Determining the diet of larvae of western rock lobster (Panulirus cygnus) using high-throughput DNA sequencing techniques.

Richard O'Rorke,Andrew G Jeffs,Anya M Waite,Peter Tsai,Shane Lavery,Haruko Takeyama,Lynnath E Beckley,Peter A Thompson,Seinen Chow,Senjie Lin

doi:10.1371/journal.pone.0042757

Abstract

The Western Australian rock lobster fishery has been both a highly productive and sustainable fishery. However, a recent dramatic and unexplained decline in post-larval recruitment threatens this sustainability. Our lack of knowledge of key processes in lobster larval ecology, such as their position in the food web, limits our ability to determine what underpins this decline. The present study uses a high-throughput amplicon sequencing approach on DNA obtained from the hepatopancreas of larvae to discover significant prey items. Two short regions of the 18S rRNA gene were amplified under the presence of lobster specific PNA to prevent lobster amplification and to improve prey amplification. In the resulting sequences either little prey was recovered, indicating that the larval gut was empty, or there was a high number of reads originating from multiple zooplankton taxa. The most abundant reads included colonial Radiolaria, Thaliacea, Actinopterygii, Hydrozoa and Sagittoidea, which supports the hypothesis that the larvae feed on multiple groups of mostly transparent gelatinous zooplankton. This hypothesis has prevailed as it has been tentatively inferred from the physiology of larvae, captive feeding trials and co-occurrence in situ. However, these prey have not been observed in the larval gut as traditional microscopic techniques cannot discern between transparent and gelatinous prey items in the gut. High-throughput amplicon sequencing of gut DNA has enabled us to classify these otherwise undetectable prey. The dominance of the colonial radiolarians among the gut contents is intriguing in that this group has been historically difficult to quantify in the water column, which may explain why they have not been connected to larval diet previously. Our results indicate that a PCR based technique is a very successful approach to identify the most abundant taxa in the natural diet of lobster larvae.

Highlights

Despite considerable research into the biology of spiny lobsters (Family Palinuridae), the larval phase of their lifecycle remains enigmatic
Further work is required to ascertain the optimal food for phyllosomata and whether oceanographic processes impact the density of optimal prey groups and/or the ability of phyllosomata to capture these prey
Attempts to culture phyllosomata have found that feed is a strong determinate of larval viability, and supplementation of feed with mussel gonad has been found to be essential for phyllosomata to advance through multiple instars (Kittaka 1997)

Summary

Introduction

Despite considerable research into the biology of spiny lobsters (Family Palinuridae), the larval phase of their lifecycle remains enigmatic. Spiny lobsters have an unusually long planktonic larval phase and P. cygnus has an estimated larval duration of 9 to 11 months that is spent in oceanic waters extending from the continental shelf margin to over 1,500 km offshore from Western Australia [3]. After this oceanic phase the larvae, which are known as phyllosomata (singular: phyllosoma), are thought to be carried shoreward by ocean currents and eventually undergo metamorphosis into nektonic post-larvae, or pueruli, which actively migrate back onshore [4]. The underpinning causal details of how these oceanic events impact lobster ecology and recruitment to the coastal benthic stock remain to be fully established

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Conclusion