ZoologicalJournal of the Linnean Society (1995), 7 74: 139-153. With 11 figures
0nychophora:past andpresent. Edited by M. H. Walker and D. B. Norman
The use of chromosomal data in the systematics
of vivi arous on chophorans from Australia
( ~ n ~ c f o p h o rJeripatopsidae)
a:
D. M. ROWELL AND A. V. HIGGINS
Division of Botany and Zoology, Australian National University, Canberra ACT 0200,
Australia
D. A. BRISCOE AND N. N. TAIT
School of Biological Sciences, Macquarie University,Sydney, N.S. W: 2709, Australia
A chromosomal analysis of populations of viviparous Australian onychophorans has uncovered
a large radiation in karyotypic form. Chromosome numbers of 18, 26, 30, 32, 33 and 34
were observed, and these classes could be further subdivided on the basis of internal size
relationships. Given the practical difficulties with the systematics of this group, the use of
chromosomal data promises to be particularly enlightening. This is the first time any karyotypic
data have been published for onychophorans since a single species was analysed in 1900.
ADDITIONAL KEY WORDS:-Australian fauna - chromosomes
-
species radiations.
CONTENTS
Introduction . . . . . . . . .
Material and Methods . . . . . . .
Collection sites and specimens examined .
Karyotyping . . . . . . . .
Standardization . . . . . . .
Karyotypicdistance . . . . . .
Results
. . . . . . . . . .
. . . . . . . . .
Discussion
Chromosomal variation . . . . .
Affinities among the 2 n = 34 karyomorphs
Conclusions .
.
Acknowledgements . . . . . . .
References
. . . . . . . . .
INTRODUCTION
Population genetic theory predicts that de novo genetic mutations stand a
higher chance of fixation in small isolated populations. The lifestyle
0024-4082/95/050 139+15 $08.00/0
139
0 1995 The Linnean Society of London
characteristics of members of the Onychophora fit this pattern of population
distribution well. Onychophorans are small, slow-moving but apparently longlived organisms, restricted to relatively moist, dark microhabits. Hence the
propensity for population isolation and fragmentation, with associated
inbreeding, is high. Allozyme analysis has revealed relatively low
intrapopulation polymorphism (Hebert et al., 1991) and extensive fixation for
alternative alleles among populations (Briscoe & Tait, unpublished data), both
indicative of fragmented populations with local inbreeding. These conditions
are also conducive to the fixation of chromosomal rearrangements; however,
as only one population of one species has been analysed cytologically
(Peripatopsis balfouri; Montgomery, 1900), no information is available as to
the extent of karyotypic variation in the phylum.
Electrophoretic and morphological data indicate that there is considerable
variation in the Australian Onychophora (Tait & Briscoe, 1990; Briscoe &
Tait, 1993; Briscoe & Tait, unpublished data), and collections of onychophorans
from eastern Australia have revealed that specimens recently collected from
many populations do not conform to any of the species descriptions given
by Ruhberg (1985) in her revision of the group. This complicates any analysis
of variation in the group, as it is often not possible to ascribe specimens to
any formal taxon. Thus it is necessary to give some background on the
current taxonomy and the important characters in the taxa which form the
subject of this study.
Both viviparous and oviparous species of Australian onychophorans have
been reported. All oviparous forms identified to date possess an ovipositor
in the female. Euperipatoides Saenger, 1869 is the only widespread genus of
viviparous onychophorans described from the eastern margin of the Australian
mainland, distributed from southern Queensland to southern New South
Wales, and also in western Tasmania (Ruhberg, 1985). The only other
viviparous genera from this region include Austroperipatus Bouvier, 1915
(which possesses an ovipositor), Cephalofovea Ruhberg et al., 1988 and
Tmrnanipatus Ruhberg et al, 1991 which are restricted in their distribution
to northern Queensland, the region west of Sydney, New South Wales, and
northeastern Tasmania respectively. Currently only one species of
Euperipatoides, E. leuckartii Saenger 1869, is described. Unfortunately, the type
specimen of E. leuckartii has been lost (Ruhberg, 1985) and the type locality
is given only as northwest of Sydney (Saenger, 1869).
Recently, specimens have been reported from a number of populations
which have distinct and complex head structures in males (Tait & Briscoe,
1990). The morphology of these structures is likely to feature prominently
in a revised taxonomy of the Australian Onychophora. The taxonomic status
of populations of eastern Australian onychophorans is currently being
evaluated by morphology (A. Reid, PhD thesis).
Given the simple morphology of onychophorans and the difficulties
encountered historically in developing a viable taxonomy for the group (see
Tait et al., 1990), more and independent data sets would be particularly
valuable. This paper reports on an extensive karyotypic radiation of viviparous
onychophorans. These data, together with allozyme data and morphology,
will contribute to a more comprehensive understanding of species delineation
and taxon relationships.
KARYOLOGY OF AUSTRALIAN ONYCHOPHORA
I
I
I
I
I
QLD
I
I
I
I
I-------------------------&
I
- - -
I
I
I
I
I
I
I
I
Gibralter Ra
ARMIDALE
4
Siding Sp
Gloucester Tops
NSW
L Pedder
HOBART
Figure 1. Location of collection sites in southeastern Australia (large lettering). Armidale,
Sydney, Brisbane, Hobart and Canberra (small lettering) included for reference. Asterisks
indicate localities where E. leuckartii has been previously reported (from Ruhberg, 1985).
MATERIAL AND METHODS
Collection sites and specimens examined
Specimens were collected by the authors from 19 localities (Table 1 and
Fig. 1). These collections include only two described species, and consequently
in this paper the populations will be referred to by locality designations, and
species name where this can be determined. Females from many of these
collections bore live young in the laboratory, and viviparity has been verified
in all populations by dissection (A. Reid, personal communication).
Specimens collected from the Tinderry Range, Gloucester Tops and
Gibralter Range have distinctive head structures in males. The Tinderry
Mountains and Gloucester Tops populations have been designated as taxa
G and E elsewhere (Tait & Briscoe, 1990), while Gibralter Range specimens
have a distinctive but morphologically similar head structure to taxon D
from Mt Warning in northern New South Wales. Similarly, specimens from
Siding Springs, a moist volcanic outcrop on the drier western slopes of New
South Wales, have a head structure resembling that of Taxon A from Barwick
River near Armidale.
We have identified the population of onychophorans from Mt Tomah
which lacks a head structure as Euper$atoides leuckartii.
Also included in the analysis are data from Cephalofovea tomahmontis from
the type locality of Mt Tomah, New South Wales, which belongs to the
only currently described genus of Onychophora possessing a head structure
(Ruhberg et al., 1988).
In the laboratory, testes were removed for karyology and bodies frozen at
-80°C for allozyme electrophoresis. Where possible, voucher specimens of
both sexes were preserved whole from each population for future
morphological investigation.
Kayotyping
Karyotypes were produced from the testes of adult and subadult males as
outlined in Rowel1 (1985), but with a 10 min hypotonic treatment with insect
saline diluted 2 : 1 with distilled water rather than 1 : 1 as specified. Colchicine
was used initially, but it was found that concentrations of 0.05% or more
resulted in degraded preparations while lower concentrations appeared to
have no effect. Consequently, the karyotypes produced varied greatly in their
degree of contraction, and so all comparisons were made using standardized
measurements.
The application of a number of C-banding methods which have been used
successfully in other invertebrates and vertebrates did not reveal any
informative bands. Extremely small C-bands were sometimes visible in the
centromeric region of some chromosomes, but they were not sufficiently
large to be consistently reproduced photographically.
Standardization
Karyotypes were standardized as follows. Chromosomes were measured by
hand from photographic prints. For each karyotype, the average chromosomal
length was measured, and each individual chromosome measurement divided
by the average length. Internal size relationships could then be assessed by
plotting the standardized lengths on a bar graph, from largest to smallest.
KARYOLOGY OF AUSTRALIAN ONYCHOPHORA
Karyotypic distance
Where chromosome number was equal, the average size deviation or
karyotypic distance (D,), between population pairs was calculated using the
following formula:
where X l i and X2, are standardized measurements of chromosomes of the
ith size rank for populations 1 and 2 respectively, and N is the number of
chromosomes in each karyotype. D, is the square root of this value.
Using this procedure, it is possible to produce distance matrices for all
painvise comparisons of populations with the same chromosome number.
However, given that little is known of constraints on chromosome lengths,
the validity of comparing distance matrices derived from populations with
different chromosome numbers is dubious.
In populations which share the chromosome number 2n = 34, the distance
matrix was used to cluster populations using the Fitch-Margoliash method
(Fitch & Margoliash, 1967), available on the PHYLIP package (Felsenstein,
1989).
RESULTS
Chromosome numbers for the 19 populations are shown in Table 1.
The chromosomes have localized centromeres, although these were often
TABLE1. Collection sites, chromosome counts and head structure characteristics for the
populations used in this study. Note that two sympatric species have been collected from
Mt Tomah
Population
Black Mountain, ACT
Black Range, NSW
Molonglo Gorge, NSW
Pebbly Beach, NSW
Sassafras Rd, NSW
Tallaganda State Forest, NSW
Monga, NSW
Big Badja Mountain, NSW
Mt Tomah, NSW
(E. leuckartit)
Kanangra, NSW
Mt Dromedary, NSW
Surprise Valley, Tas
Lake Pedder, Huon R, Tas
Crotty Kelly Basin Rd, Tas
Gloucester Tops, NSW
(taxon E, Tait & Briscoe, 1990)
Gibralter Range, NSW
Tindeny Range, NSW
(taxon G, Tait & Briscoe, 1990)
Siding Springs, NSW
Mt Tomah, NSW
(C. tomahmontis)
Grid Ref
Head
Structure
Sex
chromosomes
absent
absent
absent
absent
absent
absent
absent
absent
absent
X,X,Y
none
none
XY
XY
XY
XY
XY
XY
absent
absent
absent
absent
absent
shallow pit
XY
XY
XY
none
XY
none
modified papillae
deep pit
with spines
modified papillae
deep pit
none
XY
XY
XY
A
'k.
Figure 2. Male meiosis from Black Mt. (2n = 33), diplotene. Note the presence of 16 elements,
one consisting of three chromosomes [arrow).
difficult to see. However, this was confirmed from anaphase preparations, in
which it was clear that there were localized spindle fibre attachment sites.
The existence of markedly heteromorphic chromosome pairs in the
majority of populations was interpreted as representing an XY mode of sex
determination. In six of the populations, no dimorphism was apparent.
However, in some cases (Lake Pedder, Black Range, Molonglo Gorge),
dimorphism in populations with similar morphology and the same chromosome
number implies that X and Y chromosomes may have been present, but
this heteromorphism was not detectable with the resolution achieved.
Three chromosomes appear to be associated at meiosis in the Black Mt.
population (Fig. 2), which has one chromosome fewer than the closest
populations analysed. This may represent an X,X,Y or XY,Y, system, derived
from a fusion or interchange between an autosome and a sex chromosome.
Figures 3 to 7 show sample karyotypes of 2n = 18, 26, 30, 32 and 34
forms respectively. Two general karyotypic forms are apparent. In the
Gibralter Range population (Fig. 5), two distinct size classes are visible. There
are five pairs of large chromosomes gradually decreasing in size, the smallest
still 70°/o of the size of the largest, and then a large drop in size to the
smaller group, which again decreases in size gradually. This bimodal pattern
is repeated in the Tinderry Range population, although the two karyotypes
differ considerably (Fig. 10). In contrast, all of the other karyotypes show a
gradual and relatively uniform size decrease. This is well illustrated by Figure
8 which shows the standardized chromosomal lengths averaged for 19 cells
from the Tallaganda (2n = 34) population, with standard errors included.
KARYOLOGY OF AUSTRALIAN ONYCHOPHORA
Figures 3-5. Karyograms of male specimens Fig. 3. Lake Pedder, Tas. (2n = 18). Note the
size disribution of the chromosomes compared with the more northern forms (see Fig. 9).
Size range 2.3-4.6 pm; Fig. 4, Gloucester Tops (2n = 26). Size range 1.0-3.2 pm; Fig. 5 ,
Gibralter Range (2n = 30). Note the dual symmetry in size distribution. Size range 0.9-6.7
Pm.
Figure 9 shows a graphical comparison of chromosomal lengths in the
Crotty-Kelly Basin Road and Lake Pedder populations (both 2 n = 18) and
Figure 10 a similar comparison between the 2 n = 30 karyotypes from Gibralter
Range and Tindeny Mountains. A matrix of chromosomal distances for the
2 n = 34 populations is given in Table 2 , and these form the basis of the
dendrogram shown in Figure 11.
D. M. ROWELL et a1
Figures (i-7. Fig. ti, Karyogram of a male specimen of Euperipatoides leuckartii from M t Tomah
:2n = 32, XY). Size range 0.8-3.0 pm; Fig. 7, Karyogram of a male specimen of E. leuckartii
from Big Badja Mt. (2n = 34. XY). Size range 0.5-3.2 pm.
DISCUSSION
Chromosomal variation
Montgomery (1900) observed a complement of 2 n = 28 chromosomes in
the South African peripatopsid Peripatopsis balfouri. This is within the range
observed here but, although they are currently ascribed to the same family,
little is known regarding the phylogenetic affinities of this species with those
under study.
The use of.standardized measures is necessary for peripatopsid chromosomal
data because, owing to colchicine sensitivity, it is not possible to produce
cells at exactly the same level of contraction. In the karyograms produced
KARYOLOGY OF AUSTRALIAN ONYCHOPHORA
.2
0
1 2
;
.: -,
t;
;
..
Ill
$1
1 1 I.! [ : ; I 1 l , - , l ( i 1;
I < !!4:'~b:!I:':~:!:!:~t.!->:'l,
1 , ;'~.''t,;II.;I
,;:I.!.:.!.g
('hromnsomr nurnt~chr
Figure 8. Length distribution and standard deviation in chromosome lengths based on
measurements from 19 cells from the Tallaganda State Forest population (2n = 34, XY).
"
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
Chromosome number
Figure 9. Comparison of chromosome length distributions for Crotty-Kelly Basin Road and
Lake Pedder populations (2n = 18).
(Figs 3-7), homologue status was assigned on the basis of similarity in size
and, where visible, centromere position. However, centromeres were not
always visible and some size disparity did occur in chromosomes that were
clearly homologous (see pair 1 in Fig. 5 ) . Thus, it was considered most
appropriate to represent every chromosome individually in the standardized
2
,
.Gibralter
I
Range
1
n
Chromosome number
Figure 10. Comparison of chromosome length distributions for Gibralter Range and Tindeny
Mountains populations ( 2 n = 30). Gibralter Range values are the average for 10 cells standard errors shown.
T.+BI.E
2. Matrices of karyotypic distance measurements for the 2 n = 34 populations
Tallaganda
Big Badja
Mt.
Monga
--
Big Badja Mt.
Monga
Sassafras
Mt Dromedary
Motonglo Gorge
Pebbly Beach
Black Range
C. tomahmontis
0.105
0.076
0.097
0.098
0.07!)
0.105
0.1 15
0.196
-
-
Mt
Molonglo
Sassafras Dromedary
G.
-
-
Pebbly
Beach
Black
Ra.
-
0.140
0.1 10
0.122
0.093
0.1 18
0.135
0.228
length plots, rather than to average homologue lengths. Nevertheless, the
value of standardized length measure comparisons is dependent on two
assumptions. These are:
(i) That internal size relationships remain constant within a karyotype
irrespective of stage of contraction; i.e. that a standardized plot from a
prophase cell, for example, can be compared directly with a plot from a
metaphase cell.
(ii) That the overall lengths of the various karyotypes are the same at the
same level of condensation; no material has been added or lost since
divergence.
Because peripatopsids are generally rare, only small samples were available
from many sites, and chromosomal preparations were often at different
mitotic stages. Consequently, it is important that assumption (i) is fulfilled.
KARYOLOGY OF AUSTRALIAN ONYCHOPHORA
Black Range
Mt Dromedary
-
Sassafras
Pebbly Beach
Tallaganda
-
-
Monga
Molonglo Gorge
I
Big Badja Mt.
I
I
C. tomahmontis
1
I
I
I
0.00
0.075
0.15
Figure 11. Fitch-Margoliash tree based on the chromosomal size deviations for all 2 n = 34
populations shown in Table 2.
Unfortunately, sufficiently large samples were not available in this study to
test this assumption, but it is supported by the size of the standard errors
in Figure 8, which are small, despite the fact that the cells measured were
clearly in different states of condensation. The assumption of concerted
condensation has also been made in other studies of chromosomal evolution
(Martin & Hayman, 1965).
Although assumption (ii) is also difficult to test, the presence of only very
minute amounts of C-banding material indicates that the gain or loss of
heterochromatin is not a major factor in the chromosomal evolution of this
group.
Another factor which may introduce bias is that sex chromosomes were
observed to vary among populations more than autosomes, but because they
could not be distinguished in all populations, it was necessary to include
them in the plots, ordered by size. Consequently, chromosomes of the same
rank in different populations may not be homologous, artificially increasing
apparent differences. However, this may be countered somewhat by the fact
that the ordering of chromosomes without any clear evidence as to homology
may result in an underestimate of the karyological dissimilarity, especially
when several pairs are of similar length.
Martin & Hayman (1965) used mean percent length of each chromosome
to compare four species of the family Macropodidae. In their study they
used as a working hypothesis that chromosomes of the same size were the
same for the purposes of evolutionary inference. The dangers of this approach
are obvious, particularly in the absence of G-band data. For this reason, the
use of overall size distributions as in the present study is considered to be
a more reliable way of visualizing similarity than the use of data based on
individual chromosomes.
1.50
D. M. ROWELL et al.
Given these caveats, tentative conclusions can be drawn regarding the
relationship between chromosomal distance measurements, and relationships
among the species and populations. If it is assumed that chromosomal
evolution in onychophoran groups sharing the same chromosome number
proceeds primarily by small, random but numerous changes, such as minor
duplications, deletions, and translocations, then the chromosomal distance
measurements will become greater as populations diverge. However, as it is
not possible to distinguish homologues with certainty, once sufficient changes
have occurred to change the rank order position of a chromosome pair, the
chromosomal distance may change markedly and unpredictably, and then
gradually increase as before. This 'ratchet' effect, with gradual increases in
chromosomal distance over time, interrupted by sudden perturbations when
the relative rank of chromosomes changes means that, even with a constant
mutation rate, the measure will not accurately reflect the extent of evolutionary
divergence, except within the inter-ratchet periods. If, however, the group
under study falls within this period, distance and divergence time will be
correlated.
Six distinct groups can be defined on the basis of chromosomal distance,
the number and size distributions of the chromosomes and morphology of
the animals-2n = 18, 26A, 26B, 30A, 30B and 32-34. Each of these groups
shows a discrete geographic distribution, and it is suggested that each is a
distinct evolutionary lineage. These are discussed below.
2n = 18 group
This group, confined to Tasmania, (Crotty-Kelly Basin Road, Surprise
Valley and Lake Pedder) is of special interest. Although all specimens
collected at Crotty-Kelly Basin Road were previously reported to belong to
E. leuckartii (Ruhberg, 1985), none of our specimens from these three localities
conforms morphologically to E. leuckartii from mainland Australia, which has
here been placed in the 2n = 32-34 grouping below.
This group shows the lowest chromosome count in this study. Although
conservative in number, there is, nevertheless, marked variation within the
group. In the southern form (Lake Pedder) the difference in size between
the smallest and largest chromosomes is less than twofold, and no sex
chromosomes are visible (Fig. 3). In contrast, the northern forms (CrottyKelly Basin Road, Surprise Valley) have clearly defined sex chromosomes
and a distinctive size distribution, with a sixfold difference between the
smallest and largest chromosomes (Fig. 9).
Karyotypic distance measures show a marked difference between Lake
Pedder and Crotty-Kelly Basin Road (D, = 0.29), but less difference between
Lake Pedder and Surprise Valley (D, = 0.18). The Crotty-Kelly Basin Road
population and Surprise Valley populations are more similar to each other
(D, = 0.13) than either is to Lake Pedder.
The extent of the chromosomal differences between the Lake Pedder
population and the two more northern forms is believed to be sufficient to
recognize the southern form as a biological species distinct from the northern
forms. This is an encouraging finding, because electrophoretic analysis also
indicates the existence of distinct northern (Crotty-Kelly Basin Road and
Surprise Valley) and southern (Lake Pedder) forms (Briscoe & Tait, 1993).
KARYOLOGY OF AUSTRALIAN ONYCHOPHORA
1\51
However, in the case of Crotty-Kelly Basin Road and Surprise Valley, whose
populations share their general karyotypic form and heteromorphic sex
chromosomes, neither the chromosomal nor the electrophoretic differences
are considered to be sufficient to draw such a conclusion. It is not yet
known whether the differences among the three populations are reflected in
their morphology.
2n = 26 group, types A &? B
These two groups are comprised of the populations from Siding Springs
and Gloucester Tops respectively. As mentioned previously, specimens from
both sites possess distinctive male head structures, however the morphology
of these structures differs markedly. In males from Siding Springs the
structures consist of modified papillae only, with no alteration to the shape
of the head, while in Gloucester Tops males there is little modification of
the papillae but a distinct pit behind the antennae with a ridge posteriorly
(Tait & Briscoe, 1990). The degree of morphological divergence between
these two groups would suggest that the shared chromosome number is quite
possibly a result of convergence and, until further morphological data become
available, it is considered appropriate to refer to the Siding Springs and
Gloucester Tops forms as '2n = 26 type A' and '2n = 26 type B' respectively.
The two populations differ by an average karyotypic distance of 0.16.
2n = 30 Group, types A &? B
Karyotypes of 2n = 30 were observed for Tinderry Range and Gibralter
Range populations. Both show a tendency to dual symmetry; that is, the
chromosomes fall into two distinct size groups (Fig. lo), reminiscent of the
karyotypes of birds and many reptiles. However, on the basis of the
considerable differences in internal size relationships of the chromosomes,
and the morphology of the animals, these are considered to represent two
distinct forms or lineagesPb2n = 30 type A' (Tinderry Range) and '2n = 30
type B' (Gibralter Range).
The head structure of males of the Gibralter Range population is similar
to that of a species previously described from Mt Warning (Briscoe & Tait,
1990), consisting of a group of large papillae between the antennae. In
contrast, the head structure of the Tinderry Range males is extreme, consisting
of a deep pit containing two evertable spines in the male and a smaller pit
lacking spines in the female (Tait & Briscoe, 1990).
The average variance of the two karyotypes calculated from the data of
Figure 10 is 0.43 and, although such comparisons should be treated with
caution, this is the largest disparity observed for any two onychophorans.
2n = 32-34 group
This group contains the only populations on mainland Australia which do
not possess distinctive head structures in the male. Given the apparently low
rate of migration of members of this phylum, the large geographical range
occupied by the 2n = 32-34 group (over 450 km north to south) suggests
that it is the product of an ancient radiation. Despite the shared chromosome
number of 2n = 34 with the Mt Tomah population of C. tomahmontis, the
latter is not considered to be part of this grouping.
15'1
D. M. ROWELL
et
al.
On the basis of the shared count of 2n = 34 between the Mt Tomah
population of C. tomahmontis and eight of the populations in this group, this
number may represent the ancestral complement for the two groups. Certainly,
within the 2n = 32-34 group the evidence supports a 2n = 34 ancestral form,
on the basis of the very wide distribution of this karyotype. If the ancestral
number is 34 for this group, a plausible explanation for the origin of the
2n = 33 karyomorph (Black Mt.) is that it is derived from a 2n = 34 karyotype
via a fusion between an autosome and a sex chromosome, resulting in a
chain of three chromosomes at meiosis as shown in Figure 3. Similarly, the
2n = 32 forms, which have a more limited distribution, may be the product
of the fixation of an autosomal fusion or the redistribution of one autosomal
pair among the other autosomes. The latter is more probable as there is no
evidence of a fusion in size plots for Kanangara and Mt Tomah populations.
It should be noted, however, that geographic range data are not complete,
and populations may well exist in areas which have not yet been sampled.
Cephalofovea tomahmontis is distinctly different from the other populations
on the basis of its head structure and disposition of crural papillae (Ruhberg
et al., 1988). This is borne out here, as the chromosomal distances between
C. tomahmontis and the other populations are high (Table 2).
Afinities among the 2n
=
34 karyomorphs
The relationships in the dendrogram shown in Figure 11 can be explained
in terms of geography and the predicted behaviour of the distance measure.
Cephalofovea tomahmontis is morphologically quite distinct from the rest of the
populations and, as would be predicted from this, occupies a basal position.
The inclusion of the Tallaganda, Monga, Pebbly Beach, Big Badja Mt. and
Molonglo Gorge populations into an exclusive grouping is consistent with
their geographic distributions, as the distances between adjacent populations
in this cluster are small (Fig. I).
Mt Dromedary, Sassafras and Black Range have the greatest geographic
separation, being the most southerly, northeasterly and northwesterly sites
respectively. Nevertheless, Mt Dromedary and Sassafras cluster together. As
discussed above, at a certain point in the divergence of populations, the
relative positions of specific chromosomes may change, resulting in an
unpredictable change in the genetic distance. Thus, the internal consistency
of the tree up to the level of Mt Dromedary and Sassafras would suggest
that the homologous chromosomes appear in the same rank order in the
other five populations, but that in either the Sassafras or Mt Dromedary
populations the order may have changed-the ratchet has 'clicked'. Thus,
these two populations which show the greatest pairwise geographical separation
are actually grouped together, owing to a reduction in the chromosomal
distance value. The population from Black Range, which occupies a deep
position on the tree, is the most westerly of the populations. O n the
assumption that the ratchet has already clicked once at the Sassafras/Dromedary
level, the same phenomenon may well have occurred in this population, and
its placement in an evolutionary sense cannot be inferred with any confidence.
KARYOLOGY O F AUSTRALIAN ONYCHOPHORA
153
CONCLUSIONS
The data presented here show that, in contrast to the relatively conserved
morphology in this family, there has been extensive karyotypic evolution
and repatterning in the viviparous Peripatopsidae of eastern Australia. The
results of this study demonstrate that karyotypic data are particulary valuable
for distinguishing amongst morphologically very similar populations, and that
groupings based on chromosomal data are consistent with those inferred
from morphological data.
The fact that the dendrogram produced is consistent with geographic
distribution and evolutionary phenomena lends support to the assumptions
that chromosomal evolution in this group is
behind the method-i.e.
characterized by the accumulation of many small rearrangements which alter
comparative chromosome lengths over time. Because C-banding has proved
to be uninformative, other techniques are currently being explored in order
to identify the particular types of chromosomal changes that are involved in
the karyotypic evolution of the Australian Onychophora.
ACKNOWLEDGEMENTS
The authors wish to thank Ray Cameron, Mandy Reid, Ian Scott and
Simon Conroy for assistance with field collection. Mandy Reid also provided
helpful comments on the manuscript. This work was supported by an
Australian Research Council grant to DMR.
REFERENCES
Briscoe DA, Tait NN. 1993. Rare and localised species-peripatus. In: Smith SJ, ed. Tasmanian
Wilderness and World Heritage Values. Royal Society of Tasmania, Hobart, pp. 136-138.
Felsenstein J. 1989. PHYLIP-phylogenetic inference package (Version 3.2). Chdistics 5: 164-166.
Fitch W, Margoliash E. 1967. Construction of phylogenetic trees. Science 155: 279-284.
Hebert PDN, Billington N, Finston TL, Boileau MG, Beaton MJ, Barrette RJ. 1991. Genetic
variation in the onychophoran Plicatoperipatus jamaicensis. Heredity 67: 221-229.
Martin PG, Hayrnan DL. 1965. A quantitative method for comparing the karyotypes of related species.
Evolution 19: 157-161.
Montgomery TH. 1900. The spermatogenesis of Peripatopsis (peripatus) balfouri up to the formation of
the spermatid. Zoologische Jarhbiicher. Abteilung fuer Allgemeine Zoologische und Physiologie der Tiere 14:
277-368.
Rowel1 DM. 1985. Complex sex-linked fusion heterozygosity in the Australian huntsman spider Delena
cancerides (Araneae: Sparassidae). Chromosoma 93: 169-1 76.
Ruhberg H. 1985. Die Peripatopsidae (Onychophora). Systematik. Okologie. Chorologie und
Phylogenetische Aspekte. Zoologica 137: 1-183.
Ruhberg H, Tait NN, Briscoe DA, Storch V. 1988. Cephalofovea tomahmontis n. gen., n. sp., an
Australian peripatopsid (Onychophora) with a unique cephalic pit. ZooloNcher Anzeiger 221: 117133.
Saenger H. 1869. Peripatus capemis Sr and Peripatus leuckartii n. sp. Transactions of the Second Congress
of Russian Naturalists in Moscow pp. 239-262; translated into French; Bouvier EL 1901. Apropos
d'un travail de H. Sanger sur les Peripates. Bulletin Societ2 Philomatique de Paris 3: 9-36, PI. 12-13.
Tait NN, Briscoe DA. 1990. Sexual head structures in the Onychophora: unique modifications for
sperm transfer. Journal of Natural History 24: 1517-1527.
Tait NN, Stutchbury RJ, Briscoe DA. 1990. Review of the discovery and identification of Onychophora
in Australia. Proceedings of the Linnean Society of New South Wales 112: 153-171.