Combining microscopy and digital holography (DH) offers the unique advantage of simultaneously capturing complete 3D information about the specimen, and under coherent illumination it has been investigated for continuous PC acquisition and identification of biological specimens. Sunflower (Helianthus annuus L.) broomrape Orobanche cumana Wallr.( O. cumana) is a parasitic angiosperm, totally devoid of chlorophyll, that infects the roots of sunflower plants, causing significant damages to this strategic oilseed crop in Black Sea countries as Turkey, Romania, Ukraine, Bulgaria, including Republic of Moldova (RM). The long-term impact of the broomrapes is even more serious: their seeds may easily spread to other fields, and can persist in soil up to 20 years, leading to an accelerated increase in the infested areas. The aim of this research is to develop three-dimensional microscopy (3D) system to enhance the resolution and contrast in imaging of O. cumana seeds, occurring in different geographical regions of RM, Romania, Spain and Ukraine. Successful application of the modern technology allowed studying the broomrape seed in various layers, like surface and under surface of seed that can be the reliable means to distinguish related taxa and their geographical localization.