Co-production of 11α-hydroxyprogesterone and ethanol using recombinant yeast expressing fungal steroid hydroxylases

Background Bioethanol production from sustainable sources of biomass that limit effect on food production are needed and in a biorefinery approach co-products are desirable, obtained from both the plant material and from the microbial biomass. Fungal biotransformation of steroids was among the first industrial biotransformations allowing corticosteroid production. In this work, the potential of yeast to produce intermediates needed in corticosteroid production is demonstrated at laboratory scale following bioethanol production from perennial ryegrass juice. Results Genes encoding the 11?-steroid hydroxylase enzymes from Aspergillus ochraceus (11?-SHAoch) and Rhizopus oryzae (CYP509C12) transformed into Saccharomyces cerevisiae for heterologous constitutive expression in p425TEF. Both recombinant yeasts (AH22:p11?-SHAoch and AH22:p509C12) exhibited efficient progesterone bioconversion (on glucose minimal medial containing 300 ?M progesterone) producing either 11?-hydroxyprogesterone as the sole metabolite (AH22:p11?-SHAoch) or a 7:1 mixture of 11?-hydroxyprogesterone and 6?-hydroxyprogesterone (AH22:p509C12). Ethanol yields for AH22:p11?-SHAoch and AH22:p509C12 were comparable resulting in ?75% conversion of glucose to alcohol. Co-production of bioethanol together with efficient production of the 11-OH intermediate for corticosteroid manufacture was then demonstrated using perennial ryegrass juice. Integration of the 11?-SHAoch gene into the yeast genome (AH22:11?-SHAoch+K) resulted in a 36% reduction in yield of 11?-hydroxyprogesterone to 174 ?mol/L using 300 ?M progesterone. However, increasing progesterone concentration to 955 ?M and optimizing growth conditions increased 11?-hydroxyprogesterone production to 592 ?mol/L product formed. Conclusions The progesterone 11?-steroid hydroxylases from A. ochraceus and R. oryzae, both monooxygenase enzymes of the cytochrome P450 superfamily, have been functionally expressed in S. cerevisiae. It appears that these activities in fungi are not associated with a conserved family of cytochromes P450. The activity of the A. ochraceous enzyme was important as the specificity of the biotransformation yielded just the 11-OH product needed for corticosteroid production. The data presented demonstrate how recombinant yeast could find application in rural biorefinery processes where co-production of value-added products (11?-hydroxyprogesterone and ethanol) from novel feedstocks is an emergent and attractive possibility.publishersversionPeer reviewe

The Investigational Drug VT-1129 Is a Highly Potent Inhibitor of Cryptococcus Species CYP51 but Only Weakly Inhibits the Human Enzyme

Cryptococcosis is a life-threatening disease often associated with HIV infection. Three Cryptococcus species CYP51 enzymes were purified and catalyzed the 14α-demethylation of lanosterol, eburicol, and obtusifoliol. The investigational agent VT-1129 bound tightly to all three CYP51 proteins (dissociation constant [K(d)] range, 14 to 25 nM) with affinities similar to those of fluconazole, voriconazole, itraconazole, clotrimazole, and ketoconazole (K(d) range, 4 to 52 nM), whereas VT-1129 bound weakly to human CYP51 (K(d), 4.53 μM). VT-1129 was as effective as conventional triazole antifungal drugs at inhibiting cryptococcal CYP51 activity (50% inhibitory concentration [IC(50)] range, 0.14 to 0.20 μM), while it only weakly inhibited human CYP51 activity (IC(50), ∼600 μM). Furthermore, VT-1129 weakly inhibited human CYP2C9, CYP2C19, and CYP3A4, suggesting a low drug-drug interaction potential. Finally, the cellular mode of action for VT-1129 was confirmed to be CYP51 inhibition, resulting in the depletion of ergosterol and ergosta-7-enol and the accumulation of eburicol, obtusifolione, and lanosterol/obtusifoliol in the cell membranes

Older onset essential tremor : more rapid progression and more degenerative pathology

There are few data on rate of progression in essential tremor (ET). To quantify the rate of tremor progression in a cross-sectional sample of 348 ET cases in an epidemiological study; characterize the relationship between age of tremor onset and rate of tremor progression in that sample; and characterize the relationship between age of tremor onset, rate of tremor progression, and severity of underlying brain changes in 9 cases from a brain repository. Rate of tremor progression was defined as tremor severity divided by duration. The degeneration index = number of torpedoes per section divided by Purkinje cell linear density. In the epidemiological study, older age of tremor onset was associated with faster rate of tremor progression (P < 0.001). In the brain repository, older age of tremor onset was associated with higher degeneration index (P = 0.037), and higher degeneration index was associated with faster rate of tremor progression (P = 0.018). In a large clinical sample, older age of onset was associated with more rapid tremor progression. In a brain bank, older age of onset was associated with more degenerative pathology in the cerebellum. As in several neurodegenerative disorders, in older onset cases, it is possible that the disease advances more rapidly

Diagnostic accuracy of 1p/19q codeletion tests in oligodendroglioma:a comprehensive meta-analysis based on a Cochrane Systematic Review

Codeletion of chromosomal arms 1p and 19q, in conjunction with a mutation in the isocitrate dehydrogenase 1 or 2 gene, is the molecular diagnostic criterion for oligodendroglioma, IDH mutant and 1p/19q codeleted. 1p/19q codeletion is a diagnostic marker and allows prognostication and prediction of the best drug response within IDH‐mutant tumours. We performed a Cochrane review and simple economic analysis to establish the most sensitive, specific and cost‐effective techniques for determining 1p/19q codeletion status. Fluorescent in situ hybridisation (FISH) and polymerase chain reaction (PCR)‐based loss of heterozygosity (LOH) test methods were considered as reference standard. Most techniques (FISH, chromogenic in situ hybridisation [CISH], PCR, real‐time PCR, multiplex ligation‐dependent probe amplification [MLPA], single nucleotide polymorphism [SNP] array, comparative genomic hybridisation [CGH], array CGH, next‐generation sequencing [NGS], mass spectrometry and NanoString) showed good sensitivity (few false negatives) for detection of 1p/19q codeletions in glioma, irrespective of whether FISH or PCR‐based LOH was used as the reference standard. Both NGS and SNP array had a high specificity (fewer false positives) for 1p/19q codeletion when considered against FISH as the reference standard. Our findings suggest that G banding is not a suitable test for 1p/19q analysis. Within these limits, considering cost per diagnosis and using FISH as a reference, MLPA was marginally more cost‐effective than other tests, although these economic analyses were limited by the range of available parameters, time horizon and data from multiple healthcare organisations

Development of mental health first aid guidelines for suicidal ideation and behaviour: A Delphi study

<p>Abstract</p> <p>Background</p> <p>Suicide is a statistically rare event, but devastating to those left behind and one of the worst possible outcomes associated with mental illness. Although a friend, family member or co-worker may be the first person to notice that a person is highly distressed, few have the knowledge and skills required to assist. Simple guidelines may help such a person to encourage a suicidal individual to seek professional help or decide against suicide.</p> <p>Methods</p> <p>This research was conducted using the Delphi methodology, a method of reaching consensus in a panel of experts. Experts recruited to the panels included 22 professionals, 10 people who had been suicidal in the past and 6 carers of people who had been suicidal in the past. Statements about how to assist someone who is thinking about suicide were sourced through a systematic search of both professional and lay literature. The guidelines were written using the items most consistently endorsed by all three panels.</p> <p>Results</p> <p>Of 114 statements presented to the panels, 30 were accepted. These statements were used to develop the guidelines appended to this paper.</p> <p>Conclusion</p> <p>There are a number of actions which are considered to be useful for members of the public when they encounter someone who is experiencing suicidal thoughts or engaging in suicidal behaviour. These guidelines will be useful in revision of curricula of mental health first aid and suicide intervention training programs. They can also be used by members of the public who want immediate information about how to assist a suicidal person.</p

Emergent dynamic chirality in a thermally driven artificial spin ratchet

Modern nanofabrication techniques have opened the possibility to create novel functional materials, whose properties transcend those of their constituent elements. In particular, tuning the magnetostatic interactions in geometrically frustrated arrangements of nanoelements called artificial spin ice1, 2 can lead to specific collective behaviour3, including emergent magnetic monopoles4, 5, charge screening6, 7 and transport8, 9, as well as magnonic response10, 11, 12. Here, we demonstrate a spin-ice-based active material in which energy is converted into unidirectional dynamics. Using X-ray photoemission electron microscopy we show that the collective rotation of the average magnetization proceeds in a unique sense during thermal relaxation. Our simulations demonstrate that this emergent chiral behaviour is driven by the topology of the magnetostatic field at the edges of the nanomagnet array, resulting in an asymmetric energy landscape. In addition, a bias field can be used to modify the sense of rotation of the average magnetization. This opens the possibility of implementing a magnetic Brownian ratchet13, 14, which may find applications in novel nanoscale devices, such as magnetic nanomotors, actuators, sensors or memory cells

Predictors of COVID-19 vaccine hesitancy in the UK household longitudinal study

Vaccine hesitancy could undermine efforts to control COVID-19. We investigated the prevalence of COVID-19 vaccine hesitancy in the UK and identified vaccine hesitant subgroups. The ‘Understanding Society’ COVID-19 survey asked participants (n = 12,035) their likelihood of vaccine uptake and reason for hesitancy. Cross-sectional analysis assessed vaccine hesitancy prevalence and logistic regression calculated odds ratios. Overall vaccine hesitancy was low (18% unlikely/very unlikely). Vaccine hesitancy was higher in women (21.0% vs 14.7%), younger age groups (26.5% in 16–24 year olds vs 4.5% in 75 + ) and those with lower education levels (18.6% no qualifications vs 13.2% degree qualified). Vaccine hesitancy was high in Black (71.8%) and Pakistani/Bangladeshi (42.3%) ethnic groups. Odds ratios for vaccine hesitancy were 13.42 (95% CI:6.86, 26.24) in Black and 2.54 (95% CI:1.19, 5.44) in Pakistani/Bangladeshi groups (compared to White British/Irish) and 3.54 (95% CI:2.06, 6.09) for people with no qualifications versus degree. Urgent action to address hesitancy is needed for some but not all ethnic minority groups

Downgraded curriculum? An analysis of knowledge in new curricula in Scotland and New Zealand

The development, since 2000, of new National Curricula across the Anglophone world signals a number of policy trends, including: a move from the explicit specification of content towards a more generic, skills-based approach; a greater emphasis on the centrality of the learner; and [ostensibly] greater autonomy for teachers in developing the curriculum in school. These policy shifts have attracted some criticism, especially from social realist writers, who claim that the new curricula downgrade knowledge. This paper offers a contribution to this debate; an empirically-based analysis of two new curricula, New Zealand&rsquo;s Curriculum Framework and Scotland&rsquo;s Curriculum for Excellence. We conclude that, while these curricula continue to accord considerable importance to knowledge in their statements of policy intent, the social realist critique is at least partially justified, since both curricula are characterised by a lack of coherence and mixed messages about the place of knowledge

Evaluation of a brief anti-stigma campaign in Cambridge: do short-term campaigns work?

<p>Abstract</p> <p>Background</p> <p>In view of the high costs of mass-media campaigns, it is important to understand whether it is possible for a media campaign to have significant population effects over a short period of time. This paper explores this question specifically in reference to stigma and discrimination against people with mental health problems using the <it>Time to Change </it>Cambridge anti-stigma campaign as an example.</p> <p>Methods</p> <p>410 face-to-face interviews were performed pre, during and post campaign activity to assess campaign awareness and mental health-related knowledge, attitudes and behaviours.</p> <p>Results</p> <p>Although campaign awareness was not sustained following campaign activity, significant and sustained shifts occurred for mental health-related knowledge items. Specifically, there was a 24% (p < 0.001) increase in persons agreeing with the statement: <it>If a friend had a mental health problem, I know what advice to give them to get professional help</it>, following the campaign. Additionally, for the statement: <it>Medication can be an effective treatment for people with mental health problems</it>, there was a 10% rise (p = 0.05) in the proportion of interviewees responding 'agree' or 'strongly agree' following the campaign. These changes, however, were not evident for attitudinal or behaviour related questions.</p> <p>Conclusions</p> <p>Although these results only reflect the impact of one small scale campaign, these preliminary findings suggest several considerations for mass-media campaign development and evaluation strategies such as: (1) Aiming to influence outcomes pertaining to knowledge in the short term; (2) Planning realistic and targeted outcomes over the short, medium and long term during sustained campaigns; and (3) Monitoring indirect campaign effects such as social discourse or other social networking/contact in the evaluation.</p