The small heat shock gene ('shsp') cluster of 'Drosophila buzzatii' was sequenced and the gene order and DNA sequence were compared with those of the 'shsps' in 'Drosophila melanogaster'. The 'D. buzzatii shsp' cluster contains an inversion and a duplication of 'hsp26'. A phylogenetic tree was constructed based on 'hsp26' genes from several 'Drosophila' species of the 'Sophophora' and 'Drosophila subgenera'. The tree shows first a separation of the 'Sophophora' and the 'Drosophila subgenera' and then the 'Drosophila' subgenus is divided into the Hawaiian 'Drosophila' and the 'repleta/virilis' groups. Only the latter contain a duplicated 'hsp26'. Comparing the gene organisation of the 'shsp' cluster shows that all the 'Drosophila' subgenus species contain the inversion. Putative heat shock elements (HSE) were found in the promoters of all the 'shsp' and putative regulator elements for tissue specific expression were found in the promoter of 'hsp23', 'hsp27' and one of the 'hsp26' genes. 'hsp23' was found to be polymorphic for four non-synonymous changes that all lead to exchange of a Valine. The duplicated 'hsp26' gene in 'D. buzzatii' '(phsp26)' was polymorphic for two non-synonymous changes. The allele frequencies of these variants were determined in nine 'D. buzzatii' populations covering most of its distribution in Australia using high-resolution melting curves. The allele frequencies of one of the 'hsp23' variants showed a significant linear regression with longitude and the pooled frequency of the four Valine changes of 'hsp23' in the nine populations showed a significant linear regression with longitude and with a composite measure of climatic variables.