The pesticide Fenoxycarb (IUPAC: ethyl [2-(4-phenoxy-phenoxy)ethyl] carbamate, CAS No. 72490-01-8) is a carbamate used to control various insect pests in crops and ornamental cultures 1. It regulates insect growth by mimicking the juvenile hormone, preventing the insect from reaching maturity 2. However, it has also been observed in other arthropods 3–5. At the molecular level, it has been observed that it mediates cell proliferation inhibition and apoptosis in insects (Ayisha Banu and Manogem, 2022) and upregulates the genes involved in juvenile hormonesynthesiss in spiders (Yang et al., 2022). It is considered not harmful to vertebrates and non-target species, affecting insects and other arthropods 5, 7–9. It was reported below the detection level (0.0004 mg/L) 24 to 48 h following aerial application to experimental ponds 10. Additionally, it has been considered that nominal concentrations of 0.20, 0.80, 3.2, 13, and 50 µg/L brackets predicted environmental levels 11. Furthermore, Fenoxycarb is considered environmentally safe because of its rapid degradation 10, with a dissipation time 50 (DT50) of 4.13 days in the water column and 15 days in sediment (PPDB database: http://sitem.herts.ac.uk/aeru/ppdb/en/Reports/304.htm) 1. It has a low drift to adjacent lands from fields where it is applied 12, although it may end up in surface waters due to spray drift, runoff, or drainage 13. It has been banned in the EU but is in use in the UK, USA, and Australia. European Food Safety Authority considered it with no areas of concern in the areas of mammalian toxicology and consumer risk assessment for using fenoxycarb in apples and pears 14. However, while a low risk was assessed for birds and mammals, soil non-target macro and micro-organisms, and terrestrial non-target plants, a high risk was identified for aquatic organisms with a data gap in the risk to aquatic invertebrates considering the mode of action and the most sensitive growth stages 14. Although it is considered with low toxicity upon oral, dermal or inhalation exposure, it has been proposed to be classified as a chemical with limited evidence of a carcinogenic effect because it can produce lung and liver tumors in mice by inducing peroxisome proliferation, a mechanism less sensitive in humans 14. Because of it, fenoxycarb is labeled as very toxic for aquatic life and suspected of causing cancer.
In vertebrates, no effect on reproduction in sheep was observed 15, but it has been described that in cultured rat cortical neurons exposed for one week, Fenoxycarb considerably decreased ATP levels, mitochondrial membrane potential, and glucose consumption 16. Furthermore, it inhibits rat brain acetylcholinesterase and nicotinic acetylcholine receptors expressed in Xenopus laevis oocytes 17. On the other hand, some reports show that it can negatively affect egg production and the hatching rate in the collembola Yuukianura szeptyckii 18, although previous studies on Folsomia candida did not show such an effect 19. Similarly, adverse effects such as inhibition of molting and body length growth were observed when the shrimp Neocaridina davidi was exposed for two weeks to concentration as low as 10 µg/L 3, while the crab Rhithropanopeus harrisii experienced delayed metamorphosis at 48 µg/L of Fenoxycarb (Cripe et al., 2003). Taking together these results, additional studies are required to know the impact of this pesticide on ecosystems.
There is a lack of information on the effects of Fenoxycarb on non-arthropod freshwater invertebrates, but it is expected to be harmless to them. However, the paucity of knowledge of the physiology of invertebrates, especially regarding the endocrine system, requires confirmation of this extreme. This work aims to test the toxicity of Fenoxycarb at the transcription level, in the freshwater gastropod Physella acuta (Draparnaud, 1805) by exposing the animals for one week and analyzing the transcription profile with an array covering different relevant cellular pathways.
The freshwater snail Physella acuta, also known as Physa acuta, is a hermaphroditic and cosmopolitan species. It lives in lakes and ponds and lays its eggs in an egg mass that requires around two weeks to develop. The hatched juveniles grow for two months until they reach the adult stage, mate, and lay eggs. The species is easily cultured in the laboratory, so it is used in toxicity studies as representative of the gastropods 20,21. Recently, we designed an array to study the response to toxicants at the gene expression level in this species (Prieto-Amador et al., 2021). It included 34 genes and 4 reference genes. In addition, we extended it to 40 target genes, including some of them to analyze the alterations at the transcriptional activity level in several cellular processes. The sequence of nine genes is described for the first time for this species and extends the number of genes that can be used as biomarkers. The sequences code for proteins related to the endocrine system (galanin receptor type 2 [GalR2], estrogen-related receptor [ERR], membrane progestin receptor-beta [MPR], estradiol 17-beta-dehydrogenase 8 [Hsd17b8], and retinoic acid receptor [RXR]), DNA repair (poly-ADP-ribose polymerase I [PARP1], DNA repair protein XRCC3 [XRCC3], nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha [IkBa]), and stress response (heat shock protein 70 B2-like [Hsp70 B2]). Overall, the array allows analysis of alterations in the endocrine system, detoxification mechanisms, DNA repair, the nervous system, apoptosis, oxidative stress, stress, epigenetics, the immune system, energy metabolism, and lipid transport. In this way, the array shows changes in the main mechanisms involved in the response to stress and detoxification but also the response of processes involved in long-term effects, such as epigenetic modification mechanisms and DNA repair, that would be activated in case of any genotoxic effect of the compound.
Safety at the environmental level is a concern of all the products used as pesticides. The search for new pesticides with a reduced impact in non-target species demands testing in these species because some of them can have low impact. However, the lack of knowledge about the physiology of invertebrates requires experimental work to confirm it. It will also provide additional information about the physiology of the invertebrates, decreasing the gap with the vertebrates and favoring the use of invertebrates as alternative methods that reduce the use of vertebrates in the test of toxicity. As stated, the work aims to elucidate the putative mechanisms involved in Fenoxycarb toxicity in P. acuta exposed to predicted environmentally relevant concentrations for one week by analyzing different processes involved in the mid- and long-term response, and some of them, such as the endocrine system and the immune system, involved in relevant physiological mechanisms to survive.