Contents
-Home
-Welcome Letter
-Purpose of Workshop
-Program
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Appendix
-Participants
-Steering Committee
-Program Committee

Workshop Report
-Preface
-Final Report

and Environmental and Public Health Issues

Acacia Alcivar-Warren

Aquatics Section, Department of Environmental and Population Health

Tufts University School of Veterinary Medicine

North Grafton, MA 01536

acacia.warren@tufts.edu

ABSTRACT

It is recommended that increased funding for biotechnology research and development be directed to expand the US shrimp aquaculture industry and address environmental and public health issues. Most of the shrimp consumed by Americans is imported from developing countries. Preliminary results have shown that various pollutants (i.e. heavy metals, PCBs/pesticides, PAHs, exotic viruses, etc.) are present in tissues of both wild and cultured shrimp from various countries. Some of these pollutants have the potential to affect the function of reproductive, endocrine and immune systems of animals, including humans. Key research areas to expand industry development include (1) development of disease-resistant and pollutant-free broodstock representing various shrimp species, (2) enhancement of the rate of genetic improvement of domesticated shrimp by using both traditional quantitative genetics (selective breeding) and modern marker-assisted selection (MAS) methodologies, and (3) expansion of the knowledge base on shrimp immunology, endocrinology and toxicology. Key research areas to address environmental and public health issues include: (1) assessment of the genetic and ecosystem risks associated with the intentional or accidental release of both genetically modified shrimp and domesticated stocks that are inbred and/or disease-susceptible; (2) measurement of trace concentrations of pollutants present in shrimp used for human consumption, and (3) determination of potential public health risks associated with the presence of potential endocrine disruptors and other pollutants in shrimp.

KEYWORDS: ShrimpMap, immune response, disease resistance, pollutants, heavy metals, PCBs, PAHs, White Spot Syndrome Virus, Taura Syndrome Virus, Litopenaeus vannamei, biocomplexity

INTRODUCTION

Most of the shrimp consumed by Americans is imported from developing nations. This has caused unprecedented strain between supporters of the shrimp industry and local communities who are angrily reacting to the destruction of their public environmental resources and future livelihood. As society reacts to pressures on food production, choices between economic and societal goals will become more prominent political issues. To satisfy the increased demand for shrimp products in the U.S., efforts should be made to expand the domestic shrimp aquaculture industry. This could be achieved by applying biotechnology to the development of healthy and fast growing stocks. These technologies include biosecure, zero-exchange production systems and application of selective breeding and genomic technologies to enhance the rate of genetic improvement of domesticated shrimp. However, the risks to the environment, human health and other social concerns must be carefully evaluated before they are widely adopted.

Biotechnology has already played a major role in shrimp aquaculture (Argue and Alcivar-Warren, 1999). Benefits include: development of microsatellite markers to trace lineages in Litopenaeus vannamei and Penaeus monodon breeding programs (Wolfus et al., 1997; Xu et al., 1999), use of microsatellites and other genetic markers to analyze genetic diversity in wild and cultured L. vannamei and P. monodon (Wolfus et al., 1997; Xu et al., 2001), development of microsatellites and expressed sequence tags (ESTs) to construct a genetic map for shrimp (Alcivar-Warren, 2001; Dhar et al., 2000), cloning of immune response and disease resistance genes to study their expression under different environmental conditions (Alcivar-Warren, 2001), and improved diagnostic tools for Taura Syndrome Virus (TSV), White Spot Virus (WSV), Yellow Head Virus (YHV) and Infectious Hypodermal and Hematopoietic Necrosis Virus (IHNNV), all of which cause serious economic losses to the industry (Lightner 1996).

In the future, biotechnology is expected to play an even greater role in the improvement of the productivity and profitability of the U.S. shrimp aquaculture industry, development of more rigorous seafood safety standards for protection of human health, and conservation of biodiversity. To aid the expansion of the US domestic shrimp industry, funding for research and development is needed for (1) development of disease-resistant and pollutant-free stocks representing various shrimp species, (2) enhancement of the rate of genetic improvement of domesticated shrimp by using both traditional quantitative genetics (selective breeding) and modern marker-assisted selection (MAS) methodologies, and (3) expansion of the knowledge base on shrimp endocrinology, immunology and toxicology.

In order to identify the genes responsible for important quantitative traits and to perform marker-assisted selection, a linkage map (ShrimpMap) for L. vannamei is being developed. The map is based on both Type I (ESTs) and Type II (mostly microsatellites; and a few randomly amplified polymorphic DNA, or RAPD; and amplified fragment length polymorphism, or AFLP) markers. Most microsatellites were isolated from L. vannamei, a few are few from P. monodon. Currently, 173 microsatellites out of 1479 clones are being tested in reference mapping families (Meehan et al., 2001), The ESTs were isolated from L. vannamei tissues and a transcript database (ShrimpESTbase) has been initiated containing ESTs/cDNAs from TSV-, and WSV-challenged shrimp along with somatic and germinal cell tissues, developmental stages, and shrimp with high and low levels of environmental pollutants. The database will provide a snapshot of genes involved in the immune system and disease resistance of shrimp. It will also be used both for gene discovery and comparative mapping purposes using microarrays or gene-chip technologies. The markers will also be important as a management tool for tracing the pedigree of genetically improved stocks and better managing shrimp resources. So far, ~700 differentially expressed ESTs have been isolated from TSV-challenged L. vannamei by mRNADD, ~59% of which are unique, with no significant homology to other genes in the GenBank database. A few clones showed sequence similarities to immuno-regulatory genes from other species including a novel transmembrane domain of BMP/Tolloid genes, EGF-like and T cell receptor-like domains, Ig lambda-like domain, among others. Most interestingly, 43% of selected ESTs contained small size microsatellites and a large number (84%) contained flanking sequences to design primers for allele amplification by PCR. To date, 33% of ESTs have proven to be polymorphic (Alcivar-Warren, 2001). Tufts University researchers are collaborating with Drs. Shaun Moss and Brad Argue at OI and Dr. Jeffrey Lotz at the Gulf Coast Research Laboratory (GCRL) in Mississippi, in a project to map the QTL for TSV resistance, high growth and sex. More resources are needed to develop additional ESTs to construct a high resolution map in order to map those traits to a relatively small portion of the chromosome and to identify the genes and mutations responsible for these and other economically important traits in various shrimp species.

To complement the linkage map, there is a need to develop a physical map to facilitate comparative mapping to study synteny among species. This will require the availability of large-insert DNA libraries. Due to the large number and the small size of penaeid shrimp chromosomes, it has been difficult to identify L. vannamei chromosomes individually using conventional methods (reviewed in Argue and Alcivar-Warren, 1999). More research is needed to enumerate shrimp chromosomes using modern cytogenetic tools for assignment of genetic markers to the physical map. This would facilitate studies on chromosomal rearrangements in shrimp. Ultimately, functional analyses of genes responsible for quantitative traits are needed to demonstrate that the isolated genes are indeed responsible for the quantitative trait. This could be done using transgenic technologies. Understanding the genome organization of shrimp species will be important to accelerate the rate of genetic improvement of domesticated shrimp, learn about the structure and evolution of genomes, and understand the health and environmental interactions in wild shrimp populations.

Key research areas to address environmental and public health issues include: (1) assessment of the genetic and ecosystem risks associated with the intentional or accidental release of both genetically modified shrimp and domesticated (inbred and/or disease-susceptible) stocks; (2) measurement of trace concentrations of pollutants present in shrimp used for human consumption, and (3) determination of potential public health risks associated with the presence of endocrine disrupters and other pollutants in shrimp. Today, long-term, transgenerational studies on the potential effect of some environmental pollutants (i.e. potential endocrine disruptors) on animal health are lacking. Very preliminary results using a small number of samples have shown that certain pollutants [i.e. heavy metals, polychlorinated byphenils (PCBs), polyaromatic hydrocarbons (PAHs), exotic viruses, etc.] are present in wild and cultured shrimp obtained from Ecuador, El Salvador and The Philippines (Alcivar-Warren et al., 1999, Alcivar-Warren et al., 2001; Lightner et al., 1997). Some of these pollutants (i.e. cadmium, mercury, dieldrin) are of concern as they have the potential to impact the function of reproductive, endocrine and immune response systems of animals, including humans (EPA, 1999). Cadmium and mercury are also known to cause genetic changes in gastropods very rapidly (Nevo et al., 1986). Funding for long-term monitoring of heavy metals, PCBs and PAHs in shrimp consumed in the U.S. is needed. While heavy metals may be accumulating in wild shrimp, preliminary evidence for bioaccumulation of cadmium in cultured shrimp has also been found in domesticated shrimp grown in a raceway system in the U.S. (Alcivar-Warren and Meehan, 2001). More basic research is needed to isolate the genes involved in bioaccumulation of heavy metals in shrimp. This basic research will provide the tools needed to perform controlled studies to determine whether or not chemical residues are important predisposing factors for enhanced growth and/or disease.

Additional population based studies using a large number of samples from different ecosystems are also needed to address interactions of genetics, pollutants and susceptibility to diseases in a more holistic way. Recent information suggests that genetic differentiation and prevalence of viral diseases in wild shrimp populations are associated with the health of the ecosystem (i.e. mangrove status) and/or intensity of shrimp culture systems (Xu et al., 2001). Genetic differences between cultured and wild stocks were also demonstrated, suggesting the need to assess the genetic risks to wild populations caused by the intentional or accidental release of cultured stocks known to be inbred and/or disease-susceptible (Xu et al., 2001).

Recommendations

It is recommended that funding for biotechnology research and development be increased in order to expand the shrimp aquaculture industry in the US and address environmental and public health issues of concern (Table 1). The areas of research marked with asterisks (*) indicate that funding should also be provided to address environmental and public health issues. These issues should be addressed first before recommendations for long-term funding priorities are made.