10.5061/DRYAD.WDBRV15M8
Gueuning, Morgan
0000-0001-8574-9640
University of Neuchâtel
Praz, Christophe
University of Neuchâtel
Frey, Juerg
0000-0001-7377-2931
Agroscope
Data from: Ultraconserved yet informative for species delimitation: UCEs
resolve long-standing systematic enigma in Central European bees
Dryad
dataset
2020
mitochondrial introgression
Swiss Federal Office for Agriculture (FOAG)*
Swiss Federal Office for Agriculture (FOAG)
2020-08-19T00:00:00Z
2020-08-19T00:00:00Z
en
205244953 bytes
2
CC0 1.0 Universal (CC0 1.0) Public Domain Dedication
Accurate and testable species delimitation hypotheses are essential for
measuring, surveying and managing biodiversity. Today, taxonomists often
rely on mitochondrial DNA barcoding to complement morphological species
delimitations. Although COI barcoding has largely proven successful in
assisting identifications for most animal taxa, there are nevertheless
numerous cases where mitochondrial barcodes do not necessarily reflect the
species history. For instance, what is regarded as one single species can
be associated with two distinct DNA barcodes, which can point either to
cryptic diversity or to deep within-species mitochondrial divergences with
no reproductive isolation. In contrast, two or more species can share
barcodes, for instance due to mitochondrial introgression. These intrinsic
limitations of mitochondrial DNA barcoding can only be addressed with
nuclear genomic markers, which are expensive, labour intensive, poorly
repeatable, and often require high-quality DNA. To overcome these
limitations, we examined the use of ultraconserved nuclear genetic
elements (UCEs) as a quick and robust genomic approach to address such
problematic cases of species delimitation. This genomic method was
assessed using six different bee species complexes suspected to harbour
cryptic diversity, mitochondrial introgression, or mitochondrial
paraphyly. The sequencing of UCEs recovered between 686 and 1860
homologous nuclear loci and provided explicit species delimitation
hypotheses in all investigated species complexes. These results provide
strong evidence for the suitability of UCEs as a fast method for species
delimitation even in recently diverged lineages. Furthermore, this study
provided the first conclusive evidence for both mitochondrial
introgression among distinct species, and mitochondrial paraphyly within a
single bee species.
UCE library preparation Whole body DNA extractions were performed
overnight in a proteinase K buffer at 56°C and purified using a Qiagen
Biosprint 96 extraction robot following the manufacturer’s protocol.
Extracts were quantified using Qubit v3 (Thermofisher Scientific) and 50
ng DNA per specimen were sonicated to 500 bp fragment length using a
Bioruptor ultrasonicator (Diagenode). Two independent dual-indexed
libraries each containing 96 specimens were constructed using a Kapa Hyper
prep kit (Roche) using one fourth of the manufacturer’s recommended
volumes (as described in Branstetter, Longino, Ward, & Faircloth,
2017). PCR amplifications were performed in the recommended volumes. PCR
products were quantified using a Qubit v3 and each row of a 96-well PCR
plate were pooled equimolarly (i.e. for total of 8 pools). Libraries were
UCE enriched using the Hymenopteran v2 hybridization kit (UCE Hymenoptera
2.5Kv2 Principal/Full, myBaits, Arborbiosci). Each enrichment was
performed on a single pool of 12 specimens using 500 ng. The enrichment
protocol followed the manufacturer’s recommendations with a hybridization
step of 24 h at 65°C, followed by a PCR amplification with 14 cycles.
Pools were sequenced on a Miseq using the Illumina v3 kits (2 x 300 bp;
Illumina). Bioinformatic processing of UCE data Fastq reads were
demultiplexed on the Miseq and data from all runs were merged and
processed mainly using PHYLUCE tools (Faircloth, 2016). Raw data were
cleaned with illumprocessor (Faircloth, 2016), a tool wrapped around
trimmomatic (Bolger, Lohse, & Usadel, 2014). Clean reads were
assembled with SPAdes v3.12.0 (Nurk et al., 2013) using the single-cell
flag (“--sc”), careful option (“--careful”) and a coverage cutoff value of
five (“--cov-cutoff”). Obtained contigs were mapped against the
corresponding UCE reference file using Lastz (Harris, 2007) and matching
reads were extracted and aligned by species complex using MAFFT (Katoh
& Standley, 2013). Alignments were edge-trimmed using the PHYLUCE
“seq-cap” program; a strategy recommended for closely related species
(< 30-50 MYA) (Faircloth, 2016). Loci shared by less than 75% of
the maximum number of specimens were filtered out. Remaining alignments
were concatenated and saved in fasta format. An additional filtering step
was applied to remove specimens with more than 90% missing data.