Pathogenicity of Metarhizium anisopliae (Metsch.) Sorokin and Beauveria bassiana (Balsamo) Vuillemin,to three adult fruit fly species: Ceratitis capitata (Weidemann), C. rosa var. fasciventris Karsch and C. cosyra (Walker) (Diptera :Tephritidae)

The pathogenicity of two isolates of Beauveria bassiana and 12 of Metarhizium anisopliae towards adult fruit flies, Ceratitis capitata and Ceratitis rosa var. fasciventris was tested in the laboratory. Fruit flies were exposed to dry conidia evenly spread on velvet material covering the inner side of a cylindrical plastic tube. All isolates tested were pathogenic to both species of fruit flies. Mortality ranged from 7 to 100% in C. capitata and from 11.4 to 100% in C. rosa var. fasciventris at 4 days post-inoculation. Six isolates, M. anisopliae ICIPE 18, 20, 32, 40, 41 and 62, were highly pathogenic to both C. capitata and C. rosa var. fasciventris. The LT90 values of the most pathogenic isolates ranged between 3-4 days in both insects. Because of the difficulties in rearing C. cosyra, only the isolates that were highly pathogenic to both C. rosa var. fasciventris and C. capitata were tested against adult C. cosyra. They caused mortality of between 72-78% at 4 days post-inoculation. The LT90 values in all the isolates did not exceed 4 days. One of the most pathogenic isolates, M. anisopliae ICIPE 20, was evaluated against C. capitata and C. rosa var. fasciventris in cage experiments using three autoinoculators (maize cob, cheesecloth and Petri dish) in an autoinoculative device consisting of plastic mineral bottle. Mortality of between 70-93% was observed in flies of both species that were captured from the cages and held under laboratory conditions. These results indicate the possibility of fruit fly suppression with entomopathogenic fungi using an autoinoculative device.     

Comparative analysis of microsatellite loci in four fruit fly species of the genus Ceratitis (Diptera: Tephritidae)

The possibility to cross-species amplify microsatellites in fruit flies of the genus Ceratitis was tested with the polymerase chain reaction (PCR) by analysing 23 Ceratitis capitata (Wiedemann) microsatellite markers on the genomic DNA of three other economically important, congeneric species: C. rosa (Karsch), C. fasciventris (Bezzi) and C. cosyra (Walker). Twenty-two primer pairs produced amplification products in at least one of the three species tested. The majority of the products were similar, if not identical in size to those expected in C. capitata. The structures of the repeat motifs and their flanking sequences were examined for a total of 79 alleles from the three species. Sequence analysis revealed the same repeat type as the homologous C. capitata microsatellites in the majority of the loci, suggesting their utility for population analysis across the species range. A total of seven loci were differentially present/absent in C. capitata, C. rosa, C. fasciventris and C. cosyra, suggesting that it may be possible to differentiate these four species using a simple sequence repeat-based PCR assay. It is proposed that medfly-based microsatellite markers could be utilized in the identification and tracing of the geographical origins of colonist pest populations of the four tested species and in the assessment of their risk and invasive potentials; thereby assisting regulatory authorities in implementing quarantine restrictions and other pest control measures.     

Developmental rates at constant temperatures of three economically important Ceratitis spp. (Diptera: Tephritidae) from southern Africa

Three species of Ceratitis MacLeay are of economic importance in southern Africa. To learn more about the influence of temperature on the development of these species, the developmental rates of South African populations of Ceratitis (Ceratitis) capitata (Wiedemann), C. (Pterandrus) rosa Karsch, and C. (Ceratalaspis) cosyra (Walker) were compared at constant temperatures of 14, 18, 22, 26, and 30 degrees C. The duration of each life stage and the percentage survival of the immature life stages of each species were determined. One linear and three nonlinear developmental rate models (Briére, Lactin, and Logan-6) were found to fit the data well and were used to generate the minimum, optimum, and maximum developmental thresholds, in addition to the life cycle thermal constants for the three species. These parameter values were 9.6, 28.5, 33.0, and 338 for C. capitata, 9.7, 28.8, 33.2, and 376 for C. cosyra, and 8.6, 27.7, 33.0, and 429 for C. rosa, respectively. The parameters for C. capitata are similar to those found by other researchers for this species in Reunion but the parameters for C. rosa differ substantially from published values for a Reunion population of this species, suggesting that these are different biotypes. The similarities between the developmental parameters for C. capitata and C. cosyra do not support known differences in the distribution of these species so other limiting factors such as relative humidity and the availability of host species may be important. This finding therefore cautions against basing predictions of potential global distributions of species solely on life table or climatic parameter values.     

Mortality in Three African Tephritid Fruit Fly Puparia and Adults Caused by the Entomopathogenic Fungi, Metarhizium anisopliae and Beauveria bassiana

The pathogenicity of 13 isolates of Metarhizium anisopliae and two isolates of Beauveria bassiana to Ceratitis capitata and Ceratitis var. rosa fasciventris exposed as late third instar larvae in sand was evaluated in the laboratory. All isolates caused a significant reduction in adult emergence and a corresponding large mortality on puparia of both species. All isolates also induced large deferred mortality in emerging adults following treatment as late third instar larvae. On C. capitata , seven isolates ( M. anisopliae ICIPE 18, 20, 32, 60 and 69 and B. bassiana ICIPE 44 and 82) caused significantly higher mortality on puparia than other isolates. With the exception of ICIPE 32, the other four isolates of M. anisopliae above were the most pathogenic against C. r. fasciventris . Dose-response study carried out with these isolates of M. anisopliae on the two species of flies above plus another species, Ceratitis cosyra showed that the dose-mortality regression lines of ICIPE 18 and 20 were steeper with lower LC 50 values when compared with ICIPE 60 and 69 on the three species. When these two isolates were evaluated with regard to their pathogenicity to different pupal age, adult emergence was found to increase with increasing pupal age with a corresponding decrease in mortality in puparia and emerging adults in the three species of fruit flies. M. anisopliae ICIPE 18 and 20 were equally pathogenic to all pupal ages tested in C. capitata and C. cosyra but ICIPE 18 was more pathogenic to older puparia of C. r. fasciventris than ICIPE 20. Our results suggest that soil inoculation with M. anisopliae under mango trees might form an important component of integrated pest management strategies in areas where these three species of fruit fly coexist.     

Genetic differentiation in Spanish populations of Ceratitis capitata as revealed by abundant soluble protein analysis

Ceratitis capitata is one of the most important pest species in the tropical and temperate regions of the world, however, genetic knowledge of this species is still very limited. In the present study, we have attempted for the first time an analysis of the genetic variability in seven natural populations of C. capitata by means of abundant soluble proteins, combining high resolution techniques such as polyacrylamide gel electrophoresis and silver staining methods. A total of 66 polypeptides have been analyzed. The results show the existence of a decreasing trend in the ‘levels’ of polymorphism from the southern areas to the northern that is correlated with the latitude of the sampling areas, most probably due to the colonization process of the Iberian Peninsula. On the other hand, and with relation to the ‘pattern’ of the variability, a geographic differentiation (not strictly latitudinal) is detected when an UPGMA clustering method was applied to the data. Moreover, principal component anaysis has revealed that a part of this differentiation could be explained in relation with environmental factors such as annual rainfall and minimum temperature in winter months. This is suggesting that selection (added to the historical process) could be playing an important role in the process of geographic differentiation, ‘shaping’ the pattern of that variability. No host-related differentiation has been observed in these samples. This revised version was published online in July 2006 with corrections to the Cover Date.     

Comparison of the mitotic karyotypes of Ceratitis capitata, Ceratitis rosa, and Trirhithrum coffeae (Diptera: Tephritidae) by C-banding and FISH.

The sex chromosomes of the tephritid fruit fly Ceratitis capitata (Wiedemann) are heteromorphic. The male-determining region was located on the Y chromosome by deletion mapping using unbalanced offspring from several translocation strains. In addition, we showed that only 15% of the Y chromosome is required for male determination and male fertility. Based on this result, we expected to find Y-chromosomal length polymorphism in natural populations. Using fluorescence in situ hybridization with two repetitive DNA probes that label the Y chromosome, no obvious size differences were detected in seven wild-type strains and three mutant strains. As the medfly is probably of East African origin, we also analyzed two wild-type strains established recently from pupae sampled in Kenya. The Y chromosomes show a polymorphism in the hybridization pattern of a repetitive Y-specific medfly clone. However, the overall size of the Y chromosome is similar to that of the other strains. Besides C. capitata, the tephritid fruit flies Ceratitis (Pterandrus) rosa Karsch and Trirhithrum coffeae Bezzi also emerged from pupae sampled in Kenya. Their karyotype was analyzed by C-banding. Furthermore, the ribosomal genes were mapped to the sex chromosomes in these two species. Key words : Ceratitis capitata, Tephritidae, C-Banding, FISH, rDNA.     

Molecular evaluation of nominal species in the Ceratitis fasciventris, C. anonae, C. rosa complex (Diptera: Tephritidae)

Ceratitis fasciventris, C. anonae and C. rosa form a complex of economically important fruit fly pests infesting a variety of crops in African countries. Hitherto only adult males of these species can be distinguished easily by morphological characters. Other stages cannot, and for some taxa the taxonomic interpretation and species boundaries remain unclear. In order to clarify phylogenetic relationships and taxonomic status of these species, sequences of mitochondrial (16S, COI, ND6) and nuclear markers (period, ITS1) were analysed in specimens of the three morphospecies throughout the distribution of the complex. Maximum likelihood trees did not recover monophyletic groups corresponding to the morphospecies. Conversely, ND6 and COI divided West African C. fasciventris specimens in two consistent and bootstrap supported clades, involving specimens from Benin and from Mali/Ivory Coast, while the nuclear gene fragments per and ITS1 recovered a well-supported clade corresponding to C. fasciventris from Kenya/Uganda. Hence, the phylogenetic relationships and taxonomic interpretation of the complex appear more intricate than previously hypothesised. The current molecular data do not allow to identify C. fasciventris, C. anonae and C. rosa as distinct phylogenetic species but rather suggest that the morphospecies C. fasciventris is itself a complex of cryptic taxa.     

Ecological niches and potential geographical distributions of Mediterranean fruit fly (Ceratitis capitata) and Natal fruit fly (Ceratitis rosa)

Aim To predict and compare potential geographical distributions of the Mediterranean fruit fly (Ceratitis capitata) and Natal fruit fly (Ceratitis rosa). Location Africa, southern Europe, and worldwide. Methods Two correlative ecological niche modelling techniques, genetic algorithm for rule‐set prediction (GARP) and a technique based on principal components analysis (PCA), were used to predict distributions of the two fly species using distribution records and a set of environmental predictor variables. Results The two species appear to have broadly similar potential ranges in Africa and southern Europe, with much of sub‐Saharan Africa and Madagascar predicted as highly suitable. The drier regions of Africa (central and western regions of southern Africa and Sahelian zone) were identified as being less suitable for C. rosa than for C. capitata. Overall, the proportion of the region predicted to be highly suitable is larger for C. capitata than for C. rosa under both techniques, suggesting that C. capitata may be tolerant of a wider range of climatic conditions than C. rosa. Worldwide, tropical and subtropical regions are highlighted as highly suitable for both species. Differences in overlap of predictions from the two models for these species were observed. An evaluation using independent records from the adventive range for C. capitata and comparison with other predictions suggest that GARP models offer more accurate predictions than PCA models. Main conclusions This study suggests that these species have broadly similar potential distributions worldwide (based on climate), although the potential distribution appears to be broader for C. capitata than for C. rosa. Ceratitis capitata has become invasive throughout the world, whereas C. rosa has not, despite both species having broadly similar potential distributions. Further research into the biology of these species and their ability to overcome barriers is necessary to explain this difference, and to better understand invasion risk.     

Genetic and molecular investigations on the endogenous mobile elements of non-drosophilid fruitflies

A syndrome of abnormal genetic effects, resembling Drosophila hybrid dysgenesis, occurs in Ceratitis capitata when strains of different origin are mated. The pattern of abnormal traits observed appears to be the phenotypic expression of a complex interacting dysgenic system of inducer and suppressor effects; probably more than one system is activated in the crosses. This suggests that different systems of mobile elements occur in different strains and populations of C. capitata. Using a PCR primer specific to the ITR sequence of a deleted element, full length mariner elements were isolated from C. capitata, Ceratitis rosa, and Trirhithrum coffeae. Very high similarities were found in inter- and intraspecific comparisons of the elements. The majority of these elements contained deletions and frame-shifts. However, one clone Ccmar1.18, from C. capitata, was found to possess an uninterrupted ORF coding for 338 amino acids with ∼60% similarity to the Mos1 element of Drosophila mauritiana. Database searches and phylogenetic analyses showed that the mariner elements isolated in the present study are representatives of Robertson's mellifera mariner subfamily. The copy numbers of the elements within each species are very different, ranging from about 10 in T. coffeae to 5000 in C. rosa. This revised version was published online in August 2006 with corrections to the Cover Date.     

Patterns of variation within and between Greek populations of Ceratitis capitata suggest extensive gene flow and latitudinal clines

Four natural Greek populations of Mediterranean fruit fly, Ceratitis capitata (Wiedemann), was studied for genetic variability at 25 enzyme loci. The comparison of polymorphism within and between populations shows two loci with high between-population heterozygosity (HT) and very high fixation index (F(ST)) values, suggesting the presence of balancing selection. The gradual decline of common allele frequency of the polymorphic loci tested indicated that latitudinal clines are present in Greece. Indirect estimates of gene flow based both on Wright's method (Nm*) and on the Slatkin's method (Nm*), which depends on the frequencies of rare alleles found in only one population, revealed a substantial amount of gene flow (Nm = 3.493 and Nm* = 3.197). These estimates of gene flow may well explain why the "introduced" Greek populations of C. capitata, in spite of their low genetic variability, display the same polymorphic loci. Gene flow in combination with natural selection and genetic drift may have played an important role to genetic differentiation in this species in Greece.     

Systematic revision of the subgenus Ceratitis MacLeay s.s. (Diptera, Tephritidae)

The fruit fly genus Ceratitis MacLeay (Diptera, Tephritidae) comprises several important pest species attacking a wide range of unrelated fruits. This is the third paper in a series revising the genus, and deals with representatives of the subgenus Ceratitis s.s . Eight species are recognized: Ceratitis brachychaeta Freidberg, C. caetrata Munro, C. capitata (Wiedemann), C. catoirii Guerin-Meneville, C. comuta (Bezzi), C. malgassa Munro, C. manjakatompo Hancock and C. pinax Munro. Lectotypes are designated for C. catoirii and C. pinax . A key for both sexes is provided, with illustrations of cephalic bristles, mesonotal and wing patterns, and shape of aculeus. All species are Afrotropical, except for 67. capitata , which has become widespread. The phylogeny and biogeography of Ceratitis s.s . are discussed, based on a cladistic analysis, and showing indications that the mediterranean fruit fly, C. capitata , may have its origins in eastern Africa. The relationship of this subgenus within the genus Ceratitis s.l . is also outlined.     

The isolation of microsatellite loci in the Mediterranean fruitfly Ceratitis capitata (Diptera: Tephritidae) using a biotin/streptavidin enrichment technique

The Medfly (Ceratitis capitata) is a polyphagous dipteran pest which has spread from North Africa to the countries of the Mediterranean Basin and has also invaded tropical and subtropical regions throughout the world. Colonizing populations typically possess low levels of genetic variability. Microsatellites provide an effective means of investigating the population structure of such genetically depauperate populations, however, microsatellite markers traditionally require a long phase of development in new taxa. We used a biotin/streptavidin capture technique to isolate microsatellites directly from C. capitata genomic DNA and we describe here the identification of seven polymorphic microsatellite markers in C. capitata.     

Intron size polymorphism of the Adh1 gene parallels the worldwide colonization history of the Mediterranean fruit fly, Ceratitis capitata

The exon-primed intron-crossing (EPIC) PCR technique was used to analyse the size variation at the first intron of the Ceratitis capitata Adh ₁ gene. A total of 27 samples from 16 natural populations was analysed from five geographical regions in the species range: Africa, Mediterranean Basin, Latin America, Hawaii and Australia. The Adh ₁ first intron varies extensively in length with at least 18 size variants ranging from 1400 bp to 3450 bp. These variants can be grouped into four distinct size categories: short, medium, long and very long. The majority of these variants are present only in the African populations. Only a subset of the ancestral variants appear to have succeeded in migrating from Africa during the medfly colonization process. The medfly population structure inferred from the intron size polymorphism is congruent with that observed from the analysis of allozyme variation. The geographical dispersal of the medfly from its source area is associated with a gradual and great reduction in intron variability which parallels the trend of decreasing variability evaluated at 26 biochemical loci. The intron phylogenetic tree is in agreement with allozyme data in portraying the dynamic population history of the medfly. Stochastic evolutionary forces such as drift, bottleneck effects and migration seem to have played the major roles in the dispersion pattern of Adh ₁ intron variation during the colonization of the medfly.     

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.     

Survival and development of different life stages of three Ceratitis spp. (Diptera: Tephritidae) reared at five constant temperatures

Fruit flies (Diptera: Tephritidae) are the most damaging pests on fruit crops on Réunion Island, near Madagascar. Survival and development of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann), the Natal fruit fly, C. rosa Karsch and the Mascarenes fruit fly, C. catoirii Guérin-Mèneville were compared at five constant temperatures spanning 15 to 35 degrees C. Durations of the immature stages of C. capitata, C. rosa and C. catoirii ranged from 14.5-63.8, 18.8-65.7 and 16.8-65.8 days, respectively, at 30-15 degrees C. The lower developmental threshold and thermal constant were calculated using the temperature summation model. The thermal constant for total development of the immature stages of C. capitata, C. rosa and C. catoirii were 260, 405 and 356 DD, respectively. Species differed mainly during the larval stages and ovarian maturation period, with smaller differences in the egg stage. Ceratitis rosa appeared to be better adapted to low temperatures than the two other species as it showed a lower larval developmental threshold of 3.1 degrees C compared to 10.2 degrees C for C. capitata and 8.9 degrees C for C. catoirii. Overall, C. catoirii had a low survival rate within the range of temperatures studied. The different responses of the three Ceratitis species to various temperatures explain to some extent their distribution on the island. The results obtained will be used for optimizing laboratory rearing procedures and for constructing computer simulation models to predict fruit fly population dynamics.     

Estimates of gene flow from rare alleles in natural populations of medfly Ceratitis capitata (Diptera: Tephritidae)

Gene flow based on the spatial distribution of rare alleles at 25 gene loci was estimated in 15 populations of Ceratitis capitata (Wiedemann) from different parts of the world. Estimates of Nm, the number of migrants exchanged per generation among populations in different regions of the world, appeared to be quite similar, ranging from 3.36 in tropical Africa to 2.94 in the New World and 2.72 in Mediterranean basin populations. This suggests that gene flow among neighbouring populations of medfly is quite extensive. The genetic differentiation in American, Mediterranean and African populations was related to major climatic differences between North and South. These differences arise mainly from five loci that showed gene frequency patterns suggestive of latitudinal clines in allele frequencies. The clinal variation was such that tropical-subtropical populations were more heterozygous than temperate populations. It was concluded that gene flow, counteracting the forces of natural selection and genetic drift, determines the extent to which geographical populations of C. capitata are differentiated.     

Contribution of natural food sources to reproductive behaviour, fecundity and longevity of Ceratitis cosyra, C. fasciventris and C. capitata (Diptera: Tephritidae)

The influence of food sources comprising the natural diet on the reproductive behaviour, fecundity and longevity of three African fruit flies Ceratitis cosyra (Walker), C. fasciventris (Bezzi) and C. capitata (Wiedemann) was investigated. Three natural food sources, varying in protein and sugar content, were evaluated. These included bird droppings (farm chicken), aphid honeydew and guava (Psidium guajava L.) juice. For C. fasciventris and C. capitata, flies fed on a protein-rich diet displayed higher frequency of calling, mating and oviposition than flies fed on a protein-poor diet, whilst for C. cosyra, quality of diet significantly influenced the mating behaviour of the flies, but not the calling and oviposition behaviour. Net fecundity rates were lowest for C. fasciventris and C. capitata when fed only on guava juice (0.1, 2.6 eggs per female, respectively), and higher for those on a diet of honeydew only (9.5, 33.8 eggs per female, respectively) and a combined diet of guava, honeydew and chicken faeces (11.8, 25.8 eggs per female, respectively). For C. cosyra, due to low numbers of eggs collected, no significant differences in fecundity between diets could be detected. All species fed only on a diet of chicken faeces since emergence died within the first three days of adult life without laying eggs, but when carbohydrates were provided by addition of guava juice and honeydew, the longevity of the flies was sustained for more than four weeks after adult emergence. The practical implications of these findings for control purposes are discussed.     

Cryptic diversity and gene flow among three African agricultural pests: Ceratitis rosa,Ceratitis fasciventris and Ceratitis anonae (Diptera,Tephritidae)

The ‘Ceratitis FAR complex’ is a species complex of African fruit flies (Diptera, Tephritidae) including the major agricultural pest Ceratitis rosa and the morphologically similar Ceratitis fasciventris and Ceratitis anonae. To resolve their intra‐ and interspecific genetic relationships and to estimate gene flow within this complex, we surveyed allelic variation at 16 microsatellite loci in 27 African populations of the three morphospecies. Interpopulation genetic distances and individual Bayesian assignments distinguished five genotypic clusters: two involving C. rosa (R1, R2; that may occur in sympatry), two involving C. fasciventris (F1, F2; with parapatric distributions) and one involving C. anonae (A). Intra‐ and interspecific patterns of genetic differentiation were not hierarchically structured and genetic differentiation between conspecific clusters (F1–F2 and R1–R2) was higher or comparable with differentiation between heterospecific clusters (e.g. F1‐A or R2‐A). In some cases, gene flow estimates among morphospecies or among heterospecific genotypic clusters were significantly different from zero, showing the lack of reproductive isolation. Genetic differentiation between genotypic clusters was partly supported by morphological differences observed a posteriori in male secondary sexual characters. These results suggest important revisions to current models of ecological niche requirements and invasion risk of the major agricultural pest C. rosa and provide a basis for a taxonomic re‐interpretation of the FAR complex.     

An integrated genetic and cytogenetic map for the Mediterranean fruit fly, <Emphasis Type="Italic">Ceratitis capitata</Emphasis>, based on microsatellite and morphological markers

A genetic map based on microsatellite polymorphisms and visible mutations of the Mediterranean fruit fly (medfly), Ceratitis capitata is presented. Genotyping was performed on single flies from several backcross families. The map is composed of 67 microsatellites and 16 visible markers distributed over four linkage groups. Fluorescence in situ hybridization of selected microsatellite markers on salivary gland polytene chromosomes allowed the alignment of these groups to the second, fourth, fifth and sixth chromosome. None of the markers tested showed segregation either with the X or the third chromosome. However, this map constitutes a substantial starting point for a detailed genetic map of C. capitata. The construction of an integrated map covering the whole genome should greatly facilitate genetic studies and future genome sequence projects of the species.     

Effect of soil temperature and moisture on survival and infectivity of Metarhizium anisopliae to four tephritid fruit fly puparia

The infectivity of 4 isolates of Metarhizium anisopliae to puparia of Ceratitis capitata treated as late third-instar larvae in unsterilized soil was investigated in the laboratory under controlled temperature and moisture. At 20-30 degrees C, mortality in puparia was highest at water potential of -0.1 and -0.01 mega Pascal (MPa) and lowest at water potential of -0.0055 and -0.0035 MPa in all the isolates. In wetter soil however, isolates ICIPE 20 and 60 caused significantly higher mortality than ICIPE 18 and 69. The survival of conidia in drier soil (-0.1 MPa) was not adversely affected at all temperatures. However, in wet soil (-0.0035 MPa) there was drastic reduction in colony counts in ICIPE 18 and 69 at 25 and 30 degrees C but conidial density in ICIPE 20 and 60 remained at the initial level at 14 days after inoculation at all temperatures. When ICIPE 20 was evaluated against three other fruit fly species (Ceratitis cosyra, Ceratitis rosa, and Ceratitis fasciventris), significant reduction in adult emergence and higher pupal mortality occurred in C. cosyra and C. fasciventris than in C. rosa at a combination of 15 and 20 degrees C and -0.1 and -0.0035 MPa. However, at higher temperature and the same moisture level, the isolates were equally pathogenic across the 3 species. It is probable that in addition to pathogen cycling and multiplication from dead infected insects in the soil, a balance between microbial degradation and replenishment of inoculum of virulent isolates occur through fluctuations in, and intricate interactions between temperature and moisture levels. This study is indicative of the potential of using isolate ICIPE 20 for soil inoculation against pupariating third-instar larva of fruit flies, thus providing a novel alternative to chemical soil application.