DOI: 10.5061/dryad.s1rn8pk95
title: Metabarcoding of canopy arthropods reveals negative impacts of forestry insecticides on community structure across multiple taxa

description: 1. Insecticides used to combat outbreaks of forest defoliators can
 adversely affect non-target arthropods. Forest use insecticides
 typically suppress Lepidoptera larvae which are the keystone of the canopy
 community of deciduous oak forests. The abrupt removal of this dominant
 component of the food web could have far-reaching implications for forest
 ecosystems, yet it is rarely investigated in practice owing to several
 methodological shortcomings. The taxonomic impediment and the biased
 nature of arthropod sampling techniques particularly impede the assessment
 of insecticide impacts on diverse communities. 2. To tackle this issue, we
 propose an experimental approach combining pyrethrum knockdown sampling
 and species determination via DNA metabarcoding, using community
 subsampling to derive estimates of species abundances. We applied this
 protocol to investigate the short-term effects of the insecticides
 diflubenzuron (DFB) or Bacillus thuringiensis var. kurstaki (BTK) on
 canopy-dwelling arthropod communities in German oak woodlands. 3. Our
 approach allowed us to include most of the detected diversity and
 integrate species abundances in our analyses. By classifying arthropod
 species into assemblages based on their expected sensitivity rather than
 coarse taxonomic groupings, we could unveil substantial effects of DFB
 across multiple taxa five weeks after application. 4. Although strong
 effects on single species appear related to direct toxicity, substantial
 impacts of DFB on parasitoids and xylophagous beetles suggest that
 anti-defoliator treatments can have previously unsuspected indirect
 effects on some components of forest arthropod communities. The impacts of
 BTK on community structure were consistent with but much weaker than that
 of DFB. 5. Synthesis and applications. Comparing diversity patterns in the
 arthropod communities of sprayed and unsprayed oak canopies, our results
 show that selective insecticides can alter species diversity in presumably
 non-sensitive taxa. Even though the ecological significance of these
 impacts has yet to be assessed in an operational setting, their existence
 calls for increased regulatory scrutiny on indirect effects. As community
 approaches become more attainable with the rapid development of DNA
 metabarcoding, we suggest the inclusion of community level
 endpoints as regulatory requirements for the approval of forest use
 insecticides.
description: <strong>Sampling</strong>. Arthropods were sampled by
 pyrethrum knockdown using Swingfog SN-50 fogging machines and a 1%
 pyrethrum suspension in petroleum white oil. After 30 minutes exposure,
 all arthropods fallen on four 15 m<sup>2</sup> tarpaulin
 sheets placed below the crown were collected and stored in ethanol.  Forty
 oak trees in 10 blocks distributed over three stands with high gradation
 of the defoliator <em>Lymantria dispar</em> (Lepidoptera:
 Erebidae) were sampled. In each block, one tree had been treated with
 Bacillus thuringiensis var. kurstaki, one with diflubenzuron, and two were
 not sprayed (controls). Spraying was conducted mid-May at the maximum
 legal application rate. All trees were sampled once five weeks after
 spraying. The abundance of <em>Lymantria dispar</em> was
 monitored on each tree by counting caterpillars below tree
 bands. <strong>Species
 determination</strong>. Arthropods were sorted at high taxonomic
 rank (mostly order and sub-order; A) and split into two groups based on
 how easily they can be identified to species by available expert
 taxonomists (C1). Taxa belonging to the 'challenging' group were
 determined via DNA metabarcoding after samples were fractioned into 12
 subsamples per tree (B). Sequence read abundances were filtered to remove
 artefactual and contaminant sequences and species abundances were
 estimated at the tree level by pooling the subsample level incidence data
 (C2). Data processing protocols are provided in details
 in the associated publication.
description: The dataset includes four files, each representing an
 intermediate step of the data processing pipeline (see letters
 corresponding to each file in the method description). 
 (A) Sorting data (subsample level; only taxa later indentified by
 metabarcoding); file name: JAPPL-2021-00659_A_Data.csv
 (B) Raw metabarcoding species data with sequence reads abundance
 (subsample level); file name: JAPPL-2021-00659_B_Data.csv
 (C1) Derived metabarcoding data with species abundance estimates
 (tree level); file name: JAPPL-2021-00659_C1_Data.csv
 (C2) Morphologically identified taxa with species abundance (tree
 level); file name: JAPPL-2021-00659_C2_Data.csv Note
 that due to a pyrethrum dosing mistkaes, sampling failed on four blocks
 and these samples were excluded from the analyses as they were not
 comparable with the other samples. The excluded samples are only included
 in the dataframe A. Each data file comes with a data
 structure file (prefix Str) detailing each variable.