Phytoplasma cultivation from a micropropagated plant collection

In contrast to mycoplasmas, which cause an array of disorders in animals and humans, phytoplasmas resisted all attempts to culture them in cell-free media. Despite reduced genome size in comparison to their ancestors, they retain an independent metabolism that allows them to survive in environments as diverse as plant phloem and insect haemolymph. This versatility is a unique property among microbes, shared only with some animal- or plant-infecting viruses and a few other microorganisms such as the causal agent of malaria. Very recently the proof that phytoplasmas can now be grown on laboratory media was provided employing specific commercially available media. Several key points were important to achieve this result: plant tissue selected for isolation must contain viable phytoplasmas; release of phytoplasmas must be achieved from the sieve-tubes in the presence of sieve-tube sap coagulation mechanisms to prevent loss on trauma, the incubation period must be of sufficient time to allow evidence of phytoplasma growth to manifest itself. Inadequacy on any of these points would result in failure so there are many reasons, apart from a suitable medium, that could be responsible for previous failures. Besides the initial seven phytoplasmas, six additional strains were grown: ‘Candidatus Phytoplasma trifolii’ (strain PWB, ribosomal group 16SrVI-A), ‘Ca. P. aurantifolia’ (strain WBDL, ribosomal group 16SrII-B), ‘Ca. P. pruni’ (strain CX, ribosomal group 16SrIII-A), ‘Ca. P asteris’ (strain KVE, ribosomal group 16SrI-C), ‘Ca. P. phoenicium’ (strain PEY, ribosomal group 16SrIX-C), and ‘Ca. P. prunorum’ (strain PLNV6, ribosomal group 16SrX-B). Additional types of phytoplasmas are in the initial isolation stage, but it appears that all the available phytoplasma strains can be cultured. Contrary to the prevailing dogma in plant pathology, therefore, phytoplasmas, like mycoplasmas, can indeed be grown independently from the host(s). The results will produce a more detailed knowledge about basic mechanisms that regulate the survival of phytoplasmas, which are among the smallest known living organisms, but can induce among the most severe epidemics in agriculture.