WAVLD Symposium Handbook_V4.indd - csiro

World Association of Veterinary Laboratory Diagnosticians – 13 th International Symposium, Melbourne, Australia, 11-14 November 2007
1030 - 1230 Concurrent Session 1.4 - New Technologies & Platforms
The United States National Animal Health Laboratory Network (NAHLN)
B. M. Martin*, United States Department of Agriculture, Animal and Plant Health Inspection Service, Veterinary Services, National
Veterinary Services Laboratories,
1800 Dayton Ave., Ames, Iowa, 50010, USA;
T. F. McElwain, Washington Animal Disease Diagnostic Laboratory and Animal Health Research Center, College of Veterinary
Medicine, Washington State University,
155N Bustad Hall, Pullman, Washington 99164, USA
Introduction
The United States National Animal Health Laboratory Network (NAHLN) was established in 2002 to enhance
the early detection of, response to, and recovery from animal health emergencies, including bioterrorist
events, newly emerging diseases, and foreign animal disease outbreaks that threaten the Nation’s food
supply and public health. The NAHLN is a collaborative effort between the United States Department of
Agriculture (USDA) and the American Association of Veterinary Laboratory Diagnosticians (AAVLD). From
an initial group of 12 laboratories the NAHLN has expanded to 54 laboratories in 45 states.
Elements upon which the NAHLN was founded included:
• Standardized, rapid diagnostic techniques
• A secure communications, alert mechanism, and reporting system
• Modern equipment and trained personnel
• Training, proficiency testing, and quality assurance programs
• Facilities that meet biocontainment and security requirements
• Scenario testing in support of regional and national training exercises
Discussions & conclusions
A program review of the NAHLN was initiated in 2007 to identify stakeholder perspectives concerning the
objectives of the network, how well those objectives were being met, and whether changes in objectives are
needed. The report indicates that the original objectives are appropriate and valid and that the NAHLN has
made significant progress. Key accomplishments are summarized below.
• Standardized, rapid diagnostic assays have been validated and deployed to NAHLN laboratories for
avian influenza (AI), exotic Newcastle disease (END), and classical swine fever (CSF). NAHLN laboratories
are also participating in USDA supported surveillance efforts for bovine spongiform encephalopathy (BSE),
chronic wasting disease (CWD), and scrapie.
• A centralized national data system has been developed that receives standardized data sets from
participating NAHLN laboratories and supports automated transmission of data via Health Level Seven (HL7)
messaging. Efforts are now focused on ensuring that NAHLN laboratories routinely transmit electronic test
result messages.
• A “Train the Trainer” program for foot and mouth disease (FMD), CSF, AI, and END rapid assays was
developed and implemented. Not only has the program increased the number of laboratory personnel
prepared to respond to a national animal health emergency, but it provides a cadre of trainers available to
teach others. Successful implementation of this program was a significant step for the NAHLN and its
mission of ensuring sufficient diagnostic capability and capacity to address an animal health emergency.
• USDA’s National Veterinary Services Laboratories (NVSL) serves as the reference laboratory for NAHLN
and has provided training and proficiency testing programs for NAHLN laboratory personnel. The NVSL also
provides support through production and distribution of reagent standards and quality control panels.
• One of the major successes of the NAHLN has been the collaboration with other Federal and state
organizations to achieve common goals. The NAHLN has become the animal health laboratory backbone of
the United States emergency response and recovery program, and has enabled implementation of national,
standardized surveillance for high priority diseases.
References
1. American Association of Veterinary Laboratory Diagnosticians Website:
http://www.aavld.org/mc/page.do
National Animal Health Laboratory Network Website: http://aphis.usda.gov/animal_health/nahln
Wed 14 November
Wed 14 November
World Association of Veterinary Laboratory Diagnosticians – 13 th International Symposium, Melbourne, Australia, 11-14 November 2007
DEVELOPING AN APPROACH FOR RAPID IDENTIFICATION OF EMERGING BIOLOGICAL THREATS
Bill Colston, Ray Mariella, Reginald Beer, Klint Rose, Kevin Ness, Elizabeth Wheeler, Tom Slezak, Shea Gardner, Peter Williams, Amy
Hiddessen, Monica Borucki, Ben Hindson, Chris Bailey, and Crystal Jaing (UC Lawrence Livermore National Laboratory)
In our era of rapid, easy world wide travel, infectious diseases, both naturally occurring and
intentionally introduced, hold increasing potential to cause disease, disability and death. Their
prevention and control is fundamental to individual, national and global health and security.
Lessons learned from the recent SARS outbreak teach us that without the ability to rapidly identify
and characterize a previously unknown or emerging pathogen, our country’s ability to mount a
timely and effective response to a nationwide bioterrorism event is unlikely. In recent years the
biodefense community has focused on short-term production of assays for the specific
identification of a short list of known pathogens. While necessary, this focus has created a gap in
our defensive and public health arsenal. We are currently ill-prepared to deal with novel pathogens
(natural or engineered), complex mixtures of organisms, or detection of virulence regardless of the
organism conferring it. Particularly in the case of viral agents, persistent technology gaps exist in
this process, including sample handling and preparation and highly-multiplexed assays that can
detect and identify both known and unknown viruses. Also, in the case of viral infectious agents,
this problem is compounded by our near-total lack of knowledge of “normal” viral backgrounds in
environmental, human, and agricultural samples.
To address this challenge, we have begun developing an approach to create a translational
measurement capability that will allow rapid, high-throughput viral screening. This approach
includes (1) Sample preparation capabilities to isolate virus particles from the numerous other
inter- and intra-cellular materials present in a nasopharyngeal sample. Viruses, by their very
nature, cannot live and replicate by themselves, and must rely on the complex biochemical
machinery of the host cells. Thus, detection of viral signatures will rely on removing the great
number of interfering cellular components of the host cells, including proteins, organelles, and the
cells’ own genetic materials. (2) A long-term detection capability to isolate and individually test
every viral particle in a sample in a high-throughput, massively parallel microfluidic system. This
will be done by isolating each virus particle in its own picoliter size ‘individual biochemical
laboratory’ or microdroplet for sensitive PCR analysis. (3) Development of comprehensive, specific
multiplex PCR assays of known organisms that can be carried out in a microdroplet and will be
capable of family level identification following amplicon analyisis. The goal is to ultimately be able
to screen all known viral genomes to develop a minimum set of signatures that can be used to
characterize an unknown virus.
We will present new developments in microfluidic engineering, highly multiplexed biological assays
and advances in bioinformatics aimed at providing a broad capability for identification and
characterization of previously known and unknown viruses. Specifically, we have currently
demonstrated:
• Application of acoustic, electrophoretic and electrophoretic techniques to demonstrate a
continuous size selective sorting of particles in a flowing microfluidic stream. We have
developed a 3D theoretical simulation for a multi-field separator that combines acoustic
focusing capability with electrophoresis, and have used this simulation to design and test a
physical prototype.
• Development of preliminary viral discovery assays using theoretical calculations coupled
with wet-bench validation. To accomplish this, we designed novel algorithms and built
software to select highly conserved primer sets, optimized to work in multiplex format. We
have demonstrated reproducible, specific PCR amplification of predicted fragments from a
viral DNA genome (vaccinia virus, lister strain) using 10 bp primer sets.
•
Demonstration of Real-time, Taq-man-based sub-nanoliter PCR using digital microfluidics. Our
system detected a single copy of viral genomic DNA encapsulated in ten-picoliter droplets, with a
much earlier cycle threshold than conventional devices.