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1.1. PREAMBLE:

Pharmacognosy is defined as the study of drugs and drug

substances as well as potential drugs and drug substances from

natural sources and also search for drugs and drug substances from

natural sources (American Society of Pharmacognosy). From the wide

range of natural sources, marine is the source of choice for the region

of Goa. Being located almost on every side by the water body, a search

for newer drugs from this is but apt. The word marine is

representative of all those that are related to seas and oceans both

inside as well as around them.

Before going into the details of the literature a brief introduction

about the topic is being presented. In this, the topic is mainly divided

into three aspects, Proteins, GFP-like proteins and Zoanthids. To

begin with proteins and their importance is discussed in brief followed

by the discussion on GFP-like proteins and their speciality and

concluding with the description of marine zoanthids.

1.2. PROTEINS AND THEIR IMPORTANCE:

Proteins are the most abundant biological macromolecules,

occurring in all cells and in all parts of the cells. These are polymers

containing combinations of 20 different amino acid residues joined to

its neighbour by a specific type of covalent bond. They occur in great

variety and ranging in size from relatively small peptides to huge

polymers with molecular weights in millions, together may be found in

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a single cell. Moreover, proteins exhibit enormous diversity of

biological function (Fig.1.1. & Fig.1.2.) and are the most important

final products of the information pathways. They also play the role of

molecular instruments through which genetic information is

expressed[1].

Every protein has a specific order of arrangement of amino acids

and can be described at four levels of structural hierarchy (Fig.1.3.). A

description of all covalent bonds (mainly peptide bonds and disulphide

bonds) linking amino acid residues in a polypeptide chain is its

primary structure which clarifies the sequence of amino acid residues.

Secondary structure refers to residues giving rise to recurring

structural patterns. Tertiary structure describes all aspects of the

three-dimensional folding of a polypeptide. When a protein has two or

more polypeptide subunits, their arrangement in space is referred to

as quaternary structure[1].

Fig.1.1. The light produced by fireflies is

the result of a reaction involving the

protein luciferin and ATP, catalyzed by

the enzyme luciferase

Figure.1.2.The protein keratin, formed by

all vertebrates, is the chief structural

component of hair, scales, horn, wool,

nails, and feathers

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At the higher levels of structure (quaternary), they are also

classified as fibrous proteins, having polypeptide chains arranged in

long strands or sheets and globular proteins, having polypeptide

chains folded into a spherical or globular shape. Fibrous proteins,

usually consists of largely of a single type of secondary structure;

globular proteins often contain several types of secondary structures.

The former, play the role of structures, provide support, shape and

external protection to vertebrates, whereas the latter are mostly

enzymes and regulatory proteins[1].

Super secondary structures, also called motifs or simply folds,

are particularly stable arrangements of several elements of secondary

structure and the connections between them. Many examples of

recurring domains or motif structures are available (Fig.1.4.), and

these reveal that protein tertiary structure is more reliably conserved

than primary sequence. Proteins with significant primary sequence

Fig.1.3. Levels of structure in proteins. The primary structure consists of a sequence of

amino acids linked together by peptide bonds and includes any disulfide bonds. The

resulting polypeptide can be coiled into units of secondary structure, such as an _ helix.

The helix is a part of the tertiary structure of the folded polypeptide, which is itself one of

the subunits that make up the quaternary structure of the multisubunit protein, in this

case hemoglobin1.

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similarity, and/or with demonstrably similar structure and functions

are said to be in the same protein family. A strong evolutionary

relationship is usually evident within a protein family[1].

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Fig.1.4. Organisation of proteins based on motifs. Shown above are just a small number of the

hundreds of known stable motifs. They are divided into four classes: all α, all β, α/β, and α+β.

Structural classification data from the SCOP (Structural Classification of Proteins) database

(http://scop.mrc-lmb.cam.ac.uk/scop) are also provided. The PDB identifier is the unique number

given to each structure archived in the Protein Data Bank (www.rcsb.org/pdb).

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Two or more families with little primary sequence similarities

sometimes make use of the same major structural motif and have

functional similarities; these families are grouped as superfamilies[1].

The simple string of letters denoting the amino acid sequences of a

given protein belies the wealth of information this sequence holds.

The study of molecular evolution generally focuses on families of

closely related proteins. Usually, the families chosen for analysis have

essential functions in cellular metabolism that must have been

present in the earliest viable cells, thus greatly reducing the chance

that they were introduced relatively recently by lateral gene transfer.

The premise is simple; if two organisms are closely related, the

sequence of their genes and proteins should be similar or the

sequences increasingly diverge as the evolutionary distance between

two organisms’ increases. The members of protein families are called

homologous proteins or homologs (Fig.1.5.). Homologs present in the

same species are called as paralogs and from different species called

as orthologs. The process of tracing evolution involves first identifying

suitable families of homologous proteins and then using them to

reconstruct evolutionary paths[1].

Fig.1.5. A signature sequence in the EF-1_/EF-Tu protein family. The signature sequence (boxed)

is a 12-amino-acid insertion near the amino terminus of the sequence. Residues that align in all

species are shaded yellow. Both archaebacteria and eukaryotes have the signature, although the

sequences of the insertions are quite distinct for the two groups. The variation in the signature

sequence reflects the significant evolutionary divergence that has occurred at this site since it

first appeared in a common ancestor of both groups

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For most efforts to find homologies and explore evolutionary

relationships, protein sequences (derived either directly from protein

sequences or from the sequencing of the DNA encoding the protein)

are superior to non-genic nucleic acid sequences (those that do not

encode a protein or functional RNA). Knowledge of the sequence of

amino acids in a protein can offer insights into its three-dimensional

structure and its function, cellular location and evolution. Most of

these insights are derived by searching for homologies. Thousands of

sequences are known and available in data bases accessible through

the internet. A comparision of a newly obtained sequence with this

large bank of stored sequences often reveals relationships (Fig.1.6.)

both surprising and enlightening[1].

Fig.1.6. Evolutionary tree derived from amino acid sequence comparisons. A bacterial

evolutionary tree, based on the sequence divergence observed in the GroEL family of proteins.

Also included in this tree (lower right) are the chloroplasts (chl.) of some nonbacterial species.

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1.3. GREEN FLUORESCENT PROTEIN-LIKE PROTEINS AND THEIR

SPECIALITY:

Bioluminescence (biofluorescence) is the capacity of living

organisms to emit visible light[2]. In doing so they utilized a variety of

chemiluminescent reaction systems. Many a times the word

phosphorescence has been erroneously used to describe marine

bioluminescence. Some terrestrial species (eg., fireflies) have the same

ability, but this adaptation has been most extensively developed in the

oceans. Bioluminescent species occur in only five terrestrial phyla,

and only in one of these (Arthropoda, which includes the insects) are

there many examples. In contrast, bioluminescence occurs in 14

marine phyla, many of which include numerous luminescent species.

All oceanic habitats, shallow and deep, pelagic and benthic, include

bioluminescent species, but the phenomenon is commonest in the

upper 1000m of the pelagic environment[3].

Bioluminescence involves the oxidation of a substrate (luciferin) in

the presence of an enzyme (luciferase). The distinctive feature of the

reaction is that most of the energy generated is emitted as light rather

than as heat. There are many different and unrelated kinds of

luciferin, and biochemical and taxonomic criteria indicate that

bioluminescence has been independently evolved many times. Marine

animals, are unusual, however, in that many species in at least seven

phyla use the same luciferin. This compound is known as

coelenterazine because it was first identified in jellyfish (coelenterates)

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and its molecular structure is derived from a ring of three amino acids

(two tyrosines, and a phenylalanine). Nevertheless, many other

marine organisms use different luciferins. In some animals (eg.,

jellyfish) the luciferin/luciferase system can be extracted in the form of

a stable ‘photoprotein’ that will emit light when treated with

calcium[4,5].

1.3.1. History:

The first report was made by Davenport and Nicol, which

described the green fluorescence of the light organs of Aequorea

victoria (Fig.1.7. & Fig.1.8.) in 1955. The photoprotein responsible for

this bioluminescence was identified as Aequorin along with the

discovery of a companion protein Green Fluorescent Protein (GFP)

(Fig.1.9.) by Shimomura et.al., in 1961[6]. Though the

chemiluminescence (in vitro) of aequorin is blue, the bioluminescence

(in vivo) of Aequorea victoria is found to be green and the reason is

identified, as the radiation-less energy transfer between aequorin and

GFP, in vivo, by Morin and Hastings in 1969[7]. In 1979, Shimomura

et. al., proteolyzed denatured GFP, analysed the peptide that retained

visible absorbance and correctly proposed that the chromophore

(Fig.1.10) is a 4-(p-hydroxybenzylidene) imadazolidin-5-one attached

to the peptide backbone through the 1-and 2- positions of the ring[8].

The curcial break-through came with the cloning of the gene by

Prasher et.al., and the demonstrations by Chalfie et.al., and Inouye

and Tsuji, that expression of the gene in other organisms creates

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fluorescence[9,10,11]. Therefore, the gene contains all the information

necessary for the post-translational synthesis of the chromophore, and

no-jellyfish-specific enzymes are needed.

Fig.1.7. Jellyfish Aequorea victoria

Fig.1.8. The light-emitting organs are located along the edge of the umbrella

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Fig.1.10. Reactions in Chemiluminescence of Aequorin

Fig.1.9. Green Fluorescent Protein

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1.3.2. Chemistry:

The structure of GFP has been reported to have been solved

using seleniomethionyl-substituted protein and multi-wavelength

anomalous dispersion (MAD) phasing methods. The electron density

maps produced by the MAD phasing are clear, revealing a dimer

comprised of two quite regular barrels with 11strands on the outside

of cylinders (Fig.1.11). These cylinders have a diameter of about 30 Å

and a length of about 40 Å. Inspection of the density within the

cylinders revealed modified tyrosine side chains as part of an irregular

helical segment. Small sections of α-helix also form caps on the end of

the cylinders (Fig.1.12). This motif or folding arrangement, with a

single α-helix inside a very uniform cylinder of β-sheet structure,

represents a new protein class (α+β), as it is not similar to any known

protein structure. Two protomers pack closely together to form a

dimer in the crystal. The protein is comprised of 238 amino acids and

has a molecular weight of 26.9 KDa. Its wild type

absorbance/excitation peak is at 395 nm with a minor peak at 475

nm with extinction coefficients of roughly 30,000 and 7,000 M-1 cm-1

respectively. The emission peak is at 508 nm[11].

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Fig.1.11. The overall shape of the protein and its association into dimers. Eleven strands of β-sheet

(green) form the walls of a cylinder. Short segments of α-helices (blue) cap the top and bottom of the

'β-can' and also provide a scaffold for the fluorophore which is near geometric center of the can.

This folding motif, with β-sheet outside and helix inside, represents a new class of proteins. Two

monomers are associated into a dimer in the crystal and in solution at low ionic strengths. This

view is directly down the two-fold axis of the non-crystallographic symmetry.

Fig..No.1.12. A topology diagram of the folding pattern in GFP. The -sheet strands are shown in

light green, α-helices in blue, and connecting loops in yellow. The positions in the sequence that

begin and end each major secondary structure element are also given. The anti-parallel strands

(except for the interactions between stands 1 and 6) make a tightly formed barrel.11

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1.3.3. Chromophore Biosynthesis:

The production of visual colour is related to the formation and

maturation of a chromophore system. The part of the biomolecule

responsible for the production of any sort of colour or luminescence is

called chromophore. It is a covalently unsaturated group (eg:- C=C,

C=O, NO2 etc), responsible for absorption of energy from a radiant

source and brings about electronic transitions within the molecule.

When these transitions are of -* or n-* type, the chromophore

system that is formed, produces colour in the visible region of the

electromagnetic spectrum. However, when these transitions are not

stable and electrons drop back to the ground state from the excited

state emitting energy in the form of radiation, then it is called

fluorescence and the chromophore is then called the fluorophore[12].

In GFPs the chromophore is generated only under conditions

permissive of protein folding; that is, the polypeptide must be able to

obtain its native three-dimensional structure to become visible

fluorescent. It is generated in the presence of molecular oxygen. This

autocatalytic mechanism (Fig.1.13) is initiated by an intrachain ring

closure that leads to the formation of a cyclopentyl group from the

backbone of the original peptide (Ser65, Tyr66 and Gly67)(Fig.1.14).

The fluorophore originates from an internal Ser-Tyr-Gly sequence

which is post-translationally modified to a 4-(p-hydroxybenzylidiene)-

imidazolidin-5one structure. Studies on recombinant GFP expression

in Escherichia Coli led to a proposed sequential mechanism initiated

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by a rapid cyclization between Ser65 and Gly67 to form a imidazolin-

5-one intermediate, followed by a much slower (hours) rate-limiting

oxygenation of the Tyr66 side chain by O2[13].

Fig.1.14. Model of the fluorophore and its environment superposed on the MAD-phased electron

density map at 2.2 Å resolution. The clear definition throughout the map allowed the chain to be

traced and side chains to be well placed. The density for Ser65, Tyr66 and Gly67 is quite consistent

with the dehydrotyrosine - imidazolidone structure proposed for the fluorophore. Many of the side

chains adjacent to the fluorophore are labeled.

Fig.1.13. Fluorophore formation in GFP. Folding of proteins promotes cyclization which

is followed by debydration of the ring and oxidation of the tyrosine

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Combinatorial mutagenesis suggests that the Gly67 is required for

formation of the fluorophore. While no known co-factors or enzymatic

components are required for this apparently autocatalytic process, it is

rather thermosensitive with the yield of fluorescently active to total

GFP protein decreasing at temperatures greater than 30ºC. However,

once produced, GFP is quite thermostable[11].

1.3.4. Physical & Chemical Characteristics:

GFP is very resistant to denaturation requiring treatment with

6M guanidine hydrochloride at 90C or pH of <4.0 or >12.0. Partial to

near-total renaturation occurs within minutes following reversal of

denaturing conditions by dialysis or neutralization. Circular

dichroism predicts significant amounts of -sheet structure that is

subsequently lost on denaturation. Over a non-denaturing range of

pH, increasing pH leads to a reduction in fluorescence by 395nm

excitation and an increased sensitivity to 475nm excitation.

Reduction of purified GFP by sodium dithionite results in a rapid loss

of fluorescence that slowly recovers in the presence of room air. While

insensitive to sulfhydryl reagents such as 2-mercaptoethanol,

treatment with the sulfhydral reagent dithiobisnitrobenzoic acid

(DTNB) irreversibly eliminates fluorescence[11].

The remarkable cylindrical fold of the protein seems ideally

suited for its function. The strands of -sheet are tightly fitted to each

other like staves in a barrel and form a regular pattern of hydrogen

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bonds. Together with the short helices and loops on the ends, the

‘can’ structure forms a single compact domain and does not have

obvious clefts for easy access of diffusable ligands to the fluorophore.

Photochemical damage by the formation of singlet oxygen through

intersystem crossing is reduced by the structure. The tightly

constructed -can would appear to serve this role well and also

provide overall stability and resistance to unfolding by heat and

denaturants[11].

1.3.5. Physiological Functions:

The pigments containing these proteins display slow decay

rates, characterized by half-lives of 20days. The slow turnover of

GFP-like proteins implies that the associated energetic costs for being

colourful are comparatively low. Moreover, high in vivo stability makes

GFP-like proteins suitable for functions requiring high pigment

concentration such as photoprotection. The underlying mechanism,

however, remains controversial, as some FPs have spectral properties

that appear to be inappropriate for photoprotecting tissue by

modulating the intracellular light climate, other FPs are spectrally well

suited to fluorescence energy transfer and dissipation of light energy

via radiative and nonradiative pathways. An antioxidant function has

recently been suggested[14].

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1.3.6. GFP Homologs Comprise a Superfamily:

The group of structural homologs of Green Fluorescent Protein

(GFP) that all share the GFP-like “beta-can” fold are regarded as a

superfamily following the criteria proposed by the Protein Information

Resource(http://pir.georgetown.edu/pir,http://www.otherinfo/sfdef.h

tml.) The reason for such a classification is that this group unites at

least two clearly definable protein families. The first one consists of

G2FP domains, which are incapable of autocatalytic chromophore

synthesis and are found within multi-domain proteins of the

extracellular matrix. The second one includes fluorescent and/or

coloured proteins capable of synthesizing the chromophore auto-

catalytically and which are not found in a multi-domain context[1].

1.4. ZOANTHID TAXONOMY:

Zoanthids are hexacorallians belonging to the order Zoantharia, of

the class Anthozoa, under the phylum Coelenterata/Cnidaria.

1.4.1. Phyllum Coelenterata/Cnidaria:

Coelenterata have a pronounced radial symmertry, the body being

star-like, with the organs arranged symmetrically on lines radiating

from a common centre. The word “polyp” is fregquently applied to the

individual coelenterate animal or zooid, was originally introduced on a

fancied resemblance of hydra to a small cuttle fish (Fr – Poulpe,

Lat – Polypus)[15].

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The body of the Coelenterata, then consists of body-wall enclosing

a single cavity called “Coelenteron”. The body wall consists of an inner

and an inner and an outer layer of the cells, the “endoderm” and

“ectoderm” respectively. Between the two layers there is a substance

chemically allied to mucin and usually of jelly-like consistency, called

the “mesoglea”. The mesoglea may be very thin and inconspicuous, as

it is in Hydra and many other sedentary forms, or it may become very

thick as in the jelly-fishes and some of the sedentary Alcyonaria[15].

Another character, of great importance, possessed by all

coelenterate is the “Cnidae”. These are organelle-like capsules with

eversible tubules. There are three types of cnidae – nematocysts,

ptychocysts and spirocysts. Three other features sometimes

considered to be diagnostic of cnidria are radial symmetry, planula

and polyp stages in developmonent, but all have exceptions. Although

many cnidarians exhibit radial symmetry, some are directionally

asymmetric, and many have a biradial or bilateral organisations. The

motile stage between embryo and settled juvenile in any given

cnidarians life cycle is typically termed a planula, and although this

stage is usually ciliated, sausage-shaped, and non-feeding, deviations

of this pattern have been well documented. Polyp forms are even more

variable than planulae being solitary or colonial; if colonial,, polyps

may be monomorphic or polymorphic; they may or may not have a

mineralized skeleton; they may be benthic or pelagic; and tentacles,

although commonly present, may be absent. The phylum Coelenterata

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is further divided into three classes Hydrozoa, Anthozoa and

Scyphozoa[15].

1.4.2. Class Anthozoa:

The class Anthozoa comprises of two reciprocally monophyletic

lineages, Octocorallia and Hexacorallia. All members of Anthozoa are

exclusively polypoid, and may be colonial, clonal or solitary skeleton-

less or with a mineralic and/or protinaceous skeleton. Anthozoa

currently contains 7,500 extant species[15].

Phylogenetic analysis of morphological data, has suggested atleast

three diagnostic apomorphies for Anthozoa: actinopharynx,

siphonoglyph and mesenteries (Fig.1.15). The actinopharynx

(=stomadeum, gullet) is an ectoderm-lined tube that projects into the

gastro vascular cavity (=coelenteron); this structure is found in all

Anthozoa, with one known exception, the black coral sibopathes. The

siphonoglyph (=sulcus) is a densely ciliated, often more highly

glandular region of the actinopharynx; it is single, paired or rarely

absent (e.g., in ptychodactarian sea anemones; the presence of a

siphonoglyph in antipatharians is disputed), and in asexually-derived

individuals, there may be more than two. The siphonoglyph reflects

the plane of bilateral symmetry for the polyp. Bilateral symmetry is

further defined by the mesentries (the term septa, which has been

used for these structures has been reserved for mesentreries of

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scleratinians), radially-arrayed sheets of tissue that extend all or part

of the way

from the body wall to the actinopharynx. Mesenteries are arranged in

cycles (members of each cycle form more or less simultaneously, and

bear the gametogenic tissue and epithelia-muscular cells that are

concentrated as retractor muscles. The free edge of a mesentry is

typically elaborated into a mesenterial filament with abundant gland

cells, nematocysts and cilia[15].

1.4.3. Sub-Class Hexacorallia:

The anthozoan subclass Hexacorallia comprises all scleractinian

and black corals, tube anemones, and sea anemones in the broadest

sence (i.e., orders Actinaria, Antipatharia, Ceriantharia,

Fig.1.15. Sketch showing the internal anatomy of Anthozoa.

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Corallimorpharia, Scleractinia and Zoanthidae). Hexacorallia

currently contains about 4,300 extant species. Most hexacorallians

have hexamerous symmetry, although eight or ten-part symmetry are

not uncommon. All members of Hexacorallia have spirocysts, a type of

cnida with a single-walled capsule and a tubule composed of tiny

entangling sub-threads[15].

1.4.4. Order Zoanthidea:

Members of order Zoanthidea (=Zoantharia, Zoanthinaria) are

clonal, soft bodied polyps with two rows of marginal tentacles. Their

internal anatomy and mesenterial arrangement is distinctive among

hexacorallians, and the group is presumed to monophyletic, although

no published studies have examined this question explicitly[15].

Zoanthideans have traditionally been grouped into two suborders:

Macrocnemina and Brachycnemina, which differ in the arrangement of

the mesenteries. The fifth pair of mesentry from the dorsal directive is

incomplete in the case of Brachycnemina and complete in the

macrocnemina. The families under the sub-order Brachycnemina –

Sphenopidae and Zoanthidae; the families under the sub-order

Macrocnemina – Epizoanthidae, Abyssoanthidae and

Neozoanthidae[15].

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1.4.5. Family - Zoanthidae:

Zoanthidae comprises of three genera Zoanthus, Isaurus and

Acrozoanthus which include variable number of species depending

upon the authors[15].

1.5. WHAT ARE ZOANTHIDS?

Zoanthids (Fig.1.16) belong to the same class Anthozoa as sea

anemones. Zoanthid taxonomy is undergoing some review so the

number of known zoanthid species range from 200 to 60 depending on

how the species are defined[16].

1.5.1. Where seen?

These tiny but tough flower-like animals often carpet rocky and

rubbly areas. Some are adapted be regularly exposed to the air at low

tide. These animals are often the first to settle on any vacated space

in a reef[16].

Fig.1.16. A picture of zoanthis polyps

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1.5.2. Features:

Zoanthids look like tiny anemones. But with sea anemones are

solitary polyps, most zoanthids live in colonies (Fig.1.17) like corals do.

They don’t produce a hard skeleton like the hard coral colonies.

Instead, their skin is leathery and composed partly of chitin (the same

substance that insect exoskeletons are made of)[16].

The typical polyp has a cylindrical body coloumn, topped by a

smooth, flat oral disc that is edged by short tentacles, usually in two

rows close to one another. The oral disc is often in a contrasting

bright colour from the usually brown or drab tentacles, usually in two

rows close to one another. When exposed to low tide, however, the

animals retracts its tentacles into its body column and then looks like

a strange blob of jelly[16].

Zoanthids may have three different living arrangements

(Fig.1.18). Each zoanthid polyp may be solitary but located near one

another. These polyps are large with thick, fleshy polyps on tall

Fig.1.17. A picture of mat of zoanthids

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coloumns. Or the zoanthid polyps are joined to one another by stolons

(tube-like structures that spread across the ground like a root or

runner) – the “liberae” form. Or the zoanthid polyp may be embedded

in a common mat of tissue – “coenenchyme”. The tissue may be

strengthened by incorporating sand. The colony may form mats on

the sand or encrust rocky areas – the “intermedea” and “immerse”

forms[16].

The shape of the same zoanthid species may vary depending on

where they are found. Those inhabiting arrears with strong waves

Fig.1.18.Diagram of colony and polyp structure forms of zoanthids. a) ”immersae” form,

with polyps deeply embedded in a well-developed coenenchyme; b) “intermediate” form,

intermediate in form, ususally with well-developed, thick polyps; c) “liberae” form, with

free-standing polyps extending well above a thin coenenchyme (stolons). Often with space between oral disks.

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tend to be short and hug the surface. Others found in deeper, calmer

waaters are taller, with longer tentacles[16].

1.5.3. Toxic Flowers:

Some zoanthids contain powerful toxins to protect themselves

against predators. The most toxic marine poison, palytoxin, was

discovered in a Zoanthid (Fig.1.19). Minute quantities of palytoxin can

paralyse and even kill. So zoanthids should not be handled with open

wounds on hand or mouth or eyes should be touched after handling

them. It is believed that the toxins are not produced by the animal

but by bacteria that live in symbiosis with the polyps. However, some

animals have adapted to the poison and even eat zoanthids. These

include the common hairy crab, filefishes and nudibranchs[16].

Fig.1.19. Colourful zoanthids which are toxic flowers

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1.5.4. What do they eat?

Most zoanthids feed on amphipods (Fig.1.20), plankton, some also

feed on finer particles. Many harbour zooxanthellae (symbiotic algae)

inside their bodies. These carry out photosynthesis and may

contribute nutrients to the host polyp[16].

1.5.5. Zoanthid Babies:

Zoanthids generally produce asexually – new polyps bud to enlarge

the colony. However, they also reproduce sexually. The polys may

produce sperm or eggs, but usually only one at a time[16].

Fig.1.20. Zoanthids eating amphipods

Fig.1.21. Zoanthid releasing eggs during spawing

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Eggs and sperms are released synchronously for external fertilization

(Fig.1.21.), in mass spawing similar to that practiced by hard

corals[16].

1.5.6. Status and Threats:

Zoanthids are not listed among the threatened animals. However,

like other creatures of the intertidal zone, they are affected by human

activities such as reclamation and pollution. Trampling by careless

visitors, and over collection also have an impact on local

population[16].