ASCIDIAN NEWS*
Gretchen
Lambert
12001 11th
Ave. NW, Seattle, WA 98177
206-365-3734 gretchen.lambert00@gmail.com
home page:
http://depts.washington.edu/ascidian/
Number
73 June 2014
I taught an
Ascidian Taxonomy Workshop with Noa Shenkar March 17-21 at the Inter-University
Institute in Eilat, Red Sea coast of Israel, followed by a few days working on
some of the samples in Tel Aviv University’s museum collection. I greatly
enjoyed learning about the interesting projects Noa’s students are working on,
snorkeling in the Red Sea, and sightseeing around Tel Aviv and southern Israel,
and I am very appreciative that my trip was supported by the Israel Taxonomy
Initiative.
I am now looking
forward to teaching a two week workshop with Rosana Rocha June 17-July 2 at the
Smithsonian Tropical Research Institute (STRI) at Bocas del Toro, Panama. This
will be the 4th ascidian workshop at Bocas; the first was in 2006. It is very
gratifying to see that many of today’s biologists working on ascidians have
taken our workshops over the years. In July Susanna López-Legentil (Univ. of N. Carolina) and
I will conduct an ascidian survey of many marinas and some aquaculture sites
along the North Carolina coast, a region that has not been surveyed for
ascidians for many decades. We will report our findings at the next Invasive
Sea Squirt meeting (see below).
There are 79
new publications listed at the end of this newsletter.
*Ascidian News is not part of the scientific
literature and should not be cited as such.
1. From Mary Carman: The next Intl. Invasive Sea Squirt Conference
(IISSC5) will be held at Woods Hole, Massachusetts Oct 29-31, 2014. Please save the date! Submit abstracts to: mcarman@whoi.edu. Registration at https://www.whoi.edu/main/sea-squirt-conference-v
.
2. Congratulations
to Mari
Carmen Pineda, whose
Ph.D. thesis entitled “A global wanderer: Biology, phylogeography and
resilience of the introduced ascidian Styela
plicata”, defended September 25, 2012 and advised by Xavier Turon and
Susanna Lopez-Legentil, was recently awarded a Doctoral Extraordinary award by
the University of Barcelona. (http://www.ub.edu/irbio/ENnoticia.php?id=222)
3. Charles Lambert’s last publication has finally appeared! Lambert, C. C. 2014. Obtaining gametes
and embryos of ascidians. In: Carroll, D. J. and Stricker, S. A. (eds.),
Developmental Biology of the Sea Urchin and Other Marine Invertebrates. Humana
Press, pp. 27-33. Charley finished the final revisions for this chapter before
he died 3 years ago, and was very pleased with it. He was a whiz at getting
gametes to fertilize and develop, so if you are having methodological problems
obtaining gametes or raising embryos from either solitary or colonial species,
this chapter will be very useful.
4. From Noa Shenkar
(Zoology Dept., Tel Aviv University, Israel): In March 2014, an Ascidian Taxonomy workshop was held for the first
time at the Inter-University Institute in Eilat, Israel, taught by myself and
Gretchen Lambert. 16 participants successfully learned general aspects of
ascidian biology and ecology, and how to identify the local species. My new
laboratory at Tel-Aviv University is continuing to study non-indigenous
ascidians in the Mediterranean and biodiversity of ascidians in tropical
regions.
The WoRMS Ascidiacea World Database is
being updated and revised constantly. http://www.marinespecies.org/ Please e-mail PDF files of new species
publications to noa.shenkar@gmail.com. Any comments and
corrections to the database are greatly appreciated.
5. The 8th Intl. Tunicata meeting will be held
on July 13-17, 2015 in Aomori city, Japan. Aomori city is located at the
northern end of the Japan main island, being surrounded by well-known scenic
places such as the Shirakami Mountains World Heritage Site and Lake Towada.
July in Aomori is a pleasant season as the temperature averages approximately
22o C. The organizers are Kazuo
Inaba, Director, Shimoda Marine Research Center, University of Tsukuba, and
Gaku Kumano, Asamushi Research Center for Marine
Biology, Tohoku University, Asamushi. kumano@m.tohoku.ac.jp
6. It is with great sadness that we report the death of Dr. Ivan Goodbody on April 16, 2014, at
his home near Aberdeen, Scotland. Ivan worked for many decades on ascidian
physiology and taxonomy and produced many important publications, at least 36
of which are either specifically on ascidians or including ascidians. His
lengthy 1974 review of the physiology of ascidians still stands as the standard
reference, as are his numerous taxonomic papers of Caribbean ascidians. Ivan
and his wife Charlotte worked closely together for many years. They produced a
CD of color photos of many Caribbean ascidians in 2006.
I am very grateful to Charlotte and her
family for contributing the following biography and tribute:
Ivan Goodbody was born and raised in Dublin,
Ireland where he developed an interest in Natural History at an early age. His interest in Ascidians began during his
undergraduate studies at Trinity College Dublin, thanks to a challenging exam
question on the life cycle of Botrylloides. He received a First Class Honours degree in
Zoology and spent a year at Oxford University studying Ornithology (a lifelong
passion). He then moved to the
University of Aberdeen, Scotland where he completed a
Ph.D. on the Biochemistry and Physiology of Nitrogen Excretion in
Ascidians. In Aberdeen he met and
married Charlotte Fraser, with whom he celebrated 60 years of marriage in September,
2013.
In 1955 Ivan and Charlotte moved to Jamaica,
where he took up an appointment of Lecturer in Zoology at the University
College of the West Indies (later University of the West Indies). In 1962 he was appointed Senior Lecturer and
became Professor and Head of department in 1964.
When he moved to Jamaica Ivan’s research
involved the taxonomy and the ecology of ascidians. In his early research in
Jamaica he demonstrated the phenomenon of continuous breeding in certain
ascidians and also discovered mass mortality in tropical marine invertebrates
when exposed to fresh water runoff from heavy tropical rains. His scientific publications drew attention
to the diversity of ascidians in the Caribbean, putting him in touch with
ascidian researchers elsewhere in the world, notably Patricia Mather in
Brisbane, Robin Millar at Oban in Scotland and Donald Abbott in Monterey. Furthermore, his many publications on
tropical marine invertebrates and the publication of his Ph.D. thesis on the
metabolism of nitrogen in ascidians drew attention to his research leading to
an invitation from Sir Frederick Russell to contribute a review on ascidian
physiology to Advances in Marine Biology.
This review has stood the test of time and continues to be widely cited.
His research on ascidians was further
enhanced when he was invited to undertake a study of ascidians in Belize in
Central America, as part of a major tropical marine research program set up by
the United States National Museum of Natural History. The programme's field research laboratory was
based in the central sector of the Meso-American Barrier Reef at Carrie Bow
Cay, Belize. In the fifteen years during
which he participated in this program, he identified 70species from the Pelican
Cays and 40 from Twin Cays and documented his findings in 8 scientific
papers. He regarded this work on
ascidians as the culmination of his career as an ascidiologist.
While ascidian biology remained his primary
research interest throughout his scientific career, Ivan put great energy and drive into building applied research and educational
programs at UWI. Along with other
colleagues in the Zoology Department, he was instrumental in the founding of
the Port Royal marine lab in the 1950s which continues to operate as a key
research and teaching facility in marine biology. Under his leadership, research programs into
pollution in Kingston Harbour, aquaculture and fisheries all flourished at
UWI. He successfully promoted and managed
multi-disciplinary research, believing such collaborations as key to addressing
complex environmental questions.
Throughout his career he was a tireless advocate for conservation and
habitat preservation. He served on many
national committees and received numerous awards in Jamaica in recognition of
his many and diverse contributions to research, conservation and
education.
After living a in
Jamaica for 52 years, Ivan and Charlotte returned to Aberdeen where Ivan became
Honorary Research Fellow in the School of Biological Sciences at the University
of Aberdeen. His passion for ascidian
biology never waned, and he continued writing up his research from his new home
base.
Ivan’s career and contributions to the
University of the West Indies (UWI) are captured in this excellent tribute from
his former colleagues in Jamaica:
http://myspot.mona.uwi.edu/marcom/newsroom/entry/5671
WORK
IN PROGRESS
1. Introduced ascidians in
South and Northern Chile.
Xavier Turon. Centre for Advanced Studies
of Blanes (CEAB), Spanish Research Council.
xturon@ceab.csic.es
I had the opportunity to spend a couple of
months in 2013 in Chile, in the framework of a biodiversity project led by
Prof. Ivan Cañete (Univ. of Magallanes, Punta Arenas), with the participation
of Prof. Javier Sellanes (Universidad Católica del Norte, Coquimbo). Our goal
was to assess ascidian diversity in these two areas, with particular emphasis
on introduced ascidians. Punta Arenas is a hub for shipping activities through
the Magallanes Strait, and the Coquimbo region hosts important aquaculture
facilities. We surveyed ports and artificial structures,
scuba dove (an unforgettable experience in the freezing waters of Patagonia)
and took video footages from the surface. I was very glad to coincide with
Rosana Rocha (Univ. Federale do Paraná) in Punta Arenas and share with her the
morphological study. The samples are now being genetically barcoded by Susanna
López-Legentil (Univ. of N. Carolina). The results so far point to a relatively
poor ascidian fauna (ca. 25 species), but clearly differentiated between the
two regions surveyed. No species was found at both latitudes. In addition,
while there was a high prevalence of introduced ascidians in the Coquimbo
region, no invasive ascidian was found in the Punta Arenas region, which was
indeed a surprising result given our exhaustive sampling of ports and
artificial structures.
2. From Patrick Frank, Dept. of Chemistry,
Stanford Univ. : pfrank@slac.stanford.edu
We have extended
our XAS studies of sulfur in the blood cells of Ascidia ceratodes to the Bodega
Bay population. Free sulfate dominates total sulfur (~50%) but alkyl sulfate
esters are again richly present. Biological thiol - disulfide fractions are
similar to those recently reported for the A. ceratodes population in Monterey
Bay, and are likewise consistent with the participation of thiol in the
reduction of blood cell vanadium. A suite of other sulfur secondary metabolites
included functional groups similar to benzothiophene and thianthrene. Blood
plasma also, surprisingly, included a complex mixture of sulfur compounds. The
study includes single animals, which display much wider variations in
signet-ring vacuolar pH than the previously examined aggregated samples. I'll
be reporting these results in more detail at a symposium talk for Ken Kustin at
the August ACS meeting in San Francisco.
3. From Benedikte Vercaemer, Dawn Sephton and Claudio DiBacco Fisheries & Oceans
Canada, Dartmouth, NS, Canada. Benedikte.Vercaemer@dfo-mpo.gc.ca; Dawn.Sephton@dfo-mpo.gc.ca; Claudio.DiBacco@dfo-mpo.gc.ca
A suspicious marine tunicate (Didemnidae) specimen collected in
October 2013 by a recreational diver off the coast of Parrsboro, north of Minas
Basin, Nova Scotia (NS) was brought to the Bedford Institute of Oceanography
(BIO) for identification. The tunicate was sampled from a depth of about 10 m,
distributed over an area of about 100 sq. ft. (i.e., the size of a room) and
attached to small rocks representative of hard substrate in the area. Taxonomic
(BIO) and molecular identification (Dr. Sarah Clark, Dalhousie University)
confirmed the sample as the invasive tunicate Didemnum vexillum, which represents the first detection of this
species in Eastern Canada. Annual rapid assessment surveys conducted in
Passamaquoddy Bay, New Brunswick and areas near Eastport, Maine since 2009 as
well as southwestern NS (2013) did not detect this species. Didemnum species have never been
detected along the Bay of Fundy or in nearshore coastal habitats of Nova Scotia
despite DFO biofouling monitoring initiated in 2006.
There is considerable suitable habitat for D. vexillum throughout the Maritimes and given its known establishment in Eastport, Maine since 2003, it is likely that other regions of the Bay of Fundy are also affected. The DFO Aquatic Invasive Species group (BIO) completed a rapid assessment survey in early April 2014 to confirm the presence of D. vexillum in the Minas Basin, upper Bay of Fundy, and to collect samples for taxonomic confirmation. Molecular screening of 41 tissue samples collected from 23 stations sampled (10-30 m depth) in the Minas Basin and Minas Channel were confirmed to be D. vexillum. DFO recently initiated a bay-wide assessment to delineate and map the distribution of D. vexillum throughout the Bay of Fundy as well as regions of the southern Scotian Shelf, including Georges Bank, German Bank and Browns Bank.
4. Investigations into the reproductive
biology of non-native species of ascidians in UK waters. Anaëlle
Lemasson, Master’s
project. Supervisors: John D.D. Bishop (MBA); Andy Foggo (Plymouth
University). analem@mba.ac.uk
Asterocarpa humilis, the compass ascidian, and Corella eumyota, the orange-tipped ascidian,
are two non-native species recently identified on UK shores. They are solitary
brooders originating from the southern hemisphere. Since their first record in
the Northern Hemisphere, in France during the 2000s, they have rapidly spread
through the coasts of France and the UK. As part of my Master’s thesis,
undertaken with Plymouth University (UK) and the Marine Biological Association,
I am studying various aspects of their reproductive biology in UK waters. The
project holds three sub-studies: the brooding seasonality of A. humilis, the seasonal variation in
the fecundity potential of A. humilis
and C. eumyota, and larval dispersal
and settlement preferences of A. humilis,
C. eumyota and seven other species of
ascidian commonly found on UK shores.
A. humilis broods
larvae throughout the year in UK waters, with a possible peak in winter. Early
results for the seasonal variation in fecundity indicate the ability of both
species to release thousands of larvae at a time, with an apparent decrease in
fecundity from February to April for both species. Future work will also
compare their reproductive output with ones of typical solitary broadcast
spawners in order to determine whether their brooding strategy is costly in
terms of fecundity. The larval dispersal and settlement experiment, not
completed yet, will provide crucial information regarding the species’
dispersal potentials and settlement substrate choices, which will lead to a
better understanding the type of environment they are likely to colonize in the
field.
5. From Rosana
Rocha, Univ. Federal do Parana, Curitiba, Brasil. rmrocha@ufpr.br
I have two new doctoral candidates working
on ascidians, and a summary of their research projects follows:
Rodolfo Corrêa de Barros: Population genetics and phylogeography of Styela
canopus Savigny, 1816.
Styela canopus is a small and solitary ascidian with high external morphological
variability. It’s a fouling animal found on artificial substratum in harbors
and disturbed environments. Styela canopus is
registered in several places around the world and in Brazil it is found from
Ceara to Santa Catarina. The main goal of this project is to understand the
geographical distribution of S. canopus through
the study of genetic relationships among populations and investigate patterns
of dispersal and invasion. I will use COI and ITS1 molecular markers. So, I
will need help from researchers to collect samples of S. canopus
from distinct populations around the world. If you can help, please contact me:
rodolfo@ufpr.br. Since early detection is an important tool
in controlling bioinvasions, this project will also aim to establish protocols
for molecular identification of S. canopus, S. plicata
and C. intestinalis in samples of seawater.
Livia de Moura Oliveira:
Morphology and systematics of the genera Didemnum
Savigny, 1816 and Polysyncraton Nott,
1892.
Didemnum Sluiter, 1816 is the most diverse genus
in the Family Didemnidae with about 230 species. Polysyncraton Nott,
1892 is the second largest genus in the family with 97 known species, and both
genera have similar zooids. This study intends to test the monophyly of these
two genera and to clarify the phylogenetic relationship between them, using
morphology and molecular data, including secondary structure models. Many
samples will come from the Brazilian coast, where we already know that there
are many new species to be described. But I would also like to include species
from different geographical origins to have a better representation of both
genera. If you can help, please contact me: lihmoura.oliveira@yahoo.com.br
6. Guidelines for the nomenclature of genetic elements
in tunicate genomes.
A. Stolfi, Y. Sasakura, D. Chalopin,
Y. Satou, L. Christiaen, C. Dantec, T. Endo, M. Naville, H. Nishida, B. J.
Swalla, J.-N. Volff, A. Voskoboynik, D. Dauga, and P.
Lemaire. patrick.lemaire@crbm.cnrs.fr [A manuscript recently submitted for a
special issue of Genesis on tunicates. I thank all the authors for permission
to include it in AN. And a reminder to all readers of AN: as stated on the
first page of every newsletter, Ascidian News is not part of the scientific
literature and should not be cited as such.]
Tunicates are invertebrate members of the chordate phylum, and are
considered to be the sister group of vertebrates. Tunicates are composed of
ascidians, thaliaceans and appendicularians. With the advent of inexpensive
high throughput sequencing, the number of sequenced tunicate genomes is
expected to rise sharply within the coming years. To facilitate comparative
genomics within the tunicates, and between tunicates and vertebrates,
standardized rules for the nomenclature of tunicate genetic elements need to be
established. Here we propose a set of nomenclature rules, consensual within the
community, for predicted genes, pseudogenes, transcripts, operons, transcriptional
cis-regulatory regions, transposable elements, and transgenic
constructs. In addition, the document proposes guidelines for naming transgenic
and mutant lines.
7. Chemical
and pharmacological activities of a simple ascidian.
C.Stella Packiam,
Assistant Professor of Chemistry under
the guidance of Dr. R. Jothibai Margret,
Dept. of Chemistry, Pope’s College, Sawyerpuram, Tirunelveli and coguidance of
Dr. V.K. Meenakshi, Dept. of Zoology, A.P.C.Mahalaxmi College for Women,
Tuticorin, Tamilnadu, India.
GC-MS analysis, IR Spectral studies and
spectrophotometric studies are in progress in a simple ascidian Ascidia sydneiensis. Ascidians have
tremendous potential in pharmaceutical and biomedical field. Ascidia
sydneiensis is commonly called as crevice ascidian. It has been subjected
to Infrared (IR) spectral study which indicates the presence of aromaticity,
hydroxyl and carbonyl group in the ethanolic extract. GC-MS studies revealed
the presence of ten chemical constituents -Tetradecanoic acid,
Bis-(2-methylpropyl)ester of 1,2-benzenedicarboxylic acid, n-Hexadecanoic acid,
9-Hexyl-heptadecane, Diisooctyl ester of 1,2- benzenedicarboxylic acid,
3-ethyl-5-(2-ethylbutyl)-octadecane, Squalene, Cholest-5-en-3-ol(3∝)-carbonochloridate, Cholesterol,
2-[[2-[(2-ethylcyclopropyl)methyl]cyclopropyl]methyl]-methylester of
cyclopropaneoctanoic acid.
Infra red spectrum for ethanolic extract
of Ascidia sydneiensis (Figure-1) shows broad band at 3416.97 cm-1
which is due to the presence of moisture or hydroxyl groups in the
compound and that at 2930.39 cm-1 is characteristic for C-H
stretching vibration indicating aliphatic chain. The band at 2350.80 cm-1
for O-H stretching gives evidence for the presence of carboxylic acid
and the band at 1620.06 cm-1 for C=O stretching vibration indicates
carbonyl group. The strong bands above 3000 cm-1
shows evidence for aromatic rings. Estimation of the total carbohydrates,
proteins, lipids, phenols and flavonoids from Ascidia sydneiensis were
analysed by Spectrophotometer. The results showed that the specimen was
rich in lipids (25.33%) followed by flavonoids (19.01%), proteins (10.61%),
phenols (10.09%) and carbohydrates (5.58%). Ascidia sydneiensis possess
higher percentage of lipid and lower carbohydrate. Following observations
suggest that this commonly available ascidian has active biochemical potential
possessing antioxidant, anti-inflammatory and antimicrobial properties for
curing various ailments which can lead to the isolation of new and novel
compounds.
8. Chemical
Investigation and anticancer activities of selected
tunicates. S. Sankaravadivu, Assistant Professor of Chemistry, under the guidance of Dr.
R. Jothibai Margret, Dep.t of Chemistry, Pope’s College, Sawyerpuram,
Tirunelveli and coguidance of Dr. V.K. Meenakshi, Dept. of Zoology,
A.P.C.Mahalaxmi College for Women, Tuticorin, Tamilnadu, India.
GC-MS analysis, IR
Spectral studies and spectrophotometric studies are in progress in a colonial
ascidian Ecteinascidia venui. GC-MS analysis
of the ethanolic extract of the animal revealed the presence of twelve
components. The
peak with retention 10.38 corresponds to 1-(2-Ethyl-3-cyclohexenyl)ethanol,
14.26 to (E,E)-methyl ester of 9,12-Octadecadienoic acid, 14.34 to (E)-methyl
ester of 9-Dodecenoic acid, 16.96 to 9-Octadecenal, 18.71 to
(Z)6,(Z)9-Pentadecadien-1-ol, 19.39 to (Z,Z)-9,12-Octadecadien-1-ol, 19.46 to
13-methyl-Oxacyclotetradecane-2,11-dione, 19.73 to Eicosane, 21.13 to
Tetradecyloxirane, 22.14 to (R)-(-)-14-Methyl-8-hexadecyn-1-ol, 22.52 to
Nonadecane and 23.90 to
1-Iodo-2-methylundecane. These chemical components have not been
reported elsewhere from ascidians specially Ecteinascidia
venui and hence can be considered as a first report. Infrared spectrum for ethanolic extract of Ecteinascidia venui indicated a broad
band at 3409.83 cm-1 due to the presence of moisture or hydroxyl
group in the compound. The band at 1632.09 cm-1 is characteristic of
carbonyl group and that at 1412.10 cm-1 shows the presence of C-H
bending. Presence of peaks above 3000 cm-1 indicates aromaticity and
C-H stretching. Analysis of
carbohydrates, proteins, lipids, phenols and flavonoids were determined by
Spectrophotometry. A maximum of 45.82% flavonoids, 20.68% proteins, 9.77%
lipids, 9.21% phenols and 4.57% carbohydrates were observed in Ecteinascidia venui. These studies
clearly indicate that Ecteinascidia venui
is rich in many biochemical compounds.
1. Benthic
Ecology Meeting, Jacksonville Florida, March 19-22, 2014
Unraveling
the diversity of the ascidian microbiome. López-Legentil, S.1;
Pineda, M.C.2; Webster, N.2; Turon, X.3;
Erwin, P.M.1 1Department of Biology & Marine Biology, and
Center for Marine Science. 5600 Marvin K. Moss Ln, Wilmington NC 28409, USA. lopezlegentils@uncw.edu; 2Australian Institute of
Marine Science, Townsville Mail Centre, Qld 4810, Australia; 3Center
for Advanced Studies of Blanes (CEAB-CSIC), Accés Cala S. Francesc 14, 17300
Blanes, Girona, Spain.
Symbiotic microbes are known to play a
critical role in host metabolism and in nutrient cycles; however, the ascidian
microbiome remains largely unexplored. Here, we provide the most comprehensive
characterization to date of the ascidian microbiome by investigating the
diversity, structure and specificity of microbial symbionts in 42 Great Barrier
Reef ascidians using 16S rRNA gene tag pyrosequencing. Results revealed high
bacterial biodiversity (3,217 OTU0.03) and the widespread occurrence
of ammonia-oxidizing Archaea. The ascidian microbiota was clearly
differentiated from free-living seawater microbial communities and included
symbiont lineages shared with other invertebrate hosts as well as unique,
ascidian-specific phylotypes. Several rare seawater microbes were markedly
enriched in the ascidian microbiota, suggesting that the rare biosphere of
seawater may act as a conduit for horizontal symbiont transfer among hosts.
However, most OTUs (71.2%) were rare and specific to a single host while core
communities were comprised of only 7 OTUs, indicating an overall high degree of
host-specificity. We hypothesize that the complex ascidian microbiota revealed
herein is maintained by a dynamic tunic microenvironment, offering periodic
windows of optimal conditions for different metabolic pathways. For example,
ample chemical substrate (ammonia-rich host waste) and physical habitat (high
oxygen, low irradiance) that support nitrification processes.
2. XV Meeting of the Italian Association
of Developmental and Comparative Immunobiology (IADCI), Univ. of Ferrara,
Ferrara, February 12-14, 2014.
a. The production of amyloid requires
cross-talk between immunocytes in the compound ascidian Botryllus
schlosseri. L. Ballarin1, R. Girardello2, A. Grimaldi2, N.
Franchi1,
M. De Eguileor2 1Dept .of Biology, Univ. of Padua, Padua,
Italy; 2Dept.
of Biotechnology and Life Sci., Univ. of Insubria, Varese, Italy ballarin@bio.unipd.it
Two main immunocyte types are present in the
hemolymph of the compound ascidian Botryllus schlosseri, i.e.,
phagocytes and cytotoxic morula cells (MCs). Previous studies have demonstrated
that MCs work as sentinel cells able to sense foreign molecules and, as a
consequence, release cytokines which activate phagocytes leading to the
synthesis and release of rhamnose-binding lectin. The latter, in turn, acts as
a chemotactic factor for phagocytes and opsonizes foreign particles stimulating
their clearance. In addition, upon the contact with foreign molecules, MCs can
degranulate and release the content of their vacuoles, mainly phenoloxidase
(PO) and its polyphenol substrata. In a recent investigation on Botryllus cytotoxic
cells, we found abundance of amyloid inside MC vacuoles which likely, once
released, act as a scaffold to prevent the diffusion of PO and cytotoxicity to
the whole organism. In addition, the study of the molecular cascade leading to
the synthesis of amyloid, is revealing a non-classical
pathway in which both the phagocytes and MC are involved.
b.
Phagocytosis-induced apoptosis in the compound ascidian Botryllus
schlosseri. F. Schiavon, N. Franchi,
L. Ballarin, Dept. of Biology, Univ. of Padua, Padua, Italy.
Colonies of the ascidian Botryllus
schlosseri contain three blastogenetic generations: functional zooids,
their buds and budlets on buds. Generation change or
take-over (TO) occur cyclically and assure the recurrent renewal of the colony.
During these events, lasting 24 - 36 h, diffuse apoptosis occurs in zooid
tissues, as indicated by morphological, biochemical and molecular
investigations. Tissues are rapidly infiltrated by circulating phagocytes,
selectively recruited by dying cells, which recognize and greedily ingest them.
Using hemocytes as selected cell population for investigation, we studied the
transcription rate of three recently characterized genes involved in apoptosis,
bsbax, bsaif1 and bsparp1, during the TO as compared to
colony developmental phases far from it. In addition, we observed that the
massive ingestion leads phagocytes themselves to undergo apoptosis, probably as
a consequence of the oxidative stress related to the sustained respiratory burst,
as suggested by biochemical analysis. Therefore, a large fraction of
circulating phagocytes needs to be replaced by new, young hemocytes, entering
the circulation at the end of the generation change.
c. New evidences of conserved pathways in
complement system dynamics from the colonial ascidian Botryllus schlosseri. N.
Franchi, L. Ballarin, Dept. of Biology, Univ. of
Padua, Padua, Italy.
In recent years, it has been widely
demonstrated that complement, although often depicted as a ‘first line of defense’,
is more than just a defender against microbial intruders and acts as a tightly
integrated surveillance system. It is not only important against
microorganisms, but also for the clearance of apoptotic cells and corpses. Botryllus
schlosseri belongs, as vertebrates, to the phylum Chordata and, for this
phylogenetic trait, it is unanimously considered a
reliable model organism for the studies of the evolution of the immune system.
Moreover it is also characterized by a peculiar life cycle with a cyclical,
massive apoptosis. This two key features render B.
schlosseri a good research tool for the study of the evolution of the
complement system. Here we report the first results on the expression of BsC3
and BsFactorB, both components of the alternative pathway (AP) of complement
activation, which form the AP C3 convertase.
Since studies on mammalian models have
shown that 80 % of the observed complement response is derived from AP
convertase-mediated C3 amplification, even if initially induced by the
classical pathway (CP), studies of complement activation in an organism that
lack the adaptive immunity, as B. schlosseri, could lead to a better
comprehension of the AP cascade and the behavior of C3 convertase not only in
invertebrates, but also in vertebrates, mammals included. As in mammals, BsC3
is highly transcribed at basal level and over-expressed after incubation with
non self (zymosan) while BsFactorB always shows limited expression. In the
presence of compstatin, a 13-residue cyclic peptide able to inhibit the
activation of C3 by C3 convertases, the percentage of phagocytosing hemocytes
collapses. In the presence of both zymosan and compstatin, the transcription of
BsC3 by hemocytes increases with respect to cells exposed only to zymosan: this
suggests the presence of a conserved molecular
machinery able to control and modulate B. schlosseri as well as the
mammalian complement.
d.
Evidences for antimicrobial peptides in the colonial ascidian Botryllus
schlosseri. F. Schiavon, N. Franchi, L. Ballarin. Dept. of Biology, Univ. of Padua, Padua, Italy
Invertebrates have acquired many
mechanisms of defence in order to overwhelm the risk of pathogen attack. In
particular, the presence of various types of antimicrobial peptides (AMP)
guarantees an efficient response, making them able to kill both
gram positive and negative bacteria, fungi and viruses. In ascidians,
AMPs have been isolated and described in different species, such as in Styela
clava (Clavanins and Styelins) and in Ciona intestinalis (the recent
Ci-MAM). Previous studies have revealed molecules, including phenoloxidase and
rhamnose-binding lectins, with an antimicrobial effect also in Botryllus
schlosseri. Here, we report that the growth of some bacteria strains is
highly inhibited by extracts of haemocytes, showing alterations in their
surface. Moreover, molecular analyses allowed us to identify a Botryllus sequence
similar to Styelins that is abundantly transcribed in phagocytes.
e.
Characterization of proxiredoxins’ coding genes in Ciona intestinalis. R.
Benevenia1,
D. Ferro2,
L. Ballarin1,
G. Santovito1. 1Dept. of Biology, Univ. of Padua, Padua, Italy; 2Institute
for Evolution and Biodiversity, Westfälische Wilhelms-Universität, Münster,
Germany.
Ascidians represent an interesting model from
an evolutionary and ecotoxicology point of view, because of their large
distribution in temperate sea and their phylogenetic position of invertebrate
chordates. Immune responses imply an increase in oxygen consumption with a
consequent risk of oxidative stress. With the aim to study the components of
the antioxidant defense system in the solitary ascidian Ciona intestinalis,
we characterized gene sequences encoding peroxiredoxins (Prxs), non-selenium
peroxidases that are able to reduce hydrogen peroxide, organic hydroperoxides
and peroxynitrite. Thus they represent a class of important antioxidant enzymes, that protect cells against oxidative stress. In the
GeneBank database five Prx sequences are present, Prx2, 3, 4, 6a and 6b. The
multi-alignment analysis, conducted with orthologous sequences of vertebrates
and invertebrates, showed that in Ciona’s Prxs the amino acids essential
for their enzymatic activity are highly conserved, namely the catalytic tetrad
consisting of proline, threonine, cysteine and arginine. Preliminary
phylogenetic reconstruction indicates that Prx3 and 4 emerge as sister group of
Prxs of the respective vertebrates groups (or isoforms), while Prx2, 6a and 6b
have an uncertain position. A partial confirmation of phylogenetic results was
obtained with the analysis of homology modeling, according to which Prx3 and 4
present a structure similar to that of two vertebrate proteins, Bos taurus Prx3
and Larimichthys crocea Prx4, respectively, while Prx2 and 6 six have a
three dimensional structure similar to that of two invertebrate proteins, Ancylostoma
ceylanicum Prx1 and Arenicola marina Prx6, respectively. The
transcription of all these genes, measured by qRT-PCR, resulted variable in
different organs (intestine, ovary, pharynx, stomach). In particular, the ovary
is the organ that expresses the highest level of messenger for all Prxs.
Preliminary data were also collected about the possible circadian expression of
Prx.
3. The 9th Intl. Vanadium Symposium will be held at Padova, Italy, from
June 29- July 3. The topics will cover coordination chemistry, speciation,
biological chemistry, toxicology, enzymology, catalysis, therapeutic
applications, batteries, novel V-containing materials, and environmental/occupational
exposure to V agents. Dr. Tatsuya Ueki will give an invited presentation on the
accumulation of vanadium in ascidians. http://www.chimica.unipd.it/V9/
.
THESIS ABSTRACTS
1. Studies on the antiproliferative activity of a chosen ascidian of
Tuticorin coast. Ph.D. thesis by M. Paripooranaselvi,
Assistant Professor, Dept. of Zoology, A.P.C. Mahalaxmi College for Women, Tuticorin, under the guidance of Dr. V.K. Meenakshi. Submitted to
Manonmaniam Sundaranar University, Tirunelveli, Tamil
Nadu, India. mparipooranaselvi@gmail.com
Phallusia
nigra are common
persistent biofoulants found mostly attached to cement blocks, rocks, pilings
and oyster cages in Tuticorin harbour area. The thesis consists of two parts. Part I Biofouling deals with an
assessment of the species
composition, distribution and seasonal variation in the occurrence of
ascidian biofoulers at three different stations - Tuticorin Green Gate Barge,
Harbor installations and Tuticorin Green Gate intertidal rocky shores. 50 species of ascidians belonging to
20 genera coming under 9 families were identified in all the three stations
together and the species composition was rich in station 3 with the total
number of 44 species. Part II deals
with the in vitro and in vivo antiproliferative and
immunomodulatory studies of the ethanolic extract of simple ascidian Phallusia nigra against DLA, EAC, S-180,
HLCA-549 and MDA-MB-231 bearing mice. Acute oral toxicity studies using the
ethanolic extract of P. nigra to Swiss albino mice administered
with 2000 mg/kg body weight did not show any mortality. 100% cytotoxicity to
the above mentioned cells were observed at a concentration of 0.60, 0.40, 0.60,
0.80 and 0.60 mg/ml respectively. Administration of the extract increased
median survival time, % life span, non viable cells, hemoglobin, RBC,
lymphocyte, eosinophil, bone marrow cellularity, β-Esterase positive cells,
antibody titer and the plaque forming cells in spleen. A reduction in the body
weight, relative organ weight, solid tumor volume, packed cell volume, viable
cells, WBC, neutrophils, serum Gamma Glutamyl Transpeptidase (GGT), cellular
Glutathione (GSH) and restoration of NO levels were observed on treatment with
the extract in DLA, EAC, S-180, HLCA-549 and MDA-MB-231 tumor bearing mice. The
activity was comparatively greater than that observed for the standard drug -
Vincristin in all the parameters tested. Quantitative hemolysis of sheep red
blood cells, lymphocyte proliferation, NK cytotoxic activity and phagocytosis
rate increased on treatment with the extract indicating activation of immune
function.
2. Chemical screening and
pharmacological evaluation of ascidians, Ph.D. thesis by S. Gomathy, Assistant Professor, Dept. of Zoology, A.P.C.
Mahalaxmi College for Women, Tuticorin
under the guidance of Dr. V.K. Meenakshi. submitted to
Manonmaniam sundaranar University, Tirunelveli, Tamil Nadu, India. gomathyapcm@gmail.com
Ascidians,
an important group of marine, sedentary organisms found distributed along the
Indian coast throughout the year has been screened for their chemical
constituents and pharmacological properties. The thesis consists of five
chapters. Chapter 1 deals with the study of the distribution of ascidians at 3
different stations Kanyakumari, Thoothukudi and Mandapam along the Southeast
coast of India during January to December 2011. 53 species of ascidians
belonging to 22 genera coming under 9 families were identified in all the three
stations together. Chapter 2 deals with Chemical screening and pharmacognostic
studies of Microcosmus exasperatus. The specimen of
Microcosmus exasperatus was analyzed
macroscopically, microscopically and extracted using different solvents such as
petroleum ether, benzene, chloroform, ethanol, methanol, methylene chloride and
water. Alkaloids, terpenoids, steroids, coumarins, tannins,
saponins, flavonoids, quinones and anthraquinones were observed. Ethanolic and
Methanolic extract of Microcosmus
exasperatus has been subjected to GC – MS analysis which showed the
presence of twenty and nine compounds respectively. HPTLC studies revealed the
presence of phenolic compounds such as Gallic acid, Ferulic acid and Caffeic
acid. Rutin, Isoquercitrin and Quercetin were some of the flavonoids
identified. Fat soluble vitamins D3, A, K and water soluble vitamins
riboflavin, thiamine were noticed in high concentration. In
Chapter 3, studies on hepatoprotective activity of Microcosmus exasperatus
has been performed in Carbon
tetrachloride induced liver damaged Wistar albino rats which indicated an
increase in body weight, protein, albumin, globulin, total conjugated,
unconjugated bilirubin, GGT, GPX, GR, SOD, CAT, GSH and decrease
in the level of LPO, SGPT, SGOT, ALP in serum. Antidiabetic studies in
alloxan induced diabetic rats in Chapter
4 recorded an increase in insulin
level, protein, albumin, globulin, SOD, CAT, GPX, GSH, GR and a decrease in blood
glucose, urea, creatinine, HbAlc, SGPT, SGOT, ALP, TC, TG, LDL – C, VLDL, HDL,
PL and LPO activities in serum. Antifertility activity of the ethanolic extract of Microcosmus exasperatus in male albino rats in Chapter
5 noted a significant
decrease in the weight of body, testis and the accessory sex organs. A dose
related reduction in sperm count of testis, epididymis, motility and increase
in the level of abnormal sperms were observed. The serum biochemical parameters
and liver marker enzymes did not show any significant variation.
3. Studies on a few aspects of ascidians from the Gulf of Mannar. Ph.D. thesis by S. Senthamarai,
Research Scholar, PG and Research Dept. of Zoology, V.O. Chidambaram College,
Tuticorin. Submitted to Manonmaniam Sundaranar University, Tirunelveli, Tamil
Nadu, India under the guidance of Dr. V.K. Meenakshi.
Marine biodiversity of the Gulf of
Mannar is poorly known. Knowledge of the ascidian biodiversity of the ocean
makes it one of the greatest resources for drug discovery. The thesis consists
of two parts. Part 1 deals with the biodiversity of Gulf of Mannar ascidians, which
includes the section on Taxonomy giving detailed descriptions of the newly
recorded species. The biological resources study of ascidians of Gulf of Mannar
could give as many as 26 species of ascidians as new records to Indian waters
belonging to 3 families and 9 genera. Of the 26 species, one species - Aplidium digitalis is new to science.
The total number of species of ascidians from Gulf of Mannar water has now gone
up to 99. All the 26 species which are
new to India have been dealt in detail with photographs and camera Lucida
diagrams. Part 2 deals with the biomedical applications of a selected ascidian
from the Gulf of Mannar and contains two sections. In the first section the
screening of antibacterial activity of the simple
ascidian Microcosmus exasperatus against
MTCC and clinical isolates of human pathogens affecting the various tracts has
been carried out. The present study reveals that the crude Methylene Chloride
extract was more active, exhibiting a broad spectrum antibacterial activity
than the selected commercially available antibiotics. Maximum zone of
inhibition (23 mm) was observed against Streptococcus
pneumoniae (655) of respiratory tract and Actinomyces sp. of wound pathogen. The second section
focuses the in vitro and in vivo antitumor
and immunomodulatory studies of the ethanolic extract of simple ascidian Microcosmus exasperatus against DLA
bearing mice. Acute oral toxicity studies using the ethanolic extract of
M. exasperatus to Swiss albino
mice administered with 2000 mg/kg body weight did not show any
mortality. 100% cytotoxicity to DLA cells was observed at a concentration of
0.80 mg/ml. Administration of the extract of M. exasperatus increased
median survival time, % of life span, non viable cells, heamoglobin, RBC,
lymphocyte, eosinophil, bone marrow cellularity, β-esterase positive
cells, antibody titer and the plaque forming cells in spleen. A reduction in
the body weight, relative organ weight, solid tumor volume, packed cell volume,
viable cells, WBC, neutrophils, serum Gamma Glutamyl Transpeptidase (GGT),
cellular Glutathione (GSH) and restoration of NO levels in DLA tumor bearing
mice. The activity was comparatively greater than that observed for the
standard drug (Vincristin) in all the parameters tested.
4. Phenotypic plasticity and
adaptation potential to salinity in early life stages of the tunicate Ciona intestinalis sp. B. Elin
Renborg, PhD thesis. Awarded April 29, 2014 from University of Gothenburg,
Tjarno Marine Biological Laboratory; Jon Havenhand supervisor.
See Renborg,
E., Johannesson, K. and Havenhand, J. 2014. See: Renborg, E., Johannesson, K.
and Havenhand, J. 2014. Variable salinity tolerance in ascidian larvae is
primarily a plastic response to the parental environment. Evol.
Ecol. 28: 561-572. Abstract:
Both phenotypic plasticity and local
genetic adaptation may contribute to a species’ ability to inhabit different
environmental conditions. While phenotypic plasticity is usually considered
costly, local adaptation takes generations to respond to environmental change
and may be constrained by strong gene flow. The majority of marine species have
complex life-cycles with pelagic stages that might be expected to promote gene
flow and plastic responses, and yet several notable examples of local
adaptation have been found in species with broadcast larvae. In the ascidian,
Ciona intestinalis (Linnaeus, 1767),—a common marine species with broadcast
spawning and a short larval stage—previous studies have found marked
differences in salinity tolerance of early life-history stages among
populations from different salinity regimes. We used common-garden experiments
to test whether observed differences in salinity tolerance could be explained
by phenotypic plasticity. Adult ascidians from two low salinity populations
[2–5 m depth, ~25 practical salinity units (PSU)], and two full salinity
populations (25–27 m depth, ~31 PSU) were acclimated for 2–4 weeks at both 25
and 31 PSU. Gametes were fertilized at the acclimation salinities, and the
newly formed embryos were transferred to 10 different salinities (21–39 PSU)
and cultured to metamorphosis. Adult acclimation salinity had an overriding and
significant effect on larval metamorphic success: tolerance norms for larvae
almost fully matched the acclimation salinity of the parents, independent of
parental origin (deep or shallow). However we also detected minor population
differences that could be attributed to either local adaptation or persistent
environmental effects. We conclude that differences in salinity tolerance of C.
intestinalis larvae from different populations are driven primarily by
transgenerational phenotypic plasticity, a strategy that seems particularly
favourable for an organism living in coastal waters where salinity is less
readily predicted than in the open oceans.
5. Ecology and
genetics of invasive ascidians in the Western Mediterranean. Víctor Ordóñez. Ph.D.
thesis 2013, Univ. of Barcelona, Catalonia, Spain. Advisors: Marta
Pascual, Xavier Turon. xturon@ceab.csic.es
Biological invasions are of special
concern to marine biologists. The Mediterranean is one of the seas’ most
threatened from this point of view. The main goal of this PhD thesis is to
study ecological and genetic aspects of the biology of four invasive ascidians
in the western Mediterranean. In the first chapter, we delve into the post-border
processes of the ascidian Microcosmus
squamiger, by analysing its population genetic structure. This ascidian has
a relatively high capacity of post-border dispersion, which allows gene flow
between populations on a small scale. Moreover there are two genetic clusters
related to the two original sources of the introduced worldwide populations.
These genetic pools are mixed at the population level but not at the individual
level. In the second chapter, we evaluate biological interactions in early-life
history stages of Microcosmus squamiger
and Styela plicata, among themselves
and with mussels. No interactions between the two ascidians were found in
fertilization or larval settlement and metamorphosis processes. In contrast,
larvae of both ascidians were consumed by juvenile mussels, and the ascidians
also presented shifts in larval behavior in the presence of the mussels. This
highlights the importance of the resident community for the establishment of
introduced species. In the third chapter, we study the life cycle and
population genetic structure of the invasive Didemnum vexillum, which is reported for the first time in western
Mediterranean. This ascidian showed a marked seasonal growth and reproductive
cycle, negatively influenced by warm summer temperatures. Additionally,
introduced populations in the Mediterranean showed low diversity, suggesting a
bottleneck process in the introduction. Finally, in the fourth chapter, we study the life cycle of Clavelina oblonga. This ascidian was in
fact introduced in the Mediterranean for more than 80 years now, but it was
erroneously described as a new species, C.
phlegraea, purportedly native to the Mediterranean. Their reproductive and
growth cycle was markedly seasonal, favored by warm summer temperatures, and hibernating
in colder months.
6. Species
invasion in the marine fouling communities of British Columbia: factors that
influence invasion dynamics and how they may affect Botrylloides violaceus. Jocelyn Nelson, M.S. thesis, Univ. of British
Columbia, Vancouver, BC, Canada. Advisors Drs. Christopher D. G. Harley and
Thomas W. Therriault. jocelyn.nelson@gmail.com
Species
invasion has been recognized as a major threat to biodiversity. Knowledge of
the factors that limit the establishment and spread of non-indigenous species
(NIS), such as biotic resistance and unfavourable environmental conditions, are
important to their effective management. To test the biotic resistance and
environmental favourability hypotheses in the fouling communities of British
Columbia (BC), 22 locations were compared using settlement tiles in a
large-scale survey. Biotic resistance is believed to be stronger in more
diverse communities, therefore NIS richness and abundance were compared to
native species richness and environmental conditions to investigate their
importance using generalized and linear mixed models. Invader taxonomic group
may influence biotic resistance, and environmental tolerances vary by species,
therefore factors that affected Botrylloides violaceus presence and
abundance were investigated as a case study. The biotic resistance hypothesis
was not supported for NIS richness or NIS abundance, but could not be fully
discounted due to a trend toward a negative slope between native species
richness and B. violaceus presence and abundance, and the absence of
predator data. Environmental variables affected NIS: salinity had a positive
influence on NIS richness, NIS abundance, and B. violaceus presence, and
temperature had a positive effect on B. violaceus presence and
abundance. Salinity had a positive impact on native species richness as well,
supporting the environmental favourability hypothesis. This suggests that
knowledge of relevant environmental conditions is more important for the
management of invasive species than the species richness of vulnerable
communities.
Environmental
conditions are not static, so species invasion must be considered in the
context of climate change. To understand how climate change may influence
species invasion, B. violaceus presence and abundance in BC were
compared to a range of abiotic conditions. This comparison informed a GAMLSS
model that used linear trends from historical shore station data to project
potential abundance in BC forward 50 years. Overall, the abundance of B.
violaceus in BC was projected to increase. A larger increase in B.
violaceus abundance was forecast for locations where conditions increased
into the range favourable for growth. If temperature and salinity become more
favourable for B. violaceus, as projected, climate change could
intensify the invasion.
The full text
can be found online at http://circle.ubc.ca/handle/2429/46606.
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