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HEREDITARY BASIS OF MELANOSIS INHYBRID FISHES 1
MYRON GORDON
National Research Council Fellow in the Zoological ScitmC68, Cornell University I
In reviewing the work on the generic hybrids between Platypoecilus maculatus and X iphophorus hellerii, one is confronted byevidence which seems to point to a definite correlation betweenmelanotic conditions in the hybrid fishes and their characteristiccolor pattern. In some cases, normal and pathological hybridsoccur together in the same brood, born and reared in the sameaquarium. But it is only in hybrids of certain specific colorpatterns that melanosis appears. Since the various color patternshave been shown to be heritable, the evidence seems to indicatethat heredity plays an important r6le in determining the physicalcondition of each generic hybrid.
Abnormal conditions in certain hybrids were recognized by theearly fish hybridizers, Losslein (1912) and Haffner (1912). It isonly recently (1928) that a confirmatory note, with respect tothe origin of the pathological conditions arising under controlledbreeding practices, has been presented by Haussler, writing fromthe Institut fUr experimentelle Krebsforschung, Heidelberg. Hisresults and interpretation agree in the more important detailswith those of Kosswig (1928, 1929).
Experiments in hybridization have been in progress at CornellUniversity since 1925, but the major part of the genetic work has
1 This is the second of a series of three papers, the first of which appeared in thepreceding iBBUe of THE AMERICAN JOURNAL OF CANCER (15: 732, April 1930). Becauseof the close interrelationship of these papers, the figures are numbered consecutivelythroughout, so that there may be in this paper an occasional reference to figures appearingin the other two papers. The third of the series follows directly after this.
I The work upon which this paper is based was conducted in the laboratories ofAnimal Biology at Cornell University. It was supported in part by a grant from theHeckscher Foundation for the Advancement of Research. The writer is indebted toDr. H. D. Reed, Dr. A. C. Fraser, and Dr. G. C. Embody for permission to use some ofthe data, contained in this report, which was obtained during the period of his &8llOCiationwith them as Heckscher Research Assistant.
Published as Paper No. 175, Department of Plant Breeding, Cornell University,Ithaca, New York.
1495
1496 MYRON GORDON
been concerned with the study of inheritance of sex and colorpattern factors in Platypoecilus (Gordon 1927, 1928j Fraser andGordon 1928, 1929).
While it is not the purpose of this communication to present afinished story of the generic hybrids and the origin of melanosis,it seems desirable, at this time, to call the attention of Americanworkers to these phenomena. Interpretations of results will beoffered as far as available data permit. In many instances the dataare admittedly inadequate as a basis for conclusive statements.
It may be of interest to note that Platypoecilu8 maculaiu« andXiphoph01'U8 hellerii belong to closelyrelated genera (Hubbs 1924).Possibilities for their mating in nature seem to be entirely open;both species have been taken by Meek (1904) in the waters of theRio Papaloapan system, which drains the Vera Cruz area ofMexico. The only known hybrids, however, have had theirorigin in aquaria.
MATERIALS
A pair of Xiphophorus helleriiand several pairs of Platypoecilu8maculatus, of various strains, were purchased, early in 1925, froma breeder of tropical aquarium fishes. The stocks were multipliedby inbreeding j some of the offspring were used in the intergenericcrosses, while others were used for crosses between strains.
METHODS
1. Breeding ProcedureThe procedure for mating Xiphophorus with Platypoecilus is
quite simple, and no different from that of mating fishes of thesame species. Because of the viviparous method of reproductionin each, it is necessary merely to place the two sexes, each from adifferent genus, together in an aquarium. Practical fish breedershold that virgin fishes only may be induced to produce generichybrids. While the truth of the claim has not been tested, thepolicy has been practised here. On several occasions, however,fishes which had been used for generic crosses first have afterwardsbeen mated successfullywith those of their own species.
The gestation period for generic hybrids is slightly longer andmore variable than for fishes produced from crosses within eitherspecies. In Platypoecilus it is about thirty days. A detailedaccount of the methods used in propagating viviparous tropicalaquarium fishes has been written by Gordon (1926).
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1497
S. Use of Known Genetic RacesFrom a long series of intervarietal crosses, it has been possible
to isolate eight different genetic races in Platypoecilus. Fordetails, the reader may be referred to papers by Gordon (1927,1928) and Fraser and Gordon (1929). Each genetic race ofPlatypoecilus, with one exception, has been crossed with Xiphophorus in order to determine which color variety is concerned withmelanosis in the hybrids. Some races tested in these experimentshave been previously used by other workers (Table I).
3. Morphologic Study of Oolor PatternsMter the eight different genetic races of Platypoecilus had
been isolated by genetic methods of breeding and selection, astudy was made of the structural color elements which distinguisheach genetic race. All the types may be distinguished on the basisof the presence and distribution of two kinds of color-bearing cells:melanophores and erythrophores (see Table III).
In an extended study of the morphology of the heritable colorpatterns in Platypoecilus and Xiphophorus, the writer (1929)analyzed all the parental types which had been used in these intergeneric crosses. (An abridged summary appears in Table III.)These data on the morphology of the normal color types may beused as a standard in the detection of the deviations in the heritablepattern characters known to be possessed by the hybrids. In all,1333 sexually mature fishes (including 282 spotted and 169 blackPlatypoecilus) were observed; yet in none of these was there anyindication of melanosis such as characterizes melanotic hybrids.
ANALYSIS OF DATA
1. Relation of Color Pattern to Physical Oondition of HybridIn Tables I and II an attempt has been made to include a
summary of all published work on the hybridization of Xiphophorus and Platypoecilus. An analysis of the results of both fishfanciers and geneticists reveals the fact that whenever the Pulchraor Rubra varieties of Platypoecilus are used in a generic cross,melanosis develops in some of the hybrid offspring. When theNigra (black-banded variety) Platypoecilus is used, the resultinggeneric hybrids have more melanin pigment than either parent,but there are no definite neoplastic growths. If, however, theseblack F1 generic hybrids are back-crossed to Xiphophorus or bredamong themselves (inbred), melanosis appears in the offspring of
.... ~ QD
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1916
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,...
.B
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1922
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....
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1922
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....
....
....
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....
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1927
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11.
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....
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12.
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1928
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....
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....
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1929
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....
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6·
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....
....
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8·
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ids
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Spe
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....
....
....
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....
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anos
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24.
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....
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ipho
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....
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....
....
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ater
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-fin
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ott
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enty
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29.
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....
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....
....
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30.
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rap
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....
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-fin
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id..
....
....
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19
28
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ids
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anot
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see
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31.
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orus
...•
••.
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-fin
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id.•
....
•..
....
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19
28
·In
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32.
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ty..
..R
ed-f
inhy
brid
....
....
....
.G
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.1
92
8·
No
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ease
inre
dpi
gmen
t33
.R
edp
laty
....
....
.R
ed-f
inhy
brid
....
....
....
.G
ordo
n..
.1
92
9·
No
incr
ease
inde
gree
ofpi
gmen
tati
on
Fl
Hyb
rids
Inb
red
toG
etth
eS
econ
dG
ener
atio
n
34.
Nig
raFl
....
....
..N
igra
Fl..
....
....
....
....
Bel
lam
y..
1922
Mel
anot
icof
fspr
ing
seen
35.
Nig
raF
l...
....
...
Nig
raFl
....
....
....
....
..K
o88W
ig..
1928
Mel
anot
icof
fspr
ing
seen
36.
Red
-fin
Fl..
....
...
Red
-fin
Fl..
....
....
....
...
Gor
don
...
19
29
·N
opa
thol
ogic
alco
ndit
ione
;se
eP
late
XII
I
•R
efer
sto
yea
rof
expe
rim
ent.
no
tof
publ
icat
ion.
tNum
bers
inpa
rent
hese
sre
fer
toF
lhy
brid
sof
Tab
leI.
1500 MYRON GORDON
the next generation (Kosswig 1929). On the other hand, whenvarieties other than Pulchra, Rubra and Nigra are used, no comparable pathologic conditions appear in generic hybrids.
The question then arises, what are the factors involved in thePulchra, Rubra and Nigra varieties which give rise to pathologicalgeneric hybrids? The answer may be gained by a study of themorphology and genetics of pigmentary equipment in these fishes.
2. Consideration of the Color Patterns in PlatypoecilusIn 1927 the writer showed, genetically and morphologically,
that there were two kinds of melanophores as well as erythrophoresin the Rubra platy. The pigmentary elements which are responsible for the large black spotted condition in the Rubra platyare macromelanophores (Sp); while the micromelanophores (St)
TABLE III
Morphological Analysis of Color Patterns in Platypoecilus
Melanophores Erythrophore.
Genetic Race of PlatypoecilusMacromelano- Micromelano- Rjo Rophores, Spots phores, Stipplcs Red fins Red body
1. *Gold platy........ , ... , .. o, - - Rf -2. * Red platy ................. - - Rf R3. * Stipple platy.......... , .... - St Rf -4. Stipple, crescent............ - St Rf -5. * Stipple, one-spot ..... 0 •••• 0 - St Rf -6. * Stipple, twin-spot ...... 0 ••• - St Rf -7. *Red spotted 0 •••••••••••••• Sp - Rf R8. Nigra ................. 0 ••• Sp St Rfor rf -9. Pulchra ................. 00 Sp St Rf or rf -
10. *Rubra .................... Sp St Rf R
• Indicates that this type was used in a generic cross at the Cornell laboratory (seeTable IV).
are the small black pigment cells responsible for the olivaceouscolor and stippling of immature and female Rubras. Micromelanophores are present in male Rubras, also, but they arecovered over by erythrophores.
The Pulchras differ from the Rubras in that they lack the dominant genetic factor for red body color (R); erythrophores arepresent in the dorsal and part of the caudal fin, indicating thepresence of the factor Rf for red fins. Some are recessive forred fins (rf).
The Nigras have practically the same pigmentary equipment as
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1501
the Pulchras except that the black color elements are arranged ina different manner. The macromelanophores in the Pulchras andRubras are scattered irregularly over the entire body, while inNigras they are confined to a broad band on either side of the body.Bellamy has given the genetic symbol N to this character. According to Kosswig, Nigras possess erythrophores in the dorsal andcaudal fins (Rf). This variety also occurs without red fins (rJ).
The facts reviewed above are listed in Table III, items 8, 9and 10. This indicates that Rubras, Pulchras and Nigras possessthe following chromatophores: macromelanophores (Sp or N),micromelanophores (St) , and erythrophores in the dorsal andcaudal fins (Rf). Since all these varieties, when crossed withXiphophorus, give rise to melanotic hybrids, it may be assumed,tentatively, that the above factors, Sp, N, St and R], are concerned in melanosis which develops in the generic hybrids.
3. Consideration of the Factor for Red Body Color (R)
Owing to the fact that both Pulchras (r) and Rubras (R) giverise to melanotic hybrids, it appears that the differentiating factor(R) for erythrophores on the body proper is not concerned withdevelopment of melanosis. As further evidence of this, thefollowing data are offered.
A pure red platy (Fig. 8) differs from the Rubra platy (Fig. 16)in that it lacks both types of melanophores (Sp and St). Itsdevelopment as a distinct genetic race is traced by Fraser andGordon (1929). By crossing a red platy male with a Xiphophorusfemale (Plate X), thirteen red hybrids were obtained and rearedto maturity. In none of these was there the slightest indicationof melanosis; but the development of the red pigment cells inthe generic hybrids was accelerated. These cells appear earlier inthe young, and are more numerous in the female hybrids than inPlatypoecilus.
4. Consideration of the Factor for Red Fins (RJ)
To show that the red fin factor (Rf) may be eliminated fromconsideration in the development of melanosis, the followingevidence, gathered from experiments made independently in thislaboratory, is offered. It may be said that these data confirmthe work of Kosswig for the most part. The experiments areconcerned with the cross of the gold platy (red-finned) male and aXiphophorus female.
1502 MYRON GORDON
(a) Description oj the Gold Platy: The gold platy (Fig. 7) issharply contrasted with all other color varieties in that it lacksnot only both types of melanophores (sp, st), but also erythrophoreson the body proper (r). The entire body is covered with xanthophores, which account for its striking yellow color. In the dorsalfin and a small area immediately below it, as well as in the anteriorpart of the caudal fin, there will be found a considerable number oferythrophores. This indicates the presence of the factor for redfins (RI).
(b) Description oj the Pigmentary Pattern in XiphophoTUs (St'):The pigmentary system of Xiphophorus, as compared with that ofthe gold platy, is presented below. The Xiphophorus used here is
Xiphophorus female (St')1. Presence of the stipple complex
a. Integumentary melanophoresb. Subcutaneous melanophores
2. Mid-ventral black stripe3. Red zigzag lateral bands4. Green caudal fin sword in males5. Lack of the red-fin complex
Gold Platypoecilus male (RI)1. Lack of the stipple complex
2. Lack of mid-ventral black stripe3. Lack of lateral bands4. The species lacks a sword5. Presence of the red-fin complex:
a. Red dorsal finb. Red caudal finc. Red saddle on back
similar to those used in other generic crosses. The analysis ofthe color pattern in Xiphophorus is given at this time because astudy of the inheritance of its entire pattern may be traced tobetter advantage in the hybrid progeny originating from the crossof a gold platy with Xiphophorus than from those arising fromother types of generic crosses.
The integumentary melanophores of Xiphophorus are of thestipple type (St') , comparable to the micromelanophores ofPlatypoecilus (St). The subcutaneous melanophores refer topigment cells found about the blood vessels. In cleared specimensthese cells are seen to form a distinctive pattern of their own(Fig. 112), a pattern absent in the gold platy but found in a modified form in the stippled types of Platypoeeilus,
A faint but definite black stripe runs along the mid-ventralline of the female Xiphophorus from the anus to the tip of thecaudal fin. In the male this line runs continuously from the anusto the tip of the sword-like extension of its caudal fin, and thenback along the dorsal border of this extension (Fig. 112). Hausslerbelieves the mid-ventral line is involved in the development of
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1503
melanotic growths in some of the hybrid offspring which resultfrom crossing Xiphophorus with Platypoeeilus.
Several bands of erythrophores form a zigzag pattern on thesides of Xiphophorus, Some red cells, forming a row of red dots,may be found in the dorsal fins (Figs. 61, 63, 65, 67 and 69). Theerythrophores are never scattered over the body as they are inPlatypoecilus,
(c) The Generic Hybrids of the First Generation: Eight hybridswere obtained from the cross of the gold platy male and theXiphophorus female, three of which were males and five females(Plate X). All were fertile, and were used for other crosses.
These IIgolden" hybrids (Fig. 62) had the complete stipplepattern (St') of their Xiphophorus female parent. The midventral black stripe was also present in the hybrids, males ofwhich had the black border on the dorsal as well as on the ventraledge of the swords. The swords (Fig. 72) were much shorter thanthose found in the normal Xiphophorus. No significant modification, in point of view of pathologic conditions, was seen in thehybrids possessing the above characters.
The most striking thing about these generic hybrids is theextensive development of the red pigment cells. It would seemthat the red in the dorsal fin, the red in the caudal fin, and the redof the saddle (area beneath the dorsal fin), characteristic of thegold platy, is extended in every direction. The only region notaffected is that of the belly. The fact that the entire body ofthe hybrid is not all red is of great significance because it denotesthe differentiating element between the operation of the geneticfactor Rf for red fins and R for red body color. Even among theseII red extended" hybrids there was no indication of melanoticgrowths.
(d) The Hybrids Crossed Back to Original Species: These goldenhybrids were crossed back both to Xiphophorus and Platypoecilus.In the cross with Xiphophorus the red area was found to be stillfurther extended, but in the cross with Platypoecilus (R!) nofurther extension was detected. Kosswig's data indicate comparable results, except that in the cross back to Platypoecilus(1929d) he has detected two individuals, males, in which he seesthe beginning of red swellings. More information on these twomales is desirable.
(e) The Second Generation: Twenty individuals have been obtained by inbreeding the II golden" generic hybrids (Plate XIII).
72
1504 MYRON GORDON
They are extremely variable as to size and sexual maturity. Hereattention will be given to the heritable color patterns only, andtheir relation to melanotic conditions.
Fifteen of the twenty F2 fishes possess the complete stipplecomplex, St', and five lack it, st. This seems to indicate simplemendelian inheritance for the stippling factor of Xiphophorus(St'). Gordon (1927) has shown that the stippling pattern ofPlatypoecilus (St) is controlled by a single dominant autosomalfactor. Since the mid-ventral line of Xiphophorus is present inall the stippled F2 fishes and does not appear in the non-stippled,it seems that this character is closely associated with the stipplingcomplex, if, indeed, both are not capable of being referred to thesame genetic factor.
Only three individuals out of twenty lacked the red-fin complex(Rf) of Platypoecilus (Fig. 80). Seventeen had it in varyingdegrees of intensity (Figs. 81, 82, 83 and 84). The distribution of17 : 3 does not establish whether there are one or more geneticfactors contributing to its phenotypic expression.
In one of the gold (non-stippled) F2 hybrids (Fig. 83), a definitebut incomplete twin-spot pattern appeared. Since the fish,apparently a female, was sterile, it is impossible to say whetherthis is or is not a genetic change. More probably it is a smallsomatic change, since these patterns sometimes appear in straightcrosses of Platypoecilus, and when a breeding test is applied tothem, the new pattern does not reappear in their progeny.
In none of the twenty fishes was there the slightest indicationof melanosis. This leads to the view that there is nothing inthe genetic make-up of the gold platy which gives evidence of itscontributing to the development of melanotic overgrowths ingeneric hybrids. Experimental results similar to those describedabove have been presented by Kosswig (see Table II).
5. Consideration of the Factors for Patterns InvolvingM icromelanophores
All the varieties of Platypoecilus-Pulchra, Rubra and Nigrawhich have been involved in the production of melanotic neoplasms have been found to carry the factor for micromelanophores(St). The purpose here has been that of discovering whetherthe factor (St) for micromelanophores, and certain others relatedto it, are essential for the development of melanosis in generichybrids.
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1505
(a) The Stippled and Crescent Patterns: From the descriptionand figure of the Platypoecilus used by Gerschler (1914) in hisgeneric crosses, it is quite evident that he was dealing with astippled platy (St). This fish had an additional pattern upon itscaudal peduncle which Gerschler called H albmond, which is thecrescent (C) of American aquarists (Fig. 12). In the numerouscrosses which he made with the stippled platy (Fig. 15) andXiphophorus, no melanotic hybrids were detected in either thestippled (St) or the stippled plus crescent (St C) group.
(b) The Twin-spot Pattern: In this laboratory an intergenerichybrid resulting from the cross of the gold platy male and aXiphophorus female was crossed back to stippled platy (St) withthe twin-spot pattern (T) (Fig. 9). Eighteen back-cross progenyshowed stippling plus twin-spot (St, T), while fourteen showedstippling only (St, t). In neither of these two classes was therethe slightest indication of melanosis.
(c) The One-spot Pattern: In another back cross, to be describedmore fully below, a similar type of generic hybrid was crossed witha Rubra platy female. Among the offspring there were somethat lacked macromelanophores but which had the one-spotpattern, the factor for which was derived from the Rubra (PlateXII). In these one-spot hybrids (Figs. 73, 75) no melanoticconditions were detected.
(d) Kosswig's Pattern: Kosswig (1929a) mentions einen schwarzenFleck, which appears on the caudal peduncle of Platypoecilus.This pattern undergoes no changes when involved in a genericcross. Unfortunately Kosswig does not explain the structuralnature of this character, nor is a figure provided, so that it isimpossible to know which pattern is dealt with.
(e) Structural Basis of Pattern Formation: The caudal pedunclepatterns, crescent (C), twin-spot (T), and one-spot (0), are composed of micromelanophores arranged in various ways about themuscles of the posterior part of the body. The kind of pigmentcell found in these patterns is identical with that of the stipplepattern (St), namely micromelanophores. The type of distributionof these color elements determines the characteristic pattern; andthis in turn depends upon specific heritable factors.
Since it appears that none of the pattern factors, St, T, 0,or C, of Platypoecilus are affected by Xiphophorus modifyinggenes, as shown by hybrids possessing these factors, it may beassumed that none of them is essential for the development ofmelanosis in generic hybrids.
1506 MYRON GORDON
6. Consideration of the Factors for Patterns InvolvingMaeromelanophores
In previous sections of this paper it was shown that whenthe factors involving erythrophores Rand Rf, and those involvingmicromelanophores St, C, T, and 0, are introduced in generichybrids, no melanotic conditions arise. Consideration of thefactors controlling development of macromelanophores will nowbe made: Sp for black spotting and N for the black-bandedcondition.
(a) Behavior of M acromelanophores in Hybrid Fishes: Wheneither the Pulehra or Rubra platy is crossed with Xiphophorusand their hybrids are obtained, remarkable changes take place inthese fishes with respect to the increased rate of development anddistribution of the maeromelanophores. From the day of theirbirth, these spotted hybrids appear abnormally pigmented, indicating an accelerated rate of growth of the macromelanophores atan early stage. As the animals increase in size, the developmentof melanophores progresses rapidly. The intensity of melanosisfinally reached by the hybrid fishes varies with the individual.This subject is discussed in an accompanying paper by Reed andGordon. In brief, three states of melanosis are recognized.
The first state of melanosis is expressed in the development ofan excessive number of macromelanophores in the dermal regions.The result is that large portions of normal dermal tissues arecompletely replaced by the spotted type of black pigment cells.
In the second state of melanosis, not only does replacement ofnormal tissues by macromelanophores take place in the dermis,but the subcutaneous tissues are invaded, also. Externally, thesecond state of melanosis is made evident by loss of parts, such aspartial or complete breakdown of the fins. In some cases, the finrays remain exposed as jagged stumps, all the soft tissues havingbeen sloughed off.
The third state of melanosis is expressed in the formation ofdefinite black swellings on the body. Upon histologic study ofthese melanotic tumors, there is found to be good evidence ofneoplastic activity on the part of the macromelanophores.
(b) The Work of the Early Hybridizers: Losslein (1912) andHaffner (1912) were the first successfully to obtain generic hybridsbetween Platypoecilus and Xiphophorus, They crossed Pulehrafemales (with macromelanophores) with swordtail males and were
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1507
surprised to find that only the male offspring were blotched withblack pigment cells, while the female hybrids were completelywithout macromelanophores. The reason for criss-cross inheritance of maeromelanophores in the mother-to-son manner appearsmore lucid now, since Bellamy (1922, 1928) and Gordon (1927)have both shown that the factor for the large black pigment cellsfollows the ZW type of sex inheritance in Platypoecilus.
From the illustrations presented by Haffner in 1913, it seemsclear that both Losslein and Haffner had obtained hybrid fishes,some of which were suffering from some type of melanosis.
In later years, the reciprocal crosses (Xiphophorus females andRubra males) were made successfully by a number of independentfish breeders. These are listed in Table I. Among the hybridprogeny obtained by these breeders, conditions of what appearsto be melanosis are reported as occurring in female as well as inmale fishes.
(c) Review of More Recent Work in Hybridization: Haussler(1928) has published some important preliminary data on the crossof a Xiphophorus female with a Rubra male. Haussler obtainedsix melanotic hybrids, one of which, a male, had a melanoticgrowth on the ventral side of the caudal peduncle. Hausslercrossed this male back to a Rubra female and obtained 197 spottedand 59 non-spotted offspring. Of the former, he reared 40, eightmales of which developed melanomas on the ventral side of thecaudal peduncle.
Haussler calls attention to the pigmentation of the midventral black line in Xiphophorus, a feature which has beendescribed in section 4b and figured in Plate XVII. Since many ofhis melanotic hybrids had overgrowths in this region, it is hisbelief that they arise from the operation of the pigmentary factorsfor the mid-ventral line, in combination with factors for thedevelopment of the spotting elements in Platypoecilus, He wouldassume further, since all the melanomas observed by him appearedin males only, that testicular hormones are involved in the process.For the latter hypothesis he reserves final judgment because ofthe small number of cases observed.
In the melanotic hybrids obtained in this laboratory it wasobserved that melanotic overgrowths do appear at the regionindicated by Haussler (Figs. 108, 114). But from the observationof the breakdown of the fins in some hybrid fishes, and the appearance of great melanotic growths on the head (Figs. 68, 109) and
.... 01 o 00
TA
BL
EIV
Gen
eric
Cro
sses
Mad
eat
Cor
nell
Uni
vers
ity
Mel
anos
is
Cro
ssin
Tab
les
Ped
igre
eF
emal
eP
edig
ree
Mal
eN
ewP
edig
ree
Sp
ott
edS
tipp
led
Sti
pp
led
Gol
dT
ota
lR
ed1
23
--
18X
,9-1
P,1
1-1
425
56
11S
,'Sp
,R
,St
19P,
122-
2X
,l1
9-1
112
330
123
45Sp
,R
,St
St'
20P,
124-
1X
,l1
8-1
116
76
3c'3
C'
12S
p,R
St'
,C'
21X
,6
2,1
P,8
9-1
112
08
8S
t'st
22X
,11
9-3
P,6
0-1
514
513
13sr
R,
stB
ack
cros
ses
29P
,I9
-5X
P,2
5-1
156
60
17
418
Sp,
R,
StM
elan
otic
30P
,12
2-1
XP
,12
0-1
112
47
1611
0,10
02
0,1
047
Sp,
R,
St,°
St'
31X
P,1
20
-3X
,11
9-1
213
017
17S
t'S
t'32
XP
,12
0-2
P,14
3-11
129
16t1
2T28
St'
St,
T33
XP
,12
0-1
P,6
0-1
412
616
1127
St'
St,
RSe
cond
gene
rati
on36
XP
,12
0-1
XP
,12
0-1
212
515
520
St'
St'
246
X=
Xip
hoph
orus
,P
=P
laty
poec
ilus
.X
P=
hybr
idof
Xip
hoph
orus
and
Pla
typo
ecil
us.
Fo
rex
plan
atio
nof
gene
tic
sym
bols
see
lege
ndto
Pla
teII
(Am
.J.
Can
cer
15:
773,
Apr
il19
31).
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1509
upon the upper as well as the lower part of the caudal peduncle(Fig. 110), it seems that the region of hypertrophy is general overthe entire surface of the body. Kosswig (1928, 1929), also, reportslateral as well as ventral melanomas in hybrid fishes. He objectsto the hypothesis that testicular hormones are to be assumed asnecessary for melanosis development, since he has obtained melanomas in female as well as male hybrid fishes. Kosswig's conclusions on these points are borne out by the work in this laboratory.
(d) New Data on the Rubra-Swordtail Hybrids: In this laboratoryin 1926 a Xiphophorus female was successfully bred to a Rubramale (a spotted, stippled, red Platypoecilus). The male washeterozygous for the macromelanophore (spotted) factor and thiswas reflected in the proportional number of melanotic and nonmelanotic hybrid fishes obtained in the first generation: there werefive melanotic fishes and six normals (cross number 18, Table IV).A male melanotic hybrid, showing the first state of melanosis,was crossed back to a Rubra female (heterozygous for the macromelanophore factor). Seven of the eighteen offspring obtainedwere melanotic. One of these seven developed a large melanotictumor in the tail region (Figs. 108 and 114), while six did not gobeyond the first state of melanosis (cross number 29, Table IV).
In another experiment (number 19, Table IV), the reciprocalof the first cross (number 18) was made successfully; a Rubrafemale was crossed to a swordtail male. The Rubra female wasof a special homozygous stock with respect to the sex-linkedfactor for macromelanophores. For details of the origin of femalesof this stock, see Fraser and Gordon (1929).
From the above cross (number 19, Table IV) forty-five hybridfishes were obtained, everyone of which showed unmistakablesigns of melanosis: thirty of these hybrids did not go beyond whatis here termed the first state of melanosis (Fig. 88); twelve wereclearly in the second state of melanosis (Fig. 66) ; and three showedthe development of large melanotic tumors (Fig. 68). The latterupon histological study revealed neoplastic features (see accompanying paper by Reed and Gordon).
It is believed, since all intermediate conditions of melanosis arefound in the above group of melanotic fishes, that all of the hybridsreflect different states of the same fundamental type of pathologicaldisturbance of their pigmentary system. In this group of hybridfishes, characterized by the possession of macromelanophores, the
1510 MYRON GORDON
incidence of melanotic tumors, as such, is 6.7 per cent, but formelanosis it is 100 per cent.
Identically the same Rubra female, which had been usedpreviously for the generic cross with the swordtail and which hadproduced 100 per cent melanotic hybrids, was crossed again, thistime to a Platypoecilus male. The latter was a stippled platyand thus comparable in color pattern to the Xiphophorus maleused in the first cross. The Rubra female produced two newbroods which contained thirty-two individuals. The latter wereall spotted and red but did not possess more than the normalnumber of maoromelanophores typical of Platypoecilus (Fig. 16,Plate II).
The above data show that the genus Xiphophorus introducescertain factors which appear to interact with the spotted factor
. (Sp) of Platypoecilus, resulting in the abnormal development ofmaerornelanophores in generic hybrids. This is essentially thesame kind of conclusion reached by Haussler, Kosswig, and Winge.An extension of this explanation will be offered later.
(e) Heritable FactOTS fOT Melanosis: If it be true that Xiphophorus contributes modifying factors which act on the macromelanophores (Sp) of Platypoecilus, it should be possible todetect them in their hybrids by an appropriate breeding test.From the back crosses of Preuss; Kosswig, and Haussler, it isquite clear that melanotic hybrids actually do transmit factorswhich are concerned with the development of melanotic individualsthat appear in the next generation.
Demonstration of transmission of hereditary modifying factorsmay also be made by the use of non-melanotic generic hybrids.Hybrids resulting from the cross of gold platy male and Xiphophorus female are non-melanotic, yet they carry modifyingfactors which, when combined with the factor for macromelanophores of Platypoecilus, bring about melanotic fishes in the nextgeneration. These results will follow from the cross of a goldengeneric hybrid (non-melanotic) by a Rubra platy (Sp) (Plate XII).
The Rubra female used in this cross comes from a unique stock.The gene for black spotting (Sp) is carried on the W chromosome,while its recessive allelomorph (sp) is carried on the Z. Theorigin and behavior of females of this genetic constitution havebeen presented by Fraser and Gordon (1929). (It has been shownby these workers that these Rubra females, when crossed to recessive males of their own species, give mother-to-daughter and
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1511
father-to-son type of inheritance instead of the normal crisscross type of inheritance.)
As a result of crossing a Rubra female (W 8p Z.p) with an F 1
golden generic hybrid, forty-seven fishes were produced (crossnumber 30, Table IV); twenty-three were black spotted, whiletwenty-four were not, indicating the segregation of the factor forblack spots (Sp). Of the twenty-three individuals in the spottedclass, seventeen were reared to twelve months and over; thesewere females, as expected. Seven are melanotic, six are not, andfour are difficult to classify. In none of the seven is there adefinite tumor-like growth, but they all express the first state ofmelanosis (see Figs. 76 and 78). While it is probable that manymodifying factors are operating in Xiphophorus which influencethe degree of melanosis in the hybrids, it appears hopeless to tryto establish the exact number of these factors without increasedfacilities to obtain a greater number of individuals.
(f) Nature of Modifying Factors of Xiphophorus: All thePulchras and Rubus which have been used in generic crosses withXiphophorus have carried factors for both types of melanophores,micromelanophores (St) and macromelanophores (Sp). Theirhybrids have all been melanotic. When a Platypoecilus is usedhaving only the factor for micromelanophores (St) , no melanotichybrids are produced. It would follow logically that if the Spfactor for macromelanophores is responsible for pigment abnormalities in generic hybrids, melanotic hybrids should beproduced equally well by the crossing of a spotted (non-stippled),Platypoecilus with Xiphophorus, This was done (see Table IV,number 20), and melanotic hybrids were produced (Plate XI).This cross has been instructive in giving a clue as to the possiblefactors that Xiphophorus may contribute for the production ofmelanotic hybrids. Before entering into the details of this crosssome previous work must be reviewed.
In a short illustrated statement concerning pigment inheritancein Platypoecilus, Gordon (1928) has indicated that the factor formicromelanophores (St) interacts with the factor for macromelanophores (Sp). This was demonstrated by crossing a spotted(Sp) female with a stippled (St) male. The spotted plus stippled(Sp St) offspring which resulted showed a far greater number ofmacromelanophores than the spotted, non-stippled female parent.Where formerly the macromelanophores were confined chiefly tothe caudal peduncle of the spotted platy, they now extend over
1512 MYRON GORDON
the entire body of the spotted plus stippled platy; the latter arenot melanotic, however (see Plate XI, Figs. 11 X 15 = 16).
A study was made of the comparative effects of the factors formicromelanophores, St' of Xiphophorus and St of Platypoeeilus,upon the factor for macromelanophores, Sp of Platypoeeilus.This was accomplished by making the generic cross previouslyoutlined. A Xiphophorus male, carrying the stippling factor St',was crossed with a Platypoecilus female having the factor forspotting Sp (Figs. 20 X 69 = 70). An increase in the number ofmacromelanophores and the extension of the range of the spottingelements took place in the generic hybrids similar to that whichoccurred in fishes of the intervarietal cross. But there is a significant difference between the results of the two crosses. Thismay be expressed in the following manner:
1. The stipple factor (St) of Platypoecilus extends the scopeof activity of the factor for spots (Sp); this increase in the numberof macromelanophores is restricted to normal limits (Plate XI,Figs. 11 X 15 = 16).
2. The stipple factor (St') of Xiphophorus also extends thescope of activity of the factor for spots (Sp) of Platypoecilus; butthe process of macromelanophore development becomes uncontrolled and results in melanotic conditions (Plate XI, Figs. 20X 69 = 70).
The question of whether the stipple factor (St') of Xiphophorusis responsible both for extension of the range of the maeromelanophores of Platypoecilus in generic hybrids and for their neoplastictendencies has been subject to an experimental test. (The followingback cross has already been presented under section 6c of thispaper and is illustrated by Plate XII.)
Platypoecilus (Rubra) female X F1 non-melanotic generic hybrid
1. Heterozygous for the macro- 1. Recessive for the macromel-melanophore factor: Sp 8p. anophore factor: 8P sp.
2. Heterozygous for the micro- 2. Heterozygous for the micro-melanophore factor: St 8t. melanophore factor: St' at'.
Classification of the forty-seven offspring:a. With macro- (Sp) and micromelanophores (St or St'), 17;
5 melanotic.b. With macro- (Sp) but no micromelanophores, 6; 2 melanotic.c. With micromelanophoresonly (St or St'), 21; 0 melanotic.d. With no micro-or macromelanophores (8t 8p), 3; 0 melanotic.
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1513
From the observation that melanotic offspring occur in thetwo macromelanophore groups-five in the group (a) containingmicromelanophores also (Sp St), and two in the group (b) withoutmicromelanophores (Sp st)-it appears that the dominant stipplingfactor (St') of Xiphophorus is not concerned in the development ofmelanosis. The action of the St' gene of Xiphophorus appears tobe homologous with the action of the St gene of Platypoecilus inmerely extending, within normal limits, the range of macromelanophores. Since these conclusions have been derived fromconsideration of relatively few individuals, no final statements arepossible.
It is of interest to note, in passing, that the Xiphophorus maleused in the cross previously referred to (Fig. 69, Plate XI) isheterozygous for an autosomal factor for the crescent pattern(0'). In the unspotted, non-melanotic generic hybrids, shownin Fig. 90, this character appears in one half of the individuals.No morphologic changes were detected in this character, whichindicates that there is no effect of Platypoecilus factors upon it.This crescent (e'), like the crescent (C) of Platypoecilus, is composed of micromelanophores.
(g) The Hereditary Factor for the Black-banded Pattern: BothBellamy (1922) and Kosswig (1928,1929) were successful in rearingthe intergeneric hybrids of black Platypoecilus (Fig. 17) andXiphophorus. The cross was also made in the Cornell Universitylaboratory, but the offspring failed to mature. According to theprevious authors, abnormalities of much the same type occur ingeneric hybrids with the use of black platy as with the use ofRubra. This becomes more pronounced when the F1 hybrids arebred for the second generation or back-crossed to Xiphophorus.Kosswig declares that the degree of abnormality in the hybridsis determined by the number of Xiphophorus modifying factors.Tumors are more common among the F:z and back-cross individuals.Kosswig arranges his melanotic hybrids in a series of seven groupsaccording to the intensity of pigmentation acquired. In explanation of the variability of tumor formation he assumes a group ofmodifying factors, contributed by Xiphophorus. These factors,he believes, interact with the factor for the black pattern ofPlatypoecilus which, it will be recalled, is composed of the samelarge type of melanophores as are found in the spotted pattern ofRubras. This is further indication that the macromelanophores ofPlatypoecilus are particularly susceptible to the influence ofXiphophorus modifying factors.
1514 MYRON GORDON
SUMMARY AND CONCLUSIONS
An analysis of the results of both fish fanciers and geneticistsreveals the fact that whenever the Pulchra, Rubra and, undercertain conditions, Nigra (black) varieties of Platypoecilus areused in generic crosses with Xiphophorus, melanosis occurs insome of the hybrid offspring.
On the other hand, when varieties of Platypoecilus, such asstippled, one-spot, twin-spot and crescent, are used in genericcrosses with Xiphophorus, no comparable abnormal growthsappear.
The question has been asked, what are the factors involved inthe first group of varieties of Platypoecilus which give rise to thedevelopment of melanosis in hybrid offspring?
By a series of genetic tests and morphologic studies of the pigmentary equipment of the isolated strains, it has been possible toshow that each variety of Platypoecilus possesses certain characteristic color pigment cells or chromatophores which previousgenetic work has shown to be heritable. (This has been summarized in Table III.)
Tests of two red. strains of Platypoecilus (possessing erythrophores) have shown in crosses with Xiphophorus that, while thehybrid offspring are redder than either parent, melanosis does notappear.
The results of the work here presented indicate that all thosevarieties of Platypoecilus which give rise to the development ofmelanotic neoplasms in their hybrids possess macromelanophores:while those which merely possess micromelanophores give rise tonormal offspring.
In a crucial test, a Platypoecilus possessing only maeromelanophores (the hereditary factor for macromelanophores being carriedin the heterozygous condition) was crossed with Xiphophorus,with the result that melanotic over-growths developed in all thosehybrids which possessed macromelanophores j other hybrids of thesame brood, born and reared in the same aquarium, which didnot possess macromelanophores were normal.
On the other hand, there are many records of the results ofcrossing Platypoecilus possessing micromelanophores with Xiphophorus, which indicate that melanosis does not develop in thesehybrids.
It appears that the spotted variety of Platypoecilus providesthe cells (macromelanophores) which develop abnormally in generic
HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1515
hybrids, while the genus Xiphophorus contributes the factors whichact upon the macromelanophores of Platypoecilus.
Previous genetic and morphologic work with Platypoecilus bythe writer have shown that macromelanophoree are heritable andmay be referred to a sex-linked dominant factor (Sp). Micromelanophores have also been shown to be heritable, but they mustbe referred to a dominant autosomal factor (St).
No conclusions have been reached with respect to the determination of the nature of Xiphophorus factors which areinvolved in the development of melanosis in the generic hybrids.Several heritable factors of Xiphophorus have been eliminated asbeing involved in the process.
The development of melanotic neoplasms may be prevented ingeneric hybrids of Platypoecilus and Xiphophorus by avoiding theuse of Platypoecilus varieties carrying the heritable factor formacromelanophoree.
REFERENCES
BELLAMY, A. W.: Breeding experiments with the viviparous Teleosts:Xiphophorus helleri and Platypoecilus maoulatus, Anat. Reo. 23:98,1922.
BELLAMY, A. W.: Bionomic studies on certain teleosts (Poeciliinae).I. Statement of problems, description of material, and general noteson life histories and breeding behavior under laboratory conditions,Genetics 9: 513-529, 1924.
BELLAMY, A. W.: Bionomic studies on certain teleosts (Poeciliinae).II. Color pattern inheritance and sex in Platypoecilus, Genetics 12:226-232, 1928.
FRASER, A. C., AND GORDON, MYRON: A third linkage group in Platypoecilus, Anat. Rec. 41: 106, 1928 (a).
FRASER, A. C., AND GORDON, MYRON: Identity of genes and sex factors inintergeneric killifish hybrids, Anat. Rec. 41: 108, 1928 (b).
FRASER, A. C., AND GQRDON, MYRON: The genetics of Platypoeoilus II.The linkage of two sex-linked characters, Genetics 14: 160-179,1929.
GERSCHLER, M. W.: nber alternative Vererbung Kreuzung von Cyprinodontiden-Gattungen, Ztschr. f. Ind. Abstam. u. Verer. 12: 73-96,1914.
GOODRICH, H. B.: Mendelian inheritance in fish, Quart. Rev. Biol, 4:83-99, 1929.
GORDON, MYRON: Tropical aquarium fish-culture in industry and inscientific research, Trans. Am. Fish Soc. 56: 227-237, 1926.
PLATE X
(FIOUBJl8 61-66, 7, 8, 16)
Tml Fm.8'1'-oJ!lNEBATION HYBBID8
Xiphophorus female, 61, by gold platy, 7, yields reddish Fahybrid, 62.Xiphophorus female, 63, by red platy; 8, yields red F1 hybrid, 64-Xiphophorus female, 66, by Rubra (red, spots, &tipples) 16, yields melanotic F1
hybrid, 66.These are the results, to the fim generation, after crossing similar Xiphophorus
females to three genetically different Platypoecilus malee. Each resulting intergenerichybrid is different acoording to the basic genetic factors for pigment cells inherited fromthe Platypoecilus male parent and modified by factors contributed by the Xiphophorusfemale parent. Only the la8t, Fig. 66, &hOWl the melanotic condition; its male parentcontributed the factor for macromelanophoreB, the black spotting oolor cells; and thefemale parent furnished the modifying factors which disturb the normal development oftheee pigment cells.
Water color. btl Wil1Ielmina BrOtll1l.
1516
.:
" .-,
II
tv.,,..
«,-,~....
>< ><CD
><
-an
HEREDITARY BASIS OJ' MELANOSIS IN HYBRID FISHES 1517
GORDON, MYRON: Genetics of a viviparous top-minnow, Platypoecllus;the inheritance of two kinds of melanophores, Genetics 12: 253,1927.
GORDON, MYRON: Pigment inheritance in a Mexican killifish; interactionof factors in PlatypoeciIus, J. Hered. 19: 551-556, 1928.
GORDON, MYRON: Morphology of heritable color patterns in Platypoecilus, Xiphophorus and hybrid fishes, Thesis in Cornell University Library, Ithaca, N. Y., 1-97, pI. 1-114, 1929.
GRAM:SC~, E. (Secretary): II Naturfreund" Vereinsnachrichten, Hannover,Wcbnschr. f. Aquar. u, Ter. 10: 97, 1913.
HAFFNER, K.: Eine Kreuzung zwischen Xiphophorus helleri var. GUntheriund Platypoecilus maculatus var. pulchra, Blat. f. Aquar. u. Ter. 23:595-596, 1912.
HAFFNER, K.: Nochmals die Kreuzung zwischen Xiphophorus strigatusRegan (= X. helleri Gilntheri) manchen X Platypoecllus maeulatusvar. pulchra., Blat. f. Aquar. u. Ter. 24: 533-535, 1913.
HAUSSLER, G.: Uber Melanombildungen bei Bastarden von Xiphophorushelleri und Platypoecilus maculatus var. rubra, KIin. Wcbnschr. 7:1561-1562, 1928.
HUBBS, C. L.: Studies of the fishes of the order Cyprinodontes II. Ananalysis of the genera of the PoeciIiidae, Museum of Zoology, Univ.Michigan, 13: 5-11, 1924.
INNES, WAf. T.: Goldfish varieties and tropical aquarium fishes, Innes &Son, Philadelphia, 1921.
KOSSWIG, CURT: Uber Kreuzungen zwischen den Teleostiem Xiphophorushelleri und Platypoecilus maeulatus, II., Ztschr. f. Ind. Abet. u,Verer. 47: 150-158, 1928.
KOSSWIG, CURT: V'ber die veriinderte Wirkung von Farbgenen desPlatypoecilus in der Gattungskreuzung mit Xiphophorus, Ztschr. f.Ind. Abst. u. Verer. 50: 63-73, 1929 (a).
KOSSWIG, CURT: Melanotische GeschwuIstbiIdungen bei Fishbastarden,Verhandl. d. Deutsche Zool. Gesellsch. 1929: 90-98, 1929 (b).
KOSSWIG, CURT: Das Gen in fremder Erbmasse, Der Zuchter 1: 152-157,1929 (c).
KOSSWIG, CURT: Zur Frage der Geschwulstbildung bei Gattungsbastardender Zahnkarpfen, Xiphophorus und PlatypoeciIus, Ztschr. f. Ind.Abet. u. Verer. 52: 114-120, 1929 (d).
LOSSLEIN, F.: Vereinsnachrichten der II Aquarien und Terrarienabteilungder naturhistorischen GeseIIschaft," Blat. f. Aquar. u, Ter. 23:346-347, 1912.
MEEK, S. E.: The fresh water fishes of Mexico, north of the Isthmus ofTehuantepec, Field Col. Mus. Zool. Series 5: 1-252, 1904.
MELLON, I.: Fishes in the home, Dodd Mead & Oo., New York, 1927,pp.I-170.
PREUSS, A.: Kreuzung eines Xiphophorus-Bastardmiinchen mit Platypoecilus maculatus var. Pulehra, Blat. f. Aquar. u, Ter. 24: 608,770, 1913.
PLATE XI
(FIGU1UIlS 11, lIS. 16. 19, 20. 67-70)
Tum RECIPROCAL Caotw:s
The first cro8B. 19 X 67 ... 68, is the reciprocal of that shown in Plate 10 (65 X 16- 66), and here. also, melanotic hybrids occur.
The second cro8B, 20 X 69 .. 70. shows that only the spotted, Sp, factor, mustbe present in the Platypoecilua parent for the production of the FI melanotic hybrid.Note the small number of macromelanophores in the PI female, and compare it withthe extensive development of pigment in the F1 hybrid.
The third is reproduced from Gordon (1928) to show the normal behavior of thespotted. Sp, factor and the interaction of it with the factor for stipples, St, to producean FI type which has more macromelanophorea than its PI female parent. Yet thereis no indication of melanotic conditions here.
A somewhat comparable extension of the range of macromelanophores is seen inthe second CfOlB, 20 X 69 .. 70. This indicates that the stippling factor of Xiphophorus,St', and the stippling factor of Platypoecilua, St. both interact with the factor Sp formacromelanophores of Pla.typoecilus.
All the figures of fishesin this plate except 15 and 16 are life size. The latter are 1.5life size. Figure 69 of Xiphophorus and figure 15 of Platypoecilua are in general comparable in color pattern. When they are mated to similar females, 20 and 11. melanoticand non-melanotic individuals result.
Water colora by Wilhelmina Brown.
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HEREDITARY BASIS OF MELANOSIS IN HYBRID FISHES 1519
ROTH, P. W.: An interesting fish hybrid, Aquatic Life 1: 55-57, 1916.TURNER, C. L.: The gonads of some Platypoecilus-Xiphophorus hybrids,
Anat. Rec. 44: 243, 1929.WINGE, 0.: Arvelighedslaere, Hagerups, Copenhagen, 1928, pp. 1-390.
73
PLATE XII
(Figures 71-78)
THE BACK CROBS
The Platypoecilus female parent, Fig. 71, possesses macromelanophores (thedominant hereditary factor for macromelanophores-Sp-heing carried on the Wchromosome while the recessive-sp-is carried on the Z). It. possesses micromelanophores, also (the autosomal factor for these cells is in the heterozygous condition, also,St st).
The non-melanotic F, generic hybrid does not possess maorornelanophores, but itpossesses micromelanophores (the factor for these cells is in the heterozygous condition St: st').
Mother-to-daughter inheritance is observed for macrornelanophorea (Figs. 76, 77,and 78 are females), while father-to-son inheritance is seen for the recessive, no macromelanophores. (Figs. 73, 74, and 75 are males.)
Among the daughters which have inherited macrornelanophores, some inheritedmicromelanophores at the same time, Sp St (77 and 78), while some did not, Sp st(example 76).
Since melanotic individuals occur among those which do not possess micrornelanophores, it appears that the factor for mieromelanophores, St', is not essential to thedevelopment of melanosis.
Example number 73 lacks ruicromelanophores but possesses the one-spot characterinherited through its mother. Examples 74 and 75 possess micromelanophores and 75has the one-spot character, also. (Examples 74 and 75 appear dark on the photographs,but this is owing to the fact that the red registers darkly in photographs when no filteris used.) Examples 73, 74, and 75 are not melanotic.
1520
PL
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PLATE XlII
(Figures 72, 79-84)
THE SECOND GENERATION ~'ROM THE FERTILE INTERGEN~;RIC HYIlRILlS
Each of the F1 hybrids is heterozygous for the stippling and red-fin complex factors.The stippled F 2 hybrids are shown in Figs. 80, 81, and 82; Figs. 83 and 84 are non
stippled, golden in color.The two individuals shown in Fig. 80 are without the red-fin complex, while all
the rest have it.Figure 83 shows an imperfect twin spot character. The nature of this unusual
character is discussed in the text.There is no indication of melanosis in any of these.All the fishes were photographed to scale. The series shows size variability.
1522
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