Vertebrate Research Articles

Tuta absoluta-Specific DNA in Domestic and Synanthropic Vertebrate Insectivore Feces.

The ecology of greenhouse pests generally involves parasitoid or predatory insects. However, we investigated whether the leaf miner Tuta absoluta (Meyrick, 1917) (Lepidoptera: Gelechiidae) is part of the diet of domestic and synanthropic vertebrate animals, such as birds, reptiles, and mammals, and that take part in an ecosystem that contains a high density of tomato greenhouses. Feces from domesticated partridges, common quails, and chickens, as well as from wild lizards were collected within tomato greenhouses, and fecal pellets from bats, swallows, common swifts, and house martins living in the vicinity of tomato greenhouses were collected outside. The efficiencies of three different DNA extraction methods were compared on bird, reptile, and mammal stool samples, and the DNA extracts were analyzed using probe real-time PCR for the presence of T. absoluta DNA. The results showed that bats fed on the pest, which was also part of the diet of several bird species: partridges and common quails kept within tomato greenhouses and swallows and common swifts living outside but in the vicinity of tomato greenhouses. In addition, fecal samples of three lizard species living near tomato crops also tested positive for T. absoluta DNA. The results suggest that aerial foraging bats and insectivorous birds are part of ecosystems that involve leaf miners and tomato greenhouses.

The (Re-)Emergence and Spread of Viral Zoonotic Disease: A Perfect Storm of Human Ingenuity and Stupidity.

Diseases that are transmitted from vertebrate animals to humans are referred to as zoonotic diseases. Although microbial agents such as bacteria and parasites are linked to zoonotic events, viruses account for a high percentage of zoonotic diseases that have emerged. Worryingly, the 21st century has seen a drastic increase in the emergence and re-emergence of viral zoonotic disease. Even though humans and animals have coexisted for millennia, anthropogenic factors have severely increased interactions between the two populations, thereby increasing the risk of disease spill-over. While drivers such as climate shifts, land exploitation and wildlife trade can directly affect the (re-)emergence of viral zoonotic disease, globalisation, geopolitics and social perceptions can directly facilitate the spread of these (re-)emerging diseases. This opinion paper discusses the "intelligent" nature of viruses and their exploitation of the anthropogenic factors driving the (re-)emergence and spread of viral zoonotic disease in a modernised and connected world.

Bridging the gap between decomposition theory and forensic research on postmortem interval.

Knowledge of the decomposition of vertebrate animals has advanced considerably in recent years and revealed complex interactions among biological and environmental factors that affect rates of decay. Yet this complexity remains to be fully incorporated into research or models of the postmortem interval (PMI). We suggest there is both opportunity and a need to use recent advances in decomposition theory to guide forensic research and its applications to understanding the PMI. Here we synthesise knowledge of the biological and environmental factors driving variation in decomposition and the acknowledged limitations among current models of the PMI. To guide improvement in this area, we introduce a conceptual framework that highlights the multiple interdependencies affecting decay rates throughout the decomposition process. Our framework reinforces the need for a multidisciplinary approach to PMI research, and calls for an adaptive research cycle that aims to reduce uncertainty in PMI estimates via experimentation, modelling, and validation.

The Use of Zebrafish and in Vivo PAL as a Novel Discovery Platform for Psychoactive Agents

Our research is utilizing the zebrafish model to study molecular processes and animal models of human diseases. These studies discussed here start to assess how different bath salts and methamphetaminecause death upon overdose by addressing the molecules that may bind to the illicit drugs. We have accomplished this by developing an in vivo photo-affinity labelling (PAL) system and studying the drugs in our system. The zebrafish has been lauded as a unique and powerful tool to examine development in a vertebrate animal and address molecular aspects of human disease for decades. We will examine recent published data from our laboratory utilizing the PAL system we developed.

Effect of second warm ischemia elimination on kidney graft function: an experiment and clinical study

Objective: to evaluate the effectiveness of a new device for second warm ischemia (SWI) elimination in kidney transplantation (KT).Materials and methods. The study included clinical and experimental stages. The clinical stage included 63 patients out of 219 who underwent KT at Botkin Moscow City Clinical Hospital between July 2018 and August 2022. The inclusion criteria were kidneys from donation after brain death (DBD) donors with expanded criteria or kidneys from donation after circulatory death (DCD) donors, and an SWI time greater than 45 minutes. The first group consisted of 24 recipients operated on using the new SWI elimination device. The second retrospective control group consisted of 39 patients where sterile ice bags were used at the implantation stage. The groups had no statistically significant differences in the main recipient and donor characteristics, as well as in perioperative parameters. Also, from November 2021 to April 2022, 23 kidney autotransplantation experiments in female Landrace pigs were performed. The animals were cared for in accordance with the European Convention for the Protection of Vertebrate Animals used for Experimental and other Scientific Purposes (Strasbourg, 18 March 1986). Efficiency of different SWI elimination techniques was compared on two experimental models: standard donor (group 1, n = 12) and asystolic donor (group 2, n = 11).Results. In the clinical trial group, mean graft temperature (tm) before reperfusion was statistically significantly lower in group 1 using the special SWI elimination device: 6.4 ± 1.7 °C (95% CI 3.2–8.5) versus 22.1 ± 2.3 °C (18.1–24.6), р < 0.001. The risk of delayed graft function (DGF) was 3.86 times higher (95% CI 1.11–13.43) with the standard SWI elimination technique. In the experimental group, in the subgroups using the new device (n = 12), graft tm before reperfusion was 5.1 ± 0.4 °C (95% CI 4.5–5.8), whereas in the ice bag subgroups (n = 11), tm was 29.3 ± 1.3 °C (95% CI 27.7–30.8), which was significantly higher (p < 0.001). The overall 1-week survival of the experimental animals was significantly higher in the SWI elimination device subgroup (logrank p = 0.036).Conclusion. The developed device is effective in eliminating SWI of renal graft.

In light of breathing: environmental light is an important modulator of breathing with clinical implications.

In vertebrate animals, the automatic, rhythmic pattern of breathing is a highly regulated process that can be modulated by various behavioral and physiological factors such as metabolism, sleep-wake state, activity level, and endocrine signaling. Environmental light influences many of these modulating factors both indirectly by organizing daily and seasonal rhythms of behavior and directly through acute changes in neural signaling. While several observations from rodent and human studies suggest that environmental light affects breathing, few have systematically evaluated the underlying mechanisms and clinical relevance of environmental light on the regulation of respiratory behavior. Here, we provide new evidence and discuss the potential neurobiological mechanisms by which light modulates breathing. We conclude that environmental light should be considered, from bench to bedside, as a clinically relevant modulator of respiratory health and disease.

Control by the brain of vitamin A homeostasis

Vitamin A is a micronutrient essential for vertebrate animals maintained in homeostatic balance in the body; however, little is known about the control of this balance. This study investigated whether the hypothalamus, a key integrative brain region, regulates vitamin A levels in the liver and circulation. Vitamin A in the form of retinol or retinoic acid was stereotactically injected into the 3rd ventricle of the rat brain. Alternatively, retinoids in the mouse hypothalamus were altered through retinol-binding protein 4 (Rbp4) gene knockdown. This led to rapid change in the liver proteins controlling vitamin A homeostasis as well as vitamin A itself in liver and the circulation. Prolonged disruption of Rbp4 in the region of the arcuate nucleus of the mouse hypothalamus altered retinol levels in the liver. This supports the concept that the brain may sense retinoids and influence whole-body vitamin A homeostasis with a possible "vitaminostatic" role.

Applying data science advances in disease surveillance and control

Applying data science advances in disease surveillance and control Dr. David S. Ebert and Dr. Aaron Wendelboe explain how a cohesive, multidisciplinary, and multi-tiered approach can support a more predictive model in disease surveillance and control. Public health disease surveillance is being conducted in countless settings, including healthcare, vertebrate and invertebrate animals, wastewater, air quality, transportation, and commercial activities, but, attaining the goal of early disease detection has been somewhat elusive. For instance, one of the few key shortcoming of public health preparedness efforts is the insufficient collaboration between multidisciplinary experts, such as data scientists, computer engineers, anthropologists, social scientists, and systems engineers. To address these gaps in knowledge and preparedness, we are responding in a multi-tiered approach with a One Health perspective that will be economically feasible and sustainable. The authors have also engaged a broad set of stakeholders, created broad multidisciplinary teams, are combining relevant data sources in innovative ways that will serve as early indicators, are using advanced technologies for early diagnosis, and advancing analytic methods to maintain high specificity for true event identification.

Origins of US Government Principles for the Utilization and Care of Vertebrate Animals Used in Testing, Research, and Training.

The US Government Principles for the use of animals in research are a landmark statement of ethical values and guidance for the biomedical research community. However, when The Principles were introduced, a context was not provided for their source or foundation. The US Government Principles were formulated with input from the Council of Europe, World Health Organization, and US Interagency Research Animal Committee. The Principles continue to provide an ethical foundation for the biomedical research community.

Study of analgesic and myotropic spasmolytic activity of alkylcarb

The work is devoted to the search for new biologically active substances in a series of alkylamides of dihydroquinoline-3-carboxylic acid. Analgesic properties of 30 newly synthesized substances were studied, which made it possible to identify the leader compound (provisional name alkylcarb) and recommend its effectiveness for further research as an antispasmodic agent for pain relief. It has been established experimentally, that the substance alkylcarb exhibits concentration-dependent vasodilatory properties in vitro on segments of the thoracic aorta of rats. The data, obtained in the work, justify the prospect of using the leader substance as a new analgesic and antispasmodic drug. The research expands and deepens knowledge about the pharmacological properties of alkylamide derivatives of dihydroquinoline-3-carboxylic acid.
 The aim of the work was to conduct screening studies to establish the analgesic activity of a newly synthesized series of chemical compounds among alkylamides of dihydroquinoline-3-carboxylic acid and to study the myotropic spasmolytic activity of the leader substance.
 Materials and methods. Analgesic properties of alkylamides of dihydroquinoline-3-carboxylic acid were studied on outbred white mice in the "acetic acid convulsions" test. During the experiment, the animals were treated in accordance with the International Principles of the European Convention for the Protection of Vertebrate Animals Used for Experiments and Other Scientific Purposes (Strasbourg, March 18, 1986). The studied substances were administered intragastrically to experimental animals at a dose of 10 mg/kg in the form of a finely dispersed aqueous suspension, stabilized with Tween-80. Voltaren at a dose of 8 mg/kg, recommended for preclinical studies, and analgin at a dose of 50 mg/kg were chosen as reference drugs.
 The studies of the contractile activity of smooth muscle vessels were carried out on segments of the thoracic aorta of rats of both sexes weighing 180-200 g. The studies of dilator reactions were carried out against the background of preliminary contraction with phenylephrine at a concentration of 10-6 mol/l. The antispasmodic efficiency of the new compound was determined in comparison with the classic antispasmodic drotaverine.
 The statistical processing of the results was carried out using the package of statistical analysis of electronic spreadsheets Exel, with the help of the program "Statgraphics Plus v. 3.0.” and the standard package of statistical programs "Statistica, V. 6.0". We used the Student's test, a non-parametric analog of univariate variance analysis - the Kruskal-Wallis test, and the Mann-Whitney test. Differences were considered statistically significant at p<0.05.
 Results. The study of the analgesic activity of substances АO1-АO30 in the "acetic acid convulsion" test in mice showed that a compound АO26 (provisional name alkylcarb) has the most pronounced analgesic activity when administered intragastrically. In the "acetic acid convulsion" test, alkylcarb (10 mg/kg, per os) probably reduces the number of convulsions, caused by acetic acid. The level of activity of this compound is comparable to the activity of diclofenac (8 mg/kg, per os) and exceeds analgin (50 mg/kg, per os). The substance alkylcarb relieved vasospasm at the level of the comparison drug drotaverine in in vitro experiments on a model of an isolated fragment of the thoracic aorta of rats against the background of previous constriction with phenylephrine.
 Conclusion. Today, the search for new highly effective non-opioid analgesics is an urgent problem of modern pharmacology, since painkillers used in clinical practice do not meet the requirements of efficiency and safety. In this regard, in recent years, scientists of the National Pharmaceutical University have been intensively searching for new, highly effective substances with antinociceptive, anti-inflammatory and antipyretic effects among alkylamides of dihydroquinoline-3-carboxylic acid.

Identification of the Tembusu Virus in Mosquitoes in Northern Thailand.

Among emerging zoonotic pathogens, mosquito-borne viruses (MBVs) circulate between vertebrate animals and mosquitoes and represent a serious threat to humans via spillover from enzootic cycles to the human community. Active surveillance of MBVs in their vectors is therefore essential to better understand and prevent spillover and emergence, especially at the human-animal interface. In this study, we assessed the presence of MBVs using molecular and phylogenetic methods in mosquitoes collected along an ecological gradient ranging from rural urbanized areas to highland forest areas in northern Thailand. We have detected the presence of insect specific flaviviruses in our samples, and the presence of the emerging zoonotic Tembusu virus (TMUV). Reported for the first time in 1955 in Malaysia, TMUV remained for a long time in the shadow of other flaviviruses such as dengue virus or the Japanese encephalitis virus. In this study, we identified two new TMUV strains belonging to cluster 3, which seems to be endemic in rural areas of Thailand and highlighted the genetic specificities of this Thai cluster. Our results show the active circulation of this emerging flavivirus in Thailand and the need for continuous investigation on this poorly known but threatening virus in Asia.

Avicenna's pharmacopeia for the treatment of animal bites.

Envenomation is a common medical problem. The Canon of Medicine written by Avicenna is one of the reliable sources of Persian medicine. The present study aims to identify Avicenna's clinical pharmacology approach and the pharmacopeia used for the treatment of animal envenomations and also to evaluate the related data in light of the current medicine. The Canon of Medicine was searched using related Arabic keywords for the contents about the treatment of animal bites. A literature search was conducted in scientific databases including PubMed, Scopus, Google Scholar, and Web of Science to obtain relevant data. Avicenna recommended one hundred and eleven medicinal plants for the treatment of bites of vertebrate and invertebrate venomous animals including snakes, scorpions, spiders, wasps, and centipedes. He mentioned different methods of administrating these drugs including oral drugs, lotions, sprayed drugs, slow-dissolving tablets in the mouth, and enemas. Moreover, he paid special attention to pain relief in addition to specific treatments for animal bites. In the Canon of Medicine, Avicenna recommended several medicinal plants alongside analgesics for the management and treatment of animal envenomations. The current research elucidates the clinical pharmacology and pharmacopeia of Avicenna for the treatment of animal envenomations. Further research is encouraged to evaluate the efficacy of these therapeutic agents for the treatment of animal bites.

Relationships between neuro-endocrine, electrocardiogram, and gastric mucosal damage parameters in naїve and stressed rats

Introduction The first scientific publication on general adaption syndrome, or as we know today biologic stress has been published in Nature in 1936 by the 29-year old Hans Selye.1 Szabo et al2 in the anniversary review "Stress" is 80 Years Old” conclude that despite the extensive and multidisciplinary research on stress during the last 80 years, a lot of basic and clinical research is needed to better understand the manifestations, central and peripheral molecular regulators of stress response, especially the modes of prevention/management of distress or its transformation into eustress and the treatment of stress-related diseases. In the vast majority of publications on stress, the HPA-, HPG- and autonomous systems are the objects of research, while the place in the general adaptation syndrome of such important hormones as calcitonin and PTH has been studied only in a few publications. Another methodological shortcoming of most studies is that the subjects of analysis are limited to a single neuro-endocrine system. Therefore, we set ourselves the goal: to analyze relationships between some adaptation hormones, HRV, calcitonin, and PTH as well as electrocardiogram and gastric mucosal damage in naїve and post stressed rats. Material and methods Ethics approval All animals were kept in room having temperature 22±2ºC, and relative humidity of 44-55% under 12/12 hours light and dark cycle with standard laboratory diet and water given ad libitum. Studies have been conducted in accordance with the rules and requirements of the “General Principles for the Work on Animals” approved by the I National Congress on Bioethics (Kyїv, Ukraine, 2001) and agreed with the provisions of the “European Convention for the Protection of Vertebrate Animals used for Experimental and other Scientific Purposes” (Council of Europe No 123, Strasbourg 1985), and the Law of Ukraine “On the Protection of Animals from Cruelty” of 26.02.2006. The removal of animals from the experiment was carried out under light inhalation (ether) anesthesia by decapitation. Participants The experiment is at 38 rats Wistar line: 18 males (Weight Mean=227 g; SD=25 g) and 20 females (Mean=214 g; SD=27 g). Study design and procedure Due to the purposeful formation of groups, the potential predictors of post-stress reactions of the neuro-endocrine-immune complex and the metabolome3,4,5 were almost identical both in mean values and, to a lesser extent, in variance (SD). In particular, the hypoxic test (sec) was: 136±59 and 133±81; swimming test (min): 19±11 and 19±17; HRV Stress index4 (units) as (AMo/2*Mo*MxDMn)1/3: 0,14±0,08 and 0,14±0,05 in intact animals and those exposed to acute stress. Results and discussion Adhering to the algorithm of the Truskavetsian Scientific School of Balneology, we recalculated the current values of the parameters in Z-scores. This approach makes it possible to adequately compare deviations from the norm of parameters expressed in different units and with different variability.10,11,12 This is all the more important in view of sexual dimorphism in endocrine parameters. In addition to drastically higher levels of testosterone (41.8±1.7 vs 3.53±0.24 nM/L), males have higher levels of calcitonin (36±6 vs 21±4 ng/L), but lower levels of parathyroid hormone (154±12 vs 185±3 ng/L), adrenals mass (44±5 vs 65±5 mg), corticosterone (340±45 vs 466±57 nM/L), and aldosterone (587±8 vs 639±24 pM/L). Conclusion The condition of the gastric mucosa and myocardium as essential targets of stressors is determined by the damaging and protective effects of adaptive hormones and the autonomic nervous system.

Ancestral glycoprotein hormone-receptor pathway controls growth in C. elegans.

In vertebrates, thyrostimulin is a highly conserved glycoprotein hormone that, besides thyroid stimulating hormone (TSH), is a potent ligand of the TSH receptor. Thyrostimulin is considered the most ancestral glycoprotein hormone and orthologs of its subunits, GPA2 and GPB5, are widely conserved across vertebrate and invertebrate animals. Unlike TSH, however, the functions of the thyrostimulin neuroendocrine system remain largely unexplored. Here, we identify a functional thyrostimulin-like signaling system in Caenorhabditis elegans. We show that orthologs of GPA2 and GPB5, together with thyrotropin-releasing hormone (TRH) related neuropeptides, constitute a neuroendocrine pathway that promotes growth in C. elegans. GPA2/GPB5 signaling is required for normal body size and acts through activation of the glycoprotein hormone receptor ortholog FSHR-1. C. elegans GPA2 and GPB5 increase cAMP signaling by FSHR-1 in vitro. Both subunits are expressed in enteric neurons and promote growth by signaling to their receptor in glial cells and the intestine. Impaired GPA2/GPB5 signaling causes bloating of the intestinal lumen. In addition, mutants lacking thyrostimulin-like signaling show an increased defecation cycle period. Our study suggests that the thyrostimulin GPA2/GPB5 pathway is an ancient enteric neuroendocrine system that regulates intestinal function in ecdysozoans, and may ancestrally have been involved in the control of organismal growth.

Xenopus laevis (Daudin, 1802) as a Model Organism for Bioscience: A Historic Review and Perspective.

In vitro systems have been mainly promoted by authorities to sustain research by following the 3Rs principle, but continuously increasing amounts of evidence point out that in vivo experimentation is also of extreme relevance. Xenopus laevis, an anuran amphibian, is a significant model organism in the study of evolutionary developmental biology, toxicology, ethology, neurobiology, endocrinology, immunology and tumor biology; thanks to the recent development of genome editing, it has also acquired a relevant position in the field of genetics. For these reasons, X. laevis appears to be a powerful and alternative model to the zebrafish for environmental and biomedical studies. Its life cycle, as well as the possibility to obtain gametes from adults during the whole year and embryos by in vitro fertilization, allows experimental studies of several biological endpoints, such as gametogenesis, embryogenesis, larval growth, metamorphosis and, of course, the young and adult stages. Moreover, with respect to alternative invertebrate and even vertebrate animal models, the X. laevis genome displays a higher degree of similarity with that of mammals. Here, we have reviewed the main available literature on the use of X. laevis in the biosciences and, inspired by Feymann's revised view, "Plenty of room for biology at the bottom", suggest that X. laevis is a very useful model for all possible studies.

Drought shifts soil nematodes to smaller size across biological scales

Drought events are increasingly affecting the planet's biodiversity. While shrinking body size in response to drought has been observed in many vertebrate animals, whether this rule applies to microscopic animals and the mechanisms during this process remains largely unknown. To address this knowledge gap, we conducted a regional-scale investigation and a microcosm experiment to systematically evaluate the impact of drought stress on the body size of the most abundant soil animals on Earth – nematodes – across various biological scales, including community, population and individual levels. Our results showed that nematode body size declined with drought stress at all biological scales, including a community shift toward smaller-sized species, a smaller body size at the population scale, and a decrease in size-at-age of individuals. Additionally, we designed a Petri dish experiment to examine the reversible plasticity of body size under drought stress using a drought-tolerant nematode species. We found that while nematode body size could not be fully reversed when drought stress was alleviated in the offspring generation, offspring from parents that experienced severe drought conditions could acquire tolerance, leading to a relatively smaller reduction in overall body size compared to those from parents that suffered no or light drought conditions. Overall, our study suggests that the increasing frequency of drought events at the global scale will lead to a reduction in soil nematode body size, potentially causing far-reaching consequences for additional changes in the climate, as well as nutrient cycling in soils.

A review of shark conservation and management legal frameworks in the Philippines

Sharks are among the oldest groups of vertebrate animals, dating back to over 400 million years ago. In recent decades, fishing pressure and demand for shark products and derivatives, such as fins, meat, oil, have increased significantly, driving global declines in shark populations. Sharks are a valuable resource in the Philippines and are used for both extractive and non-extractive purposes. The primary objective of this paper is to outline international, national, and local legal frameworks that enable the management and conservation of sharks in the Philippines. This review commences by providing some background regarding the politico-legal context of the Philippines, as well as information related to shark populations, fisheries and research in this country. This paper also aims to make a case that the Philippine government is in a unique position to build on the momentum of past initiatives by passing comprehensive national shark legislation and continuing to support regulation of shark trade globally. While a national legal framework on its own cannot solve all the issues related to shark fisheries and management, it will help create and institutionalize an anchor for existing and future conservation, research, governance, and communication initiatives.

Left Atrial Ligation in the Avian Embryo as a Model for Altered Hemodynamic Loading During Early Vascular Development.

Due to its four-chambered mature ventricular configuration, ease of culture, imaging access, and efficiency, the avian embryo is a preferred vertebrate animal model for studying cardiovascular development. Studies aiming to understand the normal development and congenital heart defect prognosis widely adopt this model. Microscopic surgical techniques are introduced to alter the normal mechanical loading patterns at a specific embryonic time point and track the downstream molecular and genetic cascade. The most common mechanical interventions are left vitelline vein ligation, conotruncal banding, and left atrial ligation (LAL), modulating the intramural vascular pressure and wall shear stress due to blood flow. LAL, particularly if performed in ovo, is the most challenging intervention, with very small sample yields due to the extremely fine sequential microsurgical operations. Despite its high risk, in ovo LAL is very valuable scientifically as it mimics hypoplastic left heart syndrome (HLHS) pathogenesis. HLHS is a clinically relevant, complex congenital heart disease observed in human newborns. A detailed protocol for in ovo LAL is documented in this paper. Briefly, fertilized avian embryos were incubated at 37.5 °C and 60% constant humidity typically until they reached Hamburger-Hamilton (HH) stages 20 to 21. The egg shells were cracked open, and the outer and inner membranes were removed. The embryo was gently rotated to expose the left atrial bulb of the common atrium. Pre-assembled micro-knots from 10-0 nylon sutures were gently positioned and tied around the left atrial bud. Finally, the embryo was returned to its original position, and LAL was completed. Normal and LAL-instrumented ventricles demonstrated statistically significant differences in tissue compaction. An efficient LAL model generation pipeline would contribute to studies focusing on synchronized mechanical and genetic manipulation during the embryonic development of cardiovascular components. Likewise, this model will provide a perturbed cell source for tissue culture research and vascular biology.

ECHA reports progress on reducing animal tests

In vitro tests and other nonanimal methods are on the rise for ensuring the safety of chemicals in the European Union, according to a report from the European Chemicals Agency (ECHA). The report finds that read-across is the most common nonanimal method for assessing chemical risks under the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) regulation. Read-across uses data from chemicals with known toxicity to assess the safety of other chemicals with similar properties. The report also finds a notable increase since 2019 in the use of in vitro test methods that involve human cells, tissues, or organs. Under REACH, in vitro tests are used primarily to obtain data on skin and eye irritation and skin sensitization. Other nonanimal methods include waiving the need for certain toxicity data and using computational models to predict toxicity based on chemical structures. Testing chemicals on vertebrate animals is permitted under REACH only

Understanding the formation of efflorescence on beeswax models housed at the Natural History Museum of Florence

The Natural History Museum of Florence, and in particular “La Specola”, houses several ceroplastics collections produced in Florence in the XVIII-XIX centuries by Clemente Susini and co-workers. The collections include models of fruits and plants, vertebrate and invertebrate animals, and over 1400 anatomical waxes. All models are mainly constituted by beeswax. Although extremely resistant, the models undergo a wide range of degradation processes. Periodically, a white crystalline efflorescence, commonly called “wax bloom”, appears on the surface of the objects obviously impairing their aesthetical features.Many mechanisms have been hypothesized about the formation of these white exudates, but non univocal conclusions have yet been drawn. Our gas chromatography-mass spectrometry results indicate that wax bloom strongly depends on the composition of the models, being related to the presence of oils and fats originally used as plasticizers or pigments’ media. Thermal and X-ray measurements indicate that a poor solid-solid miscibility between the efflorescence compounds and other beeswax components leads to phase separation followed by migration of the immiscible materials towards the model surface. Besides, X-ray diffraction reveals an increased structural order of models as compared to recent beeswax certainly correlated with the increased brittleness determined by rheology measurements.