Collection and preservation of leafhoppers
More than 3,000 bulk samples of sap-feeding hemipteran insects were obtained between 1998-2018 through field work by the last author, his students and colleagues during surveys that aimed to document poorly studied insect faunas in various parts of the world and to obtain representatives of all major lineages of Cicadellidae for use in phylogenetic and systematic studies. This material was supplemented by the first author’s collections in Europe between 2001-2018. Specimens were collected using various methods including sweeping and vacuuming of vegetation, night collecting at lights, and in Malaise (flight intercept) traps. Specimens were collected directly into 95% ethanol in the field, returned to the laboratory and stored in -20ºC freezers at the INHS. Voucher specimens were also pinned for species identification and reference. Some samples included undescribed species from under-investigated areas, and they are waiting to be described in the context of other projects. In 2018, screening was carried out on a subset of 227 samples from independent sampling events in 28 countries (six continents) worldwide (Argentina, Australia, Bolivia, Brazil, Cameroon, Chile, China, Czech Republic, Ecuador, French Guiana, Ghana, Kyrgyzstan, India, Italy, Madagascar, Mongolia, Nicaragua, Papua New Guinea, Peru, Republic of Congo, Serbia, South Africa, Swaziland, Switzerland, Taiwan, Thailand, United States, Zambia). The land cover of the sampling events was analyzed using thematic maps within a geographic information system (QGIS 3.8, 2019) (Figure 1). Although 98% of the collections were intentionally obtained from natural areas or patches of native vegetation within more anthropogenic landscapes, we evaluated the land cover of a larger area including each sampling site using the raster layer Cropland and Pasture area (resolution 10 x 10 Km) (Ramankutty et al., 2008).
In total, the 227 samples encompassed about 1,000 specimens, with each species (or morphospecies) represented by 1 to 20 specimens belonging to the phloem-feeding leafhopper subfamily Deltocephalinae (except 1 sample belonging to the related hemipteran family Membracidae), which includes most of the previously documented vectors of phytoplasmas (Table S1). At least one specimen from each sample was selected randomly (with preference for males when present because species identification usually requires examination of male genitalia) for the molecular analyses.