Proc. Indian Acad, Sci. (Anim. Sci.). Vol. 96, No.3. May 1987. pp. 153-169. Printed in India.

,e

Nutritional modulation of reproduction in two phytophagous insect pests S S KRISHNA Department of Zoology, University of Gorakhpur, Gorakhpur 273009. India Abstract. In Earias vinella (F.) (a major noctuid pest of malvaceous crops) changes in the larval food quality (developing seeds, rnesocarp. epicarp of okra fruit or the entire fruit; ovary of shoeflcwer] in either or both sexes led to pronounced disparities in the reproductive performance of the emerging moths. Highest breeding potential was observed when males and females obtained their nourishment. during postembryonic period. from developing seeds of okra of 0-8 days old and such nutrition must be made available for the caterpillars at least for the first 3 days of their lives to attain enhanced level of reproductive efficiency later as moths. During adult life, a carbohydrate nutrient was mandatory for these mated females to realize full fecundity which got tremendously boosted when the sugar was raffinose. Appreciable increase in ovarian weight with marked improvement in oviposition OCCUlTed in Trlbolium castaneum (Herbst) (a serious tcnebrionid pest of cereals and other plant-derived storedcommodities) when both larvae and adults ate whole flour instead of semolina, both enriched with yeasl. This became further augmented when the flour ingested by these beetles during adult life was previously extracted in 100'\ ethanol and then reinforced with yeast. However. whole wheat flour or yeast alone given as food to such reared adults caused a sharp fall in the egg deposition of females. Number of eggs laid by mated females also differed considerably when their imaginal diet consisted of one of certain selected nutrients. oilseeds or spices-all supplemented with yeast. Egg hatchability was always 100% Keywords. potential.

I.

Earias iittel!a; Tribolium c(/slaneum; larval; adult nutrition; reproductive

Introduction

It is well known that reproduction in insects is influenced by a variety of external and internal factors and, amongst them, nutrition seems to be the most crucial single factor in affecting the total egg output in many ways in a majority of the species (Engelmann 1970). Its profound impact on the breeding potential of the adult can be a reflection of differences in the nourishment acquired by the concerned individual during its immature (larval/nymphal) and/or imaginal stage (Wigglesworth 1960; Johansson 1964; Adiyodi and Adiyodi 1974; Nayar 1977). Study of such effects of nutrition on the reproductive performance of the spotted bollworm. Earias »ittella (F.) (a notorious noctuid pest of malvaceous crops like cotton and okra in the tropics) (Butani and Jotwani 1984) and the red flour beetle, Tribolium castaneurn (Herbst) (a major tenebrionid cereal and other plant-derived stored product pest of cosmopolitan distribution (SokololT 1974; King and Dawson 1977) have been reported in diverse periodicals, during the past little over a decade by this author and his co-workers (Viswapremi and Krishna 1974b, 1975; Krishna ei al 1977; Singh and Krishna 1980, 1983; Narayan Singh and Krishna 1980,1981,1983; Mani et a/1986). Findings from all these contributions are incorporated in this communication so as to constitute one comprehensive body of information. 153

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S S Krishna

2. Materials and methods

Laboratory stock cultures of E. vittella raised on tender seeds separated from 0--8 days old fruits of okra (Abelmoschus esculentus Moench) and of T. castaneum reared on whole wheat flour containing 5% (wjw) powdered yeast (BPC product, Alembic Chemical Works Co. Ltd., Baroda) supplied the required numbers ofnewbom larvae ( < 24 h old) of these insects which served as the starting material for inclusion in the different investigations. Cultures of E. vittella were maintained at temperatures ranging between 25° and 28°C and relative humidity (RH) varying from 80--100% while those of T. castaneum were held within a thermal range spanning between 30° and 3rC and RH values fluctuating between 70% and 80%. All experiments described in this study and related to each of these two species were conducted at the same temperature and humidity ranges as were chosen for maintaining their cultures. These tests were adequately replicated and, wherever desirable, the data were subjected to appropriate statistical analysis (Paterson 1939; Snedecor 1961). For the sake ofconvenience, details of the technical procedures involved in every experiment are separately given below for the two selected pest species. 2.1

E. vittella

In all the trials set up here, single-reared, freshly emerged adult moths of either sex were employed. Pairing, mating and individual oviposition tests were performed in muslin-covered glass containers (70 mm diameter; 90 mm height) each provided with a hanging glass capillary filled with 15% sucrose solution (unless otherwise stated) that formed the food of the adults. A small section of the epicarp portion of okra fruit serving as oviposition substrate was placed at the bottom of the container. In most cases, the daily monitoring of egg laying was limited to the first 4 days of oviposition (the period when mated females in their life time deposited most of their eggs) although in a few experiments it was continued further even up to the death of the females. The hatchability of the eggs was also ascertained. 2.1a Effect of variation in larval nutrition: Three types of experiments were designed to examine this issue. In the first type, the caterpillars were nourished right from the day of their birth on 0--8 days old whole fruit of okra or on its epicarp, mesocarp or seeds removed from it to yield male and female moths which were subsequently single-paired (both sexes belonging to the same food regimen) and the reproductive potential of the females assessed in relation to such nutritional differences. In the second type, the reproductive efficiency of moths associated with larvae allowed to feed only during their first 24 or 72 h of their lives on normal and fresh tender seeds taken out of okra fruit and then shifted to a diet consisting of normal and ripened seeds excised from 9-11 days old fruits was compared with that of counterpart adults when both sexes of reproducing pairs were reared all through on the latter victual or on petroleum ether-extracted tender seeds 72 h after they derived nourishment from normal and tender seeds. Extraction of the seeds in petroleum ether was performed in a soxhlet extraction assembly. For this purpose, 100 tender seeds collected from fresh okra fruits (age as mentioned above) were first properly ground using a mortar and pestle in 25 ml of the solvent. Later, this crude seed extract was wholly transferred to the soxhlet assembly containing the same solvent and subjected to repetitive extractions within

Earias and Tribolium reproduction

155

the apparatus to ensure removal of all ether-soluble constituents. Such extracted seeds were subsequently taken out of the assembly and air-dried to eliminate completely the odour of ether present in them before offering them as diets to the caterpillars. In the third type, okra seed diet-reared newly eclosed males or females were coupled with emerging members of the opposite sex grown on epicarp diet and the breeding capacity of females in these pairs determined. Pairs of individuals reared on the ovary component of shoeflower (Hibiscus rosasinensis L.) or on tender seeds or epicarp of okra fruit were also arranged to serve identical purposes. . 2.1b Effect of variation in adult nutrition: Here male and female moths, developed on okra seed diet and paired as before for determination of the reproductive potential of this pest, were during their adult lives, allowed to imbibe only distilled water or solutions of one of 5 selected carbohydrates (raffinose, sucrose, fructose, galactose and glucose) provided through the hanging glass capillary and replenished daily. Although the concentration of all these sugars was maintained at 15%, the trisaccharide raffinose was tested at 5 different lower strengths (0'1, 0·25, 0-5. 1·0 and 5'0%) as well. 2.1 c Effect a/variation in larval and adult nutrition: Consideration of this aspect led to a comparison of reproductive competency of mated females included in pairs fed daily on fresh 15% solution of glucose and whose larval and pupal lives were raised on okra seeds with those associated with couples ingesting raffinose of similar concentration and developed from individuals reared on epicarp diet. 2.2 T. castaneum This part of the investigation mainly related to determination of (i) ovarian weight and reproductive potential of the female reared and maintained on whole wheat flour or semolina supplemented with 5% yeast (w/w) and (ii) the influence of adult diet on the reproductive potential of the female. For studying the first aspect, 10 larvae procured from thelaboratory culture were collectively permitted to continue their postembryonic development up to pupal stage on whole wheat flour or on semolina enriched with yeast provided in sufficient amounts within glass containers (30 mm diameter; 105mm height) covered at the top with muslin cloth. A male and a female pupa, distinguished on the basis of sexual dimorphic characters akin to those described for T. confusum (Chapman 1918), formed from such group-reared larvae were transferred as a single pair to a small muslin-capped glass vial (10 mm diameter; 50 mm height) consisting of adequate quantity of larval food for their eventual emergence into adults. The eclosed females held on yeast-added whole wheat flour or semolina were sacrificed as virgins in preoviposition state within 24 h of their emergence or as egg laying mateds on the 5th day of their adult life to obtain their ovaries (of one side only) made available by dissections performed on a clean microslide containing minute amounts of distilled water under a stereoscopic binocular microscope. Fresh weight of these ovaries removed from each of such 5 females in the different age groups associated with whole wheat flour or semolina were quickly determined separately on a single pan semi-micro electrical balance and the mean fresh weight of this reproductive tissue in respect of each experimental condition was calculated.

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S S Krishna

For evaluating the reproductive capacity of females reared on these two diets, a male and a female beetle, both associated with the same food and freshly emerged from single-paired pupae were coupled to facilitate mating and subsequent oviposition by the females. These tests were performed in glass vials similar to those described already for accommodation of pupae for their metamorphosis into adults. The diets, given in sufficient amounts and replenished only at the end of 20 days to the paired beetles inside the vials were identical to those fed by them during their larval lives. Oviposition was monitored daily for a period of 40 days since emergence of these insects into adults. The decision to terminate the experiments at the end of an arbitrarily selected 40-day duration,· evidently for practical convenience, was mainly based on the already known facts about the extended longevity of these beetles (Feakin 1976; King and Dawson 1977). The hatchability of the eggs laid by the females was also noted. Investigations into the second aspect entailed utilization of single pairs of newborn male and female insects raised in the laboratory, as in previously described tests, on whole wheat flour supplemented with yeast and provision of one of the following food items fortified with a specified amount of yeast (wjw) during their imaginal lives: (i) normal whole wheat flour with or without 5% yeast or only yeast, (ii) whole wheat flour extracted in distilled water, chloroform, diethyl ether or 100% ethanol (extraction procedure using solvent extraction assembly same as already described earlier with the difference that for each extraction 10 g of flour and 200 ml of a solvent were used) and then supplemented with 5% yeast, (iii) certain carbohydrates [starch, sucrose, glucose (BDH, England), fructose (E. Merck, Germany)] or a protein [vitamin-free casein (leN Pharmaceuticals Inc., USA)] or a lipid [cholesterol (Sisco Research Laboratories Pvt. Ltd., India)], all enriched with 5% yeast, (iv) a mixture of some vitamins of the B group [Glaxo Laboratories (India) Ltd.] consisting of 80 J1.g of thiamin, 30 J1.g of riboflavin, 230 J1.g of niacin, 2 J1.g of calcium pantothenate and 0·63 J1.g or 63J1.g of folic acid per gram of yeast and (v) certain oilseeds, viz cotton seed tGossypium arboreum), ground nut (Arachis hypogea), linseed iLinum usittatisisnum), sesamum (Sesamum indicum), mustard (Brassica campestris) or toria (Brassica napus) or some spices such as cardamom (Ammonum subulatum), cinnamon (Cinnamomum zeylanicum), clove (Syzygium aromaticum), coriander (Coriandrum sativum), black pepper (Peper nigrum) or cuminseed (Cuminum cyminum) each material mixed with 5% yeast. Every diet, before offering it to these tenebrionids for feeding, was powdered and sieved properly so as to maintain uniformity in its particle size. With the exception of fructose which, on account of its hygroscopic nature, was replenished daily, all other foods were renewed only once (on the 10th day) during the experiment. The tenure of these oviposition trials, whose general layout was similar to that described in the previous set of tests, was, however, limited to 20 days following emergence of the adults. As before, egg viability was also recorded. 3. Observations 3.1

E. vittella

A break-up of the oviposition data indicating summarized information about certain specific features of this reproductive activity in this moth in relation to larval rearing

157

Earias and Tribolium reproduction

Table 1. Data relating to length of oviposition period (LOP). peak oviposition day (POD). mean oviposition period (MOP) and mean egg number (MEN)jmated female in E. oittella in pairs reared on whole fruit of okra or on its components. Rearing mediumokra fruit Seeds (separated from fruit) Whole fruit Mesocarp Epicarp

Number of individuals tested

12 10 12 15

LOP· Min

Max

POD··

MOP

MEN

12(2) 3 (1) 6(1)

16(3) 14(3) 12(2) 15(3)

1(83) 1(77) 1(56) 6(18)

14·4 10·1 10·1 12-8

882 625 375 151

9(1)

"Figures in parentheses relate to number of females showing the corresponding minimum (min) and maximum (max) lengths of oviposition period for each rearing medium. ··Figures in parentheses relate to the highest mean number of eggs (adjusted to the nearest integer) 1aid on the corresponding oviposition day by females for each rearing medium.

on whole fruit of okra or on its components is given in table I. A greater proportion of females in pairs reared on seeds or on epicarp have a longer oviposition period than counterparts associated with couples grown on other two diets. The mean daily oviposition reached its peak value on the first day of egg laying itself in all the females except those in pairs raised on epicarp diet where it occurred only on the 6th day of oviposition. The overall mean fecundity was highest in females belonging to seed-reared couples and this was nearly 6 times more than the lowest mean fecundity score recorded for individuals reared on epicarp diet. From the results presented above, one can assume that differences observed in the total length of oviposition period of moths in relation to their rearing media can themselves be responsible as an indirect effect of the dietary variations in the rearing media to produce differences in the total number of eggs laid by the females reared variously. Hence a correct evaluation of the direct effect of the dietary differences in the rearing media on the reproductive potential of these moths will be possible only on the basis of comparison of the total number of eggs laid by the females during the same length of oviposition period. Accordingly, this evaluation was based on the total eggs laid by the females during a period of 10 days when one-half to all of the total number of moths that emerged from each rearing medium completed laying more than 65% of the aggregate number of eggs deposited by them during their oviposition period. The observations (table 2) again clearly indicated the competency of females in seed-reared pairs to lay maximum number of eggs during this stipulated oviposition period. Interestingly, this was significantly different even from the total egg output recorded for individuals raised on whole fruit which also contained its seeds. Having realized that nutrition obtained during larval life from developing seeds is unquestionably superior to that provided by other parts of okra fruit for E. vittella to achieve highest reproductive capability, it was of interest to ascertain the time-bound compulsory requirement for the various seed-resident nutritive constituents for the caterpillar to complete its development and to subsequently function as a sexually mature and efficiently productive adult. Table 3 summarizes the data relating to this aspect of the investigation. Mated females in pairs whose nutrition during the first 3 days of their larval lives was provided by developing seeds and then by ripened seeds of okra were only fit enough to lay significantly greater number of eggs than their

158

S S Krishna Table 2. Number of eggs laid during the first 10 days of oviposition by mated females of E. »ittella reared on whole fruit of okra or on its components*. Rearing medium-okra fruit/components

Number of individuals tested

Mean number of eggs laid

12 5 9 14

675'1 545·0 355·9 119-4

Seeds (separated from fruit) Whole fruit Mesocarp Epicarp Mean square: (1) Treatment (ii) Error F(I~~)

710367·3 4507·2 157·6

"Data subjected to analysis of variance for samples of unequal size (Snedecor 1961).

Table 3. Egg deposition during the first 4 days of oviposition and hatchability of eggs laid by mated females of E. rittella in pairs reared on various okra seed-related dietary regimens (data pooled from 5 females per test)*. Mean number of eggs laid

Mean number of viable eggs deposited

Pairs reared during the first 72 h of their larval lives on normal and tender seeds and later on normal and ripened seeds

267·00 a

227-20 a

Pairs reared during the first 24 h of their larval lives on normal and tender seeds and later on normal and ripened seeds

174·40 b

153'40ab

170'40 b 160·00 b

102·80 b

Mean

192'95

155·70

LSD

(I~~)

105·19

114·74

(5%)

76·34

83-27

Dietary regimen

Pairs reared during the first 72 h of their larval lives on normal and tender seeds and later on petroleum ether-extracted seeds Pairs reared all through on normal and ripened seeds

139·4Ob

"Means in the same vertical column followed by the same letter do not differ significantly at the I~I" or 5y'. level by the least significant difference test (LSD) (paterson 1939).

counterparts reared on other dietary combinations (P < 0·01 or 0,05). Further, hatchability of eggs deposited by these highly fecund females was markedly higher (P < 0·01 or 0-05) than that recorded for eggs released by mateds in couples developed from larvae and pupae maintained on remaining dietary regimens tested here excepting the one where the caterpillars during the first 24 h of their lives were deriving nourishment from developing seeds and later from ripened seeds of okra (P>0·05). An examination into how far larval dietary regimen consisting of (i) tender seeds of okra fruit for one sex and epicarp of this fruit for the other sex or (ii) ovary of shoeflower would affect egg deposition and egg viability in this noctuid species yielded additional interesting results. The findings (table 4) compared with such data

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Earias and Tribolium reproduction

Table 4. Estimates of egg output and egg viability in E. vittella in tests where males and females of copulating pairs were held during rearing, on varied or on similar dietary regimens (data pooled from 5 females per test)*.

Dietary regimen Okra fruit-developing Okra fruit-s-epicarp for Okra fruit-epicarp for seeds for females Okra fruit-developing epicarp for females Ovary of shoeflower for Mean LSD (1%) (5%)

seeds for both sexes both sexes males and developing

Mean number of total eggs laid

Mean number of viable eggs laid

243-80 a 158·00 b

176·40 a 70·60 b

154·80 b

107·60 b

145·20 b 103·00 b 160·96 86·00 63-06

75·60 b 80·60b 102·16 80·49 59·01

seeds for males and both sexes

*Same notations as in table 3. Table 5. Number of eggs laid by mated females of E. vittella in pairs fed on different carbohydrate diets or distilled water during. their adult lives (data pooled from 5 females per test)", Adult food Raffinose Galactose Sucrose Fructose Glucose Distilled water Mean LSD (1%) (5%)

Mean number of eggs laid

604·6 a 509-8 ab 445·4 b 420·4 be 317-4 c 164·8 d 410·4 155'1 114·5

*Same notations as in table 3..

procured from moths where both sexes were raised on developing seeds or on epicarp clearly revealed statistically significant increments in mean egg output and egg viability in females following mating between the sexes reared exclusively on okra seed diet. Nourishment obtained by pairs from other food combinations tested here gave rise to mateds having pronouncedly low and statistically similar reproductive capacity. The influence of adult diet (one of various selected sugars or plain distilled water) on the reproductive potential of this insect was also assessed. Notwithstanding absence of any major difference in longevity or in length of oviposition period between moths ingesting a carbohydrate diet and those feeding on water, a significantly greater number of eggs were laid by the females fed on anyone of the test sugars than on water (table 5). Amongst the various carbohydrates, raffinose at 15% strength helped the individuals to deposit maximum number of eggs and oviposition by the moths was equally at a significantly higher level when they ingested this trisaccharide even when maintained at as Iowa concentration as 1% (P <0,01) (table 6).

160

S S Krishna Table 6. Number of eggs laid by mated females of E. vittella in pairs fed on different concentrations of raffinose solutions during their adult lives (data pooled from 5 females per test]". Concentration

Mean number of egg~ laid

15·00 5{jQ 1·00 (}50 0·25 0·10 Mean LSD (1%)

604·6 a 56% a 534·6 a 316·2 b 293'2 b 254·8 b 428·8 147·7

* Any two means followed by the same letter do not differ significantly at the 1~/~ level" by the least significant difference (LSD) test. Table 7. Number of eggs laid by mated females of E. villella in pairs raised on epicarp or developing seeds during pre-imaginal stages and adults given raffinose or glucose solution respectively (data pooled from 5 females per test). Experimental condition *Epicarp-reared and raffinose given in adult stage *Seed-reared and glucose given in adult stage

Mean number of eggs laid (±SE)

427-2± 18·66· 322-0± 20·50

*Okra fruit component. "Significantly different at 1~~ level from the value just below in the column (t test) (Paterson 1939). SE, Standard error.

In the light of informations presented above, egg yield values in this moth was compared when raffinose was provided as adult food for pairs whose immature stages were reared on epicarp diet with couples fed on glucose during their imaginal lives and whose immature stages were raised on developing okra seeds. Number of eggs deposited by moths associated with the former dietary regimens was decidedly greater (P < 0,01) (table 7) than that laid by those related with the latter set of foods.

3.2 T. castaneum Mean fresh weight of ovaries of females reared and fed on whole wheat flour, whether unmated and remaining in preoviposition state « 24 h old beetles) or mated and ovipositing (S-day old beetles), was comparatively higher than that determined for counterpart individuals that ingested semolina following their development on the same diet (table 8). Also, egg output in these beetles was more if they were held, right from their birth, on whole wheat flour instead of on semolina (figure 1). Females whose adult food, like those of males which mated with them, consisted of

161

Earias and Tribolium reproduction Table 8. Weight (jlg) of ovaries (left side only) of unmated or mated females of T. castaneum reared and maintained on whole wheat flour or semolina supplemented with yeast (data pooled from 5 females per test). Mean fresh weight of ovaries (± SE) Diet + 5~<, yeast

Unmated"

Whole wheat flour Semolina

180±37·51 140±24·56

600±70·90 540±5H2

a< 24 hold femalesin preoviposition state. "5 day old ovipositing females which mated with males belonging to the same dietary regimen. SE. Standard error.

100 r - - - - - - - - - - - - ,

80

..... :J

.....0. 60 :J 0

en en

QI

c

40

0

QI

~

20

vV

s

Figure l. Histogrammic representation of mean egg output by a mated female of T, castaneum in a pair reared on whole wheat flour (W) or semolina (S) enriched with yeast and adults maintained during a 40-day experimental period on the same diet as their larvae (data pooled (rom 5 females per test).

whole wheat flour extracted in 100% ethanol instead of in water, ethyl ether or in chloroform and then reinforced with yeast showed, subsequent to mating, maximal egg deposition possessing high statistical significance (table 9). Curiously enough, this egg yield value was significantly higher (P < 0'01) than that obtained from mateds allowed to eat normal wholewheat flour fortified with yeast.

162

S S Krishna Table 9. Number of eggs laid during a 20-day period. commencing from the day following emergence, by mated females of T. ca~taneum in pairs fed on normal or differently extracted whole wheat flour during their adult lives (data pooled from 5 females per test)". Diet (enriched with

5% yeast)

Mean number of total eggs laid

IOOo/~ ethanol-extracted whole wheat flour Normal whole wheat flour Ether-extracted whole wheat flour Water-extracted whole wheat flour Chloroform-extracted whole wheat flour Mean LSD(J%) (5~',,)

131·20a

57-20 b 13-80c

10·60 c 8·80 c 44·32 22·56 16·54

*Same notations as in ta ble 3.

Low productivity by females compelled to feed on flour extracted in water, chloroform or ether, despite these diets supplemented with yeast, might possibly be due to this cereal getting depleted, consequent to such treatments, of a variety of useful substances like carbohydrates, proteins, lipids and vitamins, many of which are likely to be of great nutritional value for these beetles in regulating their total egg output as in several insects (Wigglesworth 1960; Johansson 1964; Engelmann 1970; Adiyodi and Adiyodi 1974; Nayar 1977). It was, therefore, considered proper to enquire into this issue also wherein the egg laying capabilities of these tenebrionids fed during their adult lives on one of certain specific organic nutrients enriched with yeast were compared with those of counterpart individuals ingesting yeastcontaining normal whole wheat flour and the data presented in table 10. Although normal whole wheat flour proved to be the best amongst the various foods tested enabling the females to exhibit maximum oviposition (P
163

Earias and Tribolium reproduction Table 10. Number of eggs laid during a 20-day period, commencing from the day following emergence, in mated females of T. castaneum in pairs fed on whole wheat Dour or on different organic nutrients during their adult lives (data pooled from 5 females per test}", Diet (all excepting vitamin B complex mixture**) enriched with yeast Whole wheat Dour Starch Vitamin-free casein Glucose Sucrose Fructose Cholesterol Vitamin B complex mixture'?" Mean LSD (1%) (5/~)

Mean number of total eggs laid 57·20 a 19-60 b 19-00b 9-40 be 4·80c 4·40c

HOc 1·80 c 14·82 14-62 10·87

*Same notations as in table 3. **For details concerning preparation and composition of this diet, see § 2. Table II. Number of eggs laid during a 20-day period, commencing from the day following emergence, by mated females of T. castaneum in pairs fed during their adult lives on whole wheat flour or on vitamin B complex mixture** having low or high folic acid content (data pooled from 5 females per test)". Diet Whole wheat flour + 5% yeast Vitamin B complex mixture** with high folic acid content (63 pg/g of total diet) Vitamin B complex mixture** with low folic acid content (0-63 pg!g of total diet) Mean LSD (1%)

(5%)

Mean number of total eggs laid

57-20a 43-40 a 1-80 b 34'13 21-53 15·36

*Same notations as in table 3_ "Same notations as in table 10.

maintained on whole wheat flour plus yeast. A significantly greater egg laying was noticed in females in tests where the beetles fed on yeast - vitamin B complex mixture with increased folic acid content (PO·05). All egg yield data from mated females of T. castaneum put to variable nutritional experiences, as outlined above, were obtained when yeast was never omitted in the imaginal food of these beetles. Whether such inclusion of yeast in a diet fed by these insects is mandatory to improve the egg output of females and whether individuals

164

S S Krishna

that ingest only yeast can exhibit reproductive efficiency similar to those eating food with or without this ingredient are questions demanding clearer answers though Sokoloff and Ho (1962) were the first to throw some light on this matter. A probe into these issues made in the present investigation indicated a definite rise in the egg output by females if they belong to pairs whose whole wheat flour diet, instead of being offered as such, was supplemented with yeast (P
Diet Whole wheat flour+ 5% yeast Whole wheat flour Yeast Mean LSD (1'1,,)

57·20 a 36·80 b !·60c 31·86 20·47

14·67

(5~{,)

"Same notations as in table 3. Table 13. Number of eggs laid during a 20-day experimental period, commencing from the day following emergence, in mated females of T. castaneum in pairs fed on different oilseeds or whole wheat flour during their adult lives (data pooled from 5 females per testj", Diet (enriched with 5~{ yeast)

Mean number of total eggs laid

Whole wheat flour Ground nut Cotton seed Scsamum Linseed Mustard Toria Mean LSDW.,) (5'};;) *Same notations as in table 3.

57-20 a 42·20 ab

35·00 b 17-40 c 15-80c 4-20 c

4·20c 25'14 29-84 22·12

Earias and Tribolium reproduction

165

laying in comparison to cotton seed (P < 0'05) or remaining oilseeds (P < 0'01). But sesamum, in which postembryonic development of these beetles was unsuccessful [Pajni and Virk 1978), when eaten by these insects during their adult lives, facilitated the mated females to lay eggs which, interestingly, was about 50% of that recorded from females in pairs whose food was cotton seed plus yeast. Amongst the various yeast-supplemented spices tested, only coriander enabled these beetles to be somewhat productive-the mean number of eggs laid per female being 10·8. All the remaining diets in this category proved incompetent even to sustain longevity of these tenebrionids till the close of the 20-day experimental period. Egg viability in all the trials arranged here was 100%. 4. Discussion 4.1

E. vittella

A conspicuous contrast in the reproductive efficiency of this pest was noticed between females associated with pairs reared on tender okra seeds (0-8 days old) and those in couples grown on epicarp of this fruit or on ovary of shoeflower. A profound influence of larval nutrition on egg production in the adult (Wigglesworth 1960) is implicit in these findings. The considerably high fecundity exhibited by the moths developed on seeds is evidently due to the effect of a superior nutritional quality of the diet, at least in regard to free amino acids and water-soluble proteins (Mani et al 1986) which, in addition to accelerating the overall development (Mehta and Saxena 1973; Vishwapremi and Krishna 1974a), boosts the reproductive potential as well. These nutrient reserves are plausibly accumulated more readily in relatively larger quantities during larval life' by the adult females reared solely on seeds and subsequently utilized by them resulting in their greatly enhanced egg output. The fact that the number of eggs laid by females developed on whole fruit, in which such seeds were also present, was still significantly less than that deposited by individuals reared only on seeds separated from the fruit lends additional support to this postulation. That such nutrition becomes an obligatory requirement for the caterpillars, at least for the first 3 days of their lives, to enable the resultant females to become maximally productive can be inferred by comparison of egg yield and egg hatchability values obtained from these individuals, whose subsequent larval and pupal phases were allowed to continue on ripened okra seeds (9-11 days old), with those recorded from females associated with pairs held on other single or mixed dietary regimens for different prescribed periods of their postembryonic development. Further, the mandatory need for both sexes of a copulating pair to avail nutrients from okra seeds during their larval development to facilitate this noctuid species to achieve highest reproductive potential (as manifested through significant increments in mean egg output and mean egg viability in the female belonging to this test couple) clearly indicates that not only in females but also in males is there an apparent impact of nutrition on the reproductive activity, once considered to be of negligible magnitude in insects (Wigglesworth 1960). So far as the role of adult nutrition in the regulation of reproduction in E. oittella was concerned, the results showed a marked elevation in egg deposition when the

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adult moths were fed on a sugar solution instead of plain water, although they were all reared under identical conditions and did not have any pronounced difference in longevity or in length of oviposition period. This unquestionably suggests that carbohydrate nutrient is very essential during imaginal period in promoting the egg laying capacity of mated females as in the butterfly, Colias philodice eurytheme (Stern and Smith 1960) plausibly through better utilization of dietary protein reserves in yolk formation during egg production (Engelmann 1970). The greatest contrast in the aggregate number of eggs deposited between raffinose- and glucose-fed moths further implies that, like in the syrphid Sphaerophoria scutellaris (Lal and Haque 1955), different sugars variably regulate the reproductive potential of the spotted bollworm. The higher egg output by females ingesting fructose or galactose instead of glucose, notwithstanding that all the 3 sugars are monosaccharides, corroborates this interpretation. The maximum egg output by moths fed on raffinose can then reasonably be assumed to be due to the combined effect of fructose, galactose and glucose (all of which will be obtained from the ingested raffinose subsequent to its complete hydrolysis) on the reproduction of this pest. The fact that eggs deposited by moths that imbibed raffinose in adult stage but reared on epicarp diet was significantly greater than those laid by seed-reared and glucose-ingested individuals is a pointer to this possibility and highlights the importance of this trisaccharide as an efficiently compensating nutritional factor in the diet of this insect. Although the need for a carbohydrate nutrient during adult life to realize full fecundity in females has now been experimentally established in E. vittella, it is still not known whether variations in the effect of different sugars ingested by the adults on the female's oviposition are due to superior nutritional value of some of these saccharides or on account of disparities in food quantities consumed. Also the question whether the moths in nature obtain through feeding all the necessary sugars, quality- and/or quantitywise, from the foliage orflowers of one or more of the plants naturally infested by them or they are impelled by instinct to visit certain other plants falling outside their normal host range to fulfil their carbohydrate requirements for reproduction is still open.

4.2

T. castaneum

The possession of more heavily weighing ovaries and occurrence of higher egg output by mated females, held right from their birth, on whole wheat flour supplemented with yeast instead of on similarly enriched semolina clearly indicate that the former cereal foodstuff is the most favourable diet for T. castaneum so far as its reproductive potential is concerned - an observation which supports the earlier findings of Good (1933) and Hamalainen and Loschiavo (1977) who, in their publications have, however, compared the fecundity of this insect on natural and enriched whole wheat flour with cereal products other than semolina. The observation that whole wheat flour reinforced with yeast, as a food is relatively nutritionally superior to semolina to effectuate the red flour beetle mated females to lay more eggs evidently led to an examination of the ability of these females to oviposit if this flour, after being extracted in one or the other of different solvents, was provided as adult food to reproductive pairs with addition of yeast. It was found that females in pairs fed on 100% ethanol-extracted whole wheat flour

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were capable of depositing the highest number of eggs significantly surpassing the egg output figures obtained not only from counterpart mateds ingesting this cereal food subsequent to extraction in water, ether or in chloroform but also, interestingly, even from those eating yeast-added normal whole wheat flour. While no forthright statement can be made presently to explain this unexpected shootup in the number of eggs laid by mateds fed during their adult lives on ethanol-extracted whole wheat flour, it is, however, presumed that such a chemical treatment of this cereal aided in the removal of certain not yet characterized ingredients that possibly function as natural checks for the females in their reproduction. specifically in relation to oviposition. This postulation may also serve as a basis for explaining, in part, the occurrence of poor egg laying in adult females associated with couples ingesting water-, ether- or chloroform-extracted whole wheat flour wherein such detrimental ethanol-soluble constituents were fully present with the simultaneous depletion of several nutritionally important chemical compounds, specially starch and casein, both of which were found to be very important in the reproduction of this insect (table 10). If this assumption is correct, it would not be out of place here to suggest that future investigations into the reproductive biology of T. castaneum may be focussed towards chemical characterization and identification of these ethanolsoluble constituents and their judicious application in pest management programmes aimed at limiting the population build-up of this beetle on stored commodities. The poor fecundity (in terms of egg deposition scores) exhibited by the mated females of this species when both sexes, as adults, were fed on water-, chloroform- or etherextracted whole wheat flour enriched with yeast, notwithstanding ingestion of yeastadded whole wheat flour during their larval life, indicates absence of any benefit provided to the imagines in their reproduction via larval nutrition and thus supports the inference drawn along similar lines by Reynolds (1944) in his studies on T. destructor. Oviposition was significantly curtailed if the adult food of the mated females consisting only of yeast-supplemented B complex vitamins had folic acid content as low as 0·63 ug instead of 63 flg/g of total diet. It appears that folic acid in appreciable concentration is also a necessary constituent in the adult diet of T. castaneum for aiding the beetle's reproduction like other components of B vitamins such as riboflavin, thiamin and niacin, all 3 of which have been reported by Hamalainen and Loschia vo (1977) to increase the fecundity of this species. Addition of yeast to whole wheat flour given as adult diet caused a marked improvement in the egg output by the mated females. From this it is inferred that yeast, in which existence of some growth factors necessary for normal development of T. confusum has already been discovered (Charbonneau and Lemonde 1960a, b), presumably contains also certain as yet unidentified components responsible, additively, for the promotion of oviposition in T. casta Ileum. The reproductive potential of these beetles, subsequent to feeding them on different selected oilseeds or spices in their adult lives, did not always coincide with the comparative dietary efficiency of these products for the larval growth of this insect (Pajni and Virk 1978). This implies the occurrence of variability in the nutritional competency of these materials insofar as larval development and adult reproduction in this pest are concerned. A 100% viability of eggs in all test conditions clearly indicates absence of any effect of nutritional difference on egg hatchability in this tenebrionid species.

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Acknowledgement

This work was supported by funds from the University Grants Commission and Indian National Science Academy, New Delhi. References Adiyodi K G and Adiyodi R G 1974 Comparative physiology of reproduction in arthropods; in Ad~ances in comparative physiology and biochemistry (ed) 0 Lowenstein (London: Academic Press) voL 5, pp 37-107 Butani D K and Jotwani M G 1984 Insects in vegetables (Delhi: Periodical Expert Book Agency) p 356 Chapman R F 1918 The confused flour beetle (Tribolium confusum Duval); Minn. State Entomol. Rpt. 17 73-94 Charbonneau Rand Lemonde A 1960a Unidentified growth factors in brewer's yeast. I. Necessity of these factors for Tribolium confusum Duval; Can. J. Zool. 38 87-90 Charbonneau Rand Lemonde A 1960b Unidentified growth factors in brewer's yeast. II. Some chemical and physical properties of these factors; Can. J. Zool. 38 443-448 Engelmann F 1970 The physiology of insect reproduction (New York: Pergamon Press) p 307 Feakin S D 1976 Insect pests of stored paddy and milled rice; PANS, Second edition, Manual No.3, p 244 Good N E 1933 Biology of the flour beetle Tribolium confusum Duv., and Triboliurn ferruqenium Fab.; J. Agric. Res. 46 327-334 Hamalainen M K and Loschiavo S R 1977 Effects of synthetic B-vitamin and natural enrichment of flour on larval development and fecundity of Tribolium confusum and T. castaneum; Eniomoi. Exp. App/. 21 29-37 Johansson A S 1964 Feeding and nutrition in reproductive processes in insects; in Insect reproduction (ed) K C Highnam (London: Symp. Roy. Ent. Soc. No.2) pp 43-55 King C E and Dawson P S 1977 Tribolium castaneum; in Diseases, pests and weeds in tropical crops (eds) J Kranz. H Schmutterer and W Koch (Berlin and Hamburg: Verlag Paul Parey) pp 394-395 Krishna S S, Vishwapremi K K C and Shahi K P 1977 Studies on the reproduction in Earias fabia Stoll (Lepidoptera: Noctuidae): Oviposition in relation to adult nutrition, mating and some environmental factors; Entomon 2 11-16 Lal R and Haque E 1955 Effect of nutrition under controlled conditions of temperature and humidity on longevity and fecundity of Sphaerophoria scutellaris (Fabr.) (Syrphidae: Diptera). Efficacy of its maggots as aphid predators; Indian J. Entomol. 17317-325 Mani H C. Pathak P H and Krishna S S 1986 Effect of larval food quality on egg number and egg viability in Eariasfabia Stoll (Lepidoptera: Noctuidae); Z. Angew. Zool. 73115-121 Mehta R C and Saxena K N 1973 Growth of the cotton spotted bollworm, Earias fabia (Lepidoptera: Noctuidae) in relation to consumption, nutritive value and utilization of food from various plants; Entomol. Exp. Appl. 1620-30 Narayan Singh R and Krishna S S 1980 Effect of some common oilseeds and spices serving as adult food on the reproductive potential of Tribolium castaneum (Hbst.) (Coleoptera: Tenebrionidae); Entomon 5 161-162· Narayan Singh R and Krishna 5 S 1981 Effects of certain dietary and exteroceptive factors associated with the developing pupa of Tribolium castaneum (Herbst) in the insect's reproductive programming; Mitt. Zool. Mus. Berl. 57 5-10 Narayan Singh R and Krishna S 5 1983 The impact of variation in adult food on the reproductive programming in Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae); New Entomol. 32 23-30 Nayar K K 1977 Invertebrate reproduction (New Delhi: Oxford and IBH Publishing Co.) p 239 Pajni H Rand Virk N 1978 Comparative dietary efficiency of common spices and oilseeds for the larval growth of Triboiium castaneum Herbst (Coleoptera: Tenebrionidae); Entomon 3 135-137 Paterson D D 1939 Statistical technique ill agricultural research (New York: McGraw Hill Book Co.) p 263 Reynolds J M 1944 The biology of Tribolium destructor (Uytt.), 1. Some effects of fertilization and food factors on fecundity and fertility; Ann. Appl. Biol. 31 132-142 Singh Raghvendra P and Krishna S S 1980 Variation in the development and reproductive programmes of Eariasfabia Stoll (Lepidoptera: Noctuidae) reared on ripened or on dilTerently extracted developing seeds of okra fruit; Kan_ Univ_ Res. J. (Sci). 1 107-114

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Singh Raghvendra P and Krishna S S 1983 Field and laboratory studies on certain aspects of orientation and reproductive behaviour of Earias [abia Stoll (Lepidoptera: Noctuidae); in Insect interrelations in forest and aqro-ecosystems (eds) P K Sen-Sarma, S K Sangal and S K Kulshrestha (Dehra Dun: Jugal Kishore and Co.) pp 147-160 Snedecor G W 1961 Statistical methods (Bombay: Allied Pacific Private Ltd.) p 534 Sokoloff A 1974 The biology of Tribolium (New York: Oxford University Press) Vol. 4 p 610 Sokoloff A and Ho F K 1962 Productivity of Tribolium castaneum and T. confusum in various media; TIB 540--42 Stern V M and Smith R F 1960 Factors affecting egg production and oviposition in populations of Colias philodice eurytheme Boisduval (Lepidoptera: Pieridae); Hilqardia 29411-454 Vishwapremi K K C and Krishna S S 1974a Variations in the development program of Earias fabia reared on whole or excised fruits of okra or on their components; Ann. Entomol. Soc. Am. 67 147-148 Vishwapremi K K C and Krishna S S 1974b Variation in the reproductive potential of Eariasfabia Stoll (Lepidoptera: Noctuidae) reared on whole fruit of okra or on its components; Proc. Indian Natl. Sci. Acad. 840 440-445 Vishwapremi K K C and Krishna S S 1975 The influence of some carbohydrate diets as adult food or group interaction during immature stages on the programming of oviposition "in the spotted bollworm, Earias fabia Stoll (Lepidoptera: Noctuidae); Proc. Indian Natl. Sci. Acad. 841 141-147 Wigglesworth V B 1960 Nutrition and reproduction in insects; Proc. Nutr. Soc. 19 18-23