DNA-based discrimination and frequency of phytoplasma infection in the two hawthorn-feeding species, Cacopsylla melanoneura and Cacopsylla affinis, in northwestern Italy.

A molecular tool, focused on the mitochondrial Control Region (CR), was developed to discriminate the two hawthorn-feeding psyllid species, Cacopsylla melanoneura (Förster) and C. affinis (Löw), and to estimate their frequencies in mixed populations. The test was carried out in paired and single-tube amplifications and validated analysing 52 male specimens previously determined by morphological analysis. The frequencies of the two species in mixed populations in the Aosta Valley (northwestern Italy) were analysed. The presence and type of 16SrX-group phytoplasmas was detected by nested PCR and RFLP tests in both species. C. melanoneura was the predominant species (86.5%; 80.4-91.2 CI); of these, 0.9% of the samples were positive for 'Ca. Phytoplasma mali' and 1.8% for 'Ca. Phytoplasma pyri'. One of 21 specimens of C. affinis was positive for 'Ca. Phytoplasma pyri'. The test also allowed us to identify two genetic variants of C. melanoneura, depending on the presence or absence of a 56 bp indel; these were named WI (with indel) and WOI (without indel), respectively. Further analyses were carried out on C. melanoneura specimens collected in apple orchards at six different locations in northern Italy where different levels of transmission efficiency have been described. Our preliminary observations suggest that some differences might exist between the two genetic variants in their ability to transmit phytoplasmas and to colonise different host plants.