Effect of post-teneral diets on the performance of sterile Anastrepha ludens and Anastrepha obliqua fruit flies

Effect of post-teneral diets on the performance of sterileAnastrepha ludens and Anastrepha obliqua fruit fliesP. Liedo1, D. Orozco2, L. Cruz-Lopez1, J. L. Quintero2, C. Becerra-Perez1, M. del Refugio Hernandez2,A. Oropeza1 & J. Toledo1

1 El Colegio de la Frontera Sur (ECOSUR), Tapachula, Chiapas, Mexico

2 Programa Moscamed Moscafrut, Tapachula, Chiapas, Mexico

Introduction

Anastrepha fruit flies are important pests of fruits in

the Americas (Aluja 1994). In Mexico, the technology

has been developed to apply the sterile insect tech-

nique (SIT) for the control of the Mexican fruit fly,

Anastrepha ludens (Loew) and the West Indies fruit fly

or mango fly, Anastrepha obliqua (Macquart). Over

100 million sterile flies of each species are produced

weekly at the MOSCAFRUT facility, in Metapa,

Chiapas, to be released in different places of

Mexico and in the southern USA (Orozco et al.

2004).

The advantages and benefits of using the SIT as

part of an area-wide integrated pest control have

been well acknowledged and the application of this

technique for the control of fruit flies has expanded

rapidly (Dyck et al. 2005). However, since the early

days of its development, the possible harmful effects

that mass-rearing, sterilization and handling, could

Keywords

Mexican fruit fly, pre-release feeding, sterile

insect technique, West Indies fruit fly

Correspondence

Pablo Liedo, ECOSUR, Carretera Antiguo

Aeropuerto Km 2.5, Tapachula, Chiapas,

Mexico 30700. E-mail: [email protected]

Received: May 12, 2010; accepted: June 23,

2010.

doi: 10.1111/j.1439-0418.2010.01568.x

Abstract

The effect of post-teneral diets on the mating performance, pheromone

production and longevity of sterile Anastrepha ludens and Anastrepha obli-

qua fruit flies (Diptera: Tephritidae) was investigated. Enriched pre-

release diets improved male mating performance. Wild and sterile males

fed on mango and orange fruits achieved the greatest copulatory suc-

cess, followed by those fed on a 3 : 1 sugar : yeast (SY) diet. Males fed

only on a sugar diet achieved the lowest number of matings. The mean

numbers of copulas achieved by wild males were significantly higher

than those by sterile males fed on a sugar-only diet, while there were

no significant differences between wild males and sterile males fed on

yeast diets. There was a trend of reduced mean number of copulas as

the proportion of yeast was reduced in the diet, but differences were

not significant. Pre-release diets had a significant effect on pheromone

production. Males fed on a 3 : 1 SY diet produced the greatest amount

of the three main pheromone components in A. ludens males and two

major components in A. obliqua males, followed by males fed on fruits

or pasteurized fruit juice. Males fed on a sugar only diet produced the

lowest amounts. The longevity response to post-teneral diets was com-

plex. The most yeast-rich diet and the poorest diet (sugar only) resulted

in the lowest life expectancies. Flies fed on 24 : 1 SY diets showed the

highest life expectancies for both males and females of the two species.

Considering the tradeoffs between mating performance and longevity,

the 24 : 1 SY diet would be recommended for programmes integrating

the sterile insect technique, but the effects of these diets on field sur-

vival and dispersal still need to be investigated.

J. Appl. Entomol.

J. Appl. Entomol. 137(Suppl. 1) (2013) 49–60 ª 2010 Blackwell Verlag, GmbH 49

have on the performance of sterile insects were

recognized (Knipling 1955, 1959). Methods have

been developed to overcome or minimize some of

these limitations. The total quality control approach,

colony management, larval diets, irradiation doses,

atmospheres and ages for sterilization are examples

of improvements at the mass-rearing factory (Ohi-

nata et al. 1977; Chang et al. 2001; FAO/IAEA/

USDA 2003; Toledo et al. 2004; Calkins and Parker

2005; Caceres et al. 2007; Liedo et al. 2007; Resilva

et al. 2007; Rull et al. 2007; Collins et al. 2008).

Development of pupal packing and shipping, adult

emergence and release methods, use of hormones,

nutrients and semiochemicals to improve mating

performance and implementation of field cage evalu-

ation of mating behaviour, are examples of post-fac-

tory improvements (Sivisnki et al. 1994; Shelly and

McInnis 2001; Hendrichs et al. 2002; Shelly et al.

2002b, 2007).

There is a large body of evidence that the addition

of protein to the diet of adult fruit flies enhances

their mating performance and other fitness attri-

butes. This could have positive effects in the applica-

tion of the SIT (Blay and Yuval 1997; Papadopoulos

et al. 1998; Yuval et al. 1998, 2002, 2007; Field and

Yuval 1999; Taylor and Yuval 1999; Kaspi and Yuval

2000; Kaspi et al. 2000; Shelly and Kennelly 2002;

Shelly et al. 2002a, 2005, 2006; Shelly and McInnis

2003; Cresoni-Pereira and Zucoloto 2006; Manrak-

han and Lux 2006; Perez-Staples et al. 2007, 2008a,

2009; Gavriel et al. 2009). In the case of Anastrepha

fruit flies, there is some evidence on the positive

effect of protein on their sexual behaviour (Aluja

et al. 2001, 2008; Cresoni-Pereira and Zucoloto

2006; Perez-Staples et al. 2008b). The purpose of this

research was to assess the effect of various pre-

release diets on the mating performance, pheromone

production and longevity of sterile A. ludens and

A. obliqua males.

Materials and Methods

Insects

Wild A. ludens flies were obtained as larvae from

infested sour oranges (Citrus aurantium L.) and wild

A. obliqua were obtained as larvae from infested yel-

low mombin fruits or jobos (Spondias mombin L.) col-

lected in the Soconusco region of Chiapas, Mexico.

The mass-reared non-irradiated and irradiated flies

were obtained as pupae from the MOSCAFRUT

mass-rearing facility at Metapa, Chiapas, Mexico.

Sterile flies were c-irradiated as pupae at 80 Gy

2 days before adult emergence. The mass-reared flies

were from strains with approximately 130 and 110

generations under mass rearing conditions, for

A. ludens and A. obliqua, respectively.

Mating performance

Initially, a series of five field cage mating tests were

carried out with A. ludens and one with A. obliqua.

The tests were done following the quality control

protocol commonly used at the MOSCAFRUT facility

(FAO/IAEA/USDA 2003).

After emergence, adult flies were sorted by sex

and placed in groups of 40 individuals in 30

cm · 30 cm · 30 cm Plexiglas cages. Before the tests,

these cages were maintained at the laboratory at

24 � 2�C, 70 � 10% RH, 550 � 50 lux light inten-

sity, with a 12 : 12 L : D photoperiod. Water and

food were provided ad libitum. The types of foods

were: (i) orange, Citrus sinensis (L.) Osbeck cv Valen-

cia or mango, Mangifera indica L. cv Ataulfo (�200 g

pieces); (ii) full diet, which was the standard 3 : 1

sugar : hydrolysed yeast laboratory diet; and (iii)

only sugar (standard cane sugar, dry sucrose). Wild

females always were fed a full diet.

Five field cages, 2.9 m diameter · 2 m height (Cal-

kins and Webb 1983), were set up at a mango orch-

ard near Tapachula, Chiapas, Mexico (14� 55¢ 08.9¢¢N, 92� 16¢34.2¢¢W and 137 msl). Six small orange

and mango potted trees (�1.5 m tall) were placed at

the periphery and central part of each field cage.

Each field cage was considered as a replicate. Thir-

teen males per treatment and 39 wild females were

released per field cage in the five A. ludens tests

(table 1). Twelve males per treatment and 48 wild

females in the A. obliqua test (table 2). Forty-eight

hours before the test, flies were individually marked

by gluing a small piece of paper printed with a num-

ber on their thorax for treatment identification. This

type of mark does not interfere with the sexual

activity of the flies (Meza et al. 2005). Mating pairs

were vial collected and the number and type of mat-

ings was recorded. The five tests for A. ludens were

made between 15:00 and 19:00 hours, and the test

for A. obliqua was between 06:00 and 12:00 hours.

In the tests for A. ludens we first compared wild

males fed on orange with sterile males fed on sugar

only and full diet. In the second, third and fourth

tests we compared the effect of orange, sugar only

and full diet on the performance of sterile, fertile

and wild males, respectively. In the fifth test the

effect of two fruits, mango and orange and full diet

was compared on sterile males. For A. obliqua only

Pre-release feeding on sterile Anastrepha male performance P. Liedo et al.

50 J. Appl. Entomol. 137(Suppl. 1) (2013) 49–60 ª 2010 Blackwell Verlag, GmbH

one test was done where wild males were fed on

orange and sterile males were fed on orange, full

diet and only sugar.

The age of the flies, at the time of the test was,

18–19 days for the wild flies and 12 days for the

mass-reared fertile and sterile flies in the case of A.

ludens, and 15 days for wild and 8 days for mass-

reared sterile A. obliqua flies.

A second set of tests were carried out to look at

the effect of different sugar : hydrolysed yeast (SY)

ratios. Laboratory and field cage conditions were as

described above. Groups of 200 males were provided

one of the following four diets: Only sugar (1 : 0),

and 24 : 1, 9 : 1 and 3 : 1 SY ratios. In the case of A.

ludens, eight sterile males from each diet treatment,

eight wild males and 40 wild females fed with the

standard 3 : 1 SY diet were released per field cage.

In the case of A. obliqua, 10 sterile males of each diet

treatment, 10 wild males and 50 wild females fed on

full diet were released in each field cage. Six repli-

cates were carried out per species.

Pheromone analysis

Pheromone volatiles emitted by males were collected

using an air-entrainment technique. Ten males were

confined in a 100 ml glass entrainment container

[4.8 cm inner diameter (ID) · 12.5 cm long]. Vola-

tiles were drawn from the container, using purifier

air that had previously passed through an activated

charcoal trap, onto a glass volatile collection trap

(4 mm ID · 40 mm long) containing 50 mg of Super

Q adsorbent (Alltech Associates, Deerfield, IL) (Heath

and Manukian 1992). Air was drawn through the

trap at a rate of 1 l/min by a vacuum pump. At the

conclusion of each air entrainment, which lasted 4 h,

the volatiles were eluted from the adsorbent with

200 ll of methylene chloride (Baker, HPLC grade)

and 100 gg of tridecane was added as an internal

standard for subsequent quantification. The samples

were kept at )20�C before analysis.

Volatiles were analysed by means of gas chromatog-

raphy-mass spectrometry using a Varian Star 3400 CX

gas chromatograph linked to a Varian Saturn 4D mass

spectrometer (GC-MS). The samples were analysed

using a fused silica column (30 m · 0.25 mm) coated

with poly (5%-diphenyl-95%-dimethylsiloxane) pro-

grammed from 50�C to 250�C at 15 �C/min. The

carrier gas was helium. The injector port temperature

was held at 200�C. Compounds were identified using

their retention times, Kovat index (KI), and mass

spectra and comparing these data with those of

synthetic standards. Synthetic standards of farnesene

(mixture of isomers that includes (E,E)-a-farnesene)

and (Z)-3-nonenol were supplied by Aldrich (Toluca,

Mexico).

Adult A. ludens sterile males were exposed to five

types of food: (i) standard laboratory full diet (3 : 1

SY dry mixture); (ii) pulp of orange fruits cv. Valen-

cia; (iii) pulp of mango fruits cv Ataulfo; (iv) com-

mercial pasteurized peach juice (Herdez �); and (v)

sugar only. These males were placed in 30 cm ·30 cm · 30 cm Plexiglas cages after emergence.

Every day a sample of 10 males per treatment was

taken and this was repeated during six consecutive

days, from 6- to 12-day-old flies. Four known main

pheromone components were quantified, (E,E)-a-

farnesene, suspensolide, anastrephin and epianastre-

phin.

Table 1 Mean number of matings (SE) achieved by Anastrepha ludens males from different strains in five separate field cage tests (five replicates

per test) after exposure to different food sources1

Sterile and wild males Sterile males Fertile males Wild males Sterile males/two fruits

Treatment No. of matings Treatment No. of matings Treatment No. of matings Treatment No. of matings Treatment No. of matings

W-O 14.4 (0.927)a S-O 8.2 (1.241)a F-O 10.0 (0.316)a W-0 10.6 (0.678)ab S-O 8.2 (0.964)ab

S-F 5.2 (1.114)b S-F 5.2 (0.374)b F-F 5.8 (0.735)b W-F 11.2 (0.707)a S-F 6.8 (0.867)b

S-S 5.2 (1.020)b S-S 4.8 (0.800)b F-S 5.2 (1.068)b W-S 6.4 (1.077) b S-M 10.2 (0.611)a

1Notation for treatments: the first letter corresponds to the strain and the second to the type of food. Strains: W: wild, S: mass-reared sterile, F:

mass-reared fertile. Food type: O: orange, M: mango, F: full diet (3 : 1 SY hydrolysate) and S: sugar only. Means follow by the same letter within

each column were not statistically different at P > 0.05.

Table 2 Mean number of matings (SE) achieved by A. obliqua sterile

and wild males exposed to different adult food sources, with wild

females in field cages

Treatment No. of matings (SE)

Wild – Orange 8.8 (1.16)a

Sterile – Orange 5.2 (1.62)ab

Sterile – Full 3.8 (1.02)b

Sterile – Sugar 3.0 (0.89)b

P. Liedo et al. Pre-release feeding on sterile Anastrepha male performance


Adult A. obliqua males were exposed to three food

sources: (i) standard laboratory 3 : 1 SY full diet: (ii)

pulp of mango fruits cv Ataulfo; and (iii) sugar only.

Pheromone components were collected from males

that were 8 to 10 days old. Three known com-

pounds, z-3-nonenol, (Z, E)-a-farnesene, and (E, E)-

a-farnesene, and one unknown compound, were

quantified.

Longevity

The effect of sugar : hydrolysed yeast ratios on the

longevity of mass-reared non-irradiated A. ludens

and A. obliqua fruit flies was evaluated under labora-

tory conditions by comparing four diets (only sugar

1 : 0, and 24 : 1, 9 : 1, and 3 : 1 S: : Y). Aluminium

frame, mesh covered, 80 cm · 50 cm · 15 cm cages,

were used, three cages per treatment. In each cage

about 2000 adult flies were released. Food, according

to each treatment, and water, were provided ad libi-

tum. Dead flies were removed daily from the cages.

The number and sex of the dead flies were recorded.

Data analysis

The data from the mating tests were analysed by

Fisher’s PLSD statistical test and P values were cal-

culated for pair-wise comparisons in each test. For

the pheromone analysis, the relative amount of

each compound was estimated from the GC graphs.

These figures were square root transformed for anal-

ysis of variance (anova). In the case of A. obliqua, a

Kruskal–Wallis test was used. Life tables were con-

structed for demographic analysis of longevity (Ca-

rey 1993). Survival data were analysed using the

Cox proportional hazard model (Everitt and Pickles

2004). Independent analyses were done for males

and females. Survival of flies exposed to 3 : 1, 9 : 1

and 24 : 1 SY diets was compared with survival of

flies fed with only sugar (control). Treatments were

compared with 95% CI. Statistical analyses were

performed using R software (R v.2.9.2. 2009, the R

Foundation for Statistical Computing, http://www.

r-project.org/).

Results

Mating

A total of 713 copulas were observed in the five ini-

tial tests with A. ludens, 56.1% were achieved by

males fed on fruit (even though only 40% of males

had been exposed to fruit), 28.7% by males fed on

full diet (33% of males on this sugar-yeast hydroly-

sed diet) and 15.1% by males on a sugar only diet

(27% of males present). The mean number (SE) of

matings recorded for A. ludens males for different

combinations of strain (wild, mass-reared fertile and

mass-reared sterile) and food type (orange and

mango fruit pieces, sugar only and full diet) in the

five initial tests is shown in table 1.

In the first of the five tests, with wild males fed

on orange and sterile males fed on sugar only or full

diet, the difference between wild males and sterile

males was significant (P < 0.0001), with the wild

ones achieving significantly more matings. The dif-

ference between sterile males fed on a full diet or

sugar only was not significant (P > 0.05). In order to

separate the effect of ‘wild’, from the effect of ‘fruit’,

the second test was designed using only mass-reared

sterile males (table 1). In this second test, the differ-

ences in matings between orange fruit and full diet

and orange fruit and sugar fed males were significant

in favour of fruit feeding (P = 0.0328 and P =

0.0181, respectively). The difference between full

diet and sugar only fed males was not significant

(P > 0.05). In the third test, mass-reared not steril-

ized males showed a similar pattern (table 1). The

difference in mating success between males fed on

fruit and males fed on full diet and sugar only was

significant (P = 0.0023 and P = 0.0009, respectively),

whereas the difference between males fed on full

diet and males fed on sugar only was not significant

(P > 0.05). Wild males showed a slightly different

mating pattern in the fourth test. There was no sig-

nificant difference between orange fruit and the full

diet (P > 0.05) and full diet was significantly differ-

ent from sugar only fed males (P = 0.0341) (table 1).

In the last test, while sterile males fed on mango

showed a significantly better mating performance

than those fed on the full diet (P = 0.0213), there

was no significant difference between males fed on

mango and those fed on orange, nor between those

fed on orange and those fed on full diet (P > 0.05)

(table 1).

In the field cage test with A. obliqua, a total of 104

matings were observed, 42.3% were by wild males

fed on orange, 25.0% by sterile males fed on orange,

18.3% by sterile males fed on full diet and 14.4% by

males fed on sugar only. The mean numbers of mat-

ings (SE) are shown in table 2. There was no signifi-

cant difference between sterile and wild males fed

with orange, while orange fed wild males performed

significantly better than sterile males fed on the full

diet or sugar. No significant differences were found

among sterile insect treatments.



Results from the second series of field cage mating

tests with males exposed to different SY ratios are

shown in table 3. A total of 123 and 86 matings

were observed for A. ludens and A. obliqua, respec-

tively. In both species, the mean numbers of matings

by wild males were significantly different from those

by sterile males fed on a sugar-only diet. There were

no significant differences between wild males and

sterile males fed on yeast diets, although the mean

number of matings tended to decline as the propor-

tion of yeast was reduced in the diet.

Pheromone

In A. ludens, diet induced differences in the amount

of the main pheromone compounds produced by

males were significant for (E,E)-a-farnesene

(F = 4.27, d.f. = 4, 12, P = 0.0223), anastrephin (F =

4.20, d.f. = 4, 12, P = 0.0234) and epianastrephin

(F = 5.57, d.f. = 4, 12, P = 0.0089). Differences in

the amount produced of suspensolide were not sig-

nificant (F = 2.42, d.f. = 4, 12, P = 0.1057). In A. ob-

liqua, differences were significant for Z-3-nonenol

(H = 3.86, d.f = 1, P = 0.05) and the unknown com-

pound (H = 6.49, d.f. = 2, P = 0.039), but non-sig-

nificant in the other two compounds (H = 4.36,

d.f. = 2, P = 0.113 and H = 4.27, d.f. = 2, P = 0.118

for Z,E-a-farnesene, and E,E-a-farnesene, respec-

tively).

In most cases, males fed with the standard full diet

produced the greatest amount of each compound

and males fed with only sugar produced the lowest

amount (figs 1 and 2). A. ludens males fed with

mango, orange or peach juice produced an interme-

diate amount of each compound. The only exception

was with (E, E)-a-farnesene in A. obliqua, where

males fed on mango produced greater amounts than

those fed on the full diet.

Longevity

Anastrepha ludens longevity response to pre-release

diets was complex. In males, the lowest expectation

of life was observed on the most yeast rich diet

(3 : 1 SY), followed by the non-yeast diet (sugar-

only). In females, the lowest life expectancy was

observed on the 3 : 1 diet, followed by the 9 : 1 diet

and the sugar only diet. The effect of the four SY

ratios on the survival of A. ludens can be seen in

fig. 3. According to the Cox proportional hazard

model and the Likehood Ratio test, differences in

survival were highly significant (table 4). The SY

ratio that resulted in the highest life expectancy was

the 24 : 1 ratio for both males and females.

The daily mortality rates (qx) during the first

30 days of A. ludens adult life are shown in fig. 4. In

males, the sugar only diet showed the lowest mortal-

ity rates during the first 20–24 days, then mortality

increased and was the highest after 30 days. In

females, the mortality rates under full diet (3 : 1 SY)

were the highest during the initial 30 days. The 9 : 1

and 24 : 1 SY diets showed the lowest mortality rates

through most of the lifespan of males and females.

The survival response of A. obliqua to the four SY

diets are shown in fig. 5 and the daily mortality rates

(qx) in fig. 6. Differences between flies fed on sugar

only and all other treatments were significant, except

with A. obliqua males fed on the 24 : 1 SY diet, where

the difference was non-significant (table 4). The most

protein rich diet (SY 3 : 1) showed the lowest life

expectancies for both, males and females. The highest

life expectancy was observed with the SY 24 : 1 diet

for both males and females.

Correspondingly, the highest mortality rates for

both A. obliqua males and females were found in the

3 : 1 SY diet and the lowest mortality rates were

observed in males and females fed on sugar only

and the 24 : 1 SY diet. The cross-over of the mortal-

ity schedules observed in A. ludens (fig. 4) was not

seen in A. obliqua females during the first 40 days of

adult life and was observed in males around 37 to

40 days old (fig. 6).

Discussion

Mating performance of A. ludens and A. obliqua males

was significantly affected by adult pre-release diet.

Fruit, and to a lesser extent, yeast enriched diets,

improved mating performance of wild, mass-reared

and sterile males. Generally, this is in agreement with

what has been reported for other fruit fly species

(Blay and Yuval 1997; Kaspi and Yuval 2000; Kaspi

Table 3 Mean number (SE) of matings achieved by wild and sterile

males of A. ludens and A. obliqua fed on diets with different SY

hydrolysate ratios in tests conducted in field cages

Strain – SY ratio A. ludens A. obliqua

Wild –3 : 1 5.00 (0.58)a 4.50 (0.81)a

Sterile –3 : 1 4.67 (0.61)ab 3.50 (1.34)ab

Sterile –9 : 1 4.33 (0.42)ab 2.50 (1.02)ab

Sterile –24 : 1 3.50 (0.76)ab 2.33 (0.56)ab

Sterile –1 : 0 3.00 (0.77)b 1.50 (0.43)b

Means followed by the same letter were not statistically different at

P > 0.05 according to Fisher’s PLSD.



a

b b b b

a

b b b b

a

ab ab ab

b

α-Farnesene

0

10

20

30

40

50

60

70

Sugar Full diet Orange Mango Peach juice

Treatments

Am

ount

(ng/

day)

Suspensolide

0

10

20

30

40

50

60


Treatments

Am

ount

(ng/

day)

Anastrephin

0

5

10

15

20

25

30

35


Treatments

Am

ount

(ng/

day)

Epianastrephin

0

20

40

60

80

100

120

140

160


Treatments

Am

ount

(ng/

day)

Fig. 1 Amount of main pheromone com-

pounds produced by sterile A. ludens males

exposed to five different sources of adult

food.



et al. 2000; Aluja et al. 2001; Yuval et al. 2002, 2007;

Shelly and McInnis 2003; Cresoni-Pereira and Zucol-

oto 2006; Shelly et al. 2006; Perez-Staples et al. 2007,

2008a, 2009; 2008a, 2009; Prabhu et al. 2008; Gavriel

et al. 2009). In the case of Anastrepha fruit flies, this is

in agreement with Aluja et al. (2001) for A. obliqua.

In A. ludens, although the mean numbers of copulas

by males from enriched diets were greater than the

number of matings by males fed on a sugar only diet,

these authors found no significant differences. This

could be attributed to particular experimental condi-

tions. Our results confirm that enriched diets improve

male mating performance in these two species and

this seems to be a universal phenomenon in pest

tephritid fruit flies.

The best performance for sterile males was

observed when they were provided with fruit. In

contrast, differences between the mating perfor-

mance of males fed on a full diet or on sugar only

were not significant in most cases. The better perfor-

mance of males fed on fruits could be explained as a

nutritional and/or as a microbiological effect. Also, it

could be attributed to the effect of possible semio-

chemicals as has been reported for Ceratitis capitata

(Wiedemann) and several Bactrocera species (Shelly

et al. 1996, 2005; Shelly and McInnis 2001; Papado-

pulous et al. 2006; Shelly and Edu 2007).

There was a non-significant trend for the mean

number of matings to increase as the yeast concen-

tration in the diets increased. The best mating per-

formance was achieved by males exposed to the diet

with the highest ratio (3 : 1, or 25% yeast). Prabhu

et al. (2008) found that 25% yeast was the optimal

concentration for mating performance of Bactrocera

tryoni (Froggatt). Above this concentration, the per-

cent of matings showed a decreasing tendency. We

did not test greater yeast concentrations because our

own unpublished observations and results by Cre-

soni-Pereira and Zucoloto (2001) on A. obliqua have

indicated that these adversely affect survival.

Post-teneral diets increased the amount produced

of three out of four main pheromone compounds in

A. ludens and two out of four in A. obliqua. Epsky

Z-3 nonenol

0

5

10

15

MangoSugarFull diet

MangoSugarFull diet

MangoSugarFull diet

MangoSugarFull diet

Treatment

Am

ount

(ng/

day) a

b

(Z,E)-α-farnesene

050

100150200250300

Treatment

Am

ount

(ng/

day)

(E,E)-α-farnesene

020406080

100

Treatment

Am

ount

(ng/

day)

Unkown

020406080

100120

Treatment

Am

ount

(ng/

day)

a

b

c

Fig. 2 Amount of main pheromone compounds produced by sterile

A. obliqua males exposed to three different sources of adult food.

Anastrepha ludens females

0

0.10.2

0.30.4

0.5

0.60.7

0.80.9

1

1 7 13 19 25 31 37 43 49 55 61 67 73 79 85 91 97 103109

Age (days)

Surv

ival

(lx)

¨3–1¨9–1¨24–1Sugar

Anastrepha ludens males

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

1 9 17 25 33 41 49 57 65 73 81 89 97 105 113 121 129 137

Age (days)

Sur

viva

l (lx

)

¨3–1¨9–1¨24–1Sugar

32.9Sugar

38.724 : 1

35.29 : 1

30.33 : 1

Life expectancy

33.0Sugar

35.724 : 1

32.09 : 1

26.93 : 1

Life expectancy

Fig. 3 Survival of male and female mass-reared A. ludens flies fed on

post-teneral diets with different SY ratios: 1 : 0 (only sugar), 3 : 1,

9 : 1 and 24 : 1.



and Heath (1993) found similar effects of food on

pheromone production in A. suspensa (Loew). These

results are consistent with our field cage mating

tests, confirming that adult nutrition has an effect

on mating performance and suggest that this effect

could be mediated, at least in part, by the effect of

food on pheromone production. However, the

amount produced by males fed on the full diet, was

greater than the amount produced by males fed on

fruits or fruit juice, whereas mating performance of

males fed on fruits was better than those fed on full

diet. This suggests that although there is an effect of

adult diet on pheromone production, there is not a

linear association between pheromone production

and mating success. Landolt and Sivinski (1992) also

found that overripe fruit increased pheromone call-

ing behaviour in A. suspensa. Thus, these results sug-

gest that the contribution of diet to male fitness may

be realized through several pathways – contribution

to quantity and quality of pheromones, increased

levels of courtship activity in the pre-copulatory

phase and possibly other post copulatory effects as

well (e.g. Aluja et al. 2008; Gavriel et al. 2009).

As in the case of mating performance, the results

obtained here on pheromone production suggest the

there might be nutritional, microbial or semiochemi-

cal factors that could have an effect on pheromone

synthesis. Because of possible basic and applied

implications, we believe the effect of fruits on mat-

ing behaviour and the relationship between phero-

mone production and mating performance, justify

further research.

The diets with the highest and the lowest protein

content (3 : 1 SY and sugar only) adversely affected

male and female longevity in both species. Sugar fed

flies showed low mortality rates in early adult life,

but in old flies (>40 days old), mortality rates of sugar

fed flies were higher than those fed on yeast enriched

diets. Mortality rates of flies fed on the richest diet

Table 4 Z-values obtained from survival analysis using the Cox pro-

portional hazard model, comparing each SY hydrolysate ratio with the

sugar only diet (control) for both sexes of Anastrepha species

Species Sex 3:1 SY 9:1 SY 24:1 SY

Likelihood

ratio test

A. ludens Male 5.165*** )7.717*** )16.424*** 515.3***

A. ludens Female 19.235*** 1.989* )9.740*** 830.4***

A. oblique Male 30.845*** 20.377*** )1.873 (NS) 1459.0***

A. oblique Female 36.801*** 19.288*** )5.164*** 2045.0***

***Highly significant differences, P < 0.0001; *significant difference,

P < 0.05; NS: non-significant difference, P > 0.05.

Anastrepha ludens males

00.010.020.030.040.050.060.07

0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30

Age (days)

Mor

talit

y ra

te (q

x) SY 3 : 1

SY 9 : 1SY 24 : 1

Sugar

Anastrepha ludens females

0

0.02

0.04

0.06

0.08

0.1

0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30

Age (days)

Mor

talit

y ra

te (q

x) SY 3 : 1

SY 9 : 1SY 24 : 1

Sugar

Fig. 4 Daily mortality rates (qx) of male and female mass-reared A. lu-

dens flies fed on post-teneral diets with different SY ratios: 1 : 0 (only

sugar), 3 : 1, 9 : 1 and 24 : 1.

Anastrepha obliqua males

0.0

0.10.2

0.3

0.40.5

0.6

0.7

0.80.9

1.0

Age (days)

Sur

viva

l rat

e (lx

)

Sugar¨3–1¨9–1¨24–1

Anastrepha obliqua females

0.00.1

0.20.3

0.40.50.6

0.70.8

0.91.0

1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 55 58 61 64 67

1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 55 58 61 64 67

Age (days)

Sur

viva

l rat

e (lx

)

Sugar¨3-1¨9-1¨24-1

21.8Sugar

22.024 : 1

17.79 : 1

16.13 : 1

Life expectancy

26.4Sugar

27.224 : 1

21.99 : 1

18.73 : 1

Life expectancy

Fig. 5 Survival of male and female mass-reared A. obliqua flies fed

on post-teneral diets with different SY ratios: 1 : 0 (only sugar), 3 : 1,

9 : 1 and 24 : 1.



(3 : 1 SY) were among the highest throughout the

lifespan of males and females in both species. This

longevity paradox was reported for A. ludens (Carey

et al. 2008) and studies on other fruit flies have

shown the adverse effect of yeast enriched diets on

male longevity (Carey et al. 1999; Jacome et al. 1999;

Kaspi and Yuval 2000; Prabhu et al. 2008). The

reduced longevity of flies fed on the sugar only diet

could be interpreted as malnutrition or depletion of

reserves (Warburg and Yuval 1996; Nestel et al.

2005). The adverse effect of yeast enriched diets could

be explained as the trade-offs between longevity and

reproduction. Protein enriched diets stimulate repro-

duction, and reproductive effort (egg production for

females, calling, intrasexual interactions, courtship

and sperm and accessory gland substances production

for males, among others) could result in higher mor-

tality (Carey et al. 1999, 2001, 2002a,b, 2008; Muller

et al. 2001). The adverse effect of pre-release diets on

longevity could be overcome by the ability of flies to

forage for natural food sources (Maor et al. 2004). Ut-

ges et al. (unpublished data) found that A. ludens and

A. obliqua flies fed on fruit and sugar before release

showed the greater recapture and dispersal distance,

compared with flies fed on the full or sugar only diets

in release–recapture field tests.

In terms of SIT application, since sterile males

have only limited sperm available for very few

matings, improved mating competitiveness should

have priority over extended lifespan.

After careful analysis of pre-release feeding trade-

offs, Yuval et al. (2007) concluded that yeast enriched

diets will be highly recommended for C. capitata SIT

programmes. Following the same approach and con-

sidering mating performance, laboratory longevity

and field survival and dispersal, the 24 : 1 SY diet

could be recommend for A. ludens and A. obliqua SIT

programmes. The mating competitiveness of males

fed on this 24 : 1 SY diet was similar to those males

fed on the most protein rich diet, without the detri-

mental effect on longevity.

Further research to understand the mechanisms

underlying the enhanced mating performance of

fruit fed males, similar to those by Papadopolous

et al. (2006) and development of methods for pre-

release feeding using fruits or fruit products, are

highly recommended.

Acknowledgements

We thank R. Bustamante, E. de Leon, J. A. Escobar-

Trujillo, M.P. Perez-Gomez, R. Rincon, G. Rodas, S.

Rodrıguez, S. Salgado, and J. L. Zamora-Palomeque

for technical assistance. We also thank J. Valle-Mora

for advice on statistical analysis and B. Yuval for

review of earlier versions. This research was sup-

ported by the Food and Agriculture Organization/

International Atomic Energy Agency (FAO/IAEA)

through contract 12860/RBF, the MOSCAFRUT facil-

ity (SENASICA – SAGARPA-IICA) and ECOSUR. The

Consejo Nacional de Ciencia y Tecnologıa (CONA-

CYT) and ECOSUR provided support through sabbati-

cal leave to PL.

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Effect of post-teneral diets on the performance of sterile Anastrepha ludens and Anastrepha obliqua fruit flies

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