Back to Basics: A Simplified Improvement to Multiple Displacement Amplification for Microbial Single-Cell Genomics

Microbial single-cell genomics (SCG) provides access to the genomes of rare and uncultured microorganisms and is a complementary method to metagenomics. Due to the femtogram-levels of DNA in a single microbial cell, sequencing the genome requires whole genome amplification (WGA) as a preliminary step. However, the most common WGA method, multiple displacement amplification (MDA), is known to be costly and biased against specific genomic regions, preventing high-throughput applications and resulting in uneven genome coverage. Thus, obtaining high-quality genomes from many taxa, especially minority members of microbial communities, becomes difficult. Here, we present a volume reduction approach that significantly reduces costs while improving genome coverage and uniformity of DNA amplification products in standard 384-well plates. Our results demonstrate that further volume reduction in specialized and complex setups (e.g., microfluidic chips) is likely unnecessary to obtain higher-quality microbial genomes. This volume reduction method makes SCG more feasible for future studies, thus helping to broaden our knowledge on the diversity and function of understudied and uncharacterized microorganisms in the environment

Microbial single-cell omics: the crux of the matter

Single-cell genomics and transcriptomics can provide reliable context for assembled genome fragments and gene expression activity on the level of individual prokaryotic genomes. These methods are rapidly emerging as an essential complement to cultivation-based, metagenomics, metatranscriptomics, and microbial community-focused research approaches by allowing direct access to information from individual microorganisms, even from deep-branching phylogenetic groups that currently lack cultured representatives. Their integration and binning with environmental ‘omics data already provides unprecedented insights into microbial diversity and metabolic potential, enabling us to provide information on individual organisms and the structure and dynamics of natural microbial populations in complex environments. This review highlights the pitfalls and recent advances in the field of single-cell omics and its importance in microbiological and biotechnological studies

Single-Cell (Meta-)Genomics of a Dimorphic Candidatus Thiomargarita nelsonii Reveals Genomic Plasticity

The genus Thiomargarita includes the world's largest bacteria. But as uncultured organisms, their physiology, metabolism, and basis for their gigantism are not well understood. Thus, a genomics approach, applied to a single Candidatus Thiomargarita nelsonii cell was employed to explore the genetic potential of one of these enigmatic giant bacteria. The Thiomargarita cell was obtained from an assemblage of budding Ca. T. nelsonii attached to a provannid gastropod shell from Hydrate Ridge, a methane seep offshore of Oregon, USA. Here we present a manually curated genome of Bud S10 resulting from a hybrid assembly of long Pacific Biosciences and short Illumina sequencing reads. With respect to inorganic carbon fixation and sulfur oxidation pathways, the Ca. T nelsonii Hydrate Ridge Bud S10 genome was similar to marine sister taxa within the family Beggiatoaceae. However, the Bud S10 genome contains genes suggestive of the genetic potential for lithotrophic growth on arsenite and perhaps hydrogen. The genome also revealed that Bud 510 likely respires nitrate via two pathways: a complete denitrification pathway and a dissimilatory nitrate reduction to ammonia pathway. Both pathways have been predicted, but not previously fully elucidated, in the genomes of other large, vacuolated, sulfur-oxidizing bacteria. Surprisingly, the genome also had a high number of unusual features for a bacterium to include the largest number of metacaspases and introns ever reported in a bacterium. Also present, are a large number of other mobile genetic elements, such as insertion sequence (IS) transposable elements and miniature inverted-repeat transposable elements (MITEs). In some cases, mobile genetic elements disrupted key genes in metabolic pathways. For example, a MITE interrupts hupL, which encodes the large subunit of the hydrogenase in hydrogen oxidation. Moreover, we detected a group I intron in one of the most critical genes in the sulfur oxidation pathway, dsrA. The dsrA group I intron also carried a MITE sequence that, like the hupL MITE family, occurs broadly across the genome. The presence of a high degree of mobile elements in genes central to Thiomargarita's core metabolism has not been previously reported in free-living bacteria and suggests a highly mutable genome

Updates to the recently introduced family Lacipirellulaceae in the phylum Planctomycetes: isolation of strains belonsging to the novel genera Aeoliella, Botrimarina, Pirellulimonas and Pseudobythopirellula and the novel species Bythopirellula polymerisocia and Posidoniimonas corsicana

Eight novel strains of the phylum Planctomycetes were isolated from different aquatic habitats. Among these habitats were the hydrothermal vent system close to Panarea Island, a public beach at Mallorca Island, the shore of Costa Brava (Spain), and three sites with brackish water in the Baltic Sea. The genome sizes of the novel strains range from 4.33 to 6.29 Mb with DNA G+C contents between 52.8 and 66.7%. All strains are mesophilic (Topt 24–30 °C) and display generation times between 17 and 94 h. All eight isolates constitute novel species of either already described or novel genera within the family Lacipirellulaceae. Two of the novel species, Posidoniimonas polymericola (type strain Pla123a$^{T}$ = DSM 103020$^{T}$ = LMG 29466$^{T}$) and Bythopirellula polymerisocia (type strain Pla144$^{T}$ = DSM 104841$^{T}$ = VKM B-3442$^{T}$), belong to established genera, while the other strains represent the novel genera Aeoliella gen. nov., Botrimarina gen. nov., Pirellulimonas gen. nov. and Pseudobythopirellula gen. nov. Based on our polyphasic analysis, we propose the species Aeoliella mucimassa sp. nov. (type strain Pan181$^{T}$ = DSM 29370$^{T}$ = LMG 31346$^{T}$ = CECT 9840$^{T}$ = VKM B-3426$^{T}$), Botrimarina colliarenosi sp. nov. (type strain Pla108$^{T}$ = DSM 103355$^{T}$ = LMG 29803$^{T}$), Botrimarina hoheduenensis sp. nov. (type strain Pla111$^{T}$ = DSM 103485$^{T}$ = STH00945$^{T}$, Jena Microbial Resource Collection JMRC), Botrimarina mediterranea sp. nov. (type strain Spa11$^{T}$ = DSM 100745$^{T}$ = LMG 31350$^{T}$ = CECT 9852$^{T}$ = VKM B-3431$^{T}$), Pirellulimonas nuda sp. nov. (type strain Pla175$^{T}$ = DSM 109594$^{T}$ = CECT 9871$^{T}$ = VKM B-3448$^{T}$) and Pseudobythopirellula maris sp. nov. (type strain Mal64$^{T}$ = DSM 100832$^{T}$ = LMG 29020$^{T}$)

Additions to the genus Gimesia: description of Gimesia alba sp. nov., Gimesia algae sp. nov., Gimesia aquarii sp. nov., Gimesia aquatilis sp. nov., Gimesia fumaroli sp. nov. and Gimesia panareensis sp. nov., isolated from aquatic habitats of the Northern Hemisphere

Thirteen novel planctomycetal strains were isolated from five different aquatic sampling locations. These comprise the hydrothermal vent system close to Panarea Island (Italy), a biofilm on the surface of kelp at Monterey Bay (CA, USA), sediment and algae on Mallorca Island (Spain) and Helgoland Island (Germany), as well as a seawater aquarium in Braunschweig, Germany. All strains were shown to belong to the genus Gimesia. Their genomes cover a size range from 7.22 to 8.29 Mb and have a G+C content between 45.1 and 53.7%. All strains are mesophilic (Topt 26-33 °C) with generation times between 12 and 32 h. Analysis of fatty acids yielded palmitic acid (16:0) and a fatty acid with the equivalent chain length of 15.817 as major compounds. While five of the novel strains belong to the already described species Gimesia maris and Gimesia chilikensis, the other strains belong to novel species, for which we propose the names Gimesia alba (type strain Pan241wT = DSM 100744T = LMG 31345T = CECT 9841T = VKM B-3430T), Gimesia algae (type strain Pan161T = CECT 30192T = STH00943T = LMG 29130T), Gimesia aquarii (type strain V144T = DSM 101710T = VKM B-3433T), Gimesia fumaroli (type strain Enr17T = DSM 100710T = VKM B-3429T) and Gimesia panareensis (type strain Enr10T = DSM 100416T = LMG 29082T). STH numbers refer to the Jena Microbial Resource Collection (JMRC)

Desenvolvimento de cultivares de soja com tolerância à deficiência hídrica.

Caracterização dos ambientes. Caracterização genética. Estratégias de melhoramento. Lançamento comercial de cultivares tolerantes à seca

Macroporous Silicone Chips for Decoding Microbial Dark Matter in Environmental Microbiomes

Natural evolution has produced an almost infinite variety of microorganisms that can colonize almost any conceivable habitat. Since the vast majority of these microbial consortia are still unknown, there is a great need to elucidate this “microbial dark matter” (MDM) to enable exploitation in biotechnology. We report the fabrication and application of a novel device that integrates a matrix of macroporous elastomeric silicone foam (MESIF) into an easily fabricated and scalable chip design that can be used for decoding MDM in environmental microbiomes. Technical validation, performed with the model organism Escherichia coli expressing a fluorescent protein, showed that this low-cost, bioinert, and widely modifiable chip is rapidly colonized by microorganisms. The biological potential of the chip was then illustrated through targeted sampling and enrichment of microbiomes in a variety of habitats ranging from wet, turbulent moving bed biofilters and wastewater treatment plants to dry air-based environments. Sequencing analyses consistently showed that MESIF chips are not only suitable for sampling with high robustness but also that the material can be used to detect a broad cross section of microorganisms present in the habitat in a short time span of a few days. For example, results from the biofilter habitat showed efficient enrichment of microorganisms belonging to the enigmatic Candidate Phyla Radiation, which comprise ∼70% of the MDM. From dry air, the MESIF chip was able to enrich a variety of members of Actinobacteriota, which is known to produce specific secondary metabolites. Targeted sampling from a wastewater treatment plant where the herbicide glyphosate was added to the chip’s reservoir resulted in enrichment of Cyanobacteria and Desulfobacteria, previously associated with glyphosate degradation. These initial case studies suggest that this chip is very well suited for the systematic study of MDM and opens opportunities for the cultivation of previously unculturable microorganisms

A Systematic Analysis of Temporal Trends in the Handgrip Strength of 2,216,320 Children and Adolescents Between 1967 and 2017

Objective: To estimate national and international temporal trends in handgrip strength for children and adolescents, and to examine relationships between trends in handgrip strength and trends in health-related and sociodemographic indicators. Methods: Data were obtained through a systematic search of studies reporting temporal trends in the handgrip strength for apparently healthy 9–17 year-olds, and by examining large national fitness datasets. Temporal trends at the country-sex-age level were estimated by sample-weighted regression models relating the year of testing to mean handgrip strength. International and national trends were estimated by a post-stratified population-weighting procedure. Pearson’s correlations quantified relationships between trends in handgrip strength and trends in health-related/sociodemographic indicators. Results: 2,216,320 children and adolescents from 13 high-, 5 upper-middle-, and 1 low-income countries/special administrative regions between 1967 and 2017 collectively showed a moderate improvement of 19.4% (95%CI: 18.4 to 20.4) or 3.8% per decade (95%CI: 3.6 to 4.0). The international rate of improvement progressively increased over time, with more recent values (post-2000) close to two times larger than those from the 1960s/1970s. Improvements were larger for children (9–12 years) compared to adolescents (13–17 years), and similar for boys and girls. Trends differed between countries, with relationships between trends in handgrip strength and trends in health-related/sociodemographic indicators negligible-to-weak and not statistically significant. Conclusions: There has been a substantial improvement in absolute handgrip strength for children and adolescents since 1967. There is a need for improved international surveillance of handgrip strength, especially in low- and middle-income countries, to more confidently determine true international trends. PROSPERO registration number: CRD42013003657