The use of DNA markers to identify and explore the genetic diversity of lionfishes Pterois miles and

Pterois volitans

Rebecca M. Hamner

Advisor: Wilson Freshwater

Pterois miles Species Pterois volitans10 Dorsal Fin Rays 11

6 Anal Fin Rays 7

Shorter Pectoral Fin Size Longer

Smaller Spot size Larger

Indian Ocean, S. Africa to Red Sea,

East to Sumatra

Distribution West Pacific Ocean from Japan to Australia,off western Australia, South Pacific to Pitcairn Group

P. miles P. volitansMixedInvasive Pterois sp.

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 19

Aquarium 13

Aquarium 15

Aquarium 20

Aquarium 07

P. volitans 01

P. volitans 02

P. volitans 03

P. antennata 01

P. antennata 02

P. mombasae 01

P. mombasae 02

D. zebra 01

D. zebra 02

P. radiata 01

P. radiata 02

D. brachypterus 01

D. brachypterus 03

D. brachypterus 02

H. hilgendorfii

0.005 substitutions/site

Pterois milesClade

Pterois volitansClade

■ Meristic ID P. miles■ Meristic ID P. volitans■ Meristic ID Unknown

100

100

100

85

87

10077

100

100

10099

94

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 19

Aquarium 13

Aquarium 15

Aquarium 20

Aquarium 07

P. volitans 01

P. volitans 02

P. volitans 03

P. antennata 01

P. antennata 02

P. mombasae 01

P. mombasae 02

D. zebra 01

D. zebra 02

P. radiata 01

P. radiata 02

D. brachypterus 01

D. brachypterus 03

D. brachypterus 02

H. hilgendorfii

0.005 substitutions/site

Pterois milesClade

Pterois volitansClade

■ Meristic ID P. miles■ Meristic ID P. volitans■ Meristic ID Unknown

100

100

100

85

87

10077

100

100

10099

94

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 19

Aquarium 13

Aquarium 15

Aquarium 20

Aquarium 07

P. volitans 01

P. volitans 02

P. volitans 03

P. antennata 01

P. antennata 02

P. mombasae 01

P. mombasae 02

D. zebra 01

D. zebra 02

P. radiata 01

P. radiata 02

D. brachypterus 01

D. brachypterus 03

D. brachypterus 02

H. hilgendorfii

0.005 substitutions/site

Pterois milesClade

Pterois volitansClade

■ Meristic ID P. miles■ Meristic ID P. volitans■ Meristic ID Unknown

100

100

100

85

87

10077

100

100

10099

94

Current Data

Disagreement between mitochondrial DNA and meristic identities

14 nuclear DNA loci lack species specific variation for P. miles and P. volitans

Act-2 AldB Cam-3

CK CKA7 ChymB-6

ITS LDH LDHA6

Mlc-3 Ops-1 RP1

RP2 SSU

Objective I

Investigate hybridization

using Randomly Amplified

Polymorphic DNAs (RAPDs) to

develop a nuclear DNA marker

to distinguish between Pterois

miles and Pterois volitans

Methods

Randomly Amplified Polymorphic DNAs (RAPDs)

Primer sitesCopied DNA fragments

Template DNA

1. Polymerase Chain Reaction5’ 3’

1 2 3 4

A ? B

2. Gel Electrophoresis

longest fragments

shortest fragments

Size marke

r

Objective II

Examine the genetic diversity

of North Carolina lionfish to

determine their source

population

?

Methods

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 19

Aquarium 13

Aquarium 15

Aquarium 20

Aquarium 07

P. volitans 01

P. volitans 02

P. volitans 03

P. antennata 01

P. antennata 02

P. mombasae 01

P. mombasae 02

D. zebra 01

D. zebra 02

P. radiata 01

P. radiata 02

D. brachypterus 01

D. brachypterus 03

D. brachypterus 02

H. hilgendorfii

0.005 substitutions/site

Pterois milesClade

Pterois volitansClade

■ Meristic ID P. miles■ Meristic ID P. volitans■ Meristic ID Unknown

100

100

100

85

87

10077

100

100

10099

94

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 19

Aquarium 13

Aquarium 15

Aquarium 20

Aquarium 07

P. volitans 01

P. volitans 02

P. volitans 03

P. antennata 01

P. antennata 02

P. mombasae 01

P. mombasae 02

D. zebra 01

D. zebra 02

P. radiata 01

P. radiata 02

D. brachypterus 01

D. brachypterus 03

D. brachypterus 02

H. hilgendorfii

0.005 substitutions/site

Pterois milesClade

Pterois volitansClade

■ Meristic ID P. miles■ Meristic ID P. volitans■ Meristic ID Unknown

100

100

100

85

87

10077

100

100

10099

94

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

P. miles 01

P. miles 06

P. miles 03

P. miles 04

Aquarium 04, 09, 14, 21

Aquarium 27, 28

Aquarium 01, 02, 08, 24, 25

P. miles 02

P. miles 05

P. miles 08

P. miles 07

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 03, 05, 06, 10,11, 16, 17, 18, 22, 23, 26Long Island 01, 03

Aquarium 19

Aquarium 13

Aquarium 15

Aquarium 20

Aquarium 07

P. volitans 01

P. volitans 02

P. volitans 03

P. antennata 01

P. antennata 02

P. mombasae 01

P. mombasae 02

D. zebra 01

D. zebra 02

P. radiata 01

P. radiata 02

D. brachypterus 01

D. brachypterus 03

D. brachypterus 02

H. hilgendorfii

0.005 substitutions/site

Pterois milesClade

Pterois volitansClade

■ Meristic ID P. miles■ Meristic ID P. volitans■ Meristic ID Unknown

100

100

100

85

87

10077

100

100

10099

94

ABI 3100 Genetic Analyzer

NC lionfish

Phylogenetic tree or Cluster

analysis

?

PCR & Sequence Reaction

Questions?

Leo (NC lionfish) at CMS

Mer = mMito = vNuc = v

Mer = vMito =

mNuc = m

Mer = mMito = vNuc = m

Mer = vMito =

mNuc = v

Nuclear and mitochondrial

DNA disagreement

-Hybridization-Incomplete

lineage sorting

Meristic and DNA disagreement

Meristic characters not fixed for P. miles

and P. volitans

Mer = meristic countsMito = mitochondrial DNANuc = nuclear DNA

2. Gel Electrophoresis

Partis, L. and R. J. Wells. 1996. Identification of fish species using random amplified polymorphic DNA (RAPD). Molecular and Cellular Probes, 10: 435-441.