Assessment of the effects of 2,4-D-dichlorophenoxyacetic acid-based herbicide exposure in eggs and embryos of golden apple snails using OCT.

This study aimed to evaluate the effects of herbicide 2, 4-D-dichlorophenoxy acetic acid on golden apple snail eggs and embryos. Additionally, the study assessed the applicability of optical coherence tomography (OCT), a non-invasive depth cross-sectional microscopic imaging technique, as a novel method, to the best of our knowledge, for studying morphological changes in golden apple snail eggs and embryos, in comparison to the conventional approach of using white light microscopy. The study revealed that the herbicide 2,4-D-dichlorophenoxy acetic acid affected the hatchery rate and morphological changes of the eggs and embryos. The lethal concentration (LC50), representing the concentration of a substance that is expected to cause death in half of the population being studied, of the golden apple eggs and embryos increased with longer exposure time and higher concentrations. The estimated median effective concentration (EC50), which denotes the concentration producing the desired effect in 50% of the exposed golden apple embryos, exhibited a similar trend of change as the LC50. When compared to the microscopic study, it was observed that OCT could be employed to investigate morphological changes of golden apple snail eggs and embryos, enabling evaluation of alterations in both 2D and 3D structures.

The effects of commercial 2,4-D herbicide on game fish species: Natural lake water vs. laboratory system water.

Aquatic herbicides with active ingredient 2,4-dichlorophenoxyacteic acid (2,4-D) are commonly used to control and combat aquatic non-native species that cause detrimental impacts including habitat destruction, strained resources among biota, and biodiversity loss. While many (eco)toxicology studies are performed in the laboratory under highly controlled circumstances, these studies may disregard the nuances and disorder that come with the complexity of natural aquatic ecosystems. Therefore, we conducted a series of laboratory experiments using laboratory system water, different lake waters, and different water parameters to determine the effects of ecologically relevant concentrations of 2,4-D (0.00-4.00 ppm a.e.) on the development and survival of two freshwater game species (Sander vitreus and Esox lucius). For 2,4-D exposures using different water sources, there were significant main effects of 2,4-D concentration and water source on walleye embryo and larval survival, however, there was no significant interaction between 2,4-D exposure and water source. For 2,4-D exposures and pH (5-9 pH), there were significant main effects of 2,4-D concentration and pH on walleye and northern pike embryo survival and a significant interaction between 2,4-D exposure and pH. Our results indicate that 2,4-D exposures in controlled laboratory system water can predict similar outcomes as 2,4-D exposures in natural lake water. Moreover, individual water parameters, such as pH, play a significant role in the toxicity of 2,4-D. Taken together, these results suggest that highly controlled laboratory studies are a useful tool for predicting impacts on survival of non-target fish in natural waters, but it is crucial for management agencies to consider individual water sources and specific lake water parameters in herbicide risk assessments to minimize the impacts to non-target organism.

Agent orange exposure and prostate cancer risk in the million veteran program.

BACKGROUND: The US government considers veterans to have been exposed to Agent Orange if they served in Vietnam while the carcinogen was in use, and these veterans are often deemed at high risk of prostate cancer (PCa). Here, we assess whether presumed Agent Orange exposure is independently associated with increased risk of any metastatic or fatal PCa in a diverse Veteran cohort still alive in the modern era (at least 2011), when accounting for race/ethnicity, family history, and genetic risk. PATIENTS AND METHODS: Participants in the Million Veteran Program (MVP; enrollment began in 2011) who were on active duty during the Vietnam War era (August 1964-April 1975) were included (n = 301,470). Agent Orange exposure was determined using the US government definition. Genetic risk was assessed via a validated polygenic hazard score. Associations with age at diagnosis of any PCa, metastatic PCa, and death from PCa were assessed via Cox proportional hazards models. RESULTS AND INTERPRETATION: On univariable analysis, exposure to Agent Orange was not associated with increased PCa (hazard ratio [HR]: 1.02, 95% confidence interval [CI]: 1.00-1.04, p = 0.06), metastatic PCa (HR: 0.98, 95% CI: 0.91-1.05, p = 0.55), or fatal PCa (HR: 0.94, 95% CI: 0.79-1.09, p = 0.41). When accounting for race/ethnicity and family history, Agent Orange exposure was independently associated with slightly increased risk of PCa (HR: 1.06, 95% CI: 1.04-1.09, <10-6) but not with metastatic PCa (HR: 1.07, 95% CI: 0.98-1.15, p = 0.10) or PCa death (HR: 1.02, 95% CI: 0.83-1.23, p = 0.09). Similar results were found when accounting for genetic risk. Agent Orange exposure history may not improve modern PCa risk stratification.

Effects of pasture intensification and sugarcane cultivation on non-target species: A realistic evaluation in pesticide-contaminated mesocosms.

Conventional soil management in agricultural areas may expose non-target organisms living nearby to several types of contaminants. In this study, the effects of soil management in extensive pasture (EP), intensive pasture (IP), and sugarcane crops (C) were evaluated in a realistic-field-scale study. Thirteen aquatic mesocosms embedded in EP, IP, and C treatments were monitored over 392 days. The recommended management for each of the areas was simulated, such as tillage, fertilizer, pesticides (i.e. 2,4-D, fipronil) and vinasse application, and cattle pasture. To access the potential toxic effects that the different steps of soil management in these areas may cause, the cladoceran Ceriophania silvestrii was used as aquatic bioindicator, the dicot Eruca sativa as phytotoxicity bioindicator in water, and the dipteran Chironomus sancticaroli as sediment bioindicator. Generalized linear mixed models were used to identify differences between the treatments. Low concentrations of 2,4-D (<97 µg L-1) and fipronil (<0.21 µg L-1) in water were able to alter fecundity, female survival, and the intrinsic rate of population increase of C. silvestrii in IP and C treatments. Similarly, the dicot E. sativa had germination, shoot and root growth affected mainly by 2,4-D concentrations in the water. For C. sancticarolli, larval development was affected by the presence of fipronil (<402.6 ng g-1). The acidic pH (below 5) reduced the fecundity and female survival of C. silvestrii and affected the germination and growth of E. sativa. Fecundity and female survival of C. silvestrii decrease in the presence of phosphorus-containing elements. The outcomes of this study may improve our understanding of the consequences of exposure of freshwater biota to complex stressors in an environment that is rapidly and constantly changing.

Meta-analytical review of antioxidant mechanisms responses in animals exposed to herbicide 2,4-D herbicide.

The 2,4-Dichlorophenoxyacetic acid (2,4-D) is a low-cost herbicide to eradicate broadleaf weeds. Since the development of 2,4-D resistant transgenic crops, it has been described as one of the most widely distributed pollutants in the world, increasing concern about its environmental impacts. This study aimed to elucidate the antioxidant system response in animals exposed to 2,4-D by different routes of exposure. It focused on determining if tissue, phylogenetic group, and herbicide formulation would influence the antioxidant mechanisms. A careful literature search of Scopus, WoS, and Science Direct retrieved 6983, 24,098, and 20,616 articles, respectively. The dataset comprised 390 control-treatment comparisons and included three routes of exposure: transgenerational, oral, and topical. The data set for transgenerational and oral exposure revealed oxidative stress through a decrease in enzymatic activities and the level of molecules of the antioxidant system. In contrast, topical exposure increased the oxidative stress. Tissue-specific analyses revealed that the transgenerational effects reduced hepatic catalase (CAT) activity. Oral exposure caused a variety of effects, including increased CAT activity in the prostate and decreased activity in various tissues. Mammals predominate in the transgenerational and oral groups, showing a significantly reduced activity of the antioxidant system. In contrast, in the topical exposure, an increased activity of oxidative stress biomarkers was observed in fish, earthworms, and mollusks. The effects of the 2,4-D formulation on oxidative stress responses showed significant differences between pure and commercial formulations, with oral exposure resulting in decreased activity and topical exposure increasing responses. In summary, orally exposed animals exhibited a clear decrease in enzyme activities, transgenerational exposure elicited tissue-specific prompted biochemical reductions, and topical exposure induced increased responses, emphasizing the need for unbiased exploration of the effects of 2,4-D on biomarkers of oxidative stress while addressing publication bias in oral and topical datasets.

Ecological Implications on Aquatic Food Webs Due to Effects of Pesticides on Invertebrate Predators in a Neotropical Region.

Predation presents specific behavioral characteristics for each species, and the interaction between prey and predator influences the structuring of the food web. Concerning insects, predation can be affected in different ways, such as exposure to chemical stressors, e.g., pesticides. Therefore, analyses were carried out of the effects of exposure to insecticide fipronil and the herbicide 2,4-D on predation, parameters of food selectivity, and the swimming behavior of two neotropical predatory aquatic insects of the families Belostomatidae (giant water bugs) and larvae of Libellulidae (dragonfly). These predatory insects were exposed for 24 h to a commercial formulation of the chlorophenoxy herbicide, 2,4-D at nominal concentrations of 200, 300, 700, and 1400 µg L-1, and to a commercial formulation of the phenylpyrazole insecticide, fipronil at nominal concentrations of 10, 70, 140, and 250 µg L-1. In a control treatment, the insects were placed in clean, unspiked water. At the end of the exposure, the maximum swimming speed of the predators was evaluated. Afterward, the predators were placed in clean water in a shared environment for 24 h with several prey species, including the cladoceran Ceriodaphnia silvestrii, larvae of the insect Chironomus sancticaroli, the amphipod Hyalella meinerti, the ostracod Strandesia trispinosa, and the oligochaete Allonais inaequalis for 24 h. After this period, the consumed prey was counted. The results reveal that predators from both families changed prey consumption compared with organisms from the control treatment, marked by a decrease after exposure to fipronil and an increase in consumption caused by 2,4-D. In addition, there were changes in the food preferences of both predators, especially when exposed to the insecticide. Exposure to fipronil decreased the swimming speed of Belostomatidae individuals, possibly due to its neurotoxic effect. Exposure to the insecticide and the herbicide altered prey intake by predators, which could negatively influence the complex prey-predator relationship and the functioning of aquatic ecosystems in contaminated areas.

A mixture of glyphosate and 2,4-D herbicides enhances the deleterious reproductive outcomes induced by Western diet in obese male mice.

The consumption of Western diet (WD) - enriched in fats and sugars - is associated with overweight, obesity and male reproductive disorders. In addition to WD intake, crops and dairy products display residues of herbicides, including glyphosate and 2,4-D that are widely applied worldwide. The concomitant exposure to WD and herbicides - mimicking contemporary scenarios - is not fully investigated. Thus, we evaluated the effects of glyphosate and 2,4-D, alone or in mixture, on WD-induced alterations in the male genital system. Male C57BL6J mice were submitted to WD (chow containing 20% lard, 0.2% cholesterol, 20% sucrose, and high sugar solution with 23.1 and 18.9 g/L of D-fructose and D-glucose) for 6 months. Concomitantly to WD, the animals received glyphosate (0.05, 5, or 50 mg/kg/day), 2,4-D (0.02, 2 or 20 mg/kg/day) or their mixture (0, 05 + 0.02, 5 + 2, or 50 + 20 mg/kg/day) by intragastrical administration (5×/week). Doses were based on Acceptable Daily Intake (ADIs) or No Observed Adverse Effect Level (NOAEL) values. Herbicide exposure did not alter the WD-induced obesity, hypercholesterolemia and hyperglycemia. WD induced sperm cell abnormalities, reduced the number, volume and area of Leydig cells, enhanced the frequency of epididymal abnormalities, decreased the proliferation in both germinal and epididymal epithelia, and reduced the number of androgen receptor (AR) positive epididymal cells. Remarkably, the herbicide mixtures promoted such WD-induced effects: increased the frequency of sperm cell and epididymal abnormalities (absence of sperm, cytoplasmic vacuoles, and clear cell hypertrophy) (5 + 2 and 50 + 20 doses); decreased Leydig cell nuclei volume and area (5 + 2 and 50 + 20 doses), reduced epididymal cell proliferation (all mixtures), and AR expression (50 + 20 dose). In addition, herbicide mixtures reduced serum testosterone levels (5 + 2 and 50 + 20 doses). Our findings indicate that the mixture of glyphosate and 2,4-D herbicides, mimicking environmentally relevant scenarios, promotes WD-induced changes in the male genital system.

Fate and toxicity of 2,4-D and fipronil in mesocosm systems.

2,4-D and fipronil are among Brazil's most used pesticides. The presence of these substances in surface waters is a concern for the aquatic ecosystem health. Thus, understanding the behavior of these substances under environmentally relevant conditions is essential for an effective risk assessment. This study aimed to determine the degradation profiles of 2,4-D and fipronil after controlled application in aquatic mesocosm systems under influencing factors such as environmental aspects and vinasse application, evaluate pesticide dissipation at the water-sediment interface, and perform an environmental risk assessment in water and sediment compartments. Mesocosm systems were divided into six different treatments, namely: control (C), vinasse application (V), 2,4-D application (D), fipronil application (F), mixture of 2,4-D and fipronil application (M), and mixture of 2,4-D and fipronil with vinasse application (MV). Pesticide application was performed according to typical Brazilian sugarcane management procedures, and the experimental systems were monitored for 150 days. Pesticide dissipation kinetics was modeled using first-order reaction models. The estimated half-life times of 2,4-D were 18.2 days for individual application, 50.2 days for combined application, and 9.6 days for combined application with vinasse. For fipronil, the respective half-life times were 11.7, 13.8, and 24.5 days. The dynamics of pesticides in surface waters resulted in the deposition of these compounds in the sediment. Also, fipronil transformation products fipronil-sulfide and fipronil-sulfone were quantified in water 21 days after pesticide application. Finally, performed risk assessments showed significant potential risk to environmental health, with RQ values for 2,4-D up to 1359 in freshwater and 98 in sediment, and RQ values for fipronil up to 22,078 in freshwater and 2582 in sediment.

Co-exposure to environmental microplastic and the pesticide 2,4-dichlorophenoxyacetic acid (2,4-D) induce distinctive alterations in the metabolome and microbial community structure in the gut of the earthworm Eisenia andrei.

Microplastics (MPs) are recognized as emergent pollutants and have become a significant environmental concern, especially when combined with other contaminants. In this study, earthworms, specifically Eisenia andrei, were exposed to MPs (at a concentration of 10 µg kg-1 of soil), herbicide 2,4-D (7 mg kg-1 of soil), and a combination of the two for 7 and 14 days. The chemical uptake in the earthworms was measured, and the bacterial and archaeal diversities in both the soil and earthworm gut were analyzed, along with the metabolomic profiles. Additionally, data integration of the two omics approaches was performed to correlate changes in gut microbial diversity and the different metabolites. Our results demonstrated that earthworms ingested MPs and increased 2,4-D accumulation. More importantly, high-throughput sequencing revealed a shift in microbial diversity depending on single or mixture exposition. Metabolomic data demonstrated an important modulation of the metabolites related to oxidative stress, inflammatory system, amino acids synthesis, energy, and nucleic acids metabolism, being more affected in case of co-exposure. Our investigation revealed the potential risks of MPs and 2,4-D herbicide combined exposure to earthworms and soil fertility, thus broadening our understanding of MPs' toxicity and impacts on terrestrial environments.

A Thorny Problem: Glyphosate and 2,4-D Associated with Neurobehavioral Effects for Ecuadorian Adolescents.

Among adolescents in an area known for its rose and other flower crops, exposures to 2,4-D and glyphosate (but not DEET) were associated with lower scores on neurobehavioral assessments. Notably, these kids did not work in floriculture.

Modifying effects of 2,4-D and Glyphosate exposures on gut-liver-adipose tissue axis of diet-induced non-alcoholic fatty liver disease in mice.

Nonalcoholic fatty liver disease (NAFLD), which is linked to western diet (WD) intake, affects 30% of the world's population and involves the crosstalk of liver steatosis, hypertrophy/inflammation of adipose tissue and deregulation of gut microbiome. Glyphosate and 2,4-D are some of the most applied herbicides worldwide, and their roles in NAFLD have not been investigated. Thus, the present study evaluated whether glyphosate and 2,4-D, in single or mixed exposure, alter WD-induced NAFLD in a mouse model. Male C57Bl/6 mice (n = 10/group) received a fat (30% lard, 0.02% cholesterol), and sucrose-rich diet (20%) and high sugar solution (23.1 and 18.9 g/L of fructose and glucose) for 6 months. Simultaneously, animals received glyphosate (0.05 or 5 mg/kg/day), 2,4-D (0.02 or 2 mg/kg/day), or their combination (0.05 +0.02 or 5 +2 mg/kg/day) by intragastrical administration (5 ×/week). Doses were based on the Acceptable Daily Intake (ADIs) or No Observed Adverse Effect Level (NOAEL) levels. Herbicide exposures featured differential responses. WD-induced obesity, hypercholesterolemia, and hyperglycemia remained unaltered. Compared to the group receiving only WD, only the concomitant exposure to WD and 2,4-D (2 mg) enhanced the percentage of mice with moderate/severe hepatic inflammation, CD68 macrophage infiltration, and malondialdehyde levels in the liver. In line, this herbicide modulated immune response- (including Cd4, C8b, Cd28, Cxcr3, Cxcr6) and oxidative stress-related (such as Gsta1, Gsta2, Gsta4, Gstm1, Gstm2, Gstm3, Gstm4, Nqo1, Gpx2) genes in the hepatic transcriptome analysis. This exposure also enriched pro-inflammatory Deferribacteres phylum in fecal microbiome. In general, the herbicide mixtures did not feature the same effects attributed to 2,4-D isolated exposure. Our findings indicate that 2,4-D, at a dose within the toxicological limits, was able to induce disturbances in mainly at the liver and gut axes involved in NAFLD development in male mice.

Cyanobacterial degradation of herbicide 2,4-dichlorophenoxyacetic acid (2,4-D): Its response to the oxidative stress induced by the primary degradation product 2,4-dichlorophenol (2,4-DCP).

Excessive use of herbicides in agricultural fields has become a major environmental concern due to the negative effects on the ecosystem. Microbial degradation has been well-known as an effective approach for combating such non-natural substances in soil. In the present study, the degradation of 2,4-Dichlorophenoxyacetic acid (2,4-D) as a result of metabolic activities of a cyanobacterium Nostoc muscorum Meg 1 was investigated using GC-MS analysis. After seven days of 2,4-D exposure, the main residue obtained was 2,4-dichlorophenol (2,4-DCP) at RT: 8.334 (confirmed using NIST library). The effects of 2,4-DCP were studied in a cyanobacterium Nostoc muscorum Meg 1 isolated from a rice field where 2,4-D is commonly used. Exposure to 2,4-DCP at 20, 40, and 80 ppm significantly increased ROS production in the cyanobacterium by 74, 107, and 211 % (p < 0.001). With rising 2,4-DCP concentrations in the surroundings, lipid peroxidation and protein oxidation in the organism correspondingly increased, indicating cellular injury. The mRNA and protein contents, and also the activities of different oxidant neutralizing enzymes such as CAT, SOD, GR, and GPx and the non-enzymatic antioxidants (proline, GSH, thiol and phytochelatin content) were found augmented in 20 ppm 2,4-DCP exposed cultures. However, in the presence of 40 and 80 ppm 2,4-DCP, most enzymatic and non-enzymatic antioxidants were severely compromised. At higher exposures, the organism's attempt to mitigate the oxidants was still visible, as both proline and TSH levels increased. SEM and TEM analysis aided in visualizing the effects of 2,4-DCP on the morphology and ultrastructures of the organism.

Impact of 2,4-D and glyphosate-based herbicides on morphofunctional and biochemical markers in Scinax squalirostris tadpoles (Anura, Hylidae).

Understanding the effects of pesticides on non-target organisms is essential to assess the impact of these xenobiotics on the environment, allowing for a more informative and safer usage. The present study sought to evaluate the response of Scinax squalirostris tadpoles when exposed to different concentrations of two herbicides, DEZ® (i.e., dichlorophenoxyacetic acid or 2,4-D) and Roundup® Original (i.e., glyphosate). We collected 140 tadpoles between Gosner's 25 and 34 stages in a preservation area of the South American jelly palm Butia odorata. The animals were separated into eight groups and maintained in aquariums: acclimatization control (17 days), exposure control (24 days), and six exposure groups (7 days), including three concentration groups of each pesticide (4 µg/L, 15 µg/L, and 30 µg/L 2,4-D acid equivalent; 65 µg/L, 250 µg/L, and 500 µg/L glyphosate acid equivalent). Markers of body condition (length, body mass, K and Kn index) and oxidative balance (superoxide dismutase, catalase, glutathione S-transferase, TBARS, and carbonyl proteins) were analyzed. After 24 days (17 days of acclimation plus 7 days of exposure), tadpoles exposed to 15 µg/L of 2,4-D and 65 µg/L of glyphosate grew at higher than expected concentrations. They also had less lipoperoxidation than control tadpoles and higher superoxide dismutase, catalase, and glutathione S-transferase activity, specifically at the highest herbicide concentrations (2,4-D: 30 µg/L; glyphosate: 500 µg/L of glyphosate). Only the highest concentration of 2,4-D determined an increase in the levels of carbonyl proteins, indicating oxidative damage induced. DEZ® required more antioxidant defenses and induced a concentration-dependent answer of carbonylated proteins, suggesting oxidative stress and more toxic potential. These results may help government agencies make more conscious decisions regarding the usage of these chemicals and consider a balance between the conservation of amphibian species and agribusiness economic sustenance.

Assessment of the impact of glyphosate and 2,4-D herbicides on the kidney injury and transcriptome changes in obese mice fed a Western diet.

The development of chronic kidney disease has been associated with comorbidities resulting from the consumption of Westernized dietary (WD) patterns, including obesity and other metabolic dysfunctions. Kidneys also have a crucial role in the metabolism and excretion of xenobiotics, including herbicides. There is limited knowledge regarding the simultaneous exposure to WD and glyphosate (glypho) and 2,4-D, the most used herbicides globally. Thus, this study examined whether exposure to glypho and/or 2,4-D, either individually or in mixed, could impact the early effects of WD intake on kidney histology and gene expression in a rodent model. Male C57BL6J mice were fed a WD containing 20% lard, 0.2% cholesterol, 20% sucrose, and high sugar solution with 23.1 and 18.9 g/L of D-fructose and D-glucose for six months. During this period, the mice also received glypho (0.05 or 5 mg/kg/day), 2,4-D (0.02 or 2 mg/kg/day), or a mixture of both (0.05 +0.02, 5 +2 mg/kg/day) via intragastric administration five times per week. The doses were within or below the established regulatory limits. While single or mixed exposures did not alter WD-induced obesity, tubular lipid vacuolation, or increased serum creatinine levels; the exposure to higher doses of the mixture (5 +2) reduced the mesangial matrix area and tubular cell proliferation, while increasing the density of F4/80 macrophages in the renal interstitium. In terms of transcriptomic analysis, the herbicide mixture altered the expression of 415 genes in the kidney, which were found to be associated with immune response processes, particularly those related to phagocyte activity. While discrete, findings indicate that herbicide mixtures, rather than single exposures, might induce minor deleterious effects on the kidneys of obese mice under WD intake.

2,4-dichlorophenoxyacetic acid (2,4-D) exposure during postnatal development alters the effects of western diet on mouse prostate.

Western diet (WD), abundant in saturated fats and simple carbohydrates, has been associated with the development of prostate diseases. In addition, 2,4-dichlorophenoxyacetic acid (2,4-D), an herbicide used in agricultural and non-agricultural settings, may interfere with the endocrine system impacting reproductive health. The association of both factors is something common in everyday life, however, there are no relevant studies associating them as possible modulators of prostatic diseases. This study evaluated the action of the herbicide 2,4-D on the postnatal development of the prostate in mice fed with WD. Male C57Bl/6J mice received simultaneously a WD and 2,4-D at doses of 0.02, 2.0, or 20.0 mg/kg b.w./day for 6 months. The prolongated WD intake induced obesity and glucose intolerance, increasing body weight and fat. WD induced morphological changes and increased PCNA-positive epithelial cells in prostate. Additionally, the WD increased gene expression of AR, antioxidant targets, inflammation-related cytokines, cell repair and turnover, and targets related to methylation and miRNAs biosynthesis compared to the counterpart (basal diet). 2,4-D (0.02 and 2.0) changed prostate morphology and gene expression evoked by WD. In contrast, the WD group exposed to 20 mg/kg of 2,4-D reduced feed intake and body weight, and increased expression of androgen receptor and genes related to cell repair and DNA methylation compared to the negative control. Our results showed that 2,4-D was able to modulate the effects caused by WD, mainly at lower doses. However, further studies are needed to elucidate the mechanisms of 2,4-D on the obesogenic environment caused by the WD.

Embryonic abnormalities and genotoxicity induced by 2,4-dichlorophenoxyacetic acid during indirect somatic embryogenesis in Coffea.

Indirect somatic embryogenesis (ISE) is a morphogenetic pathway in which somatic cells form callus and, later, somatic embryos (SE). 2,4-dichlorophenoxyacetic acid (2,4-D) is a synthetic auxin that promotes the proliferation and dedifferentiation of somatic cells, inducing the ISE. However, 2,4-D can cause genetic, epigenetic, physiological and morphological disorders, preventing the regeneration and/or resulting abnormal somatic embryos (ASE). We aimed to evaluate the toxic 2,4-D effect during the Coffea arabica and C. canephora ISE, assessing the SE morphology, global 5-methylcytosine levels (5-mC%) and DNA damage. Leaf explants were inoculated in media with different 2,4-D concentrations. After 90 days, the friable calli were transferred to the regeneration medium, and the number of normal and abnormal SE was monthly counted. The increase of the 2,4-D concentration increased the number of responsive explants in both Coffea. At 9.06, 18.08 and 36.24 µM 2,4-D, C. arabica presented the highest values of responsive explants, differing from C. canephora. Normal and abnormal SE regeneration increased in relation to the time and 2,4-D concentration. Global 5-mC% varied at different stages of the ISE in both Coffea. Furthermore, the 2,4-D concentration positively correlated with global 5-mC%, and with the mean number of ASE. All ASE of C. arabica and C. canephora exhibited DNA damage and showed higher global 5-mC%. The allotetraploid C. arabica exhibited greater tolerance to the toxic effect of 2,4-D than the diploid C. canephora. We conclude that synthetic 2,4-D auxin promotes genotoxic and phytotoxic disorders and promotes epigenetic changes during Coffea ISE.

The acute effects of fipronil and 2,4-D, individually and in mixture: a threat to the freshwater Calanoida copepod Notodiaptomus iheringi.

The magnitude of copepods' responses to pesticides, individually and in mixture, is little understood. The aims of this study were to evaluate: (i) the effects of the pesticides fipronil and 2,4-D, individually and in mixture, on the freshwater copepod Notodiaptomus iheringi; and (ii) the survival and the feeding rate of copepods after the exposure. Acute toxicity tests using the commercial formulations of fipronil and 2,4-D, individually and in mixture, were performed. The LC10-48h, LC20-48h, and LC50-48h of fipronil to N. iheringi were 2.38 ± 0.48, 3.08 ± 1.14, and 4.97 ± 3.30 µg L-1, respectively. For 2,4-D the LC10-48h, LC20-48h, and LC50-48h were 371.18 ± 29.20, 406.93 ± 53.77, and 478.24 ± 107.77 mg L-1, respectively. Morphological damages on the copepods exposed to pesticides were observed at all concentrations. Fungal filaments covering dead organisms were presented at the treatment highest concentration (R5:7.43 ± 2.78 µg L-1 fipronil). The mixture of the pesticides presented synergistic effects on the mortality of N. iheringi. Post-exposure tests showed no difference between the treatments and the control on the mortality and on the feeding rate for 4 h. However, since delayed toxicity of pesticides can occur, longer post-exposure tests using N. iheringi should be tested. N. iheringi is a key species in the aquatic Brazilian ecosystem and showed sensitivity to fipronil and 2,4-D; thus, more studies with this species assessing other responses are recommended.

Elucidating the effects of acute and chronic exposure to 2,4-Dichlorophenoxyacetic acid on fathead minnow (Pimephales promelas) innate immunity.

Aquatic herbicides, such as 2,4-dichlorophenoxyacetic acid (2,4-D) formulations, are commonly used for invasive species management throughout the United States. Ecologically relevant concentrations of 2,4-D can impair essential behaviors, reduce survival, and act as an endocrine disruptor; however, there is limited knowledge of its effects on the health of non-target organisms. Here, we investigate the acute and chronic exposure impacts of 2,4-D on adult male and female fathead minnow (Pimephales promelas) innate immune function. We exposed both adult male and female fathead minnows to three different ecologically relevant concentrations of 2,4-D (0.00, 0.40, and 4.00 mg/L) and took blood samples at three acute time points (6, 24, and 96 h) and one chronic time point (30 days). We found that male fatheads had higher total white blood cell concentrations when exposed to 2,4-D at the acute time points. For the females, only proportions of specific cell types were altered when exposed to 2,4-D at the acute time points. However, we did not observe any significant impacts of chronic exposure to 2,4-D on any innate immune responses for either males or females. Overall, this study is the first step in answering an important question for game fisheries and management agencies while providing insight to future studies that investigate the impacts of herbicide exposure to freshwater fish health and immunity.

Effects of glyphosate, 2,4-D, chlorantraniliprole, and imidacloprid formulations, separately and in mixtures in Caiman latirostris hatchlings.

The present study demonstrated the potential of glyphosate (GLY), 2,4-dichlorophenoxyacetic acid (2,4-D), imidacloprid (IMI) and chlorantraniliprole (CAP) separately and in mixtures to induce oxidative stress and DNA damage in Caiman latirostris hatchlings. Under controlled condition, an embryonic exposure to these pesticides was done at concentrations recommended for soybean crops. Treatments were: negative control, GLY, 2,4-D, IMI, CAP, mixture 1 (M1): GLY + 2,4-D, M2: IM I + CAP and M3: GLY + 2,4-D + IMI + CAP. At hatching, blood samples were taken for the evaluation of genotoxicity, oxidative damage to lipids and DNA, the enzymatic activity of catalase (CAT) and superoxide dismutase (SOD), and the expression level of their corresponding genes (catalase: cat and superoxide dismutase: sod). It has been shown that IMI, M2 and M3 induced a significant inhibition of CAT activity while no effect was observed on SOD. In turn, lipid peroxidation was significantly higher in individuals exposed to IMI, and to all the mixtures. Besides, genotoxicity and oxidative DNA damage were observed in all exposed groups. The results of mRNA expression showed no difference at transcription levels. In the same way, no alterations in growth parameters were recorded at hatching. Regarding to the mixtures, we observed a potentiating action of IMI on M3 in lipid peroxidation as well as independent action on oxidative DNA damage and genotoxicity parameters. Our results highlight the importance of investigating the effect of pesticides and their mixtures considering the potential consequences to caimans living in natural environments.

A comprehensive study demonstrating the influence of the solvent composition on the phytotoxicity of compounds, as exemplified by 2,4-D-based ILs with a choline-type cation.

BACKGROUND: Growing concern for the protection of the environment and existing ecosystems has resulted in increasing consideration of phytotoxicity tests as valid ecotoxicological indicators of the potential hazards of the use of ionic liquids (ILs) or any other chemical. The objective of this study was to gain a detailed understanding of the influence of the solvent composition of spray solutions on the phytotoxic effect of foliar application of ionic pairs with weak (choline 2,4-dichlorophenoxyacetate, [Chol][2,4-D]), medium (N-hexylcholine 2,4-dichlorophenoxyacetate, [C6 Chol][2,4-D]) and good (N-dodecylcholine 2,4-dichlorophenoxyacetate, [C12 Chol][2,4-D]) surface-active properties. RESULTS: Experimental results unambiguously demonstrated that the biological activity of the test salt solutions, particularly [Chol][2,4-D] and [C6 Chol][2,4-D], can be strongly affected by the addition of an organic solvent, such as methanol, ethanol, dimethylformamide (DMF) or dimethylsulfoxide (DMSO) compared to solutions in pure water. However, the observed tendency is less pronounced for the compound exhibiting good surface activity, [C12 Chol][2,4-D]. CONCLUSIONS: The collected findings show that caution is warranted in the exploitation or modification of methodologies for assessing phytotoxicity to ensure the reliable interpretation of obtained results for environmental risk assessment or building quantitative structure-activity relationship (QSAR) models. © 2023 Society of Chemical Industry.