Cross-species chromosome painting can directly visualize syntenies between diverged karyotypes and, thus, increase our knowledge on avian genome evolution. DNA libraries of chicken (Gallus gallus, GGA) macrochromosomes 1 to 10 were hybridized to metaphase spreads of 9 different species from 3 different orders (Anseriformes, Gruiformes and Passeriformes). Depending on the analyzed species, GGA1–10 delineated 11 to 13 syntenic chromosome regions, indicating a high degree of synteny conservation. No exchange between the GGA macrochromosome complement and microchromosomes of the analyzed species was observed. GGA1 and GGA4 were distributed on 2 or 3 chromosomes each in some of the analyzed species, indicating rare evolutionary rearrangements between macrochromosomes. In all 6 analyzed species of Passeriformes, GGA1 was diverged on 2 macrochromosomes, representing a synapomorphic marker for this order. GGA4 was split on 2 chromosomes in most karyotypes, but syntenic to a single chromosome in blackcap (Passeriformes). GGA5/10 and also GGA8/9 associations on chromosomes were found to be important cytogenetic features of the Eurasian nuthatch (Passeriformes) karyotype. Fusion of GGA4 and GGA5 segments and of entire GGA6 and GGA7, respectively, was seen in the 2 analyzed species of Gruiformes. Consistent with the literature, our inter-species chromosome painting demonstrates remarkable conservation of macrochromosomal synteny over 100 million years of avian evolution. The low rate of rearrangements between macrochromosomes and the absence of detectable macrochromosome-microchromosome exchanges suggests a predominant role for rearrangements within the gene-dense microchromosome complement in karyotypic diversification.