Multiple holins contribute to extracellular DNA release in Pseudomonas aeruginosa biofilms

Bacterial biofilms are composed of aggregates of cells encased within a matrix of extracellular polymeric substances (EPS). One key EPS component is extracellular DNA (eDNA), which acts as a ‘glue’, facilitating cell–cell and cell–substratum interactions. We have previously demonstrated that eDNA is produced in Pseudomonas aeruginosa biofilms via explosive cell lysis. This phenomenon involves a subset of the bacterial population explosively lysing, due to peptidoglycan degradation by the endolysin Lys. Here we demonstrate that in P. aeruginosa three holins, AlpB, CidA and Hol, are involved in Lys-mediated eDNA release within both submerged (hydrated) and interstitial (actively expanding) biofilms, albeit to different extents, depending upon the type of biofilm and the stage of biofilm development. We also demonstrate that eDNA release events determine the sites at which cells begin to cluster to initiate microcolony formation during the early stages of submerged biofilm development. Furthermore, our results show that sustained release of eDNA is required for cell cluster consolidation and subsequent microcolony development in submerged biofilms. Overall, this study adds to our understanding of how eDNA release is controlled temporally and spatially within P. aeruginosa biofilms

The FLUXNET2015 dataset and the ONEFlux processing pipeline for eddy covariance data

The FLUXNET2015 dataset provides ecosystem-scale data on CO2, water, and energy exchange between the biosphere and the atmosphere, and other meteorological and biological measurements, from 212 sites around the globe (over 1500 site-years, up to and including year 2014). These sites, independently managed and operated, voluntarily contributed their data to create global datasets. Data were quality controlled and processed using uniform methods, to improve consistency and intercomparability across sites. The dataset is already being used in a number of applications, including ecophysiology studies, remote sensing studies, and development of ecosystem and Earth system models. FLUXNET2015 includes derived-data products, such as gap-filled time series, ecosystem respiration and photosynthetic uptake estimates, estimation of uncertainties, and metadata about the measurements, presented for the first time in this paper. In addition, 206 of these sites are for the first time distributed under a Creative Commons (CC-BY 4.0) license. This paper details this enhanced dataset and the processing methods, now made available as open-source codes, making the dataset more accessible, transparent, and reproducible.Peer reviewe

Climate control of terrestrial carbon exchange across biomes and continents

Peer reviewe

Comparison of N2O emissions and gene abundances between wastewater nitrogen removal systems

Biological nitrogen removal (BNR) systems are increasingly used in the United States in both centralized wastewater treatment plants (WWTPs) and decentralized advanced onsite wastewater treatment systems (OWTS) to reduce N discharged in wastewater effluent. However, the potential for BNR systems to be sources of nitrous oxide (N2O), a potent greenhouse gas, needs to be evaluated to assess their environmental impact. We quantified and compared N2O emissions from BNR systems at a WWTP (Field\u27s Point, Providence, RI) and three types of advanced OWTS (Orenco Advantex AX 20, SeptiTech Series D, and Bio-Microbics MicroFAST) in nine Rhode Island residences (n = 3 per type) using cavity ring-down spectroscopy. We also used quantitative polymerase chain reaction to determine the abundance of genes from nitrifying (amoA) and denitrifying (nosZ) microorganisms that may be producing N2O in these systems. Nitrous oxide fluxes ranged from -4 × 10-3 to 3 × 10-1 mmol N2O m-2 s-1 and in general followed the order: centralized WWTP \u3e Advantex \u3e SeptiTech \u3e FAST. In contrast, when N2O emissions were normalized by population served and area of treatment tanks, all systems had overlapping ranges. In general, the emissions of N2O accounted for a small fraction (\u3c 1%) of N removed. There was no significant relationship between the abundance of nosZ or amoA genes and N2O emissions. This preliminary analysis highlights the need to evaluate N2O emissions from wastewater systems as a wider range of technologies are adopted. A better understanding of the mechanisms of N2O emissions will also allow us to better manage systems to minimize emissions

Comparison of SARS-CoV-2 evolution in paediatric primary airway epithelial cell cultures compared with Vero-derived cell lines

SARS-CoV-2 can efficiently infect both children and adults, albeit with morbidity and mortality positively associated with increasing host age and presence of co-morbidities. SARS-CoV-2 continues to adapt to the human population, resulting in several variants of concern (VOC) with novel properties, such as Alpha and Delta. However, factors driving SARS-CoV-2 fitness and evolution in paediatric cohorts remain poorly explored. Here, we provide evidence that both viral and host factors co-operate to shape SARS-CoV-2 genotypic and phenotypic change in primary airway cell cultures derived from children. Through viral whole-genome sequencing, we explored changes in genetic diversity over time of two pre-VOC clinical isolates of SARS-CoV-2 during passage in paediatric well-differentiated primary nasal epithelial cell (WD-PNEC) cultures and in parallel, in unmodified Vero-derived cell lines. We identified a consistent, rich genetic diversity arising in vitro, variants of which could rapidly rise to near fixation within two passages. Within isolates, SARS-CoV-2 evolution was dependent on host cells, with paediatric WD-PNECs showing a reduced diversity compared to Vero (E6) cells. However, mutations were not shared between strains. Furthermore, comparison of both Vero-grown isolates on WD-PNECs disclosed marked growth attenuation mapping to the loss of the polybasic cleavage site (PBCS) in Spike, while the strain with mutations in Nsp12 (T293I), Spike (P812R) and a truncation of Orf7a remained viable in WD-PNECs. Altogether, our work demonstrates that pre-VOC SARS-CoV-2 efficiently infects paediatric respiratory epithelial cells, and its evolution is restrained compared to Vero (E6) cells, similar to the case of adult cells. We highlight the significant genetic plasticity of SARS-CoV-2 while uncovering an influential role for collaboration between viral and host cell factors in shaping viral evolution and ultimately fitness in human respiratory epithelium