National Wild Fish Health Survey - Laboratory Procedures Manual

Version 2.0 - June 2004




This publication was initially produced in 2001 for the U.S. Fish and Wildlife Service, to describe the protocols and procedures utilized in the National Wild Fish Health Survey (NWFHS). It is intended for use by Fish Health Centers, but may also be helpful to fishery professionals, the technically oriented fishery industry, and students of biological or fishery sciences.

Distribution will be to select libraries and fishery-related agencies in the United States. Copies may be requested from the U.S. Fish and Wildlife Service by writing to:

Kimberly True
U.S. Fish and Wildlife Service
California-Nevada Fish Health Center
24411 Coleman Fish Hatchery Road
Anderson, CA 96007

All recipients will be required to provide feedback on their use of this Manual and offer suggestions for improvements. This material may be updated in the future if significant changes are made in the Survey procedures.

Because sections in this publication may not have undergone full editorial processing and complete peer review and may be published elsewhere in the formal literature, they should not be cited without approval of the author.

NOTE: The individual chapters comprising this Manual are to be used only within the context of the entire Manual. While chapters are presented as individual PDF files, they are not intended to be used separately. It is recommended to print all chapters and assemble the Manual in a loose-leaf binder.

Please read Chapter 1 - Overview of the National Wild Fish Health Survey in its entirety prior to using this Manual or the procedures listed within. Chapter 1 explains the intent and guiding principles of the Survey including: target pathogens, diagnostic and corroborative protocols, and data management in the NWFHS Database.



There are many published sources for laboratory procedures used in the diagnosis of finfish diseases. The National Wild Fish Health Survey (NWFHS) Laboratory Procedure Manual is not intended to be comprehensive in its treatment of this large subject area.Many of the major fish diseases that occur within the United States and throughout the world have not been documented in wild fish populations. Consequently, the NWFHS (Survey) has targeted major fish pathogens that are considered a risk to both natural and artificially produced fish populations. The NWFHS Laboratory Procedure Manual (Manual) contains protocols for these major pathogens, and also provides a general scheme of approach to allow detection of new or exotic agents through the disciplines of virology, bacteriology, parasitology and histology. Pathogens of Regional Importance (PRI) are included for informational purposes, however specific protocols have not been included, in most cases, for these pathogens.

The Survey includes methodologies that provide the highest sensitivity for detection of fish pathogens in subclinical wild fish populations.For this reason, Polymerase Chain Reaction1 (PCR) technology is included in the Survey as an additional tool to corroborate detection of specific diseases (Whirling Disease, Bacterial Kidney Disease, Ceratomyxosis, and viral agents such as IHNV and VHSV). In this application of PCR as a corroborative method, it is not the intent of the Survey to imply that PCR should replace gold standard methodologies for detecting or confirming a specific fish pathogen. Rather, PCR is being utilized as a secondary detection tool to corroborate the presence of a pathogen that has been detected by standard methods. The intent of the Survey is to test the PCR technique along side standard detection methods such as Pepsin-Trypsin Digest, ELISA, and virology. This information and field-testing will add information to our knowledge of both fish diseases and the performance of molecular tools as detection methodologies. Traditional confirmation tests such as histology (for Whirling Disease and ceratomyxosis), and serum neutralization for viral agents are also included for the Survey. Furthermore, the data collected from PCR testing will be evaluated annually to determine the appropriate use of this tool and its place in the Survey as methodologies evolve.

The foremost purpose of this Manual is to provide a working document of very detailed information for the U.S. Fish and Wildlife Service - Fish Health Center staff and clients regarding the daily procedures in which we conduct the National Wild Fish Health Survey. Where appropriate, the methods herein follow those described in Suggested Procedures for the Detection and Identification of Certain Finfish and Shellfish Pathogens, 1994, 4th edition. American Fisheries Society, Fish Health Section, Bethesda, Maryland, commonly referred to as the AFS Blue Book. Additional protocols are included for the Enzyme Linked Immunosorbent Assay (ELISA) and Polymerase Chain Reaction (PCR) techniques. In several instances protocols cited from other investigators in the published literature have been referenced as well. As with most laboratory manuals, this document will be updated as new and improved procedures become available.

1The PCR Process is covered by Patents owned by Hoffman-LaRoche, Inc.



This CD contains the Table of Contents and 15 chapters that comprise the National Wild Fish Health Survey - Laboratory Procedures Manual.  The Manual is presented as individual PDF files for each chapter to facilitate viewing and printing of the procedures.  Please read Chapter 1 - Overview of the National Wild Fish Health Survey prior to printing and using individual chapters in this Manual.

NOTE: The PDF files are for PRINTING ONLY. The Manual files do not allow users to make changes, or extract text from these documents.



Mention of brand names or trademarks, or any specific equipment in the text of this Manual is not an endorsement by the U.S. government or the Fish and Wildlife Service of any particular product. The products mentioned serve only as descriptive models for the reader; any comparable product can be used at the discretion of the reader.



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