A qPCR assay for Bordetella pertussis cells that enumerates both live and dead bacteria

Bordetella pertussis is the causative agent of whooping cough, commonly referred to as pertussis. Although the incidence of pertussis was reduced through vaccination, during the last thirty years it has returned to high levels in a number of countries. This resurgence has been linked to the switch from the use of whole-cell to acellular vaccines. Protection afforded by acellular vaccines appears to be short-lived compared to that afforded by whole cell vaccines. In order to inform future vaccine improvement by identifying immune correlates of protection, a human challenge model of B. pertussis colonisation has been developed. Accurate measurement of colonisation status in this model has required development of a qPCR-based assay to enumerate B. pertussis in samples that distinguishes between viable and dead bacteria. Here we report the development of this assay and its performance in the quantification of B. pertussis from human challenge model samples. This assay has future utility in diagnostic labs and in research where a quantitative measure of both B. pertussis number and viability is required

Fragmentation Characteristics of Collision-Induced Dissociation in MALDI TOF/TOF Mass Spectrometry

The identification of proteins by tandem mass spectrometry relies on knowledge of the products produced by collision-induced dissociation of peptide ions. Most previous work has focused on fragmentation statistics for ion trap systems. We analyzed fragmentation in MALDI TOF/TOF mass spectrometry, collecting statistics using a curated set of 2459 MS/MS spectra, and applying bootstrap resampling to assess confidence intervals. We calculated the frequency of 18 product ion types, the correlation between both mass and intensity with ion type, the dependence of amide bond breakage on the residues surrounding the cleavage site, and the dependence of product ion detection on residues not adjacent to the cleavage site. The most frequently observed were internal ions, followed by y ions. A strong correlation between ion type and the mass and intensity of its peak was observed, with b and y ions producing the most intense and highest-mass peaks. The amino acids P, W, D and R had a strong effect on amide bond cleavage when situated next to the breakage site, whereas residues including I, K and H had a strong effect on product ion observation when located in the peptide but not adjacent to the cleavage site, a novel observation

Microbial growth in simulated martian environments

In this study, four new simulants have been developed, and their associated fluid chemistries have been derived for use in a series of microbiological simulation experiments. These experiments will determine if aqueous environments on Mars, past or present, could potentially support microbial life and identify any key geochemical biosignatures that may arise as a result of that life

Testing the habitability of distinct simulated martian environments

Habitability of martian waters would have been partially determined by the chemistry arising from interactions with martian lithologies. In this study, the habitability of groundwater chemistries (based on basaltic, iron- and sulfur-enriched lithologies) and the resulting variation in biosignatures was investigated, with microbes from anaerobic estuarine sediment used as an inoculum. The microbial community was monitored by cell counts and 16S rRNA gene profiling. Changes in fluid and precipitate chemistries were measured using ICP-OES and IC, with changes over geological timescales modelled using CHIM-XPT. The fluid chemistries were shown to be habitable, with distinct patterns in cell abundance and growth phases between the chemistries. However, the same genera dominated (Acetobacterium, Desulfovibrio and Desulfosporomusa) regardless of the initial fluid chemistry. In the biotic test group, changes in fluid chemistry were the same in the three chemistries, with an enhanced concentration of aluminium and iron and the removal of sulfate. However, geochemical modelling of the fluids under abiotic conditions over geological timescales revealed similar changes to those in the biotic test groups. Therefore, these samples require further analysis to assess whether we can identify any potentially unambiguous biosignatures that could develop between geologically distinct sites

Characterising the Transfer of Biomarkers within the Phobos-Mars System

Procedural and analytical developments required for impact and heat investigation into the detection of biomarkers transported from Mars to Phobos

Improving the efficacy of selenium fertilizers for wheat biofortification

© 2019, The Author(s). Increasing the selenium (Se) concentration of staple crops by fertilization is a valuable pathway to increase Se in the human diet, thus preventing Se deficiency. A pot trial was set up to investigate whether the application of 3.33 µg kg−1 of Se (equivalent to 10 g ha−1) to wheat can be made more efficient by its co-application with macronutrient carriers, either to the soil or to the leaves. In the soil, Se was applied either on its own (selenate only) or as a granular, Se-enriched macronutrient fertilizer supplying nitrogen, phosphorus, potassium or sulfur. Selenium was also applied to leaves at head emergence with, or without, 2% w/v N fertilizers. With grain Se concentrations varying from 0.13–0.84 mg kg−1, soil application of selenate-only was 2–15 times more effective than granular Se-enriched macronutrient fertilizers in raising grain Se concentrations. Co-application of foliar Se with an N carrier doubled the Se concentration in wheat grains compared to the application of foliar Se on its own, the majority of which was in the highly bioavailable selenomethionine fraction. Results from this study demonstrate the possibility of improving the efficacy of Se fertilizers, which could enrich crops with Se without additional application costs in the field

The identification of sulfide oxidation as a potential metabolism driving primary production on late Noachian Mars

The transition of the martian climate from the wet Noachian era to the dry Hesperian (4.1–3.0 Gya) likely resulted in saline surface waters that were rich in sulfur species. Terrestrial analogue environments that possess a similar chemistry to these proposed waters can be used to develop an understanding of the diversity of microorganisms that could have persisted on Mars under such conditions. Here, we report on the chemistry and microbial community of the highly reducing sediment of Colour Peak springs, a sulfidic and saline spring system located within the Canadian High Arctic. DNA and cDNA 16S rRNA gene profiling demonstrated that the microbial community was dominated by sulfur oxidising bacteria, suggesting that primary production in the sediment was driven by chemolithoautotrophic sulfur oxidation. It is possible that the sulfur oxidising bacteria also supported the persistence of the additional taxa. Gibbs energy values calculated for the brines, based on the chemistry of Gale crater, suggested that the oxidation of reduced sulfur species was an energetically viable metabolism for life on early Mar

Characterisation of dust emissions from machined engineered stones to understand the hazard for accelerated silicosis

Engineered stones are novel construction materials associated with a recent upsurge in silicosis cases among workers in the stonemason industry. In order to understand the hazard for the short latency of lung disease among stonemasons, we simulated real-time dust exposure scenario by dry-machining engineered stones in controlled conditions, capturing and analysing the respirable dust generated for physical and chemical characteristics. Natural granite and marble were included for comparison. Cutting engineered stones generated high concentrations of very fine particles ( 80% respirable crystalline silica content, in the form of quartz and cristobalite. Engineered stones also contained 8–20% resin and 1–8% by weight metal elements. In comparison, natural stones had far lower respirable crystalline silica (4- 30%) and much higher metal content, 29–37%. Natural stone dust emissions also had a smaller surface area than engineered stone, as well as lower surface charge. This study highlighted the physical and chemical variability within engineered stone types as well as between engineered and natural stones. This information will ultimately help understand the unique hazard posed by engineered stone fabrication work and help guide the development of specific engineering control measures targeting lower exposure to respirable crystalline silica.Chandnee Ramkissoon, Sharyn Gaskin, Leigh Thredgold, Tony Hall, Shelley Rowett, Richard Gu

Colour Peak: An analogue environment for late Noachian Mars

The martian surface cannot sustain liquid water today, but there is evidence water was present during the Noachian era. The transition of the martian climate into the Hesperian would have resulted in saline and sulfuric waters. Terrestrial analogue environments that possess a chemistry like these proposed waters can be used to develop an understanding of organisms that could have persisted. Here we present the chemistry and microbiome of Colour Peak, a sulfidic and saline spring system located within the Canadian High Arctic. Nucleic acids were extracted from the microbes in the sediments and the microbiome was characterised by the amplification and sequencing of 16S rRNA gene amplicons. The elemental composition of the fluids and sediment was determined by ICP-OES and compared with brines determined from the chemistry of the “Rocknest” sample at Yellowknife Bay, Gale Crater (Mars) by thermochemical modelling. Gibbs energy values were calculated from this fluid chemistry to identify potentially viable metabolisms. Analysis of the chemistries of the Colour Peak fluids confirmed a composition like the thermochemically modelled fluid, providing justification for the classification of Colour Peak as an appropriate analogue environment to investigate the habitability of former martian waters. Profiling of the Colour Peak microbial community revealed domination by bacteria associated with oxidation of reduced sulfur species and carbon dioxide fixation. Gibbs energy values calculated using the modelled martian fluid chemistry demonstrated that oxidation of reduced sulfur species was also viable in this chemical environment under aerobic and anaerobic conditions. These results demonstrate microbial sulfide oxidation is thermodynamically viable using both modelled and environmental proxies for former martian aqueous environments. This study highlights that metabolisms utilising the oxidation of reduced sulfur species could have been thermodynamically viable in ancient martian aqueous environments. Further work is needed to test this viability and the subsequent potential for biosignature formation

A genetic case-control study confirms the implication of SMAD7 and TNF locus in the development of proliferative vitreoretinopathy

PURPOSE: Proliferative vitreoretinopathy (PVR) is still the major cause of failure of retinal detachment (RD) surgery and although the risk for developing this complication is associated with some clinical characteristics, the correlation is far from absolute, raising the possibility of genetic susceptibility. The objective of this study was to analyze the genetic contribution to PVR in patients undergoing RD surgery, the Retina 4 Project. METHODS: A candidate gene association study was conducted in 2006 in a Spanish population of 450 patients suffering from primary rhegmatogenous RD. Replication was carried out in a larger population undergoing RD surgery at several European centers among 546 new patients. Single nucleotide polymorphism (SNP) of 30 genes known to be involved with inflammation were analyzed. For replication stage, those genes previously detected as significantly associated with PVR were genotyped. Distribution of allelic and haplotypic frequencies in case and control group were analyzed. Single and haplotypic analysis were assessed. The Rosenberg two-stage method was used to correct for single and multiple analyses. RESULTS: After correction for multiple comparisons, four genes were significantly associated with PVR: SMAD7 (P = 0.004), PIK3CG (P = 0.009), TNF locus (P = 0.0005), and TNFR2 (P = 0.019) In the European sample, replication was observed in SMAD7 (P = 0.047) and the TNF locus (P = 0.044). CONCLUSIONS: These results confirm the genetic contribution to PVR and the implication of SMAD7 and TNF locus in the development of PVR. This finding may have implications for understanding the mechanisms of PVR and could provide a potential new therapeutic target for PVR prophylaxis