The fly factor phenomenon is mediated by interkingdom signaling between bacterial symbionts and their blow fly hosts

We tested the recent hypothesis that the"fly factor"phenomenon(food cur-rently or previously fed on by flies attracts more flies than the same type of food kept inccessible to flies)is mediated by bacterial symbionts deposited with feees or regur-gitated by feeding flies.We allowed laboratory-reared black blow flies,Phormia regina(Meigen),to feed and de fecate on bacterial Luria-Bertani medium solidified with agar,and isolated seven morphologically distinct bacterial colonies.We identified these us-ing matrix-assisted laser desorption/ionization mass spectrometry and sequencing of the 165 rRNA gene.In two-choice laboratory experiments,traps baited with cultures of Pro-teus mirabilis Hauser,Morganella morganii subsp.sibonii Jensen,or Serratia marcescens Bizio,captured significantly more flies than corresponding control jars baited with tryptic soy agar only.A mixture of seven bacterial strains as a trap bait was more attractive to flies than a single bacterial isolate(M.m.siboni).In a field experiment,traps baited with agar cultures of P:mirabilis and M.m siboni in combination captured significantly more flies than lraps baited with either bacterial isolate alone or the agar control.As evident by gas chromatography-mass spectrometry,the odor profiles of bacterial isolates differ,which may explain the additive effect of bacteria to the attractiveness of bacterial trap baits.As"generalist bacteria,"P mirabilis and M.m.sibonii growing on animal protein(beef liver)or plant protein(tofu)are similarly effective in attracting flies.Bacteria-derived airborne semiochemicals appear to mediate foraging by flies and to inform their feeding and oviposition decisions.     

Effective mosquito and arbovirus surveillance using metabarcoding

Effective vector and arbovirus surveillance requires timely and accurate screening techniques that can be easily upscaled. Next‐generation sequencing (NGS) is a high‐throughput technology that has the potential to modernize vector surveillance. When combined with DNA barcoding, it is termed ‘metabarcoding.’ The aim of our study was to establish a metabarcoding protocol to characterize pools of mosquitoes and screen them for virus. Pools contained 100 morphologically identified individuals, including one Ross River virus (RRV) infected mosquito, with three species present at different proportions: 1, 5, 94%. Nucleic acid extracted from both crude homogenate and supernatant was used to amplify a 269‐bp section of the mitochondrial cytochrome c oxidase subunit I (COI) locus. Additionally, a 67‐bp region of the RRV E2 gene was amplified from synthesized cDNA to screen for RRV. Amplicon sequencing was performed using an Illumina MiSeq, and bioinformatic analysis was performed using a DNA barcode database of Victorian mosquitoes. Metabarcoding successfully detected all mosquito species and RRV in every positive sample tested. The limits of species detection were also examined by screening a pool of 1000 individuals, successfully identifying the species and RRV from a single mosquito. The primers used for amplification, number of PCR cycles and total number of individuals present all have effects on the quantification of species in mixed bulk samples. Based on the results, a number of recommendations for future metabarcoding studies are presented. Overall, metabarcoding shows great promise for providing a new alternative approach to screening large insect surveillance trap catches.     

Assay of old-world screw-worm fly, Chrysomya bezziana, labelled with 32P

Techniques for ³²P labelling of larvae and adults of the screw-worm fly, Chrysomya bezziana, are described. Egg masses of labelled flies were readily identified. At the doses used for field releases, oviposition activity, fecundity, fertility and longevity of female flies were not adversely affected. Radioactive egg masses were recovered from sentinel animals following field release of labelled flies.

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Résumé Cet article décrit les techniques utilisées pour marquer avec le ³²p les larves et adultes de Chrysomya bezziana. Les pontes des mouches marquées ont été aisément identifiées. Aux doses utilisées pour le relâchement des insectes dans la nature, ni l'activité de ponte, ni la fécondité, fertilité et longévité des femelles n'ont été défavorablement modifiées. Les pontes radioactives ont été recueillies sur des animaux-pièges mis en place après le lâcher des mouches marquées.

     

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.     

Seasonal variation of tsetse fly species abundance and prevalence of trypanosomes in the Maasai Steppe,Tanzania

Tsetse flies, the vectors of trypanosomiasis, represent a threat to public health and economy in sub‐Saharan Africa. Despite these concerns, information on temporal and spatial dynamics of tsetse and trypanosomes remain limited and may be a reason that control strategies are less effective. The current study assessed the temporal variation of the relative abundance of tsetse fly species and trypanosome prevalence in relation to climate in the Maasai Steppe of Tanzania in 2014–2015. Tsetse flies were captured using odor‐baited Epsilon traps deployed in ten sites selected through random subsampling of the major vegetation types in the area. Fly species were identified morphologically and trypanosome species classified using PCR. The climate dataset was acquired from the African Flood and Drought Monitor repository. Three species of tsetse flies were identified: G. swynnertoni (70.8%), G. m. morsitans (23.4%), and G.pallidipes (5.8%). All species showed monthly changes in abundance with most of the flies collected in July. The relative abundance of G. m. morsitans and G. swynnertoni was negatively correlated with maximum and minimum temperature, respectively. Three trypanosome species were recorded: T. vivax (82.1%), T. brucei (8.93%), and T. congolense (3.57%). The peak of trypanosome infections in the flies was found in October and was three months after the tsetse abundance peak; prevalence was negatively correlated with tsetse abundance. A strong positive relationship was found between trypanosome prevalence and temperature. In conclusion, we find that trypanosome prevalence is dependent on fly availability, and temperature drives both tsetse fly relative abundance and trypanosome prevalence.     

Factors affecting the efficacy of a vinegar trap for Drosophila suzikii (Diptera; Drosophilidae)

Drosophila suzukii, Matsumura, is a relatively new pest in the United States attacking a variety of fruit crops. Studies were conducted to develop a standardized, economical trap for monitoring. Laboratory bioassays found that flies were attracted to dark colours ranging from red to black. Similarly, fly catches in 237‐ml plastic ‘spice’ jars with ten 0.48‐cm holes and baited with apple cider vinegar were significantly higher in jars with red or black than white caps. The use of an alternating set of three, horizontal red, black and red bands (‘Zorro’ trap) significantly increased fly catches compared with the use of all‐red or all‐black strips. This increase was associated with a significantly higher proportion of flies first landing on the side near the openings of the trap instead of on the cap with the ‘Zorro’ trap compared with the other traps. Laboratory data were used to develop a predictive model to define total fly capture as a function of trap colour/colour pattern, cumulative area of entry holes and the length of the trapping portion of the trap. Total fly catches by the ‘Zorro’ trap were compared with other red and clear plastic traps in five field trials conducted in several cultivated and uncultivated sites. Comparisons included a commercial red‐capped 200‐ml trap with two 0.63‐cm holes, an all‐red spice jar with ten 0.48‐cm holes and clear and red 473‐ml and clear 946‐ml plastic cups with six or ten 0.48‐ or 0.63‐cm holes. The model was successfully validated, suggesting that performance of cup traps can be predicted based on a few characteristics. The current ‘Zorro’ trap did not catch most of the flies among trap designs, but showed some advantages, including the durability and potential to recycle the plastic, small size, commercial availability and its greater selectivity for D. suzukii than the other traps tested.     

Electronic traps for detection and population monitoring of adult fruit flies (Diptera: Tephritidae)

During the last decades, the economic importance of tephritid fruit flies (FF) has increased worldwide because of recurrent invasions and expansions into new areas, and reduced control capabilities of current control systems. Efficient monitoring systems, thus, are required to provide fast information to act promptly. With this aim in mind, we developed two electronic trap (e‐trap) versions for adult FF: one with specific volatiles for male and female adult Ceratitis capitata, and the second, based on the attraction of adult FF to yellow colour, targeting Dacus ciliatus, Rhagoletis cerasi and Bactrocera oleae. In the case of B. oleae, the female pheromone and ammonium bicarbonate were added as synergists. In the two versions, attracted FF were retained in the trap on glued surfaces. Real‐time images of the surfaces were automatically taken and transmitted to a server. We tested the two e‐trap versions in insect‐proof cages, where flies were released and recaptured, and in commercial orchards throughout the Mediterranean: C. capitata in peach orchards in Italy; R. cerasi in cherry orchards in Greece; B. oleae in olive orchards in Spain and in Greece; and D. ciliatus in melons in plastic tunnels in Israel. The e‐trap showed excellent abilities to transmit real‐time images of trapped FF and a high specificity for trapping different FF species. The ability of the entomologist to correctly classify FF from images in the office was >88%. In addition, average number of flies/trap in e‐trap grids did not differ from numbers reported on grids of conventional traps that were operating simultaneously. The e‐traps developed and tested in this study provide the basis for the real‐time monitoring of FF were no olfactory attractants are available, and for the surveillance of alien FF incursions where generic, but not specific, olfactory attractants exists.     

Evidence for sexual isolation as a prezygotic barrier to gene flow between morphologically divergent species of Rhagoletis fruit flies

1. Certain groups of fruit flies in the genus Rhagoletis (Diptera: Tephritidae) are exemplars for sympatric speciation via host plant shifting. Flies in these species groups are morphologically similar and overlap in their geographic ranges, yet attack different, non‐overlapping sets of host plants. Ecological adaptations related to differences in host choice and preference have been shown to be important prezygotic barriers to gene flow between these taxa, as Rhagoletis flies mate on or near the fruit of their respective host plants. Non‐host‐related assortative mating is generally absent or present at low levels between these sympatrically diverging fly populations. 2. However, some Rhagoletis taxa occasionally migrate to ‘non‐natal’ plants that are the primary hosts of other, morphologically differentiated fly species in the genus. These observations raise the question of whether sexual isolation may reduce courtship and copulation between morphologically divergent species of Rhagoletis flies, contributing to their prezygotic isolation along with host‐specific mating. 3. Using reciprocal multiple‐choice mating trials, we measured sexual isolation among nine species pairs of morphologically differentiated Rhagoletis flies. Complete sexual isolation was observed in eight of the nine comparisons, while partial sexual isolation was observed in the remaining comparison. 4. We conclude that sexual isolation can be an effective prezygotic barrier to gene flow contributing to substantial reproductive isolation between many morphologically distinct Rhagoletis species, even in the absence of differential host plant choice and host‐associated mating.     

Performance of light‐emitting diode traps for collecting sand flies in entomological surveys in Argentina

The performance of two light‐emitting diode traps with white and black light for capturing phlebotomine sand flies, developed by the Argentinean Leishmaniasis Research Network (REDILA‐WL and REDILA‐BL traps), were compared with the traditional CDC incandescent light trap. Entomological data were obtained from six sand fly surveys conducted in Argentina in different environments. Data analyses were conducted for the presence and the abundance of Lutzomyia longipalpis, Migonemyia migonei, and Nyssomyia whitmani (106 sites). No differences were found in presence/absence among the three types of traps for all sand fly species (p>0.05). The collection mean of Lu. longipalpis from the REDILA‐BL didn´t differ from the CDC trap means, nor were differences seen between the REDILA‐WL and the CDC trap collection means (p>0.05), but collections were larger from the REDILA‐BL trap compared to the REDILA‐WL trap (p<0.05). For Mg. migonei and Ny. whitmani, no differences were found among the three types of traps in the number of individuals captured (p>0.05). These results suggest that both REDILA traps could be used as an alternative capture tool to the original CDC trap for surveillance of these species, and that the REDILA‐BL will also allow a comparable estimation of the abundance of these flies to the CDC light trap captures. In addition, the REDILA‐BL has better performance than the REDILA‐WL, at least for Lu. longipalpis.     

Evaluation of ammonia,human sweat,and bovine blood as attractants for the female canyon fly,Fannia conspicua (Diptera: Muscidae), in southern California

Human sweat, liquid ammonia, and bovine blood are known to be attractive to some hematophagous flies. These materials were evaluated with and without carbon dioxide (CO₂) for their ability to increase capture of female “canyon flies” (Fannia conspicua Malloch) using CDC‐type suction traps (without light). Ammonia acted synergistically with CO₂ to increase trap catch 89.9% over CO₂ alone. There was no synergistic effect of human sweat or bovine blood with CO₂. In the absence of CO₂, none of the three materials increased trap catch of female canyon flies relative to non‐baited traps. Implications for canyon fly control and further trap improvement are discussed.     

First use of pheromones to detect phenology patterns and density relationships of pine sawflies in German forests

1 Monitoring studies of pine sawflies with pheromone traps were performed for the first time in Germany. Pheromone traps baited with species‐specific pheromone substances were installed in pine forests at different locations in Bavaria, Brandenburg and Lower Saxony during two years. 2 It was possible to track the flight phenology of Diprion pini, Gilpinia pallida and Neodiprion sertifer reliably and to get information about the number of generations of these species in 1997 and 1998. 3 A clear relationship between trap catch and population density could not be detected, but qualitative changes in trap catch caused by different density levels were observed. 4 For D. pini, trap catches were different among endemic populations of different forest types. Furthermore, catches of males reflected the results from the regular cocoon collections by foresters during the previous winter. 5 For N. sertifer, trap catches in endemic populations were well separated from trap catches on sites with higher sawfly densities. However, no significant correlation between trap catch and sawfly density or defoliation level could be found. These results suggest that the efficacy of the pheromone traps probably varied with biological features (sex ratio, density level, immigration) of the particular population.     

Age‐specific changes in sperm levels among female tsetse (Glossina spp.) with a model for the time course of insemination

The age, insemination and ovulation status of tsetse flies Glossina pallidipes Austen (n = 154369) and Glossina morsitans morsitans Westwood (n = 19659), captured over 11 years in Zimbabwe, are assessed by ovarian dissection. Instantaneous rates of insemination increase exponentially with age in both species; 90% insemination levels are reached after 5 days post‐emergence in G. m. morsitans and 7 days in G. pallidipes, varying little with season. More than 95% of both species have ovulated by the age of 8 days and 99% by 12 days. Older flies that have not ovulated are > 100‐fold more likely to be caught in October and November than in other months. A 500‐fold decrease in trap catches did not result in any detectible decrease in the probability of females being inseminated. The proportion of partially filled spermathecae rises for approximately 6 days then declines, consistent with some flies having mated more than once. For flies caught on electric nets, with wings undamaged during capture, wing‐fray data are used to extend ovarian age estimates up to 11 ovulations. Among these flies, the volume of sperm in the spermathecae declines little in flies that have ovulated up to seven times; thereafter, it declines by approximately 1% per ovulation. The time course of insemination and the mating frequency of females are important considerations in modelling tsetse fly populations, as well as for the dynamics of interventions involving the release of genetically‐modified insects, which should not be seriously compromised by the limited levels of polyandry currently observed.     

Kairomone attractant for the leafmining fly, Liriomyza bryoniae (Diptera, Agromyzidae)

A field test carried out in an industrial greenhouse in Lithuania revealed the attractiveness of synthetic methyl salicylate (MeSa) towards an economically important leafmining tomato pest, Liriomyza bryoniae (Kaltenbach) (Diptera, Agromyzidae). The behavioural reaction of the flies depended very much on the simultaneous presence of both olfactory and visual stimuli. The attractiveness depended on the colour of a sticky trap: MeSa attracted significantly more flies (ca. 2.2 times) when placed in yellow traps than in aluminium foil colour ones, when catches in such traps were compared to a corresponding control. L. bryoniae is the first species within the Agromyzidae family attracted by MeSa. The attractant was attributed to kairomones, as the compound is known as a plant-produced volatile. MeSa can be an effective extra-tool for increasing the attractiveness of traps. It should be evaluated in future whether such trap/bait combination is effective for the mass trapping of L. bryoniae leafminers in greenhouses (closed area).     

Larval fat body and its relationship to protein storage and ovarian development in adults of the screw-worm fly Chrysomya bezziana

The vast majority of females of the Old World screw-worm fly, Chrysomya bezziana Villeneuve (Diptera: Calliphoridae), from field populations and from normally maintained laboratory cultures are autogenous. Anautogenous C. bezziana females which are smaller in size are produced when the larval feeding period is curtailed. Autogenous and anautogenous C. bezziana were compared with reference to abdominal larval fat body (LFB), ovarian development and protein relationships. At emergence, autogenous C. bezziana have larger amounts of LFB relative to body size than anautogenous C. bezziana. Volume of LFB is directly correlated with body size in females showing autogeny but there is no correlation with size in females in the anautogenous size classes. Males and females of equal sizes have the same volume of LFB and their rates of LFB depletion are similar and temperature-dependent. In autogenous females, about 70% of the LFB disappears before vitellogenesis begins. LFB disappears more rapidly in smaller flies. In anautogenous C. bezziana females, access to a source of protein from the time of adult emergence reduced the rate of LFB depletion, but in autogenous flies the depletion rate is not affected by diet.LFB protein accounts for about 24% of total protein in newly emerged autogenous females of C. bezziana. The protein content of the mature ovaries of non-protein-fed wild C. bezziana is 85% of that of the original LFB protein. Non-protein-fed anautogenous C. bezziana of the same age have negligible quantities of ovary protein.

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Résumé Les femelles provenant de populations naturelles ou d'élevages traditionnels en laboratoire de la mouche Chrysomya bezziana sont en large majorité autogènes. Des femelles de C. bezziana non-autogènes et de taille réduite sont obtenues lorsque la durée de l'alimentation larvaire est abrégée. Les relations entre corps gras abdominal des larves (CGL = LFB dans le texte anglais), développement ovarien et réserves protéiniques ont été comparées chez les femelles autogènes et non-autogènes. A l'émergence, les femelles autogènes ont un CGL rapporté au poids corporel plus abondant que les femelles non-autogènes. Le volume du CGL est en relation directe avec la taille des femelles autogènes, mais cette corrélation ne se vérifie pas chez les femelles non-autogènes. A taille égale, mâles et femelles ont un volume égal de CGL, lequel s'épuise à la même vitesse, dépendant de la température. Chez les femelles autogènes, 70% environ du CGL disparaît avant le début de la vitellogenèse. Le CGL décroît plus rapidement ches les mouches de petite taille. Chez les femelles autogènes, le taux d'utilisation du CGL n'est past affecté par l'alimentation; chez les femelles non-autogènes en revanche, l'accès à une source de protéines après émergence des adultes réduit le taux d'utilisation du CGL.Les protéines provenant du CGL représentent quelque 24% du total des protéines ches les femelles autogènes fraîchement émergées. Chez C. bezziana, prélevé dans des populations naturelles et n'ayant pas reçu d'appoint en protéines, les ovaires mûrs contiennent 82% des protéines du CGL d'origine. Au même âge, les femelles non-autogènes n'ayant reçu aucun appoint en protéines ont des quantités insignifiantes de protéines ovariennes.

     

Ingestion of food by the adult screw-worm fly, Chrysomya bezziana (Diptera, Calliphoridae)

When females of the screw-worm fly, Chrysomya bezziana Villeneuve, were given a choice of solutions of protein, sucrose and water, their protein intake was substantial only on days 2, 3 and 4 after emergence, and during the 3 days following the first oviposition. Protein consumption by males remained consistently low for 10 days following emergence.Protein ingestion by laboratory-bred autogenous and anautogenous females, autogenous wild females of C. bezziana, and females of the anautogenous blowly species, Chrysomya megacephala (F), was compared. During the first ovarian cycle (days 1–6), protein solution accounted for 9.5% and 9.4% of the total liquid intake of laboratory-bred and wild anautogenous females of C. bezziana. Protein ingestion by C. megacephala females during the first ovarian cycle represented 23% of the total liquids. During the second ovarian cycle in C. bezziana, protein ingestion represented 21% of total liquids.The presence of dry sucrose or sucrose solution of high concentration depressed protein intake during the first ovarian cycle in C. bezziana.Most feeding by females occurred during the photophase with early morning and late evening peaks in carbohydrate ingestion and a late afternoon peak in protein ingestion.The ecological implications of the results are discussed.

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Résumé Quand les femelles de Chrysomya bezziana Villeneuve ont le choix entre des solutions de protéine, de sucrose et d'eau, la consommation de protéines n'est importante que lesond, troisième et quatrième jours après l'émergence, et pendant les 3 jours suivant la première ponte. La consommation de protéines par les mâles reste faible pendant les 10 jours qui suivent l'émergence.Les consommations de protéines par des femmelles de Chrysomya bezziana (sauvages autogènes, elevées en laboratoire autogènes et anautogènes) et par des femelles de l'espèce anautogène Chrysomya megacephala F. ont été comparées. Pendant le premier cycle ovarien (jours 1 à 6) les solutions de protéine ingérées par les femelles autogènes (issues du laboratoire et sauvages de Chrysomya bezziana) correspondent respectivement à 9,5% et 9,4% du liquide total absorbé. Pendant le premier cycle ovarien la quantité de protéine ingérée par les femelles de Chrysomya megacephala représente 23% du liquide absorbé total. Pendant le second cycle ovarien de Chrysomya bezziana, les protéines ingérées représentent 21% de l'absorption totale de liquide.La présence de sucrose sec ou en liquide à concentration élevée réduit la consommation de protéines pendant le premier cycle ovarien de Chrysomya bezziana.L'essentiel de l'alimentation des femelles en carbohydrates a lieu pendant la photophase avec des pics tôt le matin et tard le soir, et avec un pic en find d'après midi pour les protéines.Les conséquences écologiques de ces résultats sont discutées.

     

DNA barcoding for identification of sand flies (Diptera: Psychodidae) in India

About 50 species of sand flies have been reported to be prevalent in India. We explored the utility of the DNA barcode approach towards species identification of these medically important insects. A total of 62 specimens belonging to seven morphologically identified species of two genera, Phlebotomus and Sergentomyia, collected from Puducherry Union Territory, Maharashtra and Rajasthan states of India were subjected to the analysis. Neighbor‐joining (NJ) analysis of DNA barcode sequences identified the individuals of seven morphological species into eight distinct species, as presented in the designed NJ tree. This methodology delineated morphologically identified species, S. bailyi, into two genetically isolated groups. Also, this study characterizes DNA barcodes of P. argentipes and P. papatasi, the vector species of leishmaniasis in India, for the first time.     

Comparison of different trapping methods for surveillance of mosquito vectors of West Nile virus in Rhône Delta,France

Five trapping methods were compared for monitoring potential vectors of the West Nile virus in four areas in the Camargue Plain of France: carbon dioxide traps, bird‐baited traps, gravid traps, resting boxes, and human landing catches. A total of 73,721 specimens, representing 14 species, was trapped in 2006. Results showed significant differences in species and abundance between the type of traps. Many more specimens were collected using CO₂ traps than any other method, with an average of 212 specimens per night per trap (p<0.05). Culex pipiens was the most abundant species collected (36.8% of total with CO₂ traps), followed by Aedes caspius (22.7%), Anopheles hyrcanus (18.3%), Culex modestus (18.3%), and Aedes detritus (3.2%). Bird‐baited traps captured only eight specimens per night per trap on average, mainly Cx. pipiens (89.9%). The species collected and their abundance are influenced by the trap location, at ground or canopy level. Culex pipiens was twice as abundant in the canopy as on the ground, whereas it was the opposite for Ae. caspius, An. hyrcanus, and Ae. detritus. Culex modestus was equally abundant at both levels. Resting boxes and gravid traps were much less efficient, capturing around 0.3 specimens per night per trap. Results are discussed in relation to West Nile virus surveillance.     

Identification of forensically important blow fly species (Diptera: Calliphoridae) in China by mitochondrial cytochrome oxidase I gene differentiation

Abstract Unambiguous and rapid sarcosaphagous insect species identification is an essential requirement for forensic investigations. Although some insect species are difficult to classify morphologically, they can be effectively identified using molecular methods based on similarity with abundant authenticated reference DNA sequences in local databases. However, local databases are still relatively incomplete in China because of the large land area with distinct regional conditions. In this study, 75 forensically important blow flies were collected from 23 locations in 16 Chinese provinces, and a 278‐bp segment of the cytochrome oxidase subunit I gene of all specimens was successfully sequenced. Phylogenetic analysis of the sequenced segments showed that all Calliphorid specimens were properly assigned into nine species with relatively strong supporting values, thus indicating that the 278‐bp cytochrome oxidase subunit one region is suitable for identification of Calliphorid species. The clear difference between intraspecific threshold and interspecific divergence confirmed the potential of this region for Calliphorid species identification, especially for distinguishing between morphologically similar species. Intraspecific geographic variations were observed in Lucilia sericata (Meigen, 1826) and Lucilia caesar (Linnaeus, 1758).     

Field evaluation of the relative attractiveness of enriched ginger root oil (EGO)lure and trimedlure for African Ceratitis species (Diptera: Tephritidae)

The males of some fruit flies (Diptera: Tephritidae) are known to be attracted to specific parapheromones. The trapping results between trimedlure (TML) and enriched ginger root oil (EGOlure) were compared at two experimental sites in Morogoro (Central Tanzania) for a period of 12 weeks co‐inciding with the main citrus season. Both attractants captured a comparable diversity of fruit flies, except that EGOlure also attracted fruit flies, such as Ceratitis cosyra, not normally found in TML‐baited traps. Both EGOlure and TML attracted mainly or exclusively male fruit flies, but the catches with EGOlure were equal or superior to those with TML. It is concluded that EGOlure should be considered as a suitable alternative for TML in detection, monitoring and control programs for African fruit flies of the genus Ceratitis. It has the added advantage that it combines the attractiveness with regard to species spectrum of both TML and terpinyl acetate.     

Molecular identification of mosquitoes (Diptera: Culicidae) in southeastern Australia

DNA barcoding is a modern species identification technique that can be used to distinguish morphologically similar species, and is particularly useful when using small amounts of starting material from partial specimens or from immature stages. In order to use DNA barcoding in a surveillance program, a database containing mosquito barcode sequences is required. This study obtained Cytochrome Oxidase I (COI) sequences for 113 morphologically identified specimens, representing 29 species, six tribes and 12 genera; 17 of these species have not been previously barcoded. Three of the 29 species ─ Culex palpalis, Macleaya macmillani, and an unknown species originally identified as Tripteroides atripes ─ were initially misidentified as they are difficult to separate morphologically, highlighting the utility of DNA barcoding. While most species grouped separately (reciprocally monophyletic), the Cx. pipiens subgroup could not be genetically separated using COI. The average conspecific and congeneric p‐distance was 0.8% and 7.6%, respectively. In our study, we also demonstrate the utility of DNA barcoding in distinguishing exotics from endemic mosquitoes by identifying a single intercepted Stegomyia aegypti egg at an international airport. The use of DNA barcoding dramatically reduced the identification time required compared with rearing specimens through to adults, thereby demonstrating the value of this technique in biosecurity surveillance. The DNA barcodes produced by this study have been uploaded to the ‘Mosquitoes of Australia–Victoria’ project on the Barcode of Life Database (BOLD), which will serve as a resource for the Victorian Arbovirus Disease Control Program and other national and international mosquito surveillance programs.