Development of physio-chemical pretreatments and mixed microbial cultures for the conversion of lignocellulosic biomass to useful products

There is increasing interest in producing biofuels; biofuels are preferable to fossil fuels as the biomass from which they are derived is seen as a renewable source, as opposed to fossil fuels which are a finite resource. “First Generation” biofuels are derived from food crops such as grains and sugar cane. The use of food crops is not sustainable in this age of increasing food insecurity. A promising alternative appears to be what is termed “Second Generation” feedstocks, such as energy crops like Miscanthus spp., and agricultural by-products. The problem with the use of second generation feedstocks is firstly that the sugars are locked up in the cell wall polymers (CWP), which need to be released by physio-chemical pre-treatments, that are costly and time consuming. The second problem is that not all the sugars that are released from CWP are able to be utilised by wild type product-forming organisms. However, model chassis organisms can be genetically modified to utilise these sugars and /or produce enzymes to degrade biomass which reduces the time and costs involved in the process. While engineering these organisms to utilise a range of monosaccharides has already been successful, engineering them to produce degradation enzymes is proving to be problematic. A potentially more effective system is to use co-cultures of both cellulose-degrading and product-forming organisms. Since this is a novel approach it is not known whether the two organisms are able to live together without any adverse effects. The aims of this study were firstly to determine whether mixed cultures of both cellulose-degrading and potential product-forming organisms could survive in the presence of one another, secondly whether the cellulose-degrading organisms could degrade potential feedstock down into their monosaccharide building blocks and thirdly whether the potential product-forming organisms could survive and utilise these monosaccharides for growth and potential fermentation. It was discovered that C. hutchinsonii can degrade both paper and Triticum aestivum straw polymers into their monosaccharide components and that B. subtilis can survive on the sugars released by C. hutchinsonii. It was also discovered that C. hutchinsonii and B. subtilis 168 can only tolerate an ethanol concentration of up to 2% (v/v) and that this is below the baseline for a biofuel system to be economically viable. Likewise, C. hutchinsonii and B. subtilis 168 have an even poorer tolerance for butanol; growth is inhibited by < 1% butanol in its growth media. A series of physio-chemical pre-treatments were developed in order to make the monosaccharides present in the cell wall polymers more accessible to microbial saccharification. Sequential pre-treatments, both physical milling and chemical hydrolysis in tandem, had the greatest effect on the bio chemistry of the biomass, but that these physio-chemical pre-treatments produced inhibitory compounds in the medium that retarded microbial growth. Attempts were made to genetically modified Bacillus subtilis 168 to produce lactic acid and ethanol by over expressing the native ldh gene under the highly-expressed promoter of the cspD gene and by integrating the fused pdc:adh gene from Z. mobilis under the same promoter. Transformation of B. subtilis to over express LDH was successful, with PCR confirmation of the correct insertion and enzyme activity for the ldh both in vitro and in vivo, with the latter producing more lactic acid aerobically than the wild type. Transformation of B. subtilis to express pdc:adh and subsequent production of ethanol was not successful

Hospital admission patterns in children with CAH: admission rates and adrenal crises decline with age

Objective: To examine patterns of hospitalisation for acute medical conditions in children with congenital adrenal hyperplasia (CAH). Design: A retrospective study of hospitalisation using administrative data. Setting. All hospitals in NSW, Australia. Patients: All patients admitted with CAH and a random sample of admissions in patients aged 0 to 18 years without adrenal insufficiency (AI). Main Outcome Measures: Admissions and comorbidities by age and sex. Results: Of 573 admissions for medical problems in CAH children, 286 (49.9%) were in males, and 236 (41.2%) had a principal diagnosis of CAH or had an adrenal crisis (AC). 37 (6.5%) ACs were recorded. An infection was found in 43.5% ( = 249) of the CAH patient admissions and 51.7% ( = 1613) of the non-AI group, \u3c 0.001. Children aged up to one year had the highest number of admissions ( = 149) and six ACs (four in males).There were 21 ACs recorded for children aged 1–5 years. Older CAH children had fewer admissions and fewer ACs. No in-hospital deaths were recorded. Conclusions. Admission for medical problems in CAH children declines with age. An AC was recorded in 6.5% of the admissions, with the majority of ACs occurring in the 1 to 5 years age group and there were no deaths

Development of the Screening Tool for Everyday Mobility and Symptoms (STEMS) for skeletal dysplasia

Background: Skeletal dysplasia are genetic disorders of cartilage and bone, characterized by impairments commonly resulting in short stature, altered movement biomechanics, pain, fatigue and reduced functional performance. While current tools quantify functional mobility performance, they have not been standardly used in this population group and do not capture patient-reported symptoms such as pain or fatigue. This study evaluated a new tool, the Screening Tool for Everyday Mobility and Symptoms (STEMS), designed to accurately and objectively assess functional mobility and associated symptomology for individuals with skeletal dysplasia. Methods: Individuals aged 5-75 years with a skeletal dysplasia completed the STEMS, the Functional Mobility Scale (FMS) and Six Minute Walk Test (6MWT). The correlation among the STEMS, use of mobility aides, FMS and 6MWT normalised for leg length was calculated. One-way analysis of variance compared the STEMS symptomatology to normalised 6MWT distance. Results: One hundred and fifty individuals with skeletal dysplasia (76 achondroplasia, 42 osteogenesis imperfecta, 32 other; 74= 18 years) participated. Almost two thirds of the group reported pain and/or fatigue when mobilising at home, at work or school and within the community, but only twenty percent recorded use of a mobility device. The STEMS setting category demonstrated highly significant correlations with the corresponding FMS category (r=-0.983 to -0.0994, all p<0.001), and a low significant correlation with the normalised 6MWT distance (r=-0.323 to -0.394, all p<0.001). A decreased normalised 6MWT distance was recorded for individuals who reported symptoms of pain and/or fatigue when mobilising at home or at work/school (all p <= 0.004). Those who reported pain only when mobilising in the community had a normal 6MWT distance (p=0.43-0.46). Conclusions: The Screening Tool for Everyday Mobility and Symptoms (STEMS) is a useful new tool to identify and record mobility aide use and associated self-reported symptoms across three environmental settings for adults and children with skeletal dysplasia. The STEMS may assist clinicians to monitor individuals for changes in functional mobility and symptoms over time, identify individuals who are functioning poorly compared to peers and need further assessment, and to measure effectiveness of treatment interventions in both clinical and research settings

P450 oxidoreductase deficiency: A systematic review and meta-analysis of genotypes, phenotypes and their relationships

Context: P450 oxidoreductase deficiency (PORD) is a rare genetic disorder that is associated with significant morbidity. However there has been limited analysis of reported PORD cases. Objective: To determine, based on the cohort of reported PORD cases, genotype-phenotype relationships for skeletal malformations, maternal virilisation in pregnancy, adrenal insufficiency and disorders of sexual development (DSD). Data Sources: PubMed and Web of Science from January 2004 to February 2018. Study Selection: Published case reports/series of patients with PORD. Eligible patients were unique, had biallelic mutations and their clinical features reported. Data Extraction: Patient data were manually extracted from the text of case reports/series. A malformation score, representing the severity of skeletal malformations, was calculated for each patient. Data Synthesis: Of the 211 patients published in the literature, 90 patients were eligible for inclusion. Over 60 unique mutations were identified in this cohort. Four groups of mutations were identified, through regression modelling, as having significantly different skeletal malformation scores. Maternal virilisation in pregnancy, reported for 21% of patients, was most common for R457H mutations. Adrenal insufficiency occurred for the majority of patients (78%) and was typically mild, with homozygous R457H mutations being the least deficient. DSD affected most patients (72%) but were less common for males (46XY) with homozygous R457H mutations. Conclusions: PORD is a complex disorder with many possible mutations affecting a large number of enzymes. By analysing the cohort of reported PORD cases, this study identified clear relationships between genotype and several important phenotypic features

Selecting Grassland Species for Saline Environments

In Australia, around 5.7 million hectares of agricultural land are currently affected by dryland salinity or at risk from shallow water tables and this figure is expected to increase over the next 50 years (LWRA, 2001). Most improved grassland species cannot tolerate the combined effects of salt and waterlogging and, therefore, the productivity of sown grasslands in salt-affected areas is low. However, there is potential to overcome the lack of suitably adapted fodder species by introducing new, salt and waterlogging-tolerant species and by diversifying the gene pool of proven species. Potential species include exotic, naturalised and native Australian grass, legumes, herb and shrub species that are halophytes and non-halophytes. A collaborative national project in southern Australia commenced in 2004 with the objective of evaluating a range of forage species for saline environments

OR13-2 Burosumab Resulted in Greater Improvement in Rickets Than Conventional Therapy in Children with X-Linked Hypophosphatemia (XLH)

XLH is characterized by excess FGF23, hypophosphatemia, skeletal deformities, and growth impairment. For the last 40 years, XLH has been treated with multiple daily doses of oral phosphate and active vitamin D (Pi/D). Burosumab, a fully human monoclonal antibody to FGF23, has been approved by the FDA for the treatment of XLH in patients ≥1 year-old. In this Phase 3 trial (NCT02915705), 61 children with XLH (1-12 years old) were randomized (1:1) to receive subcutaneous burosumab starting at 0.8 mg/kg every 2 weeks or continue Pi/D titrated and individualized for each subject by investigators. Eligibility criteria included a Total Rickets Severity Score (RSS) ≥2.0 despite prior treatment with Pi/D (>7-day washout before baseline). The primary endpoint was healing of rickets at Week 40 assessed by radiologists blinded to treatment using the Radiographic Global Impression of Change (RGI-C). The mean ± SE daily oral phosphate dose from baseline to Week 40 was 37.8 ± 3.2 mg/kg, with >99% compliance reported based on days of dosing. Compared with Pi/D, 40 weeks of burosumab resulted in a greater LS mean ± SE increase in serum phosphorus (0.92 ± 0.08 vs 0.20 ± 0.06 mg/dL), TmP/GFR (1.19 ± 0.11 vs -0.16 ± 0.05 mg/dL), and 1,25(OH)2D (30 ± 4 vs 19 ± 4 pg/mL). At Week 40, rickets improved in both groups; RGI-C global score was significantly higher in burosumab subjects than in Pi/D subjects (LS mean ± SE: +1.9 ± 0.1 vs +0.8 ± 0.1; p<0.0001). More burosumab subjects had substantial healing (RGI-C ≥+2.0), compared with Pi/D subjects (21/29, 72% vs 2/32, 6%; odds ratio of 39.1, p<0.0001). Improvement in the RGI-C lower limb deformity score was greater with burosumab than with Pi/D (+0.62 ± 0.12 vs +0.21 ± 0.12; p=0.02). Alkaline phosphatase decreased more with burosumab compared with Pi/D (-131 ± 13 vs 35 ± 19; p<0.0001). Consistent with decreases in rickets severity, burosumab improved growth and mobility. Standing height Z-score increased by a LS mean change (95% CI) of +0.15 (0.05, 0.25) for burosumab and +0.08 (-0.02, 0.19) for Pi/D. The 6 Minute Walk Test percent predicted distance increased with burosumab (Baseline to Week 40: 62% to 72%) and was unchanged with Pi/D (76% to 75%). Nephrocalcinosis score (range 0-4) shifted 0 in 20 Pi/D and 24 burosumab subjects; +1 in 3 Pi/D and 0 burosumab subjects; and -1 in 3 Pi/D and 2 burosumab subjects. Pre-defined adverse events (AEs) of interest, including hypersensitivity and injection site reactions, were higher in the burosumab group and were mild to moderate in severity overall. There were 4 serious AEs (3 burosumab, 1 Pi/D); none were treatment-related and all resolved. No subject discontinued study drug in either group. Data after 64 weeks of treatment will be available at the time of presentation. In this randomized Phase 3 trial, burosumab resulted in increases in growth and mobility, and significantly greater improvements in rickets than Pi/D in 1-12 year-old children with XLH

The differential diagnosis of children with joint hypermobility: a review of the literature

<p>Abstract</p> <p>Background</p> <p>In this study we aimed to identify and review publications relating to the diagnosis of joint hypermobility and instability and develop an evidence based approach to the diagnosis of children presenting with joint hypermobility and related symptoms.</p> <p>Methods</p> <p>We searched Medline for papers with an emphasis on the diagnosis of joint hypermobility, including Heritable Disorders of Connective Tissue (HDCT).</p> <p>Results</p> <p>3330 papers were identified: 1534 pertained to instability of a particular joint; 1666 related to the diagnosis of Ehlers Danlos syndromes and 330 related to joint hypermobility.</p> <p>There are inconsistencies in the literature on joint hypermobility and how it relates to and overlaps with milder forms of HDCT. There is no reliable method of differentiating between Joint Hypermobility Syndrome, familial articular hypermobility and Ehlers-Danlos syndrome (hypermobile type), suggesting these three disorders may be different manifestations of the same spectrum of disorders. We describe our approach to children presenting with joint hypermobility and the published evidence and expert opinion on which this is based.</p> <p>Conclusion</p> <p>There is value in identifying both the underlying genetic cause of joint hypermobility in an individual child and those hypermobile children who have symptoms such as pain and fatigue and might benefit from multidisciplinary rehabilitation management.</p> <p>Every effort should be made to diagnose the underlying disorder responsible for joint hypermobility which may only become apparent over time. We recommend that the term "Joint Hypermobility Syndrome" is used for children with symptomatic joint hypermobility resulting from any underlying HDCT and that these children are best described using <b>both </b>the term Joint Hypermobility Syndrome <b>and </b>their HDCT diagnosis.</p