1
CLUTCH SIZE AND FORAGING BEHAVIOUR IN
APANTELES SPP. (HYMENOPTERA: BRACONIDAE)
by
Andrew David le Masurier, B.Sc.
A thesis submitted for the degree of Doctor of Philosophy of the University of London and for the Diploma of Imperial College
Department of Pure & Applied Biology, Imperial College at Silwood Park, A scot, Berkshire. May, 1987 2
ABSTRACT
The aim of this thesis is to use the comparative method to study the adaptation of clutch size and foraging behaviour in the parasitoid, A p a n t e l e s g l o m e r a t u s (L.).
A. glomeratus is native to Europe, where its principal host is Pieris brassicae (L.) (Lepidoptera: Pieridae), but in 1883 it was introduced to America as a biological control agent of Pieris rapae (L.).
The optimal foraging behaviour of a parasitoid is affected by host distribution. Early instar larvae of P. brassicae are gregarious, whereas those of P. r a p a e are solitary. The searching efficiency of British A. glomeratus was found to be much higher for P. brassicae than for P. rapae. Apanteles rubecula
(Marshall), in comparison with A. glomeratus , had a higher searching efficiency for P. r a p a e (its only known host). It was therefore predicted that selection should have improved the searching efficiency of American wasps towards that shown by A. rubecula. However, no evidence of such an improvement was found.
The optimal clutch size of a parasitoid depends largely on host size.
P. brassicae is larger than P. r a p a e , and the offspring fitness of American
A. glomeratus ovipositing in P. r a p a e was found to be more severely density- dependent than that of British A. glomeratus parasitising P. brassicae. The optimum clutch size predicted for American A. glomeratus was consequently lower than that predicted for British A. glomeratus. However, no evidence of a lower clutch size in the American population was found.
Brood size in gregarious A p a n t e l e s species was found to correlate with host size, but solitary species do not lie on the same relationship, and are more numerous than expected on the basis of host size. The possible role of evolutionary constraints on clutch size optimisation is discussed.
It was predicted that the sex ratios of gregarious A p a n t e l e s species should be more female-biased than those of solitary species. A comparison of mean sex ratios provided evidence in support of this prediction. 3
CONTENTS
Page
A B S T R A C T 2 LIST OF FIGURES 8 LIST OF TABLES 19 CHAPTER ONE INTRODUCTION 26 CHAPTER TWO BIOLOGY OF HOSTS A N D PARASITOIDS; CULTURING METHODS; AND GENERAL EXPERIMENTAL METHODS 30 2.1. Biology of Hosts and Parasitoids 30 2.1.1. The hosts 30
2.1.1.1. Pier is brassicae 30
2.1.1.2. Pier is r a p a e 32 2.1.2. The parasitoids 33
2.1.2.1. Apanteles glomeratus 33 2.1.2.1. L Host range 33 2.1.2.1.2. Life cycle 33 2.1.2.1.3. Percentage parasitism 35 2.1.2.1.4. Clutch size and brood size 38
2.1.2.2. A p a n t e l e s r u b e c u l a 42 2.1.2.2.1. Host range 42 2.1.2.2.2. Life cycle 42 2.1.2.2.3. Percentage parasitism 47 2.1.2.2.4. Clutch size and brood size 48 2.2. Culturing Methods 48
2.2.1. The hosts 48 2.2.2. The parasitoids 49 2.3. General Experimental Methods 50 2.3.1. Experimental conditions 50 2.3.2. Standardisation of wasps 50 2.3.3. Host dissection method 50
CHAPTER THREE THE FORAGING BEHAVIOUR OF
A. GLOMERATUS FO R P.BRASSICAE 52 3.1. Introduction 52 3.2. Materials and Methods 53 4
Page
3.3. R esu lts 55 3.3.1. The effect of host patch size on the number of
hosts parasitised 55 3.3.2. The effect of time on patch on the number of
hosts parasitised 57 3.3.3. Possible factors affecting the attack rate of A. glomeratus on P. brassicae 57 3.3.3.1. E g g depletion 57 3.3.3.2. Time spent rejecting previously-parasitised hosts and cleaning 64 3.3.4. The effect of host patch size on the duration of
patch visits 76 3.4. D iscu ssio n 76
CHAPTER FOUR A COMPARISON OF THE SEARCHING
EFFICIENCIES OF A. GLOMERATUS
A N D A. RUBECULA F O R P. BRASSICAE
A N D P. R A P A E 89
4.1. Introduction 89 4.2. Materials and Methods 92
4.3. R e su lts 93 4.3.1. Experiment 1: the functional response of British
A. glomeratus to P. brassicae 93
4.3.2. Experiment 2: the functional response of British
A. glomeratus to P. ra p a e 96
4.3.3. Experiment 3: the functional response of British
A. rubecula to P. ra p a e 96 4.3.4. Experiment 4: the functional response of American
A. glomeratus to P. ra p a e 97
4.3.5. Factors affecting the asymptote of the functional
response curves 97
4.4. D iscu ssio n 103 5
Page
CHAPTER FIVE THE EFFECT OF HOST QUALITY ON THE
CLUTCH SIZE OF A. GLOMERATUS 105
5.1. Introduction 105
5.2. H o st Size 111 5.2.1. The effect of host species on parasitoid development 111
5.2.1.1. Materials and methods 111
5.2.1.2. R e su lts 113
5.2.1.2.1. Developm ent tim e 113 5.2.1.2.2. Juvenile su rvivo rsh ip 113
5.2.1.2.3. A d u lt size 116 5.2.1.2.4. C lu tch size 116 5.2.2. Comparison of the optimal clutch sizes of British and A m e rican A. glomeratus 116 5.2.2.1. Materials and methods 116 5.2.2.2. R e su lts 120 5.2.2.2.1. The effect of clutch size on juvenile survivorship 120
5.2.2.2.2. The effect of clutch size on adult size 120
5.2.2.2.3. E g g load 120 5.2.2.2.4. The effect of clutch size on
development time 124 5.2.2.2.5. Comparison of the clutch size fitness
functions 124
5.2.2.2.6. Comparison of the optimal clutch
sizes 127
5.2.3. Comparison of the observed clutch and brood sizes of
British and American A. glomeratus 134
5.2.3.1. Materials and methods 134
5.2.3.2. R e su lts 135 5.2.3.2.1. C lu tch size 135
5.2.3.2.2. Brood size 135
5.3. Host Instar Attacked 136
5.3.1. Materials and methods 136
5.3.2. R esults 136 6
Page
5.4. Previous Parasitism 141
5.4.1 Preliminary experiment 142
5.4.2. M a in experim ent 142
5.4.2.1. Materials and methods 142
5.4.2.2. R e su lts 149 5.4.2.2.1. Frequency of oviposition and probing behaviour in parasitised and
unparasitised hosts 149 5.4.2.2.2. The relationship between observed
wasp behaviour and egg laying 149
5.4.2.2.3. C lu tch sizes 152 5.4.2.2.4. Duration of oviposition behaviour 152
5.5. D iscu ssion 153
CHAPTER SIX HOST SIZE AND THE DISTRIBUTION OF SOLITARY AND GREGARIOUS BROOD S IZ E S I N APANTELES 160 6.1. Introduction 160
6.2. M ethods 161
6.3. Results 163 6.3.1. Brood size frequencies 163 6.3.2. The effect of host size on brood size 163
6.4. D iscu ssion 172
CHAPTER SEVEN REPRODUCTIVE STRATEGIES OF SOLITARY
AND GREGARIOUS APANTELES S P E C IE S 177
7.1. Introduction 177
7.2. M ate rials and M e thods 180
7.3. Results 181
7.3.1. Sex ratio 181 7.3.1.1. Sex ratios of solitary and gregarious
A p a n te le sspecies 181
7.3.1.2. Field sex ratios of A. glomeratus and
A. rubecula 184
7.3.1.3. Factors contributing to the female-biased sex
ratio o f A. glomeratus 184 7
Page
7.3.2. E g g complement 187 7.3.2.1. Comparison of the egg complements of
A. glomeratus and A. rubecula 187
7.3.2.2. Comparison of the fecundities of solitary and
gregarious A p a n te le s species 192
7.4. D iscu ssio n 192
CHAPTER EIGHT GENERAL DISCUSSION 198
ACKNOWLEDGEMENTS 212 R E F E R E N C E S 213 A P P E N D I X I 248 APPENDIX II 249 8
LIST OF FIGURES
Page Figure 2.1 Frequency distribution of the size of egg batches laid by captive P. brassicae on potted Brussels sprouts plants. 31
F igu re 2.2 Frequency distribution of clutch sizes (number of eggs laid
per oviposition) laid by British A. glomeratus in one day-
old P. brassicae larvae presented in patches of fifty on
potted Brussels sprouts plants. 40
F igu re 2.3 Frequency distribution of the size of A. glomeratus broods (number of cocoons per host) obtained from P. brassicae larvae exposed to field parasitism for 3-4 days as first
instars. 41
F igu re 2.4 Newly-hatched first instar larvae of a) A. rubecula
(dissected from the host, P. ra p a, e three days after oviposition) and b) A. glomeratus (dissected from
P. brassicae five days after oviposition). 45
F igu re 2.5 Newly-moulted second instar larva of A. rubecula
(dissected from the host P . r a p a, e nine days after
oviposition). 46
F igu re 3.1 Relationship between the number of hosts (one day-old
P. brassicae) per patch and the mean number of hosts
parasitised by A. glomeratus (+ 95% C.I.) in Experiments 1
and 2. C u rve draw n by eye. 56
F igu re 3.2 The relationship between the duration of patch visits made
by A. glomeratus and the number of hosts (one day-old
P. brassicae) parasitised during those visits, for patches of five hosts. Data from Experiment 1. Curve drawn by eye. 58 9
Page Figure 3.3 The relationship between the duration of patch visits made
by A. glomeratus and the number of hosts (one day-old
P. brassicae) parasitised during those visits, for patches of
ten hosts. Data from Experiment 1 (open circles) and
Experiment 2 (closed circles). Curve drawn by eye. 59
F igu re 3.4 The relationship between the duration of patch visits made
b y A. glomeratus and the number of hosts (one day-old P. brassicae) parasitised during those visits, for patches of twenty-five hosts. Data from Experiment 1 (open circles) and Experiment 2 (closed circles). Curve drawn by eye. 60
Figu re 3.5 The relationship between the duration of patch visits made
by A. glomeratus and the number of hosts (one day-old
P. brassicae) parasitised during those visits, for patches of
fifty hosts. Data from Experiment 1 (open circles) and
Experiment 2 (closed circles). Curve drawn by eye. 61
Figu re 3.6 The number of hosts (one day-old P. brassicae) p arasitise d by A. glomeratus in Experiments 1 and 2, as a function of
time on patch and host patch size. The curves have been
re-drawn from Figures 3.2 - 3.5. Curve 1: patches of five
hosts. Curve 2: patches of ten hosts. Curve 3: patches of
twenty-five hosts. Curve 4: patches of fifty hosts. 62
Figu re 3.7 Estimated total number of eggs laid by A. glomeratus on
patches of fifty hosts (one day-old P. brassicae), as a
function of time spent on the patch. See text for details of
estimation. Data from Experiment 1 (open circles) and
Experiment 2 (closed circles). 63 10
Page Figure 3.8 The cumulative number of occurrences of oviposition behaviour (closed circles) and probing behaviour (open
circles) made by one female A. glomeratus on a patch of
fifty one day-old P. brassicae, plotted at one minute
intervals. The wasp left the patch after thirty-eight
minutes. 65
F igu re 3.9 The estimated amount of time spent a) handling accepted hosts and b) handling rejected hosts; and the mean amount of time spent c) cleaning and d) searching, in each five
minute period spent by A. glomeratus on patches of fifty
P. brassicae. 71
F igu re 3.10 The estimated amount of time spent a) handling accepted hosts and b) handling rejected hosts; and the mean amount of time spent c) cleaning and d) searching, in each five
minute period spent by A. glomeratus on patches of twenty-
fiv e P. brassicae. 72
Figure 3.11 The estimated amount of time spent a) handling accepted hosts and b) handling rejected hosts; and the mean amount
of time spent c) cleaning and d) searching, in each five
minute period spent by A. glomeratus on patches of ten
P. brassicae. 73
F igu re 3.12 The relationship between the duration of patch visits made
by A. glomeratus on patches of five, ten, twenty-five or
fifty one day-old P. brassicae, and the number of direct
hits (fluid directly smeared onto the wasp) scored by host
larvae. Data from Experiment 2. 74
F igu re 3.13 The relationship between host (one day-old P. brassicae) patch size and the mean duration of patch visits (+ S.E.)
m ade by A. glomeratus in Experiments 1 and 2. Regression
equation: y = 0.503x + 11.9; F^ g2) = 8.5 (p < 0.01); r2 = 14.1. 75 11
Page Figure 3.14 The effect of terminating parasitism, when the attack
rates in Figure 3.6 decrease to a threshold, on the percent
parasitism imposed by A. glomeratus on patches of 5, 10, 25 or 50 one day-old P. brassicae. The results of applying the
following five threshold rates are shown, a) 0.05 hosts
parasitised per minute; b) 0.2hosts/minute; c) 0.5 hosts/minute; d) 1.0 hosts/minute; e) 2.0 hosts/minute.
Percent parasitism was estimated from Figure 3.6. 78
Figure 3.15 The relationship between host (one day-old P. brassicae)
patch size and the mean percent parasitism (+ S.E.)
imposed by A. glomeratus in Experiments 1 and 2. 79
Figure 3.16 Behavioural models of patch departure rules. The wasp has a level of responsiveness which reflects its estimate of the value of staying in a patch. When this level reaches a threshold (T), the wasp departs. The level may change in response to passing time, ovipositions (indicated by arrows) or both.
a) Responsiveness decreases with time, but is incremented by each oviposition. The magnitude of the increment
decreases with time spent on the patch (see Green,
1980, 1984 and Iw asa e t a l, 1981).
b) Responsiveness decreases with time, but is incremented by each oviposition. The magnitude of the increment
increases (up to a maximum) with time between two
successive ovipositions (see Waage, 1979).
c) Fixed GUT rule. Responsiveness decreases with time,
but is returned to its original value by each
oviposition.
d) Fixed number rule. Responsiveness is unaffected by
time, but is decremented a fixed amount by each
oviposition. 12
Page
e) Fixed time rule. Responsiveness is unaffected by
oviposition, but decreases with time.
f) Responsiveness decreases with time, and is also
decremented by each oviposition (see Iwasa e t a l, 1981). 83
F igu re 3.17 The outcome of plotting the time spent by a wasp on a host patch against the number of hosts parasitised by that wasp,
as predicted by the six theoretical patch departure rules
illustrated in Figure 3.16. In (b) and (c), curve (i) represents a high density host patch, and curve (ii) a low
density patch. See Figure 3.16 and text for explanation of
patch departure rules. 86
F igu re 4.1 Functional response of British A. glomeratus parasitising
one day-old P. brassicae larvae. The curve is the fitted random parasite equation, with T = 2 hours; P t = 1; and a’ and Tfo as given in Table 4.1. 94
F igu re 4.2 Functional response of British A. glomeratus parasitising one day-old P. ra p a e larvae. The curve is the fitted
random parasite equation, with T = 2 hours; P t = 1; and a’
and Th as given in Table 4.1. 98
F igu re 4.3 Functional response of British A. rubecula parasitising one
d ay-old P. ra p a e larvae. The curve is the fitted random parasite equation, with T = 2 hours; P t = 1; and a ’ and T^ as given in Table 4.1. 99
F igu re 4.4 Functional response of American A., glomeratus parasitising
one day-old P. ra p a e larvae. The curve is the fitted
random parasite equation, with T = 2 hours; P t = 1; and a’
and Tft as given in Table 4.1. 100 13
Page Figure 4.5 A comparison of the fitted functional response curves in Figure 4.1 - 4.4. Curves are as follows: 1 - British
A. glomeratus on P. brassicae; 2 - B ritish A. glomeratus on
P. ra p a; e 3 - B ritish A. rubecula on P. rapae\ 4 - American
A. glomeratus on P. rapae. 101
F igu re 5.1 a) Schematic representation of a hypothetical relationship between clutch size (c ) and fitness per egg
(f ( c ))• Curve A: weak within-brood density- dependence. Curve B: strong within-brood density-
dependence.
b) Relationship between parental fitness per clutch
(fitness per egg x clutch size; c f ( c ) ) and clutch size (c). The optimum clutch size (C t) is that which
maximises this function. Copt (A) is the optimum
clutch size for curve A (weak density-dependence);
and CQpt (B) the optimum for curve B (strong density
dependence). 107
F igu re 5.2 Relationship between clutch size and the proportion of a brood of British A. glomeratus which survives to adulthood.
a) W ith P. brassicae as host. Regression equation, with arcsine square-root transformation of y axis:
y = -0.0013x + 1.13; r 2 = 1.4; F (132) = 0.46 (p > 0.05).
b) W ith P. ra p a e as host. Regression equation, with arcsine square-root transformation of y axis:
y = -0.024x + 1.60; r2 = 29.5; F ^ 16^ = 6.69 (p < 0.05). 115 14
Page Figure 5.3 Relationship between clutch size and the mean head width of British A. glomeratus adults emerging from each brood.
a) W ith P. brassicae as host. Regression equation:
y = -0.0006x + 0.70; r2 = 8.0; F (1S2) = 2.78 (p > 0.05).
b) W ith P. r a p a e as host. Regression equation: y = -0.0032x + 0.70; r2 = 28.7; F (117) = 6.84 (p < 0.05). 118
F ig u re 5.4 Relationship between the size (head width) of newly-
emerged British A. glomeratus females and their egg complement. Correlation coefficient = 0.92. Regression equation: y = 4337x - 2225. 119
F ig u re 5.5 Relationship between the clutch size laid by American
A. glomeratus in P. ra p a eand the proportion of each brood which survived to adulthood. Regression equation, with
arcsine square-root transformation of y axis: y = -0.016x +
1.59; r 2 = 42.2; F (1 25) = 18.23 (p < 0.001). 121
F ig u re 5.6 Relationship between the clutch size laid by American
A. glomeratus in P. ra p a e and the mean head width of
adults emerging from each brood. Regression equation,
with log-transformed axes: y = -0.144x + 0.0216; r2 = 41.0;
F (i 25) = 17.37 (p < 0.001). 122
F ig u re 5.7 Mean egg load (+ S.E.) of non-ovipositing female
A. glomeratus as avfunction of female age (number of days since adult emergence). Closed circles: British
A. glomeratus from P. brassicae. Open circles: American
A. glomeratus from P. ra p a e. Means are based on dissection of four females from each population at each age. 123 15
Page Figure 5.8 Relationship between clutch size and the egg to adult development time of A. glomeratus, measured at 20 °C and
70% R.H .
a) British A. glomeratus developing in P. brassicae. b) American A. glomeratus developing in P. ra p a e. 126
F igu re 5.9 Clutch size fitness functions for A. glomeratus, showing the relationship between clutch size and an estimate of fitness per egg: juvenile survivorship x mean head width of surviving offspring.
a) British A. glomeratus developing in P. brassicae.
b) American A. glomeratus d eveloping in P. ra p a e. 129
F igu re 5.10 Clutch size fitness functions for A. glomeratus, showing the relationship between clutch size and an estimate of fitness per egg: juvenile survivorship x mean egg load of
surviving offspring.
a) British A. glomeratus d eveloping in P. b ra ssic a e .
b) American A. glomeratus developing in P. ra p a e. 131
Figure 5.11 Relationship between clutch size and a measure of parental fitness per clutch for British A. glomeratus on
P. b ra ssica e (closed circles) and American A. glomeratus on
P. ra p a e(open circles). Fitness per clutch is obtained from
clutch size x fitness per egg, using proportion surviving x
mean egg load as an estimate of fitness per egg. Values of
fitness per clutch are scaled relative to the highest fitness
obtained. Regression equation for British A. glomeratus:
y = 0.0098x + 0.04; r 2 = 69.1. L in e through poin ts fo r
A m e rica n A. glomeratus draw n by eye. 132
F igu re 5.12 Relationship between oviposition time and clutch, size for B ritish A. glomeratus ovipositing in one day-old (first
instar) P. b ra ssica e. Correlation co e fficie n t = 0.35; p > 0.05. 137 16
Page Figure 5.13 Relationship between oviposition time and clutch size for
B ritish A. glomeratus ovipositing in four day-old (second
instar) P. brassicae. C orrelation coefficien t = 0.55; p < 0.05.
Regression equation: y = 1.03x + 13.2. 138
Figure 5.14 Relationship between oviposition time and clutch size for
B ritish A. glomeratus ovipositing in eight day-old (third in s t a r ) P. brassicae. Correlation coefficient = 0.83;
p < 0.001. R egression equation: y = 0.865x + 13.5. 139
Figure 5.15 Relationship between oviposition time and clutch size for
B ritish A. glomeratus ovipositing in one day-old (first in star) P. ra p a e. Correlation coefficien t = 0.75; p < 0.001. 140
Figure 5.16 Mean length of eggs (+ 99% C.I.) laid by British
A. glomeratus in newly-hatched P. brassicae larvae, when dissected from the host 0, 1, 2, 3 or 4 days after oviposition. 143
Figure 5.17 Percentage frequency distribution of the lengths of eggs
laid by British A. glomeratus in newly-hatched P. brassicae larvae, when dissected from the host 0, 1, 2, 3 or 4 days after oviposition. Egg lengths are given in graticule units,
where one unit = 0.022 mm. The value of n is the sample
size. 145
F igu re 5.18 A ppearance o f eggs laid by A. glomeratus in newly-hatched
P. brassicae larvae, when dissected from the host a) _ _
immediately after oviposition; b) one day after oviposition; c) two days after oviposition; d) three days after
oviposition; and e) four days after oviposition. After four
days the fully formed larva is visible in the egg, and
hatching occurs between four and five days after
oviposition. 147 17
Page Figure 5.19 Percentage frequency distribution of the length of eggs laid by British A. glomeratus in two day-old P. brassicae larvae, when dissected from the host one or two days after
oviposition. Egg lengths are given in graticule units,
where one unit = 0.022 mm. The value of n is the sample
size. 150
F igu re 6.1 Correlation between length of adult fore-wing and full- grown larval length in ninety-four species of Lepidoptera
(data collected from the literature). Correlation
co e fficie n t = 0.856; p < 0.001. 162
F igu re 6.2 Frequency distribution of brood sizes in fifty-seven
species of gregarious A p a n teles. Data collected from the literature. 164
F igu re 6.3 Regression of brood size on host size (length of adult fore-
wing) in fifty-two species of gregarious A p a n te le s . Regression equation: ln y = 1.511nx - 1.21; r2 = 31.3; F (150) = 22.8 (p < 0.001). '165
F igu re 6.4 Correlation between host size (length of adult fore-wing) and length of adult wasp in seventy-seven species of
gregarious A p a n te le s. Correlation coefficient = 0.352;
p<0.01. 167
F igu re 6.5 Relationship between a measure of total parasitoid volume per host ((Length of adult wasp)3 x brood size) and host
size (length of adult fore-wing) in forty gregarious and
sixty-four solitary species of A p a n te le s . Regression
equation fo r gre gariou s species: ln y = 1.881nx - 1.64;
r 2 = 45.0; F ^ sgj = 31.2 (p < 0.01). L in e through poin ts fo r solitary species drawn by eye. 168 18
Page Figure 6.6 The number of gregarious (a) and solitary (b) A p a n te le s species parasitising hosts in each of eight different host
size classes (host size = length of adult fore-wing). The
host size distribution of gregarious species is shifted to the
right relative to that of solitary species ( X2^ = 42.4;
p < 0.01). 169
F igu re 7.1 Sex ratio (proportion males) of A. glomeratus broods reared
from field-collected P. brassicae, plotted against brood size
(number of cocoons). All-male broods have been omitted. 188
F ig u re 7.2 Egg complement of non-ovipositing A. rubecula as a
function of female age (number of days since adult emergence). Means (+ S.E.) of four females at each age. 189
F igu re 7.3 Appearance of eggs laid by A. rubecula in first instar
P. ra p a ewhen dissected from the host a) immediately after oviposition; b) one day after oviposition; and c) two days
after oviposition. After two days, the fully-formed larva
is visible in the egg, and hatching occurs between two and three days after oviposition. 197 19
LIST OF TABLES
Page Table 2.1. Head width of instars I-V of P. brassicae and P. ra p ae. Measurements were made using a binocular dissecting
microscope fitted with an eye-piece graticule. 32
T ab le 2.2. Mean length of adult fore-wing (measured from centre of thorax to wing-tip) of laboratory-reared P. brassicae and
P. rapae. 32
T ab le 2.3. Mean adult body length (measured from the head to the tip of the abdomen) of A. glomeratus (reared from P. brassicae) and A. rubecula. Measurements were made using a compound microscope fitted with an eye-piece
graticule. 35
T ab le 2.4. Percentage parasitism imposed by A. glomeratus in various field studies conducted in Europe and India. 36
T ab le 2.5. Percentage parasitism imposed by A. glomeratus in various field studies conducted in Japan, Australia and
the U S A . 37
T ab le 2.6. Percentage parasitism of first instar P. brassicae and P. ra p a eexposed to field parasitism for 3-4 days between
Ju ly and September, 1985, at Silw ood Park. T he num ber
of hosts parasitised is the number collected from which
A. rubecula or A. glomeratus later emerged. The number
exposed is the number parasitised plus the number which
pupated. 37
T ab le 2.7. Clutch size (number of eggs laid per oviposition) of A. glomeratus recorded in the literature. 39
T ab le 2.8. Field brood sizes (number of cocoons per host) of A. glomeratus recorded in the literature. 42 20
Page Table 2.9. Frequency of P . r a p a e larvae containing different combinations of live and dead A. rubecula larvae. Hosts were dissected a few hours after the parasitoid eggs had
hatched. See Chapter Four for experimental details
(E x p e rim e n t 3). 43
Table 2.10. Mean egg-larval, pupal and total development time of A. rubecula in P. ra p a e at 20 °C and 70% R.H. Oviposition
occurred in one day-old hosts which were then reared on
cut Brussels sprouts leaves until parasitoid emergence. 44
T able 2.11. Mean head width of host larvae immediately after parasitoid emergence. Hosts were parasitised as one day- old first instars. * Mean head width of unparasitised
P. ra p a e larvae, of the same age as those parasitised by A. rubecula, at the time of A. rubecula larval emergence. 44
T able 2.12. Mean head width of male and female A. rubecula. T he parasitoids were obtained from P . r a p a e la r v a e , parasitised as one day-old first instars, and then reared
on cut Brussels sprouts leaves in plastic containers at 20 °C. Measurements were made using a compound microscope fitted with an eye-piece graticule. 47
Table 3.1. An estimate of the number of hosts parasitised in each
five minute period spent by A. glomeratus on patches of
50 P. brassicae. The estimate is based on Figure 3.5. 66
Table 3.2. Mean frequency of oviposition behaviour and probing behaviour, and the mean amount of time spent in
oviposition behaviour and probing behaviour, in each
five minute period spent by A. glomeratus on patches of
50 P. brassicae. . 67 21
Page Table 3.3. An estimate of the number of true ovipositions, pseudo- ovipositions, probes and total rejections (probes plus
pseudo-ovipositions) in each five minute period spent by
A. glomeratus on patches o f 50 P. brassicae. 67
T ab le 3.4. An estimate of the number of hosts parasitised in each five minute period spent by A. glomeratus on patches of 25 P. brassicae. The estimate is based on Figure 3.4. 68
T ab le 3.5. Mean frequency of oviposition behaviour and probing behaviour, and the mean amount of time spent in
oviposition behaviour and probing behaviour, in each
five minute period spent by A. glomeratus on patches of
25 P. brassicae. 68
T ab le 3.6. An estimate of the number of true ovipositions, pseudo-
ovipositions, probes and total rejections (probes plus pseudo-ovipositions) in each five minute period spent by
A. glomeratus on patches of 25 P. brassicae. 69
T ab le 3.7. An estimate of the number of hosts parasitised in each five minute period spent by A. glomeratus on patches of
10 P. brassicae. The estimate is based on Figure 3.3. 69
T ab le 3.8. Mean frequency of oviposition behaviour and probing
behaviour, and the mean amount of time spent in oviposition behaviour and probing behaviour, in each
five minute period spent by A. glomeratus on patches of 10 P. brassicae. 70
Table 3.9. An estimate of the number of true ovipositions, pseudo- ovipositions, probes and total rejections (probes plus
pseudo-ovipositions) in each five minute period spent by
A. glomeratus on patches of 10 P. brassicae. 70 22
Page Table 4.1. Estimates (+ S.E.) of a’ and T^ obtained by fitting the random parasite equation to the data in Experiments 1-4,
w ith P t = 1 and T = 2 hours. The asymptote ( = 2/T ^ ) is
given to the nearest integer.
Experiment 1: British A. glomeratus on P. brassicae.
Experiment 2: British A. glomeratus on P. ra p a e.
Experiment 3: British A. rubecula on P. rapae.
Experiment 4: American A. glomeratus on P. ra p a e. 96
T ab le 4.2. Mean (+ S.E.) and range of the number of eggs laid in replicates at the highest host density class (39-56 hosts
per plant) in Experiments 1-4. Experiment 1: British A. glomeratus on P. brassicae.
Experiment 2: British A. glomeratus on P. rapae. Experiment 3: British A. rubecula on P. rapae. Experiment 4: American A. glomeratus on P. ra p a e. 103
T ab le 5.1. Egg-larval, pupal and total development time of British A. glomeratus in P. brassicae an d P. ra p a e at 20 °C and 70% R .H . 113
T ab le 5.2. Mean clutch size laid by British A. glomeratus in singly- presented one day-old host larvae. 116
T ab le 5.3. Mean duration of egg-larval, pupal and total development ------time of British A. glomeratus in P. brassicae, and of
A m e rican A. glomeratus in P. ra p ae . Measured at 20 DC,
70% R .H . 124
Table 5.4. The result of fitting the function f ( c ) = exp (-K c ) to the clutch size fitness functions fjor_American _A._glomeratus
on P. ra p a e in F igu re s 5.5, 5.9b an d 5.10b. T he fu n ctio n s were fitted by non-linear least-squares regression. Linear
A regression gave a worse fit in all cases. The r values
obtained from linear regression were as follows: 47.0
(F igu re 5.5.); 49.2 (F igu re 5.9b); and 40.4 (F igu re 5.10b).
The optimum clutch size (C t) is the inverse of K. 134 23
Page Table 5.5. Mean clutch size laid by British and American A. glomeratus in the functional response experiments
described in Chapter Four, at the highest host density (50
hosts per plant) only. There is a significant effect of
treatment on clutch size (one-way A N O V A : F^2 125j = 18.2;
p < 0.001). 135
T ab le 5.6. Mean brood sizes (number of cocoons) obtained from P. brassicae larvae collected in the field at Silwood Park,
and fro m P. ra p a e collected in the field at Amherst,
Massachusetts, in 1985. 136
T ab le 5.7. Mean clutch size laid by British A. glomeratus in instars
I - I I I o f P. brassicae. 141
T ab le 5.8. Frequency of oviposition and probing behaviour shown
by British A. glomeratus when exposed to alternately
unparasitised and parasitised two day-old P. brassicae larvae. 149
T able 5.9. The number of three day-old P. brassicae la r v a e
containing 1 or 3 day-old A. glomeratus eggs. Eggs were aged by their size and appearance. Host categories are as
follows. U-O. unparasitised host - oviposition behaviour.
U-P. unparasitised host - probing behaviour. P-0,
parasitised host - oviposition behaviour. P-P. parasitised
host - probing behaviour. P = parasitised host - not re-
exposed. 151
Table 5.10. Frequency of ovipositions (i.e. egg-laying) and rejections (i.e. no egg-laying) shown by A. glomeratus to parasitised
and unparasitised two day-old P. brassicae la rv a e .
Rejections are either probes (insertion of ovipositor for 2
seconds or less) or pseudo-ovipositions (insertion of
ovipositor for more than 2 seconds without eggs being
laid). 152 24
Page Table 5.11. Mean clutch sizes (+ S.E.) laid by A. glomeratus in each of five host categories (see Table 5.9. for explanation of
categories). Sample size is in brackets after each mean. 152
T ab le 5.12. Mean duration of oviposition behaviour shown by
A. glomeratus to unparasitised newly-hatched P. brassicae
larvae, and to parasitised and unparasitised two day-old
hosts. 153
T able 5.13. Reported occurrences of observed rejection or lack of rejection of previously-parasitised hosts by A p a n te le s spp. 159
T able 6.1. Host instar in which parasitoid emergence occurs, expressed as the number of instars before the host’s final
instar, in those A p a n teles species for which information
has been collected. Host sizes refer to the length of the adult fore-wing (in mm). 171
T ab le 7.1. Sex ratios (proportion males) of gregarious A p a n te le s species recorded from the literature. 182
T able 7.2. Sex ratios (proportion males) of solitary A p a n te le s species recorded from the literature. 183
T able 7.3. Mean sex ratio (proportion males) of the gregarious
species listed in Table 7.1. and of the solitary species
listed in Table 7.2. A. thompsoni, a thelytokous species, has
been excluded from the calculation of a mean sex ratio
fo r gre gariou s species. 184
Table 7.4. Sex ratios of adults emerging from 81 mixed broods of A. glomeratus obtained from P. brassicae, and from 98
cocoons of A. rubecula obtained from P. ra p a e. H o st larvae were collected from the field at Silwood Park
fro m July to September, 1985. 184 25
Page
T ab le 7.5. The number of larvae emerging from hosts, and the numbers of male and female adults emerging from 51
mixed and 12 all-male broods of A. glomeratus obtained from field-collected P. brassicae. 186
T ab le 7.6 Sex ratio of 59 broods of A. glomeratus in which some larval mortality was detected, and of 22 broods in which
. it was not. 187
T ab le 7.7. Mean lengths of eggs dissected from the ovaries of B ritish A. glomeratus and A. rubecula. 190
T ab le 7.8. Measures of the fecundity of solitary and gregarious A p a n te le s species recorded in the literature. The values
refer to the number of eggs laid per lifetime, apart from those marked * which are based on dissection of adult
females. 191 26
CHAPTER ONE
INTRODUCTION
Natural selection is believed to result in the adaptation of organisms to their environment. The study of adaptation has recently acquired greater precision due to the use of optimisation techniques (Maynard Smith, 1978). These enable testable predictions to be made about which design features or behavioural traits will, within prescribed constraints, maximise the fitness of their owners. Two of the topics in behavioural ecology that have benefitted most from this approach are clutch size and foraging behaviour.
Until recently, most studies concerned with the evolution of clutch size have focussed on birds. Lack (1947) suggested that the optimum clutch size is that which maximises the number of young a pair can rear to maturity. Charnov &
Krebs (1974) modelled the effect of a trade-off between clutch size and parental survival. Numerous experimental studies have also been conducted on avian clutch sizes (see Lessells (1986) for a review). In the last few years, however, theoretical studies of clutch size evolution have been extended to species without parental care. Weis e t a l (1983) constructed a model to interpret the results of their experimental study of clutch size in a cecidomyiid gall midge, and Parker & Courtney (1984) developed general models for insects ovipositing in patches of limited resources. Insect parasitoids, because they lay their eggs in discrete, quantifiable units of resource, are particularly amenable to this approach. Models developed specifically with parasitoids in mind have been published by Charnov &
Skinner (1984, 1985), Iwasa et al (1984), Skinner (1985) and Waage & Godfray
(1985). These models, and the experimental tests of their predictions, are reviewed by Waage (1986) and Godfray (1987a).
Ornithologists have also been responsible for much of the early theoretical and empirical work conducted on optimal foraging behaviour (see reviews in Pyke et a l (1977), Krebs et al (1983) and Krebs & McCleery (1984)). In the simplest models, natural selection is assumed to favour animals which behave in a way that maximises their net rate of energy gain. The problem with adopting this as a theoretical currency to be maximised is that it is difficult to relate to reproductive fitness, or to measure experimentally (Pyke e t a l, 1977). These 27
problems are circumvented in studies of parasitoids foraging for hosts:
selection is then assumed to maximise the rate at which eggs are laid - a
currency directly related to reproductive fitness, and one easily measured in the laboratory. Consequently, parasitoids have become popular subjects for
theoretical and empirical studies of optimal foraging (Cook & Hubbard, 1977,
1980; H u b b a rd & Cook, 1978; C om in s & Hassell, 1979; Waage, 1979; M a rris et a l, 1986).
An important technique for testing the predictions of optimality models is the com parative m ethod (M a y n a rd Sm ith, 1978; C lu tto n -B rock & H a rv e y , 1984).
Parasitoids, because of their high diversity (over 200000 species have been
described in the Hymenoptera alone (Waage & Hassell, 1982)) are a group particularly well suited to the use of inter-specific comparisons. The aim of
this thesis is to use the comparative method to investigate how host biology
affects the adaptation of clutch size and foraging behaviour in the braconid
parasitoid Apanteles glomeratus (L.). To this end, use will be made of comparisons between different populations of A. glomeratus, exploiting
different hosts, and between different species within the genus A p a n te le s.
A p a n te le s Foerster is a very species-rich genus in the subfamily
Microgasterinae. Over 1300 species have been described, and current estimates
place the total world fauna at 5000 - 10000 species (Mason, 1981). Although
Muesebeck (1921) stated that the genus is homogeneous and not subject to
division, most taxonomists now accept Nixon’s (1965) assertion that A p a n te le s
is almost certainly of polyphyletic origin. Nixon (1965) subdivided the genus into forty-four species groups. Mason (1981) went further and reclassified the
North American species into several new, and possibly monophyletic, genera.
This new classification is yet to be universally accepted, or extended to species outside North America, and in this thesis I refer to A p a n te le s in the broad sense of Nixon (1965).
A p a n teles species are parasitoids of larval Lepidoptera. The females of most species insert their eggs into the haemocoel of early instar host caterpillars
(very rarely, in the egg). Some species are gregarious, several eggs being laid at each oviposition; whilst others are solitary, in which case only one egg is laid, and only one larva ever emerges. Larval development is completed within the host (endoparasitism), and mature parasitoid larvae emerge from late instar hosts, which subsequently die. Immediately after emergence, each 28
larva spins a cocoon within which it pupates. Most species arc arrhenotokous haplo-diploids, in which fertilised eggs develop into diploid females, and unfertilised eggs into haploid males; but a few species are thelytokous, in which males are unknown, and diploid females are produced parthenogenetically. Some species are quite polyphagous, and others are monophagous. However, most species probably fall between these two extremes: each being restricted to a small number of closely-related (often congeneric) host species.
Apanteles glomeratus is a gregarious parasitoid of pierid butterflies. It is indigenous to Europe and Asia where its principal host is the large white butterfly, Pieris brassicae (L.). In confined laboratory conditions it will also parasitise larvae of the small white butterfly, Pieris rapae (L.), but in the field,
P. ra p a e is parasitised much less frequently than P. brassicae. P. rapae larvae are more commonly attacked by Apanteles rubecula (Marshall), a monophagous solitary parasitoid, closely related to A. glomeratus (they have been placed in the same species group by Nixon (1974), and in the same genus by Mason
(1981)).
However, when P. ra p a e became an important pest of cabbage crops in the
United States - having been accidentally imported from Europe in about 1860
(Muggeridge, 1943a) - it was A. glomeratus, and not A. rubecula, w hich, in 1883, was introduced from Britain as a potential biological control agent (Swan,
1964). Despite the absence of its usual host, A. glomeratus spread rapidly from its initial release sites in Washington D.C., Iowa and Nebraska (Swan, 1964). It has persisted on P. ra p a eever since, and has three generations a year (R.G. van
Driesche, pers. comm.).
If it. is assumed that natural selection adapts the behaviour and design features of parasitoids to their hosts, European A. glomeratus should be adapted to P. brassicae, and A. rubecula to P. ra p a e. During the three hundred (approximately) generations since introduction, the American population of
A. glomeratus may then have experienced selection for A. rubecula-type traits.
A similar suggestion has been made before: Price (1980) proposed that parasites should have the potential to undergo rapid adaptation to locally- abundant host populations, and Courtney (1986) invoked this argument to suggest that American A. glomeratus may have developed ecological attributes appropriate to P. ra p a e. 29
How might differences in host biology affect the adaptation of foraging
behaviour and clutch size? One important difference between P. brassicae and
P. ra p a e concerns their larval feeding distribution. Early instar P. brassicae
are gregarious, whereas P. ra p a e larvae are solitary. Host distribution may
have an important effect on the foraging behaviour and searching efficiency
of parasitoids. Chapter Three investigates how the behaviour and searching
efficiency of British A. glomeratus is affected by the gregarious nature of its
host. The effect of host species on the searching efficiency of British
A. glomeratus is considered in Chapter Four. The searching efficiency of
B ritish A. glomeratus fo r P. ra p a e is also compared with that of A. rubecula. These are then compared with the searching efficiency of American
A. glomeratus fo r P. ra p a eto see if any predicted changes have occurred in the American population.
Another difference between the two host species is that the larvae of P. ra pae
are smaller. Chapter Five describes the effect of this difference in host
biology on the larval development of British A. glomeratus. It then compares
the predicted and observed effects of host size on the clutch sizes laid by
British and American A. glomeratus. Chapter Five also considers the influence of two other aspects of host quality - instar attacked and whether or not the host has previously been parasitised - on the clutch size laid by British
A. glomeratus in P. brassicae.
Investigation of the relationship between host size and brood size is extended
to a general comparison of A p a n te le s species in Chapter Six. Particular
consideration is given to the effect of host size on the distribution of solitary
and gregarious clutch sizes. The influence of solitary or gregarious
development on two components of a parasitoid’s reproductive strategy - sex
ratio and egg complement - is explored in Chapter Seven.
The final chapter attempts to synthesise these studies, and discusses their
implications for evolutionary theory and biological control.
First, however, it is necessary to describe the biology of the four experimental species in more detail. This is the subject of Chapter Two. 30
CHAPTER TWO
BIOLOGY OF HOSTS AND PARAS1TOIDS; CULTURING METHODS; AND GENERAL EXPERIMENTAL METHODS
2.1. BIOLOGY OF HOSTS AND PARASITOIDS
2.1.1. The hosts
2.1.1.1. Pieris brassicae
The biology of this species has been thoroughly reviewed by Feltwell (1981), and is also covered by Richards (1940) and Courtney (1986).
Eggs are laid in batches of up to 140 (Figure 2.1.) on the underside of leaves of various cruciferous foodplants, especially B ra ssica spp. and T ropaeolu m
(R ich ard s, 1940). T he eggs hatch after six days (at 20 6C), and the firs t larvae to emerge remove the caps from neighbouring eggs, facilitating synchronous h atch in g (G ardiner, 1974).
There are five larval instars, lasting about twenty-five days in total (at 20°C).
The first two instars, each lasting for 3-4 days, feed gregariously in tightly- packed groups. Third, fourth and fifth instars are more loosely aggregated and may disperse if the food plant is defoliated. Table 2.1. shows the head width of each of the five instars. Eighty-six per cent of the total food consumption occurs in the final instar (David & Gardiner, 1962), and the full- grown larva is about 40mm long, and weighs about 460mg (David & Gardiner, 1962). Mature caterpillars disperse in search of pupation sites, which are usually on walls, fence posts or plant stems. The pupal stage lasts about twelve days (at 20 °C).
Wild adults have fore-wing lengths (measured from the centre of the thorax to the wing-tip) of about 34mm (Meyrick, 1927; South 1941), but laboratory- reared specimens tend to be slightly smaller (Table 2.2.). In Britain, there are u su ally two generations a year (R ich ard s, 1940; South, 1941). O ve rw in te rin g occurs as a pupa, and diapause is induced by short day-length during the larval stage (D a v id & G ard in e r, 1962). iue . Feue y srbuton of h sz of g bths laid batches egg f o size the f o n tio u istrib d cy en Frequ 2.1 Figure Number of egg batches 100 - 0 4 20 60- - 0 8 - . - o p - by captive captive by N ( o N < o m o o . asi e a ssic ra b P. 0 1 VO O VO ac size batch g g E n otd usel srus plants. sprouts ls sse ru B potted on 31 VO o ~- r~ - r o — OO < OO o O , 2 n o 121-130 32 Head width (mm) In star P. brassicae P. rapae I 0.44 0.36 II 0.68 0.48 I I I 1.17 0.88 IV 1.54 1.43 V 2.50 2.00 Table 2.1. Head width of instars I-V of P. brassicae and P. ra p a e. Measurements were made using a binocular dissecting microscope fitted with an eye-piece graticule. n Mean fore-wing length (mm) S.E. P. brassicae 35 31.3 0.3 P. ra p a e 37 21.5 0.4 Table 2.2. Mean length of adult fore-wing (measured from centre of thorax to wing-tip) of laboratory-reared P. brassicae and P. ra pae. 2.1.1.2. Pier is rapae The biology of this species is described in detail by Richards (1940) and Courtney (1986). Eggs are laid singly on the underside of leaves of cruciferous plants, especially B ra ssica spp., T ropaeolu m and mustards (S isy m b riu m spp.) (R ich ard s, 1940). They hatch after five days (at 20°C). Larvae are solitary and first instars will eat conspecific eggs (Courtney, 1986). There are five instars, lasting a total of about twenty-three days (at 20 °C). The head width of each instar is shown in Table 2.1. Eighty-five per cent of the total food consumption occurs in the final instar (Parker & Pinnell, 1973). The full-grown larva is about 30mm long, and weighs about 200mg (Smith & Smilowitz, 1976). 33 Pupation occurs mainly on walls, fence posts and plant stems, and lasts ten days (at 20 °C). The fore-wing length of wild adults is about 24mm (Meyrick, 1927; South, 1941), but laboratory-reared specimens are slightly smaller (Table 2.2. ). In Britain, there are normally three generations a year (Richards, 1940). 2.1.2. T he p arasitoids 2.1.2.1. Apanteles glomeratus Recent reviews of the biology of this species are in Feltwell (1981) and Laing & Levin (1982). 2.1.2.1.1, H o st range The recorded host range of A. glomeratus is very large. Thompson (1953), for example, lists fifty-seven host species, but many of these are now known to have been due to m isid e n tificatio n s (R ich ard s, 1940; N ix o n , 1972; Shaw , 1981; L a in g & Le vin , 1982). A. glomeratus is now thought to be restricted to a few species of Pieridae, and to show a high degree of local host-specificity. In Europe, its principal host is Pieris brassicae', secondarily P. ra p a e (Richards, 1940; Blunck, 1957; Radzievskii, 1980) or, in continental Europe only, A poria c ra ta e g i (L.) (the black-veined w hite) (O sipenko, 1978; D a n ile v sk ii, 1965; W ilbert, 1959; R a d z ie v sk ii, 1980). In Japan it parasitises m a in ly Pieris rapae cru civo ra, but is also recorded from Pieris napi (L.) (the green-veined white) (Sato, 1976, 1978; Sato & O hsaki, 1987) and, in H o k k a id o , Aporia crataegi (Sato & Ohsaki, 1987). In North America, Australia and New Zealand, where P. brassicae, P. napi and A. crataegi are absent, it is known almost exclusively from P. ra p a e. 2.1.2.1.2 L ife cycle In the laboratory, A. glomeratus will oviposit in all five instars of P. brassicae and P. ra p a e (M oiseeva, 1960; Johannson, 1951; P arker & P in n ell, 1973; Shapiro, 1976). However, hosts older than first instars can deter parasitoid attack with a defensive head-jerking behaviour (Adler, 1920; Hamilton, 1935), and fourth and fifth instar P. ra p a e encapsulate parasitoid eggs (Parker & Pinnell, 1973). Furthermore, development is not fast enough to allow successful larval emergence if oviposition occurs in fifth instar hosts 34 (Matsuzawa, 1958; Sato, 1980). First instars arc probably attacked most often (A d ler, 1920; K le in , 1932; H am ilton , 1935; R ich ard s, 1940; M u gge rid ge , 1943b). Eggs hatch after 4-5 days (at 20°C). First instar larvae are mandibulate, but the mandibles are small in comparison with those of A. rubecula (F igu re 2.4.). The larvae go through three instars, feeding initially as true parasites on host haemolymph; later on the fat body (Fuhrer & Keja, 1976; Karnavar, 1984; Junnikkala, 1985). Larval emergence occurs after about twenty-four days (at 20°C: see Chapter Five) from full-grown host larvae (Richards, 1940; Parker & Pinnell, 1973; Shapiro, 1976). Upon emerging, the larvae spin individual cocoons within which they pupate. The host is inactive after parasitoid egression, and dies within twenty-four hours. Adult wasps emerge from the cocoons after about eight days and mate on emergence (see Chapter Seven). Sex determination is haplo-diploid: fertilised eggs develop into diploid females; unfertilised eggs into haploid males (arrhenotokous parthenogenesis). Adults are about 2.6mm long (Table 2.3.). Unfed, wasps die within twenty-four hours, but they may live for up to thirty days if fed on honey (David & Gardiner, 1952; Sato, 1975) or sugar (Muggeridge, 1943b; Matsuzawa, 1958). In the field, they probably feed on nectar (M u gge rid ge , 1943b; Shapiro, 1976; Yastrebov, 1979). Females emerge with a complement of 500-800 mature eggs (Sato, 1975; Hubbard, 1977), this number increasing to 1000-2000 after 2-11 days feeding (M oiseeva, 1960; Sato, 1975; Shapiro, 1976; H u b b ard , 1977; K ita n o , 1978). E g g complement is considered in more detail in Chapters Five and Seven. Laing & Levin (1982) state that there are two generations a year in Europe and North America, but there are usually three in southern England (Richards, 1940), Russia (Shapiro, 1976) and Massachusetts (R.G. van Driesche, pers. comm.). Diapause is pre-pupal (Tagawa et a l, 1984), and is induced by short day-length (D a n ile v sk ii, 1965). 35 n Mean body length (mm) S.E. A. glomeratus 167 2.61 0.02 A. rubecula 78 3.07 0.03 Table 2.3. Mean adult body length (measured from the head to the tip of the abdomen) of A. glomeratus (reared from P. brassicae) and A. rubecula. Measurements were made using a compound microscope fitted with an eye-piece graticule. 2.1.2.1.3. Percentage parasitism Table 2.4. shows the percentage parasitism recorded in various field studies conducted in Europe and India, where A. glomeratus is primarily a parasitoid of P. brassicae. Table 2.5. shows the same for studies conducted in Japan, Australia and the United States, where P. ra p a e is the principal host. In Europe, percentage parasitism of P. ra p a e is less than that of P. brassicae (Table 2.4.). However, where P. brassicae is absent, P. ra p a e is much more heavily parasitised (Table 2.5.). Table 2.6. shows the percentage parasitism imposed by A. glomeratus on P. brassicae and P. ra p a eat Silwood Park in 1985. First instar hosts on potted Brussels sprouts plants were placed out in a 40m x 40m plot of Brussels sprouts for 3-4 days, and then returned to the laboratory. This short exposure time permitted parasitism by A p a n te le, s but prevented parasitism by Phryxe vulgaris (Fallen), a solitary tachinid parasitoid which oviposits in fourth and fifth instar hosts. It also reduced the impact of hyperparasitoids: in 1984, 53% of A. glomeratus broods obtained from field-collected fifth instar P. brassicae were hyperparasitised, mainly by Tetrastichus sp. (H ym enoptera: Eu lophidae ). In the laboratory, caterpillars were reared through to pupation or parasitoid emergence. This procedure was carried out throughout July - September, 1985. 36 Locality Host Percentage parasitism Reference USSR P. brassicae 58 - 92 Radzievskii (1980) tt P. rapae 1 - 10 it fl A. crataegi 8 - 22 tl II P. brassicae 80 - 90 Shapiro (1976) tl P. brassicae 80 Yastrebov (1979) tl P. rapae 16 M II P. brassicae 50 Moiseeva (1960) II P. rapae 10 II t! A. crataegi 25 Osipenko (1978) Yugoslavia P. brassicae 1 - 2 Vukasovic (1926) Germany A. crataegi 20 - 30 Wilbert (1959) France P. brassicae 95 Gautier (1919) II P. rapae 2 11 Britain P. brassicae 54 Richards (1940) II P. rapae 3 II II P. brassicae 84 Moss (1933) II P. rapae 18 if II P. brassicae 60 - 80 J. Bradley (pers. comm.) n P. rapae 5 11 India P. brassicae 50 - 67 Singh Rataul (1976) Table 2.4. Percentage parasitism imposed by A. glomeratus in various field studies conducted in Europe and India. 37 Locality Host Percentage parasitism Reference Japan P. rapae crucivora 58 Sato & Ohsaki (1987) ft P. melete 16 ll tl P. napi 100 ll Australia P. rapae 13 - 49 Hassan (1976) tl P. rapae 10 - 70 Hamilton (1979) USA P. rapae 60 Blunck (1957) II P. rapae 20 Pimentel (1961) ll P. rapae 11 Oatman (1966) n P. rapae 12 - 60 Parker et al (1971) It P. rapae 15 - 80 R.G. van Driesche (pers. comm.) it P. rapae 24 Chamberlin & Kok (1986) Table 2.5. Percentage parasitism imposed by A. glomeratus in various field studies conducted in Japan, Australia and the USA. Because of the restricted exposure time, the values in Table 2.6. underestimate the true level of generational parasitism (van Driesche, 1983). However, they do show that the percentage of P. rapae parasitised by A. glomeratus was less than that of P. brassicae. Number of hosts Apanteles sp. Host sp. Exposed Parasitised Percentage parasitism A. glomeratus P. brassicae 328 129 39.3 A. glomeratus P. rapae 289 7 2.4 A. rubecula P. rapae 289 98 33.9 Table 2.6. Percentage parasitism of first instar P. brassicae and P. rapae exposed to field parasitism for 3-4 days between July and September, 1985, at Silwood Park. The number of hosts parasitised is the number collected from which A. rubecula or A. glomeratus later emerged. The number exposed is the number parasitised plus the number which pupated. 38 2.1.2.1.4. Clutch size and brood size Table 2.7. shows values for the clutch size (number of eggs laid per oviposition) of A. glomeralus recorded in the literature. Figure 2.2 shows the frequency distribution of clutch sizes laid by British A. glomeratus in P. brassicae in the foraging experiments described in Chapters Three and Four. Only data from replicates using the highest host density (50 larvae per plant) are shown. In these experiments, one wasp was allowed to forage on a patch of hosts for up to two hours. Since first instar Pieris brassicae feed gregariously, this is a natural situation, but one in which superparasitism cannot be excluded. However, single ovipositions may result in the deposition of 30 eggs or more (Chapter Five), so it is unlikely that clutches smaller than 35 in Figure 2.2. represent superparasitism. Most clutches are in the range 11-25 eggs per host. Field brood sizes (number of larvae emerging per host; usually estimated from the number of parasitoid cocoons per host) recorded in the literature are usually larger than clutch sizes (Table 2.8.), implying a degree of superparasitism in the field. Figure 2.3. shows the frequency distribution of A. glomeratus brood sizes obtained from P. brassicae larvae exposed to field parasitism at Silwood Park in 1985 (see Section 2.1.2.1.3. for details). There is a peak in the distribution from 16-35 cocoons per host. 39 Locality Host Clutch size Reference Britain P. brassicae < 30 Hubbard (1977) If P. brassicae 19 Richards(1940) If P. rapae 16 H Norway P. brassicae 26 - 30 Johannson (1951) Canada P. rapae 15 - 35 Matheson (1907) Taiwan Pieris spp. 10 - 25 Chu (1974) Japan P. rapae crucivora 19 Kusano & Kitano (1974) If P. rapae crucivora 25 Sato (1976) ll P. napi 20 If ll P. rapae crucivora 20 - 35 Ikawa & Okabe (1985) If P. rapae crucivora 55 Sato & Ohsaki' (1987) If P. rapae crucivora 22 Matsuzawa (1958) n P. rapae crucivora 35 Ikawa & Suzuki (1982) If P. rapae crucivora 20 - 40 Ikawa & Okabe (1984) Table 2.7. Clutch size (number of eggs laid per oviposition) of A. glomeratus recorded in the literature. Number of clutches iue22 rqec dsrbto o cuc sizes clutch of distribution Frequency 2.2 Figure P. brassicae P. nme f gs ad e oioiin laid British by oviposition) per laid eggs of (number fifty on potted Brussels sprouts plants. sprouts Brussels potted on fifty larvae presented in patches of patches in presented larvae . glomeratus A. Clutch size Clutch 40 n n day-old one in 46-50 Figure2.3 Number of broods 30-1 20 10 - - vo — —< — i — o » days as first instars. first as days brassicae P. ros nme o ccos e hs) band from obtained host) per cocoons of (number broods Frequency distribution of the size of of size the of distribution Frequency VO < — (N o lra xoe ofed aaiim o 3-4 for parasitism field to exposed larvae oO vo CN NCN (N 41 VO ro m Brood size Brood CO JL, vo vo ■'T o ro N" vo -H . glomeratus A. VO TT o in VO 56-60 42 Locality Host Brood size Reference Britain P. brassicae 31 Hamilton (1935) »« P. brassicae 45 Moss (1933) ft P. rapae 30 it II P. brassicae 27 Shaw & Smith (unpubl) II P. rapae 28 Richards(1940) It P. brassicae 38 It USSR P. brassicae 30 - 40 Shapiro (1976) Algeria P. brassicae 45 Karnavar (1983) II P. brassicae 86 Karnavar (1984) India P. brassicae 29 Singh Rataul (1976) Japan P. rapae crucivora 32 Sato (1979) N. America P. rapae 28 Slansky (1978) it P. rapae 16 - 52 Matheson (1907) Table 2.8. Field brood sizes (number of cocoons per host) of A. glomeratus recorded in the literature. 2.1.2.2. Apanteles rubecula 2.1.2.2.1. Host range A. rubecula is thought to be specific to Pieris rapae (Gautier & Riel, 1921; Richards, 1940; Wilkinson, 1945; Thompson, 1953; Nixon, 1974). 2.1.2.2.2. Life cycle In the laboratory, A. rubecula will oviposit in all five instars of P. rapae (Parker & Pinnell, 1973), but Richards (1940) found that first instars were most commonly parasitised in the field. Eggs hatch after 2-3 days (at 20 °C). If more than one egg is laid in a host, only one of the hatchlings survives. Elimination of competitors appears to involve physical combat. In the functional response experiment described in Chapter Four (Experiment 3), 215 P. rapae larvae, parasitised by A. rubecula, 43 were dissected a few hours after the parasitoid eggs had hatched. Most (183) of the hosts contained only one A. rubecula larva, but 32 caterpillars contained two or three larvae (since each host was only exposed to one wasp, these larvae were sibs). Table 2.9. shows the number of hosts containing live and dead larvae. In only 8 of the 32 hosts containing more than one larva were all the larvae still alive. Of the 28 larvae found dead in these hosts, 10 showed visible signs of attack. Nine of these had been cut in two (through the thoracic segments), and the other, although still intact, had been mutilated. Another host contained one dead larva (which showed no apparent signs of attack) and two live larvae, one of which had embedded its mandibles in the thoracic segments of the other. Number of A. rubecula larvae per host Frequency Alive Dead (Number of hosts) 1 0 175 0 1 8 0 2 2 2 0 7 1 1 18 3 0 1 0 3 0 2 1 ' 2 1 2 2 Table 2.9. Frequency of P. rapae larvae containing different combinations of live and dead A. rubecula larvae. Hosts were dissected a few hours after the parasitoid eggs had hatched. See Chapter Four for experimental details (Experiment 3). Newly-hatched A. rubecula have large heads equipped with prominent mandibles (Figure 2.4.). These mandibles become proportionally much smaller when A. rubecula moults into the second instar, and are no longer clearly visible (Figure 2.5.). There are three instars in all, and larval emergence 44 occurs about twenty days after oviposition (at 20 °C: see Table 2.10.). Emergence is from 4th or 5th instar hosts (Richards, 1940; Wilkinson, 1966; Parker & Pinnell, 1973). n Mean development time (days) S.E. Egg-larval - 64 20.3 0.2 Pupal 64 7.7 0.1 Total 64 28.0 0.2 Table 2.10. Mean egg-larval, pupal and total development time of A. rubecula in P. rapae at 20 °C and 70% R.H. Oviposition occurred in one day-old hosts which were then reared on cut Brussels sprouts leaves until parasitoid emergence. A. rubecula retards host growth: at the time the larva emerges, the host is about half-grown, and has a head width indicative of a fourth instar (Table 2.11.). Unparasitised caterpillars of the same age are fifth instars at this stage (Table 2.11.). In contrast, A. glomeratus does not inhibit host growth, and emerges from full-grown fifth instars (Table 2.11.). Apanteles sp. Host sp. n Mean head width (mm) S.E. A. glomeratus P. brassicae 19 2.52 0.02 A. glomeratus P. rapae 10 2.07 0.02 A. rubecula P. rapae 24 1.43 0.03 - P. rapae 6 2.01* 0.03 Table 2.11. Mean head width of host larvae immediately after parasitoid emergence. Hosts were parasitised as one day-old first instars. * Mean head width of unparasitised P. rapae larvae, of the same age as those parasitised by A. rubecula, at the time of A. rubecula larval emergence. 45 I______I 0.1 mm Figure 2.4 Newly-hatched first instar larvae of a) A. rubecula (dissected from the host, P. rapae, three days after oviposition) and b) A. glomeratus (dissected from P. brassicae five days after oviposition). 46 Anal vesicle 0.5 mm Figure 2.5 Newly-moulted second instar larva of A. rubecula (dissected from the host, P. rapae, nine days after oviposition). 47 Upon emergence, the larva spins a cocoon within which it pupates. Adult eclosion occurs about eight days later (at 20°C: see Table 2.10.). Adults are about 3mm long (Table 2.3.), and females are slightly larger than males (Table 2.12.). Adults live for about thirty days if fed on 10% honey solution (at 20°C). Females have a complement of 50-150 mature eggs (see Chapter Seven). Sex-determination is haplo-diploid: fertilised eggs develop into females; unfertilised eggs into males. In Britain, there are four generations a year (Richards, 1940). Overwintering occurs within the cocoon, and diapause is induced by short day-length (Nealis, 1985). n Mean head width (mm) S.E. Males 37 0.763 0.004 Females 15 0.830 0.004 Table 2.12. Mean head width of male and female A. rubecula. The parasitoids were obtained from P. rapae larvae, parasitised as one day-old first instars, and then reared on cut Brussels sprouts leaves in plastic containers at 20 °C. Measurements were made using a compound microscope fitted with an eye-piece graticule. 2.1.2.2.3. Percentage parasitism The percentage parasitism imposed by A. rubecula on P. rapae is usually reported to be higher than that imposed by A. glomeratus. Richards (1940), for example, found that 25.7% of P. rapae larvae in his study site were parasitised by A. rubecula, but only 3.4% by A. glomeratus. Dempster (1967) recorded 2-20% parasitism of P. rapae by A. rubecula; and, in Canada, Wilkinson (1966) reported up to 50% parasitism. Table 2.6 shows the percentage parasitism imposed by A. rubecula on P. rapae at Silwood Park in 1985. A much higher proportion of P. rapae were parasitised by A. rubecula than by A. glomeratus. 48 2.1.2.2.4. Clutch size and brood size A. rubecula probably deposits one egg at each oviposition, although it is possible that two or more eggs may occasionally be laid. Of 215 parasitised P. rapae larvae dissected in the functional response experiment described in Chapter Four (Experiment 3), 81% contained only one A. rubecula larva. Cases of two or three larvae in one host probably resulted from superparasitism (t.e. repeated ovipositions), rather than from gregarious oviposition. Supernumerary larvae are invariably eliminated. The 98 P. rapae larvae parasitised by A. rubecula in the field during 1985 (Table 2.6.) each produced one parasitoid larva. 2.2. CULTURING METHODS 2.2.1. The hosts Pieris eggs were obtained from a culture at Glasshouse Crops Research Institute, Littlehampton, which originated from stock maintained at Cambridge University (David & Gardiner, 1952). The culturing method used followed that of David & Gardiner (1952). Adults were kept in cages (1.0m x 0.6m x 0.75m for P. brassicae; 0.45m x 0.45m x 0.45m for P. rapae) in a greenhouse at 20-30 °C. The greenhouse was illuminated by natural light, supplemented between October and March with artificial lighting on a 16hr light : 8hr dark cycle. Natural light is necessary for mating and egg-laying (David & Gardiner, 1961a, 1962). The butterflies were fed on 10% sucrose solution soaked onto yellow cotton wool, or onto white cotton wool with a yellow plastic surround: the colour is necessary to stimulate feeding (David & Gardiner, 1961b). Two 6-8 week-old potted Brussels sprouts plants (variety: Bedford Winter Harvest) were placed in each cage as oviposition sites. Plants and food were replaced every two days. Egg-laden plants were transfered to a room at 20 °C and 70% R.H., with a 16hr light : 8hr dark photoperiod. Upon hatching, larvae were placed on potted 49 Brussels sprouts plants in cages (0.70m x 0.50m x 0.45m). Plants were replaced when necessary and, for fifth instar caterpillars, supplemented with mature Brussels sprouts leaves. Pupation occurred on the walls of the cages. Emerging imagos were left to expand their wings, and were then caught and transfered to the adult cages. The temperature and photoperiod conditions prevented pupae from entering diapause. 2.2.2. The parasitoids British A. glomeratus and A. rubecula were obtained from host larvae placed out in a 40m x 40m plot of Brussels sprouts at Silwood Park between June and October, 1984 and 1985. American A. glomeratus were obtained from cocoon clusters collected in the field at Amherst, Massachusetts by R.G. van Driesche in September, 1985. British A. glomeratus was cultured on P. brassicae only; A. rubecula and American A. glomeratus on P. rapae only. For culturing, 3-4 female wasps, at least two days old, were introduced to a large (0.50m x 0.45m x 0.45m) cage under a bank of 125W fluorescent lighting. Three or four 6-8 week-old potted Brussels sprouts plants, upon each of which 50-100 first instar caterpillars had been feeding for about two hours, were then placed in the cage. After 3-4 hours, the wasps were removed and the plants returned to the Pieris culture. A high host:wasp ratio; a large cage; and a relatively short exposure time appear to be necessary to prevent an extremely male-biased sex ratio in the next generation (a problem also noted by Hubbard (1977)). Laboratory sex ratios were nevertheless male-biased. Parasitised caterpillars were reared with unparasitised caterpillars. A. glomeratus larvae emerge from hosts as they seek pupation sites, so cocoon clusters are usually easily located on the walls of the cage. A. rubecula emerges from P. rapae caterpillars on plants, and cocoons are usually found on leaves and stems. After parasitoid emergence, the cocoons were left to harden for about twenty-four hours. They were then removed, and placed in 50mm x 25mm muslin-topped glass tubes. For A. glomeratus, each brood was placed in a separate tube. For A. rubecula, about eight cocoons were placed in each tube. The tubes were transfered to a separate room, also at 20 °C and 70% R.H., and with a 16hr light : 8hr dark light-regime. 50 Upon emergence, adult A. glomeratus were fed with 10% honey solution soaked onto cotton wool. Mating was observed to occur within the first few hours after emergence, and was only rarely seen later than twenty-four hours after emergence. Food was replaced every two days. Emerging A. rubecula were sexed and transfered to new glass tubes, so that each tube contained 2-4 individuals, including at least one female and one male. The tubes were then provisioned with 10% honey solution, which was replaced every two days. As with A. glomeratus, most mating was observed to occur within the first twenty- four hours after emergence. 2.3. GENERAL EXPERIMENTAL METHODS 2.3.1. Experimental conditions All experiments (unless stated otherwise) were conducted in a controlled temperature room at 20 °C, 70% R.H., under a bank of 125W fluorescent lighting. Before being used in an experiment, hosts were placed on Brussels sprouts leaves or plants, of the same variety used for culturing (Bedford Winter Harvest). 2.3.2. Standardisation of wasps Before being used in an experiment, wasps were standardised by the following procedure. Eighteen hours before the experiment, 1-3 day-old females were removed from culture and placed in individual 50mm x 25mm muslin-topped glass tubes. They were presented with 5-10 one day-old host larvae (of the same species to be used in the experiment) on a small piece of host-damaged leaf. Each wasp was confined with hosts for thirty minutes, timed from the first observed oviposition. If no ovipositions occurred within the first thirty minutes, the wasp was discarded. After removal of the hosts, the wasps were left with 10% honey solution soaked onto cotton wool until the start of the experiment. 2.3.3. Host dissection method Most experiments involved dissection of hosts to count parasitised eggs or larvae. To determine clutch size, hosts were dissected forty-eight hours after 51 oviposition to give the eggs time to develop. New-laid eggs are small and transparent, and difficult to see; whereas two day-old eggs are approximately four times larger, and have a grainy appearance (see Chapter Five). The method of dissection was as follows. The caterpillar’s head was removed with a dissecting needle, and the body contents squeezed out into insect ringers solution. The concentration of the ringers solution was first adjusted so that it was isotonic with the egg contents. This was necessary to prevent the eggs from gaining water and bursting, or from losing water and contracting. Careful teasing of the body contents enabled eggs to be counted. Dissections were performed under a binocular dissecting microscope, allowing up to times forty-five magnification. Times thirty was sufficient to count eggs. Lower magnifications were adequate for counting larvae. 52 CHAPTER THREE THE FORAGING BEHAVIOUR OF A. GLOMERATUS FOR P. BRASSICAE 3.1. INTRODUCTION First instar P. brassicae feed gregariously in discrete patches. This chapter considers the consequences of this gregarious habit for the foraging behaviour of A. glomeratus. In particular, how does the wasp’s attack rate (in terms of the number of hosts parasitised per unit time) vary with time on the patch; what factors limit the number of hosts parasitised per patch; and how do female A. glomeratus allocate time to patches of different host density? A number of factors may limit the attack rate of A. glomeratus foraging for P. brassicae. For example, wasps may become egg-depleted during long foraging bouts. Studies of other parasitoids have shown that declining egg loads correlate with a reduction in the searching activity, and hence attack rate, of individual wasps (Collins et al, 1981). Secondly, as patch time progresses, an increasing proportion of encounters will be with previously- parasitised hosts, and although A. glomeratus usually rejects such hosts, it may consume appreciable amounts of time doing so (see Chapter Five). Thirdly, wasps may be forced to devote increasing amounts of patch time to cleaning. First instar P. brassicae respond to the presence of a parasitoid with a concerted defensive behaviour in which the caterpillars flick up their heads and exude from their mouths a sticky green fluid. If the wasp is sufficiently close, fluid is smeared over its body and it has to spend time cleaning itself. The cumulative effect of host defensive behaviour during a patch visit may be to increase the amount of time the wasps spend cleaning. Description of host-finding behaviour The first stage of host-location by A. glomeratus involves detection of the hosts’ food plant (Sato, 1979; Kitano, 1978). Olfactory cues appear to be important: Hubbard (1972) showed that female wasps responded to odors from chopped cabbage in an olfactometer, and Sato (1979), whilst finding that females responded to filter paper soaked in leaf extract, obtained no reaction to leaves wrapped in cellophane. 53 After location of the host plant, the wasp flies around it, a few centimetres from the leaves, with a characteristic bobbing search flight. After landing on a leaf the wasp responds to odors from leaf damage (Hubbard, 1972) and, at least in the case of P. rapae, to oral secretions, frass and silk from the host (Matsuzawa, 1958; Sato, 1979). The searching female walks slowly over the plant with out-stretched antennae tapping at the leaf surface. When host cues are encountered, walk speed drops, the antennae are curled and the abdomen is arched so that the ovipositor probes at the substrate. When a host larva is found, the wasp raises its wings vertically and thrusts its ovipositor into the side of the caterpillar. After a pause of up to two seconds, the wasp usually pulls away from the host, with the ovipositor still inserted, so that the abdomen is straightened. This position is normally held for 5-20 seconds. The wings are then flattened, the ovipositor withdrawn, and the wasp walks away. On other occasions, the ovipositor, after the initial 1-2 second pause following insertion, is rapidly withdrawn and the wasp leaves the host. This behaviour represents host-rejection, usually because the host has previously been parasitised (see Chapter Five). From here on I refer to the first type of behaviour as "oviposition behaviour"; and to the second as "probing behaviour". 3.2. MATERIALS AND METHODS Host patches were created by placing newly-hatched P. brassicae larvae on one leaf of a 6-8 week-old potted Brussels sprouts plant. The plants used were 20-30cm tall with 7-9 leaves per plant, and a total leaf area (determined by a video leaf area recorder) of 140-240cm A. The plants were placed in individual 40cm x 22cm cylindrical Watkins & Doncaster cages, with clear plastic sides and muslin-covered tops. They were then left for eighteen hours at 20 °C before being used in an experiment. During this time the caterpillars arrange themselves in a tight patch and create feeding damage, frass and silk. Experiment 1 Each replicate consisted of five plants, each with a different number of host larvae. The host patch sizes used were 1, 5, 10, 25 and 50 caterpillars per plant. One standardised female wasp was introduced to each cage and left until it found, and then left, the host patch. Wasps which failed to show searching behaviour during the first thirty minutes were removed. Searching 54 behaviour was defined as either on-plant searching with tapping antennae, or bobbing search flight (see Section 3.1.)* Wasps that did not search usually remained motionless on the top, or occasionally the side, of the cage. Observations of each cage were made at one minute intervals, and the position of each wasp recorded. A wasp was deemed to have found a patch if it displayed intensive searching behaviour, with curled antennae and arched abdomen (see Section 3.1.), in response to host cues. It was defined as having left the patch when it flew off the leaf and did not return within sixty seconds. The number of hosts parasitised during a patch visit, and the number of eggs laid per host, were determined by dissection of the hosts in isotonic insect ringers solution forty-eight hours after the experiment. For each replicate the following information was obtained. 1. The time spent by the wasp on the host patch 2. The number of hosts parasitised 3. The number of eggs laid per host The experiment was replicated six times at each density. Experiment 2 One potted Brussels sprouts plant, containing either 10, 25 or 50 host larvae, was used for each replicate. A Link Electronics video camera - with a 17.5-105mm focal length Fujinon TV zoom lens, fitted with a x4 close-up lens - was focussed on the host patch (through the cage wall) at a distance of approximately 30cm. This required all the hosts to be on the upper surface of the leaf, so only such patches were used. The camera was connected to a National time-lapse video tape recorder, visual display monitor and electronic timer. With the lens at maximum focal length, the image of the host patch completely filled the display monitor. The electronic timer displayed on the screen a digital clock, recording time down to hundredths of a second. 55 One standardised female wasp was introduced to the cage, and its on-patch foraging behaviour recorded on video tape. The wasp was removed when it left the patch. From play-back of the tapes the following information was recorded. 1. • The time spent by the wasp on the host patch. 2. The time spent handling hosts. 3. The amount of handling time composed of oviposition behaviour and probing behaviour. 4. The time spent cleaning or resting. 5. The number of defensive head-jerking movements made by the host larvae. Times were read directly from the display monitor. The number of hosts parasitised, and the number of eggs laid per host, were determined by dissection of the hosts in isotonic insect ringers solution forty-eight hours after the experiment. The experiment was replicated eight times at each of the three host densities. 3.3. RESULTS 3.3.1. The effect of host patch size on the number of hosts parasitised The mean number of hosts parasitised within each of the three highest host densities (10, 25 and 50 hosts per plant) did not differ significantly between Experiments 1 and 2 (see Appendix I). The results from the two experiments have therefore been pooled. The mean number of hosts parasitised in the two experiments is shown in Figure 3.1. as a function of host patch size. Within each host density (except that of one caterpillar per plant) an average of between ten and thirty per cent of caterpillars died during the forty-eight hours between experiment and dissection (there was no apparent density- dependence in this mortality - see Appendix I). The data in Figure 3.1. have been corrected for this mortality by calculating the expected number parasitised, if exactly 5, 10, 25 or 50 hosts had survived, from the proportion of dissected hosts that were parasitised (see Appendix I). This assumes that iue31 eainhp ewe te ubr f ot (n day-old (one hosts of number the between Relationship 3.1Figure Number of hosts parasitised . brassicae P. and 2. Curve drawn by eye. by drawn Curve 2. and aaiie by parasitised pr ac ad h ma nme o hosts of number mean the and patch per ) . glomeratus A. 56 + 5 CI) n xeiet 1 Experiments in C.I.) 95% (+ 57 there was no differential mortality of parasitised hosts. Replicates with host densities of one caterpillar per plant were repeated if the host died before dissection. Figure 3.1. shows that as patch size increases there is a decrease in the proportion of hosts parasitised by a wasp during a single patch visit. 3.3.2. The effect of time on patch on the number of hosts parasitised Figures 3.2.-3.5. show, as a function of time spent on the patch, the number of hosts parasitised by each of the wasps used in Experiments 1 and 2, on patches of 5, 10, 25 and 50 hosts respectively. As in Section 3.3.1. the results have been corrected for mortality between experiment and dissection. Figures 3.2.-3.5. show that the attack rate (in terms of the number of hosts parasitised per unit time) decreases with time on the patch. For comparison, the curves from Figures 3.2.-3.5. are shown together in Figure 3.6. Initial attack rates are higher the larger the host patch size. On patches of 5, 10 and 25 hosts, 100% parasitism is attained with long residence times, but on patches of 50 hosts the attack rate levels off well below 100% parasitism. 3.3.3. Possible factors affecting the attack rate of A. glomeratus on P. brassicae 3.3.3.1. Egg depletion Two to four day-old female A. glomeratus have a complement of 1000-1400 mature eggs (see Chapter Five). Figure 3.7. shows, as a function of time on patch, the total number of eggs laid by each wasp used in Experiments 1 and 2 on patches of 50 hosts. To allow for host mortality before dissection, the number of eggs laid has been estimated from the mean number of eggs per dissected, parasitised host; multiplied by the expected number of parasitised hosts in Figure 3.5. The longest patch times resulted in the oviposition of about 700 eggs. Wasps were therefore not limited by absolute egg numbers, although they may not be able to mobilise their entire egg complement within a short space of time. iue32 h eainhpbten h drto o pth iis made visits patch of duration the between relationship The 3.2 Figure Number of hosts parasitised . brassicae P. five hosts. Data from Experiment 1. Curve drawn by eye. by drawn Curve 1. Experiment from Data hosts. five by . glomeratus A. prstsd uig hs vst, o pths of patches for visits, those during parasitised ) n te ubr f ot (n day-old (one hosts of number the and 58 Number of hosts parasitised Figure 3.3 The relationship between the duration of patch visits made visits patch of duration the between relationship The 3.3 Figure e hss Dt fo Eprmn 1 oe crls and circles) (open 1 Experiment from eye. by drawn Curve Data circles). (closed 2 Experiment hosts. ten . brassicae) P. by . glomeratus A. aaiie drn toe iis fr ace of patches for visits, those during parasitised Time on patch (minutes) patch on Time n te ubr f ot (n day-old (one hosts of number the and 59 iue34 h eainhp ewe h drto o pth iis made visits patch of duration the between relationship The 3.4 Figure Number of hosts parasitised . brassicae) P. and Experiment 2 (closed circles). Curve drawn by eye. by drawn Curve circles). (closed 2 Experiment and by wnyfv hss Dt fo Eprmn 1 oe circles) (open 1 Experiment from Data hosts. twenty-five . glomeratus A. aaiie drn toe iis fr ace of patches for visits, those during parasitised n te ubr f ot (n day-old (one hosts of number the and 60 iue35 h eainhpbten h drto o pth iis made visits patch of duration the between relationship The 3.5 Figure Number of hosts parasitised . brassicae) P. it hss Dt fo Eprmn 1 oe crls and circles) (open 1 Experiment eye. by from drawn Curve Data circles). (closed 2Experiment hosts. fifty by . glomeratus A. aaiie drn toe iis fr ace of patches for visits, those during parasitised n te ubr f ot (n day-old (one hosts of number the and 61 Figure 3.6 The number of hosts (one day-old day-old (one hosts of number The 3.6Figure Number of hosts parasitised five hosts. Curve 4: patches of fifty hosts. fifty of 4:patches Curve hosts. five uv 2 pths f e hss Cre : ace o twenty- of patches 3: Curve hosts. hosts. ten of five patches of 1: 2:patches CurveCurve 3.5.- 3.2 Figures from drawn time on patch and host patch size. The curves have been re been have curves The size. patch host and patch on time by A . glomeratus n xeiet 1 n 2 a a ucin of function a as 2, and 1 Experiments in 62 . brassicae) P. parasitised iue37 siae ttl ubr f gs ad by laid eggs of number total Estimated Figure3.7 Estimated number of eggs laid ucin f ieset ntepth Setx frdtis of details for text See patch. the on spent time of function Experiment 2 (closed circles). (closed 2 Experiment ace o ffy ot (n day-old (one hosts fifty of patches siain Dt fo Eprmn 1 oe crls and circles) (open 1 Experiment from Data estimation. 63 . brassicae P. . glomeratus A. , s a as ), on 64 3.3.3.2. Time spent rejecting previously-parasitised hosts and cleaning From the data collected in Experiment 2 the total amount of time spent handling hosts (measured from the moment of initial contact by the wasp’s antennae, until departure from the host) during oviposition behaviour and probing behaviour was found for each wasp. The total amount of time spent cleaning (including resting) and searching was also found. Figure 3.8. is a typical plot obtained from analysis of the video tapes. It shows the cumulative number of separate instances of oviposition behaviour and probing behaviour, plotted at one minute intervals, made by a wasp on a patch of 50 hosts. In total, there were sixty-six separate occurrences of oviposition behaviour, although only twenty-six hosts were actually parasitised. Superparasitism is not a likely explanation for this excess of oviposition behaviour because the mean clutch size laid in this experiment was 19.2 + 1.0 (S.E.) eggs (range: 9 - 26): well within the range of clutch sizes obtained from single ovipositions (see Chapter Five). Similar findings were obtained from the other wasps used, and these results support the conclusions of the host- discrimination experiment described in Chapter Five: many instances of oviposition behaviour do not result in egg-laying. The host discrimination experiment showed that there is no significant difference between the mean duration of ovipositor insertion associated with real ovipositions (i.e. those in which egg-laying occurs) and with "pseudo-ovipositions" (see Chapter Five). It is therefore impossible to distinguish between these two types of oviposition behaviour by observation. However, the average number and duration of true ovipositions and rejections in each five minute period can be estimated by the following method. For patches of 50 hosts, the number parasitised in each five minute period can be estimated from Figure 3.5.: Table 3.1. shows the results. This method assumes that the curve in Figure 3.5. is a true indication of how an average wasp’s attack rate changes over time. The mean number of times oviposition behaviour and probing behaviour occurred in each five minute period has been calculated from the video tape analysis. Table 3.2. shows the results for patches of 50 hosts. The mean amount of time spent handling hosts during oviposition behaviour and probing behaviour, in each five minute period, is also shown in Table 3.2. An estimate of the number of host rejections - that is, pseudo-ovipositions plus probes (probing behaviour does not entail egg-laying: iue38 h cmltv nme o ocrecs f oviposition of occurrences of number cumulative The Figure3.8 Number of completed ovipositions/probes 20 40- 70-i 60- - o oo • intervals. The wasp left the patch after thirty-eight minutes. thirty-eight after patch the left wasp The intervals. fifty one day-old day-old one fifty circles) made by one female female one by made circles) eaiu (lsd ice) n poig eaiu (open behaviour probing and circles) (closed behaviour o o o oo • • • o ooo ooo 10 • • • Time on patch (minutes) patch on Time . brassicae P. 65 Ooo o 20 . glomeratus A. oo i • • • potd t n minute one at plotted , o o o oo n pth of patch a on o o oo oo 30 I • • o ooo o o I “ 40 66 see Chapter Five) - in each five minute period can then be obtained from the difference between the mean number of times oviposition behaviour occurred (Table 3.2.) and the estimated number of hosts actually parasitised (Table 3.1.); plus the number of probes (Table 3.2.). The results of these calculations are shown in Table 3.3. The numbers of true ovipositions and rejections can then be converted to times using the mean handling times associated with oviposition behaviour and probing behaviour calculated from Table 3.2. The results are shown in Figure 3.9. The same procedure has been carried out for patches of 25 and 10 hosts. Tables 3.4., 3.5. and 3.6. are equivalent to Tables 3.1., 3.2. and 3.3 respectively, but for patches of 25 hosts. Similarly, Tables 3.7., 3.8. and 3.9 are equivalent to Tables 3.1., 3.2. and 3.3., but for patches of 10 hosts. Figures 3.10. and 3.11. show the estimated total amount of time spent handling accepted and rejected hosts by an average wasp during each five minute period of patch time, on patches of 25 and 10 hosts respectively. Figures 3.9., 3.10. and 3.11. also show the mean amount of time spent cleaning and searching in each five minute period. Minutes on patch Estimated number of hosts parasitised per 5 minutes 1 - 5 9.8 6 - 10 6.2 11-15 4.0 16 - 20 3.1 21 - 25 2.4 26 - 30 1.8 31 - 35 1.4 36 - 40 1.2 Table 3.1. An estimate of the number of hosts parasitised in each five minute period spent by A. glomeratus on patches of 50 P. brassicae. The estimate is based on Figure 3.5. 67 Oviposition behaviour Probing behaviour Minutes Number Mean Mean Mean Mean on patch of wasps number time (s) number time (s) 1-5 8 10.5 135 2.3 8 6 - 10 7 7.1 113 2.3 12 11-15 6 4.8 101 1.2 5 16 - 20 6 6.0 82 5.2 21 21 - 25 6 3.2 110 2.3 12 26 - 30 4 4.5' 79 2.5 11 31 - 35 3 3.7 92 3.3 24 36 - 40 2 3.0 38 4.0 19 Table 3.2. Mean frequency of oviposition behaviour and probing behaviour, and the mean amount of time spent in oviposition behaviour and probing behaviour, in each five minute period spent by A. glomeratus on patches of 50 P. brassicae. Minutes on True ovipositions Pseudo-ovipositions Probes Total patch rejections 1 - 5 9.8 0.7 2.3 3.0 6 - 10 6.2 0.9 2.3 3.2 11 - 15 4.0 0.8 1.2 2.0 16 - 20 3.1 2.9 5.2 8.1 21 - 25 2.4 0.8 2.3 3.1 26 - 30 1.8 2.7 2.5 5.2 31 - 35 1.4 2.3 3.3 5.6 36 - 40 1.2 1.8 4.0 5.8 Table 3.3. An estimate of the number of true ovipositions, pseudo- ovipositions, probes and total rejections (probes plus pseudo- ovipositions) in each five minute period spent by A. glomeratus on patches of 50 P. brassicae. 68 Minutes on patch Estimated number of hosts parasitised per 5 minutes 1 5 10.0 6 10 4.2 1115 2.7 16 20 2.0 21 25 1.5 26 30 1.2 Table 3.4. A n estimate of the number of hosts parasitised in each five minute period spent by A. glomeratus on patches of 25 P. brassicae. The estimate is based on Figure 3.4. Oviposition behaviour Probing behaviour Minutes Number Mean Mean Mean Mean on patch of wasps number time (s) number time (s) 1 - 5 6 13.6 166 2.4 7 6 - 10 6 6.4 112 4.6 18 1 1 - 1 5 6 5.8 107 6.2 24 16 - 20 3 7.0 79 5.0 32 21 - 25 3 2.5 90 3.5 33 26 - 30 3 3.5 37 4.0 14 Table 3.5. M e a n frequency of oviposition behaviour and probing behaviour, and the mean amount of time spent in oviposition behaviour and probing behaviour, in each five minute period spent by A. glomeratus on patches of 25 P. brassicae. 69 Minutes on True ovipositions Pseudo-ovipositions Probes Total patch rejections 1 - 5 10.0 3.6 2.4 6.0 6 - 10 4.2 2.2 4.6 6.8 11 - 15 2.7 3.1 6.2 9.3 16-20 2.0 5.0 5.0 10.0 21 - 25 1.5 1.0 3.5 4.5 26 - 30 1.2 2.3 4.0 6.3 Table 3.6. A n estimate of the n u m b e r of true ovipositions, pseudo- ovipositions, probes and total rejections (probes plus pseudo- ovipositions) in each five minute period spent by A. glomeratus on patches of 25 P. brassicae. Minutes on patch Estimated number of hosts parasitised per 5 minutes 1 - 5 5.6 6 - 10 1.8 11 - 15 1.1 16 - 20 0.7 21 - 25 0.4 26 - 30 0.2 31 - 35 0.2 36 - 40 0.0 Table 3.7. A n estimate of the number of hosts parasitised in each five minute period spent by A. glomeratus on patches of 10 P. brassicae. The estimate is based on Figure 3.3. 70 Oviposition behaviour Probing behaviour Minutes Number Mean Mean Mean Mean on patch of wasps number time (s) number time (s) 1 - 5 6 6.8 84 2.7 13 6 - 10 4 3.5 38 5.8 17 11 - 15 4 1.0 42 1.0 7 16 - 20 3 3.0 95 5.0 16 21 - 25 2 2.5 79 0.5 5 26 - 30 2 1.5 49 0.0 0 31 - 35 2* 0.0 0 0.0 0 36 - 40 2 0.0 0 0.0 0 Table 3.8. Mean frequency of oviposition behaviour and probing behaviour, and the mean amount of time spent in oviposition behaviour and probing behaviour, in each five minute period spent by A. g l o m e r a t u s on patches of 10 P. brassicae. Minutes on True ovipositions Pseudo-ovipositions Probes Total patch rejections 1 - 5 5.6 • 1.2 2.7 3.9 6 - 10 1.8 1.7 5.8 7.5 1 1 - 1 5 1.1 0.0 1.0 1.0 16 - 20 0.7 2.3 5.0 7.3 21 - 25 0.4 2.1 0.5 2.6 26 - 30 0.2 1.3 0.0 1.3 31 - 35 0.2 0.0 0.0 0.0 36 - 40 0.0 0.0 0.0 0.0 Table 3.9. A n estimate of the number of true ovipositions, pseudo- ovipositions, probes and total rejections (probes plus pseudo- ovipositions) in each five minute period spent by A. glomeratus on patches of 10 P. brassicae. Time (s) engaged in each activity iue39 h siae muto tm set ) adig accepted spent a) handling time of estimated amount The Figure 3.9 hostsb) rejected handling and amount hosts; the mean and P. brassicae. P. f ie pn c cenn ad ) erhn, n ah five each searching, d) in spent c) timeof and cleaning minute period spent by by spentperiod minute 71 Minutes on patchMinutes . s u t a r e m o l g A. n ace o fifty of patches on Figure Time (s) engaged in each activity 3.10 h etmtd mut f ie spent accepteda) time handling of estimated amount The five hostsb) rejected handling and amount hosts; the mean and minute periodminute spent by of time spent c) cleaning and d) searching, in each five searching, d) in each c) cleaning spent and time of P. brassicae. P. Minutes Minutes on patch 72 . s u t a r e m o l g A. on patches on twenty- of Figure Time (s) engaged in each activity 3.11 h etmtd mut f ie spentaccepted a) time handling of estimated amount The P. brassicae. P. f ie pn c cenn ad )sacig i ec five d) eachsearching, in spent c) and cleaning time of hostsb) rejected and handling amount hosts; the mean and minute period spent by by spent period minute VO o — UO < — • — 73 Minutes on patch Minutes VO NrN CN o . s u t a r e m o l g A. o) vn JL, n ace o ten of patches on tN VO ro o ro m O VO ■VT rO i Figure Number of direct hits 3.12 100- 20 40- 60- 80- 0— - The relationship The theof duration patch visits between made 0* Data from Experiment 2. Experiment from Data (fluid directly onto smeared the scoredwasp) by host larvae. n day-old one by by 6 0 0 .glomeratus u t a r e m o l g A. ♦ • o o “r- 20 0 . e a c i s s a r b P. on patches of five, ten, twenty-five or fifty Time on patch (minutes)Time 40 r~ 74 ad h nme o direct hits of number the and , 60 ^ 8*0 10 0 hosts 5 hosts { o 25 o hosts • 50 • hosts 100 120 I Figure Duration of patch visits (minutes) 3.13 h rltosi bten ot oe day-old relationship (one host between The equation: y = ac sz n te en uain fpth visits patch of duration (+ the size mean patch and made by by made . s u t a r e m o l g A. .0x+1.; 52j14.1. 0.01); 8.5 r2 < (p = = 11.9; + 0.503x i xeiet 1 n . Regression 2. 1 and inExperiments 75 . e a c i s s a r b P. S.E.) ) 76 Figures 3.9., 3.10. an d 3.11. show that as patch time progresses, less time is spent handling new hosts and more time is spent cleaning. Cleaning may be a response to host-defence: the fluid exuded by caterpillars affects the legs and wings of the wasp, impairing its mobility. Figure 3.12. shows the relationship between patch time and the number of direct hits scored by caterpillars (i.e. the number of times fluid was observed to be directly smeared onto the wasp). The number of direct hits increases in proportion to patch time. There is no detectable difference in the rate at which direct hits are scored at different patch sizes. The cumulative effect of host defensive behaviour may be responsible for the extra time spent cleaning towards the end of a patch visit. Figures 3.9.-3.11. also reveal a slight tendency for the proportion of time spent rejecting hosts to increase with time on the patch. 3.3.4. The effect of host patch size on the duration of patch visits Figure 3.13. shows the relationship between host patch size and the mean duration of patch visits made by wasps in Experiments 1 and 2. Linear regression gives a significant positive slope, showing that wasps tend to stay longer on larger patches. However, the trend is only slight and there is considerable variation in patch time at all host densities, with a correspondingly low r2 value. 3.4. DISCUSSION Optimally foraging parasitoids should allocate more time to more profitable patches (Charnov, 1976; Cook & Hubbard, 1977; Comins & Hassell, 1979). To a foraging female A. glomeratus , large patches of P. brassicae offer a higher reward rate than small ones (Figure 3.6.), so to maximise its oviposition rate a wasp should stay longer on these larger patches. The allocation of more time to large patches by individual wasps is an example of parasitoid aggregation (Hassell, 1982a). Lessells (1985) has argued that if wasps remain on host patches until their attack rates decline to a fixed threshold (an assumption of the models of Charnov (1976), Cook & Hubbard (1977) and Comins & Hassell (1979)), the resultant aggregative behaviour will generate spatially density- dependent parasitism (provided egg or time constraints do not apply). 77 Such wasp behaviour would not produce density-dependent parasitism in this study, however. Figure 3.14. shows the effect on percent parasitism of terminating parasitism when the attack rates in Figure 3.6. decline to an arbitrarily-imposed threshold, which is fixed across all host densities. The results of applying five different threshold rates are shown. In this experiment, parasitism on patches of 50 hosts levelled off well below 100%, so to generate density-dependent parasitism, wasps would have had to have spent much longer on these patches than on the smaller ones, and to have exploited them to a much lower reward rate. In fact, wasps showed only a slight tendency to stay longer on larger patches (Figure 3.13.), and parasitism, as a result, was inversely density-dependent (Figure 3.15.). Morrison et al (1980) and Morrison & Strong (1981) found inversely density- dependent parasitism in their studies of eulophid and trichogrammatid egg parasitoids. They attributed this in part to re-encounters with previously- parasitised hosts reducing attack rates on large patches, causing wasps to emigrate before a high percentage of hosts had been parasitised. Re- encounters with previously-parasitised hosts may also explain part of the decline in attack rates shown in Figure 3.6., but a more important reason seems to be the time spent cleaning, possibly as a result of host defence (Figures 3.9.- 3.12.) (something that is clearly not a problem for an egg parasitoid). Constraints imposed by egg-mobilisation rate could also reduce the attack rate of A. glomeratus , and may explain w h y parasitism on patches of 50 hosts levelled off below 100%. Time rendered unavailable for foraging by temporary egg-depletion could be devoted to cleaning. When should the parasitoid leave the patch? It is clear from Figure 3.6. that remaining on a patch becomes less profitable as time progresses. Charnov (1976) considered the problem of when a forager should leave a patch to search elsewhere. In his model, the optimal departure time depends on the travel time to the next patch and on the reward rate expected there. Other studies have concentrated on the proximate rules foragers might use to decide when to leave a patch. The three types of departure rule most frequently discussed are as follows. Figure Percent parasitism 3.14 5 0 5 50 25 10 5 ] The effect of The terminating parasitism, the n attack rates whe or day-old50 one percent the on parasitism threshold, by imposed a to 3.6 decrease Figure in olwn fv trsod ae ae hw. a) threshold five rates shown. following are aaiie pr iue b 02 ot/iue c 0.5 c) hosts/minute. 2.0 e) hosts/minute; 1.0 hosts/minute; 0.2 d) b) hosts/minute; minute; per parasitised Percent parasitism estimatedwas Figure from . brassicae. P. .glomeratus u t a r e m o l g A. Host patch size 78 The results The of theapplying on patcheson of 5, 10, 25 3.6. 0.05 hosts Figure Percent parasitism 3.15 h rltosi bten ot oe day-old relationship (one host between The by by patchsize percent theparasitism and mean (+ .glomeratus u t a r e m o l g A. in 1 2.Experiments and Host patch size 79 S.E.) . brassicae) P. imposed 80 1. Stay until a fixed number of prey (or hosts) have been eaten (or parasitised). This is the "hunting by expectation" rule proposed by Gibb (1962) to explain the foraging behaviour of tits (Paridae). 2. Stay for a fixed time. This is Kreb’s (1973) "hunting by time expectation" hypothesis. 3. Stay until the time since the last capture exceeds a certain threshold value (the giving-up time, or GUT). Krebs et al (1974) suggested that black-capped chickadees (Parus atricappilus) employed a G U T which was fixed within a habitat but which varied between habitats, depending on their quality. Hassell & M a y (1974) and Murdoch & Oaten (1975) modelled an optimally-foraging predator which used a fixed GU T . Oaten (1977) pointed out that random variation in prey capture rates can lead to an inter-capture time exceeding the G U T early in a foraging bout, causing premature departure. His paper precipitated theoretical studies of giving-up rules which incorporate information on patch quality acquired during foraging. Iwasa et al (1981), for example, modelled a forager which estimates the number of prey left in a patch from its average rate of prey capture up to that instant. After each prey capture the estimate decreases until another prey item is caught. The estimate is then incremented by a certain amount. The magnitude of this increment decreases with time on the patch. Eventually the estimate declines to a threshold value which prompts the forager to leave the patch. Waage (1979) constructed a similar behavioural model to interpret the foraging behaviour of the ichneumonid parasitoid Nemeritis canescens (Grav.). In this model, the foraging wasp has a level of responsiveness (analogous to the estimate of remaining prey in the model of Iwasa et al (1981)) which decreases after an oviposition. When the next oviposition occurs, the level of responsiveness is incremented by an amount dependent on the time between the ovipositions. The longer this time, the larger the increment, up to a maximum. When the level of responsiveness declines to a threshold, the wasp departs. 81 Green (1980, 1984) developed theoretical giving-up rules similar to that of Iwasa et al (1981). By considering specific examples he was able to find the optimum departure rule for each case. These rules have the following general form: "leave the patch if after t minutes searching only K prey have been found" (Green, 1980). Green (1984) gave these rules a behavioural interpretation similar to that of Waage (1979). For one example he proposed a behavioural model in which the forager’s level of responsiveness (s e n s u Waage (1979)) decreases with time until a prey item is captured. It is then incremented by a fixed amount. The forager leaves the patch when the level of responsiveness declines to a threshold. Other patch-leaving rules can be generated by the same type of behavioural mechanism. A rule using a fixed G U T would be achieved if each prey capture returned the forager’s level of responsiveness to a fixed value (McNamara, 1982). A fixed number rule would be produced if the response level was unaffected by time, but was decremented a fixed amount by each prey capture. A response level which was unaffected by prey captures, but which decreased with time, would generate a fixed time rule. Iwasa et al (1981) also suggest a behavioural model in which the level of responsiveness decreases with time and is decremented by each prey capture. These behavioural models are summarised in Figure 3.16. Optimal patch departure rules Which of these rules generates the highest rate of prey capture depends on how prey are distributed among patches. If they are distributed randomly (that is, the number of prey per patch has a Poisson distribution), the fixed time rule (Figure 3.16e) is optimal (Iwasa et a U 1981; Stewart-Oaten, 1982; Green, 1984). If the prey distribution is completely regular, the fixed number rule is most adaptive (Iwasa et al, 1981), provided within-patch search is random and not systematic (Green, 1984). For prey with a binomial or negative binomial distribution, the best rule is one in which the forager’s level .of responsiveness is affected by time and prey captures. If prey have a binomial distribution, successive captures and passing time should decrease the forager’s response level, and a rule such as that in Figure 3.16f is optimal (Iwasa et al, 1981). If the prey have a negative binomial distribution, prey captures should increase the forager’s level of responsiveness, and a model such as that in Figure 3.16a is optimal (Iwasa et al, 1981; Green, 1980, 1984). 82 Figure 3.16 Behavioural models of patch departure rules. The wasp has a level of responsiveness which reflects its estimate of the value of staying in a patch. When this level reaches a threshold (T), the wasp departs. The level ma y change in response to passing time, ovipositions (indicated by arrows) or both. a) Responsiveness decreases with time, but is incremented by each oviposition. The magnitude of the increment decreases with time spent on the patch (see Green, 1980, 1984 and Iwasa et al, 1981). b) Responsiveness decreases with time, but is incremented by each oviposition. The magnitude of the increment increases (up to a maximum) with time between two successive ovipositions (see Waage, 1979). c) Fixed G U T rule. Responsiveness decreases with time, but is returned to its original value by each oviposition. d) Fixed number rule. Responsiveness is unaffected by time, but is decremented a fixed amount by each oviposition. e) Fixed time rule. Responsiveness is unaffected by oviposition, but decreases with time. f) Responsiveness decreases with time, and is also decremented by each oviposition (see Iwasa et al, 1981). Level of responsiveness Time on patch 84 Testing patch departure rule hypotheses The most direct way to test patch departure rule hypotheses is to determine how patch time and the G U T are affected by the number and timing of ovipositions (or prey captures). This was the approach taken by Waage (1979) in his study of patch time allocation by Nemeritis. In the experiments described in this chapter, however, true ovipositions cannot be distinguished from rejections, so it is not possible to measure oviposition rates and G U T s directly. However, the different rules illustrated in Figure 3.16. do generate testable predictions about the outcome of plotting patch time against the number of hosts attacked in each replicate (Iwasa et al, 1981). These predictions are summarised in Figure 3.17. The fixed number rule simply predicts that the same number of hosts should be parasitised in each replicate, irrespective of h o w long the wasp stayed on the patch (Figure 3.17d). The fixed time rule predicts that all wasps should stay on patches for a fixed amount of time, no matter how many hosts have been parasitised up to that time (Figure 3.17e). The patch assessment rules of Green (1980, 1984) and Iwasa et al (1981), in which patch time is incremented by each oviposition (Figure 3.16a), predict a linear or slightly accelerating relationship between patch time and the number of hosts attacked (Figure 3.17a). A model in which the level of responsiveness decreases both with time and with each oviposition (Iwasa et al, 1981) predicts the relationship shown in Figure 3.17f. Each of these four types of patch rule predict a relationship between patch time and the number of hosts parasitised which is common to all replicates: if wasps obey one of these rules, the points from a series of replicates should lie on one curve, irrespective of initial host density. This prediction is not true of rules based on a fixed G U T (Figure 3.16c) or of the model developed by Waage (1979) (Figure 3.16b). In these models, each oviposition increments patch time by an amount dependent on the time since the last oviposition. When ovipositions occur at a rapid rate, each successive one adds relatively little to the total patch time. The predicted relationship between patch time and number of hosts parasitised therefore depends on the host encounter rate, and on how this rate changes with time. It is reasonable to assume that encounter rates will be higher on high density patches, and that encounter rates at all densities will tend to decrease with time. If these assumptions hold, the relationships predicted by these models will be of the form shown in Figures 3.17b and 3.17c. Although points from separate replicates will not lie on a 85 Figure 3.17 The outcome of plotting the time spent by a wasp on a host patch against the number of hosts parasitised by that wasp, as predicted by the six theoretical patch departure rules illustrated in Figure 3.16. In (b) a n d (c), curve (i) represents a high density host patch, and curve (ii) a low density patch. See Figure 3.16 and text for explanation of patch departure rules. Number of hosts parasitised Time on patch 87 single curve, there should be a general relationship between patch time and number of hosts attacked within each host density. For the experiments described in this chapter, the observed relationships between patch time and the number of hosts parasitised by A. glomeratus are shown in Figures 3.2-3.5. Contrary to the predictions illustrated in Figures 3.17a, 3.17d, 3.17e a n d 3.17f, the points from different host density treatments do not follow a c o m m o n relationship. Nor does the shape of the curves in Figures 3.2-3.5. tally with any of those in Figures 3.17a, 3.17d, 3.17e a n d 3.17f. Thus the results do not support the fixed number or fixed time rules, nor the assessment rules proposed by Green (1980, 1984) and Iwasa et al (1981). They are most consistent with the predictions generated by a fixed G U T rule, or by the model of Waage (1979). There are additional sources of information parasitoids could use to decide how long to stay in each patch. For example, they may be able to make an initial assessment of patch quality; based, perhaps, on the perceived concentration of host kairomone: in his patch time model, Waage (1979) allowed the w a s p ’s initial level of responsiveness to vary according to host density. Another potentially important source of information for parasitoids is the rate of encounter with previously-parasitised hosts (van Lenteren, 1981; Morrison & Strong, 1981; Lessells, 1985); or the ratio of this rate to the rate of encountering new hosts (Morrison & Lewis, 1981). van Alphen & Vet (1986) discuss some of the other sources of information parasitoids might use to determine patch time. To investigate further the rules A. glomeratus m a y use to decide when to leave a host patch, it is necessary to conduct more detailed experiments, in which oviposition rates, rejection rates and G U T s can be directly measured. The duration of patch visits made by A. glomeratus may also be affected by the defensive behaviour of P. brassicae larvae. Premature patch departure can occur if the fluid exuded by caterpillars impairs the wasp’s mobility to such an extent that it falls from the patch. Three of the eight wasps used in Experiment 2 on patches of 50 hosts were observed to fall from the leaf onto the floor of the cage, unable to fly with fluid-covered wings. Involuntary departures such as these may contribute to the variability in patch time found in Experiments 1 and 2. However, the five wasps which were observed to fly 88 off patches of 50 'h6sts in Experiment 2 still showed considerable variation in patch time: from 24 to 108 minutes. Morrison & Lewis (1981) also found a high degree of variation in the patch time of Trichogramma pretiosum Riley, an egg parasitoid of Heliothis zea Boddie. They found a linear relationship between patch time and the number of host eggs attacked when they plotted the results for different host densities together. They concluded that patch time in T. pretiosum is determined by the rate at which host eggs are attacked, rather than by an initial assessment of host density. Their results are consistent with the predictions of the patch departure rules modelled by Green (1980, 1984) and Iwasa et al (1981) (Figures 3.16a a n d 3.17a). 89 CHAPTER FOUR A COMPARISON OF THE SEARCHING EFFICIENCIES OF A. GLOMERATUS A N D A. R U B E C U L A F O R P. B R A S S I C A E A N D P. R A P A E . 4.1. INTRODUCTION Theoretical studies of optimally foraging parasitoids assume that natural selection acts upon wasp behaviour to maximise oviposition rate (Cook & Hubbard, 1977; Comins & Hassell, 1979). The results in Chapter Three show that female A. glomeratus m a y spend over 100 minutes on patches of P. brassicae. Restriction of search to small areas may be adaptive if the host, like P. brassicae , is gregarious. If it is solitary, a different strategy will be advantageous. P. r a p a e is aggregated on a plant-to-plant basis (Harcourt, 1961; Jones & Ives, 1979), but within each plant caterpillars are solitary and dispersed. In this case the host patch corresponds to the whole plant, rather than to a small part of one leaf. Accordingly, Nealis (unpublished) found that A. rubecula immediately flew off the leaf after ovipositing in a P. r a p a e larva. Most of these flights ended in a return to the same plant. If the foraging behaviour of British A. g l o m e r a t u s is adapted to P. brassicae, its usual host, it is likely to search less efficiently for P. r a p a e , a host it rarely parasitises in the field. By the same argument, A. rubecula , when compared with A. g l o m e r a t u s , should be a more efficient parasitoid of P. r a p a e , which is its only known host. However, in the United States, A. glomeratus has been confined with P. r a p a e for about 300 generations. It might therefore be predicted that natural selection will have increased the searching efficiency of American A. glomeratus for P. r a p a e towards that expected of A. rubecula. The aim of this chapter is to test these predictions. Firstly, it is necessary to define precisely what is meant by "searching efficiency". Searching efficiency In general, an efficient parasitoid attacks a greater proportion of the available hosts, in a given period of search time, than an inefficient one (Hassell, 1982b). Searching efficiency depends on a number of behavioural components (Holling, 1959). These include responsiveness to host cues; speed of movement; 90 the proportion of attacks that are successful; and the degree of area-restricted search. Searching efficiency has acquired a more precise theoretical definition following its inclusion in models of predator-prey (or parasitoid-host) interactions. Nicholson (1933) assumed that the number of encounters with hosts (Ay made by P t parasitoids is in direct proportion to host density (Ay : N e = a N t P t (1) The constant of proportionality (a ) is a measure of the lifetime searching efficiency of the parasitoids, termed the "area of discovery". It can be rewritten in terms of an instantaneous searching efficiency (a ’), such that a = a ’ T with T equal to the total time available for search (Hassell, 1978). Equation (1) can then be rewritten as Ne = a'TNtPt (2) This predicts a linear relationship between N e and N t. Rearranging equation (2) in terms of a ’ gives a’ = N e / { N t T P t) (3) Thus a \ the instantaneous searching efficiency, is the number of encounters per host per unit time per parasitoid. Holling (1959) pointed out that time spent dealing with each host, and in cleaning or resting as a result of host encounters, will reduce the amount of time available for search. Such time "wasted" in non-searching activities he collectively called handling time (Ty. The actual search time (7y is then given by 7\ = T - T h (N e ,!Pt) 91 Substituting this expression into equation (2) gives Holling’s (1959) disc equation: N e = a’N tPt(T-Th (Ne/Pt)) or N e / P t = a ’ T N t/ ( l + a ’ T h N t) (4) This predicts that N e increases with N t at a decelerating rate, eventually reaching an asymptote set by the constraints of handling time. The searching efficiency defines the rate at which the curve approaches this asymptote. Rearranging equation (4) in terms of a ’ gives a’ = N e/(NtPt(T-Th (Ne/Pt))) which is identical to equation (3) except that search time (T s) replaces total time. Searching efficiency in Holling’s disc equation is therefore the number of encounters per host, per unit of search time, per parasitoid. Estimating a ’ It is difficult to measure the searching efficiency of a parasitoid directly. The conventional procedure is instead to obtain an indirect estimate of a ’ by conducting a functional response experiment. The functional response of a parasitoid is the relationship between N e and N t, with P t and T held constant (Soloman, 1949; Holling, 1959). Holling (1959) described three forms the functional response might take. Type I responses are of the linear form predicted by Nicholson’s (1933) model (equations (1) and (2)). The decelerating asymptotic curves predicted by the disc equation (equation (4)) are termed Type II responses by Holling (1959). Finally, the third form recognised by Holling (Type III functional responses) has a more complex, sigmoidal shape, generated by allowing a ’ to vary with By finding, experimentally, the relationship between N e and (with P t and T held constant), and then fitting to the data an equation such as Holling’s disc equation, it is possible to obtain an estimate of a ’ and T^. In practice, however, it is easier to measure the total number of hosts attacked ( A y , rather than the number of encounters (N e). The disc equation must therefore 92 be rewritten in terms of NQ. Thompson (1924) pointed out that if parasitoids searched host patches randomly, the probability of a host remaining unencountered (P ) is given by the zero term of the Poisson distribution, specified by a mean of N e / N t encounters per host. That is, PQ = exp ( - N e / N t) The number of hosts encountered one or more times (i.e. the total number attacked) is then given by N a = N t (1 - exp ( - N e / N t)) Substituting a term for N e from equation (4) gives N a = N t (1 - exp (-a ’ T P t/(\ + a 1 T h N t))) (5) an equation obtained by Royama (1971) and Rogers (1972), and termed by Rogers the random parasite equation. With P t and T held constant, equation (5) can be fitted to a plot of NQ against and an estimate of the parameters a ’ and T ^ obtained. This chapter describes the results of applying this procedure to the following four functional responses. 1. British A. g l o m e r a t u s on P. brassicae. 2. British A. glomeratus on P. rapae. 3. British A. rubecula on P. rapae. 4. American A. g l o m e r a t u s on P. rapae. 4.2. MATERIALS AND METHODS Experiment 1: British A. glomeratus on P. brassicae Newly-hatched P. brassicae larvae were placed on one leaf of a potted Brussels sprouts plant (of the same age, size, shape and leaf number as those described in Chapter Three) at one of five approximate densities: 1, 5, 10, 25 or 50 larvae per leaf. The plants were placed in individual 40cm x 22cm cylindrical 93 Watkins & Doncaster cages, with muslin tops and clear-plastic sides, and left for eighteen hours at 20°C before being used in the experiment. Each replicate consisted of five plants, each with a different density of hosts. One standardised female wasp was introduced to each cage and left for two hours, provided it showed searching behaviour within the first thirty minutes (see Chapter Three). The number of hosts parasitised during the two hours was determined by dissection of the caterpillars in isotonic insect ringers solution forty-eight hours after the experiment. The experiment was replicated six times at each host density. Experiments 2-4: using P. rapae as host Newly-hatched P. rapae larvae were placed on potted Brussels sprouts plants (of the same age, size, shape and leaf number as those used in Experiment 1) at one of five approximate densities: 1, 5, 10, 25 or 50 larvae per plant. Caterpillars were distributed evenly over the plant so that each leaf received approximately the same number of hosts. The plants were then placed in individual 40cm x 22cm Watkins & Doncaster cages, and left for eighteen hours at 20 °C: during this time the larvae create feeding damage, frass and silk. The experimental procedure was then as described for Experiment 1. Each experiment was replicated six times at each host density. 4.3. RESULTS 4.3.1. Experiment 1: the functional response of British A. glomeratus to P. brassicae Figure 4.1. shows the functional response of British A. glomeratus to its usual host, P. brassicae. The data fit a Type II functional response, with a decreasing proportion of hosts parasitised as host density increases. iue . Functonal epo e Brts ritish B f o se on resp l a n tio c n u F 4.1 Figure Number of hosts parasitised ven i e 4.1. le b a T in n e iv g ast e i wih ith w , n tio a u eq site ra a p ol ld -o y a d . r ssicae bra P. ave Th c v i te it a om d ran d fitte the is rve cu he T larvae. T s de sity en d ost H 94 = 2 hours; hours; 2 = . lmeaus eratu glom A. Pt 1 ad ' d n a a' and 1; = iii one g sitisin a r a p T^ as 95 The random parasite equation was fitted to the data using a non-linear least- squares curve-fitting procedure. The linear regression technique proposed by Rogers (1972) is simpler, but suffers from statistical problems and yields biased parameter estimates (Thompson, 1975; Cock, 1977; Hassell, 1978). Table 4.1. shows the estimates of a’ and T^ obtained by fitting the random parasite equation. The estimate of T^ indicates that constraints of handling time will, under the conditions of this experiment, cause the functional response to asymptote at about NQ = 22. The estimated value of a’ is very large: much larger than the usual values obtained for parasitoids from functional response experiments (see, for example, Hassell (1978)). This is because the functional response in Figure 4.1. approaches an asymptote very rapidly: parasitism at low values of N t is alm ost 100%. Rates of parasitism as high as this generate estimates of a’ approaching infinity. The random parasite equation states that as a’ gets large, N a/Nt approaches a maximum, dependent on the values of T^ and N t, i.e. 1 - exp(-fl’ T P t/{ 1 + a ’ Th N t)) = 1 - exp(-r P t/T h N t) Therefore, with T = 2 hours and Pt = 1; L im it ( ) a^ e o N“/N‘ = i - «p (-2/** w,) 6 Equation (6) specifies the proportion of hosts which can be parasitised, for a given value of if all the available time (in this case, two hours) is spent handling hosts, and none is spent searching. Very large estimates of a ’ are generated if the random parasite equation is fitted to data in which some va lu es o f N Q/N t exceed this theoretical maximum. This is the case with Experiment 1. If the twelve points representing 100% parasitism in Figure 4.1. are excluded from the fit, the estimated value of a’ falls from 121 hr-1 to 3.87 hr-1 (+ 16.69 S.E.), whilst T^ is altered only slightly to 0.083 + 0.033 (S.E.) hr. The estimate of a’ is therefore very sensitive to changes in NQ at low values of N p Consequently, the standard error associated with the estimate of a ’ is very large (Table 4.1.). Interestingly, if the parameters were estimated by Roger’s (1972) linear regression technique, points representing 100% parasitism would have necessarily to be removed because the method involves calculating the logarithm of the number of hosts remaining unparasitised at the end of the experiment. 96 Experiment a' (hrs"1) T h (hrs) Asymptote = T I T ^ 1 120.93 ± 10335 0.092 ± 0.022 2 2 2 0.09 ± 0.06 0.204 ± 0.234 10 3 0.25 + 0.07 0.005 ± 0.022 400 4 0.72 + 0.91 0.375 ± 0.092 5 Table 4.1. Estimates (+ S.E.) of a ’ and T ^ obtained by fitting the random parasite equation to the data in Experiments 1-4, with P t = 1 and T - 2 hours. The asymptote ( = 2 / T ^ ) is given to the nearest integer. Experiment 1: British A. glomeratus on P. brassicae. Experiment 2: British A. glomeratus on P. rapae. Experiment 3: British A. rubecula on P. rapae. Experiment 4: American A. glomeratus on P. rapae. 4.3.2 Experiment 2: the functional response of British A. glomeratus to P. r a p a e Figure 4.2. shows the functional response of British A. glomeratus to its less preferred host, P. rapae. Comparison with the curve in Figure 4.1. (see Figure 4.5.) shows that under the conditions of this experiment, British A. glomeratus parasitised a lower proportion of P. r a p a e than of P. brassicae. Table 4.1. shows the values of a' and T ^ obtained by fitting the random parasite equation. The estimate of a ’ is much lower than that from Experiment 1 because parasitism at low values of N t is well below 100%. The estimate of T ^ is larger, indicating a lower asymptote. 4.3.3. Experiment 3: the functional response of British A. rubecula to P. r a p a e Figure 4.3 shows the functional response of British A. rubecula to its only known host, P. rapae. Over the range of host densities tested, N a increases almost linearly with N t. This suggests a Type I functional response in which handling time is regarded as zero. Fitting the random parasite equation yields a correspondingly low estimate of 7^, indicating a very large asymptote (Table 4.1.). 97 A. rubecula parasitised a greater proportion of P. r a p a e than did British A. glomeratus (see Figure 4.5.)* The estimate of a ' obtained by fitting the random parasite equation was consequently larger than that estimated from Experiment 2 (Table 4.1.). 4.3.4. Experiment 4: the functional response of American A. glomeratus to P. r a p a e Figure 4.4. shows the functional response of American A. glomeratus to P. rapae.. Figure 4.5. shows that the overall amount of parasitism imposed by American A. glomeratus on P. r a p a e was no greater than that achieved by British A. glomeratus , and there is no significant difference in the percentage parasitism exerted by individuals of the two populations (for analysis, data were pooled within each of the following host density classes: 1, 4-7, 8-12, 23- 28 and 39-56. A two-way A N O V A on arcsine square-root transformed data was then performed: F^ 5Qj = 2.9; p > 0.05). The relatively high rate of parasitism at low values of N t (Figure 4.5.) produces an estimate of a ’ larger than that obtained in Experiment 2 (Table 4.1.). This estimate is also larger than that for A. rubecula (Table 4.1.). However, Figure 4.5. shows that, overall, the proportion of P. r a p a e parasitised by American A. glomeratus was lower than that parasitised by A. rubecula. The difference in percentage parasitism is significant ( A N O V A as before: F (150) = 15.5; p < 0.01). The estimate of obtained for American A. g l o m e r a t u s is slightly higher than that obtained for British A. g l o m e r a t u s on P. r a p a e (Experiment 2), indicating a lower asymptote (Table 4.1.). 4.3.5. Factors affecting the asymptote of the functional response curves The asymptotic nature of Type II functional response curves has traditionally been attributed to constraints imposed by handling time (e.g. Holling, 1959). In this context, handling time includes all time spent, as a consequence of encountering hosts, in activities other than searching {e.g. handling accepted and rejected hosts, cleaning and resting). The results in Chapter Three show Figure 4.2 Number of hosts parasitised 10 15 0 5 given given in Table parasite equation, with day-old Functional Functional response of British . e a p a r P. 4.1. ave Tecre iscurve fitted the random The larvae. T = 2 = T Host density 98 hours; .glomeratus u t a r e m o l g A. t P 1;= and parasitising one ’ a and and ^ T as iue43 Functionalresponse Britishof Figure 4.3 Number of hosts parasitised given in Table parasite equation, with day-old . e a p a r P. 4.1. lra. h cre is fitted curvethe random The larvae. 99 T = 2 hours; = 1; = and = 2 hours; . a l u c e b u r A. parasitising one a ’ and ^ T as gr 44 i rs ns of rcn erican m A f o se on resp l a n tio c n u F 4.4 igure F Number of hosts parasitised ven i e 4.1. le b a T in n e iv g ast e i wih ith w , n tio a u eq site ra a p ne ol ld -o y a d e on . rapae P. ave The ur i t it random d fitte e th is e rv cu e h T larvae. 100 T s de sity en d ost H = 2 hours; hours; 2 = . omeat s tu era m lo g A. t P 1 a d an 1; = astsng sitisin ra a p a and d n a ’ T^ as iue . A cmparsn of he it unctonal epo e ur s in es rv cu se on resp l a n tio c n fu d fitte e th f o rison a p com A 4.5 Figure Number of hosts parasitised gl rt s eratu m lo g . A . rapae\ P. gur 41 44 Cure ae s olows 1 British B - 1 s: w llo fo as are rves u C 4.4. - 4.1 re u ig F . omeaus eratu m lo g A. - iih ritish B - 3 on on on on . rapae. P. . asiae ssica ra b P. . rubecula A. 101 s de sity en d ost H 2 Brts ritish B - 2 ; on on . apae\ a p ra P. . omeat s tu era m lo g A. - rc n erica m A - 4 on 102 that time spent cleaning, possibly as a result of host defence, is an important factor limiting the number of P. brassicae parasitised by British A. glomeratus. Figure 4.3. shows that handling time is less of a constraint to A. rubecula. T his is partly because it has a much shorter oviposition time than A. glomeratus: it takes less than one second to deposit its single egg. Egg depletion may also impose an asymptote to the functional response (Murdoch, 1973; Hassell, 1982a; Hassell & Waage, 1984). To investigate the possible role of egg depletion in this study, Table 4.2. shows the number of eggs laid in Experiments 1-4. Only data from replicates using the highest host density class (39-56 hosts per plant) are shown. In each experiment, fewer eggs were laid at the lower host densities. Egg supply does not appear to have been an important factor limiting the functional response of British A. glomeratus to P. brassicae: the maximum number of eggs laid in a replicate was 590, with a mean of 328 eggs per replicate. Two to four day-old A. glomeratus have a complement of 1000-1400 mature eggs (see Chapter Five), and are capable of laying at least 700 eggs during a two hour period (see Chapter Three). By the same argument, egg- depletion would not have affected the functional response of British A. glomeratus to P. ra p a, e in which the maximum number of eggs laid per replicate was only 305 (Table 4.2.). A. rubecula is much less fecund than British A. glomeratus, co n ta in in g approximately one tenth as many eggs (see Chapter Seven). However, being solitary it lays a clutch size approximately one twentieth the size of that laid by A. glomeratus. All else being equal, it is therefore less likely to be egg- limited. Up to 30 eggs were laid by A. rubecula in Experiment 3 (although only one larva ever emerges from a host, two or more eggs were occasionally found in a host - see Chapter Two). Two to four day-old A. rubecula h ave a complement of 100-130 mature eggs (see Chapter Seven), so egg depletion is unlikely to have occurred under the conditions of Experiment 3. The maximum number of eggs laid by American A. glomeratus in Experiment 4 was 362, with a mean of 158 eggs per replicate (Table 4.2.). These figures are similar to the equivalent values for British A. glomeratus on P. rapae (Experiment 2). American wasps may be more prone to egg depletion, 103 however, because their complement of 500-600 eggs is smaller than that of B ritish A. glpmeratus (see Chapter Five). E xp erim en t n Mean number of eggs laid (+ S.E.) Range 1 4 328 + 93 161 - 590 2 6 120 + 48 27 - 305 3 6 2 1 + 2 13 - 30 4 6 158 + 54 17 - 362 Table 4.2. Mean (+ S.E.) and range of the number of eggs laid in replicates at the highest host density class (39-56 hosts per plant) in Experiments 1-4. Experiment 1: British A. glomeratus on P. brassicae. Experiment 2: British A. glomeratus on P. rapae. Experiment 3: British A. rubecula on P. rapae. Experiment 4: American A. glomeratus on P. rapae. 4.4. DISCUSSION The results of this study show that, . in comparison with the monophagous A. rubecuia, B ritish A. glomeratus searches relatively inefficiently for P. rapae. This is a host it rarely parasitises in the field. In the United States, however, A. glomeratus has been confined with P. rapae for approximately 300 generations. In spite of this, individuals of the American population of A. glomeratus showed no evidence of any improvement in their searching efficiency for P. rapae. In view of these results, the levels of field parasitism achieved by A. glomeratus in the United States are surprisingly high (see Chapter Two). A possible explanation is that the searching efficiency shown by A. glomeratus in the laboratory is sufficient to achieve these levels of parasitism, but that field parasitism in Britain is kept low by other factors. For example, A. glomeratus larvae are competitively inferior to those of A. rubecula when they occur in the same host (Richards, 1940; Parker et a l, 1971). This competition could decrease the detectable parasitism imposed by A. glomeratus. P arker et al (1971) found that in areas of the USA where A. rubecula was absent, 60% of P. rapae larvae were parasitised by A. glomeratus. In areas where A. rubecula was present, the corresponding figure was only 12%. Although common and 104 widespread in Britain, A. rubecula is still rare and localised in America. Even with rates of parasitism of up to 80% (see Chapter Two), selection might still be expected to have improved the searching efficiency of American A. glomeratus towards that shown by A. rubecula. However, there may have been insufficient genetic variation in the original stock introduced to America. Alternatively, American P. rapae may itself have changed, so that w h ilst A. glomeratus has become adapted to American P. ra p a, e it remains maladapted to British P. rapae. This study does not show which aspects of the foraging behaviour of A. rubecula confer on it a searching efficiency (for P. rapae) greater than that o f A. glomeratus. Nor does it reveal what makes British A. glomeratus a m ore efficient parasitoid of P. brassicae than o f P. rapae. To answer these questions it is necessary to conduct further experiments. 105 CHAPTER FIVE THE EFFECT OF HOST QUALITY ON THE CLUTCH SIZE OF A. GLOMERATUS 5.1. INTRODUCTION Models of parasitoid progeny allocation assume that as clutch size increases, density-dependent within-brood competition for larval resources causes the fitness of each offspring to decrease (Charnov & Skinner, 1984, 1985; Skinner, 1985; Waage & Godfray, 1985). Increases in clutch size may decrease survivorship (see Waage & Godfray (1985) for examples) or offspring size (Beg & Inayatullah (1980), Tagawa et a l (1982), Beckage & Riddiford (1983), Sato & Tanaka (1984) and Sato et a l (1986) give recent examples for A pan teles sp ecies), and numerous studies have demonstrated a relationship between adult parasitoid size and some component of fitness, such as fecundity (e.g. V arm a & Bindra, 1976a; Charnov et a l, 1981; Charnov & Skinner, 1984; Waage & Ng, 1984; Nealis et a l, 1984). In A. glomeratus, large brood sizes are also reported to increase development time (Johannson, 1951; Nealis et a l, 1984), w h ich m ay increase the vulnerability of the brood to pre-emergence predation or hyper parasitism. Figure 5.1a. shows a schematic representation of the relationship between clutch size (c) and fitness per egg (f ( c )). The shape of the function is typical of the examples described by Skinner (1985) and Waage & Godfray (1985). Two curves are shown, representing different intensities of density- dependence. The fitness gained from each host attacked is the product of clutch size and fitness per egg (c f(c )), and the optimum clutch size is that which maximises this function (Waage, 1986). The greater the intensity of density-dependence, the smaller the optimum clutch size will be (Figure 5.1b). Elaborations of this basic model have been developed to include the effects of egg-complement (Parker & Courtney, 1984; Iwasa et a l, 1984; W aage & Godfray, 1985); host encounter rate (Charnov & Skinner, 1984, 1985; Skinner, 1985); and the risk of mortality between hosts (Parker & Courtney, 1984; Iwasa et a l, 1984; Waage & Godfray, 1985). 106 Figure 5.1 a) Schematic representation of a hypothetical relationship between clutch size (c) and fitness per egg (f ( c )). C u rve A: weak within-brood density-dependence. Curve B: strong within-brood density-dependence. b) Relationship between parental fitness per clutch (fitness per egg x clutch size; c f(c )) and clutch size (c). The optimum clutch size (C t) is that which maximises this function. Copt (A) is the optimum clutch size for curve A (weak density-dependence); and CQpt (B) the optimum for curve B (strong density-dependence). 107 f(c) c f(c ) c 108 A prediction of these models is that clutch size should increase as host quality increases, since density-dependent competition for larval resources will be less intense in high-quality hosts. The quality of a host depends on a number of factors, but three important features are its size, the instar it is in when parasitised, and whether or not it has already been parasitised (Waage & Godfray, 1985). This chapter looks at the effect of these three aspects of host quality on the clutch size laid by A. glomeratus. H ost size All else being equal, larger hosts will produce shallower clutch size fitness functions (Waage & Godfray, 1985; Waage, 1986), and the optimum clutch size of a parasitoid will tend to increase with host size (Skinner, 1985; Charnov & Skinner, 1984,1985; Waage, 1986). Many studies have shown that parasitoids allocate more eggs to larger individuals of a host species (Legner, 1969; Purrington & Uleman, 1972; Luck et a l, 1982; Luck & Podoler, 1985), or to individuals of larger host species (Salt (1940), Klomp & Teerinck (1962, 1978), Kot (1979) and Pallewatta (1986) show this for various Trichogramma species). Other studies have shown that, compared with large host species, small hosts may impose greater larval mortality or may produce smaller adult parasitoids (Arthur & Wylie, 1959; Wylie, 1967; Klomp & Teerinck, 1967; Charnov et a l, 1981). Development time may also be reduced in small hosts (Legner, 1969). The two principal hosts of A. glomeratus differ in size: as full-grown larvae, P. brassicae is more than twice as large as P. rapae (see Chapter Two). It might therefore be predicted that, for a given clutch size, wasps developing in P. rapae will suffer higher mortality and will emerge smaller (with attendant effects on fecundity), and possibly earlier, than those developing in P.brassicae. These predictions are tested in Section 5.2.1. There are reasons, however, why British A. glomeratus may not lay a different clutch size in P. rapae than in P. brassicae. One is that parasitoids whose hosts continue to grow after parasitism (koinobiotic parasitoids in the terminology of Askew & Shaw (1986)) cannot assess host size directly at the time of oviposition in the way that idiobiotic (Askew & Shaw, 1986) egg-parasitoids such as Trichogramma spp. can (Waage, 1986). Furthermore, because British 109 A. glomeratus only rarely attacks P. rapae in the field, adjustment of clutch size to host species per se (mediated via host kairomones, for example) is also unlikely. In the United States, however, A. glomeratus has been confined with P. rapae for about 300 generations, so natural selection might be expected to have adapted the clutch size laid by American wasps to the smaller host species. What magnitude of change is expected? Does a transition from one host to another half as large (in weight) imply that half as many eggs should be laid? To answer this question, a comparison can be made of the optimum clutch sizes for the two A. glomeratus populations. In Section 5.2.2., the strength of density-dependent effects on the survivorship, adult size and development time of British A. glomeratus developing in P. brassicae is compared with that of American A. glomeratus in P. rapae. From these data the optimum clutch sizes are calculated. In Section 5.2.3., these are compared with observed clutch and brood sizes to see if adaptation of clutch size in the American population has occurred in the expected direction, and with the predicted magnitude. Host instar attacked For koinobiotic parasitoids such as A panteles spp., host size at parasitism is not always a good indication of host quality (Waage, 1986). Nevertheless, later instars of the noctuid Leucania separata Walk, receive more eggs than earlier instars when parasitised by Apanteles ruficrus Hal. (Tagawa et a l, 1982; Sato et a l, 1986) and A. ka riya eWatanabe (Sato & Tanaka, 1984; Satoet a l, 1986). Fuhrer & Keja (1976) found that parasitism in six day-old P. brassicae larvae ultimately resulted in greater food availability for A. glomeratus larvae than parasitism in one day-old hosts, and Nealis et al (1984) reported that A. rubecula emerged larger, the later the instar of P. rapae in which oviposition occurred. Oviposition in later instars may extend the duration of the host’s final instars, thereby increasing the quantity of food available. Although survival of hosts from parasitism to parasitoid emergence will be higher the later the instar attacked, because of the greater susceptibility of young caterpillars to death from the act of oviposition (Matsuzawa, 1958; Azuma & Kitano, 1971; Nealis et al, 1984) and because of heavy early instar mortality due to rainfall (Harcourt, 1966; Hubbard, 1977) and arthropod 110 predators (Dempster, 1967; Ashby & Pottinger, 1974), this should affect only the decision to oviposit, not the Clutch size laid. If the rate of egg deposition varies depending on the instar attacked, this could affect the optimum clutch size. Time spent laying large clutches must be discounted against the benefits (Iwasa et a l, 1984; Waage & Godfray, 1985), especially if the egg-laying rate decreases during oviposition. Clutch size has been shown to increase linearly with oviposition time in Japanese A. glomeratus parasitising P. rapae crucivora (Matsuzawa, 1958; Ikawa & Suzuki, 1982; Ikawa & Okabe, 1984), although Johannson (1951) found that long oviposition times in P. brassicae did not produce correspondingly large clutch sizes. In Section 5.3., the clutch size and oviposition rates of British A. glomeratus ovipositing in instars I-III of P. brassicae are compared. Previous parasitism Hosts that already contain parasitoid eggs or larvae will provide fewer resources for the offspring of a second female. Superparasitised hosts may also be more likely to die before parasitoid emergence (Johannson, 1951; Walker, 1967). Consequently, models of adaptive progeny allocation predict that fewer eggs should be laid in such hosts (Parker & Courtney, 1984; Skinner, 1985; Smith & Lessells, 1985), a prediction supported by a number of stu d ies (e.g. Wylie, 1965; Holmes, 1972; van Dijken & Waage, 1987). In many parasitoids, complete rejection of previously-parasitised hosts occurs (Wylie, 1965; Salt, 1961; van Lenteren, 1976; van Lenteren et a/, 1978; Escalente & Rabinovich, 1979; van Alphen, 1980). Wylie (1965) discusses some of the mechanisms parasitoids may use to detect that a host has already been parasitised. These include external examination with the antennae, and detection of internal cues by ovipositor insertion (van Alphen & Nell, 1982). Kusano & Kitano (1974) and Ikawa & Suzuki (1982) found that A. glomeratus frequently superparasitises P. rapae crucivora. By comparing the number of eggs dissected from hosts attacked once with the number from hosts attacked twice, they concluded that the second female laid a lower mean clutch size than the first. In neither study were the clutch sizes laid by the two females I l l directly measured. Observations of A. glomeratus females foraging on host patches reveal two types of behaviour associated with ovipositor insertion. These are described in Chapter Three, where they are termed "oviposition behaviour” (insertion of ovipositor for more than two seconds) and "probing behaviour" (insertion of ovipositor for two seconds or less). A hypothesis to explain these two types of behaviour is that the first represents egg-laying, whilst the second represents rejection of a previously-parasitised host. In Section 4, this hypothesis is tested, and the frequency with which A. glomeratus will parasitise previously- parasitised P. brassicae larvae is investigated. Finally, the clutch sizes laid in parasitised and unparasitised hosts are compared. 5.2. HOST SIZE 5.2.1. The effect of host species on parasitoid development 5.2.1.1. Materials and methods Inexperienced 1-3 day old female wasps were removed from the British A. glomeratus culture and placed in individual 50mm x 25mm muslin-topped glass tubes. 5-10 one day-old host larvae ( P. rapae or P. brassicae) w ere introduced to each tube on a small piece of host-damaged leaf, and each wasp was allowed to oviposit for about thirty minutes. Under these conditions, wasps will superparasitise some hosts, and a wide range of clutch sizes can be obtained. After removal from the tube, the hosts were placed in muslin- topped plastic dishes provisioned with fresh Brussels sprouts leaves. About twenty larvae were kept in each dish. Fresh leaves were provided each day, and old leaves and frass removed. The time from parasitism to larval emergence was recorded. After emergence, the cocoons from a single brood were left to harden for twenty-four hours. They were then placed in an individual 50mm x 25mm muslin-topped glass tube. The host remains were dissected in isotonic insect ringers solution under a binocular dissecting microscope, and any dead unemerged second or third instar parasitoid larvae counted. 112 After adult emergence, the number of opened and unopened cocoons, and the number of adults, were recorded, and the head widths of all emerged adults were measured under a compound microscope fitted with an eye-piece graticule. Head-width is an easily-measured index of body size, and is not subject to post-mortem distortion. The time from larval to adult emergence was recorded. Clutch size was estimated from the number of cocoons (plus any larvae that emerged but failed to spin cocoons) plus the number of dead second and third insta’r larvae found in the host after larval emergence. This ignores mortality of eggs and first instar larvae, which cannot be detected by this method. However, Ikawa & Okabe (1984) compared the regression of this estimate of clutch size on oviposition time with that of a direct measurement based on dissection of the host immediately after oviposition. They found no significant difference in the slope or intercept of the two regressions, and concluded that the clutch size laid by A. glomeratus can be reliably estimated from the number of emerged and unemerged larvae. As a crude estimate of fecundity, egg-load (the number of mature eggs in the abdomen) at emergence can be used (Charnov & Skinner, 1984; Waage & Ng, 1984). To determine the relationship between adult size and initial egg load, forty newly-emerged adult female A. glomeratus were killed and dissected in isotonic insect ringers solution. The eggs dissected from each female were counted under a binocular dissecting microscope. The head-width of each female was measured under a compound microscope fitted with an eye-piece gra ticu le. To determine the number of eggs laid during a single oviposition in P. rapae an d P. brassicae, six standardised female wasps in individual 50mm x 25mm muslin-topped glass tubes were each allowed to oviposit in 5-10 one day-old host larvae, presented singly on a small piece of host-damaged leaf. Each wasp was allowed to oviposit once in each host, and the duration of each oviposition - from insertion of the ovipositor to its withdrawal - was recorded by stopwatch. The experiment was performed once with P. brassicae as host, and on ce w ith P. rap ae, using different wasps in each case. After parasitism, caterpillars were placed in individual 50mm x 25mm muslin- topped glass tubes provisioned with fresh Brussels sprouts leaves, and left for 113 forty-eight hours. They were then dissected in isotonic insect ringers solution, and the number of eggs laid counted with the aid of a binocular dissecting microscope. 5.2.1.2. Results 5.2.1.2.1. Development time Table 5.1. shows the mean egg to adult development time of British A. glomeratus in P. brassicae and P. rapae. Egg-larval development was slightly, but significantly, shorter in P. rapae than in P. brassicae (t(57) = 4.1; p < 0.01) n Mean development time (days) S.E. H ost: P. brassicae E gg-larval 41 24.3 0.3 P upal 41 7.6 0.1 T otal 41 31.9 0.2 P. rapae E gg-larval 18 22.0 0.6 P upal 18 8.3 0.2 T otal 18 30.3 0.7 Table 5.1. Egg-larval, pupal and total development time of British A. glomeratus in P. brassicae and P. rapae at 20 °C and 70% R.H. 5.2.1.2.2. Juvenile survivorship Figure 5.2. shows the proportion of British A. glomeratus larvae surviving to adulthood in broods from P. brassicae (Figure 5.2a) and P. rapae (Figure 5.2b) as a function of clutch size. There is a significant negative effect of clutch size on the survivorship of broods in P. rap ae, but not on those in P. brassicae. Survivorship of individuals in broods of thirty or more larvae is lower in P. rapae than in P. brassicae. 114 Figure 5.2 Relationship between clutch size and the proportion of a brood of British A. glomeratus which survives to adulthood. ‘ a) W ith P. brassicae as host. Regression equation, with arcsine square-root transformation of y axis: y = -0.0013x + 1.13; r2 = 1.4; F (1,32) = °-46 (P > °'05)- b) W ith P. rapae as host. Regression equation, with arcsine square- root transformation of y axis: y = -0.024x + 1.60; r2 = 29.5; F(l,16) = 6'69 (P < °*05)* 115 Proportion surviving Clutch size a* h80 n 1 o O 1 o Proportion surviving 1 t o o o o _ n O O o OO .u o o K) o Clutch size 116 5.2.1.2.3. Adult size Figure 5.3. shows the mean adult head width of British A. glomeratus em ergin g from P. brassicae (Figure 5.3a) and P. rapae (Figure 5.3b). There is a significantly negative effect of clutch size on head width only of wasps from P. rapae. The head widths of wasps emerging from P. rap ae are smaller than those of wasps from P. brassicae, especially when brood sizes are large. Figure 5.4. shows that there is a strong correlation between the head width and egg load of newly-emerged British A. glomeratus. The relationship is approximately linear over the range of values shown. 5.2.1.2.4. Clutch size Table 5.2. shows the mean clutch sizes laid by British A. glomeratus in singly- p resented P. brassicae and P. rapae. There is no significant difference between the two means (t^47j = 0.78; p > 0.05). Host species n Mean clutch size S.E. P. brassicae 28 30.3 1.3 P. rapae 21 31.7 1.2 Table 5.2. Mean clutch size laid by British A. glomeratus in singly- presented one day-old host larvae. 5.2.2. Comparison of the optimal clutch sizes of British and American A. glomeratus. 5.2.2.1. Materials and methods To generate a clutch size fitness function for British A. glomeratus developing in P. brassicae, the data collected in Section 5.2.1. were used. To find the corresponding function for American A. glomeratus parasitising P. rap ae, the procedure outlined in Section 5.2.1.1. was repeated, with P. rapae as host, but using wasps descended from material collected in the field in Massachusetts in 1985 (see Chapter Two). 117 Figure 5.3 Relationship between clutch size and the mean head width of B ritish A. glomeratus adults emerging from each brood. a) W ith P. brassicae as host. Regression equation: y = -0.0006x + 0.70; r2 = 8.0; F(MJ) = 2.78 (p > 0.05). b) W ith P. rapae as host. Regression equation: y = -0.0032x + 0.70; r2 = 28.7; F(1>17) = 6.84 (p < 0.05). 8'0 L W Mean head width (mm) 00 _ 00 o Clutch size o oo o Ci p b\ Mean head width (mm) p l/i Clutch size gr 54 ati p bet he si head wi h) y- ly w e n f o ) th id w d a e (h e iz s e th n e e tw e b ip h s n io t la e R 5.4 igure F Egg complement 1000 1500-. - 0 0 5 - quain: = 37 - 2225. - 4337x = y : ation u eq o e nt Corlto ci 09. egression R 0.92. = t n ie ic f f e o c orrelation C t. en lem p com ts itish r B d e g r e m e .0 0.70 0.60 gl us tu a r e m lo g . A a wi h ( ) m (m th id w ead H 119 0 es and t r e g eg ir e th d n a s le a m e f l — 0.80 120 To compare the egg load of females from the two populations, the following experiment was conducted. Because egg load in A. glomeratus is known to vary with female age (Hubbard, 1977; Kitano, 1978), the egg complements of females of nine different ages were determined. Four females of a known age, without ovipositional experience, were removed from culture, killed and dissected in isotonic insect ringers solution under a binocular dissecting microscope; The number of mature eggs dissected from each female was recorded. The experiment was conducted once using British A. glomeratus, and once with American A. glomeratus. 5.2.2.2. Results 5.2.2.2.1. The effect of clutch size on juvenile survivorship Figure 5.5. shows the proportion of American A. glomeratus surviving to adulthood as a function of clutch size. There is a significant effect of clutch size on the probability of survival, and the survivorship of American A. glomeratus in large broods is less than that of British A. glomeratus in P. brassicae (compare with Figure 5.2a). 5.2.2.2.2. The effect of clutch size on adult size Figure 5.6. shows the mean head width of American A. glomeratus adults emerging from P. rapae as a function of clutch size. There is a significant and non-linear effect of clutch size on the mean head width of American wasps from P. ra p a, e and these wasps tend to be smaller than those from P. brassicae (compare with Figure 5.3a). 5.2.2.2.3. Egg load Figure 5.7. shows the mean egg load of British A. glomeratus fro m P. brassicae, and of American A. glomeratus from P. ra p a, e as a function of female age. In both populations egg load increases with age to a peak in females 6-8 days old. It then decreases, possibly because eggs are resorbed in old, non-ovipositing females (Flanders, 1942), although King et a l (1969) state that A. glomeratus does not resorb eggs. gr 55 l i hi t e he l c ie ai y erican m A by id la size tch clu e th een etw b ip sh n tio ela R 5.5 igure F Proportion surviving . lmeaus eratu glom A. rsne quaero tansor i y xs y - 01x + 16x .0 -0 = y axis: y f o n tio a rm sfo n tra are-root u sq e arcsin c ur ve o t Re eson equaton, ith w , n tio a u q e n ressio eg R . d o o lth u d a to ed iv rv su ich h w .9 r = 22 F( 5 = 82 ( < 0.001). < (p 18.23 = 25) (1 F 42.2; = r2 1.59; n in . rapae P. 121 nd t prpoto ec brood each f o ortion rop p e th d an g e . Reatons p bewen t cl ch sze l d by erican m A y b id la e siz h tc lu c e th een etw b ip sh n tio ela R 5.6 re igu F Mean head width (mm) 0 5- .5 0 7- .7 0 0 . . 6 8 - -. 0 . lmeaus eratu glom A. megng rm ec bod Re eson equaton, t log- ith w , n tio a u q e n ressio eg R brood. each from g ergin em 12) 1.7 .0) - 0.001). < p ( 17.37 (1|25)= F ransf s y - 144x + 0216; = 1.0; 41 = 2 r ; 6 1 2 .0 0 + x 4 4 .1 -0 = y es: x a d e m r fo s n a tr I------1------1 “ n in 0 0 0 80 60 40 20 . rapae P. n t men hed wi h of dults u ad f o th id w ead h ean m e th and 122 uth sze siz tch lu C • • • • • ------✓ 1 gr 57 an egg l + E. of non-ovi ti emale a m fe g in it s o ip v o - n o n f o .) .E S (+ d a lo g g e n ea M 5.7 igure F E gg load . lmeaus eratu glom A. i e t megnc) Clsd ice: iih ritish B circles: losed C ce). ergen em lt u d a ce sin rmfrom rmfrom emal fo ah po aton a ec age. each at n tio la u op p each from s le a m fe . brassicae. P. . rapae. P. s f ton of emal ( r days f o er b m u (n e g a le a m fe f o n ctio n fu a as as r bsd o dis i fur fou f o n tio c isse d on based are eans M e crls Ameian n erica m A circles: pen O mae g ( ays) (d age ale em F 123 . omeaus eratu m lo g A. . omeaus eratu m lo g A. 124 The egg load of American A. glomeratus was much lower than that of British A. glomeratus at most ages, consistent with the smaller size of A. glomeratus from P. rapae. 5.2.2.2.4. The effect of clutch size on development time Figure 5.8. shows the relationship between clutch size and egg to adult development time for British A. glomeratus in P. brassicae (Figure 5.8a), and A m erican A. glomeratus in P. rapae (Figure 5.8b). There is no significant effect of clutch'size on development time in either case. Table 5.3. shows the mean development times of wasps from the two populations: there are no significant differences between the results for British and American A. glomeratus. n Mean development time (days) S.E. B ritish A. glomeratus E gg-larval 41 24.3 0.3 Pupal 41 7.6 0.1 T otal 41 31.9 0.2 A m erican A. glomeratus Egg-larval 29 24.1 0.5 P upal 29 7.6 0.1 T otal 29 31.8 0.5 Table 5.3. Mean duration of egg-larval, pupal and total development time of British A. glomeratus in P. brassicae, and of American A. glomeratus in P. rapae. Measured at 20 °C, 70% R.H. 5.2.2.2.5. Comparison of the clutch size fitness functions The clutch size fitness function describes the fitness of an egg laid in a host as a function of clutch size. This fitness has two components: the probability of survival from egg to adult, and the fitness of the ensuing adult. Adult fitness depends on factors such as longevity, female fecundity and male mating ability; and will be related to adult size. Empirical studies of optimal clutch 125 F igure 5.8 Relationship between clutch size and the egg to adult development time of A. glomeratus, measured at 20 °C and 70% R .H . a) B ritish A. glomeratus developing in P. brassicae. b) A m erican A. glomeratus developing in P. rapae. 126 Development time (days) K> K> UJ O O O Clutch size J o • o' • I o u> Development time (days) t o o _ oo o o on -U o to o Clutch size 127 size can consider only a subset of all the possible factors affecting offspring fitness. Waage & Godfray (1985), for example, considered only egg-larval survivorship when calculating the optimum clutch sizes of six species of parasitoid. Ikawa & Okabe (1985) used adult weight in combination with survivorship. Charnov & Skinner (1984) incorporated egg load (estimated from adult size) with survivorship to find the optimum clutch sizes of two species of parasitoid. The simplest way to describe the effect of clutch size on offspring fitness in the present study is to use egg to adult survivorship only. This is shown in Figure 5.2a for British A. glomeratus on P. brassicae, and in Figure 5.5. for A m erican A. glomeratus on P. rapae. Figure 5.9. shows the effect of combining probability of survival (Figures 5.2a and 5.5.) with mean adult head width (Figures 5.3a and 5.6.) on the clutch size fitness functions of British A. glomeratus on P. brassicae (Figure 5.9a), and American A. glomeratus on P. rapae (Figure 5.9b). Because head width is highly correlated with egg load (Figure 5.4.), mean head width can be converted to mean egg load using the regression equation in Figure 5.4. Figure 5.10. shows the clutch size fitness functions generated by using probability of survival and mean egg load as an estimate of offspring fitness. The data for British A. glomeratus on P. brassicae are shown in Figure 5.10a, and for American A. glomeratus on P. rapae in Figure 5.10b. 5.2.2.2.6. Comparison of the optimal clutch sizes An empirical method of determining the optimal clutch sizes of British A. glomeratus in P. brassicae, and of American A. glomeratus in P. ra p a,e is to plot the product of clutch size (c) and fitness per egg (f(c )) against clutch size. This is done in Figure 5.11., using the most complete estimate of offspring fitness available: probability of survival x mean egg load (from Figure 5.10). The absence of density-dependent offspring fitness in broods of British A. glomeratus developing in P. brassicae produces the linear function shown. Over the range of clutch sizes generated, parental fitness continues to increase with clutch size, and there is no optimum. For American A. glomeratus in P. rapae, however, there is a discernible optimum of 20-25 eggs per host. All that can be said for British wasps is that at least 80 eggs should be placed in each clutch. 128 Figure 5.9 Clutch size fitness functions for A. glomeratus, sh o w in g the relationship between clutch size and an estimate of fitness per egg: juvenile survivorship x mean head width of surviving o ffsp r in g . a) B ritish A. glomeratus developing in P. brassicae. b) A m erican A. glomeratus developing in P. rapae. to o BJ V t • • • • • hfr'o i to p _ o b n 00 00 o -U O O o to. o o N* o o d- C/5 d Q Proportion surviving x mean head width o to ' b> o p p p Clutch size 130 Figure 5.10 Clutch size fitness functions for A. glomeratus, sh ow in g the relationship between clutch size and an estimate of fitness per egg: juvenile survivorship x mean egg load of surviving o ffsp rin g . a) B ritish A. glomeratus developing in P. brassicae. b) A m erican A. glomeratus developing in P. rapae. Proportion surviving x mean egg load 131 lth size Clutch gr 51 Rel i hi t e l c ie n a aue pae tal aren p f o easure m a and size tch clu een etw b ip sh n tio la e R 5.11 igure F Fitness per clutch i ft s e eg, i pooto s vi mean m x tch g clu in iv v r m su fro ed tain b proportion o g is sin u tch , clu egg per per ess ss itn e F fitn x e circles). siz en (op cosd crcl ) n Ameia erican m A and s) le c cir sed (clo = 098 + .4 r = 91 Lie hrug nt f r fo ts in o p gh rou th ine L 69.1. = r2 0.04; + n 8x erica .009 m 0 A = y g l d a a etmat of ines pr g. ue of o ss es e lu a fitn V est h ritish ig B h e egg. th for to per n e tio a u tiv ss e eq rela fitn n f o scaled ressio eg R te are a estim tch . clu ed an in ta per b o as ss e ad fitn lo egg ines pr l c or iih ritish B r fo tch clu per ss e fitn glmeaus eratu lom g . A a y eye. by n raw d 132 . lmeratus glom A. . lmeaus eratu glom A. on on . lmeaus: eratu glom A. on on . r sc e ssica bra P. . a ae rap P. 133 With data as scattered as those in Figure 5.11., finding the optimum clutch size by eye is a procedure prone to error and bias. Waage & Godfray (1985) used a more objective method to find the optimum clutch sizes of six species of parasitoid. They fitted the negative exponential expression: f (c ) = exp (-K c) to each clutch size fitness function, where K is a constant describing the intensity of density-dependence. The value of c which maximises the mother’s fitness from a single host is given by the inverse of K (Waage & Godfray, 1985). This function was fitted to the clutch size fitness functions for American A. glomeratus on P. rapae in Figures 5.5., 5.9b and 5.10b. In each case the negative exponential gave a better fit than a linear model. Table 5.4. shows the equations obtained, with the r2 values, and the optimum clutch sizes calculated for each of the three clutch size fitness functions. The inclusion of more detail in the measurement of offspring fitness progressively decreases the optimal solution. 134 Estimate of offspring fitness Fitted function 'opt Probability of survival f ( c ) = 1.44 exp (-0.022c) 53.1 45.5 (Figure 5.5.) Probability of survival x f ( c ) = exp (-0.025c) 56.0 40.0 Adult head width (Figure 5.9b) Probability of survival x f (c ) = 1260 exp (-0.043c) ' 52.2 23.3 E gg load (Figure 5.10b) Table 5.4. The result of fitting the function f ( c ) = exp (-K c ) to the clutch size fitness functions for American A. glomeratus on P. rapae in Figures 5.5, 5.9b and 5.10b. The functions were fitted by non linear least-squares regression. Linear regression gave a worse fit in all cases. The r2 values obtained from linear regression were as follows: 47.0 (Figure 5.5.); 49.2 (Figure 5.9b); and 40.4 (Figure 5.10b). The optimum clutch size (C t) is the inverse of 5.2.3. Comparison of the observed clutch and brood sizes of British and A m erican A. glomeratus. 5.2.3.1. Materials and methods The clutch sizes laid by British A. glomeratus in P. brassicae and P. ra p a, e and by American A. glomeratus in P. rapae, were obtained from the functional response experiments described in Chapter Four. Field brood sizes (number of cocoons) were obtained from P. brassicae larvae collected from Silwood Park (see Chapter Two), and from P. rapae larvae collected in Massachusetts by R.G. van Driesche (see Chapter Two). 135 5.2.3.2. Results 5.2.3.2.I. Clutch size Table 5.5. shows the mean clutch sizes laid by British A. glomeratus in P. brassicae and P. rapae> and by American A. glomeratus in P. rapae in the functional response experiments described in Chapter Four, using data from the highest host density (50 hosts per plant) only. Contrary to prediction, the clutch size laid by American A. glomeratus in P. rapae was significantly larger than that laid by British A. glomeratus in P. brassicae (t^125j = 5.6; p < 0.001). There was no significant difference between the clutch sizes laid by British and American A. glomeratus in P. rapae (t^125j = 0.9; p > 0.05). A. glomeratus popn. Host species n Mean clutch size S.E. B rita in P. brassicae 63 20.4 0.3 B ritain P. rapae 27 26.6 0.4 USA P. rapae 38 28.0 0.5 Table 5.5. Mean clutch size laid by British and American A. glomeratus in the functional response experiments described in Chapter Four, at the highest host density (50 hosts per plant) only. There is a significant effect of treatment on clutch size (one-way ANOVA: F(2t125) = 18'2; P < 0 0°1)- 5.2.3.2.2. Brood size Table 5.6. shows the mean field brood sizes obtained from P. brassicae collected at Silwood Park, and from P. rapae collected in Massachusetts. There is no significant difference between the means (t^131j = 0.9; p > 0.05). The brood size of British A. glomeratus in P. rap ae has not been included in the comparison because-too Tew-broods of British A. glomeratus were obtained from field-collected P. rapae larvae. 136 A. glomeratus popn. Host species n Mean brood size S.E. B ritain P. brassicae 111 26.1 1.0 U SA P. rapae 22 26.4 2.2 Table 5.6. Mean brood sizes (number of cocoons) obtained from P. brassicae larvae collected in the field at Silwood Park, and from P. rapae collected in the field at Amherst, Massachusetts, in 1985. 5.3. HOST INSTAR ATTACKED 5.3.1. Materials and methods Four standardised female wasps from the British A. glomeratus population were placed in individual 50mm x 25mm muslin-topped glass tubes. Each female was allowed to oviposit in five P. brassicae larvae, presented singly on a small piece of host-damaged leaf. Only one oviposition per host was permitted. The duration of each oviposition - from insertion of the ovipositor to its withdrawal - was measured with a stopwatch. After parasitism, hosts were placed in individual 50mm x 25mm muslin-topped glass tubes, provisioned with fresh Brussels sprouts leaves, and left for forty-eight hours. They were then dissected in isotonic insect ringers solution to determine the number of eggs laid in each. The experiment was performed once with four day-old (second instar) hosts, and once with eight day-old (third instar) hosts, using different wasps in each case. Data for one day-old (first instar) hosts were obtained from the experiment described in Section 5.2.1. 5.3.2. Results ~ Table 5.7. shows the mean clutch size laid in each of the first three instars of P. brassicae. There are no significant differences between the means (one-way ANOVA: F^2 64 j = 1.7; p > 0.05), but Figures 5.12., 5.13. and 5.14. show that the range of clutch sizes obtained was much wider in second and especially third in star P. brassicae than in first instars. gr 51 Rel i hi t e poston tme n cuth sz for size tch clu and e tim n sitio o ip v o een etw b ip sh n tio la e R 5.12 igure F Clutch size iih ritish B . assicae. ra b P. . lmeaus eratu glom A. reain coefi ent 03; > 0.05. > p 0.35; = t n ie ffic e o c orrelation C postng i ne y- d (i t nstar) in st (fir ld -o ay d e on in g sitin o ip v o 137 gr 51 Rel i hi t e poston tme nd cuth sze for e siz tch clu d an e tim n sitio o ip v o een etw b ip sh n tio la e R 5.13 igure F Clutch size iih ritish B grsi quain: = .3 + 13.2. + 1.03x = y : ation u eq n ressio eg R in star) star) in . assicae. ra b P. . lmeaus eratu glom A. reain coefi ent 05; < 0.05. < p 0.55; = t n ie ffic e o c orrelation C postng i our ol scnd (secon ld -o y a ,d r u fo in g sitin o ip v o 138 gr 51 Rel i hi t e poston tme n cuth sz for size tch clu and e tim n sitio o ip v o een etw b ip sh n tio la e R 5.14 igure F Clutch size grsi quain: = .6x 13.5. + 0.865x = y : ation u eq n ressio eg R iih ritish B in star) star) in . asiae. ssica ra b P. . l r us tu era m glo A. rl i fci = .3 p 0.001. < p 0.83; = t n ie ffic e o c n tio rrela o C postng i i day- d (hird (th ld -o y a d t h eig in g sitin o ip v o 139 Oviposition time Relationship between oviposition time and clutch size for B ritish A. glomeratus ovipositing in one day-old (first instar) P. rapae. Correlation coefficient = 0.75; p < 0.001. 141 The correlation between oviposition time and clutch size is significant only for instars II and III. There is no significant difference between the slopes in Figures 5.13. and 5.14. (t-test on slopes: t^S3j = 0.04; p > 0.05). Figure 5.15. shows the relationship between oviposition time and the clutch size laid by British A. glomeratus in one day-old (first instar) P. rapae. There is a strong correlation, even over a relatively narrow range of values. Host instar n Mean clutch size S.E. F irst 28 30.3 1.3 Second 20 28.6 2.8 T hird 19 35.2 3.7 Table 5.7. Mean clutch size laid by British A. glomeratus in instars I-III of P. brassicae. 5.4. PREVIOUS PARASITISM This section deals with the following questions: 1. Is probing behaviour restricted to previously-parasitised hosts? 2. Does oviposition behaviour occur in previously-parasitised hosts? 3. If so, are fewer eggs laid in previously-parasitised hosts than in unparasitised ones? 4. Are eggs always laid during oviposition behaviour? 5. Are eggs ever laid during probing behaviour? To determine whether eggs are laid in previously-parasitised hosts, it is necessary to distinguish between the eggs of the first and second females. Other studies have approached this problem by using genetically-marked parasitoids (Holmes, 1972; Werren, 1980, 1983; Bakker et al„ 1985), or by estimating clutch size from external signs of oviposition, such as abdominal movements (van Dijken & Waage, 1987; Suzuki et al.t 1984) or oviposition time (Ikawa & Suzuki, 1982). An alternative approach, and the method used here, is to differentiate between the eggs laid at different times by their size and appearance. After deposition, the eggs of A. glomeratus increase in size and change in appearance as embryonic development proceeds (Tawfik, 1975). van 142 Alphen & Nell (1982) used a similar technique to distinguish between the eggs laid by different females of the solitary braconid parasitoid, Asobara tabida N ees. 5.4.1. Preliminary experiment The sizes and developmental stages of eggs of different ages were determined by the following experiment. On day 0, 76 newly-hatched P. brassicae larvae were parasitised by 2-4 day-old A. glomeratus females (from the British population). Immediately after parasitism, 11 of these hosts were dissected in isotonic insect ringers solution under a binocular dissecting microscope fitted with an eye-piece graticule. The sizes (length and breadth) of the eggs were measured, and their developmental stages characterised. Ten more hosts were dissected on day 1; 28 on day 2; 10 on day 3; 10 on day 4; and 7 on day 5. A total of 977 eggs were measured, and their developmental stages characterised. Figure 5.16. shows the mean lengths of eggs dissected out on days 0-4 (by day 5 the eggs had hatched). Figure 5.17. shows the frequency distribution of egg sizes recorded on each day following oviposition. Figure 5.18. shows the appearance of the eggs on each day. Figures 5.16. and 5.17. show that the parasitoid eggs increase rapidly in length between one and three days after oviposition. Figure 5.18. shows that there is also a considerable change in appearance. These results show that it is possible to differentiate between eggs laid one and three days previously. 5.4.2. Main experiment 5.4.2.1. Materials and methods One newly-hatched P. brassicae larva was placed with forceps on a small piece of host-damaged leaf,, and was presented to a 2-4 day-old standardised female A. glomeratus (from the British population) in a 50mm x 25mm muslin-topped glass tube. On encountering the host, all the females used immediately showed oviposition behaviour ( i.e. there was no probing behaviour, and no host was ignored). The duration of oviposition - from insertion of the ovipositor to its withdrawal - was recorded with a stopwatch. As soon as oviposition was completed, the host was removed from the tube and replaced with another. gr 51 Me engt ( 9 I) ai tsh itis r B y b id la .I.) C 99% (+ s g g e f o th g n le n ea M 5.16 igure F Egg length (mm) O.i J , -J . lmeaus eratu glom A. setd fo he s 0 1 2 3 r das t poston. n sitio o ip v o r fte a ays d 4 or 3 2, 1, 0, ost h e th from issected d 12 3 4 3 2 1 0 ------r days t poston sitio o ip v o r fte a s y a d f o er b m u N n newl hat d e h tc a -h ly w e n in j------,------, 143 . brassicae P. ------ar , hen w e, a rv la , 144 Figure 5.17 Percentage frequency distribution of the lengths of eggs laid by B ritish A. glomeratus in newly-hatched P. brassicae larvae, when dissected from the host 0, 1, 2, 3 or 4 days after oviposition. Egg lengths are given in graticule units, where one unit = 0.022 mm. The value of n is the sample size. Percentage frequency - 0 4 80-i Egglength(graticule units) 145 = 143 = n I i ______l ______1 ______1 ------Day 0 Day 1------1 146 Figure 5.18 Appearance of eggs laid by A. glomeratus in newly-hatched P. brassicae larvae, when dissected from the host a) immediately after oviposition; b) one day after oviposition; c) two days after oviposition; d) three days after oviposition; and e) four days after oviposition. After four days the fully formed larva is visible in the egg, and hatching occurs between four and five days after oviposition. 147 L J OJ m m 148 This procedure was repeated until four females had each parasitised ten hosts. The forty parasitised caterpillars were placed in a muslin-topped plastic container provisioned with Brussels sprouts leaves. Another forty newly- hatched caterpillars were individually picked up with forceps in the same way as the parasitised larvae, but were transferred directly to a second plastic dish provisioned with Brussels sprouts leaves. Two days later, five larvae from each plastic container were presented individually, in the order UPUPUPUPUP (U - unparasitised host, P - parasitised host), to a 2-4 day-old standardised female wasp by the same method as before. The wasps initial behaviour on encountering each host was recorded as follows: Oviposition behaviour - ovipositor insertion lasting more than 2 seconds. Probing behaviour - ovipositor insertion lasting 2 seconds or less. (No behaviour other than oviposition or probing was observed in this experiment). In addition, the duration of oviposition was recorded by stopwatch. This procedure was repeated until six wasps had each been presented with five parasitised and five unparasitised hosts. After the wasp’s initial response to the host, the caterpillar was removed and placed in one of four plastic dishes, depending on the type of behaviour shown by the wasp, and on whether the host was parasitised or unparasitised before the experiment. Thus each dish contained one of the following categories of host: Dish 1: Unparasitised hosts - oviposition behaviour recorded Dish 2: Unparasitised hosts - probing behaviour recorded Dish 3: Parasitised hosts - oviposition behaviour recorded Dish 4: Parasitised hosts - probing behaviour recorded A fifth dish contained parasitised hosts that were not re-exposed to a wasp. The hosts were left in the dishes with fresh Brussels sprouts leaves for twenty- four hours. To check that eggs laid in two day-old caterpillars expand at the same rate as those laid in newly-hatched hosts, 20 two day-old larvae were parasitised, and then 10 dissected after one day; 10 after two days. The 149 frequency distributions of egg sizes on each day are shown in Figure 5.19. They are comparable with those in Figure 5.17. After the twenty-four hour period of egg expansion, caterpillars were dissected in isotonic insect ringers solution under a binocular dissecting microscope fitted with an eye-piece graticule. The numbers, lengths and developmental stages of all eggs were recorded. 5.4.2.2. Results 5.4.2.2.1. Frequency of oviposition and probing behaviour in parasitised and unparasitised hosts Table 5.8. shows the number of wasps which displayed oviposition or probing behaviour when presented with alternately unparasitised and parasitised two day-old hosts. There is a significant difference in the relative frequencies of the two types of behaviour shown to parasitised and unparasitised hosts (X2(j) = 25.9; p < 0.001). Initial behaviour shown by wasp Oviposition Probing Unparasitised host 29 1 Parasitised host 9 21 Table 5.8. Frequency of oviposition and probing behaviour shown by B ritish A. glomeratus when exposed to alternately unparasitised and parasitised two day-old P. brassicae larvae. 5.4.2.2.2. The relationship between observed wasp behaviour and egg-laying Table 5.9. shows the number of hosts, in each of five categories, containing eggs laid one or three days before dissection (as determined by the size and appearance of eggs). The data in Table 5.9. show that of the 30 initially unparasitised two day-old hosts presented to a wasp, 28 were parasitised during oviposition behaviour. One host was probed - and no eggs laid - and to one host the wasp displayed oviposition behaviour, but did not lay any eggs. g e .9 re ag fe nc dit buton of h lngt eg li by laid eggs f o th g len the f o n tio u ib istr d cy en u freq ge ta ercen P 5.19 re igu F Percentage frequency val of i te a e size. le p sam the is n f o e lu a v e h T e hs r gvn i gratcul t, r o uni = .2 mm. 0.022 = it n u e on ere h w its, n u le u tic a r g in given are s th g len iih ritish B setd fo h hs o o t y afer ovi ii Egg E . n sitio o ip v o r fte a ays d o tw or e on host the from issected d go eratus glom . A g e t gratcul its) n u le u tic a r (g gth len gg E n t ol ld -o y a d o tw in 150 . assicae ra b P. ar , hen w e, a rv la 151 None of the 30 parasitised hosts that were re-exposed to wasps were supcrparasitised. Twenty-one were probed (and no eggs laid), and to nine the wasp showed oviposition behaviour, but did not lay any eggs. Number of hosts containing the following: Host category 1 day-old eggs 3 day-old eggs 1 and 3 day-old eggs No eggs U -O 28 0 0 1 U-P 0 0 0 1 P-O 0 9 0 0 P-P 0 21 0 0 p 0 5 0 0 Table 5.9. The number of three day-old P. brassicae larvae containing 1 or 3 d ay -o ld A. glomeratus eggs. Eggs were aged by their size and appearance. Host categories are as follows. U-O. unparasitised host - oviposition behaviour. U-P. unparasitised host - probing behaviour. P-0, parasitised host - oviposition behaviour. P-P. parasitised host - probing behaviour. P = parasitised host - not re-exposed. In this experiment, eggs were never laid during probing behaviour, but ovipositioh behaviour did not always result in egg-laying (i.e. oviposition behaviour does not always imply oviposition). Table 5.10. summarises these results. Oviposition behaviour that did not result in egg-laying is termed "pseudo-oviposition" in Table 5.10. The data in Table 5.10. show that "pseudo- ovipositions" occur significantly more frequently in previously-parasitised hosts than in unparasitised hosts,—relative to the combined frequencies of- oviposition and probing behaviour (X2^) = 5.9; p < 0.05). 152 Ovipositions Rejections Total Probes Pseudo-ovipositions Unparasitised hosts 28 1 1 30 Parasitised hosts 0 21 9 30 Table 5.10. Frequency of ovipositions ( i.e. egg-laying) and rejections (i.e. no esg'laying) shown by A. glomeratus to parasitised and unparasitised two day-old P. brassicae larvae. Rejections are either probes (insertion of ovipositor for 2 seconds or less) or pseudo-ovipositions (insertion of ovipositor for more than 2. seconds without eggs being laid). 5.4.2.2.3. Clutch sizes Table 5.11. shows the mean clutch sizes recorded for each of the five host categories. There are no significant differences between the means (one-way ANOVA: F(3>59) = 0.18; p > 0.05). Mean clutch size (+ standard error) _ Host category 1 day-old eggs 3 day-old eggs U-O 24.6 ± 1.5 (n=28) 0 U-P 0 0 P-0 0 25.1 ± 1.8 (n=9) P-P 0 23.5 ± 1.0 (n=21) P 0 24.0 ± 1.6 (n=5) Table 5.11. Mean clutch sizes (+ S.E.) laid by A. glomeratus in each of five host categories (see Table 5.9. for explanation of categories). Sample size is in brackets after each mean. 5.4.2.2.4. Duration of oviposition behaviour ______. i__:. Table 5.12. shows the mean duration of oviposition behaviour recorded during the experiment. There are no significant differences between the means (one way ANOVA: F(277) = 1.97; p > 0.05). There was therefore no significant difference between the mean duration of oviposition behaviour shown to 153 parasitised and unparasitised hosts. The shortest oviposition behaviour recorded lasted nine seconds (in'a previously unparasitised host), considerably longer than the duration of probing behaviour (two seconds or less, by definition). Duration of ovipositor insertion (s) n Mean S.E. Newly-hatched hosts Unparasitised 40 19.2 2.5 Two day-old hosts Unparasitised 29 26.9 3.6 Parasitised 9 19.7 2.2 Table 5.12. Mean duration of oviposition behaviour shown by A. glomeratus to unparasitised newly-hatched P. bra ssicae larvae, and to parasitised and unparasitised two day-old hosts. 5.5. DISCUSSION H ost size For a given clutch size, the survivorship and adult size of British A. glomeratus developing in P. rapae are less than those of wasps in the larger host, P. brassicae. A difference in development time was also found. However, this was slight and may have been due to variation in temperature, since the two experiments were not conducted simultaneously. Furthermore, the difference was not found in the comparison of British A. glomeratus in P. brassicae w ith A m erican A. glomeratus in P. rapae. Evidence of density-dependent survivorship or adult size was found only for broods from P. rapae. Density-dependence within broods of Japanese A. glomeratus developing in P. rapae crucivora has been reported by Matsuzawa et a l (1957), Matsuzawa (1958) and Ikawa & Okabe (1985). Slansky (1978), however, found no density-dependence of adult weight in broods of up to 154 fifty American A. glomeratus in P. rapae. He suggested that the parasitoid larvae manipulate host growth to ensure that sufficient resources are available for their own development. Fuhrer & Keja (1976) found a similar effect using P. brassicae as host. They discovered that A. glomeratus larvae block the growth of fifth instar hosts, ensuring that nutrients ingested by the host are diverted to parasitoid development. They calculated that if eggs are laid in one day-old hosts, up to sixty larvae can develop in one host without exhausting the food supply. Fuhrer (1980) also found that heavily-parasitised hosts (containing more than fifty A. glomeratus) feed at a higher rate than lightly-parasitised hosts, and for a longer period than unparasitised caterpillars. Johannson (1951), also using P. brassicae as host, detected an increase in the mortality of A. glomeratus larvae only in broods larger than sixty, although a density-dependent effect on development time operated over a range of 5-50 parasitoids per host. In the present study, no density- dependence of adult size, survivorship or development time was found in broods of up to 80 A. glomeratus larvae developing in P. brassicae. Manipulation of host growth to the advantage of parasitoid development has been credited to other gregarious A p a n te les species. For example, a supernumerary host instar may be imposed (Beckage & Riddiford, 1978; Madar & Miller, 1983), or net host weight may increase with parasitoid load (Tagawa et aU 1982; Beckage & Riddiford, 1983; Sato & Tanaka, 1984; Satoet a l, 1986). Density-independence of adult parasitoid size has been found in broods of up to 192 Apanteles liparidis (Bouche) in Lymantria dispar (L.) (Burgess & Crossman, 1929); and in broods of 30-95 A. yukutatensis (Ash.) in noctuid caterpillars (Madar & Miller, 1983). The absence of density-dependence within broods of up to 80 A. glomeratus in P. brassicae precludes the calculation of an optimum clutch size for this host. For American A. glomeratus in P. rap ae, the calculated optimum depends on the clutch size fitness function adopted. Waage & Godfray (1985) found that the observed clutch-sizes, of six.parasitoid ^species-were. all lower than those predicted on the basis of density-dependent survivorship alone. Table 5.4. shows how incorporating additional elements of offspring fitness into the clutch size fitness function for A. glomeratus reduces the predicted optimum. Similarly, Waage (1986) showed that the optimum clutch size calculated for Trichogramma evanescens West, is lower - and nearer the observed value - if the clutch size fitness function includes fecundity as well as survivorship. 155 As expected, the clutch size predicted for the smaller host (P. rapae) is m uch less than that predicted for the larger species (P. brassicae). However, the clutch size laid by British A. glomeratus in P. brassicae is considerably smaller than that expected on the basis of the data shown in Figure 5.11. Assuming that the clutch size of British A. glomeratus is adapted to its usual host, there must be still further density-dependent components of offspring fitness that have not been considered. The reluctance of A. glomeratus to superparasitise P. brassicae (Section 5.4.) also implies that density-dependence must exist. Various other sources of density-dependence are possible. Hubbard (1972), for example, found that heavily-parasitised P. brassicae caterpillars were more vulnerable to avian predators. Parasitism also reduces the thickness of the host cuticle (El Sufty & Fuhrer, 1981a), rendering it more susceptible to pathogenic infection (El Sufty & Fuhrer, 1981b). If this effect is more pronounced at high parasite loads, it could be a source of density-dependence. Johannson (1951) found that, in the laboratory, parasitised P. brassicae w ere more likely to die if they contained particularly large broods of A. glomeratus, an effect also noted by Matsuzawa (1958) for P. rapae. Finally, long oviposition times, associated with laying large clutches, may impose penalties (see Section 5.3.). Despite the incompleteness of the clutch size fitness functions used, the qualitative result remains: the optimum clutch size in P. rapae is considerably less than that in P. brassicae. Indeed, the most common larval parasitoid of P. rapae in Britain, A. rubecula, lays only one egg per host. Since this species is expected to be well-adapted to its only known host, does this mean that the optimal oviposition strategy for A. glomeratus attacking P. rapae is also to lay a single egg? Chapter Six considers this question in more detail. In the functional response experiments (described in Chapter Four), there was no significant difference between the clutch sizes laid by British and A m erican A. glomeratus in P. rapae. Therefore, despite apparently strong selection pressure, no evidence of any adaptive change in the clutch size of American wasps was found. Moreover, the clutch size laid by British A. glomeratus in P. brassicae, the larger host species, was significantly smaller than that laid by either British or American A. glomeratus in P. rap ae. T h is finding is at odds with the results in Table 5.2. which show that, in the experiment described in Section 5.2.1., the clutch sizes laid by British wasps in P. brassicae and P. rap ae were not significantly different. The immediate 156 cause of this discrepancy appears to be that whereas the clutch size laid by B ritish A. glomeratus in P. rapae was similar in the two experiments, that laid in P. brassicae was markedly lower in the functional response experiment than in the experiment described in Section 5.2.1. A possible explanation of these observations is as follows. In the functional response experiments, the host encounter rate of wasps was much higher with P. brassicae than with P. rapae (see Chapter Four). Ikawa & Okabe (1985) found that the clutch size laid by Japanese A. glomeratus in P. rapae crucivora decreased from 40 to 20 eggs per host with successive ovipositions, if hosts were presented at a rate of one per minute. If they were presented at a much lower rate (one per 240 minutes), the clutch size did not decrease, but remained at about 40. This response to high host encounter rates has been demonstrated for other parasitoids, especially Trichogramma spp. (Hirose et a l, 1976; Pak & Oatman, 1982; Waage & Ng, 1984; Waage & Godfray, 1985; Waage, 1986), and is predicted by models that consider maximisation of parental fitness over more than one host (Parker & Courtney, 1984; Charnov & Skinner, 1984, 1985; Waage & Godfray, 1985). This could explain why, in the functional response experiments, smaller clutches were laid in P. brassicae than in P. ra p a; e and why, in the experiment described in Section 5.2.1. (where the rates of encounter with each host species were similar and relatively low), no significant difference was found. A mother should only reduce her clutch size in response to a high host encounter rate if by so doing she can increase the fitness of each egg she lays. This explanation therefore requires the existence of density-dependent offspring fitness in broods of 20-30 A. glomeratus in P. brassicae. The apparent lack of clutch size adaptation in the American population may be because there was insufficient genetic variation in the introduced stock, or because 300 generations represents ■ jtoo short a time for adaptation to have occurred. Although rapid adaptation of clutch size to novel environments has been reported for introduced birds, such as little owls' {Athene noctua Brehm ) in Britain (Lack, 1947), and goldfinches (Carduelis carduelis L.) in Australia (Frith, 1957), examples from insects appear to be lacking. 157 Host instar attacked No significant differences were found between the mean clutch sizes laid by B ritish A. glomeratus in instars I-III of P. brassicae. Sato (1980) obtained the same result for Japanese A. glomeratus parasitising P. rapae crucivora, alth ou gh he found that fewer eggs were laid in fourth instars. To establish whether there are differences in the fitness of offspring laid in different instars, it is necessary to compare their size and survivorship. This has not been done with A. glomeratus. Figure 5.14. shows that A. glomeratus takes about 60 seconds to lay 60-70 eggs in P. brassicae. If such lengthy ovipositions are disadvantageous, this could be a factor reducing the clutch size favoured by natural selection, even in the absence of density-dependent offspring fitness. Although there is no evidence that the rate of egg-laying decreases during an oviposition, other penalties to long oviposition times are possible. These could include increased vulnerability to host-inflicted damage (see Chapter Three), or an increased risk of host death from the trauma of oviposition. A correlation between oviposition time and clutch size is an assumption of several rate-maximisation models of insect oviposition (Charnov & Skinner, 1984, 1985; Skinner, 1985; Parker & Courtney, 1984), although there is little evidence, generally, that one exists'(Skinner, 1985). The clutch size laid by A. glomeratus in P. brassicae does correlate with oviposition time, but unless a sufficiently wide range of oviposition times are obtained, this correlation may be obscured by variability in the rate of egg-laying. Previous parasitism A. glomeratus is known to superparasitise under some conditions, such as when confined with hosts long enough for many re-encounters with previously- parasitised hosts to occur (Matsuzawa et a/, 1957; Matsuzawa, 1958; Ikawa & Okabe, 1985; see also Section 5.2.1.). This has been observed in other species of parasitoid (Bakker et a l, 1985; Waage, 1986), and it has been suggested that such behaviour represents retrospective adjustment of clutch size (Waage, 1986). Large brood sizes indicative of superparasitism have also been recorded from field-collected P. brassicae (Richards, 1940; Karnavar, 1983; see also Chapter Two). 158 Under the conditions of this study, however, the wasp’s first reaction to a previously-parasitised host was always to reject it. Superparasitism also appears to have been infrequent in the foraging experiments described in Chapters Three and Four: only 3% of clutches in the clutch size frequency distribution shown in Chapter Two exceed 35 eggs per host. Theoretical studies of parasitoid oviposition predict that the decision to superparasitise depends on the wasp’s previous rate of encounter with unparasitised hosts (Iwasa et a l, 1984; Charnov & Skinner, 1985). Consistent with this view, Ikawa & Suzuki (1982) found that the frequency with which A. glomeratus rejected previously-parasitised larvae of P. rapae crucivora depended on the number of unparasitised hosts presented to it beforehand. In the present study, a single exposure to an unparasitised host was sufficient to cause a standardised wasp to reject a subsequent parasitised one. Most studies of host discrimination in A panteles species have found no evidence of the rejection of previously-parasitised hosts (Table 5.13.). In this study, however, discrimination by A. glomeratus was rapid and accurate. Only two "mistakes" occurred in 60 trials (both rejections of an unparasitised host), and most rejections (22 out of 32) involved only a cursory insertion of the ovipositor (probing behaviour, lasting no more than two seconds). Some rejections (10 out of 32), however, took as long as a typical oviposition ("pseudo-ovipositions", lasting at least nine seconds in this experiment). Ikawa & Suzuki (1982) also observed rapid withdrawal of the ovipositor after insertion (termed "probing behaviour" here), and assumed that this behaviour, and only this behaviour, implied host-rejection. The present study supports the first part of their assumption, since no eggs were laid during probing behaviour, but not the second: long periods of ovipositor insertion may also result in host-rejection. It is not clear why some rejections should take .much longer- than others. Jackson (1966), in a study of the mymarid egg parasitoid Caraphractus cinctus Walker, also found that some rejection times were as long as normal ovipositions (although egg-laying during these protracted-insertions of the ovipositor was not entirely discounted). Long rejection times have consequences for the searching efficiency of the parasitoid (see Chapter T hree). 159 Ovipositor insertion occurred in all trials, indicating that it is the ovipositor that is responsible for detecting cues associated with previous parasitism. The universal rejection of previously-parasitised hosts in this study implies that oviposition in such hosts is disadvantageous. Johannson (1951) found that provided two ovipositions in a single P. brassicae larva occurred within six days of each other, the two broods of A. glomeratus would emerge simultaneously. The disadvantage of superparasitism must therefore be associated with larval competition, or with an increased risk of host mortality. This once again implies the existence of density-dependent offspring fitness among broods developing in P. brassicae. A panteles species Reference Rejection observed A. marginiventris (C resson) D m och et al (1984) A. plutellae K u rdj Lloyd (1940) A. fumiferanae V ier. Miller (1959); McLeod (1977) A. glomeratus (L.) Ikawa & Suzuki (1982); Kusano & Kitano (1974) A. melanoscelus R atz. Weseloh (1976) No rejection observed A. flavipes (C am .) CIBC (1979); Varma & Bindra (1976b) A. chilonis M ats. Varma & Bindra (1976b) A. ruficrus H al. Hafez (1947) A. tedellae N ix o n Munster-Swendsen (1979a) A. k a za k T el. _____ Carl (1976) A. lesbiae B lanch. Arce de Hamity (1978) A. angaleti M ues. N arayan an et a l (1956) A. medicaginis M ues. - Allen & Smith (1958) A. rubecula M arshall Nealis (unpublished) Table 5.13. Reported occurrences of observed rejection or lack of rejection of previously-parasitised hosts by A pan teles spp. 160 CHAPTER SIX HOST SIZE AND THE DISTRIBUTION OF SOLITARY AND GREGARIOUS BROOD SIZES IN APANTELES 6.1. INTRODUCTION A. glomeratus and A. rubecula, although closely-related and attacking closely- related host species, differ markedly in their oviposition strategy: whereas A. glomeratus is gregarious, laying 20-30 eggs per host, A. rubecula is solitary. Solitary larval development in parasitoids is characterised by the elimination of competitors: irrespective of how many eggs are laid in a host, only one larva usually emerges. Supernumerary larvae may be killed by physical combat, in which one larva attacks another with its mandibles; or by physiological suppression, involving toxins, nutritional deprivation, or anoxia (Salt, 1961; Fisher, 1961; Vinson & Iwantsch, 1980). In A. rubecula elimination of competing larvae appears to involve physical combat, and first instar larvae have large heads equipped with prominent mandibles (see Chapter Two). In contrast, newly-hatched A. glomeratus h ave small mandibles (see Figure 2.4). Elimination of supernumerary larvae has been reported for several other solitary A pan teles species (e.g. Zwolfer, 1964; Puttier & Dickerson, 1968; Weseloh, 1976; Munster-Swendsen, 1979a) and may involve physiological suppression (Muesebeck, 1918; Lloyd, 1940) as well as physical combat (Allen & Smith, 1958; Zwolfer, 1964). As discussed in Chapter Five, theoretical studies of adaptive progeny allocation predict that clutch size should increase as the intensity of density- dependent within-brood larval competition decreases (Parker & Courtney, 1984; Charnov & Skinner, 1984, 1985; Skinner, 1985; Waage & Godfray, 1985). Interspecific variation in the strength of density-dependence will depend upon differences in host size and the food requirements of individual larvae (Waage & Godfray, 1985). Among closely-related and similar-sized species of parasitoid, host size will have the* principal effect, and a general prediction of clutch size theory is that, in comparisons of such species, clutch size should NOTE: This chapter appears as a publication in Ecological Entomology, Volume 12 (1987). 161 correlate with host size. Within this theoretical framework solitariness represents one end of a clutch size continuum, with large gregarious broods at the other. Trichogramma spp., for example, exhibit a continuum of clutch sizes from one to sixty or more, depending on host size (Salt, 1940; Klomp & Teerinck, 1962; Pallewatta, 1986). In this chapter I investigate whether solitary and gregarious broods in A pan teles spp. form a continuum with host size in the way optimal clutch size models predict. 6.2. METHODS Information on the biology of 276 species of A pan teles was obtained from the literature. The following details were recorded. Adult parasitoid size The total body length of the adult wasp (excluding the ovipositor) was taken from the species description given in Viereck (1916), Muesebeck (1921), M uesebeck et a l (1951), Wilkinson (1927-1945), Nixon (1972, 1973, 1974, 1976) or Marsh (1979). If the description gave a size range the median value has been used here. H ost size Host records were taken from the lists of Muesebeck (1921), Muesebeck et al (1951), Wilkinson (1927-1945) and Nixon (1972, 1973, 1974, 1976). An index of host size was obtained for the usual host of each A pan teles species from the length of the adult fore-wing, measured from the centre of the thorax to the wing-tip. This has been used because host sizes were extracted from the Lepidoptera literature where measures of adult size are more frequently given than those of larvae. Wing length is strongly correlated with full-grown larval length in Lepidoptera (Figure 6.1.). In the case of polyphagous parasitoids whose usual host is not known, the host species of median size has been used. gr 61 rl i t e e t adul f wi nd f ll- fu d an g in -w e r fo lt u d a f o gth len een etw b n tio rrela o C 6.1 igure F Adult fore-wing length (mm) dat lce rm te ieat e. rl i fci t n ie ffic e o c n tio rrela o C re). tu litera the from llected o c ta a (d r ar lngt n net f peci of pi tera p o id ep L f o s ie c e sp r u -fo ty e in n 0.001. in < p th g len 0.856; = l a rv la n w gro 162 163 B rood size Brood size refers to the number of larvae emerging from a host (obtained from the number of cocoons spun after larval emergence) and is used as an estimate of clutch size, since clutch size itself (the number of eggs laid in a host by a single female) is infrequently recorded in endoparasitoids such as A panteles. Brood sizes were obtained from unpublished data collected by MR. Shaw (27 species); from the published literature (143 species); or (for A. glomeratus and A. rubecula) from this study (see Chapter Two and Chapter Five, Table 5.6.). If more than one brood size reference was found for a species, the one selected was that based on the largest sample size. 6.3. RESULTS T he 276 A pan teles species used in this study are listed in Appendix II, along with the information, collected from the literature, on their hosts, host sizes, brood sizes and adult parasitoid sizes. 6.3.1. Brood size frequencies 161 (58%) of the species listed in Appendix II are gregarious and 115 (42%) solitary. Figure 6.2. shows the frequency distribution of brood sizes for the 57 gregarious species whose brood size has been recorded. There is a broad peak of brood sizes from 12-26, with some much larger broods (up to 1200), but relatively few species with brood sizes in the range 2-11 larvae per host. When solitary species are taken into account there is clear evidence of bimodality in the brood size frequency distribution of A pan teles species: larvae tend either to develop alone, or in groups of twelve or more. 6.3.2. The effect of host size on brood size Figure 6.3. shows the relationship between host size and brood size in 52 gregarious A pan teles species. There is a significant regression, suggesting that the frequency distribution of gregarious brood sizes is partly determined by host size. There are, however, two important problems associated with the interpretation of Figure 6.3. Firstly, the significance of the relationship is lost Figure 6.2 Number of gregarious species 10 0 5- a — of gregarious distribution Frequency sizesof brood in fifty-seven species -tN r- H — 1 VO < — r- 72-76 iue63 Regression of size brood host on size(length Figure of adult6.3fore- Brood size 1000- * 100 10 2 5- 1 — - - - 2 ersin qain I y 15 I x 12; 5 31.3; 1.21; r5= - 1.51 x In = equation:Regression Iny ( F wing) in fifty-two species of gregarious gregarious of species fifty-two in wing) i , 5 0 ) “ 22' 8 ( P < 0 . 0 0 1 ) . j i r i i -j 1 2 50 20 10 5 165 Host Host size (mm) . s e l e t n a n A 166 if the species attacking the four largest hosts are removed from the analysis. Secondly, regression techniques assume that data points are independent. In multi-species comparisons this is often not the case because of phylogenetic relationships between species (Clutton-Brock & Harvey, 1984; Felsenstein, 1985). In applying a regression analysis to Figure 6.3. it is therefore necessary to make the assumption that clutch size, at least in gregarious species, has sufficient evolutionary lability for phylogenetic inertia to be relatively unimportant (Godfray, 1987a). Can host size account for the disproportionately large number of solitary species? A complicating factor is parasitoid size: Figure 6.4. shows a significant relationship between host size and adult wasp size in gregarious species. Thus large hosts not only support more parasitoid larvae, they also produce larger adult wasps. Assuming a relationship between adult and larval size in A p a n t e l e s , Figure 6.5. allows for this effect by showing a measure of total parasitoid volume per host (wasp length3 x brood size), plotted against host size for 40 gregarious and 64 solitary A p a n t e l e s species. The regression line for the gregarious species is shown and it is clear that, except at the smallest host sizes, solitary species do not lie on the same regression slope as the gregarious species. T w o points emerge from Figure 6.5.: 1. O n hosts of 1 0 m m adult fore-wing length or less, there are several solitary species, but few gregarious ones. Figure 6.6. confirms this observation with a larger sample size which includes all the species in Appendix II whose host size has been recorded. These hosts are not necessarily too small to support a gregarious brood: an extrapolation of the regression slope in Figure 6.3. indicates that hosts between 4 m m and 1 0 m m could theoretically sustain broods of 2-10 A p a n t e l e s larvae. 2. Solitary species are found on a wide range of host sizes and are not confined, as might be expected, to the smallest hosts. Although the majority of species on hosts larger than 1 0 m m are gregarious, substantial numbers of solitary species occur on hosts up to about 26 m m adult fore-wing length (see also Figure 6.6.). The body sizes of solitary species parasitising hosts larger than about 1 0 m m do not increase as host size increases, implying that these hosts are not fully consumed. iue64 orlto bten host size (length fore-wing)adult of Correlation between Figure 6.4 Length of adult wasp (mm) 0 n lnt o aut ap n eet-ee seis of species seventy-seven in wasp adultgregarious of length and . 01 . panteles. s e l e t n a Ap Correlationcoefficient 0.352;< p = 167 Host Host size (mm) (Length of adult wasp)3 x brood size Figure 6.5 10000 1000 100 10 1 - - - - * - - 2 species by eye. drawn Relationship between a measure of totalmeasure a Relationshipparasitoid between volume rgros pce: n = .8 n - .4 r = 45.0; = r2 1.64; - x 1.88 In = gregarious species: y In (lengthsixty- adult fore-wing) in forty of gregarious and F(i 3£) points solitary for through 31.2= Line (p0.01). < four solitaryspecies of per host ((Length of adult wasp)3 xsize) brood host and size 1 1 2 50 20 10 5 ------168 panteles. A Host Host size (mm) 1 ------Regression equation forequation Regression 1 ------r Figure 6.6 Number of A p a n t e l e s species species parasitising hosts in each of eight different host size h nme o geaiu () n solitary gregarious (b) (a) and of number The distribution gregarious right species of is the shifted to classes (host size = length of adult fore-wing). e host h T size relative to that of solitary species 2{7) X ( = 42.4; P < 0.01). Host Host size (mm) 169 s e l e t n a p A 170 This interpretation is supported by the data in Table 6.1. which show that of eleven solitary species for which information is available, only two, both parasitic upon hosts smaller that 10mm, emerge from their host’s final instar. This contrasts with ten out of eleven gregarious species. Since a caterpillar m a y consume 85% of its total food intake during the final instar (David & Gardiner, 1962), a parasitoid larva which emerges from an earlier instar considerably reduces the amount of food available to it. 171 A panteles species • Host species Host size Number of ii before host’s final instar Solitary species melanoscelus K atz. Lymantria dispar (L.) 25 3,4 vilripennis Curtis Chesias legatella (Denis & Schiff) 18 1 plutellae K urdj. Plutella xylostella (L.) 7 0 rubecula (Marshall) Pieris rapae (L.) 24 1 porihelriae Mues. Lymantria dispar (L.) 25 3,4 kazak Tel. Heiiothis armigera (Hb.) 17 3 etiellae V icreck Ancylostomia stercorea (Zell.) 11 1 circttmscripiiis N ees Lithocoiletis messaniella Zell. 5 0 hyphantriae R iley Hyphantria cunea D rury 13 2 machacralis Walk. Eutectona machaeralis Walk. 9 2 depressariae M ues. Depressaria pastinacella Staint. 11 2 Gregarious species glom eratus (L.) Pieris brassicae (L.) 34 0 congregatus (Say) Manduca sexta (L.) . 66 0,1,2 fulvipes Hal. Chesias legatella (Denis & Schiff.) is 0 ruficrus Hal. Leucania separata Walk. 22 0,1 m ililaris (W alsh) Pseudaletia unipuncla (H aw .) 21 0 sesnm iae Cam. Busseola fusca Fuller 16 0,1 popularis Hal. Tyria jacobeae L. 20 0 harrisinae Mues. Harrisina metallica (Stretch) 15 0 yakutatensis (Ash.) Autographa califomica (Speyer) 22 0 kariyae W atanabc Leucania separata Walk. 22 0,1 cuphydryadis M ues. Euphydryas phaeton D rury 21 2 Table 6.1. Host instar in which parasitoid emergence occurs, expressed as the number of instars before the host’s final instar, in those A p a n t e l e s species for which information has been collected. Host sizes refer to the length of the adult fore-wing (in mm). 172 6.4. DISCUSSION A prediction of optimal clutch size theory is that in comparisons of similar species, brood size should correlate with host size. This prediction is not supported by the pattern of progeny allocation in Apanteles. Although brood size in gregarious species correlates with host size there is a shortage of the small-brooded gregarious species expected on small hosts. Furthermore, solitary species are not confined to the smallest hosts but occur on species large enough to support broods of twenty or more gregarious larvae. A possible explanation for these observations is that factors other than host size have placed additional constraints on the evolution of clutch size in Apanteles. Possible factors affecting clutch size optimisation in Apanteles. 1. Encapsulation Salt (1968) suggested that one of the functions of gregariousness in parasitoids m a y be to overwhelm the host’s immune response. Work on A. glomeratus parasitising P. rapae crucivora has shown that up to half the eggs in artificially small clutches of fewer than ten eggs per host (created by interrupting oviposition) were encapsulated by host haemocytes, whereas in larger clutches encapsulation accounted for only 1-2% of eggs (Kitano & Nakatsuji, 1978; Ikawa & Okabe, 1985). Kitano (1986), reviewing this work, proposed that a m i n i m u m clutch size of nine eggs m a y be necessary to inhibit encapsulation. If this is true the host’s immune response imposes a restriction on the range of viable clutch sizes in A. glomeratus , and m a y in the past have constrained clutch size evolution in this species. The need to resist encapsulation could therefore account for the shortage of small clutch sizes in gregarious A p a n t e l e s species. This explanation seems unlikely, however, because it requires solitary species to possess an alternative method of suppressing the host’s immune response, and if this is the case, small-brooded gregarious species might be expected to have evolved using the same mechanism. It is also possible that the results reported by Kitano & Nakatsuji (1978) and Ikawa & Okabe (1985) are due to an experimental artefact: the interruption of oviposition could interfere with the injection into 173 the host of venom, thought to be an important encapsulation inhibitor in A. g l o m e r a t u s (Kitano, 1982; Wago & Kitano, 1985). 2. Host growth and development Parasitism frequently stunts host growth, but gregarious A p a n t e l e s are often able to ameliorate this by manipulating host development to their advantage (see Chapter Five). N o such control over host development has been reported for solitary species, and parasitised hosts are frequently much smaller than unparasitised hosts of the same age (Deshpande & Odak, 1971; Parker & Pinnell, 1973; Danks et al, 1979; Velasco, 1982). For example, by the time A. rubecula emerges from fourth instar P. r a p a e , unparasitised caterpillars which hatched at the same time have reached the fifth instar (See Chapter Two). Furthermore, solitary species, especially those parasitising large hosts, often emerge before the host is fully grown, whereas gregarious species seldom do (Table 6.1.). Consequently, the amount of potential host resource used by solitary species is less than that used by gregarious Apanteles. If the early egression and the retardation of host growth seen in solitary species is a constraint to progeny allocation, rather than a consequence of it, the host size index used in Figure 6.5. will overestimate the amount of resource available to a solitary larva. Hosts that appear superficially large m a y in effect be very small if the parasitoid is obliged (for example, by physiological constraints) to emerge from an early host instar or to stunt host growth. However, there is no evidence that this is in fact the case and an alternative viewpoint is that solitary species do not fully consume the host because they do not need to, not because they are constrained from doing so. Figure 6.5. shows that adult size of solitary A p a n t e l e s initially increases with host size, but then reaches a plateau at a body length of about 3.0mm. If further increases in size confer no advantages on an adult A p a n t e l e s , there will be no selective advantage to consuming all the host. Other factors, such as reduced-exposure time to predators and hyperparasitoids, will then become more important, and m a y favour the shorter development time associated with early emergence. Slansky (1986) discusses other possible advantages to a solitary parasitoid of suppressing host development. 174 This second interpretation seems the more likely. In A. rubecula , the instar emerged from depends on the instar attacked (Parker & Pinnell, 1973), implying an element of flexibility in parasitoid development. Allen (1958) reports a similar finding for another solitary species, A. medicaginis. Furthermore, in situations in which a Lepidoptera species is host to both a solitary and a gregarious A p a n t e l e s , physiological constraints to development in late instars might be expected to apply as much to the gregarious species as to the solitary one. This does not appear to be the case. Apanteles vitripennis Curtis, a solitary parasitoid of the geometrid Chesias legatella (Denis & Schiffermuller), emerges from the penultimate instar of its host, whilst the gregarious A. fulvipes Hal. is able to continue development into the final instar of the same species (Wall, 1975). The parasitoids of P. r a p a e provide another example: A. rubecula emerges from the half-grown caterpillar; A. glomeratus from the full-grown host. In this case it seems unlikely that physiological restrictions to development in final instar hosts should apply to the monophagous A. rubecula but not to A. glomeratus , which is normally a parasitoid of P. brassicae. 3. Parent-offspring conflict Most clutch size theory considers the evolution of progeny allocation from a parental perspective. Parent-offspring conflict over the clutch size occurs when the parental optimum differs from that of the offspring (Dawkins, 1976), and Godfray (1987b) has proposed that such conflict m a y impose constraints to adaptive progeny allocation in parasitoids. Using a genetic model he has shown that small parental optimum clutch sizes m a y under certain conditions be unstable because larvae will be selected to adopt fighting behaviour. Only one parasitoid will then emerge, irrespective of the number of eggs laid, and selection will act upon the parent to reduce its clutch size to a single egg. Whether or not fighting behaviour will evolve depends on the balance between the advantage to an aggressive larva of removing competitors, and the disadvantage through kin selection of killing sibs. In Godfray’s (1987b) model, fratricide among gregarious larvae is only selected for when the parental optimum clutch size is around 2-4 eggs, although the actual range of the unstable region depends on the nature of the clutch size fitness function used. 175 Parent-offspring conflict m a y in this way offer an explanation for the shortage of small-brooded gregarious Apanteles. It could also help to account for the occurrence of man y solitary species on large hosts. If these species are descended from solitary ancestors on smaller hosts, the evolution of a larger clutch size, in response to an increase in host size, may have been precluded by larval fighting behaviour. Godfray (1987b) shows that the conditions for larval tolerance to spread - a necessary first step in the evolution of gregariousness from a solitary lifestyle - are stringent: in the simplest model, females of solitary species must occasionally lay two eggs in a host; and when they do, the fitness of a larva in a pair must be greater than that of one which develops alone. Superparasitism, frequently reported in solitary A p a n t e l e s species (eg. Weseloh, 1976; Carl, 1976; Arce de Hamity, 1978; Munster- Swendsen, 1979a), is another force inhibiting the spread of tolerance (Godfray, 1987b). There are, however, a number of conditions that make the requirements for gregariousness to evolve less stringent. In particular, costs of fighting (risk of death; metabolic cost of mandibles) and female-biased sex ratios enable genes for larval tolerance to spread if the fitness of a larva in a pair is slightly less than that of a solitary larva (Godfray, 1987b). Assuming a negative exponential clutch size fitness function, this necessitates weak within-brood density-dependence and a correspondingly large parental optimum clutch size. Interestingly, the largest host species in Figure 6.5., which presumably induce the weakest density-dependence, support only gregarious species; and gregarious A p a n t e l e s tend to have a more female-biased sex ratio than solitary species, although this female-bias is more likely to have evolved as a consequence of gregariousness (see Chapter Seven). Clearly, an understanding of A p a n t e l e s phylogeny is required before the importance of evolutionary history can fully be tested. At present it is not known whether the solitary parasitoids of relatively large hosts in Figure 6.5. are descended from solitary ancestors, nor whether their ancestors are likely to have parasitised smaller host species. Without a detailed analysis of A p a n t e l e s phylogeny, showing the number of separate evolutionary transitions between solitariness and gregariousness, the data in this study must therefore be interpreted with caution. However, they do demonstrate that the nature of the brood size distribution in A p a n t e l e s is more complex than predicted by simple models of parasitoid progeny allocation, and they provide circumstantial 176 evidence for the models developed by Godfray (1987b). Similar evolutionary constraints to adaptive progeny allocation are likely to apply in other genera with comparable life-histories to A p anteles. A brood size distribution resembling that of A p a n t e l e s appears to occur in the Microgasterinae as a whole, in which the majority of species are solitary, but in which a substantial number of gregarious species occur (especially on large hosts), mainly with brood sizes in the range 10-40 per host (Shaw & Askew, 1976). Parent-offspring conflict is less likely to constrain clutch size adaptation in some other genera. For example, egg-parasitoids of Lepidoptera in the genus Trichogramma must completely consume the host contents before successful pupation can occur (Flanders, 1935). Consequently, a single larva in a large egg is obliged to over-eat and emerges deformed and inviable (Schieferdecker, 1969; Strand & Vinson, 1985). Other than in the smallest hosts the fitness of Trichogramma progeny is higher in a brood of two or more than when alone. Larval fighting behaviour has not evolved and female Trichogramma are free to lay the parental optimum clutch size (Waage & Ng, 1984). 177 CHAPTER SEVEN REPRODUCTIVE STRATEGIES OF SOLITARY AND GREGARIOUS APANTELES SPECIES 7.1. INTRODUCTION In Chapter Six it was shown that solitary and gregarious brood sizes in A p a n t e l e s are not part of a single host size-mediated continuum of brood sizes. Larval fighting behaviour may have constrained the evolution of parental optimum clutch sizes in solitary species, creating a strongly bimodal brood size distribution. Given this dichotomy in progeny allocation, the reproductive strategies of solitary and gregarious species might have been subjected to different selection pressures. In this chapter I consider two components of this reproductive strategy: sex ratio and egg complement. Sex ratio Fisher (1930) showed that, in a randomly-mating population, selection should lead to an equal parental investment in offspring of each sex. If one sex is in the minority, individuals of that sex will, on average, gain more matings than those of the majority sex. Parents that are genetically predisposed to over produce this rarer sex will be at a selective advantage because they will tend to leave more grandchildren. The population sex ratio will consequently approach equality and, as it does so, the advantage associated with producing an excess of the rarer sex will diminish. When the sex ratio reaches equality, parents producing a biased offspring sex ratio will be selected against: the evolutionarily stable sex ratio (assuming equal parental investment in males and females) is 1:1 (Charnov, 1982). Hamilton (1967) considered the effect of relaxing Fisher’s (1930) assumption of population-wide random mating. H e considered a breeding structure in which n females deposit their offspring on one resource patch. The progeny mate, at random, amongst themselves. The males then die (or at least do not gain matings beyond the patch) and the females disperse to colonise new patches. Because brothers compete with each other for mates, Hamilton (1967) termed this breeding structure "local mate competition" (LMC). The 178 evolutionarily stable sex ratio (proportion males, r) under these conditions is given by r = (n - 1) / 2/i If only one female colonises a resource patch (i.e. n = 1), the evolutionarily stable sex ratio is zero. In practice this means that a mother should lay only enough males to ensure that all her daughters are mated. Fitness, under these circumstances, is defined as the number of inseminated females produced. As the value of n increases, sons become more valuable to a mother because they can obtain matings with the daughters of other colonisers. The evolutionarily stable sex ratio consequently increases, rapidly approaching an asymptote at 0.50 males (which would be reached if mating became panmictic). The precise nature of the selection pressures acting on the sex ratio under L M C has been the subject of much discussion, and is reviewed in Charnov (1982) and Harvey (1985). Species of gregarious parasitic Hymenoptera have been extensively used in the study of sex ratio evolution, especially in tests of L M C theory (see Charnov (1982) and Waage (1986) for reviews). There are three main reasons wh y they have proved to be particularly appropriate for this type of work. Firstly, most species are arrhenotokous, so females have the capacity to control the sex of their offspring. Secondly, the resource patch is the host, which is discrete and easily identified. Thirdly, sibs develop together and mating often occurs before dispersal, and sometimes before emergence from the host (Suzuki & Hiehata, 1985); and the value of n (the number of females ovipositing in a host) is usually low. Gregarious A p a n t e l e s species have a breeding structure conducive to LMC. In A. glomeratus males emerge slightly before females and wait on the cocoon cluster for females to emerge (Tagawa & Kitano, 1981). Females that are about to emerge produce a sex pheromone from a gland at the base of the abdomen (Tagawa, 1977), and this diffuses through the cocoon wall, eliciting male courtship behaviour (rapid wing vibration). The males crowd around the cocoon from which the female is about to emerge, struggling to be the one closest to the cocoon opening. Mating usually occurs within a few centimetres of the cocoon cluster (Matheson, 1907; Tagawa & Kitano, 1981). By placing cocoon clusters in the field at high density, Tagawa & Kitano (1981) estimated that 60% of matings were between individuals from the same brood. Under 179 natural conditions the density of cocoon clusters would be lower, and so the amount of inbreeding would probably be higher. Where it has been looked for, pre-dispersal mating has been found in other gregarious A p a n t e l e s species, such as A. flavipes (Cam.) (Kajita & Drake, 1969); A. sesamiae Cam. (Ullyett, 1935); A. euphydryadis Mues. (Stamp, 1981); and A. koebelei Riley (White, 1973). Little is k nown about the breeding structure of solitary species. Cole (1970) found that male A. medicaginis Mues. seek out females by flying along sex pheromone odour plumes. In some species of Braconidae males are known to swarm (e.g. Southwood, 1957 and personal observation). In general, unless individuals emerge from gregarious hosts (as in the solitary scelionid egg parasitoids studied by Waage (1982a)) the breeding structure of solitary species is likely to approach panmixis. Selection should therefore favour unbiased sex ratios in solitary species, and female-biased sex ratios in gregarious species. In this chapter I test this hypothesis in two ways. Firstly by a general comparison of the sex ratios of solitary and gregarious A p a n t e l e s species recorded in the literature; and, secondly, by a more specific comparison of the sex ratios of A. glomeratus a-nd A. rubecula obtained from the field at Silwood Park in 1985. Egg complement In a series of papers, Price (1972, 1973, 1974, 1975) advanced the hypothesis that egg complement in parasitoids evolves to match the oviposition rate. Parasitoids with a high rate of oviposition (such as those attacking abundant early instar hosts) should allocate a greater proportion of their resources to eggs than those with a low oviposition rate. Wasps that are limited by the number of hosts they find should, the argument goes, direct resources from egg-production to factors which will increase the rate of host-finding. Consistent with this hypothesis, Price (1975) found a correlation between egg complement and host stage attacked in fifty-nine species of Ichneumonidae. Price reasoned that because of mortality during development, later host stages are rarer, and therefore harder to find, than young hosts. Most of the species in Price’s (1975) study were solitary. A n exception - 180 Agrothereutes tunetanus - had an exceptionally high fecundity. For a given host-encounter rate, solitary species will lay eggs at a lower rate than gregarious species and so, all else being equal, should allocate fewer resources to eggs and more to factors enhancing their searching ability. In this chapter the investment in eggs made by A. glomeratus females during development is compared with that made by the solitary species, A. rubecula. In addition, a general comparison is made of the fecundities of solitary and gregarious A p a n t e l e s species recorded in the literature. 7.2. MATERIALS AND METHODS Sex ratio Sex ratios of solitary and gregarious species of A p a n t e l e s were obtained from the literature. If more than one reference was found for a species the median value was chosen. Sex ratio data for A. glomeratus were obtained from P. brassicae larvae collected in the field at Silwood Park between July and September, 1985 (see Chapter Two). After being exposed to field parasitism for 3-4 days, first or second instar hosts were brought into the laboratory and reared through to pupation or parasitoid emergence. The remains of caterpillars from which parasitoids had freshly emerged were dissected in isotonic insect ringers solution under a binocular dissecting microscope. The number of dead unemerged second or third instar larvae left in each host was recorded. Mortality of earlier developmental stages is not detectable by this method (see Chapter Five). The emerged broods of A. glomeratus were placed in individual 5 0 m m x 2 5 m m muslin-topped glass tubes. Larvae that emerged from the host, but failed to spin cocoons, were counted. After adult emergence, the numbers of adult males and females in each brood were counted, along with the number of cocoons from which no adult emerged. P. r a p a e larvae were exposed to field parasitism and returned to the laboratory for rearing in the same way as P. brassicae. A. rubecula cocoons obtained from these hosts were transferred to 5 0 m m x 2 5 m m muslin-topped glass tubes and, after emergence, the adults were sexed to find the field sex ratio of A. rubecula. 181 Egg complement To obtain an estimate of the reproductive investment made by females of A. glomeratus and A. rubecula , the numbers and sizes of eggs stored in the ovaries of non-ovipositing females were recorded. To measure the egg complement of A. rubecula, four females at each of nine different ages were removed from culture and killed. Their abdomens were removed and dissected in isotonic insect ringers solution under a binocular dissecting microscope. The number of mature eggs dissected from each abdomen was recorded. The egg complement of A. glomeratus was determined in the same way (for full details of the method see Chapter Five). As an estimate of egg size, the lengths of ten eggs from the ovaries of each of ten females of each species were measured under a compound microscope (magnification: xlOO) fitted with an eye-piece graticule. Estimates of the fecundity of solitary and gregarious species of A p a n t e l e s were obtained from the literature. In most cases the measure of fecundity given is the total number of eggs laid per female per lifetime. If more than one reference was found for a species the median value was chosen. 7.3. RESULTS 7.3.1. Sex ratio 7.3.1.1. Sex ratios of solitary and gregarious A p a n t e l e s species Table 7.1. shows the sex ratios recorded for 15 gregarious A p a n t e l e s species, and Table 7.2. the sex ratios recorded for 33 solitary species. Table 7.3. shows the mean sex ratios calculated from Tables 7.1. a n d 7.2. The mean sex ratio (proportion males) of solitary species is significantly greater than that of gregarious species (t-test on arcsine square-root transformed data: t^ = 5.1; p < 0.001). 182 Species Sex ratio Reference flavipes (Cam.) 0.27 CIBC (1973) g l o m e r a t u s (L.) 0.29 Richards(1940) congregatus (Say) 0.50 Postley & Thurston (1974) ruficrus Hal. 0.33 Hafez (1947) spurius Wesm. 0.36 Wilkinson (1945) liparidis (Bouche) 0.41 Rao (1967) t h o m p s o n i Lyle 0.00 Vance (1931) d i a t r a e a Mues. 0.39 Davis (1944) chilonis Mats. 0.23 Kajita & Drake (1969) yakutatensis (Ash.) 0.22 Madar & Miller (1983) koebelei Riley 0.32 White (1973) bignellii Marshall 0.24 Wilkinson (1945) A p a n t e l e s sp. 2 0.25 CIBC (1969) A p a n t e l e s sp. 3 0.49 Garthwaite & Desai (1940) A p a n t e l e s sp. 5 0.13 Cock & Godfray (1985) Table 7.1. Sex ratios (proportion males) of gregarious A p a n t e l e s species recorded from the literature. 183 Species Sex ratio Reference marginiventris (Cresson) 0.50 Boling & Pitre (1970) coleophorae (Wilkn.) 0.55 Ford (1943) melanoscelus Ratz. 0.51 Weseloh (1984) d i g n u s Mues. 0.50 Platner & Oatman (1972) plutellae Kurdj. 0.36 Wilkinson (1939a) p h a l o n i a e (Wilkn.) 0.43 Ford (1943) fumiferanae Vier. 0.46 Miller (1959) r u b e c u l a (Marshall) 0.47 Richards(1940) scuttelaris Mues. 0.75 Platner & Oatman (1972) s u b a n d i n u s Blanch. 0.75 Platner & Oatman (1972) gracilariae (Wilkn.) 0.50 Ford (1943) carbonarius (Wesm.) 0.49 Ford (1943) tedellae Nixon 0.51 Munster-Swendsen (1979a) c o m e s (Wilkn.) 0.73 Ford (1943) forbesi Vier. 0.50 Puttier & Dickerson (1968) xanthostigmus (Hal.) 0.70 Zwolfer (1964) p r a e p o t e n s (Hal.) 0.47 Ford (1943) k a z a k Tel. 0.42 Carl (1976) * p r inceps (Wilkn.) 0.45 Ford (1943) starki Mason 0.50 Goyer & Schenk (1970) angaleti Mues. 0.33 Narayanan et al (1956) b o r d a g e i Giard 0.20 Notley (1948) m a r i t i m u s Wilkn. 0.52 Ford (1943) medicaginis Mues. 0.49 Allen & Smith (1958) brittanicus Wilkn. 0.51 Ford (1943) i m p e r a t o r Wilkn. 0.36 Ford (1943) t a s m a n i c a Cam. 0.50 Dumbleton (1935) belippae Rowher 0.42 Chatterji & Sarup (1961) laricellae Mason 0.50 Eidt & Sippell (1961) hyphantriae Riley 0.42 Morris (1976) victor Wilkn. 0.51 Ford (1943) machaeralis Walk. 0.50 Garthwaite & Desai (1940) circumscriptus Nees 0.55 Ford (1943) Table 7.2. Sex ratios (proportion males) of solitary A p a n t e l e s species recorded from the literature. 184 n Mean sex ratio S.E. Gregarious species 14 0.32 0.03 Solitary species 33 0.50 0.02 Table 7.3. Mean sex ratio (proportion males) of the gregarious species listed in Table 7.1. and of the solitary species listed in Table 7.2. A. thompsoni , a thelytokous species, has been excluded from the calculation of a mean sex ratio for gregarious species. 7.3.1.2. Field sex ratios of A. glomeratus and A. rub e c u l a Table 7.4. shows the sex ratios of A. glomeratus and A. rubecula recorded from host larvae collected in the field at Silwood Park in 1985. The sex ratios of the two samples are identical, but whereas that of A. glomeratus is significantly different from 1:1 ( X 2^) = 47; P < 0.001), that of A. rubecula is not (X2(1) = 2.6; p > 0 .05), due to the smaller sample size. A. glomeratus A. rubecula Number of males 736 41 Number of females 1024 57 Total adults 1760 98 Proportion male 0.418 0.418 Table 7.4. Sex ratios of adults emerging from 81 mixed broods of A. g l o m e r a t u s obtained from P. brassicae, and from 98 cocoons of A. rubecula obtained from P. rapae. Host larvae were collected from the field at Silwood Park from July to September, 1985. 7.3.1.3. Factors contributing to the female-biased sex ratio of A. glomeratus. The significant female bias in the sex ratio of A. glomeratus could be due to a biased sex ratio at oviposition, or to differential mortality of the sexes during development. 185 1. Differential mortality Smith & Shaw (1980) pointed out that in haplo-diploid species lethal recessive mutations will usually be expressed only in males, since lethal alleles very rarely occur in homozygotes (homozygosity is a necessary condition for the expression of recessive alleles in diploid females). They argued that the mutation rate per genome per generation (m) can be high enough to cause enough male mortality to significantly bias the sex ratio in favour of females. They showed that the probability of a haploid genome acquiring one or more lethal recessive alleles, either by maternal inheritance or by new mutation, is given by P where P = 1 - exp(-3m) The probability that a male does not die of a lethal recessive allele is therefore exp(-3m). Assuming that the frequency with which lethal recessive alleles occur in homozygous form is negligable, females never die from their effects, so the survival rate of haploids relative to diploids, as a result of lethal genes alone, is exp(-3m). By comparing survivorship from larval emergence to adult emergence in mixed and all-male broods of four species of gregarious Apanteles (A. chares Nixon, A. abjectus Marshall, A. zygaenarum Marshall and A. ruficrus Hal.), Smith'& Shaw (1980) estimated the ratio of male : female pupal and pre-pupal survivorship at 0.9, giving a value of m equal to 0.035. Smith & Shaw (1980) suggest that lethal recessive alleles are the most likely cause of this differential mortality. 1 Table 7.5 shows the results of performing a similar analysis on the broods of A. glomeratus obtained from the field in 1985. Of the broods for which it was possible to sex all the individuals that emerged, 51 were mixed and 12 all-male. The haploid survival rate is calculated from the number of adult males which emerged in all-male broods, expressed as a proportion of the total number of larvae in all-male broods which emerged from hosts. Assuming that this survival rate also applies to mixed broods, the number of male larvae in these broods can be deduced from the number of adult males. The number of female larvae can then be deduced, and hence the diploid survival rate. 186 Mixed broods All-male broods Number of broods 5\ 12 Numb e r of larvae 1194 244 Numb e r of adult males 424 201 Num b e r of adult females 644 Estimated haploid survival rate = 0.8238 Estimated diploid survival rate = 0.9485 Ratio of haploid : diploid survival = 0.8685 Table 7.5. The number of larvae emerging from hosts, and the numbers of male and female adults emerging from 51 mixed and 12 all-male broods of A. glomeratus obtained from field-collected P. brassicae. The calculated ratio of haploid : diploid survival is 0.8685. This gives a value of m equal to 0.047, and would generate a sex ratio of 0.465 proportion males, assuming an unbiased sex ratio at oviposition and no other sources of differential mortality. The actual sex ratio of the 51 mixed broods of A. glomeratus used in Table 7.5. w a s 0.397 proportion males, which is significantly different from an expected value of 0.465 ( X 2^) = 19.5; p < 0.001). Differential pupal and pre-pupal mortality m a y therefore explain some, but not all, of the observed female bias in A. g l o m e r a t u s . Another possible source of bias is differential survival during egg and larval development. Of the 81 mixed broods of A. glomeratus in Table 7.4., detectable mortality of second and/or third instar larvae occurred in 59, whilst 22 showed no evidence of within-brood larval mortality. Table 7.6. shows the total number of males and females emerging from broods with and without mortality. There is no significant difference between the sex ratios of the two groups (X2^) = 1-6; p > 0.05), indicating no detectable differential mortality of second and third instar larvae. 187 With mortality Without mortality Number of males 548 188 Number of females 789 235 Total adults 1337 423 Proportion male 0.410 0.444 Table 7.6 Sex ratio of 59 broods of A. glomeratus in which some larval mortality was detected, and of 22 broods in which it was not. 2. Sex ratio bias at oviposition Sex ratio theory predicts that under conditions of local mate competition sex ratios at oviposition should be female-biased, and that as the number of females ovipositing in a host increases, the sex ratio of the emerging adults should become less female-biased. The number of A. glomeratus females which oviposited in a particular host can be estimated from the size of the brood. Brood sizes over 35 probably represent superparasitism (see Chapter Two). Figure 7.1. shows the sex ratio of field-collected A. glomeratus broods as a function of brood size. There is no evidence of an increase in the proportion of males emerging as brood size increases over a range of 8 to 62 cocoons per host. Figure 7.1. reveals a large amount of variation in the sex ratios of A. glomeratus broods: from 0.00 to 0.92 proportion males. A heterogeneity chi- square test on the data is highly significant ( = 248; p < 0.001), indicating significant inter-brood variation in sex ratio. 7.3.2. Egg complement 7.3.2.1. Comparison of the egg complements of A. g l o m e r a t u s and A. rubecula Figure 7.2. shows the mean egg complement of female A. rubecula as a function of female age. The number of mature eggs increases with female age to a peak of about 120 eggs 3-4 days after emergence. The egg load then decreases, presumably because eggs are resorbed in old non-ovipositing females (Flanders, 1942). Comparison with Figure 5.7. shows that the egg complement of Figure 7.1 Sex ratio Sex (proportionmales) of Figure 7.1 Sex ratio (number of cocoons).(number All-male broods omitted.been have from field-collected from . e a c i s s a r b P. Brood sizeBrood 188 . s u t a r e m o l g A. ,plotted size against brood broods reared broods Figure 7.2 Egg complement of non-ovipositing complement Egg Figure 7.2 Number of eggs per female 100 150 50- -, Means Means f eae g (ubr f as ic aut emergence). since adult days of (number age female of 1 3 7 2 4 20 14 12 7 4 3 2 1 0 i l i l i l i “ i l l l i i S.E.) + ( of four females at each age. Female age Female (days) 189 .rubecula l u c e b u r A. as a function 190 A. rubecula is much smaller than that of A. glomeratus. Newly-emerged A. rubecula have about 70 eggs compared with about 800 in British A. g l o m e r a t u s of the same age (American A. glomeratus have smaller egg loads). Specific comparisons of egg complement are complicated by differences in adult size. Egg load in parasitoids has often been shown to increase with female size (see Chapter Five), and in newly-emerged A. glomeratus the relationship between egg load (y) and female head width (x) can be described by the equation y = 4337* - 2225 (see Figure 5.4.). The mean head width of female A. r u b e c u l a was found to be 0.830mm (range = 0.804 - 0.852mm) (see Chapter Two). If egg load in A. rubecula depends on female size in the same way as that of A. glomeratust newly-emerged females should have an average complement of about 1375 mature eggs. To compare the total investment in eggs made by females of the two species during development, a measurement of investment per egg is required. As a crude estimate of this, Table 7.7. shows the mean length of eggs stored in the ovaries of A. glomeratus and A. rubecula. The eggs of A. rubecula are about 2.15x longer than those of A. glomeratus giving a volume ratio (assuming similar shapes) of approximately 1:10. To make a total investment in eggs comparable with that of A. glomeratus , A. rubecula females should produce one tenth as m a n y eggs. Correcting for body size they should therefore mature about 140 eggs during development (i.e. 0.1 x 1375). This expected value is of the same order of magnitude as the observed egg complement of about 70 eggs. n Mean egg length (mm) 99% C.I. A. g l o m e r a t u s - __ 100 0.13 0.003 A. rubecula 100 0.28 0.002 Table 7.7. Mean lengths of eggs dissected from the ovaries of British A. glomeratus and A. rubecula. 191 Fecundity Reference Gregarious species A. flavipes (Cam.) 133 Moutia & Courtois (1952) A. ruficrus Hal. 216 Hafez (1947) A. sesamiae Cam. 76 Varma et al (1979) A. s p urius Wesm. 92 Shapiro (1960) A. liparidis (Bouche) 130 Shapiro (1956) A. harrisinae Mues. >500 Clausen (1978) A. diatraea Mues. 95 Varma et al (1979) A. obliquae Walk. 82 Lall (1958) A. thompsoni Lyle 230 * Vance (1931) A. chilonis Mats. 115 * Kajita & Drake (1969) A. glomeratus (L.) 1300 Moiseeva (1960) Solitary species A. marginiventris (Cresson) 82 Kunnulaca & Mueller (1979) A. melanoscelus Ratz. 516 Reeks & Smith (1956) A. d i g n u s Mues. 152 Cardona & Oatman (1971) A. plutellae Kurdj. 80 Velasco (1982) A. fumiferanae Vier. 102 Miller (1959) A. scutellaris Mues. 156 Platner & Oatman (1972) A. subandinus Blanch. 300 Cardona & Oatman (1975) A. angaleti Mues. 91 Subba Rao & Gopinath (1961) A. medicaginis Mues. 87 Allen & Smith (1958) A. hyphantriae Riley 41 Morris (1976) Table 7.8. Measures of the fecundity of solitary and gregarious A p a n t e l e s species recorded in the literature. The values refer to the number of eggs laid per lifetime, apart from those marked * which are based on dissection of adult females. 192 13.2.2. Comparison of the fecundities of solitary and gregarious A p a n t e l e s species. Table 7.8 shows measurements of the fecundity of eleven gregarious and ten solitary A p a n t e l e s species recorded in the literature. In comparison with the other species listed in Table 7.8., A. glomeratus has an exceptionally high fecundity. Apart from this species the reported fecundities of solitary and gregarious species are similar. 7.4. DISCUSSION Sex ratio Consistent with the predictions of Hamilton’s (1967) theory of local mate competition, gregarious species of A p a n t e l e s tend to have a more female-biased sex ratio than solitary species. Shaw & Smith (unpublished) recently obtained a similar result from a study of broods of solitary and gregarious A p a n t e l e s species collected in the field. They found that solitary species showed a slight female bias which they attributed to the expression of lethal recessive alleles in males. They interpreted the extra female bias of gregarious species as an adaptive response to LMC. Evidence of differential mortality between larval and adult emergence was found in broods of A. glomeratus. The sex ratio generated by this differential mortality alone - 0.465 proportion males - is less female-biased than the observed value of 0.397 proportion males. Similarly, Smith & S h a w ’s (1980) estimate of haploid r diploid survival, based on the combined data of four species, generated a predicted sex ratio (0.474 proportion males) less female- biased than that observed (0.390 proportion males). To demonstrate that this extra bias is, as Shaw & Smith (unpublished) suggest, due to local mate competition, it is necessary to determine the primary sex ratio. This involves either accounting for all possible sources of differential mortality during development, or directly measuring the sex ratio at oviposition. The first of these alternatives is difficult to apply to A. glomeratus because mortality of eggs and first instar larvae cannot be detected by dissection of hosts after parasitoid emergence. The second 193 alternative is also problematic because the eggs of A. g l o m e r a t u s cannot readily be sexed. Whereas the eggs laid by some species of parasitoid can be sexed by observing external signs of fertilisation (Cole, 1981; Suzuki et al, 1984; Pallewatta, 1986; van Dijken & Waage, 1987), no such signs are visible during oviposition by A. glomeratus. The potential exists to sex the eggs of haplo- diploid species by counting chromosomes. A n attempt to do this with A. g l o m e r a t u s eggs and larvae was made using the chromosome stain proprionic orcein. Although chromosomes could be detected, considerable refinement of the technique is required before they can reliably be counted, and before entire clutches can be sexed. Even if a female-biased primary sex ratio can be demonstrated, there ma y be hypotheses alternative to L M C to account for it. For example, sexual asymmetries in larval competitiveness m a y select for a female-biased sex ratio in the absence of L M C (Waage & Godfray, 1985; Waage, 1986; Godfray, 1986). Figure 7.1. shows that the sex ratios of A. glomeratus broods from field- collected hosts are very variable, and that they show no tendency to change as brood size changes. This has also been noted by Ikawa & Okabe (1985) for A. glomeratus parasitising P. rapae crucivora in the laboratory, and by S.C. Littlewood (pers. comm.) for A. glomeratus obtained from field-collected P. brassicae. L M C theory would predict an increase in the proportion of males in a brood as brood size increases, if large broods imply oviposition by two or more females. Suzuki et al (1984) and Ikawa & Okabe (1985) attribute the variability of sex ratio in A. glomeratus to the rapid rate of oviposition. They argue that this precludes the fine control of egg fertilisation required to produce a more precise sex ratio. This implies that the observed variation represents random fluctuation about a mean, but the significance of the heterogeneity chi-square test applied to the data in Figure 7.1. suggests that this is not the case. Rather, wasps may vary the sex ratio they produce from host to host. Alternatively, or additionally, different individuals ma y lay different sex ratios. Orzack (1986) and Orzack & Parker (1986) found that different genotypes of the gregarious pteromalid Nasonia vitripennis (Walk.) laid significantly different sex ratios, and Shaw & Smith (unpublished) found significant between-brood variation in the sex ratios of fifteen out of twenty-four gregarious A p a n t e l e s species, including A. glomeratus. 194 To compare the sex ratios of A. glomeratus and A. rubecula , a larger sample of A. rubecula is required. From the results obtained in this study for A. glomeratus , and by Smith & Shaw (1980) for four other gregarious species of A p a n t e l e s , A. rubecula should show a slight female bias (about 0.47 proportion males) due to the expression of lethal recessive alleles between larval and adult emergence. Egg complement A comparison of the fecundities of solitary and gregarious species of A p a n t e l e s provides no evidence in support of the hypothesis that solitary species, because of an intrinsically lower oviposition rate, should invest a lower proportion of their resources in eggs than should gregarious species. Although A. rubecula has a much smaller egg complement than A. glomeratus , its total reproductive investment is comparable because each egg is larger. W h y does A. rubecula produce a few large eggs and A. glomeratus many small ones? The eggs of A. glomeratus contain little yolk (Tawfik, 1975), and most of the nutrients required for embryonic development are absorbed from the host’s haemolymph (King et al, 1969) across the egg’s hydropic chorion (Flanders, 1942). Between oviposition and hatching the eggs increase in volume by about ninety times (Sato, 1980; see also Figure 5.18.) as a consequence of this nutrient absorption (King et al, 1969). The larger size of A. rubecula eggs is presumably a consequence of more parentally-derived nutrients being placed in each egg before oviposition. Development after oviposition is more rapid (at 2 0 °C hatching occurs 2-3 days after oviposition, compared with 4-5 days for A. glomeratus ) and involves less egg expansion (compare Figure 7.3. with Figure 5.18.). The newly-hatched larva of A. rubecula is also larger than that of A. glomeratus (see Chapter Two). Rapid embryonic development and large larval size m a y be advantageous for combative larvae. Older larvae are often reported to be at a competitive advantage in contests with conspecifics (Fisher, 1961; Weseloh, 1976; Munster- Swendsen, 1979a; van Alphen & Nell, 1982; Bakker et al, 1985) or with larvae of other species (Fisher, 1961; Force & Messenger, 1968; Wylie, 1972). Large larvae may also be more effective than smaller ones. In non-combative species, such as A. glomeratus^ there may be less of an advantage to rapid embryonic development or large first instar larvae, and so selection m a y favour the 195 production of man y small eggs. 196 Figure 7.3 'Appearance of eggs laid by A. rubecula in first instar P. r a p a e when dissected from the host a) immediately after oviposition; b) one day after oviposition; and c) two days after oviposition. After two days, the fully-formed larva is visible in the egg, and hatching occurs between two and three days after oviposition. 198 CHAPTER EIGHT GENERAL DISCUSSION A central tenet of evolutionary biology is that adaptation is a product of natural selection. Within constraints, the behaviour and design features of organisms are assumed to evolve so that their contribution to the organism’s reproductive success is maximised. Natural selection is believed to be an optimising process. Experimental investigations of optimality models test not the assumption that adaptation has occurred, but the specific constraints and selection pressures hypothesised in the model (Maynard Smith, 1978). This approach has been criticised by Lewontin (1978) and by Gould & Lewontin (1979) on the grounds that the omnipotence of natural selection is rendered untestable, and that insufficient importance is attached to non-adaptive differences between organisms. It is, however, difficult to disprove the hypothesis of adaptation for any particular trait, since its falsification depends on experiments which fail to find an adaptive function. In such cases there is always the possibility that subsequent experiments will detect a previously overlooked functional explanation (Clutton-Brock & Harvey, 1979). Nevertheless, adaptationist arguments should avoid the uncritical use of functional explanations (Williams, 1966; Maynard Smith, 1978). Otherwise, as Gould & Lewontin (1979) point out, these arguments can degenerate into no more than a test of the observer’s ingenuity in conjuring up plausible stories. For example, observations made to test a functional hypothesis frequently fail to match with prediction. The hypothesis may then be adjusted to account for these observations. It would not then be valid to claim support for the revised hypothesis from the original data: further tests are necessary. It is important to have an adequate methodology for testing optimality models (Williams, 1966; Maynard Smith, 1978; Clutton-Brock & Harvey, 1979). T w o methods are commonly employed: quantitative tests can be made of specific predictions; or comparisons can be made of the trait in different species, 199 populations or individuals (Maynard Smith, 1978). A n example of the first approach is the analysis of optimal clutch size (see Chapter Five). The clutch size which maximises parental fitness can be found experimentally, and then compared with the actual clutch size laid. A problem with this approach is that no model can account for all the constraints operating on the system. In an analysis of optimal clutch size, parental fitness m a y be affected by factors other than those considered by the investigator. There may also be components of within-brood density-dependence additional to those measured in the experiment. Consequently, the model’s predictions are unlikely to give a close quantitative fit with observation, and it is then up to the experimenter’s intuition to decide whether the fit is good enough to justify continued faith in the model (Maynard Smith, 1978). For example, Parker & Stuart (1976) developed a fitness gain rate model - similar to those used in the study of optimal patch departure time (Charnov, 1976) or optimal clutch size (Charnov & Skinner, 1984, 1985) - to predict optimal copula duration in the dungfly, Scatophaga stercorea L. The model gave a qualitatively close, but quantitatively inexact, fit with observation. In this case, the authors decided that the fit was sufficiently convincing to support their hypothesis, and invoked a cost of sperm production to account for the discrepancy (Parker, 1978). It is often differences between species which .provoke investigation in the first place (Clutton-Brock & Harvey, 1984), and most functional hypotheses generate predictions about the evolution of traits in different populations or species (Maynard Smith, 1978). Comparative studies are therefore, a useful means of investigating adaptation, and because they tend not to rely on a close fit with quantitative predictions, they can often provide more convincing evidence than quantitative tests. For example, models of optimal progeny allocation predict that clutch sizes should be larger in species experiencing weaker within-brood density-dependence. Waage & Godfray (1985) tested a simple quantitative model which assumed that parental fitness is maximised per clutch, and that the clutch size fitness function can be described by the negative exponential expression, f(c) = exp (-ATc), where c is clutch size, and K is the strength of density-dependence (see Chapter Five). A prediction of this model is that clutch size should be equal to the inverse of K. This prediction was tested for six host-parasitoid combinations (Waage & Godfray, 1985). Not surprisingly, this unrealistically simple model failed in each case to predict the -7 200 observed clutch size. Strictly speaking, the model was falsified (Maynard Smith, 1978). However, a comparison of the six host-parasitoid combinations reveals a clear negative relationship between the strength of density- dependence (K ) and clutch size, consistent with the model’s qualitative prediction (larger clutches where density-dependence is weaker). Clutton-Brock & Harvey (1984) distinguish between comparisons involving a few species and those which include many species. The advantage of comparing only a few species (or populations of a single species) is that it is possible to conduct a detailed experimental study. In Chapter Four, for example, the searching efficiencies of A. glomeratus and A. rubecula are compared. Empirical estimates of searching efficiency are critically dependent on the experimental conditions, so such a comparison is possible only if detailed experiments are conducted under identical conditions. It would be impracticable to extend the comparison to more than a few species. A disadvantage of this type of comparison is that it is often impossible to tell which of an array of ecological differences between the species is responsible for differences in the trait under study (Clutton-Brock & Harvey, 1979). For example, a comparison of two closely-related parasitoid species, which attack two different hosts, one larger than the other, might reveal that the parasitoid of the larger host laid a larger clutch size than the species parasitising the smaller host. A n example of this is A. glomeratus on P. brassicae compared with A. rubecula on P. rapae. Such a result would be consistent with the hypothesis that host size is an important determinant of clutch size in parasitoids. However, factors other than host size could be responsible for the difference in clutch size. For example, the parasitoid of the larger host might require a larger clutch size to suppress a host immune response more hostile than that possessed by the smaller host. The inclusion of more species in the analysis will show if there is any generality in the observed relationship, and will enable the effects of confounding variables to be removed (Clutton-Brock & Harvey, 1979, 1984). Multi-species comparisons have been used to generate and test a wide range of functional hypotheses (see Clutton-Brock & Harvey (1979, 1984) and Felsenstein (1985) for recent reviews). However, they are not without problems. Some of these are discussed by Clutton-Brock & Harvey (1979, 1984). A n important difficulty is that values for individual species are 201 usually taken from the literature, with the consequence that the quality of the data varies between species. Little confidence should be attached to the values of individual data points, and when one or two points exert a strong influence on the overall relationship, results should be interpreted with caution. In Chapter Six, for example, the statistical significance of the relationship between host size and brood size in A p a n t e l e s spp. (Figure 6.3.) is lost if the values for the four largest hosts are removed. Statistical analyses of multi-species comparisons assume that data points are independent. However, phylogenetic relationships between species will usually violate this assumption. This problem is discussed by Clutton-Brock & Harvey (1984) and Felsenstein (1985) (see also Chapter Six). A problem with all comparative studies is that the evidence they provide, based as it is on correlation, is necessarily circumstantial. A correlation between two variables, such as host size and brood size, gives no indication of the direction of causality, nor whether a causal relationship exists at all. This can frustrate attempts to test optimality models, since several hypotheses can- predict the same relationship (Waage, 1986). In Chapter Seven, for example, it was shown that the sex ratios of gregarious A p a n t e l e s species are, on average, significantly more female-biased than those of solitary species. A causal effect of oviposition strategy on sex ratio is predicted by at least two adaptive hypotheses: Hamilton’s (1967) local mate competition hypothesis, and Godfray’s (1986) sexual asymmetries hypothesis (see also Waage (1986)). A non-adaptive explanation is also possible: the primary sex ratio could be unbiased in both solitary and gregarious species, but competition within gregarious broods could cause differential mortality, creating a female-biased secondary sex ratio. Finally, causality could operate in the opposite direction. The possibility that a female-biased sex ratio could make a species more likely to evolve gregariousness was discussed in Chapter Six (see Godfray, 1987b). Despite these problems, the comparative method is a potentially powerful means of investigating adaptation. Although experimental studies tend to be more successful at identifying causal relationships, confounding variables are often as difficult to discount in experiments as in comparisons, and in many areas of investigation comparative studies provide the strongest evidence. For example, Waage’s (1982) comparative analysis of sex ratios in solitary scelionid egg parasitoids is probably the study most frequently cited as evidence in 202 support of L M C theory. The aim of this thesis has been to apply the comparative method to the study of how behavioural traits affecting the reproductive success of female parasitoids are optimised by natural selection. The lifetime reproductive success of a female parasitoid is determined by the product of two components: the number of hosts encountered, and the fitness gained from each host. Fitness per host can conveniently be defined as the number of potential grandchildren obtained from each host, since this incorporates both the number and fitness of surviving offspring. A n important factor affecting fitness per host is clutch size. The study of clutch size evolution is based on the use of optimisation techniques, and the simplest theoretical models assume that natural selection maximises parental fitness per clutch (after Lack, 1947). These simple models have so far found little quantitative support from empirical studies: observed parasitoid clutches tend to be smaller than predicted (Charnov & Skinner, 1984; Waage & Godfray, 1985; Takagi, 1985). There are at least three possible reasons for this consistent deviation of observation from prediction. Firstly, there could be faults in the design of the experiments. It is important that studies aimed at predicting the optimum clutch size of a parasitoid should consider as many components of offspring fitness as possible. In Chapter Five, it was shown how incorporating additional elements of offspring fitness progressively decreased the predicted optimum for A. g l o m e r a t u s ovipositing in P. rapae. Moreover, the experiments described in Chapter Five found no evidence of a relationship between clutch size and the adult size, juvenile survivorship or development time of A. glomeratus developing in P. brassicae , over a range of clutch sizes in excess of that normally laid. This suggests that important elements of density-dependent fitness remain undetected. Possible sources of these are discussed in Chapter Five. 203 Secondly, there m a y be faults in the theory. The premise of the model - that natural selection maximises fitness per clutch - is likely to be true only if hosts are severely limiting. More realistic models predict that as the rate of host encounter increases, parasitoids should reduce clutch size to increase fitness per egg (Charnov & Skinner, 1984, 1985; Parker & Courtney, 1984; Skinner, 1985; Waage & Godfray, 1985). This prediction has not been tested quantitatively, but qualitative agreement has been found. In the experiments described in Chapter Four, A. glomeratus laid a lower mean clutch size in P. brassicae than in P. r a p a e (see Chapter Five). It was suggested that this difference could have been due to a difference in host encounter rate. Further experiments could show whether or not this is so. Finally, there ma y be constraints on the parent’s ability to lay larger clutches. In Chapter Six, it was shown that solitary species of A p a n t e l e s exploit hosts large enough to support gregarious broods. They do not fully consume the available resource, and appear not to lay an optimal clutch size. The possibility that this is due to constraints imposed by parent-offspring conflict was discussed. Fitness per host depends also on sex ratio, because the expectation of grandchildren through sons and daughters m a y not be equal (see Chapter Seven). The optimisation of clutch size and sex ratio have usually been analysed separately, but are not in fact independent, because changes in sex ratio ma y affect the optimum clutch size (Waage, 1986). Waage & N g (1984) incorporated both clutch size and sex ratio in a simulation model of the optimal oviposition strategy of the facultatively gregarious egg parasitoid, Trichogramma evanescens (Westw.), and Godfray (1986) constructed general analytical models seeking simultaneously to optimise clutch size and sex ratio. This type of approach is important when dealing with small-brooded species such as Trichogramma , but separate analysis of clutch size and sex ratio is adequate for large-brooded species such as A. glomeratus (Werren, 1984; Godfray, 1986). In interspecific comparisons there m a y also be an effect of clutch size on sex ratio. Local mate competition or sexual asymmetries could select for a female-biased sex ratio in gregarious species, whilst selection favours an equal sex ratio in solitary species (see Chapter Seven). 204 A third factor affecting fitness per host is host quality. This will be affected by the size, age and species of host, and by whether or not the host has previously been parasitised. In Chapter Five, it was shown that for A. glo'rritratus, P. brassicae is a higher quality host than P. rapae. Wasps developing in P. brassicae had higher juvenile survivorship, larger adult size, and were more fecund than those developing in P. rapae. Differences in host quality affect the optimum clutch size. In Chapter Five, comparison of the strengths of density-dependent offspring fitness predicted a lower optimum clutch size for American A. glomeratus ovipositing in P. r a p a e than for British wasps attacking P. brassicae. The most important aspect of host quality is probably host size, and in Chapter Six it was shown that there is a positive correlation between host size and brood size in gregarious A p a n t e l e s species. Host quality m a y also influence the optimum sex ratio. Charnov et al (1981) suggest that female offspring suffer relatively more than males w h e n developing in poor quality hosts, and that mothers should, as a consequence, lay proportionally more males in hosts of low quality. A prediction of this hypothesis is that wasps which encounter various host species, which differ in quality, should produce different progeny sex ratios in each. Thus if A. g l o m e r a t u s could distinguish between P. brassicae and P. r a p a e , it should lay a more female-biased sex ratio in P. brassicae than in P. rapae.. As discussed in Chapter Five, however, host size at oviposition gives little information about the ultimate quantity of resource available, and A. glomeratus does not discriminate between the two species with respect to clutch size. It is therefore unlikely that there will be a difference in sex allocation. Only seven broods of British A. g l o m e r a t u s were obtained from field-collected P. r a p a e (see Chapter Two), yielding fifty-eight females and forty males: a sex ratio (0.408 proportion males) not significantly different from the 0.418 proportion males (Chapter Seven) obtained from P. brassicae. The only reference in the literature to the sex ratio of A. glomeratus from two sympatric host species is in Richards (1940), who recorded 0.45 proportion males from P. brassicae , and 0.29 proportion males from P. rapae. The difference is significant, and was consistent over two generations, but is the reverse of that predicted by Charnov et a V s (1981) host size hypothesis. 205 When a host has already been parasitised once, the most adaptive clutch size and sex ratio of the superparasite depends on the sex and progeny allocation made by the first female (Holmes, 1970, 1972; Werren, 1980, 1983; Suzuki & Iwasa, 1980; Parker & Courtney, 1984). The sex ratio of the second female is expected to be less female-biased than that of the first. This is a prediction of L M C theory, and of the host size hypothesis - a previously-parasitised host is effectively a smaller host. Werren (1984) modelled the combined effects of host size and L M C on the sex ratio. These models predict that the progeny sex ratio of A. glomeratus should increase with brood size - if increasing brood size implies increasing numbers of foundresses. N o evidence in support of this prediction was found in Chapter Seven. Previously parasitised hosts - or hosts low in quality for any other reason - may be rejected altogether. In Chapter Five, it was shown that A. glomeratus invariably rejected previously-parasitised hosts w h e n presented with alternately unparasitised and parasitised P. brassicae. This can be interpreted as a clutch size decision, since a rejected host is effectively allocated a clutch size of zero (Waage, 1986). It can also be thought of in terms of optimal foraging. In one of the seminal papers on optimal foraging, MacArthur & Pianka (1966) distinguished between two components of foraging: selecting which prey (or host) patches to search and, within patches, which prey (or host) individuals to accept. — The problem of how foragers should make the latter decision has been analysed with the use of optimal diet models (see Pyke et al (1977) for a review). These predict that low-quality prey items should be added to the diet when the rate of encounter with high-quality prey decreases below a threshold (Krebs & McCleery, 1984). Charnov & Skinner (1984, 1985) extended this approach to parasitoids encountering parasitised and unparasitised hosts, and Iwasa et al (1984) presented.a general model of host choice when host quality varies. Optimal diet models assume that the quantity maximised by natural selection is the rate of gain of fitness. When high quality prey are abundant, foragers should ignore low quality prey to avoid wasting time. Applying this concept to parasitoids encountering parasitised and unparasitised hosts assumes that it takes less time to reject a host than to oviposit in one. In ma n y cases this will be true. The results in Chapters Three and Five show that most rejections of previously-parasitised P. brassicae by A. glomeratus involve only a cursory insertion of the ovipositor, lasting two seconds or less ("probing behaviour"). 206 However, in Chapter Five it was shown that a substantial number of rejections take as long as a normal oviposltion. In such cases it is more appropriate to view host-rejection as an egg-saving (clutch size) decision than as a time saving (foraging) decision. The other principal question tackled by optimal foraging theory - ho w animals should allocate time to different patches - has also been treated as a gain rate maximisation problem (Charnov, 1976; Cook & Hubbard, 1977; Comins & Hassell, 1979; Lessells, 1985). It is central to the second component of a female parasitoid’s reproductive success: the number of hosts encountered. The rate of host encounter is affected by a number of factors, including where the parasitoid searches; how long it stays in each patch; how quickly it moves from host to host and from patch to patch; its reactive distance; its handling time; and the speed with which it rejects unsuitable hosts. Optimality models have concentrated mostly on the problem of patch time allocation (see Chapter Three). The factors affecting host encounter rate collectively determine the searching efficiency of the parasitoid, and in this respect have been studied by population ecologists, interested in the dynamics of the parasitoid-host interaction, and by biological control practitioners wishing to assess the efficacy of parasitoid species as natural enemies. A widely-held view is that parasitoids are capable of regulating host populations by imposing density-dependent parasitism (Murdoch & Oaten, 1975; Hassell, 1978, 1985; Hassell & Waage, 1984). A n important factor determining whether parasitism is spatially density-dependent is the behavioural response of wasps to host density. Aggregation of parasitoids in areas of high host density is an important factor promoting density-* dependence (Hassell & May, 1973; Murdoch & Oaten, 1975; M a y & Hassell, 1981). The allocation T>y searching parasitoids of more time to high host density patches, leading to aggregation, is predicted by optimal foraging models, and has been observed in a number of empirical studies (e.gv Hassell, 1971, 1978, 1982; Munster-Swendsen, 1979b; Waage, 1983). The functional response of a parasitoid (see Chapter Four) places an upper limit on the number of hosts parasitised within a patch in a given period of time, and all else being equal, tends to produce inversely density-dependent patterns of parasitism (Murdoch & Oaten, 1975; Oaten & Murdoch, 1975; 207 Hassell, 1978). The balance between the opposing effects of aggregation and the m a x i m u m attack rate per patch determines the spatial distribution of parasitism (Huffaker et aly 1968; Hassell et al, 1985; Hassell, 1986). In Chapter Three, it was shown that the rate of encounter of A. glomeratus with P. brassicae decreases with time on the patch because increasing amounts of time are spent in non-searching activities, such as handling previously-parasitised hosts and cleaning. This effect is stronger on high density host patches. Consequently, for a given period of time, a lower proportion of the available hosts are parasitised on high density patches than on low density patches (see also Chapter Four, Figure 4.1.). In the laboratory experiments described in Chapter Three, there was only a slight tendency for wasps to stay longer on larger host patches. Parasitism was therefore inversely density-dependent: the effect of a m a x i m u m attack rate per patch overrided the aggregative response. In the field, however, Hubbard (1972, 1977) found that aggregation of A. g l o m e r a t u s to areas of high P. brassicae density was in some years sufficient to overcome the effects of a Type II functional response, and to generate spatially density-dependent parasitism. Morrison & Strong (1980, 1981) and Morrison et al (1980) give examples of field data in which the probability of a parasitoid visiting a host patch increases with the size of the patch, but in which the conditional probability of any one host being attacked, given that the patch has been found, decreases with increasing patch size. The resulting distribution of parasitism was inversely density-dependent. Waage (1983) found that aggregation of the ichneumonid Diadegma eucerophaga Horstmann led to density-independent parasitism of Plutella xylostella (L.). Lessells (1985) reviewed forty-eight studies of the spatial distribution of parasitism, and found approximately equal proportions reporting density-dependent, inversely density-dependent and density-independent parasitism. Density-dependent parasitism is often stated to be a desirable feature of classical biological control programmes (Huffaker et al, 1968; DeBach, 1974; Murdoch, 1975; Huffaker & Messenger, 1976; Hassell, 1978; Waage & Hassell, 1982; Greathead, 1986). . A potentially useful biological control agent is therefore thought to be one which aggregates to areas of high host density, and which has a high m a x i m u m within-patch attack rate. A high m a x i m u m attack rate is more likely in species with a short handling time (Holling, 1959; 208 Hassell, 1978) and a relatively high fecundity (Waage & Hassell, 1982; Hassell, 1982). However, Murdoch et al (1984, 1985) and Reeve & Murdoch (1985) point out that spatially density-dependent parasitism is neither necessary for, nor characteristic of, many successful examples of biological control. Other features of natural enemies ma y be more important. A large clutch size, with low developmental mortality, and a female-biased sex ratio increases the recruitment of female parasitoids, and tends to lower the host equilibrium (Waage & Hassell, 1982; Hassell, 1986). A n y density-dependent changes in these features will also contribute to the stability of the host-parasitoid interaction (Hassell & Waage, 1984). Density-dependent changes in sex ratio m a y be a particularly important stabilising phenomenon (Hassell et al, 1983). Other attributes of a parasitoid will also have implications for the effectiveness of the species as a biological control agent. Female wasps should be responsive to host kairomones, for example. They should also be able to discriminate between parasitised and unparasitised hosts (van Lenteren, 1981). Their eggs and larvae must be capable of resisting encapsulation. In general, a biological control agent should be well adapted to its host (Waage & Hassell, 1982). In particular, it should be adapted to the distribution and density of the host, since this affects the optimisation of foraging behaviour (see Chapters Three and Four), clutch size (Chapter Five), sex ratio (Waage, 1982a,b) and fecundity (Chapter Seven). Indeed, it is only if all else is equal that a large clutch size and a high fecundity are necessarily desirable. In practice, all else is not equal. A large clutch size often means lower offspring fitness (see Chapter Five). High fecundity m a y be achieved only nt the expense of searching ability (Price, 1975). Consequently, the most suitable candidates for biological control agents are likely to be those that optimise their clutch size, sex ratio, fecundity and foraging behaviour to suit the target host species (Waage & Hassell, 1982). A p a n t e l e s species have been widely used* in biological control programmes. Clausen (1978) lists thirty-one species which had been introduced up to 1968. According to Greathead (1986), sixteen species have successfully established as biological control agents, and of these, fourteen give effective control. A. g l o m e r a t u s has a long history of use as a biological control agent of P. rapae. The first introduction was from Britain to the U S A in 1883 (Laing & Levin, 209 1982). It became widely established, and m ay exert effective control in some local areas (Oatman, 1966; Sutherland, 1966; R.G. van Driesche, pers. comm.). Generally, however, economic control is not achieved (Reid & Cuthbert, 1960; Oatman, 1966; Parker et «/, 1971; Chamberlin & Kok, 1986), and pesticide applications are still required (van den Bosch et al, 1983). P. r a p a e remains one of the most important cabbage pests in the United States (van den Bosch et alt 1983; Chamberlin & Kok, 1986). P. r a p a e was accidentally imported into N e w Zealand in about 1930 (Muggeridge, 1943a), and A. glomeratus was subsequently dispatched from Britain in 1931-32 (Muggeridge, 1933). However, establishment did not occur until a second introduction was made, in 1938-9, of material collected in America (Muggeridge, 1943b). This led Muggeridge (1943c) to suggest that the American population of A. glomeratus m a y have been pre-adapted to P. r a p a e by virtue of its prior association with this host. P. r a p a e reached Australia in 1939 (Muggeridge, 1943a). A. glomeratus was introduced from Canada in 1942 and from Britain in 1943. It rapidly became established, but, as in North America, does not provide effective economic control (Hassan, 1976). With respect to clutch size and searching efficiency, this study found no evidence in support of the suggestion made by Muggeridge (1943c), and in a different context by Courtney (1986), that the American population of A. glomeratus should be better adapted'to P. r a p a e than the British population. In Chapter Five it was shown that, in comparison with British A. glomeratus developing in P. brassicaet American wasps in P. r a p a e suffer higher, density- dependent, juvenile mortality, and emerge as smaller, markedly less fecund adults. This difference, it was argued, should create a selection pressure for reduced clutch size in American wasps. The experiments described in Chapter Four showed that British A . gl o m e r a t u s search less efficiently for P. r a p a e than for P. brassicae, and that A. rubecula has a comparatively high searching efficiency for P. rapae. In this case, selection is expected to favour an increased searching efficiency in American wasps, approaching that shown by A. rubecula. Despite these results and predictions, and despite approximately three hundred generations of exposure to P. r a p a e , American A. glomeratus , when compared with British A. glomeratus attacking P. r a p a e , showed no evidence of having evolved a smaller clutch size (Chapter Five), or a higher 210 searching efficiency (Chapter Four). Although American A. glomeratus may have become adapted in other ways, the lack of adaptation of clutch size and searching efficiency is likely to be a severe constraint on the effectiveness of American wasps as biological control agents of P. rapae. Parker (1970) and Parker et al (1971) invoked several additional reasons to account for the ineffectiveness of A. glomeratus in the United States. These included poor synchrony with host phenology; high egg and larval encapsulation rates; heavy hyperparasitism; and competition with A. rubecula , which is a superior larval competitor (Richards, 1940; Parker et al, 1971). A. rubecula is potentially a more effective control agent than A. glomeratus. Not only does it have a higher competitive ability as a larva, and a higher searching efficiency as an adult, it is also reported to aggregate in areas of high host density in the field (Richards, 1940). Furthermore, hosts parasitised by A. rubecula are killed before they are half-grown. They therefore cause much less damage than caterpillars parasitised by A. g l o m e r a t u s , which emerges from the full-grown host. The potential of A. rubecula as a biological control agent of P. r a p a e was overlooked until quite recently. It was discovered in British Columbia in the early 1960s, presumably having been imported accidentally (Wilkinson,. 1966). Introductions were made from British Columbia to Missouri (Puttier et al, 1970) and California (Oatman & Platner, 1972). Establishment did not occur in either locality, although effective control of P. r a p a e was achieved in Missouri using repeated releases of A. rubecula and Trichogramma evanescens , together with host eggs (Parker et al, 1971; Parker & Pinnell, 1972). A. rubecula did become established when introduced to Ottawa (Corrigan, 1982). The failure of the species to establish in southern latitudes has been attributed to the wasps entering a photoperiodically-induced diapause at lethally-high autumn temperatures (Oatman & Platner, 1972; Nealis, 1985). Poor adaptation of exotic parasitoids to novel environments may often limit the success of biological control programmes (van den Bosch et al, 1983). The biological control of P. r a p a e in the United States appears to be hampered by the lack of adaptation of A. glomeratus to the host, and of A. rubecula to the climate. A cited advantage of natural enemies over insecticides is their capacity to coevolve with the host over evolutionary time (Greathead, 1986). 211 Evidence for more rapid post-colonisation adaptation, of the sort envisaged by Price (1980), appears to be rare in the literature (Peschken, 1972). The ability of introduced insect populations to evolve rapidly m a y be limited because the original introductory stock is often based on a small number of founders (Messenger et al, 1976). This may create a genetic bottleneck, and a population with low genetic diversity. In conclusion, it is clear that studies on the behaviour and evolutionary biology of parasitoids have implications for understanding their role in population regulation and classical biological control (Waage & Hassell, 1982). Models of host-parasitoid population dynamics have long incorporated searching efficiency and handling time (Hassell, 1978). More recently, the effects of clutch size, sex ratio and developmental mortality have also been included (Hassell, 1986). A n evolutionary understanding of clutch size and sex ratio may also provide practical advantages in the mass-rearing of parasitoids for inundative release programmes. For example, Apanteles flavipes (Cam.), a gregarious parasitoid of pyralid stalk borers, is used in the biological control of sugar cane pests in South America. Macedo et al (1983) describe the production of ten million wasps per month for continuous release against D i a t r a e a spp. in Brazil. The high-density rearing associated with this type of mass production creates problems concerned with maintaining quality control. Sex ratios are often male-biased, and adult wasps small and unfit (Waage, 1986). A n appreciation of how wasps adjust clutch size and sex ratio in response to host and parasitoid densities m a y help to improve mass production techniques (Waage, 1986; Pallewatta, 1986). 212 ACKNOWLEDGEMENTS I thank Mike Hassell for making available the facilities at Silwood Park; and m y supervisor, Jeff Waage, for his help, advice and encouragement on all aspects of this work. I also benefitted from helpful discussion with Charles Godfray, Mike Hassell and Mick Crawley. Carlos Garcia and Mike Hassell assisted with the curVe- fitting procedures described in Chapter Four. I a m grateful to Robert Zwart, Neil Gilbert and Roy van Driesche for supplying A. glomeratus cocoons from overseas; to Gwen Marsh and Chris Paine of the Glasshouse Crops Research Institute for providing Pieris eggs; and to Mark Shaw for permission to use his unpublished data in Chapter Six. The work was supported by a research studentship from the Natural Environment Research Council. 213 REFERENCES Adler, S. 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Commonwealth Institute of Biological Control, Technical Bulletin No. 4, 121-142. 248 A P P E N D IX I Host mortality in Experiments 1 and 2 of Chapter Three Table (i) shows the mean number of hosts dissected (AO in Experiments 1 and 2, in each host density (D) treatment (excluding that of one caterpillar per plant), and the calculated percentage survival of hosts between experiment and dissection (mean N as a percentage of D). Experiment 1 Experiment 2 D M ean N Mean % survival M ean N Mean % survival 5 4.5 90.0 - - 10 8.7 87.0 8.9 89.0 25 22.5 90.0 18.3 73.2 50 40.1 80.2 43.1 86.2 T able (i) An estimate of the total number of hosts attacked during each replicate (N a), correcting for this mortality, was obtained by multiplying the proportion of dissected hosts found to be parasitised (N p /N ) by initial host density (D), i.e .: Na = D .N p /N Table (ii) shows the mean values of N a calculated for each host density treatment used in Experiments 1 and 2. For each of the three common host densities (10, 25 and 50 caterpillars per plant), the mean values of N a do not differ significantly between the two experiments (two-way ANOVA: F^2 35 j = 0.12; p > 0.05). Experiment 1 Experiment 2 D n M ean N a S.E. n M ean N a S.E. 5 6 3.3 0.7 - - - 10 6 8.2 0.9 8 6.8 0.8 25 6 16.2 3.0 7 17.9 2.4 50 6 23.5 3.7 8 21.0 4.3 T able (ii) 249 APPENDIX II L ist o f A panteles species used in Chapter Six The following is a list of all the A pan teles species taken from the literature which are known to be solitary or gregarious, together with information (where available) on their hosts, host sizes, wasp sizes and brood sizes. Host size is the length of the adult fore-wing (in mm). Wasp size is the total length of the adult (in mm). Brood size is the number of cocoons per host. Species known to be gregarious, but whose actual brood size is unknown, are marked w ith a G. A pan teles sp. Host sp. Host size W asp size B rood s a b jectu s Eligmodonta ziczac 22 2.5 15 acasta Allophyes oxyacanthae 22 62 acherontiae Ackerontia lachesis 65 2.5 1226 acraeae Acraea acerata 20 2.5 1 acron yctae Acronycta oblinita 2.5 G acuminatus Mellicta athalia 21 42 aethiopicus Utethesia pulchella 17 2.5 G a ffinis Cerura vinula 35 . 2.5 G aletiae Alabama argillacea G amabilis Noctua pronoba 27 2.5 12 am ericanus Erinnys ello 53 G anarsiae Anarsia lineatella 2.7 1 ancilla Colias hyale 26 2.5 G angaleti Pectinophora gossypiella 9 1 anisotae Anisota senatoria 2.0 G anomidis Anomis flava 14 14 anthelae Anthella ocellata - 2.7 1 anti pod a Spodoptera mauritia 15 2.0 G arcticus Arctia agestis 14 2.6 G argynnidis Speyeria cybele 43 G atalan tae Vanessa atalanta 34 G atella e Atella phalanthra 2.3 G 250 A partteles sp. H ost sp. H ost size W asp size B rood ater Archips podanus 11 2.4 G australiensis Antheraea eucalypti 70 3.5 938 b a d g leyi 3.0 G baoris Parnana mathias 19 1.9 G bataviensis Odonestis plagifera 2.5 G bedelliae Bedellia somnulentella 4 G belippae Belli pa lohor 3.0 r belliger Pseudaletia unipuncta 21 2.2 l berberis 2.5 G bign ellii Euphydryas aurinia 21 2.1 39 bisulcata Stenodactyla concursa 2.2 1 bordagei Leucoptera coffeella 3 1.7 1 brevicornis Cleoceris viminalis 16 2.4 8 brittanicus Aristotelia inopella 5 2.7 1 butalidis Sc yt hr is fuscoaenea 7 2.7 1 caberae Bupalus piniarius 19 2.7 1 cacao Lymantria ampla 2.5 G cacoeciae Archips semiferanus 10 4 c a ffre y i 2.2 G ca jae Arctia cajae 31 2.9 18 callidu s Parasemia plantaginis 17 3.0 G callunae Anarta myrtilli 13 2.8 1 canarsiae Ancylis comptana 6 1 capucinae Calyptra thalictri 3.0 G carbonarius Bucculatrix nigricomella 4 1.8 1 carpatus Tinea pellionella 6 2.5 1 cassianus Eurema nicippe 21 1 cerialis Ascotis selenaria 22 2.7 1 charadrae Charadra deridens 2.2 G chares Calotois pennaria 24 2.8 9 chilonis Chilo partellus 13 23 circumscriptus Lithocolletis messaniella 5 2.5 1 cleora Apeira syringaria 20 3.0 19 coleophorae Coleophora serratella 2.5 1 251 A pan teles sp. Host sp. * Host size W asp size B rood com es Bucculatrix cristatella 3 1.8 1 compressiventris Arctia villica 27 2.9 141 com pressus G congestus 2.2 G congoensis Dichocrocis crocodora 24 congregatus Manduca sexta 66 2.5 150 coni fera e Swammerdamia lutarea 6 2.5 1 corvinus Swammerdamia lutarea 6 2.4 1 coryphe Hemaris fuciformis 24 3.1 1 cram bi Crambus zeellus 2.0 G cupreus Lycaena phlaeas 14 2.4 2 cyan iridis Lycaena pseudargiolus 17 2.8 1 cynthiae Euphydryas cynthia 24 3.6 G delicatu s Hemerocampa leucostigma 18 2.8 1 depressariae Depressaria pastinacella 11 1 detrectans 3.0 1 diacrisiae Hemerocampa leucostigma 18 G diatraea Diatraea saccharalis 14 64 d iffic ilis Macrothylacia rubi 30 G dignus Keifera lycopersicella 1 dilectu s Caloptilia syringella 6 2.6 1 diparopsidis Diparopsis castanea 16 G ed w a rd sii Vanessa atalanta — 34 2.6 1 electrae Hemileuca electra G em pretiae Sibine stimulea G endem us Abraxus grossulariata 22 2.6 1 enephus Erannis defoliaria 21 3.0 1 erionotae Pelopidas thrax 33 2.6 69 errator . Eupithecia virgaureata .11 3.2 1 etiellae Ancylostomia stercorea 1 euchaetis Euchaetis egle G eucosm ae Diacrisia mundata 2.5 G eulipis Rheumaptera hastata 16 2.5 G euphydryadis Euphydryas phaeton 21 2.2 28 252 A pan teles sp. H ost sp. H ost size Wasp size Brood size eu ryale 2.5 1 expu lsu s Amyna puncium 2.0 G fa b ia e 2.0 G fa lca tu s Parastichtis monoglypha 25 3.5 G ferrugineus Archanara geminipuncta 16 2.0 G fisk e i G flaviconchae Pseudaletia unipuncta 21 2.2 G flavicornis Erynnis Juvenalis 20 2.0 G flavicoxalis Lymantria obfuscata 2.5 1 fla v ip e s Chilo partellus 13 2.2 35 flaviventris Erinnys ello 53 627 flavovariatus 3.3 G florid anus Leucania latiuscala 2.3 G forbesi Lacinopolia renigera 1 formosus Abraxus grossulariata 22 3.2 1 fra tern u s As pi tales ochrearia 16 1.9 100 fu lvip es Chesias legatella 18 3.0 21 fumi feranae Choristoneura fumiferana 9 1 gabrielis Pionea forficalis 14 2.4 11 gades Stauropus fagi 31 3.0 G g a lleriae Galleria mellonella 18 2.7 1 gastropachae Gastropacha quercifoliella 36 2.2 21 geryon is _ Procris geryon 12 2.2 G glom eratu s Pieris brassicae 34 2.7 27 glyphodes - Glyphodes sericae - - 3.1 G gonopterygis ' Gonopteryx rhamni - 33 2.9 1 gracilariae Caloptilia syringella 6 2.7 1 grenadensis Pseudaletia unipuncta 21 1 g r iffin i Agrotis gladiaria G harrisinae Harrisina metallica 15 G harti Pyrausta penitalis 1 hem ileucae Hemileuca maia G hesperidivorus Erynnis tristis 1.7 G h ydriae Calocalpe undulata 18 2.2 G 253 A pan teles sp. Host sp. Host size Wasp size Brood size hyphantriae Hyphantria cunea 13 3.8 1 h yposidrae 2.0 G im m unis 2.7 1 im perator Epermenia chaerophyllella 6 2.7 1 im portunus Nephopteryx rhodobasalis 2.0 1 inclusens Euproctis chryssorhoea 19 2.7 17 indiensis Lymantria obfuscata 3.0 1 in fim u s 2.2 1 inquisitor Lamprosema diemenalis 12 2.2 1 isolde Polyploca ridens 19 2.4 G ju cu n du s 3.0 1 ju ju b a e 2.5 G ju n ip tera e Thera juniperata 14 2.8 1 junoniae Junonia coenia 26 2.6 1 kariya e Leucania separata 22 60 k a za k Heliothis armigera 17 3.5 1 koebelei Euphydryas edithe 26 20 lacteicolor Euproctis chryssorhoea 19 2.5 1 lacteus Homoeosoma nimbella 9 3.2 1 laetus Caloptilia semi fascia 5 2.4 1 laeviceps Trichoplusia ni 17 G laevigatus Anacampsis populella 8 3.0 1 laevissimus - Evetria sylvestrana 7 2.7 1 lamprosemae Lamprosema diemenalis 12 2.5 1 laricellae Argyresthia laricella - 5 1 la teralis Anthophila fabriciana 6 2.7 1 laverna Pyrausta aurata 8 2.0 9 lenea Oncocera semirubella 14 3.2 1 leptoura _ z. Jiypsipyla robusta 3.6 1 lesbiae Colias lesbia 24 1 leucaniae Leucania straminea 18 2.4 G lim batus Abraxus grossulariata 22 2.7 19 lineola Evergestis forficalis 14 2.5 16 lip a rid is Lymantria dispar 25 3.2 24 254 A pan teles sp. Host sp. - Host size Wasp size Brood size longicauda 3.0 1 lunatus Papilio polyxenes 45 3.0 1 luteipennis 2.0 G lycophron Melitaea didyma 23 2.5 G lym antriae Lymantria dispar 25 3.0 1 machaeralis Eutectona machaeralis 9 2.6 1 malevolus Hyblaea puera 2.2 G marginiventris Spodoptera frugiperda 17 1 maritimus Bucculatrix maritima 4 2.2 1 medicaginis Colias philodice 24 1 megathymi Megathymus yuccae 30 G melanoscelus Lymantria dispar 25 2.8 1 melittaerum Melitaea cinxia 24 3.2 20 m endranae 2.0 G m ilitaris Pseudaletia unipuncta 21 72 m lan je 2.5 G murtfeldtae 1 nanus 2.0 1 nem oriae Eupithecia miserulata 1.9 1 neomexicanus 3.5 .1 nepitae Nepita conferta 3.0 G nigricornis Ctenucha brunnea 2.4 G nonagriae Phragmatiphila truncata ~ -- G nothus Anticlea badiata 16 1.8 21 numen Eupithecia intricata- 12 3.0 1 obliquae Diacrisia obliqua - 3.0 G obscurus Ebulea crocealis - : 12 3.6 1 ocneriae Lymantria dispar 25 2.2 G octonarius Oenistis quadra 22 2.4 G o fella Acronicta rumicis 19 3.0 75 onaspis Cnaemidophorus rhododactyla11 2.3 G ordinarius Dendrolimus pini 37 3.1 G ornigis Phyllonorycter lucidicostella 2.2 1 ovestes Philudoria potatoria 30 . 2.8 G 255 A pan teles sp. Host sp. Host size Wasp size Brood size pallipes Autographa gamma 21 2.7 71 paludicolae Exelastis atomosa 2.2 1 papaipemae Papaipema nebris 2.5 G parallelus Hemithea aestivaria 16 2.5 1 paranthrenidis Paranthrene robiniae 3.8 G parasae Parasa lepida 15 2.0 G parasitellae Triaxomera parasitellae 9 2.9 1 parastichtidis Rusina bicolorago G phaloniae Phalonia smeathmanniana 9 3.0 1 phigaliae Phigalea titea G phobetri Halisidota tesselaria G pholisorae Pholisora catullus 2.5 1 phytometrae Phytometra chalcites G pieridis Aporia crataegi 35 2.5 17 pilicornis Amblyptilia punctidactyla 10 3.1 1 pinicola Thera variata 14 3.4 1 pistrinariae Mylothris chloris 31 2.5 G plutellae Plutella xylostella 7 3.0 1 podunkorum Pyrausta futilalis G politus Scolecocampa liburna G polychrosidis Polychrosis liriodendrana 1 popu laris Tyria jacobaea 20 3.0 14 porthetriae Lymantria dispar 25 3.2 1 praepotens Operophtera brumata 15 2.9 1 prince ps Coleophora virgaureae 5 2.8 1 prosper 2.0 G puera Hyblaea puera 33 3.2 -- r p y ra lid is Nomophila noctuella 15 2.2 G pyrau stae Pyrausta futilalis G rad ia n tis Euxoa radians 2.1 G reinhardi Plusia gamma 21 3.0 G risilis Gonepteryx rhamni 33 . 3.2 1 robiniae Recurvaria robiniella 2.0 1 rubecula Pieris rapae 24 3.1 1 256 A pan teles sp. Host sp. Host size Wasp size Brood size ru bripes Geometra papilionaria 28 3.0 G ru ficru s Leucania separata 22 2.2 24 ru focoxalis Pseudaletia unipuncta 21 2.5 G ru idis Eutectona machaeralis 9 2.2 1 sa g a x Sylepta derogata 18 2.5 24 sa lta to r Anthocaris card amines 23 3.5 1 sarrothripae Acleris permutana 9 G scabriculus Earias clorana 11 2.8 1 schae feri Lymantria dispar 25 2.2 12 sch izu rae Schizura unicornis G scitulus Acronycta oblinita 2.5 G scutellaris Pthorimaea operculella 7 2.6 1 sesamiae Busseola fusca 16 105 seteb is 7 sm erinthi Smerinthus geminatus G sotad es 3.1 1 spurius Bis ton strataria 27 2.9 15 stantoni Margoronia laticostalis 2.5 1 sta rk i _ Eucosma rescissoriana - 1 subandinus Pthorimaea operculella 7 1 sybyllaru m Ladoga Camilla 30 2.9 G talidicida Talides sergestus 24 G taprobanae Stauropus alternus 2.7 G tasmanica Tortrix postvittana 3.0 1 ted ella e Epinotia tedella 6 1 tetricu s Maniola jurtina 25 2.2 33 theclae Everes comyntas 13 2.3 G thorn psoni Pyrausta nubilalis 16 2.2 G thurberiae Thurberiphaga diffusa — 3.8 1 tibia lis Maniola jurtina 25 2.6 39 tirath abae Tirathaba complexa 3.0 1 triangulator Pseudoterpna pruinata 19 2.2 G ugand aensis " Archips occidentalis 21 G ultor Euproctis chryssorhoea 19 2.5 G 257 A pan teles sp. Host sp. Host size Wasp size Brood vanessae Cynthia cardui 33 2.5 G vernaliter 2.1 1 victor Goniodoma limoniella 5 2.7 1 villanus Arctia villica 27 2.7 12 viminetorum Elachista trapeziella 4 2.1 1 vitripennis Chesias legatella 18 3.2 1 w ebsteri 2.3 G xanthostigmus Swammerdamia lutarea 6 2.9 1 xylinus Agrotis c-nigrum 20 2.5 G yakutatensis Autographa californica 22 29 zygaenarum Zygaena filipendulae 17 2.6 13 A pan teles sp. 1 Hypsipyla robusta 33 A pan teles sp. 2 Hypsipyla robusta 23 A pan teles sp. 3 Hyblaea puera 40 A pan teles sp. 4 Galleria mellonella 18 1 A pan teles sp. 5 Darna trima 10 122