Reviewing the animal literature: how to describe and choose between different types of literature reviews.

Before starting any (animal) research project, review of the existing literature is good practice. From both the scientific and the ethical perspective, high-quality literature reviews are essential. Literature reviews have many potential advantages besides synthesising the evidence for a research question. First, they can show if a proposed study has already been performed, preventing redundant research. Second, when planning new experiments, reviews can inform the experimental design, thereby increasing the reliability, relevance and efficiency of the study. Third, reviews may even answer research questions using already available data. Multiple definitions of the term literature review co-exist. In this paper, we describe the different steps in the review process, and the risks and benefits of using various methodologies in each step. We then suggest common terminology for different review types: narrative reviews, mapping reviews, scoping reviews, rapid reviews, systematic reviews and umbrella reviews. We recommend which review to select, depending on the research question and available resources. We believe that improved understanding of review methods and terminology will prevent ambiguity and increase appropriate interpretation of the conclusions of reviews.

Are the Lives of Animals Well-spent in Laboratory Science Research? A Study of Orthopaedic Animal Studies in Turkey.

BACKGROUND: As in all fields of medicine, animal studies are widely performed in orthopaedics and have increased in number over time. However, it is not clear to what extent these studies provide a basis for future research or advancements in clinical science. Concerns about the reliability and translational ability of animal studies have been reported, and major orthopaedic journals and organizations are encouraging the reduction of unnecessary experiments on animals. QUESTION/PURPOSES: (1) What proportion of animal studies conducted for orthopaedic research in Turkey were never published? And of those that were published, how long did it take to publish? (2) What proportion of those studies were published in journals with an Impact Factor of 2 or more? (3) What proportion of those published papers were never cited or cited only once? (4) What was the contribution to science of an animal euthanized for orthopaedic research in Turkey? METHODS: We reviewed all oral and poster presentations at the Turkish National Congress of Orthopaedics and Traumatology from 2009 to 2017 (retrieved from the archives of Acta Orthopaedica et Traumatologica Turcica), as well as all postgraduate theses in orthopaedics from 1991 to 2017 (retrieved from the archives of the National Thesis Center of the Council of Higher Education) to identify all orthopaedic studies that involved animals. We searched the keywords "animal studies," "experimental studies," and "orthopaedics" in these archives. We defined animal research as orthopaedic studies based on animal models. From this search and using that definition, 252 studies were identified. Of those, 4% (9) were excluded as they were thesis studies with no abstract in the archives. Thus, a total of 243 animal studies performed in Turkey were included for analysis in this retrospective study. The abstracts of these studies were examined to determine the study model (such as bone fracture models, tendon healing models, cartilage models) and number of euthanized animals. Between 1991 and 2017, 9412 vertebrate animals were euthanized for these studies. We searched PubMed, Google Scholar, ResearchGate, and ORCID to determine whether these papers were subsequently published, in which journal, and how long after the initial presentation publication occurred. The Web of Science 2019 database was used to determine the Impact Factor of the journals, the total citation count of each study, and the mean annual citation for each study (citations per year). For purposes of this analysis, we divided journals into those with an Impact Factor of 2 or more, 4 or more, and those with an Impact Factor below 2. The mean annual citation per euthanized animal (citations per animal per year) was calculated to determine the contribution of a euthanized animal to science. RESULTS: A total of 42% (101 of 243) of the animal studies in Turkey were never published. For all published studies, the mean time to publication was 2.2 ± 2.6 years (95% CI 1.7 to 2.6). The proportion of studies published in orthopaedic journals with an Impact Factor of 2 or more was 14% (34 of 243). Among the 142 published papers, 38% (54) were either never cited or were cited only once, and the mean citations per year was 1.1 ± 1.7 (95% CI 0.7 to 1.3). The mean citations per animal/year among the 142 published studies was 0.03 ± 0.04 (95% CI 0.02 to 0.04). CONCLUSION: In the 243 theses and national congress presentations, 9412 animals were euthanized. Based on the low percentage of papers using animals that were euthanized and the very low proportion of studies published in higher-Impact Factor journals or garnering more than a single citation, in aggregate, little seems to have been gained from the loss of animal life. Future studies should try to replicate or refute our results in other countries. CLINICAL RELEVANCE: Orthopaedic researchers should try to reduce their use of unnecessary animal studies, for example, by reporting on the use of the "3Rs" (replacement, reduction, and refinement) in the development of an animal study design, as well as through following universal guidelines so that a study might have a clinical impact. Researchers should not conduct an animal study until they are convinced that the expected results are quite likely to deliver substantial benefit to people or to advance science in a meaningful way; although this seems intuitive, our results suggest that this may not be taking place. Ethics committees in Turkey should consider more detailed questioning before approving animal studies. If our results are replicated elsewhere, then a broader look at how these approvals are conducted should be performed.

New European Union statistics on laboratory animal use - what really counts!

Seven years after the last release, the European Commission has again collated and released data on laboratory animal use. The new report is the first to correspond to the requirements of the new Directive 2010/63/EU. Beside minor problems in reporting, the new reporting format is a major step forward, with additional new categories like severity allowing insight into animal use related questions that goes far beyond the previous reports. An in-depth analysis confirms a slight decrease in animal use from 2015 to 2017, but also compared to the 2005, 2008 and 2011 reports, though the new reporting scheme makes this comparison difficult. Notable success is evident for replacing rabbit pyrogen testing but, in general, the implementation of accepted alternative methods lags behind expec-tations. Beside the roughly 10 million animals per year covered in the report, about 8 million animals were identified that fall under the Directive but are not included in this number. Their omission downplays the impact of REACH on animal use. The report, second to none in its detail internationally, represents an important instrument for benchmarking and strategi-cally focusing activities in the 3Rs.

Experiments done in Black-6 mice: what does it mean?

Low replicability of animal experiments is perceived as a major hurdle in the field of biomedicine. Attempts to enhance the replicability and to reduce the variability in basic research has led to the recommendation to use isogenic mice. The C57BL/6 strain has evolved as a gold standard strain for this purpose. However, C57BL/6 mice are maintained as substrains by multiple vendors. Evidence exists that the subtle differences between these mouse lines have not been systematically investigated and are often ignored. In the present study, we characterized the female mice of two closely related substrains (C57BL/6J and C57BL/6N) from three vendors in Europe (Charles River Laboratories, Envigo, Janvier Labs) in a battery of behavioral tests. Our data show and confirm substantial behavioral differences between the C57BL/6J and C57BL/6N mice. Importantly, the substrain differences were largely affected by the origin of the animals, as a significant effect of vendor or interaction between the substrain and vendor occurred in all tests. This work highlights the importance of adhering to precise international nomenclature in all publications reporting animal experiments. Moreover, the generalization of research findings from a single mouse substrain can be seriously limited due to genetic drift and environmental variables occurring at different vendors. However, heterogenization of samples, by including animals of different substrains, can enhance generalizability. These issues need to be seriously addressed to improve reproducibility, replicability, and the translational potential of the mouse models.

Invited review: Reproducible research from noisy data: Revisiting key statistical principles for the animal sciences.

Reproducible results define the very core of scientific integrity in modern research. Yet, legitimate concerns have been raised about the reproducibility of research findings, with important implications for the advancement of science and for public support. With statistical practice increasingly becoming an essential component of research efforts across the sciences, this review article highlights the compelling role of statistics in ensuring that research findings in the animal sciences are reproducible-in other words, able to withstand close interrogation and independent validation. Statistics set a formal framework and a practical toolbox that, when properly implemented, can recover signal from noisy data. Yet, misconceptions and misuse of statistics are recognized as top contributing factors to the reproducibility crisis. In this article, we revisit foundational statistical concepts relevant to reproducible research in the context of the animal sciences, raise awareness on common statistical misuse undermining it, and outline recommendations for statistical practice. Specifically, we emphasize a keen understanding of the data generation process throughout the research endeavor, from thoughtful experimental design and randomization, through rigorous data analysis and inference, to careful wording in communicating research results to peer scientists and society in general. We provide a detailed discussion of core concepts in experimental design, including data architecture, experimental replication, and subsampling, and elaborate on practical implications for proper elicitation of the scope of reach of research findings. For data analysis, we emphasize proper implementation of mixed models, in terms of both distributional assumptions and specification of fixed and random effects to explicitly recognize multilevel data architecture. This is critical to ensure that experimental error for treatments of interest is properly recognized and inference is correctly calibrated. Inferential misinterpretations associated with use of P-values, both significant and not, are clarified, and problems associated with error inflation due to multiple comparisons and selective reporting are illustrated. Overall, we advocate for a responsible practice of statistics in the animal sciences, with an emphasis on continuing quantitative education and interdisciplinary collaboration between animal scientists and statisticians to maximize reproducibility of research findings.

Laboratory animal science course in Switzerland: participants' points of view and implications for organizers.

Switzerland has implemented a mandatory training in laboratory animal science since 1999; however a comprehensive assessment of its effects has never been undertaken so far. The results from the analysis of participants in the Swiss Federation of European Laboratory Animal Science Associations (FELASA) Category B compulsory courses in laboratory animal science run in 2010, 2012, 2014 and 2016 showed that the participants fully appreciated all elements of the course. The use of live animals during the course was supported and explained by six arguments characterized with cognitive, emotional and forward-looking factors. A large majority considered that the 3R (replacement, reduction and refinement) principles were adequately applied during the course. Responses to an open question offered some ideas for improvements. This overall positive picture, however, revealed divergent answers from different subpopulations in our sample (for example, scientists with more hindsight, scientists trained in biology, or participants from Asian countries).

Survey of environmental enhancement programs for laboratory primates.

Animal welfare regulations in the United States require that nonhuman primate environmental enhancement plans be made in accordance with currently accepted professional standards; however, little information is available for quantifying common practice. Here we report the results of a 2003 survey that was sent to individuals overseeing enrichment programs at a variety of primate research institutions. The surveys requested information on program administration and management, implementation standards, procedures, and constraints pertaining to major categories of environmental enrichment, as well as intervention plans for animals exhibiting behavioral pathologies. Data were obtained on the management of 35,863 primates in 22 facilities. Behavioral scientists performed program oversight at the majority of facilities. Most programs reported recent changes, most commonly due to external site visits, and least commonly resulting from internal review. Most facilities' institutional animal care and use committees (IACUCs) included of individuals with behavioral expertise, and about two-thirds reported that enrichment issues could influence research protocol design. While most primates were reported to be housed socially (73%), social housing for indoor-housed primates appears to have changed little over the past 10 years. Research protocol issues and social incompatibility were commonly cited constraints. Implementation of feeding, manipulanda, and structural enrichment was relatively unconstrained, and contributions to these aspects of behavioral management generally included individuals in a wide variety of positions within a facility. In contrast, enrichment devices were used on a less widespread basis within facilities, and positive reinforcement programs that involved dedicated trainers were rare. We suggest that altering the role of the IACUC would be a productive avenue for increasing the implementation of social housing, and that an emphasis on prevention rather than intervention against behavioral pathology is warranted. The data from this survey may be useful for anticipating future program evaluations, establishing more effective internal evaluations, and assessing program progress and resource allocation.

Biomedical journals: keeping up and reading critically.

By extrapolation from studies of physicians, knowledge and practice of laboratory animal medicine and science are expected to become progressively more outdated the longer practitioners are out of school. Keeping up with current literature and practice is a challenge that necessitates the use of many different sources of continuing education. Both veterinarians and physicians consistently list journals as the most beneficial source of new information. Accordingly, they must select from the veterinary and biomedical literature articles that report original studies and systematic reviews and recognize and respond to valid new knowledge to improve diagnostic and therapeutic approaches and maintain consistent clinical skills. Other objectives include selecting journals for general information and for information relevant or specific to one's field of research. Lastly, candidates for board certification need to read articles from journals that potentially provide the basis for questions on the examination. 'High-impact' journals should be identified, and articles should be reviewed critically. In a survey of recent candidates for laboratory animal medicine board examination, these journals included Contemporary Topics (now JAALAS), Comparative Medicine, ILAR Journal, and Laboratory Animals. Strategies for coping with the challenge of staying current with the literature include wise use of technology, journal clubs, and consultation with colleagues. A laboratory animal practitioner can become a better scientist and clinician by evaluating the research performed by others. Thorough, critical review of biomedical literature is paramount to these goals.

Life history and aging of captive-reared California sea hares (Aplysia californica).

Although the California sea hare, Aplysia californica, is well known from neurobiological studies and is raised in the laboratory for this purpose, various aspects of its life history in the laboratory, such as aging dynamics, are unknown. Therefore we collected life history data on 4 cohorts of eggs from hatchery-reared animals and performed an actuarial analysis of mortality data. Temperature was controlled at 13 to 15 degrees C, the photoperiod was a 14:10-h light:dark cycle, and the seawater O2 concentration, pH, and salinity were held at optimized levels. The feeding protocol for 3 cohorts was unrestricted access to the red macroalga Gracilaria ferox, whereas the remaining cohort was fed standard hatchery rations of G. ferox 4 times per week. Growth was sigmoidal in each cohort and resulted in linear growth rates of 1.25 to 3.62 g/d during the exponential phase; these rates were not influenced by feeding level. Sexual maturity occurred at approximately 160 g, at ages ranging from 144 to 241 d. Egg production was highly variable in the different cohorts. Mean lifespan of cohorts fed ad libitum was approximately 228 d. In contrast, the cohort fed standard rations lived an average of 375 d and showed a lower initial mortality rate, suggesting that calorie restriction on a single-species diet prolongs lifespan in California sea hares.

Report on primate supply for biomedical scientific work in the UK. EUPREN UK Working Party.

A Working Party of the UK group of European Primate Resources Network (EUPREN) considered primate supply for scientific work in the UK. Through a questionnaire, which achieved a very good response, it obtained details of primate use, sources and breeding in the UK and it put forward options to ensure that animal welfare is the best possible whilst ensuring continued supply. The questionnaire showed that contract research laboratories and pharmaceutical companies use about 80% of the 4233 primates used annually at the moment, with the rest accounted for by academic establishments and public sector laboratories. Fifty-four per cent are cynomolgus macaques (Macaca fascicularis), of which nearly 90% are captive-bred outside the European Union (EU), the remainder being bred in the UK. Nearly 90% of cynomolgus macaques are used by only five institutions. Thirty-seven per cent of primates used are marmosets (Callithrix jacchus jacchus), all of which are bred in the UK. Most of the rest are rhesus macaques (Macaca mulatta), about half of which are captive-bred outside the EU, the other half being bred in the UK. Overall primate use has increased from about 3000 per year in 1990 and users predict that requirements for all species except baboons (Papio sp.) will be maintained or increase. Marmoset breeding in the UK is already closely matched to use, and it could be increased reasonably easily if necessary. Some of the existing breeding centres of macaques in the UK would be prepared to consider expanding to supply others, although investment and imported breeding stock would be needed and it is likely that a large investment would be needed to breed a significant fraction of the macaque use in the UK. A further problem is that the users of only about 10% of the cynomolgus macaques said that they could replace this species by rhesus macaques, which are easier to breed in the UK. The questionnaire showed that much of the use of macaques would be transferred to other countries equally remote from the natural source countries of the animals, if constraints on primate use became more severe in the UK. Users felt that it is unlikely that much of the work could be transferred to the natural source countries themselves. A review of the literature revealed a paucity of information on the effects of transport on primate welfare. The importance of obtaining this information before making decisions about alternative means of supply is stressed. Current schemes for the accreditation of primate breeders were reviewed. A list of options is presented for discussion. Users vary so much in their requirements that it is unlikely that one means of supply will be applicable to all. Animal welfare will benefit and supply will be more certain if cooperation between those concerned (preferably through the UK group of EUPREN) is maintained.