Detection of DNA damage in haemocytes of zebra mussel using comet assay

Pavlica, Mirjana; Klobučar, Göran; Mojas, Nina; Erben, Radovan; Papeš, Dražena

doi:10.1016/s1383-5718(00)00162-5

Cited by 127 publications

(52 citation statements)

References 28 publications

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“…Bivalves are widely distributed sessile filter feeders which are good bioindicators of aquatic pollutants (Venier et al, 1997;López-Barea and Pueyo, 1998;Le Pennec and Le Pennec, 2001) and have often been used as test organisms in studies of aquatic pollution (Wilson et al, 1998;Narbonne et al, 1999;Pavlica et al, 2001;Pruski and Dixon, 2002).…”

Section: Introductionmentioning

confidence: 99%

“…The comet assay has been utilized to determine the ability of aquatic contaminants to induce DNA damage but its applicability has been the subject of intense investigation (Pavlica et al, 2001). The comet assay derives its name from the comet-like shape that cells with damaged DNA acquire due to the migration of broken DNA fragments during electrophoresis, the size of the comet tail reflecting the extent of DNA damage.…”

Section: Introductionmentioning

confidence: 99%

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Comet assay comparison of different Corbicula fluminea (Mollusca) tissues for the detection of genotoxicity

2005

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The comet assay was used to study the sensitivity of the widely distributed freshwater bivalve mollusk Corbicula fluminea to the DNA-damaging alkylating-agent methylmethane sulfonate (MMS). This study was undertaken to ascertain if C. fluminea is a good bioindicator of pollutants in aquatic environments and identify which C. fluminea tissue is most effective and practical for genotoxicity studies. The mollusks were exposed to 0.6, 1.2 or 2.4 X 10 -4 M MMS for 40 min and their hemolymph, gill tissue and digestive gland tissue assessed for the level of DNA damage and the time needed for the tissues to recovery. Regression analysis showed a direct linear dose-response relationship between MMS concentration and the number of damaged cells for hemolymph and digestive gland tissue but a quadratic relationship for gill tissue, which made the interpretation the gill tissue results difficult. The basal level of DNA damage to gill tissue was very high, possibly because gill is the organs most directly exposed to environmental toxins and mutagenic agents. Although all three types of tissue produced useful results, hemolymph and digestive gland tissue produced more reproducible and reliable results. Hemolymph was the best sample type in that it was easy to obtain and handle, while gill tissue required more manipulation to obtain cell suspensions. Our results indicate that C. fluminea is an optimal bioindicator for the determination genotoxic contaminants in aquatic environments.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Comet assay comparison of different Corbicula fluminea (Mollusca) tissues for the detection of genotoxicity

2005

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“…Occupational exposure to xenobiotics might have resulted in their covalent binding to DNA, which might lead to chromosome alterations and could be an initial event in the process of chemical carcinogenesis (Fairbairn et al, 1995;Shah et al, 1997). Comparably fewer studies have been conducted with aquatic invertebrates and these have been restricted to bivalve species such as the marine mussel (Mytilus edulis) (Accomando et al, 1999), the zebra mussel (Dreissena polymorpha) (Pavlica et al, 2001), Mediterranean mussel (Mytilus galloprovincialis) (Frenzilli et al, 2001) and oyster (Crassostrea americanus) (Nacci et al, 1996). This assay has been carried out in fishes such as bullhead (Ameriurus nebulosus) Common carp (Cyprinus carpio) (Pandrangi et al, 1995), brown trout (Salmo trutta) (Belpaeme et al, 1996), flounder (Pleuronectes americanus) (Nacci et al, 1996), rainbow trout (Oncorhynchus mykiss) (Devaux et al, 1999), butterfish (Pholis gunnellus) (Bombail et al, 2001), zebra fish (Danio rerio) (Schnurstein and Braunbeak, 2001) tilapia (Tilapia mossambica) (Banu et al 2001) and Channa punctatus (Pandey et al, 2006) etc.…”

Section: Comet Assaysmentioning

confidence: 99%

Genotoxicity Testing in Pesticide Safety Evaluation

Mohanty¹,

Mohanty²,

Jena³

et al. 2011

Pesticides in the Modern World - Pests Control and Pesticides Exposure and Toxicity Assessment

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“…Trabajos in vitro e in vivo realizados en células de tejido branquial, hemocitos y hepatocitos de individuos del género Mytilus, reportan el efecto tóxico de compuestos como ciclofosfamida, peróxido de hidrógeno (H 2 O 2 ), cobre (Majone et al, 1988) y benzo(a)pireno (Dolcetti & Venier, 2002); observándose cambios en las frecuencias de MN relacionados con la concentración y tiempos de exposición, así como lesiones ocurridas a nivel de cadenas simples o dobles del DNA evaluadas mediante modificaciones de la técnica del Ensayo Cometa (Pavlica et al, 2001;Venier et al, 1997). Sin embargo, pocos son los estudios que evalúan la interacción de los cambios ambientales y la presencia de contaminantes con la biología del organismo, factores que estarían produciendo alteraciones en el ciclo celular y en la organización del DNA (Bascomb, 1982;Anitha et al, 2000).…”

Section: Introductionunclassified

Aplicación de dos biomarcadores para el análisis de lesiones en el DNA de bivalvos marinos

et al. 2006

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ResumenEl presente trabajo describe la estandarización de las técnicas del Test de Micronúcleo (MN) y Ensayo Cometa para la evaluación del daño en el DNA de bivalvos marinos. La importancia de estas técnicas radica en que permiten evaluar la respuesta temprana al estrés ocasionado por agentes contaminantes y cambios ambientales. Tejidos branquiales de Semimytilus algosus y Aulacomya ater fueron utilizados para tratamientos in vivo e in vitro. Los tejidos fueron expuestos a los agentes mutagénicos Mitomicina C en concentraciones de 0,02; 0,04 y 0,06 x 10 -6 M y H 2 O 2 en una concentración de 100 μM con tiempos de evaluación de 1, 3, 6, 24 y 48 h. Diferentes soluciones para la colecta del tejido y aislamiento celular fueron evaluados obteniendo una mayor viabilidad celular en la solución salina CMFS, pH 7,3; en frío. El Test de Micronúcleo se modificó en la forma de obtener el disgregado celular, realizando la hipotonización en Citrato de sodio 0,9%, fijación en metanol por segundos posterior al frotis y la tinción con Giemsa 2%. La estandarización del Ensayo Cometa Alcalino, descrita por Wilson et al. (1998), se realizó modificando la suspensión celular en agarosa LMP (1% en solución Kenny, pH 7,5), la exposición a solución de lisis (pH 10 con DMSO y Tritón) con variaciones en el tiempo de 1 a 16 h, y la electroforesis en solución alcalina pH 13,7 y tinción con Bromuro de etidio y Nitrato de plata para geles de agarosa. Los cambios realizados en ambas técnicas permitieron obtener un alto número celular con morfología definida, observándose macrolesiones como la formación de MN y aberraciones nucleares, y la determinación cualitativa de microlesiones observadas por fragmentación y migración del DNA. Palabras claves: daño del DNA, micronúcleo, Ensayo Cometa alcalino, estrés, bivalvos AbstractThis work describes the standardization of Micronucleus Assay (MN) and Comet Assay techniques used for the evaluation of the DNA damage on marine bivalves. These techniques are important because they can evaluate the early stress response caused by pollutant agents and environmental changes. Branchial tissues of Semimytilus algosus and Aulacomya ater were used for in vivo and in vitro treatments. Tissues were exposed to the mutagenetic agents Mitomicine C in concentrations of 0,02; 0,04 and 0,06 x 10 -6 M and H 2 O 2 in concentration of 100 μM evaluated at 1, 3, 6, 24 and 48 h. Different solutions for the tissue collection and cell isolation were evaluated, obtaining more cell viability with cold CMFS pH 7,3 saline solution. Micronucleus Assay were modified in the obtaining of suspension cell, we used 0,9% Sodium citrate hipotonic solution, fixation in methanol for seconds after the spread and dyed with 2% Giemsa. The Alkaline Comet Assay protocol described by Wilson et al. (1998) was standardized modifying the single-cell suspension embedded in LMP Agarose (1% in Kenny solution, pH 7,5), the exposure at lysing solution (pH 10 with DMSO and Triton) with changes from 1 to 16 h, electrophoresis in alkaline solution pH 13,7 and d...

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Detection of DNA damage in haemocytes of zebra mussel using comet assay

Cited by 127 publications

References 28 publications

Comet assay comparison of different Corbicula fluminea (Mollusca) tissues for the detection of genotoxicity

Comet assay comparison of different Corbicula fluminea (Mollusca) tissues for the detection of genotoxicity

Genotoxicity Testing in Pesticide Safety Evaluation

Aplicación de dos biomarcadores para el análisis de lesiones en el DNA de bivalvos marinos

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