Anim. Behav., 1991, 42, 389-394

Trophallaxis in the honeybee, Apis mellifera (L.) as related to the profitability of food sources

W A L T E R M. F A R I N A & J O S U E A. NIJ lqEZ Departamento de Ciencias Biolrgieas, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos

Aires, Pab. II--Ciudad Universitaria, CP 1428 Buenos Aires, Argentina

(Received 7 September 1990; initial acceptance 12 October 1990; final acceptance 8 January 1991; MS. number: A5874)

Abstract. Trophallaxis by honeybees was studied in two series of experiments. Honeybees were fed to satiation with sucrose solutions from 10 to 50% w/w (sugar concentration experiment), or with different amounts of a 50% w/w sucrose solution (crop load experiment). By increasing the concentration of solutions, the transfer rate was increased up to a maximum value for the 30-40% sucrose solution. With increasing crop load, maximum transfer rate was attained for satiated bees. The amount of sucrose transferred during trophallactic contact was related to the amount of sucrose carried in the honey sac for both experimental series. Results are discussed in relation to the foraging and recruiting patterns displayed by foragers.

Trophallaxis is the mouth-to-mouth exchange of food between individuals of the same or different castes in social insect colonies (Wheeler 1923). Trophallactic contacts are frequently observed in honeybees, where two individuals participate: a donor, or D-bee, and a recipient, or R-bee (Free 1956, 1957). The D-bee offers food regurgitated from its honey sac by opening its mandibles; the R-bee takes the offered food by protruding its pro- boscis and placing it between the mouth parts of the D-bee (Korst & Velthuis 1982). The D-bee may offer food spontaneously or after active solicitation from the R-bee.

It has already been established that the profit- ability of food sources (e.g. flow rates or sugar concentrations of solutions, is represented in the liveliness of the recruiting dance (yon Frisch 1965; Nflfiez 1970), although the possibility that the same kind of information may be exchanged through trophallactic contacts has not yet been discussed. In this paper, we studied the effects of qualitative and quantitative changes in the food afforded to the D-bee on trophallaxis.

M A T E R I A L S AND M E T H O D S

We performed these experiments from February to April 1989, at the end of the nectar flow season, in the apiary of the Argentine Beekeeper Society (S.A.D.A.), Gonzfilez Catfin (34.6 ~ South), Province of Buenos Aires, Argentina.

Experimental Procedure

We conditioned honeybees, from a hive sited 50 m from the laboratory, to a feeder tray (a plastic petri dish, 5 cm in diameter, with 3-ram holes in the cover). The dish was filled with 50% weight/weight (w/w) sucrose solution and situated at a window of the laboratory. Sucrose solutions used at the food source and in experiments were marked by mixing 80 gl of vanilla essence into each litre of solution.

In vials, we captured two foragers that had been conditioned for about 1 h at the feeder tray, before they began to drink. We arbitrarily assigned one bee as the D-bee and the other as the R-bee. We then fed the D-bee through a graduated capillary, with a known amount and concentration of sucrose solution; we did not feed the R-bee. We weighed the vials containing the bees to the nearest 0-1 mg (Wi) and then attached them to an arena (see Fig. 1). We observed trophallactic interactions in the arena, which measured 4 x 6 • 1 cm and was divided into two halves by a sliding door. At opposite walls of the arena, we could attach plastic vials (1 cm diameter • 2.5 cm length), which had sliding doors to retain bees (Fig. 1). Ten minutes after capture, we opened the three sliding doors, thus allowing bees to come into contact with each other. If no exchange occurred, after a trophallactic contact of at least of 1 s or after a maximum of 10 rain, we isolated D- and R-bees by closing the main sliding door of the arena. After the bees returned to the

0003-3472/91/090389 + 06 $03.00/0 �9 1991 The Association for the Study of Animal Behaviour

389

390 Animal Behaviour, 42, 3

Viol J l

dllmmll[t.t~"

S l i d i n c d o o r ~ j

l c m

Cover

Figure 1. Diagram of experimental arena.

vials we weighed them (W 0. We disregarded tro- phallactic contacts shorter than 1 s in duration (see Korst & Velthuis 1982).

Measurements in control bees showed a loss of body mass weight during the observation period ( Y = - 0 . 1 + 0 ' 0 9 7 X , N=55, P<0.01; linear regression between loss of mass in mg and obser- vation time in minutes). We did not consider this loss of mass during experiments.

We recorded the following values. Trophallactic time (Tt): the time spent engaged in food exchange behaviour; exchanged volume: the mean difference between final (Wf) and initial weights (Wi) of the two bees by specific weight of solution; and trophallactic responsiveness (Tr): number of trophallactic trials • 100/total trials. Using the parameters of exchanged volume (pl) and trophallactic time (s), we could estimate the transfer rate in gl/s (slope, b, of linear regression between these parameters), for each concentration and crop load.

Sugar concentration experiment

We fed D-bees until satiation with 10, 20, 30, 40 and 50% w/w sucrose solution.

Crop load experiment

We fed D-bees with 5, 10, 20, 40 gl and until satiation (57.5 p.1 approximately) of 50% w/w sucrose solution. Data at satiation with 50% w/w sucrose solution were the same for both experimental series.

We used a total of 150 different pairs of bees in both experiments.

R E S U L T S

Sugar Concentration Experiment Trophallactic responsiveness was similar and

remained high for every sucrose concentration studied (heterogeneity G-test, see Table I). Neither trophallactic time nor exchanged volume were dependent on the concentration of the sucrose solution fed (one-way ANOVA, Table I). Transfer rate increased with sucrose concentration to a maximum with the 30% sucrose solution (Fig. 2).

If only the weight of sucrose transferred (mg) per second (sucrose transfer rate) is taken into account (e.g. for 10% solution: transfer ratel0o/o • specific weight 10,/o • 10/100 = 1. l 15 gl/s • 1-038 mg/gl x 10/ 100=0-116 mg/s), the maximum was attained for the 40-50% solution (Fig. 2).

Crop Load Experiment Trophallactic responsiveness was dependent on

donor crop load (heterogeneity G-test, see Table II). When the load was 5 pl, no trophallactic con- tacts occurred during the observation period. Satiated bees, however, showed a trophallactic responsiveness of up to 88-9%. We found no depen- dence between crop load and trophallactic time (one-way ANOVA), although exchanged volume was dependent on crop load (Table II).

Transfer rate and particularly its correlation coefficient r, increased with crop load (Fig. 3).

Farina & N~t~ez: Trophallaxis in honeybees

Table I. Mean (_ sE) values of trophallactic responsiveness (Tr) , trophallactic time (T 0 and exchanged volume (Ev) at different concentrations

391

%w/w

10 20 30 40 50

n/N 8/11 8/9 9/11 9/15 16/18 Crop load (lal) 49.0+2.0 49.2_+2.2 53.3_+1.2 57.8+1.8 57.5_+1.5 T r (%)* 72.7 88.9 81.8 60.0 88.9 Tt (s)1" 5+1.3 12+2.3 10+1.1 9-+0.6 13-+2.7 Ev (Ixl)~ 6.1_+1-5 17-2-+3.2 11.4-1-2.0 7-7+1.3 14.0_+3.0

n/N= number of assays with food exchange/number of replicates. *G r = 7.96, MS (heterogeneity G-test). i F = 1.72, df= 4,45, NS (one-way ANOVA). :~F= 2.45, df= 4,45, NS (one-way ANOVA).

/[ •\ ~1.. 1"5 / ",,

/,(}' \\

I-0 I I I I Jr IO 20 30 40 50

Sucrose concentrofion (% W/W)

Figure 2. Estimated rates for different sucrose concen- trations. The transfer rate (0) for each concentration is the slope of linear regression, b + s~, between exchanged volume and trophallactic time. Means of sucrose transfer rates (O): slopes in mg of sucrose transferred/s. The corresponding regression equations were: 10% solution, Y=0.395+l.lI5X, r=0.96, P<0-001, N=8; 20%, Y=0.739+l.330X, r=0.954, P<0.001, N=8; 30%, Y=4.166+I.611X, r=0.895, P<0.001, N=9; 40%, Y=6.545+l.493X, r=0.775, P<0.05, N=9; 50%, Y=0.201 + 1.066X, r =0.964, P<0.001, N= 16.

0-5 2

0.1

With higher crop load, the exchanged volume was strongly dependent on the time D- and R-bees were in trophallactic contact.

D,SCUSS,ON We defined the limits of our study with the follow- ing observation of von Frisch (1968). 'It is not yet understood which signals are used when there is no dance or scent.'

Recruiting efficiency, i.e. the number of recruited foragers, depends on the profitability of the food source in terms of the concentration of the sugar solution (von Frisch 1965) as well as the flow rate at a constant concentration (Nfifiez 1971). A quanti- tative study of this dependence (Ntifiez 1966, 1971) allowed us to define four ranges of flow rates for a 50% w/w sucrose solution that are characterized by behavioural responses correlated with the profit- ability of the food source (Nfifiez 1975). In the first range, above 6 ~ 6 5 gl/min (the maximum intake rate of an individual bee; Nfifiez 1966), solution availability exceeds the bee's perception threshold for food availability. Only the participation of recruited gatherers afforded an estimation of the profitability of the food source. The number of recruited bees at the food sources and the average crop load attained are behavioural parameters correlated with source productivity (Nfifiez 1971). In the second range, between 8-10~tl/min and 60-65~d/min, the individual bee can assess profitability through the time needed to fill its crop, 1-7 min according to the flow rate at the food source (Nfifiez 1966). In the third range, between 0'3 and 8-10gl /min, a correlation between flow rate and crop load was found (Nfifiez 1966). Finally, below 0.3 ~l/min, a quantitative dependence was observed between flow rate and length of the subsequent foraging pause in the hive (Nfifiez, unpublished data).

Here, we studied trophallactic behaviour when the individual bee could assess profitability through the concentration of sucrose solution, as observed by many authors (Boch 1956; von Frisch 1965). We also looked at behaviour in the third flow range,

392 Animal Behaviour, 42, 3

Table II. Mean ( _+ sE) values of trophallactic responsiveness (Tr), trophallactic time (Tt) and exchanged volume (Ev) compared among different crop loads with a 50% w/w sucrose solution

Crop load (gl)

Satiation 5 l0 20 40 (57.5• 1.5)

n/N 0/6 6/20 8/30 8/24 16/18 T r (%)* 0.0 30-0 26.7 33.3 88.9 Tt (s)t - - 6_+1.1 12_+2.2 7_+1.6 13+2.7 E., (gl):~ - - 2 .5_+0-5 8.1+2.1 4 '1___1'3 14'0+3'0

n/N= number of assays with food exchange/number of replicates. *G r = 130.8, P < 0.005 (heterogeneity G-test). t F= 1-68, df= 3,34, Ns (one-way ANOVA). :~F= 3.82, df= 3,34, P < 0.02 (one-way ANOVA).

1.0

oA

0.5 g #

O

S ' L

- - ~ I I I I I

5 I0 20 40 60 Crop lood (pA)

Figure 3. Transfer rate ( � 9 for different donor crop loads, b_+ sE, of a 50% w/w sucrose solution. Estimated re- gression equations were: 10 gl, Y= 1.351 +0.198X, r= 0'504, NS, N=6; 20 ~1, Y=--0'0665+0'757X, r=0-782, P<0.05, N=8; 40~1, Y=-0.619+0'702X, r=0.876, P < 0.01, N= 8; satiated bees (approximately 57.5 gl), Y= -0.201 + 1.066X, r =0-964, P< 0.001, N= 16.

when individual bees could assess profitability through the flow rate and represented it through the crop load attained during the foraging trip (Nflfiez 1966).

Satiated D-bees showed maximal responsive- ness for trophallactic contact. Independent of the sucrose concentration of the load, a high percent- age of trials ended with solution transfer (Table I). It is important to mention that satiated D-bees were frequently observed spontaneously offering a drop of solution, even when no begging occurred. Transfer rate increased with concentration, up to a peak for the 30% sucrose solution (1.61 gl/s), largely overriding the intake rate of the bee (Nflfiez 1966). This means that, as concentrat ion increases,

the D-bee transfers the sugar solution with increas- ing speed, and the time needed by the R-bee to ingest it exceeds the trophallactic time recorded. On the other hand, the decrease in transfer rate at even higher concentrations could be explained through a saturation threshold attained by bees in their gustatory receptors (Whitehead & Larsen 1976) and to the higher viscosity of solution (Fig. 2).

When the crop load of D-bees decreased, transfer rate decreased. The decreasing readiness of D-bees to dispense solution to begging R-bees could explain the low values of the correlation coefficient between exchanged volume (Ev) and trophallactic time (Tt), i.e. sucrose transfer (Ev) was less frequently associated with time of trophallactic display (T 0.

By comparing the two experimental series, we found an amazing correspondence between the amount of sucrose transferred and the amount of sucrose in the donor crop, independent of concen- tration (Fig. 4a). For example, when a D-bee stored 20 pl of 50 % solution in its crop (12.3 mg of sucrose), it transferred an average of 5'0 mg of the sucrose to the R-bee. When a D-bee stored 49.2 gl of a 20% solution (10.6 mg of sucrose), it transferred 3-7 mg (the difference between these values was not sig- nificant). Moreover, as the sucrose load of D-bees increased, the amount transferred during the trophallactic contact also increased up to a load of 10-12 mg in both experiments. It could be assumed that recruiting intensity depends essentially on the amount of sucrose carried by the forager. This assumption is in agreement with Nflfiez (1966) and suggests that the actual input variable of the intake control system in the individual forager is the rate of

Farina & N(t~ez." Trophallax& in honeybees 393

E

o (/3

iO Ca)

lO 20 30

5 0 (b)

~ 3o

..f --.%.7 ~ 2o

J i I l I I I t 0 I0 20 30

Sucrose load (mg)

Figure 4. (a) Sucrose load transferred by trophallactic exchange as related to sucrose loaded in donor crop: (�9 crop load experiment ( � 9 concentration experiment. Means of sucrose loads transferred increase significantly up to 10.6 mg of sucrose load (t-test: 3.0 versus 5.1 mg in crop, t = 3.214, df= 12, P < 0-01; 5.1 versus 6.1 rag, t = 4.22, df= 12, P < 0.01; 6.1 versus 10.6 mg, t=2.593, df= 12, P<0.05; the other differences among successive pairs are not significant). (b) Percentage of the sucrose transferred (sucrose transferred x 100/sucrose load) for different sucrose loads; (O) crop load experiment; (0) concentration experiment. Percentages transferred are different among sucrose-loads: GT=75'04, P < 0.005 (heterogeneity G-test).

intake of sucrose (mg/s). It also raises the question of a quantifying function of this system to evaluate profitability of the food sources.

I f sucrose transferred is expressed as the percent- age of sucrose load in the donor crop, a considerable decrease is found for the highest loads. Thus, D-bees reduce the percentage of sucrose load (mg) trans- ferred in the first food exchange at high sucrose loads (Fig. 4b), retaining a higher percentage of the load in their crops. This suggests an increase in the degree of parti t ioning of the sugar load, stimulating an increased number of hive-mate beggers with sugar solution.

Food partit ioning between hive-mates combined with an increased sucrose transfer rate could be a reliable information strategy during recruiting for field food sources. Sucrose transfer rate and the percentage of sucrose load transferred attained maximum and minimum values respectively in the 30-50% concentration range, suggesting that food sources within this concentration range would be more attractive and induce higher levels of recruit- ment in the hive. Further experimental studies are needed to analyse this possibility.

The D- and R-bees we used were foraging bees; non-foraging R-bees may behave differently in the

394 Animal Behaviour, 42, 3

arena, as well as in a normal context (the hive). In addition, we used only one colony of honeybees: there could be considerable differences in this behaviour between colonies, especially if the bees are unrelated, or of different subspecies.

In our experimental design we deliberately ignored the fact that different loads are attained from natural food sources (Nfifiez 1970). Prelimi- nary experiments introducing this variable (W. M. Farina, unpublished data) suggest that time spent at the food source is positively correlated with the D-bees' delay in starting a trophallactic contact. Environmental factors (distance from the food source, wind-speed, ambient temperature) experi- enced by the foragers in the field should increase the time-cost during the trip, and would then be expected to modulate the bees' readiness to give the nectar load at the hive. In other words, a 'hard earned load' would be transferred only after 'hard begging'.

Through this begging/giving process, the transfer speed and the amount of the sugar load transferred, it would be possible for bees at the hive to be aware of food availability at sources exploited by other foragers (Nfifiez 1982). The bees at the hive would avoid foragers with low flow rates and choose foragers with high rates. On the other hand, foragers would be aware of the needs of the hive through the begging intensity of their hive-mates, reinforcing their foraging activity (Lindauer 1948; Seeley 1986).

A C K N O W L E D G M E N T S

We are deeply indebted to Dr Hayo H. W. Velthuis (Utrecht, The Netherlands) for valuable comments and corrections on the manuscript and to M. Giurfa, C. Lazzari, F. Roces and two anonymous referees for comments on the manuscript and many valuable suggestions. The Argentine Beekeeper Society (S.A.D.A) provided facilities and technical assistance. The work was supported by a grant

from CONICET (National Research Council, Buenos Aires, Argentina) to J.A.N.

R E F E R E N C E S

Boch, R. 1956. Die Tfinze der Bienen bei nahen und fernen Trachtquellen. Z. vergl. Physiol., 38, 136167.

Free, J. B. 1956. A study of the stimuli which release the food begging and offering responses of worker honey- bees. Br. J. Anim. Behav., 4, 94-101.

Free, J. B. 1957. The transmission of food between worker honeybees. Br. J. Anim. Behav., 5, 41-47.

von Frisch, K. 1965. Tanzsprache und Orientierung der Bienen. Berlin: Springer-Verlag.

von Frisch, K. 1968. The role of dances in recruiting bees to familiar sites. Anita. Behav., 16, 531-533.

Korst, P. J. A. M. & Velthuis, H. H. 1982. The nature of trophallaxis in honeybees. Insectes soc., 2, 209-221.

Lindauer, M. 1948. fOber die Einwirkung von Durf-und Geschmacksstoffen sowie anderer Faktoren auf die T~inze der Bienen. Z. vergl. Physiol., 31,348-412.

Nfifiez, J. A. 1966. Quantitative Beziehungen zwischen den Eigenschaften von Futterquellen und dem Verhal- ten yon Sammelbienen. Z. vergl. Physiol., 53, 142 164.

Nfifiez, J. A. 1970. The relationship between sugar flow and foraging and recruiting behaviour of honeybees (Apis mellifera L.). Anita. Behav., 18, 527 538.

Nfifiez, J. A. 1971. Beobachtungeng an sozialbezogenen Verhaltensweisen von Sammelbienen. Z. Tierpsychol., 28, 1-18.

Nfifiez, J. A. 1975. El comportamiento de la abeja Apis mellifera L.: un estudio cuantitativo del comporta- miento. In: Progresos en Biologia (Ed. by F. D. Barbieri & A. Legname), pp. 225-234. Tucumfin: Fundaci6n Miguel Lillo.

N~fiez, J. A. 1982. Honeybee foraging strategies at a food source in relation to its distance from the hive and the rate of sugar flow. J. Apicult. Res., 21, 139-150.

Seeley, T. 1986. Social foraging by honeybees: how colonies allocate foragers among patches of flowers. Behav. Ecol. Sociobiol., 19, 343-354.

Wheeler, W. M. 1923. Social Life among Insects. London: Constable.

Whitehead, A. T. & Larsen, J. R. 1976. Electrophysiologi- cal responses of galeal contact chemoreceptors of Apis mellifera to selected sugars and electrolytes. J, Insect. Physiol., 22, 1609-1616.