OIE Reference Laboratory Reports
Activities in 2011
Name of disease (or topic) for
which you are a designated OIE
Reference Laboratory:
Address of laboratory:
Infection with Mikrocytos mackini
Pacific Biological Station
3190 Hammond Bay Road
Nanaimo, BC
CANADA, V9T 6N7
Tel.:
250-756-7034
Fax:
250-756-7053
e-mail address:
Gary.Meyer@dfo-mpo.gc.ca
website:
Name (including Title and
Position) of Head of Laboratory
(Responsible Official):
Dr Stewart Johnson
Name(including Title and
Position) of OIE Reference
Expert:
Gary Meyer
Name (including Title and
Position) of writer of this report
(if different from above):
Gary Meyer
Annual reports of OIE Reference Centres, 2011
1
Infection with Mikrocytos mackini
Part I: Summary of general activities related to the disease
1.
Test(s) in use/or available for the specified disease/topic at your laboratory
Test
2.
For
Specificity
Total
Histology
Parasite visualisation
Group
1200
Tissue imprints
Parasite visualisation
Group
0
PCR
Molecular test
Group
800
qPCR
Molecular test
Type
(Currently undergoing validation)
3000
DIG-ISH
Molecular test for
parasite visualisation
Group
0
Electron microscopy
Parasite visualisation
Type
0
Production and distribution of diagnostic reagents
The reagents required for light microscopy and the molecular tests are available commercially. Positive control
samples are available to diagnostic laboratories and researchers in all OIE Member Countries upon request.
Positive control samples are used with each batch of tests performed in our laboratory.
Part II: Activities specifically related to the mandate
of OIE Reference Laboratories
3.
International harmonisation and standardisation of methods for diagnostic testing or the
production and testing of vaccines
a)
Establishment and maintenance of a network with other OIE Reference Laboratories
designated for the same pathogen or disease and organisation of regular inter-laboratory
proficiency testing to ensure comparability of results
Not applicable.
b)
2
Organisation of inter-laboratory proficiency testing with laboratories other than OIE
Reference Laboratories for the same pathogens and diseases to ensure equivalence of
results
1.
Proficiency test panels for histological detection of M. mackini were developed for Canada’s National
Aquatic Animal Health Laboratory System (NAAHLS).
2.
Reproducibility of a new qPCR test (see section 5) is currently being assessed with participation from
CSIRO’s Australian Animal Health Laboratory in Geelong, and two labs within Canadian National
Aquatic Animal Health Laboratory System (Gulf Fisheries Centre in Moncton, NB, and Freshwater
Insitute in Winnipeg, MB). This is being organized by C. Abbott at the Pacific Biological Station (the
OIE reference laboratory).
Annual reports of OIE Reference Centres, 2011
Infection with Mikrocytos mackini
4.
Preparation and supply of international reference standards for diagnostic tests or vaccines
Histological sections and DNA tissue samples of Pacific oysters (Crassostrea gigas) from British Columbia
infected with M. mackini were supplied as reference standards to the following laboratories:
1.
5.
Mrs. Cara Brosnahan, Ministry of Agriculture and Forestry, National Centre for Bio security and
Infectious Disease Campus, 66 Ward Street, Upper Hutt, 5018, New Zealand.
Research and development of new procedures for diagnosis and control
Development of a new TaqMan qPCR assay specific to M. mackini has been completed and full diagnostic
validation following OIE guidelines is in progress. The assay targets the ITS2-28S region of M. mackini rDNA
and can detect as little as 65 copies of a plasmid containing the target sequence in a matrix of host tissue. Assay
specificity was confirmed by the absence of amplification in Mikrocytos sp. and a broad range of other shellfish
pathogens and commercially important shellfish hosts. Due to the focal nature of infections, qPCR sensitivity was
evaluated across several tissue types, which were selected based on historical observations of the distribution of
infections in the host. Tissue types sampled were: mantle; palps; adductor muscle; a mid-body cross section
including gill, digestive gland, mantle, connective tissue, and gonad; and hemolymph from 62 Pacific oysters.
Statistical analyses of qPCR results from the five tissue types of positive individuals showed that the mid-body
slice was the optimal tissue to sample for detection of M. mackini by qPCR. Performance characteristics of the
assay (diagnostic sensitivity and specificity) will be derived this year from data generated from over 700 oysters.
Repeatability and reproducibility are also being measured following OIE guidelines.
6.
Collection, analysis and dissemination of epizootiological data relevant to international disease
control
Dr. Ralph Elston (Aqua Technics, 455 W. Bell Street, Sequim, WA USA 98383) detected M. mackini in
Kumamoto oysters (C. sikamea) from Humboldt Bay California in February 2011 using histology and
conventional PCR techniques; which represents a new host record and extension of the geographical range.
Denman Island Disease in Kumamoto Oysters – Impact and Management. Abstract and presentation by Dr. Ralph
Elston at the Pacific Coast Shellfish Growers Association and National Shellfisheries Association Meeting in
Salem, Oregon, USA in September 2011.
7.
Maintenance of a system of quality assurance, biosafety and biosecurity relevant to the
pathogen and the disease concerned
Canada’s National Aquatic Animal Health Program (NAAHP) lists Mikrocytos mackini as a reportable disease.
The National Aquatic Animal Health Laboratory System (NAAHLS) is in the process of implementing a quality
assurance and management system designed to the internationally recognized standards of ISO / IEC 17025:2005.
8.
Provision of consultant expertise to OIE or to OIE Member Countries
Provided an updated chapter to the OIE on infection with Mikrocytos mackini for the 2012 Manual of Diagnostic
Tests for Aquatic Animals.
9.
Provision of scientific and technical training to personnel from other OIE Member Countries
None in 2011.
10. Provision of diagnostic testing facilities to other OIE Member Countries
On one occasion (August 2011) provided confirmatory testing (qPCR and DNA sequencing of 18S and ITS genes)
on samples that originated from the USA which further confirmed the presence of M. mackini in Kumamoto
oysters (C. sikamea) collected from Humboldt Bay , California in February 2011.
Annual reports of OIE Reference Centres, 2011
3
Infection with Mikrocytos mackini
11. Organisation of international scientific meetings on behalf of OIE or other international bodies
None in 2011.
12. Participation in international scientific collaborative studies
None in 2011.
13. Publication and dissemination of information relevant to the work of OIE (including list of
scientific publications, internet publishing activities, presentations at international conferences)

Presentations at international conferences and meetings
Lowe G., Meyer G., Abbott M.G., Johnson S.C., Abbott C.L. Development of qPCR assay for Mikrocytos mackini
and assessment of optimum tissue for diagnostic testing. European Association for Fish Pathologists International
Conference, 12-16 October 2011, Split, Croatia.

Scientific publications in peer-reviewed journals
Abbott C.L., Gilmore S.R., Lowe G., Meyer G. & Bower S. 2011. Sequence homogeneity of internal transcribed
spacer rDNA in Mikrocytos mackini and detection of Mikrocytos sp. in a new location. Dis. Aquat. Org., 93, 243250.

Other communications
An Overview of Past and Present Research on Denman Island Disease (Mikrocytos mackini). Abstract and
presentation by Gary Meyer at the Pacific Coast Shellfish Growers Association and National Shellfisheries
Association Meeting in Salem, Oregon, USA in September 2011.
_______________
4
Annual reports of OIE Reference Centres, 2011