E. Robardet and F. Cliquet
Anses, Nancy laboratory for rabies and
wildlife, WHO CC for Research and Management
in Zoonoses Control, OIE RL for Rabies, EU
RL for Rabies and Rabies Serology
An annual activity questionnaire was sent to
National Reference laboratories (NRLs) last
January 2011 to collect and collate data on
methods used and results of test carried out
in the Community for rabies control
(Commission regulation N° 737/2008).
Questionnaires from 22 NRLs were received
back. This document reviews the analysis
performed at the scale of the European Union
in 2010.
GENERAL DATA
Reference laboratory network includes 27
National Reference Laboratories (one of them
is the European Union Reference Laboratory)
and 127 regional laboratories. For the full
year 2010, 75 873 Fluorescent Antibody
tests, 3 788 Rabies Tissue Culture Infection
Tests, 1 962 Mouse Inoculation Tests, 2 448
RT-PCR and 2157 Real Time PCR were performed
for rabies diagnosis.
In the frame of oral vaccination campaign
follow-up, 27 221 wildlife serology tests
and 39 366 tetracycline (TTC) detection
tests were carried out.
Figure 1: Accreditation in the field of
rabies in NRLs
QUALITY ASSURANCE
Twenty two on 25 laboratories (2 NRLs
not included in the data set) have at
least an accreditation for one of the
techniques relatives to
rabies.Fluorescent Antibody test (FAT)
harbours the highest frequency of
laboratories with accreditations among
rabies techniques in European Union
level (63%) (Figure1) as this method is
the gold standard for rabies diagnosis.
FAT is followed by FAVN test with 52% of
accredited laboratories. Rabies Tissue
Culture Infection Test (RTCIT) and
RT-PCR are also well represented with
respectively 33% and 22% of accredited
laboratories.
RABIES DIAGNOSIS TESTS
All laboratories (except one laboratory
not allowed to work with live rabies
virus and acquiring BSL3 laboratory)
currently use FAT (Table 1). RTCIT and
Mouse Inoculation Test (MIT) are not
systematically performed as confirmatory
test. Six laboratories use neither of
these two techniques. Molecular biology
techniques are more frequently used than
MIT (32% of laboratories perform MIT
while 44% perform RT-PCR and 56% perform
Real Time PCR) and typing is undertaken
in 28% of laboratories.
Table 1: Rabies diagnosis tests performed in
2010 in the frame of rabies surveillance
TECHNIQUES USED FOR ORV MONITORING IN
2010
Eleven EU countries were involved in
oral vaccination programmes in 2010.
Bait titration were carried out in NRLs
of involved countries except for four
countries where the titration was
undertaken in another laboratory. Three
different techniques of serology are
used for monitoring efficacy of oral
vaccination campaigns: ELISA, RFFIT and
FAVN test. ELISA is the test the most
frequently used (73% of laboratories
performing serology tests) followed by
RFFIT (18%). Only one laboratory (9%)
performs FAVN test. Tetracycline
detection was undertaken in every
country either by NRL or regional
laboratories.
RESULTS OF ORV MONITORING
It should be noted that data of rabies
antibody and tetracycline determination
percentages should be interpreted taking
into account the strategy of oral
vaccination adopted.
Levels of tetracycline detection in fox
teeth were found highly heterogeneous
among countries varying from 12% to 91%
(Figure 3). Same trends were observed in
seroconversion rates varying from 17% to
73% when using ELISA test and from 45%
to 82% for RFFIT (Figure 2). The country
performing FAVN test obtained 65% of
seroconverstion.
Fig 2: Level of tetracycline detection and
seroconversion rate in target population in
2010
CONCLUSION
• A low level of typing was observed on
isolates currently found positive for
rabies. As recommended by EC, OIE and WHO,
every positive sample should be typed. If
technique is not available in a NRL, contact
should be taken to perform analysis in
specialised NRLs.
• RTCIT and MIT are not systematically used
as confirmatory test for FAT while molecular
biology techniques (RT-PCR and Real Time PCR)
are more frequently used. Inter-
laboratory tests on diagnosis techniques
ensure comparability of rabies data within
Members States and should continue.
Participation in proficiency testing is also
part of quality assurance issues.
• In EU, evaluation of fox seroconversion
levels is performed using three different
tests (FAVN test, RFFIT and ELISA. A high
variability in serology and tetracycline
detection results is observed.
These data should be interpreted in view of
oral vaccination strategy used. However,
these data suggest the need to harmonize the
technique used for TTC detection and for
serology. Particularly for serology, the
technique of choice for serology follow-up
should be stated to obtain more comparable
results for monitoring oral vaccination
campaigns.