FRAME and the Royal Commission on Environmental Pollution: common recommendations for assessing risks posed by chemicals under the EU REACH system

This document discusses recommendations made by FRAME and the Royal Commission on Environmental Pollution (RCEP) with regard to the current European Commission proposals on the Registration, Evaluation and Authorisation of Chemicals (REACH) system for assessing the risks of chemicals to humans, wildlife and the environment. Of several common aims and recommendations, the two most important are: a) the greater use of non-animal testing methods, especially computational prediction methods (for example, [quantitative] structure-activity relationships, expert systems and biokinetic modelling) for prioritising chemicals for hazard assessment; and b) the greater use of intelligent exposure-based targeted risk assessment, with less emphasis being placed on tonnage-triggers. FRAME has produced a decision-tree testing scheme to illustrate the way in which these approaches could be used, together with in vitro test methods. This scheme has been slightly modified to take account of proposals subsequently made by the RCEP. In addition, FRAME points out that new and improved computational methods are needed through more coordinated research, and that these and existing methods need to be validated. The similarities between the independent publications of FRAME and the RCEP add weight to the recommendations that each have made concerning the implementation of the REACH system.     

FRAME and the Royal Commission on Environmental Pollution: common recommendations for assessing risks posed by chemicals under the EU REACH system

This document discusses recommendations made by FRAME and the Royal Commission on Environmental Pollution (RCEP) with regard to the current European Commission proposals on the Registration, Evaluation and Authorisation of Chemicals (REACH) system for assessing the risks of chemicals to humans, wildlife and the environment. Of several common aims and recommendations, the two most important are: a) the greater use of non-animal testing methods, especially computational prediction methods (for example, [quantitative] structure-activity relationships, expert systems and biokinetic modelling) for prioritising chemicals for hazard assessment; and b) the greater use of intelligent exposure-based targeted risk assessment, with less emphasis being placed on tonnage-triggers. FRAME has produced a decision-tree testing scheme to illustrate the way in which these approaches could be used, together with in vitro test methods. This scheme has been slightly modified to take account of proposals subsequently made by the RCEP. In addition, FRAME points out that new and improved computational methods are needed through more coordinated research, and that these and existing methods need to be validated. The similarities between the independent publications of FRAME and the RCEP add weight to the recommendations that each have made concerning the implementation of the REACH system.     

Genetic laboratory practices related to testing of the GJB2 (Connexin-26) gene in the United States in 1999 and 2000

Given the rapidly growing area of molecular genetic laboratory testing, we sought to assess changing issues over a 2-year period of time pertaining to the availability of testing for GJB2 mutations associated with non-syndromic hearing loss. Laboratory assessments carried out by telephone interviews with directors or other key personnel revealed variations among laboratories in informed consent practices, evaluation of test requests for appropriateness, and the reporting of results. From 1999 to 2000, referral patterns shifted as did sources for reimbursement, policies regarding evaluation of incoming test requests, and reporting procedures. We propose that these results reflect changes occurring as a result of a new test moving from the research to the clinical phase.     

White‐crowned sparrow males show immediate flexibility in song amplitude but not in song minimum frequency in response to changes in noise levels in the field

The soundscape acts as a selective agent on organisms that use acoustic signals to communicate. A number of studies document variation in structure, amplitude, or timing of signal production in correspondence with environmental noise levels thus supporting the hypothesis that organisms are changing their signaling behaviors to avoid masking. The time scale at which organisms respond is of particular interest. Signal structure may evolve across generations through processes such as cultural or genetic transmission. Individuals may also change their behavior during development (ontogenetic change) or in real time (i.e., immediate flexibility). These are not mutually exclusive mechanisms, and all must be investigated to understand how organisms respond to selection pressures from the soundscape. Previous work on white‐crowned sparrows (Zonotrichia leucophrys) found that males holding territories in louder areas tend to sing higher frequency songs and that both noise levels and song frequency have increased over time (30 years) in urban areas. These previous findings suggest that songs are changing across generations; however, it is not known if this species also exhibits immediate flexibility. Here, we conducted an exploratory, observational study to ask whether males change the minimum frequency of their song in response to immediate changes in noise levels. We also ask whether males sing louder, as increased minimum frequency may be physiologically linked to producing sound at higher amplitudes, in response to immediate changes in environmental noise. We found that territorial males adjust song amplitude but not minimum frequency in response to changes in environmental noise levels. Our results suggest that males do not show immediate flexibility in song minimum frequency, although experimental manipulations are needed to test this hypothesis further. Our work highlights the need to investigate multiple mechanisms of adaptive response to soundscapes.     

An overall strategy for the testing of chemicals for human hazard and risk assessment under the EU REACH system

In its White Paper, "Strategy for a Future Chemicals Policy," published in 2001, the European Commission (EC) proposed the REACH (Registration, Evaluation and Authorisation of CHemicals) system to deal with both existing and new chemical substances. This system is based on a top-down approach to toxicity testing, in which the degree of toxicity information required is dictated primarily by production volume (tonnage). If testing is to be based on traditional methods, very large numbers of laboratory animals could be needed in response to the REACH system, causing ethical, scientific and logistical problems that would be incompatible with the time-schedule envisaged for testing. The EC has emphasised the need to minimise animal use, but has failed to produce a comprehensive strategy for doing so. The present document provides an overall scheme for predictive toxicity testing, whereby the non-animal methods identified and discussed in a recent and comprehensive ECVAM document, could be used in a tiered approach to provide a rapid and scientifically justified basis for the risk assessment of chemicals for their toxic effects in humans. The scheme starts with a preliminary risk assessment process (involving available information on hazard and exposure), followed by testing, based on physicochemical properties and (Q)SAR approaches. (Q)SAR analyses are used in conjunction with expert system and biokinetic modelling, and information on metabolism and identification of the principal metabolites in humans. The resulting information is then combined with production levels and patterns of use to assess potential human exposure. The nature and extent of any further testing should be based strictly on the need to fill essential information gaps in order to generate adequate risk assessments, and should rely on non-animal methods, as far as possible. The scheme also includes a feedback loop, so that new information is used to improve the predictivity of computational expert systems. Several recommendations are made, the most important of which is that the European Union (EU) should actively promote the improvement and validation of (Q)SAR models and expert systems, and computer-based methods for biokinetic modelling, since these offer the most realistic and most economical solution to the need to test large numbers of chemicals.     

An overall strategy for the testing of chemicals for human hazard and risk assessment under the EU REACH system

In its White Paper, Strategy for a Future Chemicals Policy, published in 2001, the European Commission (EC) proposed the REACH (Registration, Evaluation and Authorisation of CHemicals) system to deal with both existing and new chemical substances. This system is based on a top-down approach to toxicity testing, in which the degree of toxicity information required is dictated primarily by production volume (tonnage). If testing is to be based on traditional methods, very large numbers of laboratory animals could be needed in response to the REACH system, causing ethical, scientific and logistical problems that would be incompatible with the time-schedule envisaged for testing. The EC has emphasised the need to minimise animal use, but has failed to produce a comprehensive strategy for doing so. The present document provides an overall scheme for predictive toxicity testing, whereby the non-animal methods identified and discussed in a recent and comprehensive ECVAM document, could be used in a tiered approach to provide a rapid and scientifically justified basis for the risk assessment of chemicals for their toxic effects in humans. The scheme starts with a preliminary risk assessment process (involving available information on hazard and exposure), followed by testing, based on physicochemical properties and (Q)SAR approaches. (Q)SAR analyses are used in conjunction with expert system and biokinetic modelling, and information on metabolism and identification of the principal metabolites in humans. The resulting information is then combined with production levels and patterns of use to assess potential human exposure. The nature and extent of any further testing should be based strictly on the need to fill essential information gaps in order to generate adequate risk assessments, and should rely on non-animal methods, as far as possible. The scheme also includes a feedback loop, so that new information is used to improve the predictivity of computational expert systems. Several recommendations are made, the most important of which is that the European Union (EU) should actively promote the improvement and validation of (Q)SAR models and expert systems, and computer-based methods for biokinetic modelling, since these offer the most realistic and most economical solution to the need to test large numbers of chemicals.     

Influence of adjuvant radiotherapy on circulating epithelial tumor cells and circulating cancer stem cells in primary non-metastatic breast cancer

Background: There is an unmet need to identify biomarkers that directly reflect response to adjuvant radiotherapy (RT). Circulating epithelial tumor cells (CETCs) represent the liquid component of solid tumors and are responsible for metastatic relapse. CETC subsets with cancer stem cell characteristics, circulating cancer stem cells (cCSCs), play a pivotal role in the metastatic cascade. Monitoring the most aggressive subpopulation of CETCs could reflect the aggressiveness of the remaining tumor burden. There is limited data on the detection and monitoring changes in CETC and cCSC numbers during RT in early breast cancer.Methods: CETC numbers were analyzed prior to, at midterm and at the end of RT in 52 primary non-metastatic breast cancer patients. Hormone receptor status was determined in CETCs prior to and at the end of RT. For the identification of cCSCs cell suspensions from the peripheral blood of patients were cultured in vitro under conditions favoring growth of tumorspheres.Results: Hormone receptor status in CETCs before RT was comparable to that in primary tumor tissue. Prior to RT numbers of CETCs correlated with aggressiveness of primary tumors. cCSCs could be successfully identified and monitored during RT. Prior to RT patients treated with neoadjuvant chemotherapy had significantly higher numbers of CETCs and tumorspheres compared to patients after adjuvant chemotherapy. During RT, the number of CETCs decreased continuously in patients after neoadjuvant chemotherapy but not after adjuvant chemotherapy.Conclusion: Monitoring the number of CETCs and the CETC subset with cancer stem cell properties during RT may provide additional clinically useful prognostic information.     

Delayed-Incubation Membrane-Filter Test for Fecal Coliforms

A delayed-incubation membrane-filter technique for fecal coliforms was developed and compared with the immediate fecal coliform test described in Standard Methods for the Examination of Water and Wastewater (13th ed., 1971). Laboratory and field evaluations demonstrated that the delayed-incubation test, with the use of the proposed vitamin-free Casitone holding medium, produces fecal coliform counts which very closely approximate those from the immediate test, regardless of the source or type of fresh-water sample. Limited testing indicated that the method is not as effective when used with saline waters. The delayed-incubation membrane-filter test will be especially useful in survey monitoring or emergency situations when the standard immediate fecal coliform test cannot be performed at or near the sample site or when time and temperature limitations for water sample storage cannot be met. The procedure can also be used for analyzing the bacterial quality of water or waste discharges by a standardized procedure in a central examining laboratory remote from the sample source.     

Evidence-Based Laboratory Medicine: Is It Working in Practice?

The principles of Evidence-Based Medicine have been established for about two decades, with the need for evidence-based clinical practice now being accepted in most health systems around the world. These principles can be employed in laboratory medicine. The key steps in evidence-based practice, namely (i) formulating the question; (ii) searching for evidence; (iii) appraising evidence; (iv) applying evidence; and (v) assessing the experience are all accepted but, as yet, translation into daily clinical and laboratory practice has been slow. Furthermore, the demand for evidence-based laboratory medicine (EBLM) has been slow to develop.There are many contrasting observations about laboratory medicine, for example (i) there is too much testing vs insufficient testing; (ii) testing is expensive vs laboratories are expected to generate income; and (iii) test results have little impact on outcomes vs test results are crucial to clinical decision making. However, there is little evidence to support any of these observations. Integrating the principles of EBLM into routine practice will help to resolve some of these issues by identifying (a) where laboratory medicine fits into the care pathway; (b) where testing is appropriate; (c) the nature and quality of evidence required to demonstrate the clinical utility of a test; (d) how the test result impacts on clinical actions; (e) where changes in the care pathway will occur; and (f) where benefit/value can be achieved. These answers will help to establish the culture of EBLM in clinical and laboratory practice.     

From Evidence to Best Practice in Laboratory Medicine

Laboratory tests offer value if they provide benefit to patients at acceptable costs. Laboratory testing is one of the most widely used diagnostic interventions supporting medical decisions, yet evidence demonstrating its value and impact on health outcomes is limited. This contributes to wide variations in test utilisation including underdiagnosis, overdiagnosis and misdiagnosis, which may impact the quality and the clinical- and cost-effectiveness of care and patient safety. Therefore implementing evidence into the care of patients is a moral and social imperative to laboratory professionals and all health care staff.This review investigates the reasons research does not get into practice, or only does with a very long delay. Apart from reviewing the common barriers to implementation, it also discusses the drivers of inappropriate test utilisation. By reviewing the theoretical and practical aspects of implementation science, recommendations are made for approaches that are thought to be most effective and that can be adopted to close the gap between evidence and practice, and to facilitate evidence-based laboratory medicine. Passive dissemination of the evidence and educational interventions are insufficient and do not offer sustainable solutions. A multifaceted and individualised implementation strategy, including individually tailored academic detailing, reminder systems, clinical decision support systems, feedback on performance, and participation of doctors and laboratory professionals in quality improvement activities addressing test selection and interpretation and in clinical audits, has greater potential for success. Examples of these initiatives at the laboratory and clinical interface are provided with links to valuable resources.

‘Knowing is not enough; we must apply.Willing is not enough; we must do.’JW von Goethe

     

Genetic screening for the next decade: application of present and new technologies.

Molecular genetic technology is diffusing from the research laboratory to the clinical laboratory, where it has already begun to influence prenatal diagnosis and counseling. In the very near future, this technology will be applied more generally, using population-based screening strategies. Pilot programs are beginning to evaluate the technical feasibility and efficacy of recombinant DNA techniques for newborn screening follow-up. DNA-based population screening is being considered for heterozygous carriers of an autosomal recessive disorder such as cystic fibrosis in order to identify carrier couples at risk of having an affected child. We will review the current DNA methodologies in the context of three genetic disorders: sickle-cell disease, Duchenne muscular dystrophy, and cystic fibrosis. We will then consider the requirements for implementation of these new technologies. We will conclude that implementation will require two key factors: machines and people. Machines are required to automate molecular genetic procedures, which are currently personnel-intensive, so that the expense can be reduced and the procedures made more cost-effective. The people who are required are health professionals knowledgeable in the clinical aspects of the target disorders, as well as in the DNA laboratory testing. These professionals will be able to facilitate sample acquisition and information exchange among the laboratory, the primary health care provider, and the families requesting consultation.     

Anticipated effects of abiotic environmental change on intraspecific social interactions

Social interactions are ubiquitous across the animal kingdom. A variety of ecological and evolutionary processes are dependent on social interactions, such as movement, disease spread, information transmission, and density-dependent reproduction and survival. Social interactions, like any behaviour, are context dependent, varying with environmental conditions. Currently, environments are changing rapidly across multiple dimensions, becoming warmer and more variable, while habitats are increasingly fragmented and contaminated with pollutants. Social interactions are expected to change in response to these stressors and to continue to change into the future. However, a comprehensive understanding of the form and magnitude of the effects of these environmental changes on social interactions is currently lacking. Focusing on four major forms of rapid environmental change currently occurring, we review how these changing environmental gradients are expected to have immediate effects on social interactions such as communication, agonistic behaviours, and group formation, which will thereby induce changes in social organisation including mating systems, dominance hierarchies, and collective behaviour. Our review covers intraspecific variation in social interactions across environments, including studies in both the wild and in laboratory settings, and across a range of taxa. The expected responses of social behaviour to environmental change are diverse, but we identify several general themes. First, very dry, variable, fragmented, or polluted environments are likely to destabilise existing social systems. This occurs as these conditions limit the energy available for complex social interactions and affect dissimilar phenotypes differently. Second, a given environmental change can lead to opposite responses in social behaviour, and the direction of the response often hinges on the natural history of the organism in question. Third, our review highlights the fact that changes in environmental factors are not occurring in isolation: multiple factors are changing simultaneously, which may have antagonistic or synergistic effects, and more work should be done to understand these combined effects. We close by identifying methodological and analytical techniques that might help to study the response of social interactions to changing environments, highlight consistent patterns among taxa, and predict subsequent evolutionary change. We expect that the changes in social interactions that we document here will have consequences for individuals, groups, and for the ecology and evolution of populations, and therefore warrant a central place in the study of animal populations, particularly in an era of rapid environmental change.     

Strategies and testing methods for identifying mutagenic risks

The evolution of testing strategies and methods for identification of mutagenic agents is discussed, beginning with the concern over potential health and population effects of chemical mutagens in the late 1940s that led to the development of regulatory guidelines for mutagenicity testing in the 1970s and 1980s. Efforts to achieve international harmonization of mutagenicity testing guidelines are summarized, and current issues and needs in the field are discussed, including the need for quantitative methods of mutagenic risk assessment, dose-response thresholds, indirect mechanisms of mutagenicity, and the predictivity of mutagenicity assays for carcinogenicity in vivo. Speculation is offered about the future of mutagenicity testing, including possible near-term changes in standard test batteries and the longer-term roles of expression profiling of damage-response genes, in vivo mutagenicity testing methods, and models that better account for differences in metabolism between humans and laboratory model systems.     

Susceptibility testing of anaerobic bacteria: A review of current methods and future prospects

The current NCCLS document, M11 A2, describes two methods for susceptibility testing of anaerobic bacteria. The reference method utilizes an agar dilution procedure, which is labor intensive and not convenient for testing individual patient isolates. The broth microdilution method does not support the growth of 15–40% clinical isolates and demonstrates poor correlation with the reference method for some members of the Bacteroides fragilis group with β-lactam agents and clindamycin. Etest is a new technique that incorporates an antibiotic gradient onto a plastic strip and utilizes agar media. This method is easily performed, permits growth of all anaerobes, and provides quantitative MICs for rapidly growing strains after overnight (20 hr) incubation. This method is convenient and reliable and enables the laboratory to provide the clinician with MIC data for individual patient isolates within a clinically relevant time period.     

Quality Assurance and Standardization in Immunohistochemistry. A Proposal for the Annual Meeting of the Biological Stain Commission,June, 1991

Quality assurance, quality control, proficiency testing, reagent documentation and validation are standard parts of everyday practice in clinical laboratories throughout the United States. Immunohistochemical stains employ reagents and principles in common with immunoenzyme methods utilized in the clinical laboratory. However, immunohistochemistry has not routinely been subjected to similar standardization and quality assurance procedures that manufacturers and pathologists alike have applied to essentially the same techniques in the clinical laboratory environment. The current proposal was invited by the Biological Stain Commission with the charge of incorporating the findings of previous workshops on quality control in immunohistochemistry into a practical design for implementation. The status of quality assurance, quality control and standardization in immunohistochemistry is reviewed and a phased strategy for implementation is proposed.     

Quality Assurance and Standardization in Immunohistochemistry. A Proposal for the Annual Meeting of the Biological Stain Commission, June, 1991

Quality assurance, quality control, proficiency testing, reagent documentation and validation are standard parts of everyday practice in clinical laboratories throughout the United States. Immunohistochemical stains employ reagents and principles in common with immunoenzyme methods utilized in the clinical laboratory. However, immunohistochemistry has not routinely been subjected to similar standardization and quality assurance procedures that manufacturers and pathologists alike have applied to essentially the same techniques in the clinical laboratory environment. The current proposal was invited by the Biological Stain Commission with the charge of incorporating the findings of previous workshops on quality control in immunohistochemistry into a practical design for implementation. The status of quality assurance, quality control and standardization in immunohistochemistry is reviewed and a phased strategy for implementation is proposed.     

Global Proteinuria Guidelines: Are We Nearly There Yet?

Assessment of albumin and/or protein excretion in the urine is a key step in the early detection and appropriate management of chronic kidney disease. The approach to testing for albuminuria/proteinuria in the community is variable and often suboptimal. It is hampered by: variation in laboratory measurement; lack of standard reference materials and testing procedures; variable definitions and units of reporting; conflicting recommendations and practices regarding who to test; and uncertainty over when and how testing is most appropriately done. This review discusses the current status of proteinuria guidelines around the world and the key clinical issues that need to be addressed before a unifying global guideline can be developed.     

Allen Denver Russell Memorial Lecture, 2006. The use of microbicides in infection control: a critical look at safety, testing and applications

Microbial pathogens continue as major threats to health. Indeed, many ongoing societal changes are enhancing our vulnerability and exposure to several frank and opportunistic pathogens. This, together with rampant antimicrobial resistance and reduced prospects for newer drugs and vaccines, is forcing a higher reliance on microbiocides in infection prevention and control. That this reliance may not be well-founded becomes apparent from a closer look at current ways of testing and registering microbiocides, their label claims as well as human and environmental safety of certain widely used microbicidal chemicals. Many methods to test microbiocides for registration are flawed and/or entail test conditions irrelevant to field use. Pathogens listed on product labels may not be among those amenable to interruption through microbiocide use. The wide variations and discrepancies in existing national/regional regulations for registering microbiocides for sale stifle innovation. This is a critical look at the above-mentioned issues with emphasis on chemicals meant for use on environmental surfaces and medical devices. It highlights better ways to test microbiocides and to attain global harmonization of testing and product registration. It also details the known and potential dangers of microbiocide use and what to consider in choosing such formulations for optimal safety and effectiveness. End users are advised to be more critical and prudent in the selection and application of microbicidal chemicals, manufacturers are encouraged to explore infection control products and technologies that are safer in the workplace and for the environment, and regulators are urged to review and update the requirements and procedures for premarket review of microbiocide efficacy data and label claims. Independent investigations are also urgently needed to document the proportion of nosocomial infections that would be amenable to prevention through chemical disinfection of environmental surfaces.     

How to Test Mandatory Text Templates: The European Patient Information Leaflet

The structure of patient information leaflets (PILs) supplied with medicines in the European Union is largely determined by a regulatory template, requiring a fixed sequence of pre-formulated headings and sub-headings. The template has been criticized on various occasions, but it has never been tested with users. This paper proposes an alternative template, informed by templates used in the USA and Australia, and by previous user testing.The main research question is whether the revision better enables users to find relevant information. Besides, the paper proposes a methodology for testing templates. Testing document templates is complex, as they are “empty”. For both the current and the alternative template, we produced a document with bogus text and real headings (reflecting the empty template) and a real-life document with readable text (reflecting the “filled” template). The documents were tested both in Dutch and in English, with 64 British and 64 Dutch users. The test used a set of scenario questions that covers the full range of template (sub)topics; users needed to indicate the text locations where they expected each question to be answered. The revised template improved findability of information; this effect was strongest for the “filled” template with readable text. When participants were shown both filled templates, there was a clear preference for the revised template. A closer analysis of the findability data revealed question-specific effects of topic grouping, topic ordering, subtopic granularity and wording of headings. Most of these favoured the revised template, but our revision led to adverse effects as well, for instance in the new heading Check with your doctor. Language-specific effects showed that the wording of the headings is a delicate task. Generally, we conclude that document template designs can be analyzed in terms of the four parameters grouping, ordering, granularity and wording. Furthermore, they need to be tested on their effects on information findability, with template translations requiring separate testing. The methodology used in this study seems an appropriate one for such tests. More specifically, we find that the new patient information leaflet template proposed here provides better information findability.     

Survey of the Diagnostic Retooling Process in National TB Reference Laboratories, with Special Focus on Rapid Speciation Tests Endorsed by WHO in 2007

Background

Successful integration of new diagnostics in national tuberculosis (TB) control programs, also called ‘retooling’, is highly dependent on operational aspects related to test availability, accessibility and affordability. This survey aimed to find out whether recommendations to use new diagnostics lead to successful retooling in high TB endemic countries, using immunochromatographic tests (ICTs) for TB culture speciation as a case study. ICTs are recommended to accurately confirm the presence of bacteria of the Mycobacterium tuberculosis complex in liquid culture isolates.

Methods and Findings

Questionnaires were sent to national TB reference laboratories (NRLs) in 42 high TB endemic countries to address their access to information on ICT implementation, logistics related to availability, accessibility and affordability of ICTs, and testing algorithms. Results from 16 responding countries indicated that half of the NRLs were aware of the contents of WHO guidance documents on liquid culture and ICT implementation, as well as their eligibility for a negotiated pricing agreement for ICT procurement. No major issues with availability and accessibility of ICTs were raised. When asked about testing algorithms, ICTs were not used as stand-alone or first test for TB culture identification as recommended by WHO.

Conclusions

The low response rate was a limitation of this survey and together with NRLs managers'' unawareness of global guidance, suggests a lack of effective communication between partners of the global laboratory network and NRLs. TB tests could become more affordable to high TB endemic countries, if the possibility to negotiate lower prices for commercial products is communicated to them more successfully. NRLs need additional guidance to identify where available technologies can be most usefully implemented and in what order, taking into account long-term laboratory strategies.