Abstract Book of EAVLD2012 - eavld congress 2012

S3 - O - 03
DIAGNOSIS OF Q FEVER IN DAIRY CATTLE BY PHASE-SPECIFIC MILK-SEROLOGY
Böttcher J 1 , Frangoulidis D 2 , Schumacher M 1 , Janowetz B 1 , Gangl A 1 , Alex M 1
1 Bavarian Animal Health Service, Poing, Germany,
2
Bundeswehr Institute of Microbiology, Munich, Germany
Q fever, serology
Introduction
Dairy cattle herds are frequently endemically infected by C.
burnetii the causative pathogen of Q fever. Shedding of C.
burnetii at calving and in milk is of special concern. Chronic
infection frequently results in an increased level of milk-shedding.
The diagnostic value of phase-specific serology was assessed by
a longitudinal study on milk samples and puerperal swabs in an
endemically infected dairy cattle herd with about 100 dairy cows.
PhII titer
10000
1000
100
10
PhI-serology
negative
positive
Materials & methods
Cows were kept in two groups (MS and RB) with close contact
within one barn. The same calving boxes are used for both
groups. Individual milk samples were collected from March 2010
until December 2011; since August 2010 puerperal swab were
collected from a proportion of cows within 16 h after calving.
Samples: 405 milk samples, 59 puerperal, 9 samplings (MS) and
465 milk samples, 64 puerperal swabs, 10 samplings (RB).
Phase-specific ELISAs were performed as described (1) with
minor modifications: Milk samples were diluted 1/5 log 10 and the
titer was calculated at 20% (OD%) of the positive control (serum
control diluted 1/400).
Frequency of positive samples
1,4
1,2
1
0,8
0,6
0,4
0,2
0
swabs
milk samples
1003 1004 1007 1008 1009 1010 1011 1012 1101 1102 1103 1104 1105 1106 1107 1108 1109 1110 1111 1112
52 46 98 0 46 57 57 52 0 54 58 51 52 0 56 45 47 0 52 46
0 0 0 5 11 2 10 9 5 9 8 5 0 7 5 3 6 8 11 6
Sampling (YYMM) and number of milk-samples and swabs, respectively
Fig. 1: Frequency of positive qPCR (CI95%) was plotted over
year/month of sample collection (YYMM, top), the number of milk
samples (middle) and the number of analysed puerperal swabs (bottom).
Cut-offs for milk&swabs were ≥10 C.b./ml milk and ≥1 C.b./swab.
qPCR (C.b. log10/swab and ml milk, resp.)
10
8
6
4
2
0
1003
1004
puerperal swabs
milk
1007
1008
1009
1010
1011
1012
1101
1102
1103
1104
Sampling (YYMM)
Fig. 2: Amount of C. burnetii-target in qPCR-positive swabs and milk
samples. qPCR scored positive if the titer exceeded 1 C.b./swab and ml
milk, respectively.
Milk samples and puerperal swabs were analysed by quantitative
(q) PCR (2). Target was quantified by a standard curve derived
from diluted C. burnetii reference isolate Nine Mile RSA493
(kindly provided by C. Heydel, Giessen).
The status of milk-shedding was assessed by two procedures: (1)
Mean shedding per cow was calculated (C.b./ml) and classified
as 1000 C.b./ml. (2) Shedding
pattern was defined as chronic shedding (CS; >3 times ≥10
C.b./ml) irrespective of negative results in between. Intermittent
shedding (IS3, IS2, IS1) with 3, 2 and 1 positive result, and
negative, if all samples gave less than 10 C.b./ml. Classification
relied on a minimum of three samples except for IS3, this status
required four samplings.
1105
1106
1107
1108
1109
1110
1111
1112
1
1003
1004
1007
1009
1010
1011
1012
1102
1103
1104
Milk sampling (YYMM)
Fig. 3: PhII-titers in cows (500. Only one multiparous CScow
remained just below 500. Positive predictive value for PhI 500
was 40%, whereas it was 25% for PhI 100 and PhII 100 , respectively.
In contrast to CS-cows, IS1/IS2-cows were characterized by a
striking instability of PhI- and PhII-results. Even after positive
puerperal swabs seroconversion in milk was not a regular finding.
PhI - /PhII + -pattern in young cows indicates the episode of
puerperal shedding at herd level and PhI 500 is a suitable
screening for heavy milk shedders.
Acknowledgements
The present study was supported financially by the Free State Bavaria and the
Bavarian Joint Founding Scheme for the Control and Eradication of Contagious
Livestock Diseases (Bayerische Tierseuchenkasse).
References
1. Böttcher, J., Vossen, A., Janowetz, B., Alex, M., Gangl, A., Randt A., and Meier,
N. (2011): Insights into the dynamics of endemic Coxiella burnetii infection in
cattle by application of phase-specific ELISAs in an infected dairy herd.
Veterinrary Microbiology, 151, 291-300.
2. Böttcher, J., Frangoulidis, D., Schumacher, M., Janowetz, B., Gangl, A., Alex,
M. (2012): Diagnostic value of Coxiella burnetii phase I and II antibody titers in
individual milk samples of cows. In preparation.
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