Paratuberculosisor Johne’s disease is a chronic intestinal disease in domestic and wild ruminants. It affects global dairy economy and is caused byMycobacterium aviumsubsp.paratuberculosis(MAP). The objective of this study was to analyze strain diversity in MAP-positive fecal samples by using a particular single nucleotide polymorphism (SNP) distinguishing between cattle (C-) and sheep (S-) type MAP and analysis of SNPs withingyrAandgyrBgenes differentiating between Types I, II, and III. Moreover, mycobacterial interspersed repetitive unit and variable-number tandem repeat (MIRU-VNTR) analysis using eight established loci was performed. A total of 90 fecal samples from diseased animals presenting diarrhea and/or weight loss, originating from 59 bovine herds across 16 cantons of Switzerland were screened by PCR for the MAP-specific F57 and IS900genes and were further subtyped. 96.7% and 3.3% of the samples contained C- and S-type MAP, respectively. Ten INRA Nouzilly MIRU-VNTR (INMV) profiles, with a discriminatory index of 0.802, calculated based on 65 epidemiological independent genotypes, were detected: INMV 1 (33.8%), INMV 2 (23.1%), INMV 6 (16.9%), INMV 9 (9.2%), INMV 116 (4.6%), INMV 3 (3.1%), INMV 5 (3.1%) and INMV 72 (1.5%), including two novel INMV profiles, namely INMV 253 (3.1%; S-type III) and INMV 252 (1.5%; C-type). INMV 1, INMV 2, and INMV 6 comprised almost 75% of the F57- and IS900-positive samples. Typing data from 11 herds suggest that there are some herds with intra-herd diversity of genotypes. The results of this study indicate a heterogeneity of MAP in Switzerland.