August 2013 - October 2013, Vol. 3, No. 4; 2712-2721 . E- ISSN: 2249 –1929

Journal of Chemical, Biological and Physical Sciences

An International Peer Review E-3 Journal of Sciences Avail able online at www.jcbsc.org Section B: Biological Science

CODEN (USA): JCBPAT Research article

On The Occurrence of One New Species and Three New Records of the Genus Aplidium from Indian Waters

V.K. Meenakshi 1*and S. Senthamarai 1

1 Department of Zoology, A.P.C. Mahalaxmi College for Women, Tuticorin, Tamilnadu, India.

Received: 30 August 2013; Revised: 19 September 2013; Accepted: 28 September 2013

Abstract: Ascidians which are pharmacologically important marine sedentary organisms were collected from mussel landing centers, trawl discards and by SCUBA diving from the Gulf of Mannar waters of India. They were identified up to the species level. The survey reports one species - Aplidium digitalis n.sp., as new to science and three species Aplidium filiforme Kott, 1992, Aplidium lunacratum Kott, 1992 and Aplidium macrolobatum Kott, 1992 of the family Polyclinidae as new records to Indian waters. The total number of Aplidium species reported from Indian water could be raised from three to seven. Key to the species of genus Aplidium recorded from India is given with detailed descriptions of all the new records. Keywords: Compound ascidian, Aplidium digitalis n.sp., Aplidium filiforme , Aplidium lunacratum , Aplidium macrolobatum , Gulf of Mannar.

INTRODUCTION

The subphylum Urochordata of phylum Chordata includes sac like marine animals called Sea squirts or ascidians. They are cosmopolitan in distribution found from the shallow intertidal rocky shores to the depth of the ocean. About 2800 species of ascidians have been reported from different parts of the world 1. India lags far behind the rest of the world in taxonomical study of ascidians and only a meager number of 86 species has been described in detail. From the Gulf of Mannar waters ten families of

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zzOn the.... Meenakshi and Senthamarai. ascidians have been reported 2. Among these, the family Polyclinidae includes compound ascidians with usually massive colonies. Their zooids are relatively large, with seven or more rows of stigmata and the body divided into a thorax, abdomen and post abdomen. From Tuticorin coast, three genera of the family Polyclinidae - Polyclinum, Sidnyum and Aplidium has been reported. The genus Polyclinum is represented by six species - Polyclinum madrasensis 3, Polyclinum indicum 4, Polyclinum constellatum 5, Polyclinum fungosum 6, Polyclinum nudum 7, Polyclinum saturnium 5; Sidnyum by one species - Sidnyum pentatrema 8 and Aplidium by three species - Aplidium multiplicatum9, Aplidium brevilarvacium10 and Aplidium distaplium7, so far 11-14 . In the present study a survey of ascidians from the Gulf of Mannar waters was carried out and identified up to the species level in order to add them to the faunistic list of Indian ascidians.

MATERIALS AND METHODS

The methodology suggested by Dr. Patricia Kott, Queensland Museum, Australia was followed for collection, narcotization, preservation and identification 15 . Collections were done from mussel landing centers, trawl discards and by SCUBA diving from the Gulf of Mannar waters along the south east coast of India. The collected samples were narcotised with a few crystals of menthol and fixed in a mixture of 40% formaldehyde and sea water in the ratio 1:10. A dissecting microscope was used to observe the entire surface of the colony. The zooids of the individual colony was separated and accurately identified. Diagrams were drawn with the help of Camera Lucida for interpretation of the results. Identification of the collected ascidians to the species level was carried out based on the key to identification of ascidians2.

RESULTS AND DISCUSSION

The present survey adds one species Aplidium digitalis n.sp., as new to science and three more species Aplidium filiforme 7, Aplidium lunacratum 7 and Aplidium macrolobatum 7 as new records of the genus Aplidium to Indian waters. The diagnostic characters of all the species has been described in detail with suitable plates, figures and key to identification. Systematic position: Phylum: Chordata; Subphylum: Urochordata; Class: Ascidiacea; Order: Enterogona; Suborder: Aplousobranchia; Family: Polyclinidae; Genus: Aplidium .

APLIDIUM DIGITALIS N.SP.

[Figure - 1 (A to F), Plate - 1, 2] Occurrence: This species was collected from mussel landing centre of Melakurumpanai, Kanniyakumari District. Distribution: India. Holotype: ( AS 912) deposited in the ascidian collections of the Museum of the Department of Zoology, A.P.C. Mahalaxmi College for Women, Tuticorin, Tamilnadu, India. Description: Colony is massive and gelatinous, 2 cm long, dome shaped cushion attached by the base on to the surface of mussel shell. Sand particles are present on the outer surface of the colony. Test is transparent without any embedded sand. It consists of a large number of opaque pigment cells. Zooids can be easily removed from the soft test. Outer surface of the test is wrinkled. Systems are circular. At the place of the common cloacal openings, sand particles are very small. The distance between common cloacal openings is 3 mm. Faecal pellets were not observed. Colonies are sandy brown in colour.

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zzOn the.... Meenakshi and Senthamarai.

A B C

E

D F

Figure 1: Aplidium digitalis n.sp. A. thorax, B. thorax with abdomen, C. post-abdomen showing gonads with gonoduct, D. post-abdomen showing gonads with gonoduct, E. post-abdomen showing tip, F. larva. Scale: A to F - 0.12 mm

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Plate : 1. Aplidium digitalis n.sp., Colony 2. Aplidium digitalis n.sp., Larva

Zooids are narrow, thread like, orange in colour and measure about 1 cm long. In preservative the colour fades. Thorax and abdomen are equal in length. Some opaque cells are scattered in the zooids. Posterior abdomen is very long. The branchial siphon is 6 lobed. The lobe varies in shape, flat or triangular. There is a thick sphincter muscle band at the base of branchial siphon. Atrial opening is wide in first row and atrial lip separated from the rim of the opening. Short atrial lip is bifid and finger shaped. 7 thick muscle bands arise from the branchial sac and extend to the post-abdomen as long ventral bands on each side. Transverse muscles are thick in between the stigmatal rows. The prebranchial area is wide. Neural gland opens in a funnel shaped opening. Branchial sac is long and narrow. There are 10 rows of stigmata with 9 per row. Stigmata are very small and oval. Branchial papilla is small tongue shaped. Length of the oesophagus and position of the stomach varies. It is situated either in the middle or posterior end of abdomen. Stomach has 24 to 26 deep longitudinal folds. Duodenum is short and wide. The anal opening is opposite to the 6 th or 7 th row of stigmata. Female follicle is in the anterior end of the posterior abdomen and single row of male follicles are arranged behind the female follicle. The posterior abdomen is long. The posterior end of the zooid is bifid. Larvae are present in the peribranchial cavity. They are oval in shape. There are 3 long wide ampullae at the anterior end. Adhesive organs are large and cup shaped. Anterior end of the adhesive organ is cushion like. The tail is wound round the head three fourth the way. Remarks: This species resembles A. multiplicatum in the gelatinous nature of the test but differs from it in having dome shaped colonies, presence of a thin layer of sand on test and entire stomach folds. A. brevilarvacium and A. filiforme have lobed colonies compared to massive cushions in the present species. A. distaplium can be easily distinguished by 4 rows of stigmata and 7 stomach folds. . A. lunacratum has only 5 stomach folds whereas the present species has 24-26 folds. A. digitalis n.sp. differ from A. macrolobatum in having dome shaped colonies, zooid with narrow thorax and greater number of stomach folds. The special features of the present species are massive dome shaped colonies, gelatinous transparent test with pigment cells and absence of faecal pellets. Systems are circular and the arrangement of zooids is marked by sand particles. The presence of a narrow thorax with 10 rows of stigmata, 9 per row and an atrial lip separate from rim of aperture with a bifid finger like tip are also characteristics of this species. There are 24-26 uninterrupted stomach folds and a bifid posterior abdomen. In view of the above features the present species was treated as new and named Aplidium digitalis .

APLIDIUM FILIFORME 7 KOTT, 1992

[Figure - 2 (A & B), Plate - 3] Aplidium thomsoni :10 Kott, 1963, p. 97.

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zzOn the.... Meenakshi and Senthamarai. Aplidium filiforme 7 Kott, 1992, p. 542.

A B

Figure 2: Aplidium filiforme A. thorax with abdomen, B. post-abdomen,

Plate 3: Aplidium filiforme Larva

Occurrence: This species was collected from trawl discards of Sadaimudiyanvalasai, Ramanathapuram District (AS 1646). Distribution: Australia, India. Description: The colony consists of upright, flat topped sandy lobes that rise from the surface of a common basal test mass. They form a sandy cushion up to 3 mm in diameter on the upper surface of each lobe. Zooids are arranged in circular systems with one common cloacal opening. Each system consists of a wide cloacal aperture in the centre surrounded by branchial apertures. Sand is absent around the cloacal openings. Systems are usually quite conspicuous, even when sand is present on the upper surface. The arrangement of sand is distinct with small particles surrounded by large. Test is firm and transparent. Small orange pigment cells and prochloron cells are sparsely distributed in the test. Faecal pellets were not observed. Colony is sandy orange in colour. Zooids are transparent, minute, thread like about 2.6 mm long with 0.4 mm thorax, 0.5 mm abdomen and 1.7 mm post-abdomen. Branchial siphon is short with 6 triangular lobes. Atrial aperture is opposite to the first row of stigmata with a large flat atrial lip from the rim of the aperture. Both the apertures are sessile without sphincter muscles. About 20 longitudinal muscle bands extend from 2716 J. Chem. Bio. Phy. Sci. Sec. B; Aug. 2013-Oct.2013; Vol.3, No.4; 2712-2721.

zzOn the.... Meenakshi and Senthamarai. thorax to the post-abdomen. Transverse muscles are present in the thorax. The branchial sac is long and narrow. Prebranchial area is wide with crescent shaped neural gland and slit like opening of dorsal tubercle. Tentacles are numerous, small and uniformly distributed at the base of branchial siphon. Branchial sac is with 8-9 rows of long and narrow stigmata 12-13 per row. Oesophageal neck is very long and tube like. Short stomach has 16 parallel longitudinal folds. Duodenum and post stomach are wide in nature. The smooth anal opening is opposite to the 3 rd row of stigmata. Posterior abdomen is long, cylindrical in shape with rounded tip. Only male follicles were found in the posterior abdomen arranged in double rows throughout. 3 to 5 larvae of different stages of development were observed in the peribranchial cavity. They are oval in shape with 4 long ampullae and 3 median cushion like adhesive organ at the anterior end. Tail is wound half way round the head. Remarks: The present species resembles A. brevilarvacium in the presence of lobed colonies, atrial lip from the rim of aperture but differs in having only 8 rows of sitgmata. A. distaplium differs from A. filiforme in the nature of colony and in the presence of only 4 rows of stigmata. A. lunacratum is distinguished from the present species by the 5 stomach folds. A. macrolobatum has massive sheet like colonies compared to lobed colonies in the present species. A. digitalis n.sp. has gelatinous colonies, zooids with greater number of stomach folds (24-26), a bifid atrial lip and posterior end of post-abdomen. The colony from Indian water resembles A. filiforme Kott, 1992 in having sandy lobes, wide common cloacal aperture, atrial lip from the rim of aperture and long oesophageal neck but it differs in having lesser stigmatal rows, number per row and lesser stomach folds.

APLIDIUM LUNACRATUM 7 KOTT, 1992

[Figure - 3 (A to D), Plate - 4] Psammaplidium ordinatum 16 Herdman and Riddell, 1913, p. 885. Psammaplidium ordinatum :10 Kott, 1963, p. 99. Aplidium lunacratum 7 Kott, 1992, p. 558. Occurrence: This species was collected from a depth of 3-4 meters from Tiruchendur, Tuticorin District. (AS 1942) Distribution: Australia, Victoria, New South Wales, India. Description: Colonies are massive with stalked heads, irregular and sandy measuring 3 cm X 4 cm. Sand is found throughout the colony uniformly. Crowded and relatively large sand grains adhere to the ridges between the systems and usually to the outside of the colony. Common cloacal apertures are on small protuberance surrounded by a shallow concave area. Zooids arranged in circular system. Small round common cloacal apertures are scattered 0.75 - 1 cm apart. Colour in life and preservative are sandy. They are tough and brittle being attached by the stalked head to the surface of mussel shell. Pigment cells are in the branchial and atrial aperture and siphons. Algal cells and faecal pellets were not observed. Zooids are arranged in compartments of the circular system perpendicular to each other. The test is firm and transparent. Zooids are 4 mm long when relaxed with 2 mm post-abdomen. Grey colour pigment cells were observed on the surface of the branchial and atrial aperture. The remaining part of the zooid is transparent in nature. The branchial aperture is terminal having 6 fringed branchial lobes on a dome shaped branchial siphon. The atrial aperture is small situated from the anterior end opposite to the 5th row of the stigmata. Small rim of sphincter muscle is on the branchial and atrial siphon. A flat atrial lip with divided tip extends out a little from the upper rim at the level of 3 rd row of the stigmata. About 20 longitudinal muscle bands extend from the thorax. Transverse muscles are present. Stigmata 2717 J. Chem. Bio. Phy. Sci. Sec. B; Aug. 2013-Oct.2013; Vol.3, No.4; 2712-2721.

zzOn the.... Meenakshi and Senthamarai. are in 11-12 rows of 11-12 per row arranged in long and wide branchial sac. The prebranchial area is wide. Numerous simple long branchial tentacles are distributed uniformly. Endostyle extends from the anterior end to the posterior end of the thorax. Branchial papillae are absent. Oesophagus is long tube like. The stomach is small narrow with 5 deep folds situated in the middle of the abdomen. A short duodenum and a posterior stomach are present. Bilobed anal opening is at the level opposite to the 9 th row of the stigmata. The posterior abdomen is very long without gonads in the present specimen and the tip of the post-abdomen is smooth rounded and paddle shaped. No larvae were observed.

A B C D

Figure 3: Aplidium lunacratum , A. thorax, B. abdomen, C. post-abdomen, D. tip of post-

abdomen. Scale: A to D - 0.12 mm

Plate 4: Aplidium lunacratum , Colony

Remarks: The present species resembles A. digitalis n.sp. in the presence of atrial lip separate from the rim of aperture. It differs from A. filiforme and A. brevilarvacium in having only five stomach folds and more stigmatal rows but exhibits similarities in the nature of lobed colonies. A. multiplicatum which has already been reported from India can be distinguished by the absence of sand particles in the test. The present species also differs from A. distaplium in having more number of rows of the stigmata and A. macrolobatum which has an atrial lip from the rim of aperture. The colony from Indian waters resembles A. lunacratum Kott, 1992 in having circular system, common cloacal apertures on prominances, atrial lip separate from aperture and 5 stomach folds but it differs in having lesser stigmatal row and number per row. 2718 J. Chem. Bio. Phy. Sci. Sec. B; Aug. 2013-Oct.2013; Vol.3, No.4; 2712-2721.

zzOn the.... Meenakshi and Senthamarai. APLIDIUM MACROLOBATUM 7 KOTT, 1992

[Figure - 4 (A to D), Plate - 5] Aplidium lobatum :10 Kott, 1963, p. 97 (part, colonies from Heron Island and Sarina). Aplidium macrolobatum 7 Kott, 1992, p. 561.

A B

C D

Figure 4: Aplidium macrolobatum A. thorax, B. thorax with abdomen, C. post-abdomen showing gonads, D. tip of post-abdomen. Scale: A to D - 0.22 mm

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Plate 5: Aplidium macrolobatum, Colony

Occurrence: This species was collected from Mussel landing centre of Keezhakurumpanai, Kanniyakumari District (AS 1362). Distribution: Australia, India. Description: Colonies are large, thick sand embedded investing sheet up to 12 cm X 5 cm size and black in colour. The test includes sand particles and shell pieces. Colony attached by the basal flat surface to the mussel shell. The outer surface of the test has larger sand particles while the embedded sand is of small size. Margin of colony extend into rounded and flattened lobes. Common cloacal apertures are large, depressed from the surface with circular or elongated oval openings. Around the system fine sand particles are present. The depth of the common cloacal opening is 3 mm. Test is thick, hard, glassy and gelatinous with brown colour pigment cells and faecal pellets. Zooids are embedded in the test and arranged in circular systems. Green colour prochloron cells are present in the test and upper surface of zooids. Zooids are transparent, minute thread like and about 0.5 to 1 cm long. Thorax and abdomen are equal in length and posterior abdomen is long. Branchial siphon is long with 6 triangular lobes. Atrial lip with trilobed tip extends out from the upper rim of atrial aperture. Narrow muscle bands are present in the atrial lip. Sphincter muscles are present in branchial and atrial siphons. Six pairs of strong longitudinal muscle bands present in thorax which extend to the middle of the post-abdomen. Thick strong transverse muscle bands extend in between the stigmatal rows. Prebranchial space is narrow. Opening of the dorsal tubercle is funnel shaped. Dorsal lamina is straight and smooth. Many smooth, curved tentacles are present at the base of branchial siphon. Wide and short branchial sac having 7 to 10 rows of stigmata with 15 per row. Stigmata are long and narrow. Small branchial papillae and internal longitudinal vessels present. Oesophagus is long. Stomach wall has 12 deep parallel longitudinal folds. Abdomen has short duodenum and long rectum. Anus opens at the base of the sixth or seventh row of stigmata. Atrial siphon situated at the base of the first stigmatal row. Peribranchial area is packed with two or three fertilized eggs. Male follicles are arranged in the long posterior abdomen in double rows. Round female follicles are present in the post-abdomen. Tip of the posterior abdomen is small finger like. No larva was observed. Remarks: The present species resembles A. brevilarvacium in the nature of colony and presence of algal cells in the test. A. distaplium differs in having only 4 rows of stigmata and 7 stomach folds. A. filiforme can be distinguished by the lobed colonies, smaller zooids and narrow thorax. A. lunacratum has massive colonies with stalked head, protruding common cloacal cavity and only 5 stomach folds. A. digitalis n.sp., differs from the present species in having dome shaped gelatinous colonies and 24- 26 stomach folds. The colony from Indian water resembles A. macrolobatum 7 Kott, 1992 in all respects except for the difference in colony size and stomach folds.

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zzOn the.... Meenakshi and Senthamarai. KEY TO THE SPECIES OF APLIDIUM RECORDED FROM INDIA 1. Sand embedded in test ………………………………………………………………….… 2 2. Sand not embedded in test ……………………...……………………..….. A. multiplicatum 3. Stigmata in 4 rows ……………………………………………….…….…….. A. distaplium 4. Stigmata in more than 4 rows …………………………………..……..…………...……… 3 5. Five deep stomach folds …………..……...……………….……...…………. A. lunacratum 6. More than five stomach folds ………………………………..………..……….………..… 4 7. Atrial lip separate from the rim of aperture …………...... …… A. digitalis n.sp. 8. Atrial lip from the rim of aperture ……………………..……..………………..…..……… 5 9. Atrial lip trilobed ……..…………………..………………….…………... A. macrolobatum 10. Atrial lip flat …………………………………………………………….………………… 6 11. Algal cells present in the test ……………..…….………………...…….. A. brevilarvacium 12. Algal cells absent in the test ………………………...………………………….. A. filiforme ACKNOWLEDGEMENT

The authors express their sincere gratitude to the University Grants Commission, New Delhi F. No. 39-588/2010(SR) for financial assistance and to Dr. T.K. Renganathan, Retired Professor, Department of Zoology, V.O.Chidambaram College, Tuticorin, for introducing to the field ascidian systematics.

REFERENCES

1. G. Lambert, Canadian Journal of Zoology, 2005, 83 , 34-50. 2. V.K. Meenakshi, T.K. Renganathan, S. Senthamarai and J. Jayalakshmi, Marine Biodiversity – Taxonomy of Indian Ascidians. Final Technical Report of Major Research Project submitted to the Ministry of Environment and Forests, New Delhi, 2003, 1-37. 3. V.O. Sebastian, Current Science, 1952, 21 , 316-317. 4. V.O. Sebastian, Washington Academy of Science, 1954, 44 (1), 18-24. 5. J.C. Savigny, Systeme de la classe des Ascidies, in Memoirs sur les-animaux sans vertebres . (Paris), 1816, 1-239. 6. W.A. Herdman, Report on the Scientific Results of the Voyage of HMS Challenger, 1886, 14 (38), 1-425. 7. P. Kott, Memoirs of Queensland Museum, 1992, 32 (2), 375-620. 8. C. Monniot, Bulletin du Museum national d’Histoire Naturelle , Paris, 1972, 61, 939-948. 9. C.P. Sluiter, Siboga Expedition , 1909, 56, 1-112. 10. P. Kott, Australian Journal of Marine and Freshwater Research , 1963, 14 (1), 70-118. 11. R. Krishnan, M.R. Chandran and T.K. Renganathan, Geobios new Reports , 1989, 8, 70-74. 12. V.K. Meenakshi, Journal of Marine Biological Association of India , 1998, 40 , 201-205. 13. T.K. Renganathan, Geobios new Reports , 1984, 3, 155-156. 14. V.K. Meenakshi and S. Senthamarai, European Journal of Zoological Research , 2012, 1, 65-69. 15. P. Kott, Memoirs of Queensland Museum, 1985, 23, 1-440. 16. W.A. Herdman and W. Riddell, Memoirs of the Australian Museum, 1913, 4(7), 873-889.

Corresponding author: V.K. Meenakshi; Department of Zoology, A.P.C. Mahalaxmi College for Women, Tuticorin,

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