Adaptor protein-2 sigma subunit mutations causing familial hypocalciuric hypercalcaemia type 3 (FHH3) demonstrate genotype-phenotype correlations, codon bias and dominant-negative effects

The adaptor protein-2 sigma subunit (AP2sigma;2) is pivotal for clathrin-mediated endocytosis of plasma membrane constituents such as the calcium-sensing receptor (CaSR). Mutations of the AP2sigma;2 Arg15 residue result in familial hypocalciuric hypercalcaemia type 3 (FHH3), a disorder of extracellular calcium (Ca<inf>o</inf>²⁺) homeostasis. To elucidate the role of AP2sigma;2 in Ca<inf>o</inf>²⁺ regulation, we investigated 65 FHH probands, without other FHH-associated mutations, for AP2sigma;2 mutations, characterized their functional consequences and investigated the genetic mechanisms leading to FHH3. AP2sigma;2 mutations were identified in 17 probands, comprising 5 Arg15Cys, 4 Arg15His and 8 Arg15Leu mutations. A genotype-phenotype correlation was observed with the Arg15Leu mutation leading to marked hypercalcaemia. FHH3 probands harboured additional phenotypes such as cognitive dysfunction. All three FHH3-causing AP2sigma;2 mutations impaired CaSR signal transduction in a dominant-negative manner. Mutational bias was observed at the AP2sigma;2 Arg15 residue as other predicted missense substitutions (Arg15Gly, Arg15Pro and Arg15Ser), which also caused CaSR loss-of-function, were not detected in FHH probands, and these mutations were found to reduce the numbers of CaSR-expressing cells. FHH3 probands had significantly greater serum calcium (sCa) and magnesium (sMg) concentrations with reduced urinary calcium to creatinine clearance ratios (CCCR) in comparison with FHH1 probands with CaSR mutations, and a calculated index of sCa × sMg/100 × CCCR, which was ≥ 5.0, had a diagnostic sensitivity and specificity of 83 and 86%, respectively, for FHH3. Thus, our studies demonstrate AP2sigma;2 mutations to result in a more severe FHH phenotype with genotype-phenotype correlations, and a dominant-negative mechanism of action with mutational bias at the Arg15 residue.

Optimisation of singlet oxygen production from photosensitiser-incorporated, medically-relevant hydrogels

Photodynamic therapy and photodynamic antimicrobial chemotherapy are widely used, but despite this, the relationships between fluence, wavelength of irradiation and singlet oxygen (1O2) production are poorly understood. To establish the relationships between these factors in medically-relevant materials, the effect of fluence on 1O2 production from a tetrakis(4-N-methylpyridyl)porphyrin (TMPyP)-incorporated 2-hydroxyethyl methacrylate: methyl methacrylate: methacrylic acid (HEMA:MMA:MAA) copolymer, a total energy of 50.48 J/cm², was applied at varying illumination power, and times. 1O2 production was characterised using anthracene-9,10-dipropionic acid, disodium salt (ADPA) using a recently described method. Using two light sources, a white LED array and a white halogen source, the LED array was found to produce less 1O2 than the halogen source when the same power (over 500-600 nm) and time conditions were applied. Importantly, it showed that the longest wavelength Q band (590 nm) is primarily responsible for 1O2 generation, and that a linear relationship exists between increasing power and time and the production of singlet oxygen.

Adjusting HIV Prevalence Estimates for Non-participation: an Application to Demographic Surveillance

Introduction: HIV testing is a cornerstone of efforts to combat the HIV epidemic, and testing conducted as part of surveillance provides invaluable data on the spread of infection and the effectiveness of campaigns to reduce the transmission of HIV. However, participation in HIV testing can be low, and if respondents systematically select not to be tested because they know or suspect they are HIV positive (and fear disclosure), standard approaches to deal with missing data will fail to remove selection bias. We implemented Heckman-type selection models, which can be used to adjust for missing data that are not missing at random, and established the extent of selection bias in a population-based HIV survey in an HIV hyperendemic community in rural South Africa.

Methods: We used data from a population-based HIV survey carried out in 2009 in rural KwaZulu-Natal, South Africa. In this survey, 5565 women (35%) and 2567 men (27%) provided blood for an HIV test. We accounted for missing data using interviewer identity as a selection variable which predicted consent to HIV testing but was unlikely to be independently associated with HIV status. Our approach involved using this selection variable to examine the HIV status of residents who would ordinarily refuse to test, except that they were allocated a persuasive interviewer. Our copula model allows for flexibility when modelling the dependence structure between HIV survey participation and HIV status.

Results: For women, our selection model generated an HIV prevalence estimate of 33% (95% CI 27–40) for all people eligible to consent to HIV testing in the survey. This estimate is higher than the estimate of 24% generated when only information from respondents who participated in testing is used in the analysis, and the estimate of 27% when imputation analysis is used to predict missing data on HIV status. For men, we found an HIV prevalence of 25% (95% CI 15–35) using the selection model, compared to 16% among those who participated in testing, and 18% estimated with imputation. We provide new confidence intervals that correct for the fact that the relationship between testing and HIV status is unknown and requires estimation.

Conclusions: We confirm the feasibility and value of adopting selection models to account for missing data in population-based HIV surveys and surveillance systems. Elements of survey design, such as interviewer identity, present the opportunity to adopt this approach in routine applications. Where non-participation is high, true confidence intervals are much wider than those generated by standard approaches to dealing with missing data suggest.

Optimization of singlet oxygen production from photosensitizer-incorporated, medically relevant hydrogels

Photodynamic therapy and photodynamic antimicrobial chemotherapy are widely used, but despite this, the relationships between fluence, wavelength of irradiation and singlet oxygen ((1) O2 ) production are poorly understood. To establish the relationships between these factors in medically relevant materials, the effect of fluence on (1) O2 production from a tetrakis(4-N-methylpyridyl)porphyrin (TMPyP)-incorporated 2-hydroxyethyl methacrylate: methyl methacrylate: methacrylic acid (HEMA: MMA:MAA) copolymer, a total energy of 50.48 J/cm(2) , was applied at varying illumination power, and times. (1) O2 production was characterized using anthracene-9,10-dipropionic acid, disodium salt (ADPA) using a recently described method. Using two light sources, a white LED array and a white halogen source, the LED array was found to produce less (1) O2 than the halogen source when the same power (over 500 - 600 nm) and time conditions were applied. Importantly, it showed that the longest wavelength Q band (590 nm) is primarily responsible for (1) O2 generation, and that a linear relationship exists between increasing power and time and the production of singlet oxygen. 

Being counted? Examining the prevalence of looked-after disabled children and young people across the UK.

Since the 1970s, there has been growing academic interest in children and young people living in state care and, more recently, in the lives of disabled children. However, there has been little attention on the lives of disabled children who are looked after by the state. This paper compares and critiques what is known about the numbers of disabled children who are looked after in England, Northern Ireland, Scotland and Wales. We discuss the conceptual and methodological limitations of systematically collecting data on disabled children in state care across the United Kingdom. We argue that to ensure the rights of disabled children in state care are identified, acknowledged and upheld, ‘being counted’ is a fundamental first step.

Factors that impact on the use of mechanical ventilation weaning protocols in critically ill adults and children: a qualitative-evidence synthesis

Background:
Prolonged mechanical ventilation is associated with a longer intensive care unit (ICU) length of stay and higher mortality. Consequently, methods to improve ventilator weaning processes have been sought. Two recent Cochrane systematic reviews in ICU adult and paediatric populations concluded that protocols can be effective in reducing the duration of mechanical ventilation, but there was significant heterogeneity in study findings. Growing awareness of the benefits of understanding the contextual factors impacting on effectiveness has encouraged the integration of qualitative evidence syntheses with effectiveness reviews, which has delivered important insights into the reasons underpinning (differential) effectiveness of healthcare interventions.

Objectives:
1. To locate, appraise and synthesize qualitative evidence concerning the barriers and facilitators of the use of protocols for weaning critically-ill adults and children from mechanical ventilation;

2. To integrate this synthesis with two Cochrane effectiveness reviews of protocolized weaning to help explain observed heterogeneity by identifying contextual factors that impact on the use of protocols for weaning critically-ill adults and children from mechanical ventilation;

3. To use the integrated body of evidence to suggest the circumstances in which weaning protocols are most likely to be used.

Search methods:
We used a range of search terms identified with the help of the SPICE (Setting, Perspective, Intervention, Comparison, Evaluation) mnemonic. Where available, we used appropriate methodological filters for specific databases. We searched the following databases: Ovid MEDLINE, Embase, OVID, PsycINFO, CINAHL Plus, EBSCOHost, Web of Science Core Collection, ASSIA, IBSS, Sociological Abstracts, ProQuest and LILACS on the 26th February 2015. In addition, we searched: the grey literature; the websites of professional associations for relevant publications; and the reference lists of all publications reviewed. We also contacted authors of the trials included in the effectiveness reviews as well as of studies (potentially) included in the qualitative synthesis, conducted citation searches of the publications reporting these studies, and contacted content experts.

We reran the search on 3rd July 2016 and found three studies, which are awaiting classification.

Selection criteria:
We included qualitative studies that described: the circumstances in which protocols are designed, implemented or used, or both, and the views and experiences of healthcare professionals either involved in the design, implementation or use of weaning protocols or involved in the weaning of critically-ill adults and children from mechanical ventilation not using protocols. We included studies that: reflected on any aspect of the use of protocols, explored contextual factors relevant to the development, implementation or use of weaning protocols, and reported contextual phenomena and outcomes identified as relevant to the effectiveness of protocolized weaning from mechanical ventilation.

Data collection and analysis:
At each stage, two review authors undertook designated tasks, with the results shared amongst the wider team for discussion and final development. We independently reviewed all retrieved titles, abstracts and full papers for inclusion, and independently extracted selected data from included studies. We used the findings of the included studies to develop a new set of analytic themes focused on the barriers and facilitators to the use of protocols, and further refined them to produce a set of summary statements. We used the Confidence in the Evidence from Reviews of Qualitative Research (CERQual) framework to arrive at a final assessment of the overall confidence of the evidence used in the synthesis. We included all studies but undertook two sensitivity analyses to determine how the removal of certain bodies of evidence impacted on the content and confidence of the synthesis. We deployed a logic model to integrate the findings of the qualitative evidence synthesis with those of the Cochrane effectiveness reviews.

Main results:
We included 11 studies in our synthesis, involving 267 participants (one study did not report the number of participants). Five more studies are awaiting classification and will be dealt with when we update the review.

The quality of the evidence was mixed; of the 35 summary statements, we assessed 17 as ‘low’, 13 as ‘moderate’ and five as ‘high’ confidence. Our synthesis produced nine analytical themes, which report potential barriers and facilitators to the use of protocols. The themes are: the need for continual staff training and development; clinical experience as this promotes felt and perceived competence and confidence to wean; the vulnerability of weaning to disparate interprofessional working; an understanding of protocols as militating against a necessary proactivity in clinical practice; perceived nursing scope of practice and professional risk; ICU structure and processes of care; the ability of protocols to act as a prompt for shared care and consistency in weaning practice; maximizing the use of protocols through visibility and ease of implementation; and the ability of protocols to act as a framework for communication with parents.

Authors' conclusions:
There is a clear need for weaning protocols to take account of the social and cultural environment in which they are to be implemented. Irrespective of its inherent strengths, a protocol will not be used if it does not accommodate these complexities. In terms of protocol development, comprehensive interprofessional input will help to ensure broad-based understanding and a sense of ‘ownership’. In terms of implementation, all relevant ICU staff will benefit from general weaning as well as protocol-specific training; not only will this help secure a relevant clinical knowledge base and operational understanding, but will also demonstrate to others that this knowledge and understanding is in place. In order to maximize relevance and acceptability, protocols should be designed with the patient profile and requirements of the target ICU in mind. Predictably, an under-resourced ICU will impact adversely on protocol implementation, as staff will prioritize management of acutely deteriorating and critically-ill patients.

Regular medication reviews for children and young people with ADHD

The National Institute for Health and Care Excellence's (2008) guidelines for the diagnosis and management of attention deficit hyperactivity disorder (ADHD) recommend a full clinical and psychological assessment by an appropriately trained clinician; this should include a detailed developmental and psychiatric history. Stimulant medications, which are Schedule II controlled drugs, are the most commonly prescribed medicines in the UK and across the world for the management of ADHD. Children and young people with a diagnosis of ADHD receiving these stimulant medications are required to attend regular review appointments with a consultant child and adolescent psychiatrist or specialist nurse under shared care guidelines with general practices, and it has long been recognized that appropriately educated nurses can assist in the management of ADHD. Owing to the pharmacological action of the stimulant medication on neurotransmission, there is potential for misuse and dependence. A growing body of evidence suggests that adolescents with ADHD can become involved in drug diversion and that the topic should be explored during assessment. The level of misuse of prescribed stimulants is increasing, and adolescents and young people with ADHD may misuse to enhance cognitive function for academic purposes. The following scenario highlights some of the challenges and opportunities for independent nurse prescribers working in child and adolescent mental health services.

Mysis salemaai in Ireland: new occurrences and existing population declines

We report two new occurrences of Mysis salemaai, a conservationally important glacial relict at the southern limit of its range, in Castlewellan Lake and Lough Scolban, in the North of Ireland. This increases the number of lakes in Ireland where the species has been recorded to thirteen. We consider lake area and maximum lake depth as factors that might determine the long-term survival of M. salemaai populations and show that these populations tend to occur in relatively large, deep, lakes. We also show that population densities in Lough Neagh and Lough Erne are declining.

Using particle dispersal models to assist in the conservation and recovery of the overexploited native oyster (Ostrea edulis) in an enclosed sea lough

Oyster populations around the world have seen catastrophic decline which has been largely attributed to overexploitation, disease and pollution. While considerable effort and resources have been implemented into restoring these important environmental engineers, the success of oyster populations is often limited by poor understanding of site-specific dispersal patterns of propagules. Water-borne transport is a key factor controlling or regulating the dispersal of the larval stage of benthic marine invertebrates which have limited mobility. The distribution of the native oyster Ostrea edulis in Strangford Lough, Northern Ireland, together with their densities and population structure at subtidal and intertidal sites has been documented at irregular intervals between 1997 and 2013. This paper revisits this historical data and considers whether different prevailing environmental conditions can be used to explain the distribution, densities and population structure of O. edulis in Strangford Lough. The approach adopted involved comparing predictive 2D hydrodynamic models coupled with particle tracking to simulate the dispersal of oyster larvae with historical and recent field records of the distribution of both subtidal and intertidal, populations since 1995. Results from the models support the hypothesis that commercial stocks of O. edulis introduced into Strangford Lough in the 1990s resulted in the re-establishment of wild populations of oysters in the Northern Basin which in turn provided a potential source of propagules for subtidal populations. These results highlight that strategic site selection (while inadvertent in the case of the introduced population in 1995) for the re-introduction of important shellfish species can significantly accelerate their recovery and restoration.

Biocontrol agents promote growth of potato pathogens, depending on environmental conditions

There is a pressing need to understand and optimize biological control so as to avoid over-reliance on the synthetic chemical pesticides that can damage environmental and human health. This study focused on interactions between a novel biocontrol-strain, Bacillus sp. JC12GB43, and potato-pathogenic Phytophthora and Fusarium species. In assays carried out in vitro and on the potato tuber, the bacterium was capable of near-complete inhibition of pathogens. This Bacillus was sufficiently xerotolerant (water activity limit for growth = 0.928) to out-perform Phytophthora infestans (~0.960) and challenge Fusarium coeruleum (~0.847) and Fusarium sambucinum (~0.860) towards the lower limits of their growth windows. Under some conditions, however, strain JC12GB43 stimulated proliferation of the pathogens: for instance, Fusarium coeruleum growth-rate was increased under chaotropic conditions in vitro (132 mM urea) by >100% and on tubers (2-M glycerol) by up to 570%. Culture-based assays involving macromolecule-stabilizing (kosmotropic) compatible solutes provided proof-of-principle that the Bacillus may provide kosmotropic metabolites to the plant pathogen under conditions that destabilize macromolecular systems of the fungal cell. Whilst unprecedented, this finding is consistent with earlier reports that fungi can utilize metabolites derived from bacterial cells. Unless the antimicrobial activities of candidate biocontrol strains are assayed over a full range of field-relevant parameters, biocontrol agents may promote plant pathogen infections and thereby reduce crop yields. These findings indicate that biocontrol activity, therefore, ought to be regarded as a mode-of-behaviour (dependent on prevailing conditions) rather than an inherent property of a bacterial strain.

New insights into the FLPergic complements of parasitic nematodes: Informing deorphanisation approaches

FMRFamide-like peptide (FLP) receptors are appealing as putative anthelmintic targets. To date, 31 flp-encoding genes have been identified in Caenorhabditis elegans and thirteen FLP-activated G-protein coupled receptors (FLP-GPCRs) have been reported. The lack of knowledge on FLPs and FLP-GPCRs in parasites impedes their functional characterisation and chemotherapeutic exploitation. Using homology-based BLAST searches and phylogenetic analyses this study describes the identification of putative flp and flp-GPCR gene homologues in 17 nematode parasites providing the first pan-phylum genome-based overview of the FLPergic complement. These data will facilitate FLP-receptor deorphanisation efforts in the quest for novel control targets for nematode parasites.