Cytogenetic analysis of Malpighian tubule and salivary gland polytene chromosomes of Bactrocera oleae (Dacus oleae) (Diptera: Tephritidae).

Photomaps of the Malpighian tubule and the salivary gland polytene chromosomes of Bactrocera oleae (Dacus oleae) are presented and compared with those of the fat body. Five polytene chromosomes (10 polytene arms) corresponding to the five autosomes of the mitotic nuclei, as well as a heterochromatic mass corresponding to the sex chromosomes, are observed in the nuclei of the three somatic tissues. The most prominent features of each polytene chromosome, the reverse tandem duplications, as well as the rather unusual ectopic pairing of the telomeric regions of different chromosome arms, are described. The constancy of the banding pattern based on the analysis of the three larval tissues is discussed.     

Olive fruit fly [Bactrocera (Dacus) oleae (Rossi) (Diptera: Tephritidae)] adult rearing diet without antibiotic

The olive fruit fly [Bactrocera (Dacus) oleae (Rossi) (Diptera: Tephritidae)] adult diet since its development in the 1960’s regularly incorporates antibiotic. Considering recent findings on the importance and function of the indigenous microbial flora of insects, the effects of antibiotic removal were measured on the survival and egg laying of wild flies derived from McPhail trappings and from field infested olive fruits. In the first case wild flies fed no antibiotic laid significantly greater numbers of eggs for two generations (in 5 out of 10 and 2 out of 10 counting dates for G1 and G2 respectively), while there were no significant differences in survival in either test (P = 0.221 for P generation, P = 0.988 for G1 generation from McPhail traps, P = 0.056 for flies from infested fruits). Percent egg–pupa recovery and adult emergence were not significantly affected by lack of antibiotic. An antibiotic‐free strain has been maintained for eight generations, showing acceptable performance when compared to the long‐reared standard ‘Laboratory’ strain. Overall results suggest the feasibility of an adult diet free of antibiotic without negative effects on colony survival and performance.     

Mitotic and polytene chromosome analysis in Dacus oleae (Diptera: Tephritidae).

The present study constitutes the first attempt to construct a photographic map of the polytene chromosomes of Dacus oleae, a pest of the olive tree that causes serious financial damage in all olive oil producing countries. The map was constructed by using the larval fat body cells, the chromosomes of which are representative of the polytene chromosomes of other polytene tissues. In addition, the mitotic chromosomes of brain ganglia were examined, permitting tentative correlations between mitotic and polytene elements. This investigation shows that D. oleae is suitable for cytogenetic analysis in both mitotic and polytene chromosomes, a fact that may prove very useful for obtaining more detailed genetic information on the pest's natural populations.     

Genetic and cytogenetic analysis of the olive fruit fly Bactrocera oleae (Diptera: Tephritidae)

The genetic and cytogenetic characteristics of one of the major agricultural pests, the olive fruit fly Bactrocera oleae, are presented here. The mitotic metaphase complement of this insect consists of six pairs of chromosomes including one pair of heteromorphic sex chromosomes, with the male being the heterogametic sex. The analysis of the polytene complements of three larval tissues, the fat body, the salivary glands and the Malpighian tubules of this pest has shown (a) a total number of five long chromosomes (10 polytene arms) that correspond to the five autosomes of the mitotic nuclei and a heterochromatic mass corresponding to the sex chromosomes, (b) the constancy of the banding pattern of the three somatic tissues, (c) the absence of a typical chromocenter as an accumulation of heterochromatin, (d) the existence of reverse tandem duplications, and (e) the presence of toroid tips of the chromosome arms. The in situhybridization of genes or DNA sequences to the salivary gland polytene chromosomes of B. oleaeprovided molecular markers for all five autosomes and permitted the establishment of chromosomal homologies among B. olea, B. tryoniand Ceratitis capitata. The heat shock response of B. oleae, as revealed by heat-inducible puffing and protein pattern, shows a higher thermotolerance than Drosophila melanogaster.     

Genetic differences among natural and laboratory-reared populations of the olive-fruit fly Dacus oleae (Diptera: Tephritidae)

Twelve wild populations of the olive-fruit fly Dacus oleae from around the Mediterranean basin and two laboratory populations were examined for their electrophoretic profiles at two esterase loci. The genetic distances among wild and laboratory populations were consistently greater than the distances among wild populations. There was a pattern in this genetic differentiation, in that the most common allele decreased in frequency during culture in the laboratory, thus causing an increase of heterozygosity in laboratory populations. The findings are discussed in the light of assessing the genetic quality of mass-laboratory (or factory) cultures of economically important insects.

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Zusammenfassung Die elektrophoretischen Profile zweier Esterase-Loci von zwölf natürlichen Populationen der Olivenfliege Dacus oleae aus Ländern des Mittelmeerbeckens und von zwei Laboratorium-Populationen wurden untersucht. Die genetischen Abstände zwischen natürlichen und Laboratorium-Populationen waren konsistent grösser als die Abstände zwischen natürlichen Populationen. Bei dieser genetischen Differenzierung zeigte sich eine Regelmässigkeit. Die häufigsten Allele nahmen während der Zucht im Laboratorium in ihrer Häufigkeit ab, was zu einer Zunahme der Heterozygosis in Laboratorium-Populationen führte.Diese Resultate werden diskutiert im Zusammenhang mit der genetischen Qualität von Massenzuchten wirtschaftlich wichtiger Insekten im Laboratorium und in der Industrie.

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On leave of absence from Department of Biology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 4J1.

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This is paper No. VII of the series The Genetics of Dacus oleae     

Effect of larval density and host species on preimaginal development of Bactrocera tau (Diptera: Tephritidae)

The fruit fly, Bactrocera tau (Walker) (Diptera: Tephritidae), is an important pest of fruit and vegetable crops. In this study, host preference of B. tau females and the effects of host species and larval density on larval survival, pupal weight, adult emergence, and developmental duration were investigated on cucumber (Cucumis sativus L.), sponge gourd (Luffa cylindrical L. (Roem)), bitter gourd [Momordica charantia (Cucurbitaceae) L.], guava [Psidium guajava (Myrtaceae) L.], and tangerine [Citrus reticulata (Rutaceae) (Blanco)]. The results showed that females preferred to cucumber over other host species. Larval feeding experience affected subsequent host oviposition preference of adult females. Host species and initial larval density affected certain aspects of the biology of B. tau. Larval density negatively affected insect performance. Survival rates at low densities were significantly higher than that at high densities. Total developmental duration reduced at high larval densities. Cucumber was more suitable to larval growth. Larvae on cucumber grew faster and the puparia were heavier than that on other host species. Larval survival, pupation rate and adult emergence were higher on cucumber compared with those in other host species. Oviposition preference of adult females was correlated with performance of their offspring.     

Insecticidal activity of Citrus aurantium peel extract against Bactrocera oleae and Ceratitis capitata adults (Diptera: Tephritidae)

Citrus aurantium (L.) peel extracts in petroleum ether were evaluated for toxicity against olive fruit fly, Bactrocera oleae (Gmelin) and medfly, Ceratitis capitata (Wiedemann) adults. Bactrocera oleae flies were more susceptible to the extract than C. capitata in contact and residual bioassays. Fumigation bioassay had no effect. Both sexes of B. oleae were equally susceptible in both types of bioassays. However, males of C. capitata were more susceptible than the conspecific females. LD₅₀ values (concentration causing 50% mortality) after 96 h for the males and females of B. oleae were 44.8 and 40.1 μg/insect in contact bioassay through topical application. Whereas, LD₅₀ values for the males and females of C. capitata were 38.8 and 67.8 μg/insect respectively. LC₅₀ values after 96 h for the males and females of B. oleae were 18.8 and 17.8 μg/cm² in Petri dish residual bioassay. Whereas, LC₅₀ values for the males and females of C. capitata were 70.6 and 147.1 μg/cm² respectively.
Fractionation of the extract on a silica gel column with three different polarity solvents resulted in three fractions with only the intermediate polarity solvent fraction having substantial insecticidal activity. Toxicity of the mixtures of active and inactive fractions was equal to the original extract. Our results indicate that C. aurantium has potential for controlling insect pests.     

Structure and morphogenesis of the eggshell and micropylar apparatus in the olive fly, Dacus oleae (Diptera: Tephritidae)

The egg of the olive fly, Dacus oleae (Diptera, Tephritidae), is laid inside olives and the larva eventually destroys the fruit. The oocyte is surrounded by several distinct layers which are produced during choriogenesis. The chorion covering the main body of the egg outside of the vitelline membrane includes a "wax" layer, an innermost chorionic layer, an endochorion consisting of inner and outer layers separated by pillars and cavities similar to their counterparts in Drosophila melanogaster, as well as inner and outer exochorionic layers. The anterior pole is shaped like an inverted cup, which is chiefly hollow around its base and has very large openings communicating with the environment. Holes through the surface of the endochorion result from deposition of endochorionic substance around follicular cell microvilli. An opening at the apex of the cup provides an entrance for sperm entering the micropylar canal, which traverses the endochorion and continues into a "pocket" in a thickened vitelline protrusion. The micropylar canal is formed by deposition of endochorion and vitelline membrane around an elongated pair of follicular cell extensions. These extensions later degenerate and leave an empty canal about 5 microns in diameter and the narrower pocket about 1 micron in diameter. Respiration is thought to be facilitated by openings at the base of the anterior pole as well as by openings through the "plastron" around the main body of the shell.     

Cage study of spinosad-based bait efficacy on Bactrocera cucurbitae, Dacus ciliatus, and Dacus demmerezi (Diptera: Tephritidae) in Reunion Island

On Reunion Island, Bactrocera cucurbitae (Coquillett), Dacus ciliatus (Loew), and Dacus demmerezi (Bezzi) cause severe damage to Cucurbit crops. The aim of the study was to test in field cages the effectiveness of Synéis-appat (Dow AgroSciences), a spinosad-based bait (0.02% of spinosad) on both attraction and mortality of young adults (6-9 d old) of these three species. The effects of gender were also evaluated for all species whereas the effects of protein deprivation were tested with B. cucurbitae only. For the first 15 min after application, significantly more B. cucurbitae adults (21.7 +/- 1.8%) were attracted to the bait than D. demmerezi (7.6 +/- 2.4%) and D. ciliatus (2.7 +/- 1.4%); the subsequent response (30-75 min after bait application) of D. demmerezi was statistically similar to that recorded for B. cucurbitae; whereas the response ofD. ciliatus to the bait was consistently significantly lower. Adult mortality was significantly higher for B. cucurbitae (94.6 +/- 0.7%) than for D. demmerezi (85.7 +/- 2.1%), and was significantly higher for the latter than for D. ciliatus (60.4 +/- 4.4%). Sex had no significant effect on the mortality rate for each species. The efficiency of the bait was the same for B. cucurbitae adults regardless whether or not the diet included proteins. Overall, Synéis-appat appears to be more effective against B. cucurbitae and B. demmerezi than against D. ciliatus. In Reunion Island, this bait could constitute a useful component in the framework of Integrated Pest Management.     

The importance of dietary amino acids on the reproduction and longevity of adult Dacus oleae (Gmelin) (Diptera Tephritidae)

When the amino-acid mixture of an effective chemically defined diet was replaced by single amino acids, keeping the total nitrogen at the same level, the egg production of Dacus oleae was minimal with all the 19 amino acids tested. Male survival was adversely affected by the amino acids : alanine, aspartic acid, glutamic acid, glycine, hydroxyproline, lysine, serine and tyrosine, while female survival was shortened when the amino acids : glycine, hydroxyproline and lysine were added. The creation of amino-acid imbalances, by deleting the 19 amino acids individually, from the complete amino-acid mixture, showed that the amino acids : arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, serine, threonine, tryptophane and valine were indispensable to the adult Dacus oleae flies, as far as egg production is concerned. Survival of the male flies was significantly shortened when the amino acids : alanine, hydroxyproline and tryptophane were omitted. Significant differences in longevity between males and females were scored, when the amino acids : alanine, aspartic acid, cystine, glycine and tryptophane, were omitted.     

Inhibition of larval growth of Dacus oleae (Diptera: Tephritidae) by streptomycin II. Effect of treating the parents

Inhibition of growth of Dacus oleae larvae in untreated green olive fruits occurred in a high percentage when the parents were treated, in the laboratory, with 1% streptomycin sulphate plus a spreader-wetter in the following ways: — (a) brief immersion of the olives in which the parent insects grew as larvae; (b) brief immersion as fully grown larvae; (c) spraying of adults; (d) allowing parent females to oviposit for a few hours in treated olives, prior to ovipositing in untreated ones. In the last case, the degree of inhibition of progeny growth was related to the time between the maternal treatment and the birth of the progeny.

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Résumé Le développement larvaire de Dacus oleae dans des olives vertes non-traitées est inhibé à un pourcentage élevé quant les parents ont été traités, au laboratoire, avec 1% de sulfate de streptomycine associé à un produit dispersant-mouillant, selon les modalités suivantes: (a) immersion brève des olives dans lesquelles les parents se sont développés comme larves; (b) immersion brève des larves à développement complet; (c) pulvérisation des adultes; (d) oviposition des femelles pour quelques heures dans des olives traitées, avant de pondre dans des olives non-traitées. Dans ce dernier cas, l'inhibition de la descendance fut influencée par le temps séparant le traitement de la mère et la naissance de la descendance.

     

Amino-acid synthesis in adult Dacus oleae (Gmelin) (Diptera Tephritidae) determined with [U-14C] glucose

The ability of adult Dacus oleae for amino-acid synthesis from [U-14C] glucose was investigated. The relatively high specific activity radiometric measurements indicated that both sexes were able to synthesize the amino acids: alanine, aspartic acid, cystine, glutamic acid, glycine, hydroxyproline, proline and tyrosine; therefore, these amino acids are considered as nutritionally dispensable for D. oleae. On the other hand, the amino acids: arginine, histidine, leucine, isoleucine, lysine, methionine, phenylalanine, serine, threonine, and valine, showed a very low specific activity and therefore are considered as nutritionally indispensable. It was not possible to conclude about tryptophane, since the acid hydrolysis destroyed this amino acid.     

Bacteria associated with four species of Dacus (Diptera: Tephritidae) and their role in the nutrition of the larvae

Summary The bacteria associated with Dacus tryoni (Froggatt), Dacus jarvisi (Tryon), Dacus neohumeralis (Hardy) and Dacus cacuminatus (Hering) were examined. Bacteria were isolated from the surface of freshly-laid eggs, from within surface sterilised pupae, from heads and abdomens of wild and laboratory-maintained flies, and from decomposed fruits in which the wild larvae were feeding. A more diverse flora was associated with D. tryoni and D. jarvisi (15 and 14 species, respectively) than with D. neohumeralis and D. cacuminatus (9 and 6 species, respectively). Most of the bacteria belonged to the family Enterobacteriaceae and while there were similarities of bacterial associations between fly species there was no evidence of a strict symbiotic association of a particular bacterium or bacteria with each species of fly. The larvae of D. jarvisi were unable to develop normally in an artificial medium containing unhydrolysed protein and free of bacteria and on a medium containing casein and Serratia liquefaciens (isolated from the flies and shown to secrete protease) the larvae died. On the same casein medium containing Enterobacter cloacae (isolated from the flies and shown to be protease negative) the larvae developed normally. Larvae of D. tryoni and D. jarvisi were devoid of protease and cellulase activity, but contained some amylase activity. The significance of these results in terms of current hypotheses concerning symbioses between tephritids and bacteria is discussed.     

Respiration and nutrition of radiation-sterilized female Dacus cucumis (Diptera: Tephritidae)

The oxygen consumption of flies irradiated with between 10 kilorads (krad) and 80 krad was not demonstrably different between doses, or when compared with untreated controls.Carbohydrate intake and initial body weights of irradiated flies were less than normal flies. However, the 10 krad irradiated flies accumulated a greater volume of fat body than normal flies, presumably because of failure of vitellogenesis in the former. Accumulation of fat body by 40 krad flies was initially retarded compared with 10 krad flies, but they later accumulated fat body at the same rate as the 10 krad flies. It is suggested that this was due to an initial hormonal disturbance caused by the higher dose.No degenerative changes were noted in the midgut of the 10 krad or 40 krad flies. The protein intake of these flies was similar to that of normal flies.

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Zusammenfassung Einige Autoren haben wahrscheinlich machen können, daß sterilisierende Strahlendosen bei einigen Insektengruppen den Stoffwechsel beeinflussen. Jedoch sind für die Dipteren wenig Beweise für solche durch Bestrahlung hervorgerufene Störungen veröffentlicht worden. Die in dieser Arbeit beschriebenen Versuche waren darauf gerichtet, die Einflüsse der Bestrahlung auf einige physiologische Systeme von weiblichen Dacus cucumis zu bestimmen.Auf den Sauerstoffverbrauch konnte eine Wirkung von Strahlendosen von 80 Kilorad (krad) nicht festgestellt werden.Beim Vergleich von unbestrahlten Fliegen mit solchen, die 10 oder 40 krad erhalten haben, wurde gefunden, daß in beiden Fällen die bestrahlten Tiere zunächst weniger als normale wogen, daß aber später die Wachstumsrate der bestrahlten Fliegen anstieg, so daß ihr Gewicht sich schließlich dem der normalen anglich.Der Verbrauch von Eiweißlösung durch mit 10 und 40 krad bestrahlten Fliegen war dem der normalen ähnlich. Jedoch verbrauchten die bestrahlten Fliegen eine geringere Menge Zuckerlösung als die normalen. Es wird angenommen, daß darin der Ausfall des Kohlenhydratbedarfs durch die sterilen Ovarien zum Ausdruck kommt.Die mit 10 krad bestrahlten Fliegen bildeten ein größeres Fettkörpervolumen als die normalen. Es wird vermutet, daß dies eine kombinierte Folge der Nicht-Auflösung des pupalen und des gesteigerten. Aufbaus des adulten Fettkörpers bei Ausschaltung funktionierender Ovarien darstellt.Der Aufbau des Fettkörpers war anfänglich bei den mit 40 krad bestrahlten Fliegen gegenüber den mit 10 krad bestrahlten gehemmt, aber nach einer Verzögerung war die Zuwachsrate den mit 10 krad bestrahlten ähnlich. Es wird angenommen, daß das auf einer anfänglichen Hormonstörung bei den mit 40 krad bestrahlten Fliegen beruhen konnte.

     

Sex chromosomes and associated rDNA form a heterochromatic network in the polytene nuclei of Bactrocera oleae (Diptera: Tephritidae)

The olive fruit fly, Bactrocera oleae, has a diploid set of 2n?=?12 chromosomes including a pair of sex chromosomes, XX in females and XY in males, but polytene nuclei show only five polytene chromosomes, obviously formed by five autosome pairs. Here we examined the fate of the sex chromosomes in the polytene complements of this species using fluorescence in situ hybridization (FISH) with the X and Y chromosome-derived probes, prepared by laser microdissection of the respective chromosomes from mitotic metaphases. Specificity of the probes was verified by FISH in preparations of mitotic chromosomes. In polytene nuclei, both probes hybridized strongly to a granular heterochromatic network, indicating thus underreplication of the sex chromosomes. The X chromosome probe (in both female and male nuclei) highlighted most of the granular mass, whereas the Y chromosome probe (in male nuclei) identified a small compact body of this heterochromatic network. Additional hybridization signals of the X probe were observed in the centromeric region of polytene chromosome II and in the telomeres of six polytene arms. We also examined distribution of the major ribosomal DNA (rDNA) using FISH with an 18S rDNA probe in both mitotic and polytene chromosome complements of B. oleae. In mitotic metaphases, the probe hybridized exclusively to the sex chromosomes. The probe signals localized a discrete rDNA site at the end of the short arm of the X chromosome, whereas they appeared dispersed over the entire dot-like Y chromosome. In polytene nuclei, the rDNA was found associated with the heterochromatic network representing the sex chromosomes. Only in nuclei with preserved nucleolar structure, the probe signals were scattered in the restricted area of the nucleolus. Thus, our study clearly shows that the granular heterochromatic network of polytene nuclei in B. oleae is formed by the underreplicated sex chromosomes and associated rDNA.     

Alcohol dehydrogenase allele frequencies in Bactrocera oleae (Dipt., Tephritidae): effect of ethanol addition in larval diet

Abstract: The effect of ethanol in larval medium on Bactrocera oleae larvae was examined at four concentrations. Ethanol exerted a differential effect on the three alcohol dehydrogenase allele frequencies. While originally being at equilibrium under laboratory conditions, after three generations of larval development in a diet containing ethanol at 1% concentration, Adh -F allele frequency increased, that of Adh -I dropped significantly and the frequency of Adh -S remained unaltered. Adh -S allele seems to be adapted in nature where only minor quantities of alcohol are present in the insects' natural host, while Adh -I is best adapted in the alcohol-free laboratory culture medium. The frequency of Adh -F allele remains unaltered when feral populations are introduced in the laboratory.     

Biology of Bactrocera carambolae (Diptera: Tephritidae) on four hosts

Bactrocera carambolae is a quarantine pest found in Brazil, restricted to the states of Amapá, Pará and Roraima. This fruit fly can potentially cause extensive socioeconomic and environmental damage in the country, if it disperse into areas where fruit is grown for exporting. The objective of this work was to study the biology of B. carambolae on fruits of Averrhoa carambola L. (Oxalidaceae), Psidium guajava L. (Myrtaceae), Spondias mombin L. (Anacardiaceae) and Eugenia stipitata McVaugh (Myrtaceae). The following parameters were investigated: duration of egg-larva, pupal, egg-adult, pre-oviposition, oviposition and post-oviposition periods, pupal weight and viability, sex ratio, fecundity, fertility and longevity. All parameters except pupal weight, oviposition and post-oviposition period, egg fertility and sex ratio were influenced by the host plant on which the larvae were reared. The carambola fruit fly completes its development on all those hosts studied here, with the highest fecundities on A. carambola and P. guajava.