Page 36 - Volume 4, Issue 2, July 2011

Thiram Bioaccumulation in

Mytilus galloprovincialis

65

for all cell types in relation to control. Injec-

tion and/or DMSO does not cause additional

DNA damage as proved by a series of exper-

iments in tissues of

Mytilus edulis

and

Mytilus

galloprovincialis

(data not shown)

Discussion

Even though thiram is toxic to fish (19), bi-

valves (

Unio tumidus

) were able to withstand

0.1 mg/L thiram for 3 days without mortal-

ities (15). Taking into account the apparent

lack of thiram LC50 for bivalves in bibliogra-

phy, the aforementioned concentration was

used as the lowest one which can elicit bio-

chemical responses. This concentration elic-

ited responses in some of

M. galloprovincial-

is

tissues as mentioned later in this section.

Usually, the determination of thiram is

associated with its decomposition to car-

bon disulfide (CS

2

) in acidic medium, fol-

lowed mainly by spectrometry (5, 12, 38)

and head space gas chromatography (1, 34).

The drawback of these methods is that they

are time consuming and they also lack of se-

lectivity or sensitivity Another way to de-

tect DTCs is to employ Liquid Chromatog-

raphy (LC) and capillary electrophoresis (CE)

with UV and/or electrochemical detection.

Recently Blasco

et al.

(10) applied a quan-

titative matrix solid-phase dispersion and

liquid chromatography-atmospheric pres-

sure chemical ionization mass spectrome-

try (LC-APCI-MS) method for the simultane-

ous analysis of DTCs and their degradation

products in plants. Among the compounds

analyzed was thiram which was detected by

both APCI and ESI methods with different

corresponding ions each time. This method

was also applied here and was able to de-

tect concentration-related differences in the

way mussels bioaccumulate thiram.

Even though the concentrations test-

ed were high, only certain signs of morbidi-

ty (profuse mucus secretion and delayed re-

action to stimuli) were observed. Similarly,

high concentrations of agrochemicals have

not produced mortalities in bivalves (37).

Thiram is not considered genotoxic

in

vivo

, despite being Ames positive (19).

In vit-

ro

alkaline filter elution has also produced

positive results (increase in SSB) in rat and

human cells (9). The discrepancy between

in vivo

and

in vitro

results for thiram is fur-

ther mirrored in the experiments conducted

by Villani

et al.

(45), where exposure of mice

to the maximum tolerated dose caused a

borderline increase in SSB in lymphocytes

and no increase in splenocytes. In contrast,

concomitant exposure of human lympho-

cytes to thiram

in vitro

caused a significant

increase in SSB. Our results for

Mytilus gal-

loprovincialis

can therefore mostly relate to

the

in vitro

outcomes of Bjørge

et al.

(9) rath-

er than to the

in vivo

ones of Villani

et al.

(45).

Bivalves possess unique physiology and bio-

chemistry quite different fromhigher organ-

isms. As a result, innocuous substances for

higher organisms may be biologically active

for bivalves and vice versa.

The effect of thiram (SSB increase) was

different in each tissue as shown in Figures

4 and 5 and there was a significant interac-

tion between dose and tissue (Table 2). The

route of exposure (water-spiking) may be an

important factor for gill susceptibility since

gill cells would be the first ones to come into

contact with thiram. The direct contact of

the large surface area of gills with the dilut-

ed pollutants may also contribute to this (3).

This difference may also stem from the cell

subpopulations: whereas the main gill cell

type and the agranular haemocytes of

Myti-

lus galloprovincialis

were sensitive to the

genotoxic effects of benzo[a]pyrene (BaP),

the agranular ones which are present at var-

ied concentrations were relatively refractory

to these effects (44). Regarding the digestive

gland, it is considered to be the main organ

of metabolism of organic xenobiotics and

the main site of biotransformation enzyme

activities (8). As a result a number of reactive

intermediates produced may directly attack

vicinal digestive gland DNA. Furthermore,

the highly condensed chromatin of diges-

tive gland nuclei which creates additional al-

kali labile sites also contributes to % tail DNA

(33). This higher baseline damage may have

masked here the effect of thiram, at least in