Comparative study about the effects of pollution on glass and yellow eels (Anguilla anguilla) from the estuaries of Minho, Lima and Douro Rivers (NW Portugal)

Ecotoxicology and Environmental Safety

Volumn 73, Issue 4, 2010, Pages 524-533

  Gravato, Carlos ^a   Guimarães, Laura ^a   Santos, Joana ^a   Faria, Melissa ^a   Alves, Anabela ^a   Guilhermino, Lúcia ^a,b  

^a UNIVERSITY OF PORTO   (Portugal)

^b UNIVERSITY OF PORTO   (Portugal)

Author keywords

Biomarkers; Field populations; Glass and yellow eels; Health condition; Oxidative stress; Pollution

Indexed keywords

ADENOSINE TRIPHOSPHATASE (POTASSIUM SODIUM); ANTIOXIDANT; BIOLOGICAL MARKER; CHOLINESTERASE; GLUTATHIONE TRANSFERASE; LACTATE DEHYDROGENASE;

BIOMARKER; COMPARATIVE STUDY; EEL; ENZYME ACTIVITY; ESTUARINE POLLUTION; MORPHOMETRY; POLLUTION EFFECT;

ANGUILLA ANGUILLA; ARTICLE; BODY HEIGHT; CHEMICAL ANALYSIS; COMPARATIVE STUDY; CONTROLLED STUDY; EEL; ENZYME ACTIVITY; ENZYME INHIBITION; ESTUARY; HEALTH STATUS; LIPID PEROXIDATION; MORPHOMETRICS; NONHUMAN; PORTUGAL; QUALITATIVE RESEARCH; RIVER; SPECIES COMPARISON; WATER POLLUTION; WEIGHT REDUCTION;

ANGUILLA; ANIMALS; ANTIOXIDANTS; BODY SIZE; CHOLINESTERASES; ENVIRONMENTAL MONITORING; FEMALE; GLUTATHIONE TRANSFERASE; HYDRO-LYASES; LIPID PEROXIDATION; MALE; OXIDATIVE STRESS; PORTUGAL; RIVERS; SODIUM-POTASSIUM-EXCHANGING ATPASE; WATER POLLUTION;

DOURO ESTUARY; LIMIA ESTUARY; MINHO ESTUARY; PORTO [PORTUGAL]; PORTUGAL; VIANA DO CASTELO [PORTUGAL];

ANGUILLA ANGUILLA;

EID: 77951652923     PISSN: 01476513     EISSN: 10902414     Source Type: Journal    
DOI: 10.1016/j.ecoenv.2009.11.009     Document Type: Article

References (94)

1. Andersson T., Förlin L., Härdig J., Larsson A. Physiological disturbances in fish living in coastal waters polluted with bleached kraft pulp mill effluents. Can. J. Fish. Aquat. Sci. 1988, 45:1525-1536.
2. Baker M.A., Cerniglia G.J., Zaman A. Microtiter plate assay for the measurement of glutathione and glutathione disulfide in large numbers of biological samples. Anal. Biochem. 1990, 190:360-365.
3. Barata C., Baird D.J., Soares A.M.V.M., Guilhermino L Biochemical factors contributing to response and sensitive clones of Daphnia magna Straus exposed to ethyl parathion. Ecotoxicol. Environ. Saf. 2001, 49:155-163.
4. Bird R.P., Draper A.H. Comparative studies on different methods of malondyhaldehyde determination. Methods Enzymol. 1984, 90:105-110.
5. Bradford M. A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 1976, 72:248-254.
6. Buet A., Banas D., Vollaire Y., Coulet E., Roche H. Biomarker responses in European eel (Anguilla anguilla) exposed to persistent organic pollutants. A field study in the Vaccarès lagoon (Camargue, France). Chemosphere 2006, 65:1846-1858.
7. Cairrão E., Couderchet M., Soares A.M.V.M., Guilhermino L. Glutathione-S-transferase activity of Fucus spp. as a biomarker of environmental contamination. Aquat. Toxicol. 2004, 70:277-286.
8. Cajaraville M.P., Bebianno M.J., Blasco J., Porte C., Sarasquete C., Viarengo A. The use of biomarkers to assess the impact of pollution in coastal environments of the Iberian Peninsula: a practical approach. Sci. Total Environ. 2000, 247:201-212.
9. Castro M., Santos M.M., Monteiro N.M., Vieira N. Measuring lysosomal stability as an effective tool for marine coastal environmental monitoring. Mar. Environ. Res. 2004, 58:741-745.
10. Clairborne A. Catalase activity. CRC Handbook of Methods in Oxygen Radical Research 1985, 283-284. CRC Press, Boca Raton, FL. R.A. Greenwald (Ed.).
11. Colombo G., Grandi G. Sex differentiation in the European eel: histological analysis of effects of sex steroids on the gonad. J. Fish Biol. 1995, 47:394-413.
12. Comoglio L., Amin O., Roque A., Betancourt-Lozano M., Anguas D., Haro B.M. Evaluation of sublethal biomarkers in Litopenaeus vannamei on foodborn exposure to methyl parathion. Ecotoxicol. Environ. Saf. 2005, 62:66-74.
13. Cribb A.E., Leeder J.S., Spielberg S.P. Use of a microplate reader in an assay of glutathione reductase using 5,5'-dithiobis(2-nitrobenzoic acid). Anal. Biochem. 1989, 183:195-196.
14. Cunha I., Hoff P., Van de Vijver K., Guilhermino L., Esmans E., De Coen W. Baseline study of perfluorooctane sulfonate occurrence in mussels, Mytilus galloprovincialis, from north-central Portuguese estuaries. Mar. Pollut. Bull. 2005, 50:1128-1132.
15. Damásio J., Guilhermino L., Soares A.M.V.M., Riva M.C., Barata C. Biochemical mechanisms of resistance in Daphnia magna exposed to the insecticide fenitrothion. Chemosphere 2007, 70:74-82.
16. De Coen W.M., Janssen C.R., Segner H. The use of biomarkers in Daphnia magna toxicity testing V. in vivo alterations in the carbohydrate metabolism of Daphnia magna exposed to sublethal concentrations of mercury and lindane. Ecotoxicol. Environ. Saf. 2001, 48:223-234.
17. Désaunay Y., Guérault D. Seasonal and long-term changes in biometrics of eel larvae: a possible relationship between recruitment variation and North Atlantic ecosystems productivity. J. Fish Biol. 1997, 51:317-339.
18. Diamantino T.C., Almeida E., Soares A.M.V.M., Guilhermino L. Lactate dehydrogenase activity as an effect criterion in toxicity tests with Daphnia magna Straus. Chemosphere 2001, 45:553-560.
19. Doyotte A., Mitchelmore C.L., Rinisz D., McEvoy J., Livingstone D.R., Peters L.D. Hepatic 7-ethoxyresorufin O-deethylase activity in the eel (Anguilla anguilla) from the Thames Estuary and comparisons with other United Kingdom estuaries. Mar. Pollut. Bull. 2001, 42:1313-1322.
20. Ellman G.L., Courtney K.D., Andres V., Feather-Stone R.M. A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem. Pharmacol. 1961, 7:88-95.
21. Fenet H., Casellas C., Bontoux J. Laboratory and field-caging studies on hepatic enzymatic activities in European eel and rainbow trout. Ecotoxicol. Environ. Saf. 1998, 40:137-143.
22. Ferreira M., Antunes P., Gil O., Vale C., Reis-Henriques M.A. Organochlorine contaminants in flounder (Platichthys flesus) and mullet (Mugil cephalus) from Douro estuary, and their use as sentinel species for environment monitoring. Aquat. Toxicol. 2004, 69:347-357.
23. Ferreira, M., Antunes, P., Gil, O., Vale, C., Reis-Henriques, M.A., 2002. Testis-ova in mullet (Mugil cephalus) exposed to organic contaminants in the Douro estuary. In: Keller, R., Dircksen, H., Sedlmeier, D., Vaudry, H. Bonn (Eds), Proceedings of the 21st Conference of European Comparative Endocrinologists, August 26-30, pp. 81-85.
24. Ferreira M., Moradas-Ferreira P., Reis-Henriques M.A. The effect of long-term depuration on phase I and phase II biotransformation in mullets (Mugil cephalus) chronically exposed to pollutants in River Douro Estuary, Portugal. Mar. Environ. Res. 2006, 61:326-338.
25. Feunteun E. Management and restoration of European eel population (Anguilla anguilla): an impossible bargain. Ecol. Eng. 2002, 18:575-591.
26. Filho D.W. Fish antioxidant defences - a comparative approach. Braz. J. Med. Biol. Res. 1996, 29:1735-1742.
27. Filho D.W., Tribess T., Gáspari C., Cláudio F.D., Torres M.A., Magalhães A.R.M. Seasonal changes in antioxidant defenses of the digestive gland of the brown mussel (Perna perna). Aquaculture 2001, 203:149-158.
28. Gravato C., Faria M., Alves A., Santos J., Guilhermino L. Biomonitoring studies performed with European eel populations from the estuaries of Minho, Lima and Douro Rivers (NW Portugal). Advanced Environmental Monitoring 2008, 390-401. Springer, The Netherlands. Y.J. Kim, U. Platt (Eds.).
29. Griffith O.W. Determination of glutathione and glutathione disulfide using glutathione reductase and 2-vinyl-pyridine. Anal. Biochem. 1980, 106:207-212.
30. Guerreiro, N., Pereira, P.B., 2002. Instituto da Água, p. 506.
31. Guilhermino L., Barros P., Silva M.C., Soares A.M.V.M. Should the use of inhibition of cholinesterase as a specific biomarker for organophosphate and carbamate pesticides be questioned?. Biomarkers 1998, 3:157-163.
32. Guilhermino L., Lopes M.C., Carvalho A.P., Soares A.M.V.M. Acetylcholinesterase activity in juveniles of Daphnia magna Straus. Bull. Environ. Contam. Toxicol. 1996, 57:979-985.
33. Guimarães L., Gravato C., Santos J., Monteiro L.S., Guilhermino L. Yellow eel (Anguilla anguilla) development in NW Portuguese estuaries with different contamination levels. Ecotoxicology 2009, 18:385-402.
34. Habig W.H., Pabst M.J., Jakoby W.B. Glutathione-S-transferases, the first enzymatic step in mercapturic acid formation. J. Biol. Chem. 1974, 249:7130-7139.
35. Hoque M.T., Yusoff F.M., Law A.T., Syed M.A. Effect of hydrogen sulphide on liver somatic index and Fulton's condition factor in Mystus nemerus. J. Fish Biol. 1998, 52:23-30.
36. INAG, 2000. Planos das bacias Hidrográficas dos rios Luso-Espanhois-Síntese. Caracterização e Diagnóstico. Instituto da Água, Direcção de serviços de recursos Hídricos, Divisão de Recursos Subterrâneos, 398 pp.
37. Knapen D., Bervoets L., Verheyen E., Blust R. Resistance to water pollution in natural gudgeon (Gobio gobio) populations may be due to genetic adaptation. Aquat. Toxicol. 2004, 67:155-165.
38. Knights B. Risk assessment and management of contamination of eels (Anguilla spp.) by persistent xenobiotic organochlorine compounds. Chem. Ecol. 1997, 13.
39. Laffaille P., Caraguel J.-M., Legault A. Temporal patterns in the upstream migration of European glass eels (Anguilla anguilla) at the Couesnon estuarine dam. Estuarine Coastal Shelf Sci. 2007, 73:81-90.
40. Langston W.J., Chasman B.S., Burt G.R., Pope N.D., McEvoy J. Metallothionein in liver of eels Anguilla anguilla from the Thames estuary: an indicator of environmental quality?. Mar. Environ. Res. 2002, 53:263-293.
41. Lecomte-Finiger R. Growth history and age at recruitment of European glass eels (Anguilla anguilla) as revealed by otolith microstructure. Mar. Biol. 1992, 114:2054-2210.
42. Lemaire P., Livingstone D.R. Pro-oxidant/antioxidant processes and organic xenobiotics interactions in marine organisms, in particular the flounder Platichthys flesus and mussels Mytilus edulis. Trends Comp. Biochem. Physiol. 1993, 1:1119-1150.
43. Levesque H.M., Moon T.W., Campbell P.G.C., Hontela A. Seasonal variation in carbohydrate and lipid metabolism of yellow perch (Perca flavescens) chronically exposed to metals in the field. Aquat. Toxicol. 2002, 60:257-267.
44. Lima I., Moreira S.M., Rendón-Von Osten J., Soares A.M.V.M., Guilhermino L. Biochemical responses of the marine mussel Mytilus galloprovincialis to petrochemical environmental contamination along the North-western coast of Portugal. Chemosphere 2007, 66:1230-1242.
45. Lindström-Seppä P., Pesonen M. Biotransformation enzymes in fish as tools for biomonitoring the aquatic environment. Acta Biol. Hung. 1986, 37:85-95.
46. Livingstone D.R., Garcia Martinez P., Michel X., Narbonne J.F., O'Hara S., Ribera D., Winston G.W. Oxyradical generation as a pollution-mediated mechanism of toxicity in the common mussel, Mytilus edulis L., and other molluscs. Funct. Ecol. 1990, 4:415-424.
47. Livingstone D.R., Mitchelmore C.L., Peters L.D., O'Hara S.C., Shaw J.P., Chesman B.S., Doyotte A., McEvoy J., Ronisz D., Larsson D.G., Forlin L. Development of hepatic CYP1A and blood vitellogenin in eel (Anguilla anguilla) for use as biomarkers in the Thames Estuary, UK. Mar. Environ. Res. 2000, 50:367-371.
48. Lloret J., Gil de Sola L., Souplet A., Galzin R. Effects of large-scale habitat variability on condition of demersal exploited fish in the north-western Mediterranean. ICES J. Mar. Sci. 2002, 59:1215-1227.
49. MacCleave J.D., Brickley P.J., O'Brien K.M., Kistner D.A., Wong M.W., Gallagher M., Watson S.M. Do leptocephali of the European eel swim to reach continental waters? Status of the question. J. Mar. Biol. Assoc. UK 1998, 78:285-306.
50. Marchand J., Quiniou L., Riso R., Thebaut M-T., Laroche J. Physiological cost of tolerance to toxicants in the European flounder Platichthys flesus, along the French Atlantic coast. Aquat. Toxicol. 2004, 70:327-343.
51. McCormick S.D. Methods for non-lethal gill biopsy and measurement of Na+,K+-ATPase activity. Can. J. Fish. Aquat. Sci. 1993, 50:656-658.
52. Mohandas J., Marshall J.J., Duggins G.G., Horvath J.S., Tiller D. Differential distribution of glutathione and glutathione related enzymes in rabbit kidney. Possible implications in analgesic neuropathy. Cancer Res. 1984, 44:5086-5091.
53. Monteiro M., Quintaneiro C., Nogueira A.J., Morgado F., Soares A.M.V.M., Guilhermino L. Impact of chemical exposure on the fish Pomatoschistus microps Kroyer (1838) in estuaries of the Portuguese Northwest coast. Chemosphere 2007, 66:514-522.
54. Moreira S.M., Moreira-Santos M., Guilhermino L., Ribeiro R. An in situ postexposure feeding assay with Carcinus maenas for estuarine sediment-overlying water toxicity evaluations. Environ. Pollut. 2006, 139:318-329.
55. Moreira S.M., Moreira-Santos M., Ribeiro R., Guilhermino L. The 'Coral Bulker' fuel oil spill on the North coast of Portugal: spatial and temporal biomarker responses in Mytilus galloprovincialis. Ecotoxicology 2004, 13:619-630.
56. Mucha A.P., Bordalo A.A., Vasconcelos M.T.S.D. Sediment quality in the Douro river estuary based on trace metal contents, macrobenthic community and elutriate sediment toxicity test (ESTT). J. Environ. Monitor. 2004, 6:585-592.
57. Mucha A.P., Teresa M., Vasconcelos S.D., Bordalo A.A. Vertical distribution of the macrobenthic community and its relationships to trace metals and natural sediment characteristics in the lower Douro estuary, Portugal. Estuarine Coastal Shelf Sci. 2004, 59:663-673.
58. Ohkawa H. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal. Biochem. 1979, 95:351-358.
59. Orbea A., Ortiz-Zarragoitia M., Sole M., Porte C., Cajaraville M.P. Antioxidant enzymes and peroxisome proliferation in relation to contaminant body bordens of PAHs and PCBs in bivalve molluscs, crabs and fish from the Urdaibai and Plentzia estuaries (Bay of Biscay). Aquat. Toxicol. 2002, 58:75-98.
60. Pacheco M., Santos M.A. Induction of EROD activity and genotoxic effects by polycyclic aromatic hydrocarbons and resin acids on the juvenile eel (Anguilla anguilla L.). Ecotoxicol. Environ. Saf. 1997, 38:252-259.
61. Palstra A.P., van Ginneken V.J.T., Murk A.J., van den Thillart G.E. Are dioxin-like contaminants responsible for the eel (Anguilla anguilla) drama?. Naturwissenschaften 2006, 93:145-148.
62. Payne J.F., Mathieu A., Melvin W., Fancey L.L. Acetylcholinesterase, an old biomarker with a newfuture? Field trials in association with two urban rivers and a paper mill in Newfoundland. Mar. Pollut. Bull. 1996, 32:225-231.
63. Peña-Llopis S., Pena J.B., Sancho E., Fernández-Vega C., Ferrando M.D. Glutathione-dependent resistance of the European eel Anguilla anguilla to the herbicide molinate. Chemosphere 2001, 45:671-681.
64. Peters L.D., Doyotte A., Mitchelmore C.L., McEvoy J., Livingstone D.R. Seasonal variation and estradiol-dependent elevation of Thames estuary eel Anguilla anguilla plasma vitellogenin levels and comparisons with other United Kingdom estuaries. Sci. Total Environ. 2001, 279:137-150.
65. Quintaneiro C., Monteiro M., Pastorinho R., Soares A.M.V.M., Nogueira A.J., Morgado F., Guilhermino L. Environmental pollution and natural populations: a biomarkers case study from the Iberian Atlantic coast. Mar. Pollut. Bull. 2006, 52:1406-1413.
66. Quintaneiro C., Querido D., Monteiro M., Guilhermino L., Morgado F., Soares A.M.V.M. Transport and acclimation conditions for the use of an estuarine fish (Pomatoschistus microps) in ecotoxicity bioassays: effects on enzymatic biomarkers. Chemosphere 2008, 71:1803-1808.
67. Regoli F., Pellegrini D., Winston G.W., Gorbi S., Giuliani S., Virno-Lamberti C., Bompadre S. Application of biomarkers for assessing the biological impact of dredged materials in the Mediterranean: the relationship between antioxidant responses and susceptibility to oxidative stress in the red mullet (Mullus barbatus). Mar. Pollut. Bull. 2002, 44:912-922.
68. Regoli F., Principato G. Glutathione, glutathione-dependent and antioxidant enzymes in mussels, Mytilus galloprovincialis, exposed to metals under field and laboratory conditions: implications for the use of biochemical biomarkers. Aquat. Toxicol. 1995, 31:143-164.
69. Regoli F., Winston G.W., Gorbi S., Frenzilli G., Nigro M., Corsi I., Focardi S. Integrating enzymatic responses to organic chemical exposure with total oxyradical absorbing capacity and DNA damage in the European eel Anguilla anguilla. Environ. Toxicol. Chem. 2003, 22:2120-2129.
70. Ribeiro C.A.O., Vollaire Y., Sanchez-Chardi A., Roche H. Bioaccumulation and the effects of organochlorine pesticides, PAH and heavy metals in the Eel (Anguilla anguilla) at the Camargue Nature Reserve, France. Aquat. Toxicol. 2005, 74:53-69.
71. Robinet T., Feunteun E. Sublethal effects of exposure to chemical compounds: a cause for the decline in Atlantic eels?. Ecotoxicology 2002, 11:265-277.
72. Roche H., Buet A., Jonot O., Ramade F. Organochlorine residues in european eel (Anguilla anguilla), crucian carp (Carassius carassius) and catfish (Ictalurus nebulosus) from Vaccarès lagoon (French National Nature reserve of Camargue)-effects on some physiological parameters. Aquat. Toxicol. 2000, 48:443-459.
73. Rodrigues P., Reis-Henriques M.A., Campos J., Santos M.M. Urogenital papilla feminization in male Pomatoschistus minutus from two estuaries in northwestern Iberian Peninsula. Mar. Environ. Res. 2006, 62:258-262.
74. Rodríguez-Fuentes G., Gold-Bouchot G. Environmental monitoring using acetylcholinesterase in vitro. A case study in two Mexican lagoons. Mar. Environ. Res. 2000, 50:357-360.
75. Schlezinger J.J., Stegeman J.J. Induction of cytochrome P450 1A in the American eel by model halogenated and non-halogenated aryl hydrocarbon receptor agonists. Aquat. Toxicol. 2000, 50:375-386.
76. Sheppard K., Simkiss K. The effects of heavy metal ions on Ca2+ ATPase extracted from fish gills. Comp. Biochem. Physiol. B 1978, 61:69-72.
77. Simon J. Age, growth, and condition of European eel (Anguilla anguilla) from six lakes in the River Havel system (Germany). ICES J. Mar. Sci. 2007, 64:1414-1422.
78. +-ATPase, metallothionein and P450 1A1 in dab Limanda limanda in the Germain Bight: indicators of sediment contamination?. Mar. Ecol. Prog. Ser. 1992, 91:105-115.
79. Stagg R.M., Rusin J., Brown F. Na+,K+-ATPase activity in the gills of the flounder (Platichthys flesus) in relation to mercury contamination in the Firth of Forth. Mar. Environ. Res. 1992, 33:255-266.
80. Tesch F.W. Age and growth rates of North Atlantic eel larvae (Anguilla spp.), based on published length data. Helgolander Meeresun. 1998, 52:75-83.
81. Tietze F. Enzymic method for quantitative determination of nanogram amounts of total and oxidized glutathione. Anal. Biochem. 1969, 27:502-522.
82. Torres M.A., Testa C.P., Gáspari C., Masutti M.B., Panitz C.M.N., Curi-Pedrosa R., Almeida E.A., Di Mascio P., Filho D.W. Oxidative stress in the mussel Mytella guyanensis from polluted mangroves on Santa Catarina Island, Brazil. Mar. Pollut. Bull. 2002, 44:923-932.
83. Valavanidis A., Vlahogianni T., Dassenakis M., Scoullos M. Molecular biomarkers of oxidative stress in organisms in relation to toxic environmental pollutants. Ecotoxicol. Environ. Saf. 2006, 64:178-189.
84. van der Oost R., Beyer J., Vermeulen N.P.E. Fish bioaccumulation and biomarkers in environmental risk assessment: a review. Environ. Toxicol. Pharmacol. 2003, 13:57-149.
85. van der Oost R., Goksoyr A., Celander M., Heida H., Vermeulen N.P.E. Biomonitoring of aquatic pollution with feral eel (Anguilla anguilla) II. Biomarkers: pollution-induced biochemical responses. Aquat. Toxicol. 1996, 36:189-222.
86. Vassault A. Lactate dehydrogenase. Methods of Enzymatic Analysis. Enzymes: oxireductases, transferases 1983, 118-126. Academic Press, New York. M.O. Bergmeyer (Ed.).
87. Vieira L.R., Sousa A., Frasco M.F., Lima I., Morgado F., Guilhermino L. Acute effects of Benzo[a]pyrene, anthracene and a fuel oil on biomarkers of the common goby Pomatoschistus microps (Teleostei, Gobiidae). Sci. Total Environ. 2008, 395:87-100.
88. Watson T.A., Beamish F.W.H. Effects of zinc on branchial ATPase activity in vivo in rainbow trout, Salmo gairdneri. Comp. Biochem. Physiol. C 1980, 66:77-82.
89. Watson T.A., Beamish F.W.H. The effects of zinc on branchial adenosine triphosphatase enzymes in vitro from rainbow trout, Salmo gairdneri. Comp. Biochem. Physiol. C 1981, 68:167-173.
90. Wendelaar Bonga S.E. The stress response in fish. Physiol. Rev. 1997, 77:591-625.
91. Wilson J.M., Reis-Santos P., Fonseca A.-V., Antunes J.C., Bouça P.D., Coimbra J. Seasonal changes in ionoregulatory variables of the glass eel Anguilla anguilla following estuarine entry: comparison with resident elvers. J. Fish Biol. 2007, 70:1239-1253.
92. Winston G.W., Di Giulio R.T. Prooxidant and antioxidant mechanisms in aquatic organisms. Aquat. Toxicol. 1991, 19:137-161.
93. Wirgin I., Waldman J.R. Resistance to contaminants in North American fish populations. Mutat. Res. 2004, 552:73-100.
94. Zar J.H. Biostatistical Analysis 1996, Prentice-Hall International, Inc., USA. third ed.