Breeding of water buffaloes is an important zootechnic reality in the Latium region due to the international level of their DOP products ("mozzarella di bufala campana"). The species Bubalus bubalis is genetically different from the buffalo species situated in our region and that is breeded for another productive target. In order to increase our knowledge on the susceptibility of Mycobacterium avium subsp. paratuberculosis (MAP) to this species, we carried out a study on 130 buffaloes of a herd in the province of Latina. In 1998 three animals presented the typical symptoms of paratuberculosis infection and serological and cultural tests showed positive results. Diagnostic controls have been periodically repeated during the following years and revealed another 3 positive animals which led to the establishment of a sanitary control program for this farm. 108 sera and 75 fecal samples have collected from all buffaloes older than 2 years without any clinical signs related to paratuberculosis. Both, AGID and ELISA, tests showed negative results, whereas the cultural approach on HEYM medium revealed 3 animals with positive fecal samples. All isolates were typed as MAP. These 75 fecal samples were analised with two different PCR approaches (home-made and the commercial kit ADIVET^TM PARATUB) to test for other non- or low-shedding subjects. Parameters and conditions of the two PCR protocols were worked out using 30 previous tested bovine fecal samples of an international proficiency test and therefor used as positive and negative controls. The results of both PCR methods correlated completely with the controls so that we are now carrying out PCR tests on DNA extracted from the 75 buffalo fecal samples to value the sensitivity and specificity of our approach and to validate these PCR systems as rapid and reliable diagnostic tools for our purpose.