Heat shock-induced physical changes of megaplasmids in Rhodococcus sp. strain DK17

Rhodococcus sp. strain DK17 possesses three megaplasmids (380 kb pDK1, 330 kb pDK2, and 750 kb pDK3). The alkylbenzene-degrading genes (akbABCDEF) are present on pDK2 while the phthalate operons which are duplicated are present on both pDK2 (ophA'B'C'R') and pDK3 (ophABCR). DK17 with an optimal temperature of 30°C showed no growth at 37°C. When transferred to 30°C, however, the 37°C culture began to grow immediately, indicating that 37°C is not lethal but stressful for DK17 growth. In addition, when exposed to 37°C even for a short time, a part of DK17 cells lost the ability to degrade o-xylene (a model compound of alkylbenzenes). When two hundred colonies were randomly selected for colony PCR for pDK2-specific akbC, ophC', or pDK3-specific ophC, a total of 29 colonies were found to have lost at least one of the three genes. PFGE analysis clearly showed that all the mutants have different megaplasmid profiles from that of DK17 wild type, which are divided into five different cases: Type I (10 mutants, pDK2 loss and acquisition of a new ~700 kb plasmid), Type II (9 mutants, pDK2 loss), Type III (8 mutants, pDK3 loss and acquisition of a new ~400 kb plasmid), Type IV (1 mutant, pDK3 loss), and Type V (1 mutant, pDK2 and pDK3 loss and acquisition of the ~400 kb and ~700 kb plasmids). The above results showing that growth temperature changes can induce physical changes in bacterial genomes suggest that environmental changes in habitats including temperature fluctuations affect significantly the evolution of bacteria.