Proceedings of the 10th International Colloquium on Paratuberculosis

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#97 The EU ParaTBTools Project – Thematic Area 3: The inactivation <strong>of</strong> Mycobacterium avium subsp. paratuberculosis in milk and dairy products John Anthony Donaghy, Michael Rowe Food Microbiology Branch, Agri-Food & Biosciences Institute, Belfast, N. Ireland, United Kingdom The objectives <strong>of</strong> Theme Area 3 <strong>of</strong> <strong>the</strong> EU ParaTBTools project were to develop improved processing and manufacturing parameters for milk and dairy products to achieve improved inactivation <strong>of</strong> Map and to standardize culture and molecular methods for <strong>the</strong> detection/isolation/recovery <strong>of</strong> Map from dairy matrices. The Workpackages (WP’s 7, 8 & 9) involved research institutes from Ireland, UK, Germany, France, Ne<strong>the</strong>rlands and Belgium. In WP7 molecular and culture methods were developed for <strong>the</strong> detection/enumeration <strong>of</strong> Map from yoghurt, Gouda, Danish Blue and Munster cheeses. Real-Time PCR using an IS900-based TaqMan assay and a commercial Map DNA extraction kit was <strong>the</strong> most effective molecular method. Culture methods using HEYM supplemented with <strong>the</strong> antibiotic supplements PANTA, nisin (125 U) and penicillin (12 IU) was most suitable for <strong>the</strong> recovery <strong>of</strong> Map from Yoghurt, Danish Blue and Gouda cheeses. However, this medium was less suited for <strong>the</strong> recovery <strong>of</strong> Map from a raw milk cheese. In WP 8 eleven laboratories participated in a dairy products ring trial using <strong>the</strong> developed methodologies according to <strong>the</strong> NordVal validation method. Dairy samples contaminated with ‘medium’ and ‘low’ levels <strong>of</strong> Map were prepared and distributed to participant laboratories. Average sensitivities across all laboratories at medium and low levels <strong>of</strong> inocula were 95%, 98%, 100%, 84% and 96% for faecally-contaminated milk, yogurt, Gouda, Danish Blue and Munster cheese respectively. Calculated specificities for all matrices tested were in <strong>the</strong> range 95-100%. The sensitivity and specificity <strong>of</strong> <strong>the</strong> culture methods were lower than those recorded for molecular methods. In WP 9 <strong>the</strong> inactivation <strong>of</strong> Map during <strong>the</strong> manufacture <strong>of</strong> Gouda, Danish Blue, a smear ripened raw milk s<strong>of</strong>t cheese and yogurt has been assessed. The inactivation <strong>of</strong> Map during high temperature short time pasteurisation <strong>of</strong> milk in combination with homogenization (up-stream and down-stream) and bact<strong>of</strong>ugation are under investigation. 287
#65 A deer model for heritable resistance to MAP infection and Johne’s disease Frank Griffin, Rory O’Brien, Simon Liggett, Colin Mackintosh University <strong>of</strong> Otago, New Zealand; AgResearch Invermay, New Zealand Objective: To identify breeds <strong>of</strong> deer with polarized phenotypes for susceptibility or resistance to MAP Infection. Materials and Methods: An ELISA test was used to screen naturally infected deer stud herds to identify <strong>the</strong> relative prevalence <strong>of</strong> infection from eight uniquely different deer breeds (genotypes). Seropositive animals were necropsied and detailed microbiological and histopathological examinations carried out to confirm <strong>the</strong>ir infection status. Cross-bred hybrids were also examined. Followup studies were carried out using embryos from <strong>the</strong> distinct genotypes studied in <strong>the</strong> herd screen. Results: The different deer breeds had MAP seroprevalence ranging from 0 - 80%. Animals with a given genotype displayed similar infection rates when farmed in different herds. The influence <strong>of</strong> genotype was explored fur<strong>the</strong>r using embryos derived from different breeds as purebred or hybrid lines. The advantage using embryos is that exposure risk from <strong>the</strong> dam was controlled as all embryos were transferred randomly into test negative recipient hinds. Hybridising parental breeds that displayed low infection prevalence (0%) with breeds showing high prevalence (58%) produced <strong>of</strong>fspring with intermediate prevalence (24%) suggesting Mendelian heritability <strong>of</strong> resistance traits. Conclusion: High levels <strong>of</strong> line breeding are carried out within <strong>the</strong> New Zealand deer industry where some <strong>of</strong> <strong>the</strong> top performing stags are overrepresented in <strong>the</strong> gene pool. Selection criteria have been largely focused solely on superior production (venison and antler) and temperament. This selective breeding strategy within deer studs appears inadvertently to have selected animals with genotypes that display extremes <strong>of</strong> susceptibility or resistance to mycobacterial infections (tuberculosis and paratuberculosis). The purebred lines <strong>of</strong> deer that display <strong>the</strong>se extreme phenotypes are being studied to identify <strong>the</strong> genes that contribute to susceptibility/resistance using immunoassays for innate and adaptive immunity. Polymorphisms in Immune response genes (SNPs) and <strong>the</strong> mononuclear cell transcriptome <strong>of</strong> <strong>the</strong>se animals is being studied using 454 sequencing. 288
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Book of Abstracts
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Proceedings <stron
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Grant Support and Sponsorship <stro
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Scott Wells - Planning Committee Ch
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Monday, August 10, 2009 - All sessi
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Kari Røste Lybeck, Anne Kristine S
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1120-1140 #88: Influence of
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Concurrent Session B - Rm 175 Wille
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Thursday, August 13 Thursday, Augus
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Diagnostics and Genotyping Convenor
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#229 Detecting Johne’s Disease he
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Kalis CHJ, Hesselink JW, Barkema HW
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MATERIALS AND METHODS Sampling For
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To better determine the</st
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#107 Multilocus Short Sequence Repe
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Fig. 1. Dendogram (showing genetic
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#105 MIRU-VNTR genotyping o
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Diagnostics and Genotyping Poster A
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Missouri). Serum from a cow with do
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#7 Significance of
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RESULTS Of the 327
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Direct fecal nested Map PCR testing
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#16 Histopathological and immunohis
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#19 Development of
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#36 Genetic diversity of</s
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#60 Evaluation of
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#66 Comparison of
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(b) IS900 Nested PCR, using p90-p91
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(d) f57 Real Time PCR. The figure o
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• MIRU (3 loci); • VNTR-MIRU (7
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indexes, compared to each single an
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#108 Evaluation of
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#110 Detection and molecular confir
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endoscopic criteria and histopathol
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Bull TJ, McMinn EJ, Sidi-Boumedine
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#131 Seroprevalence of</str
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could be partly attributed to a sma
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#146 Analysis of v
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#162 Comparison of
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#187 Comparison of
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#191 Potentiating day-old blood sam
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samples from the s
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(MAA) (MAP87 and MAP3776) and 1 fro
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#196 Inter- and intra-subtype genot
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RESULTS In 601 (29%) of</st
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#211 Culture of my
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#242 Application and field validati
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#251 Diagnosis of
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Table 2. Paratuberculosis PCR resul
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#254 Programmes on Paratuberculosis
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Fig. 1. European countries specifyi
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A bulk milk quality assurance progr
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Republic of Lithua
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#276 Elimination of</strong
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other projects and
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Woodbine KA, Schukken YH, Green LE,
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Host Response and Immunology Keynot
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#33 MERKAL AWARD LECTURE No interfe
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#75 Interleukin 17 and interleukin
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#173 Local and systemic roles for b
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#23 Role of Mycoba
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Host Response and Immunology Poster
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#28 Intestinal MAP Infection via Pe
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#73 Age susceptibility of</
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#95 Role of nitric
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#134 Analysis of <
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#157 Study of <str
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#163 Immunohistochemical expression
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#193 Development of</strong
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#209 Murine macrophages carrying an
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#224 Application of</strong
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Table 1 UK and Cyprus MAP survey Re
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#228 Mycobacterium avium ssp. parat
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#145 MERKAL AWARD LECTURE Multinomi
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#50 Assessment of
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Table 1. Posterior median (CrIs) <s
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#55 Birth clusters of</stro
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from their dam, se
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#88 Influence of b
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#104 Relation between faecal shedde
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Epidemiology Poster Abstracts 109 1
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#22 The Prevalence of</stro
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CONCLUSION IS1311-based nested Ma/M
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#42 Seroprevalence survey o
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#77 Characterisation of</st
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Environmental samples were collecte
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REFERENCES 1) Chiodini RJ, Van Krui
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#113 Effect of soi
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#136 Error-adjusted, variance parti
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Latium Region Viterbo districts RES
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#144 The suitability of</st
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#178 Age structure of</stro
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A statistically significant lower f
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#183 Sero-prevalence of</st
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#218 Johne’s disease in t
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#244 A survey to estimate t
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Control Programs Keynote Lecture Re
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#202 Milk quality assurance for par
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#141 Control of pa
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#198 Use of small
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Control Programs Poster Abstracts 1
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RESULTS Loss of al
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#40 Managing Bovine Johnes Disease
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#74 Economic analysis of</s
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#87 Evaluation of
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separately for each existing herd <
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#98 An inter-laboratory ring-trial
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#120 Paratuberculosis vaccine inter
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The statement provides: • A stand
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COMPLEMENTARY ASPECTS On-Farm Disea
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#168 Paratuberculosis in Iran: past
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#180 Behavioural change in owners <
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the fact that <str
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#182 An integrated risk based appro
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etween 0 to 10, depending on <stron
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#203 Milk quality assurance for par
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#234 Johne’s disease vaccine: a c
Page 238 and 239: Pathogenomics and MAP Biology Conve
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Page 246 and 247: Pathogenomics and MAP Biology Persp
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Page 250 and 251: #52 Is ‘Indian Bison Type’ Myco
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Page 266 and 267: Abstract not available at press tim
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Page 272 and 273: Perspectives Talk: Public Health My
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Page 276 and 277: A total of 206 qua
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Page 286 and 287: #76 Towards improved reporting stan
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Page 292 and 293: Paratuberculosis ELISA Reliable, si
Page 294 and 295: PRIONICS AG WAGISTRASSE 27A PHONE +
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Proceedings of the 10th International Colloquium on Paratuberculosis

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