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Abstract Drosophila melanogaster has been used as an in vivo model organism for the study of genetics and development since 100 years ago. Recently, the fruit fly Drosophila was also developed as an in vivo model organism for toxicology studies, in particular, the field of nanotoxicity. The incorporation of nanomaterials into consumer and biomedical products is a cause for concern as nanomaterials are often associated with toxicity in many in vitro studies. In vivo animal studies of the toxicity of nanomaterials with rodents and other mammals are, however, limited due to high operational cost and ethical objections. Hence, Drosophila, a genetically tractable organism with distinct developmental stages and short life cycle, serves as an ideal organism to study nanomaterial-mediated toxicity. This review discusses the basic biology of Drosophila, the toxicity of nanomaterials, as well as how the Drosophila model can be used to study the toxicity of various types of nanomaterials.
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Drosophila melanogaster as a model organism to
study nanotoxicity
Cynthia Ong, Lin-Yue Lanry Yung, Yu Cai, Boon-Huat Bay & Gyeong-Hun Baeg
To cite this article: Cynthia Ong, Lin-Yue Lanry Yung, Yu Cai, Boon-Huat Bay & Gyeong-
Hun Baeg (2015) Drosophila melanogaster as a model organism to study nanotoxicity,
Nanotoxicology, 9:3, 396-403, DOI: 10.3109/17435390.2014.940405
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ISSN: 1743-5390 (print), 1743-5404 (electronic)
Nanotoxicology, 2015; 9(3): 396–403
!2014 Informa UK Ltd. DOI: 10.3109/17435390.2014.940405
Drosophila melanogaster as a model organism to study nanotoxicity
Cynthia Ong
, Lin-Yue Lanry Yung
, Yu Cai
, Boon-Huat Bay
, and Gyeong-Hun Baeg
Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore,
Department of Chemical & Biomolecular
Engineering, Faculty of Engineering, National University of Singapore, Singapore, and
Temasek Life Sciences Laboratory, National University of
Singapore, Singapore
Drosophila melanogaster has been used as an in vivo model organism for the study of genetics
and development since 100 years ago. Recently, the fruit fly Drosophila was also developed
as an in vivo model organism for toxicology studies, in particular, the field of nanotoxicity.
The incorporation of nanomaterials into consumer and biomedical products is a cause for
concern as nanomaterials are often associated with toxicity in many in vitro studies. In vivo
animal studies of the toxicity of nanomaterials with rodents and other mammals are, however,
limited due to high operational cost and ethical objections. Hence, Drosophila, a genetically
tractable organism with distinct developmental stages and short life cycle, serves as an ideal
organism to study nanomaterial-mediated toxicity. This review discusses the basic biology
of Drosophila, the toxicity of nanomaterials, as well as how the Drosophila model can be used
to study the toxicity of various types of nanomaterials.
Drosophila melanogaster,in vivo model
organism, nanomaterials, toxicity
Received 24 February 2014
Revised 18 June 2014
Accepted 19 June 2014
Published online 22 July 2014
Drosophila melanogaster was first introduced by Thomas Hunt
Morgan as a model for research since 100 years ago (Morgan,
1910). From 1910 to 1960s, genetic approaches directed the
research carried out in Drosophila, while genetic concepts
developed during these 50 years led the way to novel discoveries
in the biological systems (Bellen et al., 2010). Subsequently, the
Drosophila was used to study human diseases and therapeutic
strategies (Pandey & Nichols, 2011). Recently, the Drosophila
was also successfully developed as a model organism in
toxicology studies (Bhargav et al., 2008; Coulom & Birman,
2004; Dean, 1985; Hosamani, 2013; Siddique et al., 2013), and
the new term Drosophotoxicology was proposed (Rand, 2010).
Among other compounds, the use of the Drosophila in inorganic
mercury toxicity testing has identified toxicity effects on various
physiological functions of Drosophila and possible signaling
cascades associated with inorganic mercury toxicity, providing a
better understanding of the mechanism mediating inorganic
mercury toxicity (Paula et al., 2012).
In light of the recent success of the Drosophila model in
toxicology studies, there has been an enhanced interest in the use
of Drosophila for the understanding of nanomaterial-mediated
toxicity. Nanomaterial is defined as any material with the size of
1–100 nanometers in one or more dimensions (Powell & Kanarek,
2006b; SCENIR (Scientific Committee on Emerging and Newly
Identified Health Risks)). Their small size and unique properties
have spurred the incorporation of nanomaterials in a myriad of
consumer and biomedical products (Eby et al., 2009; Hemmati
et al., 2009; Lin et al., 2013; Tripp et al., 2007). Despite the
extensive use of nanomaterial today, there is still limited
understanding of nanomaterial-mediated toxicity in vivo. The
Drosophila model presents an interesting alternative in the study
of nanotoxicity. A recent editorial has also encouraged the use
of the fruit fly Drosophila in nanotoxicity and nanomedicine
research in view of the possible novel scientific knowledge and
technological breakthroughs that Drosophila can bring about
(Vecchio, 2014). In this review, we will discuss in greater depth
the toxicity of nanomaterials, the basic biology of Drosophila and
how the Drosophila model was used to study nanotoxicity.
Toxicity of nanomaterials
Engineered nanomaterials can be classified into carbon-based,
polymer-based, silicon, ceramic, metal or metal oxide materials.
The minute size of nanomaterials, which are between 1 and 100
nanometers, brings about a high surface area to volume ratio,
resulting in novel properties that are absent in their bulk form
(Powell & Kanarek, 2006a). The physicochemical properties
such as high conductivity, strong optical scattering properties,
strong absorbance and ease of functionalization give rise to
many nanomaterial-related consumer and biomedical products
(Jang et al., 2010; Lim et al., 2011; Ong et al., 2013; Panahifar
et al., 2013). However, recent in vitro studies have revealed that
nanomaterials could result in toxicity. Toxicity refers to the extent
of a substance to cause harm to an organism and it is closely
related to substance exposure, distribution, metabolism, inter-
action with macromolecules and the toxic end point (Hodgson,
2004). For instance, titanium dioxide nanoparticles (TiO
which are one of the most common nanomaterials used in
consumer products, were found to be cytotoxic and genotoxic in
human epidermal cells (A431) after 48 h and 6 h exposure
to 80 mg/ml of TiO
NPs, respectively (Shukla et al., 2011).
Correspondence: Gyeong-Hun Baeg, Department of Anatomy, Yong Loo
Lin School of Medicine, National University of Singapore, Block MD10,
4 Medical Drive, Singapore 117594, Singapore. Tel: +65 6516 7973. Fax:
+65 67787643. E-mail:
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The increase in reactive oxygen species (ROS) production and
effect on cellular calcium homeostasis were attributed to be the
cause of the nanotoxic effects observed (Shukla et al., 2011;
Simon et al., 2011). Some studies on TiO
NPs toxicity revealed
varying toxicity profile in different crystalline structure of TiO
NPs. TiO
NPs can exist in the anatase form and rutile form in
a mix of 80/20, and anatase was found to result in more ROS
production and toxicity than rutile, especially after UV irradiation
(Petkovic et al., 2011; Sayes et al., 2006). Carbon nanotubes
also caused genotoxicity in human bronchial epithelial cells
and mesothelial cells. Dose-dependent DNA damage in human
bronchial epithelial cells was detected by the comet assay
(Lindberg et al., 2009, 2013). Gold nanoparticles (AuNPs),
which are inert in its bulk form, are cytotoxic as well. AuNPs
induce oxidative stress-mediated genotoxicity in MRC-5 human
lung fibroblast cells. The presence of autophagosomes, which
are concomitantly observed with AuNPs uptake under the
transmission electron microscope and proteomic analysis, indi-
cates that autophagy is induced by nanoparticles and may be a
cellular defense mechanism against oxidative stress (Li et al.,
2008, 2010a, 2011). Alteration in micro RNA expression and
epigenetic processes was also implicated during AuNP exposure
(Ng et al., 2011). Recently, more in vivo studies of nanomaterials
have been performed for a better understanding of nanomaterial-
mediated toxicity. Silver nanoparticles (AgNPs) of size
7.9 ± 0.95 nm were intravenously injected (0.5 mg/kg and
5.0 mg/kg AgNPs) at a single dose into rabbits and monitored
for up to 28 days. AgNP accumulation was observed in liver,
kidney, spleen, lung, brain, testis and thymus, indicating that
AgNPs can be transported through the blood circulation. Notably,
excretion of silver was mainly found in the feces than urine,
suggesting that biliary excretion may be the main mechanism
for the removal of AgNPs from the body (Lee et al., 2013).
On the other hand, anatase TiO
NPs exposure to newborn mice
by intranasal instillation as a single dose (1 mg/g body weight)
on postnatal day 4 or three doses on postnatal days 4, 7 and 10
was shown to induce inflammation and inhibit lung development.
As the study endpoint is at 14 days of age, a longer study period
is required to determine the risk of respiratory problems later
in life after TiO
NPs exposure (Ambalavanan et al., 2013).
Nanotoxicity studies using non-mammalian in vivo models
have also enabled a greater understanding of the toxicity effects
of nanomaterials. In Caenorhabditis elegans (roundworm),
reduced growth rate, decreased lifespan, declined reproduction
and defective embryogenesis were observed after the ingestion of
highly soluble amide-modified single-walled carbon nanotube.
The observed toxicity effects were shown to be resulted from
defective endocytosis, decreased citrate cycle and nuclear trans-
location of DAF-16 transcription factor, indicating the molecular
mechanism underlying the nanotoxicity (Chen et al., 2013).
The Danio rerio (zebrafish) model has also shown to be useful.
Exposure to various concentrations of 62 nm silica nanoparticles
to 4–96 h old embryos can pose adverse effects on the mortality,
hatching rate and larval locomotor activity in a dose-dependent
manner. Malformations of embryos such as pericardial edema,
yolk sac edema, tail and head malformation were also observed by
the treatment of silica nanoparticles (Duan et al., 2013). Together,
these in vitro and in vivo studies have provided important insights
into our understanding of nanomaterial-mediated toxicity.
Biology of D. melanogaster
Drosophila melanogaster is often used to study genetics due to
its simple genetic makeup consisting of only four chromosomes.
The complete Drosophila genome has previously been sequenced
and annotated (Adams et al., 2000). Approximately 13 600 genes
were identified, of which 95% of the genes were encoded on three
of the four chromosomes (Rand, 2010). A systemic analysis
further revealed that 77% of distinct genes related to human
diseases matches Drosophila sequences (Reiter et al., 2001).
Studies have also shown that Drosophila proteins involved in the
regulation of gene expression and metabolism exhibit close
similarity to human counterparts. Furthermore, genomic analysis
revealed a good correspondence of Drosophila biosynthetic
networks of important biochemical pathways to human (Adams
et al., 2000).
Drosophila is easy to maintain, propagate and manipulate.
Flies can be kept in vials and fed on food medium consisting
of cornmeal, glucose, agar and fungicide (Rand, 2010). The
whole life cycle of Drosophila is relatively rapid and takes only
approximately 10–12 days at room temperature. The Drosophila
development is divided into various stages: embryo, larva, pupa
and adult (Figure 1). Eggs are laid on the food, and embryogenesis
takes place within the egg. In less than 24 h, the first instar larva
hatches and begins feeding. This feeding and growth phase
will last for four days. About a 200-fold increase in weight of the
larva is expected during the growth phase and is largely due to
the endoreplication of larval tissues. However, the larval tissues
will not be the part of the adult fly as these tissues are broken
down during metamorphosis in the pupa stage. The imaginal
discs, which are made up of diploid cells of undifferentiated
epithelium, will eventually contribute to the development of adult
fly structures. At the end of third instar stage, the larvae stop
feeding and leave the food in search of an area for pupariation.
During the pupa stage, metamorphosis occurs for four days,
following which adult flies eclose. Adult female flies are normally
larger than adult male flies with females weighing 1.4 mg and
males 0.8 mg. Females are ready to mate in less than 24 h after
eclosion and can lay up to 100 eggs per day. Adult flies live about
two months after eclosion (Pandey & Nichols, 2011; Stocker &
Gallant, 2008).
Drosophila as a model to study the toxicity
of nanomaterials
Despite the increase in the number of in vivo nanomaterial
toxicity studies being carried out today, it is still insufficient
to address important issues in the field of nanotoxicology. Key
questions such as what are the long-term effects of nanomaterial
Figure 1. The whole life cycle of the fruit fly Drosophila is relatively
rapid and takes only approximately 10–12 days at 25 C. The Drosophila
development is divided into various stages: embryo, larva (first instar,
second instar and third instar), pupa and adult.
DOI: 10.3109/17435390.2014.940405 Toxicity study in Drosophila 397
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exposure and what is the exact molecular mechanism underlying
nanomaterial-mediated toxicity remain to be addressed. Many
nanotoxicology studies are often limited to in vitro models as
in vivo models are expensive and difficult to carry out, and often
raise ethical objections.
Drosophila can live up to 40–60 days after eclosion, and thus
nanotoxicity can be assessed at various ages of adult flies.
Furthermore, due to its short lifespan, chronic nanotoxicity studies
can easily be carried out using specific tissues or organs of
subsequent filial generations to examine the effects of nanoma-
terials on genome stability, development, reproduction and
viability (Greenspan, 2004). The various developmental stages
of Drosophila are also essentially good models for toxicology
studies. For instance, at the embryonic stage, developmental
studies regarding cell fate determination, neuronal development
and organogenesis upon exposure to toxins can be carried out
while the wandering third instar larva can be used for develop-
mental, physiological and behavioral studies. Furthermore, the
identification of how toxins affect the imaginal discs during the
late larval through the pupa stage will also be useful to understand
the adverse effects of toxins on endoreplication and morpho-
logical changes from larval to adult stage (Pandey & Nichols,
2011; Stocker & Gallant, 2008). Notably, the anatomical struc-
tures such as the brain, heart, lung, kidney, liver, gut and
reproductive tract of the adult Drosophila are physiologically
very similar to those of humans (Pandey & Nichols, 2011).
For example, the Drosophila fat body functions like the human
liver. The fat body utilizes lipoprotein particles for the transpor-
tation of lipids to peripheral tissues, while lipid metabolism in
Drosophila is regulated by insulin signaling (Canavoso et al.,
2001; Diangelo & Birnbaum, 2009). The respiratory system of
Drosophila, named Drosophila tracheal system, is a branched
network of epithelial tubes that ramifies throughout the body
for the transport of oxygen and other gases. Considering
these physiological similarities of organs between Drosophila
and humans, Drosophila can serve as a suitable model for organ-
specific toxicology studies.
Recommendations by the European Centre for the Validation
of Alternative Methods indicated that D. melanogaster is an
ideal model organism to study nanomaterial-mediated toxicity
(Ahamed et al., 2010). The short life cycle, the distinct
developmental stages, the availability of various tools and
reagents, known genome sequence and the physiological similar-
ity of Drosophila with humans make Drosophila an excellent
in vivo model organism to rapidly test toxicity in whole organism
and elucidate the molecular mechanisms underlying the toxicity.
The most direct method to evaluate nanomaterial toxicity in the
Drosophila model is through the determination of survivorship
after nanomaterial exposure. As nanomaterials can gain entry into
the human body by several ways such as oral, dermal and
inhalation routes, it is essential that such entries are modulated
in the Drosophila model (Li et al., 2010b).
One possible route of nanomaterial exposure to Drosophila
is through ingestion. For instance, nanomaterials can be added
directly to the standard Drosophila food at various concentrations.
Newly enclosed male adult flies were transferred directly to the
food containing AgNPs (20 nm) after 6 h starvation, and survivors
were accounted every 24 h for a period of 10 days. A significant
dose-dependent decrease in survival rate was observed for flies
fed with AgNPs as compared to control flies. However, food
containing similar dose of AgNO
did not affect the survival
rate of flies, suggesting that the toxicity was caused specifically
by AgNPs (Tian et al., 2013). Survivorship can also be
investigated at various developmental stages. When Drosophila
eggs were exposed to AgNP-food, a significant effect on
survivorship was observed during the adult stage. Many of the
resulting larvae failed to pupate, and time to pupation was
delayed. In addition, there was a decrease in the number of flies
leaving the pupa stage and emerging as adult flies compared to
control flies (personal communication). CdSe-ZnS quantum dots
(QDs) are toxic to Drosophila. Ingestion of CdSe-ZnS QDs
caused a strong decline in lifespan when compared to controls.
Lifespan refers to the length of time between eclosion and death.
Notably, different types of coating on nanomaterials can contrib-
ute to different toxicological profile. CdSe-ZnS QDs coated with
poly(maleic anhydride octadecene) and polyethylene glycol were
more toxic than those coated with mercaptoundecanoic acid or
poly(maleic anhydride octadecene) alone (Galeone et al., 2012).
The lifespan of Drosophila can also be used to study how various
metrics can affect the toxicity of nanomaterials, as exemplified
by the finding that the total number of particles but not the
total surface area of ingested citrate-capped AuNPs shorten the
lifespan of Drosophila (Pompa et al., 2011b). Importantly,
however, not all nanomaterials are toxic to Drosophila, indicating
the robustness and reliability of nanotoxicity study in Drosophila.
Organically modified silica nanoparticles (20 nm) and gallium
phosphide nanowires (80 nm) have no significant effects on the
development and viability of larvae and adult Drosophila
(Adolfsson et al., 2013; Barandeh et al., 2012). Likewise,
Gellan Gum-PEI nanocomposites showed no significant effects
on the survivorship of Drosophila (Goyal et al., 2011). Submicron
size insulin-small lipid nanoparticles, developed for insulin
delivery, were also non-toxic to Drosophila even after chronic
exposure from egg to adult, providing important preliminary
evidence of the suitability of insulin-small lipid nanoparticles
for oral insulin delivery in patients (Fangueiro et al., 2013).
Nanomaterials can also be introduced to Drosophila via dry
physical exposure, which is equivalent to dermal exposure to
human. Dry nanomaterial such as carbon black and single-walled
nanotubes were added as a powder to the bottom of sealed glass
vials in the absence of food and water. Nanomaterials were found
to adhere strongly to the surface of the flies, resulting in mortality
in all exposed flies within a few hours. As nanomaterials were
found to partially block spiracle openings in Drosophila, defects
in respiration were considered the primary cause of the mortality
during dry exposure (Lehmann, 2001; Liu et al., 2009).
Nanotoxicity study in Drosophila can also be carried out in the
methodology of inhalation exposure, using a nebulizer-based
method that exposes Drosophila to aerosolized nanoparticles,
which are small enough to enter the spiracle openings of
Drosophila. This study has shown that different sizes and types
of nanoparticles such as FluoSpheres (24, 100 and 210 nm), silver
(20 nm) and CdSe/ZnS (5.7 ± 0.5 nm) can be delivered to the
Drosophila respiratory system (Posgai et al., 2009). Subsequent
toxicity testing such as survivorship can then be studied. Findings
from this mode of exposure can thus be a good preliminary study
to investigate nanomaterial inhalation toxicity in human.
The availability of simple yet versatile ways of administration,
and the reproducibility and specificity of nanomaterial-mediated
toxicity, make Drosophila an excellent model organism to study
nanotoxicity in vivo. However, it is worth to note that relevant
doses of nanomaterials should be administered to investigate the
toxicity of nanomaterials in vivo. Doses that are too low or high
may not yield meaningful conclusions regarding the toxicological
profile and mechanisms of nanomaterial as such concentrations
may never be encountered by humans through occupational or
daily exposure. Furthermore, the method of administration is vital
to consider as different routes of entry of nanomaterials may
induce toxicity in different ways. For instance during inhalation
398 C. Ong et al. Nanotoxicology, 2015; 9(3): 396–403
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exposure of AuNPs in rats, AuNPs was found to accumulate
mostly in the lungs, while intravenously injected AuNPs
were found to be accumulated more in the liver and spleen
(Balasubramanian et al., 2010; Yu et al., 2007). Therefore, the
differential accumulation of AuNPs due to the different routes
of entry may account for varying organ specific toxicity.
Oxidative stress
Numerous in vitro studies showed that increased ROS production
after exposure to nanomaterials is one of the most common
causes of nanomaterial-mediated toxicity (Jaeger et al., 2012;
Ng et al., 2013; Passagne et al., 2012; Pichardo et al., 2012).
Excess ROS may cause damage to proteins, lipids and DNA
in cells, eventually leading to diseases such as cardiovascular
diseases and neurological disorders (Turrens, 2003). Higher doses
of nanomaterials can induce greater oxidative stress. However,
it is often challenging to investigate oxidative stress in an in vivo
mammalian model due to ethical issues of exposing high doses
of nanomaterials to rodents. Drosophila, which raises less ethical
concern, thus can serve as a suitable in vivo model to study
nanomaterial-induced oxidative stress.
Oxidative stress was identified to be the primary cause of
toxicity induced by nanomaterials. Intracellular ROS level in
Drosophila after oral ingestion of various sizes of AuNPs (5, 15,
40 and 80 nm) was measured from the homogenate of the
resulting flies using the 2,7-dichlorofluoresceindiacetate (DCF-
DA) dye. There was a significant increase in ROS level in
Drosophila exposed to AuNPs as compared to unexposed control
flies. However, the different sizes of AuNPs had no effect on ROS
production, indicating that the total surface area of AuNPs is not
an important parameter in inducing oxidative stress (Vecchio
et al., 2012b). Treatment of flies with high doses of amorphous
silica nanoparticles (10 and 100 mg/ml) also caused a time and
dose-dependent increase in oxidative stress in larval midgut,
as demonstrated by flow cytometry using DCF dye and
lipid peroxidation assay that measures malonyl dialdehyde.
Consistently, increased antioxidant activities of superoxide
dismutase (SOD) and catalase, which are thought to be related
to the cellular defense of Drosophila against nanomaterials, were
detected (Pandey et al., 2013). Similar experiments with AgNPs
also revealed the high induction of both oxidative stress and
antioxidant activity in third-instar larvae. Western blotting
analysis using the larval tissue extract showed an increase in
the expression of heat-shock protein (HSP) 70, which plays an
important role in biological stress and is a valuable marker for the
evaluation of the adverse effects of AgNPs (Ahamed et al., 2010).
Ingestion of CdSe/ZnS QDs also showed to increase the
transcription of hsp70 and hsp83 genes in the larva (Brunetti
et al., 2013).
Interestingly, however, when vitamin C or vitamin C palmitate
was added to AgNP-food for ingestion during the larval stage, a
significant increase in survivorship, development and mating
success was observed. This suggests that the antioxidant vitamin
C can suppress the induction of oxidative stress caused by
nanomaterial treatment. In support of this, a short-term 24-hour
exposure of flies to AgNPs with vitamin C was shown to reduce
SOD levels and increase Glutathione levels as compared to flies
exposed to AgNP alone (Posgai et al., 2011). Finally, alumina
nanoparticles decreased the frequency of oscillations in the local
interneurons (LNs) and synchronizations in paired LNs, and these
adverse effects of the nanomaterials on the central nervous system
of Drosophila were thought to be caused by oxidative stress
(Huang et al., 2013). All these studies suggest that increased
levels of oxidative stress are the primary cause of nanomaterial-
mediated toxicity across phyla.
Specific damage to the genetic material (genotoxicity) is often the
cause of various diseases such as cancer and hereditary genetic
disorders. Hence, there is an increased interest in the field of
nanotoxicology to study the effects and interactions of nanoma-
terials with the genome of organisms. Drosophila that was
traditionally used for genomic discovery is an ideal model to
examine genotoxicity of nanomaterials in vivo, as demonstrated
by the pioneering studies of Muller and Auerbach in investigating
the genotoxic effects of X-ray and mustard gas compounds,
respectively (Auerbach, 1947; Muller, 1928). The known genome
sequence, high genetic homology to humans and short lifespan
are also advantageous for the use of Drosophila as a model in
assessing acute and/or chronic genotoxicity.
Exposure of the f lies to 15 nm sodium citrate-capped AuNPs
resulted in DNA fragmentation in the gastrointestinal (GI)
tissue, as demonstrated by the terminal transferase dUTP nick-
end-labeling (TUNEL) assay. A significant higher occurrence of
DNA damage was encountered in AuNP-treated Drosophila
(8%) than the unexposed controls (51%) (Pompa et al.,
2011a). Notably, larger AuNPs (40 and 80 nm) were found to
induce less genotoxic effects as compared to smaller AuNPs (5
and 15 nm), suggesting that smaller AuNPs may enter the tissue of
the GI tract more efficiently than larger AuNPs (Vecchio et al.,
2012b). The p53 is one of the key molecules involved in the
regulation of cellular and genomic integrity, the inhibition of cell
growth and apoptosis (Marcel et al., 2011). Quantitative real-time
PCR expression profiling revealed an increase in p53 gene
expression in Drosophila upon AgNP treatment as compared to
controls, corresponding to the increase in DNA damage observed
in the TUNEL assay (Pompa et al., 2011a; Vecchio et al., 2012b).
Long-term mutagenic effects of AuNPs were investigated through
F0, F1 and F2 generations. While F0 generations were fed with
AuNP-food, F1 and F2 generations were kept in untreated food.
Systematic screening of the phenotype of F1 generation identified
flies with malformed wings, eyes or thorax. The F1 flies with the
malformed phenotype were then crossed with wild-type flies to
obtain F2 generation flies. Interestingly, some F2 flies were
observed to have severely impaired body parts that include
malformed eyes or wings. Therefore, this study suggests that
AuNPs are genotoxic and induce genetic mutations in germline
cells, causing chronic genotoxicity on subsequent generations
(Vecchio et al., 2012a). Genetic damage can also be induced by
AgNPs in Drosophila. The expression of p53 and p38 was found
to be up-regulated upon AgNPs ingestion during the third-instar
larval stage, indicating the effects of AgNPs on DNA integrity and
cell viability (Ahamed et al., 2010). CdSe-ZnS QDs also induce
genotoxicity in Drosophila. More TUNEL-positive nuclei were
observed in hemocytes exposed to the QDs than those of control
flies (Galeone et al., 2012).
Genotoxicity can be investigated using the wing somatic
mutation and recombination test (SMART). The SMART assay
enables to identify somatic recombination and other genomic
aberrations, such as point mutations, deletions and chromo-
somal aberrations (Graf et al., 1984). Genotoxic effects induced
can be observed as an increase in the number of mutant spots,
which are caused by the expression of two recessive markers,
namely, multiple wing hairs or flare-3 on the wings. Third
instar larvae fed with AgNP-food had an increase in occurrence
of small single mutant spots as compared to the negative
control. By contrast, balanced heterozygous flies, which have
abolished somatic recombination, had no difference in the
number of mutant spots, indicating that the genotoxic effects
are due to somatic recombination. On the other hand, silver
nitrate at various concentrations failed to induce genotoxicity,
DOI: 10.3109/17435390.2014.940405 Toxicity study in Drosophila 399
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suggesting that genotoxic effects are due to AgNPs and
not Ag
(Demir et al., 2011). Dose-dependent genotoxic
effects of cobalt nanoparticles were also detected by the
SMART assay in Drosophila (Vales et al., 2013). Similarly,
not all nanomaterials are genotoxic. Gallium phosphide
nanowires, multi-walled carbon nanotubes, zirconium oxide,
aluminum oxide and TiO
failed to induce genotoxicity in
Drosophila (Adolfsson et al., 2013; de Andrade et al., 2014;
Demir et al., 2013).
Drosophila also serves as a model for fecundity testing due to its
rapid life cycle and numerous offsprings (Stocker & Gallant,
2008; Tiwari et al., 2011). In particular, fecundity testing
in toxicology studies is increasingly being investigated in
Drosophila (Gupta et al., 2007; Kislukhin et al., 2012). In the
case of nanomaterials, the effect of AgNPs on fertility was
investigated for a long-term exposure of up to eight filial
generations. Both males and females (parental generation) were
fed with food containing 5 mg/L of AgNP (29 ± 4 nm) for 10
days. F1 progenies were then transferred to new AgNP-food to
obtain the F2 generation. This was repeated till F8 generation.
While no significant changes in the number of F1 flies were
observed, a gradual decrease in the number of eclosed flies was
observed in the subsequent generations. However, fecundity was
found to improve from F4 generation and returned to normal at
F8 generation. Adaptability by decreasing development time and
thus less ingestion of AgNP-food in the larval stage were
suggested as the main reason for the improvement in fecundity
(Panacek et al., 2011). Similar decline in fecundity upon AgNP
or AuNP exposure was also observed in other studies
(Armstrong et al., 2013; Pompa et al., 2011a; Posgai et al.,
2011; Vecchio et al., 2012b). Fecundity can also be measured
by accounting the number of eggs laid by females fed with
AuNP-food during larval stage (Vecchio et al., 2012a). Taken
together, these observations indicate that Drosophila can serve
as an in vivo model to study both short-term and long-term
effects of nanomaterials on fecundity. In Drosophila, anatomical
structures and cellular characteristics of germline stem cells
and their stem cell microenvironments (niches) in male and
female reproductive organs have been well described, and thus
subsequent detailed studies for understanding the molecular
and cellular mechanisms of fecundity can be carried out in
Drosophila (Fuller & Spradling, 2007).
Metabolic defects
The adult Drosophila fed with AgNP-food during larval stage
exhibits defects in cuticle development and melanization.
Drosophila that ingested AgNPs has non-pigmented soft cuticle
as compared to controls (Figure 2) (Armstrong et al., 2013; Gorth
et al., 2011; Key et al., 2011; Panacek et al., 2011; Philbrook
et al., 2011; Posgai et al., 2011). The pigmentation defect is due
to the influence of AgNPs on copper transporters. Excess Ag
results in competitive inhibition of copper uptake at the copper
transporters, causing a depletion of copper in cells. As the copper-
dependent tyrosinase is required for melanin synthesis, its decline
in activity may be the reason for AgNP-induced pigmentation
defect (Armstrong et al., 2013). Indeed, the phenotypic modifi-
cation was first reported by Rapoport in 1939, and the reduction
in body pigmentation was thought to be related to treatment
with silver salts (Stefano, 1943). Hence, more comprehensive
studies are required to better evaluate if pigmentation defects
are due to AgNPs or silver salts. Nonetheless, these suggest
that Drosophila can also be used to study metabolic disorders
caused by nanomaterials.
Conclusion and future directions
The ease of synthesis and manipulation of nanomaterial today has
given rise to the development of a variety of nanomaterial-related
products. However, many in vitro studies have pointed out that
these nanomaterials are potentially cytotoxic and capable of
resulting in oxidative stress, followed by genotoxicity. Several
in vivo studies of nanotoxicity in mammals also provided
supporting evidence of cytotoxicity, but the number of in vivo
studies are still limited due to reasons such as high operational
cost and ethical issues.
Many studies to determine the effects of nanomaterials on the
survivorship, oxidative stress production, genotoxicity and fec-
undity have been carried out in Drosophila, and this Drosophila
model has proven to be a useful for nanotoxicological studies
(Figure 3). There are many advantages of using Drosophila as a
model organism. The genetically tractable organism Drosophila is
easy to maintain in the laboratory and inexpensive to keep and has
a short life cycle and high genetic similarity with humans. The
Drosophila model is advantageous over other non-mammalian
model organisms. In particular, compared with zebrafish and
C. elegans,Drosophila allows nanomaterials to be administered
through various routes including oral ingestion, direct dermal
exposure and respiratory exposure, while the route of nanomater-
ial exposure in zebrafish and C. elegans is mainly through water-
borne or food-exposure, respectively. Drosophila organs and
genes are more homologous to humans than those of C. elegans.
In addition, while the sexual reproducing Drosophila is more
appropriate for the aspect of toxicology study, the hermaphrodite
characteristic of C. elegans makes it unsuitable for the study of
mating behaviors and toxicology effects on the reproductive
systems. Zebrafish is also another attractive model for nanotox-
icology study, but it requires special space and equipment to
maintain and has a relatively long life cycle. Despite the many
benefits of the Drosophila model for the study of nanotoxicology,
there are several limitations to this model. Since insects are very
susceptible to stress, different raising and handling practices
such as temperature, humidity, day/night cycle and density may
result in varying biological effects on flies, possibly confounding
the toxic effects observed in nanomaterials (Matsumoto et al.,
2003). In addition, the effect of nanomaterials on the feeding
pattern and food intake of Drosophila is not well established and
remains challenging to be investigated (Deshpande et al., 2014).
Hence, nanotoxicity effects observed through the ingestion of
Figure 2. Drosophila treated with AgNPs exhibit defects in cuticle
development and melanization. Drosophila exposed to 5 mg/L of AgNPs
via ingestion has unpigmented cuticle (right) as compared to normal
control Drosophila (left).
400 C. Ong et al. Nanotoxicology, 2015; 9(3): 396–403
Downloaded by [NUS National University of Singapore] at 17:20 17 April 2016
nanomaterials may be confounded by the change in the food
consumption pattern of the flies. The stability of various
nanomaterials during the different routes of exposure to
Drosophila remains to be studied. As nanomaterials are affected
by its local environment, the physio-chemical properties of
nanomaterials may differ in different conditions such as fly
food recipes and fly food temperature (Pfeiffer et al., 2014).
Finally, even with a high structural homology and genetic
similarity between Drosophila and human, it still remains
challenging to translate the nanotoxicology findings to effects
of nanomaterial on human health. Nonetheless, nanotoxicological
studies in the Drosophila model can yield important lessons
for understanding nanotoxicity effects in vivo and provide
paradigms for researchers working in mammalian systems.
Fundamental new knowledge obtained about the nanotoxicity
effects on whole organism is expected to advance the general field
of nanotoxicology.
In Drosophila, numerous genetic tools and reagents to identify
genes and/or dissect their function are available. Drosophila
transgenic RNA interference (RNAi) project has generated tens
of thousands transgenic animals with an RNAi hairpin under
UAS/Gal4 system that allows to inhibit certain gene function in a
cell or tissue specific manner in vivo; the Drosophila RNAi
Screening Center ( has generated a set of 21 000
double-stranded RNAs that covers every annotated gene in the
Drosophila genome and has developed high-throughput RNAi
screening of culture cells; a collection of 463 chromosomal
deficiency lines is available; and the Berkeley Drosophila
Genome Project has attempted to disrupt every annotated
Drosophila gene by the insertion of a single transposable element
and now several thousand mutant lines are available. In addition,
numerous in vivo reporter lines that accurately reflects the
activation of specific signaling in living organism. For example,
the ROS reporter expressing green fluorescent protein can be used
to monitor and identify tissues and organs in vivo, which are
susceptible to oxidative stress induced by nanomaterials (Sykiotis
& Bohmann, 2008). These reagents and methodologies allow
the opportunity to design systemic functional genomic approaches
to many cell biological processes caused by nanomaterials, such
as ROS induction and genomic aberration. Finally, Drosophila
can be used as a rapid, reliable, robust and cost-efficient in vivo
model for fast assessing the possible adverse effects of
nanomaterials. Taken together, Drosophila can serve as an
excellent model organism for evaluating and studying nanotoxi-
city in vivo, and thus this tiny animal presents limitless
possibilities in the study of nanotoxicity.
The authors would like to thank Ms Song-Lin Bay for her assistance in
preparing the artwork for the figures used in this article and Ms You Fang
from the Department of Chemical and Biomolecular Engineering,
National University of Singapore, for synthesizing the AgNPs used in
this study.
Declaration of interest
The authors declare that they have no conflict of interest.
This research was funded by the National Research Foundation
(NRF), Prime Minister’s Office, Singapore, under its Campus for
Research Excellence and Technological Enterprise (CREATE) pro-
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... And though humans have 23 pairs of chromosomes, and the fruit fly only has four, the fruit fly still shares over 75% of the genes connected to disease in humans, lending to its role as a useful model for biomedical research (Reiter et al., 2001). The fruit fly also has a life cycle from embryo to adult of only 10-12 days and females can generate upwards of 100 eggs per day, both traits which allow for extremely fast generation time of genetic mutants and transgenic animals for experimentation (Nichols, 2006;Ong et al., 2015). Owing to this ability for translation and ease of experimentation, Drosophila have at times been on the very forefront of discoveries later translated to humans (e.g., factors controlling embryonic development or circadian rhythm) and have garnered six Nobel prizes in Physiology or Medicine involving use of the fruit fly model (Axel, 2004;Baptista et al., 2021;Bargiello et al., 1984;Hardin et al., 1990;Hoffmann, 2011;Huang, 2018;Lewis, 1998;Liu et al., 1992;Morgan, 1916;Muller, 1928;Nichols, 2006;Price et al., 1998;Siwicki et al., 1988;Tolwinski, 2017;Vosshall et al., 1994;Zehring et al., 1984). ...
Abnormal pain affects ~50 million adults nationwide. With many of the current treatment options for chronic pain, such as opioid analgesics, carrying side effects such as the threat for addiction, research into safer and more effective options for chronic pain relief is crucial. Abnormal alterations in nociceptive sensitivity, which is the sensitivity of peripheral sensory neurons that detect noxious stimuli, can underlie, and perpetuate chronic pain. However, much is still unknown about the mechanism of how these abnormal alterations in sensitivity occur. To help elucidate genetic components controlling nociceptive sensitivity, the Drosophila melanogaster larval nociception model has been used to characterize well-conserved pathways through the use of genetic modification and/or ultraviolet (UV) irradiation injury to alter the sensitivity of experimental animals. We have continued to build upon this knowledge to reveal a more complete system for how nociceptive sensitivity can be altered, even without injury, by investigation into the potential roles of other novel genes/signaling pathways including, Arm, a component within the Wnt/Wg signaling pathway. Our findings indicate Arm to be a facilitator in controlling nociceptive sensitivity in the absence of injury, by maintaining baseline sensitivity. In an effort to also explore the mechanisms of the primary nociceptors (nociceptors which directly detect noxious stimuli), we conducted bioinformatic analysis of RNA transcripts derived specifically from the nociceptors of larvae after UV injury. Results from this effort led to the discovery of a downregulation in serine proteases during peak allodynia (when something not normally noxious becomes so) development. Results also led to the hypothesis that upregulated Rgk1 and AnxB11 were involved in recovery of the nociceptor from hyperalgesia. This was supported by the knockdown of Rgk1 and AnxB11 having led to nociceptor hypersensitivity in larvae. And in an effort to move the methodology of our field forward, and because the larval stages of fruit fly development are relatively brief, we developed a methodology that allows longer term experimentation of nociceptive sensitization after injury in adult fruit flies. Ultimately, our research uncovered components involved in nociceptive sensitivity, which will hopefully lead to uncovering better treatment options for abnormal pain in the future.
... Drosophila or fruit flies or vinegar flies are one of the best known genetic system for studying developmental biology, genetics, medicine, human disease, and stem cell research etc. [8] Fruit flies are good model organisms for various studies because they are inexpensive, easy to maintain in the lab, have a simplified genetic architecture, and have a fast generation time, (allowing for fast experiments with large samples). There are many basic processes that are shared between Drosophila and humans. ...
Introduction: Isolation of genomic DNA is an initial step in molecular biology techniques. The quality of isolated DNA depends on procedures and chemicals, as well as source and types of the sample used. Several existing procedures are expensive and time consuming. In this study, we isolated high quality genomic DNA with an inexpensive and least time consuming procedure using Drosophila melanogaster flies, larvae, and pupae. Methods: Drosophila melanogaster samples were collected from pre-cultured bottles, and genomic DNA was extracted using a proposed novel and PCR-ready method from three different pools of flies [PF1, PF2, and PF3], similarly from larvae and pupae [PL1, PL2, PL3, PP1, PP2, and PP3, respectively]. Isolated genomic DNA was subjected to PCR amplification with different dilutions using the COI gene and further amplicons were used for RAPD and DNA sequencing. Results: The high quality of isolated genomic DNA was confirmed by 0.8% agarose gel electrophoresis and the purity and quantity of the DNA isolated from single fly, larva and pupa was similar to the purity and quantity of the DNA isolated using the NucleoSpinR Tissue kit method. Isolated genomic DNA was successfully amplified when the template was diluted in the ratio of 1:10. Further successful RAPD amplification and sequencing analysis of the COI gene confirms the efficiency of the downstream application of the proposed novel method. Conclusion: The present Novel and PCR ready rapid DNA isolation method will be potentially beneficial, and it can be successfully used for quick isolation of high molecular weight DNA from Drosophila flies larvae and pupae for DNA barcoding, identification of new species, genotyping, RAPD analysis, etc. Moreover, it can also be easily scaled up for bulk preparations.
... There are four stages in the life cycle of Drosophila: embryo, larva, pupa, adult. The development time from egg to adult in ideal conditions is 8.5 days (Calap-Quintana et al., 2017;Ong et al., 2015;Pandey and Nichols, 2011;Reiter et al., 2001). The flies used in these studies typically followed a standard Drosophila corn-sucrose yeast medium diet and were reared at around 22 • C with a 12:12 h light/dark cycle. ...
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Exposure to particulate matter (PM) air pollution increases risk of adverse human health effects. As more attention is brought to bear on the problem of PM, traditional mammalian in vivo models struggle to keep up with the risk assessment challenges posed by the countless number of unique PM samples across air sheds with limited or no toxicity information. This review examines the utility of three higher throughput, alternative, in vivo animal models in PM toxicity research: Danio rerio (zebrafish), Caenorhabditis elegans (nematode), and Drosophila melanogaster (fruit fly). These model organisms vary in basic biology, ease of handling, methods of exposure to PM, number and types of available assays, and the degree to which they mirror human biology and responsiveness, among other differences. The use of these models in PM research dates back over a decade, with assessments of the toxicity of various PM sources including traffic-related combustion emissions, wildland fire smoke, and coal fly ash. This article reviews the use of these alternative model organisms in PM toxicity studies, their biology, the various assays developed, endpoints measured, their strengths and limitations, as well as their potential role in PM toxicity assessment and mechanistic research going forward.
... and were transferred to new bottles every 3-4 days. Eggs hatched into flies within 10-12 days as expected from the literature (Ong et al., 2014). The newly hatched flies were stored at 18 • C until they were used. ...
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Copper oxide (CuO) NPs are widely used and subsequently released into terrestrial ecosystems. In the present study, bioaccumulation and effects of CuO NPs and dissolved Cu was examined in the fruit fly Drosophila melanogaster after 7 and 10 days dietary exposure at concentrations ranging between 0.09 and 1.2 mg Cu ml⁻¹ for dissolved Cu and between 0.2 and 11 mg Cu ml⁻¹ for CuO NPs. Both Cu forms were bioaccumulated and affected survival and climbing in flies, but not egg-to-adult development. Dissolved Cu caused higher mortality than CuO NPs (CuO NPs 10-days LC50 was 2 times higher), whereas NPs affected climbing and decreased the number of eggs laid per female, potentially affecting fruit fly population size in terrestrial environments. Thus, the study indicates that CuO NPs might cause effects that are different from dissolved Cu due to differences in the mechanism of uptake or toxicity. Therefore, we need to consider relevant sublethal endpoints when assessing these CuO NPs to ensure that we do not overlook long-term effects consequences.
... However, the database used by either of these in silico studies were generated from cell culture-based experiments, which fail to closely mimic the relevant in vivo situations. As Drosophila melanogaster (fruitfly) and Caenorhabditis elegans (nematode) have been recognized as two powerful model systems that can be used to explore the molecular and cellular basis of ENMs-induced toxicity on both organismal and population scales [59,60]. Because various toxicity endpoints (e.g., mortality, developmental deficits, locomotor performance, lifespan and healthspan, and oxidative stress and gene expression levels) can feasibly be examined in vivo using these two models, we can theoretically establish more informative and comprehensive databases on nanobio interactions and identify stronger predictors for realworld/in vivo nanotoxicity assessments. ...
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Background Silver nanoparticles (AgNPs) are considered a double-edged sword that demonstrates beneficial and harmful effects depending on their dimensions and surface coating types. However, mechanistic understanding of the size- and coating-dependent effects of AgNPs in vitro and in vivo remains elusive. We adopted an in silico decision tree-based knowledge-discovery-in-databases process to prioritize the factors affecting the toxic potential of AgNPs, which included exposure dose, cell type and AgNP type (i.e., size and surface coating), and exposure time. This approach also contributed to effective knowledge integration between cell-based phenomenological observations and in vitro/in vivo mechanistic explorations. Results The consolidated cell viability assessment results were used to create a tree model for generalizing cytotoxic behavior of the four AgNP types: SCS, LCS, SAS, and LAS. The model ranked the toxicity-related parameters in the following order of importance: exposure dose > cell type > particle size > exposure time ≥ surface coating. Mechanistically, larger AgNPs appeared to provoke greater levels of autophagy in vitro, which occurred during the earlier phase of both subcytotoxic and cytotoxic exposures. Furthermore, apoptosis rather than necrosis majorly accounted for compromised cell survival over the above dosage range. Intriguingly, exposure to non-cytotoxic doses of AgNPs induced G2/M cell cycle arrest and senescence instead. At the organismal level, SCS following a single intraperitoneal injection was found more toxic to BALB/c mice as compared to SAS. Both particles could be deposited in various target organs (e.g., spleen, liver, and kidneys). Morphological observation, along with serum biochemical and histological analyses, indicated that AgNPs could produce pancreatic toxicity, apart from leading to hepatic inflammation. Conclusions Our integrated in vitro, in silico, and in vivo study revealed that AgNPs exerted toxicity in dose-, cell/organ type- and particle type-dependent manners. More importantly, a single injection of lethal-dose AgNPs (i.e., SCS and SAS) could incur severe damage to pancreas and raise blood glucose levels at the early phase of exposure.
... AgNPs have been the subject of extensive studies due to their ease of handling and storage in biological uses (Natsuki et al., 2015). Prominent medicinal uses of AgNPs are identified as antibacterial activity (Yadav et al., 2017;Lalitha et al., 2013), anticancer therapy (Jeyaraj et al., 2013;Ong et al., 2015), anti-microbial activity (Prabhu & Poulose, 2012;Durán et al., 2016;Mittal et al., 2013;Rudramurthy et al., 2016;Martins et al., 2014;Hajipour et al., 2012;Patil et al., 2015;Le et al., 2019), anti-fungal therapy (Taware et al., 2020;Tran & Le, 2013), antiviral activities (Kango et al., 2013), wound healing (Galdiero et al., 2014), wound dressing (Chernousova & Epple, 2013), implanted material (Wilkinson et al., 2011), tissue engineering, medical devices, diagnostic applications in biosensing (Rafique et al., 2017), anti-permeability test, and dental preparations (Li et al., 2014). ...
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Nanomaterials have been attracting the attention of many researchers because of their size, high stability , affinity, and selectivity nature. Over the past decades, considerable intensive studies on many metal and metal-oxide nanomaterials have drawn consideration through their significant properties like size, shape, surface mass proportion, and their reactivity; all these properties are fundamental cornerstones for the turn of events and use of nanomaterials and nanoscale gadgets in biomedical areas. There is also a vast scope for a broad range of biofunctional applications such as antibacterial, antiviral, antifungal, antitumor, bioimaging, tissue engineering, biosensors, gene, and drug delivery. The authors review the nature, forms, and synthesis of nanomaterials here, with a thorough biological synthesis assessment. They also address the development of nanoparticles by microorganisms in depth, and this chapter also includes updates on different biological and biomedical applications of these bionanomaterials.
... Insects possess a highly successful immune system that rapidly identifies pathogens and parasites and either kills them directly or immobilises them thus ensuring the survival of the host [1]. A wide range of structural and functional similarities exist between the insect immune response and the innate immune response of mammals [2,3] and, as a result, a wide variety of insects (Galleria mellonella, Drosophila melanogaster, Manduca sexta, Bombyx mori) is now used as in vivo models for assessing microbial virulence or for evaluating the in vivo efficacy and toxicity of antimicrobial compounds [4][5][6][7]. Larvae of the greater wax moth (Galleria mellonella) are widely used in academia and industry and in many cases generate results comparable to those that can be obtained using mammals [8][9][10]. Larvae have the advantage of being inexpensive to purchase and house, easy to manipulate, and being free from the legal and ethical restrictions that hinder vertebrate use [11]. ...
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Larvae of the greater wax moth, Galleria mellonella, are a convenient in vivo model for assessing the activity and toxicity of antimicrobial agents and for studying the immune response to patho-gens and provide results similar to those from mammals. G. mellonella larvae are now widely used in academia and industry and their use can assist in the identification and evaluation of novel antimicrobial agents. Galleria larvae are inexpensive to purchase and house, easy to inocu-late, generate results withing 24-48 hours and their use is not restricted by legal or ethical con-siderations. This review will highlight how Galleria larvae can be used to assess the efficacy of novel antimicrobial therapies (photodynamic therapy, phage therapy, metal-based drugs, tria-zole-amino acid hybrids) and for determining the in vivo toxicity of compounds (e.g., food pre-servatives, ionic liquids) and/or solvents (polysorbate 80). In addition, the disease development processes associated with a variety of pathogens (e.g., Staphylococcus aureus, Listeria monocyto-genes, Aspergillus fumigatus, Madurella mycotomatis) in mammals are also present in Galleria larvae thus providing a simple in vivo model for characterising disease progression. The use of Galleria larvae offers many advantages and can lead to acceleration in the development of novel antimicrobials and may be a prerequisite to mammalian testing.
Nanotechnology has proved a boon as it brings a revolutionary change in the world with the tremendous advancements in the field of science such as engineering, biotechnology, analytical chemistry, and agriculture. The synthesis of the metal nanoparticle is a developing area of exploration in present-day material science and innovation. Among the techniques involved in the synthesis of metallic nanoparticles, the biological methods or the green synthesis are reliable due to its cost-effective, environment friendly, and use of non-harmful materials in the processing. Therefore, the current work is focused on the biosynthesis of silver nanoparticles with the leaf extract of holy plant Aegle marmelos by utilizing 1mM silver nitrate solution. From the bio-reduction reaction mixture, the synthesis of silver nanoparticles was studied by UV-Visible spectrophotometer at the range of 200 nm to 800nm and discovered maximum absorbance at 460nm. Further, the SNPs were analyzed for the protein or other organic groups by Fourier Transformed Infrared spectrophotometer (FTIR). The dried silver nanoparticles were further characterized by utilizing a scanning electron microscope (SEM) to observe the actual size, shape, and distribution of particles. This result showed the actual size of the bioengineered silver nanoparticles ranges from 25-30nm. The EDX result showed a peak of Ag that confirmed its presence in the suspension. The main objective of this study is to develop a fast, environmentally friendly, and advantageous technique for the synthesis of silver nanoparticles. By standardizing the biosynthesis techniques, silver nanoparticles can be synthesized in bulk with a safe environment and it may be utilized for different clinical applications including control of pathogenic microorganisms and so on.
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In recent decades there has been growing interest of material chemists in the successful development of functional materials for drug delivery, tissue engineering, imaging, diagnosis, theranostic, and other biomedical applications with advanced nanotechnology tools. The efficacy and safety of functional materials are determined by their pharmacological, toxicological, and immunogenic effects. It is essential to consider all degradation pathways of functional materials and to assess plausible intermediates and final products for quality control. This review provides a brief insight into chemical degradation mechanisms of functional materials like oxidation, photodegradation, and physical and enzymatic degradation. The intermediates and products of degradation were confirmed with analytical methods such as proton nuclear magnetic resonance (1H NMR), gel permeation chromatography (GPC), UV-vis spectroscopy (UV-vis), infrared spectroscopy (IR), differential scanning calorimetry (DSC), mass spectroscopy, and other sophisticated analytical methods. These analytical methods are also used for regulatory, quality control, and stability purposes in industry. The assessment of degradation is important to predetermine the behavior of functional materials in specific storage conditions and can be relevant to their behavior during in vivo applications. Another important aspect is the evaluation of the toxicity of functional materials. Toxicity can be accessed with various methods using in vitro, in vivo, ex vivo, and in silico models. In vitro cell culture methods are used to determine mitochondrial damage, reactive oxygen species, stress responses, and cellular toxicity. In vitro cellular toxicity can be measured by MTT assay, LDH leakage assay, and hemolysis. In vivo studies are performed using various animal models involving zebrafish, rodents (mice and rats), and nonhuman primates. Ex vivo studies are also used for efficacy and toxicity determinations of functional materials like ex vivo potency assay and precision-cut liver slice (PCLS) models. The in silico tools with computational simulations like quantitative structure-activity relationships (QSAR), pharmacokinetics (PK) and pharmacodynamics (PD), dose and time response, and quantitative cationic-activity relationships ((Q)CARs) are used for prediction of the toxicity of functional materials. In this review, we studied the principle methods used for degradation studies, different degradation pathways, and mechanisms of functional material degradation with prototype examples. We discuss toxicity assessments with different toxicity approaches used for estimation of the safety and efficacy of functional materials.
Zinc oxide (ZnO) finds numerous applications in the field of cosmetics, biomedical and food industry. Often ZnO in cosmetics is a mixture of ZnO nanoparticles (ZnO NP) and microparticles (MP). Hence along with ZnO NPs, safety of ZnO MPs is an important aspect of cosmetics and skin care products. Present study is the first report of the ZnO MP mediated effects on Drosophila melanin pathway and its constituent amines. Results of the study indicate that ZnO MP does not reduce the synthesis of melanin resulting in normal pigmentation of the flies. Biochemical estimation of melanin, tyrosine and dopa revealed absence of any adverse effect on their levels confirming the safety of ZnO MP for the melanin pathway and its components. Moreover phenol oxidase enzyme which is crucial for both melanization and immunity was found to be functioning normally upon exposure to ZnO MPs. All these results are indicator that ZnO MP is a safe candidate for topical and oral administration as far as melanin pathway of Drosophila is concerned.
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In recent years, the advent of nanomaterial use has increased exposure rates and raised health concerns. However, the toxicology profiles of many nanomaterials are far from complete for various reasons. In this study, Drosophila melanogaster, commonly called fruit flies, were exposed to one of the most widely used nanomaterials, silver nanopowder (Ag NP), to assess its toxicity and determine if D. melanogaster would be a good model organism for nanotoxicology studies. Comparison of developmental progression amongst groups of flies ingesting different Ag NP concentrations (0.05%/~90 ppm-5.0%/~9000 ppm), revealed that hatch rates were unaffected, but that larval progression was impeded at any dosage of Ag NP. At 0.3% Ag NP an approximate LD50 was observed. Additionally, a distinctive phenotype was observed among emergent adults (F1 generation) that arose from larvae exposed to Ag NP which included reduced body pigmentation accompanied by shortened life span and abnormal climbing behavior. The phenotype prompted speculation that Ag NPs may affect the dopamine and/or the stress response pathway(s).
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The physico-chemical properties of colloidal nanoparticles (NPs) are influenced by their local environment, as, in turn, the local environment influences the physico-chemical properties of the NPs. In other words, the local environment around NPs has a profound impact on the NPs, and it is different from bulk due to interaction with the NP surface. So far, this important effect has not been addressed in a comprehensive way in the literature. The vicinity of NPs can be sensitively influenced by local ions and ligands, with effects already occurring at extremely low concentrations. NPs in the Hückel regime are more sensitive to fluctuations in the ionic environment, because of a larger Debye length. The local ion concentration hereby affects the colloidal stability of the NPs, as it is different from bulk owing to Debye Hückel screening caused by the charge of the NPs. This can have subtle effects, now caused by the environment to the performance of the NP, such as for example a buffering effect caused by surface reaction on ultrapure ligand-free nanogold, a size quenching effect in the presence of specific ions and a significant impact on fluorophore-labelled NPs acting as ion sensors. Thus, the aim of this review is to clarify and give an unifying view of the complex interplay between the NP's surface with their nanoenvironment.
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Food intake is a fundamental parameter in animal studies. Despite the prevalent use of Drosophila in laboratory research, precise measurements of food intake remain challenging in this model organism. Here, we compare several common Drosophila feeding assays: the capillary feeder (CAFE), food labeling with a radioactive tracer or colorimetric dye and observations of proboscis extension (PE). We show that the CAFE and radioisotope labeling provide the most consistent results, have the highest sensitivity and can resolve differences in feeding that dye labeling and PE fail to distinguish. We conclude that performing the radiolabeling and CAFE assays in parallel is currently the best approach for quantifying Drosophila food intake. Understanding the strengths and limitations of methods for measuring food intake will greatly advance Drosophila studies of nutrition, behavior and disease.
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Graphene, a two-dimensional carbon sheet with single-atom thickness, have attracted the scientific world for its potential applications in various field including the biomedical areas. In the present study the graphene copper nanocomposite (GCNC) was synthesized, characterized and evaluated for its toxic potential on third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg(9) . The synthesized GCNC was analyzed by X-ray diffraction (XRD), scanning/transmission electron microscopy (SEM/TEM), atomic force microscopy (AFM), and fourier transform infrared spectroscopy (FTIR). The GCNC in 0.1% DMSO was sonicated for 10 min and the final concentration of 0.033, 0.099, 0.199 and 3.996 µg/µl of diet were established. The third instar larvae were allowed to feed on it separately for 24 and 48 hrs. The hsp70 expression was measured by O-nitrophenyl-β-D-galactopyranoside assay, tissue damage by trypan blue exclusion test and β-galactosidase activity was monitored by in situ histochemical β-galactosidase staining. Oxidative stress was monitored by performing lipid peroxidation assay and total protein estimation. Ethidium bromide/acridine orange staining was performed on midgut cells for apoptotic index and the comet assay was performed for the DNA damage. The results of the present study showed that the exposure of 0.199 and 3.996 µg/µl of GCNC were toxic for 24 hr of exposure and for 48 hr of exposure: 0.099, 0.199 and 3.996 µg/µl of GCNC was toxic. The dose of 0.033 µg/µl of GCNC showed no toxic effects on its exposure to the third instar larvae for 24 hr as well as 48 hrs. This dose can be considered as No Observed Adverse Effect Level (NOAEL).
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Gold nanoparticles (AuNPs) have diverse applications in the biomedical industry such as in diagnosis, labeling, delivering and sensing. Despite their prevalent medical use, nanotoxicity induced by AuNPs is still largely unknown. We have previously shown that AuNPs could exert cytotoxic effects on lung fibroblasts. In this study, we investigated the in vitro toxicological effects of AuNPs in small airway epithelial cells (SAECs) which are the first cells of contact for inhaled NPs and compared expression of metallothionein (MT), a reactive oxygen species scavenger, in SAECs and lung fibroblasts in vitro. Transmission electron microscopy (TEM) and energy-dispersive X-ray (EDX) spectroscopy study revealed cellular uptake of aggregates of AuNPs into the cytoplasm at the ultrastructural level. A significant increase in lipid peroxide as well as substantial DNA damage and cytotoxicity was observed in AuNP-treated cells. For MT expression, AuNPs induced down-regulation of the MT-1X isoform in SAECs, but up-regulation of the MT-1X and MT-2 A isoforms in MRC5 lung fibroblasts. The present study suggests that AuNPs could induce oxidative stress-related cytotoxicity and genotoxicity in SAECs.
Drosophila melanogaster has long been a prime model organism for developmental biologists. During their work, they have established a large collection of techniques and reagents. This in turn has made fruit flies an attractive system for many other biomedical researchers who have otherwise no background in fly biology. This review intends to help Drosophila neophytes in setting up a fly lab. It briefly introduces the biological properties of fruit flies, describes the minimal equipment required for working with flies, and offers some basic advice for maintaining fly lines and setting up and analyzing experiments.
Abstract Drosophila was the most important model organism used in the fields of medicine and biology over the last century. Recently, Drosophila was successfully used in several studies in the field of nanotoxicology. However, only a part of its potential has been exploited in this field until now. In fact, apart from macroscopic observations of the effect due to the interaction between nanomaterials and living organism (i.e. lifespan, fertility, phenotypic aberrations, etc.), Drosophila has the potential to be a very useful tool to deeply analyze the molecular pathways involved in response to the interactions at nano-bio level. The aim of this editorial is to encourage the use of Drosophila by the different research groups working in the fields of nanotoxicology and nanomedicine, in order to define the effects induced by nanomaterials at molecular level for their subsequent exploitation in the field of nanomedicine.