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We evaluated how fire impacts the ecological coherence of Aleppo pine forests and their biodiversity over a 40-year period. The study area forms part of an insular ecosystem of Zakynthos Island (Zante) in western Greece, which forms part of the Ionian Islands. Post-fire effects were studied for both plant and bird diversity at 20 sampling plots, using stratified random sampling, during the summer of 2012. The plots were selected based on the frequency of burning since the 1970s. Sites were categorized as: 1) no burning for >40 years, 2) burnt once in the 1970s, 3) burnt twice, first in the 1970s and again in 2000–2010, 4) burnt three times, first in the 1970s, and twice more in 2000–2010, and 5) burnt four times, first in the1970s, twice more in 2000–2010, and again in 2011. A total of 79 plant species and 26 bird species were recorded at the studied sites. One-way ANOVA analysis showed that fire intensity has a significant impact on alpha floristic diversity. Average plant species richness at sites that burned in 2011 was significantly higher than in those that burned in the 1970s. Detected differences in bird species richness were not significant; however, more species were documented in the forested habitats (unburnt for >40 years, and burnt in the 1970s). The highest Jaccard similarity index was observed between the sites that had not burned for more than 40 years and the sites that burned in the 1970s. The lowest floristic similarity to non-burnt sites was observed between the recently burnt sites in 2011 and the sites that burned in the 1970s. The lowest bird similarity was detected between recently burnt sites in 2011 and sites that had not burned for more than 40 years. DCA ordination showed the presence of a clear fire gradient, from intensively burnt open sites to non-burnt forest sites. We suggest that fire is essential to maintain biodiversity in Aleppo pine forests, but only at intermediate frequency.