Dose-dependent effect of silver nanoparticles (AgNPs) on fertility and survival of Drosophila: An in-vivo study

Abstract

Silver nanoparticles (AgNPs) containing consumer products have been proliferating in the market due to its unique antimicrobial property, however, lack of in-depth knowledge about their potential effect on human health in a longer run is of great concern. Therefore, we investigated dose-dependent in vivo effect of AgNPs using Drosophila as a model system. Drosophila, a genetically tractable organism with distinct developmental stages, short life cycle and significant homology with human serves as an ideal organism to study nanomaterial-mediated toxicity. Our studies suggest that ingestion of AgNPs in Drosophila during adult stage for short and long duration significantly affects egg laying capability along with impaired growth of ovary. Additionally, dietary intake of AgNPs from larval stage has more deleterious effects that result in reduced survival, longevity, ovary size and egg laying capability at a further lower dosage. Interestingly, the trans-generational effect of AgNPs was also observed without feeding progeny with AgNPs, thereby suggesting its impact from previous generation. Our results strongly imply that higher doses of AgNPs and its administration early during development is detrimental to the reproductive health and survival of Drosophila that follows in generations to come without feeding them to AgNPs.

Fig 1. Schematic representation of AgNPs treatment conditions.

Fig 2. Characterization of AgNPs.

(A) An average hydrodynamic size of mono-dispersed sample containing AgNPs is 252.9 nm by DLS measurements (B) TEM image of AgNP_S in suspension on Cu grid are in the range of 20–100 nm and spherical in shape. Majority of these particles fall in the size range of 20-50nm (C) X-ray diffraction measurements displayed the average crystallite size to be ~ 47 nm which is obtained by using Scherrer’s formula. (D) Zeta potential value of -27.4 mv displayed moderate stability of AgNPs.

Fig 3. Dose dependent effect of AgNPs on egg laying and ovary development.

(A) Egg laying capability of flies aged for 3, 10 and 30 days without (control) and with AgNPs at different doses (5mg/L, 25mg/L, 50mg/L and 250mg/L) (B) A notable difference in the size of ovary was observed in the flies aged on control and AgNPs supplemented food for 10 days and (C) 30 days in a dose-dependent manner. Significance was calculated by using an analysis of variance (ANOVA) followed by Tukey-Kramer post hoc test (MSD α_0.05 = 116.87).

Fig 4. AgNPs ingestion during larval stage interferes with the growth and survival of F1 progeny.

(A) Eggs reached to pre-pupal stage (B) pupa formation (C) adult eclosion rate and (D) longevity of F1 flies in P+L+ condition. Comparable results were observed for P-L+ and P+L+ conditions; P-L- represents control. Significance was calculated by using an analysis of variance (ANOVA) followed by Tukey-Kramer MSD post hoc test (MSD α_0.05: Pre-pupal count = 14.62; Pupal count = 27.06; Adult eclosion = 27.06; Longevity = 35.34).

Fig 5. Impact of AgNPs ingestion at larval stage on egg laying and ovary development.

(A) Egg laying capability of F1 female flies reduces substantially at higher dose. (B) Improper growth of ovary in P-L- (control) and P-L+ (at different doses) females. Statistical significance was calculated by using an analysis of variance (ANOVA) followed by Tukey Kramer MSD post hoc test (MSD α_0.05 = 85.4).

Fig 6. Trans-generational effect of AgNPs.

(A) Hatching rate of first instar larvae of F2 generation in the absence of AgNPs. (B) Percentage mortality during 2^nd and 3^rd larval instars and (C) Percentage of F2 adults eclosed. Significance was calculated by analysis of variance (ANOVA) followed by Tukey-Kramer MSD post hoc test (MSD α_0.05: Larval mortality = 27.6; F2 adult eclosion = 12.09).