J.MoI.Evol.8,283-294

Journalof MolecularEvolution

(1976)

© by Springer-Verlag 1976

Evolution of Flightless Land Birds on Southern Continents: Transferrin Comparison Shows Monophyletic Origin of Ratites ELLEN M. PRAGER 1, ALLAN C. WILSON 1, DAVID T. OSUGA 2, and ROBERT E, F E E N E Y 2 1

Department of Biochemistry, University of California, Berkeley, Cal. 94720, USA

2

Department of Food Science and Technology, University Of California, Davis, Cal. 95616, USA

Received May 29, 1976; July 20, 1976

Summary. A biochemical approach was used to study the evolution of ratite birds, i.e., the ostriches, rheas, cassowaries, emus, and kiwis. Quantitative immunological comparison of transferrin from ratites, tinamous, and other flying birds indicates that all the ratites and tinamous are allied phylogenetically and that they are of monophyletic origin relative to other birds. To explain the current geographic distribution of ratites and the magnitude of the transferrin distances, it is supposed that the ancestors of these flightless birds walked across land bridges between the southern continents during Cretaceous times.

Key words: Protein Evolution/Transferrin/Immunology/Micro-Complement Fixation/Phylogeny/Flightlessness/Ostrich/Rhea/Tinamou/Continental Drift

The

large

flightless

birds

of

the

southern

continents

have

in-

trigued evolutionary biologists for over a century. These birds, k n o w n as r a t i t e s , i n c l u d e t h e l i v i n g o s t r i c h e s , r h e a s , c a s s o w a r i e s , e m u s , a n d k i w i s , as w e l l as t h e r e c e n t l y e x t i n c t m o a s and elephant birds. Their geographic distribution is i n d i c a t e d in F i g u r e I. A s i d e f r o m b e i n g c o m p l e t e l y flightless, the ratites share a few anatomical features that distinguish them

*Supported in part by grants GB-42028X from NSF and GM-21509 from NIH to A.C.W. and by grants HD-OOI22 from NIH and GA-12607 from NSF to R.E.F. The following abbreviation is used in this work: MC'F = micro-complement fixation.

283

Africa

Fig.l Geographic distribution of ratites South America J and tinamous. The numbers within the Australia RH~EA 9 OSTRICH dashed lines indicate, in millions of New?udinea 0__~ years ago, the times of disappearance i New | of land bridges between the continent~ ,CASSOWARY EMU Z~d [INAMOU1 and islands shown (Cracraft, 1972, 1973; Maxson et al., 1975). The I Antarctica ] ostriches, rheas, emus, and cassowaries, weighing 80-140 kg, are often known collectively as the "large ratites", in contrast to the 2-3 kg kiwis. The tinamous, unlike the ratites, are able to fly; they are included in this figure since their phylogenetic relationship to the ratites is demonstrated. The extinct flightless moas and elephant birds (see text) existed in New Zealand and Madagascar, respectively

:501

lao',

,17°",

from all other birds except tinamous ( P a r k e s & C l a r k , 1966; Sibley & Ahlquist, 1972; C r a c r a f t , 1974) .I U n t i l t h e t h e o r y of c o n t i n e n t a l drift became widely accepted there was much uncertainty r e g a r d i n g t h e o r i g i n of r a t i t e s . Their flightlessness and anatomical similarities could have arisen by either convergent or divergent evolution. According to t h e c o n v e r g e n t evolution hypothesis, t h e r a t i t e s on a g i v e n land mass arose from ancestors that flew there and later became flightless and convergently s i m i l a r in a n a t o m y a n d w a y of l i f e to the r a t i t e s o n o t h e r c o n t i n e n t s ; t h u s r a t i t e s c o u l d b e of polyphyletic origin. Alternatively, a c c o r d i n g to t h e d i v e r g e n t evolution hypothesis, the c o m m o n a n c e s t o r of r a t i t e s w a s f l i g h t l e s s a n d w a l k e d a c r o s s l a n d b r i d g e s to t h e v a r i o u s l a n d m a s s e s n o w i n h a b i t e d b y r a t i t e s ; t h u s r a t i t e s c o u l d b e of m o n o p h y l e t i c o r i g i n . A s s o o n as g e o l o g i s t s became convinced that l a n d b r i d g e s e x i s t e d b e t w e e n all t h e s o u t h e r n l a n d m a s s e s during the Cretaceous period (see F i g u r e I), i.e., d u r i n g t h e e a r l y s t a g e s of b i r d e v o l u t i o n , it b e c a m e e a s y to a c c e p t t h e hypothesis of m o n o p h y l e t i c origin. Although scarcely any new morphological evidence has been uncovered in t h e l a s t 2 d e c a d e s and although the available morphological e v i d e n c e is a m b i g u o u s , ornithologists n o w t e n d to f a v o r t h e m o n o p h y l y hypothesis.

1 The ratites all lack a keel on the sternum; on this basis the 4 ratite orders (Struthioniformes, Rheiformes, Casuariiformes, Apterygiformes) are grouped as the Ratitae in contrast to the remaining 23 avian orders (Wetmore, 1960), comprising the Carinatae, all of whose members have a keel. The most salient features shared by ratites and tinamous but absent in all other carinate birds are the palaeognathous palate and the unique structure of the bill (Parkes & Clark, 1966; Sibley & Ahlquist, 1972; Cracraft, 1974).

284

Much effort has been acterization of various

d e v o t e d to t h e p u r i f i c a t i o n and charproteins, particularly egg white pro-

teins, from ratites and tinamous ( O s u g a & F e e n e y , 1968) a n d t h e c o m p a r i s o n of r a t i t e a n d t i n a m o u p r o t e i n s w i t h t h o s e of

to

other birds and reptiles by chemical, electrophoretic, and qualitative immunological methods ( M i l l e r & F e e n e y , 1964; O s u g a & F e e n e y , 1968; F e e n e y & A l l i s o n , 1969; S i b l e y & A h l q u i s t , 1972; S i b l e y & F r e l i n , 1972; S i b l e y e t al., 1974; W i l s o n e t al., 1964). W h i l e t h o s e b i o c h e m i c a l results 2 are generally consistent with

the monophyly

hypothesis,

they

do

not

suffice

to d e m o n -

strate ratite monophyly rigorously. The quantitative immunological c o m p a r i s o n s of t r a n s f e r r i n reported below are consistent both with the monophyly hypothe s i s a n d w i t h t h e i d e a t h a t t h e a n c e s t o r s of r a t i t e s w a l k e d a c r o s s l a n d b r i d g e s in l a t e also demonstrate a probable

Cretaceous times. These comparisons phylogenetic relationship between

ratites and the South American tinamous. As discussed in a n a c c o m p a n y i n g article (Ho et al., 1976, and references t h e r e i n ) , c o m p a r i s o n of t h e a m i n o a c i d s e q u e n c e s of h o m o l o g o u s p r o t e i n s a m o n g l i v i n g s p e c i e s p e r m i t s t h e c o n struction of molecular phylogenies and thereby the determination

2 Electrophoretic methods, though useful for comparing proteins from closely related species, particularly members of the same genus, generally cannot yield quantitative measures of the amount of difference between proteins from phylogenetically more distantly related species such as members of different orders. On the basis of such electrophoretic comparisons of the egg white proteins of representatives of all avian orders (Sibley, 1970; Sibley & Ahlquist, 1972; Sibley & Frelin, 1972; Sibley et al., 1974) as well as comparison of the electrophoretic mobility of the tryptic peptides of ovalbumin from ratites, tinamous, and a few other carinate birds (Sibley & Frelin, 1972), Sibley and coworkers concluded that the large ratites were closely related, with the cassowary and emu particularly close; they could not place the kiwi especially close to any other avian group and suggested a possible kiwi-tinamou alliance as well as a possible tinamou-galliform alliance. Though immunoelectrophoresis and immunodiffusion can give a more realistic appraisal of the similarities between proteins, the immunological studies of Feeney's group (Miller & Feeney, 1964; Osuga & Feeney, 1968) were somewhat limited in scope, involving, principally, antisera to whole egg white and purified ovotransferrin from the chicken and the cassowary and including species from only a few avian orders besides the ratites and tinamous. The overall conclusions of Feeney and coworkers were that all the ratites formed one assemblage, with the emu most closely related to the cassowary, and that the tinamou was related to the ratites. Further quilitative evidence (Wilson et al., 1964) in favor of a single assemblage of ratites and tinamous came from comparative electrophoretic and thermostability studies with the heart form of lactate dehydrogenase. 285

of e v o l u t i o n a r y r e l a t i o n s h i p s among e x t a n t species d e s p i t e ina d e q u a t e fossil e v i d e n c e and a m b i g u o u s i n t e r p r e t a t i o n of m o r p h o l o g i c a l data. The d e g r e e of a m i n o acid s e q u e n c e diff e r e n c e a m o n g p r o t e i n s is m o s t r e a d i l y a s s e s s e d on a large scale by the q u a n t i t a t i v e i m m u n o l o g i c a l t e c h n i q u e of m i c r o c o m p l e m e n t f i x a t i o n (MC'F) (Champion et al., 1974, 1975; Ho et al., 1976), and we now r e p o r t M C ' F c o m p a r i s o n s of t r a n s f e r r i n from r a t i t e s w i t h the h o m o l o g o u s p r o t e i n from r e p r e s e n t a t i v e species of all o r d e r s of birds.

M A T E R I A L S AND M E T H O D S A n t i s e r a w e r e e l i c i t e d in r a b b i t s to t r a n s f e r r i n s p u r i f i e d from egg w h i t e or s e r u m of 5 r a t i t e s (ostrich, rhea, c a s s o w a r y , emu, kiwi), one tinamou, and 10 a d d i t i o n a l s p e c i e s (Adelie and E m p e r o r p e n g u i n s , a r c t i c loon, g r e a t c r e s t e d grebe, owl, Wester: gull, pigeon, b l a c k b i r d , chicken, and duck). D e t a i l s of p r o t e i n p u r i f i c a t i o n , a n t i s e r u m p r o d u c t i o n , and i m m u n o l o g i c a l m e t h o d s are g i v e n in an a c c o m p a n y i n g a r t i c l e (Ho et al., 1976), as are the s c i e n t i f i c names of these 16 species r e p r e s e n t i n g 14 of the 27 avian orders.

RESULTS In T a b l e I we show the r e s u l t s of M C ' F tests c a r r i e d out w i t h the 16 d i f f e r e n t antisera. Each a n t i s e r u m was t e s t e d w i t h s a m p l e s d e r i v e d from r e p r e s e n t a t i v e s of each of the 27 b i r d orders. By i n s p e c t i o n of T a b l e I we see in the u p p e r r i g h t h a n d corner a c l u s t e r i n g of low v a l u e s a m o n g the r a t i t e s and t i n a m o u r e l a t i v e to the other birds. A b s t r a c t e d from T a b l e 1 and p r e s e n t e d in T a b l e 2 are the a v e r a g e s of r e c i p r o c a l immun o l o g i c a l d i s t a n c e m e a s u r e m e n t s o b t a i n e d w i t h the a n t i s e r a to the 16 t r a n s f e r r i n s . The c l u s t e r i n g of low v a l u e s a m o n g the r a t i t e s and t i n a m o u a p p e a r s in the lower r i g h t - h a n d c o r n e r of this table. We have used the data in T a b l e 2 to c o n s t r u c t the t r a n s f e r r ~ p h y l o g e n y shown in F i g u r e 2. S a l i e n t f e a t u r e s to note are I. t~ close a s s o c i a t i o n of the c a s s o w a r y and emu, c o n s i s t e n t w i t h p r e v i o u s m o r p h o l o g i c a l and b i o c h e m i c a l evidence; 2. the a l m o s t s i m u l t a n e o u s d i v e r g e n c e into 3 l i n e a g e s at 2 p o i n t s (tinamou, ostrich, other r a t i t e s and, later, kiwi, rhea, c a s s o w a r y + emu) and, m o s t important, 3. the m o n o p h y l e t i c a s s e m b l a g e of the r a t i t e s and t i n a m o u r e l a t i v e to all other birds. Our c o n c l u s i o n , b a s e d on t r a n s f e r r i n i m m u n o l o g i c a l distancel that the r a t i t e - t i n a m o u a s s e m b l a g e is m o n o p h y l e t i c is supporte~ by M C ' F tests w i t h serum a l b u m i n and o v a l b u m i n . W i t h a n t i s e r a to rhea a l b u m i n the a v e r a g e i m m u n o l o g i c a l d i s t a n c e b e t w e e n the

286

S7 OSTRICH 2F~CASSOWARY'~I

A i~

IO0

i

i

,

I

I

50

r

Phylogenetic

I

I

I

I

of

transferrin

immunological

dis-

tances in Table 2. The heavy

B~ROS

i

relationships

ratites and tinamous based on

I------ 1 -

TINAMOU OTHER

74

t i

KLW~

Fig.2

B

~MO -71

J'-~q.~,-~-

I

-

t I

i

i

0 0 IMMUNOLOGICALDISTANCE

i

i

I

50

i

I

vertical arrows indicate the start i

,

I

I00

of the divergence of the reference species among themselves;

this

section of the phylogeny is presented in detail elsewhere (Ho et al., 1976). Phylogeny A, constructed according to the method of Fitch & Margoliash (1967), shows the branching order of the lineages leading to the present-day as the immunological

distance units of transferrin

have occurred along each lineage.

The reconstructed

distances

tree appear in Table 2. The percent standard deviation, goodness of fit of the output versus input data, phylogenetic

species as well

change calculated to for the entire

a measure of the

is 12.1% for the complete

tree. Phylogeny B, which shows only branching order,

into account alternative phylogenies

(Ho et al.,

1976)

takes

having similar

percent standard deviations and does not give credence to common ancestral lineages along which only a few units of transferrin

change may have occurr-

ed. The thick lines on the cladogram indicate those sections which have been collapsed. We have also constructed a phylogeny based on the data in Table 2 according to the Farris

(1972) method.

The resulting tree was far less satisfac-

tory, having a percent standard deviation of 29.6%. While the FitchMargoliash

(1967)

and similar

(Sarich,

iterative averaging procedures,

1969a,b)

methods are essentially

the Farris technique

is based on the as-

sumption that each input datum is either a true estimate or an underestimate, but never an overestimate, units.

Since,

however,

estimates as underestimates on immunological

of the actual distance between 2 taxonomic

immunological

data are as likely to include over-

of distances between species, Farris trees based

data are often inferior to Fitch-Margoliash

structed from the same data

(Prager et al.,

1976; Prager,

trees con-

unpub,

calcu-

lations)

rhea

and

1974), and

all

Using and

other

while other

antisera

tinamou

rhea,

while

tinamou

3prager,

ratites

the

orders to

both

and

average of

rhea were

tinamous

distance birds

48

were

57

an

of

20

average

unpublished observations,

37

units

Brush 4

immunological

representatives

assemblage

is

ovalbumin,

is

between

75

(Prager

ratites

and

(Prager

et

found

distance

orders of

units

that

al.,

al., the

1974). 3

ostrich

units

from

the

ratite-

outside units

et

tinamous

from

the

the

rhea.

using antisera to albumin from loon

(Gavia arctica), herring gull (Larus argentatus), cowbird and duck (Anas platyrhynchos) .

(Molothrus ater),

4 A.H. Brush, personal

communication.

287

Table i •

Immunological

distances

&

among bird transferrlns

Species

Antiserum AP

Struthio camelus Rhea americana b

EP

Loon G

Owl

Struthioniformes--Ostrich,

224

195

246

220

143

Rheiformes--Rhea,

139

124

143

163

201

137

152

166

169

240

141

154

143

161

232

146

154

193

161

146

170

128

191

132

198

35

17

1

40

48

31

17

0

ND

NE

51

47

55

4

76

ND

ND

O

NE

12

47

50

82

0

27

37

68

32

36

49

7~

63

92

62

96

Casuarius aruensis --Emu, Dromiceius novae-hollandiae Apterygiformes--Kiwi, Apteryx australis Tinamiformes--Tinamou, Eudromia elegans c Gaviiformes--Common loon, Gavia immer Casuariiformes--Cassowary,

--Arctic loon, Gavia arctica Podicipediformes--Western

grebe, Aechmophorus occi-

dentalis --Great crested grebe, Podiceps cristatus ND Sphenisciformes--Adelie penguin, Pygoscelis adeliae O --Emperor penguin, Aptenodytes forsteri 13 Procellariiformes--Albatross, Diomedea immutabilis 31 Pelecaniformes--Snake

bird, Anhinga anhinga

61

Ardea cocai Anas platyrhynchos Falconiformes--Falcon, Falco sparveria Galliformes--Chicken, Gallus gallus Ciconiiformes--Heron,

Anseriformes--Duck,

Rallus limicola gull, Larus occidentalis --Herring gull, Larus argentatus Columbiformes--Pigeon, Columba livia Psittaciformes--Parrot , Psittacus erithacus Cuculiformes--Cuckoo, Coccyzus americanus Strigiformes--Owl, Asio otus

35

32

55

61

84

103

112

152

128

174

56

36

50

61

94

115

118

129

129

18(

Gruiformes--Rail,

71

52

62

60

71

Charadriiformes--Western

67

49

ND

58

8[

65

ND

60

ND

N[

59

59

83

68

llC

60

45

52

57

91

88

64

96

99

IO~

71

67

82

86

[

Caprimulgus parvulus Apodiformes--Swift, Chaetura pelagica Coliiformes--Mousebird, Colius sp. e

69

56

58

54

71

74

59

68

59

8(

77

ND

80

ND

N[

Trogoniformes--Trogon,

Aphaloderma narina Megaceryle alcyon Piciformes--Woodpecker, Colaptes auratus Passeriformes--Blackbird, Agelaius phoeniceus

99

92

103

96

13(

Coraciiformes--Kizgfisher,

58

52

61

60

9[

81

93

Iii

87

14~

119

111

92

94

ii


4~ v ~J

r,j ,--i r~

R

R

r~

•

rj~

290

,~

0

0

o ~

o

~

~

o

~

~

o

0 0

R

r~

•

4~

.~

~

~

-~

DISCUSSION Continental

Drift

The r e s u l t s we r e p o r t here m a y be c o n s i d e r e d in r e l a t i o n to c u r r e n t t h e o r i e s of c o n t i n e n t a l d r i f t (Cracraft, 1972, 1973; M a x s o n et al., 1975). In F i g u r e I we have s u m m a r i z e d the d r i f t of the s o u t h e r n c o n t i n e n t s , once all c o n n e c t e d via A n t a r c t i c a . In a p r e v i o u s study (Maxson et al., 1975) s u b s t a n t i a l a g r e e m e n t was o b t a i n e d on the times of d i v e r g e n c e of the A u s t r a l i a n and N e w W o r l d m a r s u p i a l s and b y l i n e frogs b e t w e e n i m m u n o l o g i c a l e s t i m a t e s u s i n g the a l b u m i n m o l e c u l e as an e v o l t i o n a r y clock and e s t i m a t e s from c o n t i n e n t a l drift. S i m i l a r l y , if one assumes a rate of a v i a n t r a n s f e r r i n e v o l u t i o n of I-1.2 u n i t s of i m m u n o l o g i c a l d i s t a n c e per m i l l i o n years (Prager et al., 1974), the i m m u n o l o g i c a l d i s t a n c e s shown in F i g u r e 2 from the o s t r i c h and t i n a m o u to s p e c i e s on o t h e r c o n t i n e n t s are c o n s i s t e n t w i t h the times of d i s a p p e a r a n c e of land b r i d g e s shown in F i g u r e I. However, the d i s t a n c e s b e t w e e n the kiwi and rhea and from t h e s e 2 s p e c i e s to the emu and c a s s o w a r y are, on the average, r o u g h l y half the v a l u e s p r e d i c t e d from c o n t i n e n t a l d r i f t and the rate of t r a n s f e r r i n e v o l u t i o n . T h e s e d i s c r e p a n c i e s are p r o b a b l y due to the s t a t i s t i c a l l i m i t a t i o n s (Maxson et al., 1975) of the t r a n s f e r r i n clock, p a r t i c u l a r l y since avian t r a n s f e r r i n e v o l v e s m o r e s l o w l y and i r r e g u l a r l y than m a m m a l i a n a l b u m i n or t r a n s f e r r i n (Prager et al., 1974; Ho et al., 1976), and to p o s s i b l e c o n v e r g e n c e and c o n s e r v a t i s m (Cronin & Sarich, 1975) at the t r a n s f e r r i n locus in the kiwi and r h e a lineages.

Morphological

Phylogeny

C r a c r a f t ' s c l a d o g r a m (Cracraft, 1973, 1974), b a s e d on a n a l y s i s of d e r i v e d m o r p h o l o g i c a l c h a r a c t e r states, agrees w i t h our t r a n s f e r r i n p h y l o g e n y (Figure 2) in s h o w i n g the r a t i t e - t i n a m o u a s s e m b l a g e to be m o n o p h y l e t i c . However, his c l a d o g r a m d i f f e r s f r o m ours r a t h e r c o n s p i c u o u s l y in p l a c i n g the d i v e r g e n c e of the o s t r i c h and rhea at the last b r a n c h i n g p o i n t along the e n t i r e c l a d o g r a m . Since a c c o r d i n g to the p r e v i o u s l y a v a i l a b l e e v i d e n c e and that p r e s e n t e d here the r a t i t e s form a m o n o p h y l e t i c g r o u p and since (cf. F i g u r e 3 below) the c o m m o n a n c e s t o r of the o s t r i c h and r h e a could not have flown b e t w e e n A f r i c a and South A m e r i c a , C r a c r a f t ' s c l a d o g r a m a l o n g w i t h c o n s i d e r a t i o n s of c o n t i n e n t a l d r i f t (Figure I) i m p l i e s that the o s t r i c h and rhea d i v e r g e d from a c o m m o n a n c e s t o r at least 80-90 m i l l i o n y e a r s ago. His c l a d o g r a m f u r t h e r i m p l i e s that the o t h e r r a t i t e s and the t i n a m o u l i n e a g e d i v e r g e d off m u c h earlier, 1OO m i l l i o n or m o r e y e a r s ago. G i v e n our t r a n s f e r r i n r e s u l t s as w e l l as the

291

Flying Birds

Tinamous

X

Fig.3

Ratites

/ --N0n-fliers

Sustained F l i e r s - -

Conjectural phylogenetic model for the origin of flight in 2 stages. The dark, striped, and clear areas represent flightlessness,

non-

sustained flight, and sustained flight, respectively. The number of species in the ratite-tinamou --Non-fliers assemblage is less than I% of the nearly 9000 living bird species. Though Figure 3 suggests that the loss of all flying ability among the ratites occurred only once, in an ancestor common to all the ratites, Figure 2 is consistent with the possibility that flightlessness among these birds may have evolved more than once - for instance, once along the lineage leading to the ostrich and again along that leading to the remaining ratites Clearly it would be most parsimonious for flightlessness to have developed only once, prior to the beginning of the break-up of the southern continents and prior to any intra-ratite divergence, as Cracraft (1972, 1973, 1974) suggests. Molecular phylogenies constructed on the basis of comparisons of additional proteins such as ovalbumin and albumin permit further testing of this hypothesis. As discussed elsewhere (He et al., 1976) and illustrated here in Figure 2, flightlessness in ratites has arisen entirely independently from its development in penguins, the only other flightless avian order. (There have of course been occasional instances of flightlessness developing in selected species or subspecies within a number of bird orders, due primarily to selective pressures on islands) ---Non-sustained

Fliers

a v a i l a b l e d a t a o b t a i n e d w i t h a l b u m i n and o v a l b u m i n , we s u b m i t that such a n c i e n t times for the start of the i n t r a - r a t i t e d i v e r g e n c e are i m p r o b a b l e . W e r e the p o s i t i o n of the o s t r i c h s h i f t e d on C r a c r a f t ' s c l a d o g r a m , his b r a n c h i n g o r d e r c o u l d o t h e r w i s e in b r o a d o u t l i n e be r e c o n c i l e d w i t h our t r a n s f e r r i n p h y l o g e n y as w e l l as w i t h c u r r e n t t h e o r i e s c o n c e r n i n g the d r i f t of the s o u t h e r n c o n t i n e n t s .

Origin

of F l i g h t

Our t r a n s f e r r i n p h y l o g e n y m a y h a v e a b e a r i n g on t h e o r i e s as to h o w f l i g h t o r i g i n a t e d . The o l d e s t k n o w n f o s s i l bird, Archaeopter was p r o b a b l y able to fly (Ostrom, 1974). H o w e v e r , it is u n c e r t a i n w h e t h e r this a n i m a l was c a p a b l e of susta;ined f l i g h t (Ostrc 1974). T h e t r a n s f e r r i n r e s u l t s s u g g e s t that t h e r e are, phyloger e t i c a l l y speaking, 2 m a j o r g r o u p s of l i v i n g birds, n a m e l y the r a t i t e s and t i n a m o u s on the one h a n d and the r e m a i n i n g b i r d s or the o t h e r hand. C o n s i d e r a t i o n s h o u l d t h e r e f o r e be g i v e n to the s c h e m e shown in F i g u r e 3 for the o r i g i n of flight. T i n a m o u s , t h o u g h able to fly, a p p e a r i n c a p a b l e of s u s t a i n e d f l i g h t (Hudson, 1920; Wagner, 1949; P e a r s o n & P e a r s o n , 1955; V a n T y n e 292

Berger, This

1961;

finding

Figure

3.

resent birds

Lancaster, is

accounted

Ratites

a lineage before

and

to

fly

for

by

further

at all,

of

and

tinamous

have

may

(Ostrom,

1974)

restrial

rather

sustained

never is

easily

that than

the

from

capacity that

Thus

been

common

that

ancestor and was

of of

able

in

then

to lost

the to

ratites

sustained

to

other

ability

of

Ostrom's all

rep-

sustained

the

ancestor

with

model

leading for

ratites

capable

1973).

the model,

retained

the

reconciled

arboreal

to

the

tinamous

manner.

Vigil,

phylogenetic

off

assumes

while

in a n o n - s u s t a i n e d hypothesis

the

1965;

according

branched

acquisition

fly This

Johnson,

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the

flight. 5 The model ability

1964;

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suggestion

birds

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fly only

ter-

in n o n -

bursts.

Acknowledgements.

We thank C.Y.-K. Ho for purifying the 2 penguin ovo-

transferrins, carrying out the immunization program with them, and providing us with aliquots of the resulting antisera, and we thank V.M. Sarich for advice and assistance. We are especially grateful to A.H. Brush, a fellow investigator in trying to determine evolutionary relationships from comparative molecular data, for communicating his ovalbumin results to us, particularly those he has obtained with anti-rhea ovalbumin, and to R.K. Colwell, M. Foster, and O.P. Pearson for information concerning the flying ability of tinamous. Additionally, we appreciate comments made by K.C. Parkes and T. Uzzell. Finally, we thank officials of the governments of the United States, Australia, New Zealand, and the state of California for granting us their approval to collect, receive, and possess avian materials.

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