An investigation of a Mollicute-like organism inhabiting the human gastrointestinal tract

The microflora inhabiting the human gastrointestinal tract can be considered an essential 'metabolic organ', in a symbiotic relationship with its host. Due to the low cultivability and inappropriate sampling methodology the microflora is poorly explored and ill-defined. Preliminary, molecular-based research at the University of Waikato revealed the presence of 16S rRNA gene sequences originating from novel Mollicute-like species inhabiting the human GI tract. A ~830bp 'consensus' sequence representing these novel Mollicute-like sequences was classified within the Mollicute Genus Anaeroplasma the type species of which is Anaeroplasma abactoclasticum. It also displayed near exact matches with 16S rRNA sequences obtained from the human GI tract and matches of high similarity to those from the mouse GI tract in the NCBI database. This thesis describes an attempt to design and create primers that would amplify and characterize full-length versions of these Mollicute-like sequences from samples obtained from the mucosal surface of the human gastrointestinal tract. Primers sets targeted extended 5' and 3' versions of these novel 'known' sequences and were designed from sequence matches found in the preliminary work and other related sequences from the NCBI database. The attempt to amplify a full-length version of these novel Mollicute-like sequences was proven to be unsuccessful. No sequences were classified within the Genus Anaeroplasma, although 81% of amplicons from the 5' extending primer sets were classified within the same division as the Mollicutes, the Firmicutes, only 6% of the sequenced amplicons from the 3' extending primer set belonged to this division. Phylograms containing these 'relevant' sequences and the 'consensus' sequence grouped the 'consensus' sequence separately, indicating a lower relatedness than would have been seen if any of the amplicons contained the 'consensus' sequence

Residential sorting across Auckland neighbourhoods

This paper addresses the extent to which people in Auckland exhibit residential location patterns that differ between groups, i.e. the extent to which they are spatially sorted. To measure patterns of residential location, the paper uses the index of segregation, an isolation index, Gini coefficients, Ellison & Glaeser and Maurel & Sédillot concentration measures, Moran’s I and Getis and Ord’s G*. Results are presented based on a classification of the population in different ways: ethnicity, income, education, age and country of birth. Both city-wide and local measures are considered. We find that ethnic-based sorting is the strongest indicator of residential sorting patterns, but soring by income, education and age is also present. Sorting by income and qualifications is strongest at the top and, to lesser extent, at the bottom of the income and qualifications range. Age segregation is most pronounced for older residents. Clustering is strongest within a range of up to one kilometre and declines significantly over greater distances. Local analysis by means of Getis and Ord’s G* calculations suggest significant ethnic clustering. Apart from Maori and Pacific Islanders, ethnic groups tend to locate way from each other, as confirmed with cross-Moran’s I calculations. When considering interactions between ethnicity and income we find that the location of ethnicity income subgroups is more strongly related to neighbourhood income

Mortality and quality of life in the five years after severe sepsis

Peer reviewedPublisher PD

Novel biocatalytic modules for the cell-free conversion of cellodextrins to glucaric acid

Cell-free biocatalysis offers a versatile platform for the biomanufacturing of bulk or specialty chemicals due to the flexibility in assembling enzymes from different organisms in synthetic reaction pathways. Current challenges of this approach include costly enzyme preparation, low enzyme stability and efficient enzyme recycling. To overcome these challenges, we present a molecular toolbox that facilitates the simple construction of enzymes as low-cost and recyclable biocatalytic modules. The toolbox is composed of three interchangeable components: (i) inorganic matrices; (ii) matrix-specific solid-binding peptides (SBPs); and (iii) thermostable enzymes. SBPs are short amino acid sequences that can be fused genetically to proteins and direct the orientated immobilization of the resulting protein fusion onto solid matrices (1, 2). The biocatalytic module design relies on the affinity of the SBP for inorganic matrices. Single enzyme biocatalytic modules can be prepared easily consisting of one type of enzyme immobilized per matrix while a multiple enzyme biocatalytic module consists of multiple enzymes immobilized simultaneously onto the matrix. The modules can be combined rationally to generate product-specific reaction pathways and their subsequent removal from the reaction medium allows for a ‘pick, mix, and reuse’ approach, which can be optimized easily for low-cost cell-free biomanufacturing. Recently, we have shown that it is possible to assemble single and multiple enzyme biocatalytic modules using thermostable polysaccharide-degrading enzymes and that the enzymes retain their specific hydrolytic activities upon several rounds of recycling at high temperatures (2). Here, we applied the biocatalytic modules concept for the cell-free conversion of cellodextrins to glucaric acid, via a more complex seven enzyme synthetic pathway. Glucaric acid is one of the 12 top candidates for bio-based building blocks and is a precursor for polymers, including nylons and hyperbranched polyesters (3). Its bioproduction from cellodextrins, which can be derived from organic waste, provides a sustainable alternative to the fossil-derived production of polymers. Initially, single enzyme biocatalytic modules were prepared with a silica-specific SBP fused to two enzymes of the synthetic pathway allowing for their selective immobilization onto an inexpensive silica-based matrix. The SBP mediated the binding of each enzyme onto the matrix with over 85% immobilization efficiency. When comparing the enzyme activities of the biocatalytic modules against the free enzymes, 85 and 93% of their initial activities were retained upon immobilization, respectively. Furthermore, co-immobilization of these two enzymes as a multiple enzyme module resulted in similar immobilization yields. Performance of both enzymes in the multiple enzyme module in a successive reaction revealed that they retained 70% of their activity when compared to the free enzymes. Currently, the silica-specific SBP has been incorporated into other 5 enzymes of the pathway and we are proceeding with the construction of the single and multiple enzyme biocatalytic modules and pathway assembly. (1) Care A, Bergquist PL, Sunna A (2015) Trends in Biotechnology, 33: 259-268. (2) Care A, Petroll K, Gibson ESY, Bergquist PL, Sunna A (2017) Biotechnology for Biofuels 10:29. (3) Werpy T and G Petersen (2004). Results of Screening for Potential Candidates from Sugars and Synthesis Gas. National Renewable Energy Lab

Synthetic biocatalytic modules for enhanced transformation of biological waste products

Many insoluble materials can be used as carriers for the immobilisation of enzymes. Solid-binding peptides (SBPs) are short amino acid sequences that can act as molecular linkers to direct the orientated immobilisation of proteins onto solid materials without impeding their biological activity [1]. Silica-based materials like silica and zeolite have been found to be suitable matrices for enzyme immobilisation in industrial processes. They are inexpensive, offer high mechanical strength and stability, are chemically inert and can be deployed over a wide range of operating conditions. We have constructed biocatalytic modules that are based on the incorporation of a silica-binding SBP (‘linker’) sequence into several genes for thermostable enzymes to facilitate the immobilisation of the proteins onto silica-based matrices, enabling the hydrolysis of both simple and complex polysaccharides. We have shown also that the procedure is suitable for the construction of complex enzymological pathways. In proof of concept experiments, the linker (L) sequence was attached to the N- or C-terminus of three thermostable hemicellulases isolated from thermophilic bacteria using genetic engineering techniques [2]. The resulting L-enzymes remained active after fusion and displayed the same pH and temperature optima but differing thermostabilities in comparison to their corresponding enzymes without linker. The linker facilitated the rapid and simple immobilisation of each L-enzyme onto zeolite, resulting in the construction of ‘single enzyme biocatalytic modules’. All three L-enzymes co-immobilised onto the same zeolite matrix resulted in the formation of ‘multiple enzyme biocatalytic modules’, which were shown to degrade various hemicellulosic substrates effectively in a ‘one-pot’ reaction. Cell-free synthetic biology circumvents many of the limitations encountered by in vivo synthetic biology by operating without the constraints of a cell. It offers higher substrate and enzyme loading and the facile optimisation of enzyme ratios. Some of the challenges of this approach include costly enzyme preparation, biocatalyst stability, and the need for constant supplementation with co-factors. To overcome these challenges, we have developed a molecular toolbox that facilitates the construction of biocatalytic modules with predefined functions and catalytic properties. It consists of three interchangeable building blocks: (a) low-cost inorganic matrices (e.g., silica, zeolite), (b) matrix-specific SBPs and (c) thermostable enzymes. The rational combination of these building blocks allows for flexibility and a ‘pick, mix’ and re-use’ approach with multiple biocatalytic modules available for the assembly of natural and non-natural pathways. Individual immobilised enzymes can be combined rationally to assemble recyclable and product-specific reactions. We present preliminary results relating to the construction of two synthetic pathways for the conversion of organic wastes such as coffee and plant biomass. The pathway assembly process allows for rapid evaluation for proof of concept and for assessing the parameters for a synthetic pathway, which are very labour- and time-intensive by the in vivo approach. [1] Care, A, Bergquist, PL, Sunna, A. (2015) Trends Biotech. 33: 259-268 [2] Care, A, Petroll, K, Gibson, ESY, Bergquist, PL, Sunna, A. (2017) Biotech. Biofuels. 10: 2

Association of Blood Biomarkers With Acute Sport-Related Concussion in Collegiate Athletes: Findings From the NCAA and Department of Defense CARE Consortium

Importance: There is potential scientific and clinical value in validation of objective biomarkers for sport-related concussion (SRC). Objective: To investigate the association of acute-phase blood biomarker levels with SRC in collegiate athletes. Design, Setting, and Participants: This multicenter, prospective, case-control study was conducted by the National Collegiate Athletic Association (NCAA) and the US Department of Defense Concussion Assessment, Research, and Education (CARE) Consortium from February 20, 2015, to May 31, 2018, at 6 CARE Advanced Research Core sites. A total of 504 collegiate athletes with concussion, contact sport control athletes, and non-contact sport control athletes completed clinical testing and blood collection at preseason baseline, the acute postinjury period, 24 to 48 hours after injury, the point of reporting being asymptomatic, and 7 days after return to play. Data analysis was conducted from March 1 to November 30, 2019. Main Outcomes and Measures: Glial fibrillary acidic protein (GFAP), ubiquitin C-terminal hydrolase-L1 (UCH-L1), neurofilament light chain, and tau were quantified using the Quanterix Simoa multiplex assay. Clinical outcome measures included the Sport Concussion Assessment Tool-Third Edition (SCAT-3) symptom evaluation, Standardized Assessment of Concussion, Balance Error Scoring System, and Brief Symptom Inventory 18. Results: A total of 264 athletes with concussion (mean [SD] age, 19.08 [1.24] years; 211 [79.9%] male), 138 contact sport controls (mean [SD] age, 19.03 [1.27] years; 107 [77.5%] male), and 102 non-contact sport controls (mean [SD] age, 19.39 [1.25] years; 82 [80.4%] male) were included in the study. Athletes with concussion had significant elevation in GFAP (mean difference, 0.430 pg/mL; 95% CI, 0.339-0.521 pg/mL; P < .001), UCH-L1 (mean difference, 0.449 pg/mL; 95% CI, 0.167-0.732 pg/mL; P < .001), and tau levels (mean difference, 0.221 pg/mL; 95% CI, 0.046-0.396 pg/mL; P = .004) at the acute postinjury time point compared with preseason baseline. Longitudinally, a significant interaction (group × visit) was found for GFAP (F7,1507.36 = 16.18, P < .001), UCH-L1 (F7,1153.09 = 5.71, P < .001), and tau (F7,1480.55 = 6.81, P < .001); the interaction for neurofilament light chain was not significant (F7,1506.90 = 1.33, P = .23). The area under the curve for the combination of GFAP and UCH-L1 in differentiating athletes with concussion from contact sport controls at the acute postinjury period was 0.71 (95% CI, 0.64-0.78; P < .001); the acute postinjury area under the curve for all 4 biomarkers combined was 0.72 (95% CI, 0.65-0.79; P < .001). Beyond SCAT-3 symptom score, GFAP at the acute postinjury time point was associated with the classification of athletes with concussion from contact controls (β = 12.298; 95% CI, 2.776-54.481; P = .001) and non-contact sport controls (β = 5.438; 95% CI, 1.676-17.645; P = .005). Athletes with concussion with loss of consciousness or posttraumatic amnesia had significantly higher levels of GFAP than athletes with concussion with neither loss of consciousness nor posttraumatic amnesia at the acute postinjury time point (mean difference, 0.583 pg/mL; 95% CI, 0.369-0.797 pg/mL; P < .001). Conclusions and Relevance: The results suggest that blood biomarkers can be used as research tools to inform the underlying pathophysiological mechanism of concussion and provide additional support for future studies to optimize and validate biomarkers for potential clinical use in SRC

Physical restraint in residential child care : the experiences of young people and residential workers

There have long been concerns about the use of physical restraint in residential care. This paper presents the findings of a qualitative study which explores the experiences of children, young people and residential workers about physical restraint. The research identifies the dilemmas and ambiguities for both staff and young people, and participants discuss the situations where they feel physical restraint is appropriate as well as their concerns about unjustified or painful restraints. They describe the negative emotions involved in restraint but also those situations where, through positive relationships and trust, restraint can help young people through unsafe situations

Projections of multi-morbidity in the older population in England to 2035: estimates from the Population Ageing and Care Simulation (PACSim) model

Background models projecting future disease burden have focussed on one or two diseases. Little is known on how risk factors of younger cohorts will play out in the future burden of multi-morbidity (two or more concurrent long-term conditions). Design a dynamic microsimulation model, the Population Ageing and Care Simulation (PACSim) model, simulates the characteristics (sociodemographic factors, health behaviours, chronic diseases and geriatric conditions) of individuals over the period 2014–2040. Population about 303,589 individuals aged 35 years and over (a 1% random sample of the 2014 England population) created from Understanding Society, the English Longitudinal Study of Ageing, and the Cognitive Function and Ageing Study II. Main outcome measures the prevalence of, numbers with, and years lived with, chronic diseases, geriatric conditions and multi-morbidity. Results between 2015 and 2035, multi-morbidity prevalence is estimated to increase, the proportion with 4+ diseases almost doubling (2015:9.8%; 2035:17.0%) and two-thirds of those with 4+ diseases will have mental ill-health (dementia, depression, cognitive impairment no dementia). Multi-morbidity prevalence in incoming cohorts aged 65–74 years will rise (2015:45.7%; 2035:52.8%). Life expectancy gains (men 3.6 years, women: 2.9 years) will be spent mostly with 4+ diseases (men: 2.4 years, 65.9%; women: 2.5 years, 85.2%), resulting from increased prevalence of rather than longer survival with multi-morbidity. Conclusions our findings indicate that over the next 20 years there will be an expansion of morbidity, particularly complex multi-morbidity (4+ diseases). We advocate for a new focus on prevention of, and appropriate and efficient service provision for those with, complex multi-morbidity

Effect of QUiPP prediction algorithm on treatment decisions in women with a previous preterm birth: a prospective cohort study.

OBJECTIVE:The QUiPP algorithm combines cervical length, quantitative fetal fibronectin (qfFN) and medical history to quantify risk of preterm birth. We assessed the utility of QUiPP to inform preterm birth prevention treatment decisions. DESIGN:A prospective cohort study with a subsequent impact assessment using the QUiPP risk of birth before 34 weeks gestation. SETTING:A UK TERTIARY REFERRAL HOSPITAL: SAMPLE: 119 women with previous spontaneous preterm birth (sPTB) or preterm premature rupture of membranes (PPROM) before 34 weeks gestation. METHODS:Cervical length and qfFN were measured at 19+0 - 23+0 weeks gestation. Clinical management was based on history and cervical length. After birth, clinicians were unblinded to qfFN results and QUiPP analysis was undertaken. MAIN OUTCOME MEASURES:Predictive statistics of QUiPP algorithm using 10% risk of sPTB before 34+0 weeks as treatment threshold. RESULTS:Fifteen of 119 women (13%) had PPROM or sPTB before 34 weeks. Of these 53% (8/15) had QUiPP risk of sPTB before 34+0 weeks above 10%. Applying this treatment threshold in practice would have doubled our treatment rate (20% vs 42%). QUIPP threshold of 10% had positive likelihood ratio (LR) of 1.3 (95% CI 0.76-2.18), and negative LR of 0.8 (95% CI 0.45-1.40) for predicting sPTB before 34+0 weeks. CONCLUSIONS:Use of the QUiPP algorithm in this population may lead to substantial increase in interventions without evidence that currently available treatment options are beneficial for this particular group. This article is protected by copyright. All rights reserved