A putative Type IIS restriction endonuclease GeoICI from Geobacillus sp. – A robust, thermostable alternative to mezophilic prototype BbvI.

Screening of extreme environments in search for novel microorganisms may lead to the discovery of robust enzymes with either new substrate specificities or thermostable equivalents of those already found in mesophiles, better suited for biotechnology applications. Isolates from Iceland geysers’ biofilms, exposed to a broad range of temperatures, from ambient to close to water boiling point, were analysed for the presence of DNA-interacting proteins, including restriction endonucleases (REases). GeoICI, a member of atypical Type IIS REases, is the most thermostable isoschizomer of the prototype BbvI, recognizing/cleaving 5 -GCAGC(N8/12)-3 DNA sequences. As opposed to the unstable prototype, which cleaves DNA at 30°C, GeoICI is highly active at elevated temperatures, up to 73°C and over a very wide salt concentration range. Recognition/cleavage sites were determined by: (i) digestion of plasmid and bacteriophage lambda DNA (λ); (ii) cleavage of custom PCR substrates, (iii) run-off sequencing of GeoICI cleavage products and (iv) shotgun cloning and sequencing of λ DNA fragmented with GeoICI. Geobacillus sp. genomic DNA was PCR-screened for the presence of other specialized REases-MTases and as a result, another putative REase- MTase, GeoICII, related to the Thermus sp. family of bifunctional REases-methyltransferases (MTases) was detected.

Arapidly progressing, deadly disease of Actias selene (Indianmoonmoth) larvae associated with a mixed bacterial and baculoviral infection.

The outbreak of an infectious disease in captive-bred Lepidoptera can cause death of all the caterpillars within days. A mixed baculoviral–bacterial infection observed among Actias selene (Hübner 1807), the Indian moon moth (Insecta: Lepidoptera: Saturniidae), larvae was characterized and followed by a photographic documentation of the disease progression. The etiological agents were determined using mass spectrometry and polymerase chain reaction (PCR). It appeared that the disease was caused by a mixed infection of larvae with a baculovirus and Morganella morganii. A molecular phylogenetic analysis of the virus and microbiological description of the pathogenic bacterium are presented.