Real-time monitoring of oriental fruit moth,Grapholita molesta,populations using a remote sensing pheromone trap in apple orchards

A fusion of information technology (IT) and sex pheromone monitoring provides a remote sensing IT-pheromone trap to monitor Oriental fruit moth, Grapholita molesta, populations in apple orchards. Once a male of G. molesta is attracted to its sex pheromone lure in the trap, an infrared sensor installed at the funnel-shaped orifice generates an electric signal. The signal is processed in a central processor and then transferred to an internet site via a code division multiple access protocol. The signal also contains information about when each male is caught. Daily trapping information from different localities is archived in a website. The accuracy of IT-pheromone traps in detecting male catches was shown by a high correlation (r = 0.956) between the generated IT signals and actual numbers of males caught in the trap in apple orchards. Using this IT-pheromone trap, G. molesta in apple orchards was monitored for one year. These data were compared with monitoring data obtained from a conventional wing type-based sticky trap containing the identical sex pheromone lure. Both showed four characteristic adult peaks from April to September and were significantly correlated (r = 0.695). IT-pheromone traps also gave real-time signals of male catches in the field. These real-time signals of male catches showed a characteristic diel attraction rhythm from 4 pm to midnight. The diel rhythm of the male response to the sex pheromone started earlier in the evening in the spring season compared to mid and late seasons. This study provides a novel sex pheromone trap for G. molesta to monitor its population in field conditions in real-time without visiting or counting. The field monitoring data can be accessed any time through a designated internet website.     

Electrophysiological and behavioral response of the oriental fruit moth <Emphasis Type="Italic">Grapholita</Emphasis><Emphasis Type="Italic">molesta</Emphasis> (Busck, 1916) (Lepidoptera: Tortricidae) to the volatiles of apple fruits at different stages of development

The oriental fruit moth, Grapholita molesta, occurs in Southern Brazil throughout the year, and migrates from peach to apple orchards. Because moths rely on volatile organic compounds (VOCs) during the host-location process, variations in the emission of these compounds during fruit maturation can influence the time of infestation and preference of the moths for a particular genotype. The aim of this work was to identify VOCs emitted by the apples “Eva” and “Gala” at different stages of development and to determine the behavioral and electrophysiological responses of G. molesta to these compounds. For this purpose, VOCs from immature, maturing, and mature fruits of both cultivars were collected and analyzed by gas chromatography and gas chromatography-mass spectroscopy. The response of the antennae of virgin males and females and mated females to volatiles released by the three fruit stages was registered by gas chromatography coupled to an electroantennography. A dual-choice behavioral test for the different combinations of insect groups and fruit stage was also performed. Amongst the volatiles released by mature fruits, twelve compounds elicited a response. The antennae of the oriental fruit moth did respond to isoamyl hexanoate and α-farnesene emitted by “Eva” maturing fruits. In general, virgin females did not respond to volatiles in olfactometer bioassays and mated females were attracted to volatiles released by mature fruits. Our results show that the variation in the emission during the maturation of fruits can influence the orientation of G. molesta.     

不同栽培管理梨园梨小食心虫发生程度研究

梨小食心虫Grapholitha molesta(Busck)是梨园中的一种重要害虫。本论文采用性诱剂诱集法研究了不同栽培管理条件下梨园梨小食心虫发生的情况。结果表明,在5种不同种植模式的果园中,梨小食心虫在单植桃园中发生最重,试验期间梨小食心虫的日平均诱蛾量为10.9头/盆,与其他4种栽植模式果园的诱捕量均呈显著差异,且混有桃树的果园中梨小食心虫的发生数量多,而单植梨园、梨苹果混栽园、单植苹果园的梨小食心虫发生相对较轻。果实套袋的管理方法也可以显著降低梨小食心虫的发生数量,非套袋梨园的日平均诱捕量为13.8头/盆,是套袋梨园的1.52倍。试验还表明,不同品种和不同树龄的梨树对梨小食心虫的抗虫性均存在显著差异。酥梨比巴梨的抗虫性差,试验期间酥梨园的日平均诱捕量为12.6头/盆,是巴梨园的2.21倍,而40年老酥梨园日平均诱蛾量为12.5头/盆,是20年酥梨园诱蛾量的2.5倍。     

Olfactory responses of Grapholita molesta to different developmental stages of immature fruit of peach,plum, and apple

The Oriental fruit moth, Grapholita molesta (Busck) (Lepidoptera: Tortricidae), is known to shift its host use from stone fruits (the primary hosts) to pome fruits (secondary hosts) as the season progresses. Grapholita molesta recognizes potential hosts by semiochemicals from fruits, but few studies have addressed the olfactory responses of G. molesta to host fruits during the seasonal transition of the moth from stone to pome fruits. So, we investigated the olfactory responses of larvae and gravid females of G. molesta to different fruit stages (collected on May 31, June 11, and June 25) of immature peach, plum, and apple fruits, using both multiple and two choice tests. In the multiple choice tests, for fruits collected after June 11, larvae showed a significant preference for peach as their first choice, compared to plum or apple. The final choice rate (when larvae had stayed on the fruit for more than 1 h) of larvae was also highest for peach fruits collected on June 11 and June 25. In two choice tests (Y-tube test), the response rate and overall choice rate of larvae and mated ovipositing adult females were also highest on peaches collected on May 31 (larvae) and June 25 (adults), respectively. In conclusion, we found that both the adults and larvae of G. molesta were highly attracted to immature peach fruits in laboratory choice tests.     

Mating disruption barriers in pome fruit for improved control of oriental fruit moth Grapholita molesta Busck (Lep., Tortricidae) in stone fruit under mating disruption

Oriental fruit moth (OFM) Grapholita molesta Busck (Lep., Tortricidae) is a very important pest in commercial orchards in Victoria, Australia. Pheromone‐mediated mating disruption (MD) applied in stone fruit orchards successfully controlled OFM populations for many years, but damage to shoot tips and fruit at the edge of peach blocks located adjacent to pear blocks under insecticide treatments has become problematic. To improve protection of stone fruit against edge damage and outbreaks of OFM, all orchards were treated with sex pheromone dispensers for MD on an area‐wide basis. Area‐wide MD treatment, including all pome and stone fruit orchards in a discrete area, successfully controlled edge infestations of OFM, but was expensive. To reduce the cost of OFM control, sex pheromone dispensers for MD were applied as barrier treatments to 54–60 m of neighbouring pears adjacent to peaches under MD. Detailed monitoring of the OFM population, shoot tip and fruit damage assessments indicated that application of MD barriers on pears during two consecutive seasons provided sufficient control of OFM on peaches. This MD barrier treatment was able to reduce the number of OFM caught in all experimental peach blocks, with damage to shoot tips and fruit giving similar results to MD treatment of the whole neighbouring pear block. Extending the MD treatment area for 54–60 m into the neighbouring pear block significantly reduced the edge damage in MD‐treated peaches in the first season and almost eliminated OFM damage in the second season.     

High Genetic Diversity and Structured Populations of the Oriental Fruit Moth in Its Range of Origin

The oriental fruit moth Grapholita ( = Cydia) molesta is a key fruit pest globally. Despite its economic importance, little is known about its population genetics in its putative native range that includes China. We used five polymorphic microsatellite loci and two mitochondrial gene sequences to characterize the population genetic diversity and genetic structure of G. molesta from nine sublocations in three regions of a major fruit growing area of China. Larval samples were collected throughout the season from peach, and in late season, after host switch by the moth to pome fruit, also from apple and pear. We found high numbers of microsatellite alleles and mitochondrial DNA haplotypes in all regions, together with a high number of private alleles and of haplotypes at all sublocations, providing strong evidence that the sampled area belongs to the origin of this species. Samples collected from peach at all sublocations were geographically structured, and a significant albeit weak pattern of isolation-by-distance was found among populations, likely reflecting the low flight capacity of this moth. Interestingly, populations sampled from apple and pear in the late season showed a structure differing from that of populations sampled from peach throughout the season, indicating a selective host switch of a certain part of the population only. The recently detected various olfactory genotypes in G. molesta may underly this selective host switch. These genetic data yield, for the first time, an understanding of population dynamics of G. molesta in its native range, and of a selective host switch from peach to pome fruit, which may have a broad applicability to other global fruit production areas for designing suitable pest management strategies.     

Adjusting the phenology model of codling moth (lepidoptera: tortricidae) in Washington State apple orchards

Studies were conducted with codling moth, Cydia pomonella L., to fit cumulative curves for the occurrence of injured fruits and male moth catches in sex pheromone-baited traps as a function of accumulated degree-days after the start of moth flight. Twelve data sets were collected from seven apple, Malus domestica Bordhausen, orchards in Washington State from 2003 to 2006. Cumulative data were grouped across years for orchards either treated with sex pheromone dispensers or untreated and fit to logistic equations for both the first and second generation. No significant differences were found for the cumulative curves of moth flight or egg hatch between pheromone-treated and untreated orchards; thus, these data were combined. These new logistic models for moth flight and egg hatch were compared with a widely used distributed-delay model originally developed in Michigan. The cumulative flight curves for the logistic and distributed-delay models were statistically different (slopes) for the first but not the second generation. Cumulative egg hatch in the logistic model was significantly different from the distributed-delay model (intercepts and slopes) for both generations. Most strikingly, the timing of 50% egg hatch during the first generation was delayed 100 DD in the logistic model. The potential impact of this change in the characterization of codling moth's phenology on the effectiveness of insecticide programs targeting eggs and newly eclosed larvae was examined. Possible explanations for this significant difference between the models are discussed.     

Larval performance of the oriental fruit moth across fruits from primary and secondary hosts

A common characteristic of many invasive herbivorous insects is their ability to utilize a broad range of host plants. By using various hosts in phenological succession, multivoltine herbivores may increase the number of successful annual generations, at the same time as potentially increasing their overall fitness. To achieve such success, herbivores must be able to develop efficiently on the nutritional resources offered by their hosts. The oriental fruit moth Cydia (= Grapholita) molesta (Busck) (Lepidoptera: Tortricidae) is one of the most damaging invasive insect species. Peach (Prunus persica) is its primary host, whereas the pome fruits apple (Malus × domestica Borkh) and pear (Pyrus communis) are considered as secondary hosts. In many parts of its geographical range, including southern Europe, populations of the moth switch from peach to apple or pear orchards during the growing season. The present study tests whether this temporal switch is supported by the physiological capability of the larvae with respect to developing efficiently on fruits of these taxonomically‐related host plants. Larvae are reared on peach, apple or pear fruits; several life‐history traits are measured; and correlations between the traits are calculated. The results obtained show that larvae do not have the same physiological capability with respect to using apple or pear fruits as hosts compared with using peach fruit. Pear fruit in particular is a sub‐optimal diet. These findings suggest that, in the case of continuous geographical expansion, concomitantly with global warming, apple orchards might support oriental fruit moth populations better than pear orchards, and that the switch onto novel hosts might be accompanied by restricted population growth.     

Simultaneous mating disruption of two Grapholita species in apple orchards

Two Grapholita species (G. molesta and G. dimorpha) cause serious losses in apple production by means of direct fruit damage and destruction. These two congeners occurred at similar seasons in the same orchards. Furthermore, the two species occurred in similar population sizes in apple orchards, though G. molesta occurred much more in total occurrence density than did G. dimorpha in a pear orchard. The two species share two major sex pheromone components, (Z)-8-dodecenyl acetate (Z8-12:Ac) and (E)-8-dodecenyl acetate (E8-12:Ac), but differ with regard to the relative mixture ratio necessary to create conditions of maximal attraction. To develop a simultaneous mating disruption (SMD), a mass ratio of 95:5 of Z8-12:Ac and E8-12:Ac was selected due to its high cross-calling activity and subsequent high mating disruption against both species. When the mixture was applied to orchards, it resulted in significant mating disruption against both species. However, mating disruption efficacy was slightly higher in G. dimorpha than in G. molesta. Application dose and dispenser type affected efficacy duration of SMD. In addition, the high density occurrence of G. molesta significantly reduced the SMD efficacy. Using a wax-type of dispenser, an application of SMD in 100?g per hectare dispersed in early spring recorded >90% mating disruption efficacy through the entire growing season against both Grapholita species.     

Cross-amplification of microsatellites from the codling moth Cydia pomonella to three other species of the tribe Grapholitini (Lepidoptera: Tortricidae)

This study examined cross-species amplification of 33 microsatellite markers, previously developed for Cydia pomonella, in three related fruit moth species of the same tribe (Grapholitini), namely Grapholita molesta, Grapholita funebrana and Grapholita lobarzewskii. Eight microsatellite loci yielded polymorphic products for G. molesta, nine for G. funebrana and 11 for G. lobarzewskii. At all these loci, the number of alleles ranged between four and 11 in G. molesta, and between four and nine in G. funebrana and G. lobarzewskii each. The successful cross-amplified loci can be used for research on population genetics and gene flow of the three target species.     

Monitoring oriental fruit moth (Lepidoptera: Tortricidae) with the Ajar bait trap in orchards under mating disruption

Studies in Oregon, California, Pennsylvania and Italy evaluated the relative performance of the Ajar trap compared with several other traps for the capture of Grapholita molesta (Busck), in pome and stone fruit orchards treated with sex pheromone dispensers for mating disruption. The Ajar is a delta‐shaped trap with a screened jar filled with an aqueous terpinyl acetate plus brown sugar bait solution (TAS) that opens inside the trap and is surrounded by a sticky liner. The TAS‐baited Ajar trap was evaluated with and without the addition of a sex pheromone lure and compared with a delta trap baited with a sex pheromone lure and a bucket trap filled with the TAS bait. Although the Ajar trap had a 90% lower evaporation of the TAS bait than the bucket trap, both of them caught similar numbers in the majority of the field tests of both sexes of G. molesta. The addition of the sex pheromone lure did not increase moth catches by the TAS‐baited Ajar trap. The TAS‐baited Ajar trap caught significantly greater numbers of moths than the sex pheromone‐baited delta trap in 18 of the 20 orchards. Few hymenopterans were caught in orange TAS‐baited Ajar traps, but the catch of flies and other moths relative to the target pest remained high. Flight tunnel and field tests evaluated the effect of several screen designs on the catches of G. molesta and non‐target species. All exclusion devices significantly reduced the catch of larger moths. However, designs that did not reduce the catch of male G. molesta did not reduce the catch of muscid flies. Exclusion devices with openings <7.0 mm significantly reduced the catch of female G. molesta. The addition of (E)‐β‐farnesene, (E)‐β‐ocimene or butyl hexanoate septa lures to TAS‐baited Ajar traps significantly increased total moth catch. The addition of (E)‐β‐ocimene also significantly increased female moth catch.     

Comparative analysis of peach and pear fruit volatiles attractive to the oriental fruit moth,Cydia molesta

Cydia molesta is an important pest of peach and pear fruits late in growing season. We identified and quantified volatiles from immature and mature fruits of peach and pear using gas chromatography-mass spectrometry. Antennally, active compounds were identified by gas chromatography-electroantennogram detection and were further tested in the field. Consistent electroantennographic activity was obtained for nine compounds from headspace collections of the mature fruits of two peach and two pear varieties. Esters were the major components, but qualitative and quantitative differences were found among four odor profiles. Blends mimicking pear fruit volatiles were more attractive to both sexes than blends mimicking peach fruit volatiles in both orchards. Our finding indicated that mixtures mimicking peach and pear fruit volatiles attracted both females and males of C. molesta, and these host volatiles may also be involved in the host switch behavior from peach to pear in mid-late season.     

Monitoring oriental fruit moth and codling moth (Lepidoptera: Tortricidae) with combinations of pheromones and kairomones

Experiments were conducted in North and South America during 2012–2013 to evaluate the use of lure combinations of sex pheromones (PH), host plant volatiles (HPVs) and food baits in traps to capture the oriental fruit moth, Grapholita molesta (Busck), and codling moth, Cydia pomonella (L.), in pome and stone fruit orchards treated with sex pheromones. The combination of the sex pheromone of both species (PH combo lure) significantly increased G. molesta and marginally decreased C. pomonella captures as compared with captures of each species with either of their sex pheromones alone. The addition of a HPV combination lure [(E,Z)‐2,4‐ethyl decadienoate plus (E)‐β‐ocimene] or acetic acid used alone or together did not significantly increase the catch of either species in traps with the PH combo lure. The Ajar trap baited with terpinyl acetate and brown sugar (TAS bait) caught significantly more G. molesta than the delta trap baited with PH combo plus acetic acid in California during 2012. The addition of a PH combo lure to an Ajar trap significantly increased catches of G. molesta compared to the use of the TAS bait or PH combo lure alone in 2013. Female G. molesta were caught in TAS‐baited Ajar traps at similar levels with or without the use of additional lures. Ajar traps baited with the TAS bait alone or with (E)‐β‐ocimene and/or PH combo lures caught significantly fewer C. pomonella than delta traps with sex pheromone alone. Ajar traps with 6.4‐mm screened flaps caught similar numbers of total and female G. molesta as similarly baited open Ajar traps, and with a significant reduction in the catch of non‐targets. Broader testing of HPV and PH combo lures for G. molesta in either delta or screened or open Ajar traps is warranted.     

Effects of different sex pheromone compositions and host plants on the mating behavior of two Grapholita species

The two congener species Grapholita molesta and Grapholita dimorpha share two major sex pheromone components: cis-8-dodecenyl acetate (Z8-12Ac) and trans-8-dodecenyl acetate (E8-12Ac). In fact, commercial sex pheromone lures composed of only these two major components attract the males of both species. In this study, we aimed to determine the reproductive isolation components of these two species by analyzing the effects of the minor sex pheromone components and host plants. First, different ratios of the two major sex pheromone components were greatly favored by either male species. Sex pheromone gland extracts of G. dimorpha contained a lesser proportion of Z8-12Ac than that of G. molesta. In the three (apple, pear, and peach) orchards investigated in this study, a larger number of G. molesta males were attracted to the 95:5 pheromone mixture (Z8-12Ac and E8-12Ac, respectively), while a larger number of G. dimorpha males were attracted to the 85:15 mixture. Second, there was a significant variation in male attractions in different host plants. G. molesta males were more attracted to the sex pheromone lure in the apple orchards than that in the pear and peach orchards. In contrast, G. dimorpha males were more attracted to the lures in the pear and peach orchards than that in the apple orchard. Third, the minor sex pheromone components were important for reproductive isolation. Among the four minor components tested, addition of (Z)-8-dodecenol (Z8-12OH) to the major sex pheromone components significantly suppressed male attraction in G. dimorpha and slightly elevated male attraction in G. molesta. The discriminating effect of Z8-12OH was further validated using male electroantennogram analysis. These results suggest that reproductive isolation between two congeners can be achieved by variations in the minor sex pheromone components and in the host plants, as well as by changes in the ratio of the two major components.     

Simultaneous identification of Grapholita molesta Busck and Grapholita dimorpha Komai by PCR‐RFLP

Although several molecular diagnostic techniques are available for the identification of the apple‐feeding pests Grapholita molesta Busck and Grapholita dimorpha Komai, these pests are severely affecting apple orchards in Korea. These two pests may be misidentified or the available molecular diagnostic techniques may not facilitate the simultaneous identification of the morphological features of both species. In this study, we developed a multiplex assay for these two species using the polymerase chain reaction – restriction fragment length polymorphism (PCR‐RFLP) method. Sixty‐two specimens were collected from apples presumed infested with moth larvae and from pheromone traps from 2013 to 2014. Both species were identified morphologically, and a partial region of the cytochrome b gene was sequenced to design primers for PCR‐RFLP. Digestion profiles of G. molesta and G. dimorpha, using the Sau3A1 restriction enzyme, were characterized using three DNA fragments each for G. molesta (363 bp, 91 bp and 31 bp) and G. dimorpha (220 bp, 234 bp and 31 bp). The RFLP assay developed for both species in this study was more efficient and accurate than other currently used diagnostic assays and would be helpful to identify field‐collected specimens for pest control research.     

A comparison of various mating disruption technologies for control of two internally feeding Lepidoptera in apples

Sex pheromone mating disruption (MD) is an approach used to control several moth pest species of pome fruit by disrupting the ability of the males to find females and consequently prevent mating. The following experiments were performed to determine the effectiveness of several new and experimental sex pheromone MD technologies, and dispenser densities for simultaneous control of the codling moth (CM), Cydia pomonella (L.), and the oriental fruit moth (OFM), Grapholita molesta (Busck) (both Lepidoptera: Tortricidae), in Pennsylvania apple orchards. In one study, three MD approaches to control CM and oriental fruit moth – CM and OFM Disrupt Micro‐Flakes, Isomate CM/OFM TT, and both a CideTrak OFM and a CideTrak CM dispenser containing both codlemone and pear ester – and an insecticides‐only treatment were compared over the course of 2 years. In the other studies, the efficacy of several CheckMate Duel dispenser densities (i.e., 250, 375, 425, and 500 dispensers ha^?1) were compared against Isomate CM/OFM TT, and an insecticides‐only treatment. The CideTrak CM/pear ester combination and Isomate CM/OFM TT treatments both substantially reduced CM captures in traps in 2007 and 2008. Meanwhile, OFM trap shutdown was highest in the CheckMate Duel densities of 375 (99.9 ± 0.08%) and 500 dispensers ha^?1 (98.9 ± 0.07%) and the Isomate CM/OFM TT treatment (98.0 ± 1.13%), and lowest in the 250 dispensers ha^?1 density treatment (94.3 ± 3.23%). In orchards where OFM is the dominant pest species, a CheckMate Duel dispenser density of 375 ha^?1 is necessary for effective control, whereas higher densities are needed to control CM.     

The role of winter weather in the population dynamics of spring moths in the southwest Korean peninsula

Moth monitoring in a coniferous forest in southwestern Korea was carried out for 3 years in order to investigate the effects of winter weather factors on moth survival and emergence in early spring. A total of 686 individuals in 51 species of 4 families were collected during 3 spring sampling periods. The species richness observed for all 3 years was similar, though their abundance was differed. A total of 15 weather variables were analyzed including temperature and precipitation during the 4 months preceding April, and cumulative sums of 5-degree days with base temperatures ranging from 6 to 10 °C. Due to the multicollinearity of dependent variables, we adopted the eigenvectors of major axes from principal component analysis for multiple stepwise regression analysis. The species richness and abundance were significantly related to the first and second axes. Both axes were positively related with precipitation and temperature during the preceding 2 months, as well as the average winter temperature. This indicated that the effects of temperature and precipitation during winter played an important role in moth emergence in early spring, though the weather conditions of each month during winter varied significantly compared to the pattern of adult emergence.     

Vegetative and reproductive phenological traits of Rhizophora mucronata Lamk. and Sonneratia alba Sm.

Mangrove phenology is important in understanding the past, present and future response of mangrove species to impacts of climate change. Our study is the first long term direct observation of the phenology of Rhizophora mucronata and Sonneratia alba in Kenya. Objective of the study was to determine, interpret and document the timing of the various phenoevents and phenophases, and to establish relationships between phenology and the climatic variables. Phenological traits were investigated in six monospecific mangrove stands in Gazi Bay, south of Mombasa, for 2 years. Leaf emergence, leaf fall, flower bud, flower and fruit initiation data were recorded every fortnight in 54 shoots of 9 trees at each site. Continuous leaf emergence and loss characterized by multimodal peaks was observed for the two species. Leaf emergence and leaf fall peaked in the wet months and was reduced in the dry months. There was a relationship between the leaf emergence and drop with the reproductive phenology in the two species. Mean leaf longevity for R. mucronata and S. alba was 12.8 ± 1.2 and 4.9 ± 0.5 months respectively. The reproductive cycle took approximately 16–20 months in R. mucronata and 4–5 months in S. alba. Bud initiation in R. mucronata was seasonal and occurred in October and September. Buds were observed for 8–11 months developing slowly on the shoots. Shifts were observed in the timing of flower initiation, and the flowering period lasted in total for 4–5 months. The time period from flower appearance to the developmental start of immature propagules lasted about 1 month at reforested sites and up to 5 months in stands of natural vegetation. In R. mucronata initiation of immature propagules was not seasonal and varied among the sites. Fruiting in S. alba was short and seasonal without overlap, and shifts were observed in the timing of flowering, flowering and fruiting peaks. Abscission of reproductive parts started in July and in June, respectively, in the years 2005 and 2006. Fruiting was observed starting in August in both years, and fruiting peak was reached in October in 2005 and one month earlier in 2006. In R. mucronata vegetative and reproductive phenophases significantly correlated with climatic variables, whereas in S. alba only leaf emergence and leaf fall correlated with temperature. The flowering plasticity in the reproductive phenology of the two species indicates possible sensitivity to certain climatic and environmental triggers. Our results also indicate that R. mucronata trees have a distinctly higher investment in the reproductive cycle than S. alba.     

Different types of fruit damages of three internal apple feeders diagnosed with mitochondrial molecular markers

Three tortricid pests, Grapholita dimorpha (Komai), G. molesta (Busck), and Carposina sasakii (Matsumura), are known as internal apple feeders in Korea. To identify young larvae, this study developed two types of molecular markers from their mitochondrial DNA (mtDNA) sequences. To this end, six different loci of mtDNA were sequenced in G. dimorpha: cytochrome oxidase subunit I (460 bp), cytochrome oxidase subunit II (446 bp), cytochrome b (308 bp), NADH dehydrogenase 3 (585 bp), NADH dehydrogenase 4 (ND4, 835 bp), and 16S rRNA (1300 bp). These sequences were compared with those of G. molesta and C. sasakii in order to develop PCR–RFLP and diagnostic primers. ND4 locus was selected to be used for developing a PCR–RFLP marker. ND4-Swa I digests showed two bands for G. dimorpha, one band for G. molesta, and three bands for C. sasakii. On the other hand, species-specific diagnostic PCR primers were developed using ND4 locus. These markers were then applied to diagnose larvae infesting apples to determine species-specific fruit damage patterns, in which G. dimorpha, G. molesta, and C. sasakii showed different feeding behaviors in terms of their main feeding sites in apple fruits.