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Venom glands in scatophagid fish
Tox7eon, 1970, Vol. 8, pp. 171-178. Pergamon Press. Printed in Great Britain.
VENOM GLANDS IN SCATOPHAGID FISH ANN M. CADiIItON and R. ENDEAN Department of Zoology, University of Queensland, Brisbane, Australia (Acceptedjorpublication 29 July 1969) Abstract-Venom glands have been demonstrated in scatophagid fishes for the first time. Six specimens of Scatopitaigus argus, from 62 mm to 296 mm in standard length, possessed a pair of venom glands accommodated in paired antero-lateral grooves in each fin spine. The venomglands ofthe larger specimenswere shorter than those ofthe smallerSshes. In Selertotoca multifasciata specimens up to 98 mm in standard length possessed venom glands but glands were lacking in specimens greater than approximately 140 mm in standard length. In both species scales extended relatively further distally along the fin spines in larger specimens than in smaller specimens. The glands of both species were aggregations of large gland cells in the thickened epidermis of the integumeatary sheath which filled the spine grooves. The glands were not encapsulated in connective tissue sheaths. Elongate supporting cells occurred among the venom gland cells some of which possessed unusual rod-like bodies in their cytoplasm. INTRODUCTION
HALSTeAn and MITCI~LI. (1963) listed the family Scatophagidae among those fishes reputed to be venomous . However, no anatomical or histological demonstrations of venom glands in scatophagids have been made . There are only two genera in the family, Scatophagus Cuvier and Valenciennes and Selenotoca Myers. Although some species of Scatophagus are popular aquarium fishes and considered harmless (INNFS,1946), it is noteworthy that aborigines of the Northern Territory, Australia, refer to these fish as `poison bone' (WHITLEY, 1963). Scatophagus argus (Linnaeus, 1766) is said to inflict more painful wounds than do allied species (M ARCFiAi .L, 1964). S. argus (Fig . 1) is a widely ranging species of the Indo-west Pacific region (WEBER and nE BEAUFORT, 1936) and, in Australia, has been recorded from the Northern Territory, Queensland and New South Wales (MARSHALL, 1964). In southern Queensland it is known as the spotted butter-fish and grows to a total length of 32 cm (MARSHALL, 1964). It is common knowledge in Queensland that the fin spines of Selerwtoca multifasciata (Richardson,1844) can cause painful wounds, references to which have appeared in popular works (`MEn>co', 1950 ; MARSHALL, 1964 ; GRANT, 1965). An envenomation involving this species was recorded by WHITLEY (1963) . Some Queensland fishermen are of the opinion that small specimens of S. multifasclata sting more severely than do larger specimens. This species (Fig . 2) ranges from New Guinea, the Aru Islands and New Caledonia to Queensland and New South Wales (WESERR and DE BEAUFORT, 1936). It is netted frequently in bays, estuaries and fresh water streams. Its common names include butter-fish and Johnny Dory and it grows to a total length of 41 cm (MARSHALL, 1964). It was decided to ascertain whether a venom apparatus is posse.4sed by adult andjuvenile specimens of Scatophagus argus and Selenotoca multifasciata . 171
17 2
ANN M. CAMERON and R. ENDFAN MATERIALS AND METHODS
Juvenile and adult specimens of Scatophagus argus were obtained from Cairns, north Queensland and juvenile and adult specimens of Selenotoca multifasciata were obtained from the Tweed River near the border between New South Wales and Queensland. Measurements were made of standard length and the lengths of undamaged fin spines for each specimen examined . Several specimens of both species were anaesthetized with ethyl carbamate and each undamaged fin spine, together with its investing integumentary sheath, was cut through near its articulation using bone cutters. Each spine and its associated sheath was then fixed for times which ranged from 1 to 24 hr in one ofthe following fixatives Baker's formol calcium, Bouin's fluid, Bouin-Duboscq fixative, Heidenhain's `Susa' fixative, Zenker acetic fixative, Zenker formol fixative and basic lead acetate-formaline fixative. Subsequently, the tissues were washed, decalcified, washed, dehydrated, cleared, embedded, sectioned and dyed as described in an earlier paper (CAMERON and Erro~r+N, 1966). RESULTS
Scatophagus argus Specimens ranged in size from 62 mm to 296 mm in standard length and each possessed 11 dorsal, 4 anal and a pair of ventral fin spines . Each spine possessed a pair of lateral grooves, one on each side of the spine . No other traumatizing organ which might form part of a venom apparatus was detected . Serial transverse sections of many stings revealed the following structures in each : a T-shaped spine embedded in dermal collagenous connective tissue, a basement membrane and an epidermis of stratified epithelium, thickened in the region of the spine grooves . Dermal scales were present in the proximal parts of the integumentary sheath but did not extend to the distal end ofany spine . However, they extended relatively further distally in larger specimens than they did in smaller specimens . Aggregations of gland cells occurred in the epidermis filling the spine grooves (Fig. 3). Because of their position and organization these aggregations were considered to be venom glands and the individual gland cells, venom-producing cells (p.177). The dimensions ofthe two glandular aggregations associated with each spine were measured from serial transverse sections and those of the longer gland recorded . The distance between the distal end of the gland and the tip of the spine was also noted in each case. The ratio of venom gland length to spine length for a total of 69 glands from six specimens had a mean value of 020 and ranged from 001 to 045 . The longest gland measured was 10,360 ~. in length and was associated with the fifth dorsal spine of a specimen 125 mm in standard length . It was 041 times the length of the spine bearing it. The shortest gland measured was 340 ~ in length and was associated with the eighth dorsal spine of the 296 mm standard length specimen. It was 001 times the length of the spine . There was no consistent relationship between the longest gland possessed by a specimen and the serial order of the spine bearing it. The distal terminations of venom glands varied in their distance from the distal ends of the spines bearing them. This distance ranged from 10 to 3230 ~.. All the stings sectioned possessed venom glands . From Table 1 it can be seen that the larger specimens possessed relatively shorter glands than did the smaller specimens . Measurements made of gland width and gland depth within the spine groove indicated that the venom glands were wedge-shaped in cross section and possessed tapered ends, the greatest cross-sectional area being approximately half-way along their lengths . In all cases the extent of the venom glands along the spines was limited proximally by the encroachment of scales (Fig. 4).
Venom Glands in Scatophagid Fish
173
TAHLa 1. VENOMGLAND LHNGTHS AND RATI03 OF GLAND LENGTH TO SPINS LENGTH IN 6 SPSCIMHNS of S. argus
Standard length of specimen (mm) 62 Spine
80
D, Dio Dii Al A, As A, L.V .
R.V. Mean ratio Range in ratio Standard deviation
125
211
296
Gland Gland Gland Gland Gland Gland length Ratio length Ratio length Ratio length Ratio length Ratio length Ratio
(~)
D, D, D, D, D, D, D, De
96
(w)
1880 2900 3550 ?A00 4420 3890 3310 3280 2410 1160 1820 2540 2990 2190 580 3390
0"38 Oß4 026 0"18 Oß5 Oß4 0"30 Oß3 0"28 018 0"28 0"36 0"37 0"27 0"09 027
029
(~)
2710 5380 6820 5910 5450 5270 4120 3810 3120 2]80 3360 4770 3770 3180 5030 4880
0"39 0"45 0"40 Oßl Oß0 0"31 027 029 0"28 0"24 0"37 040 0" 31 027 0"28 027
(w)
2260 2920 3840 3870 5190 3340 4230 2330 2010 1770 760 2250 2140 2400 1700 3380 3340
0"32
0"25 021 0" 19 018 026 0" 19 023 0" 17 017 0" 16 006 025 0"19 0" 20 0"12 0"18 0" 19
(w)
400 5890 10360 8400 2050 2950 3950
0,04 0"18 0"41 Oß9 0"10 0"16 0"15
(r~)
5570 4360 4890 3010 3840 4390
0"15 0"11 018 0" 12 0"20 0"11
980 4020 3000 3190 340 1740 2410
0"19
0.20
0"15
0"06
006 009 006 0"07 0"01 0"05 0"OS
009-0ß8
0"24-0"45
006-0"26
004--0"41
0" 11-0"20
001-0~09
0"08
006
0"OS
0"14
0"04
0"02
Dl _ ll indicates dorsal spines ; A l..., indicates anal spines ; L.V . indicates left ventral spine; R.V. indicates right
ventral spine; - indicates stingnot sectioned.
The venom gland cells were irregularly shaped but tended to be columnar in the deepest portion of a gland, abutting on the groove in the spine which accommodated them (Fig. 5). Deep in the spine groove no basal layer of germinal cells separated the venom-producing cells from the basement membrane of the epidermis but such a basal layer was present elsewhere . Each venom cell possessed a spherical nucleus which was centrally situated. These nuclei were difficult to colour with both Delafield's haematoxylin and Weigert's iron haematoxylin . Mature venom cell cytoplasm coloured red with eosin, yellow-green with Lillie's allochrome stain, orange to red with Mallory's triple stain, deep blue with mercuric bromphenol blue reagent and yielded a weakly positive PAS reaction. The cytoplasm appeared homogenous after each of the fixatives employed and no granular secretory products were observed. However, angular rod-like bodies (Fig. 5) occurred in some of the venom cells. They coloured with dyes as did the venom cell cytoplasm but more intensely so. They yielded a strongly positive reaction for protein with mercuric bromphenol blue reagent but were PAS negative . Among the venom cells were elongate epidermal cells with cigar-shaped nuclei (Fig. ~. These will be termed supporting cells. Stages in the development of venom cells from small, undifferentiated epidermal cells possessing basophilic cytoplasm were apparent in some regions of the glands, particularly
174
ANN M . CAMERON and R . ENDEAN
at their proximal and distal ends. As the epidermal cells increased in size they lost their affinity for haematoxylin and acquired, progressively, an affinity for eosin. Irregularly shaped gland cells with a granule-packed cytoplasm were distributed sparsely in the epidermis peripheral to the venom glands ofsome spines . The cytoplasm of these cells responded to dyes as did that ofthe venom cells but always less intensely so. In some sections, spherical to pear-shaped goblet cells were observedin theepidermis. Theircytoplasmcoloured pale blue with Mallory's stain, faint blue after the mercuric bromphenol blue test for protein and was PAS-positive . It was less eosinophilic than the venom cells cytoplasm. At the outer edge ofthe epidermis some of these goblet cells were observed to be discharging their secretions via short necks. Measurements on all the cell types described are listed in Table 2 . TABLE 2. MEASItrefluflNTS ON THE VENOM CELLS, THE GRANULAR CELLS, THB MUCOU3 CELLS AND THE 3UPPORTINO CELLS OF Scatopl4agus argus Measurement
Venom cell long axis Venom cell short anis Venom cell nucleus diameter Long axis of rod Short axis of rod Supporting cell nucleus long axis Supporting cell nucleus short axis Granular cell long axis Granular cell short axis Mucous cell long axis Mucous cell short axis
measurements
No . of
Mean (~)
20 20 20 l8 18 10 10 20 20 10 10
31 13 3~9 12~2 3~8 7~1 1~7 23 16 8~1 6~5
Range (rs)
20-46 8-20 3~3- 3~7 4~9-17~9 1~6- 6~5 4~9-11~4 1~1- 2~4 14-39 9~8-21 6~5- 9~8 4~3- 9~8
S.D.
7~41 3~42 0~21 5~30 1~40 3~81 0~11 7~13 2~71 0"74 0~70
Selenotoca multifasciata Specimens examined ranged from 44 to 343 mm in standard length . Each possessed twelve dorsal, four anal and a pair of ventral fin spines . Each fin spine possessed a pair of lateral grooves, one on each side . No other traumatizing organ which might form part of a venom apparatus was detected . Scales were present in the integumentary sheaths of all fin spines and this squamipinniform condition extended relatively further distally in large specimens than in smaller ones. Serial transverse sections of stings revealed paired glandular
FIG . 3. PHOTOMICROGRAPH OF A TRANSVERSE SECTION OP THE FIFTH DORSAL STING OF A SPECIMEN OF S. atglL4 ôO mm IN STANDARD LENGTH . Formol calcium fixation, 10 t', dyed with haematoxylin and eosin . Yellow filter. x 173. FIG. S. PHOTOMICROGRAPH OF PART OF A TRANSVERSE SECTION OF THE NINTA DORSAL STING OF A SPECIMEN OF S. argaS 125 mm IN STANDARD LENGTH SHOWRVG CYTOPLASMIC RODS IN THE VENOM CELLS . Bouin fixation, 10 tc, dycd with haematoxylin and eosin. x 168. FIG . 6. PHOTOMICROGRAPH OF A TRANSVERSE SECTION OF THE SECOND DORSAL STING OF A SPECIMEN of S. nlult~fasciata 98 mm nv sTANDARD LENGTH. Bouin fixation, l0 h, dyed with haematoxylin and eosin . x 173.
Fra. 7. PHOTOMICROGRAPH OF PART OF THE SAME BECTION AS IN FYG. 6, sHOwING crroPLASxuc RCDS irr THE vENOM CELLS . x 168.
FICr . 1 . LIVING SPECIMEN OF SCOIOp%!Qg[IS OI'gUS 97 Illill IN STANDARD LENGTH . FIG . 2 . LIVING SPECIMEN OF
Selexoloca mullifasciata 63
Yellow filter.
I17Ili IN STANDARD LENGTH .
Tox. f.p .
174
FIG . H .
PHOTOMICROGRAPH OF PART OF THE SAME SECTION AS IN FIG . G, SHOWING GOBLET CELLS PERIPHERAL TO THE VENOM GLAND .
Venom Glands in Scatophagid Fish
17 5
1mm
FIG.
~i.
SBMI-DIAGRAMMATIC REPRESENTATION OF THE EXTENT OF THE VHNOM GLANDS ON A TYPICAL SPINE OF S. Q7g(L4 AND THE P09rITON OP THE SCALES IN THE INTEGUMENTARY SHEATH.
A=transverse section near the distal end of the spine showing twin venom glands and no scales ; B=transverse section in the mid region of the venom glands showing scales in the integumentary sheath ; C=transverse section proximal to one venom gland; D=diagram showing unequal extent of the paired venom glands on a spine 17 mm long and the positions of the sections A, B and C. The scale line refers to D only. The dashed line indicates the distal extent of the scales. d=dermis ; ep=epidermis ; gl=venom gland; sc=scale ; sp=spine.
17 6
ANN M. CAMERON and R. ENDFAN
aggregations, regarded as venom glands (p.177) associated with all the fin spines ofspecimens 50, 51, 61 and 98 mm in standard length . No such glands were detected in any of the fin spines possessed by specimens 141, 153 and 218 mm in standard length . Essentially, the arrangement of the venom apparatus was similar to that of Scatophagus argus. The longest gland measured was one on the eighth dorsal spine of a specimen 98 mm in standard length . The ratio of the length of this gland to that of the spine was 031 . The ratio of gland length to spine length ranged from 006 to 031 . As can be seen from Fig. 6 the presence of an aggregation of closely packed gland cells distinguished the stratified epithelium filling each of the spine grooves from that elsewhere in the integumentary sheath . In transverse section this aggregation was fan-shaped (Fig. 6). The venom cells were irregular in shape although they tended to be columnar in the deepest portion of the gland abutting on the groove in the spine which accommodated them. They possessed spherical nuclei, each with a prominent nucleolus . Elongated angular bodies, termed rods, were contained in the cytoplasm of some of the venom cells (Fig. 7). Usually, only one rod was present in a gland cell but, occasionally, as many as four were noted in a single cell. The longest rod measured was virtually as long as the long axis of the venom cell containing it. Data on venom cell size and rod size are presented in Table 3. TABLE 3 . DIMENSIONS OF VENOM CELLS, THEIR NUCLEI AND THEIR RODS IN THE VENOM GLANDS OF A SPECIMEN OF S . 7RüItlfQSC(Q10 9H mm IN STANDARD LENGTH
Measurement
No. of measurements
Venom cell long axis Venom cell short axis Venom cell nucleus diameter Long axis of rod Short axis of rod
10 10
8
11
9
Mean (~) 290 131 3~7 186 2~7
Range (~)
S.D .
16-39 9-16 2~9~~2 3~3-299 1 ~5-3~3
659 233 039 983 066
Venom cell cytoplasm was eosinophilic the rods being more eosinophilic than the cytoplasm surrounding them. With Mallory's triple stain the venom cell cytoplasm coloured variously from orange to purple but the rods were invariably coloured bright red with this stain . After exposure to mercuric bromphenol blue reagent the venom cell cytoplasm was coloured deep blue and the rods even more deeply so . Venom cell cytoplasm was faintly PAS-positive but the rods were PAS-negative. Among the venom cells were elongate supporting cells with cigar-shaped to ovoid nuclei (Fig. 7). These nuclei measured 6~5 ~, by 1 ~3 to 2~5 ,~. Developmental stages of the venom cells from small, undifferentiated epidermal cells were observed . The cytoplasm of such undifferentiated cells was haematoxylinophilic . As the cells increased in size they lost their cytoplasmic alilnity for haematoxylin and coloured with eosin . Rods, which always coloured more intensely than the surrounding cytoplasm, appeared in some cells. Among the outermost layers of epidermal cells, peripheral to the wedge ofvenom gland cells, were spherical to pear-shaped goblet cells (Fig. 8) with a mean diameter of 14~5~.. They ranged in diameter from 8~0 to 26 ~ and s=492 for 11 measurements . The cytoplasm of the goblet cells was less eosinophilic than that of the venom cells. It was coloured pale blue with Mallory's triple stain, faintly blue after the mercuric bromphenol blue test for proteins was employed and was PAS-positive.
Venom Glands in Scatophagid Fish DISCUSSION
17 7
All teleost venom glands so far described are aggregations of large epidermal gland cells (clavate cells) among which elongate supporting cells occur (PAWLOWSKY, 1913, 1914, 1927; REED, 1924; TANGE, 1957). This organization is consistent regardless of whether the aggregations are isolated within spine grooves by dermal connective tissue capsules as in the Scorpaenidae (CArsIItON arid ENDEAN, 1966), or are merely thickenings of the epidermal portion of the integumentary sheath with concentration of clavate cells in the region of the fin spine, as in catfish such as Galeichthysfells (Hn>:.srEnD et al., 1953). Clavate cells occur in a wide variety of forms but are always readily distinguishable from mucus-producing cells, another major glandular component of teleost epidermis. All the available data suggest that teleost venom gland cell cytoplasm is of a basic proteinaceous nature (ENDEAN, 1961 ; CAMERON and ENDEAN, 1966). In both Selenotoca multifasciata and Scatophagus argus there are aggregations of large epidermal gland cells, comparable with the clavate cells of other teleosts and occupying twin grooves in each fin spine. These aggregations are not encapsulated by dermal connective tissue capsules as are the venom glands of scorpaenids but their location in pronounced spine grooves is similar to that in the scorpaenid type of venom gland (CAMERON arid ENDEAN, 1966). The organization of the glandular aggregations, as thickenings of the epidermal portion of the integumentary sheath in the vicinity of the fin spines, is also similar to that of the venom glands of the catfish mentioned above. The clavate cell cytoplasm of both scatophagids is of a basic proteinaceous nature . The features described-disposition of glandular aggregations of large clavate cells with basic proteinaceous cytoplasm, separated by elongate supporting cells, in the vicinity of the fin spines-are concluded to represent avenom apparatus in each ofthe scatophagids studied. The possession of such a venom apparatus would account for the painful wounds which the fin spines are capable of causing. In Selenotoca multifasciata possession of a venom apparatus is confined to juveniles of a size group (presumably related to age) the upper limit of which lies between 98 and 141 mm in standard length. Although only six specimens of Scatophagus argus were studied, the data suggest that fish larger than those examined may not possess venom glands, or if they do, that such glands may be of lesser extent than those described (Table 1). It may be significant that both species are squamipinniform and this condition is not confined to the soft parts of the fins but extends along the spines . The presence of scales in the integumentary sheaths of the spines would probably impede the release of venom into punctures caused by the spines . Also, it was noted that in both specimens the scales extend relatively further distally in larger fish than they do in smaller specimens . Their relatively smaller size exposes juvenile fish of any species to greater potential predation than that to which the relatively larger adults are subjected. Venom glands are a possible defence mechanism against predation of such juveniles. Presumably, the nonvenomous adults of a species with venomous juveniles are defended by other mechanisms including their increased body size. The condition in Selenotoca multifasciata (and probably in Scatophagus argus also) parallels that in a species of surgeonfish, Acanthurus triostegus sandvicensis (family Acanthuridae). RANDALL (1961) found that the second dorsal, second anal and the pelvic spines of the acronurus larvae of this species can inflict stings on man similar to bee stings . He postulate3 that this venomous condition is a larval adaption because it is lost on the third day of transformation . Of great interest are the cytoplasmic rods of the venom cells of Selenotoca multifasciata
178
ANN M. CAMERON and R. ENDEAN
and Scatophagus argus. Because of their remarkably angular shape it was thought initially that they were artefacts. However, they were subsequently shown to be present in glands fixed in different fixatives and in glands of fish of different sizes. They appear to be proteinaceous elaborations of the cytoplasm but more data are required to resolve their present enigmatic nature. It would probably be instructive to compare their ultrastructure with that of other proteinaceous cytoplasmic crystals, for example those in frog egg cytoplasm which attain a long axis of the order of 7 tc \ "" ARD, 1962) and those which occur in the interstitial cells of human testis and attain measurements of20 by 2-3 ~ (FAwcEZ-r and BURGOS, 1960). Worthy of attention is the fact that the Chaetodontidae, of which the scatophagids are sometimes considered a sub-family (WESER and DE BEAUFORT, 1936), are also listed by HAISTEAD and MITCHELL (1963) among fish suspected to be venomous . It would be of interest to examine chaetodontids for the presence of a venom apparatus. Acknowledgements-The authors wish to thank Mr. D. McCoLM and Mr . W. $rAHLUM for photographing the living scatophagids illustrated in this paper and are indebted to Mr . N. DEAN for supplying a specimen of Scatopht~qus argus. The work was supported by a grant from the University of Queensland Research Grants Committee.
REFERENCES CAMERON, A. M. and ENDEAN, R. (1966) The venom apparatus of the scorpion fish Notesthes robrrsta . Toxlcon 4, 111 . ENDEAN, R. (1961) A study of the distribution, habitat, behaviour, venom apparatus and venom of the stonefish . Aust . J. nwr. Freshwat . Res. 12, 177. FAWCErT, D. W. and BvRCos, M. H. (1960) Studies on the fine structure of the mammalian testis-II. The human interstitial tissue. Am. J. Anat . 107, 245. GRANT, E. M. (1965) Guide to Fishes . Brisbane : S. G. Reid. HAL91'EAD, B. W., KvNtxosu, L. S. and HESARn, H. G. (1953) Catfish stings and thevenom apparatus of the Mexican catfish Galeichthysjells (Linnaeus) . Trans. Am . mittust. Soc. 72, 297. HALSTEAD, B. W. and MrrcBELL, L. R. (1963) A review of the venomous fishes of the Pacific area . In : Venomaus arrd Poisonous Animals and Noxious Plants of the Pacific Region, p. 173 (KEEGAN, H. L. and MACFARLANE, W. V., Eds.). Oxford : Pergamon Press. INNES, W. T. (1946) Exotic Aquarium Fishes, 7thed. Philadelphia : Innes. MARSBALL, T. C. (1964) Fishes of the Great Barrier Reef and Coastal Waters of Qrreensland. Sydney : Angus and Robertson. `MEDICO' (1950) Fish wounds and first aid. Fisherman's Luck and Fair Sport 1, 4. PAwtowstc3r, E. N. (1913) Sur la structure des glandes a venin de certains poisons et en particulier de celles de Plotosus . C. r. Sfanc. Soc. Blol . 74, 1033 . PAWLOWSKY, E. N. (1914) Ù'ber den Bau der Giftdrûsen bei Plotosus und anderen Fischen. Zool. Jb. 38, 427. PAWLOWSKY, E. N. (1927) G{fttiere und ihrer Gljtigkeit. Jena : G. Fischer. RANDALL, J. E. (1961) A contribution to the biology of the convict surgeonfish of the Hawaiian Islands, Acanthurus triostegus sandvlcensis. Pacif. Sct. 15, 215. REED, H. D. (1924) The morphology of the dermal glands in nematognathous fishes . Z. Morph . Antimop. 24, 227. TANGE, Y. (1957) Beitrag zur Kenntnis der Morphologie des Giftapparates bei den japanischen FischenXVI. Zusammenfassende Betrachtung über den Giftapparat. Yokohama n:ed. Bull. 8, 62 . WARD, R. T. (1962) The origin ofprotein and fatty yolk in Rann piplens-II. Electron microscopical and cytochemical observations of young and mature ot5cytes . J. Cell Biol. 14, 309. WEesR, M. and DE BeauFORr, C. F. (1936) The Fishes of the Indo-Australian Archipelago-VII, Perciformes (continued). Leiden : E. J. Brill. WBtrLEV, G. P. (1963) Dangerous Australian fishes . Supplement to Bull. Post-Graduate Comrrr . Med. Unlv, Sydney 18 Suppl. 41 .
VENOM GLANDS IN SCATOPHAGID FISH ANN M. CADiIItON and R. ENDEAN Department of Zoology, University of Queensland, Brisbane, Australia (Acceptedjorpublication 29 July 1969) Abstract-Venom glands have been demonstrated in scatophagid fishes for the first time. Six specimens of Scatopitaigus argus, from 62 mm to 296 mm in standard length, possessed a pair of venom glands accommodated in paired antero-lateral grooves in each fin spine. The venomglands ofthe larger specimenswere shorter than those ofthe smallerSshes. In Selertotoca multifasciata specimens up to 98 mm in standard length possessed venom glands but glands were lacking in specimens greater than approximately 140 mm in standard length. In both species scales extended relatively further distally along the fin spines in larger specimens than in smaller specimens. The glands of both species were aggregations of large gland cells in the thickened epidermis of the integumeatary sheath which filled the spine grooves. The glands were not encapsulated in connective tissue sheaths. Elongate supporting cells occurred among the venom gland cells some of which possessed unusual rod-like bodies in their cytoplasm. INTRODUCTION
HALSTeAn and MITCI~LI. (1963) listed the family Scatophagidae among those fishes reputed to be venomous . However, no anatomical or histological demonstrations of venom glands in scatophagids have been made . There are only two genera in the family, Scatophagus Cuvier and Valenciennes and Selenotoca Myers. Although some species of Scatophagus are popular aquarium fishes and considered harmless (INNFS,1946), it is noteworthy that aborigines of the Northern Territory, Australia, refer to these fish as `poison bone' (WHITLEY, 1963). Scatophagus argus (Linnaeus, 1766) is said to inflict more painful wounds than do allied species (M ARCFiAi .L, 1964). S. argus (Fig . 1) is a widely ranging species of the Indo-west Pacific region (WEBER and nE BEAUFORT, 1936) and, in Australia, has been recorded from the Northern Territory, Queensland and New South Wales (MARSHALL, 1964). In southern Queensland it is known as the spotted butter-fish and grows to a total length of 32 cm (MARSHALL, 1964). It is common knowledge in Queensland that the fin spines of Selerwtoca multifasciata (Richardson,1844) can cause painful wounds, references to which have appeared in popular works (`MEn>co', 1950 ; MARSHALL, 1964 ; GRANT, 1965). An envenomation involving this species was recorded by WHITLEY (1963) . Some Queensland fishermen are of the opinion that small specimens of S. multifasclata sting more severely than do larger specimens. This species (Fig . 2) ranges from New Guinea, the Aru Islands and New Caledonia to Queensland and New South Wales (WESERR and DE BEAUFORT, 1936). It is netted frequently in bays, estuaries and fresh water streams. Its common names include butter-fish and Johnny Dory and it grows to a total length of 41 cm (MARSHALL, 1964). It was decided to ascertain whether a venom apparatus is posse.4sed by adult andjuvenile specimens of Scatophagus argus and Selenotoca multifasciata . 171
17 2
ANN M. CAMERON and R. ENDFAN MATERIALS AND METHODS
Juvenile and adult specimens of Scatophagus argus were obtained from Cairns, north Queensland and juvenile and adult specimens of Selenotoca multifasciata were obtained from the Tweed River near the border between New South Wales and Queensland. Measurements were made of standard length and the lengths of undamaged fin spines for each specimen examined . Several specimens of both species were anaesthetized with ethyl carbamate and each undamaged fin spine, together with its investing integumentary sheath, was cut through near its articulation using bone cutters. Each spine and its associated sheath was then fixed for times which ranged from 1 to 24 hr in one ofthe following fixatives Baker's formol calcium, Bouin's fluid, Bouin-Duboscq fixative, Heidenhain's `Susa' fixative, Zenker acetic fixative, Zenker formol fixative and basic lead acetate-formaline fixative. Subsequently, the tissues were washed, decalcified, washed, dehydrated, cleared, embedded, sectioned and dyed as described in an earlier paper (CAMERON and Erro~r+N, 1966). RESULTS
Scatophagus argus Specimens ranged in size from 62 mm to 296 mm in standard length and each possessed 11 dorsal, 4 anal and a pair of ventral fin spines . Each spine possessed a pair of lateral grooves, one on each side of the spine . No other traumatizing organ which might form part of a venom apparatus was detected . Serial transverse sections of many stings revealed the following structures in each : a T-shaped spine embedded in dermal collagenous connective tissue, a basement membrane and an epidermis of stratified epithelium, thickened in the region of the spine grooves . Dermal scales were present in the proximal parts of the integumentary sheath but did not extend to the distal end ofany spine . However, they extended relatively further distally in larger specimens than they did in smaller specimens . Aggregations of gland cells occurred in the epidermis filling the spine grooves (Fig. 3). Because of their position and organization these aggregations were considered to be venom glands and the individual gland cells, venom-producing cells (p.177). The dimensions ofthe two glandular aggregations associated with each spine were measured from serial transverse sections and those of the longer gland recorded . The distance between the distal end of the gland and the tip of the spine was also noted in each case. The ratio of venom gland length to spine length for a total of 69 glands from six specimens had a mean value of 020 and ranged from 001 to 045 . The longest gland measured was 10,360 ~. in length and was associated with the fifth dorsal spine of a specimen 125 mm in standard length . It was 041 times the length of the spine bearing it. The shortest gland measured was 340 ~ in length and was associated with the eighth dorsal spine of the 296 mm standard length specimen. It was 001 times the length of the spine . There was no consistent relationship between the longest gland possessed by a specimen and the serial order of the spine bearing it. The distal terminations of venom glands varied in their distance from the distal ends of the spines bearing them. This distance ranged from 10 to 3230 ~.. All the stings sectioned possessed venom glands . From Table 1 it can be seen that the larger specimens possessed relatively shorter glands than did the smaller specimens . Measurements made of gland width and gland depth within the spine groove indicated that the venom glands were wedge-shaped in cross section and possessed tapered ends, the greatest cross-sectional area being approximately half-way along their lengths . In all cases the extent of the venom glands along the spines was limited proximally by the encroachment of scales (Fig. 4).
Venom Glands in Scatophagid Fish
173
TAHLa 1. VENOMGLAND LHNGTHS AND RATI03 OF GLAND LENGTH TO SPINS LENGTH IN 6 SPSCIMHNS of S. argus
Standard length of specimen (mm) 62 Spine
80
D, Dio Dii Al A, As A, L.V .
R.V. Mean ratio Range in ratio Standard deviation
125
211
296
Gland Gland Gland Gland Gland Gland length Ratio length Ratio length Ratio length Ratio length Ratio length Ratio
(~)
D, D, D, D, D, D, D, De
96
(w)
1880 2900 3550 ?A00 4420 3890 3310 3280 2410 1160 1820 2540 2990 2190 580 3390
0"38 Oß4 026 0"18 Oß5 Oß4 0"30 Oß3 0"28 018 0"28 0"36 0"37 0"27 0"09 027
029
(~)
2710 5380 6820 5910 5450 5270 4120 3810 3120 2]80 3360 4770 3770 3180 5030 4880
0"39 0"45 0"40 Oßl Oß0 0"31 027 029 0"28 0"24 0"37 040 0" 31 027 0"28 027
(w)
2260 2920 3840 3870 5190 3340 4230 2330 2010 1770 760 2250 2140 2400 1700 3380 3340
0"32
0"25 021 0" 19 018 026 0" 19 023 0" 17 017 0" 16 006 025 0"19 0" 20 0"12 0"18 0" 19
(w)
400 5890 10360 8400 2050 2950 3950
0,04 0"18 0"41 Oß9 0"10 0"16 0"15
(r~)
5570 4360 4890 3010 3840 4390
0"15 0"11 018 0" 12 0"20 0"11
980 4020 3000 3190 340 1740 2410
0"19
0.20
0"15
0"06
006 009 006 0"07 0"01 0"05 0"OS
009-0ß8
0"24-0"45
006-0"26
004--0"41
0" 11-0"20
001-0~09
0"08
006
0"OS
0"14
0"04
0"02
Dl _ ll indicates dorsal spines ; A l..., indicates anal spines ; L.V . indicates left ventral spine; R.V. indicates right
ventral spine; - indicates stingnot sectioned.
The venom gland cells were irregularly shaped but tended to be columnar in the deepest portion of a gland, abutting on the groove in the spine which accommodated them (Fig. 5). Deep in the spine groove no basal layer of germinal cells separated the venom-producing cells from the basement membrane of the epidermis but such a basal layer was present elsewhere . Each venom cell possessed a spherical nucleus which was centrally situated. These nuclei were difficult to colour with both Delafield's haematoxylin and Weigert's iron haematoxylin . Mature venom cell cytoplasm coloured red with eosin, yellow-green with Lillie's allochrome stain, orange to red with Mallory's triple stain, deep blue with mercuric bromphenol blue reagent and yielded a weakly positive PAS reaction. The cytoplasm appeared homogenous after each of the fixatives employed and no granular secretory products were observed. However, angular rod-like bodies (Fig. 5) occurred in some of the venom cells. They coloured with dyes as did the venom cell cytoplasm but more intensely so. They yielded a strongly positive reaction for protein with mercuric bromphenol blue reagent but were PAS negative . Among the venom cells were elongate epidermal cells with cigar-shaped nuclei (Fig. ~. These will be termed supporting cells. Stages in the development of venom cells from small, undifferentiated epidermal cells possessing basophilic cytoplasm were apparent in some regions of the glands, particularly
174
ANN M . CAMERON and R . ENDEAN
at their proximal and distal ends. As the epidermal cells increased in size they lost their affinity for haematoxylin and acquired, progressively, an affinity for eosin. Irregularly shaped gland cells with a granule-packed cytoplasm were distributed sparsely in the epidermis peripheral to the venom glands ofsome spines . The cytoplasm of these cells responded to dyes as did that ofthe venom cells but always less intensely so. In some sections, spherical to pear-shaped goblet cells were observedin theepidermis. Theircytoplasmcoloured pale blue with Mallory's stain, faint blue after the mercuric bromphenol blue test for protein and was PAS-positive . It was less eosinophilic than the venom cells cytoplasm. At the outer edge ofthe epidermis some of these goblet cells were observed to be discharging their secretions via short necks. Measurements on all the cell types described are listed in Table 2 . TABLE 2. MEASItrefluflNTS ON THE VENOM CELLS, THE GRANULAR CELLS, THB MUCOU3 CELLS AND THE 3UPPORTINO CELLS OF Scatopl4agus argus Measurement
Venom cell long axis Venom cell short anis Venom cell nucleus diameter Long axis of rod Short axis of rod Supporting cell nucleus long axis Supporting cell nucleus short axis Granular cell long axis Granular cell short axis Mucous cell long axis Mucous cell short axis
measurements
No . of
Mean (~)
20 20 20 l8 18 10 10 20 20 10 10
31 13 3~9 12~2 3~8 7~1 1~7 23 16 8~1 6~5
Range (rs)
20-46 8-20 3~3- 3~7 4~9-17~9 1~6- 6~5 4~9-11~4 1~1- 2~4 14-39 9~8-21 6~5- 9~8 4~3- 9~8
S.D.
7~41 3~42 0~21 5~30 1~40 3~81 0~11 7~13 2~71 0"74 0~70
Selenotoca multifasciata Specimens examined ranged from 44 to 343 mm in standard length . Each possessed twelve dorsal, four anal and a pair of ventral fin spines . Each fin spine possessed a pair of lateral grooves, one on each side . No other traumatizing organ which might form part of a venom apparatus was detected . Scales were present in the integumentary sheaths of all fin spines and this squamipinniform condition extended relatively further distally in large specimens than in smaller ones. Serial transverse sections of stings revealed paired glandular
FIG . 3. PHOTOMICROGRAPH OF A TRANSVERSE SECTION OP THE FIFTH DORSAL STING OF A SPECIMEN OF S. atglL4 ôO mm IN STANDARD LENGTH . Formol calcium fixation, 10 t', dyed with haematoxylin and eosin . Yellow filter. x 173. FIG. S. PHOTOMICROGRAPH OF PART OF A TRANSVERSE SECTION OF THE NINTA DORSAL STING OF A SPECIMEN OF S. argaS 125 mm IN STANDARD LENGTH SHOWRVG CYTOPLASMIC RODS IN THE VENOM CELLS . Bouin fixation, 10 tc, dycd with haematoxylin and eosin. x 168. FIG . 6. PHOTOMICROGRAPH OF A TRANSVERSE SECTION OF THE SECOND DORSAL STING OF A SPECIMEN of S. nlult~fasciata 98 mm nv sTANDARD LENGTH. Bouin fixation, l0 h, dyed with haematoxylin and eosin . x 173.
Fra. 7. PHOTOMICROGRAPH OF PART OF THE SAME BECTION AS IN FYG. 6, sHOwING crroPLASxuc RCDS irr THE vENOM CELLS . x 168.
FICr . 1 . LIVING SPECIMEN OF SCOIOp%!Qg[IS OI'gUS 97 Illill IN STANDARD LENGTH . FIG . 2 . LIVING SPECIMEN OF
Selexoloca mullifasciata 63
Yellow filter.
I17Ili IN STANDARD LENGTH .
Tox. f.p .
174
FIG . H .
PHOTOMICROGRAPH OF PART OF THE SAME SECTION AS IN FIG . G, SHOWING GOBLET CELLS PERIPHERAL TO THE VENOM GLAND .
Venom Glands in Scatophagid Fish
17 5
1mm
FIG.
~i.
SBMI-DIAGRAMMATIC REPRESENTATION OF THE EXTENT OF THE VHNOM GLANDS ON A TYPICAL SPINE OF S. Q7g(L4 AND THE P09rITON OP THE SCALES IN THE INTEGUMENTARY SHEATH.
A=transverse section near the distal end of the spine showing twin venom glands and no scales ; B=transverse section in the mid region of the venom glands showing scales in the integumentary sheath ; C=transverse section proximal to one venom gland; D=diagram showing unequal extent of the paired venom glands on a spine 17 mm long and the positions of the sections A, B and C. The scale line refers to D only. The dashed line indicates the distal extent of the scales. d=dermis ; ep=epidermis ; gl=venom gland; sc=scale ; sp=spine.
17 6
ANN M. CAMERON and R. ENDFAN
aggregations, regarded as venom glands (p.177) associated with all the fin spines ofspecimens 50, 51, 61 and 98 mm in standard length . No such glands were detected in any of the fin spines possessed by specimens 141, 153 and 218 mm in standard length . Essentially, the arrangement of the venom apparatus was similar to that of Scatophagus argus. The longest gland measured was one on the eighth dorsal spine of a specimen 98 mm in standard length . The ratio of the length of this gland to that of the spine was 031 . The ratio of gland length to spine length ranged from 006 to 031 . As can be seen from Fig. 6 the presence of an aggregation of closely packed gland cells distinguished the stratified epithelium filling each of the spine grooves from that elsewhere in the integumentary sheath . In transverse section this aggregation was fan-shaped (Fig. 6). The venom cells were irregular in shape although they tended to be columnar in the deepest portion of the gland abutting on the groove in the spine which accommodated them. They possessed spherical nuclei, each with a prominent nucleolus . Elongated angular bodies, termed rods, were contained in the cytoplasm of some of the venom cells (Fig. 7). Usually, only one rod was present in a gland cell but, occasionally, as many as four were noted in a single cell. The longest rod measured was virtually as long as the long axis of the venom cell containing it. Data on venom cell size and rod size are presented in Table 3. TABLE 3 . DIMENSIONS OF VENOM CELLS, THEIR NUCLEI AND THEIR RODS IN THE VENOM GLANDS OF A SPECIMEN OF S . 7RüItlfQSC(Q10 9H mm IN STANDARD LENGTH
Measurement
No. of measurements
Venom cell long axis Venom cell short axis Venom cell nucleus diameter Long axis of rod Short axis of rod
10 10
8
11
9
Mean (~) 290 131 3~7 186 2~7
Range (~)
S.D .
16-39 9-16 2~9~~2 3~3-299 1 ~5-3~3
659 233 039 983 066
Venom cell cytoplasm was eosinophilic the rods being more eosinophilic than the cytoplasm surrounding them. With Mallory's triple stain the venom cell cytoplasm coloured variously from orange to purple but the rods were invariably coloured bright red with this stain . After exposure to mercuric bromphenol blue reagent the venom cell cytoplasm was coloured deep blue and the rods even more deeply so . Venom cell cytoplasm was faintly PAS-positive but the rods were PAS-negative. Among the venom cells were elongate supporting cells with cigar-shaped to ovoid nuclei (Fig. 7). These nuclei measured 6~5 ~, by 1 ~3 to 2~5 ,~. Developmental stages of the venom cells from small, undifferentiated epidermal cells were observed . The cytoplasm of such undifferentiated cells was haematoxylinophilic . As the cells increased in size they lost their cytoplasmic alilnity for haematoxylin and coloured with eosin . Rods, which always coloured more intensely than the surrounding cytoplasm, appeared in some cells. Among the outermost layers of epidermal cells, peripheral to the wedge ofvenom gland cells, were spherical to pear-shaped goblet cells (Fig. 8) with a mean diameter of 14~5~.. They ranged in diameter from 8~0 to 26 ~ and s=492 for 11 measurements . The cytoplasm of the goblet cells was less eosinophilic than that of the venom cells. It was coloured pale blue with Mallory's triple stain, faintly blue after the mercuric bromphenol blue test for proteins was employed and was PAS-positive.
Venom Glands in Scatophagid Fish DISCUSSION
17 7
All teleost venom glands so far described are aggregations of large epidermal gland cells (clavate cells) among which elongate supporting cells occur (PAWLOWSKY, 1913, 1914, 1927; REED, 1924; TANGE, 1957). This organization is consistent regardless of whether the aggregations are isolated within spine grooves by dermal connective tissue capsules as in the Scorpaenidae (CArsIItON arid ENDEAN, 1966), or are merely thickenings of the epidermal portion of the integumentary sheath with concentration of clavate cells in the region of the fin spine, as in catfish such as Galeichthysfells (Hn>:.srEnD et al., 1953). Clavate cells occur in a wide variety of forms but are always readily distinguishable from mucus-producing cells, another major glandular component of teleost epidermis. All the available data suggest that teleost venom gland cell cytoplasm is of a basic proteinaceous nature (ENDEAN, 1961 ; CAMERON and ENDEAN, 1966). In both Selenotoca multifasciata and Scatophagus argus there are aggregations of large epidermal gland cells, comparable with the clavate cells of other teleosts and occupying twin grooves in each fin spine. These aggregations are not encapsulated by dermal connective tissue capsules as are the venom glands of scorpaenids but their location in pronounced spine grooves is similar to that in the scorpaenid type of venom gland (CAMERON arid ENDEAN, 1966). The organization of the glandular aggregations, as thickenings of the epidermal portion of the integumentary sheath in the vicinity of the fin spines, is also similar to that of the venom glands of the catfish mentioned above. The clavate cell cytoplasm of both scatophagids is of a basic proteinaceous nature . The features described-disposition of glandular aggregations of large clavate cells with basic proteinaceous cytoplasm, separated by elongate supporting cells, in the vicinity of the fin spines-are concluded to represent avenom apparatus in each ofthe scatophagids studied. The possession of such a venom apparatus would account for the painful wounds which the fin spines are capable of causing. In Selenotoca multifasciata possession of a venom apparatus is confined to juveniles of a size group (presumably related to age) the upper limit of which lies between 98 and 141 mm in standard length. Although only six specimens of Scatophagus argus were studied, the data suggest that fish larger than those examined may not possess venom glands, or if they do, that such glands may be of lesser extent than those described (Table 1). It may be significant that both species are squamipinniform and this condition is not confined to the soft parts of the fins but extends along the spines . The presence of scales in the integumentary sheaths of the spines would probably impede the release of venom into punctures caused by the spines . Also, it was noted that in both specimens the scales extend relatively further distally in larger fish than they do in smaller specimens . Their relatively smaller size exposes juvenile fish of any species to greater potential predation than that to which the relatively larger adults are subjected. Venom glands are a possible defence mechanism against predation of such juveniles. Presumably, the nonvenomous adults of a species with venomous juveniles are defended by other mechanisms including their increased body size. The condition in Selenotoca multifasciata (and probably in Scatophagus argus also) parallels that in a species of surgeonfish, Acanthurus triostegus sandvicensis (family Acanthuridae). RANDALL (1961) found that the second dorsal, second anal and the pelvic spines of the acronurus larvae of this species can inflict stings on man similar to bee stings . He postulate3 that this venomous condition is a larval adaption because it is lost on the third day of transformation . Of great interest are the cytoplasmic rods of the venom cells of Selenotoca multifasciata
178
ANN M. CAMERON and R. ENDEAN
and Scatophagus argus. Because of their remarkably angular shape it was thought initially that they were artefacts. However, they were subsequently shown to be present in glands fixed in different fixatives and in glands of fish of different sizes. They appear to be proteinaceous elaborations of the cytoplasm but more data are required to resolve their present enigmatic nature. It would probably be instructive to compare their ultrastructure with that of other proteinaceous cytoplasmic crystals, for example those in frog egg cytoplasm which attain a long axis of the order of 7 tc \ "" ARD, 1962) and those which occur in the interstitial cells of human testis and attain measurements of20 by 2-3 ~ (FAwcEZ-r and BURGOS, 1960). Worthy of attention is the fact that the Chaetodontidae, of which the scatophagids are sometimes considered a sub-family (WESER and DE BEAUFORT, 1936), are also listed by HAISTEAD and MITCHELL (1963) among fish suspected to be venomous . It would be of interest to examine chaetodontids for the presence of a venom apparatus. Acknowledgements-The authors wish to thank Mr. D. McCoLM and Mr . W. $rAHLUM for photographing the living scatophagids illustrated in this paper and are indebted to Mr . N. DEAN for supplying a specimen of Scatopht~qus argus. The work was supported by a grant from the University of Queensland Research Grants Committee.
REFERENCES CAMERON, A. M. and ENDEAN, R. (1966) The venom apparatus of the scorpion fish Notesthes robrrsta . Toxlcon 4, 111 . ENDEAN, R. (1961) A study of the distribution, habitat, behaviour, venom apparatus and venom of the stonefish . Aust . J. nwr. Freshwat . Res. 12, 177. FAWCErT, D. W. and BvRCos, M. H. (1960) Studies on the fine structure of the mammalian testis-II. The human interstitial tissue. Am. J. Anat . 107, 245. GRANT, E. M. (1965) Guide to Fishes . Brisbane : S. G. Reid. HAL91'EAD, B. W., KvNtxosu, L. S. and HESARn, H. G. (1953) Catfish stings and thevenom apparatus of the Mexican catfish Galeichthysjells (Linnaeus) . Trans. Am . mittust. Soc. 72, 297. HALSTEAD, B. W. and MrrcBELL, L. R. (1963) A review of the venomous fishes of the Pacific area . In : Venomaus arrd Poisonous Animals and Noxious Plants of the Pacific Region, p. 173 (KEEGAN, H. L. and MACFARLANE, W. V., Eds.). Oxford : Pergamon Press. INNES, W. T. (1946) Exotic Aquarium Fishes, 7thed. Philadelphia : Innes. MARSBALL, T. C. (1964) Fishes of the Great Barrier Reef and Coastal Waters of Qrreensland. Sydney : Angus and Robertson. `MEDICO' (1950) Fish wounds and first aid. Fisherman's Luck and Fair Sport 1, 4. PAwtowstc3r, E. N. (1913) Sur la structure des glandes a venin de certains poisons et en particulier de celles de Plotosus . C. r. Sfanc. Soc. Blol . 74, 1033 . PAWLOWSKY, E. N. (1914) Ù'ber den Bau der Giftdrûsen bei Plotosus und anderen Fischen. Zool. Jb. 38, 427. PAWLOWSKY, E. N. (1927) G{fttiere und ihrer Gljtigkeit. Jena : G. Fischer. RANDALL, J. E. (1961) A contribution to the biology of the convict surgeonfish of the Hawaiian Islands, Acanthurus triostegus sandvlcensis. Pacif. Sct. 15, 215. REED, H. D. (1924) The morphology of the dermal glands in nematognathous fishes . Z. Morph . Antimop. 24, 227. TANGE, Y. (1957) Beitrag zur Kenntnis der Morphologie des Giftapparates bei den japanischen FischenXVI. Zusammenfassende Betrachtung über den Giftapparat. Yokohama n:ed. Bull. 8, 62 . WARD, R. T. (1962) The origin ofprotein and fatty yolk in Rann piplens-II. Electron microscopical and cytochemical observations of young and mature ot5cytes . J. Cell Biol. 14, 309. WEesR, M. and DE BeauFORr, C. F. (1936) The Fishes of the Indo-Australian Archipelago-VII, Perciformes (continued). Leiden : E. J. Brill. WBtrLEV, G. P. (1963) Dangerous Australian fishes . Supplement to Bull. Post-Graduate Comrrr . Med. Unlv, Sydney 18 Suppl. 41 .