Acrosome formation in Ceratitis capitata (Diptera, Tephritidae)

The stages of morphogenesis of the acrosome of Ceratitis capitata are well defined. This organelle is formed by the Golgi complex and, as it matures, takes up a position laterally in relation to the anterior region of the sperm nucleus. An interstitial membrane marks the area of contact between nucleus and acrosome in the spermatid, and is found even in mature sperm cells. The acrosome contains hydrolytic enzymes, as detected by acid phosphatase reaction.     

Choriogenesis in the medfly Ceratitis capitata (Wiedermann) (Diptera : Tephritidae)

Four chorionic stages (11, 12, 13, 14) can be discerned in the medfly Ceratitis capitata (Diptera : Tephritidae) by light microscope. More detailed staging (stages 11A, 11B, 12A, 12B, 12C, 13A, 13B, 13C, 14A and 14B) is possible only by electron microscope. Throughout these stages, the first chorionic layer (wax layer) is formed at stage 11A, followed by the formation of the innermost chorionic layer (stage 12A), the inner endochorion, the pillars and the cavities of the first trabecular layer (stage 12), creation of the second trabecular layer and the chorionic network (stage 13), and finally the secretion of the exochorionic layers (stage 14). Pulse-chase autoradiography has revealed that the follicle cells are responsible for the synthesis of all proteinaceous layers. No extensive regional complexity is observed besides the existence of the micropylar apparatus and aeropyles in the anterior pole. Biochemical analysis has revealed several eggshell proteins and their stage-specific synthesis: two intermediate molecular-weight proteins are the major chorion proteins; in addition, there are 2 more groups comprising 6 proteins, which might be characteristic for the different chorion layers as can be deduced by their stage-electrophoretic pattern.     

Cross-resistance to insecticides in a malathion-resistant strain of Ceratitis capitata (Diptera: Tephritidae)

Resistance to malathion has been reported in field populations of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), in areas of Spain where an intensive use of this insecticide was maintained for several years. The main goal of this study was to determine whether resistance to malathion confers cross-resistance to different types of insecticides. Susceptibility bioassays showed that the malathion-resistant W-4Km strain (176-fold more resistant to malathion than the susceptible C strain) has moderate levels of cross-resistance (three- to 16-fold) to other organophosphates (trichlorphon, diazinon, phosmet and methyl-chlorpyrifos), the carbamate carbaryl, the pyrethroid lambda-cyhalothrin, and the benzoylphenylurea derivative lufenuron, whereas cross-resistance to spinosad was below two-fold. The W-4Km strain was selected with lambda-cyhalothrin to establish the lambda-cyhalothrin-resistant W-1Klamda strain (35-fold resistant to lambda-cyhalothrin). The synergistic activity of the esterase inhibitor DEF with lambda-cyhalothrin and the increase in esterase activity in the W-1Klamda strain suggests that esterases may be involved in the development of resistance to this insecticide. Our results showed that resistance to malathion may confer some degree of cross-resistance to insecticides currently approved for the control of Mediterranean fruit fly in citrus crops (lambda-cyhalothrin, lufenuron, and methyl-chlorpyrifos). Especially relevant is the case of lambda-cyhalothrin, because we have shown that resistance to this insecticide can rapidly evolve to levels that may compromise its effectiveness in the field.     

Cold treatment of Ceratitis capitata (Diptera: Tephritidae) in oranges using a larval endpoint

South Africa currently exports fresh citrus (Citrus spp.) fruit to Japan using an in-transit cold treatment protocol of 14 d or 12 d at temperatures <0 degrees C for treatment of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) in 'Clementine' mandarins (Citrus reticulata Blanco) and other citrus types, respectively. To reduce the risk of chilling injury with this treatment, research was conducted with temperatures >0 degrees C. Earlier South African research had shown that young (6-d-old) larvae were slightly more tolerant of cold treatment and that there were no significant differences between cold tolerance of these larvae in different citrus types [oranges, Citrus sinensis (L.) Osbeck; grapefruits, Citrus paradisi Macfad.; lemons, Citrus limon (L.) Burm.f.; and mandarins). Due to their ready availability, 'Valencia' oranges were used in this study. When 62,492 larvae in total were treated in three replicates at a mean temperature of 1.5 degrees C for 16 d, there were three larval survivors. The trial was therefore repeated with oranges using a 16-d period at a mean temperature of 1.0 degrees C and a mean of 1.4 degrees C for the hourly maximum probe readings. Three replicates were again conducted and the resultant mean mortality in the control was 8.1% of 21,801 larvae, whereas the cold treatment mortality was 100% of 71,756 larvae. This treatment at a mean temperature of 1 degree C exceeded the Japanese confidence level requirement and also exceeded the Probit-9 mortality level, but not at a confidence level of 95%. These data support the establishment of a treatment protocol of 16 d at temperatures <1.4 degrees C, commencing once all fruit pulp probes reach a temperature of 1 degree C or lower.     

Toxicity of citrus essential oils against Ceratitis capitata (Diptera: Tephritidae) larvae

Citrus peel essential oils are considered to constitute the most important resistance factor of citrus fruits against fruit flies. Essential oils were obtained from three sweet orange varieties, one bitter orange and one lemon variety. Yield, chemical composition and toxicity against neonates of the Mediterranean fruit fly were determined. Based on chemical analysis, the toxicity of commercially purchased major and minor components (monoterpenes and sesquiterpenes) of essential oils was determined. In addition, fractions were prepared to evaluate the role of minor components in the toxicity of crude essential oils. Limonene was by far the most abundant ingredient (96.2–97.4%) in all sweet orange varieties and in bitter orange, while the concentration of limonene was much lower in lemon essential oils (74.3%). Orange and bitter orange essential oils were more toxic than lemon essential oils. The toxicity of orange and bitter orange essential oils was similar to that of their major component limonene. In tests of commercially purchased chemicals, the oxygenated components of essential oils were more toxic than hydrocarbons but their low concentration in citrus essential oils could not affect the toxic activity of essential oils. The presence of α-pinene and β-pinene seems to account for the lower toxicity of lemon essential oils in relation to other citrus essential oils. The importance of understanding the toxicity of essential oils in relation to their composition and their role regarding the resistance of citrus fruits to Ceratitis capitata infestation is discussed.     

Conservation of nucleolar structure in polytene tissues of Ceratitis capitata (Diptera: Tephritidae)

Nucleolar structure was studied in mitotic and three polytene tissues of the Mediterranean fruit fly, Ceratitis capitata using in situ hybridization with a tritium-labelled rDNA probe and silver staining. In mitotic metaphase chromosomes nucleolar organiser regions were localised in the short arms of both sex chromosomes. In polytene nuclei of trichogen cells, salivary glands and fat body rDNA was detected within nucleoli. Nucleoli in these tissues have a similar structure with rDNA labelling concentrated in a central core. Silver staining resulted in very heavy staining of polytene nucleoli and interphase nucleoli in diploid cells. Silver staining of nucleolar organisers in metaphase chromosomes is weak or absent although the X chromosome has numerous dark silver bands in other locations. The results suggest that nucleolar structure is conserved in polytene tissues contrasting with the variability of autosomal banding patterns and sex chromosome structure. They also indicate that silver staining is not necessarily specific for nucleolar regions.     

Inability of Ceratitis capitata (Diptera: Tephritidae) to overwinter in the Judean hills

The overwintering potential of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), in cold winter areas within its northern distribution is a key element in understanding its ecology. Recent studies have suggested that although originating in tropical Africa, the fly has become adapted to the cold weather that prevails within its northernmost areas of distribution. We address the question of whether the Mediterranean fruit fly has expanded its overwintering range to include the mountains of central Israel. Doing so would imply that the fly has developed either a behavioral or a physiological mechanism to cope with low temperature and/or damp conditions in combination with cold. We monitored adult populations year round, sampling fruit, calculating expected emergence days for overwintering flies, and studying adults captured within dense and sparse apple orchards. We also performed several manipulative experiments to study preimago ability to survive the winter under natural or seminatural conditions. The study was conducted in the central mountains of Israel at 700-m altitude from 1994 to 2003. Comparison experiments also were conducted at 400 m and at sea level. Our results show 1) no adults captured during the winter and spring, 2) an absence of new infestations during the winter and spring, and 3) inability of preimago stages to overwinter in the central mountains of Israel. Thus, we conclude that the fly does not overwinter in the central mountains of Israel. We discuss the ecological and applied significance of our findings.     

Chemosterilants as control agents of Ceratitis capitata (Diptera: Tephritidae) in field trials

Lufenuron is a chitin synthesis inhibitor, which is able to impede Mediterranean fruit fly, Ceratitis capitata (Wiedemann), reproduction. In laboratory trials, following ingestion of lufenuron, the eggs laid by female Ceratitis capitata were prevented from hatching. In field trials in Valencia, Spain, lufenuron showed its effectiveness by reducing C. capitata wild populations and its continuous application to several generations of fruit fly resulted in increased pest control. This field trial was conducted in an isolated valley some 80 ha in size, over a continuous four-year period. In order to maintain the sterilizing effect in the field throughout the whole year, a new lufenuron bait gel was developed. This bait gel was introduced in to delta traps suspended in trees at a density of 24 traps ha-1, and these traps were replaced once a year during the field trial. Monitoring of the adult C. capitata population was conducted to assess the effects of the chemosterilant treatment. In the first year of treatment with sterilizing traps, a reduction of the C. capitata population was observed, indicating that the traps reduce the population right from the first generation. In the second, third and fourth years, a continuous and progressive reduction of the adult Mediterranean fruit fly population was observed. Therefore, the successive application of chemosterilization treatment has a cumulative effect on reducing the fly population year after year. Aerial treatment using malathion does not produce this cumulative effect, and consequently every year it is necessary to start again with the same number of flies as the year before. The possibility of using the chemosterilant method alone or combined with the sterile insect technique is discussed.     

Effective sampling range of a synthetic protein-based attractant for Ceratitis capitata (Diptera: Tephritidae)

Studies were conducted in Honduras to determine effective sampling range of a female-targeted protein-based synthetic attractant for the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae). Multilure traps were baited with ammonium acetate, putrescine, and trimethylamine lures (three-component attractant) and sampled over eight consecutive weeks. Field design consisted of 38 traps (over 0.5 ha) placed in a combination of standard and high-density grids to facilitate geostatistical analysis, and tests were conducted in coffee (Coffea arabica L.),mango (Mangifera indica L.),and orthanique (Citrus sinensis X Citrus reticulata). Effective sampling range, as determined from the range parameter obtained from experimental variograms that fit a spherical model, was approximately 30 m for flies captured in tests in coffee or mango and approximately 40 m for flies captured in orthanique. For comparison, a release-recapture study was conducted in mango using wild (field-collected) mixed sex C. capitata and an array of 20 baited traps spaced 10-50 m from the release point. Contour analysis was used to document spatial distribution of fly recaptures and to estimate effective sampling range, defined by the area that encompassed 90% of the recaptures. With this approach, effective range of the three-component attractant was estimated to be approximately 28 m, similar to results obtained from variogram analysis. Contour maps indicated that wind direction had a strong influence on sampling range, which was approximately 15 m greater upwind compared with downwind from the release point. Geostatistical analysis of field-captured insects in appropriately designed trapping grids may provide a supplement or alternative to release-recapture studies to estimate sampling ranges for semiochemical-based trapping systems.     

Sexual behavior of mutant strains of the medfly Ceratitis capitata (Diptera: Tephritidae)

Males of the mutant strains (blind, vestigal-winged) of the Mediterranean fruit fly (medfly), Ceratits capitata (Wiedmann) showed differences in behavior compared with control (mass-reared) males. Mutant males made fewer mating attempts and achieved fewer matings than control males. Vestigal-winged females copulated less frequently with both mutants. Blind males climbed rather than jumped onto females and copulated in very low numbers compared with control and vestigal males. Blind females copulated normally with control, males and in very low numbers with both types of mutant males.     

Feeding behavior of Ceratitis capitata (Diptera,Tephritidae): influence of carbohydrate ingestion

Experiments were conducted on Ceratitis capitata larvae and adults to determine the influence of sugar (glucose and sucrose) ingestion on some aspects of the feeding behavior of this species. The results indicate that larval choice of a diet containing glucose or sucrose is not influenced by the rearing diet, by carbohydrate deprivation or by previous experience (induction). Carbohydrate deprivation did not alter the discrimination threshold of the larvae for sucrose. In adult females, the discrimination threshold for sucrose was unchanged when the flies were submitted to carbohydrate deprivation during the adult phase although ingestion by deprived females was 35% higher than ingestion by the control group. However, adults submitted to carbohydrate deprivation during the larval phase suffered a profound reduction in discrimination threshold for sucrose.     

Improvement of egg hatch of Ceratitis capitata (Diptera: Tephritidae) for enhanced output

Embryonated eggs of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) genetic sexing strain (GSS), VIENNA 8 were treated with low concentrations of five disinfectants-formaldehyde, iodine, sodium hypochlorite, peracetic acid, and quaternary ammonium--for decontamination and egg hatch improvement. The newly laid eggs were successfully treated with formaldehyde at 100 ppm for 1 min with 74.2% hatching and with quaternary ammonium at 150 ppm for 1 and 2 min with 70.4 and 69% hatching, respectively. Increased formaldehyde concentration may have affected the embryos, because it resulted in a decrease in the hatching percentage. However, egg viability was not impaired and hatch was not affected by quaternary ammonium treatment compared with controls and eggs treated with other disinfectants. Quaternary ammonium shows promise for decontaminating eggs and improving egg hatch.     

Field evaluation of female attractants for monitoring Ceratitis capitata (Diptera: Tephritidae) under a range of climatic conditions and population levels in Western Australia

Field trials were conducted in Western Australia to compare captures of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann), in a standard male-targeted trap (Lynfield trap baited with Capilure) with a synthetic, female-targeted attractant marketed as BioLure. BioLure was also compared with other fenale attractants (orange ammonia, liquid protein bait) and tested in plastic McPhail, Tephri, and Lynfield traps. The possibility of using one trap to monitor female and male C. capitata populations was also tested by combining BioLure in a trap with the male attractant, Capilure. The results of these experiments show that BioLure outperformed the female-targeted system currently used for monitoring female C. capitala (liquid protein in MePhail trap). More male C. capitata were caught in the standard male-targeted trap, but more females were caught in traps baited with BioLure irrespective of trap type, climate, host tree, or population level. Combined lure traps caught equivalent total numbers of C. capitata to the standard male-targeted trap, but fewer females were captured. Tephri traps caught more flies than McPhail traps, but McPhail traps caught equivalent proportions of females. We compared the performance in commercial orchards of the standard male-targeted trap with a female-targeted trap (McPhail with BioLure). We found that the male trap detected C. capitata more often, caught more flies, triggered the economic threshold more often (66% of the time) and was more cost effective. The male-targeted trap is recommended for use on commercial orchards if cost is limiting. However, using both male and female-targeted traps increases the chance of detecting flies and triggering the economic threshold level. The synthetic female attractant is recommended for replacement of protein hydrolysate lures and may be used in either Tephri or McPhail traps.     

Sexual selection on multivariate phenotype in wild and mass-reared Ceratitis capitata (Diptera: Tephritidae)

Males with a larger thorax and narrower face were found to be favoured by directional sexual selection in a previous field cage experiment where Ceratitis capitata males from the Seib 6-96 genetic sexing strain competed with wild males from Alto Valle (Patagonia) for the possession of wild females. Targets of sexual selection, however, might differ between wild and laboratory females as a response to adaptation to mass-rearing conditions. To evaluate possible divergences on the targets of sexual selection as a by-product of adaptation to mass-rearing conditions, field cage tests were performed with both wild and laboratory females. To avoid possible bias due to correlation among the measured traits (eye length [EL], face width [FW], head width [HW], and thorax length [TL]), a multivariate analysis was applied. Consistent with the previous experiment, the results indicated that TL and FW are probable targets of directional sexual selection independently of female strain. However, laboratory females were less selective than wild ones. Additionally, correlational sexual selection was detected acting on the multivariate phenotype. The effects of correlational selection overlap with those of directional selection on each single trait. The analysis of mating pair characteristics showed patterns that do not match the expectations for a random mating system. The current analysis indicates that during mating pair formation two processes overlap. On the one hand, sexual selection favours males with larger size (TL) and narrower faces (FW). This effect occurs in both wild and laboratory females. In addition, assortative mating based on both phenotype and origin was also observed.     

Bacteria Associated with the Oesophageal Bulb of the Medfly Ceratitis capitata (Diptera:Tephritidae)

Extracellular Gram negative bacteria were found to be commonly associated to the oesophageal bulb of Ceratitis capitata with Klebsiella oxytoca and Enterobacter agglomerans as the most common species. All the isolates tested in vitro, except one, were sensitive to the antibacterial material present on the medfly laid egg surface. Received: 3 May 2001 / Accepted: 7 June 2001     

Transgenic sexing system for Ceratitis capitata (Diptera: Tephritidae) based on female-specific embryonic lethality

Fruit fly pest species have been successfully controlled and managed via the Sterile Insect Technique (SIT), a control strategy that uses infertile matings of sterile males to wild females to reduce pest populations. Biological efficiency in the field is higher if only sterile males are released in SIT programs and production costs are also reduced. Sexing strains developed in the Mediterranean fruit fly Ceratitis capitata (medfly) through classical genetics are immensely beneficial to medfly SIT programs but exhibit reduced fertility and fitness. Moreover, transfer of such classical genetic systems to other tephritid species is difficult. Transgenic approaches can overcome this limitation of classical genetic sexing strains (GSSs), but had resulted so far in transgenic sexing strains (TSSs) with dominant lethality at late larval and pupal stages. Here we present a transgene-based female-specific lethality system for early embryonic sexing in medfly. The system utilizes the sex-specifically spliced transformer intron to restrict ectopic mRNA translation of the pro-apoptotic gene hid^Ala5 to females only. The expression of this lethal effector gene is driven by a tetracycline-repressible transactivator gene tTA that is under the control of promoters/enhancers of early-acting cellularization genes. Despite observed position effects on the sex-specific splicing, we could effectively establish this early-acting transgenic sexing system in the medfly C. capitata. After satisfactory performance in large scale tests, TSSs based on this system will offer cost-effective sexing once introduced into SIT programs. Moreover, this approach is straight forward to be developed also for other insect pest and vector species.     

Male diet and age influence to inhibit female remating in Ceratitis capitata (Diptera: Tephritidae)

This study evaluated the influence of age and adult nutritional status of Ceratitis capitata males on their ability to inhibit female remating. Their roles and that of copula duration on the amount of sperm transferred to female spermathecae were also analysed. After emergence, adults were kept in separate groups according to their diets (either high protein – 6.5 g of brewer’s yeast, or low protein – 3.5 g of brewer’s yeast) and their age at the time of use in experiments (4, 8, 12, 16 and 20 days old). The results demonstrated that: (i) male age was not a factor that influenced remating when females mated with well‐nourished males; however, the youngest males (4 days old) in the low‐protein group were less efficient in preventing female remating; (ii) 12‐ and 16‐day‐old males fed on a high‐protein diet transferred and produced more sperm than males of other groups; (iii) there was no correlation between copula duration and the amount of sperm transferred to the female; the longest copula durations were observed among low‐protein‐diet/20‐day‐old males. These results suggest that age and nutrition influence the ability of the males to inhibit female remating. The sterile insect technique is most effective when females do not remate after coupling with sterile males, and therefore, the capacity to inhibit female remating is an important characteristic of males released in the field.     

The ceratotoxin gene family in the medfly Ceratitis capitata and the Natal fruit fly Ceratitis rosa (Diptera: Tephritidae)

Ceratotoxins (Ctxs) are a family of antibacterial sex-specific peptides expressed in the female reproductive accessory glands of the Mediterranean fruit fly Ceratitis capitata. As a first step in the study of molecular evolution of Ctx genes in Ceratitis, partial genomic sequences encoding four distinct Ctx precursors have been determined. In addition, anti-Escherichia coli activity very similar to that of the accessory gland secretion from C. capitata was found in the accessory gland secretion from Ceratitis (Pterandrus) rosa. SDS-PAGE analysis of the female reproductive accessory glands from C. rosa showed a band with a molecular mass (3 kDa) compatible with that of Ctx peptides, also slightly reacting with an anti-Ctx serum. Four nucleotide sequences encoding Ctx-like precursors in C. rosa were determined. Sequence and phylogenetic analyses show that Ctxs from C. rosa fall into different groups as C. capitata Ctxs. Our results suggest that the evolution of the ceratotoxin gene family might be viewed as a combination of duplication events that occurred prior to and following the split between C. capitata and C. rosa. Genomic hybridization demonstrated the presence of multiple Ctx-like sequences in C. rosa, but low-stringency Southern blot analyses failed to recover members of this gene family in other tephritid flies.     

Attractiveness of 79 compounds and mixtures to wild Ceratitis capitata (Diptera: Tephritidae) in field trials

Seventy-nine volatile compounds chosen from those emitted by Mediterranean fruit fly males, fruits and food baits, were tested for their attractiveness to females of Ceratitis capitata (Wiedemann) in field trials with a wild population. To correct for density fluctuations, the results were presented as a percentage of males caught in the same type of traps baited with trimedlure, a known male attractant. The mixtures of corn steep liquor with ammonia compounds and amines were the most effective attractants for females. Other compounds exhibiting C. capitata attraction were a mixture ethyl acetate+acetic acid+ethanol for both sexes, and p-cymene, 1,3-diethylbenzene and borneol for females.