Fast and sensistive on-site isothermal assay (LAMP) for diagnosis and detection of three fruit tree phytoplasmas

Over the years, realtime PCR outflanked endpoint PCR in phytopathogen
diagnostics, mainly because of the increase in sensitivity and timesaving
aspects of the technique. However, a time consuming 16S rRNAbased
nested PCR method is still the gold standard for phytoplasma diagnosis. This is also the case for phytoplasma detection in Malus, Pyrus and Prunus, the three main host plants of apple proliferation (AP), pear decline (PD) and European stone
fruit yellows (ESFY) phytoplasma, respectively. The last decade, loop mediated
isothermal amplification (LAMP) (Notomi et al. 2000 ) is gaining a lot in significance and is also for phytoplasmas expected to become a widely
used reliable diagnostic tool. High specificity and sensitivity which also
requires a less stringent need for DNA purification, and the short analysis time
and the limited equipment requirements makes the LAMP method a fast and
affordable alternative with great point of care diagnostic potential. In this paper, we present a LAMP primer set for the ribosomal group 16SrX, containing the important fruit tree phytoplasmas AP, PD and ESFY. The primers were developed and validated for fast and sensitive detection and general use for diagnosis. We foresee that the LAMP technique will also have its application in on site
diagnosis of the fruit tree phytoplasmas during inspections and surveys.