] 142
Specialia
prises t h e a u t o s o m e t o w h i c h t h e X is t r a n s l o c a t e d . R e p lication of o n l y t h e s h o r t arm, t h e a c t u a l X, w a s t a k e n into account. I n b o t h t h e lots of c o n t i n u o u s l y labelled cells (31/2 a n d 21/2 h 3 H - T d R ) , one X w a s h o t in m o s t of t h e plates. A t o t a l of 483 m e t a p h a s e s w i t h one late r e p l i c a t i n g X were scored. Of these, t h e l o n g - n e c k e d X (X 0 was late replic a t i n g in 57.8% (279 cells) a n d t h e s h o r t - n e c k e d X (Xs} d i s p l a y e d t h e s a m e p r o p e r t y in t h e r e s t of t h e 42.2% (204 cells) m e t a p h a s e s (figures 2a, b). A s s u m i n g t h a t t h e X - c h r o m o s o m e i n a c t i v a t i o n t a k e s place a t r a n d o m , t h e p e r c e n t a g e v a l u e of e a c h t y p e of t h e late r e p l i c a t i n g X d e v i a t e s c o n s i d e r a b l y f r o m t h e e x p e c t e d value of 50%. A simple Z2 t e s t reveals t h a t t h i s difference b e t w e e n t h e 2 X ' s is s t a t i s t i c a l l y v e r y s i g n i f i c a n t 0r 2 = 11.64; p >
o.ool). T h e p r e r e q u i s i t e for t h e cytological e v a l u a t i o n of r a n d o m n e s s of X - i n a c t i v a t i o n in m a m m a l i a n females 4 is t h e o c c u r r e n c e of 2 m o r p h o l o g i c a l l y d i s t i n g u i s h a b l e b u t n o r m a l X - c h r o m o s o m e s in a female. T h e r e f o r e t h e t w o female s y s t e m s m o s t l y used for t h i s p u r p o s e are m u l e s as well as mice h e t e r o z y g o u s for C a t t a n a c h ' s X - a u t o s o m e t r a n s location. I n b o t h t h e i n s t a n c e s , h o w e v e r , c o n t r a d i c t o r y r e s u l t s h a v e b e e n o b t a i n e d ~-10. R e c e n t l y , in C a t t a n a c h mice, T a k a g i a n d Sasaki 11 as well as W a k e e t al. 12 h a v e s h o w n t h a t w h e r e a s in a d v a n c e d stages of e m b r y o n i c dev e l o p m e n t t h e f r e q u e n c y of late r e p l i c a t i n g X was r a n -
EXPERIENTIA 33/9
dom, in e a r l y e m b r y o n i c d e v e l o p m e n t p a t e r n a l X was p r e f e r e n t i a l l y late replicating. These results are in acc o r d a n c e w i t h earlier f i n d i n g s of C h a n d l e y 9, w h o obs e r v e d selective i n a c t i v a t i o n of p a t e r n a l X in C a t t a n a c h mice. The p r e s e n t r e p o r t o n t h e M u n t j a c X ' s also illust r a t e s p r e f e r e n t i a l late r e p l i c a t i o n of t h e X I c h r o m o s o m e . W e do n o t k n o w t h e p a t t e r n of i n h e r i t a n c e of t h e s e 2 X ' s a n d h e n c e c a n n o t d e r i v e t h e i r p a r e n t a g e . Moreover, a l t h o u g h t h e results are clear-cut, it m a y be p r e m a t u r e to d r a w decisive e v i d e n c e in f a v o u r of n o n - r a n d o m n e s s of X - i n a c t i v a t i o n . The p r e s e n t d a t a could as well be a n exa g g e r a t e d e x p r e s s i o n of w h a t m i g h t h a v e b e e n a c h a n c e d e f l e c t i o n a t t h e t i m e of d i f f e r e n t i a t i o n of t h e X - i n a c t i v a t i o n in t h e p r i m o r d i a l cells. A l t e r n a t i v e l y p r e f e r e n t i a l d i s t r i b u t i o n (or selection) of one t y p e of X in a h i g h l y d i f f e r e n t i a t e d tissue c a n n o t b e ruled o u t as a possibility. 6
B.B. Mukherjee, A. B. Mukherjee and A. B. Mukherjee, Nature 228, 1321 (1970). 7 B.B. Mukherjee and A. K. Sinha, Proc. Nat. Acad. Sci. 51,252 (1964). 8 J.L. Hamerton, F. Giannelli, F. Collins, J. Hallett, A. Fryer, V. M. McGuire and R. V. Short, Nature 222, 1277 (1969). 9 A.C. Chandley, Nature 221, 70 (1969). 10 H . J . Evans, C. E. Ford, M. F. Lyon and J. Gray, Nature 206, 900 (1965). 11 N. Takagi and M. Sasaki, Nature 256, 640 (1975). 12 N. Wake, N. Takagi and M. Sasaki, Nature 262, 580 (1976).
N o a s s o c i a t i o n b e t w e e n Gilbert's s y n d r o m e , the ABO b l o o d g r o u p s a n d the h a p t o g l o b i n p h e n o t y p e s 1, 2 R. P l a t z e r a n d J. B i r c h e r
Department o] Clinical Pharmacology, University o] Berne, Murtenstrasse 35, CH-3070 Bern (Switzerland), 9 February 7977 Summary. The d i f f e r e n t AB0 blood g r o u p s a n d h a p t o g l o b i n p h e n o t y p e s are as f r e q u e n t in a well-defined g r o u p of s u b j e c t s w i t h G i l b e r t ' s s y n d r o m e as in a p p r o p r i a t e s a m p l e s of t h e general p o p u l a t i o n . G i l b e r t ' s s y n d r o m e is generally c h a r a c t e r i z e d b y mild u n c o n j n g a t e d h y p e r b i l i r u b i n e m i a in t h e a b s e n c e of evid e n c e for h e p a t i c or h e m o l y t i c disease. A l t h o u g h a familial incidence w i t h a u t o s o m a l d o m i n a n t t r a n s m i s s i o n a n d v a r i a b l e e x p r e s s i v i t y h a s b e e n n o t e d a-s, t h e m o d e of i n h e r i t a n c e h a s n o t b e e n s a t i s f a c t o r i l y e s t a b l i s h e d . Since g e n e t i c m a r k e r s m a y be useful in s u c h s i t u a t i o n s , t h e r e c e n t l y r e p o r t e d association b e t w e e n slow a c e t y l a t o r phenotypes and Gilbert's syndrome appears notew o r t h y ~ I n a search for f u r t h e r g e n e t i c m a r k e r s , t h e
association b e t w e e n G i l b e r t ' s s y n d r o m e , t h e AB0 blood g r o u p s y s t e m a n d t h e h a p t o g l o b i n p h e n o t y p e s was inv e s t i g a t e d in t h e s u b j e c t s p r e v i o u s l y p u b l i s h e d 9. Material and methods. The r e c o r d s of t h e 27 p a t i e n t s w i t h G i l b e r t ' s s y n d r o m e i n v e s t i g a t e d for t h e a c e t y l a t o r p h e n o t y p e 9 were e x a m i n e d for b l o o d g r o u p d e t e r m i n a t i o n s . I n 22 cases s e r u m (stored a t - - 2 0 ~ w a s available for det e r m i n a t i o n s of s e r u m h a p t o g l o b i n p h e n o t y p e s . D e t e r m i n a t i o n s were d o n e b y s t a r c h gel electrophoresis. The diagnosis of G i l b e r t ' s s y n d r o m e was b a s e d on a) long-
AB0 blood groups and haptoglobin phenotypes in patients with Gilbert's syndrome
1
Analyzed proportions
Gilbert's syndrome
Controls
Z~*
0 :A 0 :(B+AB) A :(B+AB) A :{0+B+AB) Hp2-1:Hp2-2 Hp2-1:Hp 1-1 Hp2-2:Hpl-1 Hp2-1:(Hpl-l+Hp2-2)
13:11 13:3 11:3 11:16 12:7 12:3 7:3 12:10
1067:1274 1067:366 1274:366 1274:1433 1376:970 1376:431 970:431 1376:1401
0.703 0.410 0.006 0.432 0.160 0.127 0.163 0.218
2 3 4 5 6 7 8 9
*Z~ corresponding to p < 0.05 is 3.9.
Acknowledgments. The haptoglobin phenotype determinations were kindly performed by the Control Laboratory, Transfusion Service of the Swiss Red Cross, Wankdorfstrasse 10, CH-3014 Berne. Supported by the Swiss National Science Foundation. W.T. Foulk, H, R. Butt, C. A. Owen, F. F. Whitcomb and H. L. Mason, Medicine 38, 25 (1959). E. Meulengracht, Klin. Wschr. 18, 118 (1939). N. Alwall, Acta med. stand. 723, 560 (1946). W. Damasluk and K. Singer, Arch. intern. Med. 67, 259 (1941). L.W. Powell, E. Hemingway, B. H. Billing and S. Sherlock, New Engl. J. Med. 277, 1108 (1967). E. Penner, W. R. Mayr, S. Djawan, H. Seyfried and M. Pacher, Schweiz. reed. Wschr. 106, 860 (1976). J. Bircher, R. Platzer, A. Ktipfer and R. Preisig, Gastroenterology 71, 896 (1976).
15.9. 1977
Specialia '
s t a n d i n g or i n t e r m i t t e n t u n c o n j u g a t e d h y p e r b i l i r u b i n emia, b) exclusion of h e p a t i c disease b y clinical e x a m i n a t i o n a n d b y n o r m a l f i n d i n g s of t h e following l a b o r a t o r y t e s t s : f a s t i n g t o t a l s e r u m bile acids, GOT, a l k a l i n e p h o s p h a t a s e , s e r u m p r o t e i n electrophoresis, h e p a t i t i s B s - a n t i gen, B S P - t e s t s 1~ a n d galactose e l i m i n a t i o n c a p a c i t y n. No p a t i e n t h a d o v e r t hemolysis. T h e d i s t r i b u t i o n of A, B, 0 a n d A B blood g r o u p s a n d H p 1-1, H p 2-1 a n d H p 2-2 h a p t o g l o b i n p h e n o t y p e s in t h i s s a m p l e was c o m p a r e d w i t h a series i n v e s t i g a t e d for forensic p u r p o s e s TM. T h e a v a i l a b l e n u m b e r of cases allowed a n a l y s i s of g r o u p s 0, A a n d B + A B c o m b i n e d as well as of H p 2-1, H p 2-2, a n d H p 1-1 + H p 2-2 c o m b i n e d . Differences in t h e p r o p o r t i o n of b l o o d g r o u p s a n d h a p t o g l o b i n p h e n o t y p e s in G i l b e r t ' s s y n d r o m e a n d in t h e cont r o l series were t e s t e d for s t a t i s t i c a l significance a c c o r d i n g t o W o o l f 13. Results. T h e i n c i d e n c e of t h e d i f f e r e n t genetic t r a i t s is d e t a i l e d in t h e t a b l e . No differences could b e d e t e c t e d between the patients with Gilbert's syndrome and the control population.
Chromosomes
1143
Discussion. A s s o c i a t i o n s b e t w e e n diseases a n d some gen e t i c p o l y m o r p h i s m s , a l t h o u g h rare, are well k n o w n p h e n o m e n a 14. T h e y are u s u a l l y i n t e r p r e t e d as suggestive e v i d e n c e for genetic factors c o n t r i b u t i n g to t h e p a t h o genesis of t h e diseases in q u e s t i o n . T h e p r e s e n t s t u d y is similar t o t h e r e s u l t s o b t a i n e d b y t e s t i n g t h e H L - A histocompatibility antigens in subjects with Gilbert's syndrome, by showing that the AB0 blood group system a n d t h e h a p t o g l o b i n p h e n o t y p e s c a n n o t be used as genetic m a r k e r s in t h i s c o n d i t i o n s . T h e a s s o c i a t i o n b e t w e e n a c e t y l a t o r p h e n o t y p e a n d G i l b e r t ' s s y n d r o m e o b s e r v e d in t h e s a m e s a m p l e of p a t i e n t s b e c o m e s t h e m o r e r e l e v a n t . 10 W. Haecki, J. Bircher and R. Preisig, J. Lab. clin. Med. 88, 1019 (1976). 11 ' J. Bircher, R. Blankart, A. Halpern, W. Haecki, J. Laissue and R. Preisig, Eur. J. clin. Invest. 5, 72 (1973). 12 M. Beringer, Inaugural Dissertation Zfirich 1967. Orell Ftissli AG, Zfirich. 13 B. Woolf, Ann. hum. Genet. 21, 397 (1957). t4 O. Mayo, in: Biochemical Genetics in Man, p. 347. Academic Press, London 1972.
of the b l a c k a b a l o n e ( H a l i o t i s c r a c h e r o d i i ) 1
j. Minkler 2
Biomedical Sciences Division, Lawrence Livermore Laboratory, University o[ Cali/ornia, Livermore (Cali[ornia 94550, USA), J March 1977 Summary. T h e c h r o m o s o m e s of t h e b l a c k a b a l o n e I-Ialiotis c r a c h e r o d i i are d e s c r i b e d for t h e first time. T h e diploid n u m b e r is 36, w i t h n o a p p a r e n t s e x u a l d i m o r p h i s m . S e v e r a l g a s t r o p o d k a r y o t y p e s are d e s c r i b e d in t h e literat u r e a p p e a r i n g f r o m 19053 to t h e r e c e n t p a s t 4-6. T h e s e are m a i n l y of snails a n d n u d i b r a c h s . C y t o g e n e t i c s t u d i e s o n a b a l o n e are lacking. 4-year-old 500 g specimens, 1 m a l e a n d 1 female, were o b t a i n e d f r o m t h e i n t e r t i d a l zone n e a r D i a b l o C a n y o n , California, a n d used for t h i s s t u d y . 5 m l of a 0.1% Colcemid s o l u t i o n (in sea w a t e r ) were i n j e c t e d
Fig. A. Karyotype of a male Haliotis cracherodii.
Fig. B. Karyotype of a female Haliotis cracherodii. Bar equals 10 [xm.
i n t o t h e c e n t e r of t h e foot. 4 h later, t h e a n i m a l was dissected 3 a n d t i s s u e f r a g m e n t s were o b t a i n e d f r o m t h e gill, m a n t l e , d i g e s t i v e gland, g o n a d a n d blood. T h e tissues were p l a c e d in H a n k s b a l a n c e d s a l t s o l u t i o n c o n t a i n i n g 0.3% t r y p s i n for s u b s e q u e n t m i n c i n g . A f t e r c e n t r i f u g a t i o n t h e cell p e l l e t w a s s u s p e n d e d in 5 m l 0.075 M KC1 a n d allowed t o sit for 20 m i n a t r o o m t e m p e r a t u r e . F o l l o w i n g 3 f i x a t i o n s in a b s o l u t e m e t h a n o l : glacial acetic acid (3:1) t h e cells were d r o p p e d o n t o slides, air dried, a n d s t a i n e d in G i e m s a (10% in G u r r ' s Buffer). T h e d i g e s t i v e g l a n d was t h e o n l y tissue t o yield mitoses. Of 21 t o t a l c o u n t a b l e m e t a p h a s e s , 15 h a d 36 c h r o m o somes, 3 h a d 35, 2 h a d 34 a n d 1 h a d 18. 4 m a l e a n d 3 f e m a l e k a r y o t y p e s were p r e p a r e d . E x a m p l e s are s h o w n in figures A a n d B. T h e s e s h o w 16 m e t a c e n t r i c , 16 submetacentric, and 4 acrocentric chromosomes. The chromosomes are small, t h e l a r g e s t b e i n g a p p r o x i m a t e l y 4 ~zm long. No s e x u a l d i m o r p h i s m is o b s e r v a b l e in t h e s e k a r y o t y p e s . Sex d e t e r m i n a t i o n m a y b e a t t h e i n t r a c h r o m o s o m a l level. I t is also possible t h a t t h e s e k a r y o t y p e s could repr e s e n t t e t r a p l o i d s , especially since t h e s e ceils c a m e f r o m t h e liver-like d i g e s t i v e gland. F u r t h e r w o r k will b e necess a r y to clarify t h e s e a m b i g u i t i e s .
1 This work was performed under the auspices of the U. S. ERDA Contract No. W-7405-ENG-48. 2 I thank Dr Florence Harrison for her expert dissections. 3 W.M. Smallwood, Gegenbaurs morph. Jb. 33, 73 (1905). 4 C.M. Patterson, Malacol. Rev. 6, 141 (1973). 5 L. Raghunathan, Malacologia, 15, 447 (1976). 6 Y. Minichev, Zool. ZH 53, 1255 (1974).
Specialia
prises t h e a u t o s o m e t o w h i c h t h e X is t r a n s l o c a t e d . R e p lication of o n l y t h e s h o r t arm, t h e a c t u a l X, w a s t a k e n into account. I n b o t h t h e lots of c o n t i n u o u s l y labelled cells (31/2 a n d 21/2 h 3 H - T d R ) , one X w a s h o t in m o s t of t h e plates. A t o t a l of 483 m e t a p h a s e s w i t h one late r e p l i c a t i n g X were scored. Of these, t h e l o n g - n e c k e d X (X 0 was late replic a t i n g in 57.8% (279 cells) a n d t h e s h o r t - n e c k e d X (Xs} d i s p l a y e d t h e s a m e p r o p e r t y in t h e r e s t of t h e 42.2% (204 cells) m e t a p h a s e s (figures 2a, b). A s s u m i n g t h a t t h e X - c h r o m o s o m e i n a c t i v a t i o n t a k e s place a t r a n d o m , t h e p e r c e n t a g e v a l u e of e a c h t y p e of t h e late r e p l i c a t i n g X d e v i a t e s c o n s i d e r a b l y f r o m t h e e x p e c t e d value of 50%. A simple Z2 t e s t reveals t h a t t h i s difference b e t w e e n t h e 2 X ' s is s t a t i s t i c a l l y v e r y s i g n i f i c a n t 0r 2 = 11.64; p >
o.ool). T h e p r e r e q u i s i t e for t h e cytological e v a l u a t i o n of r a n d o m n e s s of X - i n a c t i v a t i o n in m a m m a l i a n females 4 is t h e o c c u r r e n c e of 2 m o r p h o l o g i c a l l y d i s t i n g u i s h a b l e b u t n o r m a l X - c h r o m o s o m e s in a female. T h e r e f o r e t h e t w o female s y s t e m s m o s t l y used for t h i s p u r p o s e are m u l e s as well as mice h e t e r o z y g o u s for C a t t a n a c h ' s X - a u t o s o m e t r a n s location. I n b o t h t h e i n s t a n c e s , h o w e v e r , c o n t r a d i c t o r y r e s u l t s h a v e b e e n o b t a i n e d ~-10. R e c e n t l y , in C a t t a n a c h mice, T a k a g i a n d Sasaki 11 as well as W a k e e t al. 12 h a v e s h o w n t h a t w h e r e a s in a d v a n c e d stages of e m b r y o n i c dev e l o p m e n t t h e f r e q u e n c y of late r e p l i c a t i n g X was r a n -
EXPERIENTIA 33/9
dom, in e a r l y e m b r y o n i c d e v e l o p m e n t p a t e r n a l X was p r e f e r e n t i a l l y late replicating. These results are in acc o r d a n c e w i t h earlier f i n d i n g s of C h a n d l e y 9, w h o obs e r v e d selective i n a c t i v a t i o n of p a t e r n a l X in C a t t a n a c h mice. The p r e s e n t r e p o r t o n t h e M u n t j a c X ' s also illust r a t e s p r e f e r e n t i a l late r e p l i c a t i o n of t h e X I c h r o m o s o m e . W e do n o t k n o w t h e p a t t e r n of i n h e r i t a n c e of t h e s e 2 X ' s a n d h e n c e c a n n o t d e r i v e t h e i r p a r e n t a g e . Moreover, a l t h o u g h t h e results are clear-cut, it m a y be p r e m a t u r e to d r a w decisive e v i d e n c e in f a v o u r of n o n - r a n d o m n e s s of X - i n a c t i v a t i o n . The p r e s e n t d a t a could as well be a n exa g g e r a t e d e x p r e s s i o n of w h a t m i g h t h a v e b e e n a c h a n c e d e f l e c t i o n a t t h e t i m e of d i f f e r e n t i a t i o n of t h e X - i n a c t i v a t i o n in t h e p r i m o r d i a l cells. A l t e r n a t i v e l y p r e f e r e n t i a l d i s t r i b u t i o n (or selection) of one t y p e of X in a h i g h l y d i f f e r e n t i a t e d tissue c a n n o t b e ruled o u t as a possibility. 6
B.B. Mukherjee, A. B. Mukherjee and A. B. Mukherjee, Nature 228, 1321 (1970). 7 B.B. Mukherjee and A. K. Sinha, Proc. Nat. Acad. Sci. 51,252 (1964). 8 J.L. Hamerton, F. Giannelli, F. Collins, J. Hallett, A. Fryer, V. M. McGuire and R. V. Short, Nature 222, 1277 (1969). 9 A.C. Chandley, Nature 221, 70 (1969). 10 H . J . Evans, C. E. Ford, M. F. Lyon and J. Gray, Nature 206, 900 (1965). 11 N. Takagi and M. Sasaki, Nature 256, 640 (1975). 12 N. Wake, N. Takagi and M. Sasaki, Nature 262, 580 (1976).
N o a s s o c i a t i o n b e t w e e n Gilbert's s y n d r o m e , the ABO b l o o d g r o u p s a n d the h a p t o g l o b i n p h e n o t y p e s 1, 2 R. P l a t z e r a n d J. B i r c h e r
Department o] Clinical Pharmacology, University o] Berne, Murtenstrasse 35, CH-3070 Bern (Switzerland), 9 February 7977 Summary. The d i f f e r e n t AB0 blood g r o u p s a n d h a p t o g l o b i n p h e n o t y p e s are as f r e q u e n t in a well-defined g r o u p of s u b j e c t s w i t h G i l b e r t ' s s y n d r o m e as in a p p r o p r i a t e s a m p l e s of t h e general p o p u l a t i o n . G i l b e r t ' s s y n d r o m e is generally c h a r a c t e r i z e d b y mild u n c o n j n g a t e d h y p e r b i l i r u b i n e m i a in t h e a b s e n c e of evid e n c e for h e p a t i c or h e m o l y t i c disease. A l t h o u g h a familial incidence w i t h a u t o s o m a l d o m i n a n t t r a n s m i s s i o n a n d v a r i a b l e e x p r e s s i v i t y h a s b e e n n o t e d a-s, t h e m o d e of i n h e r i t a n c e h a s n o t b e e n s a t i s f a c t o r i l y e s t a b l i s h e d . Since g e n e t i c m a r k e r s m a y be useful in s u c h s i t u a t i o n s , t h e r e c e n t l y r e p o r t e d association b e t w e e n slow a c e t y l a t o r phenotypes and Gilbert's syndrome appears notew o r t h y ~ I n a search for f u r t h e r g e n e t i c m a r k e r s , t h e
association b e t w e e n G i l b e r t ' s s y n d r o m e , t h e AB0 blood g r o u p s y s t e m a n d t h e h a p t o g l o b i n p h e n o t y p e s was inv e s t i g a t e d in t h e s u b j e c t s p r e v i o u s l y p u b l i s h e d 9. Material and methods. The r e c o r d s of t h e 27 p a t i e n t s w i t h G i l b e r t ' s s y n d r o m e i n v e s t i g a t e d for t h e a c e t y l a t o r p h e n o t y p e 9 were e x a m i n e d for b l o o d g r o u p d e t e r m i n a t i o n s . I n 22 cases s e r u m (stored a t - - 2 0 ~ w a s available for det e r m i n a t i o n s of s e r u m h a p t o g l o b i n p h e n o t y p e s . D e t e r m i n a t i o n s were d o n e b y s t a r c h gel electrophoresis. The diagnosis of G i l b e r t ' s s y n d r o m e was b a s e d on a) long-
AB0 blood groups and haptoglobin phenotypes in patients with Gilbert's syndrome
1
Analyzed proportions
Gilbert's syndrome
Controls
Z~*
0 :A 0 :(B+AB) A :(B+AB) A :{0+B+AB) Hp2-1:Hp2-2 Hp2-1:Hp 1-1 Hp2-2:Hpl-1 Hp2-1:(Hpl-l+Hp2-2)
13:11 13:3 11:3 11:16 12:7 12:3 7:3 12:10
1067:1274 1067:366 1274:366 1274:1433 1376:970 1376:431 970:431 1376:1401
0.703 0.410 0.006 0.432 0.160 0.127 0.163 0.218
2 3 4 5 6 7 8 9
*Z~ corresponding to p < 0.05 is 3.9.
Acknowledgments. The haptoglobin phenotype determinations were kindly performed by the Control Laboratory, Transfusion Service of the Swiss Red Cross, Wankdorfstrasse 10, CH-3014 Berne. Supported by the Swiss National Science Foundation. W.T. Foulk, H, R. Butt, C. A. Owen, F. F. Whitcomb and H. L. Mason, Medicine 38, 25 (1959). E. Meulengracht, Klin. Wschr. 18, 118 (1939). N. Alwall, Acta med. stand. 723, 560 (1946). W. Damasluk and K. Singer, Arch. intern. Med. 67, 259 (1941). L.W. Powell, E. Hemingway, B. H. Billing and S. Sherlock, New Engl. J. Med. 277, 1108 (1967). E. Penner, W. R. Mayr, S. Djawan, H. Seyfried and M. Pacher, Schweiz. reed. Wschr. 106, 860 (1976). J. Bircher, R. Platzer, A. Ktipfer and R. Preisig, Gastroenterology 71, 896 (1976).
15.9. 1977
Specialia '
s t a n d i n g or i n t e r m i t t e n t u n c o n j u g a t e d h y p e r b i l i r u b i n emia, b) exclusion of h e p a t i c disease b y clinical e x a m i n a t i o n a n d b y n o r m a l f i n d i n g s of t h e following l a b o r a t o r y t e s t s : f a s t i n g t o t a l s e r u m bile acids, GOT, a l k a l i n e p h o s p h a t a s e , s e r u m p r o t e i n electrophoresis, h e p a t i t i s B s - a n t i gen, B S P - t e s t s 1~ a n d galactose e l i m i n a t i o n c a p a c i t y n. No p a t i e n t h a d o v e r t hemolysis. T h e d i s t r i b u t i o n of A, B, 0 a n d A B blood g r o u p s a n d H p 1-1, H p 2-1 a n d H p 2-2 h a p t o g l o b i n p h e n o t y p e s in t h i s s a m p l e was c o m p a r e d w i t h a series i n v e s t i g a t e d for forensic p u r p o s e s TM. T h e a v a i l a b l e n u m b e r of cases allowed a n a l y s i s of g r o u p s 0, A a n d B + A B c o m b i n e d as well as of H p 2-1, H p 2-2, a n d H p 1-1 + H p 2-2 c o m b i n e d . Differences in t h e p r o p o r t i o n of b l o o d g r o u p s a n d h a p t o g l o b i n p h e n o t y p e s in G i l b e r t ' s s y n d r o m e a n d in t h e cont r o l series were t e s t e d for s t a t i s t i c a l significance a c c o r d i n g t o W o o l f 13. Results. T h e i n c i d e n c e of t h e d i f f e r e n t genetic t r a i t s is d e t a i l e d in t h e t a b l e . No differences could b e d e t e c t e d between the patients with Gilbert's syndrome and the control population.
Chromosomes
1143
Discussion. A s s o c i a t i o n s b e t w e e n diseases a n d some gen e t i c p o l y m o r p h i s m s , a l t h o u g h rare, are well k n o w n p h e n o m e n a 14. T h e y are u s u a l l y i n t e r p r e t e d as suggestive e v i d e n c e for genetic factors c o n t r i b u t i n g to t h e p a t h o genesis of t h e diseases in q u e s t i o n . T h e p r e s e n t s t u d y is similar t o t h e r e s u l t s o b t a i n e d b y t e s t i n g t h e H L - A histocompatibility antigens in subjects with Gilbert's syndrome, by showing that the AB0 blood group system a n d t h e h a p t o g l o b i n p h e n o t y p e s c a n n o t be used as genetic m a r k e r s in t h i s c o n d i t i o n s . T h e a s s o c i a t i o n b e t w e e n a c e t y l a t o r p h e n o t y p e a n d G i l b e r t ' s s y n d r o m e o b s e r v e d in t h e s a m e s a m p l e of p a t i e n t s b e c o m e s t h e m o r e r e l e v a n t . 10 W. Haecki, J. Bircher and R. Preisig, J. Lab. clin. Med. 88, 1019 (1976). 11 ' J. Bircher, R. Blankart, A. Halpern, W. Haecki, J. Laissue and R. Preisig, Eur. J. clin. Invest. 5, 72 (1973). 12 M. Beringer, Inaugural Dissertation Zfirich 1967. Orell Ftissli AG, Zfirich. 13 B. Woolf, Ann. hum. Genet. 21, 397 (1957). t4 O. Mayo, in: Biochemical Genetics in Man, p. 347. Academic Press, London 1972.
of the b l a c k a b a l o n e ( H a l i o t i s c r a c h e r o d i i ) 1
j. Minkler 2
Biomedical Sciences Division, Lawrence Livermore Laboratory, University o[ Cali/ornia, Livermore (Cali[ornia 94550, USA), J March 1977 Summary. T h e c h r o m o s o m e s of t h e b l a c k a b a l o n e I-Ialiotis c r a c h e r o d i i are d e s c r i b e d for t h e first time. T h e diploid n u m b e r is 36, w i t h n o a p p a r e n t s e x u a l d i m o r p h i s m . S e v e r a l g a s t r o p o d k a r y o t y p e s are d e s c r i b e d in t h e literat u r e a p p e a r i n g f r o m 19053 to t h e r e c e n t p a s t 4-6. T h e s e are m a i n l y of snails a n d n u d i b r a c h s . C y t o g e n e t i c s t u d i e s o n a b a l o n e are lacking. 4-year-old 500 g specimens, 1 m a l e a n d 1 female, were o b t a i n e d f r o m t h e i n t e r t i d a l zone n e a r D i a b l o C a n y o n , California, a n d used for t h i s s t u d y . 5 m l of a 0.1% Colcemid s o l u t i o n (in sea w a t e r ) were i n j e c t e d
Fig. A. Karyotype of a male Haliotis cracherodii.
Fig. B. Karyotype of a female Haliotis cracherodii. Bar equals 10 [xm.
i n t o t h e c e n t e r of t h e foot. 4 h later, t h e a n i m a l was dissected 3 a n d t i s s u e f r a g m e n t s were o b t a i n e d f r o m t h e gill, m a n t l e , d i g e s t i v e gland, g o n a d a n d blood. T h e tissues were p l a c e d in H a n k s b a l a n c e d s a l t s o l u t i o n c o n t a i n i n g 0.3% t r y p s i n for s u b s e q u e n t m i n c i n g . A f t e r c e n t r i f u g a t i o n t h e cell p e l l e t w a s s u s p e n d e d in 5 m l 0.075 M KC1 a n d allowed t o sit for 20 m i n a t r o o m t e m p e r a t u r e . F o l l o w i n g 3 f i x a t i o n s in a b s o l u t e m e t h a n o l : glacial acetic acid (3:1) t h e cells were d r o p p e d o n t o slides, air dried, a n d s t a i n e d in G i e m s a (10% in G u r r ' s Buffer). T h e d i g e s t i v e g l a n d was t h e o n l y tissue t o yield mitoses. Of 21 t o t a l c o u n t a b l e m e t a p h a s e s , 15 h a d 36 c h r o m o somes, 3 h a d 35, 2 h a d 34 a n d 1 h a d 18. 4 m a l e a n d 3 f e m a l e k a r y o t y p e s were p r e p a r e d . E x a m p l e s are s h o w n in figures A a n d B. T h e s e s h o w 16 m e t a c e n t r i c , 16 submetacentric, and 4 acrocentric chromosomes. The chromosomes are small, t h e l a r g e s t b e i n g a p p r o x i m a t e l y 4 ~zm long. No s e x u a l d i m o r p h i s m is o b s e r v a b l e in t h e s e k a r y o t y p e s . Sex d e t e r m i n a t i o n m a y b e a t t h e i n t r a c h r o m o s o m a l level. I t is also possible t h a t t h e s e k a r y o t y p e s could repr e s e n t t e t r a p l o i d s , especially since t h e s e ceils c a m e f r o m t h e liver-like d i g e s t i v e gland. F u r t h e r w o r k will b e necess a r y to clarify t h e s e a m b i g u i t i e s .
1 This work was performed under the auspices of the U. S. ERDA Contract No. W-7405-ENG-48. 2 I thank Dr Florence Harrison for her expert dissections. 3 W.M. Smallwood, Gegenbaurs morph. Jb. 33, 73 (1905). 4 C.M. Patterson, Malacol. Rev. 6, 141 (1973). 5 L. Raghunathan, Malacologia, 15, 447 (1976). 6 Y. Minichev, Zool. ZH 53, 1255 (1974).
