-1-
BIOLOGY AND BEHAVIOUR OF TELENOMUS SPP. (HYMENOPTERA: SCELIONIDAE) EGG PARASITOIDS, ATTACKING SPODOPTERA SPP. (LEPIDOPTERA: NOCTUIDAE)
BY
SIVASUBRAMANIAM RAVEENDRANATH B.Sc. (Agric.) Sri Lanka
A thesis submitted for the degree of Doctor of Philosophy
of the University of London and for the Diploma of membership of the Imperial College.
Dept, of Pure and Applied Biology Imperial College, Silwood Park, Ascot,
Berkshire February 1987 -2-
ABSTRACT
The main objective of this laboratory based study is to compare the biology and behaviour of eight Telenomus populations obtained from different parts of the world and from the findings to recommend which strain and/or species will be more suitable to control the eggs of
Spodoptera spp. in the field.
The developmental times of the eight Telenomus populations were studied on Spodoptera littoralis and S. frugiperda and it was found that the developmental time of the Telenomus population from the Hawaiian region was longer than the others. Longevity was measured for each population in the presence and absence of the host. In all the Telenomus populations mated males are short lived and the longevity of mated females was reduced considerably when exposed to hosts.
The fecundity of these Telenomus populations was studied on S_. littoralis and S. frugiperda. In both host species, the highest and lowest potential fecundity was recorded in the Telenomus populations from Barbados and Hawaii respectively.
Cross breeding experiments were carried out between the Telenomus populations from various sources to determine their genetic compatibility.
Findings from these experiments indicate that there are two distinct biological species, T. nawaii (Hawaiian region) and T. remus (Barbados and elsewhere). Therefore further experiments on the biology and behaviour of these two biological species were carried out with additional host species -3-
S. exigua and exempta. The fecundity of both the biological species was
significantly reduced when they were exposed to exigua. Although more
females than males were observed in these naturally inbreeding populations,
a male biased sex ratio was observed with increased parasitoid:host ratio
of the two biological species.
In a multilayered eggbatch, both of these biological species were able
to attack only a small proportion of the eggs in the lower layer. Finally
the searching efficiency of the two biological species was compared on the host eggs laid on Brussels sprout and onion plants. It was found that,
although T. nawaii finds hosts more rapidly, the searching efficiecy
(attack rate) was higher for T. remus. The two parasitoid species showed no behavioural differences in attacking eggs laid on these two plants.
Based on the biological parameters tested in this study T. remus
Barbados is a better choice than T. nawaii in many aspects and could be recommended for future use. ACKNOWLEDGEMENTS
In the course of this work over a period of more than three years, many have helped me in various ways. It is difficult now to apportion my
indebtedness and some past assistance may be inadvertently overlooked in this ultimate expression of gratitude.
I wish to record my sincere thanks to my supervisor Dr. D.J. Greathead and my advisor Dr. J.K. Waage for their unstinted support, guidance and invaluable help throughout the course of this work.
My sincere thanks are also due to Professor M.P. Hassell and to Dr. D.J.
Greathead for providing facilities at Silwoodpark and at the C.I.B.C. repectively.
I wish to acknowledge the friendship and occasional helping hand given to me by members of C.I.B.C. Sincere thanks are also due to to Drs. M.J.W.
Cock and D.J.O'Donnell for the valuable discussions related to various aspects of this work. I wish to express my gratitude to Mrs. A. Greathead for spending her time reading the typescript of this document. Statistical advice given by Drs. M.J. Crawley and T. Ludlow is greatly acknowledged.
Sincere thanks go to the following: Mrs. R. Warner (IOV), Miss Sandra
Simith (TDRI, PortonDown) and Miss Stella Simith (ICI) for providing
Spodoptera cultures and the Ministry of Agriculture and Fisheries (UK) for granting permission to culture them in the United Kingdom. C. Hamilton and
D. Girling for finding literatures through the computer data bank at C.I.E. -5-
I am very grateful to my best friend/ Aris, for the colour photographs and the occasional helping hand given by him during the course of this study. The friendship afforded to me by members of Insect Ecology group at
Silwood Park, and my colleagues Carlos, Trevor, Andy, Saysi and Elena, deserve special mention.
I am most grateful to the financial help from the Association of the
Commonwealth Universities and to Professor S. Rajaratnam, Batticaloa
University for granting study leave for me to persue this study. -
Finally I would like to thank my parents, my wife Easwary and our children Dus hi and Abi for the support, encouragement and joy they have given me during the period of this study. TABLE OF CONTENTS PAGE
Title page • • • • 1
Abstract • • • • 3
Acknowledgements • • • • 5
Table of contents • • • • 6
List of Tables • • • • 10
List of Figures • • • • 13
List of Plates • • • • 14
CHAPTER 1 GENERAL INTRODUCTION • • • • 15
Spodoptera as a pest • • • • 15
1.1 Taxonomy and diversity • • • • 15
1.2 Distribution • • • • 16
1.3 General life history • • • • 16
1.3.1 eggs • • • • 16
1.3.2 larval stage • • • • 17
1.3.3 Pupal stage • • • • 19
1.3.4 Adult stage • • • • 19
1.4 Food plants • • • • 19
1.5 Nature of damage and economic losses 20
1.6 Control methods 21
1.6.1 Chemical control 21
1.6.2 Biological control 22
1.6.2.1 Pathogens 22
1.6.2.1.1 Viruses 23
1.6.2.1.2 Bacteria 23
1.6.2.2 Predators 24
1.6.2.3 Parasitoids 24 -7-
1.6.3 Other methods of control .... 25
1.7 Importance of Telenomus spp. in control .... 26
1.7.1 Telenomus egg parasitoids associated with Spodoptera .... 26
1.7.2 Biology and Ecology of Telenomus .... 27
1.7.3 Introduction and Results .... 29
1.8 Aims of this study .... 36
CHAPTER 2 CULTURING OF HOST AND PARASITOIDS .... 38
2.1 Culturing of host .... 38
2.2 Rearing of parasitoids .... 40
CHAPTER 3 MORPHOLOGY .... 43
3.1 Introduction .... 43
3.2.Materials and Methods .... 44
3.2.1 Male genitalia .... 44
3.2.2 Female antenna .... 46
3.3 Results and discussion .... 46
3.3.1 Male genitalia .... 46
3.3.2 Female antenna .... 50
CHAPTER 4 GENERAL BIOLOGY .... 51
4.1 Introduction .... 51
4.2 Materials and Methods .... 52
4.2.1 Developmental time .... 52
4.2.2 Longevity .... 52
4.3 Results and discussion .... 53
4.3.1 Developmental time .... 53
4.3.1.1 Effect of host species on development time .... 53
4.3.1.2 Effect of temperature on the dev. time of the two biological species 60 -8-
4.3.2 Longevity .... 66
4.3.2.1 Males .. .. 66
4.3.2.2 Females .... 69
4.3.2.3 Females with host .... 70
CHAPTER 5 REPRODUCTIVE BIOLOGY .... 71
5.1 Introduction .... 71
5.2 Materials and Methods .... 72
5.2.1 Potential fecundity .... 72
5.2.2 Actual fecundity .... 74
5.3 Results and Discussion .... 74
5.3.1 Potential fecundity .... 74
5.3.2 Actual fecundity .... 81
5.3.3 Comparing actual and potential fecundity .... 90
5.3.4 Size and fecundity .... 93
5.3.5 Rate of increase .... 93
5.3.6 Sex ratio .... 97
CHAPTER 6 CROSS BREEDING .... 101
6.1 Introduction .... 101
6.2 Materials and Methods .... 102
6.3 Results and Discussion .... 105
CHAPTER 7 BEHAVIOUR OF THE TWO BIOLOGICAL SPECIES OF Telenomus .... 113
7.1 Introduction .... 113
7.2 Materials and Method .... 114
7.2.1 Effect of plant species in host location .... 115
7.2.1.1 Results and Discussion .... 117
7.2.1.1.1 Host locating efficiency of the parasitoids .... 118
7.2.1.1.2 Effect of plant species 122 -9-
7.2.1.1.3 Effect of host species .... 125
7.2.2 Efficiency of the parasitoids .. .. 127
7.2.2.1 Method .... 127
7.2.2.2 Results and discussion .... 130
7.2.2.2.1 Percentage parasitism .... 130
7.2.2.2.2 Area of discovery .... 130
7.3 Effect of the structure of the host egg mass on the oviposition of Telenomus .... 135
7.3.1 Method .... 136
7.3.2 Results and discussion .... 138
CHAPTER 8 GENERAL DISCUSSION .... 140
BIBILIOGRAPHY .... 149
Appendix I Raw data for rate of increase experiments .... 162
Appendix II Raw data for host finding experiments .... 164
Appendix III Raw data for searching efficiency experiments 168 -10-
List of Tables
Distribution of common Spodoptera spp 18
Records of Telenomus spp. against Spodoptera spp 30
Introduction of Telenomus spp. against Spodoptera spp. by the C.I.B.C. (India) 31
Introduction of Telenomus spp. against Spodoptera spp. by the C.I.B.C. (Barbados) 32
Introduction of Telenomus spp. against Spodoptera spp. by the C.I.B.C (Trinidad). 33
Source of Spodoptera spp 39
Source of Telenomus spp 42
Morphological description of Telenomus associated with Spodoptera 45
Length and width of male genitalia of T. remus Barbados, T. nawaii and T. solitus 48
The developmental times of the eight Telenomus populations on two host species 54
The overall mean developmental times of the eight Telenomus populations 55
Anova: comparing the developmental times of eight Telenomus populations on two host species 55
Developmental times of the two biological species of Telenomus on four host species 57
The overall mean developmental times of the two biological species of Telenomus on four host species 58
Anova: comparing developmental times of the two bioTogical species of Telenomus on four host species 58 -11-
4.7 Size of host eggs 59
4.8 Anova: Mean hh SE size of host eggs 59
4.9 Developmental times of the two biological species at five temperatures 61
4.10 longevity of males 67
4.10a Anova: longevity of mated males 67
4.10b Anova: longevity of unmated males 67
4.11 longevity of females 68
4.11a Anova: longevity of females reared in groups 68
4.11b Anova: longevity of females reared individually 68
4.11c Anova: longevity of females with hosts 68
5.1 Anova: Potential fecundity of eight Telenomus populations on two host species 79
5.2 Potential fecundity of eight Telenomus populations reared on two host species 79
5.3 Anova: comparing the potential fecundity of two biological species reared from four host species .... 80
5.4 Potential fecundity of the two biological species of Telenomus on four host species .... 80
5.5 Anova: comparing the actual fecundity of eight Telenomus populations on two host species .... 82
5.6 Actual fecundity of Telenomus on two host species .... 83
5.6a Mean ovipositional time of the eight Telenomus populations .... 89
5.7 Intrinsic rate of increase of eight Telenomus populations on two host species .... 95
5.8 Sex ratio of eight Telenomus populations on two host species .... 96
6.1 Time taken for mating between different crosses .... 107
6.2 Hybridization experiments: percentage -12-
female progeny in F1 and F2 generation 108
Hybridization avperiments: potential fecundity of F1 progeny resulting from various crosses 111
Comparing the fecundity of F1 progeny with the fecundity of mid parent 112
Cumulative number of parasitoids found found the egg batches after six hours 119
Number of parasitoids found on the egg batch after six hours 120
Comparing overall mean number of parasitoids arriving at the host eggs laid on different plant species after six hours 123
The percentage of parasitism of host eggs on different plant species 128
Anova: Comparing percentage parasitism of host eggs 129
The area of discovery of T. remus and T. nawaii 132
Anova: comparing area of discovery, A of the two biological species 129
Anova: comparing area of discovery, a of the two biological species 134
Proportion host eggs parasitized by the two biological species 137
Anodev: comparing proportion host eggs parasitized by the two biological species 134 -13-
LIST OF FIGURES
Figure Page
3.1 Structure of the male genitalia of T. remus Barbados, T. nawaii and T. solitus .... 47
4.1 A Relationship between temperature and the development time of T. remus Barbados .... 63
4.1 B Relationship between temperature and the development time of T. nawaii .... 65
5.1 Potential fecundity of the eight Telenomus populations reared on S. littoralis and S. frugiperda .... 76
5.2 Number of progeny produced by Telenomus per female per day on S. littoralis .... 86
5.3 Number of progeny produced by Telenomus per female per day on S. frugiperda .... 88
5.4 Relationship between size and potential fecundity .... 92
5.5 Progeny sex ratio of T. remus and T. nawaii under different host/parasitoid ratio .. .. 100
7.1 Experimental cage used in host finding experiments .. .. 116
7.2 The cumulative mean number of T. remus and T. nawaii arriving on the host eggs laid on different plant species .... 121
7.3 The cumulative mean number of parasitoids arriving on different plant species with time .. .. 124
7.4 The cumulative (mean + se) number of parasitoids arriving on different host species with time 126 LIST OF PLATES
PLATE Page
I Structure of the female antenna of T. remus Barbados, T. nawaii and T. solitus .... 49
II A dissected female of T. remus. .... 73
III Parasitized and unparasitized eggs of Spodoptera littoralis 104 -15-
CHAPTER 1
INTRODUCTION
1 Spodoptera spp. as Pests
1.1 Taxonomy and diversity.
The genus Spodoptera Guenee (Lepidoptera; Noctuidae) is composed of species the larvae of which are popularly known as armyworms because of their habit of swarming from field to field during the caterpillar phase, although Brown & Dewhurst (1975) reported that all the Spodoptera species do not behave in this manner (e.g. S. littoralis Eoisduval). The species of this genus occur throughout the tropics and sub-tropics, and the presence of some Spodoptera spp. in the temperate region has also been reported by Todd & Poole (1980). The same authors have characterised and constructed a key for 14 species from the Western hemisphere.
Brown & Dewhurst (1975) reported that the species now included in
Spodoptera were until recently distributed in three separate genera,
Laphyama Guenee, Prodenia Guenee, and Spodoptera Guenee. They further wrote that, after the synonymity of these three genera under Spodoptera, there are at least 22 species in this genus including the 14 species from the Western hemisphere -16-
1.2 Distribution.
S.exigua (Hdbner) is the most widespread species in the genus, known from every continent except South America, and extending from the equator northwards to Finland and Sweden, and southwards to Australia and New
Zealand. S. frugiperda (J.E.Smith) is found throughout most of the Western
Hemisphere from Southern Canada to Chile and Argentina, and it is very abundant in the Caribbean Islands. There are records of S. exempta
(Walker) from most of the African countries south of the Sahara. This species is mainly confined to Africa, Asia and Australia. S. littoralis is very common in the Mediterranean countries, Africa and Europe. A detailed list of the distribution of the common Spodoptera species is given in Table
1.1.
1.3 General life history.
The life history of the Spodoptera species is reviewed briefly based on the information published in the literature (Luginbill 1928, Hatting 1941,
Wafa et al. 1972).
1.3.1 Eggs.
There are certain features by which the eggs of a species within this genus may be distinguished. They are characteristically laid in batches ranging from 100 to 800 or more, usually on the lower surface of the leaves. The eggs may be in a single flat layer, or in two or more layers.
Tanada & Beardsley (1958) reported up to five layers in S. mauritia
Boisduval. The eggs are placed very close together, and usually scales -17- from the moth cover the mass. The scales are very fine and are deposited from the body of the female while she is in the act of oviposition. The older females which have used up most of their supply of hair scales, produce a less complete covering. Brown & Dewhurst (1975) reported that the colour of the scales deposited is diagnostic for each species and provides one useful character for identification.
1.3.2 Larval stage.
The larval stage of armyworms is the most crucial both in terms of crop damage and potential for biological control. There are usually 5 to 6 larval instars. Larval characters have been used to identify and separate armyworm species (Brown & Dewhurst 1975). The length of each individual instar and the total length of the life of the larvae are dependent primarily on temperature. During the warmer months larvae are very active,
grow rapidly and have shorter instars and reach maturity sooner than do
larvae living in cooler weather. Phase variation (a type of polymorphism) during the larval stage is a common occurrence in some armyworm species.
Faure (1943 a, b) observed that the occurrence of dark caterpillars in the
field was often associated with high population density, and he showed experimentally that in S. exigua and S. exempta the effect could be
reproduced in the laboratory by crowding. Increased rate of development,
synchronization of development and increased food tolerance are some of the
important characters associated with phase variation in armyworms. High
density outbreak populations have more dispersive adults and larvae, and
the latter are darker in colour than the normal populations (Brown 1962) -18-
Table 1.1 Distribution of Spodoptera spp.
Species Locality
S.mauritia (Boisd.) Madagasger, Tanzania, Uganda, Pakistan Paddy armyworm India, Bangaladesh, SriLanka, S.E.Asia S.China, Phillipines, Indonesia, Hawaii, Pacific Islands,Fiji. (CIE map 162)
S.exempta (Walker) Africa,Madagasger,India, SriLanka, Papua African armyworm New Guinea, Burma, Malaya, Java, Hawaii, Borneo,Australia (CIE map 53)
S.littoralis (Boisd.) Africa, Israel, Egypt, Jordan, Lebanon, Cotton leafworm Syria, Turkey, Crete, France, Spain (CIE map 232)
S.litura (Fabricus) South and East old world tropics inclu Rice cutworm ding Pakistan, India, SriLanka, China, Korea, Pacific Islands, Hawaii and Fiji (CIE map A61)
S.exigua (Hubner) Africa, S.E.Asia, S.Europe, Australia, Beet armyworm Southern USA, Canada. (CIE map A302)
S.frugiperda North America, Central America, South Fall armyworm America, West Indies ( Trinidad, Jamaica, Barbados, St. Lucia. (CIE map 68 revised) -19-
1.3.3 Pupal stage.
The colour of the pupae of armyworm species varies from golden yellow to reddish brown. Pupation normally occurs in the soil. If the soil is loose, the larva burrows into it a depth varying from 3 to 8 centimeters and constructs a loose cocoon by tying together particles of soil with silk. The cell in which the larva pupates is oval in outline. When the soil is too firm for the larva to burrow in, it may pupate at the base of a plant by merely bringing dead foliage and sand particles together and making a frail cocoon. Pupae of S. frugiperda have also been found on the top of maize plants and especially in the husk of the maize ear (Luginbill
1928).
1.3.4 Adult stage.
Moths of armyworm species are not active during the greater part of the day but remain concealed, mainly under the foliage and among refuse. The colour and the shape of these moths have been used by taxonomists to separate different armyworm species. Activity begins a little before sunset, at sunset or just as the light begins to fade. Mating and oviposition usually occur during the night. The number of eggs laid by individual moths varies within and between species. As many as 1200 eggs per individual moth have been recorded for S. exempta and S. littoralis.
Among the factors that influence the longevity of adults are food and temperature. The average longevity of the moths of most armyworm species in the laboratory varies from 7 to 13 days.
1.4 Food plants -20-
The larvae of Spodoptera spp. feed on a large number of plants, although different Spodoptera spp. show some preference in attacking host plants. A detailed list of host plant species attacked by Spodoptera spp. is given by
Brown & Dewhurst (1975). Brown (1962) reported that, as a serious pest, S. exempta is virtually confined to graminaceous food plants. In this respect
S. frugiperda tends to resemble S.exempta (Luginbill 1928), although it has a far greater range of alternative food plants belonging to families other than Gramineae. S. exigua is less confined to grasses and is known to feed on plants belonging to at least 36 different families (Brown & Dewhurst
1975). In many of the published reports of food plants it is difficult to distinguish between S. littoralis and S. litura. Moussa et al. (1960) reported that S. litura feed on host plants approximately amounting to 112 species from 44 different families. It seems likely that any plant eaten by
S. littoralis would also be eaten by S. litura and vice versa.
1.5 Nature of damage and economic losses.
Climatic factors influence the sudden outbreaks of these insect pests in the tropics (Wiltshire 1946). The damage done by the caterpillars of
Spodoptera spp. is principally to the foliage. There is a definite pattern of attack shown by all Spodoptera spp. The first instar larva seldom eats entirely through a leaf, but it usually eats the green tissue from one side, leaving the membraneous epidermis on the other side intact. The whitish gnawed areas stand out prominently against the dark green background and this appearance is useful to determine the presence of young larvae in the crop. During the second and third instars, although some skeletonizing occurs, the larvae begin to make holes in the leaves and eat from the edges of the leaves inward. Luginbill (1928) reported that the -21- mature larvae of S. frugiperda damage corn ears, and the damage is similar to that of Heliothis sp. The economic losses due to S. frugiperda are well documented. Mitchell (1979) reported that the average annual crop losses due to this pest in the United States exceed 300 million dollars; during particularly severe outbreaks such as occurred in 1975, 1976, and 1977, losses attributed to this pest exceeded 500 million dollars annually.
1.6 Control methods.
1.6.1 Chemical control.
Chemical pesticides are useful and powerful tools for the management of
Spodoptera spp. populations. Many are effective dependable and adaptable for use in a wide variety of situations. Selective chemicals appear to offer an almost ideal means of pest control. Many insecticides and poison baits have been used against the larvae of Spodoptera spp. and it has not been possible to make an extensive study of the literature. (For example, during the period from 1973 to 1985, there are abstracts for about 312 references in Review of Applied Entomology on the use of insecticides against S_. littoralis alone). Therefore, the number of references on the use of insecticides to control the whole range of Spodoptera spp. throughout the world must be considerable. Effective insecticides to date include organochlorines, organophosphates, carbamates and synthetic pyrethroids. Although insecticides can be effective against Spodoptera spp., the development of strains resistant to insecticides with time, has been reported in the literature (Mitchell 1979, El Guindy et uL 1982,
Ingram 1975, Shaban & Elmalla 1983). However, Brown (1970) stated that resistance to insecticides is a problem unlikely to be encountered in S -22- exempta, since consecutive generations of the same population of S. exempta are unlikely to be subjected to the same chemical treatment* The author argues that owing to its migratory habit, one generation living on a cultivated crop and treated with insecticides may be followed by one or more generations in rangeland where chemical control is unlikely to be applied, so that there will be no continuity of selection to produce resistance.
An important disadvantage with application of pesticides in an agroecosystem is the consequent reduction of natural enemies of these and other pests. These effects have been demonstrated by resurgence of target pests and the change of status of formerly economically unimportant species into important pests. In Egypt, for example, the widespread use of chlorinated hydrocarbons in cotton fields was followed by resurgences of the target pest S. littoralis, and the unleashing of non target pests that generally had been of little importance, such as S. exigua, aphids and spider mites (FAO 1983).
1.6.2 Biological control.
1.6.2.1 Pathogens.
When naturally occurring mortality agents are assessed, it is readily seen that insect pathogens are important biotic entities which aid in regulating the abundance of many insect pest species (Burges & Hussey
1971). Besides causing outright death, insect pathogens may interfere with insect development and reproduction and lower insect resistance to attack by pesticides. Viruses and bacteria are the most important pathogens in -23- regulating the populations of armyworms.
1.6.2.1.1 Viruses.
Jack (1930) reported that viruses are the most effective and quickest natural factor reducing S. exempta numbers. The contribution of nuclear polyhedrosis virus (NPV) in the natural mortality of S. frugiperda has been reported by Ashley et al. (1980) and (ruxa 1982). However, the attempts made to control £>_. exempts with virus in Africa were not successful (Brown
& Swaine 1965). From their experiments, these authors found that the polyhedral virus is very specific to S. exempts and it did not attack other armyworm species, S. littoralis and S. exigua. Honsy et al. (1983) have shown that a sprayed viral formulation gave control of S. littoralis comparable with that obtained using egg mass collection or conventional insecticides and consideration is therefore now being given to the development of virus production on a commercial scale in Egypt.
1.6.2.1.2 Bacteria.
Bacteria, in particular Eacillus thuringiensis Eerliner (BT), can be effective as control agents for insect pests (Tanada 1959). Although previous attempts with BT failed to control S. littoralis (Moore & Navon
1973), Sneh et al. (1981) showed that all six instars of S. littoralis were killed by one active isolate of BT. Ignoffo et al. (1977) reported that higher dosage of BT is essential to control S. exigua. Salama & Zaki
(1983) raised doubts on the combined use of BT with larval parasites to control S. littoralis, as the biology of a larval parasite, Zele chloropthalma Nees Ichneumonidae is affected when it develops on the larvae -24- of S. littoralis fed on a diet containing BT.
1.6.2.2 Predators.
A great many vertebrate predators are known to destroy Spodoptera spp. and a detailed list of predators is given by Brown (1962). Of the predators, toads (Bufo spp. Bufonidae) may be important as control agents in some cases, but the most generally distributed and frequently reported are birds. In Africa the birds known to attack S. exempta are white storks
(Ciconia ciconia (L)) and crows (Corvus spp. Corvidae). However Brown
(1970) believes the numbers of these birds are never sufficient to make an
impression on really large outbreaks of hundreds of acres. The only introduction of a predator which is claimed to have been successful in controlling armyworms is that of the Indian mynah (Acridotheres tristis L.
Stumidae) into Hawaii, where 20 years after its introduction Zimmerman
(1953) considered it to be a more important control agent than insect parasitoids. Kamal (1951) reported that in Egypt the most important predators are insects (especially beetles and ants). Carabid beetles, especially Calosoma spp. have been reported preying on armyworms. The author also observed that even S. litura eggs parasitized by Telenomus nawaii Ashmead Scelionidae were also attacked by some predators.
1.6.2.3 Parasitoids.
Armyworms are attacked by a relatively large number of parasitoids. A detailed list of parasites attacking armyworms is given by Brown (1962).
According to his report at least 36 species of Hymenoptera and 23 species of Diptera (22 of them tachinids) are known to attack S. exempta. Ashley -25-
(1979) reported that fiftythree species of parasitoids from 43 genera and
10 families have been reared from the larvae of fall armyworm, S. frugiperda, and among those the most frequently recovered parasitoids are
Apanteles marginiventris Cresson Braconidae and Chelonus insularis
(texanus) Cresson Braconidae. In Cyprus three braconids and two ichneumonids are known to attack different stages of S. exigua and S. littoralis (Ingram 1981). It is interesting to note that there are no reports of the presence of egg parasitoids of Spodoptera spp. in Africa.
Brown (1970) reported that the fact that the combined effect of all these parasitoids fails to keep S. exempta in check suggests that there is no great future for biological control of Spodoptera spp. by insect parasitoids in Africa. In Queensland, Jarvis (1921) recorded 34.5 % of S. exempta larvae killed by parasitoidss, and Bell (1936) reported as much as
53 % destroyed by a tachinid species alone. There is no doubt that parasitoids exercise a large measure of control in Hawaii. Several species of parasitoids have been introduced into this island, and the relative scarcity of armyworms since 1942 has been attributed to their effects
(Pemberton 1933, 1948, 1964). The role of Telenomus spp. egg parasitoids in reducing the armyworm populations will be discussed separately.
1.6.3 Other methods of control.
Several practices which may be included under this heading have been
claimed as effective in certain circumstances. Among them hand picking of
larvae and eggs or trenching in front of advancing larvae are some of the
common methods that have been widely used by farmers. However Whellan
(1954) considers all these methods to be inferior to treatment with a good
insecticide. Agronomic methods such as flooding of rice fields were found -26- to be very effective in controlling S. frugiperda in the United States
(Luginbill 1928). Light traps and pheromone traps are now being used to monitor the armyworm populations and forewarn farmers of changes in population abundance (Honsy et al. 1983/ Brown 1970).
1.7 Importance of Telenomus spp. in control.
1.7.1 Telenomus spp. egg parasitoids associated with
Spodoptera spp.
According to the literature there are at least five species of Telenomus associated with Spodoptera spp. They are,
1) Telenomus nawaii Ashmead
2) Telenomus spodopterae Dodd
3) Telenomus remus Nixon
4) Telenomus minutissimus Ashmead
5) Telenomus solitus Johnson
T. nawaii was first discovered by Y.Nawa in Japan in 1904 from unidentified lepidopteran eggs and was described by Ashmead. It was accidentally introduced into Hawaii (Pemberton 1933) where it is known to attack and destroy about 90% of the egg masses of S. mauritia in the field.
Pemberton (1933) believes that T. nawaii came from Fiji.
T. spodopterae was described from Java and is known to attack the eggs of Scodoptera spp. It differs from T. remus in having large fore wings
(Nixon 1937) It was introduced into Egypt to control S. litura (Kamal -27-
1951).
T. remus was described by Nixon in 1937 from Spodoptera mauritia egg masses collected in Malaysia. The Telenomus sp. from the Spodoptera spp. egg masses collected in Papua New Guinea by staff of the Commonwealth
Institute of Biological Control (C.I.B.C.) Indian station in 1963 was identified as T. remus by the Commonwealth Institute of Entomology
(C.I.E.). However the same species was identified as T. nawaii by the
United States Department of Agriculture (U.S.D.A.).
T. minutissimus was originally described from Trinidad as a parasite of
Dactylopius sp. (Ashmead 1895) but this is generally considered an error and the host is thought to have been Spodoptera sp. Although the same species is known from Puerto Rico, the pre release survey by staff of the
C.I.B.C. West Indian station in Trinidad indicated that this species is absent from Trinidad. Furthermore, the Telenomus spp. specimens from the
Caribbean region were determined as T.remus by N.F. Johnson (Yaseen et al.
1981). However, based on the information gathered from individuals, Cock
(1985) reported that the presence of T. minutissimus in the Caribbean cannot be ruled out and suggested that this species is probably less effective against Spodoptera spp.
T. solitus was first collected from unidentified noctuid eggs on potato foliage near Salala, Guatemala, by E.R. Oatman in 1977. Johnson (1983) has described this species from the eggs of Trichoplusia ni Htlbner Noctuidae.
It has been introduced into the United States to control beet armyworm, S. exigua (McCurty 1983).
1.7.2 Biology and ecology of Telenomus spp. -28-
The basic biologies of T_. remus (reared on S. littoralis) and T. nawaii
(reared on S. mauritia) were described by Gerling (1972) and Pemberton
(1933) respectively.
T. remus.
T.remus is a solitary arrhenotokous parasitoid. At 25° C, the life cycle takes 10 days. It must oviposit in host eggs that are less than 48 hours old or the larvae will not develop. The longevity of this wasp under natural condition is unknown. In laboratory studies Schwartz & Gerling
(1974) found that females in groups lived six times longer (18 days) than solitary ones but the males did not show such difference in the longevity.
The authors also reported that a single female of this species would lay about 160 eggs throughout her lifetime. Wojcick et al. (1976) tested thirtynine species of Lepidoptera of various families in the laboratory and
found that T. remus will oviposit on eleven species of Noctuidae and one
species of Pyralidae.
T. nawaii.
T. nawaii is also a solitary arrhenotokous parasitoid. The
developmental duration of this species varies from 13 to 17 days. It has
not been studied under controlled environment. This species could live up
to 20 days even if it was deprived of host. The potential fecundity of
this species varied from 63 to 84 eggs. The actual fecundity ranged from
16 to 132 per parasitoid with an average of 89. In each case the
parasitoid laid all of its eggs during the first few days and never after
the 7th day. It prefers freshly laid host eggs for oviposition. -29-
1.7.3 Introductions and results
Telenomus spp. were introduced into various countries for biological control by many workers during the early part of this century (Pemberton
1933, Lever 1943, Kamal 1951). There were no attempts to introduce these parasitoids during the period from 1940 to 1963 due to the widespread use of chemical pesticides. However the adverse effects of chemical pesticides such as the development of resistant strains of armyworm and the risk of
environmental pollution have led to a revival of the use of biological
control agents against armyworm in many parts of the world (Yaseen 1979).
As a part of the integrated control programme of armyworm, the introduction of Telenomus spp. was undertaken by the C.I.B.C. through its network of
stations throughout the world. Details of these introductions and the
results are listed in Table 1.2. Surveys of natural enemies of S. litura
in Western Samoa indicate that 54% of S. litura eggs were parasitized by T. remus on taro (Colocasia esculenta L. Schott Araceae) plantations (Braune
1982). In India T. remus introduced from Papua New Guinea was recorded parasitizing about 60 percent of eggs of S. litura on cauliflower (Brassica
oleraceae var. botrytis L. Cruciferae)(Patel et al. 1979). Successful
control of S. frugiperda was achieved in Barbados by the release of T.
remus introduced from India (Alam 1979). The establishment of T. remus in
Earbados and the recoveries reported from India, Pakistan and Caribbean
countries (CARDI 1982) indicate the potential of this parasitoid in the
future control of armyworm, Table 1.2 Records of Telenctnus egg parasitoids against Spodoptera spp. (excluding the Introductions by C.I.B.C. stations)
Parasitoid Introduced from Introduced to Target Pest Remarks Reference
1. Telenanus nawaii Hawaii Guam S.litura effective against S.exempts V.P.Rao (1971) and S.mauritia Sweezy (19*10)
Tahiti Fiji S.litura Lever (19*13)
Fiji Egypt S.litura very effective against S.litura Kamal (1951)
Fiji Hawaii S.mauritia Accidentally introduced Pemberton (1933)
Hawaii C.I.B.C.lhinidad S.frugiperda fail to produce viable offspring with Cock (1982) T.remus
2. Telenanus spodopterae Java Egypt S.litura very effective when locating the egg masses of the host Kamal (1951)
J Table 1.2 Introduction of Telenctnus egg parasitoids by C.I.B.C. stations
A) Fran India
Parasitoid sp. Year* Introduced to Target pest Remarks References
Telenomus "remus" 1966,1967 Cyprus Spodoptera spp. results not known Determined as Telenanus nawaii by USDA 1971 Guam S.litura
1975 St. Helena Spodoptera spp
197^ fteuritius
1969 Israel S.littoralis results not known Rao (1971)
1971,1972 Barbados S.frqgiperda Established for some years and destroys over 80$ Spodoptera populat- ions in Barbados Alam (1979)
1975 C.I.B.C. for multiplication and further distri- Trinidad bution.
1975 New Zealand S.litura effective against S.litura Boardman (1977)
*Years refer to C.I.B.C. annual report Table 1.2 (B) From Barbados
Parasitoid Year Introduced to Target pest Results References
Telenanus ranus Nixon 1976 C.I.B.C. Trinidad Origin( Papua New Guinea) 1981/82 Antigua S.frugiperda recoveries reported CARDI (1982)
1981/82 Dominica
1981/82 Montserrat
1981/82 St.Kitts Spodoptera spp. recoveries reported CARDI (1982)
1981/82 St. Vincent
* Years refer to CIBC annual report Table 1.2 (C) Fran TRINIDAD
Parasitoid Year* Introduced to Target pest Remarks Rreference
1)Telenomus remus (PNG) 1975/76 Antigua Spodoptera spp. recovries reported CARDI (1982) Papua New Guinea strain 1975 Florida Spodoptera spp. not established VJaddill & Whitcomb (1982'
from 1976 et seq. Trinidad fields Spodoptera spp. recoveries reported Yaseen (1979)
1979/80 Surinam Spodoptera spp. recoveries reported Segeren et al.(1979)
1978/79 Sri Lanka Spodoptera exigua parasites were not effective on the Ravel (1978) host eggs laid on onion plants 1975,1979/80 St.Helena Spodoptera spp.
1975 Bermuda
1975,1984/85 Peru
1976,1977/78,1981 Nicaragua Spodoptera spp. results not known
1976,1978/79,1980 Msxico
1976,1977/78 Colombia
*Years refer to CIBC annual reports Table 1.2 (C) continued
Parasitoid Year* Introduced to Target pest Remarks Reference
T.remus (PNG) 1981/82,1984/85 Bolivia Spodoptera spp
1981/82 Netherlands
1977/78 Guatemala Spodoptera spp. results not known
1977/78 Guadeloupe
1977/78 Bahamas
1978/79 El Salvador Spodoptera spp.
1978/79,1981/82 Venezuela
1980/81 St.Kitts
1980/81 Guyana
1980/81 C.I.B.C. Pakistan S.litura recoveries reported
1981/82 Cape Verde
1982 Western Australia S.litura
* Years refer to C.I.B.C annual reports. Table 1.2 (C) continued
Parasitoid Year* Introduced to Target pest Remarks Reference
2) Telencmus remus 1983 Efr’azil Spodoptera spp. Dominican Republic strain 1983 India Spodoptera litura results not known
1983 Bolivia
1985 Nicaragua Spodoptera spp. results not known
3) Telenomus solitus 1985 Msxico Johnson. 1985 Peru
1985 Bolivia Spodoptera spp. results not known
•Years refer to C.I.B.C. annual reports -36-
1.8 Aims of this study.
The aims of this laboratory based study are to compare the biology and
behaviour of eight Telenomus populations from different parts of the world
and from the findings to recommend which strains and/or species will be more suitable to control the eggs of Spodoptera spp. in the field.
It has been shown by several workers that the biological parameters such
as rate of increase and searching efficiency are important for biological
control agents. Therefore studies were carried out on these aspects of the
Telenornus spp. populations to assess their value as biological control
agents. Moreover it was also hoped to find out if there had been any
changes in the biology of Telenomus spp. strains introduced to different
countries to adapt to local conditions.
Since there is some controversy regarding the taxonomic status of the
Telenornus spp. associated with Spodoptera spp., the taxonomy of these
species was also considered in this study. Because of the taxonomic
problems of Telenomus spp. associated with Spodoptera spp., there were
doubts about which species had been introduced to which country and it was
hoped that studies on biosystematics and cross breeding of these
populations from different sources would be useful to solve this problem.
As there were instances, particularly in my country (Sri Lanka), where
Telenomus remus introduced from Trinidad failed to parasitize S. exigua
eggs laid on onion plant (Allium cepa L. Allidiceae), studies were carried
out to test their efficiency of attacking the host eggs laid on different plant species.
Although the results of the experiments on aspects of biosystematics and crossing are discussed in different chapters the experiments were done at the same time on the Telenomus spp. populations as they arrived. -38-
CHAPTER 2
CULTURING OF HOST AND PARASITOIDS
Techniques for the culture of hosts and parasitoids are dealt with in this chapter. Precise details of experimental methods are given in appropriate chapters.
2.1 Culturing of host
Four Spodoptera spp. were used in this study and their origin is given in Table 2.1. Of the four species three were successfully cultured on an artificial diet described by Hoffman et al. (1966). The fourth species,
Spodoptera exempta, was cultured on wheat plants because it rejected the artificial diet. These insects were reared in the quarantine room at 25 +
1°C and 70% +_ 5% relative humidity with 16:8 dark:lignt, photoperiod.
Adult moths were kept in Watkins & Doncaster cages measuring 30 cm high and 22 cm in diameter. Ten pairs of moths were placed in each cage. Two cages were set up every week. The lid and the bottom of each cage was covered with Whatman No 1 qualitative filter paper, and vertical filter paper strips were hung inside the cages for oviposition. The adults were fed on 50% honey in water solution every day. The eggs were laid in tightly packed batches covered with hairs and scales from the tip of the female's abdomen. The egg masses were collected every morning for continuous culturing of the host species and for experiments with parasitoids. -39-
Table 2.1 Source of host species •
Host Obtained from
S. littoralis Tropical Developmet and Research Institute, Porton Down, Salisbury, Wiltshire, UK.
S. frugiperda Institute of Virology, Mansfield Road, Oxford, UK.
S. exiaua Imperial Chemical Industries, Jealott's Hill, Bracknell, Berkshire, UK.
S . exempta Desert Locust Control Organization for Eastern Africa, P.O.Box 30023, Nairobi, Kenya. -40-
For continuous culturing, a batch of about 300 eggs was soaked in a 0.2% bleach solution (sodium hypochlorite) and gently brushed to detach eggs.
The eggs were collected by filtration and then left to soak in a Petri dish containing 10% formaldehyde solution for half an hour, after which the eggs were throughly rinsed with distilled water. The separated eggs were collected on to filter tissues (nappy liners) to dry for about 20 minutes before being placed in 250 cc clear plastic pots. Hatching usually occured in four days and newly hatched larvae were transferred with the aid of a fine brush into clean 250 cc plastic pots with 100 cc diet. Two pots containing 50 to 100 larvae were set up every week. To avoid contamination and cannibalism the larvae were reared individually in 30 cc plastic cups after the fifth day from hatching and up to the pupal stage. Before pupation they stop feeding and tunnel into the remaining diet and hollow out a small chamber in which they pupate. The pupae were then collected and kept in plastic boxes with damped vermiculite for adult emergence.
2.2 Rearing of parasitoids.
As the egg parasitoids (Telenomus spp.) obtained from various sources were reared from different Spodoptera spp. they were exposed to the egg masses of S. littoralis to develop populations on a standard host species.
The source and the host species of the parasitoids are listed in Table 2.2.
The parasitoids were cultured in a quarantine room at 25° _+ 1°C and 70% +
5% RH with 16:8 dark : light regime. Ten females of each parasitoid population were exposed to a batch of about 400 fresh eggs of S. littoralis for a period of 24 hours. The egg masses were removed after being parasitized by Telenomus spp. and kept in separate glass vials (2.5 cm x
7.5 cm) for further development. On the fourth day the parasitized eggs -41-
tum black and the unparasitized ones hatched. The hatching host larvae were removed with a fine brush, as they tended to feed on the parasitized
eggs. The adult parasitoids were fed on pure honey drops smeared on the
wall of the glass vials. Parasitoids were standardized by allowing an
equal number of males and females together for 24 hours before being used
for other experiments. - 42-
Table 2.2 Source of parasitoids
Parasitoid Obtained from Host Date
T.remus (TRB) CIBC (Barbados) S.frugiperda 7/2/1984 field collected (J.E.Smith)
T.remus (TRT) CIBC (Trinidad) S.sunia 13/2/1984 lab. culture (Guenee)
T.remus (TRI) CIBC (India) S.litura lab. culture (Fabricius) 14/3/1984
T.remus (TRP) Phillipines S.mauritia 7/5/1984 field collected (Boisduval)
T.remus (TRQ 1) Queensland S.mauritia 26/9/1984 field collected
T.remus (TRQ 2) Queensland (Cairns) S.litura 26/9/1984 field collected
T.remus (TRM)* Mexico S.exigua (Htlbner) 14/3/1985 lab. culture
T.nawaii (TN) Hawaii S.mauritia 29/6/1984 field collected
I I
* This species has been described as T.solitus by N.F.Johnson. -43-
CHAPTER 3
MORPHOLOGY
3.1 Introduction.
The taxonomy of Telenomus spp. associated with Spodoptera spp. is in considerable confusion. Telenomus sp. collected from Papua New Guinea by the C.I.B.C. Indian Station was identified as T. remus Nixon by the C.I.E., yet apparently the same species was identified as T. nawaii Ashmead by the
U.S.D.A. Moreover F.W.Muesebeck (in Wojcek et al. 1976) wrote that T. remus is probably a synonym of T. minutissimus Ashmead while Nixon (1937)
in his original description of T. remus says that probably it will turn out
to be T. spodopterae Dodd. T. solitus, another species known to attack S.
exigua in Mexico was descibed by Johnson (1983). He reported that T.
solitus differs from T. remus in having large digital teeth in the male
genitalia.
Thus, according to the literature there are at least five different
species of Telenomus associated with Spodoptera spp. The morphological
descriptions of these species are summarized in Table 3.1. It is clear
from the table that the descriptions of some species are incomplete.
The structure of male genitalia and the female antennae are being used
to identify and separate Telenomus spp. by taxonomists. Therefore studies -44- were made on these structures of the two biological species of Telenomus as determined by experimentation and the population from Mexico.
3.2 Materials and methods.
Since there are biological differences between the population from the
Hawaiian (TD. nawaii) region and others (T. remus Earbados and elsewhere), detailed studies were made on the structure of the male genitalia and the female antennae of the two biological species of Telenomus. T.remus from
Barbados was selected to represent the rentus group. T. solitus from Mexico was also included in this study because, although Johnson (pers. comm.) observed differencs in the digital teeth on the male genitalia between T. remus and T. solitus, they interbred.
3.2.1 Male genitalia.
Ten males from each Telenomus population were removed from the cultures with the aid of a pooter. They were then placed in a test tube with 10% potassium hydroxide. After heating for 5-10 minutes to clear, the specimens were thoroughly washed with distilled water and transferred into a thick watch glass. These specimens were then examined under a binocular microscope and the male genitalia were carefully separated from the gaster with the aid of two fine pins. The separated male genitalia were dehydrated with a series of ethanol (30% to absolute) and xylene before being mounted on a slide. The slides were examined at x400 magnification.
Length and width of the male genitalia were measured using a callibrated eye piece graticule. Of the ten males used the three best specimens from each population were used for measurements. Table 3.1 Summary of the description of Telenomus spp. egg parasltolds associated with Spodoptera spp
T. minutissimus T. solitus T. spodopterae T. remus T. nawaii
Described by W. H. Ashmead (1895) Norman F. Johnson (1983) Alan P. Dodd (1914) G. E. J. Nixon (1937) W. H. Ashmead (1904) Origin Trinidad Gautemala Java Malaysia Japan Host spp. Dactyloplus sp.(Trinidad) Trichoplusla ni Spodoptera sp. S. mauritla S. mauritla (Hawaii) Anicla infecta (Guyana) S. exigua (Mexico) S. litura (Egypt) S. litura (India) S. litura (Fiji) Spodoptera sp. (Dominican republic) Morphology External Length 0.5 mm(female) TL 1.24 - 1.5 mm 0.6 mm (Female) 0.55 mm (male & 0.45 - 0.5 mm (male & TL 1.05 - 1.38 mm female) female)
Colour(female) brown brown brownish brownish dark brown -45 (male) not described yellow not described pale yellow pale yellow
Antenna(female) 11 segments(club 4segs.) llsegs. (clava 5 segs.) not described 11 segs.(club 4 segs) llsegs.(clava 5 segs.) claval formula 1.2.2.1 (male) 12segs. 12segs. not described 12segs. 12 segs.
Head Broad, much wider than smooth, broader than 11111e wider than transverse, wider than wider than the thorax. the thorax the thorax the thorax the thorax Eyes oval, sparsely pilose heavily setose not hairy not very large not hairy
Wings hyaline, ciliated clear, basal vein fore wings are fore wings greyish not described venation 1ight brown weakly pigmented narrow for this hind wing very narrow hind wing narrow genus beyond nervature Internal male genitalia not described large digital teeth not described digital teeth short not described broad aedeagal lobe and broad
TL Total length ( follows Johnson 1983). -46-
3.2.2 Female antenna.
The special structures known as plate sensilla in the clava of the antennae were found to be useful to identify and separate different
Telenomus species (Bin 1981). The structure of the female antennae of these populations was studied under scanning electron microscope/ using the method described by Johnson ( 1984). Ten females were used for each population.
3.3 Results and discussion.
3.3.1 Male genitalia.
The structures of the male genitalia of the three populations used in
this study are shown in Figure 3.1. The length and width (at the widest part) of the male genitalia is given in Table 3.2. The mean (+ SE) length
(0.114 _+ 0.0008 mm) of the male genitalia of T. nawaii is slightly longer
than that of T. remus and T. solitus. But the difference is not
significant (p>.05)(t test n=3).
Among the adult characters Nixon (1937) suggested that the structure of
the male genitalia is useful to identify and separate Telenomus spp. This
character has been widely used by the other taxonomists (Johnson 1984, Bin
1981). Among the Telenomus spp. associated with Spodoptera spp., Nixon
(1937) found that T. remus reared from S. mauritia has short digital teeth.
Johnson (pers. comm.) claims that the digital teeth of Telenomus solitus
reared from Trichoplusia ni are larger than that of Telenomus remus. Fig. 3.1 Structure of the male genitalia (x 400) terminology of the male genitalia follows Snodgrass (1941)
A. T. nawaii B. T. remus C. T. solitus
I a av- I
al-
al aedeagal lobe di - digitus av aedeago-volsellar shaft dt - digital teeth -48-
However findings from the present studies indicate that these three species
(reared on S. littoralis) have short digital teeth in the male genitalia
and therefore the large digital teeth observed by Johnson may be due to the host species, Trichoplusia ni. The external characters of Telenomus
solitus fits very well with T. remus Barbados. Moreover it should also be
noted that E.R. Oatman has collected Telenomus sp. (identified as T.
solitus by Johnson) in Guatemala, where T. remus had already been
introduced by the C.I.B.C. West Indian station in 1977. This creates more
confusion on the identity of T.solitus. Although it was thought that it would be useful to study the male genitalia of these populations reared on
Trichoplusia ni, it has not been tried due to the unavailability of this
host species during the period of this study.
Table 3.2 Size of the male genitalia of the three
Telenomus populations.(n=3)
Parasitoid Length Width Number of
digital
mean + s.e mean + s.e teeth
T. remus 0.112 .0008 0.036 .0002 3 pairs
T. nawaii 0.114 .0009 0.037 .0001 3 pairs
T. solitus 0.112 .0007 0.037 .0002 3 pairs -4 9-
Plate 1 Antennomeres A8-A11 (from left to right)
seen ventrally showing a 5 segmented
clava characterized by claval formula
1 : 2 : 2 : 1 .
PS Plate sensilla X 800
1 Telenomus so 1itus (Mexico)
2 Telenomus remus (Barbados)
3 Telenomus nawaii (Hawaii)
-50-
3.3.3 Female antenna.
The structure of the female antennae of these three species are shown in
Plate I. The terminology of antennae follows Bin (1981). He constructed a formula (claval formula) based on the number of plate sensilla located in the ventral part of the claval segments of the antenna. No morphological differences were observed between these populations. In all the three species used in this study, the female antennae are 11 segmented, clava 5 segmented; claval formula (A11- A8) 1:2:2;1.
Therefore, in this study T. solitus is considered as a _T. remus population from Mexico, as the former is identical in the morphology and readily crossed with T. remus population from Barbados and produced viable offspring (Chapter 6). Although there is no distinct difference in the morphology among these populations, T. nawaii could be separated from the others due to its inability to interbreed with them. In this study materials other than T. nawaii are referred to as T. remus populations of the country they arrived from and the acronyms used are given in Table 2.2. -51-
CHAPTER 4
GENERAL BIOLOGY
4.1 Introduction
Experiences in the past have indicated that many biological control attempts were unsuccessful due to lack of knowledge concerning the biology of the natural enemy. DeBach (1979) wrote that basic research on the biology of a natural enemy of a host insect can be a key to success in any given project and cited many examples. Kot (1979) reported that the longevity, developmental duration and fecundity of the parasitoids are of key importance in the potential of different species to reduce pest populations.
Even populations of parasitoids of the same species from different geographical region are often characterized by biological differences.
Selecting the appropriate races or strains may be important in classical biological control (Messenger et al.1976) because of their high degree of ecological specialization involving climatic tolerances, searching abilities or even host relationships (Simmonds 1963).
In augmentative biological control a knowledge of the developmental duration, longevity and fecundity of the parasitoid is particularly important to the mass production, shipment and pre release preparation. As
Telenomus species and strains have been, and will be, candidates for both classical and augmentative release, studies were made on the developmental duration and longevity of different Telenomus populations and the results are discussed in this chapter -52-
4.2 Materials and methods.
All the experiments (unless specified otherwise) related to developmental duration and longevity were conducted at 25 + 1°C, 70 + 5% relative humidity and a photoperiod of 16:8 hours light:dark.
4.2.1 Developmental time.
The developmental time of each Telenomus population was studied on S. littoralis and S. frugiperda. The developmental time of the two biological species of Telenomus was studied on two additional host species, S. exigua and S. exempt a. Five mated females and 50 fresh host eggs were used for each treatment and replicated three times. Mated females were allowed to parasitize fresh host eggs (laid on filter paper) in Petri dishes.
Parasitized eggs were removed after 24 hours and placed in separate glass tubes measuring 2.5 cm x 7.5 cm for further observations.
The developmental time of the two biological species of Telenomus reared on S. littoralis was studied at five different temperatures, 15, 20, 25,
27, and 30°c. Parasitized host eggs were incubated separately in glass tubes (2.5 cm x 7.5 cm) at the temperatures indicated above. In both experiments the glass tubes were checked daily and the emergence of the parasitoids and their sex recorded.
4.2.2 Longevity
Adult longevity of the males and females of each Telenomus population reared on S. littoralis was studied. The following criteria were tested under longevity tests -53
a) mated males
b) unmated males
c) mated females in groups
d) mated females individually
e) mated females with hosts
Seventy-five individuals were used in each category except for females with host eggs, where thirty individuals were used. The parasitoids were
reared individually in 5 cc ventilated glass tubes and provided with honey.
The longevity of the females in a group was studied by rearing 15 females
in 2.5cmx7.5cm glass tubes. This trial was replicated 5 times. The
longevity of the females with host species was studied by providing fresh
host eggs daily until the parasitoids died. The number of survivors with
time was recorded and the results were analysed using Analysis of Variance.
Duncan's multiple range test was used to compare the means.
4.3 Results and discussion.
4.3.1 Developmental time.
4.3.1.1 Effect of host species on developmental time.
The developmental time of each Telonornuus population reared on different
host species is shown in Table 4.1. In all the Telenomus populations,
males emerged significantly earlier than the females. The time difference
between the male and female emergence ranged from 6-24 hours. Within each
host species, the developmental time of T. nawaii was significantly
(F= 1861.08, p<0.01) longer than the others (Table 4.2 and 4.3). The mean
developmental t\mes of T- na-uja-ic weve a n d /4 -o Table 4.1 The mean + SE developmental duration of Telenontus populations on S. littoralis and on S. frugiperda • S. littoralis S. frugiperda Parasitoid Sex N X SEN XSE IN male 54 13.79 0.07 46 13.8 0.10 female 93 14.42 0.10 96 14.46 0.12 TEB male 36 10.32 0.08 44 10.68 0.09 female 107 11.05 0.10 86 11.37 0.06 TRT male 31 10.22 0.06 43 10.51 0.14 female 88 11.19 0.11 83 11.52 0.09 TRI male 53 10.23 0.08 42 10.48 0.10 female 87 11.26 0.10 101 11.37 0.09 TRP male 34 10.05 0.10 36 10.8 0.12 female 89 11.24 0.12 75 11.59 0.13 TFQ 1 male 28 10.34 0.17 39 10.93 0.14 female 75 11.19 0.07 66 11.73 0.07 TRQ 2 male 48 10.32 0.21 52 10.86 0.17 female 91 11.51 0.08 83 11.39 0.07 TPM male 29 10.11 0.11 44 10.45 0.24 female 86 11.29 0.08 87 11.67 0.06 -55- Table 4.2 The overall mean developmental time of Telenomus populations Telenomus Developmental time TRB 10.86b TRT 10.83b TRI 10.84b TRP 10.90b TRQ 1 10.94b TRQ 2 10.89b TRM 10.86b TN 14.10a Means followed by the same letter within each column are not significantly (p<0.05) different (Duncan's new multiple range test). Table 4.3 ANOVA Comparing the developmental times of the eight Telenomus populations reared on S. littoralis and S. frugiperda. Factor DF SSMS F Telenomus (A) 7 107.60 107.60 1861.08 p<0.01 Spodoptera (B) 1 2.50 2.50 299.53 p<0.01 Sex (C) 1 21.10 21.10 2559.90 p<0.01 A X B 7 0.80 0.11 13.04 p<0.01 B X C 1 0.02 0.02 2.80 p>0.05 A X C 7 0.70 0.10 12.00 p<0.01 A X B X C 7 0.07 0.07 1.15 p>0.05 Residual 64 0.53 0.008 Total 95 133.32 -56- and S_. frugiperda respectively. The mean developmental times of all the other populations ranged from 10.66 to 10.84 days on _S. littoralis and 10.95 to 11.25 days on S. frugiperda respectively. Pemberton (1933) observed that the developmental time of T. nawaii reared on S. mauritia ranged from 13-17 days whereas Kamal (1951) reported that T. nawaii took 10-11 days to complete development on S. litura during warmer months in Egypt. However neither of these studies was conducted in controlled environmental conditions and hence it is very difficult to conclude whether the difference in the developmental time observed by these two authors is due to environmental factors or to host species. The developmental times of T. nawaii reared on S. littoralis, S. frugiperda, S. exigua and S.exempta do not differ significantly (Table 4.4 and 4.5). Developmental times for all parasitoid populations except T. nawaii were significantly (F=299.53, p<0.01) longer on S. frugiperda (Table 4.6). The host eggs were measured to find out whether their size could be associated with the delay in development but the results indicated that there was no significant size difference between the eggs of the four host species (Table 4.7 and 4.8). Table 4.4 The mean (+ SE) developmental duration of the two biological species of Telenotnus on four different host spp S. littoralis S. frugiperda S. exigua S. exempta Parasitoid Sex N X SE N X SE NX SE N X SE T. nawaii male 54 13.79 0.07 46 13.80 0.10 44 13.75 0.10 48 13.22 0.11 female 93 14.42 0.10 96 14.46 0.12 88 14.35 0.09 87 14.43 0.08 T. remus male 36 10.32 0.08 44 10.68 0.09 36 10.26 0.09 25 10.34 0.05 ( (Barbados) female 107 11.05 0.10 86 11.37 0.06 93 11.15 0.08 83 11.12 0.12 -58- T&ble 4.5 The overall mean developmental time of T. remus Barbados and T. nawaii on four different Spodoptera species. Parasitoid Host T. rerrtus T. nawaii S. littoralis 10.70b 14.11a S. frugiperda 11.10a 14.09a S. exigua 10.71b 14.08a S. exempta 10.67b 14.06a Means followed by the same letter within each oolum are not significantly (p<0.05) different (Duncan’s new multiple range test). Table 4.6 ANCVA Cfcmparing the developmental times of the two biological species of Telenaras reared on four host species. Factor DF SS MS F Telenanus (A) 1 131.17 131.17 5813.62 p<0.01 Spodoptera (B) 3 0.24 0.08 3.54 p<0.05 Sex (C) 1 5.12 5.12 226.90 p<0.01 A X B 3 0.45 0.15 6.90 p<0.01 B X C 3 0.01 0.003 0.13 p>0.05 A X C 1 0.24 0.24 10.45 p<0.01 A X B X C 3 0.09 0.03 1.46 p>0.05 Residual 32 0.72 0.022 Total 47 138.04 -59- Table 4.7 ANOVA: Size of host eggs Factor DF SS MS F ratio host size 3 1.4x10‘? 4.6x10”! 0.511 ns Error 96 8.6x10^ 9.0x10 * Total 99 1.0x1(3 Table 4.8 Mean (+ SE) size (diameter in mm) of the eggs of four Spodoptera spp. Spodoptera spp. Size (in mm) X SE S. littoralis 0.446 0.004 S. frugiperda 0.441 0.002 S. exigua 0.436 0.003 S. exempta 0.439 0.002 -60- 4.3.1.2 The effect of temperature on the developmental time of the two biological species. The rates of development of T. remus (Barbados) and T. nawaii were regressed on temperature and the results are shown in Figs. 4.1 A and 4.1B respectively. Males and females of each Telenomus populations developed successfully from egg to adult emergence over a temperature range of 15°C to 30°C. The rates of development of these two species were significantly correlated with temperature. The rates of development were two and a half times faster at 30°C than at 15°C. The progeny production in both parasitoids was reduced to 51% and 52% (Table 4.9) at 15°C and 30°C compared with an averge of more than 80% at other temperatures. The faster developmental rate with increase in temperature has also been reported for Telenomus podisi Ashmead (Yeargen 1980) and for other hymenopteran parasitoids such as Trichogramma spp. Trichogrammatidae (Yu et al. 1984) and Aphytis spp. Aphelinidae (Rosen & DeEach 1979). Arun Kumar et al. (1984) suggested that reduced rate of development at lower temperatures would be beneficial in the mass production of Telenomus remus and showed that the Corcyra cephalonica Stainton Pyralidae eggs parasitized by T. remus could be stored at 5°C for two weeks without any detrimental effect to the embryo. Table 4.9 The mean +_ SE developmental duration of T. remus (Barbados) and T. nawaii at different temperatures. T. remus (barbados) T.nawaii (Hawaii) Males Females Males Females Temperature ° C NXSE NX SEN X SENX SE 0) 15 21 22.4 0.12 52 23.9 0.12 26 25.1 0.16 43 25.9 0.79 -A 20 41 16.6 0.04 95 17.2 0.14 51 18.2 0.25 90 19.6 0.19 25 48 10.2 0.08 85 11.14 0.01 53 12.2 0.21 91 13.1 0.18 27 46 9.1 0.05 93 10.06 0.16 40 11.15 0.19 94 12.2 0.21 30 25 8.2 0.16 48 8.8 0.13 32 9.4 0.22 44 9.86 0.12 - 6 2 - Figure 4.1 A Effect of constant temperature on the duration and rate of development of T .remus on S . • — • developmental time ma 1 e • -• developmental time female g — B rate of development male □— □ rate of development female regression equation for rate of development; Male Y = - 0.035 + 0.004 X (r = 0.98) Female Y = - 0.04 + 0.005 X (r = 0.99) Both regressions are significant (p<0.001) RATE OF DEVELOPMENT OF RATE DEVELOPMENTAL TIME (IN DAYS) (IN TIME DEVELOPMENTAL TEMPERATURE - 6 4 - Figure 4.1 B Effect of constant temperature on the duration and rate of development of T .n awa i i on S . □ ------□ developmenta 1 male time □ — □ developmental time fema1e • — ^ rate of development ma 1 e •— • rate of development female regression equation for rate of development; Male Y = - 0.03 + 0.004 X (r = 0.99) Female Y = - 0.028 + 0.005 X (r = 0.99) Both regressions are significant (p<0.001) I I a RATE OF DEVELOPMENT OF RATE DEVELOPMENTAL TIME (IN DAYS) DAYS) f (IN TIME DEVELOPMENTAL 6 6 o 6 6 A N) N) CTl 00 O N> H ;mperature -66- Longevity. 4.3.2.1 Males. The mean longevities of mated and unmated males are given in Table 4.10. In all the populations, the mated males lived for a very short period (1-3 days) compared with unmated ones which lived for a period ranging from 3-10 days. The difference in the longevities of mated males was not significant (F=1.91, p>0.05) between Telenomus populations (Table 4.10a). The shortest lifespan of the mated males may have resulted from continuous mating. Schwartz & Gerling (1974) also found that the mated males lived for a very short period. There was significant (F= 20.02, p<0.01) difference in longevity between the mated and unmated males of each Telenomus population (Table 4.10b). The longevities of unmated males of the T. remus populations from Queensland (TRQ1) and Philippines (TRP) were significantly (p<0.05) greater than the others. It is interesting to note the two T. remus populations from the same country (Queensland) have a significant difference in their longevities. The longest lifespan of the unmated males may facilitate their search for females in the field, -67- Table 4.10 Mean longevity of males (N=75). Parasitoid Mated males Unmated males TRB 1.68 6.83bc TRT 1.42 7.79abc TRI 1.85 7.52abc TRP 2.00 8.32ab TRQ1 1.63 8.72a TRQ2 1.46 7.32abc TPM 1.73 6.57bc TN 1.41 6.43bc Means followed by the same letter within each column are not significantly (p<0.05) different (Duncan's new multiple range test). Table 4.10 a ANOVA longevity of unmated males Factor DF SSMS F ratio Telenomus 7 355.31 50.76 20.02 p<0.01 Error 592 1502.98 2.53 Total 599 1858.29 - Table 4.10 b ANOVA longevity of mated males Factor DF SS MS F ratio Telenomus 7 30.94 4.42 1.91 ns Error 592 1657.87 2.84 Total 599 1688.81 -68- Table 4,11 Mean longevity of females. Parasitoid In groups Solitary With hosts (N=75) (N=75) (N=30) TRB 13.24 12.40b 3.00c TRT 12.27 10.08cd 2.93c TRI 12.37 12.80b 3.17c TRP 12.23 11.95b 4.33b TRQ1 13.65 14.88a 2.57c TRQ2 12.75 11.47b 2.97c TRM 12.38 11.77b 3.17c TN 13.32 11.25c 5.57a Means followed by the same letter within each column are not significantly (p<0.05) different (Duncan's new multiple range test). Table 4.11 a ANOVA Longevity of mated females with host. Factor DF SSMS F ratio Telenomus 7 207. 10 29.59 26.55 p<0.01 Error 232 258.57 1.11 Total 239 465.66 Table 4.11 b Anova Longevity of females reared as individuals Factor DF SS MS F ratio Telenomus 7 1022.4 146.06 15.0 p<0.01 Error 592 5763.23 9.74 Total 599 6785.62 Table 4.11 c ANOVA Longevity of females reared in groups Factor DF SS MS F ratio Telenomus 7 191.00 27.3 1.81 ns Error 592 8903.68 15.04 Total 599 9094.68 ns - not significant -69- 4.3.2*2 Females. The mean longevities of the Telenomus populations reared under different conditions are shown in Table 4.11. The females lived for a period ranging from 4-25 days whether they were reared in groups or as individuals. The longevity of the females reared in groups did not significantly (F=1.81, p>0.05) differ between the Telenomus populations (Table 4.11a). However significant differences (F=15/ p<0.01) were observed in longevity between the populations when reared as individuals (Table 4.11b). T. remus population from Queensland (TRQ1) lived for a significantly longer period (14.88 days) than the others. Among the others the only population that varied significantly was T. reirtus from Trinidad, which lived for a shorter period (10.08 days). Although Schwartz & Gerling (1974) reported that the females of T. remus (from India) reared in groups (at 25°C) lived six times (on average 18 days) longer than the ones reared as individuals, the present studies show that the longevity of the females was not affected by whether they were reared as individuals or in groups. Contrary to the findings of Schwartz & Gerling (1974) some of the T. remus populations (TRQ1, TRI, TRM) used in this study lived for a longer period when they were reared as individuals. Although T. remus populations from Queensland (TRQ2), Philippines, Barbados and T. nawaii lived for a longer period when they were reared in groups, they did not show a six fold increase in their longevity as observed by Schwartz & Gerling (1974). -70- 4.3.2.3 Females with hosts. There were significant differences between the longevity of females which were allowed or denied oviposition (F=26.55, p<0.01)(Table 4.11c). In all the Telenomus populations the longevity of mated females was significantly reduced when they were exposed to hosts. T. nawaii and T. remus from Philippines lived for a longer period than the other Telenomus populations. It has been shown (Powell & Sheppard 1982) that the longevity of Trissolcus basalis Wollastson Scelionidae was several times shorter when they were exposed to its host Nezara viridula (L) Pentatomidae. The same effect has been shown for Diadegma fenestralis (Holmgren) Ichneumonidae (Legaspi 1984). However Orr et al. (1985) found that the females' of another scelionid, Telenomus chloropus Thompson, which were allowed to oviposit freely, did not show any difference in longevity with and without hosts and suggested that this species is less affected by the energy spent on oviposition. -71- CHAPTER 5 REPRODUCTIVE BIOLOGY 5.1 Introduction. It is widely accepted among biocontrol workers (Van Emden 1974, Coppel & Mertins 1970, Murdoch et al. 1985) that an effective parasitoid should have a rapid rate of increase for successful biological control. This character incorporates a short generation time and a high potential fecundity. Therefore knowledge of the reproductive rates of parasitoids is important for evaluating their potential effectiveness as biological control agents. Fecundity has been used in biosystematic studies to separate Aphytis spp. (Rosen & DeBach 1979). The authors suggested that a comparative study on the fecundity of different species on a given host, under uniform conditions, may serve to indicate biological differences between them and may even aid in their separation. They have also commented on the limitations of this character when used in separating different species. Moreover fecundity plays a major role in the mass multiplication of parasitoids for inundative releases. Certain aspects of the fecundity of T. remus (Gerling & Schwartz (1974), Ramdass (1982) and T. nawaii (Pemberton 1933) has been reported. Aside from these reports very little is known about the reproductive biology of Telenomus spp associated with Spodoptera spp. The fecundity, sex ratio and the intrinsic rate of increase of each Telenomus population considered in this study is discussed in detail in this chapter -72- 5.2 Materials and methods. Parasitoids and host eggs were collected from the stock culture maintained in the quarantine room for fecundity experiments. The fecundities of the eight Telenomus populations reared on both S. littoralis and S. frugiperda were studied. As the findings from the cross breeding experiments indicated that there are two distinct biological species (T. nawaii and T. remus), the fecundity of these two species were studied on two additional host species, Sh exigua and J3. exempta. Fecundity was determined by two methods. The experiments were carried out at 25jH°C and 70+5% relative humidity and replicated ten times. 5.2.1 Potential fecundity. Potential fecundity was measured by counting eggs in ovaries and oviducts by dissection. One hundred mated females from each Telenomus population reared from S. littoralis and S. frugiperda were maintained on a diet of honey. They were then mounted in Berlese fluid and covered with a cover slip. The abdomen of the wasp was squashed by tapping the cover slip (Plate II). This was done in batches of ten at regular intervals from emergence for six days. Few females were dissected after fifteen days. The number of eggs in the ovaries was counted with the aid of a binocular compound microscope. The potential fecundity of the two biological species on additional host species was not estimated at regular intervals, instead it was estimated at the age of 48 hours when the parasitoids produce their maximum complement of eggs -73- Plate II A dissected female of Telenomus viewed under a phase contrast microscope . Eggs are stained with acid fuchsin. (X 400) -74- 5.2.2 Actual fecundity. This was determined by providing fresh egg masses of the host daily to a mated female parasitoid until its death. Single layered egg masses were used in this study in order to maintain homogeneity between replicates. Forty-eight hours old wasps were used in these studies as they produce the maximum complement of eggs at this age. Freshly laid eggs of S. littoralis and S. frugiperda were placed in separate glass vials measuring 2.5 cm X 7.5 cm smeared with honey. The mated female wasp of each Telenomus population was introduced individually into the glass vials with the host eggs. The host egg mass was replaced daily with fresh egg masses until the death of the wasp. The parasitized egg masses were kept separately in glass vials for further observation. Spodoptera sp. larvae (ie from unparasitized eggs) were removed from the container soon after emergence, to prevent predation on unemerged parasitoids. The female wasps were dissected after they died to count the number of eggs remaining in the abdomen. The number of eggs parasitized, number of progeny and the sex ratio were recorded. 5.3 Results and discussion. 5.3.1 Potential fecundity. The potential fecundity of each Telenomus population reared from S. littoralis and S. frugiperda is shown in Fig 5.1. The graphs were drawn separately for each population for clarity. In all the Telenomus populations studied, the potential fecundity increased with time for a period of two days from emergence Similar findings were observed in - 7 5 - Figure 5.1 The mean + SE (N = 10) potential fecundity of Te1enomus populations reared on S .litto- ralis ( • ) and S .frugiperda ( □ )with time. The graphs were drawn separately for each population for clarity. A T.remus (Earbados) B T .remus (Trinidad) C T.remus (India) D T .remus (Philippines) E T .remus (Queens land 1 ) F T .r emus (Queensland 2 ) G T .remus (India) H T . nawai i (Hawaii) 180 A B 150 ■ • v f 120 V// I 90 / 60 30 1 | u □ 180 -9 p -9 150 A 1 1 1 t 120 / / / / / / / / / j 7/ J 90 1 60 30 ' ' ■ ■ ■ I ■ I " " l~ ■ I 23456701234567 TIME (IN DAIS) POTENTIAL FECUNDITY POTENTIAL TIM -78- another scelionid parasitoid, Telenomus gifuensis Ashmead (Hidaka 1958). After two days from emergence the fecundity remained almost at a constant level up to a period of six days (measurements were taken only up to this period). Flanders (1943) proposed that egg production in parasitic Hymenoptera was inhibited when females did not encounter hosts. This was not the case in Telonomus populations used in this study, as there was no change observed in the fecundity after 15 days even if they were deprived of hosts. On both host species, the highest and lowest fecundity was recorded in T. remus (Barbados) and T. nawaii (Hawaii) respectively. The results were analysed using a two way Anova (Table 5.1) and the mean differences (Table 5.2) between Telenomus populations were compared using Duncan's multiple range test, which indicates that the fecundity of T. nawaii, varies significantly (p<0.05) from the other populations used in this study. However the mean fecundity of the T. remus populations did not vary significantly. Based on informations from individuals and from literature, Cock (1985) suggested that the presence of T. minutissimus in the Caribbean cannot be ruled out and mentioned that this species is probably less effective than the T.remus strain from Papua New Guinea. Therefore, the highest fecundity recorded in field collected T. remus from Barbados may be due to the mixing of that population with the indigenous populations already present in Barbados. Gadgil & Bossert (1970) predicted that greater availibility of a resource will lead to a greater reproductive effort. Therefore it could be argued that perhaps the diversity (and as a consequence the abundance) of host species in Barbados may have led the parasitoid to evolve with a high fecundity. -79. Table 5.1 ANOVA Comparing the potential fecundity of each Telenomus population on two host spp. Factor DF SSMS F Telencxnus (A) 7 2647.9 378.3 5.56* Spodoptera (B) 1 74.3 74.3 1.09 ns A X B 7 174.5 24.9 0.36 ns Error 144 9794.1 68.0 Total 159 12690.7 * Significant P<0.01 Table 5.2 Mean potential fecundity of Telenomus reared on S.littoralis (SL) and S.frugiperda (SF) Parsitoid SL SF mean TRB 151.3a 149.9a 150.6a TRT 146.2a 147.4a 146.8a TRI 147.0a 148.3a 147.6a TRP 144.7a 142.5a 143.6a TFQ1 146.9a 141.8a 144.8a TRQ2 140.1bc 144.0a 142.0a TRM 142.2ab 142.8a 142.5a TN 138.6c 137.8b 138.1b Mean 144.6 144.4 144.12 Means followed by the same letter within each column are not significantly(p<.05) different(Duncan's new multiple range test). -80- Table 5.3. ANOVA Comparing the potential fecundity of two Biological species reared from four host species. Factor DF SS MS F Spodoptera (A) 3 8218.0 2739.3 57.3** Telenomus (B) 1 2354.5 2354.5 49.25** A X B 3 74.5 24.8 0.5 NS Error 72 3442.8 47.8 Total 79 14089.8 ** Significant P<0.01 Table 5.4 Mean potential fecundity of Telenomus reared from four host species. Host spp T.remus Barbados T.nawaii Mean S. littoralis 151.3 137.8 144.95 a S. frugiperda 149.9 137.3 143.85 a S.exempta 148.8 135.3 142.05 a S.exigua 123.5 115.8 119.65 b Mean 143.32A 131.87B The means followed by the same letter within each column are not significantly (p>0.05) different (Duncan's new multiple range test). -81- In the second experiment the potential fecundity of T. nawaii and T. remus (Barbados) was compared on four different host species. It was found that the potential fecundity of these two populations was significantly (F=57.3, p<0.01) reduced when they were reared on S. exigua. The results are shown in Table 5.3 and 5.4. Highest fecundity for both parasitoids was recorded on S_. littoralis followed by S. frugiperda and S. exempta. Ramdass (1982) showed that the fecundity of T. remus reared on S. exigua and S. litura was significantly lower than when they were reared on Agrotis ipsilon (Hafnagel) Noctuidae. Kamal (1951) reported that T. nawaii was more prolific when it was exposed to Agrotis ipsilon. 5.3.2 Actual fecundity. Actual (lifetime) fecundity is based on the mean number of host eggs parasitized by a single female parasitoid throughout her lifetime. The highest and lowest actual fecundity on both host species, was recorded for TV. remus populations from Barbados and Mexico respectively (Table 5.5 and 5.6). The overall mean fecundity of all the Telenomus populations on S. frugiperda was significantly (F=15.4,p<0.01) lower than on S. littoralis. It is interesting to note that the mean actual fecundity of TRQ 1 (Queensland 1) was significantly higher than TRQ 2. The Telenomus populations, TRQ 1 and TrQ 2 were obtained on S. mauritia and S. litura respectively. Ramdass (1982) showed that the fecundity of T. remus was significantly reduced when it was exposed to S. litura. Therefore the variation in fecundity between the two Queensland populations may have been caused by the host species, S.litura. The significant interaction between the host and parasitoid indirectly indicates the preference shown by the Telenomus populations for different Spodoptera spp. Although both the host species were equally acceptable to the Telenomus populations from Trinidad - 8 2 - Table 5.5 Anova Comparing the actual fecundity of Telecomus on two host species. Factor d . f s . s m . s F Spodoptera 1 1600.0 1 600.0 15.4** Telenorcus 7 16397.3 2342.47 22.5** Tele X Spodo. 7 2604.2 372.02 3.58* Error 144 1 496 1 . 3 103.89 Total 159 35562.8 Table 5.6 Mean (actual) fecundity of Telenomus on two host species and the number of eggs remaining in the ovary after oviposition. £.littoral is S.frugiperda Mean Parasitoid Fecun- Fggs re- Fecun- Eggs re- Fecun- Eggs re- dity maininn dity maini ng dity maining T.r Parbados 141.3a 2.5 145.7a 1.8 143.5a 2.2 T.r Trinidad 138.5a 3.1 137.6a 0.4 138.1a 1.7 T.r India 133.9a 2.7 121.5b 2.1 127.7b 2.4 T.r Philippines 132.8a 3.4 135.7a 1.7 134.3a 2.5 T.r Queensland 1 140.8a 1.8 139.2a 2.2 140.0a 2.0 T.r Queensland 2 133.6a 4.0 123.8d 2.8 128.7b 3.4 T.r Mexico 122.0b 3.2 103.3e 6.7 112.7cd 4.4 T.n Hawaii 125.1b 2.4 110.6de 0.7 117.9c 1.5 Mean (Host) 133.5A 127.2B The means followed by the same letter within each column are not, significantly (p>0.05) different (Duncan’s new multiple range test). -84- and Queensland (TRQ 1), T_. remus populations from Barbados and Philippines preferred the eggs of S. frugiperda for oviposition and laid more eggs. The other four populations (TN, TPM, TPQ 2, TRI) preferred S. littoralis to S. frugiperda and laid more eggs on the former. Although the Telenomus populations from Barbados were reared on S. littoralis in the laboratory, the preference shown by the Barbados populations towards S. frugiperda could be expected as this population arrived from an area where S. frugiperda is abundant. However, this does not explain the preference of the Philippines population. The daily production of progeny by each Telenomus population exposed to S. littoralis and S. frugiperda is represented in Figs. 5.2 and 5.3 respectively. Schwartz & Gerling (1974) reported that an average of 60-70 progeny were produced by one day old T. remus on S. littoralis and the egg prodution declined sharply, yielding 20 progeny or less for each successive day of the female's life. The results from the present studies are in agreement with their findings. However one of the populations from Queensland (TRQ 1) produced more than 90 progeny on the first day. Pemberton (1933) observed that T. nawaii, laid all of its eggs during the first few days and never after the seventh day. It is seen from the Figs. 5.2 and 5.3 that a similar pattern is shown by T. nawaii in the present studies. The overall mean (+ se) ovipositional period was 6.07 + 0.30 days for T. nawaii (Table 5.6 a). This is significantly (F=8.59, p<0.01) longer than the others. Among the other Telenomus populations, the only population significantly varied is T. remus from Philippines which took (mean +_ SE) 4.04 _+ 0.06 days to complete egg laying. Although these parasitoids produce their full complement of eggs after 48 hours from emergence, they never laid the whole amount on the first day even if they were provided with ample hosts to oviposit - 8 5 - Figure 5.2 Mean +_ SE number of progeny produced by Telenomus per female per day on Spodoptera littorali s (N=10). Numbers in parenthesis represent the adult females alive at that particular time. A T .remus (Barbados) B T.remus (Trinidad) C T.remus (Mexico) D T .remus (Philippines) E T .remus (Queensland 1 ) F T .r emus (Queensland 2 ) G T .remus (India) H T.nawaii (Hawaii) MEAN NUMBER OF PROGENY/ FEMALE/ DAY FEMALE/ PROGENY/ OF NUMBER MEAN NUMBER OF DAYS - 8 7 - Figure 5.3 Mean _+ SE number of progeny produced by Telenomus per female per day on Spodoptera frugiperda (N=10). Numbers in parenthesis represent the adult females alive at that particular time. A T . remus (Barbados ) B T . remus (Trini dad) C T .remus (Mexico) D T.remus (Philippines) E T .r emus (Queens land 1 ) F T .r emus (Queens land 2) G T .r emus (India) H T.nawaii (Hawaii) MEAN NUMBER OF PROGENY / FEMALE / DAY -89- Table 5.6 a Overall mean ovipositional period of the eight Telenomus populations• (N=2 0) Parsitoid Mean ovipositional time (in days) TRB 3.22c TRT 2.81c TRI 3.18c TRP 4.04b TRQ1 2.81c TRQ2 2.97c TRM 3.20c TN 6.07a Means followed by the same letter within each column are not significantly(p<.05) different(Duncan's new multiple range test). -90- 5.3.3 Comparing actual and potential fecundities. In all the Telenomus populations the actual fecundity was always significantly (F=19.2, p<0.01) lower than the potential fecundity. The possible reasons for the difference may be due to the following factors. 1) natural egg mortality of the parasitoid 2) some female wasps do not lay their full complement of eggs as shown in Table 5.6. 3) and finally laying more than one egg into a host egg also could be considered to explain the differences between the two fecundities, because it has been reported that T. remus could readily deposit more than one 'egg per host egg and is unable to recognize the parasitized host (Gerling & Schwartz 1974). However Earl & Graham (1984) reported that although T. remus occasionally lays more than one egg per host egg it could discriminate between parasitized and unparasitized hosts. Therefore the differences between the two fecundities may have resulted from any of those behaviours listed above. - 9 1 - Figure 5.4 Relationship between wasp size (body length) and potential fecundity for T .remus (A) and T . nawai i(B ) . Both regressions are insigni ficant (for T.remus, Y = 45.5 + 176.5X, P > . 0 5 , r = 0.329 ; for . nawa ii , Y = 5 3.43 + 140.09X, P >.05/ r = 0.401). FECUNDITY •51 •53 E Z (in I S mm) SIZE (inSIZE ) m m •55 - 2 9 - •57 •59 -93- 5.3.4. Size and fecundity. The relationship between size (body length) and potential fecundity for the two biological species of Telenomus (TRB and TN) is shown in Fig. 5.4. Although significant positive correlations between size and fecundity has been shown in gregarious hymenopteran parasitoids (Pak & Oatman 1982/ Waage & Ming 1984, Takagi 1985), the results from the experiments with Telenomus spp. indicate an insignificant positive correlation between size and fecundity. This suggests that the body size of the female parasitoid is not a good predictor of the potential fecundity for these two Telenomus populations. 5.3.5 Rate of increase. Life tables incorporating the rate of development, reproduction and survival into an intrinsic rate of increase value r have been developed m for several insects (Messenger 1964, Smith & Hubbes 1986). The r value m has been used to differentiate between different Trichogramma spp. (Orphanides & Gonzalez 1971). The intrinsic rate of natural increase (r ) is a measure of the growth rate of a population per female. Age specific life tables were used to calculate the rate of increase (r ) of these populations reared on S. littoralis and S. frugiperda. The experiments were conducted at 25° + 1°C, 75% RH and 16:8 light:dark photoperiod. The F1 progeny were counted and the number of males and females recorded separately (Section 5.2.2). Only female progeny were considered for life table studies. The intrinsic rates of increase (r ) °f these populations were determined, with the aid of a computer, based on the following formula (Southwood 1966): -94- £ e~r-x lx m x = 1 X where x pivotal age in days 1 age specific survival m^ age specific fertility r rate of increase m The rates of increase (r ) for Telenomus populations were used to m ------compare their potential growth in two host species, S. littoralis and S. frugiperda. Table 5.7 shows the r^ for the Telenomus populations on the two different host species (raw data in Appendix I). The overall mean _+ SE values of r for all the Telenomus populations on S. littoralis and S. m ------— ------— frugiperda were 0.422 _+ 0.017 and 0.420 +_ 0.013 respectively. The difference between the host species did not vary significantly (t=0.06, P>0.05). The highest rate of increase was observed in T. remus from Queensland (TEQ 1). No statistical tests were performed to compare the difference between the Telenomus populations, due to insufficient replications. However, the results showed that the r value for T. nawaii was 1.32 times lower than that for T. remus (Barbados). The lower rate of increase in T. nawaii is expected due to it's lower fecundity and the longer developmental time -95- Table 5.7 The intrinsic rate of increase (r ) of the in Telenanus populations on two Spocioptera species. Parasitoids Intrinsic rate of increase (rm > S.littoral is S.frugiperda TRB 0.4432 0.4418 TFT 0.^380 0.4385 TRI 0.4293 0.4174 TKP 0.4220 0.4329 TRQ1 0.4560 0.4550 TFQ2 0.4390 0.4180 TRM 0.4322 0.4187 TN 0.3201 0.3460 Mean 0.422 0.420 Table 5.8 Sex ratio (Males and females) of the inbreeding Telenomus populations resulted from three consecutive generations. S.littoralis S.frugiperda Parasitoid Male Female x 2 Probability Male Female x 2 Probability T.remus Barbados 260 276 0.47 p>.05 226 239 0.36 p>.05 T.remus Trinidad 242 298 7.58 p<.01 219 234 0.49 p>.05 T.remus India 218 261 3.86 p<.05 187 201 0.50 p>.05 T.remus Philippines 194 215 1.07 p>.05 199 207 0.15 p>.05 T.remus Queensland 1 165 195 2.5 p>.05 305 324 0.57 p>.05 T.remus Queensland 2 183 201 0.84 p>.05 213 221 0.16 p>.05 T.remus Mexico 148 163 0.72 p> .05 249 256 0.09 p>.05 T.nawaii Hawaii 192 207 0.53 p>.05 232 248 0.53 p>. 05 -97- 5.3.6 Sex ratio. Since virgin females invariably produce males and mated females normally produce both sexes, the sex of the egg laid by a mated female is determined by whether or not it has been fertilized prior to oviposition. In this study the progeny sex ratio of each Telenomus population was studied on two host species, £>. littoralis and S. frugiperda. The sex ratio of each of these populations was observed for three consecutive generations and the results are shown in Table 5.8. In all the populations reared on both the host species the number of females was higher than the number of males. However, the populations from India and Trinidad reared on S. littoralis 2 were the only populations which varied significantly (X test Table 5.8) from the sex ratio 1:1. The sex ratios of the two biological species were studied on S. littoralis under different host/ parasitoid ratios. The number of host eggs used in this study was 80 and the parasitoid numbers varied from 1 to 32. Although the sex ratio was female biassed when the parasitoid density was 1 the sex ratio shifted towards a male biassed one at the parasitiod density 16 and 32. Hamilton (1967) proposed that a female biased sex ratio should evolve where local mate competition and sib mating are regular features of mating behaviour. Under these conditions females should produce only sufficient males to fertilize all their female offspring but the sex ratio will stabilize at 1:1 with increasing numbers of colonizing females. This hypothesis was supported by data for gregarious parasitic Hymenoptera (Werren 1983) and some solitary egg parasitoids (Waage 1982) where clusters of host eggs are likely to be parasitized by a single parasitoid female. The progeny sex ratio of T. remus Barbados and jP. nawaii were female biased at lower female parasitoid densities. However with increased number of female parasitoids the sex ratio shifted towards a -98- male biased one. At the highest parasitoid density the experimentally obtained sex ratios are above the asymptote level of 0.5 predicted by Hamilton's equation (Figure 5.5) Patch sex ratio (proportion males) of T . r emus ( • ) and T . n a w a i i ( □ ) . Feans + SS are for 5 replicates. The solid line X •J is the predicted sex ratio curve given by the equa tion of Hamilton ( 1 97S ) for the expected sex ratios of haplodiploid organisms with local mate competition within mating group. • ---- X = Cn-i3C2n— i3/nC*nj SEX_RAT10 £ proportion males] 0-7- UBR FWSS/ EG BATCH EGG / OF WASPS NUMBER 100 I I -101- CHAPTER 6 CROSS BREEDING 6.1 Introduction. Taxonomic separation of the Telenomus spp. associated with Spodoptera spp. is difficult, because of their minute size and the relative scarcity of reliable diagnostic characters (Johnson pers. comm.). Mayr (1963) defined sibling species as "morphologically similar or identical natural populations that are reproductively isolated". Sibling species can be used to test the validity of the biological versus the morphological species concept (Mayr 1963). Such species can be recognized as distinct only through biosystematic research. Hybridization tests are therefore considered to be the ultimate proof of the specific status of closely related populations. Harland & Atteck (1933) suggested that solutions to taxonomic problems involving Trichogramma could be found only by making extensive studies of their biological characteristics, including crossing experiments. Hybridization experiments have been widely used by several workers and proved to be very useful in correcting misconceptions concerning synonymy and determining the evolutionary proximity or distance between different populations of Trichogramma (Fazaludin & Nagarkatti 1971, Nagarkatti & Nagaraja, 1968, Oatman et al. 1968). Similar studies have been conducted with other parasitic Hymenoptera such as Aphytis spp. (Rao & DeBach 1969a, -102- b) and Gryon spp. (Sankaran and Nagaraja, 1975). Although crossing experiments between the Telenomus spp. associated with Spodoptera spp. were conducted by Yaseen et al. (1981)/ they did not study the effects of these crosses on the fertility of the F1 progeny in detail. However they observed that T.nawaii, from the Hawaiian region would not interbreed with T. remus from Papua New Guinea. As there is little difference in the morphology of the different Telenomus populations obtained from various sources attempts were made to make crosses among them. Crossing experiments with the eight Telenomus populations obtained from different parts of the world are discussed in detail in this chapter. 6.2 Materials and method. The sources of Telenomus populations used in this study are listed in Table 2.2. Since these Telenomus populations have arrhenotokous parthenogenetic reproduction, the female progeny of a cross are those derived from fertilized (diploid) eggs and the males are from unfertilized (haploid) eggs. Therefore the percentage of females produced may be taken as an index of reproductive compatibility or incompatibility between the two parental sexes. Cultures of the different Telenomus populations were maintained on Spodoptera littoralis eggs at 25°+1°C and 70%+5% relative humidity. Experiments were conducted at the same temperature and humidity. -103- Crossing experiments were conducted as follows: The eggs of S.littoralis were exposed to different Telenomus populations, and the parasitized eggs were carefully separated from the egg masses without damaging the egg shell. The parasitized eggs (Plate 3) of S.littoralis could be easily recognized by their change of colour (cream to black). Separated eggs were then placed individually in 5 cc glass vials provided with honey for the emerging adult. On emergence the adults were sexed, vials with females were separated from those with males. This could be easily done by the shape of their antennae, clavate in females and monoliform in males. Matings between adult parasitoids of Telenomus populations from various sources were accomplished by placing a freshly emerged virgin female of a given population (for instance, T. remus Barbados) with one or two males of another population (for instance T. remus India), in a 5 cc glass vial. This was replicated ten times. The mating time (where mating occurred) between each cross was recorded. Reciprocal crosses (for instance, T. remus Barbados males with T. remus India females) were also performed between these populations. After the mating all the females were released individually into 7.5 cm x 2.5 cm glass vials containing sufficient number of freshly laid eggs of S. littoralis for oviposition until the death of the parasitoids. The total number of progeny and the number of female progeny were recorded in the F1 generation. The F1 individuals were selfed and their ability to produce viable offspring in the F2 generation was also tested. Records were taken on the size and potential fecundity of the females in the F1 progeny using the methods described in chapter 5. Ten females were used in each experiment. _ 104 - -TO Plate III Difference between the parasitized and unparasitized eggs of Spodoptera ^ r a £i s• A Parasitized eggs B Unparasitized eggs -105- After the crossing experiments between the five Telenomus populations (T. remus Barbados, T. remus Trinidad, T. remus India, T. remus Philippines, T. nawaii Hawaii) obtained at the beginning of this study, the Telenomus populations from Hawaii and Barbados were maintained in the laboratory. Since the Telenomus populations, from Queensland and Mexico arrived during the latter part of this study, crossing experiments with these populations were performed with T. nawaii and with T. remus Barbados only. 6.3 Results and discussion. Both males and females are sexually receptive immediately after emergence. Schwartz & Gerling (1974) have shown that in T.remus the males are attracted by an odour produced by the females. The odour appears to be consist of at least three chemicals. They reported that upon approaching a conspecific female, the male soon became excited, fluttered its wings, and displayed some rapid movements. The female responded by ceasing all movements and remained stationary. The male then mounted her dorsum and copulated. Successful matings occurred between all T. remus populations (TRB, TRT, TRI, TRP, TRQ 1, TRQ 2, TrM). The time taken for copulation between different crosses is given in Table 6.1. Where mating occured there was no significant difference (F=1.28, p>0.05) in the mating time between the different crosses. However, it is important to note that T. nawaii did not mate with the other Telenomus populations considered in this study. In all the crosses the females of the other Telenomus populations reject the males of T. nawaii preventing them from mounting by forceful movements of their hind legs. In the reciprocal crosses males of the other Telenomus -106- populations were rejected by the females of T. nawaii in the same manner. Van den Assem et al. (1982) observed a similar type of reaction shown by the females of Mellitoba spp. when they were approached by an alien male. Similar findings on mating behaviour were observed in cross breeding studies with Aphytis spp. (Rao & DeBach 1969a). In their studies attempts were made to induce the females to remain in more or less quiet state to promote mating by an alien male. However the results from their findings were not encouraging, as they obtained very few viable offspring in the F1 generation. In hybridization experiments with Trichogramma spp., the males of some species succeeded in copulating with the alien females (Nagarkatti & Nagaraja 1968). However, despite this successful copulation, the females died within few hours after copulation. The authors speculated that this effect may be due to physiological incompatibility between the species, involved as suggested by Mayr (1963). The percentages of female progeny obtained from crosses in the F1 and F2 generations are shown in Table 6.2. Although limited numbers of females were used in the crossing experiments the absence of any female progeny in the F1 generation, in all the crosses involving T. nawaii, is a clear indication that this species is reproductively isolated from the populations used in this study. In all other crosses except TRB x TKP, TRB x TPM there were more than 50% female progeny in the F1 generation. This is strong evidence supporting their compatibility. 107- Tafole 6.1 Time taken (in s e c o n d s ) for mating b e t w e e n different crosses (Mean of ten replicates). TFB TFT TRI TFP TFQ 1 TFQ 2 TFMTN TRB 8.2 7.6 7.2 8.6 9.0 7.5 7.6 N TFT 7.8 8.6 8.4 7.6 - - - N TRI 7.6 8.1 7.4 8.1 - - - N TFP 7.0 8.1 7.1 8.1 - - - N TFQ 1 8.2 - - - 8.9 - - N TFQ 2 8.0 - - - - 8.1 - N TFM 7.2 - - - - - 8.4 N IN N N NN N N N 8.9 F=1.28, (Df 25,234.') p>0.05 Not significant, -108- cr7 TEE TRT TRI TRF TRQ1 TEC2 TRK TN TEE F1 71 60 6C 63 70.4 50 53 0 F2 52 54 63 57 45 60 61 - TRT F1 61 64 51 71.3 __ _ 0 F2 49 51 52.5 49.6 - - - - TRI F1 rO5o 61 70 73 — _- 0 F2 52 60 52 56 - - - - TEP F1 45 67.3 69 65 — 0 F2 51 51.8 54.6 57.1 - - -- TP 01 F1 70 _• 71.6 — _ 0 F2 52. 5 - - - 61.2 - - - TRQ2 F1 51 —— _ 63 - c F2 57 -- - - 53 - - TP. I' F1 49 —— _ _ _ 76 0 F2 53. 7 - - --- 65 - n t ::- F1 C 0 0 n 0 0 72 r~- r> i* cL ------w r Tab: Hybridization experiments; outcome of crosses between Telenomus populations from different geographic areas Hurrbers represent the percentage female progeny in F^ and F^. 0 Ho hybrids Crosses net attempted -109- Yaseen et al. (1981) observed that T. remus from Papua New Guinea (PNG) would not interbreed with T. nawaii. They also found that T.remus from the Dominican Republic would readily cross with T. remus from Papua New Guinea. Thus, the results from my crossing experiments are in agreement with their findings. Although the Telenomus population from Trinidad is claimed by Muesebeck (in Wojcik et al. 1976) to be T\ minutissimus, the morphological similarities and interbreeding ability with the other populations indicate that they are conspecific. Telenomus solitus (population from Mexico) readily crosses with T. remus population from Barbados. However, Johnson (pers. cornn.) insists that the digital teeth in the male genitalia of T. solitus are significantly larger than those of T. rernus, and therefore the two cannot be considered conspecific. From the cross breeding experiments, the only species that shows complete reproductive isolation from all others is T. nawaii. The morphological differences, however slight, and the other biological characters such as developmental duration and fecundity also help to separate this as a good species. The potential fecundity of the F1 progeny is summarized in Table 6.3. In all crosses where T. rernus from Barbados was used as a female parent there is significant increase in the fecundity of the F1 progeny compared to the mean fecundity of the two parents involved in that particular cross (t test p<.05) (Table 6.4). Hybrid vigour or heterosis may account for this increase in fecundity. Increased fecundity in F1 hybrids was observed in crossing experiments between poulations of same species of Drosophila having different geographic origins (Vetukiv 1954, 1956). He found that the F1 hybrids between the populations were significantly superior to the parental populations. A similar effect in the fecundity of F1 hybrids has also been observed in other parasitic hymenopterans such as Aphytis maculicornis Masi (Kashimudin & DeBach 1964) and Muscidifurax raptor Girault and Sanders Pteromalidae(Leaner 1972). One might argue that it is improper to compare the results obtained in Drosophila with Telenomus as the latter shows arrhenotokous parthenogenesis. However, I believe that comparing the results on fecundity is justified because this is a female character, always showing diploidy -111 THE TRT TRI_ TRP TRQ1 TRC2 TRf TM TOPillL. 151.3 157.4 158.5 153.6 158.ii 155.6 159 M TRT 140.6 146.3 149 149.4 - - - M TRI 143.7 141.5 147 140 -- - M TRP 137.8 145.3 141.6 144 - • - - M '■ TRQ1 149.2 - - - 147.4 -- H m______ TRQ2 145.2 -- -- 142.9 - M TRK 13?.9 - --- - 143.7 M TM M M i'l NM M T.T 138.5 Table 6.3 Hybridization experiments; outcome of crosses between Teleronus populations from different geographic areas, numbers represent the potential fecundity of the f progeny resulted from various crosses. M lie hybrids Crosses not attempted -112- Table 6.4 Comparing the fecundity of F1 with midparent (N=10). (Method adopted from Vetukiv 1956). Crosses Fecundity Midparent F1 X Female X Male A.V* SE SE t value THE X TRT 148.8 1.93 157.4 1.02 2.87" THE X TRI 149.1 2.07 158.5 1.61 2.524 THE X TRP 1^7.7 2.01 153-6 0.72 2.19* TR5 X TRQ1 149.4 2.21 158.4 1.80 2.23* THE X TF.Q2 145.4 2.35 155.6 0.91 2.04- THE X TRM 147.5 1.95 159.1 2.71 2.48* rnr“\ TET X i r. d 148.8 1.98 140.6 1.24 -2.7 TRI X THE 149.1 2.07 143.7 0.77 -1.0° THE X THE 147.7 2.01 137.8 2.69 -2.10 TRC1X THE 148.4 2.21 149.2 1.42 -0.06 TRC2X THE 145.4 2.35 145.2 1.23 -0.07 TEM X TEE 147.5 1.95 138.9 2.87 -1.78 TFT X TRI 146.6 2.17 149.6 1.39 0.65 TRT X TRP 145.1 2.12 149.6 1.24 1.36 TRI X rppT*ilii. 146.6 2.03 141.5 3.72 -0.87 TRI X TR? 145.5 2.17 140.7 1.07 -1.48 TF.P X TFI 145.5 2.17 141.6 6.29 -0.46 TRP X TET 145.1 2.12 145.3 5.19 -0.03 * Significant at 5% level -113- CHAPTER 7 BEHAVIOUR OF THE TWO BIOLOGICAL SPECIES OF TELENOMUS 7.1 Introduction. Host location or host finding is defined as the parasitoid's perception of and orientation to their hosts, from a distance, by responses to stimuli produced or induced by the host or its products (Weseloh 1981). Apart from the host species, the plants where the host species are located have also been found to influence the parasitoid host finding behaviour. Plants provide chemical cues to which parasitoids respond, helping them to locate the appropriate host habitat (Vinson 1976). For example, the response of aphid parasitoids to odours from food plants of their hosts has been shown by Powell & Zhang (1983). On the other hand morphological characteristics and exudates of plants may also be deleterious to host finding by certain parasitoids. Rabb & Bradley (1968) found that the eggs of Manduca sexta (Johanson) were readily parasitized by Telenomus sphingis Ashmead and Trichogramma minutum Riley when they occurred on several plant species such as tomato (Lycopersicon esculentum Miller Solanaceae), "jimsonweed", and pepper (Capsicum annum L. Solanaceae) but were not parasitized to any great extent when they occurred on tobacco (Nicotiana tabacum L. Solanaceae ) plants. The authors suggested that the reduced rate of parasitism on tobacco plants is due to the presence of sticky exudates on the trichomes of the tobacco leaves. Vinson (1976 and references therein) reported on similar interference with parasitism or escape by an otherwise acceptable -114- host because of its location within a plant. Jordan & van Lenteren (1978) reported that Encarsia fonr.osa (Ga’nan) Aphelinidae was unable to control Trialeurodes vaporariorum Westwood Aleyrodidae on hairy cucumber Cucumis sativus L. Cucurbitaceae plants compared to tomato and egg plant Solanum melongina L. solanaceae. Ravel (1978) found that, although T.remus introduced from Trinidad attacked S. exigua eggs laid on beetroot Beta vulgaris L. Chenopodiaceae, castor Ricinus sp. Euphorbiaceae, and some weeds, it failed to attack S. exigua on onion plant, so he suggested that the odour of onion plants might have prevented this parasitoid from attacking the host eggs. The efficiency of the two biological species of Telenomus in attacking Spodoptera spp. eggs laid on different plant species was studied and the details are discussed in this chapter. 7.2 Materials and methods. Experiments were carried out to determine whether the species of host plant on which the eggs of Spodoptera spp. are laid, affects their susceptibility to attack by the parasitoid species. Two plant species (onion and Brussels sprout Brassica oleraceae L. var. gemmifera Zenker) two parasitoid species (T. remus Barbados and T. nawaii) and two host species (S. littoralis and S. exigua) were used in this study. S. exigua and onion plants were included, since findings from Sri Lanka (Ravel 1978) indicated that T. remus introduced from Trinidad failed to attack S. exigua eggs on onion plants. These experiments were rim at 25+1°c in a room lighted with fluorescent lights and with two fluorescent bulbs at about 5 cm directly over the experimental cage. Onion and Brussels sprout plants were grown in separate 10 cm diameter plastic pots. The plants were grown in 3:1 ratio (i.e. 3 onion plants/pot and 1 Brussels sprout plant/pot) in order to -115- maintain equal leaf area. These plants were placed in slightly modified ventilated perspex cages designed hy Scopes et al. (1975) (modification shown in Fig.7.1). Five pairs of moths were released into this cage for oviposition for a 24 hours. The plants with egg masses of Spodoptera were then transferred into another similar cage for experiments with parasitoids. The number of host eggs per egg mass varied in replicates. Two separate experiments were carried out to compare the efficiency of these parasitoids on the host eggs laid on different plant species. 7.2.1 Effect of the plant species in host location. 7.2.2 Efficiency of the parasitoid within host species The treatments for the experiment are as follows; P 1 H 1 Pa 1 P 1 H 1 Pa 2 P 1 H 2 Pa 1 P 1 H 2 Pa 2 P 2 H 1 Pa 1 P 2 H 1 Pa 2 P 2 H 2 Pa 1 P 2 H 2 Pa 2 P 3 H 1 Pa 1 P 3 H 1 Pa 2 P 3 H 2 Pa 1 P 3 H 2 Pa 2 „ 116— Fig. 7.1 Ventilated perspex cage designed by Scopes et al (1975) modified (D.O.Donnel CH3C UK) by providing a sleeve and a permanent door covered with poly- esterine mesh in the front wall. A front door B sleeve C ventilation hole D airline with T junctions E plastic tray -117- Where P 1 : Onion plant (no choice) P 2 : Brussels sprout (no choice) P 3 : Onion and Brussels sprout (choice) H 1 : _S. littoralis H 2 : S. exigua Pa 1: T. remus Barbados Pa 2: T. nawaii Plants with one host egg batch were used in these experiments. Plant species were kept separately and together in the experimental cages for no choice and choice experiments respectively. 25 mated female parasitoids (48 hours old) for each replicate were given experience by allowing them to lay few eggs into Spodoptera eggs placed in Petri dishes before being released into the experimental cage. Each treatment was replicated 6 times. In the first experiment, the number of parasitoids that found the egg masses was recorded at half hour intervals for a period of six hours. The parasitoids were removed from the host eggs every half hour. The cumulative numbers of parasitoids that located the host eggs within that period were compared. Pairwise comparisons were made between treatments using "t" test. 7.2.1.1 Results and discussion. When the parasitoids were released into the experimental cages containing plants with host eggs, the parasitoids started to fly around inside the cage and tried to settle on the cages walls. On some occasions they accidentally settled on the plants. It is not clear what cues they -113- used to locate the host eggs, although previous work with T. remus suggested that it responded to chemicals emanating from the host egg mass (Nordlund et_ al. 1983). Initially both species of parasitoids walked aimlessly, sometimes flying a short distance but with time they started appearing on the plants. When the parasitoids arrived near the host egg mass the behaviour changed suddenly and the movement slowed down or stopped. At the same time they began to vibrate their antennae rapidly. In all the treatments, the number of parasitoids that located the host eggs within six hours was less than 50% of the total number released at the beginning of the experiment. However, in one treatment where a choice of plants was provided, 52% of the total number of T. nawaii located the host eggs. 7.2.1.1.1 Host locating efficiency of the parasitoids. The cumulative mean number of parasitoids that located the host eggs is shown in Table 7.1 and in Figure 7.2. In all the treatments where the plants with egg batches were exposed to the parasitoids separately (no choice), T. nawaii started finding the host egcs more quickly than T. remus on both plants. When a choice of plants was offered, although both parasitoids started locating the host eggs at the same rate for a period of four hours, the the cumulative mean number of T. nawaii that located the eggs was higher than that of T. remus after six hours and as a result at the end of the experimental time the cumulative mean number of T. nawaii that arrived on the eggs was significantly higher than the cumulative number of T. remus (t=2.24, p<0.05). Table 7.1 Cumulative number of parasitoids found the host egg batch after 6 hours (Experiment 7.2.1) Host species Treatment r2 X SE Ri R3 R4 R6 TOO 12 11 13 14 11 9 11.66 0.72 TOB 8 13 16 11 6 12 11.0 1.46 TOQB 13 8 12 9 13 17 12.0 0.73 Spodoptera TRO 7 11 1 7 13 8 8.5 1.75 littoralis TN - T. nawaii TRB 7 9 11 10 6 13 9.33 1.05 TR - T. rerrus TRCB 14 13 12 10 10 12 11.83 0.65 0 - Onion B - Brussels sprout TOO 10 14 10 9 11 10 10.0 0.76 TOB 4 12 10 12 14 13 10.5 1.40 TOOB 9 10 15 12 12 20 13.0 1.63 Spodoptera TRO 8 7 10 9 10 8 8.66 0.49 exigua TRB 8 9 5 9 9 7 7.83 0.65 TROB 10 10 14 12 9 12 11.66 0.75 liable 7.1 a Number of parasitoids found on the egg batches after 6 hours (Experiment 7.2.2) Host species Treatment X SE Ri R2 ”3 R4 R6 TNO 9 8 7 10 11 8 8.83 0.68 Spodoptera TNB 6 8 12 7 10 7 8.33 1.05 IN - T. nawaii littoralis TR - T. remus TRO 6 5 7 8 5 9 6.66 0.66 0 - Onion plant B - Brussels TRB 5 8 4 7 5 7 6.0 0.72 sprout to 0 1 TOO 10 6 11 10 9 12 9.66 0.84 Spodoptera TOB 9 13 10 8 11 5 9.33 1.11 exigua TRO 5 9 4 8 7 9 7.0 0.83 TRB 8 6 8 7 8 7 7.83 0.65 -121 - S • jj t_tO£ajj_s S* Qxigua i------1------r — i------i— 12 . lo s s' 4- 2- 0. 12 10 8- 6 4 2 0 14. 12 10 8 6 4 2 0 TIME ( IN HOURS) The cumulative number of T.reaus and T.nawai i arriving on the host eggs laid on different plant species with time • — § T.reraus (Barbados)- _ ~ T -122- 7.2.1.1.2 Effect of plant species. Table 7.1 shows the cumulative mean number of parasitoids which were found on the host eggs laid on different plant species after six hours. The arrival of parasitoids on the host eggs with time is represented graphically in Figure 7.3. In all the treatments, S. littoralis and S. exigua eggs laid on onion plants were located quicker than when they were laid on Brussels sprout plants by both parasitoids, except in one treatment where S. exigua eggs on Brussels sprout plants were located more quickly than they were on onion plants by T. remus. However, even in this occasion, at the end of the experiment, the number of T. remus finding S_. exigua eggs on onion plants was higher than on Brussels sprout plants. The host eggs are usually laid on the lower surface of the leaves of the plant. However, due to the nature of leaf structure, the eggs on the onion plants are more exposed than those laid on Brussels sprout plants. Therefore the reason for locating the eggs more quickly on onion than on Erussels sprout may be due to the oviposition site on the plant. Although eggs on the onion plants were located more rapidly than on Brussels sprout plants within the experimental time, the cumulative number of parasitoids arriving on the plant species did not significantly vary between the plant species (Table 7.2). A similar effect was also observed in the choice experiments. Table 7.2 Cotiparing the overall mean + se number of parasitoids arrived at the host eggs laid on different plant species after six hours. Factor Treatment mean + se nimber t value Probability of parasitoids BvsO B vs OB OvsOB Brussels 9.66 + 0.70 sprouts (B) 0.027 p > .05 NS Plant Onion (0) 9.70 + 0.73 1.87 P > .05 NS 123 (N=16) Chion & Brussels 1.79 p > .05 NS sprout (OB) 12.0 + 0.55 T. remus Barbados 9.50 + 0.46 Parasitoid 2.24 * P < .05 (N=24) T. nawaii 11.52 + 0.49 S. littoralis 11.0 + 1.33 Best 0.196 P > .05 NS (N=24) S. exigua 10.5 + 1.22 t * Significant at 5% level Figure 7.3 The cumulative mean number of parasitiods arriving on CUMULATIVE NUMBER OF PARASITOIDS different plant species with time. Standard error bars are omitted for clarity. • Onion• —• Brussels•— sprout □ Onion□ and Brussels sprout -124 - -125- As mentioned in the introduction. Ravel (1978) reporting on field studies conducted in Sri Lanka stated that T. remus did not attack eggs of S. exigua on onion, but in the present study both T. remus and T. nawaii successfully parasitized S. exigua eggs on onion plants. However in Ravel's study the eggs were laid by S. exigua that had been reared on onion plants, and in the present study S. exigua was reared on artificial diet. There is evidence that during oviposition female moths deposit substances from their larval host plant in their eggs (Duffey^ 1970, Rothschild 1971) and it could be possible that the presence of onion plant substances in the eggs of S. exigua that has been reared on onion prevents attack by T. remus. Further studies must be made on the chemical composition of eggs of S. exigua reared on different plants and on artificial diet in order to clarify this point. 7.2.1.1.3. Effect of host species. The cumulative number of parasitoids arriving on the host eggs is shown in Figure 7.4. In all the treatments the number of parasitoids finding S. littoralis eggs was higher than for S. exigua, except in a choice experiment where the number of T. nawaii finding S. exigua was higher than for S. littoralis. However, the overall comparison between these two host species, based on the number of parasitoids that found the egg masses, indicated that there was no significant difference between them (t=0.196, p>.05). An increased tendency to find more S. littoralis eggs compared to S. exigua could be due to the greater number of eggs per egg mass and the greater amount of scale covering the egg mass in S. littoralis. It has - 126 Telenomus n a w a i 1 T . remus (Barbados) 12 10 8 6 4 2 0 12 10 8 € 4 2 0 14 12 10 8 6 4 2 0 7 arriving on different host species with time O—□ S.littoralis S . e x i gr u a - 127- been shown that T. remus females responded to chemicals emanating from the abdominal tips of _S. frugiperda (Nordlund et al. 1983) from which the scales covering the egg mass are obtained. Therefore the more scales in S. littoralis may have produced more odour, which in turn may have attracted more parasitoids. Further studies on the chemical composition and attractiveness of the scales of these two host species are necessary to clarify this point. Experiment 2 7.2.2 Efficiency of the parasitoids. 7.2.2.1 Method. As findings from the first experiments indicated that there was no significant difference between the plant species the choice treatment was omitted in the second experiment. In the second experiment, the parasitoids were allowed to remain on the eggs, and the number of parasitoids on the egg mass was recorded at the end of the experiment (i.e. after six hours). The parasitized eggmasses were removed from the plant and kept separately for further observation on the numbers of eggs parasitized and progeny emerged. The number of host eggs per egg mass varied in replicates. Table 7.3 Ihe (mean + se) percentage parasitism of host eggs on different plant species (N=6) S. littoralis S. exigua Onion Brussels sprout Onion Brussels sprout T. remus 91.06 + 3.04 86.23 + 4.12 88.66 + 2.52 90.16 + 3.09 T. nawaii 80.01 + 3.19 82.31 + 2.34 79.71 + 3.02 87.63 + 3.85 Table 7.3 a ANOVA: percentage parasitism of host eggs Factor DF SS MS F ratio Telenomus (A) 1 397.73 397.73 8.69 p<0.01 Spodoptera (B) 1 21.03 21.03 0.45 ns Plant (C) 1 52.21 52.21 1.14 ns A x B 1 18.99 18.99 0.41 ns A X C 1 75.23 75.23 1.64 ns B X C 1 83.79 83.79 1.83 ns A X B X C 1 22.44 22.44 0.49 ns Residual 40 1829.30 45.72 Total 47 2500.45 Table 7.4 a ANOVA: Comparing areas of discovery, a, of the two biological species of Telenornus. Factor DFSS MS F ratio Telenomus (A) 1 0.24 0.24 25.8** p<0.01 Spodoptera (B) 1 0.009 0.009 0.96 ns Plant (C) 1 0.009 0.009 0.96 ns A X 3 1 0.03 0.03 3.22 ns B X C 1 0.007 0.007 0.77 ns A X C 1 0.026 0.026 2.79 ns A X B X C 1 0.014 0.014 1.55 ns Residual 40 0.375 0.0093 Total 47 0.710 ns - not significant -130- 7.2.2.2 Results and discussion. 7.2.2.2•1 Percentage parasitism. The percentage parasitism by T. remus and T. nawaii was used as a parameter to compare the efficiency of these parasitoids. The percentage parasitism by both parasitoids on host eggs on different plant species is shown in Table 7.3. The percentage of Spodoptera spp. eggs parasitized by T. remus was significantly higher than that parasitized by T. nawaii (F=8.69/ p<0.01). However there was no significant difference in the percentage of host eggs parasitized between either of the plant species (F=1.14, p>0.05) or Spodoptera species (F=0.45, P>0.05)(Table 7.3 a). The results observed in the actual fecundity experiments (chapter 5) indicated that the rate of oviposition in T. remus Earbados was higher than in T. nawaii. Therefore the reason for the higher overall percentage parasitism in T. remus may be due to its higher rate of oviposition. 7.2.2.2.2 Area of discovery. An important parameter that is commonly used to compare the efficiency of parasitoids is their area of discovery. This was first proposed by Nicholson (1933) and Nicholson & Bailey (1935). This parameter measures the area effectively searched by the parasitoids during a period of time. Since this study was mainly aimed at the effect of plant species on the parasitoids, the experiment was conducted by allowing a known number of parasitoids (25) to a single egg mass. The area of discovery of these two parasitoids were calculated separately for each replicate, using -131- Nicholsonian equation. 1 N A = — log e -- P N-Na Where, A = effective area of discovery P = density of total number of parasitoids N = initial host density N = Number of hosts attacked cL Since the number of parasitoids foraging on the egg mass was not recorded until the end of the experiment, there is an uncertainty on the actual number of parasitoids searching. Therefore the effective areas of discovery, A, based on the total number of parasitoids (25) was calculated for T. remus and T. nawaii. The area of discovery of T. remus and T. nawaii is shown in Table 7.4. The results were analysed using a 3 way Anova ( Table 7.4 a) and it was found that T. remus had a significantly higher area of discovery than T. nawaii (F=9.57, p<.01). The higher area of discovery of T. remus Barbados may have been associated with its higher rate of oviposition than that of T. nawaii. However, neither the host species nor the plant species have significant effect on the area of discovery of the parasitoids. Table 7.4 The (mean + se) area of discovery of T. remus and T. nawaii (N = 6) S. littoralis S. exigua Brussels Brussels sprout Onion sprout Onion A 0.09 + 0.014 0.109 + 0.015 0.103 + 0.011 0.092 + 0.007 T. remus a 0.382 + 0.04 0.406 + 0.025 0.310 + 0.028 0.346 + 0.022 132- A 0.071 + 0.006 0.068 + 0.009 0.095 + 0.014 0.066 + 0.006 T. nawaii a 0.206 + 0.027 0.205 + 0.03 0.294 + 0.085 0.178 + 0.014 A = Effective area of discovery (based on total number of wasps released N = 25) a = area of discovery (based on the number of wasps searching) -133- The area of discovery, a, based on the number of parasitoids foraging on the host egg mass at the end of the experiment was also compared and found that here again the area of discovery of T. remus was significantly higher than T. nawaii (F = 25.8, PC0.01)(Table 7.4b). Results of individual replicates (Appendix II) indicated that the area of discovery of both parasitoids decreased with increased parasitoid density on some occassions. Hassell and Varley (1969) proposed that the searching efficiency as measured by Nicholson's area of discovery a, did not remain constant with varying parasitoid densities but that it declined at a constant rate, m, the mutual interference constant. It has been reported that scelionid females show aggressive behaviour towards other females while ovipositing on the same host egg mass (Hidaka 1958, Wilson 1961). However previous findings (Pemberton 1933, Gerling & Schwartz 1974) and casual observations in this study suggested that neither T. remus nor T. nawaii do interfere with conspecific females while ovipositing on the same egg mass. Therefore the reduction in area of discovery may be due to the late arrival of some of the wasps, although their contribution towards parasitism is negligible they were included to calculate the per capita area of discovery of the parasitoids. -134 - Table 7.4 b ANOVA: comparing areas of discovery, A, of the two biological species of Telenomus. Factor DF S3 MS F ratio Telenomus (A) 1 0.0067 0.0067 9.57** p<0.01 Spodoptera (B) 1 0.0002 0.0002 0.28 ns Plant (C) 1 0.0005 0.0005 0.71 ns A X B 1 0.0005 0.0005 0.71 ns B X C 1 0.0007 0.0007 1.00 ns A X C 1 0.0010 0.0010 1.42 ns A X B X C 1 0.0020 0.002 2.85 ns Residual 40 0.3000 0.0007 Total 47 0.4116 Table 7.5 a ANODEV Efficiency of the two biological species of Telenomus in attacking the host eggs laid in layers. Source DF Deviance Mean Deviance F ratio Position (A) 2 235.8 117.90 58.90 p<0.01 Spodoptera (B) 3 2.9 0.96 0.50 ns Telenomus (C) 1 0.7 0.70 0.34 ns A X B 6 19.4 3.2 1.60 ns A X C 2 12.0 6.0 3.20 p<0.05 B X C 3 4.5 1.5 0.70 ns A X B X C 6 20.1 3.35 1.63 ns Residual dev. 96 192.2 1.99 ns - not significant -135- Moreover the results from the host finding experiments (where the parasitoids were removed at half hourly intervals) showed that the overall cumulative mean + se number of T. nawaii and T. remus that arrived at the egg batches after 6 hours were 11.52 _+ 0.49 and 9.5 +_ 0.46 (Table 7.1) respectively. However, in the second experiment where the parasitoids were allowed to remain on the host eggs the numbers of T. nawaii and T. remus were reduced to 9.04 +• 0.42 and 6.75 _+ 0.31 (7.1 a) respectively. Since the experimental conditions for both studies were the same, the reduction in the number of parasitoids in the second experiment may be due to the presence of wasps which might have prevented the arrival of new wasps, or the other possibility is that the wasps which arrived at the egg mass may have left the patch due to unavailability of suitable hosts for oviposition. Therefore detail studies on the interference effect is essential to confirm this. Experiment 3 7.3 Effect of the structure of the host egg mass on the oviposition of Telenomus spp. The moths of Spodoptera spp. usually lay their eggs in 1-3 layers and cover them with scales. Therefore the effectiveness of Telenomus spp. egg parasitoids is mainly dependent on how they exploit the host eggs in the lower layer of the eggmass. The success of parasitization of the lower layer eggs was examined to test their accessibility to the parasitoid. Two parasitoid species, T. remus Barbados and T. nawaii and the egg masses of four host species, S. littoralis, S. frugiperda, S. exigua and S_. exempta -136- were used in this study. 7.3.1 Method. Five 48 hour old mated females were exposed to egg masses with three layers. The parasitoids were allowed to parasitize the egg masses in Petri dishes for a period of 24 hours. The parasitized eggs were then removed from the Petri dishes and kept in 2.5 on x 7.5 cm glass tubes for further observation. The larvae emerging from the unparasitized eggs were removed from the tubes. The experiment was replicated 5 times. The number of parasitoid emergence holes in the host eggs was taken as a measure of accessibility to the parasitoid. Since it was difficult to maintain the number of host eggs in replicates the analysis is based on the proportion parasitism. Analysis of deviance (ANODEV), where binomial errors are assumed instead of normal errors, is used to analyse the results. Table 7.5 The proportion of Spodoptera spp. eggs parasitized by T. remus Barbados and T. nawaii in relation to their position (A) Spodoptera spp. vs Position S. littoralis S. frpgiperda S. exigua S. exempta Top layer 0.97 0.94 0.96 0.97 Middle layer 0.89 0.91 0.93 0.92 bottom layer 0.80 0.84 0.77 0.75 (B) Spodoptera vs Telenanus 137 S. littoralis S. fV’ugiperda S. exigua So exempta T. rerrus 0.86 0.87 0.88 0.85 T. nawaii 0.88 0.89 0.83 0.90 (C) Telenanus spp. vs position T. remus T. nawaii Top layer 0.961 0.969 Middle layer 0.89 0.94 Bottom layer 0.81 0.77 -138- 7,3.2 Results and Discussion. Table 7.5 shows the proportion host eggs parasitized by T. remus and T. nawaii, in each layer and on different host species. In all the Spodoptera spp., the proportion of eggs that were attacked by the parasitoids on the top layer was higher than the proportion that was attacked in the lower layers. The results, analysed using ANODEV, indicated that the proportion of eggs parasitized in different layers varied significantly between layers (F=58.9, p<0.01). There was no significant difference in the proportion host eggs parasitized either between the Spodoptera spp. (F=0.5, p>0.05) or Telenomus spp. (F=0.7, p>0.05). The significant (F= 3.11, PC0.05) interaction between Telenomus spp. and the layers indirectly indicated that the proportion of host eggs parasitized in the middle layer by T. nawaii was higher than that were parasitized by T. remus in that position. However, the proportion of host eggs parasitized in the upper and lower layers did not vary between parasitoids. There are contradictory reports on the efficiency of these parasitoids attacking Spodoptera spp. eggs laid in multilayered egg masses. Pemberton (1933) found that T. nawaii successfully parasitized the eggs of S_. mauritia laid in multilayers and observed that this parasitoid was able to penetrate its ovipositor through the gaps between the host eggs to reach the eggs that were laid in the lower layer. Tanada & Beardsly (1953) observed that the presence of scales and several layers of eggs in the eggmasses, prevented S. mauritia being parasitized by T, nawaii, although the scales of S. frugiperda are known to attract T. remus Nixon (Nordlund et. al. 1983). Schwarz & Gerling (1974) reported that T. remus could equally parasitize each layer of the egg masses of S. littoralis. However -139- Braune (1982) claims that the effectiveness of T. remus was limited in large compact egg masses, as it could attack only a lesser proportion of eggs in the lower layer compared to the the upper layer. Findings from the present study indicate that both parasitoids, were not very efficient in attacking the host eggs laid in the bottom layer in a three layered egg mass -140- CHAPTER 8 GENERAL DISCUSSION In this final chapter an attempt will be made to bring out the features of general interest which emerge from this study, and thus place it in perspective. Detailed studies were made on the biology and the crossbreeding ability of Telenomus spp. populations parasitizing eggs of Spodoptera spp. from Barbados, Trinidad, India, Philippines, Mexico, Queensland and Hawaii with the following objectives; 1) to determine the systematic relationship between populations. 2) to study their biological differences. 3) to evaluate their potential for biological control. As discussed in chapter 3, the taxonomy of Telenomus spp. associated with Spodoptera spp. is in considerable confusion. Taxonomic separation of the Telenomus spp. associated with Spodoptera spp. is difficult owing to their minute size and the relative scarcity of reliable morphological characters. Eiosystematics has provided vital clues in solving taxonomic problems related to parasitic hymenopterans such as Aphytis spp. (Rao and DeBach 1969) and Trichogramma spp. (Nagarkatti & Nagaraja 1968). As suggested by Gordh (1977), biological control workers probably adhere to -141- the biological species concept more than do other entomologists, because subtle differences in the genetic constituents of a population can alter the biological attributes of it and thus alter its effectiveness in biological control. Experimental hybridization tests are fundamental to any biosystematic study because they provide a quantitative assessment of the genetic relationships among populations. For instance, hybridization studies combined with careful morphological analysis have resulted in the publication of keys to the new world species of Trichogramma (Nagaraja & Nagarkatti 1973). Hybridization studies were conducted on Telenanus spp. populations from different geographical areas to determine their systematic relationship. Findings from the crossbreeding studies showed that successful mating occurred between all T_. remus populations (TRB, TRT, TRI, TRP, TRQ 1, TRQ 2, TRM) and produced viable offspring in the F1 and F2 generations (Table 6.2). However, it is important to note that T. nawaii did not mate with any of these populations and as a result it produced an all male progeny in the F1 generation (Table 6.3). These findings suggests that T. nawaii is reproductively isolated from the other populations used in this study. Yaseen et_ al. (198T) also observed that T. nawaii from the Hawaiian region would not interbreed with T. remus from Papua New Guinea. The findings from the cross breeding studies along with the biological differences suggest that there are two distinct biological species, _T. nawaii and T. rernus. Despite the successful mating and the production of viable progeny between T. remus and T. solitus Johnson, Johnson (pers. comm.) insisted that the population from Zlexico is a separate species because of the structural difference in the digital teeth of male -142- genitalia. The structure of the digital teeth in the two biological species and the population from Mexico reared on S. littoralis was studied, and it was found that rhe structure of the digital teeth of the male genitalia did not vary between them. It is premature to draw conclusions from these findings since Johnson used Trichoplusia ni as the host species whereas _S. littoralis was used in these studies. Therefore I have not formally synonimized these species because this requires more detailed studies of the morphological characters. The voucher specimens of the Telenomus populations used in this study are vested with C.I.E and with Dr N.F.Johnson and I believe that the biological differences observed in these populations will be useful in future taxonomic studies to clarify their taxonomic status. The biological characteristics such as developmental time, longevity and the fecundity of the eight Telenomus spp. populations were studied and it was found that the population from the Hawaiian region (T. nawaii) had a lower fecundity (Table 5.1) and a longer developmental time (Table 4.2) compared to the other populations. In all the populations mated males lived for a significantly shorter period than the unmated ones. The longevity of females exposed to host eggs was reduced significantly in all the populations. The results on the longevity of males are in agreement with the findings of Schwartz & Gerling (1974). Although the same authors reported that the females of T. remus (from India) reared in groups lived six times longer (on average 18 days) than the ones reared as solitary, the present study shows that the longevity of the females was not affected by whether they were reared in groups or as individuals -143- The fecundity of each population was studied on two host species, S. littoralis and S. frugiperda. The results showed that the potential fecundity of T. nawaii was significantly lower than the other populations on both of them. The potential fecundity of the other populations did not vary significantly among them. However the highest fecundity was recorded for T. remus Barbados. The actual fecundity based on the number of host eggs parasitized by a single female until her death, showed that here again T. remus Barbados had the highest fecundity. The actual fecundities of the T. remus populations from India, Mexico and Queensland (TRQ 2) varied significantly from the other remus populations. It is interesting to note that the two populations from Queensland had a significant difference in fecundity. Ramdass (1982) has shown that the fecundity of T. remus was reduced when it was exposed to S. exigua and to S. litura. Therefore the host species, S. litura may have been responsible for the reduced fecundity observed in TRQ2 as this population was obtained on S. litura. The other Telencmus populations showed similarities in many aspects. However it is worth mentioning the increased rate of oviposition and the longest lifespan of one of the two T. remus populations from Queensland (TPQ 1). Therefore based on the biological differences and the inability to produce viable offspring with other populations, T. nawaii was separated as a good biological species. The crossability and the similarities in the biological characters suggests that all the other populations are the same species. Having established the presence of two biological species, the biological characters and behaviour of these two biological species were -144- studied in detail to compare their efficiencies so as to recommend the most suitable candidate for future biological control of Spodoptera spp. The population from Barbados was used to represent T. remus group. It is widely accepted by many biological control workers that an efficient biocontrol agent should possess a rapid rate of increase and high searching efficiency for successful supression of its host. Among the biological parameters studied T. remus had a higher fecundity and shorter developmental time compared to T. nawaii. The combined effect of the two parameters is that T. rernus has a faster intrinsic rate of increase (r ) than T. nawaii: the rate of increase of T. remus was 1.32 times higher than T. nawaii and hence in this respect T. remus is superior to T. nawaii. The rate of oviposition in T. nawaii was significantly slower than in T. remus and as result the former lived for a longer period when exposed to host eggs. The extended period of longevity of females may be beneficial for T. nawaii in enabling it to find and destroy more Spodoptera spp. eggs in the field. However, the number of egcs laid during the extended period of life was not sufficient to enable it to have a greater impact than T. remus on the host population (Figs. 5.2 and 5.3). Moreover in an integrated pest management programme against Spodoptera spp. it is very likely that insecticides will be one of the components and hence the extended longevity of T. nawaii may be detrimental due to its longer period of exposure to insecticides, whereas a species such as T. remus is less susceptible as it completes oviposition within three days. However field evaluations are necessary to clarify this aspect. The rate of host finding by T. nawaii was greater than for T. remus. This behavioural ability might help T. nawaii to find host eggs -145- comparatively more quickly than T. reraus. However, even so if the encountered egg batch is large the rate of exploitation will be minimal due to its low rate of oviposition. Although the host finding rate of T. remus was significantly slower than T_. nawaii, within the host eggbatch the attack rate was higher than that of the latter due its higher oviposition rate. Both biological species were found to be very effective in attacking single layered egg batches of S. littoralis, S. frugiperaa, S. exempts and S_. exigua. Laboratory findings from this study showed that the percentage parasitism of S. exigua and S. littoralis by these two parasitoids exceeded 80% (Table 7.3) on many occasions. However both the parasitoids become egg limited when a single parasitoid encounters a large egg mass with 800-1000 eggs, which is very common in species like S. littoralis and S. mauritia. As a result, the Spodoptera spp. larvae emerging from the unparasitized eggs may be sufficient to maintain high pest populations and may also reduce the growth rate of the parasitoid population by destroying parasitized eggs encountered within its egg mass. Moreover the eggs of Spodoptera moths are usually laid in 1-3 layers and the structure of the egg mass is also one of the factors which might affect the efficiency of these parasitoids. As observed in chapter 7 the efficiency of both parasitoid species is rather limited in attacking eggs in the lower layer. Therefore, it is unlikely that they will alone have a sufficient impact on the pest population during an outbreak to bring about a collapse. As mentioned in chapter 1, Ravel (1978) reported that T. remus introduced from Barbados failed to attack S. exicua eggs laid on onion plants. However findings from the present study showed that the S. exigua -146- eggs laid on onion plants were successfully parasitized by the T. remus population from Barbados. As discussed in chapter 7 the possible reason for the differences between these observations may have been caused by the methods used to rear S. exigua. Therefore further studies must be made on the chemical composition of eggs of S. exigua reared on different plants and on artificial diet in order to clarify this point. The other important finding from the hybridization experiment is the recognition of positive heterosis in potential fecundity in the F1 hybrids resulting from the crosses where T. remus Barbados was used as the female parent. The occurrence of heterosis in F1 hybrids has been shown in other parasitic Hymenoptera. Legner (1972) showed two types of positive heterosis and an example of negative heterosis in the hybrids resulted from the crosses between different populations of the parasitoids of synanthropic flies. Kashimudin & DeEach (1976) produced hybrids of Aphytis maculicornis which had greater fecundity and produced more progeny than either of the parental population. Ashley et al. (1974), however reported that Trichogramma, hybridized and selected for temperature adaptation, was poorer than unselected parasites in field cage studies tests with Trichoplusia ni on cotton plants. DeBach & Hagen (1964) suggested that practical advantage might be taken of positive heterosis in the biological control technique of inundation, where large numbers of laboratory reared parasitoids are mass released against the insect pest population. Finally the variations and the effects of host species on the biological characteristics between the populations from different geographical areas and the positive heterosis shown in the F1 progeny resulting from the crosses where T. remus Earbados was used as a female parent suggests a need -147- for more detailed studies in this respect. There is plenty of scope for more studies of this kind, preferably broader and in greater depth. These are more interesting if they can be run side by side with observations from the field because the natural conditions are important as they determine evolutionary pressures and direct population change. The theoretical potential of positive heterosis in fecundity indicates further research and the possible expansion of the number of Telenomus strains presently being utilized for mass releases against Spodoptera spp. The possibility of using hybridization technique for production of synthetic strains that are superiorly adapted might be explored in future. Since the behavioural experiments were confined to T. nawaii and T. remus population from Barbados, it was not possible to comment on these characters of the other T. remus populations. Thus, a comparative study on behavioural characters, between populations from different geographic areas might help to find a population with additional desirable characters. For example in addition to the increased rate of oviposition, higher fecundity and the longest lifespan, the population from Queensland (TRQ1) might possess a higher searching efficiency. Among the eight populations tested in this study it is interesting to note that none of the populations showed reproductive compatibility with T.nawaii. Therefore, this type of biological study should be made on more samples, especially from Japan and Fiji, to elucidate distribution and origin of T. nawaii. Therefore, based on the biological parameters tested in this study T. remus Barbados is a better choice than T. nawaii in many aspects and could be recommended for future use. However, the inferior qualities such as the lower fecundity and the longer developmental time of T. nawaii may be compensated for by its quick host finding ability, and it may be better -148- suited in conditions where the size of the host egg masses encountered are small. -149 BIBLIOGRAPHY Alam, M.M. (1979) Attempts at the biological control of major insect pests of maize in Earbados, W.I. Proceedings. Caribbean Food Crops Society 15, 127-135. Arunkumar, D., Divakar, B.J. and Pawar, A.D. 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APPENDIX I Raw data used in section 5.3.5 (Intrinsic rate of increase) S. littoralis S. frugiperda X 1 X m X mx X 0-10 0-10 11 1 66.9 11 1 69.5 TRB 12 0.9 21.8 12 0.7 21.6 13 0.6 10.6 13 0.3 11.1 14 0.2 4.0 14 0.1 2.0 0-10 0-10 mm 11 1.0 63.3 11 1.0 65.1 12 0.9 19.6 12 0.8 23.1 TFT 13 . 0.9 11.88 13 0.7 10.7 14 0.4 3.33 14 0.1 2.3 15 0.3 2.66 — 0-10 0-10 _ 11 1.0 59.8 11 1.0 57.0 12 1.0 14.0 12 0.7 13.1 TRI 13 0.7 10.5 13 0.4 10.1 14 0.5 6.0 14 0.3 0.8 15 0.3 1.3 0-10 0-10 11 1.0 51.9 11 1.0 59.9 12 1.0 14.3 12 0.9 18.8 TRP 13 0.9 14.11 13 0.3 10.5 14 0.5 6.4 14 0.5 8.8 15 0.3 2.3 15 0.2 3.0 16 0.1 1.0 -163- APPENDIX I Ocntd. Raw data used in section 5.3.5 (Intrinsic rate of increase) S.littoralis S.frugiperda X 1 m X 1 m X XX X 0-10 0-10 . 11 1.0 79.7 11 1.0 78.1 IRQ 1 12 - 1.0 22.5 12 1.0 21.6 13 0.9 2.0 13 0.5 5.8 14 0.4 1.0 14 0.3 4.5 0-10 0-10 11 1.0 65.1 11 1.0 55.3 IRQ 2 12 0.9 19.7 12 0.8 13.11 13 0.8 11.8 13 0.6 9.83 14 0.2 1.5 14 0.2 4.0 0-10 0-10 11 1 60.5 11 1.0 54.7 IRM 12 0.9 18.0 12 0.7 19.3 13 0.7 7.85 13 0.6 12.3 14 0.3 2.0 14 0.3 8.33 0-13 _ 0-13 14 1.0 47.8 14 1.0 49.1 15 1.0 15.87 15 1.0 15.1 IN 16 0.8 5.25 16 1.0 7.1 17 0.7 3.8 17 0.7 2.3 18 0.4 1.7 18 0.7 1.8 19 0.3 0.7 19 0.6 0.8 20 0.2 0.3 20 0.2 0.7 Appendix II Raw data for experiments in chapter 7 Table I Raw data for experiment 7.2.2 A) On Spodoptera littoralis T.remus (onion plant) T.remus (Brussels sprout) R, *2 R4 R6 R i "2 *3 R4 R6 p 6 5 7 8 5 9 5 8 4 7 5 7 164 N 153 121 149 157 140 110 119 120 124 107 161 122 N 141 98 144 150 119 108 101 112 100 103 89 a 152 a 0.42 0.33 0.45 0.39 0.38 0.44 0.38 0.34 0.41 0.47 0.58 0.19 N = Nunber of hosts available N = Nunber of hosts attacked a P = Nimber of Parasitoids a = area of discovery Table I contd. A) On Spodoptera littoralis T. nawaii (onion plant) T. nawaii (Brussels sprout) R1 R2 R3 R4 R5 R6 R1 R2 R3 R4 R5 R6 p 9 7 8 10 11 8 6 8 12 7 10 7 N 107 117 88 119 123 142 114 117 128 140 133 102 N a 101 92 73 85 95 108 93 106 113 107 105 80 a 0.32 0.19 025 0.13 0.14 0.18 0.28 0.3 0.18 0.21 0.16 0.13 N = Number of hosts available N = Number of hosts attacked a P - Number of Parasitoids a = area of discovery Table I Oantd, B) Qi Spocjptera exigua T.nawaii (onion plant) T.nawaii (Brussels sprout) - R1 R2 R3 R4 R5 R5 R1 R2 R3 R4 R5 R6 p 10 6 11 10 9 12 9 13 10 8 11 5 N 117 111 102 108 97 105 101 117 121 123 105 103 N 166 a 87 78 91 90 83 80 75 109 96 106 101 100 a 0.14 0.20 0.20 0.18 0.22 0.14 0.15 0.21 0.16 0.25 0.30 0.71 N = Number of hosts available N = Number of hosts attacked P = Number of Parasitoids a = area of discovery Table I contd, B) on Spodoptera exigua T.remus (onion plant) T.remus (Brussels sprout) R i ”2 *3 R4 R6 R i *2 R4 * p 5 9 4 8 7 9 8 6 8 7 8 7 N 70 75 87 107 90 116 109 120 142 138 113 108 •167- N 61 70 67 99 81 108 103 107 109 127 102 98 a a 0.41 0.3 0.42 0.32 0.33 0.28 0.36 0.37 0.18 0.36 0.29 0.35 N = Number of hosts available N = Number of hosts attacked P = Number of Parasitoids a = area of discovery Appendix III Raw data far host finding experiment (7.2.1) Table A) O n Spocfetera exigua T.nawaii (onion plant) T.nawaii (Brussels sprout) Time R R3 R4 *6 R i R2 R3 4 *5 R6 R i "2 0 0 0.0 0 0 0 0 0 0 0 0 0 0 0 0 0.5 0 0 0 1 0 0 0 0 0 0 1 0 1.0 1 1 1 1 0 0 0 0 1 0 1 1.5 2 2 1 1 2 1 1 1 1 1 2 1 2 2 2 1 1 1 2 2 1 2.0 1 2 168 i 1 0 3 1 2 1 3 i 2.5 1 3 1 2 1 1 2 1 3.0 2 2 1 1 1 0 0 1 1 i 1 0 2 3.5 2 1 1 0 0 1 0 2 0 2 2 1 4.0 0 0 0 0 0 0 1 1 1 0 1 2 4.5 0 1 1 1 1 2 1 2 2 2 3 1 5.0 1 1 1 0 2 2 0 1 2 1 0 1 5.5 0 1 2 0 2 1 0 0 0 0 0 0 6.0 0 0 0 0 0 0 0 0 0 Table A Cn Spodcptera exigua oontd, T.remus (cnicn plant) T.remus (Brussels sprout) Time R! ** *3 R4 R6 R 1 R2 "3 R4 0.0 0 0 0 0 0 0 0 0 0 0 0 0 0.5 0 0 0 0 0 0 0 0 0 0 0 0 1.0 1 0 0 0 1 0 0 0 0 0 0 0 1.5 1 1 1 0 1 1 1 2 1 1 1 1 2.0 0 1 1 0 1 1 0 1 1 1 1 1 2.5 2 0 1 1 1 1 1 1 2 3 2 1 3.0 1 1 2 2 1 2 2 1 1 1 1. 0 3.5 2 1 2 1 1 1 1 1 0 1 1 0 4.0 0 0 1 1 1 1 2 1 0 0 2 2 691. 4.5 0 0 0 1 0 1 1 1 0 0 . 0 1 5.0 1 1 1 1 1 0 0 0 0 0 1 0 5.5 0 1 0 2 0 0 0 0 0 1 0 0 6.0 0 1 1 0 2 0 0 1 0 1 0 1 Table B Ch Spodgptera littoralis T. nawaii (Brussels sprout) T. nawaii (onion plant) Time R1 R2 R3 R4 R5 R6 R1 R2 R3 R4 R5 R6 0.0 0 0 0 0 0 0 0 0 0 0 0 0 0.5 2 0 2 1 1 0 0 0 1 0 0 2 1.0 1 0 2 3 0 1 2 1 3 3 1 1 1.5 1 1 2 2 1 1 1 1 3 1 1 0 2.0 2 2 1 2 1 1 1 4 1 2 2 2 -170 2.5 0 0 1 3 3 1 4 1 0 1 2 2 3.0 0 1 0 1 2 0 1 1 0 3 1 1 3.5 2 1 0 1 1 0 1 2 2 2 1 0 4.0 1 1 0 0 0 0 0 0 0 1 0 0 4.5 1 0 2 0 1 0 1 1 0 0 1 0 5.0 2 0 1 1 1 0 0 2 0 0 1 0 5.5 0 1 2 1 0 2 0 0 1 1 1 1 6.0 0 1 0 1 0 0 1 0 0 0 0 0 Table B oontd, CH S. littoralis T.remus (onion plant) T.rerrcus (Brussels sprout) Time *2 R4 R6 R i *2 “ 3 R4 0.0 0 0 0 0 0 0 0 0 0 0 0 0 0.5 1 0 0 0 0 0 0 0 0 0 0 0 1.0 1 3 0 0 1 0 0 0 0 0 0 1 1.5 1 2 0 1 1 2 1 0 1 1 1 1 2.0 1 1 1 1 3 1 1 1 2 1 1 1 171 2.5 1 1 0 2 2 1 1 1 3 1 0 2 3.0 0 1 0 2 1 0 0 2 1 1 0 3 3.5 0 2 0 2 1 0 0 3 1 2 0 1 4.0 0 1 0 1 2 0 1 1 2 1 2 2 4.5 1 0 0 1 1 1 2 0 1 1 1 1 5.0 1 0 0 1 1 1 1 0 0 2 1 1 5.5 0 0 0 0 0 1 0 0 0 0 0 0 6.0 0 0 0 0 0 1 0 1 0 0 0 0