Lucilia Sericata Research Articles

Predicting the Geographic Distribution ofLucilia sericataandLucilia cuprina(Diptera: Calliphoridae) in South Africa

Lucilia sericata (Meigen, 1826) and Lucilia cuprina (Wiedemann, 1830) (Diptera: Calliphoridae: Luciliinae) have medical, veterinary and forensic importance. Knowing their distribution in South Africa would allow more effective management and utilisation of these flies. Their predicted geographic distributions in South Africa were modelled using maximum entropy analysis of selected climatic variables. The most important environmental variables in modelling the distributions were the magnitude of monthly rainfall and the magnitude of the monthly maximum temperature for L. sericata and the seasonal variation in monthly mean humidity and magnitude of monthly rainfall for L. cuprina. A clear geographical bias was shown in museum records and supports the need for focused surveys. There was no correlation between the predicted distribution of L. cuprina and sheep farming in South Africa, nor between the predicted distribution of L. sericata and human population density. Although their patterns of occurrence differed, both species are widely distributed in South Africa and therefore one cannot identify these flies by locality alone — morphological or molecular identification is necessary.

Urban blow flies (Diptera: Calliphoridae) in four cities of the Colombian Caribbean coast

The purpose of this study was to evaluate the species composition of the family Calliphoridae from samples collected in four different cities in the Colombian Caribbean coast. Van Someren-Rydon traps were used baited with human faeces, rotten fish and fermented fruit were used. Six traps were placed in each city (two traps per type of bait), for a total of 24 traps. They were left for 72 hours in each site and samples were collected every 12 hours (day and night). 5654 individuals were identified, belonging to the subfamilies Chrysomyinae and Luciliinae. The identified species were Cochliomyia macellaria, Chrysomya albiceps, Chrysomya megacephala, Lucilia eximia, Lucilia sericata and Chloroprocta idioidea, expanding the range of distribution for the last two species. The best bait was the rotten fish and the best time to collect these species was during daylight.

Identifying 1st instar larvae for three forensically important blowfly species using “fingerprint” cuticular hydrocarbon analysis

Calliphoridae are known to be the most forensically important insects when it comes to establishing the minimum post mortem interval (PMImin) in criminal investigations. The first step in calculating the PMImin is to identify the larvae present to species level. Accurate identification which is conventionally carried out by morphological analysis is crucial because different insects have different life stage timings. Rapid identification in the immature larvae stages would drastically cut time in criminal investigations as it would eliminate the need to rear larvae to adult flies to determine the species. Cuticular hydrocarbon analysis on 1st instar larvae has been applied to three forensically important blowflies; Lucilia sericata, Calliphora vicina and Calliphora vomitoria, using gas chromatography–mass spectrometry (GC–MS) and principal component analysis (PCA). The results show that each species holds a distinct “fingerprint” hydrocarbon profile, allowing for accurate identification to be established in 1-day old larvae, when it can be challenging to apply morphological criteria. Consequently, this GC–MS based technique could accelerate and strengthen the identification process, not only for forensically important species, but also for other entomological samples which are hard to identify using morphological features.

Lucimycin, an antifungal peptide from the therapeutic maggot of the common green bottle fly Lucilia sericata

We report the identification, cloning, heterologous expression and functional characterization of a novel antifungal peptide named lucimycin from the common green bottle fly Lucilia sericata. The lucimycin cDNA was isolated from a library of genes induced during the innate immune response in L. sericata larvae, which are used as therapeutic maggots. The peptide comprises 77 amino acid residues with a molecular mass of 8.2 kDa and a pI of 6.6. It is predicted to contain a zinc-binding motif and to form a random coil, lacking β-sheets or other secondary structures. Lucimycin was active against fungi from the phyla Ascomycota, Basidiomycota and Zygomycota, in addition to the oomycete Phytophtora parasitica, but it was inactive against bacteria. A mutant version of lucimycin, lacking the four C-terminal amino acid residues, displayed 40-fold lower activity. The activity of lucimycin against a number of highly-destructive plant pathogens could be exploited to produce transgenic crops that are resistant against fungal diseases.

Cryopreservation of embryos of Lucilia sericata (Diptera: Calliphoridae).

Embryos of Lucilia (Phaenicia) sericata (Meigen) (Diptera Calliphoridae), the green blowfly, were successfully cryopreserved by vitrification in liquid nitrogen and stored for 8 yr. Embryos incubated at 19 degrees C for 17 h after oviposition were found to be the most appropriate stage to cryopreserve. Removal of the embryonic surface water was done using 2-propanol before the alkane treatment to permeabilize the embryo. Exposure to 2-propanol for > 10 s caused necrotic tissue damage in the embryos. Among the alkanes used, hexane was found to be a superior permeabilizing solvent compared with heptane or octane, with embryo hatching rates on par with the controls. Treatment with the vitrification solution for < 12 min was insufficient to vitrify the embryos. Treatment time in the solution beyond 15 min reduced embryo viability. However, the percentage of embryos vitrifying upon exposure to liquid nitrogen vapor remained constant after 12 min of treatment. Long-term storage was initiated in 2004, and the mean hatch percentage recorded then for the short-term cryopreserved embryos was 9.51%. When the long-term stored samples were retrieved in 2012, 8.47% of the embryos hatched, 66.36% larvae pupariated, and 36.96% of the pupae eclosed. Recent optimization of the technique has resulted in a hatch rate of 34.08 +/- 15.5%, of which 67.5% of the larvae pupariated and 72% of the pupae eclosed to normal flies.

Quantifying the Temperature of Maggot Masses and its Relationship to Decomposition

Numerous Calliphoridae species have been observed to form larval aggregations during the feeding stage of development, resulting in localized increases in temperature. This study investigates the relationship between maggot numbers in a mass and heat generation. Single-species aggregations (Lucilia sericata) of various sizes (50-2500 individuals) were reared in the laboratory at a constant ambient temperature of 22°C. Internal mass temperatures were recorded every 5 min throughout the feeding stage of development. Results showed that mass temperatures increased with mass numbers (p-value < 0.001), ranging from 2.5 to 14°C above ambient. A minimum mass size of 1200 produced overall temperatures that were significantly warmer than ambient, diverging away from 22°C after c. 26 h. These results indicate that the microclimate of a mass has the potential to differ significantly from ambient, which may be influencing larval development rates and should therefore be factored into mPMI estimates to increase accuracy.

Wound Myiasis in a Sixty-Two-Year-Old Man

Introduction: Wound myiasis is one of the common types of myiasis that demonstrates obligatory parasitic behavior. Case Presentation: A 62-year-old homeless man presented with an abnormally growing and ruptured neck mass containing worms. He had wound myiasis caused by the larva of Lucilia sericata in his previously punctured neck. Discussion: Lucilia sericata can cause both facultative and obligatory myiasis. Immunocompromised patients with diabetes, malignancies, and patients without any kind of nursery are at risk of wound myiasis. In addition, aggressive treatment is needed to prevent the secondary infections in case of obligatory infestation.

Synanthropy and temporal variability of Calliphoridae living in Cosenza (Calabria, southern Italy).

The aim of this study was to investigate the synanthropy of Diptera: Calliphoridae, insects of forensic importance, in Calabria (southern Italy). The study lasted 2 years, from February 2010 to January 2012, and was carried out in three areas of Cosenza province representing “urban”, “rural”, and “wild” environments. Bottle traps baited with pork were used to catch Calliphoridae. Six species were identified, Calliphora vicina Robineau-Desvoidy 1830, Calliphora vomitoria (L.), Chrysomya albiceps (Wiedemann 1819), Lucilia ampullacea Villeneuve 1922, Lucilia caesar (L.), and Lucilia sericata (Meigen 1826). Data on phenologies in the study areas are reported for these species and the Synanthropy Index was calculated to evaluate their relationship with the human environment.

Effect of freezing on the initial colonization of the carcass with necrophagous organisms

This study was aimed to determine whether freezing of cadaver prior to the free exposure affects the species composition and the rate of its initial colonization with necrophagous organisms. Two experiments were realized in Smecno town, the Central Bohemian Region of the Czech Republic, in which carcasses of domestic fowl (Gallus gallus f. domestica L.) weighing about 1.5 kg were obtained and treated the same way, only half of them were frozen before exposure in June and July 2013. Pre-frozen and fresh carcasses colonized the same kinds of blowflies (Diptera, Calliphoridae): Calliphora vicina Robineau-Desvoidy, 1830, Lucilia sericata (Meigen, 1826), Lucilia illustris (Meigen, 1826), Lucilia ampullacea Villeneuve, 1922, Phormia regina (Meigen, 1826), and Protophormia terraenovae (Robineau-Desvoidy, 1830). Percentage rate of each species was almost the same in both versions but we found differences in the total number of individuals (larvae) decomposing carcasses and differences in the process of decomposition of carcasses: fresh carcass decomposition was predominantly anaerobic (putrefaction) and started from the digestive system to the outside of the body (inside-out). Pre-frozen carcasses decomposed predominantly aerobic (decay) and started from the surface of body inwards (outside-in). Utilization of our results in forensic practice is discussed.

Larval Distribution and Behavior of Chrysomya rufifacies (Macquart) (Diptera: Calliphoridae) Relative to Other Species on Florida Black Bear (Carnivora: Ursidae) Decomposing Carcasses.

Larval interactions of dipteran species, blow flies in particular, were observed and documented daily over time and location on five black bear carcasses in Gainesville, FL, USA, from June 2002 - September 2004. Cochliomyia macellaria (Fabricius) or Chrysomya megacephala (Fabricius) larvae were collected first, after which Chrysomya rufifacies (Macquart) oviposited on the carcasses in multiple locations (i.e., neck, anus, and exposed flesh) not inhabited already by the other blow fly larvae. Within the first week of decomposition, C. rufifacies larvae grew to ≥12mm, filling the carcasses with thousands of larvae and replacing the other calliphorid larvae either through successful food source competition or by predation. As a result, C. macellaria and C. megacephala were not collected past their third instar feeding stage. The blow fly species, C. megacephala, C. macellaria, Lucilia caeruleiviridis (Macquart), Phormia regina (Meigen), Lucilia sericata (Meigen), and C. rufifacies, completed two developmental cycles in the 88.5-kg carcass. This phenomenon might serve to complicate or prevent the calculation of an accurate postmortem interval.

Multiple actions of Lucilia sericata larvae in hard‐to‐heal wounds

In Europe ≈15,000 patients receive larval therapy for wound treatment annually. Over the past few years, clinical studies have demonstrated the success of larvae of Lucilia sericata as debridement agents. This is based on a combination of physical and biochemical actions. Laboratory investigations have advanced our understanding of the biochemical mechanisms underlying the beneficial effects of larval secretions, including removal of dead tissue, reduction of the bacterial burden, and promotion of tissue regeneration. The present article summarizes our current understanding of the microbiological, immunological, and wound healing actions of larval therapy, and the molecules involved in these beneficial effects. Future studies will focus on the isolation, identification, and (pre)clinical testing of the effective molecules of L. sericata larvae. These molecules may be candidates for the development of new agents for the treatment of several infectious and inflammatory diseases, including chronic wounds.

Proteolytic activity regarding Sarconesiopsis magellanica (Diptera: Calliphoridae) larval excretions and secretions

Sarconesiopsis magellanica (Diptera: Calliphoridae) is a medically important necrophagous fly which is used for establishing the post-mortem interval. Diptera maggots release proteolytic enzymes contained in larval excretion and secretion (ES) products playing a key role in digestion. Special interest in proteolytic enzymes has also been aroused regarding understanding their role in wound healing since they degrade necrotic tissue during larval therapy. This study was thus aimed at identifying and characterising S. magellanica proteolytic enzyme ES products for the first time. These products were obtained from first-, second- and third-instar larvae taken from a previously-established colony. ES proteins were separated by SDS-PAGE and their proteolytic activity was characterised by zymograms and inhibition assays involving BAPNA (Nα-benzoyl-dl-Arg-p-nitroanilide) and SAPNA substrates, using synthetic inhibitors. The protein profile ranged from ∼69kDa to ∼23kDa; several of them coincided with the Lucilia sericata ES protein profile. Serine-protease hydrolysis activity (measured by zymogram) was confirmed when a ∼25kDa band disappeared upon ES incubation with PMSF inhibitor at pH 7.8. Analysis of larval ES proteolytic activity on BAPNA and SAPNA substrates (determined by using TLCK and TPCK specific inhibitors) suggested a greater amount of trypsin-like protease. These results support the need for further experiments aimed at validating S. magellanica use in larval therapy.

Antennal sensilla of the green bottle fly, Lucilia sericata (Meigen) (Diptera: Calliphoridae)

Lucilia sericata (Meigen) is a cosmopolitan synanthropic fly of forensic and medical importance, which can work as a mechanic vector of pathogens or cause myiasis of both human and sheep. As essential olfactory organs, antennae of adult L. sericata were examined with stereoscopic microscope, scanning electron microscope, and laser scanning confocal microscope. On antennal scape and pedicel, both microtirchiae and several bristles are detected, while another two structures, setiferous plaques and pedicellar buttons, are also found on the antennal pedicel. Seven subtypes of antennal sensilla are observed on antennal funiculus including one subtype of trichoid sensilla, two subtypes of basiconic sensilla, two subtypes of coeloconic sensilla, and two subtypes of sensory pits. Size and density of the former four types of sensilla on antennal funiculus are measured. Three distinctive sensillar characters of L. sericata are detected, which may contribute to greater olfactory sensitivity of this species and their wide distribution throughout the world. Unlike the common poreless pedicellar button with mechanoreceptor function, every pedicellar button in L. sericata is perforated by three pores, which might indicate potential chemoreceptor function of this structure. Besides, another unique feature is greater number of setiferous plaques in genus Lucilia than calliphorids of other genera. Expect for the common sensory pits with basiconic or basiconic-like sensilla in them, sensory pits filled with rarely described coeloconic-like sensilla are founded in L. sericata as well. After comparison with previous equivalent findings, the functions of these specific structures are discussed according to the life history of this calliphorid.

Degradation of MSCRAMM target macromolecules in VLU slough by Lucilia sericata chymotrypsin 1 (ISP) persists in the presence of tissue gelatinase activity

Venous leg ulcer slough is unpleasant to the patient and difficult to manage clinically. It harbours infection, also preventing wound management materials and dressings from supporting the underlying viable tissues. In other words, slough has significant nuisance value in the tissue viability clinic. In this study, we have sought to increase our knowledge of slough by building upon a previous but limited analysis of this necrotic tissue. In particular, slough has been probed using Western blotting for the presence of proteins with the capacity to engage microbial surface components recognising adhesive matrix macromolecules. Although the samples were difficult to resolve, we detected fibrinogen, fibronectin, IgG, collagen, human serum albumin and matrix metalloproteinase-9. Furthermore, the effect of a maggot-derived debridement enzyme, chymotrypsin 1 on macromolecules in slough was confirmed across seven patient samples. The effect of chymotrypsin 1 on slough confirms our thesis that this potential debridement enzyme could be effective in removing slough along with its associated bacteria, given its observed resistance to intrinsic gelatinase activity. In summary, we believe that the data provide scientists and clinicians with further insights into the potential molecular interactions between bacteria, wound tissue and Lucilia sericata in a clinically problematic yet scientifically interesting wound ecosystem.

Insects colonising carcasses in open and forest habitats of Central Europe: Search for indicators of corpse relocation

Several traces may reveal the post-mortem relocation of a corpse. Insects are particularly useful for that purpose. The use of insects for inferring the transfer of a corpse rests on a premise that particular species colonise corpses in different habitats. However, only some insects reveal a strong preference for a given type of habitat. In order to find those insects which colonise corpses exclusively in open habitats, as opposed to forest habitats, a pig carrion study was made in rural open and rural forest habitats of Central Europe. Lucilia sericata (Diptera: Calliphoridae), Dermestes frischi, Dermestes laniarius (Coleoptera: Dermestidae), Omosita colon, some species of Nitidula (Coleoptera: Nitidulidae) and Necrobia rufipes (Coleoptera: Cleridae) were found to breed exclusively in open habitats. Only Oiceoptoma thoracicum (Coleoptera: Silphidae) avoided definitely breeding in open habitats. Sarcophaga caerulescens (Diptera: Sarcophagidae) regularly bred in open habitats but rarely bred in forests. Accordingly, L. sericata, D. frischi, O. colon, species of Nitidula and supposedly N. rufipes may be classified as indicators of corpse relocation from rural open to rural forest habitats of Central Europe. Only O. thoracicum may be classified as an indicator of the relocation in an opposite direction.

An Initial in Vitro Investigation into the Potential Therapeutic Use of Lucilia Sericata Maggot to Control Superficial Fungal Infections

In this work an attempt was performed to investigate the in vitro ability of Lucilia sericata maggots to control fungi involved in superficial fungal infections. A novel GFP-modified yeast culture to enable direct visualization of the ingestion of yeast cells by maggot larvae as a method of control was used. The obtained results showed that the GFP-modified yeasts were successfully ingested by Lucilia sericata maggots and 1mg/ml of Lucilia sericata maggots excretions/ secretions (ES) showed a considerable anti-fungal activity against the growth of Trichophyton terrestre mycelium, the radial growth inhibition after 10 days of incubation reached 41.2 ±1.8 % in relation to the control, these results could lead to the possible application of maggot therapy in the treatment of wounds undergoing fungal infection.

Applicability of partial characterization of cytochrome oxidase I in identification of forensically important flies (Diptera) from China and Egypt

Precise species identification of every insect sample collected from criminal scenes play an essential role in the accurate estimation of postmortem interval. The morphological similarity poses a great challenge for forensic entomologists. DNA-based method can be used as a supplemental means of morphological method. In the present study, we demonstrate the applicability of the 304-bp cytochrome oxidase I gene fragment in molecular identification of forensically important Diptera. We analyzed 75 specimens belonging to 19 species of 3 families originating from China (Calliphoridae: Chrysomya megacephala, Chrysomya albiceps, Chrysomya rufifacies, Chrysomya nigripes, Aldrichina grahami; Lucilia bazini, Lucilia caesar, Lucilia cuprina, Lucilia sericata, Lucilia porphyrina; Muscidae: Musca autumnalis, Musca domestica, Fannia canicularis, Stomoxys calcitrans; Sarcophagidae: Sarcophaga albiceps, Sarcophaga dux, Helicophagella melanura) and Egypt (Calliphoridae: C. megacephala, C. albiceps, L. sericata; Muscidae: M. domestica, F. canicularis, S. calcitrans, Synthesiomyia nudiseta; Sarcophagidae: Sarcophaga argyrostoma). This region was amplified using polymerase chain reaction followed by direct sequencing of the amplification products. Nucleotide sequence divergences were calculated using the Kimura two-parameter distance model and a neighbor-joining phylogenetic tree generated. Intraspecific variation ranged from 0-0.8 % and interspecific variation occurred between 1-19 %. Although all examined specimens were assigned to the correct species and formed distinct monophyletic clades, the data of the phylogenetic analysis were not completely in accordance with the traditional morphological classification. As both C. nigripes and A. grahami unexpectedly joined with Muscidae and Sarcophagidae groups respectively. Moreover, both Calliphorinae and Luciliinae clades failed to represent Calliphoridae as a separate group. Therefore, although molecular methods are beneficial especially if the available evidence are immature stage or degraded fragments of insect, we suggest using this fragment only in addition to other identification methods.

Lucifensin II, a Defensin of Medicinal Maggots of the BlowflyLucilia cuprina(Diptera: Calliphoridae)

A novel homolog of insect defensin, designated lucifensin II (Lucilia cuprina Wiedemann [Diptera: Calliphoridae] defensin), was purified from hemolymph extract from larvae of the blowfly L. cuprina. The full-length primary sequence of this peptide of 40 amino acid residues and three intramolecular disulfide bridges was determined by electrospray ionization-orbitrap mass spectrometry and Edman degradation and is almost identical to the previously identified sequence of lucifensin (Lucilia sericata Meigen defensin). Lucifensin II sequence differs from that of lucifensin by only one amino acid residue, that is, by isoleucine instead of valine at position 11. The presence of lucifensin II also was detected in the extracts of other larval tissues, such as gut, salivary glands, fat body, and whole body extract.

Expression of lucifensin in Lucilia sericata medicinal maggots in infected environments

Lucifensin, a novel larval defensin, is one of the antibacterial agents of medicinal maggots involved in maggot therapy. The goal of this study was to examine lucifensin expression in various larval tissues during Lucilia sericata development and in maggots exposed to a variety of infectious environments in vitro. In situ hybridisation revealed lucifensin expression in the salivary glands of all larval stages. Expression was occasionally detected in a few cells of the fat body and in the grease coupler of the salivary glands. Expression of lucifensin in the salivary glands was initiated 5-6 h after hatching from the egg. Maximum expression was reached about 24 h after hatching, remained strong during the second and third instars and declined at the end of the third instar, before the wandering stage. Expression of lucifensin was also investigated in maggots after oral ingestion of certain pathogens regularly found in infected chronic wounds. No differences were detected in the salivary glands after stimulation by wound bacterial isolates. However, lucifensin expression was strongly stimulated in the fat body by the presence of Staphylococcus aureus and Pseudomonas aeruginosa. Our data suggest that certain infectious environments increase lucifensin expression only in the fat body, whereas its production and antimicrobial activity in excretion/secretion products are not affected.

Evidence of active aggregation behaviour in Lucilia sericata larvae and possible implication of a conspecific mark

Vectors of aggregation are well known for some arthropod species, but not for many others. We aimed to describe larval aggregation (experiment 1) in the carrion fly, Lucilia sericata (Diptera: Calliphoridae), and to investigate the effect of food and conspecifics on larval behaviour (experiment 2). In experiment 1, 40 larvae were placed in a petri dish with a homogeneous diet for 30 min, 1 h, 3 h, 5 h or 24 h. This experiment demonstrated for the first time under controlled conditions the active aggregation of L. sericata larvae. The results indicate that the aggregation took place quickly and was reinforced with time. After only 3 h, one main aggregate comprising a majority of individuals was observed. These results also highlight the likely use by necrophagous larvae of a signal left by conspecifics as an aggregation vector. In experiment 2, we used a video-tracking system to investigate whether such an aggregative signal exists. Fed and starved larvae were tracked for 5 min in a circular area with each half marked with a different signal combination. The time spent in the signal zones, the distance travelled, the velocity, the time at the stop and the number of stops in each zone were measured. The larvae were significantly retained by a signal (mark) left by conspecifics. Together, the results of this study demonstrate the existence of a contact and/or odour-mediated signal involved in the aggregative behaviour of necrophagous larvae.