Committee experiences of using formal consensus in healthcare guidelines: a longitudinal qualitative study.

BACKGROUND: This feasibility study has the primary aim of capturing and comparing participant expectations and experiences of using a formal consensus method (FCM) and to explore whether these views change following participation within a guideline committee where FCM are used. METHODS: Twelve healthcare committee members and associated technical team members participated in semi-structured qualitative interviews before and after using FCM during guideline committee meetings. Interviews also focused on past experiences and expectations of informal consensus methods. RESULTS: Participants said formal consensus included a greater range of evidence. They described positive reactions and found it a useful way to encourage involvement by balancing group power dynamics. Group discussion time was identified as important to clarify ideas, supported by good group chairing. However, participants reported that undertaking FCM required additional resources and suggested targeting its use for low quality evidence, limited committee expertise, or where the evidence is controversial. CONCLUSIONS: FCM is an acceptable alternative to informal consensus methods that has qualities specifically helpful to healthcare guidelines such as encouraging participation, inclusivity of a broad range of evidence, and managing group dynamics. More research is required to better understand when using formal consensus is most appropriate and effective.

Professionalism of physicians at a major teaching hospital during the Fukushima nuclear disaster.

It poses a serious problem if physicians leave a hospital without having a replacement or without permission. A huge earthquake followed by a devastating tsunami seriously damaged the Fukushima-Daiichi nuclear power plant. This disaster overwhelmed a major teaching hospital in the local area and many hospital employees, including some resident physicians, left the premises. Since the threat of severe radiation exposure poses a potentially greater lifetime risk to younger individuals, letting the young resident physicians leave the hospital was not only allowed, it was actually recommended by many attending physicians and hospital administrators. The hospital administrator was required to make the difficult decision of whether to make all efforts to provide the highest level of medical care, including keeping all of the physicians on the premises, or to evacuate the resident physicians in order to preserve their health and their potential future contributions to healthcare. Consideration and compassion needed to be provided to all people, regardless of the reason they wanted to leave. From an ethical perspective, the roles of performance under these complex circumstances should be understood and embraced by us as individuals, professionals, supervisors and society as a whole.

Uptake and utilization of directly acting antiviral medications for hepatitis C infection in U.S. veterans.

New drugs therapies have revolutionized the treatment of hepatitis C virus (HCV) infection. The objectives of this study were to evaluate uptake and utilization of boceprevir and telaprevir in the Department of Veterans Affairs (VA). We evaluated whether therapies conformed to response-guided protocols, whether they replaced standard interferon plus ribavirin treatment, and whether IL-28B was used to guide treatment. We performed an administrative data-based analysis of all patients receiving pharmacologic treatment for HCV in VA from October 2009 to July 2013. There were 12 737 new HCV prescriptions in VA during this time, with 5564 boceprevir or telaprevir prescriptions (44%) and 7173 prescriptions (56%) written for standard interferon plus ribavirin treatment. Prescriptions for the new treatments heavily favoured boceprevir vs telaprevir (83% vs 17%). Sixty-two percent (62%) of boceprevir-treated patients completed their minimum-specified protocol, while 69.2% of telaprevir-treated patients completed their minimum-specified protocol. From October 2010 to July 2012, 4090 patients had an IL-28B test; less than 16% of these tests guided subsequent HCV prescriptions. Uptake of boceprevir and telaprevir was rapid; the number of patients initiating treatment approximately doubled in the period after their introduction. While new prescriptions favor boceprevir or telaprevir over standard interferon plus ribavirin therapy, there appears to still be a strong role of interferon plus ribavirin in treating HCV patients. This work can inform our understanding of how other new effective HCV therapies will be used, their diffusion, and the timing of their diffusion in actual clinical practice.

A Review of OIE Country Status Recovery Using Vaccinate-to-Live Versus Vaccinate-to-Die Foot-and-Mouth Disease Response Policies I: Benefits of Higher Potency Vaccines and Associated NSP DIVA Test Systems in Post-Outbreak Surveillance.

To rapidly return to trade, countries with OIE status, FMD-free country where vaccination is not practised, have destroyed emergency vaccinated animals, raising ethical concerns with respect to social values, the environment, animal welfare and global food security. This two-part review explores whether science could support eligibility to return to previous OIE status in 3 months irrespective of vaccinate-to-live or vaccinate-to-die policies. Here, we examine the benefits of higher potency (≥ 6 PD50 ), high-purity vaccines formulated from antigen banks for emergency use, their efficacy and performance in differentiating infected from vaccinated animals (DIVA) assays for post-outbreak surveillance. From an intensive programme of research, we conclude that high-quality, higher potency vaccines are proven to reduce FMD virus (FMDV) subclinical circulation and the risk of carriers. Broader coverage than predicted by serology suggests the potential to hold a few 'key' vaccine strains improving logistics and reducing the financial burden of antigen banks. The OIE should adopt formal definitions for emergency vaccination and emergency vaccines. In terms of supportive tools, we consider that the lack of OIE recognition of DIVA tests other than those of PANAFTOSA in cattle is a shortcoming. There is need for research on maternal antibody interference with DIVA tests and on the use of such tests to establish whether greater purification of vaccines improves performance. We consider that alignment of waiting periods for vaccinate-to-live and vaccinate-to-die in OIE Code Article 8.5.9 1 b. and c. is feasible until an acceptable level of statistical certainty for surveillance or target probability of freedom is established to substantiate the absence of FMDV infection or circulation. It is surveillance intensity rather than waiting periods that establishes the risk of residual FMDV. EU Directive 2003/85/EC implicitly recognizes this, permitting derogation of the OIE waiting periods.

A Review of OIE Country Status Recovery Using Vaccinate-to-Live Versus Vaccinate-to-Die Foot-and-Mouth Disease Response Policies II: Waiting Periods After Emergency Vaccination in FMD Free Countries.

For countries with OIE status, FMD free country where vaccination is not practised, vaccinate-to-live policies have a significant economic disincentive as the trade restriction waiting period is double that of vaccinate-to-die policies. The disposal of healthy vaccinated animals strictly for the purpose of regaining markets with debatable scientific justification is a global concern. The feasibility of aligning the waiting periods to facilitate vaccinate-to-live is explored. The first article of this two-part review (Barnett et al., 2015) explored the qualities of higher potency Foot-and-Mouth Disease (FMD) vaccines, performance of differentiating infected from vaccinated animals (DIVA) diagnostic assays particularly in vaccinates and carriers, as well as aspects of current limitations of post-outbreak surveillance. Here, the history behind the OIE waiting periods for FMD free status is reviewed as well as whether the risk of vaccinated animals and their subsequent products differ appreciably at 3 versus 6 months. It is concluded that alignment is feasible for vaccinate-to-live using higher potency FMD vaccines within the current OIE waiting period framework of 3 and 6 months blocks of time. These waiting periods reflect precedence, historical practicalities and considered expert opinion rather than a specific scientific rationale. The future lies in updated epidemiological and diagnostic technology to establish an acceptable level of statistical certainty for surveillance or target probability of freedom of FMDV (infection or circulation) not time restricted waiting periods. The OIE Terrestrial Code limits trade from a FMD free country where vaccination is not practiced to animal products and live non-vaccinated animals. The risk of FMDV in products derived from higher potency vaccinated animals is appreciably less than for countries with infected FMD status or even from a FMD free country where vaccination is practised for which the Code has Articles with guidelines for safe trade with time restrictions of 3 months or less. All these presume that key requirements in the implementation of emergency vaccination including appropriate vaccine match, vaccine application, susceptible population coverage, etc. are addressed.

Mutations in the T (brachyury) gene cause a novel syndrome consisting of sacral agenesis, abnormal ossification of the vertebral bodies and a persistent notochordal canal.

BACKGROUND: The T gene (brachyury gene) is the founding member of the T-box family of transcription factors and is vital for the formation and differentiation of the mesoderm and the axial development of all vertebrates. RESULTS: We report here on four patients from three consanguineous families exhibiting sacral agenesis, a persistent notochordal canal and abnormal ossification of the vertebral bodies, and the identification and characterisation of their underlying genetic defect. Given the consanguineous nature and the similarity of the phenotypes between the three families, we performed homozygosity mapping and identified a common 4.1 Mb homozygous region on chromosome 6q27, containing T, brachyury homologue (mouse) or T. Sequencing of T in the affected individuals led to the identification of a homozygous missense mutation, p.H171R, in the highly conserved T-box. The homozygous mutation results in diminished DNA binding, increased cell growth, and interferes with the normal expression of genes involved in ossification, notochord maintenance and axial mesoderm development. CONCLUSIONS: We have identified a shared homozygous mutation in three families in T and linked it to a novel syndrome consisting of sacral agenesis, a persistent notochordal canal and abnormal ossification of the vertebral bodies. We suggest that screening for the ossification of the vertebrae is warranted in patients with sacral agenesis to evaluate the possible causal involvement of T.

Acquisition of high quality DNA for massive parallel sequencing by in vivo chromatin immunoprecipitation.

ChIP-seq is rapidly becoming a routine technique for the determination of the genome wide association of DNA binding proteins and histone modifications. Here we provide a protocol for the isolation, purification, and immunoprecipitation of DNA fragments associated with a target transcription factor of interest. Although the method makes use of adult mouse hearts, it can, with relative ease, be adapted for the in vivo ChIP isolation of DNA from other cell and tissue sources with the intention of massive parallel sequencing.

Nuclear recruitment assay as a tool to validate transcription factor interactions in Mammalian cells.

Identification and verification of novel transcription factor interactions is an inherent step in the discovery of molecular mechanisms driving gene transcription and regulation. Co-immunoprecipitation and GST-pull down are often key techniques in the verification process. Despite wide applicability, their use may sometimes be restricted. We provide a detailed protocol for an intracellular immunofluorescence technique that may be used as an alternative or complimentary study for transcription factor interaction verification.

A DNA vaccination regime including protein boost and electroporation protects cattle against foot-and-mouth disease.

Protection against foot-and-mouth disease (FMD) using DNA technology has been documented for sheep and pigs but not for the highly susceptible species of cattle. Twenty-five Holstein Friesian cross-bred cattle were vaccinated twice, 21 days apart, with a DNA vaccine containing the capsid coding region (P1) along with the non-structural proteins 2A, 3C and 3D (pcDNA3.1/P1-2A3C3D) of O(1) Kaufbeuren alone or coated onto PLG (d,l-lactide-co-glycolide) microparticles. In some pcDNA3.1/P1-2A3C3D was also combined with an adjuvant plasmid expressing bovine granulocyte macrophage colony stimulating factor (GM-CSF). DNA vaccinations were administered intramuscularly with, or without, the use of electroporation and at 42 days post primary vaccination cattle received a protein boost of 146S FMD virus (FMDV) antigen and non-structural protein 3D. For comparison, four cattle were vaccinated with a conventional FMD vaccine and two more included as unvaccinated controls. Apart from those immunised with PLG microparticles all cattle were challenged with 10(5) TCID(50) cattle adapted O(1) Lausanne FMDV virus at day 93 post primary vaccination. All DNA vaccinated cattle regardless of regime developed good humoral and cell mediated responses prior to challenge. The best overall virus neutralising antibody, IFN-γ and clinical protection (75%) were seen in the cattle whereby the DNA was delivered by electroporation. In contrast, only 25% of cattle vaccinated with the DNA vaccine without electroporation were clinically protected. The addition of GM-CSF in combination with electroporation further improved the efficacy of the vaccine, as demonstrated from the reduction of clinical disease and virus excretions in nasal swabs. We thus demonstrate for the first time that cattle can be clinically protected against FMDV challenge following a DNA prime-protein boost strategy, and particularly when DNA vaccine is combined with GM-CSF and delivered by electroporation.

Foot-and-mouth disease marker vaccine: cattle protection with a partial VP1 G-H loop deleted virus antigen.

Contrary to the dogma that the VP1 G-H loop is essential for FMD vaccine efficacy, it has been previously shown that foot-and-mouth disease 146s antigen containing heterologous VP1 G-H loops confers complete protection in pigs and cattle. Moreover, serological evaluation of cattle vaccinated with an antigen lacking a large proportion of the VP1 G-H loop indicated that these animals should be protected against infection with FMD. Absence of this loop provides opportunity for the development of an FMD negative marker vaccine, allowing infection to be detected by antibodies against this missing region. Cattle vaccinated with this negative marker vaccine were fully protected following virus challenge 28 days post vaccination as determined by the absence of generalised lesions on their feet. Furthermore, use of our improved differentiation ELISA identified animals exposed to infection as early as 7 days post-challenge. We thus demonstrate, for the first time, the ability of this FMD negative marker vaccine to fully protect cattle from experimental challenge and rapidly distinguish animals that are subsequently exposed to infection.

Sox4 mediates Tbx3 transcriptional regulation of the gap junction protein Cx43.

Tbx3, a T-box transcription factor, regulates key steps in development of the heart and other organ systems. Here, we identify Sox4 as an interacting partner of Tbx3. Pull-down and nuclear retention assays verify this interaction and in situ hybridization reveals Tbx3 and Sox4 to co-localize extensively in the embryo including the atrioventricular and outflow tract cushion mesenchyme and a small area of interventricular myocardium. Tbx3, SOX4, and SOX2 ChIP data, identify a region in intron 1 of Gja1 bound by all tree proteins and subsequent ChIP experiments verify that this sequence is bound, in vivo, in the developing heart. In a luciferase reporter assay, this element displays a synergistic antagonistic response to co-transfection of Tbx3 and Sox4 and in vivo, in zebrafish, drives expression of a reporter in the heart, confirming its function as a cardiac enhancer. Mechanistically, we postulate that Sox4 is a mediator of Tbx3 transcriptional activity.

Characterisation and epitope mapping of neutralising monoclonal antibodies to A24 Cruzeiro strain of FMDV.

Characterisation of seven neutralising monoclonal antibodies (mAbs) produced against foot-and-mouth disease virus A(24) Cruzeiro revealed three reactivity groups. Gr-I recognised linear epitopes where as Gr-II was conformation-dependent and trypsin-insensitive. The Gr-III was also conformation-dependent, but trypsin-sensitive. Mar (mAb neutralisation resistant)-mutants could only be produced against Gr-I and Gr-III mAbs. Capsid sequence comparison of Gr-I mar-mutants with parent virus revealed changes in the G-H loop of VP1 at positions 141, 143 and 147. Similarly, a Gr-III mar-mutant showed a change from a highly conserved glycine to a tryptophan at position 148 of VP1 along with three additional changes at the N-terminus of VP1, VP2 and VP4. This residue at 148 of VP1 is located at +2 position after "RGD" and is equivalent to the position identified by the mAb recognising site 5 in serotype O viruses. This site is probably formed because of the interaction of the G-H loop with other residues in different structural proteins.

Marker vaccine potential of a foot-and-mouth disease virus with a partial VP1 G-H loop deletion.

Previous work in cattle and pigs demonstrated that protection against foot-and-mouth disease (FMD) could be achieved following vaccination with chimeric foot-and-mouth disease virus (FMDV) vaccines, in which the VP1 G-H loop had been substituted with that from another serotype. This indicated that the VP1 G-H loop may not be essential for the protection of natural hosts against FMDV. If this could be substantiated there would be potential to develop FMD marker vaccines, characterised by the absence of this region. Here, we investigate the serological responses to vaccination with a virus with a partial VP1 G-H loop deletion in order to determine the likelihood of achieving protection and the potential of this virus as a marker vaccine. Inactivated, oil adjuvanted, vaccines, consisting of chemically inactivated virus with or without a partially deleted VP1 G-H loop, were used to immunise cattle. Serum was collected on days 0, 7, 14 and 21 and antibody titres calculated using the virus neutralisation test (VNT) to estimate the likelihood of protection. We predict a good likelihood that cattle vaccinated with a vaccine characterised by a partial VP1 G-H loop would be protected against challenge with the same virus containing the VP1 G-H loop. We also present evidence on the potential of such a construct to act as a marker vaccine, when used in conjunction with a novel serological test.

Toward a global foot and mouth disease vaccine bank network.

A network of foot and mouth (FMD) vaccine banks has been initiated with the support of vaccine bank managers and technical advisors that participated in a workshop held at the Institute for Animal Health, Pirbright, in the United Kingdom in April 2006. Terms of Reference that provide guidance for coordinated activities are under consultation. Practical and economic benefits can be realised from collaboration, which will be achieved through mutually acceptable mechanisms for the exchange of information and materials relevant to vaccine banks and their management. If administrative and technical hurdles can be overcome, the network has the potential to contribute significantly to the improved control of FMD worldwide. A 'global' and interactive vaccine bank association could be created by agreeing a system of resource sharing that could orchestrate additional emergency cover with vaccine or antigen from the reserves of network members.

The importance of quality assurance/quality control of diagnostics to increase the confidence in global foot-and-mouth disease control.

The last decade international trade in animals and animal products was liberated and confidence in this global trade can increase only if appropriate control measures are applied. As foot-and-mouth disease (FMD) diagnostics will play an essential role in this respect, the Food and Agriculture Organization European Commission for the Control of Foot-and-Mouth Disease (EUFMD) co-ordinates, in collaboration with the European Commission, several programmes to increase the quality of FMD diagnostics. A quality assurance (QA) system is deemed essential for laboratories involved in certifying absence of FMDV or antibodies against the virus. Therefore, laboratories are encouraged to validate their diagnostic tests fully and to install a continuous quality control (QC) monitoring system. Knowledge of performance characteristics of diagnostics is essential to interpret results correctly and to calculate sample rates in regional surveillance campaigns. Different aspects of QA/QC of classical and new FMD virological and serological diagnostics are discussed in respect to the EU FMD directive (2003/85/EC). We recommended accepting trade certificates only from laboratories participating in international proficiency testing on a regular basis.

FMD vaccines: reflections on quality aspects for applicability in European disease control policy.

Most foot-and-mouth disease (FMD) vaccines used around the world are inactivated vaccines for prophylactic or emergency use, generally manufactured by the same basic methodology outlined in the OIE Manual and, for Europe, in the European Pharmacopoeia, and for the EU Member States in compliance with Directive 2001/82/EC of the European Parliament and of the Council of 6 November 2001 on the Community code relating to veterinary medicinal products as amended by Directive 2004/28/EC. Most of the requirements that apply to all immunological veterinary medicinal products apply equally to FMD vaccines. There are, however, some unique features of the disease and vaccines used against it that require a different approach to fulfil the requirements of the relevant legislation, if a vaccinate-to-live policy will be applied with 'authorized' vaccines. Several aspects of vaccine efficacy and safety are elaborated with emphasis on quality assurance/quality control (QA/QC). The purity of the vaccine in respect of the presence of non-structural protein antibodies could be checked indirectly by serology after vaccination. The viability of a vaccine bank approach was greatly aided by the principle of storing inactivated concentrated FMD viral antigen (Ag) over liquid nitrogen for subsequent formulation into vaccine. A worldwide Ag bank network might be an option for the far future and a solution to the problem of covering many different FMDV serotypes and strains. The producers should respect the strict FMD biosecurity rules worked out by the FAO EUFMD and described in Council Directive 2003/85/EC. Making the experience related to vaccine QA/QC available to all countries will reduce the risk of an FMD outbreak within these countries and consequently will reduce the FMD risk around the world.

Chimeric foot-and-mouth disease viruses: evaluation of their efficacy as potential marker vaccines in cattle.

Previous work in pigs, has demonstrated that full protection against foot-and-mouth disease (FMD) can be achieved following vaccination with chimeric foot-and-mouth disease virus (FMDV) vaccines, in which the VP1 G-H loop had been substituted with that from another serotype. If proven to be effective in other economically important species such as cattle, such vaccine constructs could be trialed as potential marker vaccines. Here, we determine if G-H loop chimera FMDV vaccines can: (i) protect cattle from virus challenge and (ii) induce an antibody response that would enable the identification of infection, regardless of vaccination status. Inactivated, oil adjuvanated, chimeric vaccine constructs, based on the backbone sequence of the A(12)119 serotype virus, fully protected cattle from challenge 21 days post-vaccination. Differentiation assays developed for use in this study were able to identify sub-clinical infection, which in one vaccinated animal, persisted beyond day 32 post-challenge. This paper emphasises the importance of epitopes outside of the VP1 G-H loop for protective immunity in cattle, and demonstrates that chimeric FMDV vaccines could prove to be useful marker vaccines for the future.

Evaluation of a monoclonal antibody-based approach for the selection of foot-and-mouth disease (FMD) vaccine strains.

Foot-and-mouth disease (FMD) virus exists as seven serotypes within which are numerous variants necessitating careful selection of vaccine strains. Currently, a serological assay system based on the use of polyclonal vaccine antisera is widely used for this selection. However, inherent variability in the matching antisera used makes the tests poorly reproducible and difficult to interpret. In this study, we have explored the possibility of replacing or supplementing the polyclonal antibody (PAb)-based method with one based on use of monoclonal antibodies (MAb). Panels of MAbs raised against two serotype O vaccine strains were examined for reactivity with 22 field viruses, isolated over a 10-year period between 1991 and 2001. Antigenic site 2 was found to comprise more than one epitope. The sequence variation in capsid protein VP2 harbouring antigenic site 2 was analysed and the amino acid residues at positions 79 and 134 appeared to greatly influence the binding of site 2 MAbs. Prediction of antigenic match based on MAb reactivity did not correlate closely with the results of a PAb-based "gold-standard" method and it was concluded that a wider panel of MAbs are needed that recognise all protective epitopes present on the surface of FMD virus together with a better understanding of those epitopes which are important in conferring protection.

T-box factors determine cardiac design.

The heart of higher vertebrates is a structurally complicated multi-chambered pump that contracts synchronously. For its proper function a number of distinct integrated components have to be generated, including force-generating compartments, unidirectional valves, septa and a system in charge of the initiation and coordinated propagation of the depolarizing impulse over the heart. Not surprisingly, a large number of regulating factors are involved in these processes that act in complex and intertwined pathways to regulate the activity of target genes responsible for morphogenesis and function. The finding that mutations in T-box transcription factor-encoding genes in humans lead to congenital heart defects has focused attention on the importance of this family of regulators in heart development. Functional and genetic analyses in a variety of divergent species has demonstrated the critical roles of multiple T-box factor gene family members, including Tbx11, -2, -3, -5, -18 and -20, in the patterning, recruitment, specification, differentiation and growth processes underlying formation and integration of the heart components. Insight into the roles of T-box factors in these processes will enhance our understanding of heart formation and the underlying molecular regulatory pathways.

Efficacy of particle-based DNA delivery for vaccination of sheep against FMDV.

As an alternative strategy to classical inactivated viral vaccine against FMDV, naked DNA vaccine is attractive because of safety, flexibility and low cost. However DNA vaccination is usually poorly efficient in target species. Indeed we found that naked DNA plasmids encoding for P1-2A3C3D and GM-CSF proteins did not induce any detectable immunity against FMDV in sheep. Interestingly, we demonstrate herein that formulations of DNA on poly(D,L-lactide-co-glycolide) (PLG) or in lipofectin triggered divergent types of immune responses: PLG stimulated a T cell response and could elicit significant neutralising antibody titers, whereas lipofectin generated even higher antibody titers but no significant T cell response. The DNA/PLG regimen used in five sheep protected against clinical symptoms and viraemia and prevented the carrier state in four of them. Thus formulated DNA can be remarkably efficient against FMDV in a ruminant species that is usually refractory to DNA vaccination.