Wednesday, February 1, 2012

February 1st, 2012 class

1. Introduction: Problems facing taxonomy and diversity.
2. Accelerating “taxonomy”: DNA barcoding.
3. Promoting taxonomy: Census of Marine Life.
4. Images from one CoML Project (Creefs).
5. Conclusions.
6. Test advice.

1. Introduction: Problems facing taxonomy and diversity.
Problems facing taxonomy 1
Too many species! Diversity confounds our best efforts to examine it.
Keep finding new species.
Extinction rates increasing.
Problems facing taxonomy 2
Not enough taxonomists.
Pay poor, work long.
Everyone says “important” but not considered essential.
Many groups have no active workers.
Potential solutions 1
Increasing technology and information available.
Global information systems.
Molecular experiment techniques.
Potential solutions 2
Increasing international research collaboration.
Growing awareness of biodiversity and importance.

2. Accelerating “taxonomy”: DNA barcoding.
What is “DNA barcoding”?
A DNA barcode is a short sequence, taken from standardized portions of the genome,used to identify species.
If a genome project is deep and narrow, DNA barcoding is broad and shallow.
Genome projectは深くて、狭いが、遺伝子バーコードは浅くて、広い。
Requirements of a DNA barcoding marker
A sequence/marker used to barcode should:
be easy to amplify
not possess paralogues
have conserved regions to design primers efficiently for a broad taxonomic sampling
be variable enough to distinguish species
but conserved enough within species
Choosing the correct DNA marker is critical.
Barcoding does not aim to solve phylogeny!
Reasons for DNA barcoding
1. Works with fragments.
2. Works with all stages of life: Can link male/females. Different stages of same organism. E.g. Amphipods (White & Reimer 2012)
3. Cryptic species detection. E.g. Astraptes
4. Reduces ambiguity (set DNA code).
5. Makes expertise go further.
6. Democratizes access to data. E.g. Barcode of Life project
7. Opens the way for handheld barcoders.
8. Finds new diversity.
9. Demonstrates value of museum collections. Sequencing of collections vital.
10. Speeds up discovery of new species.
Additional strong point
Does not need expert knowledge.
Weak points
1. DNA (specifically COI) does not always work for each group of organisms.
2. Handheld technology has not succeeded, despite many advances.
3. Different taxa have different DNA protocols, so standardization is difficult.
4. The “barcoding gap”
Barcoding implies that the level of DNA divergence between and within species is different.
But evolution not neat - hybrids, incomplete lineage sorting, etc.
This gap is not always present – so taxonomy comes back to “judgement”.

DNA barcoding proposed in 2003 as a “solution” to taxonomy.
Two large projects: Barcode of Life and Ocean Genome Legacy.
Encyclopedia of Life on the internet.
Common method of identification.
Gaining acceptance as a practical method to obtain much data.
But has not solved taxonomy, instead a new approach or “sub-field”.

For zoanthids (and corals), > 1 DNA marker is needed.
mt DNA evolves very slowly.
Still, better than no experts at all!

3. Promoting taxonomy: Census of Marine Life.

Scientific Framework
What has lived in the oceans?
What does live in the oceans?
What will live in the oceans?
The Census consisted of four major component programs organized around these questions.
1. Investigating the Past
Census researchers undertook the challenge of constructing the history of marine animal populations since human predation became important, roughly the last 500 years. This program component was called HMAP.
Teams of fisheries scientists, historians, economists and others conducted case studies in southern Africa, Australia, and approximately a dozen other regions.
Together, these case studies created the first reliable picture of life in the oceans before fishing.
The long historical records of marine populations help distinguish the contributions of natural fluctuations in the environment from the effects of human activities.
2. Assessing the Present
The largest component of the Census involved investigating what now lives in the world's oceans through 14 field projects.
Each sampled important kinds of biota in one of six realms of the global oceans using a range of technologies.
This included CReefs.
3. Forecasting the Future
To speak about what will live in the oceans required numerical modeling and simulation. This component program is the Future of Marine Animal Populations (FMAP).
Integrating data from many different sources and creating new statistical and analytical tools to predict marine populations and composition of ecosystems in the future
4. Living Legacy
Such a global initiative required a state-of-the-art data assimilation framework, and this is:Ocean Biogeographic Information System (OBIS).

2,700 scientists
80+ nations
540 expeditions
US$ 650 million
2,600+ scientific publications
6,000+ potential new species
30 million distribution records and counting

Example: CReefs:
Heron Island Trip
November 8th – December 1st, 2010
Census of Coral Reef Ecosystems (, part of Census of Marine Life (CoML –
Large international effort to understand biodiversity, use data for conservation.
Many researchers from different institutions, focused on different taxa, many “ignored”.
I focused on zoanthids (of course)!
Heron Island
Many seabirds nest here. Always noisy, and dangerous to walk without a hat!
Rails also live on the island.
The island is an important nesting site for green and loggerhead sea turtles.
Heron Island Research Station is run by the University of Queensland. Most of the station is very new.
There are excellent facilities for experiments, sample collection, and analyses, indoors and outside.
Scientists from all over the world, most based in Australia, but others from USA, Japan, Iceland, etc.
Most scientists had much field experience, and were good divers. Many also had boat licenses and first aid training.
There were also dive officers, who acted as boat captains, guides, diving assistants, etc.
Outreach included a professional photographer, and a professional blogger with stories and images posted every day.
A professional chef ensured everyone was well-fed. The food was amazing!
Every night at dinner, based on everyone’s ideas, weather conditions, and tides, the next day’s schedule was decided.
Boats were launched from the harbor, 3-10 people per boat. Often boats were out for over 7 hours.
Diving was always done under the buddy system, with very strict guidelines on diving protocol.
Special permits were obtained to allow specimen collection. Work underwater was intense and focused.
I was able to collect 270 specimens in 33 dives. Other groups collected up to >2000 specimens!
Samples were also collected on reef walks, and by snorkeling. Other groups used ARMS and the “carpet of death”.
Some groups found many new species, others new records for the southern GBR. This genus was found in the southern Pacific Ocean for the 1st time, likely a new species.
Another potential new species of zoanthid.
Many people also collected for other groups.
Back in the lab, data were collected, and specimens numbered. All data were given to the CReefs data manager as well.
All sites were assigned numbers, and all had GPS coordinates. 130 sites were visited in 3 weeks!
Samples will be shipped to institutions all over the world and analyzed further.
Of course, work was not 24 hours a day…
Sunset drinks were the one time of day when everyone would relax and take a break.
And the sunsets were amazing…

References cited:
1. CP Meyer, G Paulay. 2005. DNA barcoding: error rates based on comprehensive sampling. PLoS Biology 3(12) e422.
2. Consortium for Barcoding of Life homepage.
3. Census of Marine Life homepage.
4. OBIS homepage.

Wednesday, February 8th; 8:30-10:00.
10 questions, choose 7.
Open book, no cellphones, etc.
Must pass to pass class.
Arrive after 9 a.m. – out.
Advice: study!

December 21st, 2011 class

1. Introduction: What is a species?
2. Taxonomic species: Amphipods in the Ryukyus.
3. Biological species: Fish species complex!
4. OTU & barcodes: ARMS and estimating coral reef biodiversity.
5. Conclusions.
6. JCRS images.

1. Introduction: What is a species?
What is biodiversity? (Review)
Biodiversity = Number of taxa (species, genera).
OTUs = operational taxonomic units
Coral reefs are highly biodiverse, <0.2% of surface of Earth, >25% of marine species live there.
What is a species?
A species can be different, depending on who you talk to, or what meaning is implied.
Before we discuss species, we must be clear about what species concept we are discussing.
Three types are:
1. taxonomic species
A species with a scientific name.
Names are important for science & conservation.
Accuracy and authorship important.
Naming species takes time & money.
Few experts.

2. Biological species
A species proven to reproduce only with itself.
Reproductive isolation.
Species concept important.
Experiments take time & money.
Hybrids? Evolution is messy.

3. OTU/barcode species
A species defined by genetic differences.
Implies a threshold value.
Quick and easy.
Sequencing species takes time & money.
Limited by previous knowledge.
Not always “true”, barcoding gap is problematic.

It is estimated we have described 1~25% of the total marine species.
Description of all species – possible?
However, describing is important.
1. Unknown species - What kind of undescribed species exist?
We have no idea what awaits discovery.
2. “Hidden” species
Hidden species may be hidden due to their cryptic nature, or due to researchers not being able to distinguish them from other, similar species. Usually small.
Often, there are large groups of “species complexes” that are hard to make sense of.

C. Known but not described species
We know this is a zoanthid. DNA has shown us how it is related to other zoanthids (=very unique).
We also know it is undescribed, and only on Okinawa.
It is waiting to be described.

Part 2: Taxonomic species
Commensal amphipods in Okinawa
White & Reimer 2011-2012
Taxonomic species
Can be detected through observation of morphology or classic diagnostic characteristics. DNA?
Compare with previous taxonomic literature.
Only way to name a species.

Leucothoidae (139 sp.) – one family of amphipods
Anamixid clade (pronounced sexual dimorphism):
• 22 Anamixis Stebbing, 1897
• 4 Nepanamixis Thomas, 1997a
• 13 Paranamixis Schellenberg, 1938
Leucothoid clade
(minimal to moderate sexual dimorphism):
• 98 Leucothoe Leach, 1814a
• 2 Paraleucothoe Stebbing, 1899
Ecologically unique
• Endocommensal associates of sponges, ascidians, and bivalve mollusks
• Extended parental care
• Social system
• Potential eusociality

These are completely unexamined in Okinawa!
Contain two morphotypes, leucomorphs and anamorphs, previously thought to be different families of amphipods! DNA barcoding confirmed same species.
Materials and Methods
• Collect in-situ directly from host, take whole host, or coral rubble
• Traditional taxonomy
• Digital Inking
• Molecular DNA sequencing
25 new species from Okinawa in 1.5 years. More diverse in Okinawa than the GBR.

Part 3: Biological species
Fish species complex in the Pacific (Leray et al. 2010)
Biological species
Can be detected through observation of reproduction.
Also, reproductive experiments.
But these require extensive field work. Recently, molecular analyses usual, using phylogeny.
Background – Genus Dascyllus: Complex has 4 species:
1. D. trimaculatus (E. Africa to Central Pacific): 3 spots
2. D. albisella (Hawaii): white flanks
3. D. strasburgi (Marquesas Islands): gray
4. D. auripinnis (Line, Phoenix Islands): yellow fins
Each species has its own ecology and preferred habitats.
Materials and Methods
Specimens (n=563) from across the Indo-Pacific.
Microsatellite DNA data examined (data from previous research included).
Phylogenetic analyses and genotype assignment.
Previous research showed 5 clades, not 4 (Bernadi et al. 2003).
This research showed 7 (!) groups of Dascyllus, not all with clear morphology.
Appears ancient history of Indo-Pacific combined with recent evolutionary events drives speciation.
Hybridization appears to be occuring.
NOTE: unknown species are not yet valid species, or new species. They are undescribed species.

So…. Biological species
Results could lead to “finding” undescribed species or groups
This leads to…
Taxonomic species
Describing a species.
Only way to give a scientific name.
New species.

Part 4: OTU/barcode species
ARMS and crustaceans
Plaisance et al. 2011;
Brainard et al. 2010
Coral reef monitoring of other species asides from fish/corals.
Establish systematic and consistent metric to assess and monitor change.
ARMS – manmade deployment devices to look at crustaceans/molluscs, etc., simulate dead coral heads.
Easy to build, 200USD each.
Rugged, look like plastic boxes/crates.
Prototypes tested in Northwestern Hawai’i. Seemed to work well.
Deployed worldwide, and retrieved. Then processed, samples preserved, DNA sequencing (DNA barcoding).

So…. OTU/barcode species
Results could lead to “finding” undescribed species or groups
This leads to…
Biological species
Results could lead to “finding” undescribed species or groups
This leads to…
Taxonomic species
Describing a species.
Only way to give a scientific name.
New species.

References cited:
1. CP Meyer, G Paulay. 2005. DNA barcoding: error rates based on comprehensive sampling. PLoS Biology 3(12) e422.
2. KN White, JD Reimer – three papers (2011-2012). Commensal Leucothoidae in the Ryukyu Archipelago, Japan. ZooKeys (in press).
3. M Leray, R Beldade, SJ Holbrook, RJ Schmitt, S Planes, G Bernadi. 2010. Allopatric divergence and speciation in coral reef fish: the three spot dascyllus, Dascyllus trimaculatus, species complex. Evolution 64-5: 1218-1230.
4. L Plaisance, MJ Caley, R Brainard, N Knowlton. 2011. The diversity of coral reefs: what are we missing? PLoS One vol 6 issue 10 e25026.
5. R. Brainard et al. 2009. Autonomous Reef Monitoring Structures (ARMS): a tool for monitoring indices of biodiversity in the Pacific Islands. Pacific Science InterCongress, March 4, 2009.

Part 6: Japan Coral Reef Society 14th Annual Symposium
Images from the conference.