Transformation of crustacean pathogenic bacterium Spiroplasma eriocheiris and expression of yellow fluorescent protein

- A transformation method is developed for Spiroplasma eriocheiris.
- The transformation is achieved using oriC plasmid haboring S. eriocheiris specific promoters.
- Fluorescent protein useful for protein tagging is expressed in S. eriocheiris.
- Methods developed here should be useful to identify genes involved in swimming and trace those proteins.

Spiroplasma eriocheiris, the cause of crab trembling disease, is a wall-less bacterium, related to Mycoplasmas, measuring 2.0-10.0 μm long. It features a helical cell shape and a unique swimming mechanism that does not use flagella; instead, it moves by switching the cell helicity at a kink traveling from the front to the tail. S. eriocheiris seems to use a novel chemotactic system that is based on the frequency of reversal swimming behaviors rather than the conventional two-component system, which is generally essential for bacterial chemotaxis. To identify the genes involved in these novel mechanisms, we developed a transformation system by using oriC plasmid harboring the tetracycline resistant gene, tetM, which is under the control of a strong promoter for an abundant protein, elongation factor-Tu. The transformation efficiency achieved was 1.6 × 10−5 colony forming unit (CFU) for 1 μg DNA, enabling the expression of the enhanced yellow fluorescent protein (EYFP).