DNA Barcoding of Invertebrates Inhabiting Olive Orchards and Vineyards Accelerates Understudied Mediterranean Biodiversity Assessment

Abstract

The Mediterranean area has a high but unevenly studied level of invertebrate diversity. Genetic-based methods, such as DNA barcoding and metabarcoding, are proposed for biodiversity assessment; however, their application is not always straightforward. The current state of data available in genetic databases limits species identification, especially in the case of certain invertebrate groups. The aim of the study was (1) to assess the diversity of seven invertebrate groups, which are potential prey for predatory arthropods, in order to facilitate the analysis of the metabarcoding of trophic interactions and, thus, expand our knowledge on biocontrol potential, and (2) to estimate the representation of local species in BOLD and NCBI GenBank databases. The DNA barcoding results consisting of sequences for 269 specimens, collected in Zadar County, within Mediterranean part of Croatia, were used for species identification and species richness assessment through the comparison of our data with the data available in BOLD and NCBI databases, and by applying species delimitation methods. Previous barcode records enabled Lepidopteran, Hemipteran, Dipteran, and Hymenopteran species identification, while Collembolan and Oligochaetes species numbers were assessed using species delimitation. Our results showed that a high number of species were unique to the study area, especially in the case of Collembolans and Oligochaetes. We confirmed that the studied area is under-researched, which was particularly evident in taxonomically demanding groups, such as Collembolans and Oligochaetes, which are rich in rare endemic species.

Mediterranean vineyards and olive groves in Croatia harbour some rare and endemic invertebrates

“…Species from the genus Corynoptera are often examples of the cryptic diversity ( Morinière et al 2019 ). There is a higher prevalence of unrecorded and ignored species in families with the lowest body sizes, implying that the number of dipteran species in Croatia is likely to be substantially larger than previously reported and, thus, new records are not surprising ( Morinière et al 2019 , Anđelić Dmitrović et al 2022 ). This genus belongs to the family Sciaridae .…”

“…This species is found throughout Europe; however, its small size makes identification difficult. The first record of this species was obtained in vineyards with IPM using the DNA barcoding method as a part of the MEDITERATRI project ( Anđelić Dmitrović et al 2022 ). This is a new record for Croatian fauna.…”

“… Sciaridae , commonly known as Dark wing fungus gnats, are a globally common, but poorly researched dipteran family ( Evenhuis et al 2016 ), since their small size and superficial homogeneity do not make them attractive to taxonomists and collectors. The first record of this species for Croatian fauna was obtained in olive orchards with IPM using the DNA barcoding method as a part of the MEDITERATRI project ( Anđelić Dmitrović et al 2022 ). Findings such as this one underline the advantages of molecular tools in species identification, such as the DNA barcoding method ( Anđelić Dmitrović et al 2022 , Chimeno et al 2022 ).…”

“…The first record of this species for Croatian fauna was obtained in olive orchards with IPM using the DNA barcoding method as a part of the MEDITERATRI project ( Anđelić Dmitrović et al 2022 ). Findings such as this one underline the advantages of molecular tools in species identification, such as the DNA barcoding method ( Anđelić Dmitrović et al 2022 , Chimeno et al 2022 ).…”

“…Species pertaining to the following groups: Araneae and Coleoptera ; Carabidae and Chrysomelidae , Hymenoptera ; Formicidae , Hemiptera , Diptera , were collected using beating stick with a net method, Tullgren Funnel method and by hand were analysed using the integrative taxonomy approach, combining morphological identification using the above-mentioned taxonomic keys and the DNA barcoding method. Total genomic DNA isolation, PCR amplification, amplicon purification, sequencing and genetic data analysis were performed as described in Anđelić Dmitrović et al (2022) . Individuals identified by the DNA barcoding method are available in the Barcode of Life Database (BOLD) ( Ratnasingham and Hebert 2007 ).…”