Flow-cytometric quantification of microbial cells on sand from water biofilters

Rapid quantification of absolute microbial cell abundances is important for a comprehensive interpretation of microbiome surveys and crucial to support theoretical modelling and the design of engineered systems. In this paper, we propose a protocol specifically optimised for the quantification of microbial abundances in water biofilters using flow cytometry (FCM). We optimised cell detachment from sand biofilter particles for FCM quantification through the evaluation of five chemical dispersants (NaCl, Triton-X100, CaCl2, sodium pyrophosphate (PP), Tween 80 combined with PP), different mechanical pre-treatments (low and high energy sonication and shaking) and two fixation methods (glutaraldehyde and ethanol). The developed protocol was cross-compared using other established and commonly employed methods for biomass quantification in water filter samples (adenosine triphosphate (ATP) quantification, real-time quantitative PCR (qPCR) and volatile solids (VS)). The highest microbial count was obtained by detaching the biofilm from biofilter grains and dispersing clusters into singles cells using Tween 80 and sodium pyrophosphate combined with four steps of high energy sonication (27W, for 80 s each step); glutaraldehyde was shown to be the best fixative solution. The developed protocol was reliable and highly reproducible and produced results that are comparable to data from alternative quantification methods. Indeed, high correlations were found with trends obtained through ATP and qPCR (ρ = 0.98 and ρ = 0.91) measurements. The VS content was confirmed as an inaccurate method to express biomass in sand samples since it correlated poorly with all the other three methods (ρ = 0.005 with FCM, 0.002 with ATP and 0.177 with qPCR). FCM and ATP showed the strongest agreement between absolute counts with a slope of the correlation equal to 0.7, while qPCR seemed to overestimate cell counts by a factor of ten. The rapidity and reproducibility of the method developed make its application ideal for routine quantification of microbial cell abundances on sand from water biofilters and thus useful in revealing the ecological patterns and quantifying the metabolic kinetics involved in such systems

The dual endothelin converting enzyme/neutral endopeptidase inhibitor SLV-306 (daglutril), inhibits systemic conversion of big endothelin-1 in humans

Aims - Inhibition of neutral endopeptidases (NEP) results in a beneficial increase in plasma concentrations of natriuretic peptides such as ANP. However NEP inhibitors were ineffective anti-hypertensives, probably because NEP also degrades vasoconstrictor peptides, including endothelin-1 (ET-1). Dual NEP and endothelin converting enzyme (ECE) inhibition may be more useful. The aim of the study was to determine whether SLV-306 (daglutril), a combined ECE/NEP inhibitor, reduced the systemic conversion of big ET-1 to the mature peptide. Secondly, to determine whether plasma ANP levels were increased. Main methods - Following oral administration of three increasing doses of SLV-306 (to reach an average target concentration of 75, 300, 1200 ng ml− 1 of the active metabolite KC-12615), in a randomised, double blinded regime, big ET-1 was infused into thirteen healthy male volunteers. Big ET-1 was administered at a rate of 8 and 12 pmol kg− 1 min− 1 (20 min each). Plasma samples were collected pre, during and post big ET-1 infusion. ET-1, C-terminal fragment (CTF), big ET-1, and atrial natriuretic peptide (ANP) were measured. Key findings - At the two highest concentrations tested, SLV-306 dose dependently attenuated the rise in blood pressure after big ET-1 infusion. There was a significant increase in circulating big ET-1 levels, compared with placebo, indicating that SLV-306 was inhibiting an increasing proportion of endogenous ECE activity. Plasma ANP concentrations also significantly increased, consistent with systemic NEP inhibition. Significance - SLV-306 leads to inhibition of both NEP and ECE in humans. Simultaneous augmentation of ANP and inhibition of ET-1 production is of potential therapeutic benefit in cardiovascular disease

Medium shapes the microbial community of water filters with implications for effluent quality

Little is known about the forces that determine the assembly of diverse bacterial communities inhabiting drinking water treatment filters and how this affects drinking water quality. Two contrasting ecological theories can help to understand how natural microbial communities assemble; niche theory and neutral theory, where environmental deterministic factors or stochastic factors predominate respectively. This study investigates the development of the microbial community on two common contrasting filter materials (quartz sand and granular activated carbon-GAC), to elucidate the main factors governing their assembly, through the evaluation of environmental (i.e. filter medium type) and stochastic forces (random deaths, births and immigration). Laboratory-scale filter columns were used to mimic a rapid gravity filter; the microbiome of the filter materials, and of the filter influent and effluent, was characterised using next generation 16S rRNA gene amplicon sequencing and flow-cytometry. Chemical parameters (i.e. dissolved organic carbon, trihalomethanes formation) were also monitored to assess the final effluent quality. The filter communities seemed to be strongly assembled by selection rather than neutral processes, with only 28% of those OTUs shared with the source water detected on the filter medium following predictions using a neutral community model. GAC hosted a phylogenetically more diverse community than sand. The two filter media communities seeded the effluent water, triggering differences in both water quality and community composition of the effluents. Overall, GAC proved to be better than sand in controlling microbial growth, by promoting higher bacterial decay rates and hosting less bacterial cells, and showed better performance for putative pathogen control by leaking less Legionella cells into the effluent water

Diversity and Dispersal Among Eastern Continental Divide Headwater Stream Fishes in Gwinnett County, Georgia.

The eastern continental divide that bisects Georgia runs through Gwinnett County, separating headwater streams of the western Chattahoochee River watershed from headwater streams of the eastern Ocmulgee and Oconee River watersheds. This landscape feature was used to test hypotheses regarding headwater habitat, fish diversity and gene flow. Headwater habitats are dominant components of river network ecosystems delivering vital ecosystem services and biodiversity. Three headwater streams, one in the Chattahoochee watershed and two in the Oconee watershed, on Gwinnett County Park property, were sampled for differences in physical and chemical properties and fish inter- and intraspecific diversity. Our results suggest the headwater habitats are each distinguished by unique physical and temporal features. Initial hypotheses regarding expectations of fish diversity based on habitat type were not supported. However, hypotheses regarding the impact of gene flow on genetic diversity among headwater stream species were supported. Gene flow estimates and phylogenetic analyses among three well-sampled species observed in these headwater streams, Semotilus atromaculatus, Notropis lutipinnis, and Nocomis leptocephalus, suggest the Eastern Continental Divide acts as a barrier for gene flow for some species. Our findings highlight a proposed methodology for headwater stream analysis that combines habitat heterogeneity with community and species-level measures of diversity

A quantitative structure-biodegradation relationship (QSBR) approach to predict biodegradation rates of aromatic chemicals

The objective of this work was to develop a QSBR model for the prioritization of organic pollutants based on biodegradation rates from a database containing globally harmonized biodegradation tests using relevant molecular descriptors. To do this, we first categorized the chemicals into three groups (Group 1: simple aromatic chemicals with a single ring, Group 2: aromatic chemicals with multiple rings and Group3: Group 1 plus Group 2) based on molecular descriptors, estimated the first order biodegradation rate of the chemicals using rating values derived from the BIOWIN3 model, and finally developed, validated and defined the applicability domain of models for each group using a multiple linear regression approach. All the developed QSBR models complied with OECD principles for QSAR validation. The biodegradation rate in the models for the two groups (Group 2 and 3 chemicals) are associated with abstract molecular descriptors that provide little relevant practical information towards understanding the relationship between chemical structure and biodegradation rates. However, molecular descriptors associated with the QSBR model for Group 1 chemicals (R2 = 0.89, Q2loo = 0.87) provided information on properties that can readily be scrutinised and interpreted in relation to biodegradation processes. In combination, these results lead to the conclusion that QSBRs can be an alternative tool to estimate the persistence of chemicals, some of which can provide further insights into those factors affecting biodegradation

A “piano movers” problem reformulated

Abstract-It has long been known that cylindrical algebraic decompositions (CADs) can in theory be used for robot motion planning. However, in practice even the simplest examples can be too complicated to tackle. We consider in detail a "Piano Mover's Problem" which considers moving an infinitesimally thin piano (or ladder) through a right-angled corridor. Producing a CAD for the original formulation of this problem is still infeasible after 25 years of improvements in both CAD theory and computer hardware. We review some alternative formulations in the literature which use differing levels of geometric analysis before input to a CAD algorithm. Simpler formulations allow CAD to easily address the question of the existence of a path. We provide a new formulation for which both a CAD can be constructed and from which an actual path could be determined if one exists, and analyse the CADs produced using this approach for variations of the problem. This emphasises the importance of the precise formulation of such problems for CAD. We analyse the formulations and their CADs considering a variety of heuristics and general criteria, leading to conclusions about tackling other problems of this form

Program Veri¿cation in the presence of complex numbers, functions with branch cuts etc

Abstract-In considering the reliability of numerical programs, it is normal to "limit our study to the semantics dealing with numerical precision" (Martel, 2005). On the other hand, there is a great deal of work on the reliability of programs that essentially ignores the numerics. The thesis of this paper is that there is a class of problems that fall between these two, which could be described as "does the lowlevel arithmetic implement the high-level mathematics". Many of these problems arise because mathematics, particularly the mathematics of the complex numbers, is more dif¿cult than expected: for example the complex function log is not continuous, writing down a program to compute an inverse function is more complicated than just solving an equation, and many algebraic simpli¿cation rules are not universally valid. The good news is that these problems are theoretically capable of being solved, and are practically close to being solved, but not yet solved, in several real-world examples. However, there is still a long way to go before implementations match the theoretical possibilities

Predicting the effects of biochar on volatile petroleum hydrocarbon biodegradation and emanation from soil: a bacterial community finger-print analysis inferred modelling approach

We investigated the response of the dominant bacterial taxa in gravelly sand to the addition of biochar and/or mixtures of volatile petroleum hydrocarbons (VPHs) using denaturing gradient gel electrophoresis (DGGE) and sequencing of cut bands. Biochar addition alone had only weak effects on the soil bacterial community composition in batch study samples, while VPH addition had strong effects. Indirect effects of biochar on soil bacterial communities were apparent in column study samples, where biochar-enhanced sorption affected VPH spreading. Following VPH addition, cell abundance increased by no more than a factor of 2 and several Pseudomonas spp. became dominant in soil with and without biochar. We present a VPH fate model that considers soil bacterial biomass dynamics and a nutrient limited soil biomass carrying capacity. The model simulates an apparent lag phase before the onset of a brief period of intensive VPH biodegradation and biomass growth, which is followed by substantially slower VPH biodegradation, when nitrogen needs to be recycled between decaying and newly formed biomass. If biomass growth is limited by a factor other than the organic pollutant bioavailability, biochar amendment may enhance VPH attenuation in between a VPH source below ground and the atmosphere by reducing the risk of overloading the soil's biodegradation capacity