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The common green bottle blow fly Lucilia sericata (family, Calliphoridae) is widely used for maggot debridement therapy, which involves the application of sterile maggots to wounds. The larval excretions and secretions are important for consuming necrotic tissue and inhibiting bacterial growth in wounds of patients. Lucilia sericata is also of importance as a pest of sheep and in forensic studies to estimate a postmortem interval. Here we report the assembly of a 565.3 Mb genome from long read PacBio DNA sequencing of genomic DNA. The genome contains 14,704 predicted protein coding genes and 1709 non-coding genes. Targeted annotation and transcriptional analyses identified genes that are highly expressed in the larval salivary glands (secretions) and Malpighian tubules (excretions) under normal growth conditions and following heat stress. The genomic resources will underpin future genetic studies and in development of engineered strains for genetic control of L. sericata and for biotechnology-enhanced maggot therapy.

Self-inflicting injuries are common among special children. One of the complications is Oral Myiasis. 11 yr old spastic female reported with pain and swelling in palatal region due to trauma from wooden stick. Oral examination revealed, an ulcerated lesion measuring about 2 cm × 1 cm in relation to 11 and 12. 4 live maggots were removed from the ulcerated wound which after entomological examination was reported as Chrysomya Bezziana. Uniqueness of the case This case describes a non-invasive management of Oral myiasis in a special child with poor socioeconomic condition which demanded out-patient care which was clinically effective and affordable.

Forensic entomotoxicology has grown to impact judicial systems in developed countries. Where the use of insects and maggots as samples in death investigations as an alternative technique, especially following degradation or loss of the conventionally used samples. Carrion flies feed on dead bodies and may ingest toxic substances found in the dead body, especially when the body was poisoned before death. The knowledge of how the chemicals interact with the insect following ingestion is crucial to forensic entomotoxicologists. The study investigated the impact of dimethoate on the life cycle of four species of Calliphoridae flies, namely Chrysomya megacephala, Chrysomya saffranea, Chrysomya rufifacies and Chrysomya indiana. Various concentrations of dimethoate (1 ppm, 2 ppm, 3 ppm and 4 ppm) were utilized in the study. The rate of development of the carrion flies showed a negative correlation with the concentration of the chemical. This paper glares at the impact of the chemicals may pose to the insects, and how analysis of such impacts can guide forensic investigations of poisoning and help the investigators to solve the crime puzzle.

Wound healing is a complex process involving multiple biochemical and cellular events and represents a neglected public health issue. As a consequence, millions of people worldwide suffer from chronic wounds. The search for new treatment alternatives is therefore an important issue. In the context of wound healing, Maggot Debridement Therapy (MDT) is an inexpensive treatment with few contraindications and very promising results. This study aimed to evaluate the in vitro feasibility and implications of larvae (= MDT) use when combined with topical agents as a strategy to identify additive or synergistic combinations. The weight and survival rate of Cochliomyia macellaria (Fabricius) (Insecta, Diptera, Calliphoridae) larvae reared in an in vitro wound with either honey, hydrogel, 10% papain gel, essential fatty acids (EFA), collagenase, or silver sulfadiazine were evaluated after 24, 48 and 72 h. Hydrogel (for weight: 24 and 72 h; for survival: 24 h) and 10% papain gel (for weight: 48 h; for survival: 48 and 72 h) demonstrated the least interference in larval weight gain and survival rate, when compared to the control group. The results obtained in this study showed that the combined use of larvae and hydrogel or 10% papain gel may be promising to maximize the tissue repair. Honey, EFA and collagenase could be used to prepare the bed of the dry wounds to receive the MDT.

The Neem has wider range of medicinal properties and it has antimicrobial properties against Candida, Microsporum, Trichophyton, Geotrichum, Epidermophyton, Trichosporon etc., The Anti-bacterial properties in neem inhibits the activity of bacteria such as Salmonella typhi, Staphylococcus aureus, Streptococcus mutants, Mycobacterium tuberculosis, Vibrio cholerae, Mycobacterium pyogenes and Klebsiella pneumonia. The Anti-viral properties make it useful in the treatment of fowl pox, smallpox, chicken pox, vaccinia virus, warts, moderate inhibition of hepatitis B virus, Chikungunya, herpes virus and measles virus. The Insecticidal activity is effective against maggots, horn flies, head-lice, blowflies and biting flies. The neem is also being used by tribal community as poultice of leaves of Neem, Capparis sepiaria (Jal) and Tamarindus indica (Imli) is applied on eyes by Bhils community to cure night-blindness. The tribals of the Himalayan region use Neem as a valuable medicine as an antiseptic, used in the treatment of chickenpox. Small twigs are used as toothbrushes and as a prophylactic for mouth and teeth complaints. This popular article represents the truth of medicinal uses of Neem and the ethical value to the ecosystem.

Pyoderma gangrenosum (PG) is a neutrophilic dermatosis characterized by cutaneous ulceration. PG primarily affects young and middle-age adults with a preference for women and is often associated with an underlying systemic disease. Although its etiology is unclear, neutrophilic dysfunction, genetic risk factors, and chronic inflammation are thought to contribute to its pathogenesis. Patients with classic PG often present with a single or multiple ulcers commonly on the lower extremities or trunk. In addition to medical treatment, wound care to combat accumulation of necrotic tissue debris and biofilm formation is required for optimal management. Wound debridement often presents a challenge in PG because of its feature of pathergy, or exacerbation in response to incidental or iatrogenic trauma, which has been reported in up to 31% of cases.1 Maggot debridement therapy (MDT), a centuries-old tradition in medical history of placing larvae over a wound to digest necrotic tissue, has seen a recent resurgence in popularity.2 The technology behind this technique has evolved with controlled therapeutic myiasis in which the live larvae are enclosed within a nylon mesh bag so they are obscured from view and cannot escape.3 In this case series we review 2 cases of longstanding PG that were successfully debrided with MDT.

Postharvest fumigation of fruits and vegetables is an important tool for managing pests and diseases that can cause devastating loss if not properly controlled. Sulfur dioxide (SO2) may have promise as replacement for methyl bromide, which is expected to be phased out. However, SO2 is known to cause injury to small fruit such as table grapes (Vitis vinifera L.). We extend previous research on SO2 fumigation by focusing on an economically important fruit pest genus and through quantitative and qualitative measurements of highbush blueberry (Vaccinium corymbosum L.) fruit bleaching. This study assesses fruit damage due to SO2 fumigation at concentrations ranging 0–2.2 % (v/v) as well as the effectiveness of SO2 as a fumigant prior to cold storage for control of blueberry maggot, Rhagoletis mendax Curran. We show that fruit quality traits such as firmness, total soluble solid content, and titratable acidity are largely unaffected except at the highest SO2 concentration (2.2 %). SO2 caused bleaching and discoloration of blueberry fruit in a dose-dependent manner. Damage was also cultivar-dependent, with cv ‘Bluecrop’, and ‘Jersey’ more susceptible than ‘Draper’, ‘Elliott’, and ‘Liberty’. We show that R. mendax can be effectively controlled using a short-term (2 h), high concentration (22,000 μL L−1) SO2 fumigation followed by >14 d of cold storage at 0.5 °C. However, this treatment for control of R. mendax would likely result in damage, affecting fruit marketability.

Food waste, as a major part of the municipal solid waste has been generated increasingly worldwide. Efficient and feasible utilization of this waste material for productivity process is significant for both economical and environmental reasons. In the present study, Musca domestica larva was used as the carrier to conduct a bioconversion with food waste to get the value-added maggot protein, oil and organic fertilizers. Methods of adult flies rearing, culture medium adjuvant selection, maggot culture conditions, stocking density and the valorization of the waste have been explored. From the experimental results, every 1000 g culture mediums (700 g food waste and 300 g adjuvant) could be disposed by 1.5 g M. domestica eggs under proper culture conditions after emergence in just 4 days, 42.95 ± 0.25% of which had been consumed and the culture medium residues could be used as good organic fertilizers, accompanying with the food waste consumption, ∼53.08 g dried maggots that contained 57.06 ± 2.19% protein and 15.07 ± 2.03% oil had been produced. The maggot protein for its outstanding pharmacological activities is regarded as a good raw material in the field of medicine and animal feeding. Meanwhile, the maggot oil represents a potential alternative feedstock for biodiesel production. In our study, the maggot biodiesel was obtained after the procedure of transesterification reaction with methanol and the productivity was 87.71%.

miR-126 may increase angiogenesis in patients with diabetic foot ulcers (DFUs) treated with maggot debridement therapy (MDT).Real-time quantitative PCR was used to detect expression of miR-126 mRNA in the peripheral blood among the non-diabetic population, type 2 diabetes mellitus patients without DFU, and patients with DFUs of type 2 diabetes mellitus. The expression of miR-126 mRNA in the peripheral blood of patients with DFUs was observed before and after MDT. Finally, human umbilical vein endothelial cells (HUVEC) were utilized to explore miR-126 mRNA expression with maggot excretions/secretions (ES).In the patients with DFUs, the miR-126 mRNA expression level in the peripheral blood was less than that type 2 diabetes mellitus patients without DFU, and much lower than that in the non-diabetic population (P0.001). The miR-126 expression level was significantly increased in those DFU patients treated with MDT (P0.05). Finally, using HUVEC co-cultured with ES, we showed the ES increased miR-126 expression in vitro (P0.001).MDT upregulates the miR-126 expression in the peripheral blood of patients with DFUs.

Blowfly larvae aggregate on exposed carcasses and corpses and pass through three instars before wandering from the carcass and pupating. The developmental landmarks in this process can be used by forensic entomologists to estimate the time since the insects colonised the carcass, which sets a minimum post mortem interval. Large aggregations of feeding larvae generate a microclimate with temperatures up to 15°C above ambient conditions, which may accelerate larval development and affect forensic estimates of post-mortem intervals. This study investigated the effects of heat accumulated by maggot masses of Lucilia cuprina at aggregations of 20, 50 and 100 larvae, each at incubation temperatures of 18°C, 24°C and 30°C, using body length and life stage as developmental indicators. Aggregation temperatures reached up to 18.7°C above ambient temperature, with significant effects of both size and temperature of the aggregation on the development time of its larvae. Survivorship was highest for all life stages at 24°C, which is near the developmental optimum of L. cuprina. The results of this study provide a broadly applicable method of quantifying heat accumulation by aggregations of a wide range of species of forensic importance, and the results obtained from such studies will demonstrate that ambient temperature cannot be considered the only source of heat that blowfly larvae experience when they develop on a carcass. Neglect of temperatures within larval aggregations will result in an overestimation of post-mortem intervals and thus have far-reaching medicolegal consequences.

Use of sterile fly larvae (maggots) of blow flies for the treatment of many different types of skin and soft tissue wounds is called Maggot debridement therapy (MDT). The larvae of blow flies secrete a broad spectrum of compounds with diverse mechanisms of action in the gut and salivary glands called excretion/secretion (ES) products which showed to have antimicrobial activities against Gram negative and positive bacteria. Cutaneous leishmaniasis (CL) which is the common form of leishmaniasis is difficult to treat. In this study, the effect of ES from 2nd and 3rd stages of L. sericata and C. vicina larvae on in vitro Leishmania major amastigote growth in macrophage was evaluated. The effect of ES on Leishmania growth was estimated by assessing the rate of macrophage infection and the number of amastigotes per infected macrophages. In addition, the anti Leishmania activities of larval and ES of L. sericata and C. vicina on the skin lesion induced by L. major infection was evaluated in susceptible BALB/c mice. The results showed that ES of both flies reduced the number of infected macrophages; 2.6 and 1.5-fold using L. sericata ES and C. vicina ES, respectively, and inhibited amastigotes growth in macrophages; 2.03 and 1.36-fold by L. sericata ES and C. vicina ES, respectively as compared to the control group. The results showed that L. sericata ES was significantly more effective than C. vicina ES to inhibit in vitro L. major amastigotes growth, The size of lesion was significantly smaller in BALB/c mice treated with L. sericata ES than treated with C. vicina ES. The results of in vivo experiments suggested that pre-treatment with ES derived from L. sericata may have some protective effects on the development of L. major lesion. Therefore, it seems that maggot ES might be considered as a possible candidate for the treatment of cutaneous leishmaniasis.

Flies of the genus Cochliomyia frequently cause myiasis in the Americas during the hot and humid climate, which favors the development of the parasite's life cycle. A three-year-old female sambar deer (Rusa unicolor), originated from a conservation farm in the municipality of Casimiro de Abreu, Rio de Janeiro state, Brazil, was presented for a necropsy. Grossly, a nodular wound in the right lacrimal gland was observed, with many intralesional larvae. The histopathological evaluation of the injured lacrimal gland revealed maggots surrounded by areas of necrosis and inflammation. Specimens were collected for parasitological examination. Morphologically, larvae were identified as Cochliomyia macellaria (Diptera: Calliphoridae). It is suggested that adult C. macellaria was attracted to the site of infestation by glandular secretion. The present study shows the first report of secondary myiasis by Cochliomyia macellaria in deer.

Myiasis is the infestation of animals or humans by larvae from some species of dipteran flies. Depending on the tissues invaded, the maggots of these insects can produce different diseases of the skin, or mucoses (ocular, genitourinary, and oropharyngeal). Wohlfahrtia magnifica is one of the species causing myiasis; although it is a real veterinary problem, it rarely infests humans and extraordinarily in the context we describe. We herein present the case of a diabetic patient diagnosed with class IV peripheral vascular disease (Fontaine classification) who suffered infestation by W. magnifica and the management given to this pathologic process. The patient consented to the publication of this report.

To determine the role of maggot debridement therapy (MDT) on diabetic foot wound healing, we compared growth related factors in wounds before and after treatment. Furthermore, we utilized human umbilical vein endothelial cells (HUVECs) to explore responses to maggot excretions/secretions on markers of angiogenesis and proliferation. The results showed that there was neo-granulation and angiogenesis in diabetic foot wounds after MDT. Moreover, significant elevation in CD34 and CD68 levels was also observed in treated wounds. In vitro, ES increased HUVEC proliferation, improved tube formation, and increased expression of vascular endothelial growth factor receptor 2 in a dose dependent manner. These results demonstrate that MDT and maggot ES can promote diabetic foot wound healing by up-regulating endothelial cell activity.

International audience; Initially isolated from the alimentary canal of a Japanese corbicula clam, Oscillibacter valericigenes is a Gram-negative rod, of which culture remains very difficult. Herein we present the first case of bacteremia due to Oscillibacter valericigenes, in humans. A 55-year-old man was hospitalized for clinical management of multiple neglected leg wounds (colonized with maggots) that had occurred during a motorcycle accident. Following radiological confirmation of the bone infection, a transfemoral amputation was performed to limit the risk of extended infection. During hospitalization, before the amputation, the patient experienced fever, biological inflammation justifying the sampling of multiple blood cultures. Anaerobic blood culture was positive after 34 hours, without identification by routine procedure (MALDI-TOF), justifying identification by 16S DNA sequencing. In the absence of possible subculture, antibiotic sensitivity testing could not be performed. A pre-emptive treatment by piperacillin-tazobactam was introduced for 14 days. The evolution was good, except for a local disunion. Complete phylogenic analysis of the clinical strain showed that it significantly differed from the reference strain, which is distantly related to the Clostridia cluster IV. Due to the culture conditions and specialized identification method by sequencing, prevalence of O. valericigenes may be underestimated. Optimization of blood culture procedures and utilization of 16S rRNA gene sequencing are tools needed for identification of rare pathogens that could help to optimize clinical management of infected patients.

The high moisture content, low carbon/nitrogen ratio and higher oil & fat contents in the post-consumer food waste make it unfavourable for its application to the composting process. The present study is an attempt to develop favourable conditions for rapid composting of post-consumer food wastes through a novel combination of garbage enzyme and maggots’ larvae (0.04 g of maggots per gram of waste) in different combination of food waste with wheat straw and biomass waste, respectively. The in-vessel composting experiments were conducted in three polystyrene vessels out of which the Vessel 1 contained only food waste, Vessels 2 and 3 contained food waste with wheat straw and biomass wastes in the ratio 3:2, respectively. Indigenously prepared garbage enzyme and the isolated maggots’ larvae were introduced in the same amount in all the vessels to observe its effect on the composting process. The co-composting of these combinations was carried out and its comparative analysis was also performed. The maximum organic matter degradation of 77.72 ± 0.11% was found in Vessel 2 with a ‘k’ value of 0.15 d−1, while, the organic matter degradation of 76.98 ± 0.01% and 76.21 ± 0.07% with a ‘k’ value of 0.12 d−1 and 0.1 d−1 was observed in Vessels 1 and 3, respectively. A significant reduction in heavy metals along with high bioaccumulation factors was detected in maggots’ larvae. It is comprehended that applying the requisite combination of maggots’ larvae and garbage enzyme can prove to be a feasible solution for rapid and efficient composting of post-consumer food wastes with a different combination of agricultural and biomass wastes.

Decomposition of carrion is a complex ecological process wherein nutrients and energy are recycled in the environment. After feeding but prior to pupation, blow fly larvae disperse from decomposing carrion, presumably to escape unfavorable microsite conditions or perhaps predation. By leaving the immediate area, larvae are effectively transporting carrion-derived nutrients throughout the ecosystem. This dispersal is generally characterized as individual-based and moving randomly from the carcass in every direction, however, non-random, en masse maggot dispersal has been documented at least twice. Here, we report another incidence of en masse blow fly larval dispersal traveling due east in a small column. At approximately 2 m, the column stopped its unidirectional path and dispersed in a radial, 360° pattern. A subsample of 5 larvae was collected, returned to the lab, and identified as Lucilia coeruleiviridis, a common blow fly often involved in carrion decomposition in nature. These observations contribute to an increasing body of evidence that migrating fly larvae do not always disperse randomly, which may have important implications for the dispersal of carrion-derived nutrients as well as interactions with blow fly predators.

Onion maggot, Delia antiqua (Meigen), and seedcorn maggot, Delia platura (Meigen), are important pests of spring-sown onions, Allium cepa L. Larvae of both species feed on developing epicotyls and roots of young onion plants often resulting in seedling mortality. Cultural controls used in combination with the insecticide chlorpyrifos are currently the standard practice for maggot control in the western USA. However, cultural controls are only partially effective and reliance on chlorpyrifos has several potential problems including future availability and development of resistance. Insecticides including clothianidin, imidacloprid, spinosad, and thiamethoxam were evaluated in California, USA in 2011–2013 to identify efficacious alternatives to chlorpyrifos. Some insecticides were applied in multiple ways including seed treatment, in-furrow application at planting, and rototiller incorporation prior to planting. Onion plant population, vigor, and yield were measured to assess insecticide efficacy. Maggots reduced onion plant populations by more than 65% of the seeding rate in the untreated controls. Seed treatments with spinosad or clothianidin + imidacloprid were the best alternative to chlorpyrifos for minimizing onion mortality from maggot feeding. Onions treated with both seed treatments had similar or higher plant populations and bulb yields compared to chlorpyrifos. The efficacy of spinosad was greatly improved when applied as a seed treatment compared to an in-furrow application at planting or when incorporated into the soil with a rototiller prior to planting. Spinosad seed treatment increased onion plant populations by 256%, 76%, and 853% compared to untreated controls in 2011, 2012, and 2013, respectively. Conversely, in-furrow and rototill-incorporated applications of spinosad were similar to the untreated control in terms of onion plant population and yield. Seed treatments with newer chemistries could provide an efficacious alternative to chlorpyrifos for protecting onions from maggot damage in western onion production systems.

The hairy maggot blow fly, Chrysomya rufifacies (Diptera: Calliphoridae), is a forensically important fly often encountered on human and other vertebrate remains in temperate and tropic regions throughout the world including Australia, Asia, Central America and North America. C. rufifacies was reared under controlled laboratory conditions on three muscle types (i.e., porcine, equine and canine) at three temperatures (i.e., 20.8, 24.8 and 28.3°C). Rate of larval weight gain across time was statistically significant between muscle types (P≤0.0001) and approaching significance across time between temperatures (P=0.0511). This research represents the first development study for C. rufifacies from central Texas, USA and the first study to examine the impact of tissue type on its development. Furthermore, these data, when compared to those available in the literature, indicate developmental differences that could be due to genetic differences in populations or possibly methods employed during the studies. Caution should be emphasized when applying development data for this species from one region to forensic investigations in other ecoregions as such differences in development based on tissue fed upon by larvae, population genetics, and methodologies used in the studies could represent error in estimating the time of colonization.

Intersexual selection drives the evolution of reproductive traits, including ornamentation and courtship behaviour. However, most studies attempting to explain the relationships between reproductive traits are oversimplified because traits are evaluated independently, and assessment of mate choice is restricted to one sex. Further research is required to understand the extent to which interactions between traits influence mating decisions in both sexes. The small hairy maggot blowfly, Chrysomya varipes, offers an ideal opportunity to examine how interactions between foreleg ornamentation (males have conspicuous white and black hairs, whereas females have only black hairs) and complex courtship behaviour influence mating success in males and females. This study involved mate choice trials between single females and males that varied in foreleg ornamentation. Courtship investment was quantified for males, and ornamentation and body size were quantified for both sexes. Mating success was predicted by time invested in courtship, but not by either male and female ornamentation or body size alone. However, there was a significant interaction between courtship investment, ornamentation and body size. At any given courtship investment, males with more ornamentation had an increased probability of mating success. Females with more ornamentation also had an increased probability of mating success, because males invested more courtship in more ornamented females. Furthermore, the lightest males had an increased probability of mating success compared to the heaviest males, as lighter males invested more in courtship. Female weight, however, did not significantly affect the probability of mating. The results suggest that male foreleg ornamentation in C. varipes acts as an amplifier of the visual courtship display, adding to a small but growing body of evidence that multiple male and female traits can interact to influence mating decisions.

SUMMARY Antimicrobial peptides (AMP) are important components of the host innate immune response, as they exert broad-spectrum antimicrobial activities against pathogenic microbes. The AMP allow housefly larva (maggots) to live in harsh environments filled with pathogenic bacteria. In this study, maggot AMP were induced by incubation with inactivated Salmonella pullorum and crudely extracted. The concentration and antimicrobial activity of the maggot AMP were then measured. In bird experiments, chickens were artificially infected with S. pullorum, and the maggot AMP extracts were used to treat the infected chickens. The expression level of AMP was significantly enhanced by S. pullorum stimulation, and the antibiotic activity of the S. pullorum-induced AMP was significantly stronger than that of the noninduced AMP, especially against S. pullorum. In the bird experiments, based on survival rate, blood indicators, and intestinal bacterial changes, maggot AMP and antibiotics were successful in treating the S. pullorum-infected chickens. In conclusion, AMP have the potential for further development as a convenient, alternative antibiotic strategy to reduce the use of antibiotics and disease resistance.

The temporal incidence of cabbage maggot, Delia radicum L. was investigated using three exclusion cage experiments, one each during spring, summer and fall, in broccoli fields as well as two surveys, one each in three broccoli fields, and sixteen turnip plantings in central coast of California. In the cage experiments, sets of broccoli plants were exposed to natural populations of D. radicum flies for w14-d periods after plant emergence throughout the growing season. For the surveys, soil, root samples, and yellow sticky traps were collected every week from broccoli fields to determine number of eggs, maggots, feeding-injury and adults. Only roots were sampled from turnip plantings to determine feeding-injury. In all three cage experiments, feeding injury from D. radicum maggot was less during the first 14-d than w15e28 d after plant emergence (DAE). In the summer and fall, feeding injury by D. radicum was less during 29e42 and 43e56 DAE than it was 15e28 DAE. In the survey of broccoli fields, a greater number of D. radicum eggs were detected starting the fourth week after planting (WAP). Similarly, an increase in number of D. radicum maggots and feeding injury was observed at fifth and sixth WAP, respectively. However, adults were abundant throughout the growing period. In the turnip survey, increase in injury from D. radicum feeding did not appear until the fifth WAP. Overall, these studies indicate that increased incidence of D. radicum was delayed by about two to three weeks after plant emergence. The implications of these results for timing of insecticide application for D. radicum in the central coast of California are discussed.

It is of fundamental importance in forensic entomology that the factors controlling carcass temperatures during decomposition are thoroughly understood. The thermal environment to which fly larvae are exposed is the primary influence on their growth rate, and hence affects any estimate of minimum time since death using such specimens in homicide investigations. To date, much of the entomological research on maggot masses has focused on their elevation of carcass temperatures, with very little focus on the bacteria associated with larval activity. The aim of this study was to investigate the heat associated with decay and the types of bacteria present during the decomposition of a carcass, both in the presence and in the absence of maggots. Three treatments were imposed: fresh, frozen and maggot-infested, each consisting of five replicate pig carcasses. Temperature measurements and bacterial swabs were taken from the gastro-intestinal region of each pig and temperatures and bacterial communities compared between treatments. All carcasses reached average maximum temperatures above 32 °C in a temperature controlled room set at 23 °C. Treatment had no statistically significant effect on the temperatures recorded in each carcass but did significantly affect the community structure of the bacteria. However, bacterial community structure varied across time. This study suggests that bacterial metabolism plays a significant role in carcass thermogenesis, and that maggot masses, while contributing to localised heating within the carcass, may have less of a role in elevating carcass temperatures than previously assumed.

The impact of maggot mass size on body water content, net transpiration rate, and dehydration tolerance of fly pupae was examined in six species of necrophagous flies. Species that spent more time on food as larvae, produced pupae with high body water contents. Dehydration tolerance limits of pupae were modest, matching the moisture-rich conditions of decaying carrion for larvae. Protophormia terraenovae pupariates on food as it dries, and this was reflected by pupae having the highest body water content and lowest net transpiration rate. Megaselia scalaris featured the lowest body water content and highest dehydration tolerance, implying that this species is arid-suited, which matches its ability to feed and colonize on post-decay carrion. Lucilia illustris was the most sensitive to larval overcrowding, resulting in a dramatic decrease in pupal size, early dispersal from food, fed less and had fast net transpiration rates. By contrast, Lucilia sericata was the most resistant, by showing no pupal size decrease and no change in net transpiration rate. Other species were between these extremes, requiring larger maggot mass sizes to produce the effect of decreasing pupal size and increasing net transpiration rate. We conclude: (1) the pupa’s response to overcrowding and water balance profile are species-specific, varying according to pupal size and net transpiration rate as independent characteristics; (2) water balance profile of the pupae reflects the behavior and microhabitat of the larva; and (3) danger of lethal desiccation to smaller-sized pupae is circumvented by a faster developmental rate rather than enhanced water conservation.

To gain a better understanding of the molecular mechanisms regulating pupal diapause of the onion maggot Delia antiqua, PCR-based suppressive subtractive hybridization was performed to identify genes involved in summer and/or winter diapause. A total of 209 unique sequences were obtained including 89 in forward library for winter diapausing pupae and 120 in the reverse library for summer diapausing pupae. 76.4% (68/89) and 68.3% (82/120) unique sequences had significant hits to non-redundant proteins database. Gene functional annotation showed these non-redundant sequences are involved in stress response and innate immunity, metabolism and energy, information processing and regulation, binding, food storage, morphogenesis and development, cell skeleton and cycle, protein synthesis and folding. Approximately 28.2% (59/209) transcripts showed no significant similarity to any other sequence in the public databases, probably representing unique genes of the onion maggot. Semi-quantitative RT-PCR revealed that the relative expression levels of 18 genes were comparable between summer and winter diapause. This study elucidates the temporal expression of diapause-related genes in onion maggot, also provides new insights into the differences in the physiological changes in summer and winter pupae. Functional characterization of some candidate genes will further enhance the understanding of the generating, maintaining, and breaking mechanism of diapause.

Myiasis is defined as an infestation of the organs and/or tissues of human and other animals by fly maggots. Fly species that normally breed in meat or carrion (Diptera: Calliphoridae, Sarcophagidae) may become involved in cutaneous myiasis by colonizing preexisting wounds. Reports of human wound myiasis contracted in hospitals and nursing homes, especially when patients are chronically ill or bed-ridden, are not uncommon across North America and often result in cases of neglect and civil litigation. Based on a case history dealing with this latter situation and circumstances surrounding the treatment of maggot infestation, we designed an experiment to assess the effectiveness of wound cleansing solutions on maggot mortality. Treatments, consisting of four commonly used cleaning solutions (isopropyl alcohol, Dakin's solution, iodine, and hydrogen peroxide) and a control (deionized water), were applied to experimental units (n=5), with each unit consisting of groups of actively feeding Lucilia sericata maggots (Diptera: Calliphoridae). Every 24h, treatments were applied and mortality was assessed for the duration of the study (14 days). Total mean mortality increased over the duration of the experiment, with an initial large increase (10-25%) after the first treatment application, followed by a gradual increase over the remainder of the study. General differences among treatments indicated greatest mean total mortality for Dakin's solution (sodium hypochlorite) (46%), followed by isopropyl alcohol (42%), Betadine (37%), hydrogen peroxide (33%) and lowest mortality for the control (25%); however, no statistically significant differences were observed among treatments and no treatment resulted in 100% maggot mortality. Traditional wound cleansing solutions may not be sufficient for maggot infestations of pre-existing wounds and supplemental treatments may be necessary to effectively treat cases of wound myiasis.

Although oral myiasis is a rare condition in non-tropical countries comparing to those in tropical zone, the manifestation of myiasis could not be overlooked, especially among oral cancer patients who are unable to keep their mouth closed and maintain good oral hygiene. The diagnosis of myiasis is confirmed clinically by detecting the maggots and traditional management of mechanical removal of the larvae by using turpentine oil has been observed. The houseflies may carry larvae (maggots) and lay them down on exposed wounds. In this case an intruding oral cancer spreading through facial skin not covered with a surgical dressing was infested into larvae, and caused myiasis. Local application of turpentine oil was utilized to eliminate maggots but the procedure carries a risk of toxicity.

Research to quantify the potency of larval excretion/secretion from Lucilia sericata using liquid culture assays has produced contradictory results. In this study, viable counting was used to investigate the effectiveness of excretion/secretion against three marker bacterial species (Pseudomonas aeruginosa, Staphylococcus aureus, and Escherichia coli) and the effects of varying growing conditions in assays. Results demonstrate that factors such as number of larvae, species of bacteria and addition of nutrient influence its antibacterial potency. Therefore a standardised method should be employed for liquid culture assays when investigating the antibacterial activity of larval excretion/secretion from L. sericata.

Maggot debridement therapy is a nontraumatic, minimally invasive method to treat infections in a foot compromised by chronic laminitis. A mechanical strategy must first be in place to address the instability of the distal phalanx and hoof capsule. Adverse reactions to maggot debridement therapy are uncommon and the only side effect observed has been irritation or hypersensitivity at the site. Chronic laminitic cases of sepsis/necrosis within the hoof benefit from this procedure due to the noninvasive, continuous debridement and healing properties provided by the larvae.

Accidental oral myiasis are caused by ingestion of fly's eggs or direct oviposition over a compromised area of the host. Here the authors report 2 cases of accidental oral myiasis caused by direct oviposition over an edentulous area in the first patient and in the periodontium in the second patient. Both cases were treated successfully with topical applications of nitrofurazone during a 3-day period. No surgical procedures were required. Flushing the wound with nitrofurazone caused the maggots to endure a liquid-filled anaerobic environment. Healing was uneventful in the 2 cases presented and no adverse reactions were observed during the treatment after 2 months of follow-up. The maggots of both patients were identified as Cochliomyia hominivorax.

We investigated the time taken by blowflies to find and oviposit on fresh carcasses placed outdoors and indoors. Paired dead piglets, one in the open and the other in a nearby room (on the first floor of an occupied, detached, suburban house near Cologne, Germany, with a window opened 9 cm) were exposed simultaneously on nine occasions. The species visiting both locations and the number of egg batches deposited by blowflies between both locations were monitored 2, 8, 24 and 48 h after exposure. In all cases the indoor piglet carcass was exclusively infested by Calliphora vicina; only in one case, on a very hot day after a 48-h exposure did Lucilia sericata infest an indoor carcass. The outdoor piglets were infested by a variety of common corpse-visiting species: L. sericata, L. caesar, L. illustris, C. vicina and C. vomitoria. A significant difference in the number of egg batches was detected between indoors and outdoors. Furthermore, in only two of nine runs did oviposition occur within the first 24 h of exposure indoors. Ambient temperature, daylength and rainfall had no significant effect on the number of egg batches. Moreover, we observed fewer larvae on indoor piglets, too few to form maggot masses. This might result in slower larval development than in the case of outdoor piglets. We conclude that post-mortem interval (PMI) estimation for corpses found indoors must be handled carefully as oviposition might have taken place with a delay up to 24 h.

This paper describes two cases of wounds with different aetiology and dissimilar size which were treated with Larval Debridement Therapy. Two different techniques of Larval Debridement were used for each case: Free Range and BioFoam larvae. Observations and treatment were carried out in a Primary Health Centre in Barcelona, Spain. The application and observations were performed by two nurses with the collaboration of two family physicians. The Larval Therapy efficacy outcome was checked with an Ulcer Reverse Stage Scale (PUSH v2), by taking photographs and a Visual Analogue Pain Scale (VAPS) was completed by both subjects before and after the treatment. It was observed in both cases that the larval therapy caused debridement of all fibrin and collagen tissue, and the slough dropped to a normal level. As a consequence of this change the PUSH scale punctuations were improved. On the negative side, the therapy was associated with an increase in pain. The beneficial effect of applying Larval Therapy enables following a basic wound healing care plan. The larval debridement therapy did not pose any threat for the patients; they could go about their Activities of the Daily Life (ADL) without any change. These wound cases treated with larval debridement therapy are the first ones reported in the Primary Health Care field in Spain.

It has been known for centuries that application of larvae is useful to heal certain wounds by facilitating debridement of necrotic tissue. Their therapeutic use was popularised in the beginning of the 19th century, but waned in the 1940s with the advent of antiseptic wound management and antibiotics. In more recent years, larvae are once again in vogue for management of difficult wounds. The mechanism of wound debridement by larvae includes the complete wound by continuous larval motion, secretion of proteolytic enzymes and antibacterial substances, effects on epidermal growth factor and interleukin-6 (IL-6) and ingestion and digestion of bacteria and necrotic tissue. In our study, wound debridement was achieved satisfactorily in 29 of 34 patients (85%) with chronic wounds. In the remaining five patients, failures occurred due to inadequate sealing in two patients (6%), death of larvae in two patients (6%) and treatment intolerance in one patient (3%). Larval therapy should be considered as a therapeutic option in the management of certain difficult wounds.

The term myiasis is applied to the injurious action that larvae of certain diptera cause to the organism of vertebrate animals in the living or dead tissue in which they grow. Because of its great destructive potential, appropriate and preventative treatment are necessary. Among the sites of infestation, the human mouth is a common site, mainly in tropical countries. We present two cases of oral myiasis caused by Cochliomyia hominivorax spp. Ivermectin is an extremely effective semi-synthetic macrolides, in the treatment of this condition.

Maggot-therapy is the application of disinfected fly larvae to chronic wounds to debride the wound bed of necrotic tissue, reduce bacterial contamination and enhance the formation of healthy granulation tissue. Interest in the use of maggot-therapy in human medicine is growing as a result of the prevalence of antibiotic resistant bacteria. Maggot therapy, however, is used relatively little in veterinary medicine. Nevertheless, concern over antibiotic resistance and the increase in demand for organic husbandry and residue-free meat and milk, suggest that it is an option which merits further consideration.

Behavioural preferences of Drosophila larvae are often measured in a simple ‘plate test’, in which groups of larvae are allowed to migrate on an agar surface in response to variation in a stimulus. We investigated the behaviour of individual larvae in an olfactory test, alone and in groups. Using mixed groups of responsive and nonresponsive larvae, we showed that, although larvae behaved as individuals, the strength of their responses was affected by the presence of other larvae. This effect was apparently due to larvae bumping into each other, which reduced the distance they travel in the test. Larval olfactory responses were not affected by starvation: there were no significant differences in the responses of starved and not-starved larvae up to a duration of 5 h without food. However, food deprivation did significantly affect the movement of larvae, with more starved larvae leaving the start zone. We found significant differences in the olfactory responses of maggots of different ages, with responses increasing in strength with increased age but declining at the oldest age tested (late third instar). Finally, we found evidence that suggests that larvae may show a circadian modulation of the strength of their olfactory response, with a significant increase over the light:dark cycle. This response was maintained when wild-type larvae were shifted to continuous darkness but not in arrhythmic period0 mutant larvae. We hypothesize that this apparent effect is due to a central modulation of locomotor behaviour rather than to a specific effect on olfaction.

To test the hypothesis that long-term survival of sugarbeet root maggot in storage is facilitated by larvae undergoing prolonged diapause, respiration and gene expression patterns of field-collected diapausing larvae were compared with those of 1-, 2-, and 5-year laboratory-stored larvae. Additional assessments were made on post-storage survival, emergence, and reproductive fitness of stored larvae. Respirometry, carried out at 5 and 20 degrees C revealed no differences among respiration rates of initially diapausing and long-term stored larvae. A 15 degrees increase in temperature elevated respiration in both diapausing and stored larvae, with levels of CO2 release ranging between 8- and 14-fold higher at 20 degrees C than at 5 degrees C. Similarly, 6-10-fold increases in O2 consumption levels were observed at the higher temperature. A transcript with sequence similarity to the fat body protein 2 (Fbp2) gene was highly expressed in diapausing larvae, and trace levels were expressed in some samples of 1-year stored larvae. However, no expression was detected in 2- and 5-year stored larvae. Survival and emergence studies of stored larvae revealed mixed populations of diapausing (i.e., the 5-17% of larvae that did not pupate) and post-diapausing (62-84% of larvae pupated) insects, with a high incidence of pupation (62%) and emergence (47%), even after 4 years in cold storage. Therefore, extended survival of Tetanops myopaeformis larvae in long-term cold storage is facilitated by two mechanisms, with a majority of larvae in post-diapause quiescence and a smaller fraction in a state of prolonged diapause.

117:1134, 1977 54. Scholten T, Brunsdon D, Rogan W, Richards R: Three unusual cases of parasitic infections. Can Med Assoc J 117:368, 1977 55. McIver SB, Dutta PC, Freeman RS: Cutaneous myiasis by Dermatobia hominis. Can Med Assoc J 104:771, 1971 56. Corrigan SH, Corrigan CE: Three cases of creeping myiasis in Saskatchewan. Can Med Assoc J 15:403, 1925 57. Salt RW: Insects of the season 1941 in southern Alberta. Can Insect Pest Rev 20:181, 1942 58. Judd WW: Intestinal myiasis of a human caused by maggots of the flesh fly, Sarcophaga haemorrhoidalis (Fln.) (Diptera: Sarcophagidae), in Southwestern Ontario. Can Entomol 88:117, 1956 59. Ali-Khan FEA, Ali-Khan Z: Two cases of human Sarcophaga (Diptera: Sarcophagidae) myiasis in Quebec, with descriptions of the larvae. Can J Zool 52:643, 1974 60. Curtis LC: A case of aural myiasis in man. Can J Public Health 47:298, 1956 61. Israels S, Shuman B: Myiasis dermatosa in infancy. Pediatrics 4:665, 1949 62. Seamans HL: Insects of the season 1941 in southern Alberta. Can Insect Pest Rev 20:91, 1942 63. Brady MJ: Cutaneous myiasis. Arch Pediatr 40:638, 1923 64. Lewis L: Cutaneous myiasis occurring in western Canada. Can Med Assoc J 56:319, 1947 65. Walker EM: Wohlfahrtia vigil (Walker) as a human parasite (Diptera: Sarcophagidae). J Parasitol 7:1, 1920 66. Hunt BR, Edwards P: Dermal myiasis in eastern Canada. Can Med Assoc J 64:69, 1951 67. Chown G: Report of a case of cutaneous myiasis in an infant, Wohlfahrtia vigil (Walker) infection. Can Med Assoc J 14:967, 1924 68. Croll NA, W GT, Brown CA: A case of Wohlfahrtia vigil from Prince Edward Island. Can Dis Wkly Rep 2:190, 1976 69. Croll NA, W GT, Faubert GM: A case of Eristalis tenax myiasis in Quebec. Can Dis Wkly Rep 2:175, 1976

Studies were conducted in 2002 and 2003 to evaluate the phenology of the apple maggot, Rhagoletis pomonella (Walsh), in Henderson County, NC, and to compare the efficacy of various products for protection of fruit from maggot infestation. The period of peak trap capture on red spheres differed among orchards, with two orchards having peak captures in early June, and in an abandoned orchard peak capture occurred in late July–September. Surround WP, a particle film kaolin clay product, was evaluated for management of apple maggot at 28.0 kg/ha at two spray volumes (935 and 1870 L/ha) and two application intervals (7- and 14-d). The percentage of apple maggot-infested fruit in Surround treatments, regardless of the spray volume evaluated, was equivalent to that of azinphosmethyl and spinosad treatments. Application of Surround with an airblast sprayer resulted in heterogeneous coverage of fruit within the tree canopy, with higher deposition on fruit on the periphery of trees compared with those in the inner or upper canopy. In choice tests with apple maggot populations in an abandoned orchard, fewer flies alighted on Surround-treated foliage or fruit compared with the water controls, suggesting that the whitish color of Surround-treated trees interfered with visual cues used by flies to locate hosts. In an abandoned orchard with >60% of non-treated fruit infested with apple maggot, the chloronicotinyl compounds imidacloprid (10.0%) and thiacloprid (5.3%) resulted in comparable levels of control to that of azinphosmethyl (8.0%), while spinosad, indoxacarb, and thiamethoxam all had >33% infested fruit. These studies demonstrated that Surround and a number of chloronicotinyl insecticides may be alternatives to azinphosmethyl for management of apple maggot in North Carolina.

Hoof disease and injuries are common and serious problems for equines. Maggot debridement therapy (MDT) has been used to treat problematic wounds in humans, but has been used only rarely in other animals. US veterinarians who had employed MDT were surveyed to investigate their reasons for the choice of this treatment and their clinical experiences with it. Between 1997 and 2003, 13 horses were treated by eight veterinarians who used MDT to control infection or debride wounds, which could not easily be reached surgically or were not responding to conventional therapy. Seven animals were lame, and six were expected to require euthanasia. Following maggot therapy, all infections were eradicated or controlled, and only one horse had to be euthanased. No adverse events were attributed to maggot therapy for any of these cases, other than presumed discomfort during therapy. The data collected suggest that maggot therapy could be useful for treating some serious equine hoof and leg wounds.

In Thailand, the hairy maggots of the blow flies, Chrysomya rufifacies (Macquart) and Chrysomya villeneuvi Patton, are of forensic importance. Both flies are closely related species, not only in the morphological appearance of their larvae and puparia, but also on the aggressive feeding habit of the former. In our continuing studies of forensically important flies, identification of immature ones needs particular attention. In this study, we reported the morphological comparison between the puparia of these two blow fly species using scanning electron microscopy (SEM). Observation revealed that the cuticular sculpture of tubercles along the dorsal and lateral segments had markedly different features: with C. rufifacies having many sharp spines assembling only at the tip, while of C. villeneuvi bore stout spines throughout the tubercle. A larger number of globules at the bubble membrane on the dorsolateral border of the fifth segment was found in C. villeneuvi (average 225) than in C. rufifacies (average 35), and more papillae was observed on the anterior spiracle in C. villeneuvi (13-15) than in C. rufifacies (9-12). However, the morphology of distinct net-like patches of the integument and structure of the posterior spiracle of both species were almost identical. Morphological comparison in this study permitted identification of the puparia of both fly species, particularly in areas where they co-exist.

Management of onion maggot, Delia antiqua (Meigen), in onion requires the use of an insecticide applied at planting. Insecticide resistance and a dearth of available products have stimulated an effort to identify new insecticides for onion maggot control, especially chemistries that can be delivered as seed treatments. Onion seeds film-coated with fipronil, spinosad, clothianidin or thiamethoxam were evaluated for onion maggot control in muck soils located in two major onion-growing regions in New York, USA in 2001–2003. Fipronil, spinosad and clothianidin at rates of 25, 25 and 50 g (a.i.)/kg of seed, respectively, consistently provided excellent control of onion maggot (⩽5% seedling loss). Moreover, these seed treatments performed well during wet and dry seasons. In contrast, thiamethoxam at a rate of 50 g (a.i.)/kg of seed failed to control onion maggot in three of the five experiments. Cyromazine seed treatment, which is currently used by onion growers, provided an equivalent level of onion maggot control to fipronil, spinosad and clothianidin. The other commonly used treatment, chlorpyrifos in-furrow drench, performed poorly in three of the four experiments, resulting in an average seedling loss of 17%. Insecticide resistance was implicated as the reason for the failure in two of the trials where seedling loss averaged 24%. Seed treatments provide growers with a convenient, reliable and environmentally responsible method for managing economically damaging onion maggot infestations, including those that are resistant to chlorpyrifos. Fipronil, spinosad and clothianidin seed treatments should be considered for future registration on onion. Availability of these chemistries would allow rotation with the cyromazine seed treatment, which would likely prolong the longevity of all products.

The uzi fly, Blepharipa zebina, is a well-known larval endoparasitoid of the tropical tasar silkworm, Antheraea mylitta. The present study dealt with the effect of the number of maggots developing per host on host nutritional parameters, parasitoid development and reproduction. Nutritional indices for ingestion, digestion, approximate digestibility, relative consumption rate, relative growth rate, and gain in body weight declined significantly with the increase in parasitoid burden, but the efficiency of conversion of digested food recorded a significant increase. The efficiency of conversion of ingested food remained little affected. The developmental period was significantly extended in larvae parasitized with 5 and 10 maggots per larva (mpl). Cocoon shell weight decreased by 27-63.5% in parasitized groups (1, 2, and 5 mpl) while larvae parasitized with 10 mpl could not spin cocoons. The maggot development period, recovery percentage, and fecundity of the uzi fly declined significantly with the increase in number of maggots developing per host.

In the literature maggot therapy is discussed as a promising and potent form of debridement therapy. The number of maggots needed to debride a wound is estimated at 10 per cm 2 , and more in case of a higher percentage of necrosis or slough. In the authors' hospital, from March 1999 to May 2002, 16 patients were successfully treated with maggot therapy. The average maggot treatment time was 27 days, with an average of seven maggot changes. Most patients were treated for osteomyelitis, with trauma being the leading aetiological factor. In accordance with in-vitro findings, maggot therapy was found to be more effective in Gram-positive infected wounds. Gram-negative bacteria are cultured more often after maggot treatment than before it (p=0.001). The opposite effect was found for Gram-positive infected wounds (non-significant p=0.07). In vivo maggots seem to be less effective against Gram-negative infected wounds. The authors believe that a higher number of maggots is needed not only for a larger wound or a wound with a higher percentage covered with slough, but also for a wound infected with Gram-negative bacteria.

Green fluorescent protein-labeled Escherichia coli O157:H7 and Salmonella enterica serovar Enteritidis were inoculated at 10(7) CFU/g into cow, hog, or chicken manure. Ten- or 11-day-old soldier fly larvae (Hermetia illucens L.) (7 to 10 g) were added to the manure and held at 23, 27, or 32 degrees C for 3 to 6 days. Soldier fly larvae accelerated inactivation of E. coli O157:H7 in chicken manure but had no effect in cow manure and enhanced survival in hog manure. The initial pH values of the hog and chicken manure were 6.0 to 6.2 and 7.4 to 8.2, respectively, and it is surmised that these conditions affected the stability of the larval antimicrobial system. Reductions of E. coli O157:H7 populations in chicken manure by larvae were affected by storage temperature, with greater reductions in samples held for 3 days at 27 or 32 degrees C than at 23 degrees C. Pathogen inactivation in chicken manure by larvae was not affected by the indigenous microflora of chicken manure, because Salmonella Enteritidis populations in larvae-treated samples were approximately 2.5 log lower than control samples without larvae when either autoclaved or nonautoclaved chicken manure was used as the contaminated medium during 3 days of storage. Extending the storage time to 6 days, larvae again accelerated the reduction in Salmonella Enteritidis populations in chicken manure during the first 4 days of storage; however, larvae became contaminated with the pathogen. After 2 days of feeding on contaminated manure, Salmonella Enteritidis populations in larvae averaged 3.3 log CFU/g. Populations decreased to 1.9 log CFU/g after 6 days of exposure to contaminated chicken manure; however, the absence of feeding activity by the maggots in later stages of storage may be responsible for the continued presence of Salmonella Enteritidis in larvae. Transfer of contaminated larvae to fresh chicken manure restored feeding activity but led to cross-contamination of the fresh manure.

Summary The presence of maggots (fly larvae) in an open wound is a repelling sight to many and documented cases of myiasis in the literature are scant due to the rarity of such infestation in live patients. A unique case of such a presentation is elaborated in a patient who sustained a crush injury to the hand. This case serves to highlight the unique challenges faced in treating such injuries and to raise the profile of maggots and their untapped potential use in biodebridement and management of open wounds in modern day wound care practices.

The aim of this study was to simulate the low temperatures that insects could experience between the time being sampled from cadavers and their arrival in the laboratory. This was in order to investigate the effect of low temperature on development of maggots. At different stages of development, individuals of Protophormia terraenovae (Robineau-Desvoidy) reared at 24 degrees C were submitted to a temperature of 4.0+/-0.5 degrees C for a period varying from 1 to 10 days. Independent of the stage of development at which the insects were refrigerated, the treatment induced significant changes on the duration of development. The effect of low temperature on the developmental time between the return to 24 degrees C to adult emergence depended on the larval stage that was refrigerated. When first instar larvae and prepupae were refrigerated, the time to emergence at 24 degrees C decreased with an increase of duration of the refrigeration period. Time to emergence increased under the same conditions when second instar larvae and pupae were refrigerated. These results indicate that keeping larvae of P. terraenovae at 4 degrees C does not just simply lead to a cessation of metabolism but disturbs the regular development. Ten days of cooling induced an error in estimating post-mortem interval (PMI) of more than 6h.