67 results

Aquatic Animal Health Subprogram: validation of DNA-based (PCR) diagnostic tests suitable for use in surveillance programs for marteiliosis of rock oysters in Australia

Project number: 2001-630
Project Status:
Completed
Budget expenditure: $58,279.00
Principal Investigator: Rob D. Adlard
Organisation: Queensland Museum
Project start/end date: 30 Jan 2002 - 30 Jun 2005
Contact:
FRDC

Need

Marteiliosis (QX disease, aetiological agent the protozoan parasite marteilia sydneyi) typically causes serious, seasonally recurrent mortalities in farmed and wild rock oysters in eastern Australia. The disease is listed as notifiable by the OIE and is included on the Australian National List of Reportable Disease of Aquatic Animals.

The OIE has recently adopted the concept of zoning to facilitate trade and to prevent spread of disease within a country. In turn, Australia has recognised the value of zoning in its aquaculture industries with the adoption and endorsement of Zoning Policy Guidelines by Standing Committee on Fisheries and Aquaculture.

The establishment of scientifically defensible zoning and translocation policies, particularly in relation to QX disease control, is critical to the long term development of the rock oyster aquaculture industry. NSW Fisheries currently prohibits movement of oysters from known QX infected estuaries to those thought to be free of infection. However, given the many millions of rock oysters translocated annually between NSW estuaries of undetermined disease status, there is an urgent need to accurately identify free an infected zones. This, in turn, depends upon the availability of standardised, validated diagnostic tests.

Histopathology is currently viewed as the 'gold standard' for QX disease diagnosis, while preliminary comparative data (Callinan and Wesche, unpublished data) suggest that an alternative cytological method, stained tissue imprints of oyster digestive gland, has a sensitivity of 60% and specifically of 100%. Recently, however, there have been major advances in development of PCR tests for marteiliosis (Berthe et al. 2000; Kleeman and Adlard 2000). It is possible that PCR can be used to confirm presumptive/inconclusive diagnoses obtained by histopathology or cytology. PCR may also have potential as a cheap and reliable mass screening diagnostic test. In either event, however, rigorous standardisation and validation will be necessary before a PCR test can be accepted for use in zoning-related QX disease surveillance.

Objectives

1. Production of a fully validated, standard PCR diagnostic test for the presence of marteilia sydneyi in oyster tissue capable of identifying marteilia sydneyi to species level and with a high level of sensitivity.
2. Assessment of comparative cost/benefit of histological, cytological and PCR diagnostic methods for identification of marteilia sydneyi.
3. Production of an Australian and New Zealand Standard Diagnostic Procedure (ANZSDP) for marteiliosis.

Final report

ISBN: 0-9751116-1-2
Author: Robert Adlard

Aquatic Animal Health Subprogram: development of a disease zoning policy for marteiliosis to support sustainable production, health certification and trade in the Sydney rock oyster

Project number: 2001-214
Project Status:
Completed
Budget expenditure: $281,226.02
Principal Investigator: Rob D. Adlard
Organisation: Queensland Museum
Project start/end date: 6 Jun 2001 - 15 Jul 2005
Contact:
FRDC

Need

The rock oyster industry in Australia is currently valued at around $28 million annually. The current output is about half of the industry peak in the late 1970’s. For the industry to survive in the long-term requires the ability to service what may become a premium domestic market demanding a high quality product. The expansion of the industry is likely to be available only from international export, which in turn requires compliance with international regulations on oyster health with a transparent health audit trail. The rock oyster is potentially positioned for re-emerging export success, being a unique product with an extended shelf-life relative to other oyster species (e.g. the Pacific oyster, Crassostrea gigas) and this is an opportunity that should be exploited by the industry.

The techniques of surveillance and diagnosis for molluscan pathogens required by the OIE for imported oyster products are not only stringent and accepted as the worldwide standard, but are also applicable to domestic requirements within Australia. In essence, the regulations state that appropriate diagnostic tests are applied for detecting the presence of pathogens of molluscs (microscopic identification techniques with the potential for specific molecular identification using monoclonal antibodies or DNA probes) which have been collected as part of a surveillance program within delimited coastal zones. The sample size, period and frequency are determined with reference to the cycle of infection of the particular pathogen and its prepatent period. There is an initial 2 year period of surveillance before a zone can be granted a disease-free status, with ongoing surveillance required for this status to be maintained.

The development of a zoning policy framework for marteiliosis will provide a valuable opportunity to implement and field-test Australia’s zoning policy guidelines in a practical context to assist with the development of further zoning policies for diseases of aquatic animals. Considerable interest has already been expressed in the case study by State authorities and it will be discussed at an Aquatic Animal Disease Zoning Workshop in Canberra on 23 January 2001, hosted by the National Offices of Animal and Plant Health. Furthermore, the development of the zoning policy will be of direct benefit to the oyster industry by facilitating domestic and international market access, and through identifying and protecting the remaining disease-free production areas

Objectives

1. 1. The primary objective is to implement and field-test the zoning policy framework developed under Aquaplan in a practical context and to facilitate the development of further zoning policies for other significant diseases of aquatic animals. This will be conducted using marteiliosis as a case study to develop an effective zoning policy that is consistent with internationally recognised (OIE) standards. The zoning policy will aim to:* Reduce the risk of introducing this pathogen into the remaining disease-free production areas
and* Facilitate domestic and international market access for the industry.
2. 2. The sub-objectives necessary to achieve this are to:* Identify through sampling and appropriate diagnosis marteiliosis-free and marteiliosis-endemic estuaries within oyster culture areas
* Determine the specific identity of Marteilia sp. from positive samples through ultra-structural and molecular diagnostics
* Develop a rational and effective program of surveillance for marteiliosis, based on occurrence and an assessment of risk for each oyster producing estuary
* In consultation with fisheries managers and industry, develop a coastal zoning plan for marteiliosis.

Final report

ISBN: 0-9751116-3-9
Author: Robert Adlard
Final Report • 2006-02-01 • 975.12 KB
2001-214-DLD.pdf

Summary

The edible oyster industry in Australia is currently valued at around $62.5 million annually of which rock oyster production accounts for approx 56%. For the industry to survive in the long-term requires the ability to service what may become a premium domestic market demanding a high quality product.  The expansion of the industry is likely to be available only from international export, which in turn requires compliance with international regulations on oyster health with a transparent health audit trail.  The rock oyster is potentially positioned for re-emerging export success, being a unique product with an extended shelf-life relative to other oyster species (e.g. the Pacific oyster, Crassostrea gigas) and this is an opportunity that should be exploited by the industry.
 
Within Australia, the Sydney Rock Oyster industry is subjected to periodic epizootics of disease induced by a range of parasitic organisms that produce significant mortality and morbidity of commercial oyster stocks.  The most significant of these is the agent responsible for ‘QX disease’ (caused by the protistan parasite Marteilia sydneyi) affecting the Sydney rock oyster Saccostrea glomerata.  Management of this disease has been based on quarantine of affected estuaries enforced through limitation on the movement of potentially infected stock.  In this context, it was obvious that the oyster industry required a disease zoning policy based on scientifically defensible data to allow domestic best practice in oyster farming and to maximise market accessibility for the industry.  This host/parasite system then formed the basis for a test of the zoning policy framework developed under the federal government’s ‘AQUAPLAN’.
 
A number of key issues related to zoning and surveillance for specific diseases were addressed through this project.  Initially the design of field collection and the appropriate test to use for diagnosis were assessed to maximise, and allow quantification of, disease detection limits in the surveillance program.
 
1. The design of field sampling to identify disease infected oysters was critical in order to reach a statistically robust probability of disease detection.  Global animal health standards (Office Internationale des Epizooties) recommend random sampling from a zone to detect a prevalence of 2% or greater disease in a population.  This was fulfilled using a computer generated random selection of geographic co-ordinates under which individual oysters were sampled (Angus Cameron, AusVet).
 
2. The appropriate method for diagnosis of disease, another critical issue in disease surveillance programs, was assessed by comparing the sensitivity and specificity of: tissue imprints (cytology); or tissue sections (histology); or the presence of specific parasite DNA (by polymerase chain reaction - PCR).  Our analysis showed clearly that PCR was the most sensitive diagnostic test followed by cytology then histology.  PCR also detected the presence of sub-clinical infections which could not be unambiguously identified using either histology or cytology.  Confirmatory diagnosis (following PCR) at sub-clinical levels was undertaken using DNA in situ hybridisation tests designed to stain the QX organism specifically in tissue section.
 
Combined surveillance results from 2001 (NSW estuaries only), 2002-03 (NSW and Queensland estuaries) and 2004 (Queensland estuaries only) demonstrated some significant departures from the geographic distribution expected for QX disease.  In 2001 diagnosis was undertaken using cytology and no unexpected occurrences of the disease were observed, with positives recorded only from the Clarence River (1.5% of sample infected), Georges River (47% of sample infected).  In 2002 the distribution of disease was significantly different to that expected.  Initially using cytology for diagnosis there were no apparent unusual infections with Southern Moreton Bay (0.8% of sample infected), Richmond River (40.8% of sample infected), Clarence River (22% of sample infected) and Georges River (16% of sample infected) recording oysters positive for the disease.  However, when PCR techniques were used for diagnosis in estuaries that had never recorded the presence of the disease agent it became obvious that the organism was more widespread than indicated by previous diagnostic testing or previous occurrences of disease outbreaks.  In total 142 unexpected positives for Marteilia sydneyi were found in oysters scored as negative by cytological examination during surveillance in this project.  Of these, 61 were identified in oysters sampled from estuaries with no prior record of Marteilia sydneyi.  These represent oysters from Hastings River, Wallis Lake, Port Stephens, Bateman’s Bay, Tuross Lake, Narooma and Merimbula.
 
Further testing of these infections confirmed the identity of the QX organism and found it to be present in the oyster tissues at a sub-clinical level i.e. prior to reaching the oyster’s digestive gland where the parasite would normally produce spores.  At this stage of development, pathology in the oyster is reduced and the condition factor of oysters is not seriously compromised.
 
In 2003 surveillance and diagnosis using PCR techniques showed a reduced impact of QX disease with Southern Moreton Bay (0.67% of sample infected), Brunswick River (1.3% of sample infected), Richmond River (13.3% of sample infected), Clarence River (6% of sample infected) and Georges River
(0.67% of sample infected).
 
This project has had a significant impact on our understanding of QX disease in rock oysters as it applies to management.  Rather than the disease agent being limited geographically to those estuaries that experience periodic outbreaks, the agent has been identified in most rock oyster growing areas on the east coast of Australia.  As such there is the potential for outbreaks of QX disease in all commercial growing areas (indeed such an outbreak occurred in 2004, with seasonal re-occurrence in 2005, in the Hawkesbury River) and that disease is likely to be regulated through a combination of the dynamics of the parasite lifecycle and the level of oyster fitness.  Furthermore, in any aquatic system the environment will play an equally significant role in the outcomes of host/parasite interactions both through direct impact on stages (spores, infective stages) in the lifecycle of the parasite and indirectly through its impact on host fitness.
 
In the light of our new understanding of the distribution of the QX disease agent it could be argued that management through quarantine of identified QX-endemic estuaries is no longer appropriate.  However, the biology of Marteilia sydneyi (dynamics of the life cycle of the parasite, interactions with alternate hosts) and its interaction with the host oyster’s immune system are incompletely understood and the precautionary principle should be upheld especially in the case of such a serious disease.
 
While estuaries which undergo periodic outbreak should remain closed to export of oysters for relaying live in water elsewhere, local management will focus on disease seasonality and stock rotation to avoid the high risk periods in mid to late summer.  These periods should be identified with accuracy to maximise available growth periods in disease endemic areas of estuaries.  The ongoing projects to develop QX disease resistant oysters (NSW DPI and collaboration with Macquarie University) should run parallel with a program of incremental addition to the biological knowledge of this pathogen.  Specifically, an absence of our ability to maintain a laboratory based infection model hampers research on identifying those factors (pathogen-specific, oyster-specific and environment-specific) which promote disease.

Review of hatchery production technology for Sydney rock oysters

Project number: 2001-213
Project Status:
Completed
Budget expenditure: $42,032.00
Principal Investigator: John Benzie
Organisation: UNSW Sydney
Project start/end date: 13 Oct 2001 - 31 Dec 2002
Contact:
FRDC

Need

The breeding program for fast growth has been very successful and selection for disease resistance is showing good promise. Future progress of these breeding programs will be augmented with the development of genetic markers for disease resistance and fast growth. However, all this work is now on hold pending the results of this review.

Despite the best efforts of the staff at the Port Stephens Fisheries Centre, the hatchery production problems of the Sydney rock oyster have not been overcome over the past 10 years. This has failed despite input from marine pathologists within NSW Fisheries, other laboratories in Australia as well and overseas. (For recent review see Heasman, 2000). Therefore it is imperative that past research and hatchery practices are reviewed to determine if with revised and more disciplined approach the problems could be resolved.

The fact that a hatchery in Albany, WA has been able to successfully produce commercial quantities of the western strain of Sydney rock oysters spat suggests that problems encountered in NSW are surmountable. It is therefore proposed that hatchery production technology and the breeding program at the Port Stephens Fisheries Centre be reviewed afresh by an independent panel of experts to determine if a remedial course of action can be formulated.

Without commercial hatchery production of spat the NSW oyster industry is doomed to decline further, as the competing Pacific oyster industries interstate and New Zealand take advantage of genetic improvement and year round supply of spat.

References:
Heasman, M., 2000. Improved early survival of molluscs - Sydney rock oyster (Saccostrea glomerata). Final Report to Aquaculture CRC Project A.2.1. NSW Fisheries Final Report Series (in press).

Objectives

1. To critically review objectives, methodology and results to date for the Sydney rock oyster breeding program.
2. To critically review practices and procedures for Sydney rock oyster hatchery technology at the Port Stephens Fisheries Centre and problems associated with larval and post-settlement mortality.
3. Prepare a cost/benefit review of the Sydney rock oyster hatchery program.
4. Provide recommendations for either continuation or discontinuation of Sydney rock oyster hatchery R&D and breeding program.

Development of a national biotoxin strategy

Project number: 1999-332
Project Status:
Completed
Budget expenditure: $64,970.00
Principal Investigator: Ken Lee
Organisation: Department of Primary Industries and Regions South Australia (PIRSA)
Project start/end date: 29 Jun 1999 - 14 Sep 2002
Contact:
FRDC

Need

The potential for poisoning events from biotoxin contamination of shellfish is a growing concern for all shellfish producers, aquaculture managers and the general public in Australia.

Outside of shellfish consumption, the occurrence of marine biotoxins has direct implications for human health for recreational users of the marine and coastal areas in which these blooms occur.

There are also implications for other seafood products which can accumulate biotoxins, as well as for ballast water management and the shipping sector, for general environmental protection / human health, and for sewage processors (as mentioned above, there are strong linkages between algal blooms and sewage output). However, the immediate concern for seafood industries is to assess the need to undertake sampling and monitoring programs in shellfish producing areas and to determine what risks exist for other seafood industries.

There is a genuine and urgent need to update the work undertaken in 1993, and supplement this work with additional information in order to develop a consistent national approach to biotoxin monitoring. The resulting monitoring strategy will be based on an appropriat and targeted sampling strategy, developed through an assessment of need and risk, in order to support the development and implementation of effective management regimes for the shellfish industry.

Objectives

1. To design a national biotoxin monitoring strategy, in consultation with government and industry, which provides an appropriate level of protection to the seafood industry and the consumer, against biotoxin contamination.
2. To assess the implications for public health from marine biotoxins.
3. To identify those organisms that pose a biotoxin threat to marine and estuarine shellfish in Australasian waters, and identify those Australian industries at risk.
4. To review existing biotoxin monitoring programs, phytoplankton surveillance, analytical expertise and recognition of program deficiencies.
5. To identify internationally recognised practices for the management of marine biotoxins in shellfish.
6. To identify gaps in current methodology for the identification and measurement of relevant biotoxins.
7. To determine a suitable protocol for consolidation, collation and analysis of data on biotoxins to support the development of predictive and management tools.

Final report

ISBN: 0-473-08391-4
Author: Ken Lee
Final Report • 2001-11-01 • 728.06 KB
1999-332-DLD.pdf

Summary

In Australia aquaculture and wild harvest of shellfish is an economically important and growing industry.  The safety of these products as a food source is of utmost importance from both public health and economic points of view.  One of the potential problems faced by shellfish growers is the contamination of their product with marine biotoxins.  These toxins are chemical compounds that are produced by specific naturally occurring marine microalgae. Most microalgae (often referred to as phytoplankton) are actually an important food source of the shellfish.  However, if biotoxins are produced they can induce human illness if contaminated shellfish are consumed.  This is not only a problem for commercially produced or harvested shellfish, but also for recreational and subsistence shellfish gatherers.

Biotoxins are not only a problem for Australia, as most coastal countries in the world have had, or have the potential for, problems with marine biotoxin contamination in shellfish.  In order to manage this problem, many countries have monitoring programs aimed at both the detection of the species of microalgae that produce the toxins, and at the detection of toxins in the shellfish.  Phytoplankton monitoring is a faster and cheaper test than shellfish flesh testing, and provides an early warning of the potential for contamination of shellfish with marine biotoxins.  However, the two types of testing need to be performed in conjunction with each other.  Internationally, food safety regulations are based on the levels of toxins in shellfish, and it is these results that should generally be used for regulatory decisions.  

Internationally the impacts of toxic microalgae on both public health and the economy are increasing in frequency, intensity and geographic distribution.  As aquaculture expands, and its importance as both food and income sources increases for many countries, it is expected that these impacts of marine biotoxins will also increase.  As international markets become more conscious of the safety of the foodstuffs they import, they impose safety regulations and can also impose non-trade barriers.  

Australia’s shellfish industry’s market has a large domestic component, worth approximately $90M per year.  There is, perhaps, less external pressure on Australia to manage these problems.  However the domestic consumers are no less important than overseas consumers, and hence there remains the need for protection from marine biotoxins.  There is a need for controls between states, just as there is a need for controls for exported product.  The proposed strategy is for a voluntary agreement between states, and spells out the acceptable monitoring programs, controls and regulations that must be met in order to ‘export’ shellfish to another signatory state.  This “model ordinance” is fairly well accepted as an international standard for shellfish safety, along with the European Union directives, which must be met in order to export shellfish to the EU. This proposed strategy is supported by a Model Australian National Marine Biotoxin Management Plan (Cawthron Report No. 646).

A marine biotoxin monitoring program is a long-term commitment to protecting the public health of shellfish consumers, understanding more about the shellfish resource and assisting the industry to growing into the future. It requires regulatory commitment at Federal and State government level to maintain and police biotoxin standards. 

Keywords: Biotoxins, aquaculture, shellfish, microalgae, monitoring programs.

Optimal stocking density for Sydney and Pacific oyster cultivation

Project number: 1999-307
Project Status:
Completed
Budget expenditure: $78,292.00
Principal Investigator: Tony Underwood
Organisation: University of Sydney (USYD)
Project start/end date: 25 Jul 1999 - 30 Jun 2002
Contact:
FRDC

Need

A means of quantifying the optimal stocking density of oysters relative to their food resources (quantity and nutritional quality) is a necessary feature in the sustainable management of the oyster industry at a time of increasing demand both for a greater volume of production and improved production efficiency. This has been recognized by the industry as a major need for there to be further development of production. It is also urgent because of increasing scrutiny and regulation under principles of ecologically sustainable development. Maximal development can only be sustainable if local carrying capacity is well understood.

Of the various ways, in theory, of arriving at quantification of optimal densities, an approach which concentrates upon the oyster and its food, measured initially at the scale of the individual lease, is practical and feasible. Once this relationship is defined, it may then with confidence be extended to a variety of habitat conditions, since it will be based upon the fundamental physiological properties of the species.

This proposal aims to define these relationships via rigorous physiological determinations, coupled with appropriate field studies and modelling. The proposed product will be a tool of value to the oyster farmer and to those concerned with planning and approving the expansion of leases within coastal habitats.

Objectives

1. To establish a functional relationship between stocking density, individual growth rate and yield for Sydney rock and Pacific oysters in an estuarine embayment in the Port Stephens estuary.
2. To generalise this relationship for relevance to other habitats by determining the interactions between available food, the feeding physiology and the growth of these oysters.
3. To use these formal relationships to demonstrate the optimal stocking densities for oyster cultivation in a variety of different environmental conditions.
4. To investigate the influence that feral oysters have upon such optimal density estimates.
5. To relate stocking density to the quality of the marketed oysters and investigate possible economic implications.

Final report

ISBN: 1-86487-487-2
Author: Tony Underwood

Oyster depuration: a re-assessment of depuration conditions and the role of bacterial and viral indicators in determining depuration effectiveness

Project number: 1998-319
Project Status:
Completed
Budget expenditure: $284,053.03
Principal Investigator: Kenneth A. Buckle
Organisation: UNSW Sydney
Project start/end date: 28 Jun 1998 - 27 Jun 2005
Contact:
FRDC

Need

The NSW oyster industry has suffered severe economic losses during the past 20 years as the result of major outbreaks of gastroenteritis and other oyster-borne diseases. Further outbreaks will cripple the industry from immediate losses and resulting litigation.

There is an urgent need to re-examine the conditions for depuration of NSW oysters harvested in very different geographic and environmental conditions with particular reference to water temperature, salinity and turbidity during depuration. It is important to determine what limitations exist in the current technology and how they might affect purification efficiency and commercial acceptance by oyster farmers. In light of such information, there will be a need to modify the Code of Practice for Oyster Depuration. Equally important is the need to educate and train oyster farmers in proper quality assurance of oyster quality and safety, which will include optimal application of purification technology.

Current safety of oysters is assessed by the presence of E. coli as indicators of bacterial pathogens, yet the majority of oyster-borne disease is of viral origin, principally NV and more recently, HAV. Unfortunately, direct testing for human viruses in oysters is time consuming and very expensive, and will remain so for the foreseeable future. From the perspective of implementing a quality assurance program that will effectively protect public health, it is essential to have more accurate and more reliable indicator tests for the presence of human viruses in oysters. Consequently, there is a clear need to examine the relationship between the presence of bacterial indicators (E. coli), coliphages and human infectious viruses in oysters before, during and after purification. Such a study will not only clarify the behaviour of viruses during purification but could lead to a rapid coliphage assay as a more accurate indicator of human virus presence in oysters.

Objectives

1. To optimize the process of oyster depuration with respect to conditions of: initial load of contamination
water temperature, salinity, turbidity and oxygen content
oyster: water ratio
time of depuration
geographical location of oysters
and oyster eating quality.
2. Based on the findings from objective 1, and in consultation with the NSW Department of Health, NSW Fisheries and representatives of the oyster industry, develop and write modifications to the Code of Practice for the Depuration of Oysters in NSW.
3. In conjunction with the NSW Shellfish Quality Assurance Program, conduct a series of workshops aimed at education of farmers in management of the quality and safety of oysters.
4. Investigate the possibility of using bacteriophages as an additional indicator of oyster safety and the performance of oyster depuration, with specific reference to the elimination of human viruses such as Norwalk and Hepatitis A viruses. Correlate bacteriophage occurrence and behaviour in oysters during purification with traditional E. coli standards and human virus behaviour as measured by an inactivated strain of polio virus.

Symposium on parasitic diseases of aquatic animals: 10th International Congress of Protozoology

Project number: 1997-336
Project Status:
Completed
Budget expenditure: $8,350.00
Principal Investigator: Bob J. Lester
Organisation: University of Queensland (UQ)
Project start/end date: 25 Apr 1997 - 4 Jan 1999
Contact:
FRDC

Need

The overseas speakers have agreed to come, speak at the Congress and speak to at least one other group while in Australia providing there is some assistance with their travel expenses. This is a great opportunity for members of the fishing and aquaculture industry to hear about latest developments in disease research. The proposed presence of these speakers at the Congress has already attracted other experts in marine disease to come to Australia for the Congress and these also will be meeting with special interest groups while here.

Objectives

1. The objective is to bring three overseas experts to explain about current developments in marine parasitology that relate to wild and caged tuna and other fish, prawns and oysters.

Final report

Author: Bob Lester
Final Report • 1998-11-18 • 519.90 KB
1997-336-DLD.pdf

Summary

A symposium on protozoan diseases of aquatic animals was planned as a feature of the 10th International Congress of Protozoology.

Speakers invited for the symposium were: Dr Mike Hine, NIWA, NZ, an expert on oyster and fish diseases, Prof. Tim Flegel, Mahidol University, Bangkok, an expert on prawn diseases, Dr El-Matbouli, University of Munich, an expert on myxosporeans of fish, and Prof. Bob Lester, University of Queensland, to talk on white spot disease in fish and to chair the symposium.

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