Chapter III. Biological control of Coffee white stem borer, Xylotrechus quadripes Chevrolat

Transcription

1 Chapter III. Biological control of Coffee white stem borer, Xylotrechus quadripes Chevrolat

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3 117 INTRODUCTION Literature is replete with observations that coffee white stem borer, Xylotrechus quadripes, is one of the most serious pests of arabica coffee, which has the potential to destroy about 1686 acres of arabica coffee plantations and about 6,75 tonnes of coffee every year (Anonymous, 2003a). Different methods are employed to control the onslaught of the pest (Anonymous, 2000a). Chemical control (pesticides) method had been practiced in some African countries and in India (Le Pelley, 1968, Schoeman and Pasques, 1993). In spite of it, the success rate was not up to expectations. It is a fact that IPM is a package in which chemical, mechanical and biological methods are included. Biological control methods for coffee stem borer are still in its infancy. The parasitoid, Sclerodermus domesticus, a bethylid appeared to be a prolific breeder in Vietnam and liberation of this parasitoid successfully destroyed the stem borer in small areas (Duport, 1919, 1924). Since the report of Metapelma sp., an ectoparasitoid, (Hymenoptera: Eupelmidae) by Subramaniam, (1941), studies on the natural enemies of coffee white stem borer has received much importance. Le Pelley (1968) cited that there were many important larval parasitoids of the coffee stem borer belonging to the family Bethylidae, Braconidae, Eurytomidae, Evanidae and Ichneumonidae in Vietnam. He has

4 118 also stated "it seems strange that there are so many important borer parasites in Vietnam, and so few recorded from India. As borer has been carefiilly studied in Mysore, it seems unlikely that so many powerful parasites could have been missed". Keeping this in mind, a search was made for parasitoids by many workers. Campyloneurus sp. (Hymenoptera: Braconidae) (Anonymous, 1976) Gastemption sp. (Hymenoptera: Gasteruptidae) (Anonymous, 1984) and Allorhogas pallidiceps (Perkins) synonymous Doryctes strioliger Kieffer (Hymenoptera: Braconidae) (Prakasan et ai, 1986) were recorded. The biology behaviour and culturing technique of ^. pallidiceps were studied by Veeresh et al., (1992). Shylesha et al, (1992) observed the presence of Apenesia sp.. Scleroderma vigilans Westwood, Scleroderma sp. (Hymenoptera: Bethylidae), Doryctes coxalis (Turner) (Hymenoptera: Braconidae) and Eurytoma sp. (Hymenoptera: Eurytomidae) on stem borer. Working on white stem borer in Thailand, Visitpanich (1994) recorded Pristaulacus sp. (Aulacidae), Diastephanus sp. (Stephanidae) and one Ichneumonidae species as parasitoids. Recently, a braconid parasitioid, Iphiaulax sp. (Hymenoptera: Braconidae) was recorded by Venkatesha et ai, (1997). However, attempts to breed these parasitoids in the laboratory and utilize them for biological control of the pest were generally not successful.

5 119 Recent observations recorded on the parasitoid activity in the field at the Central Coffee Research Institute, revealed that A. pallidiceps, Iphiaulax sp. and Apenesia sp. were the most common ones of all natural enemies, but attempts to mass breed these parasitoids were unsuccessful (Anonymous, 2000). In a routine laboratory experiment, a female of parasitoid Apenesia sp was provided with a stem borer grub to study the natiu-e of the parasitoid. The parasitoid laid the eggs on the larvae and multiplied. Hence, studies on the laboratory rearing of this parasitoid were intensified. From the preliminary studies with respect to the searching habit, survival and robust nature, the parasitoid, Apenesia sp. appeared to be a promising candidate for the biocontrol of white stem borer. Furthermore, working on the natural enemies of white stem borer, Veeresh (1993) stated that further studies on the life cycle and percent parasitization oi Apenesia sp. need to be investigated. Nevertheless, the detailed studies on the biology of this parasitoid have not yet been done. Hence, studies on the hfe cycle, sex ratio, longevity, mating behavior, host specificity along with field liberations were made and the results of the same are presented and discussed in this chapter.

6 MATERIALS AND METHODS '.^;in!it3s;;ii<ss3ii:»a i: ;s::^is-:s a»3iais33s:uaia

7 120 MATERIALS AND METHODS A. Material Apenesia sp. collected from the coffee fields formed the material for the present investigations. B. Methods 1. Biology oi Apenesia sp. a. Recording the oviposition sites and number of eggs To study the biology oi Apenesia sp. females collected from the fields were provided with third and fourth instar white stem borer grubs kept in Kum-Kum jars or heera boxes (Fig. 10). The larvae paralyzed by the sting of the parasitoid were observed regularly for the presence of eggs oi Apenesia sp. If eggs were present, the number of eggs laid and place of egg laying on the grub were recorded. Observations were recorded on fifty female parasitoid. b. Incubation period of eggs To understand the incubation period of parasitoid eggs, infested grubs were continuously observed under a ; stereo microscope (Nikon SMZ 10) until the emergence of I instar larvae. The time taken from the paralysis of grub up to the ecolsion of I instar larvae was recorded.

8 121 c. Larval instars The larval instars were determined by studying the shape and size of the larvae and mandibular shape and size. The morphometric data were recorded, for different larval instars. Here also, fifty individuals were used to record the size. d. Sex ratio To study the sex ratio, a gravid female was allowed to lay eggs on third/fourth instar host larvae. The same female after first egg laying and completing development of the generation was again provided with fresh host larvae. At regular intervals, the female was provided with fresh larvae until its death. The number of males and females that emerged from the host larvae was recorded at regular intervals and the same was used to calculate the sex ratio. e. Preference of larvae for oviposition by the parasitoid To study the preference of female a parasitoid to lay eggs on specific instar larvae, studies were undertaken by providing different instar larvae of the stem borer. The site of egg laying and the number of eggs laid on different instars were recorded.

9 122 f. Longevity To observe the longevity, 10 mated males and females of Apenesia sp. were provided with stem borer larvae as food at regular intervals. The number of dead ones was recorded regularly, until the complete elimination of the experimental parasitoids. The experiments were repeated five times. g. Mating behavior In order to understand mating behavior, the male and female pupae were placed in plastic container. The males that emerged first were observed for their behavior and the mating behaviour was observed after the emergence of the females. Fifty pairs were observed to record the mating behavior. h. Host specificity and maternal care To study the host preference of the parasitoid, it was provided with larvae of different species of insects. They included white stem borer, X. quadripes, another Cerambycid, X. subscutellatus. Cockchafer Holotrichia sp.. Silkworm Bombyx mori, and Corcyra sp. The number of eggs laid on each species larva was recorded. In order to understand maternal care, the females were observed for their behavior after egg laying and during larval development at regular intervals.

10 Studies on searching behaviour and parasitism in laboratory and field Parasitisation and searching ability of the parasitoid are important to understand the efficiency of the wasp in infecting the host population. In order to understand it, the following experiments were conducted. i) Release of parasitoids on infested plants Seven year old, borer infested arabica coffee plants C. arabica (variety: Cauvery) were uprooted from the field and roots were trimmed. These plants were planted in earthen pots filled with soil. The plants were watered every day. Eight plants were maintained and two parasitoids were released per plant. Observations on searching and parasitism were recorded by splitting open the stem and tracing the tunnels made by the parasitoid. Numbers of dead white stem borer larvae, parasitized larvae or empty pupal cases were recorded to calculate the parasitisation percentage. ii) Release on borer infested arabica cut stems Set -1 In these experiments, borer infested arabica cut stems were used instead of whole plants. Infested arabica coffee stems of about 14 cm length and 4.5 cm diameter were collected from the field. These stems were split longitudinally into two halves to identify the visible borer stages inside.

11 124 After recording the stages, the two halves were joined together firmly using a piece of twine to facilitate periodic observations on the development of the parasitoid and its activity. Fifteen cut stems were distributed into five plastic containers (three each) and three parasitoid females were released per container. The searching and parasitisation behavior of the females were recorded. Set-2 As we could see the entry of parasitoid female through the cut edges of the stem, a new set of experiments were carried out. In these experiments, both the cut halves of stems of 14 cm length were joined together and the cut surfaces were sealed with wax to prevent entry of the parasitoid from cut surfaces. Three such cut stems each, were placed in five plastic containers and three parasitoids were released per container. Observations were recorded periodically to obtain the data on searching and parasitisation behavior. Set-3 In the earlier two experiments, only few larvae were present in a small size stem. Hence in the present experiments, still longer stems with larger diameter were used in order to obtain highfrequencyof larvae. In this set of experiments, infested stems of 30 cm length and 4 cm diameter were placed

12 in rectangular glass jars. Five stems were kept in each jar and five such jars were maintained. The parasitoid was released at the rate of one per stem. The observation on parasitism was recorded every month by splitting open five stems selected at random. Set-4 Ten white stem borer infested arabica coffee stems of size 45 cm and 5 cm diameter were placed inside wooden cages of 60 x 45 x 45 cm size, with two sliding glass windows. Ten such cages were maintained and 10 parasitoids were released per cage. The observations on parasitism and searching behavior were recorded after 5 months. iii) Field study Field parasitism was studied by releasing the parasitoids on white stem borer infested coffee plants. Borer infested plants were selected in a block and parasitoids were released at the rate of 1,2,3,4 and 5 females per plant. Five plants were maintained in each set, and the observations on parasitism and searching were recorded by uprooting the plants and splitting open the main stem at periodical intervals. The tunneling of parasitoid through the tunnel made by the white stem borer, left over of the host larvae after feeding by the parasitoid and the pupal cases of parasitoid were recorded to assess the parasitism. 125

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14 126 RESULTS a. Morphological characters of adults From the parasitised grubs of the stem borer, males and females of Apenesia sp. emerged. These were further multiplied by providing stem borer larvae in the laboratory. These laboratory multiplied males and females were utilized for morphometry and study of other characters using a stereo-binocular microscope. The adult males were winged, shiny metallic black in color (Fig. 1). The antennae were filiform, 13-segmented, and black in color. The thorax was also black in color and the legs were dull white in color. The six-segmented abdomen bore a pair of appendages on the last segment in a dorso-lateral position. The body size of males ranged from 5.0 to 8.58 mm with an average of mm length and mm with a range of 0.86 to 1.72 mm width (Table 1). Contrary to this, the females were wing less and dark brown in color (Fig. 1). The antennae were filiform, 13 segmented and yellowish brown in colour. The females measured mm with a range of 4.29 to 8.58 mm in length and mm with a range of 0.72 to 1.72 mm in width (Table 1). The head, thorax and legs were yellowish brown in color. The abdomen was broad, dark brown in color and almost the size of the head and thorax put together.

15 127 b. Oviposition behaviour The mated female recognized the host larva and crawled all over the dorsum or dorsal surface of the body. It curved its abdomen towards the anterior end of the grub, just above or around the mandible and inserted its ovipositor and paralyzed the host (Fig. 2a). A single female could paralyze 10 to 15 grubs in the laboratory, with an average of 2 to 3/day. If the paralyzed grubs were removed and replaced with fresh ones, the female paralyzed the grubs within 2 to 3 minutes after encounter. Before egg laying the adult female paralyzed the host larvae, probably by injecting some fluid in to the head capsule region. The time taken for paralyzing the host was 2 to 3 minutes after encounter. The females were ready to lay eggs about 8 to 10 days after emergence. Most of the eggs were placed on the lateral side of the host larvae (86.97 %) with some being laid on the dorsal surface (13.03 %). The eggs were placed longitudinally on the host body with the anterior end of the eggs facing towards the head portion of the host (Fig. 2b). The eggs were attached to the host at the anterior end with some sticky substance. The number of eggs laid was more on the posterior segments of the host with the highest being on the eighth segment (23.94 %) (Table 2). The period of egg laying was days with a range of 1 to 5 days. The average number of

16 128 eggs laid was eggs/day with a range of 1 to 15 eggs/day. The mated female could lay on an average eggs in one single clutch. The same female could lay eggs on another larva after the first brood had completed development. A single female could lay eggs in batches (range 1 to 5 batches) during her entire life span on different larvae. Thus, a female wasp during her life span could lay eggs (range of 10 to 123 eggs) (Table 3). It was observed that the number of eggs laid on the host larvae varied according to the size of the host. As the size of host increased, the number of eggs laid was also more (Table 4). For example a 5.72 mm long host larvae had 3 eggs, with the increase in size to mm,, the number of eggs laid increased to and on a host larvae of mm length the number of eggs laid was 35. c. Developmental stages i. Eggs The eggs were elongate oval with the portion attached to the body of the host being slightly concave, and translucent pale white in color. The eggs measured a length of mm (range 0.64 to 0.85) and width of 0.30 ± mm (range 0.29 to 0.36) (Table 5) (Fig.2b). The incubation period was days (range 2 to 5). The pre oviposition period was

17 12.48 ± 0.63 days and the egg laying period was days (Table 6). ii. I instar larva The process of eclosion started from the anterior end of the egg, which was opened by the larva inside. The newly hatched larvae were hyaline, pale yellow in color. The egg cover remained attached on the ventral surface of the larva as a pad. The first instar larvae measured mm in length (range 0.86 tol.l4 mm) and 0.37 ± mm in girth (range 0.29 to 0.43 mm) (Table 5) (Fig. 3b). The head was distinct from the body, being slightly bulged and shaped like a blunt arrow. The segments, though present were indistinguishable. Immediately on hatching the larva made the feeding puncture and attached itself to the body of the host (Fig. 3 a). The mandibles were simple, facing downwards from the sides of the body (Fig. 3c). The larva fed by sucking the body content of the host. The duration of the first instar was 0.94 ± days (range 0.83 to 1.00) (Table 6). iii. II instar larva The second instar larva was similar to the first instar larva in color, with 13 segments, which were clearly visible. 10 pairs of spiracles were present. The body length was 1.68 ± mm (range 1.21 to 2.00) and 129

18 width 0.63 ± mm (range 0.50 to 0.86) (Table 5) (Fig. 4a, 4b). The mandibles were bi-dentate and more or less horizontal in position (Fig. 4c). The duration of this instar was days (range 1 to 2) (Table 6). iv. Ill instar larva During this stage the larva was spindle shaped, distinctly segmented (13 segments) and yellowish in color. The size was mm (range 2.0 to 3.28) in length and 0.92 ± mm (range 0.86 to 1.00) in width (Table 5) (Fig. 5a, 5b). The duration of this stage was 1.88 ± days (range 1.5 to 2) (Table 6). The larva retained the exuvia as a pad beneath the body. During this stage the mandibles were tri-dentate (Fig. 5 c). V. IV instar larva By this stage, most of the host body was eaten with only the head and thorax visible. This instar stage measured a length of mm (range 3.28 to 8.58) and 1.23 ± mm (range 1.00 to 1.43) width (Table 5) (Fig. 6a,6b). The duration of this instar lasted about days (range 2.5 to 3 days) (Table 6). Before pupation, the larva detached itself from the host body (Fig. 6c). The detached larvae then spread out in the container and started spinning the cocoon. In this instar the mandibles were more chitinized and tridentate with prominent spines. The total larval period lasted 7.56 ± days (range 5.83 to 8 days) (Table.6). 130

19 131 vi. Cocoon formation The fully-grown fourth instar larva started producing a silken thread that was used for spinning the cocoon (Fig. 7a). The cocoons were white or dull white in color, oblong in shape and tough with one narrower end attached to the substratum. The size of the cocoon was mm (range 5.0 to 9.87) in length and 2.09 ± mm (range 1.43 to 2.86) in width (Table 5) (Fig. 7b, 7c). The cocoons spun by the males were bigger than that of the females. Cocoon formation took ^^"115 days (range 2 to 4 days) (Table 6). vii. Pre-pupa The pre-pupal stages could be observed when there was shortage of food in the brood. A few larvae were unable to spin the cocoon and they waste their silken threads. In the process of changing in to pre-pupae the larvae stopped moving; a constriction was developed indicating segmentation to demarcate the head, thorax and abdomen. Further development took place inside the cocoon. viii. Pupa The pupal period lasted for days (range 13 to 23 days) (Table 6). The pupae inside the cocoon were pale yellow in color. The male pupa developed wings that were black in color and hence could be easily

20 identified by the black color visible through the cocoon. The female pupa developed dark brown abdomen, which also could be identifiable. The exarate (fi-ee) pupae, of the size of mm length and mm in the width in the male and mm in length and mm in width in the case of the female in length and width of mm in male and mm in female (Table 6) (Fig. 8a, 8b). The total duration of life cycle was days (range 22 to 41 days) (Table 6). ix. Adult emergence and behaviour The males emerged a day earlier than the females. Generally, the developing adults, inside the cocoon were facingfi-eeend of the cocoon and emerges out by tearing the cocoon. After emergence the males started tearing open the female cocoons. The females were ready to mate on the day of emergence. The duration of mating was minutes. A female appeared to mate only once (Fig. 9a, 9b). d. Sex ratio The number of females was always more in any clutch observed. The sex ratio was generally 1 male to 5 females. The sex ratio was found to change between broods of the same female with the first brood having 1: 4, second 1: 3 and third brood 1: 2 (Table 7). 132

21 133 e. Use of alternate host for oviposition. In the laboratory, adult female of Apenesia sp. failed to lay eggs on the silkworm {Bombyx mori) larvae even though they were paralyzed. The parasitoid did not lay eggs on larvae of cockchafer and Corcyra sp. The egg laying was observed on larva of other cerambycids. The adult female completed its development on larvae of Xylotrechus subscutellates with an average egg laying of eggs (3 to 24) (Table 8), where as it was eggs on the larvae of^ quadripes. f. Longevity The female wasps lived for a period of days on an average with a range of 11 to 128 days. In the case of males, the average longevity was days with a range of 5-15 days (Table 9). g. Parental care The adult female, after egg laying, did not leave the vicinity but kept moving around the grub, brushing its antenna and mandible around the eggs. When the young larvae started feeding, the mother moved around the grub and collected all the debris including the fecal matter, which was then kept in one place. During cocoon formation the mother assisted the larvae by dragging away some of them to prevent overcrowding.

22 Study on parasitism in tlie laboratory and field i. Laboratory experiments The following observations were made in the study conducted using potted plants. The female parasitoid exhibited good searching capability when released on coffee plants (Fig. 11). They traveled all over the plant including stem and leaves in search of suitable entry point, which would allow access to the stem borer grubs inside. The female wasp managed to get entry through the weak spots on the main stem like ridges or exit holes made by the stem borer. Entry of the wasp could be confirmed by the presence of the yellowish powder that was thrown out while boring through the tunnels made by the grubs (Fig. 12). This powder was nothing but the excreta and wood powder used by the borer grubs to pack the tunnels. Once the wasp had access to a borer tunnel, it kept making a hollow tunnel through this till it reached a grub (Fig. 13). If the grub was of suitable size, it was paralyzed and the egg laying process started. The mother wasp remained near the brood till the next generation adults had emerged. Later, the host searching process started again. The activity of the wasp depended upon the presence of suitable grubs inside the stem. If a suitable grub was encountered at the start of the search.

23 further tunneling was stopped till the next brood emerged. The wasp traveled through all the tunnels in search of grubs and all the tunnels were searched in 35 to 45 days if no grub was encountered. Activity of the parasitoid inside the stem could be confirmed by the powder coming out of the stem (Fig. 12), presence of their tunnel (Fig. 13), remanent of parasitised grub (Fig. 14a), pupal cases of parasitoid (Fig. 14b) or more number of aduh parasitoids than the numbers released originally. The results of the four sets of laboratory experiments are summarized in Table 10. The data revealed that the parasitoids could kill the host grubs and continue the life cycle. Parasitism up to the level of % could be observed in these experiments (Table 10). ii. Field experiments In the case of field releases at 1, 2, 3, 4 and 5 parasitoids per plant, parasitoid activity could be observed even after five months of release. The observed mortality of white stem borer larvae ranged from 20 to 100 % (Table 11). In this trial, a total of 75 female parasitoids were released and 54 could be recovered after five months. In another bulk trial, 200 white stem borer infested plants were marked and for each plant 10 parasitoids were released. The observations recorded after five months yielded an average mortality of % borer stages in the field. 135

24 136 Table 1. Morphometry (in mm) of male and female ofapenesia sp. (n=50) Sex Length Width Range Mean + SE Range Mean + SE Male Female

25 137 Table 2. Frequency of eggs laid by Apenesia sp. on different segments of the host larvae (n =50) Segments of the host Number of eggs % egg laying 1 Segment Segment Segment Segment Segment Segment Segment Segment Segment Segment Segment Segment

26 138 Table 3. Fecundity of Apenesia sp. Particulars Range Mean + SE Egg laid/female/host 3-35 eggs Eggs laid/day 1-15 eggs Batches of eggs/female 1-5 batches Fecundity/female eggs

27 139 Table 4. Correlation between the size of host larvae and number of eggs laid by Apenesia sp. Length (mm) Host larvae Width (mm) Number of eggs laid

28 140 Table 5. Sizes (in mm) of difterent stages of Apenesia sp. Stages Length Width Range Mean + SE Range Mean + SE Egg I instar n instar m instar rv instar Cocoon Pupae -male Pupae- female ,

29 141 Table 6. Duration (in days) of different stages of Apenesia sp. (n=50) Stages Duration Range Mean + SE Eggs I instar n instar m instar rv instar Total larval duration Cocoon formation Pupal period Total life cycle Pre-oviposition period Egg laying period Incubation period

30 142 Table 7. Sex ratio of Apenesia sp. Sex ratio Particulars Total number of adults obtained Males Females Sex ratio A. General population :4.86 B. Brood with marked -1 brood :4 n brood :3 m brood

31 143 Table 8. Fecundity of Apenesia sp. on different host species larvae Different species larval forms No. of eggs Range Mean + SE Silk worm larvae ~ ~ ~ Cockchafer larvae ~ ~ ~ Conyra sp. ~ ~ ~ X subscutellatus larvae X quadripes larvae

32 144 Table 9. Longevity (in days) of Apenesia sp. Sex No. of individuals Range Mean + SE Male Female

33 145 Table 10. Percentage parasitization of white stem borer larvae by the parasitoid Apenesia sp. in the laboratory Particulars No. of Cut No. of parasite No. of white stem borer No. of parasitised white stem borer % Parasitism stems released larvae present larvae I'^set l"" set "* set "" set

34 146 Table 11. Percentage parasitisation of white stem borer larvae by the parasitoid Apenesia sp. in the filed Month No. of plants observed No. of parasitoid released No. of white stem borer stages No. of parasitoid white stem borer stages % parasitisation 1** month 2"^ month 3"* month ^ month * month

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36 2a. 2b Fig. 2a. Adult searching and egg laying 2b. Host larva with eggs and eggs enlarged view

37 3a. Fig.3a. I instar larvae feeding on host 3b. I instar larva 3c. Mandibles of I instar larva

38 4a. 4b. 4c. Fig. 4a. II instar larvae feeding 4b. II instar larva 4c.Mandibles of II instar larvae

39 5a. 5b. 5c. Fig. 5a. Ill instar larvae feeding on host 5b. Ill instar larva 5c. Mandibles of III instar larvae

40 6a. 6b. 6c. Fig. 6a. IV instars larvae feeding on host 6b. IV instar larva enlarged 6c. Parasite larvae detached from the host

41 7b. Fig. 7a. Cocoon formation 7b. Cocoons

42 8a. ~^^-Female 8b. Fig. 8a. Pupal development 8b. Male and Female pupae

43 9b. e'^i^b«i»^» Fig. 9a. Dragging of female during mating 9b. Mating behaviour

44 Fig. 10. Laboratory multiplication using heera boxes

45 Fig. 11. White stem borer infested plant

46 Entry point of parasitoid Parasitold tunnels Fig 12 Entry of parasitoid in borer infested stems Fig 13 Parasitoid tunnel with in the white stem borer tunnel 14a 14b Fig. 14a. Remanent of head capsule of the host larvae after feeding 14b. Pupal case of parasitoid showing its activity inside the stem

47 DISCUSSION

48 147 DISCUSSION The forest and forest products are being destroyed by insect pests especially the wood boring insects, which demand exploitation of different control measures to save them. Information on the use of biological control agents against timber pests is very meagre. Ants were the first group of insects utilized in an attempt to control insect pests by the biocontrol method (Beeson, 1941). Recent investigations have demonstrated the attempts to exploit some of the parasitoids on coffee white stem borer (Anonymous, 2000). Still, much attention has not been focused on the mass rearing, biology and parasitisation potential. Hence an attempt has been made in the present investigation to study the biology, host preference and parasitisation potentials of the recently discovered hymenopteran Apenesia sp. Apenesia sp., a bethylid wasp exhibits host preference to coleopterans, which is similar with the members of the family Bethylidac; The presence of winged males and apterous females gave us the clear idea of sexual dimorphism in the species (Fig.l). Even when the size variation was considered, it was possible to distinguish males and females Qi Apenesia sp. (Table 1). Males with 6.24 ± 0.19 nwn and females with mm body length demonstrated that the males were smaller than the females. However, Shylesha et al, (1992) working on the morphology oi Apenesia

49 148 sp. observed that both males and females were winged and were of equal size. Further more, Venkatesha et al., (1997) observed that females were usually apterous and males winged. Bridwell (1929) recorded one third of the females are winged and only a small fraction of 1 per cent of the males were apterous in the related species of Bethylid namely, Sclerodermus immigrans and S. macrogaster. On the other hand, Wheeler (1928), discussing the latter species, stated that the winged females and wingless males are rare. Hence, it is very difficult to say at this juncture unequivocally whether bethylid females are apterous and males winged, but without ambiguity one can say that in Apenesia sp. studied during the present investigation, males are winged and females are wingless (Fig. 1). The gravid females of Apenesia sp. exhibited good searching capability. They traveled all over the plant including stem and leaves in search of suitable entry point on the main stem like ridges or exit holes, which would allow access to the stem borer grubs inside. The female wasp managed to get entry through the tunnel made by the stem borer. On par with this. Khan (1990) also demonstrated similar activities oi Apenesia sp. boring through the fibrous plugs, while parasitising Serixia (S. str.) andamanica Gardner. From the extensive observations, it is clear that the place of insertion of the sting for paralyzation of the larvae at the head

50 149 capsule, which is the site of the brain. Thus, it made the host larvae immobile within a short period of time of 1 to 2 minutes. Most of the eggs were laid on the pleural projections of posterior segments of the host larvae (Table 2). It is interesting to note that the higher numbers of eggs were found on eighth segment compared to others (Table 2). Similar observation was made by Khan (1990) in Apenesia sp. parasitising S. andamanica. One to fifteen eggs/day was the range for Apenesia sp. on the coffee stem borer, whereas it was in contrary to 4 to 10 eggs/day by Apenesia sp. on S. andamanica in the Andaman. A maximum of 123 eggs was laid by Apenesia sp. on coffee white stem borer ^ quadripes and 18 eggs on S. andamanica (Khan, 1990) in her life span. Nevertheless, a range from 10 to 123 eggs could be observed in Aspenesia sp. in her life span (Table 3). Even the number of eggs laid by a female during her life span was more on coffee stem borer compared to that on S. andamancia. Allorhogas pallidiceps the other parasitoid of the coffee white stem borer had a fecundity range of eggs per female (Veeresh et al, 1992). Duport (1918), in Vietnam, has recorded up to 25 eggs of Sclerodermus domesticus on mature larvae of white stem borer. This shows that Apenesia sp. with high fecundity on coffee white stem borer is a more suitable parasitoid than A. pallidiceps or S.

51 150 domesticus for use as a biocontrol agent for the management of white stem borer. Size of the host is also important for the parasitisation. Perisierola emigrata a bethyhd deposited only few eggs upon small Ereunetis larvae and an average of almost eight on the larger larvae of Cryptophlebia (Clausen, 1940). In the present investigations also more number of eggs was deposited on the bigger size larvae. For example 3 eggs were laid on 5.72 mm length larvae and 35 eggs on mm length larvae (Table 4). From this it is clear that the females of different parasitoids including Apenesia sp. regulate the number of eggs deposited upon a single host in relation to size. It could be considered as a behavioural pattern to ensure sufficient food for the developing young ones. Length of the eggs oi Apenesia sp. was mm and width was mm (Table 5). Thus the average length and width of the other parasitoids of coffee such as A. pallidiceps (Veeresh et al. 1992) were in the same range. Even the incubation period of the eggs of all parasitoids recorded so far had the same period of 2 to 3 days (Table 6). Apenesia sp. on different hosts also had the same incubation period (Khan, 1990). Different sizes and patterns of mandibles (Fig.3c, 4c, 5c) were identified in different size larval stages. This type of size correlation has been used to

52 151 decipher the different larval stages. Thus 4 larval instars were identified in Apenesia sp. on coffee white stem borer. The first instar larva with mm length had a not so distinct chitinised mandible, which is directed downwards. In the second instar, the mandible is bidentate and horizontal and the larval size is mm in length. The third instar with length of mm is characterized by tridentate mandibles. In the case the of IV instar larvae the size had been increased to mm with much chitinized, tridentate with prominently spinned mandibles (Table 5). Similarly, Khan (1990) also reported 4 larval instars of Apenesia sp. on S. andamanica. First instar required only days to develop into the second instar, whereas the longest duration was observed in the case of the fourth instar with days. The total larval feeding period was days (5.83 to 8 days) (Table 6). Contrary to this the total larval development required 7 to 10 days in Apenesia sp. on S. andamanica (Khan 1990). The cocoons of Apenesia sp. on the host of S. andamanica were closely attached with each other in such a manner that the mass attained a single interlaced structure (Khan, 1990). But this was not seen in the case of Apenesia sp. on coffee stem borer. To spin the cocoon, Apenesia sp. took

53 2.88 ± days on stem borer (Table 6), where as in the case of ^. pallidiceps, it took only a single day (Veeresh et al, 1992). The pupal period lasted for ± 0.27 days (Table 6) in the present investigation. Contrary to this. Khan (1990) has observed a pupal duration of 5 to 9 days for Apenesia sp. on S. andamanica. This indicates that the duration of stages depend on the host species on which the parasite develops. Furthermore, irrespective of host species, the larval feeding period is usually followed by a rather prolonged resting period in the cocoon. Apenesia sp. took a total of ± days to complete the entire life cycle on stem borer X. quadripes (Table 6). Similarly, A. pallidiceps, S. vigilans and S. domesticus, the ecto-parasitoids on coffee required days to attain adult stage from the egg (Veeresh et al, 1992; Veeresh, 1993, Duport, 1918). On the other hand, Metapelma sp., an eupelmid ectoparasitoid on coffee completes life cycle in 32 to 35 days (Anonymous, 1967). The present investigation is the first report to demonstrate the emergence pattern and mating behavior of the parasitoid. The males of Apenesia sp. emerged a day earlier than the females. Thus the sexing of the individual was made easy. A similar observation was made by Venkatesha et al, (1997). It was observed that the first emerged male pierced the female 152

54 153 cocoon and dragged the female out. Soon after dragging, the male mated with the female without any pre mating rituals. The female once mated would not allow other males to mate. The mating duration was found to be ±5.3 minutes (Fig. 9a, 9b) The females predominate numerically in all species of Bethylids on which data relating to the sex ratio have been secured (Clausen, 1940). This ranged from 2 tol ins. domesticus to 5 to 1 in 5. immigram. In Apenesia sp., 5 to 1 sex ratio was observed in the present investigation. However, it was interesting to note that the different broods of a single female had different sex ratios. The first being 4 females: 1 male; second 3: 1 and the third being 2: 1 (Table 7). Decrease in the female population may be because of the development of unfertilized eggs in to males. In conjecture with this, Bridwell (1929) recorded that only males were reared from the eggs laid by the virgin females ofls". immigrans. In A. pallidiceps Veeresh et al., (1992) observed the sex ratio of seven females to one male. It is interesting to note that Apenesia sp. could oviposit only on Xylotrechus sp. larvae and not a single egg was found on other larval fonns. This shows that Apenesia sp. gets the cues only from the Xylotrechus species to lay eggs. Between two species of Xylotrechus, X. quadripes larvae got high number of eggs (Table 8). By this we can infer that X.

55 quadripes is the best host compared to other species larvae. Screening larvae of some other Xylotrechus sp. could throw more light on this aspect. In the present investigation, the males lived for 7.96 ± 0.30 days where as females could survive for ± 3.66days (Table 9). In contrast to this, in another braconid parasitoid of coffee, A. pallidiceps the males lived for 7 to 10 days and females for 34 to 40 days (Veeresh et ai, 1992). On the other hand, a evanid parasitoid of coffee stem borer, Pristaulacus sp. had average longevity of 4.4 ±1.1 days in the case of males and 7.8 ±1.2 days in 154 females (Visitpanich, 1994a). Among the three coffee stem borer parasitoids, the females of Apenesia sp. had higher longevity compared to others. This higher longevity shows the adaptability of the parasitoid to the pest environment on one hand and higher parasitism of the host on the other. Furthermore, it may be one of the important characteristics, which could be utilized to exploit this parasitoid to be used extensively as a biocontrol agent in the coffee ecosystem. Parental care is a prerequisite for the development of the young ones and for continuity of the progeny. In bethylids, maternal care of the developing progeny was exhibited in a number of species (Clausen, 1940). A similar observation was made in the present investigation, where the female scouting from egg to cocoon formation, assisted her progeny.

56 Looking into the extent of parasitism by a parasitoid, one can adjudge the parasitoid as an efficient or inefficient controller of a pest. In the present investigations, Apenesia sp. could parasitise up to % in the laboratory (Table 10) and 20 to 100% in field conditions (Table 11). Contrary to this Khan, (1990) reported that Apenesia sp. caused to % parasitism on the round head borer, Serixia andamanica. Veeresh et al, (1993) observed 19.81% parasitisation of ^. pallidiceps on coffee stem borer % parasitism in the case of Pristaulacus sp. on the same borer was shown by Visitpanich, (1994). Hence, it can be said that among the three parasitoids, Apenesia sp. seemed to be the most important species to regulate the coffee stem borer population. Although, the ultimate aim of any parasitoid or predator is the control of the host, the immediate objective should be its successful establishment in the host environment. Apenesia sp. as observed in the forgoing study, in contrast to others, not only established it-self, but also caused higher mortality of the borer. Further, it laid higher number of eggs on bigger size host. This fact might suggest that, if the releases are made at low host densities, which produce the greatest number of offspring for each parasitoid, then the probability of more successful augmentation of parasitoid and thus the chance of pest suppression could be increased. 155

57 Secondly, the adoption of any control measure should be to manage the pest well below the economic threshold level. From this point of view, Apenesia sp. appears to be a very well suited controlling agent for coffee stem borer. This is because of the fact that the parasitoid is rather easy to collect and to rear successfully in the laboratory. More over, female with high longevity, high egg production, searching behaviour, and apterous nature which is highly adapted to burrow through the packed tunnels of white stem borer can easily be released on the borer infested coffee plants to achieve a greater mortality rate of the pest. This is in turn would assist in reducing the pesticide use and protect the coffee environment and in turn the growth of coffee Industry. 156

58 f ibom«ii1tilmm»>. LMMW ri SUMMARY

59 157 SUMMARY In the Integrated Pest Management system, biological control using parasitoids and predators is an important tool. In the coffee ecosystem, the use of indigenous and exotic natural enemies in the biological control of pests like coffee berry borer and mealybugs have been explored successfully. But in the case of white stem borer X. quadripes attempts to use biocontrol agents are very less. From this point of view, the present work is the first venture in exploring the practical utility of adopting biological control against the coffee white stem borer in India. The observations have revealed that Apenesia sp. is one of the best natural parasitoids available in the field, which has not been studied to under stand its life cycle, mating behavior, sex ratio, longevity, host specificity and efficiency in parasitism. Hence present investigations were under taken. Apenesia sp. culture was maintained in the laboratory by providing the grub of X quadripes. Adult males aiapenesia sp. are identifiable by the presence of wings (winged) whereas the females by the absence of wings (apterous). Thus there is sexual dimorphism in this species. Males are invariably smaller in size with mm length than females with a length of mm. The gravid female search for the host larvae and paralyzes it before egg laying. More number of eggs are laid on latero

60 posterior side of the host larvae. The eighth segment of larvae is the most suitable site for egg laying. The average number of eggs laid is eggs/day with a range of 1 to 15 eggs. The mated female could lay on an average eggs in one single clutch. A single female could lay eggs in batches. Thus a female during her life span, could lay ± 3.06 eggs. A host larva of 5.22mm length can harbor only 3 eggs of the parasitoid. With the increase in the size of the grub the number of eggs laid also increased. For example, if the length of the host larvae was mm, the number of eggs observed was 20.22, whereas, if it was mm, the number of eggs was 35. High density of eggs ensures the parasitoid to completely utilize the host larvae. Furthermore, it is a behavioral pattern of the female to regulate the egg laying depending on the size of the host larva to ensure sufficient food for the developing young ones. The eggs are elongate, oval with the anterior attached to the body of the host larvae being slightly concave and translucent pale white in color. The length of the egg is 0.78 ± mm and width is 0.30 ± mm. This develops into first instar larvae within 5 days. There are four larval instars. These instars can be differentiated by the size and by the presence of different types of mandibles. In the first 158

61 159 instar the mandibles are not so well developed, simple and faced down words. In the second instar, it is bidentate and horizontal in position. The third instar is characterized by tridentate mandibles. On the other hand, even though fourth instar is having tridentate mandibles, these are highly chitinized and with prominent spines. There is a pupal stage and pupa is present inside the cocoon. The total duration of life cycle is days. The sex ratio is generally one male to 5 females. Females can live up to days and males for 7.96 ± 0.3 days. This shows that males are short lived than the females. This is a characteristic of female in that she can parasitise maximum number of host larvae in the life time. In the laboratory, females of Apenesia sp. could not lay eggs on the larvae of different species except Xylotrechus. Even in the genus Xylotrechus, more number of eggs are laid on Xylotrechus quadripes. This shows that Apenesia sp. is having high affinity towards X. quadripes than the other species of Xylotrechus. Thus this can be used effectively to control coffee stem borer. Parasitism up to % could be observed in the laboratory where as it was in the range of 20 to 100 % in the field conditions. Furthermore, the

62 160 parasitoid could be observed even after 5 months after release in the field indicating its survivability. With high percentage of parasitism, high longevity, more egg production, efficient searching behavior, apterous nature and ease to rear in the laboratory, Apenesia sp. appears to be the best suited parasitoid which could be QxploitQd for use as a biocontrol agent for the management oi the dreaded enemy of arabica coffee, the coffee white stem borer.