The Orchidaceae family, which is one of the most species-rich flowering plant families, includes species with highly diversified and specialized flower shapes. The aim of this study was to analyze the MADS-box genes expressed in the inflorescence of Orchis italica, a wild Mediterranean orchid species. MADS-box proteins are transcription factors involved in various plant biological processes, including flower development. In the floral tissues of O. italica, 29 MADS-box genes are expressed that are classified as both class I and II. Class I MADS-box genes include one Mβ-type gene, thereby confirming the presence of this type of MADS-box genes in orchids. The class II MIKC* gene is highly expressed in the column, which is consistent with the conserved function of the MIKC* genes in gametophyte development. In addition, homologs of the SOC, SVP, ANR1, AGL12 and OsMADS32 genes are expressed. Compared with previous knowledge on class II MIKCC genes of O. italica involved in the ABCDE model of flower development, the number of class B and D genes has been confirmed. In addition, 4 class A (AP1/FUL) transcripts, 2 class E (SEP) transcripts, 2 new class C (AG) transcripts and 1 new AGL6 transcript have been identified. Within the AP1/FUL genes, the sequence divergence, relaxation of purifying selection and expression profiles suggest a possible functional diversification within these orchid genes. The detection of only two SEP transcripts in O. italica, in contrast with the 4 genes found in other orchids, suggests that only two SEP genes could be present in the subfamily Orchidoideae. The expression pattern of the MIKCC genes of O. italica indicates that low levels at the boundary of the domain of a given MADS-box gene can overlap with the expression of genes belonging to a different functional A-E class in the adjacent domain, thereby following a "fading borders" model.