8.5 mM and a rate of 5.7 mmol L−1 d−1. In addition, up to 6.5 mM heptanoate was produced at a rate of 2.9 mmol L−1 d−1. Batch experiments were also conducted to study the individual growth of the two strains on ethanol. A. neopropionicum showed the highest growth rate when cultured with 50 mM ethanol (μmax = 0.103 ± 0.003 h−1) and tolerated ethanol concentrations of up to 300 mM. Cultivation experiments with C. kluyveri showed that propionate and acetate were used simultaneously for chain elongation. However, growth on propionate alone (50 mM and 100 mM) led to a 1.8-fold reduction in growth rate compared to growth on acetate. Our results also revealed sub-optimal substrate use by C. kluyveri during odd-chain elongation, where excessive ethanol was oxidised to acetate. Conclusions This study highlights the potential of synthetic co-cultivation in chain elongation processes to target the production of OCCAs. Furthermore, our findings shed light on to the metabolism of odd-chain elongation by C. kluyveri.

Producción Científica, Biotechnologies are a feasible alternative for indoor air pollutant abatement. Among biotechnologies, bioactive coatings consist of microorganisms embedded in polymeric matrices, allowing a direct contact between microorganisms and gas pollutants, thus enhancing their abatement. Three bioreactors (BR1, BR2 and BR3) were inoculated with a VOC-degrading enriched culture, a latex bioactive coating with the VOC-degrading enriched culture, and a latex bioactive coating with fresh activated sludge. The influence of empty bed residence time (EBRT) and inlet concentrations on the removal of toluene, α-pinene and n-hexane was assessed. BR1 and BR2 achieved steady-state toluene and pinene removals >90% down to 30 s. BR3 lower removals could be attributed to the lack of acclimation of activated sludge. When inlet concentrations were progressively reduced to 80% in BR1 and BR2, but only to 64.2% in BR3. Pinene removals reached 90.9% in BR1, and >70% in BR2 and BR3. The bacterial population was dominated by Rhodococcus, Mycobacterium, Devosia and Rhodobacteraceae members in BR1 and BR2. No significant and robust hexane removal was observed regardless of the inoculum or operational conditions, probably due to mass transfer limitations, which entailed a low dominance of organisms with this metabolic capability., Ministerio de Ciencia, Innovación y Universidades (project RTI2018-0-096441-B-I00), Junta de Castilla y León y EU-FEDER (program grant number CLU 2017–09, CL-EI-2021-07 and UIC 315), European Commission-H2020-MSCA–IF–2019 grant (ENHANCEMENT, ref. 897284)

The dataset includes the raw data on the concentration of organic micropollutants, ammonia, total organic carbon, and other relevant contaminants. This dataset also includes the processed data, such as the loading rates and kinetic constants of organic micropollutants and the loading rates of ammonia and dissolved organic matter. In addition, the qPCR data is also present in this dataset. The details of methodology were given in the published article which is linked to this dataset.

The dataset includes raw / processed data on the removal of organic micropollutants (OMPs) and other contaminants in excel format. The dataset also contains the raw data on qPCR and 16S rRNA amplicon sequencing analysis. The completed 16S rRNA amplicon sequencing data have been uploaded to BioProject PRJEB48767 repository. The details of methodology were given in the published article which is linked to this dataset.

With an ever-increasing amount of (meta)genomic data being deposited in sequence databases, (meta)genome mining for natural product biosynthetic pathways occupies a critical role in the discovery of novel pharmaceutical drugs, crop protection agents and biomaterials. The genes that encode these pathways are often organised into biosynthetic gene clusters (BGCs). In 2015, we defined the Minimum Information about a Biosynthetic Gene cluster (MIBiG): a standardised data format that describes the minimally required information to uniquely characterise a BGC. We simultaneously constructed an accompanying online database of BGCs, which has since been widely used by the community as a reference dataset for BGCs and was expanded to 2021 entries in 2019 (MIBiG 2.0). Here, we describe MIBiG 3.0, a database update comprising large-scale validation and re-annotation of existing entries and 661 new entries. Particular attention was paid to the annotation of compound structures and biological activities, as well as protein domain selectivities. Together, these new features keep the database up-to-date, and will provide new opportunities for the scientific community to use its freely available data, e.g. for the training of new machine learning models to predict sequence-structure-function relationships for diverse natural products. MIBiG 3.0 is accessible online at https://mibig.secondarymetabolites.org/.