Techniques in Aquatic Toxicology, Volume 2

Front cover......Page 1
Dedication......Page 6
Preface......Page 8
About the Editor......Page 10
Contributors......Page 12
Contents......Page 18
section one......Page 22
Introduction......Page 24
General zebra fish maintenance......Page 26
Cellular energy reserves......Page 27
Condition indices......Page 29
Reproduction......Page 30
Results and discussion......Page 31
References......Page 36
Introduction......Page 40
Materials required......Page 41
Fish......Page 43
Contaminant exposure and pathogen challenge......Page 44
Infection of salmon with L. anguillarum......Page 45
Confirmation of pathogen-induced mortality......Page 48
Results and discussion......Page 49
References......Page 57
Enhanced frog embryo teratogenesis assay: Xenopus modelusingXenopustropicalis......Page 60
Introduction......Page 61
Reagents......Page 62
Adult husbandry......Page 65
Test design......Page 66
FETAX screen procedure for aqueous solutions......Page 67
FETAX definitive procedure with MAS for aqueous solutions......Page 68
Control results......Page 69
Copper......Page 71
Bermuda pond sediment extract (complex mixture)......Page 72
Conclusions......Page 73
References......Page 74
A short-term mummichog (Fundulus heteroclitus) bioassay to asses endocrine responses to hormone-active compounds and mixtures......Page 76
Introduction......Page 77
Animal husbandry......Page 78
Short-term bioassay......Page 79
Fish sampling......Page 80
Classification of follicle stage......Page 82
Radioimmunoassay......Page 83
Vitellogenin assay......Page 85
Mummichog field collection......Page 89
Animal husbandry......Page 90
Artificial regression and recrudescence for year-round supply of fish......Page 91
Short-term bioassay......Page 92
Fish sampling......Page 93
Classification of follicle stage......Page 95
Plasma steroid extractions......Page 96
Radioimmunoassay......Page 97
Vitellogenin assay......Page 102
Statistics......Page 105
Results and discussion......Page 106
References......Page 109
Conducting dose–response feeding studies with salmonids: Growth as an endpoint*......Page 114
Fish husbandry/experimental......Page 115
B. Raising fish from eggs......Page 116
C. Raising salmon fry to presmolt juveniles......Page 117
E. Distribution of fish to experimental tanks......Page 119
F. Conducting the experiment......Page 122
Discussion/considerations......Page 123
I.A Water quality parameters......Page 124
I.B Water flow rate......Page 125
I.D Feeding fish and increasing food ration......Page 126
Appendix II Method of weighing fish......Page 127
III.A Determining the number of experimental tanks......Page 128
III.B Randomization of fish and treatments......Page 130
III.C Analyzing the results......Page 131
IV.A Fish food for experimentation......Page 132
IV.B Dosing the fish pellets......Page 133
References......Page 134
Field experiments with caged bivalves to assess chronic exposure and toxicity......Page 138
Introduction......Page 139
Test organisms and materials required......Page 140
Materials required......Page 141
Exposure chambers......Page 142
Conducting the tests......Page 144
Results and discussion......Page 149
Summary and conclusions......Page 154
References......Page 155
Introduction......Page 158
Tissue preparation......Page 159
Procedures......Page 160
system of stressed P. scaber......Page 164
Histopathology in isopod toxicity studies......Page 166
References......Page 167
Sperm cell and embryo toxicity tests using the sea urchin......Page 168
Introduction......Page 169
Reagents......Page 170
Animal collection and culture......Page 171
Pre-test phases......Page 172
Sperm cell test procedure......Page 174
Embryo toxicity test procedure......Page 175
Preparation of test solutions......Page 176
Data analysis......Page 177
Negative controls......Page 179
Quality assurance/quality control......Page 180
Sensitivity toward pure substances......Page 181
References......Page 187
Introduction......Page 190
SRB culture......Page 191
Preparation of the nutrient medium......Page 193
Determination of biomass concentration......Page 194
Sampling and analysis......Page 195
EC50......Page 196
References......Page 197
section two......Page 200
Theory......Page 202
Coral nubbins for experimental application......Page 208
Sample preparation and assay protocols......Page 209
Step 1: preparing the organism and experimental design......Page 210
Sample preparation, quality control validation, and ELISA protocol......Page 213
Statistical analysis of data......Page 220
Case study: hexachlorobenzene and corals......Page 222
References......Page 226
A non-destructive technique to measure cytochrome P4501A enzyme activity in living embryos of the estuarine fish......Page 230
Materials required......Page 231
Spawning stock......Page 233
Embryo-larval exposure and development......Page 234
Data management, storage and analysis......Page 236
Associations between EROD and developmental effects......Page 237
Method sensitivity and specificity......Page 239
Testing mixtures and environmental samples......Page 240
Investigating toxicity mechanisms......Page 241
Summary......Page 242
References......Page 244
Introduction......Page 248
Lipid analyses......Page 250
Microlipid extraction of samples......Page 251
TLC/FID analysis......Page 252
Quality assurance......Page 253
Results and discussion......Page 254
References......Page 257
Introduction......Page 260
Materials required......Page 261
3. Dissection of larvae and in vitro incubation of prothoracic glands......Page 263
4. Ecdysteroid extraction......Page 269
5. Radioimmunoassay......Page 270
Results and discussion......Page 273
References......Page 276
The electro-olfactogram: An in vivo olfactory function and sublethal neurotoxicity in fish......Page 278
Introduction......Page 279
Equipment......Page 281
Reagents......Page 283
Fish......Page 284
Microelectrode preparation......Page 285
Fish preparation......Page 286
Direct delivery of odorants and contaminants to the olfactory epithelium......Page 287
Determining an appropriate pulse duration......Page 288
Selecting odorants and concentrations......Page 290
Sources of artifacts......Page 292
Estimating thresholds for sublethal copper neurotoxicity......Page 293
References......Page 296
Introduction......Page 298
Chemicals for analysis......Page 299
Procedures......Page 300
Calculation of IC50s......Page 302
Factors that affected results......Page 303
References......Page 305
Introduction......Page 308
Materials required......Page 309
Procedures......Page 310
Results and discussion......Page 313
References......Page 320
Introduction......Page 322
Innate immunity and antimicrobial peptides......Page 323
Materials required......Page 325
Microbial conditions......Page 328
Experimental curves......Page 329
Results and discussion......Page 330
References......Page 333
Introduction......Page 336
Materials required......Page 337
Tissue biopsy......Page 338
Tissue homogenization......Page 339
DNA amplification......Page 340
Results and discussion......Page 341
References......Page 346
section three......Page 350
Introduction......Page 352
Materials required......Page 353
Selection of coral species......Page 354
Collection of gametes and larvae......Page 355
Recruitment assays......Page 357
References......Page 359
Using the stickleback to monitor androgens and anti-androgens in the aquatic environment......Page 360
Introduction......Page 361
ELISA for spiggin......Page 362
Light microscopy......Page 363
ELISA for spiggin......Page 366
Results and discussion......Page 367
References......Page 375
Introduction......Page 378
Mobility of organic compounds to surface water......Page 380
Time-weighted sampling......Page 382
Distributional concentration model (percentiles)......Page 383
Time series concentration model (daily)......Page 385
Distributional concentration model......Page 392
Time series concentration model......Page 395
References......Page 403
Design and analysis of toxicity tests for the development and validation of biotic ligand models for predicting metal bioavailability and toxicity......Page 406
Supplies......Page 407
Reagents......Page 408
Mathematical theory......Page 409
Steps in the development of a BLM......Page 412
Step 1: experimental design......Page 414
Step 2: toxicity testing/measuring......Page 415
Step 3: calculation of speciation and EC50s......Page 417
Step 5: estimation of Zn binding parameters for the biotic ligand......Page 422
Step 7: comparison of observed and predicted EC50s......Page 425
References......Page 427
Introduction......Page 430
Biosensor......Page 431
Chemicals......Page 432
Procedures......Page 433
Assay protocol......Page 434
Data interpretation......Page 436
Reference response series......Page 439
Fingerprinting algorithm and use of best-fit identification technique......Page 441
Identification of toxicant spikes......Page 442
Characterization and differentiation of industrial effluents......Page 443
References......Page 446
Aquatic in situ bioassays to detect agricultural non-point source pesticide pollution: A link between laboratory and field......Page 448
Exposure system......Page 449
Type, number, and source of organisms......Page 452
Case study......Page 455
Site selection......Page 456
Experimental design......Page 457
Additional measurements......Page 458
Case study......Page 459
General......Page 461
First experiment on exposure box design......Page 463
Second experiment on pesticide effects: toxicological and ecological evaluation......Page 464
Conclusion......Page 466
References......Page 467
Improvements to high-performance liquid chromatography/photodiode array detection (HPLC/PDA) method that measures dioxin-like polychlorinated biphenyls and other selected organochlorines in marine biota......Page 470
Tissue extraction......Page 471
Analyses of OCs by HPLC/PDA......Page 472
Analyses of OCs by GC/MS......Page 473
Sample extraction and cleanup......Page 474
OC analyses by HPLC/PDA......Page 475
Quality assurance......Page 476
Results and discussion......Page 477
Acknowledgments......Page 483
References......Page 484
Introduction......Page 486
Material required......Page 487
Procedures......Page 488
Sample collection......Page 489
Empore disc elution......Page 490
Results and discussion......Page 493
Reference......Page 497
A toxicity assessment approach for evaluation of in-situ bioremediation of PAH contaminated sediments......Page 500
Introduction......Page 501
Sources of sediment samples......Page 503
Sulfate reducing conditions slurry system studies......Page 505
Sediment toxicity tests......Page 506
Sediment manipulation methods......Page 507
Sediment dilution procedure......Page 509
Sediment aeration procedure......Page 510
Biodegradation studies with N/P amendments......Page 511
Biodegradation studies with organic amendments......Page 512
Biodegradation studies under sulfate reducing conditions......Page 513
Sediment toxicity testing......Page 521
Conclusions......Page 526
References......Page 528
Introduction......Page 532
SPME as a biomimetic sampling device......Page 534
Determining time taken to reach equilibrium......Page 535
Solid phase microextraction fiber analysis......Page 536
Results and discussion......Page 539
Procedure......Page 540
Results and discussion......Page 541
References......Page 543
Introduction......Page 546
Materials required......Page 547
Procedures......Page 548
Results and discussion......Page 550
References......Page 551
section four......Page 556
Introduction......Page 558
Tissues......Page 559
Supplies and equipment for FT-IR spectral analysis......Page 560
Other considerations......Page 561
Principal components analysis......Page 562
DNA damage index......Page 563
Comparison of mean DNA spectra......Page 564
DNA damage index......Page 565
References......Page 566
Introduction......Page 568
Material requirements and setup......Page 569
Clinical pathology......Page 572
Results and discussion......Page 573
References......Page 577
Introduction......Page 580
Why study behavior?......Page 581
Fish models in behavioral testing......Page 582
Avoidance and attractance......Page 589
Swimming patterns......Page 591
Respiratory patterns......Page 592
Social behavior and group dynamics......Page 593
Behavioral analysis systems......Page 595
Preliminary studies......Page 599
Conclusions......Page 600
Acknowledgments......Page 602
References......Page 603
Measuring metals and metalloids in water, sediment, and biological tissues......Page 612
General......Page 613
Metals in biological tissue or sediments......Page 619
Dissolved metals in water......Page 620
Metals in biological tissue or sediments......Page 621
Appendix: Analytical details for specific elements......Page 625
As (arsenic, AW 74.9216, D0 5.0, Ei 9.81, BP 615 (hydride AsH3-55,......Page 626
Ca (calcium, AW 40.08, D0 4.8, Ei 6.11, BP 1494, Ca-40 96.95%, Ca-42 0.65%,......Page 627
Cr (chromium, AW 51.996, D0 4.4, Ei 6.77, BP 2672, Cr-50 4.35%, Cr-52......Page 628
Fe (iron, AW 55.847, D0 4.2, Ei 7.90, BP 2862, Fe-54 9.59%, Fe-56 91.72%,......Page 629
Mo (molybdenum, AW 95.94, D0 6.3, Ei 7.09, BP 4639, Mo-92 14.84%,......Page 630
Pb (lead, AW 207.2, D0 3.9, Ei 7.42, BP 1750, Pb-204 1.40%, Pb-206 24.10%,......Page 631
Sn (tin, AW 118.69, D0 5.7, Ei 7.34, BP 2603 (hydride SnH4-52), Sn-112......Page 632
Zn (zinc, AW 65.38, D0 2.9, Ei 9.39, BP 907, Zn-64 48.60%, Zn-66 27.90, Zn-......Page 633
References......Page 634
Background......Page 638
Inorganic LSER variable value development......Page 639
Materials required......Page 645
Inorganic LSER variable value determination......Page 646
Results and discussion......Page 647
References......Page 649
Determining aromatic hydrocarbons and chlorinated hydrocarbons in sediments and tissues using accelerated solvent extraction and gas chromatography/mass spectrometry......Page 652
Introduction......Page 653
Equipment......Page 655
Reagents......Page 656
Standard solutions......Page 657
Sample extraction......Page 658
Sample cleanup......Page 659
Gas chromatography/mass spectrometry......Page 660
Calculations of results......Page 662
Quality assurance......Page 664
Results and discussion......Page 665
References......Page 672
Histological preparation of invertebrates for evaluating contaminant effects......Page 674
Introduction......Page 675
Materials required......Page 676
Supplies......Page 677
1. Select and prepare fixative solution......Page 678
2. Inventory samples......Page 679
3. Trim and fix tissue......Page 681
4. Remove excess fixative, decalcify tissue (optional), and place in cassettes......Page 682
4. Process and embed tissue......Page 686
5. Section and mount tissue......Page 689
6. Stain tissue......Page 691
Special histochemical staining......Page 693
Immunohistochemical staining......Page 696
Results and discussion......Page 698
Davidson’s solution......Page 702
Cason’s trichrome solution......Page 703
5% aqueous phosphotungstic acid solution......Page 704
References......Page 705
Isolation of genes in aquatic animals using reverse transcription-polymerase chain reaction and rapid amplification of cDNA ends......Page 708
Total RNA isolation......Page 709
PCR of internal fragment, sub-cloning and sequence characterization......Page 710
Optional: SSCP-PCR analysis......Page 711
Total RNA isolation protocol......Page 712
DNase I treatment......Page 713
First strand synthesis protocol......Page 714
PCR protocol......Page 715
Sub-cloning and sequence characterization protocol......Page 716
RACE protocol......Page 717
in tumor tissue samples......Page 719
Anticipated results......Page 721
SSCP application: anticipated results......Page 722
References......Page 723
Analysis of mutations in l transgenic medaka using the cII mutation assay......Page 726
Introduction......Page 727
Animals......Page 728
Tissue dissection and DNA isolation......Page 729
Overview of l cII mutation assay procedures......Page 733
Sequencing of l cII– mutants......Page 739
Considerations for conducting mutation analyses......Page 740
Examples of studies of chemical mutagenesis......Page 743
Summary......Page 747
Supplies and reagents......Page 748
Packaging extract A (BHB 2688 cell stock)......Page 751
Packaging extract B (NM 759 cell stock)......Page 752
References......Page 753
Improved methods of conducting microalgal bioassays using flow cytometry......Page 756
Test species......Page 757
Maintenance of algal cultures......Page 759
Test sample preparation......Page 761
Preparation of test solution......Page 762
Flow cytometric analysis......Page 763
Protocol 1: Chronic growth rate inhibition test......Page 764
Protocol 2: Acute esterase inhibition tests with P. subcapitata and......Page 769
Results and discussion......Page 771
Conclusions......Page 775
References......Page 776
Index......Page 778
Back cover......Page 792