DNA metabarcoding for diet analysis and biodiversity: A case study using the endangered Australian sea lion (Neophoca cinerea)

Abstract

The analysis of apex predator diet has the ability to deliver valuable insights into ecosystem health, and the potential impacts a predator might have on commercially relevant species. The Australian sea lion (Neophoca cinerea) is an endemic apex predator and one of the world's most endangered pinnipeds. Given that prey availability is vital to the survival of top predators, this study set out to understand what dietary information DNA metabarcoding could yield from 36 sea lion scats collected across 1,500 km of its distribution in southwest Western Australia. A combination of PCR assays were designed to target a variety of potential sea lion prey, including mammals, fish, crustaceans, cephalopods, and birds. Over 1.2 million metabarcodes identified six classes from three phyla, together representing over 80 taxa. The results confirm that the Australian sea lion is a wide‐ranging opportunistic predator that consumes an array of mainly demersal fauna. Further, the important commercial species Sepioteuthis australis (southern calamari squid) and Panulirus cygnus (western rock lobster) were detected, but were present in <25% of samples. Some of the taxa identified, such as fish, sharks and rays, clarify previous knowledge of sea lion prey, and some, such as eel taxa and two gastropod species, represent new dietary insights. Even with modest sample sizes, a spatial analysis of taxa and operational taxonomic units found within the scat shows significant differences in diet between many of the sample locations and identifies the primary taxa that are driving this variance. This study provides new insights into the diet of this endangered predator and confirms the efficacy of DNA metabarcoding of scat as a noninvasive tool to more broadly define regional biodiversity.

Environmental DNA (eDNA) metabarcoding reveals strong discrimination among diverse marine habitats connected by water movement

“…These observations indicate that eDNA, at least in the study area used here, is only transported over small distances (~100 to 200 m), without heavily influencing eDNA metabarcoding surveys, consistent with results from the one other vertebrate eDNA survey that has been conducted to date in a highly dynamic coastal environment (Port et al, ). However, these eDNA signals might also be originating from predator faecal matter (Berry et al, ; Deagle, Kirkwood, & Jarman, ), and, hence, not the result of eDNA signal dispersal.…”

“…Library preparation followed the protocol described in (Berry et al, ; Stat et al, ). Briefly, samples were analyzed using three metabarcoding assays targeting two fragments of the 16S rRNA gene region and one fragment of the cytochrome c oxidase subunit I (COI) gene region (Supporting information Table S2).…”

Environmental DNA metabarcoding studies are critically affected by substrate selection

“…Quantitative polymerase chain reaction (qPCR) was performed on isolated DNA to ascertain if samples exhibited inhibition and to determine the optimal level of input DNA (Murray et al, ). Three PCR assays were used for this quantitation at GI and AB: 16SFD/16S2R‐Degenerate (Berry et al, ; Deagle et al, ; herein referred to as 16S fish), Crust16S_F/Crust16S_R (Berry et al, ; herein referred to as 16S crustacean) targeting the 16S rRNA region of the mitochondrial genome, and a universal eukaryote assay: 18S1F/400R (Pochon, Nathan, Smith, & Wood, ) targeting the nuclear 18S rRNA V1‐3 hypervariable region. In addition to the 18S assay, a degenerate COI assay: m1COIintF/jgHCO2198 (Leray et al, ) was used on the Caspian Sea samples (all primer sequences, conditions and references can be found in Supporting Information Table S1).…”

Non‐specific amplification compromises environmental DNA metabarcoding with COI

“…Early eDNA metabarcoding studies targeting fishes used the cytochrome b gene (Minamoto, Yamanaka, Takahara, Honjo, & Kawabata, ; Thomsen, Kielgast, Iversen, Møller, et al, ; Thomsen, Kielgast, Iversen, Wiuf, et al, ), but more recent studies have used the 12S ribosomal rRNA locus (Hänfling et al, ; Kelly et al, ; Port et al, ; Stoeckle, Soboleva, & Charlop‐Powers, ; Ushio et al, ; Yamamoto et al, ), and also 16S rRNA (Berry et al, ; Bylemans, Gleeson, Hardy, & Furlan, ; Jeunen et al, ; Shaw et al, ; Stat et al, ). Various regions of 12S have been proposed as metabarcoding markers, including a ca.…”