laboratory training for field epidemiologists rapid diagnostic tests investigation strategies and...

E P I D E M I C A L E R T A N D R E S P O N S E Laboratory Training for Field Epidemiologists

Rapid diagnostic tests

Investigation strategies and methods

May 2007


Learning objectivesLearning objectives

At the end of the presentation participants should know:

• What a rapid diagnostic test (RDT) is

• When a RDT should be used

• How a RDT should be used

• Who should use a RDT

• Advantages and disadvantages of RDTs


Biological confirmationBiological confirmationRequires laboratory with minimal culture, immunoassayequipment

Requires time :

• Bacterial culture : 24-72 hours

• Identification : 4 -24 hours

• Antibiotic susceptibility testing : 24 hours…

• ELISA : 2-4 hours, requires several samples to decrease the cost

Requires trained people


Biological confirmationBiological confirmation

In many developing countries, in a epidemic contextrequirements for biological confirmation at peripherallevel is not possible or feasible

Media and political pressure, rumours time is critical

Rapid diagnosis tests can offer a solution

?


RDT definitionRDT definition

Results in minutes to 1-2 hours

Accurate, simple to use, low cost, easy to interpret,stable under extreme conditions, little or no processing,culturally acceptable

Include “point of care” (for doctor) and “walk away” tests(home tests)

(Source http://www.rapid-diagnostics.org)


MethodsMethods

Antigen detection

Antibody detection

Molecular detection


Antigen detectionAntigen detection

Detects bacterial, viral or parasite antigen (surfaceantigen, soluble antigen) or toxin in biological fluids(CSF, blood, urine)

Primary techniques:

• Direct agglutination: slides, cards

• Latex agglutination: slides, cards

• Immunochromatography: dipsticks


Latex agglutination testLatex agglutination test

bacterial Ag

Latex beads (= polystyrene particles)

Antibodies specific to Bacterial polysaccharide Ag

Source: WHO meningitis workshop Ouagadougou Sept 2004

Source: http://www.rapid-diagnostics.org

Dye-labelled antibody, specific for target antigen, is present on the lower end of nitrocellulose strip or in a plastic well provided with the strip

Antibody, also specific for the target antigen, is bound to the strip in a thin (test) line, and either antibody specific for the labelled antibody, or antigen, is bound at the control line

Source: http://www.wpro.who.int/rdt

Lysing agendLabled AB.

Test band(bound AB)

Control band(bound AB)

Nitrocellulose strip

BoundAB

Free labled AB

http://www.rapid-diagnostics.org/

ImmunochromatographyImmunochromatography


Blood and buffer, which have been placed on strip or in the well, are mixed with labelled antibody and are drawn up strip across the lines of bound antibody


Test band(bound AB)

Control band(bound AB)

Blood and labled Ab flushed along the strip

Parasite antigen (AG.)Captured by labled AB.

ParasitizedBlood

ImmunochromatographyImmunochromatography


If antigen is present, some labelled antibody will be trapped on the test line. Excess-labelled antibody is trapped on the control line


Captured Ag-labled Ab-complex

Captured labled Ab

Labled AB-AG-complexCaptured by bound AB of test band

Labled AB-AG-complexCaptured by bound AB ofcontrol band

Immunochromatography


Antibody detectionAntibody detection

Requires seroconversion detection:

• IgG titer elevation not possible with RDT (= qualitative)

• IgM detection (after IgG elimination or IgM capture)

Main techniques:

• Direct agglutination (red cells + antigen, latex + antigen)

• Agglutination inhibition

• Immunodot

• Immunochromatography


Molecular detectionMolecular detection

Real-time PCR ?

• DNA extraction < 1 h

• Simultaneous amplification and detection <2 h

• Cost +++


Commercialized testsCommercialized tests

Family – genus – species

Bacteria

Clostridium perfringens, Corynebacterium diphteriae, Leptospira interrogans

Mycobacterium tuberculosis, Legionella pneumophila, Neisseria meningitidis

Salmonella Typhi, Yersinia pestis, Vibrio cholerae

Virus Adenovirus, Rotavirus, Influenza virus, Dengue virus, SRV

parasites Plasmodium sp, Giardia lamblia, Cryptosporidium

Not comprehensive


Advantages Advantages

Easy to use, minimal training

Rapid – same day results possible

Shelf life up to 1-2 years without refrigeration

Limited/no instrumentation; can be performed at theperiphery of health systems without laboratory or electricity

Some tests as accurate as reference-level laboratory tests

Source: PATH RDT website: http://www.rapid-diagnostics.org


Disadvantages Disadvantages Cost per test more than traditional tests

Some have limited shelf lives therefore increased demandson procurement and distribution

Mainly produce only "yes/no" answers

Could require subjective interpretation (reader variation)

Rapid tests can be less sensitive or less accuratecompared to existing tests

Source http://www.rapid-diagnostics.org


RDT use in algorithmsRDT use in algorithms

Algorithms are decision trees or visual schemes

Include behavioural, biological, or genetic risk factors for adisease, clinical signs and symptoms, use of other tests

Consider disease incidence and prevalence, availabilityand accuracy of other tests, probable consequences ofmisdiagnosis

Algorithms may be population specific and need to beupdated periodically



AlgorithmsAlgorithms

Source WHO Rapid HIV tests: guidelines for use in HIV testing and counselling services in resource-constrained settings, 2004


Necessity of AlgorithmsNecessity of Algorithms« The selection of the rapid HIV tests and test algorithms … is a responsibility of national governments … should not be delegated, either expressly or by neglect, to commercial enterprises, donor agencies or external aid programmes. The decision on which tests to use should be made following country-level technical assessments and the evaluation of other relevant factors, such as cost, current and continued availability, shelf-life and storage requirements. In each country these assessments … should preferably be the responsibility …of referral laboratories. Before selecting rapid test kits, many countries evaluate tests by using local specimens to confirm sensitivity and specifi city in their particular settings »



CostCost

Cost analysis : cost of using this diagnostic test oralgorithm in terms of personnel, facilities, equipment,sample collection materials, reagents, etc.

Cost-benefit analysis : cost of a correct diagnosis minusthe benefits from reaching that correct diagnosis

• Averted treatment costs and losses due to illness (e.g. wages)

• Less tangible costs e.g. reduced pain and suffering to patients

• Very difficult to cost benefits accurately and there is much debate on the accuracy of cost-benefit analyses.



CostCost

Cost-effectiveness analysis : costs of a diagnostictest or algorithm compared to the health outcomeresulting from the diagnosis

Answers the following:

Relative to other tests or algorithms, is this test oralgorithm a good use of health care funds?



Cost: malaria example Cost: malaria example



AccuracyAccuracy



AccuracyAccuracyPositive and negative predictive values* for various HIV prevalences

HIV prevalence 0.1% 1% 5% 10% 30%

NPV with one non-reactive test

100%

100% 99.9%

99.9%

99.9%

PPV with one reactive test 9% 50% 83.9%

91.7%

98.5%

PPV with two reactive tests 90.8%

99.0%

99% 99.9%

100%

*A sensitivity of 99% and a specificity of 99% have been used in these calculations. Predictive values have been rounded to one decimal place

NPV= negative predictive valuePPV= positive predictive value



Barriers to use of RDTBarriers to use of RDTAcceptability

• To policymakers, clinicians, and patients

• Sufficient sensitivity and specificity for the approval of international health and donor agencies

• Adequate predictive value and ease-of-use for clinicians, require culturally appropriate specimens

• Perceived as credible, to be accepted by patients

• Users must trust and accept RDT results, if tests are perceived as too simple, results may not be trusted



Barriers to use of RDTBarriers to use of RDT

Affordability

• Many RDTs are more expensive than other tests or algorithms they are intended to replace

– Especially true if RDT is used as a replacement for a syndromic algorithm

• Affordability constraints can be reduced by

– Working to decrease the cost per test

– Carefully designing algorithms to use the tests cost-effectively

– Educating users of cost-savings for more efficient use of therapeutic drugs



Barriers to use of RDTBarriers to use of RDTAvailability

• RDT not consistently available in many developing countries

• Most tests have a limited shelf life and many countries have poorly developed procurement and distribution systems

• The consistency and quality of imported tests

– Local government regulations, quality assurance, shelf life testing, and distribution systems all need to be assessed and improved



Role of laboratoriesRole of laboratories

Central public health laboratories

• Developing algorithms

• Writing standard operating procedure (SOP)

• National External Quality Control Scheme organization

• RDT accuracy testing – a certification process

• Training courses at peripheral level


Role of laboratoriesRole of laboratoriesPeripheral laboratories need

• Well trained people

• SOP and quality assurance culture

• Basic laboratory equipment (refrigerator, pipettes and tips, water bath)

• Sample collection and transportation experience

• Data collection and registration experience

Try to involve lab specialist as much as possible


ConclusionConclusion

RDT should be used in outbreak detection and investigation

Several manufactures

Be aware of the limitations and constraints

Use algorithms

Involve laboratories at central and peripheral level


Additional resourcesAdditional resources

Medecins Sans Fontieres (video on meningitis RDT)

Malaria RDT http://www.wpro.who.int/rdt/

• guidelines, reviews, trials etc.

http://www.rapid-diagnostics.org/


Developed by the Department of Epidemic and Pandemic Alert and Response of the World Health Organization with assistance from:

European Program for Intervention Epidemiology Training

Canadian Field Epidemiology Program

Thailand Ministry of Health

Institut Pasteur

Investigation strategies and methods

laboratory training for field epidemiologists rapid diagnostic tests investigation strategies and...

Documents

laboratory training

field epidemiologists

strip antibody

target antigen

antibody specific

soluble antigen

lines of bound antibody

strip parasite antigen