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Aust ra l ian Jou rnal

of

Entomology , 1996, 35: 271 -278

27

1

The Brooks-Dyar Rule and Morphometrics of the Processionary

Caterpillar Ochrogaster Zunifer Herrich-Schaffer Lepidoptera:

Thaumetopoeidae)

GRAHAM

J.

FLOATER

Departm ent of En tomology , University of Queensland, Q ld 4072.

ABSTRACT

Vario us insect species display a unif orm geo me tric increase in size during th e larval

stage (that is they follow the B rooks-Dyar rule). Here , results of larval development are presented

for the bunny-tailed moth , Ochrogaster lunifer. The processionary larvae of this species live in

a com munal coh ort, an d mou lt en masse in the silken nest sp un at the base of their host tree (usually

a phyllodinous acacia). Th e exuviae, which remain buried in the accumulated silk and frass of the

nest, provide a life history record of the larval co ho rt. Larval exuviae were collected fro m 773 cohorts

a t 37 localities in southeastern Q ueensland between Novem ber 1993 and May 1994. Th e 6,948 exuviae

examined were from cohorts feeding on Acacia

concurrens

Pedley. Head-capsules showed a strongly

uniform geometric increase in size through eight larval instars, supporting the Brooks-Dyar rule.

The number

of

instars did not vary between trees

or

localities.

A

bimodal distribution

of

final instar

head-capsule widths was shown to b e a sexual dimorph ism, an d s imilar bimodal dis tr ibutions were

foun d fo r instars V-VII. P upal size was also sexually dimorp hic. Th e geometric size increase from

one larval instar to the next holds for both males and the larger females. Th e geometric rule was

tested using larval cohorts reared

on

A . concurrens in the greenhouse through instars

I-IV;

development was rem arkably s imilar to that in the f ield. Larval growth pa t terns of 0 lunifer a r e

very different from the structurally similar bag-shelter moth. The ability to distinguish different

instars

of

0

unifer

with a high degree of precision from field-collected exuviae will allow accura te

comparisons of developme nt, survival and dispersal of larvae in different gr oup sizes, on different

trees and in different localities.

Introduction

Ground-nesting populations of the bunny-tailed

moth, Ochrogaster lunifer Herrich-Schaffer

(Lepidoptera: Thaumetopoeidae), are common

and widespread along the eastern seaboard of

Australia (Froggatt 1896; Floater 1996a). The

processionary larvae live in a communal cohort

at the base of the host tree (usually a species of

phyllodinous acacia), moving up the trunk in

single file to feed in the canopy at night. The larvae

moult

en

masse in the silken nest spun at the base

of the tree, where the exuviae remain buried in

accumulated silk and frass. These exuviae

therefore represent a life history record of the

larval cohort.

I

reasoned that

if

different larval

stages could be distinguished within the same nest,

the moth would be an ideal subject for ecological

studies on the mortality and development of

individuals within and between populations.

The number of larval instars in insect species

is often determined from

a

frequency distribution

of larval size. A representative sample of larvae

of all ages is collected from the field, and various

measurements (often the width of the head)

recorded. Alternatively, the moulted skins of

larvae may be collected, as long as the head

capsules, or other well-sclerotised structures,

remain intact.

I f

the growth rates of larvae are

relatively uniform, the resulting frequency

distribution should consist of

a

series of peaks,

with each peak representing one instar (see Daly

1985 for review). In practice, however, the

frequency distribution is often ragged with

overlapping peaks caused by sampling error,

environmental variation, sexual dimorphism,

parasitism and genetic differences between

individual animals.

A second method of analysis involves the

Brooks-Dyar rule (Brooks 1886; Dyar 1890;

Hutchinson and Tongring 1984). The rule states

that larval head widths in successive stages

describe a regular geometric progression (Dyar

1890) with the following equation:

where X is the instar number (1, 2, 3, etc.); Y is

head-capsule width; and a and b are

constants. The equation serves both as a growth

curve and as

a

method of checking for an

overlooked instar in

a

frequency distribution (Daly

1985). To check for a hidden instar, equation

1)

is made linear by taking the natural logs of both

sides, giving:

1nY = c

+

bX. (2)

where c = In(a). The relationship between 1nY and

X should be a straight line with slope b, and

therefore a significant deviation from a straight

line indicates

a

missing instar.

While the Brooks-Dyar rule holds for many

species (e.g. Dyar 1890; Taylor 1931; Gaines and

Campbell 1935; Mackay 1978), many exceptions

exist (e.g. Kishi 1971; Craig 1975; Schmidt

et

al.

1977; Allsopp and Adams 1979; Jobin

et

al. 1992).

Here the different larval stages of

0

lunifer are

distinguished using the Brooks-Dyar rule and the

subsequent model tested using cohorts reared in

the greenhouse.

Y = aebX (1)

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212 0

.

FLOATER

Methods

0

unifer

is univoltine.

A

survey of larval exuviae

f rom one genera t ion was conducted f rom

November 1993 (when eggs hatched) to Ma y 1994

(when final instar larvae dispersed to enter

prepupa l diapause). In November 1993, over 4,000

acacia trees of various species were examined f or

egg batches at 37 localities in southeastern

Quee nsland. D etails of localities can be f ou nd in

Floater (1996a). A tota l of 773 egg batches w ere

recorded on six

Acacia

species: A .

concurrens

Pedley,

A . leiocafyx

Domin (Pedley),

A .Jimbriata

Cunn. ex

G .

Don ,

A . aulacocarpa

Cunn. ex

Bentham,

A . implexa

Bentham and

A . maidenii

F.

v. Mueller. Over 95 of trees with eggs were

A . concurrens

and the rest of the study focused

on larvae feeding on this species.

Localities w ere revisited

2

m onth s later (in late

Janu ary 1994), when larvae were in the fifth instar.

Co hor t extinction (100 mortality) is comm on

in the early instars, an d the rem ains of all extinct

cohorts were collected and examined for larval

skins. Head-capsule widths of the 10largest skins

in each extinct coho rt were then measured t o give

an estimate of larval size at the time o f,

or

shortly

before, extinction. Depending on the time of

ex t inc t ion , the head-capsu le wid ths could

therefore represent instar I, 11, I11 or VI.

Measurements were made

to

the nearest 0.1 m m

using a graticule set in a stereo microscope.

In Ju ne 1994, after the final instar larvae had

migrated from trees to pupate underground, all

remaining nests were collected from 29 of the 37

localities. Th e nest m aterial was examined in the

4001

labo rato ry. Exuviae of the penultimate (seventh)

instar (i.e. the last larval moult before the final

instar larvae leave the nest) were estimated by eye,

and th e head-capsule of each removed. Instar VII

cast skins appeared to be distinctly larger than

other skins in the nest, and were often matted

together. If doubt existed over assigning a skin,

the head-capsule was removed an d added to the

rest. For each cohort, head-capsule widths were

measured t o the nearest 0.1 m m, a nd a frequency

distribution of sizes plotted. If a discontinuity

existed in the lower end of the distribution (i.e.

there appeared to be skins in distinct size classes

below instar VII; see Fig. l) , only da ta above the

discontinuity were used in subsequent analyses.

In nests where larvae had defoliated their host-

plant an d emigrated before the penultimate mo ult,

all skins present were removed for analysis. Once

again, for each cohort, a frequency distribution

was plotted, and only data above the highest

discontinuity (i.e. skins tha t appeared to be in the

highest distinct size class) were subsequently used.

This distribution represented either instar

V

or

instar VI, depending on the stage at which

emigration to ok place. In all, 6,948 head-capsules

were measured fro m exuviae collected in Jan ua ry

and June.

T o measure the head capsule widths of known

larval instars, cohorts were reared

to

the fourth

instar stage in :he greenhou se. I n December 1994,

five egg batches were collected from Kuraby in

eastern Brisbane (this locality was not par t of the

regional survey from the year before). Each batch

was then ke pt in a small plastic tu b until the first

larval moult. Th e first instar larvae d o not feed,

1

I Ins tar

VII

Instar VI

I

1

I

I

I \

.___

1.8 2 2.2 2.4

2.6

2.8 3 3.2 3.4

3.6

3.8 4 4.2 4.4 9 6

head-capsule width

(mm)

Fig. 1.

Frequency distribution

of

head-capsule widths in a larval cohort. The distribution describes instars

V-VII,

with each

instar displaying a bimodality of sexes.

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MORPHOMETRICS

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0 LUNIFER 213

an d remain in the m at of scales deposited by the

female moth over her egg batch. When all the

larvae in a cohort had m oulted, 20 head-capsules

were removed at random from the tub for

subsequent examination.

The newly moulted second instar larvae were

transferred from the tubs to greenhouse plants.

Plants were well watered and fertilised. Each

cohort was placed, along with the scale mat, at

the base of a potted plant of

A . concurrens

whereupon the larvae ascended the plant to feed.

30 1

I n=25)

5

0

The co horts were kept on the plants for

6

weeks,

and during this time the larvae moulted four times.

After each moulting event, all skins from a cohort

were removed fro m the base of the plant. Of these,

20

skins were chosen at random and the head

capsules removed. C onseque ntly, at the end of the

experim ent, a collection of head capsules had been

made, with

20

capsules of each instar fr om each

cohort. The width of each head capsule was

measured to the nearest

0.04

mm using a stereo

microscope.

0 5 0.9 1 . 3 1 .7 2 . 1 2 . 5 2 . 9 3 . 3 3 . 7 4.1 4 . 5 4.9

head-capsule width

mm)

30 -

I

25

I1

20 w

G

c

2

1 5

ct:

\

I

10

I11

VI

0.5

0.9

1 .3 1 .7 2 .1

2.5 2.9

3 . 3 3 . 7

4.1

4 . 5

4.9

head-capsule width mm)

Fig.

2.

Frequency distribution of larval size modal head-capsule width) from field-collected cohorts, with seven peaks representing

seven instars:

upper)

Distribution o f fema le sizes; the upper female) mode head-capsule width was calculated for the largest

exuviae in each coh ort and plotted a s a single data entry;

lower)

Distribution

of

male sizes; lower mo de head-capsule widths.

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214

G.

J.

FLOATER

To measure final instar moults and to

investigate sex differences, eight nests were

collected from trees of

A . concurrens

in May 1995.

The nests contained final instar caterpillars that

were close to prepupal diapause. The resulting

pupae were removed from their silken cocoons in

September 1994, sexed and measured (length and

width). The head-capsule of the final moult

contained in the cocoon was measured also.

Consequently, the sex and size of final instar

larvae could be ascertained.

2 1

1.5

.I

Results

Sexual

dimorphism.

Female pupae, with length

24.4

k

1.3 mm (mean

SD,

n = 29) and width

9.8

0.5

mm, were significantly larger than male

pupae, with length 21.1

k

1.2 mm (n

=

25) and

width 8.2 0.6 mm. Furthermore, the bimodal

distribution of head capsule widths of final instar

larvae was shown to be the result of sexual

dimorphism. Female head capsule widths were

significantly larger 5.51 0.39 mm, n = 29) than

- . 5

1

INSTAR

2

c

w

.5

.

m

1

0 1 3 4 5 6 7 8

INSTAR

Fig. 3. A

regression showing'the geometric increase

of

larval size (represented as ln[head-capsule width]) with

instar number: upper) Female larvae; lower) Male larvae.

corresponding

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MORPHOMETRICS

OF

0 LUNIFER 275

those of male larvae (4.65 f 0.19 mm, n =

25);

ANOVA F

= 17.13;

P =

0.002.

This difference

between the sexes can be seen in earlier instars.

Fig. 1 shows the distribution of head capsule sizes

from a single nest. The distribution represents

three instars (instars

5

6,

7)

and clearly

demonstrates the bimodality of each instar, even

in instar 5 The differences in size between males

and females must be taken into account when

3

2 5

3 2

2

.

1.5

2

-

l

.

. 5

a 0

- - . 5

1

1.5

W

--

c

W

c:

N

d

n

z

d

assessing the validity of the Brooks-Dyar rule.

Consequently, in analyses of larval growth, the

upper an d lower modes in the distribution of head-

capsule widths for each instar from instar

5

to

8

are considered separately.

Larval development in field-collected cohorts. By

plotting the modal head-capsule width of cohorts

from the Janu ary an d J un e collections in 1994,

a

continuous frequency distribution of head-

1

2

3

4

5

6

7

8

INSTAR

Fig.

4. A regression showing the geometric increase in the variation of larval size with increasing instar number.

I

120

100

80

60-

L 2

G

40

IV

20

1 1

0.64 0.8 0.96 1.12 1.28

1.44 1.6

1.76 1.92

head-capsule width

(mm)

Fig. 5 .

Frequency distribution of head-capsule widths of larvae from five cohorts reared in the greenhouse. The four peaks

represent instars I-11.

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216 0

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FLOATER

capsule widths was created for instars

I-VII

(Fig.

2). Becauseof the differences in head size between

males and females, two distributions were created.

The first distribution (Fig. 2a) represents upper

(i.e. female) modes, while the second (Fig. 2b)

represents lower (male) modes.

In

both graphs, a

single mode was calculated for cohorts collected

in January (instars

I-IV),

and hence both graphs

have an identical distribution of head sizes from

0 5 to 1.7 mm.

Examining the frequency distribution of female

modes, there appear to be seven major peaks with

modes at approximately 0.6 mm (peak l), 0.85

mm(peak2), 1.1 mm(peak 3), 1.4 mm(peak4),

2.15 mm (peak

5),

2.95 mm (peak

6 ,

4.4 mm

(peak 7). Peaks 5-7 (from the June collection) are

quite distinct, and peak 7 appears to be bimodal.

Cohorts collected at some localities had distinctly

larger larvae than cohorts at other localities,

creating a dichotomy

of

sizes rather than a

continuous range of sizes. The reason for this

geographical distinction is unclear. Peaks 1-4

(from the January collection) are also quite

distinct, though a possible gap exists between

peaks 4 and

5 . The first instar moult (peak 1) is

particularly pronounced.

Assuming the seven peaks in Fig. 2a to represent

seven instars, the following categories were

created: instar

1:

0.6; instar 2: 0.8-1.0; instar

3:

1.1-1.3; instar 4: 1.4-1.7; instar 5 : 1.9-2.4; instar

6:

2.6-3.4; instar 7: 3.7-4.9. A regression was then

plotted of larval size against corresponding instar

number for each cohort (Fig. 3a). The result is a

near perfect straight line with equation InY

=

0.326X 0.845; R* = 0.994, and consequently,

the seven peaks do appear to correspond to seven

consecutive instars. Furthermore, the line predicts

the mode head-capsule width of final instar

females (instar 8) accurately (Fig. 3a). A similar

analysis

of

lower size modes (corresponding to

male head-capsule widths) gives a straight line with

equation

InY =

0.305X 0.803;

R2

0.994 (Fig.

3b). Because the increase in size from one instar

to the next is geometric, the size ratio of one instar

to the next is a constant (eb). This ratio is 1.39 for

females and 1.36 for males.

The mean larval size of males and females is not

the only variable to increase exponentially with life

stage. The range

of

larval head-capsule widths

around the mode also gets progressively larger as

the larvae moult from one instar to the next. While

the relationship is not as tight as for the modes,

Fig. 4 clearly shows the geometric increase in size

range with instar number. The straight-line

equation describing the relationship is 1nY

=

0 517X 1.22; RZ 0.961 (where

Y

is the range

of head-capsule widths in each instar).

Larval development

in

greenhouse-reared

cohorts.

From the field results, larval development of 0

lunifer

does appear to follow the Brooks-Dyar

rule, with a geometric growth rate through eight

larval instars. To test the validity of the findings

from the field-collected data, the size distributions

of greenhouse-reared larvae were compared to the

regression models calculated from the field data.

When pooled together, the size distribution of

I

i

-.6

.4

- . 2

0

. 2

.4

.6

predicted

larval

size

In

hcw)

(mm)

Fig. 6 . A

test of the Brooks-Dyar rule in

0 lunifer

comparing observed head-capsule widths

of

greenhouse-reared larvae

with those predicted for respective instars (I-IV). The solid line shows the 1:l ratio of observed to predicted sizes, on which

the points were predicted to lie.

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MORPHOMETRICS OF

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LUNZFER 2

greenhouse-reared larvae showed distinct classes

for instars 1-4, with prominent modes at 0.60,

0.84, 1.12 and 1.60 mm (Fig. 5 . The results from

the greenhouse experiment were compared with

the geometric model calculated from the field

data. A predicted head-capsule size was first

calculated as the mean of male and female head-

capsule widths for each instar from the field data.

Fig. 6 shows the strong relationship between

predicted head-capsule sizes and those observed

in the experiment for instars 1-4. The points lie

very close to a 1 relationship, and the regression

is highly significant (Y

= 0.006 +

1.022X; R 2 =

0.996), supporting the Brooks-Dyar rule. The

straight-line equation describing the relationship

between reared-larval size and instar number is InY

= 0.322X 0.835 RZ = 0.996), which lies

between the lines predicted for male and female

head-capsule widths.

Discussion

Assigning larval instars of 0 lunijer.

When

comparing larval development and survival

between cohorts on different trees and in different

localities, it is essential to be able to distinguish

between larval instars within the same cohort.

Different insect species exhibit different growth

patterns, which may aid or hinder the researchers

ability to assign exuviae to particular instars. In

the Lepidoptera, growth patterns may display

consecutive size ratios that remain constant

throughout development (Dyar 1890; Gaines and

Campbell 1935); that decrease with each moult

(Jobin et al. 1992); or that are highly variable from

one moult to the next (Schmidt et al. 1977). In

some insect species, larval development is so

irregular that assigning individuals to age classes

is simply impossible. For example, AIlsopp and

Adams (1979), in a study of the tenebrionid beetle,

Pterohelaeus darlingensis Carter, were only able

to distinguish one larval instar (the first) and that

due to its colour rather than size. The problem

with assigning independent individuals to stadia

is that even when development is uniform for each

individual, the different developmental rates

between individuals will usually produce overlaps

in instar size; particularly in later instars.

Larvae of 0 lunijer possess two features that

allow accurate assignment of larval exuviae to

respective instars. First, rate of development

appears to be relatively uniform for all larvae

within a cohort. And second, because the rate of

development follows a geometric progression,

instars remain relatively distinct. Even cohorts

reared on young, fertilised plants in the

greenhouse had a remarkably similar rate of

growth to those on older trees of varying condition

in the field. Variation in size exists within each

instar, but rarely overlaps between instars within

the same cohort. As Fig. 4 shows, the range of

larval sizes increases exponentially with instar

number. It is for this reason that in insect species

that do not follow the Brooks-Dyar rule, later

instars with large size ranges overlap with one

another. Only if the mean instar size increases

exponentially along with the range of sizes around

the mean, will later instars remain distinct.

When comparing rates of larval development

between cohorts, it is enough to distinguish the

respective peaks of male and female size for a

particular instar. However, when comparing larval

survival, the researcher must know the number of

individuals in a particular instar. Consequently,

it is more important to distinguish between the

upper and lower size bounds of an instar rather

than the position of the mode. As Fig. 1 shows,

these bounds are quite distinct in a cohort. Clearly,

some subjective judgment is inevitabIe when

assigning the one or two individuals at the

boundary between adjacent instars, but these will

be all but irrelevant in statistical comparisons of

cohort numbers. Comparing larval numbers at

instar VII has the added advantage that instar VIII

exuviae are not present in the nest. Furthermore,

in over 90 of the nests sorted, a distinct gap

existed between instar VI and instar VII, and in

cases where an overlap existed, the number of

individuals at the boundary was never more than

two.

Morphometrics and the 0 lunifer species

complex. While Common (1990) has suggested

that the name

0

lunifer

be restricted to ground-

nesting populations in eastern Australia, the name

has been used by other authors to describe the

canopy-nesting bag-shelter moth (van Schagen et

al. 1992). Although adults of the bag-shelter moth

are structurally similar to those of the bunny-tailed

moth (Common 1990), various behavioural and

morphological differences exist between the

immature stages (Floater 1996a).

The morphometrics o f both moths also suggest

that they are different species. Van Schagen et al.

(1992) described the larval morphometrics of the

bag-shelter moth, and found the following: six

larval instars;

a

decrease in the size ratio from

instar to instar, contradicting the Brooks-Dyar

rule; no relationship between size range and mean

size for each instar; and no sexual dimorphism in

larval size. In contrast, the results of my study on

the bunny-tailed moth show eight larval instars;

a

constant size ratio;

a

geometric increase in size

range from instar to instar; and marked sexual

dimorphism in larval size. It is not clear why van

Schagen et al. (1992) found a greater range (and

variance) of head capsule widths in instar I than

in instar VI, while the mean size increased almost

by an order of magnitude (instar

1:

0.57 0.15

mm S.D.; instar

VI ;

4.80 0.10 mm). The scope

of the investigation was limited (a total of 240

larvae were examined) and a more comprehensive

study of the early stages would be required to

verify whether the range of first instar sizes in the

bag-shelter moth was indeed large, or whether

a

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