Chromosome Variability in the Lemuridae ' J. EGOZCUE Department of Genetics, Oregon Regional Primate Research Center, Beaverton, Oregon Chromosome studies have been conducted in Lemur catta, L. macaco, ABSTRACT L. mongoz, L. fztlvus fulvus, and a hybrid L. fulvus fulvus x L. fulvus albifrons. Comparative analysis shows that inter- and intraspecific chramosome variability is a common finding in Lemurs. The Lemuridae are divided into three groups according to the characteristics of their chromosome complement. A reclassification of L. catta with the Hapalemurs is suggested, based on chromosomal and nonchromosomal data. Some years ago (Chu and Swomley, '61 ; Chu and Bender, '61), chromosome studies were published on several species of Lemurs, including L. catta, L. fulvus, L. variegatus, L . macaco, Hapalemur griseus, and Microcebus murinus. Interspecific chromosome variability was found to be the rule for the genus, the chromosomes ranging in number from 44-66, but intraspecific variability was seen only in L. fulvus and H . griseus. Based on these studies, a three-group division of the Lemurine lemurs was proposed: the first composed of the Hapalemurs; the second, of L. cutta, 1,.mongoz, L. fuluus rufus, and L. fuluus albifrons; the third, of L. fuluus fulvus, L. variegatus, L. macaco, and a new subspecies, L. SQ., which resemble the brown lemurs (Chu and Swomley, '61; Bender and Chu, ' 6 3 ) . We report here our studies on L . catta, L. macaco, L. mongoz mongoz, L . fulvus fulvus, and a hybrid, L. fulvus fuluus x L. fulvus albifrons. The results show that chromosome variability is even more widespread among the Lemuridae than was formerly supposed and suggest that a revision of the genus Lemur is in order. MATERIALS AND METHODS Two female L. catta, three male and three female L. macaco, one male and seven female L. mongoz, two male and one female L. fuluus fulvus, and one hybrid, L. fulvus fulvus x L. fulvus albifrons, were used i n this study. Blood (Egozcue and Vilarasau de Egozcue, '66) and kidney cultures (Melnick, '66; Hagemenas and Egozcue, in preparation) were set up for Am. J. PHYS.ANTHROP.,26: 341-348. chromosome studies, incubated for three days (blood) or two weeks (kidney) at 37"C, and chromosome preparations made up according to a technique described by us (Egozcue and Vilarasau de Egozcue, '66). Micrographs were taken at 1000 X and the karyograms constructed from metaphase prints at 2000 X. TABLE 1 Species Specimens Chromosomes Male Female 2n L e m u r catta - L.macaco 3 3 44 L. mongoz 1 7 58 L.fulvus fulvus 2 1 58 Hybrid L . fulvus fUlV7LS x L . fulvus albifrons 1 - 58 2 56 RESULTS Most of the metaphases in the preparations were normal and diploid. The chromosome number turned out to be 2n=56 in L . catta, 2n=44 in L. macaco, and 2n=58 in all other animals studied (table 1).In L. mongoz, of 232 cells counted, 186 had 58 chromosomes; in L. fulvus fulvus, of 95 cells counted, 86 had 58 chromosomes; and in the hybrid, of 48 cells counted, 42 had 58 chromosomes. IPublication no. 214 of the Oregon Regional Primate Research Center, supported in part by Grant FR 00163-07 from the National Institutes of Health. 341 342 J. EGOZCUE The chromosomes were paired and arranged visually in order of decreasing size. The sex chromosomes were determined by elimination in the males. The chromosome complement of L. catta is composed of four pairs of metacentric chromosomes, three pairs of submetacentrics and 20 pairs of acrocentrics; the sex chromosomes are acrocentric (fig. 1). Our results differ somewhat from those previously reported (Chu and Swomley, '61). There are in the complement one more pair of metacentric and submetacentric chromosomes and two Iess pairs of acrocentrics. Our results in L. macaco are identical to those of Chu and Bender ('61) (fig. 2). The chromosome complement is composed of six pairs of metacentrics, four pairs of submetacentrics and 11 pairs of acrocentrics; the X chromosome is the longest acrocentric, Fig. 1 the Y a very small acrocentric. The chromosome complements of L. mongoz, L. fulvus fuluus and the hybrid (figs. 3,4,5) are composed of two pairs of submetacentric chromosomes (pairs no. 1 and 2) and 26 pairs of acrocentrics. The X chromosome is the longest acrocentric, similar in size to pair no. 3. The Y chromosome is the smallest acrocentric. The only individually identifiable pairs are no. 1 and 2; both are submetacentric and differ considerably in size from one another. In the complement of all Lemurs studied by us, two pairs of chromosomes bear a rather long acromatic region in the midproximal third of the long arms (fig. 1, 5). The size of the chromosomes makes its visualization difficult. These pairs are found, by size, in positions ranging from no. 1618 for the long pair, and from no. 18-20 Chromosome complement of a female L. catta. CHROMOSOMES OF L E M U R I D A E Fig. 2 343 Metaphase of a female L. macaco. for the short one. In the karyograms, we opted to position them according to size rather than to assign a definite position €or each of them. DISCUSSION A comparison of all the Lemuridae studied (table 2) reveals that there is a wide range of variability in their chromosome numbers. Intraspecific variability is found in L. mongoz mongoz (2n=58, 2n=60) and in L. fulvus albifrons (2n=58, 2n=60), as well as in L. fulvus fulvus (2n=58, 2n=48). The type of chromosome variability is the same for the mongoose lemurs and the white-fronted lemurs but quite different for L. fulvus fulvus. Our specimens of L. m o n g m have one less small pair of acrocentric chromosomes than the ones studied by Chu and Swomley ('61). There is a striking difference between our counts in L. fulvus fulvus and the ones by Chu and Swomley ('61). While the karyotype of our animals is the same as in L. mongoz and L. fulvus albifrons and almost identical to that of other L. mongoz and L. fulvus rufus, the karyotype of the two animals described by Chu and Swomley ('61) is closer to the type found in L. variegatus and L. macaco. The results in the hybrid L. fulvus f u L vus x L . fulvus albifrons support our count of 2n=58 for L. fulvus fulvus and suggest that the same chromosome number and morphological characteristics (fig. 5) 344 J. EGOZCUE Fig. 3 Chromosome complement of a female L. mongoz. should be found in L. f u l v u s albifrons, since half of the complement is contributed by each of the parents and no differences were found in the chromosome complement of the hybrid. On the grounds of their results, Chu and Swomley ('61) and later Bender and Chu ('63) suggested that the Lemuridae be divided into three groups: (1) Hapalemurs; (2) L. catta, L. mongoz, L. fulvus r u f u s , and L . f u l v u s albifrons; (3) L. f u l v u s fulvus, L. variegatus, L. macaco, and L. s p . The data obtained in our studies indicate that a revision of these groups is in order without, however, altering their basic structure. The first group would consist of the lemurs with a diploid number of 2n=60 or 2n=58, with no metacentric chromosomes and with a significant predominance of acrocentrics. It would include the two subspecies of L. mongoz, the subspecies of L. f u l v u s f u l v u s studied by us, L. fulvus r u f u s , and L. fulvus albifrons. The species with a diploid number around 46, with a high number of metacentric and acrocentric chromosomes, would form the second group. These are L. f u l v u s fulvus, L. variegatus, and L. macaco. It is rather surprising that while most subspecies of L. fulvus have firstgroup type karyotypes, a single subspecies of L. fulvus fulvus should have a different chromosome complement, especially when other animals supposedly belonging to the same subspecies have first-group karyotypes like all other fulvus lemurs. CHROMOSOMES O F LEMURIDAE 345 Fig. 4 Chromosome complement of a female L . fuZvus fulvus. The third group would be composed of the Hapxlemurs. L. catta is a rather special case. Bolwig ('60) suggested that it be classified as a new genus, Odorlemur catla, based upon analysis of the brachial glands, the antebrachial organs (carpal glands), and the associated spurs of the animal. Affolter ('38), in a description of the brachial organs of Hapalemur griseus, points out the close res,emblance between such organs in Hapalemur and L. catta, although he does not think that this means they are closely related. Montagna and Yun ('62), in a detailed histological, histochemical, and anatomical study of the dermal and carpal glands and organs of L. catta, find many resemblances to similar structures in Hapalemur. .Buettner-Janusch ('63) on these grounds states that if ever they had t o be reclassified, L. catta should be mare logically placed in a taxon with Hapalemur. Based on chromosomal evidence, we suggest that L. catta be included in the Hapalemur group, characterized by a chromosome number around 56, with a low count of both metacentric and submetacentric chromosomes and a predominance of the acrocentric. The new species of lemur, L. sp., described by Chu and Swomley ('61) is chromosomally closer to the Hapalemur group although it has the general morphological characteristics of L. fuluus. As only one animal has been studied, we believe that further studies are needed before any definitive classification of this possibly new species can be made; it may prove to be only a case of individual variability. Thus, the three groups would be: ( 1 ) L. mongoz and L. fuZuus; (2) L. uariegatus and L. macaco, with the inclusion of the specimens of L. fuluus fuluus with a dip- Fig. 5 Chromosome complement of a male hybrid L. fulvus fulvus x L. fulvus albifrons. TABLE 2 Species Common name Chromosomes Specimens Reference ~~ Male Female 2n M S A X Y 60 0 4 54 A A ChuandSwomley,’61 0 4 52 A A Thisreport ~ Lemur mongoz L. mongoz L. fulvus L . fulvus fulvus L. f U l V U S T U f U S L. fulvus albifrons Hybrid L. fulvus fulvus x L. fulvus albifrons L. fulvus fulvus Mongoose lemur 2 3 Mongoose lemur 1 7 Brown lemur ...... 1 Brown lemur 2 1 Red-fronted lemur 2 White-fronted lemur 1 1 L. macaco L. macuco L. s p . L. catta L. catta Hapalemur griseus griseus H . griseus olivaceus 1 Not stated. (58) Chiarelli, ’61 2 58 0 4 52 A A Thisreport GO 0 4 54 A A ChuandSwomley,’Gl 60 0 4 54 A A ChuandSwomley,’61 58 0 4 52 A A Thisreport 48 10 6 30 A A ChuandSwomley,’Gl 46 14 4 26 M A Chu andSwomley,’61 Brown lemur L. variegatus 58 Ruffed lemur Black lemur Black lemur Ringtail lemur Ringtail lemur Gray gentle lemur 2 Tentative. 1 1 1 1 1 44 12 8 22 A A ChuandBender,’Gl 3 1 3 44 52 12 8 8 4 22 38 A M A A Thisreport Chu andSwomley,’61 56 6 4 44 A A Chu andSwomley,’Gl 2 56 8 6 40 A A Thisreport 1 54 4 6 42 A A Chu andSwomley,’61 1 58 2 4 52 - - Chu andSwomley,’61 2 1 CHROMOSOMES OF LEMURIDAE loid number of 2n=48; ( 3 ) L. catta and Hapalemur. Further investigation with more animals may disclose new cases of chromosome variability in the Lemuridae. LITERATURE CITED Affolter, M. 1938 Lcs organes cutanks brachiaux d’HapaZemur griseus. Bull. Acad. Malgache. Tananarive, 20: 77-100. Bender, M. A., and E. H. Y. Chu 1963 The chromosomes of Primates. In: Evolutionary and Genetic Biology of Primates. J. BuettnerJanusch, Ed. Academic Press, New York. Bolwig, N. 1960 A comparative study of the behaviour of various lemurs. Mem. Inst. Sci. Madagascar, Ser. A: Biologic Anirnale, 14: 205-21 7. Buettner-Janusch, J. 1963 An introduction to Primates. In: Evolutionary and Genetic Biology of Primates. J. Buettner-Janusch, Ed. Academic Press, New York. 347 Chiarelli, B. 1961 Cited by M. A. Bender and E. H. Y. Chu 1963 The chromosomes of Primates. In: Evolutionary and Genetic Biology of Primates. J. Buettner-Janusch, Ed. Academic Press, New York. Chu, E. H. Y., and M. A. Bender 1961 Chromosome cytology and evolution i n primates. Science, 133: 1399-1405. Chu, E. H. Y., and B. A. Swomley 1961 Chromosomes of lemurine lemurs. Science, 133: 1925-1926. Egozcue, J., and M. Vilarasau de Egozcue 1966 Simplified culture and chromosome preparations of primate lcukocytes. Stain Technol., 41: 173-1 77. Hagemenas, F., and J. Egozcue ( I n preparation) Simplified tissue culture and chromosome preparations in primates. Melnick, J. L. 1956 In: Diagnostic Procedure f o r Viral and Rickettsial Diseases. 2nd Ed. American Public Health Association, New York. Montagna, W., and J. S. Yun 1962 The skin of Primates. X. The skin of the Ring-tailed Lemur. Am. J. Phys. Anthrop., 20: 95-118.