HOPS: Automated detection and authentication of pathogen DNA in archaeological remains

Abstract

High-throughput DNA sequencing enables large-scale metagenomic analyses of complex biological systems. Such analyses are not restricted to present day environmental or clinical samples, but can also be fruitfully applied to molecular data from archaeological remains (ancient DNA), and a focus on ancient bacteria can provide valuable information on the long-term evolutionary relationship between hosts and their pathogens. Here we present HOPS (Heuristic Operations for Pathogen Screening), an automated bacterial screening pipeline for ancient DNA sequence data that provides straightforward and reproducible information on species identification and authenticity. HOPS provides a versatile and fast pipeline for high-throughput screening of bacterial DNA from archaeological material to identify candidates for subsequent genomic-level analyses.

Hybridisation capture allows DNA damage analysis of ancient marine eukaryotes

“…To further increase the Eukaryota signal in future studies, it may be beneficial to add a larger volume of baits (> 3 µL) to sedaDNA extracts expected (e.g., from shallow shotgun sequencing prior to enriching) to have a relatively high Eukaryota sedaDNA content. The HOPS bioinformatic tool 23 proved highly valuable in identifying and analysing ancient eukaryote sequences in our sedaDNA. The HOPS generated output of our 'Eukaryota' (taxa list a) run enabled the determination of '% eukaryote sedaDNA damage' , a parameter that can be used as a proxy of sedaDNA authenticity in the future.…”

“…The red and blue lines denote C > T substitutions in 5′ direction and G > A substitutions in 3′ direction, respectively, for all ancient alignments. Grey lines denote estimated noise 23 www.nature.com/scientificreports/ investigate how the % eukaryote sedaDNA damage varies in much older sediment records (older than Holocene) and depending on sediment properties (e.g., clay-rich sediments that appear to benefit DNA preservation 38 ). The strong positive correlation between % eukaryote sedaDNA damage amongst Shotgun, Planktonbaits1 and HABbaits1, demonstrates that DNA damage signals present in sedaDNA are preserved throughout the hybridisation capture approach.…”

“…There is a lack of high-quality reference sequences for the thousands of marine organisms occurring in the global ocean, and the threshold of ~ 250 reads per species required to analyse and plot DNA damage patterns in mapDamage 22 is often not reached in sedaDNA. Recently, Hübler et al 23 developed a new bioinformatic tool HOPS-'Heuristic Operations for Pathogen Screening'-based on the mapDamage algorithm, to identify and authenticate bacterial pathogens in ancient metagenomic samples and extract this information for further downstream analysis. In combination with hybridisation capture to generate a larger number of ancient eukaryote sequences, HOPS has the potential to allow the assessment of sedaDNA authenticity based on DNA damage profiles from key marine eukaryotes, even if only very few sequences are available (≥ 50 reads per species 23 ).…”

“…Ancient DNA authenticity assessment and damage analysis. To assess the authenticity of our Shotgun, Plank-tonbaits1 and HABbaits1 sedaDNA we ran the 'MALTExtract' and 'Postprocessing' tools of the HOPS v0.33-2 pipeline 23 . The latter included the use of the NCBI mapping and NCBI tree files (13 Nov 2019) provided with HOPS (https ://githu b.com/rhueb ler/HOPS/tree/exter nal/Resou rces).…”

“…Configurations deviating from the default HOPS settings included topMaltEx = 0.10, minPIdent = 95, meganSummary = 1, and destackingOff = 1. We processed each dataset using the 'def_anc' mode, which provided results for all filtered reads ('default') as well as all reads that had at least one damage lesion in their first 5 bases from either the 5′ or 3′ end ('ancient') 23 . Generally, HOPS determines DNA damage patterns separately for individual taxa, i.e., requires an input list of target taxa for which to compare the sedaDNA sequences identified in our samples to their modern references.…”