Edited by S.A. Shepherd, P. E. McShane and F.E. Wells

Molluscan Research (Formerly Journal of The Malacological Socicty of Australia)

Dr F.E. Wells Western Australian Museum Perth, Western Australia

Editorial Board

Dr A.G. Beu

Institute of Geological and Nuclear Sciences

Lower Hutt, New Zealand

Mr R. Burn Museum of Victoria Melbourne, Victoria

Dr L.M. Joll W.A. Fisheries Department Perth, Western Australia

Dr W.F. Ponder Australian Museum Sydney, New South Wales

Dr G. Rosenberg Academy of Natural Sciences Philadelphia, Pennsylvania, USA

Dr B.J. Smith Queen Victoria Museum Launceston, Tasmania

Dr J. Stanisic Queensland Museum Brisbane, Queensland

Dr P. Bouchet

Museum National d’ Histoire Naturelle

Paris, France

Dr T.A. Darragh Museum of Victoria Melbourne, Victoria

Mr B.A. Marshall National Museum of New Zealand Wellington, New Zealand

Dr D. Reid The Natural History Museum London, United Kingdom

Dr W.B. Rudman Australian Museum Sydney, New South Wales

Prof E.S. Upatham Mahidol University Bangkok, Thailand

Dr R.C. Willan

Museum and Art Gallery of the Northern Territory Darwin, Northern Territory

Oe be ee Led

Molluscan Research

Australasian Abalone

Edited by S. A. Shepherd, P. E. McShane and F. E. Wells

Volume 18(2) 31 October 1997

Published by The Malacological Society of Australasia A.C.N. 067 894 848

Molluscan Research

Volume 18(2) 31 October 1997

Copyright Council of the Malacological Society of Australasia

Printed by THE WESTERN AUSTRALIAN MUSEUM

Moll. Res. ISSN No. 1323-5818 31 October 1997

Contents

S. Daume, S. Brand and W. J. Woelkerling Effects of post-larval abalone (Haliotis rubra) grazing on the epiphytic diatom assemblage

of(corallineited/algac’s tremm it rome Yen T. Std 1 Vn iyi Neat A eT, RU 119 R. D. Roberts and C. M. Nicholson Variable response from abalone larvae (Haliotis iris, H. virginea) to a range of settlement cues .......... 131

H. Takami, T. Kawamura and Y. Yamashita Contribution of diatoms as food sources for post-larval abalone Haliotis discus hannai

onfatcrustose;corallinetal ga trem strtreitetr ents eterno ere nent eee eee nt Tee yee vee 143 G. A. Moss

Early juvenile growth of the abalone Haliotis australis im culture .....c.ccccccsssssssesssssessesvssesscsvesessesessecseesens 153 P. E. McShane

Differences in the relative abundance of abalone (Haliotis iris) in relation to the perceived Statusjotitwojrepional ifisheriessiniNewsZealand monna nE EE area me 161 G. P. Hawkes, R. W. Day, M. W. Wallace and T. D. Harriden

Optimum conditions using managanese as a shell marker for abalone age validation studies .….............. 169

C. L. Kitting and D. E. Morse Feeding effects of postlarval red abalone, Haliotis rufescens (Mollusca: Gastropoda)

onfencrustingtcorallinetal ac sesretresc ATA AN EE E NO nes) ners penne 183 S. A. Shepherd and M. Avalos-Borja

The shell microstructure and chronology of the abalone Haliotis corrugata u... 197 F. E. Wells and J. K. Keesing

How many juvenile abalone are there? The example of Haliotis roei „sern 209 S. A. Shepherd and J. R. Turrubiates-Morales

A practical chronology for the abalone Haliotis fulgens manra E 219 J. Reyn. Naylor and P. E. McShane

Post-settlement survival of abalone (Haliotis iris, H. australis) in turbulent floWS o... 227 S. A. Shepherd and L. Triantafillos

Studies on southern Australian abalone (genus Haliotis) XVII. A chronology of H. laevigata no.n.. 233 S. A. Shepherd and S. Huchette

Studies on southern Australian abalone (genus Haliotis) XVIII. Ring formation in H. scalaris sees... 247 K. R. Rodda, J. K. Keesing and B. L. Foureur

Variability in larval settlement of abalone on artificial collectons a...n 253 P. A. Preece, S. A. Shepherd, S. M. Clarke and J. K. Keesing

Abalone stock enhancement by larval seeding: effect of larval density on settlement and survival ........ 265

N. L. Andrew, D. G. Worthington and P. A. Brett Size-structure and growth of individuals suggest high exploitation rates in the fishery for

blacklip abalone, Haliotis rubra, in New South Wales, Australia ...scccccccccsccsssesssscesscscssseseevesceseeseccesceseeeee 275 D. R. Schiel Review of abalone culture and research in New Zealand Srne Enn 289

S. H. Hooker, R. G. Creese and A. G. Jeffs

Growth and demography of paua Haliotis iris (Mollusca: Gastropoda) in northeastern New Zealand ... 299 C. S. Friedman, R. Grindley and J. A. Keogh

Isolation of a fungus from shell lesions of New Zealand abalone, Haliotis iris Martyn

ana Ea australisaGmelinpeetersrtsi A reteset reer RA A 313

Instructions|to/atithors a ET AA E A RN alee E a R 325

THE AUSTRALASIAN SYMPOSIUM ON ABALONE FISHERIES, CULTURE AND BIOLOGY WELLINGTON, NEW ZEALAND, October 1996.

INTRODUCTION

Scoresby A. Shepherd! and Paul E. McShane?

South Australian Research and Development Institute, PO Box 120 Henley Beach, SA 5022, Australia

' shepherd.scoresby@pi.sa.gov.au * mcshane.paul@pi.sa.gov.au

This special issue of Molluscan Research contains 19 papers given at the first regional abalone symposium in Wellington, New Zealand in October 1996. In the Introduction to ‘Progress in Abalone Research’ containing the papers from the 1994 2nd International Abalone Symposium in Hobart, Day and Shepherd (1995) suggested that the most fertile topics for future research in wild fisheries for abalone were likely to be:

* the use of cultured larvae to study the early life history

e larval dispersal and larval availability to reefs

* the relation between egg production and later recruitment

e the aging of abalone shells, and

e the effect of fishing on abalone aggregations.

It is pleasing to see that many of these topics have been vigorously pursued in the short time that has elapsed since the Hobart meeting. On the first topic we have seven papers on the ecology of larvae on settlement or of post-larvae after settlement. Roberts and Nicholson show that larval settlement is not a single process, but has two distinct elements — attachment and metamorphosis — each of which may have its own suite of biochemical cues. Daume et al. demonstrate the dependence of post-larvae on crustose coralline algae (CCA) for food in the first week or two before they begin feeding on diatoms. Kitting and Morse further explore the relation between post-larvae and CCA from the point of view of the algal host ‘(a decrease in epiphytes and a stimulus for growth) and speculate on the evolution of this extraordinary symbiotic mutualism. In some elegant experiments Takami et al. show that CCA rapidly become inadequate for a growing post-larva, and must be supplemented by diatoms to promote rapid growth. Moss pursues the theme from a maricultural point of view and determines the optimal size at which the diet of a tiny abalone should switch from diatoms to macro-algae or artificial food. The last two of this septet of papers used cultured larvae to study the survival of settlers in the wild: Preece et al.

in the context of stock enhancement (a subject of increasing interest wherever stocks are declining) and Naylor and McShane in an assessment of sources of variation in post-settlement survival. The latter paper described a simple device for simulating the water flows encountered in subtidal habitats.

On the second topic Rodda et al. deployed larval collectors for 3 years to measure the recruitment of larvae above natural reefs. This study has provided insight into larval dispersal and the timing of episodic settlement on reefs. Progress will be even more when the post-larvae of the five sympatric Species of abalone can be identified to species, and the relation between larval availability and subsequent recruitment to reefs is clarified.

The third topic has been addressed obliquely in two studies that examined patterns of recruitment two species in superficially similar habitats — studies which incidentally raise intriguing questions Out the different life history strategies that have evolved in different conditions, Wells and Keesing und that weak recruitment can be offset by a very fast juvenile growth which reduces the window

in ab fo

of vulnerability to predators. In contrast, Andrew et al. found apparently much stronger recruitment, which maintained a stable fishery despite a high exploitation rate. Both studies are reminders of the perils of extrapolating conclusions from one abalone fishery to another.

The fourth topic on shell-aging has been advanced, in terms of methodology and practice, in five papers. Hawkes et al. establish the use of manganese as a time-stamp for marking shells for validation of shell-aging, and Shepherd et al. in a quartet of papers describe the rate of ring deposition in two Mexican species and two Australian ones. Each species described has a different pattern of ring deposition, and sometimes there is variation within a species, and sometimes variation due to confounding factors such as endobionts and shell abrasion. This topic will certainly attract more attention as biologists seek to apply age-structured models to abalone fisheries.

The importance of the fifth topic on effects of fishing is now increasingly recognised in the light of recent work on the fertilisation success in free-spawning invertebrates. In an unusual approach McShane shows how the perceptions of divers regarding aggregation sizes and recruitment shape

- their selective fishing behaviour — which may ultimately have consequences for recruitment to the fishery. Clearly we need a better understanding of the interaction between diver behaviour (the predator) and the behavioural biology of the prey abalone.

Of course some papers will always fall outside the best-laid prescriptions of mice and men but are still of great interest. Hooker et al. consider the growth of Haliotis iris near its northern limit, and Friedman et al. describe a fungal disease which has long been known to infest New Zealand abalone.

Finally, and fittingly, Schiel has given a useful review of research on New Zealand abalone with a critique, aptly acerbic, of research in the fledgling abalone mariculture industry.

The topical framework employed in this introduction, while not intended to be prescriptive, will nevertheless, we believe, be valid for research in wild fisheries for some years to come. Yet if we were to venture to predict a topic that will soon become significant it would be this. Given the increasing emphasis on ecosystem, as opposed to single species, management of coastal waters (Larkin 1996) where exploitation of species at different levels of the food web, from algae to primary, to secondary consumers, is ever increasing, attention must be given to the relations between abalone and all other species which share its niche. This is an ambitious order because the niche of an abalone changes in scale from the tiny (the CCA habitat) to the reef at a scale of metres, to clusters of reefs (the metapopulation dimension) at a scale of kilometres, and embraces macro-algae in their structural role as well as for food, and predators, competitors and parasites. We look forward to further Australasian studies which emphasize the ecological interactions between abalone and other reef-dwelling organisms.

Literature cited Day, R.W. and Shepherd, S.A. 1995. Fisheries biology and ecology of abalone: introduction. In Shepherd, S.A., Day, R.W. and Butler, A.J. Progress in abalone research. Marine and freshwater research 46(3): iii—v.

Larkin, P.A. 1996. Concepts and issues in marine ecosystem management. Reviews in Fish Biology and Fisheries 6: 139- 164.

Acknowledgements

All papers were reviewed in accordance with the policy of Molluscan research and we thank the 30 or so referees for their contributions to the work. Our task would have been almost impossible and this volume impoverished without their advice.

The symposium was generously supported by the National Institute of Water and Atmospheric Research Ltd, Dover Fisheries Ltd, Australian Abalone Producers Association, Ngai Tahu Abalone Ltd, Ocean Ranch (Wellington) Ltd, Promak Technology (NZ) Ltd and Sealord Shellfish Ltd. The organising committee, Paul McShane, Peter Gerring, Steve Mercer, Pete Notman, with much help from Maria Fraser, John Capa, Owen Anderson and Alan Blacklock ensured that the symposium was a success.

Moll. Res. 18: 119-130 (1997).

Effects of post-larval abalone (Haliotis rubra) grazing on the epiphytic diatom assemblage of coralline red algae

Sabine Daume', Sascha Brand, and Wm. J. Woelkerling

School of Botany, La Trobe University Bundoora, Victoria 3083

' Corresponding author. E-mail: botsd@lure.latrobe.edu.au

Abstract

Larvae of the abalone Haliotis rubra were settled on pieces of the non-geniculate coralline red alga Phymatolithon repandum. We measured a mean growth-rate of ca. 30 pm/day, 11 days after larval settlement which stayed constant throughout the experiment. The diatom assemblage on the surface of the alga was studied for 53 days after larval settlement using scanning electron microscopy. Two species of the genus Cocconeis were found to dominate the diatom assemblage on the surface of P. repandum. Up to 18 days after larval settlement, the diatom population increased exponentially. Eighteen days after settlement, post-larval abalone started to graze on the diatom Cocconeis scutellum leaving the bottom valves of the diatoms on the surface of P. repandum. As a result, the diatom population decreased markedly. The outermost cells and polysaccharide layer of P. repandum were often missing but we did not find any grazing marks on the thallus surface up to 53 days post-settlement. We discuss the hypothesis that epithallial cell sloughing of non-geniculate coralline red algae is an intrinsic mechanism to reduce microscopic epiphytes and may interact closely with grazing by post-larval abalone. We concluded that the post-larvae must find an additional food source derived from the NCA before grazing on the diatoms, because of the high growth-rate during the first and second week of rearing and the constancy throughout the experiment.

Introduction

The relationship between abalone and non-geniculate coralline red algae (NCA) (Corallinales, Rhodophyta) is well known. Abalone larvae settle on NCA (Morse & Morse 1984; 1988; Morse 1990; 1991) and remain in the coralline habitat until they are about 6 mm in shell length (Shepherd & Daume 1996). Post-larval abalone are known to feed on benthic diatoms (Kawamura et al. 1995). Diatoms Occurring on artificial plates have also been used as the initial food source in commercial abalone hatcheries. Recent interest has focused on settlement and feeding experiments of post-larval abalone on species of benthic diatoms (Kawamura & Kikuchi 1992; Kawamura & Takami 1995; Kawamura et al. 1995; Matthews & Cook 1995; Kawamura 1996). Despite the importance of diatoms in abalone feeding ecology, the naturally occurring diatom assemblage on the NCA surface has never been studied in detail.

Molluscan herbivores have a major effect on the algal community by feeding on the sporelings of Macroalgae (Dayton 1975; Menge 1976; Underwood & Jernakoff 1981). If herbivores are removed, the macroflora increases. This effect has been well documented in the past (Southward 1964; Dayton 1971) and later tested at the microflora level (Nicotri 1977; Underwood 1984) however, the influence of post-larval abalone grazing on the diatom assemblage of NCA surfaces has not been investigated.

Phymatolithon repandum (Foslie) Wilks and Woelkerling 1994 and other species of NCA frequently slough epithallial cells, and this has been interpreted as an antifouling mechanism which

120 S. Daume, S. Brand, W.J. Woelkerling

inhibits algal spore settlement (Masaki et al. 1984; Johnson & Mann 1986; Keats et al. 1993; 1997). The effects of epithallial cell sloughing on the diatom assemblage of NCA surfaces is described in this study.

Herbivores like Cellana spp., Acmaea spp., Notoacmaea spp., Patella spp. as well as Haliotis spp. are known to leave grazing marks produced by the specialized teeth of their radula while feeding on NCA (Clarkson & Shepherd 1985; Padilla 1985; Garland et al. 1985). Garland et al. (1985) described grazing marks of 6 week old post-larvae which were 1—2 cells deep. It is unknown however, if post-larval abalone younger than 6 weeks leave grazing marks on the thallus surface.

In this paper we describe:

(1) the distribution of the dominant diatom species on the surface of the NCA Phymatolithon repandum; (2) the effect of post-larval abalone grazing on the dominant diatom species from day of larval ; settlement to 53 days after settlement; (3) cell sloughing on different growth-forms of P. repandum and the effect it has on the diatom assemblage; and (4) the radula structure and teeth arrangement of post-larval abalone.

Materials and Methods

Sampling

Small rocks encrusted with NCA were collected from a boulder habitat in front of the Gloucester Reserve, Williamstown, Victoria, Australia (37° 53'S, 144° 55'E) at 3— 4 m depth. Williamstown is located at the north end of Port Phillip Bay and is close to Port Melbourne and the mouth of the Yarra River. Only the blacklip abalone, Haliotis rubra Leach 1814 is present here. Phymatolithon repandum is the dominant NCA species at this locality (unpublished data) and occurs with two different growth-forms (encrusting and warty, see Woelkerling et al. 1993). Pieces (ca. 1 cm?) of each growth-form were removed from the rocks with a razor blade or chisel. Each replicate for the diatom study and the experiment was obtained from a different plant to ensure independence.

Distribution of the dominant diatoms on P. repandum

Pieces of the encrusting and warty growth forms were freshly collected and air dried after sampling to study the composition and distribution of diatoms on the thallus surface. The samples were mounted on aluminium stubs with ‘Fotobond’ acrylic adhesive (Agfa-Gevaert Ltd). The stubs were sputter-coated with gold before viewing in a Siemens ETEC Autoscan microscope at 20 kV to determine the frequency of diatom occurrence on the two growth-forms of P. repandum. For each sample, the presence or absence of diatom species was recorded at a magnification of 1200x in 20 fields of view.

Effect of post-larval grazing

Larvae of the abalone Haliotis rubra obtained from a hatchery (Cheetham Salt, Lara, Victoria, Australia)-were allowed to settle onto surfaces of Phymatolithon repandum. Two pieces of P. repandum (ca. | cm’) representing the encrusting and warty growth forms were placed in each of 24 sterile glass jars with 300 ml UV sterilised sea water. Jars were kept at 15° C, 3000 lux with a 12 h L:D photo cycle. One hundred larvae competent to settle were added to each of 18 jars. Six jars, without larvae and only the warty growth-form, were used as a control. The shell length of four randomly chosen larvae was measured before being added to the jars. Three to four larvae were measured 11, 18 and 53 days after settlement to calculate their growth-rate.

Less than 20 larvae/cm? were maintained on each piece of NCA. Abalone were observed moving over the NCA surface through a dissecting microscope, showing side to side scraping by the radula and judged to be actively feeding.

121 Post-larval abalone grazing on diatoms of coralline surfaces

s

i i is: Cocconeis Figure 1. Diatoms (Cocconeis spp.) on the surface of Phymatolithon repandum. A: Two species of Cocconeis

i istributi : Di roliferate around scutellum (arrow) and Cocconeis sp. B: Diatoms occur in a patchy distribution. C: Diatoms p conceptacles.

122 S. Daume, S. Brand, W.J. Woelkerling

Diatom distribution under grazing pressure and cell sloughing of NCA

Pieces of both NCA growth-forms with post-larvae and pieces from the control jars (without post- larvae) were subsampled randomly 4, 11, 18, 30 and 53 days after larval settlement. The subsamples were air dried and treated for the scanning electron microscopy (SEM) study as described above. For each sample, the presence or absence of diatom species and cell sloughing was recorded at a magnification of 1200x and 150x respectively in 20 fields of view. The mean frequency (+ standard error) of diatom species and cell sloughing was calculated.

Abalone radula

To study the post-larval abalone radula, the shell was removed, the tissue of the whole animal dissolved in 6% sodium hypochlorite overnight, washed in distilled water, allowed to dry and then prepared for SEM as described above.

Data analysis

Statistical analyses were carried out using the STATISTICA (Statsoft Inc. 1995) computer package. One-way and two-way ANOVAs were carried out on untransformed data (number of fields with Cocconeis spp.) followed by Tukey‘s multiple comparison test.

Results

Distribution of the dominant diatoms on P. repandum

During the period of the study (October to December 1995) only pennate diatoms were observed on the surface of P. repandum. The genus Cocconeis was represented by two species (Cocconeis scutellum (Ehrenberg) Boyer 1927; Cocconeis sp.) (Fig. 1A) that dominated the diatom assemblage. On all samples examined, they formed 90-100% of the diatom population with a size range from 10 to 40 um. We observed a low frequency (5-10%) of diatoms on freshly sampled pieces of both growth-forms of P. repandum. All diatoms were found in a patchy distribution (Fig. 1B) and proliferated in cavities and around conceptacles (Fig. 1C).

Effect of post-larval grazing

The frequency of Cocconeis spp. on the surface of P. repandum grazed by post-larval abalone changed during the period of study (Fig. 2). Between four and 11 days after abalone larvae settled on the NCA surface, the frequency of Cocconeis spp. increased exponentially on both growth- forms of NCA but not on the controls where larvae were absent. Cocconeis spp. reached a maximum frequency of 80% on the encrusting growth-form 11 days after larval settlement (Fig. 3A). There was a highly significant difference in frequency of Cocconeis spp. between the control and the 2 treatments with post-larval abalone 11 and 18 days after settlement (2-way ANOVA, Treatment x Age Interaction, p<0.001). However, there was no significant difference in diatom frequency between the warty and the encrusting growth-form throughout the experiment (2-way ANOVA, p=0.24).

After 18 days, the Cocconeis population decreased on both treatments with post-larval abalone (Fig. 2). On the grazed surfaces, raphe-bearing valves (bottom valves) of C. scutellum, ca. 20 um in length, appeared approximately 18 days after larval settlement (Fig. 3B), suggesting that post-larval abalone were removing the top valves of the diatom and thus gaining access to the diatom cell contents.

The growth-rate of the post-larval abalone was relatively constant throughout the experiment with a mean growth-rate of 31.4 um/ day (S.E. 5.2 um) 11 days after settlement, 33.3 um/ day (S.E. 1.4 um) 18 days after settlement and 33.9 um/ day (S.E. 0.8 pm) 53 days after settlement.

Post-larval abalone grazing on diatoms of coralline surfaces 123

~-@- encrusting —i warty —4— control

g Q 17) 2 a g Q Q 8 Q Y- o > (S) S ® 3 o ® dæ — aS

30

days after settlement

Figure2. Changes in frequency of occurrence of Cocconeis spp. on the surface of Phymatolithon repandum when grazed by post-larval abalone (Haliotis rubra). Vertical bars indicate the standard error.

Cell sloughing of NCA

Cell sloughing was observed on the control pieces of P. repandum and occurred significantly more on the warty than on the encrusting growth-form of the treatment (1-way ANOVA, p<0.047). The mean frequency of cell sloughing was 16% (S.E. 4.3%) for the encrusting and 45% (S.E. 12.1%) for the warty growth-form. Cell sloughing was intense both around and between warty protuberances (Fig. 4A). Figure 4B shows an area of intensive sloughing in detail. The outermost cells become

detached in sheets of numerous cells and subsequently removes all epiphytes occurring on the surface,

Abalone radula

The post-larval abalone radula has numerous teeth per row (Fig. 5A). Each row consists of 1 median tooth with a smooth edge, 5 (2+3) strong lateral teeth on each side with sharp endings for rasping, and numerous marginal teeth with serrated edges. The marginal teeth are similar to the outermost lateral teeth, but become progressively thinner.

Each tooth has a width-to-length ratio of approximately 1:2. The teeth of Haliotis rubra have a clearance angle (angle from the coralline surface to the back of the tooth) of about 8°.

124 S. Daume, S. Brand, W.J. Woelkerling

ty

nti Whe oe

Figure 3. Abalone post-larvae grazing on the surface of Phymatolithon repandum . A: Post-larvae 11 days after settlement, Cocconeis sp. proliferating on the surface. B: Grazed surface of NCA with bottom, raphe-bearing valve of the diatom Cocconeis scutellum still attached to the surface.

Post-larval abalone grazing on diatoms of coralline surfaces

So eee

Figure 4. Epithallial cell sloughing of Phymatolithon repandum. A. Warty growth-form with intensive cell sloughing (arrows). B. Outermost layer of cells sloughing off and effectively remove diatoms growing on the surface.

126 S. Daume, S. Brand, W.J. Woelkerling

Grazing marks matching the pattern of post-larval abalone radulae were not found on the surface of P. repandum during the 53 days after settlement. The polysaccharide layer together with the outermost cells of the thallus were often missing on both the grazed and ungrazed (control) pieces.

Discussion

Distribution of the dominant diatoms on P. repandum

The low frequency of diatoms observed on freshly sampled NCA is comparable to non-living substrata such as rocks but not other macroalgae (Round et al. 1990). Non-living substrata are less favourable to diatom growth than macroalgae and seagrasses and thus support a low frequency of diatoms (Hudson & Bourget 1981).

Eckman and Norwell (1984) noticed that on the bottom of bumps, areas with a specific horseshoe vortex effect of flow, the microbial activity and mucus binding capacity is enhanced. Consequently, we expected a higher frequency of diatom occurrence on the warty growth-form. However, we did not notice any difference between the two growth-forms, neither on freshly sampled pieces or on pieces sampled throughout the experiment. The observed higher frequency of cell sloughing on the warty growth-form (warty treatment and control) could have offset the expected growth-form effect and thus the frequency of diatoms was similar on both growth-forms of Phymatolithon repandum.

In this study, species of Cocconeis were observed to be the dominant diatoms on the surface of P. repandum. In agreement with our findings, Kawamura et al. (1992) observed that in areas dominated by crustose coralline algae, diatoms with strong adhesive solitary forms such as Cocconeis spp. dominate the benthic diatom population. In addition, Hudon and Bourget (1981) found that Cocconeis spp. and Amphora spp. were the first and major colonizers on artificial plates from May to mid August (northern hemisphere). They noticed a change in diatom dominance at the beginning of September which suggests that the dominance of Cocconeis spp. may be seasonal.

Effect of post-larval grazing

Four to 11 days after larval settlement the populations of Cocconeis spp. increased exponentially on both growth-forms of NCA. We did not find any bottom valves of diatoms on the NCA surface at this young age, suggesting that post-larval abalone do not graze on Cocconeis spp. at this stage. Species of Cocconeis are known to have high adhesive strength (Kawamura et al. 1995) and young post-larvae might not be able to detach the valves from the surface of P. repandum. In addition, competing diatoms and bacteria may have been preferentially consumed at this early stage. Young post-larvae are known to feed on loosely attached diatoms. The extracellular mucus associated with these diatoms is considered to be a good food source (Kawamura and Takami 1995; Kawamura 1996). The grazing activity and metabolic processes of post-larvae up to 11 days after larval settlement, seems to allow the Cocconeis population to proliferate which ensures enough food supply for the abalone in coming weeks. In the control, without abalone grazing activity, the Cocconeis population never exceeded a 5% frequency. In agreement, Suzuki et al. (1987) and Matthews and Cook (1995) noted that under the grazing pressure of juvenile abalone, prostrate diatoms like Cocconeis spp., start to proliferate. After several days artificial plates are covered entirely with these diatoms (Suzuki et al. 1987). However, none of them observed post-larvae or juveniles grazing on Cocconeis spp.

We measured a mean growth-rate of ca. 30 um/ day 11 days after settlement which stayed relatively constant throughout the experiment. Garland et al. (1985) settled larvae of Haliotis rubra onto NCA in 1000- culture bins. The larvae grew to a mean size of 692+ 33 um in 42 days (10.5 m/day) which is considerably lower than the growth-rate we calculated. However, they did not have any measurements for the first two weeks of rearing. Kawamura and Takami (1995) measured the mean growth-rate of the post-larval abalone Haliotis discus hannai, settled in tissue culture wells with only Cocconeis scutellum as a food source. They recorded a growth-rate of 24.7+1.6 um/day

Post-larval abalone grazing on diatoms of coralline surfaces 127

Figure 5. Part of the radula of a post-larval abalone (Haliotis rubra) with 1 median (M) and 5 lateral teeth (L1-L5).

from 0 to 10 days after settlement. The growth rate decreased 10 days after settlement (7.542. 1 pm/ day) and was significantly lower than those of abalone fed on other species of diatoms. Takami et al. (this issue) settled abalone larvae of the same species onto small rocks encrusted with the NCA Lithophyllum yessoense. When treated with GeO, to reduce the diatom growth on surfaces of L yessoense, the growth-rate of the abalone did not differ from untreated L. yessoense surfaces during the first seven days of rearing. However, the growth-rate of the post-larvae H. discus hannai in this experiment was both constant and higher on the surface of L. yessoense (treated 37.0 + 0.7 um/day and untreated 40.9 + 0.7 um/day) compared to Kawamura and Takami‘s (1995) experiment, in which the larvae were grown on diatoms only. In agreement with our results, this suggests that post- larval abalone must find an additional food source derived from the NCA rather than the diatoms occurring on its surface. This food source is important during the first week of rearing.

Giraud and Cabioch (1976) describe a layer of polysaccharide material covering the external part of the thallus of a variety of different species of NCA. The cells just below that layer have uncalcified cell walls at their external pole and vesicles full of polysaccharide fibrils which can open through the plasmalemma and diffuse into the outermost layer. Garland et al. (1985) suggested that post-larval abalone (6 weeks old) graze on the surface mucus, polysaccharide layer and the epithallial cell content of the NCA. ;

Diatoms with valves which can be ruptured by the abalone radula, such as the valves of Cocconeis Spp., are considered to be a high value food source for post-larval abalone, approximately 1mm in

128 S. Daume, S. Brand, W.J. Woelkerling

shell length (Kawamura 1996). Our findings suggest that post-larvae of Haliotis rubra start feeding on diatoms about 18 days after settlement, when they are approximately 800 pm in shell length. In addition, Takami et al. (1997b) stated that post-larvae of H. discus hannai are able to access diatom cell contents once they reach about 800 pm. Takami et al. (this issue) found a higher growth-rate of Haliotis discus hannai fed on NCA with diatoms, than on NCA without diatoms, 2 weeks after settlement. In agreement with our findings this also suggests that the NCA alone provides enough food for the post-larvae until 2 weeks after settlement and /or post-larvae can not utilize the diatom food source until approximately 2 weeks after larval settlement.

Cell sloughing of NCA

Cell sloughing was observed on the control pieces and more frequently on the warty than on the encrusting growth-form of P. repandum. This difference could explain the lower frequency of diatoms on the warty growth-form in Fig. 2 and the overall low diatom frequency of the ungrazed control. It also suggests that cell sloughing intrinsically controls overgrowth by diatoms and other epiphytes when grazers are absent. Cell sloughing may also facilitate the occurrence of early successional diatom species such as C. scutellum (Hudson and Bourget 1981) by providing new, uncolonised substrata.

The polysaccharide layer together with epithallial cells of the coralline thallus were partially missing on all pieces observed. Garland et al. (1985) report that post-larvae (6 and 13 weeks old) of Haliotis rubra remove and ingest the polysaccharide layer and epithallial cells of NCA. In this study grazers in the field (before the NCA had been sampled) or the post-larval abalone (up to 53 days after settlement) might have removed the outermost cells and polysaccharide layer without leaving any obvious, deeply penetrating grazing marks.

Abalone radula

Abalone have a rhiphidoglossan radula like many primitive herbivorous prosobranchs (Crofts 1929). The teeth of post-larval abalone have a low clearance angle. A clearance angle close to 0° results in a tooth sliding across the surface rather than cutting into the surface of the algae (Padilla 1985). This might explain why we did not observe grazing marks of post-larval abalone on the surfaces of P. repandum. Given the low clearance angle of the teeth, it seems that abalone post- larvae would be more efficient grazers on flat surfaces eg. encrusting compared to a warty growth- form of NCA.

We concluded that abalone post-larvae must find an additional food source derived from the NCA before grazing on the diatoms, because of the high growth-rate during the first and second week of rearing and the constancy throughout the experiment.

Both mechanisms, the cell sloughing of P. repandum and grazing by post-larvae, seem to enhance the quality of the diatom assemblage on-NCA as a food source for post-larval abalone. Cell sloughing controls overgrowth by diatoms and other epiphytes (Keats et al. 1997) which is known to be advantageous during the early life history of abalone (Shepherd & Daume 1996). Grazing by post- larval abalone seems to encourage proliferation of Cocconeis spp. which provides a good food source for the coming weeks.

Further experiments are required to gain a better understanding of the interactions of both mechanisms on epiphytic diatom assemblages and their effect on abalone settlement and early growth. -

Acknowledgments

We are grateful to Tony Smith and Peter Rankin from Ridley Corporation Ltd., Division of Pet Products and Emerging Businesses, Victoria for the supply of abalone larvae. We thank Dr .T. Kawamura for confirming the diatom identifications, Trevor Phillips for assistance with photography, Dr. Scoresby Shepherd and Adela Harvey for helping to improve the manuscript and

Post-larval abalone grazing on diatoms of coralline surfaces 129

Stephen Madigan and an anonymous reviewer for their useful comments. This study was supported by an ARC collaborative research grant in association with the South Australian Research and Development Institute and a La Trobe University post-graduate scholarship to the first author.

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Moll. Res. 18: 131-141 (1997).

Variable response from abalone larvae (Haliotis iris, H. virginea) to a range of settlement cues

Rodney D. Roberts! ? and Christine M. Nicholson!

'Cawthron Institute, Private Bag 2, Nelson, New Zealand University of Otago, P.O.Box 56, Dunedin, New Zealand

Abstract

We exposed competent larvae of two abalone species (Haliotis iris and H. virginea) to a range of potential settlement cues, and observed a hierarchy of responses. The end point and the timing of the settlement process varied, depending on the cue provided. Response was qualitatively consistent between experiments, but showed some quantitative variation. Rocks coated with crustose coralline algae (CCA) induced 80-100% of larvae to attach and metamorphose, and 50-100% to grow peristomal shell, within 2 days. A distilled water extract of CCA induced almost 100% attachment within 2 days, but metamorphosis occurred gradually between 2 and 5 days. Shell growth was minimal, apparently inhibited by chemical interference from the extract. y-Aminobutyric acid (GABA, 1 uM) induced 90-100% attachment and 20-60% metamorphosis in H. iris within 2 days. For H. virginea, 1 M GABA induced attachment (65-100%) but typically only 0-5% metamorphosis within 2 days. KCl (10 mM) added to seawater induced attachment (50- 70%) but less than 10% metamorphosis. Different diatoms induced responses ranging from rapid or gradual induction of metamorphosis, to minimal response. Some combinations of cues produced synergistic effects. E.g., GABA (1 1M) plus diatom biofilm induced more metamorphosis (49%) than biofilm (28%) or GABA (1%) alone. Addition of CCA crude extract to the same biofilm hastened metamorphosis. Observations of attachment without subsequent metamorphosis suggest that there may be separate cues

for attachment and metamorphosis. This study highlights the complexity of the settlement response in abalone.

Introduction

Most marine invertebrates have a free swimming larval stage prior to a benthic adult phase. For many species, including abalone, the transition from swimming larva to crawling post-larva (herein referred to as “settlement”) requires a chemical cue (Crisp 1974; Pawlik 1992). Larval abalone settle and metamorphose in response to various substances including intact crustose coralline algae (CCA), extracts from CCA (Morse et al. 1980), -aminobutyric acid (GABA — Morse 1984; Yang and Wu 1994), excess potassium ion (Yool et al. 1986; Yang and Wu 1995) and cultures of benthic diatoms (Kawamura and Kikuchi 1992).

Some variations in the settlement response of abalone have been documented, e.g. Searcy-Bernal et al. (1992) found that a diatom-based biofilm induced metamorphosis more slowly than 1 uM GABA or mucus treatments; and Morse (1984) reported that concentrations of GABA exceeding 1 uM induced attachment, but inhibited metamorphosis. However, there has been little detailed comparison of settlement responses across a range of different cues. Such a comparison needs to account for variation between batches of larvae (Dineen and Hines 1994) and differences in methods between

132 R.D. Roberts, C.M. Nicholson

experiments (Morse 1992). One aim of this study was to determine the response of a single batch of abalone larvae to several cues which have previously been shown to induce settlement in other abalone (see above). Another was to quantify