How to become a good evidence based Bioveterinary practitioner

What is evidence-based practice

Within the field of bioveterinary science and veterinary practice and care, evidence and knowledge are forever changing. With continuing research consistently making fresh and exciting new break throughs for animal health, it is becoming increasingly difficult to keep up with what is recent and appropriate evidence. Evidence based medicine is a term that was first adopted in the 1990’s and was best defined as “a systemic approach to analyse published research as the basis of clinical decision making.” Although first adopted for use in the human medicine profession, evidence-based practice (EBP) has developed into many methodologies and tools that can be used by a variety of different professions within a diverse amount of clinical practices. EBP is a process where by a practitioner takes it upon themselves to evaluate and appraise the best evidence available to them to enhance their decision making within a clinical setting.  EBP is critical for practitioners to remain competent, relevant and clinically effective. However, acquiring evidence does not always mean scanning through pages of research journals and sifting through systematic reviews. Evidence can also consist of expert opinion, stakeholder characteristics and differing scenarios. Asking a colleague for their opinion on a situation can also be regarded as acquiring evidence. This being said, EBP has come a long way from making decisions solely based on the opinions of those who are deemed as experts or following through with a decision because “that’s the way its always been done”. The purpose of EBP is to encourage those working in the field of bioveterinary science to actively accumulate sources of evidence, evaluate and appraise that evidence, and then make appropriate decisions based on the knowledge and understanding they have gained that will be most beneficial to them and any stakeholders involved. The purpose of EBP is to reduce the risk of human bias, provide opportunities for clinicians and practitioners to become more individualised, define best practices that is supported by the most relevant forms of evidence and to overall improve the work practitioners and the services provided to involved stakeholders by allowing them to create sound and informed decisions using methodologies designed to decipher and appraise what is to be deemed as best evidence.

Acquiring evidence

So where do bioveterinary practitioners acquire evidence from? What constitutes to make something a good and valid source of evidence? The hierarchy of evidence is a tool designed for evidence-based practitioners that guides users in the appraisal of what can be deemed as best evidence for use in decision making. The hierarchy allows users to take a top-down approach to locating best evidence. In theory, this approach assumes that when practitioners are searching for evidence, they start by looking for a well-conducted and recent systematic review to answer the clinical question in hand. If a systematic review is not available, then the user will move down to the next tier of evidence and so on. The lower down the tier a piece of evidence falls, the lesser its ability to constitute towards what is deemed as best evidence. Although the hierarchy of evidence is an effective tool for evaluating physically published sources of evidence, it does not consider other constituents that make up EBP as a whole. Effective clinical decision making should always consider the best available evidence but make no assumptions that one type of evidence has overall superiority. The type of evidence needed by practitioners will vary depending on the clinical question in hand. Evidence needed will depend on the nature and activity of its purpose, so considerations must be made when determining what evidence is best for a particular clinical scenario. Clinical feasibility is the consideration of whether an activity or purpose is practical and practicable within a given context. Research into best available evidence may highlight a practice or activity as being superior to others, but if this activity cannot be performed by the practitioner then it is not the best form of evidence. The appropriateness of an activity goes hand in hand with evaluating the feasibility. Evaluating appropriateness relates to how ethical or suitable an activity relates to the context in which it is being used. Considerations into the benefits and effects is also paramount to determining best evidence. Evidence can be seen to be advantageous if those involved have gained a positive experience regarding the evidence used in a particular situation. A well-rounded evidence-based practitioner should always consider other stakeholders and factors that could have an influence in the type of evidence used in their decision making and clinical thinking.

 

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Tools for evidence-based practice

Methodologies and EBP guidelines are a structural and efficient way to engage in evidence-based practices. One of the most commonly used EBP guidelines is the 5 steps of evidence-based practice.

  • Step 1) Practitioners are encouraged to first convert their need for information, what ever the topic or scenario may be, into a structured and answerable question. Phrasing a question correctly allows for a more organised search for evidence and reducing the chance of any important information being missed from a search.
  • Step 2) Once a clinical question has been constructed, the search for the best available evidence can then begin. The hierarchy of evidence can be used within during this step to assess the validity of evidence that has been searched for.
  • Step 3) Critical appraisal of acquired information and evidence is then necessary to evaluate considerations that may affect the use of the evidence. These include considering the feasibility and appropriateness of the evidence as well as how beneficial it will be to all stake holders involved.
  • Step 4) This step involves integrating critically appraised and appropriate best evidence with existing clinical practices and expertise. Incorporation of past experiences with current best evidence can aid in enhancing current practices. Integration of these two factors should also involve the integration of external stakeholders such as clients and patients. Considering the needs, values and opinions of external stakeholders will help to further evaluate the feasibility and appropriateness of evidence.
  • Step 5) The final step is to evaluate the effectiveness of steps 1-4. Reflecting on the process can help an evidence-based practitioner assess the effects that evidence had on influencing their final decision. Reflection may involve consideration into different application of the process, essentially refining the process for better use in the future.

 

EBP steps

 

 How to implement EBP in the work place

It is recognised that there are barriers than can affect the process of evidence-based practice within the work place. Information available to provide best evidence is usually spread over a variety of different sources and can make it very time consuming for practitioners to find information that can be used as best evidence, as well as practitioners having limited time to analyse information especially in a clinical care environment. Challenges surrounding evaluating the value of information should also be considered, even if EBP methodologies are used. To overcome these barriers, opportunities should be given to practitioners that aid in advancing their evidence-based practice skills. Such opportunities could involve educational work shops into how to correctly use EBP methodologies and how to improve skills such as; searching for appropriate literature in a timely manner, and critical appraisal of information. Providing education on how to correctly and effectively use methodologies with in turn save practitioners time in the future. Online resources such as the evidence based veterinary medicine (EBVM) toolkit, provided by RCVS knowledge, provided free information for veterinary practitioners who wish to learn more about EBP within their field of bioveterinary science. Another resource provided by RCVS knowledge is the online Knowledge Summaries. Knowledge summaries are designed to provide concise conclusions to clinical questions that be easily and quickly accessed by staff. They are summarised resources used to address knowledge needs by providing best available evidence on defined clinical questions. Knowledge summaries are particularly useful for overcoming the barrier on not being able to able EBP in a clinical care setting.

 

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Further reading

EBP is an ever-expanding topic that has many benefits for bioveterinary practitioners if it is implemented correctly. If readers wish to expand their knowledge on the topic and explore into the processes of EBP further, the following further reading resources have been provided:

Giuffrida, M. A. (2017) ‘Practical Application of Evidence-Based Practice.’ Veterinary Clinics of North America: Exotic Animal Practice. (Evidence-Based Clinical Practice in Exotic Animal Medicine), 20(3) pp. 737–748.

La Caze, A. (2016) ‘The hierarchy of evidence and quantum theory.’ Journal of Clinical Epidemiology, 72, April, pp. 4–6.

Paravattil, B., Shabana, S., Rainkie, D. and Wilby, K. J. (2019) ‘Evaluating knowledge, skills, and practice change after an accredited evidence-based medicine course for community pharmacy preceptors.’ Currents in Pharmacy Teaching and Learning, April.

Saunders, H., Vehviläinen-Julkunen, K. and Stevens, K. R. (2016) ‘Effectiveness of an education intervention to strengthen nurses’ readiness for evidence-based practice: A single-blind randomized controlled study.’ Applied Nursing Research, 31, August, pp. 175–185.

Sullivan, K. J., Wayne, C., Patey, A. M. and Nasr, A. (2017) ‘Barriers and facilitators to the implementation of evidence-based practice by pediatric surgeons.’ Journal of Pediatric Surgery, 52(10) pp. 1666–1673.

Using in silico methods as alternatives to using protected animals within toxicology research

Introduction

Exposure to potentially harmful chemicals is always a possibility throughout varying areas of everyday life. They can be found as pesticides sprayed onto food, compounds for a new potential pharmaceutical or even as a new formula for a popular cosmetic product. Toxicology can be defined as the field of science dedicated to the investigation of the harmful effects caused by chemical substances on living organisms (Soto, 2018). Measuring the adverse effects that a specific chemical may have on a body, whether that be animal or human, is paramount for assessing the safety of a product. Measuring the dosage of chemicals is also extremely important when assessing safety. When assessing safety of chemicals, it is also imperative to evaluate how the body reacts to different dosages. The dose response curve refers to the relationship between the size of the dose given and its affect the body. In the right dosages, these affects could be very positive, but too much and the chemical could then become toxic and extremely harmful (Tsatsakis et al., 2018). Within the field of toxicology in vivo experiments involving animals have been widely used. Shukla et al., (2010) discusses how animal models have been used to investigate specific toxicological end points such as; immunotoxicity, genotoxicity and carcinogenicity. Although the outcomes of testing on animal models have provided useful information, they aren’t always consistently predictive of human biology, and they come at a large expensive. Ethical debates arise around the subject of in vivo experimentation. Where possible, the replacement of animals within research is essential. In silico is a method of testing which involves computer analysis and simulation (Raunio, 2011). Raies and Bajic, (2016) describe how in silico testing is widely used throughout toxicology to estimate the toxicity of chemicals. The aim of in silico testing is to complement existing toxicology tests by; minimising late-stage failures in drug development, prioritise chemicals and guide toxicity tests. This report will discuss how protected animals are used within research, the use of in silico technology methods as an alternative to using protected animals within toxicology research and discussions will be made into the ethics surrounding the use of research animals and why is it important to focus on the development of alternative testing methods.

The use of protected animals within research

The use of animals for scientific research has been a heavily debated subject for years. Animals have been considered good model systems for humans and human disease, therefore in vivo testing methods have been widely used within toxicology and other related research. Human obligation to consider animal ethics within research has been paramount for the development of ethical and legal guidelines that must be considered and followed by researchers (Cheluvappa et al., 2017). In 1959, William Russell and Rex Burch introduced the concept of the “3 Rs”; replacement, reduction and refinement (Ferdowsian and Beck, 2011). Miziara et al., (2012) goes on to explain how the 3 Rs were a tool designed to rationalise the use of animals in research and humanise the care of animals. Russel and Burch further argued that alternative methods such as in vitro testing and computer modelling should always be considered to replace and reduce the number of animals used where possible. Where animals must be used, techniques should be refined so as to reduce any pain or suffering caused to research animals. Such techniques included care with analgesia and postoperative antisepsis treatments. Within the UK, legislation was introduced with the purpose of protecting laboratory animals as much as possible. This legislation is known as the Animals (Scientific Procedures) Act, 1986 (ASPA). In accordance with ASPA, any living vertebrate (other than man) and any living cephalopod is deemed as “protected” once it has reached two thirds of its gestation or incubation period or is able to feed independently (‘Animals (Scientific Prodecures) Act 1986, Section 1.,’ 2019). The principles of the 3Rs and the protection of laboratory animals should also extend to the breeding, accommodation and care of any protected animals used for research (Guidance on the Operation of the Animals (Scientific Procedures) Act 1986., 2014).

There are many different factors surrounding the husbandry of laboratory animals. If husbandry is not correct, dependent on the species, it can have detrimental effects on the animal’s health and welfare. Animals can sense the seasons even when housed in windowless rooms. Reports have indicated that seasonal variation can cause immunosuppression induced by chronic stress. Other research has also shown that mice living with a 12:12-hour photoperiod were at higher risk of death from peritonitis in the summer or autumn compared with other season, highlighting the importance of understanding seasonal variation in regards to animal health (Nevalainen, 2014). A similar study conducted by (Hawkins and Golledge, 2018) describes how it is common practice to carry out scientific procedures on rats and mice during the day time. Because rodents are nocturnal, this tends to interrupt their inactive period. Testing on rodents during their inactive periods can cause cognitive defects and induced stress. Species behaviour and species enrichment are further examples as to why species specific husbandry is so significant. Alho et al., (2016) explains how a lack of feline enrichment for indoor housed cats can lead to disorders such as; anxiety, stress, obesity and feline idiopathic cystitis. As laboratory cats must be housed in doors due to health, hygiene and other relating factors that could affect research, it is paramount for laboratory workers to ensure that there is always available enrichment to enable laboratory cats to be able to exhibit their natural behaviours.

The law states that a procedure that is undertaken on a protected animal becomes regulated if that procedure has the potential of causing pain, suffering, distress or lasting harm equivalent to, or higher than, pain induced by the insertion of a hypodermic needle according to good veterinary practice. Such procedures must only be carried out if they have a scientific or educational purpose (Guidance on the Operation of the Animals (Scientific Procedures) Act 1986., 2014). The breeding of genetically modified animals also falls under the umbrella of regulated procedure. A home office report from 2017 indicates that approximately 0.28m (15%) genetically altered animals had experienced some form of potentially harmful effect due to the genetic alteration procedure (Great Britain and Home Office, 2018). Li et al., (2015) further explains how pig genomes are manipulated to increase the compatibility with human biology. This raises the question as to how reliable animal models can be if it is needed to manipulate their genome to make them more representative of human biology. Alternative methods may be a more feasible and ethical alternative. Further regulated procedures within toxicology include the assessment of multiple different toxicological endpoints including; acute oral toxicity and skin sensitisation. Oral toxicity studies involve the determination of the median lethal dose of a substance, otherwise known as LD50 testing (Zakari and Kubmarawa, 2016). This test determines the dose responsible for killing 50% of the participant animals within 24 hours. LD50 testing is a high debated method of endpoint testing. It requires a large number of animals in order to gain enough statistical data and is deemed as cruel and an inappropriate use of animals due to its questionable translatability to humans (Buesen et al., 2016). Skins sensitisation is another major toxicology endpoint that has to be assessed particularly in the development of dermatology products. Guinea pigs are often used as test models for skin sensitisation, but again the translatability of the results for human skin has been argued. Skin sensitisation also involve the application of an adjuvant that causes pain and distress to treated animals, highlighting the desirability of a replacement method (Basketter and Kimber, 2018).

 

The use of in silico methods as an alternative to using protected animals within research

Before a research project can begin it is required that a comprehensive project evaluation is conducted. This evaluation should consider ethical deliberations surrounding the use of animals for said project. The purpose of this ethical evaluation is to ensure that principles of the 3R’s (replacement, reduction and refinement) have been implemented within the study design (Guidance on the Operation of the Animals (Scientific Procedures) Act 1986., 2014). The 3Rs provide a useful strategy to rationally consider how animals are used in a project, without compromising the quality of the research being undertaken (Kandárová and Letašiová, 2011). Replacement is defined as a substitution of protected animals by non-sentient material or methods. Replacement does not necessarily mean that no animals are used at all. Alternative methods can be used to partially replace animals for example; replacing the use of animals for certain kind of test substances or for a particular range of toxicological hazard. Alternative methods implement the principles of reduction by decreasing the number of animals used within a project. Finally, refinement is defined as any decrease in the incidence or severity of procedures conducted for research. For example, in silico analysis may be able to predict the severity or lethality of a substance before it is tested on an animal. This allows researchers to refine their testing strategies, evidently reducing the amount of pain or suffering experienced by the animal (Mushtaq et al., 2018). In silico toxicology (IST) methods use computer resources to analyse, simulate, visualise or predict toxicity of chemicals and substances  (Myatt et al., 2018). IST methodologies aim to complement existing toxicology tests to potentially minimise the need for animal testing and improve toxicity prediction and safety assessment (Raies and Bajic, 2016). IST is a very fast expanding example of alternatives methods to animal testing used within the field of toxicology.

Toxicology testing is a necessary step in the process of all new product developments containing potentially hazardous chemicals or substances. For example; pharmaceutical drug design, food production, pesticide development and the making of cosmetic products. The developmental stage requires an analysis of different data sets to predict the toxicity of chemicals for potential new products. Data sets tend to fall into two main types of classification; single-label and multi-label classification (Yang et al., 2018). Single-label classification is used when each sample in a data set falls into only one class, or if the data set is binary and can be split into two classes. For example; carcinogenic or noncarcinogenic compounds. If a data set can be split into more than two classes, it then becomes a multi-class classification. An example of this would be classifying a compound on the degree of skin sensitisation (e.g. low, moderate, high) (Raies and Bajic, 2018). A study conducted by Zhang et al., (2015) describes the use of in silico methods used to predict chemical toxicity of pesticides within avian species. More than 663 diverse chemicals were assessed, and all chemicals were classified into three categories; highly toxic, slightly toxic and non-toxic. Although this study still used protected animals to obtain data, the prediction models significantly reduced the number of animals needed within the project. The high accuracy of the prediction models that were developed meant that pain and suffering was kept to a minimum as the highly toxic substances had already been predicted. The data that was recorded for the study can be used again for further avian toxicology research, reducing the number of animals needed in the future.

Although the development of in silico methods for toxicology testing is a seemingly popular and expanding field of research, there are some areas of limitation. There is a continuing need to ensure that new methods which are developed for use within toxicology are validated correctly. Clarity is needed about what to validate new methods against; is it already available test methods or accredited knowledge of adverse health effects of chemicals. A limitation to validating new test methods is the cost of potential time delays for the industry. For example, development of new in silico methods in parallel with traditional methods can add an extensive amount of time to a research project (Prior et al., 2019). Further limitations can include the need for whole organism interactions. Planchart et al., (2016) explains how zebra fish are still commonly used within toxicology as fish and humans share similar developmental and physiological responses to chemical exposures. Furthermore, although alternative models such as in silico are excellent for providing insight into reactions within specific areas of the body, there needs to be further development into in silico predictions regarding adverse effects to the whole organism. Although limitations are clearly apparent, in silico toxicology still has many benefits. Toxicology accounts for around 50% of failures in preclinical drug development, indicating the need for reliable prediction strategies to minimise these failures (Ford, 2016). A study conducted by Tambunan et al., (2019) explains how including in silico methods into their study design greatly reduced resource requirements in comparison to conventional methods. This is beneficial to the project as it saves time and expenses but also works in accordance with the 3Rs. Additionally, a differing study led by Passini et al., (2017) describes how in silico drug trials institute influential methodology to predict clinical risk of arrhythmias in human cardiotoxicity. The benefits shown by the methodology allow it to be integrated into existing cardiotoxicity assessment schemes, thus contributing to the reduction of animals used for experimentation.

Conclusion

Animal contributions to the field of toxicology research have provided extremely advantageous information and knowledge and have shaped the development of many life-saving drugs, safe and effective pesticides and many other advances in toxicology safety assessments. The use of in silico technology as an alternative to using animal models has been demonstrated as extremely effective and beneficial for both the research it has been involved with and to minimising the number of protected animals used within toxicology research, working in accordance with ASPA and respecting the 3Rs. Although in silico testing methods still must be used alongside animal models, the toxic predictability allows researchers to rapidly refine their studies. Predicting the toxicity of chemicals before they are tested on protected animals means that animals will experience less pain and suffering as researchers can already gauge how the animal may react to a certain chemical. Further research needs to be carried out to development more in silico models and testing methods to further increase the benefits in silico testing in the future, but needless to say in silico is an extremely positive alternative to using protected animals in research will continue to improve welfare and minimise the use of protected animals with research.

 

References              

Alho, A. M., Pontes, J. and Pomba, C. (2016) ‘Guardians’ Knowledge and Husbandry Practices of Feline Environmental Enrichment.’ Journal of Applied Animal Welfare Science, 19(2) pp. 115–125.

‘Animals (Scientific Prodecures) Act 1986, Section 1.’ (2019). Government Legislation.

Basketter, D. A. and Kimber, I. (2018) ‘Are skin sensitisation test methods relevant for proteins?’ Regulatory Toxicology and Pharmacology, 99, November, pp. 244–248.

Buesen, R., Oberholz, U., Sauer, U. G. and Landsiedel, R. (2016) ‘Acute oral toxicity testing: scientific evidence and practicability should govern Three Rs activities.’ Altern Lab Anim, 44(4) pp. 391–398.

Cheluvappa, R., Scowen, P. and Eri, R. (2017) ‘Ethics of animal research in human disease remediation, its institutional teaching; and alternatives to animal experimentation.’ Pharmacology Research & Perspectives, 5(4) p. e00332.

Ferdowsian, H. R. and Beck, N. (2011) ‘Ethical and Scientific Considerations Regarding Animal Testing and Research.’ PLOS ONE, 6(9) p. e24059.

Ford, K. A. (2016) ‘Refinement, Reduction, and Replacement of Animal Toxicity Tests by Computational Methods.’ ILAR Journal, 57(2) pp. 226–233.

Great Britain and Home Office (2018) Annual Statistics of Scientific Procedures on Living Animals Great Britain 2017.

Guidance on the Operation of the Animals (Scientific Procedures) Act 1986. (2014).

Hawkins, P. and Golledge, H. D. R. (2018) ‘The 9 to 5 Rodent − Time for Change? Scientific and animal welfare implications of circadian and light effects on laboratory mice and rats.’ Journal of Neuroscience Methods. (Measuring Behaviour 2016), 300, April, pp. 20–25.

Kandárová, H. and Letašiová, S. (2011) ‘Alternative methods in toxicology: pre-validated and validated methods.’ Interdisciplinary Toxicology, 4(3) pp. 107–113.

Li, P., Estrada, J. L., Burlak, C., Montgomery, J., Butler, J. R., Santos, R. M., Wang, Z.-Y., Paris, L. L., Blankenship, R. L., Downey, S. M., Tector, M. and Tector, A. J. (2015) ‘Efficient generation of genetically distinct pigs in a single pregnancy using multiplexed single-guide RNA and carbohydrate selection.’ Xenotransplantation, 22(1) pp. 20–31.

Miziara, I. D., de Matos Magalhães, A. T., Santos, M. d’Aparecida, Gomes, É. F. and de Oliveira, R. A. (2012) ‘Research ethics in animal models.’ Brazilian Journal of Otorhinolaryngology, 78(2) pp. 128–131.

Mushtaq, S., Daş, Y. K. and Aksoy, A. (2018) ‘Alternative Methods to Animal Experiments.’ Turkiye Klinikleri Journal of Laboratory Animals.

Myatt, G. J., Ahlberg, E., Akahori, Y., Allen, D., Amberg, A., Anger, L. T., Aptula, A., Auerbach, S., Beilke, L., Bellion, P., Benigni, R., Bercu, J., Booth, E. D., Bower, D., Brigo, A., Burden, N., Cammerer, Z., Cronin, M. T. D., Cross, K. P., Custer, L., Dettwiler, M., Dobo, K., Ford, K. A., Fortin, M. C., Gad-McDonald, S. E., Gellatly, N., Gervais, V., Glover, K. P., Glowienke, S., Van Gompel, J., Gutsell, S., Hardy, B., Harvey, J. S., Hillegass, J., Honma, M., Hsieh, J.-H., Hsu, C.-W., Hughes, K., Johnson, C., Jolly, R., Jones, D., Kemper, R., Kenyon, M. O., Kim, M. T., Kruhlak, N. L., Kulkarni, S. A., Kümmerer, K., Leavitt, P., Majer, B., Masten, S., Miller, S., Moser, J., Mumtaz, M., Muster, W., Neilson, L., Oprea, T. I., Patlewicz, G., Paulino, A., Lo Piparo, E., Powley, M., Quigley, D. P., Reddy, M. V., Richarz, A.-N., Ruiz, P., Schilter, B., Serafimova, R., Simpson, W., Stavitskaya, L., Stidl, R., Suarez-Rodriguez, D., Szabo, D. T., Teasdale, A., Trejo-Martin, A., Valentin, J.-P., Vuorinen, A., Wall, B. A., Watts, P., White, A. T., Wichard, J., Witt, K. L., Woolley, A., Woolley, D., Zwickl, C. and Hasselgren, C. (2018) ‘In silico toxicology protocols.’ Regulatory Toxicology and Pharmacology, 96, July, pp. 1–17.

Nevalainen, T. (2014) ‘Animal Husbandry and Experimental Design.’ ILAR Journal, 55(3) pp. 392–398.

Passini, E., Britton, O. J., Lu, H. R., Rohrbacher, J., Hermans, A. N., Gallacher, D. J., Greig, R. J. H., Bueno-Orovio, A. and Rodriguez, B. (2017) ‘Human In Silico Drug Trials Demonstrate Higher Accuracy than Animal Models in Predicting Clinical Pro-Arrhythmic Cardiotoxicity.’ Frontiers in Physiology, 8.

Planchart, A., Mattingly, C. J., Allen, D., Ceger, P., Casey, W., Hinton, D., Kanungo, J., Kullman, S. W., Tal, T., Bondesson, M., Burgess, S. M., Sullivan, C., Kim, C., Behl, M., Padilla, S., Reif, D. M., Tanguay, R. L. and Hamm, J. (2016) ‘Advancing Toxicology Research Using In Vivo High Throughput Toxicology with Small Fish Models.’ ALTEX, 33(4) pp. 435–452.

Prior, H., Casey, W., Kimber, I., Whelan, M. and Sewell, F. (2019) ‘Reflections on the progress towards non-animal methods for acute toxicity testing of chemicals.’ Regulatory Toxicology and Pharmacology, 102, March, pp. 30–33.

Raies, A. B. and Bajic, V. B. (2016) ‘In silico toxicology: computational methods for the prediction of chemical toxicity.’ Wiley Interdisciplinary Reviews. Computational Molecular Science, 6(2) pp. 147–172.

Raies, A. B. and Bajic, V. B. (2018) ‘In silico toxicology: comprehensive benchmarking of multi-label classification methods applied to chemical toxicity data.’ Wiley Interdisciplinary Reviews: Computational Molecular Science, 8(3) p. e1352.

Raunio, H. (2011) ‘In Silico Toxicology – Non-Testing Methods.’ Frontiers in Pharmacology, 2, June.

Shukla, S. J., Huang, R., Austin, C. P. and Xia, M. (2010) ‘The Future of Toxicity Testing: A Focus on In Vitro Methods Using a Quantitative High Throughput Screening Platform.’ Drug discovery today, 15(23–24) pp. 997–1007.

Soto, M. (2018) ‘Foreword.’ In Costa, P. M. (ed.) The Handbook of Histopathological Practices in Aquatic Environments. Academic Press, pp. vii–viii.

Tambunan, U. S. F., Parikesit, A. A., Ghifari, A. S. and Satriyanto, C. P. (2019) ‘In silico identification of 2-oxo-1,3-thiazolidine derivatives as novel inhibitor candidate of class II histone deacetylase (HDAC) in cervical cancer treatment.’ Arabian Journal of Chemistry, 12(2) pp. 272–288.

Tsatsakis, A. M., Vassilopoulou, L., Kovatsi, L., Tsitsimpikou, C., Karamanou, M., Leon, G., Liesivuori, J., Hayes, A. W. and Spandidos, D. A. (2018) ‘The dose response principle from philosophy to modern toxicology: The impact of ancient philosophy and medicine in modern toxicology science.’ Toxicology Reports, 5, January, pp. 1107–1113.

Yang, H., Sun, L., Li, W., Liu, G. and Tang, Y. (2018) ‘In Silico Prediction of Chemical Toxicity for Drug Design Using Machine Learning Methods and Structural Alerts.’ Frontiers in Chemistry, 6, February.

Zakari, A. and Kubmarawa, D. (2016) ‘Acute Toxicity (LD50) Studies Using Swiss Albino Mice and Brine Shrimp Lethality (LC50 and LC90) Determination of the Ethanol Extract of Stem Bark of Echinaceae angustifolia DC.’ Natural Products Chemistry & Research, 04(06).

Zhang, C., Cheng, F., Sun, L., Zhuang, S., Li, W., Liu, G., Lee, P. W. and Tang, Y. (2015) ‘In silico prediction of chemical toxicity on avian species using chemical category approaches.’ Chemosphere, 122, March, pp. 280–287.