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CONTRIBUTIONS FROM FAO REFERENCE LABORATORIES AND COLLABORATING CENTRES

FAO/OIE WORLD REFERENCE LABORATORY FOR FMD AND RINDERPEST,
PIRBRIGHT, UK.

FMD REPORT FOR JULY TO SEPTEMBER 1999

Samples received from Iran indicate outbreaks caused by serotypes Asia 1, O and A. Routine vaccination in Iran is against serotypes O and A and the Asia 1 outbreaks required immediate control by the veterinary authorities to keep them contained. It is thought likely that the virus was imported illegally with live animals from an eastern neighbour, where Asia 1 is endemic. There is concern that further illegal traffic in live animals could allow the virus to spread into Turkey or the Caucasus. A genetically new strain of serotype A is also present in Iran, different again from the Iran '96 strain. The new strain is also antigenically different from Iran '96 and will require the use of a homologous vaccine if it continues to cause outbreaks. Results are awaited from recent type A isolates received from Turkey, to show whether they are similar to Iran '96 or have been replaced by the new A strain from Iran.

There have been further outbreaks of SAT 1 and 2 in Burundi, reflecting much larger outbreaks of FMD in East Africa. Reports from Kenya indicate that there have been economically serious outbreaks of FMD, but no samples have been submitted to the World Reference Laboratory.

RINDERPEST AND PPR REPORT FOR JULY TO SEPTEMBER 1999

Country

Species

Disease

Diagnosis technique

Result

Yemen

Cattle

Rinderpest

PCR

-ve

Mozambique

Suni antelopes

Rinderpest/PPR

C-ELISA

-ve

Chad

Wildlife

Rinderpest/PPR

C-ELISA

-ve

Burkina Faso

Wildlife

Rinderpest/PPR

C-ELISA

1/13 PPR +ve

Central African Republic

Wildlife

Rinderpest/PPR

C-ELISA

2/122 PPR +ve

Bangladesh

Sheep/goats

PPR

PCR

+ve

Iran

Goats

PPR (vaccine)

Molecular characterization

Under way

 

TRAINING COURSES AT THE INSTITUTE OF ANIMAL HEALTH, PIRBRIGHT, EPIDEMIOLOGY DIVISION

The courses are aimed mainly at overseas scientists who wish to improve their knowledge of exotic virus diseases and thus extend their national diagnostic capabilities. They provide training in the principles and applications of more modern biotechnology. The lectures also form part of the post-graduate training programme. The following courses will be run at IAH on an annual basis.


Course Title:
Laboratory diagnosis of rinderpest, peste des petits ruminants (PPR) and bluetongue diseases

Date: February 2000

Scope of the Course:
The aim of the 4 week course is to provide training in laboratory techniques for the diagnosis of rinderpest, peste des petits ruminants (PPR) and bluetongue diseases. The course will provide specific training in:

Rinderpest and PPR:
- Clinical signs and epidemiology
- Collection and transport of samples
- Antigen detection by immunocapture ELISA
- Antibody detection by competitive ELISA
- Lectures on eradication strategy and current status

Bluetongue and epizootic haemorrhagic disease (EHD):
- Clinical signs and epidemiology
- Collection and transport of samples
- Virus isolation in eggs and tissue culture
- Antigen detection by immunocapture ELISA
- Antibody detection by competitive ELISA


Course Title:
ELISA techniques and cell cultures for foot and mouth disease, swine vesicular disease and vesicular stomatitis viruses

Date: May 2000

Scope of the Course:
The aim of the 4 week course is to provide training in ELISA and cell culture techniques for the diagnosis of foot and mouth disease (FMD), swine vesicular disease (SVD) and vesicular stomatitis (VS). The course will provide specific training in:
- ELISA principles and test optimisation
- Preparation of samples for both virus isolation and ELISA
- Tissue culture and virus isolation
- Sandwich ELISA for the detection of FMDV, SVDV and VSV antigens
- Competitive ELISA for detection of antibodies to SVDV
- Liquid-phase blocking ELISA for detection of antibodies to FMDV
- Discussions on the epidemiology and control of FMD


Course Title:
Application of RT-PCR for the diagnosis of foot and mouth disease (FMD) and rinderpest

Date: November 2000

Scope of the Course:
The aim of the 4 week course is to provide training in polymerase chain reaction (PCR) for the diagnosis of foot and mouth disease (FMD) and rinderpest. The course will provide specific training in:
- Principles of DNA replication
- Principles of PCR
- Preparation of samples for PCR
- Optimisation of PCR
- Confirmation of identity of PCR product
- PCR ELISA
- Discussions on the epidemiology and control of FMD and rinderpest


MODULAR TRAINING FOR INDUSTRY PROGRAMME

Title of Programme:
Principles and applications of ELISA for disease diagnosis

Date: September/October 2000

Aims and objectives:
There are three main aims for this module:
1. Provide an in-depth understanding of the principles and applications of ELISA.
2. Provide a detailed insight into the development, validation and standardization of diagnostic assays. This will allow critical evaluation of commercially produced kits and greater ability to "troubleshoot" when and if problems arise.
3. Provide a full understanding of the relative merits of various assay systems which should assist in the development of appropriate immunoassays for both research and commercial applications.

Content, Structure and Delivery of Module:
The Module will run for 4 weeks and will be composed of a mixture of the following lectures, demonstrations and practical exercises. Individual lectures will be interspersed between the various stages of the practical exercises. The module will be run once in 1999 but subject to demand may be increased to twice in later years. Complimentary modules on molecular techniques such as PCR and nucleotide sequencing may be developed in the near future.



LECTURES ON BASIC PRINCIPLES AND TECHNIQUES

Basic Immunology:
- The Immune response
- Antigens
- Antibodies
- Adaptive Immunity and clonal selection
- Polyclonal antibodies
- Monoclonal antibodies

Basic Principles of ELISA:
- Terminology
- Direct ELISA
- Indirect ELISA
- Sandwich ELISA
- Competitive ELISA
- Choice of assay system

Stages of ELISA:
- Solid-phase
- Washing
- Addition of reagents
- Incubation
- Blocking non-specific reactions
- Enzyme conjugates
- Substrate/chromogen
- Colour development and stopping reaction
- Plate reading
- Practical problems

Practical aspects:
- Developing an ELISA
- Choice of antigens and control antisera
- Validating an ELISA
- Standardisation and quality control
- Internal Quality control
- External quality assurance
- Units of measurement, dilutions and pipettinG

PRACTICAL EXERCICES

Introduction to equipment:
- Calibration and use of pipettes
- ELISA reader and soft-ware programme

Direct ELISA:
- Titration of antigen and conjugate
- Cross-reactivity of anti-species conjugates
- Interpretation of results

Indirect ELISA:
- Titration of antigen
- Titration of positive and negative sera
- Selection of test serum dilution
- Frequency distribution of negative population
- Selection of cut-off value
- Testing field sera
- Interpretation of results

Sandwich ELISA:
- Titration of trapping antibody
- Titration of capture antibody
- Detection of antigens
- Interpretation of results

Competitive ELISA:
- Titration of antigen
- Titration of detecting antibody
- Cross-reaction of anti-species conjugates
- Antigen competition
- Antibody competition
- Selection of test serum dilution
- Frequency distribution of negative population
- Selection of cut-off value
- Testing field sera
- Interpretation of results

Trouble-shooting:
- Problem solving in all assays

Immunochemical techniques:
- Purification of immunoglobulins
- Coupling antibodies to enzyme

There will be a written, practical and oral assessment at the end of the course.
 

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