Diversity and Distribution of Marine Synechococcus: Multiple Gene Phylogenies for Consensus Classification and Development of qPCR Assays for Sensitive Measurement of Clades in the Ocean

Marine Synechococcus is a globally significant genus of cyanobacteria that is comprised of multiple genetic lineages or clades. These clades are thought to represent ecologically distinct units, or ecotypes. Because multiple clades often co-occur together in the oceans, Synechococcus are ideal microbes to explore how closely related bacterial taxa within the same functional guild of organisms co-exist and partition marine habitats. Here we sequenced multiple gene loci from cultured strains to confirm the congruency of clade classifications between the 16S–23S rDNA internally transcribed spacer (ITS), 16S rDNA, narB, ntcA, and rpoC1 loci commonly used in Synechococcus diversity studies. We designed quantitative PCR (qPCR) assays that target the ITS for 10 Synechococcus clades, including four clades, XV, XVI, CRD1, and CRD2, not covered by previous assays employing other loci. Our new qPCR assays are very sensitive and specific, detecting down to tens of cells per ml. Application of these qPCR assays to field samples from the northwest Atlantic showed clear shifts in Synechococcus community composition across a coastal to open-ocean transect. Consistent with previous studies, clades I and IV dominated cold, coastal Synechococcus communities. Clades II and X were abundant at the two warmer, off-shore stations, and at all stations multiple Synechococcus clades co-occurred. qPCR assays developed here provide valuable tools to further explore the dynamics of microbial community structure and the mechanisms of co-existence

Inherent Negative Refraction on Acoustic Branch of Two Dimensional Phononic Crystals

Guided by theoretical predictions, we have demonstrated experimentally the existence of negative refraction on the lowest two (acoustic) passbands (shear and longitudinal modes) of a simple two dimensional phononic crystal consisting of an isotropic stiff (aluminum) matrix and square- patterned isotropic compliant (PMMA) circular inclusions. At frequencies and wave vectors where the refraction is negative, the effective mass density and the effective stiffness tensors of the crystal can be positive-defnite, and that, this is an inherent property of phononic crystals with an isotropic stiff matrix containing periodically distributed isotropic compliant inclusions. The equi-frequency contours and energy ux vectors as fuctions of the phase-vector components, reveal a rich body of refractive properties that can be exploited to realize, for example, beam splitting, focusing, and frequency filtration on the lowest passbands of the crystal where the dissipation is the least. By proper selection of material and geometric parameters these phenomena can be realized at remarkably low frequencies (large wave lengths) using rather small simple two-phase unit cells. Keywords: Doubly periodic phononic crystals, acoustic branch negative refraction, beam splitting, focusing, imaging, frequency filtration at large wave length

Genome Sequence of the Estuarine Synechococcus sp. Strain NB0720_010

Synechococcus spp. are unicellular cyanobacteria widely distributed in the world\u27s oceans. We report the complete genome sequence of Synechococcus sp. strain NB0720_010, isolated from Narragansett Bay, Rhode Island. NB0702_10 has several large (.3,000-amino acid) protein-coding genes that may be important in its interactions with other cells, including grazers in estuarine habitats

Optimizing de novo genome assembly from PCR-amplified metagenomes

Background Metagenomics has transformed our understanding of microbial diversity across ecosystems, with recent advances enabling de novo assembly of genomes from metagenomes. These metagenome-assembled genomes are critical to provide ecological, evolutionary, and metabolic context for all the microbes and viruses yet to be cultivated. Metagenomes can now be generated from nanogram to subnanogram amounts of DNA. However, these libraries require several rounds of PCR amplification before sequencing, and recent data suggest these typically yield smaller and more fragmented assemblies than regular metagenomes. Methods Here we evaluate de novo assembly methods of 169 PCR-amplified metagenomes, including 25 for which an unamplified counterpart is available, to optimize specific assembly approaches for PCR-amplified libraries. We first evaluated coverage bias by mapping reads from PCR-amplified metagenomes onto reference contigs obtained from unamplified metagenomes of the same samples. Then, we compared different assembly pipelines in terms of assembly size (number of bp in contigs ≥ 10 kb) and error rates to evaluate which are the best suited for PCR-amplified metagenomes. Results Read mapping analyses revealed that the depth of coverage within individual genomes is significantly more uneven in PCR-amplified datasets versus unamplified metagenomes, with regions of high depth of coverage enriched in short inserts. This enrichment scales with the number of PCR cycles performed, and is presumably due to preferential amplification of short inserts. Standard assembly pipelines are confounded by this type of coverage unevenness, so we evaluated other assembly options to mitigate these issues. We found that a pipeline combining read deduplication and an assembly algorithm originally designed to recover genomes from libraries generated after whole genome amplification (single-cell SPAdes) frequently improved assembly of contigs ≥10 kb by 10 to 100-fold for low input metagenomes. Conclusions PCR-amplified metagenomes have enabled scientists to explore communities traditionally challenging to describe, including some with extremely low biomass or from which DNA is particularly difficult to extract. Here we show that a modified assembly pipeline can lead to an improved de novo genome assembly from PCR-amplified datasets, and enables a better genome recovery from low input metagenomes

A global experiment on motivating social distancing during the COVID-19 pandemic

Finding communication strategies that effectively motivate social distancing continues to be a global public health priority during the COVID-19 pandemic. This cross-country, preregistered experiment (n = 25,718 from 89 countries) tested hypotheses concerning generalizable positive and negative outcomes of social distancing messages that promoted personal agency and reflective choices (i.e., an autonomy-supportive message) or were restrictive and shaming (i.e., a controlling message) compared with no message at all. Results partially supported experimental hypotheses in that the controlling message increased controlled motivation (a poorly internalized form of motivation relying on shame, guilt, and fear of social consequences) relative to no message. On the other hand, the autonomy-supportive message lowered feelings of defiance compared with the controlling message, but the controlling message did not differ from receiving no message at all. Unexpectedly, messages did not influence autonomous motivation (a highly internalized form of motivation relying on one’s core values) or behavioral intentions. Results supported hypothesized associations between people’s existing autonomous and controlled motivations and self-reported behavioral intentions to engage in social distancing. Controlled motivation was associated with more defiance and less long-term behavioral intention to engage in social distancing, whereas autonomous motivation was associated with less defiance and more short- and long-term intentions to social distance. Overall, this work highlights the potential harm of using shaming and pressuring language in public health communication, with implications for the current and future global health challenges

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

New tRNA-targeting transposons that hijack phage and vesicles

Genomic islands are hotspots for horizontal gene transfer (HGT) in bacteria, but, for Prochlorococcus, an abundant marine cyanobacterium, how these islands form has puzzled scientists. With the discovery of tycheposons, a new family of transposons, Hackl et al. provide evidence for elegant new mechanisms of gene rearrangement and transfer among Prochlorococcus and bacteria more broadly. © 2023 Elsevier Lt

Culture Isolation and Culture-Independent Clone Libraries Reveal New Marine Synechococcus Ecotypes with Distinctive Light and N Physiologies

Marine microbial communities often contain multiple closely related phylogenetic clades, but in many cases, it is still unclear what physiological traits differentiate these putative ecotypes. The numerically abundant marine cyanobacterium Synechococcus can be divided into at least 14 clades. In order to better understand ecotype differentiation in this genus, we assessed the diversity of a Synechococcus community from a well-mixed water column in the Sargasso Sea during March 2002, a time of year when this genus typically reaches its annual peak in abundance. Diversity was estimated from water sampled at three depths (approximately 5, 70, and 170 m) using both culture isolation and construction of cyanobacterial 16S-23S rRNA internal transcribed sequence clone libraries. Clonal isolates were obtained by enrichment with ammonium, nitrite, or nitrate as the sole N source, followed by pour plating. Each method sampled the in situ diversity differently. The combined methods revealed a total of seven Synechococcus phylotypes including two new putative ecotypes, labeled XV and XVI. Although most other isolates grow on nitrate, clade XV exhibited a reduced efficiency in nitrate utilization, and both clade XV and XVI are capable of chromatic adaptation, demonstrating that this trait is more widely distributed among Synechococcus strains than previously known. Thus, as in its sister genus Prochlorococcus, light and nitrogen utilization are important factors in ecotype differentiation in the marine Synechococcus lineage