José Roberto Pujol-Luz
Full Text Available Intra-puparial development of the females of Chrysomya albiceps (Wiedemann (Diptera, Calliphoridae. The chronology and morphological changes that take place during intra-puparial development of Chrysomya albiceps is described based on 254 specimens reared in the laboratory. Larvae were obtained from the eggs laid by a single female. The pre-pupae were separated according to the reduction of larval length and the degree of pigmentation and sclerotization of the cuticle. After pupation, 10 individuals were fixed in Carnoy's solution and preserved in 70% ethanol, 10 individuals were fixed every 3 hours up to complete the first 24 hours (n = 80, the remaining individuals were fixed every six hours up to the 90th hour (n = 110 when 54 females emerged. The pupae were immersed in 5% formic acid for 48 hours and maintained in 70% ethanol, and then dissected and analyzed. C. albiceps shows four intra-puparial stages, each of which were described and compared with those described for Musca domestica, Calliphora erythrocephala, Sarcophaga bullata, Cuterebra tenebrosa, Oestrus ovis and Dermatobia hominis. Four developmental stages may be described: (1 the larva-pupa apolysis, after three hours; (2 the criptocephalic pupa, after six hours; (3 the phanerocephalic pupa, after nine hours; (4 the pharate pupa, after nine hours. The pharate adult is completely formed after 81 hours.
Moore, HE; Pechal, JL; Benbow, ME; Drijfhout, FP
Creative Commons Attribution 4.0 International License Cuticular hydrocarbons (CHC) have been successfully used in the field of forensic entomology for identifying and ageing forensically important blowfly species, primarily in the larval stages. However in older scenes where all other entomological evidence is no longer present, Calliphoridae puparial cases can often be all that remains and therefore being able to establish the age could give an indication of the PMI. This paper examined ...
Zhu, Guang-Hui; Jia, Zheng-Jun; Yu, Xiao-Jun; Wu, Ku-Sheng; Chen, Lu-Shi; Lv, Jun-Yao; Eric Benbow, M
Preadult development of necrophagous flies is commonly recognized as an accurate method for estimating the minimum postmortem interval (PMImin). However, once the PMImin exceeds the duration of preadult development, the method is less accurate. Recently, fly puparial hydrocarbons were found to significantly change with weathering time in the field, indicating their potential use for PMImin estimates. However, additional studies are required to demonstrate how the weathering varies among species. In this study, the puparia of Chrysomya rufifacies were placed in the field to experience natural weathering to characterize hydrocarbon composition change over time. We found that weathering of the puparial hydrocarbons was regular and highly predictable in the field. For most of the hydrocarbons, the abundance decreased significantly and could be modeled using a modified exponent function. In addition, the weathering rate was significantly correlated with the hydrocarbon classes. The weathering rate of 2-methyl alkanes was significantly lower than that of alkenes and internal methyl alkanes, and alkenes were higher than the other two classes. For mono-methyl alkanes, the rate was significantly and positively associated with carbon chain length and branch position. These results indicate that puparial hydrocarbon weathering is highly predictable and can be used for estimating long-term PMImin.
Zhang, Q.; Žďárek, Jan; Nachman, R. J.; Denlinger, D. L.
Roč. 29, č. 2 (2008), s. 196-205 ISSN 0196-9781 Institutional research plan: CEZ:AV0Z40550506 Keywords : diapause hormone * neuropeptide analogs * diapause termination * pupariation * Helicoverpa zea Subject RIV: CC - Organic Chemistry Impact factor: 2.565, year: 2008
Moore, Hannah E; Pechal, Jennifer L; Benbow, M Eric; Drijfhout, Falko P
Cuticular hydrocarbons (CHC) have been successfully used in the field of forensic entomology for identifying and ageing forensically important blowfly species, primarily in the larval stages. However in older scenes where all other entomological evidence is no longer present, Calliphoridae puparial cases can often be all that remains and therefore being able to establish the age could give an indication of the PMI. This paper examined the CHCs present in the lipid wax layer of insects, to determine the age of the cases over a period of nine months. The two forensically important species examined were Calliphora vicina and Lucilia sericata. The hydrocarbons were chemically extracted and analysed using Gas Chromatography - Mass Spectrometry. Statistical analysis was then applied in the form of non-metric multidimensional scaling analysis (NMDS), permutational multivariate analysis of variance (PERMANOVA) and random forest models. This study was successful in determining age differences within the empty cases, which to date, has not been establish by any other technique.
A programme to develop genetic sexing mechanisms in the Mediterranean fruit fly (medfly), Ceratitis capitata (Wiedemann), was initiated at the IAEA Laboratories, Seibersdorf, in 1983. Because of the potential benefits arising from the elimination of females early in the developmental cycle, combined with the anticipated relative ease of inducing temperature sensitive lethal (tsl) factors, it was decided to attempt to induce and isolate tsl factors active in the egg or early larval stages. Initially, five recombination suppressor (RS) strains were isolated. The degree of recombination suppression ranged from 77.6% to 99.1%. The viability of each of the five RS strains was assessed and RS 30/55 was selected as the most suitable strain. Ethyl methanesulphonate (EMS) was used to induce the tsl factors, by feeding two-day old adult males with a suspension of EMS in a 10% solution of sugar in the drinking water supply. Temperature tolerance tests indicated a discriminating temperature of 32 deg. C when isolating tsl factors active in the egg stage and 35 deg. C when isolating such factors in the early larval stage. A total of 39 and 22 tsl factors have been isolated in the two stages, respectively. However, none has yet proved stable. Induction of tsl factors with a reduced dose of EMS is now being attempted. An alternative genetic sexing programme was initiated in 1985, based on the use of pupal colour dimorphisms. Previously, a genetic sexing strain, T:Y(wp + )101, based on a white female/brown male puparial colour dimorphism, had twice been assessed for stability under mass rearing conditions. In both cases the sexual colour dimorphism disintegrated immediately. Another similarly dimorphic strain, T:Y(wp + )30C, was developed. This strain remained stable for seven generations of mass rearing, after which it started to disintegrate. Disintegration of this strain was probably caused by accidental contamination by wild type medflies. 34 refs, 1 fig., 1 tab
José Roberto Pujol-Luz
Full Text Available Intra-puparial development of the females of Chrysomya albiceps (Wiedemann (Diptera, Calliphoridae. The chronology and morphological changes that take place during intra-puparial development of Chrysomya albiceps is described based on 254 specimens reared in the laboratory. Larvae were obtained from the eggs laid by a single female. The pre-pupae were separated according to the reduction of larval length and the degree of pigmentation and sclerotization of the cuticle. After pupation, 10 individuals were fixed in Carnoy's solution and preserved in 70% ethanol, 10 individuals were fixed every 3 hours up to complete the first 24 hours (n = 80, the remaining individuals were fixed every six hours up to the 90th hour (n = 110 when 54 females emerged. The pupae were immersed in 5% formic acid for 48 hours and maintained in 70% ethanol, and then dissected and analyzed. C. albiceps shows four intra-puparial stages, each of which were described and compared with those described for Musca domestica, Calliphora erythrocephala, Sarcophaga bullata, Cuterebra tenebrosa, Oestrus ovis and Dermatobia hominis. Four developmental stages may be described: (1 the larva-pupa apolysis, after three hours; (2 the criptocephalic pupa, after six hours; (3 the phanerocephalic pupa, after nine hours; (4 the pharate pupa, after nine hours. The pharate adult is completely formed after 81 hours.Desenvolvimento intra-pupal de fêmeas de Chrysomya albiceps (Wiedemann (Diptera, Calliphoridae A cronologia e as mudanças morfológicas que ocorrem durante o desenvolvimento intra-pupal de Chrysomya albiceps são descritos com base em 254 espécimes criados em laboratório. As larvas foram obtidas a partir os ovos postos por uma única fêmea. As pré-pupas foram separadas de acordo com a redução do comprimento larval, o grau de pigmentação e esclerotização da cutícula, depois da formação das pupas, 10 indivíduos foram fixados em solução de Carnoy e conservados em etanol
Methyl farnesoate (MF) and juvenile hormone (JH III), which respectively bind to the receptors USP and MET, and bisepoxy JH III (bisJHIII) were assessed for several activities during Drosophila larval development, and during prepupal development to eclosed adults. Dietary MF and JH III were similar...
Disruption of pupariation and eclosion behavior in the flesh fly, Sarcophaga bullata Parker (Diptera: Sarcophagidae), by venom form the ectoparasitic wasp Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae)
Rivers, D. B.; Žďárek, Jan; Denlinger, D. L.
Roč. 57, - (2004), s. 78-91 ISSN 0739-4462 R&D Projects: GA ČR GA522/01/0501 Grant - others:USDA-NRI(US) 98-35302-6659; USDA-NRICGP(US) 2001-1005 Institutional research plan: CEZ:AV0Z4055905 Keywords : ectoparasitoid * ecdysone * fly development Subject RIV: CC - Organic Chemistry Impact factor: 1.173, year: 2004
Sukontason, Kom; Piangjai, Somsak; Siriwattanarungsee, Sirisuda; Sukontason, Kabkaew L
The larval morphology and developmental rate of Chrysomya megacephala (F.) and Chrysomya rufifacies (Macquart), the two most forensically important blowfly species in Thailand, are presented. Morphological comparison of the third instar of both species revealed different characteristics (e.g., body appearance, cephalopharyngeal skeleton, dorsal cuticular spines between the prothorax and mesothorax, and feature of the posterior spiracle), thereby, allowing correct identification. A data analysis was conducted in Chiang Mai province, Northern Thailand during 2000-2001 on the developmental rate of both flies under natural ambient temperature and a natural light-dark photoperiod. The results indicated that larvae of C. megacephala developed more rapidly in April, pupariation initiated at 84 h at temperatures averaging 31.4 degrees C, and the larvae grew slower in the rainy season and winter. Similarly, rapid development of C. rufifacies larvae appeared in the summer, with a pupariation period as short as 96 h in June (average temperature 27.4 degrees C). Analysis of the median body length of C. megacephala and C. rufifacies larvae in different seasons of the years 2000-2001 in Thailand revealed that both species developed rapidly in the summer; pupariation of C. rufifacies initiated at 144 h, while C. megacephala initiated pupariation at 156 h. This information is potentially useful for estimating the postmortem interval of a corpse in forensic investigations, where the corpse becomes infesting with these fly species.
Walker, Jane; Kwon, So Yeon; Badenhorst, Paul
, larval growth is dramatically impaired, with progression to the third instar delayed for ~24 hr, and pupariation occurring only at day 14 after egg laying. Melanotic nodules appear after 4 days. Microarray analysis shows that stress response genes are induced and ecdysone-induced transcription factors...
Myška, Petr; Žďárek, Jan
Roč. 102, č. 1 (2005), s. 21-26 ISSN 1210-5759 R&D Projects: GA ČR(CZ) GA522/01/0501 Institutional research plan: CEZ:AV0Z4055905 Keywords : ecdysteroids * metamorphosis behaviour * pupariation Subject RIV: CC - Organic Chemistry Impact factor: 0.745, year: 2005
Verma, Pushpa; Cohen, Stephen M
signaling downregulates miR-965 at the onset of pupariation, linking activation of the histoblast nests to the hormonal control of metamorphosis. Replacement of the larval epidermis by adult epidermal progenitors involves regulation of both cell-intrinsic events and cell communication. By regulating both...
Shen, K; Hu, J; Wu, B; An, K; Zhang, J; Liu, J; Zhang, R
The melon fly, Bactrocera cucurbitae (Coquillett), and the pumpkin fly, Bactrocera tau (Walker), are economically important pests that attack mainly cucurbitacean fruits. The two fruit fly species have similar natural distributions, host ranges, and population growth capacities. This study was designed to assess the asymmetrical competitions through resource exploitation between the larvae of B. cucurbitae and B. tau at different density levels and temperatures, and on different hosts by comparing the relative effects of interspecific and intraspecific interactions on four life history parameters: survival rate, puparial mass, puparial duration, and developmental duration. Our results showed that intraspecific and interspecific competitions occurred under some laboratory conditions, and B. cucurbitae took advantage over B. tau at the high-density level and at low and high temperatures on pumpkin, bitter gourd, and bottle gourd when interspecific competition took place. Intraspecific and interspecific competitions mainly affected the puparial mass and the survival rate of the two fruit fly species but had no marked effect on the puparial duration or development duration.
Robinson, L A; Bryson, D; Bulling, M T; Sparks, N; Wellard, K S
Developmental data of forensically important blowflies used by entomologists to estimate minimum post mortem interval (mPMI) are established under controlled laboratory conditions for various temperature ranges throughout the stages of egg, 1st-3rd instar, puparia, and adult fly emergence. However, environmental conditions may influence the patterns of development and behaviour of blowflies, potentially impacting on these established development rates. Previous studies investigating indoor colonisation have focused on the delay to oviposition, with behaviour during the post-feeding phase in this setting often overlooked. The environment in which third instar larvae disperse when searching for a pupariation site may vary drastically at both outdoor and indoor scenarios, influencing the activity and distance travelled during this phase and possibly affecting developmental rates. This study investigated the effect of eight common domestic indoor surfaces on dispersal time, distance travelled, and behaviour of post-feeding Lucilia sericata as well as any resulting variation in development. It was found that pupariation and puparia length within a pupariation medium of sawdust (often used in laboratory settings) produced comparable results with that of carpeted environments (those deemed to be 'enclosed'). Non-carpeted environments (those which were 'exposed') produced a delay to pupariation likely due to increased activity and energy expenditure in searching for pupariation sites which enabled burial. In addition, the observed speed of travel during dispersal was seen via time lapse photography to be greater within 'exposed' conditions. Larvae which dispersed upon burnt laminate flooring were observed to travel faster than in all other conditions and showed the only significant variation (P=0.04) in the day of emergence in comparison to the control condition of sawdust. This study has demonstrated that wandering phase activity is affected by the environmental surface
Nicholas D Testa
Full Text Available The selective pressures leading to the evolution of Sexual Size Dimorphism (SSD have been well studied in many organisms, yet, the underlying developmental mechanisms are poorly understood. By generating a complete growth profile by sex in Drosophila melanogaster, we describe the sex-specific pattern of growth responsible for SSD. Growth rate and critical size for pupariation significantly contributed to adult SSD, whereas duration of growth did not. Surprisingly, SSD at peak larval mass was twice that of the uneclosed adult SSD with weight loss between peak larval mass and pupariation playing an important role in generating the final SSD. Our finding that weight loss is an important regulator of SSD adds additional complexity to our understanding of how body size is regulated in different sexes. Collectively, these data allow for the elucidation of the molecular-genetic mechanisms that generate SSD, an important component of understanding how SSD evolves.
Moore, H E; Butcher, J B; Adam, C D; Day, C R; Falko, P D
Cuticular hydrocarbons were extracted daily from the larvae of two closely related blowflies Calliphora vicina and Calliphora vomitoria (Diptera: Calliphoridae). The hydrocarbons were then analysed using Gas Chromatography–Mass Spectrometry (GC–MS), with the aim of observing changes within their chemical profiles in order to determine the larval age. The hydrocarbons were examined daily for each species from 1 day old larvae until pupariation. The results show significant chemical changes occ...
Singh, Devinder; Bala, Madhu
The postfeeding larval stage in blow flies is generally an irreversible condition when the fully grown third instar larvae stop feeding and give no response towards food. The larvae of most species then disperse away from their feeding medium and pupariate. There are several cases reported about the use of postfeeding larvae as forensic evidence. It is a matter of common observation that the postfeeding stage can be reached earlier than the expected time if food becomes unavailable. However, no information is available on whether postfeeding stage induced by scarcity of food is also irreversible. Similarly, the minimum period of development required by the larvae of different blow flies species to enable their survival as postfeeding larvae and pupariation in the absence of food is unknown. It was observed during the present studies that the larvae of two Chrysomya species must feed for at least 35 h at 28 degrees C in order to be capable of reaching the postfeeding stage and subsequent pupariation. Duration of the starvation period required to induce postfeeding behavior decreases with increasing age of larvae. In the case of Chrysomya megacephala, 35, 45, 55 and 65 h old larvae attained irreversible postfeeding stage after 30, 20, 12 and 2 h of starvation, respectively. Similarly, larvae of Chrysomya rufifacies that were 35, 45, 55 and 60 h old attained irreversible postfeeding stage after 25, 16, 6 and 2 h of starvation, respectively.
Yanmanee, Surasuk; Husemann, Martin; Benbow, Mark Eric; Suwannapong, Guntima
Chrysomya rufifacies represents an important indicator species in forensic entomology that is often used to estimate the minimum postmortem interval (PMImin) in crime scene investigation. However, developmental rates differ locally, so that estimates should be based on regionally generated development data. Therefore, we determined the developmental rates of C. rufifacies within its native range in Thailand under nine constant temperature regimes: 15, 18, 21, 24, 27, 30, 33, 36 and 39°C. Developmental times from egg to adult varied among the temperatures and were longest at 15°C (618h) and shortest at 33°C (168h). No pupae emerged at 39°C. We used linear regression models to estimate the minimum development threshold temperatures for each life stage: egg stage=9.5°C, first to second instar=10.8°C, second to third instar=11.5°C, third instar to pupariation=11.4°C, pupariation to adults=5.0°C; the minimum threshold to complete all larvae stages was 11.1°C and to complete all life stages from eggs to adult was 9.5°C. We further generated isomorphen and isomegalen diagrams that can be used to quickly estimate the PMImin for forensic applications. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.
Lynn M. Riddiford
Full Text Available The development of the adult optic lobe (OL of Drosophila melanogaster is directed by a wave of ingrowth of the photoreceptors over a 2-day period at the outset of metamorphosis, which is accompanied by the appearance of the pupal-specific transcription factor Broad-Z3 (Br-Z3 and expression of early drivers in OL neurons. During this time, there are pulses of ecdysteroids that time the metamorphic events. At the outset, the transient appearance of juvenile hormone (JH prevents precocious development of the OL caused by the ecdysteroid peak that initiates pupariation, but the artificial maintenance of JH after this time misdirects subsequent development. Axon ingrowth, Br-Z3 appearance and the expression of early drivers were unaffected, but aspects of later development such as the dendritic expansion of the lamina monopolar neurons and the expression of late drivers were suppressed. This effect of the exogenous JH mimic (JHM pyriproxifen is lost by 24 h after pupariation. Part of this effect of JHM is due to its suppression of the appearance of ecdysone receptor EcR-B1 that occurs after pupation and during early adult development.
Mądra-Bielewicz, Anna; Frątczak-Łagiewska, Katarzyna; Matuszewski, Szymon
Despite widely accepted standards for sampling and preservation of insect evidence, unrepresentative samples or improperly preserved evidence are encountered frequently in forensic investigations. Here, we report the results of laboratory studies on the survival of Lucilia sericata and Calliphora vomitoria (Diptera: Calliphoridae) intra-puparial forms in hermetic containers, which were stimulated by a recent case. It is demonstrated that the survival of blowfly intra-puparial forms inside airtight containers is dependent on container volume, number of puparia inside, and their age. The survival in both species was found to increase with an increase in the volume of air per 1 mg of puparium per day of development in a hermetic container. Below 0.05 ml of air, no insect survived, and above 0.2 ml of air per 1 mg of puparium per day, survival reached its maximum. These results suggest that blowflies reveal a single, general pattern of survival under decreasing oxygen conditions and that this pattern is a product of number of developing insects, their age and the initial amount of available air. Implications for forensic entomology are discussed.
Olekšáková, Tereza; Žurovcová, Martina; Klimešová, Vanda; Barták, Miroslav; Šuláková, Hana
Several methods of DNA extraction, coupled with 'DNA barcoding' species identification, were compared using specimens from early developmental stages of forensically important flies from the Calliphoridae and Sarcophagidae families. DNA was extracted at three immature stages - eggs, the first instar larvae, and empty pupal cases (puparia) - using four different extraction methods, namely, one simple 'homemade' extraction buffer protocol and three commercial kits. The extraction conditions, including the amount of proteinase K and incubation times, were optimized. The simple extraction buffer method was successful for half of the eggs and for the first instar larval samples. The DNA Lego Kit and DEP-25 DNA Extraction Kit were useful for DNA extractions from the first instar larvae samples, and the DNA Lego Kit was also successful regarding the extraction from eggs. The QIAamp DNA mini kit was the most effective; the extraction was successful with regard to all sample types - eggs, larvae, and pupari.
Møller, Morten Erik; Danielsen, Erik Thomas; Herder, Rachel
. Remarkably, our study shows that the same well-defined genetic program that stimulates a systemic downstream response to ecdysone is also utilized upstream to set the duration and amplitude of the ecdysone pulse. Activation of this switch-like mechanism ensures a rapid, self-limiting PG response......Steroid hormones trigger the onset of sexual maturation in animals by initiating genetic response programs that are determined by steroid pulse frequency, amplitude and duration. Although steroid pulses coordinate growth and timing of maturation during development, the mechanisms generating...... of hormone synthesis, the two key parameters determining pulse shape (amplitude and duration). We show that ecdysone has a positive-feedback effect on the PG, rapidly amplifying its own synthesis to trigger pupariation as the onset of maturation. During the prepupal stage, a negative-feedback signal ensures...
Voss, Sasha C; Magni, Paola; Dadour, Ian; Nansen, Christian
Forensic entomology is primarily concerned with the estimation of time since death and involves determination of the age of immature insects colonising decomposing remains. Accurate age determination of puparia is usually accomplished by dissection, which means destructive sampling of evidence. As part of improving abilities to correctly identify species and developmental age, it is highly desirable to have available non-destructive methods. In this study, we acquired external hyperspectral imaging (HSI) data (77 spectral bands, 389-892 nm) from the dorsal and ventral sides of individual puparia of two species of blowfly (Diptera: Calliphoridae), Calliphora dubia Macquart 1855 and Chrysomya rufifacies Macquart 1842. Puparia were dissected to determine the presence/absence of eight internal morphological development characteristics (legs, wings, labella, abdominal segments, antennae, thoracic bristles, orbital/facial bristles and eye colour and arista). Based on linear discriminant analysis and independent validation of HSI data, reflectance features from puparia could be used to successfully (1) distinguish the two species (classification accuracy = 92.5 %), (2) differentiate dorsal and ventral sides of puparia (classification accuracy C. dubia = 81.5 %; Ch. rufifacies = 89.2 %) and (3) predict the presence of these morphological characteristics and therefore the developmental stage of puparia (average classification accuracy using dorsal imaging: C. dubia = 90.3 %; Ch. rufifacies = 94.0 %). The analytical approach presented here provides proof of concept for a direct puparial age relationship (i.e. days since the onset of pupation) between external puparial reflectance features and internal morphological development. Furthermore, this approach establishes the potential for further refinement by using a non-invasive technique to determine the age and developmental stage of blowflies of forensic importance.
Full Text Available The circadian clock provides the temporal framework for rhythmic behavioural and metabolic functions. In the modern era of industrialization, work and social pressures, the clock function is often jeopardized, resulting in adverse and chronic effects on health. Understanding circadian clock function, particularly individual variation in diurnal phase preference (chronotype, and the molecular mechanisms underlying such chronotypes may lead to interventions that could abrogate clock dysfunction and improve human (and animal health and welfare. Our preliminary studies suggested that fruitflies, like humans, can be classified as early rising ‘larks’ or late rising ‘owls’, providing a convenient model system for these types of studies. We have identified strains of flies showing increased preference for morning emergence (Early or E from the pupal case, or more pronounced preference for evening emergence (Late or L. We have sampled pupae the day before eclosion (4th day after pupariation at 4 h intervals in the E and L strains, and examined differences in gene expression by RNAseq. We have identified differentially expressed transcripts between the E and L strains which provide candidate genes for studies of Drosophila chronotypes and their human orthologues.
Alonso, M.; Río, M.A. del; Jacas, J.
Ceratitis capitata is a quarantine pest. Spanish citrus exports to countries such as the USA or Japan are subjected to a mandatory quarantine treatment consisting of exposure of fruits to a low temperatures. Some citrus (''Fino'' lemon, ''Fortune'' mandarin) are very sensitive to this kind of treatment and can not be treated this way. Therefore, alternative treatments are necessary. In this study, high energy electrons were investigated as an alternative quarantine treatment against C. capitata in citrus. Survival of the different instars (egg to old pupae) of C. capitata reared in an artificial medium was assessed when exposed to different doses between 0 and 1 kGy. Both pupariation and adult emergence were almost prevented at 0.25 kGy, and no viable adults were obtained at 0.50 kGy. When artificially infested fruits (in both ''Fino'' lemon and ''Fortune'' mandarin) were exposed to 1 kGy, 100% mortality was obtained. Finally, quality (texture, color index, maturity index, juice yield, ethanol and acetaldehyde contents, physiological alterations and organoleptic characteristics) of irradiated (1 kGy) and non irradiated fruit were compared. High energy electron irradiation resulted in unacceptable damage to ''Fortune'' mandarin, but quality of ''Fino'' lemon resulted unaltered even when evaluated one month after irradiation. Therefore high energy electrons could be a useful alternative to cold quarantine treatment for ''Fino'' lemons. (author) [es
Nestel, D.; Nemny-Lavy, E.; Alchanatis, V.
The pattern of gas-exchange (CO 2 emission) was investigated for developing Mediterranean fruit fly (medfly) Ceratitis capitata (Wiedemann) pupae incubated at different temperatures. This study was undertaken to explore the usefulness of gas-exchange systems in the determination of physiological age in developing pupae that are mass produced for sterile insect technique projects. The rate of CO 2 emission was measured in a closed flow-through system connected to commercial infrared gas analysis equipment. Metabolic activity (rate of CO 2 emission) was related to pupal eye-color, which is the current technique used to determine physiological age. Eye-color was characterized digitally with 3 variables (Hue, Saturation and Intensity), and color separated by discriminant analysis. The rate of CO 2 emission throughout pupal development followed a U-shape, with high levels of emission during pupariation, pupal transformation and final pharate adult stages. Temperature affected the development time of pupae, but not the basic CO 2 emission patterns during development. In all temperatures, rates of CO 2 emission 1 and 2 d before adult emergence were very similar. After mid larval-adult transition (e.g., phanerocephalic pupa), digital eye-color was significantly correlated with CO 2 emission. Results support the suggestion that gas-exchange should be explored further as a system to determine pupal physiological age in mass production of fruit flies. (author) [es
Goodbrod, J R; Goff, M L
Rearing of Chrysomya megacephala (F.) and Chrysomya rufifacies (Macquart) in pure cultures at seven different population densities (larvae per gram of liver) demonstrated an inverse relationship between density and the duration of the larval stage. In pure cultures, larval mortality rates decreased with increasing density until an optimum density was reached (8 larvae/g liver for C. megacephala and 10 larvae/g liver for C. rufifacies), then decreased directly with density. Puparial and adult weights varied inversely with density for both species in pure cultures. Internal feeding mass temperatures were above ambient temperatures for all cultures, with maximum temperatures recorded in cultures with 20 and 40 larvae/g liver for G. rufifacies and C. megacephala, respectively. In paired encounters, larvae of C. rufifacies were cannibalistic and predatory on C. megacephala larvae after the first instar. In mixed cultures of these two species, the larval mortality of C. rufifacies remained relatively stable, whereas the larval mortality of C. megacephala increased directly with population density.
Musah, Rabi A.; Espinoza, Edgard O.; Cody, Robert B.; Lesiak, Ashton D.; Christensen, Earl D.; Moore, Hannah E.; Maleknia, Simin; Drijfhout, Falko P.
A high throughput method for species identification and classification through chemometric processing of direct analysis in real time (DART) mass spectrometry-derived fingerprint signatures has been developed. The method entails introduction of samples to the open air space between the DART ion source and the mass spectrometer inlet, with the entire observed mass spectral fingerprint subjected to unsupervised hierarchical clustering processing. A range of both polar and non-polar chemotypes are instantaneously detected. The result is identification and species level classification based on the entire DART-MS spectrum. Here, we illustrate how the method can be used to: (1) distinguish between endangered woods regulated by the Convention for the International Trade of Endangered Flora and Fauna (CITES) treaty; (2) assess the origin and by extension the properties of biodiesel feedstocks; (3) determine insect species from analysis of puparial casings; (4) distinguish between psychoactive plants products; and (5) differentiate between Eucalyptus species. An advantage of the hierarchical clustering approach to processing of the DART-MS derived fingerprint is that it shows both similarities and differences between species based on their chemotypes. Furthermore, full knowledge of the identities of the constituents contained within the small molecule profile of analyzed samples is not required. PMID:26156000
Yang, Yong-Qiang; Li, Xue-Bo; Shao, Ru-Yue; Lyu, Zhou; Li, Hong-Wei; Li, Gen-Ping; Xu, Lyu-Zi; Wan, Li-Hua
The characteristic life stages of infesting blowflies (Calliphoridae) such as Chrysomya megacephala (Fabricius) are powerful evidence for estimating the death time of a corpse, but an established reference of developmental times for local blowfly species is required. We determined the developmental rates of C. megacephala from southwest China at seven constant temperatures (16-34°C). Isomegalen and isomorphen diagrams were constructed based on the larval length and time for each developmental event (first ecdysis, second ecdysis, wandering, pupariation, and eclosion), at each temperature. A thermal summation model was constructed by estimating the developmental threshold temperature D0 and the thermal summation constant K. The thermal summation model indicated that, for complete development from egg hatching to eclosion, D0 = 9.07 ± 0.54°C and K = 3991.07 ± 187.26 h °C. This reference can increase the accuracy of estimations of postmortem intervals in China by predicting the growth of C. megacephala. © 2016 American Academy of Forensic Sciences.
Mazzanti, Morena; Alessandrini, Federica; Tagliabracci, Adriano; Wells, Jeffrey D; Campobasso, Carlo P
Puparial cases are common remnants of necrophagous flies in crime investigations. They usually represent the longest developmental time and, therefore, they can be very useful for the estimation of the post-mortem interval (PMI). However, before any PMI estimate, it is crucial to identify the species of fly eclosed from each puparium associated with the corpse. Morphological characteristics of the puparium are often distinctive enough to permit a species identification. But, even an accurate morphological analysis of empty puparia cannot discriminate among different species of closely related flies. Furthermore, morphological identification may be impossible if the fly puparia are poorly preserved or in fragments. This study explores the applicability of biomolecular techniques on empty puparia and their fragments for identification purposes. A total of 63 empty puparia of necrophagous Diptera resulting from forensic casework were examined. Samples were divided into three groups according to size, type and time of eclosion in order to verify whether the physical characteristics and puparia weathering can influence the amount of DNA extraction. The results suggest that a reliable genetic identification of forensically important flies may also be performed from empty puparia and/or their fragments. However, DNA degradation can deeply compromise the genetic analysis since the older the fly puparia, the smaller are the amplified fragments. 2009 Elsevier Ireland Ltd. All rights reserved.
Richards, Cameron S; Simonsen, Thomas J; Abel, Richard L; Hall, Martin J R; Schwyn, Daniel A; Wicklein, Martina
We demonstrate how micro-computed tomography (micro-CT) can be a powerful tool for describing internal and external morphological changes in Calliphora vicina (Diptera: Calliphoridae) during metamorphosis. Pupae were sampled during the 1st, 2nd, 3rd and 4th quarter of development after the onset of pupariation at 23 °C, and placed directly into 80% ethanol for preservation. In order to find the optimal contrast, four batches of pupae were treated differently: batch one was stained in 0.5M aqueous iodine for 1 day; two for 7 days; three was tagged with a radiopaque dye; four was left unstained (control). Pupae stained for 7d in iodine resulted in the best contrast micro-CT scans. The scans were of sufficiently high spatial resolution (17.2 μm) to visualise the internal morphology of developing pharate adults at all four ages. A combination of external and internal morphological characters was shown to have the potential to estimate the age of blowfly pupae with a higher degree of accuracy and precision than using external morphological characters alone. Age specific developmental characters are described. The technique could be used as a measure to estimate a minimum post-mortem interval in cases of suspicious death where pupae are the oldest stages of insect evidence collected. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.
Brock, Amanda R; Seto, Mabel; Smith-Bolton, Rachel K
Regeneration is a complex process that requires an organism to recognize and repair tissue damage, as well as grow and pattern new tissue. Here, we describe a genetic screen to identify novel regulators of regeneration. We ablated the Drosophila melanogaster larval wing primordium by inducing apoptosis in a spatially and temporally controlled manner and allowed the tissue to regenerate and repattern. To identify genes that regulate regeneration, we carried out a dominant-modifier screen by assessing the amount and quality of regeneration in adult wings heterozygous for isogenic deficiencies. We have identified 31 regions on the right arm of the third chromosome that modify the regenerative response. Interestingly, we observed several distinct phenotypes: mutants that regenerated poorly, mutants that regenerated faster or better than wild-type, and mutants that regenerated imperfectly and had patterning defects. We mapped one deficiency region to cap-n-collar ( cnc ), the Drosophila Nrf2 ortholog, which is required for regeneration. Cnc regulates reactive oxygen species levels in the regenerating epithelium, and affects c-Jun N-terminal protein kinase (JNK) signaling, growth, debris localization, and pupariation timing. Here, we present the results of our screen and propose a model wherein Cnc regulates regeneration by maintaining an optimal level of reactive oxygen species to promote JNK signaling. Copyright © 2017 by the Genetics Society of America.
Bhogal, Balpreet; Plaza-Jennings, Amara; Gavis, Elizabeth R
Dendritic arbor morphology is a key determinant of neuronal function. Once established, dendrite branching patterns must be maintained as the animal develops to ensure receptive field coverage. The translational repressors Nanos (Nos) and Pumilio (Pum) are required to maintain dendrite growth and branching of Drosophila larval class IV dendritic arborization (da) neurons, but their specific regulatory role remains unknown. We show that Nos-Pum-mediated repression of the pro-apoptotic gene head involution defective (hid) is required to maintain a balance of dendritic growth and retraction in class IV da neurons and that upregulation of hid results in decreased branching because of an increase in caspase activity. The temporal requirement for nos correlates with an ecdysone-triggered switch in sensitivity to apoptotic stimuli that occurs during the mid-L3 transition. We find that hid is required during pupariation for caspase-dependent pruning of class IV da neurons and that Nos and Pum delay pruning. Together, these results suggest that Nos and Pum provide a crucial neuroprotective regulatory layer to ensure that neurons behave appropriately in response to developmental cues. © 2016. Published by The Company of Biologists Ltd.
Li, Kaixia; Zhang, Xubo; Zuo, Ying; Liu, Weimin; Zhang, Jianzhen; Moussian, Bernard
The insect cuticle is an extracellular matrix that consists of the polysaccharide chitin, proteins, lipids and organic molecules that are arranged in distinct horizontal layers. In Drosophila melanogaster, these layers are not formed sequentially, but, at least partially, at the same time. Timing of the underlying molecular mechanisms is conceivably crucial for cuticle formation. To study this issue, we determined the time period during which the function of Knickkopf (Knk), a key factor of chitin organization, is required for wing cuticle differentiation in D. melanogaster. Although knk is expressed throughout metamorphosis, we demonstrate that its expression 30 h prior and 48 h after pupariation is essential for correct wing cuticle formation. In other words, expression beyond this period is futile. Importantly, manipulation of Knk expression during this time causes wing bending suggesting an effect of Knk amounts on the physical properties of the wing cuticle. Manipulation of Knk expression also interferes with the structure and function of the cuticle surface. First, we show that the shape of surface nano-structures depends on the expression levels of knk. Second, we find that cuticle impermeability is compromised in wings with reduced knk expression. In summary, despite the extended supply of Knk during metamorphosis, controlled amounts of Knk are important for correct wing cuticle differentiation and function in a concise period of time. Copyright © 2017 Elsevier Ltd. All rights reserved.
Miller, Daniel L; Ballard, Shannon L; Ganetzky, Barry
The Drosophila larval neuromuscular junction (NMJ) is a powerful system for the genetic and molecular analysis of neuronal excitability, synaptic transmission, and synaptic development. However, its use for studying age-dependent processes, such as maintenance of neuronal viability and synaptic stability, are temporally limited by the onset of pupariation and metamorphosis. Here we characterize larval NMJ growth, growth regulation, structure, and function in a developmental variant with an extended third instar (ETI). RNAi-knockdown of the prothoracicotropic hormone receptor, torso, in the ring gland of developing larvae leaves the timing of first and second instar molts largely unchanged, but triples duration of the third instar from 3 to 9.5 d (McBrayer et al., 2007; Rewitz et al., 2009). During this ETI period, NMJs undergo additional growth (adding >50 boutons/NMJ), and this growth remains under the control of the canonical regulators Highwire and the TGFβ/BMP pathway. NMJ growth during the ETI period occurs via addition of new branches, satellite boutons, and interstitial boutons, and continues even after muscle growth levels off. Throughout the ETI, organization of synapses and active zones remains normal, and synaptic transmission is unchanged. These results establish the ETI larval system as a viable model for studying motor neuron diseases and for investigating time-dependent effects of perturbations that impair mechanisms of neuroprotection, synaptic maintenance, and response to neural injury.
Curtis, C F [London School of Hygiene and Tropical Medicine (UK); Langley, P A [Bristol Univ. (UK)
The first laboratory and field experiments on genetic control were with tsetse flies, and they made use of the sterility in crosses and hybrids between closely related species of the Glossina morsitans complex. Backcrosses indicate that there are two separate sterility mechanisms involved: (a) genetic incompatibility between a mother and the products of alien genes in the embryo or larva in the uterus; (b) inability of hybrid males to inseminate due to incompatibility of their X-chromosomes with an alien Y or autosomes. The two largest sterile male release programmes have been in Tanzania and Upper Volta, and have used irradiation at the pupal or adult stage, respectively, for the production of sterility. Male tsetse are remarkably resistant to radiosterilization and, with the doses required to induce dominant lethals in more than 95% of sperms, premature senescence and lethargic behaviour of the males tends to result. With G. morsitans irradiated at the puparial stage these effects can be alleviated by the use of a nitrogen atmosphere during irradiation. If the puparia are then transferred to air at 11/sup 0/C for transport to the release site, immediate emergence occurs on re-warming after arrival. This advantageous procedure was used for the releases in Tanzania. In addition to dominant lethals, irradiation also produces chromosome translocations which cause inherited partial sterility. A homozygous translocation line was selected but this example did not have sufficient fitness to be used in a mass rearing programme. Chemosterilants can be applied by pupal dipping, adult contact with deposits or in aerosols. Studies are now in progress on their use in association with odour-baited traps or pheromone-baited decoys as a means of sterilizing the wild population and thus avoiding the costly and difficult process of mass rearing tsetse.
Flores, Micah; Crippen, Tawni L; Longnecker, Michael; Tomberlin, Jeffery K
Forensic entomologists often rely on development data associated with a given species to estimate when it colonized human or other vertebrate remains. In most instances, these development studies are based on single species reared in isolation in the laboratory. This study examined the impact of excretions and secretions (ES) associated with third-instar Chrysomya rufifacies (Macquart), a predator, on the development of its prey, Cochliomyia macellaria (F.). Not surprisingly, Ch. rufifacies ES did not impact the development of first- or second-instar C. macellaria, which are typically not preyed on by Ch. rufifacies. However, development of third-instar C. macellaria, which do experience predation, was impacted. First, larvae were longer than those in the control (deionized water, dH2O). Filtering the ES and removing the associated bacteria and byproducts >0.2 µm dampened the previous impact observed by the unfiltered ES on third-instar C. macellaria. Second, third-instar C. macellaria treated with unfiltered ES completed pupariation 8 h quicker than the controls. Filtering the ES lessened this effect by 50%. And finally, third-instar C. macellaria treated with filtered or unfiltered Ch. rufifacies ES reached adulthood ∼5 h faster than controls treated with dH2O. In summary, these data have large ramifications for forensic entomology, as multiple species being present on decomposing remains is not uncommon. Understanding the impact of associated ES produced by interspecific cohorts on associated development could lead to more precise estimates of the minimum postmortem interval for forensic investigation of decomposing remains. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: email@example.com.
George, Kelly A; Archer, Melanie S; Green, Lauren M; Conlan, Xavier A; Toop, Tes
Insect specimens collected from decomposing bodies enable forensic entomologists to estimate the minimum post-mortem interval (PMI). Drugs and toxins within a corpse may affect the development rate of insects that feed on them and it is vital to quantify these effects to accurately calculate minimum PMI. This study investigated the effects of morphine on growth rates of the native Australian blowfly, Calliphora stygia (Fabricius) (Diptera: Calliphoridae). Several morphine concentrations were incorporated into pet mince to simulate post-mortem concentrations in morphine, codeine and/or heroin-dosed corpses. There were four treatments for feeding larvae; T 1: control (no morphine); T 2: 2 microg/g morphine; T 3: 10 microg/g morphine; and T 4: 20 microg/g morphine. Ten replicates of 50 larvae were grown at 22 degrees C for each treatment and their development was compared at four comparison intervals; CI 1: 4-day-old larvae; CI 2: 7-day-old larvae; CI 3: pupae; and CI 4: adults. Length and width were measured for larvae and pupae, and costae and tibiae were measured for adults. Additionally, day of pupariation, day of adult eclosion, and survivorship were calculated for each replicate. The continued presence of morphine in meat was qualitatively verified using high-performance liquid chromatography with acidic potassium permanganate chemiluminescence detection. Growth rates of C. stygia fed on morphine-spiked mince did not differ significantly from those fed on control mince for any comparison interval or parameter measured. This suggests that C. stygia is a reliable model to use to accurately age a corpse containing morphine at any of the concentrations investigated.
Salazar-Souza, Mônica; Couri, Márcia S; Aguiar, Valeria M
Insects display different patterns of development, and blow flies have one of the most specialized patterns of intrapuparial development of all. In forensic entomology, pupae can be used as a tool to estimate the minimum postmortem time interval (minPMI). We analyzed the intrapuparial development of Chrysomya albiceps (Diptera: Caloricidade), whose larvae had been fed pig lungs and reared in a climate-controlled room at 28°C day/26°C night, 70 ± 10% RH, and 12 h of photophase and monitored daily. After the third-instar larvae abandoned their diet, the process of pupariation and pupation was monitored. At pre-established times, five pupae were collected, euthanized, and fixed in 5% formaldehyde, inside polypropylene test tubes with caps. Since they were the first, they were classified as 0 h pupae. Twelve collections occurred until the emergence of the adults, at 0, 2, 4, 6, 8, 10, 24, 30, 48, 54, 72, 78, 96, and 99 h (n = 84). The fixed pupae were dissected under the microscope, with the aid of anatomical tweezers and hypodermic needles, and photographed. The stages of metamorphosis and the morphological alterations occurring during the process were identified, described, and recorded before and after pupation. These phases were: pupation, larval pupal apolysis, cryptocephalic, phanerocephalic, pharate adult, emergence, and adult. The cryptophalic phase occurred between 4 and 6 h after pupation; the phanerocephalic phase between 6 and 10 h after; the pharate adult phase between 24 and 96 h after; and the imago/emergence phase 99 h after pupation.
Developmental rates of immatures of three Chrysomya species (Diptera: Calliphoridae) under the effect of methylphenidate hydrochloride, phenobarbital, and methylphenidate hydrochloride associated with phenobarbital.
Rezende, Fábio; Alonso, Marcela A; Souza, Carina M; Thyssen, Patrícia J; Linhares, Arício X
Entomotoxicology is focused on obtaining data on necrophagous entomofauna, for criminal investigations purposes. This study aimed to evaluate the effect of different concentrations of methylphenidate hydrochloride, phenobarbital, and their association on the developmental rate, larval and pupal survivorship, and the interval of emergence of adults of Chrysomya albiceps (Wiedemann), Chrysomya megacephala (Fabricius), and Chrysomya putoria (Wiedemann) (Diptera: Calliphoridae). Considering the therapeutic dose (TD) of methylphenidate hydrochloride (0.29 mg/Kg), the concentrations tested were 10× TD, 50× TD, and 100× TD. For phenobarbital, the concentrations used were 1× TD (=150 mg/Kg), 3.3× TD, and 6.7× TD. For the association of the drugs, the combinations used were 10× TD-methylphenidate hydrochloride plus 1× TD-phenobarbital, 50× TD-methylphenidate hydrochloride plus 3.3× TD-phenobarbital, and 100× TD-methylphenidate hydrochloride plus 6.7× TD-phenobarbital. The control group, without addition of drug, was maintained under the same conditions of temperature (25 ± 1 °C), humidity (70 ± 10%), and photoperiod (12 h). Specimens of each group were weighed every 12 h until pupariation. The developmental rate of the three Chrysomya species immatures was monitored. For C. albiceps the developmental time was delayed in 24 h for methylphenidate hydrochloride group and in 12 h for the phenobarbital and the drugs association groups. The effect was observed only at specific ages for C. megacephala, without altering the developmental time. For C. putoria, the developmental time was delayed in 12 h for methylphenidate hydrochloride group and in 24 h for the phenobarbital and the drugs association groups. The emergence interval was similar among all experimental groups, but larval and pupal viabilities were affected in different ways.
Chen, Li; Reece, Christian; O'Keefe, Sandra L; Hawryluk, Gregory W L; Engstrom, Monica M; Hodgetts, Ross B
During Drosophila metamorphosis, the 'early-late' genes constitute a unique class regulated by the steroid hormone 20-hydroxyecdysone. Their induction is comprised of both a primary and a secondary response to ecdysone. Previous work has suggested that the epidermal expression of the dopa decarboxylase gene (Ddc) is likely that of a typical early-late gene. Accumulation of the Ddc transcript is rapidly initiated in the absence of protein synthesis, which implies that the ecdysone receptor plays a direct role in induction. However, full Ddc expression requires the participation of one of the transcription factors encoded by the Broad-Complex. In this paper, we characterize an ecdysone response element (EcRE) that contributes to the primary response. Using gel mobility shift assays and transgenic assays, we identified a single functional EcRE, located at position -97 to -83 bp relative to the transcription initiation site. This is the first report of an EcRE associated with an early-late gene in Drosophila. Competition experiments indicated that the affinity of the Ddc EcRE for the ecdysone receptor complex was at least four-fold less than that of the canonical EcRE of the hsp27 gene. Using in vitro mutagenesis, we determined that the reduced affinity of the EcRE resided at two positions where the nucleotides differed from those found in the canonical sequence. The ecdysone receptor, acting through this EcRE, releases Ddc from a silencing mechanism, whose cis-acting domain we have mapped to the 5'-upstream region between -2067 and -1427 bp. Deletion of this repressive element resulted in precocious expression of Ddc in both epidermis and imaginal discs. Thus, epidermal Ddc induction at pupariation is under the control of an extended genomic region that contains both positive and negative regulatory elements. Copyright 2002 Elsevier Science Ireland Ltd.
Koo, Hyun-Na; Yun, Seung-Hwan; Yoon, Changmann [Department of Plant Medicine, College of Agriculture, Life and Environment Sciences, Chungbuk National University, Cheongju 361-763 (Korea, Republic of); Kim, Gil-Hah, E-mail: firstname.lastname@example.org [Department of Plant Medicine, College of Agriculture, Life and Environment Sciences, Chungbuk National University, Cheongju 361-763 (Korea, Republic of)
The American serpentine leafminer fly, Liriomyza trifolii (Burgess), is one of the most destructive polyphagous pests worldwide. In this study, we determined electron beam doses for inhibition of normal development of the leaf miner and investigated the effect of electron beam irradiation on DNA damage and p53 stability. Eggs (0-24 h old), larvae (2nd instar), puparia (0-24 h old after pupariation) and adults (24 h after emergence) were irradiated with increasing doses of electron beam irradiation (six levels between 30 and 200 Gy). At 150 Gy, the number of adults that developed from irradiated eggs, larvae and puparia was lower than in the untreated control. Fecundity and egg hatchability decreased depending on the doses applied. Reciprocal crosses between irradiated and unirradiated flies demonstrated that males were more radiotolerant than females. Adult longevity was not affected in all stages. The levels of DNA damage in L. trifolii adults were evaluated using the alkaline comet assay. Our results indicate that electron beam irradiation increased levels of DNA damage in a dose-dependent manner. Moreover, low doses of electron beam irradiation led to the rapid appearance of p53 protein within 6 h; however, it decreased after exposure to high doses (150 Gy and 200 Gy). These results suggest that electron beam irradiation induced not only abnormal development and reproduction but also p53 stability caused by DNA damage in L. trifolii. We conclude that a minimum dose of 150 Gy should be sufficient for female sterilization of L. trifolii. - Highlights: > Electron beam irradiation inhibited normal development of the leaf miner. > Electron beam irradiation inhibited normal reproduction of the leaf miner. > Electron beam irradiation increased levels of DNA damage. > Electron beam irradiation induced p53 stability.
Zd' arek, J; Cvacka, J; Sanda, M [Institute of Organic Chemistry and Biochemistry, AV CR, Prague (Czech Republic); Takac, P; Keszeliova, D; Simo, L; Roller, L [Institute of Zoology, SAV, Bratislava (Slovakia)
A problem in tsetse mass rearing facilities is the increased incidence of abortions of underdeveloped larvae or pupariation of larvae within the mother's uterus. We analysed the problem by investigating neural, hormonal and environmental factors controlling parturition. Input from the mother's brain is essential for normal parturition, since a female whose brain is disconnected by ligation fails to deposit her larva. The expulsion of a larva is stimulated by a putative parturition hormone present within the female's uterus. The hormone also elicits abortion when injected into neck ligated females at earlier stages of pregnancy. This report describes attempts to reveal the chemical nature of this hormone by purification of extracts of uteri of Glossina females and identification of behaviourally active fractions using a MALDI-MS instrument. Genomic (BLAST) analysis of the identified sequences did not reveal a significant match with any protein with bioactive properties in other species. However, similarity with various enzymes or structural proteins (and hypothetical proteins) was detected occasionally. In the Glossina genomic and cDNA databases no nucleotide sequence corresponding to the deduced AA sequences was found. Perhaps the deduced sequences are too short to obtain more significant hits both in protein and nucleotide databases. We also made investigations to elucidate environmental influences and physiological mechanisms associated with tsetse parturition. We found that the circadian rhythm of parturition of flies kept in Bratislava (G. m. morsitans, G. f. fuscipes and G. pallidipes) is less pronounced than under natural conditions. The loss of synchrony in the laboratory may have three possible causes: (i) genetic - absence of selective pressure, (ii) environmental - low intensity or absence of an entraining light or temperature stimulus, and (iii) physiological - impaired sensitivity to olfactory stimulation by a hypothetical 'oviposition' pheromone that