Marraffini, m invasibility of communities supplementary Materials



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Marraffini, M


INVASIBILITY OF COMMUNITIES

Supplementary Materials



Figure SM1. Experimental Setup. A. attachment of PVC frame that held experimental blocks attached to floating dock. B. View of diver (Heather Hawk) assisting with installation of a block. C. View of a tile several weeks after installation with solitary tunicates and Sabellidae tube worms that were attached using marine epoxy. D. Close up of treatment tiles in in block B showing that attachment of animals using marine epoxy. Photos A, B, D by Scott Gabara; photo C by Michelle Marraffini.

Figure SM2. Total number of recruits after 100 days plotted as a function of original species richness (A); original percent cover (B), and original percent of native species (C). Data are binned across other factors to examine the main effect in each panel.



Table SM1 Approximate sizes of adult organisms used in the experiment. Size is taken as the longest axis of a solitary animal and the diameter of a colonial animal. For anemones, this is their width in their non-contracted form.

Species

Phyla

Body Plan

Range of sizes (cm)

Botrylloides violaceus (I)

Chordata

Colonial

5-10

Watersipora subtorquata (I)

Bryozoa

Colonial

5-10

Mytilus galloprovincialis (I)

Mollusca

Solitary

7-10

Bugula neritina (I)

Bryozoa

Colonial

5-8

Ciona savignyi (I)

Chordata

Solitary

4-8

Diplosoma listerianum (I)

Chordata

Colonial

3-7

Botryllus "schlosseri"(I)

Chordata

Colonial

2-6

Ascidia ceratodes (N)

Chordata

Solitary

7-10

Balanus crenatus (N)

Arthropoda

Solitary

1-2

Corynactis californica (N)

Cnidaria

Solitary

1-2

Metridium senile (N)

Cnidaria

Solitary

3-6

Aplidium californium (N)

Chordata

Colonial

5-10

Distaplia occidentalis (N)

Chordata

Colonial

3-7

Barentsia ramosa (N)

Entoprocta

Colonial

2-6

Eudistylia polymorpha (N)

Annelida

Solitary

10-20

Mytilus californianus (N)

Mollusca

Solitary

7-10



Organisms_recruited_during_the_experiment.'>Table SM2. Organisms recruited during the experiment. Listed in order of appearance and asterisks indicate species that were not originally used in treatments.

Organism

 

 

Date of Appearance

Successful Recruit

Phyla

Lowest Taxonomic Level

Native Status

DOY

Day of Experiment

Size (mm)

Annelida

Spirorbis sp.*

Native

7/16/12

14

N/A

Chordata

Diplosoma listerianum

NIS

7/16/12

14

2

Chordata

Ascidiacea (colonial)*

Unknown

7/16/12

14

2

Chordata

Botrylloides violaceus

NIS

7/30/12

28

2

Bryozoa

Watersipora subtorquata

NIS

7/30/12

28

1

Chordata

Botryllus schlosseri

NIS

8/13/12

42

2

Chordata

Ciona savignyi

NIS

8/13/12

42

2

Annelida

Salmacina tribranchiata*

Native

8/13/12

42

2

Arthropoda

Balanus crenatus

Native

9/10/12

70

2

Bryozoa

Bugula neritina*

NIS

9/24/12

84

1

Annelida

Serpula columbiana*

Native

9/24/12

84

2


Table S3. Estimates of variation explained by the GLMMs used in this experiment. ICC refers to interclass correlation, explaining the proportion of variance explained by random intercept over the variance of the residuals, interpreted as the total variance between groups (Zuur et al., 2009). Time is in days into the experiment.

Model

Table

Time

Response

Components

ICC

Rc2

Rm2

Overall GLMM

4

100

Recruits

R, PC, PNS, R:PC, Time, Block

0.5

0.85

0.46

Early Recruitment

S2

57

Recruits

R, PC, PNS, R:PC, Time, Block

0.5

0.66

0.32

Stachowicz Comparison

7

71

Recruits

R, PC, PNS, R:PC, Time, Block

0.5

0.66

0.31

Invasions

S3

30

NIS Recruits

R, PC, PNS, R:PC, Time, Block

0.5

0.61

0.21


Table SM4. Estimated parameters from GLMM for the first 57 days. Model for cumulative recruitment during the first 57 days of the experiment.




Parameter

Estimate

Std. Error

P value

Variance

 Fixed

Intercept

1.0382

0.3650

0.00445

--

 

Richness (PR)

0.0435

0.0066

3.44e-07

--

 

Percent Cover (PC)

0.0118

0.0006

<2e-16

 

 

Percent Native Species (PPNS)

-0.018

0.0002

<2e-16

--

 

R:PC

0.0003

0.0003

<2e-16

 

 

Time

-0.0029

0.0019

<2e-16

--

 Random

Block

--

--

--

6.631e-01




Time

--

--

--

1.797e-05


Table SM5. Estimated parameters from GLMM for the first 71 days. Model for cumulative recruitment during the first 71 days of the experiment, for comparison to previous experiments (6, 7).




Parameter

Estimate

Std. Error

P value

Variance

 Fixed

Intercept

1.2907

0.3387

0.00014

--

 

Richness (PR)

-0.0349

0.0057

1.6e-09

--

 

Percent Cover (PC)

-0.0203

0.0006

<2e-16

 

 

Percent Native Species (PPNS)

-0.0034

0.0002

<2e-16

--

 

R:PC

-0.0057

0.0002

<2e-16

 

 

Time

0.0223

0.0012

<2e-16

--

 Random

Block

--

--

--

5.713e-01




Time

--

--

--

6.955-06


Table SM6. Estimated parameters from the best-fit GLMM on NIS recruitment. Modeling cumulative recruitment of NIS during the first month of the experiment.




Parameter

Estimate

Std. Error

P value

Variance

 Fixed

Intercept

-0.0585

0.4528

0.897

--

 

Richness (PR)

-0.013

0.0218

0.551

--

 

Percent Cover (PC)

-0.0256

0.0023

<2e-16

 

 

Percent Native Species (PNS)

-0.004

0.0009

8.39e-06

--

 

R:PC

-0.0051

0.0009

4.95e-08

 

 

Time

0.0676

0.0095

1.01e-12

--

 Random

Block

--

--

--

0.6764




Time

--

--

--

0.0003


Genetic Methods

When morphological identification was not feasible, genetic methods were used to confirm the identity of recruited individuals (genus or species) and organisms in community manipulations. A sample of DNA was extracted from fresh tissue using Promega© Wizard 96-well DNA extraction kits following manufacturer’s protocol, with the additional step of tissue maceration by bead beating before overnight incubation. Polymerase chain reaction (PCR) amplified the COI region with primers jgLCO1490 and jgHCO2198 [1x Green Gotaq Master Mix, Promega (Cat #PRM7123), 0.2 M BSA, 1.5 mM MgCl2, 0.2 μM of each primer] using a three-step thermocycler program (three minute at 94C followed by 30 cycles of 94C for one minute as a denaturation step, 47C for one minute as the annealing temperature, and an extension step at 72C for one minute) using universal COI primers (Geller et al. 2013). PCR products deemed successful as bright single bands visualized by agarose electrophoresis were sequenced by the Sanger method (Elim Biopharmaceuticals, Hayward, CA). Geneious© software (by Biomatters Ltd.) was used to assemble forward and reverse reads, and edited sequences were compared to known sequences in Genbank (National Center for Biotechnology Information) and a reference library developed in the San Francisco invasive species monitoring program (Geller et al. 2013). Similarity of sequences from unknown specimens to a reference sequence of 95% or greater was considered a match for identification.



Results

Ninety-three samples were successfully amplified by PCR and sent for Sanger sequencing. Of these samples 61 sequences, showed strong quality sequences that could be used for analysis (Table S4). This analysis helped resolve the identification of two species (an anemone and Balanus sp.) too small for morphological identification. These specimens had results from Genbank but the identifications were to species never recorded in this area and the matches were suspect. Comparisons to the Geller database revealed better matches and identifications of species known to inhabit these areas. Several specimens remained unresolved as the samples failed to amplify or to give a clean, high quality sequence. Further work is needed to resolve the identity of these samples. The unresolved species were not assigned native or NIS status and were removed from analyses on invasion success.

References

Geller, J. , Meyer, C., Parker, M. and Hawk, H. 2013. Redesign of PCR primers for mitochondrial cytochrome c oxidase subunit I for marine invertebrates and application in all-taxa biotic surveys. Molecular Ecology Resources 13(5) 851-861.




Table S4. Field and molecular identifications of organisms used in treatments. Shows comparisons with Genbank database based on the best sequence.

Field Identification

Number of Samples

Molecular Identification

Accession Number

Percent Identical

Final Identification

Anemone (small brown)

4

Aiptasia pulchella

HG423158.1

95

Unknown

Balanus sp.

5

Amphibalabus reticulatus

JQ035518.1

89

Unknown

Lamillaria sp.

4

Assiminea pecos

DQ533844.1

83

Lamillaria sp.

Botrylloides violaceus

7

Botrylloides violaceus

GQ365691

100

Botrylloides violaceus

Colonial Tunicate

3

Botryllus schlosseri

JN083284

100

Botryllus schlosseri

Botryllus schlosseri

3

Botryllus schlosseri

JN083238

99

Botryllus schlosseri

Bugula sp. (brown)

1

Bugula neritina

KC129832.1

99

Bugula neritina

Bugula sp. (white)

1

Bugula stolonifera

KC129849.1

100

Bugula stolonifera

Encrusting Bryozoa

2

Celleporaria oculata

AY168607.1

94

Unknown

Corynactis californica

1

Corynactis californica

AB441257.1

100

Corynactis californica

Sabellid sp.

3

Eudistylia vancouveri

HM473366.1

90

Eudistylia polymorpha

Sabellid sp. (smaller tube)

2

Megalomma splendida

HM473463

98

Megalomma splendida

Mytilus galloprovincalis

6

Mytilus galloprovincalis

GQ480284.1

100

Mytilus galloprovincalis

Sponge (yellow)

3

Haliclona xena

JN242209.1

99

Haliclona sp.

Anemone (small)

5

Metridium senile

HG423143

99

Metridium senile

Mytilus californianus

4

Mytilus californianus

GQ902188.1

99

Mytilus californianus

Watersipora sp.

2

Watersipora sutorquata

DQ417456.1

99

Watersipora sutorquata

Tube (Amphipod)

1

Ericthonius brasiliensis

JX545463

89

Amphipoda

Diplosoma listerianum

4

Diplosoma listerianum

KF791868.1

99

Diplosoma listerianum

Total

61















Table S5. Field and molecular identifications of organisms unresolved by Genbank. Shows comparisons with Genbank database and Geller lab database based on the best sequence.

Field Identification

Genbank

Identification

Accession Number

Percent Identical

Geller Lab Identification

Percent Identification

Finial Identification

Anemone (small brown)

Aiptasia pulchella

HG423158.1

95

Diadumene lineata

100

Diadumene lineata

Balanus sp.

Amphibalabus reticulatus

JQ035518.1

89

Balanus crenatus (haplotype A)

98

Balanus crenatus







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