Assessment of the developmental and neurotoxicity of the mosquito control larvicide, pyriproxyfen, using embryonic zebrafish

Abstract

In 2014, as an attempt to address the Zika health crisis by controlling the mosquito population, Brazil took the unprecedented action of applying a chemical larvicide, pyriproxyfen, to drinking water sources. The World Health Organization has established an acceptable daily intake of pyriproxyfen to be 100 micrograms per kg of body weight per day, but studies have demonstrated that at elevated doses (>5000 mg/kg), there are adverse effects in mice, rats and dogs. To better understand the potential developmental toxicity of pyriproxyfen, we utilized the embryonic zebrafish. Our results demonstrate that the concentration resulting in 50% of animals presenting adverse morphological effects (EC50), including craniofacial defects, was 5.2 μM for daily renewal exposure, and above this concentration, adverse behavioral effects were also observed in animals that followed a static exposure regimen. Thus, zebrafish data suggest that the developmental toxicity of pyriproxyfen may not be limited to insects.

The larvicide pyriproxyfen blamed during the Zika virus outbreak does not cause microcephaly in zebrafish embryos

“…This observation is consistent with the half maximal effective concentration (EC 50 ) of 1.6 μg/ml (i.e. 5 μM) that has been found in a previous study in zebrafish37. Consistently, we also treated embryos with extremely high doses of pyriproxyfen, concentrations ranging from 10 to 100 μg/ml, and noticed a high level of toxicity leading to death in 100% of the embryos as early as 2 dpf ( p  < 0.0001, Logrank test, N = 2, n = 20; Fig.…”

“…All the experimental protocols were performed under the approval of the vertebrate animal welfare assurance from the Institutional Animal Care and Use Committee (IACUC) for the use of adult zebrafish (approved on 2015/08/31). The elavl3, nestin and gfap transgenic lines were previously generated as described3743 and are routinely maintained in our fish facility at the CRCHUM.…”

Behavioral screening of the LOPAC1280 library in zebrafish embryos

“…As such, a repertoire of zebrafish-based behavioral assays across multiple life-stages (including adulthood) have been developed over the last 10–15 years (Bang et al, 2002; Brockerhoff et al, 1995; Egan et al, 2009; Kokel et al, 2010). Although different forms of larval and adult zebrafish locomotion have been leveraged within behavioral assays, spontaneous activity (tail contraction) – a behavior that occurs from late-segmentation (~17–19 hours post-fertilization, hpf) through early-pharyngula (~27–29 hpf) – represents an early, primitive form of motor activity within zebrafish embryos, providing a potential readout for rapid identification of neuroactive chemicals (Kokel et al, 2010; Raftery et al, 2014; Reif et al, 2015; Truong et al, 2016). Although previous studies have explored the biological basis of this behavior (Knogler et al, 2014; Knogler and Drapeau, 2014; Saint-Amant and Drapeau, 2001, 2000), it remains unclear whether spontaneous activity is responsive only to certain compounds with specific mechanisms of action.…”

The pyriproxyfen metabolite 4'OH- pyriproxyfen disrupts thyroid hormone signaling and enhances Musashi-1 levels in neuroprogenitors.

“…Only two studies, both using Zebrafish, have addressed the question of whether PPF could be implicated in microcephaly, each producing contradictory results (Dzieciolowska et al, 2017; Truong et al, 2016). One study (Truong et al, 2016) demonstrated that PPF induces adverse morphological changes including craniofacial defects in zebrafish at 5.2 μM (1.66 mg/L) - EC50. In the behavioral test PPF at 6.4μM (2 mg/L) and 64μM (20mg/L) significantly reduced the mobility of the zebrafish larvae compared to control animals.…”

“…In the behavioral test PPF at 6.4μM (2 mg/L) and 64μM (20mg/L) significantly reduced the mobility of the zebrafish larvae compared to control animals. The authors concluded that the developmental toxicity of PPF may not be limited only to insects (Truong et al, 2016).…”