We conducted a series of experiments to gain more insights about the toxicity of PS-MPs, using the fruit fly D. melanogaster as a model of terrestrial organisms. The results revealed that flies can really differentiate between MPs-treated and non-treated food, and exposure to PS-MPs caused sex-specific toxic effects with male flies were more sensitive to PS-MPs and could not survive after a 24-hour starvation.
The presence of defensive or harmful compounds is one of the factors that influence what and how much animals consume 36,37. In the current study, both male and female flies were able to distinguish and avoid PS-MPs-treated food regardless of PS-MPs concentrations. Moreover, flies fed on sucrose solution spiked with either 0.5 or 0.05 µg/ml of PS-MPs significantly consumed less food compared to flies fed on non-treated sucrose solution (control). Similarly, Gammarus pulex, a freshwater shrimp, avoided a diet containing acrylic microfibers when given an alternative diet containing no microfibers 38 and Achatina fulica, a terrestrial snail, consumed less food when fed on PET microplastic fibers 39. This implies that MPs have a repellent effect, though the mechanism is unknown and needs further investigation. Additionally, the reduction in feeding activity may have long-term effects on the physiology, behavior, and fitness of the flies due to difficulties with the energy budget and the entire metabolism 11,40.
In the current study, chronic exposure of D. melanogaster flies to food media spiked with different concentrations of PS-MPs for 7 days significantly reduced survival of both male and female flies when compared to controls. A similar pattern was observed in nematode worms (Caenorhabditis elegans) after 3 days of exposure to 1.0 mg L− 1 PS, but at the nanoscale 41. In contrast, dialyzed PS spheres of different sizes and concentrations have low mortality rate on D. melanogaster flies 26,42. Furthermore, at comparable dialyzed PS particle exposure concentrations, PS spheres have a low mortality rate in mice 43 and different species of bacteria and algae 44,45. Interestingly, in the current study and in a previous study that demonstrated toxic effects 46, unwashed commercial PS particle suspensions were used, which frequently contain toxic preservatives and surfactants like sodium azide 47. However, when these chemicals were removed through dialysis, mortality rates decreased. This is noteworthy and should be considered when assessing the ecological risk of MPs on living organisms.
Sex-specific responses in Drosophila to some physiological, environmental and ecological stressors are common 28,48,49. Here, we found that male flies were more sensitive to PS-MPs, with all males dying after 14 days when exposed to 0.5 g/L of PS-MPs, whereas 20% of female flies survived to day 20. Furthermore, regardless of the concentration used, nearly all male flies exposed to PS-MPs died after 24 hours starvation. While only female flies fed media containing 0.05 or 0.5 µg/L PS-MPs had significantly fewer fecal pellets. In contrast, when male and female D. melanogaster flies were chronically exposed to PET-MPs, male flies lived longer than females 28. These disparity could be attributed to the type of plastic particle (PS vs. PET) or/and to flies' different metabolic levels 50.
Desiccation resistance may change due to variations in the rate of metabolite consumption in drier environments 51. When we examined male flies died after 24h starvation period, we found their carcasses were very desiccated. This suggests that PS-MPs caused desiccation, which may have contributed to their death after a 24-hour deprivation of food. Sex-specific differences in D. kikkawai desiccation resistance have been reported 48, with female flies having higher desiccation resistance than males, which may be due to higher levels of body water, hemolymph, carbohydrates, and dehydration tolerance in females. The latter is true as female flies' fecal pellet counts considerably decreased in response to greater PS-MP concentrations in the current study, suggesting that female flies alter their physiological and metabolic states to cope with different stressors. More research is needed to gain a better understanding of the mechanisms underlying the observed desiccation caused by PS-MPs in male flies after 24h starvation period.
MPs accumulation in tissues can have a variety of negative consequences. We found fluorescent PS beads in the gut tissues, Malpighian tubules, hemolymph, body cavity of adult D. melanogaster flies. Our findings support earlier studies that showed PS-MPs could be ingested and accumulated in the midgut and hind gut of honeybees as well as transported to the trachea and Malpighian tubules 52,53. Ingested microscopic plastic also have been shown to translocate to the circulatory system of the mussel, Mytilus edulis (L.) 54 and recently have been detected in human blood and human placenta 55,56. Interestingly, after 7 days of exposure, we found PS-MPs in the compound eye and ocelli of flies, which may have an adverse effect on vision ability, as seen in the water flea Daphnia magna after 21 days of exposure to PS-MPs 57. Additionally, it was discovered that PS-MPs modulate retinal function in D. melanogaster flies by changing the receptor potential amplitude and gene expression of the phototransduction pathway at a concentration (50 g/L) 100 times greater than the highest dosage employed in the current investigation 58.
Despite the fact that both male and female flies exposed to PS-MPs excreted a large number of pellets that fell uncontrollably while walking, similar to those seen in patients suffering from diarrhea or urinary incontinence, no observable histopathological effects of PS-MPs on midgut and hind gut tissues were found. However, we did find concentration-dependent effects of PS-MPs on the cellular structure of midgut cells of D. melanogaster flies. Between columnar cells and inside the nucleus, intracellular clefts were seen, along with vacuolated cellular membrane, aberrant basement membrane folding, indentation in the nuclear membrane, matrix lysis, and rupture of mitochondrial cristae. Previous research has found comparable effects in bees that have been chronically exposed to PS-MPs 53. The blue mussel (Mytilus edulis L.) also underwent considerable histological alterations after ingesting high-density polyethylene (HDPE) particles 59. Additionally, comparable outcomes were observed in our earlier investigations, in which D. melanogaster flies and honey bees were repeatedly exposed however, to CdO or /and PbO NPs 30,60. These ultrastructural changes reflect the hallmarks of cell necrosis and apoptosis 61,62. More research is needed to investigate the potential histological and cellular changes that MPs of various shapes and sizes, either alone or in combination with other xenobiotics (metal ions, nanomaterials, and persistent organic pollutants), may induce.