Biology Reference
In-Depth Information
trout). The results of these experiments were in line with those reported in earlier studies
(Longwell et al. 1992; Hose 1994; Anderson and Wild 1994) that described links between
exposure to genotoxic agents and deleterious consequences on progeny.
Gametes were also analyzed in the amphipod
Gammarus fossarum
, males and females,
encaged on both sides of a pollution source. DNA damage was higher in spermatozoa
and ova than in hemocytes at the polluted sites (Lacaze et al. 2011). Application of the
comet assay to spermatozoa of this species required the time of lysis to be extended to 18 h
before electrophoresis, as a consequence of a high degree of compaction of sperm chroma-
tin (Lacaze et al. 2010). Such a high compaction rate is disturbing and possibly constitutes
one mechanism of protection of the genetic material.
The comet assay has been often used in multibiomarker approaches in field studies to
determine:
• The relationships between genotoxicity effects and pollution gradient as pio-
neered by Large et al. (2002) and Shaw et al. (2002)
• The inluence of seasonal parameters, duration of exposure, delay for occurrence
of lesions, and sensitivity of transplanted and native species
• The sensitivity of the comet assay compared to other genotoxicity tests
Mollusks, including bivalves such as
Mytilus
sp. and
D. polymorpha
, have often been used
in biomonitoring marine/estuarine and freshwater, respectively. In transfer experiments
using bivalves, fluctuations in DNA breaks were observed in the first 10 days (Steinert et
al. 1998a; Regoli et al. 2004). Therefore, a very short transplantation experiment is not rec-
ommended. DNA breaks in gill cells of transplanted
M. galloprovincialis
reached the lev-
els of native species within 30 days of transfer, whereas micronuclei measures remained
lower than in native species (Nigro et al. 2006). Seasonal, temporal, and intrasite varia-
tions had been registered (Nigro et al. 2006; Le Goff et al. 2006; Rocher et al. 2006; Rank
et al. 2007; Baussant et al. 2009), which underlined the importance of standardizing and
defining precise deployment and sampling protocols in field studies. DNA damage was
generally lower in winter than in summer (Shaw et al. 2000) and in subtidal mussels
compared to intertidal mussels from the same site. This may be due to sunlight and pho-
toactivation of organic pollutants, such as PAHs (Steinert et al. 1998b).
The occurrence of adaptation mechanisms in chronically exposed populations through
up-regulation of DNA repair mechanisms has been suggested. This was supported by
results in native (Shaw et al. 2002; Noventa et al. 2011) and transplanted invertebrates
(Large et al. 2002), and also in fish (McFarland et al. 1999).
Among other prospects of the assay, it is worth mentioning its usefulness to follow DNA
repair (Lemière et al. 2005b), and to evaluate the time-course variations in DNA dam-
age in the field and transplantation experiments (Frenzilli et al. 2009). It has been used to
explore the genetic mechanisms of damage involving alterations of oncogenes and anti-
oncogenes induced by oxidative stress (Mai et al. 2010). Specific damage, such as oxida-
tive injury and pyrimidine dimerization, can be highlighted using DNA repair enzymes
(Figure 13.1). The comet assay offers the possibility of analyzing epigenetic mechanisms of
toxicity in relation to gene methylation. DNA methylation plays an important role in gene
regulation and maintaining genome stability. Hypermethylation of cytosine may lead to
aberrant silencing of crucial genes, such as DNA repair genes and genes involved in cell
cycle control. Wentzel et al. (2010) coupled the comet assay with use of different restriction
endonucleases to assess the general DNA methylation status. The endonucleases used are
Search WWH ::
Custom Search