Determining putative vectors of the Bogia Coconut Syndrome phytoplasma using loop-mediated isothermal amplification of single-insect feeding media

Abstract

Phytoplasmas are insect vectored mollicutes responsible for disease in many economically important crops. Determining which insect species are vectors of a given phytoplasma is important for managing disease but is methodologically challenging because disease-free plants need to be exposed to large numbers of insects, often over many months. A relatively new method to detect likely transmission involves molecular testing for phytoplasma DNA in sucrose solution that insects have fed upon. In this study we combined this feeding medium method with a loop-mediated isothermal amplification (LAMP) assay to study 627 insect specimens of 11 Hemiptera taxa sampled from sites in Papua New Guinea affected by Bogia coconut syndrome (BCS). The LAMP assay detected phytoplasma DNA from the feeding solution and head tissue of insects from six taxa belonging to four families: Derbidae, Lophopidae, Flatidae and Ricaniidae. Two other taxa yielded positives only from the heads and the remainder tested negative. These results demonstrate the utility of combining single-insect feeding medium tests with LAMP assays to identify putative vectors that can be the subject of transmission tests and to better understand phytoplasma pathosystems.

Phytoplasmas–The “Crouching Tiger” Threat of Australian Plant Pathology

“…This approach offers the advantages of low cost and high sensitivity, low risk of cross-contamination because reaction vessels do not need to be opened and, most especially, use in a kit form that requires very little training and can be used in the field (Hodgetts et al, 2011 ; Dickinson, 2015 ; Kogovšek et al, 2015 ). Though LAMP assays have yet to be used in studies of Australian phytoplasma pathosystems they have recently proven useful in studies of Bogia coconut syndrome in nearby Papua New Guinea (Lu et al, 2016 ).…”

“…A relatively new method that is intermediate in ease of use and the level of proof involves holding individual insects in vessels from which they can feed on a sucrose solution through a parafilm barrier with subsequent PCR-based detection of phytoplasma DNA in the medium (Tanne et al, 2001 ). Though this approach involves many samples (i.e., one per insect rather than one per plant as in the case of a cage transmission test) a recent study has employed LAMP to readily handle the large numbers of samples associated with screening multiple putative vector species of Bogia coconut syndrome (Lu et al, 2016 ).…”

Jikradia olitoria([Hemiptera]:[Cicadellidae]) Transmits the Sequevar NAGYIIIβ Phytoplasma Strain Associated with North American Grapevine Yellows in Artificial Feeding Assays

“…For this reason, the detection of phytoplasma DNA in insects is not a sufficient indication of their competency to transmit phytoplasmas and was our rationale for not analyzing insects themselves for phytoplasmas. The use of sucrose feeding solutions to screen potential vectors is an established method that offers advantages over the conventional plant transmission trials: it reduces the time of the investigation by avoiding the need to wait for symptoms, reduces costs, and allows for individual assessment of insects as potential vectors (Tanne et al 2001, Bressan et al 2006, Lu et al 2016). Of 49 field-collected insect species screened, J. olitoria was the only insect that secreted a NAGY phytoplasma into the sucrose feeding solution.…”

Potential role of the alien planthopper Ricania speculum as vector of Flavescence dorée phytoplasma

“…Indeed, the ability to transmit phytoplasmas depends on the pathogen -plant host -insect vector combination, and several factors, such as feeding behavior and host-pathogen molecular recognition, are involved in mediating phytoplasma transmission capability by an insect species (Bosco and D'Amelio 2010;Galetto et al 2011a). Consider for example the two Ricanidae species suspected to transmit BCS phytoplasma in Papua New Guinea, the pathogen was detected in both species and also in artificial media used to feed insects of one of the two species, indicating that acquisition occurred but transmission did not (Lu et al 2016). Similar situations have been reported also for FD phytoplasma, which was detected in several field collected Hemiptera insect species (Psylla spp., Phlogotettix cyclops, Cixius nervosus, Metcalfa pruinosa), but in absence of effective transmission evidence (Trivellone 2019).…”

“…Interestingly, Vitis spp. and C. vitalba are well-known plant hosts of the grapevine Flavescence dorée phytoplasma (FDp, 16SrV-C and -D) (EFSA 2016) and, since two unidentified Ricaniidae species were suspected to transmit Bogia coconut syndrome (BCS) phytoplasma in Papua New Guinea (Lu et al 2016), concerns were raised on this alien species as potential vector of FDp.…”