Genome sequencing and assembly
We sequenced the Polyplax lice pooled sample on one lane of
Illumina HiSeq2000 (GeneCore, Heidelberg, Germany) using 2×100
paired-end reads. Read quality was checked using FastQC (Andrews, 2010)
and quality trimming was performed using the BBtools package
(https://jgi.doe.gov/data-and-tools/bbtools).
The resulting dataset contained 309 892 186 reads. We used SPAdes
assembler 3.10 (Bankevich et al., 2012) to build the assembly,
implementing careful options and enabling mismatch corrections. To check
for bacterial plasmid(s) we submitted the complete assembly (124 985
contigs) to the PlasmidFinder (Carattoli et al., 2014) with sensitivity
set to three different thresholds (95%, 85%, and 60%). We identified
bacterial contigs by blasting Snodgrassella alvi wkB2 genome
against the assembly using custom blast in the program Geneious (Kearse
et al., 2012). This procedure retrieved 39 contigs which were putatively
assigned to Neisseriales and their origin was further confirmed by blast
analyses of individual genes as specified below.
To sequence a complete Hoplopleura acanthopus lice metagenome, we
employed Illumina MiSeq (GeneCore, Heidelberg, Germany) and
Oxford-Nanopore (University of Urbana, Illinois, USA) technology. We
constructed the Illumina library from the total DNA of 35 individuals
and sequenced it in four runs of Illumina MiSeq using V2 500 cycle
chemistry. We used the same procedure for quality checking and filtering
as described for the Polyplax data set. The resulting number of
reads was 34 406 078. We used high molecular weight DNA from 5 H.
acanthopus as a template for Oxford-Nanopore sequencing on GridIONx5.
The total number of reads was 1 653 194. The quality of Nanopore reads
was checked using NanoPack tools (De Coster, D’Hert, Schultz, Cruts, &
Van Broeckhoven, 2018) and quality filtering was performed using
Filtlong (https://github.com/rrwick/Filtlong). To assemble theH. acanthopus metagenome, we employed a hybrid approach combining
the Illumina and Nanopore data (for details see
SupplementaryInformation/SupplementaryText).
We annotated genomes of both Neisseria -related symbionts using
RAST (Aziz et al., 2008) and deposited them in GenBank under the
accession numbers CP046107 (closed genome of the symbiont from H.
acanthopus ) and WNLJ00000000 (draft genome of Polyplax serratasymbiont in 39 contigs). To assess phylogenetic origin of individual
genes, we blasted each gene against the nr database and categorised its
putative origin based on the returned hits (for details see
SupplementaryInformation/SupplementaryText).