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Title:   Author  item
                                            Robbe Austerman, Sueleeitem
                                            Stabel, Judithitem
                                            HARRIS, B - USDA-APHISitem
                                            JOHANSEN, K - USDA-APHISitem
                                            Palmer, Mitchellitem
                                            PAYEUR, J - USDA-APHIS
  Submitted to: American Association of Veterinary Laboratory Diagnosticians
                                    Publication Type: Abstract Only
                                    Publication Acceptance Date: 10/22/2004
                                    Publication  Date: 10/22/2004
                                    Citation: Robbe Austerman, S., Stabel, J.R., Harris, B.N., Johansen, K., Palmer, M.V., Payeur, J.B. 2004. Culturing mycobacterium avium spp paratuberculosis from sheep feces in the united states [abstract]. American Association of Veterinary Laboratory Diagnosticians. p. 86.Interpretive Summary:
                                Technical Abstract: Attempts to culture feces from sheep infected with Mycobacterium avium ssp. paratuberculosis (MAP) using traditional solid media have been unrewarding.  Australia has successfully cultured their sheep MAP strains using the Bactec' 460 radiometric culture (Becton Dickinson, Franklin Lakes, NJ USA) by enhancing the media with 18% egg yolk and Panta' Plus(Becton Dickenson, Franklin Lakes, NJ USA) antibiotic mixture.  There is some evidence that sheep strains in the US are different than the Australian sheep strains although strains from both countries have not been successfully grown on traditional solid media. This study attempts to use the Australian method to culture fecal material from United States sheep known to be infected with MAP.  
	Two grams of fecal material from sheep known to be infected with MAP located in Iowa, Minnesota, South Dakota, and Wisconsin were decontaminated by the double centrifugation, double incubation method (NADC) or the Cornell method (NVSL) and 100-200 ul was added to a bactec' 460 vial containing the following additives, 1 ml of 100% egg yolk, 100 ul of Panta' plus, and 800 ul of sterile water. Samples were cultured for a total of 12 weeks. All vials having a growth index (GI) reading greater than 10 were sampled for PCR and acid fast staining at the end of 12 weeks and earlier if the vials had a GI> 200.  Negative samples were retested at the end of 12 weeks before they were considered negative. 
	 MAP was successfully cultured from all five sheep with clinical Johne's disease.  The GI was greater than 200 after 2 weeks of incubation for serveral samples. One subclinical animal took 10 weeks before the GI was greater than 10 and never had a GI of over 40. All of these animals were negative on Harold's egg yolk media. 
	It appears that the Australian method can readily grow sheep strains. Contamination was a problem. In the US, sheep are often fed stored feed rather than grass. Other antifungal agents need to be investigated to decrease the contamination rate.


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