Microhabitat Distributions of Juvenile Gastropods•

A thesis submitted to the faculty of San Francisco State University In partial fulfillment of The requirements for The degree

Master of Science In Marine Science

by

Patricia Ann McMillan

San Francisco, California

November 2003 © Copyright by Patricia Ann McMillan 2003 Microhabitat Distributions of Juvenile Hydrothermal Vent Gastropods

Patricia Ann McMillan San Francisco State University 2003

Early gastropod settlement patterns around a hydrothermal vent at 9°50' N East

Pacific Rise were determined and compared to the distribution of adults of the same species. Post-settlement processes were also examined by deploying caged and uncaged settling blocks. These blocks were deployed in 4 different zones at the vent site for a 5-month period. The zones were characterized by different intensities of hydrothermal influence. Juvenile (<1mm) gastropods were identified and counted from the blocks. The most abundant species were

Lepetodri/us spp., Eulepetopsis vitrea, C/ypeosectus delectus and Gorgofeptis sp. (C/ypeosectus and Gorgoleptis were combined due to morphological similarity). The number of juvenile Lepetodrifus spp. was higher in the area of high hydrothermal influence compared to no influence. There was a significant difference between adult and juvenile distributions for Lepetodrilus spp. and

Cfypeopsectus!Gorgoleptis across all zones. Juvenile Eufepetopsis vitrea were

more abundant on the caged than the uncaged blocks. These data suggest •· that initial settlement patterns of vent gastropods are modified by post-

settlement processes for Lepetodrilus spp. and CfypeopsectusfGorgolepUs. ACKNOWLEDGMENTS

I would like to thank: • Dr. Stacy Kim for her enthusiasm, assistance and encouragement. • Dr. Nicholas Welschmeyer, Dr. Kenneth Coale, Dr. J. Timothy Pennington, and Dr. Kenneth Johnson for their encouragement, comments and suggestions on this manuscript. • Dr. Lauren Mullineaux and Susan Mills for providing me the samples and their assistance. • MBARI for their technical support.

I am also very grateful to: • The faulty, staff and student of Moss Landing Marine Labs for their support. • Kimberly Puglise for all her help and support during our time at MLML • Dr. Wiebke Ziebis for her friendship, support and reviewing of this manuscript • Christine Whitcraft for her support and many reviews of this manuscript. • Donna Drazenovich, Maureen Shea, Val Growney, Sandy Krebsbach, and Bonnie Becker for their friendship and encouragement. • Dr. Lisa Levin and her lab for their support. • Dr. Paul Hyde, Dr. Sabina Wallach and Dr. Jean Mefferd for the gift of life! • My friends for their continued support and encouragement • My family for being themselves and always asking, "So how's the thesis going?" " My husband Tom for his love, support and patience during this endeavor.

My funding sources: • Dr. Earl H. Myers and Ethel M. Myers Oceanographic and Marine Biology Trust • David and Lucille Packard Grant for Graduate Research and Travel

v TABLE OF CONTENTS

List of Tables ...... vii

List of Figures...... viii

Introduction ...... 1

The Physical Setting: Vent Geology ...... 2 Vent Biology...... 6 Dispersal Mechanisms (to disperse or not) ...... 8 Settlement Cues and Recruitment ...... 10 General Gastropod Biology and Adult Distribution ...... 12

Method ...... 15

Statistical Analyses ...... 1 9

Results ...... 20

Juveniles ...... 21 Juveniles vs. Adults ...... 22

Discussion ...... 23

Dispersal potential...... 24 Juvenile distribution ...... 25 Role of predation ...... 28 Juvenile vs. Adult distribution ...... 29 Conclusion ...... 30

Literature Cited ...... 33

vi LIST OF TABLES

Table Page

1. Generalize description of hydrothermal vent zones at EPR . .42 2. Identified juvenile hydrothermal vent gastropod species .... 43 3. Statistical results of ANOVA ...... 44 4. Statistical results of a priori ANOVA ...... 45

vii LIST OF FIGURES

Figure Page

1. Generalized diagram of zonation pattern at an EPR black smoker vent site ...... 46 2. Study site ...... 4 7 3. Deployment of settling substrate ...... 48 4. Major species identified ...... 49 5. Other species id en tiffed ...... 50 6. Distribution of adult and juvenile Lepetodrilus spp ...... 51 7. Distribution of adult and juvenile Clypeosectus delectus and Gorgoleptis sp...... 52 8. Distribution of adult and juvenile Eu/epetopsis vitrea ...... 53

viii Introduction

Larval settlement and post-settlement processes have been studied in the rocky intertidal (e.g. Connell 1961, 1972). As space becomes available, opportunistic species colonize thereby creating an environment for subsequent species. The post-settlement processes such as space competition, migration, and predation that structure the rocky intertidal communities may also structure hydrothermal vent communities. This scenario may be analogous to that at hydrothermal vents. The ability in the rocky intertidal to do manipulative experiments and thus observe daily changes is much easier. However at hydrothermal vents, the ability to remove fauna from areas within the vent habitat and daily observations are not feasible. Therefore, the knowledge obtained from the rocky intertidal studies may provide insight into larval settlement and post-settlement processes at hydrothermal vents. This study exams settlement through the use of settling blocks to determine settlement patterns of hydrothermal vent gastropods compared to adult distributions of the same species and physical (geological) setting.

1 The Physical Setting: Vent Geology

Hydrothermal vents occur at diverging plate boundaries, where plates of the earth's crust are spreading apart. As the plates move, hot magma rises to fill in the gaps. The rising magma is not evenly distributed but is focused in disjunct fissures from which the lava may be extruded. The young crust is very porous, and a "hydrothermal circulation" develops where cold seawater penetrates cracks and fissures in the rock. This water is heated by the hot rocks and magma before reemerging at the surface. This hot hydrothermal fluid is enriched in reduced chemical compounds (sulfur species) and with some metals (e.g. iron, copper, zinc). When it exits the sea floor and mixes with the surrounding cold and alkaline seawater, "black smoker" plumes can form by the precipitation of metal sulfides. Columnar chimneys typical of hydrothermal vents on the East Pacific Rise can grow around the fluid outlet starting by the precipitation of anhydrite

(calcium sulfate) and its manifestation in sheaths (Tivey 1995).

The bulk of evidence indicates that the spreading rate at the mid-ocean ridges is proportional to the heat flux and influences the distance between vents and the length of time a vent is active. In general, the faster the spreading rate, the more closely spaced and

2 ephemeral the vents are. At slow spreading ridges (Mid-Atlantic

Ridge, MAR}, vents are 10's to 1000's of km apart (Grassle 1986;

Seyfried and Mottl 1995) and may remain active for hundreds of years

(Kim 1996). At fast spreading ridges (East Pacific Rise, EPR}, the vents are 10's to 100's of m apart and may last 10 - 100 years

(Mullineaux 1994; Kim 1996). Recent investigations on Gakkel Ridge, an ultraslow-spreading ridge in the Arctic Ocean, have found more active vents than would be expected; these were localized near volcanic centers (Edmonds et al. 2003). This recent evidence suggests that the distance between vents may not be directly related to the spreading rate.

A unique feature at hydrothermal vents is that reduced sulfur species, as well as reduced metals that are expelled with the hydrothermal fluid, provide an energy source for chemoautotrophic bacterial growth. Free-living or symbiotic sulfur-oxidizing bacteria are important primary producers in vent environments (Fowler and

Tunnicliffe 1997). They fix carbon through oxidation of reduced chemical species found in vent fluid and can provide food for higher taxa which thrive in amazing abundance at many vents. The

endosymbionts associated with clams or tubeworms similarly provide

3 their hosts with nourishment. As the symbionts require sulfide for their energy metabolism, the hosts are restricted to the vent areas.

Surrounding individual black smokers on the East Pacific Rise were five distinct faunal zones. A general diagram of this zonation pattern is in Figure 1. The zone closest to the vent opening was characterized by very high temperatures (-250°C) dominated by alvinellid polychaetes (Hessler et al. 1985; Mullineaux et al. in press;

Van Dover and Hessler 1990). However, in my research this zone was not investigated, as it is very difficult to perform experiments here. A summary table describing the zones is in Table 1. The first zone (z1) considered in this thesis was characterized by high flow and high temperatures, elevated sulfide levels (up to 330 J.!M), and low but variable oxygen levels (Johnson et al. 1988). The predominant organisms in this zone were vestimentiferan tubeworms (e.g., ). The second zone (z2) was characterized by a moderate to low diffuse flow, and temperature a few degrees above ambient

(Hessler and Smithey 1983; Hessler et al. 1988). This area was

mostly colonized by bivalves (mussels and clams). The third zone (z3)

had low flow and a temperature at most a few tenths of a degree

above ambient (Hessler and Smithey 1983). The sulfide levels were

4 low, and oxygen levels were high. The organisms that reside here were primarily suspension feeders, typically serpulid worms. The fourth zone (z4) was outside the vent field and was characterized by ambient temperature, sulfide and oxygen levels. The organisms that resided here were typical deep-sea fauna. The approximate spatial range from zone 1 to zone 4 is 3 to 20 meters.

The spatial structuring of or plant communities in distinct zonation patterns is controlled by physical, biological or chemical variables and their interactions. The more familiar zonation in the rocky intertidal zone can, to some extent, be regarded as an analogue to the zonation pattern of vent environments. In the intertidal, benthic communities are also often controlled by extreme variables and structured along desiccation and temperature gradients. Other important factors for species survival and thus community structure are biological interactions, for example, predation. This interaction of physical, chemical and biological forces structures all ecosystems.

The purpose of the present work is to examine potential biological and physical interactions in the vent community by examining the pattern of larval settlement, and corresponding adult distributions, across the physicalfchemical gradient characteristic of the vent zones.

5 Vent Biology

The isolation and distances between hydrothermal sites have always provoked discussion on the larval dispersal, settlement and recruitment of the mostly endemic and sessile species found near vents. The vent environment includes unique microhabitats with equally unique species that thrive in this extreme environment The environmental factors of temperature, oxygen availability and sulfide concentration are dramatically different at hydrothermal vent sites than at the rest of the ocean floor (Johnson et al. 1994).

Two life history features should influence larval dispersal patterns of vent species: larval developmental mode and larval life span. The first. larval development mode, varies widely for vent endemic taxa. Some mytilid mussel larvae have demersal development (Lutz et al. 1980). Other bivalve larvae have been found near the surface of the oceans over vents (Berg and Van Dover 1987).

Bouchet and Waren (1994) found evidence of ontogenetic migration to the surface in planktotrophic larvae of vent gastropods including

Phymorhynchus (turrid), Shinkailepas (limpet), and Alviniconcha and

Desbruyeresia spp. ("hairy snail"). Larvae of vestimentiferans are

6 lecithotrophic (Marsh et al. 2001), although it is unknown where they develop. Thus, the larvae of vent dwelling organisms vary widely, probably along with their dispersal patterns. The evolutionary benefits of these differing strategies for vent organisms are at present unclear.

The second life history feature, larval life span, depends partly on whether larvae are planktotrophic or lecithotrophic. Planktotrophic larvae feed on the microplankton in the water column where an adequate food supply can extend survival for long periods of time

(Hessler and Kaharl 1995). However, microplankton is generally most abundant at the surface and rare at depth (Fuhrman et al. 1989), so many planktotrophic vent larvae may swim to the surface to feed then return to the deep-sea to settle. Conversely, some mytilid mussel larvae are both planktotrophic and demersal; such larvae apparently feed in the benthic boundary layer near vents (Lutz et al. 1980).

Lecithotrophic larvae do not feed but instead are provisioned with yolk that provides nourishment during development Larvae of vestimentiferans are lecithotrophic (Marsh et al. 2001 ). Recently,

Marsh et al. (2001) fertilized and reared the vestimentiferan, Riftia pachyptila, at in situ temperature and pressure in the lab. Based on their results, they estimate a metabolic larval lifespan to be 38 days.

7 Since it is generally believed that food supply is restricted in deep water, it makes sense that most vent species would produce non­

planktotrophic larvae (Lutz 1988; Gustafson and Lutz 1994) even though such larvae probably have shorter life spans and thus more

limited dispersal potential than planktotrophic larvae. Still, several

studies have shown that some lecithotrophic larvae absorb dissolved

organic matter (DOM) (Jaeckle and Manahan 1989a & 1989b;

Manahan 1990; Shilling and Manahan 1994). The absorption of DOM

could extend the survival of larvae of vent organisms, allowing for

greater dispersal distance. In addition, the cold ambient water of the

deep-sea may lower the metabolic rate of the larvae, thereby

extending time in the water before settling (Boidron-Metairon 1995;

Mullineaux and France 1995; Craddock et al. 1997).

Dispersal Mechanisms (To disperse or not)

Dispersal distance depends on the flow field as well as larval

survival time. Flows may act to retain larvae near the parent

population or may transport larvae to new sites. A positive

consequence of retention is ensuring a suitable habitat to settle in

while a disadvantage is that gene flow is limited. An advantage of long

8 distance dispersal is potential colonization of new vent fields and enhanced gene flow between vent fields. A disadvantage is larval loss.

Since hydrothermal vents are a patchy habitat, larval retention or an ability to return to suitable habitat would seem advantageous.

The larvae of vent organisms may be retained near their originating vent if they remain in near bottom flows. The currents moving along the ridge axis have a mean velocity of <2 cm/s, although speeds of up to 15 cm/s were recorded at 10°N EPR over a 3 day period (reviewed in Mullineaux and France 1995). The currents move cross-axis at

roughly between 2 and 5 cm/s (Kim and Mullineaux 1998). The overall elliptical flow pattern would cause larvae to be retained near the

originating vent site.

A similar patchy habitat, estuarine systems, may provide insight

into another potential larval retention mechanism through larval

behavior. In estuaries, larvae vertically migrate in the water column

with the tide; during ebb tide the larvae migrate lower in the water

column to prevent being washed out to sea, and at flood tide, the

larvae migrate higher in the water column to move further into the

estuary (Cronin and Forward 1982; 1986). At hydrothermal vents,

9 mesoscale eddies (e.g. Taylor caps) can retain a water mass in the proximity of the region of origin for weeks to months (Mullineaux 1994).

As discussed above, the currents may also transport the larvae off the ridge axis. The effect of these currents and eddies would depend upon when the larvae were released. Larval vertical migration, as observed in estuaries, could also affect the distribution of larvae of hydrothermal vent species.

Another long distance dispersal mechanism may be vent plumes. Plumes are the result of the release of hot buoyant fluid. As the fluid ascends, this flow creates eddies that entrain the ambient water and particulate matter, including organisms and larvae (Kim et al. 1994; Helfrich and Speer 1995; Lupton 1995; Speer and Helfrich

1995). The plume rises until mixing with ambient water and cooling make it neutrally buoyant, at which point it spreads laterally. Larvae may be entrained and thus transported upwards via the plume, then advected horizontally in ambient currents (Mullineaux 1994).

Settlement Cues and Recruitment

For species with planktonic and thus widely-dispersing larvae, settlement cues are thought to be especially important, and the

10 physico-chemical factors that are distinctive near vents could be potential settling cues. However, the relatively small habitat targets and often harsh conditions that characterize hydrothermal sites must also make settlement in the vent habitat a challenge to the larvae of vent endemics. Nevertheless, successful recruitment to this ephemeral and widely spaced habitat must depend on the dispersal of larvae and their subsequent successful settlement.

The cues for larval settlement in vent communities are still largely unknown. Larval settlement may be triggered by water temperature (Lutz et al. 1970; Mullineaux et al. 1997; Fisher 1998).

With increasing water temperature, the concentrations of silicate and sulfide also increase with concomitant decreases in oxygen levels

(Johnson et al. 1986). Another settlement cue may be the concentration of sulfides in the water. Some organisms need high levels for symbionts; others would avoid it due to toxicity. Settlement cues in other systems include light (Koback 2001; Strasser et al.

1999), gravity, conspecifics (Koback 2001), substrate type (Gutierrez

1998), and flow dynamics.

Several recruitment studies have been conducted at hydrothermal vent communities. Van Dover et al. (1988) and

11 Mullineaux et al. (1998) conducted studies at the EPR and Tunnicliffe

(1990) conducted a study a Juan de Fuca Ridge. Van Dover et al. compared recruitment within and outside the vent area at 21°N. This study used shale as a settling substrate and found more settlement within the vent area. The Mullineaux et al. study was at 9°N and used natural basalt obtained near 18°N. This was a comparison between two vent zones at two vent sites 1km apart. They found settlement in both zones with one species, Lepetodrilus elevantus, at both sites.

Tunnicliffe deployed slate arrays and a time-lapse camera. Recovery resulted in loss of many of the mobile species. However, visual observation showed that Lepetodrilus fucensis heavily populated several of the plates.

General Gastropod Biology and Adult Distribution

The larval development mode of several hydrothermal vent gastropod species has been deduced from the larval shell (Lutz et al

1984; Lutz 1988; Gustafson and Lutz 1994). Four specific species of interest in this study, Lepetodrilus spp. Clypeosectus delectus,

Gorgoleptis spp. and Eulepetopsis vitrea, have been inferred to

12 produce lecithotrophic larvae. The food source for these species is microbial films.

Physical characteristics vary with each species. Lepetodrl/us spp. larvae have a length of 120- 200 ~-tm. and the adult size range is

6 - 20 mm, depending on the species. One species is known to be sexually mature at 1/3 of its maximum length (Gustafson and Lutz

1994). Clypeosectus delectus larvae have a maximum diameter of

200 ~-tm. and an adult size range is 5 - 8 mm (Gustafson and Lutz

1994). Depending on the species of Gorgo/eptis, the larvae have a

maximum length of 120- 130 11m and an adult size range of 3-9 mm

(Gustafson and Lutz 1994). The larvae shell of Eulepetopsis vitrea has

a maximum length of 400 11m and is often lost on adults (Gustafson

and Lutz 1994). The adult can be up to 17 mm.

The adult distribution patterns are also species specific.

Lepetodrilus spp. has been found mostly in collections from Riftia in

the Vestimentiferan zone (Mclean 1988). In a study by Mullineaux,

Peterson, and Fisher (in press), adult Lepetodrllus spp. were found in

all zones of hydrothermal vent influence with decreasing abundance

further away from a central smoker. For C/ypeosectus delectus and

13 Gorgoleptis spp., a general habitat for both these species is unknown

(Mclean 1988, 1989b). However, Mullineaux et al. (in press), found the highest concentrations in the Vestimentiferan zone (z1) and the

Suspension zone (z3). Adults were in all areas of vent influence but in very low abundance in the Bivalve zone (z2). Eulepetopsis vitrea has been found on basalt substrate, mussels, and a few individuals from washings of vestimentiferan tubewonms (Mclean 1990). In the

Mullineaux et al. study (in press), the adults were found in all areas of vent influence.

The purpose of this study was to compare the influence of

environmental factors on larval settlement of gastropods in the four

distinct zones, as well as to investigate the impact of predation on

juvenile survival and thus adult community structure. This study

examined settlement patterns to artificial substrata to address the

following questions:

1) What is the zonal/habitat distribution of juvenile gastropods at

9°50'N EPR?

2) How does juvenile distribution compare to that of the adults

(size >1 mm) of the same species?

14 3) Do settlement (larval choice) or post-settlement processes (e.g.

predation) have a greater influence on adult distribution?

Methods and Materials

The study was carried out at 9°50' N on the East Pacific Rise

(EPR) at the East Wall site (Figure 2), where several studies have

been conducted by Mullineaux et al. (1998; 2000; in press}. To

determine the early recruitment patterns of vent fauna, settling

substrates of basalt, a natural substrate at vents, were deployed. The

basalt was acquired from a quarry in Washington State (Interstate

Rock Products). The basalt blocks were 10 em on a side with buoyant

polypropylene handles looped through a small hole in one corner that

facilitated deployment and retrieval.

The blocks were placed in four zones via the submersible Alvin

on November 1994 and retrieved in April 1995 to determine the early

settlement patterns. The zones were defined as: (1) vigorous diffuse

hydrothermal flow, identified by tubeworms; (2) moderate to low levels

15 of diffuse fluid hydrothermal flow, identified by bivalves; (3) very tow or no detectable hydrothermal flow, identified by suspension-feeders; and

(4) no exposure to vent flow, identified by non-vent species. The above zones will be referred to as z1 - Vestimentiferan, z2 - Bivalve, z3

- Suspension, and z4 - Periphery, respectively.

Six replicate basalt blocks were placed in each zone; three with cages and three without cages (Figure 3) (in zone 2, only two of the caged blocks were retrieved). The cages were designed to exclude large predators such as crabs, the octopods, and fishes (Mullineaux

1996; 1997; Micheli et al. 2002). Cages were constructed of 7 mm plastic mesh in a 20 em cube with the settling blocks suspended in the center of the cage with plastic cable ties to prevent abrasion of the

recruits (Mullineaux et al. 1997). The mesh size was selected to keep

out major predators and not become blocked by microbial growth.

Mullineaux (1997) previously conducted a cage control experiment to

determine whether there was a hydrodynamic effect of the cages on

recruitment. The cage controls consisted of basalt blocks elevated 2-{l

em above the seafloor with one side of the cage mesh missing. They

found no significant difference in animal abundance between cages

16 and cage controls indicating that caging did not appear to affect settlement (Mullineaux et al. 1997).

During retrieval, the settlement blocks were placed into isolated individual slots in a sealed collection box for transport to the surface.

Upon arrival at the surface, the organisms were removed from the blocks and the transport slot and passed through a 63J1m sieve to retain any individuals that may have detached during transport.

Organisms were preserved in 80% ethanoL

Mullineaux et al. (2000, in press) identified the most conspicuous organisms obtained from the blocks. This included gastropods >1mm; their data are used here for comparisons. Small juvenile gastropods remained unidentified even though they were often the most abundant taxa. These gastropods are the subjects of this study.

The terminology that I will use is that all individual gastropods that are less than 1mm will be referred to as juveniles. This encompasses both post-larvae and juveniles with growth beyond the larval shell. Since the blocks were deployed for only 5-months, it could not be determined if larvae settled 5 minutes or 4.9 months before retrieval. Results concerning settlement were inferred from

17 examination of the juveniles <1 mm. Larger animals were termed adults, and their abundances were previously determined by

Mullineaux et aL (in press).

The small gastropods (<1mm) were sorted under a dissecting microscope using 50x to 70x magnification. Whenever possible, identification was made at this level. However, in cases where the shell-sculpting pattern on the juvenile gastropods was indiscernible, a scanning electron microscope (SEM) was used.

The organisms were prepared for SEM by coating with gold­ palladium using a Pelco SC-7 sputter coater under 0.02 mbar of pressure. Both an lSI WB6 and a Philips XL40 SEM were used for analysis, and each produced equivalent images. Digital images of each gastropod were saved and used for identification based on the morphology of the protoconch (larval shell). The species level identifications were made using taxonomic guides of Mullineaux et al.

(1996), Waren and Bouchet (1989; 1993) and Mclean (1988; 1989a;

1989b; 1990).

18 Statistical analvses

Analyses were run for the most abundant juvenile gastropods encountered, Lepetodrilus spp., Eulepetopsis vitrea, and a combination

of G/ypeosectus delectus and Gorgoleptis sp. (C/ypeosectus/

Gorgo/eptis) (Figure 4). The latter group was combined due the

inability to distinguish between them (i.e. similar juvenile

morphologies).

Parametric 2-way model 1 ANOVA was run using SYSTAT 9.0

on rank transformed count data (S. Bros pers. comm.) at a.=0.05. The

ANOVA was used to determine if there were differences between

zones (Vestimentiferan (z1), Bivalve (z2), Suspension (z3), and

Periphery (z4)), between treatments (caged and uncaged), and

interactions between zones and treatment. When a statistically

significant difference was found, an a priori ANOVA was performed

using the ranked count data (S. Bros pers. comm.). The number of

comparisons that can be done for a priori tests is limited to 'a - 1'

where 'a' is the number of levels for the independent variable (i.e.

zones). This analysis was limited to three comparisons. The three

most interesting comparisons between zones were chosen. The

19 statistical significance levels are reported without adjustment due to low replication.

To determine if there was a difference between adult and juvenile distribution across zones, Chi-square goodness of fit tests were run. Each species was examined separately across all zones.

The juvenile distribution pattern was set as the expected distribution with a=0.05 for df=3.

Results

Juveniles of eight different gastropod species were identified in this study (Table 2). Images of these species appear in Figures 4 and

5. Lepetodrilus spp., Eulepetopsis vitrea, Clypeosectus delectus and

Gorgoleptis sp. (Figure 4) were in high abundances and statistical analyses were performed on these species. Cyathermia naticoides,

Melanodrymia sp., Peltospira sp., and Rhynchopelta concentrica

(Figure 5) were found in very low abundances, eight specimens total.

These numbers were so low that statistical tests could not be

20 performed, and trends could not be distinguished. Many specimens remained unidentified (- 21 %) because the shells were in small pieces, the protoconch had been etched beyond recognition, or the shells had become decalcified.

Juveniles

Caging effects v.aried by species (Table 3). Only for

Eu/epetopsis vitrea, was there a significant cage effect, with more settlers on caged blocks (Table 3). The cages provided protection from predation, a post-settlement process. These limpets are larger than the other species in this study and therefore potentially more

"visible" to predators as larvae or recent settlers. There were no significant cage effects for Lepetodri/us spp. and C/ypeosectusl

Gorgoleptis, so the juvenile patterns observed in the different zones presumably are not affected by post-settlement predation for these three species.

The abundance of juvenile Lepetodri/us spp. varied by zone.

There was a significant higher abundance of juvenile Lepetodri/us spp. in the Vestimentiferan (z1) zone than in the Periphery (z4) zone (fable

4). The lower abundance of these young away from vents

21 suggests that characteristics of the vent habitat attract settlers.

Eulepetopsis vitrea and ClypeosectusiGorgoleptis juvenile abundances were not different across zones (Table 3), suggesting that larvae of these three species settle indiscriminately across the four zones.

There were no interactions between zone and treatment.

Juveniles vs. Adults

A Chi-square goodness of fit test tested for overall differences in abundance of adults versus juveniles. For Lepetodrilus spp., there was a statistically significant difference between the adults and juveniles across zones, (x.2=461.781, (p<0.001 )), with adults more abundant in Vestimentiferan (z1) zone and Bivalve (z2) zone but less abundant in Suspension (z3) zone (Figure 6). For Clypeosectusl

Gorgoleptis, the comparison of the distribution patterns between adults and juveniles also showed that there was a significant difference

(·l=539.552, (p<0.001)), with adults more abundant in Vestimentiferan

(z1) zone and Bivalve (z2) zone but less abundant in Suspension (z3) zone (Figure 7). In Eulepetopsis vitrea, no difference was observed

(x.2=0.052, (p=0.996)) (Figure 8).

22 Discussion

This study provides insight into the early settlement patterns for hydrothermal vent gastropods, as reflected in the distribution of small juveniles on introduced substrate. The gastropods, Lepetodrilus spp.,

Clypeosectus/Gorgoleptis, and Eulepetopsis vitrea all produce lecithotrophic larvae, but which exhibit differential settlement success across the vent spatial gradient (Figures 6-8). The short deployment time of 5-months for the settlement blocks was a useful approach to investigate these patterns. In the deep-sea, cost limitations require that experimental manipulations involve a balance between replication and bottom time. Although limited in the number of replicates of caged and uncaged treatment blocks, this experiment was still sufficient to provide insight into the role of predation early in the settler's life. At this vent site, East Wall on the East Pacific Rise, the settlement patterns were different for each gastropod species.

23 Dispersal potential

The gastropods, Lepetodrilus spp., Clypeosectus/Gorgofeptis, and Euiepetopsis vitrea, analyzed in this study have been implied to produce lecithotrophic larvae (Lutz et al. 1984; Lutz 1988; Gustafson and Lutz 1994). This development mode may well provide the ability for these larvae to disperse long distances or disperse then return to the same vent site. Marsh et al. (2001) estimated the larval lifespan of

Riftia pachyptiia to be 38 days. Kim and Mullineaux (1998) found most vent gastropod larvae in the near bottom (within 15m above bottom), both inside and outside the axial ridge. At heights of 10-45m above bottom, they also observed gastropod larvae.

Potential dispersal mechanisms include hydrothermal plumes, currents, or mesoscale eddies, as discussed earlier. Hydrothermal vent plumes ascend entraining ambient water until the water mass becomes neutrally buoyant which then spreads out laterally.

Mesoscale eddies formed by the rising plume and the Coriolis effect can break away from the plume and remain a coherent water mass while moving away (reviewed in Van Dover 2000). Currents are constrained by the ridge crest topography and flow up and down the

24 ridge axis. If larvae get into these different flow patterns, they have the ability to be transported to a different area or retained.

Vents are temporally ephemeral. They senesce, disappear and new ones appear. The larvae that are in the water column must be at least occasionally available to settle these new sites. The actual mechanisms of colonization at new sites are still unknown.

New tools, such as genetic testing, may provide some insight into larval dispersal distances. For two of the species in this study,

Lepetodrilus spp. and Eulepetopsis vitrea, genetic testing has been successfully applied to determine that these species have long distance dispersal (Vrijenhoek 1997; Vrijenhoek et al. 1998). Won et al. (2003) have also found that the diverging oceanic flows that cross mid-ocean ridges apparently isolate the sections on either side. This results in genetic differences within the same species on each side of the ridge axis.

Juvenile distribution

The zonation of the communities at hydrothermal vents has been compared to rocky intertidal zonation. Near the smoker opening, high levels of sulfides favor species that contain symbionts. These

25 symbionts utilize the sulfides, and energetically support the host organisms. At vents, the harshest physical conditions co-occur with the most abundant energy source. Conversely, in the intertidal, the harshest physical conditions are high in the intertidal above the low intertidal areas of high food availability. More organisms settle at lower levels of the shoreline than at the upper levels (Connell 1972; Menge and Branch 2001) where the organisms are least impacted by wave exposure and desiccation. The opposing trends - more settlement in the physically stressful area near vents versus more settlement in the physically benign area in the intertidal - indicate that some factor other than physical stress is influencing settlement. Lepetodrilus spp. settled predominately in the Vestimentiferan zone (z1); the most stressful environment. Clypeosectus/Gorgoleptis settled mainly in the

Suspension zone (z3) which Eulepetopsis vitrea had no preference.

Food availability is a factor that is high near vents and lower in the intertidal, and may be controlling settlement in both these habitats. For

Lepetodrilus spp., another factor may be that it contains a symbiont. A species of Lepetodrilus at Juan de Fuca Ridge was found to multiple ways to obtain nourishment which included grazing, a symbiont or a combination of the two (Fox et al. 2002).

26 Another factor affecting distribution is space competition, which is observed in many communities. In the intertidal, organisms actively remove other organisms (Benedetti-Cecchi 2000) or passively crowd out others (Connell, 1961 ). At hydrothermal vents, Micheli et al. (2002) suggest that new settlers may be dislodged from the substrate by the grazing of previous settlers or adults. The grazing also removes a food source, surface-attached microbes (Hessler and Smithey 1983; Van

Dover and Fry 1994). In the present experiment, the deployed basalt blocks were initially completely bare. This represents a maximum of space availability and surface for the development of a microbial film, the first step in microhabitat creation. This then offers maximum space and food availability and therefore maximum settlement yield. For 5- months, there were an average of eight gastropods/block of both adults and juveniles. The size range of these gastropods was <1mm to 20mm. Therefore, space competition may not have been a factor in these experiments.

Although not statistically significant, there was an interesting trend of low or no settlement in the Bivalve (z2) zone. This lack of settlement suggests that biotic forces may be contributing not only to post-settlement mortality but also to settlement patterns themselves.

27 Lenihan et al. (2001) have shown that recruitment to mussel patches at vents is lower than in areas with no mussels. In aquatic systems,

Dreissena polymorpha (zebra mussel), is found in dense beds similar to those at hydrothermal vents. Though no studies on clearance rates have been done on vent mussels, studies have been conducted on

Dreissena polymorpha. Pace et al. (1998) have reported zebra mussels reducing zooplankton biomass by >70%. This would suggest that vent mussels could consume larvae as they pass over.

Role of predation

After settlement, predators may reduce abundances of juveniles. In this study, predation was a factor only for Eulepetopsis vitrea as there were more in the caged blocks. An interesting observation is that a large number of gastropods from zone 1 caged blocks remained unidentified. Dominant predators have been shown in the intertidal to play a role in community structure (Menge and Branch

2001). Mobile predators have an impact at vents (Micheli et al. 2002).

Micheli et al. (2002) and Mullineaux et al. (in press) showed through gut analysis of the vent fish, cerberus, that it is a major predator of gastropods. Mullineaux et al. (in press) observed an

28 increase in abundance of gastropods in cages that excluded T. cerberus. Other predators include crabs and octopods. The size of mesh, 7 mm, used for the cages in this study effectively blocked these predators. Although the cages excluded predators, they also provided an additional surface for microbial film to develop. Therefore, more food was likely available on the caged blocks, which may have produced a cage effect However, Mullineaux et al. (1997) found no cage effect when comparing cage and cage control blocks utilizing the same caging design. Predation was statistically significant only for the species with the largest juvenile (Eulepetopsis vitrea). The large size may make them more obvious to predators.

Adult vs. Juvenile distribution

Lepetodrilus spp. and Clypeosectus/Gorgoleptis (the smallest juveniles) showed no effect of predation in the caging experiment

However, there was a significant difference between adults and juveniles for Lepetodrilus spp. and Clypeosectus!Gorgoleptis. There was a trend, for both species, of more juveniles in the Suspension (z3) zone. This suggests that migration and/or mortality were affecting the

29 observed distribution. Both these mechanisms can be influenced by predation, habitat unsuitability or space competition.

Migration between blocks or zones would allow species to find the appropriate microhabitat where they could survive. In this study, it was not specifically investigated whether these gastropods migrate between the zones. However, Mullineaux et al. (unpub. data) showed no change is distribution for Lepetodrilus spp. and Glypeosectusl

Gorgoleptis at the time intervals of 8- and 13-months. This suggests that within a 5-month time period, these gastropods found the appropriate habitat and remained there.

Food availability may also affect survival and therefore the observed species distributions. The amount of food available (e.g. microbial mat) differs in each zone. There is a gradient with the highest concentration of microbes in the Vestimentiferan (z1) zone and a general decrease away from the vent smoker.

Conclusion

Juvenile gastropods occurred in all areas of vent influence but rarely in the Periphery zone (z4). For Lepetodrilus spp. there was a

30 decrease in juvenile abundances between the Vestimentiferan (z1) zone and Periphery (z4) zone, suggesting variable characteristics (e.g. chemical or food availability) within the vent habitat enhance settlement for this species in the Vestimentiferan (z1) zone. Significant differences in abundance of adults and juveniles were found for

Lepetodrilus spp. and Clypeosectus!Gorgoleptis (more juveniles in z3 and more adults in z1 and z2) suggesting that post-settlement processes (e.g. predation or migration) were superimposed on the initial settlement response to vent habitat to create the adult distribution patterns. There was also a significant positive caging effect found for Eulepetopsis vitrea suggesting that, especially in this species, predation influenced adult distributions.

This study addressed post-settlement processes that have been shown to affect distribution patterns. The results raised additional questions concerning pre-settlement processes. First, the observation of low juvenile abundances in the Bivalve zone led to the question of what impact the mussels have on larval settlement in that zone.

Secondly, this study demonstrated that settlement occurred in zones of

hydrothermal influence, raising the question of what are the cues for larval settlement. Finally, the actual mechanism(s) for how larvae

31 disperse remain unknown; specifically, are the larvae retained; do they disperse then return to the same vent; or do they disperse to distant vents? In combination with research on these potential future directions, the post-settlement questions addressed in this study are important to increase our understanding of settlement at new vent sites.

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41 Table 1. Generalized descriptions of hydrothermal vent zones at East Pacific Rise. ** Zone is not sampled in this study due to difficulty in performing experiments here.

Zone# Zone Temp°C Sulfide J.tm Oxygen !J.m Dominant Organisms 0** -250 0 Alvinellid polychaetes 1 Vestimentiferan Up to 30 Up to 330 >25-0 Riftia pachyptila 2 Bivalve 2.3-7 27- 150 75-25 Mussels and clams 3 Suspension 2-2.3 0-27 110- 75 Serpulid worms 4 Periphery 2 0 -110 Deep-sea fauna

42 Table 2. Identified juvenile hydrothermal vent gastropod species (in order of abundance) which settled from November 1994 to Apri11995 at East Wall vent site. The numbers are the Individuals of each species in each zone and deployment type.

Species Zones Vestimeniferan Bivalve Suspension Periphery Caged Uncaged Caged Uncaged Caged Uncaged Caged Uncaged Lepetodrilus spp. 533 264 5 78 250 36 1 Clypeosectus!Gorgoleptis 48 3 623 59 Eulepetopsis vitrea 3 1 1 4 Melanodrymia sp. 1 2 1 Cyathermia natacoides 1 1 Peltospira sp. 1 Rhynchopelta concentrica 1 Unidentified 415 12 11 51 9

43 Table 3. Statistical results of ANOVA for comparison between zones (Vestimeniferan, Bivalve, Suspension, and Periphery) and for treatment effects (caged vs. uncaged) of the juveniles. The symbol • denotes significance at the 0.05 level.

Species Effect ss df MS F p Lepetodri/us spp. Zone 354.533 3 118.178 3.884 0.031+ Treatment 0.708 1 0.708 0.023 0.881 Zone*Treatment 112.533 3 37.511 1.233 0.333 Error 456.458 15 30.431

C/ypeosectus!Gorgoleptis Zone 192.018 3 64.006 2.623 0.089 Treatment 20.745 1 20.745 0.850 0.371 Zone*T reatment 20.965 3 6.988 0.286 0.834 Error 366.000 15 24.400

Eu/epetopsis vitrea Zone 103.695 3 34.565 2.000 0.157 Treatment 164.414 1 164.414 9.511 o.oos• Zone*Treatment 121.748 3 40.583 2.348 0.114 Error 259.292 15 17.286

44 Table 4. Statistical results of a priori ANOVA for comparison between zones of the juveniles. V=Vestimeniferan; B=Bivalve; S=Suspension; P=Perlphery. The symbol+ denotes significance at the O.OSievel.

Species Zone Effect ss df MS F p Lepetodrilus spp. Vvs. B 63.375 1 63.375 2.083 0.170 Vvs.S 102.083 1 102.083 3.355 0.087 Vvs. P 352.083 1 352.083 11.570 0.004+ Error 456.458 15 30.431

45 Zonation of resident fauna

Vestimen!iferan Bivalve Suspension-feeder Periphery

Figure 1. Generalized diagram of zonation pattern at an East Pacific Rise black smoker vent site. (Modified from Mullineaux at al. in press)

46 Biovent

East Wall

Worm Hole

9°48' +------'------..,------! 104°18'36" 104°17'24" 104°16'12"

Figure 2. East Wall vent site at 9°50'N East Pacific Rise between the Clipperton (C) and Sequieros (S) fracture zones (from Mullineaux et al., 2000). Arrow denotes study site.

47

in ;(yt.>ec•sectu's delectus D) Gorgoleptis sp..

49 Figure 5. B) Me,lar.lod'rvtnia sp. C) Peltospira sp. D) Rhynchopelta concentrica.

50 11!11 Juvenile D Adult

200

100 ~i

0 v B s p Zone figure 6. Distribution of adult and juvenile Lepetodrilus spp. across zones. V=Vestlmeniferan; B=Bivaive; S=Suspension; P=Peripheii'Y. !Error bars indicate standard errol'S.

51 22 5 - -- ~-~--- - ~ --~------

Juvenile D Adult

150 -,

0 v B s p Zone

Figure 7. Distribution of adult and juvenile Clypeosectus delectus and Gorgoleptis sp. across zones. V=Vestimeniferan; IB=Bivaive; S=Suspension; P=Periphery. Error b;:us indicate standard errors.

52 1.5 llftl Juvenile o Adult

0.0 v 8 s p Zone

Figure 8. Distribution of adult and juvenile Eulepetopsis vitrea across zones. V=Vestimeniferan; B=Bivalve; S=Suspension; P=Periphery. Error bars indicate standard errors.

53