Ethiopia is center of diversity and one of the top nine agro-ecological regions for faba bean growth with high diversity. This high diversity is an opportunity for the improvement of the crop. However, the farmer named landraces and their molecular base variation was not well studied. Therefore, the objective of this study was to analyze the genetic diversity of farmers’ faba bean varieties in different agro-ecologies (sm3; tepid sub-moist mid highlands and sm4; wet sub-moist mid highlands) of North-East Amhara National Regional State, Ethiopia. The collection for the strata is considered as populations to analyze genetic base variation through inter simple sequence repeats (ISSR) genetic marker. Six varieties were used in the study areas among which four were local and the rest two were released as high yielding (Mert bakela) and water logging resistant (yeweha bakela). Six plants from each of the four populations were analyzed using 9 ISSR primers which amplified 113 loci of which 112 were polymorphic. Primer 812, 854 and 881 had produced 15 loci each and a total of 168 bands. Primer 812 had the highest resolving power, 7.32; primer 854 had the highest band informativeness, 0.49 and primer 881 had the highest polymorphic information content, 0.95. Population 2 that is of Sm3-South Wollo stratum showed the highest spatial diversity (4.615385) and Shannon-Weavers’ Diversity (1.364). Five out of six faba bean varieties were found in population 2 and three of them were unique. However, the most abundant local faba bean varieties, yehabesha and yeferenje bakela, had higher genetic diversity. Especially Yehabesha bakela had different unique futures including wider rating for disease resistance compared to the other most abundant released variety, yeweha bakela, (p = 0.01828).