Abstract Book of EAVLD2012 - eavld congress 2012

S4 - P - 01
MOLECULAR CHARACTERIZATION OF MYCOBACTERIUM AVIUM SUBSP. PARATUBERCULOSIS
STRAINS IN A NATIONAL PARATUBERCULOSIS CONTROL PROGRAM
Michaela Altmann 1 , Eva Sodoma 1 , Petra Möbius 2 , Michael Dünser 1
1
Institute for Veterinary Disease Control, Austria Agency for Health and Food Safety, Linz, Austria
2
Institute for Molecular Pathogenesis, Friedrich Loeffler Institute, Jena, Germany
Mycobacterium avium subsp. paratuberculosis, RFLP, MIRU, VNTR, paratuberculosis
Introduction
In 2006 Austria started a national paratuberculosis (PTB) control
programme based on government regulation which affects cattle,
sheep, goat and farmed deer. PTB was declared a notifiable
disease, so animals showing clinical signs have to be tested for
MAP at the national reference laboratory. MAP strains isolated
from different herds between 2006 and 2011 and strains from red
deer were selected for molecular characterization.
Materials & methods
165 MAP field strains from 164 clinically diseased ruminants
were analysed by IS900 RFLP with two restriction enzymes
(BstEII, PstI) and MIRU-VNTR at 8 genomic loci (MIRU292,X3;
VNTR25,47,3,7,10,32) according to MÖBIUS et al. (2008) and
THIBAULT et al. (2007). These field strains included 141 MAP
cattle isolates from 73 farms and different breeds (Angus,
Aquitaine Blonde, Austrian Brown Mountain, Fleckvieh
(Simmental), Tyrolean Grey Mountain, Holstein Friesian, Jersey,
Limousin, Murbodner, Piemontese, Red Friesian, Danish
Melkrace), 19 isolates from goat and 4 isolates from red deer.
Results
The combination of RFLP and MIRU-VNTR analysis allowed a
differentiation of 18 MAP strain types classified AT1 to AT18.
Detailed results and frequency in animals and herds are shown in
Table 1. The occurrence of MAP strain types in different cattle
breeds, goats and red deer is listed in Table 2.
Table 1: RFLP and MIRU-VNTR results of 165 MAP field strains
(INMV: INRA NOUZILLY MIRU-VNTR)
RFLP INMV
MIRU VNTR
Σ
292,X3 25,47,3,7,10,32 animals
AT1 C1P1 1 4,2 3,3,2,2,2,8 16
AT2 C1P1 2 3,2 3,3,2,2,2,8 59
AT3 C18 2 3,2 3,3,2,2,2,8 18
PnewIII
AT4 C1P1 17 3,1 3,3,2,2,2,8 8
AT5 C18 2 4,2 3,3,2,2,2,8 4
PnewII
AT6 C1 2 4,2 3,3,2,2,2,8 9
PnewV
AT7 C1P7 1 4,2 3,3,2,2,2,8 1
AT8 C1P1 5 4,2 3,3,2,2,1,8 2
AT9 C18 1 4,2 3,3,2,2,2,8 15
PnewIII
AT10 C1P1 new 2,2 5,2,2,2,2,6 1
AT11 CnewI 2 4,2 3,3,2,2,2,8 1
PnewIII
AT12 C18P1 17 3,1 3,3,2,2,2,8 1
AT13 C28 1 4,2 3,3,2,2,2,8 1
PnewI
AT14 C18P1 2 4,2 3,3,2,2,2,8 5
AT15 C1 2 4,2 3,3,2,2,2,8 1
PnewIV
AT16 C1P1 12 2,2 5,2,2,2,2,8 3
AT17 CnewIII 2 4,2 3,3,2,2,2,8 19
PnewIV
AT18 C14
PnewIII
2 4,2 3,3,2,2,2,8 1
Table 2: The occurrence of MAP strain types AT1-18
Breed
AT MAP strain type (number)
Angus 2(3)
Aquitaine Blonde 5(2),6(3)
Austrian Braun Mountain 1(1),2(3),6(1)
Fleckvieh 1(2),2(10),3(1),4(5),5(1),8(2)
Austrian Grey Mountain 1(2)
Holstein Friesian 1(1),2(9),5(1),6(3),16(3)
Jersey 2(3),3(3),7(1),15(1)
Limousin 1(5),2(25),3(12),4(3),9(15),11(1)
13(1),14(5),18(1)
Murbodner 3(1),12(1)
Piemontese 1(4)
Red Friesian 2(2),3(1),6(1),
Danish Melkrace 2(1),6(1),10(1)
Goat
Red Deer
18(19)
1(1), 2(3)
Discussion & conclusions
Despite the fact that 18 different MAP types were detected in a
comparatively large and diverse sample selection, 42% of all
isolates were classified as MAP type AT2 (RFLP C1-P1; INMV
2). MAP type AT2 was found in 8 of 12 PTB affected cattle
breeds and in 2 of 4 isolates from red deer and is therefore the
predominating MAP type strain in Austria. 9 new RFLP patterns
and 1 new INMV profile could be determined within the 165
analysed field strains. Some unique occurring strains showed
association with certain cattle breeds, goat and red deer (Table
2). This is the first study carried out in Austria providing data
about MAP strain types on a national scale. Additional
information about routes of transmission between herds and the
epidemiology of PTB in Austria could be achieved by using the
national cattle data base and the results of MAP typing.
References
1. MÖBIUS, P., LUYVEN, G., HOTZEL, H., KÖHLER, H. (2008): High
Genetic Diversity among Mycobacterium avium subsp. paratuberculosis
Strains from German Cattle Herds Shown by Combination of IS900
Restriction Fragment Length Polymorphism Analysis and Mycobacterial
Interspersed Repetitive Unit Variable Number Tandem Repeat Typing. J.
Clin. Microbiology, 46, 972-980
2. THIBAULT, V., GRAYON, M., BOSCHIROLI, M. L., HUBBANS, C.,
OVERDUIN, P., STEVENSON, K., GUTIERREZ, M.C., SUPPLY, P., BIET,
F. (2007): New Variable Number Tandem Repeat Markers for Typing
Mycobacterium avium subsp. paratuberculosis and M. avium Strains:
Comparison with IS900 and IS1245 Restriction Fragment Length
Polymorphism Typing. J. Clin. Microbiology, 45, 2404-2410