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.

The increasing sensitivity of PCR has meant that in the last two decades PCR has emerged as a major tool in diet studies, enabling us to refine our understanding of trophic links and to elucidate the diets of predators whose prey is as yet uncharacterized. The achievements and methods of PCR-based diet studies have been reviewed several times, but here we review an important development in the field: the use of PCR enrichment techniques to promote the amplification of prey DNA over that of the predator. We first discuss the success of using group-specific primers either in parallel single reactions or in multiplex reactions. We then concentrate on the more recent use of PCR enrichment techniques such as restriction enzyme digests, peptide nucleic acid clamping, DNA blocking and laser capture microdissection. We also survey the vast literature on enrichment techniques in clinical biology, to ascertain the pitfalls of enrichment techniques and what refinements have yielded some highly sensitive methods. We find that while there are several new approaches to enrichment, peptide nucleic acid clamping and DNA blocking are generally sufficient techniques for the characterization of diets of predators and highlight the most important considerations of the approach.

To understand the ecosystem dynamics that underpin the year-round presence of a large generalist consumer, the Bryde’s whale (Balaenoptera edeni brydei), we use a DNA metabarcoding approach and systematic zooplankton surveys to investigate seasonal and regional changes in zooplankton communities and if whale diet reflects such changes. Twenty-four zooplankton community samples were collected from three regions throughout the Hauraki Gulf, New Zealand, over two temperature regimes (warm and cool seasons), as well as 20 samples of opportunistically collected Bryde’s whale scat. Multi-locus DNA barcode libraries were constructed from 18S and COI gene fragments, representing a trade-off between identification and resolution of metazoan taxa. Zooplankton community OTU occurrence and relative read abundance showed regional and seasonal differences based on permutational analyses of variance in both DNA barcodes, with significant changes in biodiversity indices linked to season in COI only. In contrast, we did not find evidence that Bryde’s whale diet shows seasonal or regional trends, but instead indicated clear prey preferences for krill-like crustaceans, copepods, salps and ray-finned fishes independent of prey availability. The year-round presence of Bryde’s whales in the Hauraki Gulf is likely associated with the patterns of distribution and abundance of these key prey items.

Achatinella mustelina is a critically endangered tree snail that subsists entirely by grazing microbes from leaf surfaces of native trees. Little is known about the fundamental aspects of these microbe assemblages: not taxonomic composition, how this varies with host plant or location, nor whether snails selectively consume microbes. To address these questions, we collected 102 snail faecal samples as a proxy for diet, and 102 matched-leaf samples from four locations. We used Illumina amplicon sequencing to determine bacterial and fungal community composition. Microbial community structure was significantly distinct between snail faeces and leaf samples, but the same microbes occurred in both. We conclude that snails are not 'picky' eaters at the microbial level, but graze the surface of whatever plant they are on. In a second experiment, the gut was dissected from non-endangered native tree snails in the same family as Achatinella to confirm that faecal samples reflect gut contents. Over 60% of fungal reads were shared between faeces, gut and leaf samples. Overall, location, sample type (faeces or leaf) and host plant identity all significantly explained the community composition and variation among samples. Understanding the microbial ecology of microbes grazed by tree snails enables effective management when conservation requires captive breeding or field relocation.

Despite the global significance of spiny lobsters, the biology of their planktonic larvae, especially their carnivorous diet, is very poorly understood. Unsuitable diets and feeding regimes are one of the major reasons for consistently high mortalities during larval culture. In an effort to begin to determine the nutritional requirements of phyllosoma larvae of the spiny lobster, Jasus edwardsii, the present study first characterised the proximate composition of 11 previously identified groups of their potential prey. This included gelatinous zooplankton (chaetognaths, ctenophores, tunicates, siphonophores, scyphozoans, heteropods and pteropods), crustacean zooplankton (amphipods, copepods and euphausiids) and fish larvae. The composition of the 54 samples of 30 species of zooplankton ranged over 79.2–98.1% moisture content, and 1.9–54.2% for protein, 0.4–11.2% for carbohydrate, 0.1–27.9% for lipid, and 3.9–76.4% for ash of dry weight. The results showed that, in the most, zooplankton species provide a reliable source of protein. Only a small number of zooplankton had high lipid yield, suggesting that a narrow range of zooplankton species high in lipid are likely to be targeted by late-stage phyllosomas. The results of the present study will be useful as a guide for the development of artificial diets to improve phyllosoma aquaculture.

This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. AbstractThe recent dramatic decline in settlement in the population of the spiny lobster, Panulirus cygnus, may be due to changes in the oceanographic processes that operate offshore of Western Australia. It has been suggested that this decline could be related to poor nutritional condition of the post-larvae, especially lipid which is accumulated in large quantities during the preceding extensive pelagic larval stage. The current study focused on investigations into the lipid content and fatty acid (FA) profiles of lobster phyllosoma larvae from three mid to late stages of larval development (stages VI, VII, VIII) sampled from two cyclonic and two anticyclonic eddies of the Leeuwin Current off Western Australia. The results showed significant accumulation of lipid and energy storage FAs with larval development regardless of location of capture, however, larvae from cyclonic eddies had more lipid and FAs associated with energy storage than larvae from anticyclonic eddies. FA food chain markers from the larvae indicated significant differences in the food webs operating in the two types of eddy, with a higher level of FA markers for production from flagellates and a lower level from copepod grazing in cyclonic versus anticyclonic eddies. The results indicate that the microbial food web operating in cyclonic eddies provides better feeding conditions for lobster larvae despite anticyclonic eddies being generally more productive and containing greater abundances of zooplankton as potential prey for lobster larvae. Gelatinous zooplankton, such as siphonophores, may play an important role in cyclonic eddies by accumulating dispersed microbial nutrients and making them available as larger prey for phyllosoma. The markedly superior nutritional condition of lobster larvae feeding in the microbial food web found in cyclonic eddies, could greatly influence their subsequent settlement and recruitment to the coastal fishery.

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Abstract. Changes in the offshore oceanographic processes are suspected to be the cause of a recent dramatic decline in the settlement of post-larvae of the Western Australian spiny lobster (Panulirus cygnus), which has greatly reduced the productivity from the world's second-largest spiny lobster fishery. The present study assessed whether there are differences in the nutritional condition of the larvae of P. cygnus sampled from two pairs of cyclonic eddies (CEs) and anticyclonic eddies (AEs). Morphometric and biochemical analyses were undertaken on the mid-late-stage larvae (VI, VII, VIII) sampled offshore from two pairs of adjacent counter-rotating mesoscale eddies in the Leeuwin Current off Western Australia. The results showed that larvae captured from CEs had greater average total dry mass (P , 0.001) than those from AEs. Stage VIII larvae from CEs contained more protein (P , 0.008) (38.5% AE 5.4 s.e.) and lipid (P , 0.005) (67.2% AE 12.1) than did those from AEs. The possible causes for these differences are uncertain but may be related to differences in water temperatures in CEs v. AEs influencing the ability of phyllosomas to accumulate nutritional reserves.