Morphology of immature stages of Hemipyrellia ligurriens (Wiedemann) (Diptera: Calliphoridae) for use in forensic entomology applications

In forensic investigations, all immature stages of flies (egg, larvae, and puparium) can serve as entomological evidence at death scenes. These insects are primarily used to estimate the post mortem interval (PMI), but can also be involved in the analysis of toxic substances, determining manner of death, and in indicating relocation of a corpse in homicide cases. In this study, we present the morphology of the egg, larvae, and puparium of Hemipyrellia ligurriens, a blow fly species of forensic importance in Thailand. Examination was conducted using scanning electron microscopy (SEM). The egg stage was found to display a relatively wide plastron region (or median hatch line area) that spans almost the entire length of the egg. The median hatch line is oriented in an upright position. External chorionic sculpture of the egg is present in a hexagonal pattern whose reticular boundaries are slightly elevated. In the larval stages, the most prominent morphological changes were detected upon comparison of the first to the second instar; whereas, the differences between second and third instar larvae were less obvious outside of the increase in number of posterior spiracular slits. Most of the major differences involve body size and structure of the anterior and posterior spiracles. Each anterior spiracle in both the second and third instars projects five to seven papillae apically. Each posterior spiracular disc of a third instar exhibits a complete peritreme, three spiracular slits, and a prominent button that is ventromedially located. The puparium is coarctate and features a clustered bubble membrane comprised of approximately 57 mammillate structures positioned dorsolaterally on each side of the first abdominal segment in young puparia. This feature is replaced by short, tubular respiratory horns in aged puparia. This study provides more detailed exposure of important morphological features that can be used for accurate identification of immature stages of H. ligurriens. Information presented can aid in forensic investigations involving this fly species.     

Sterilization of blow fly eggs,Chrysomya megacephala and Lucilia cuprina, (Diptera: Calliphoridae) for maggot debridement therapy application

Parasitology Research - Maggot debridement therapy (MDT) is an application of sterile laboratory-reared blow fly larvae to remove necrotic tissue and disinfect wounds for medical conditions. For...     

Effects of temperature on development of Phormia regina (Diptera: Calliphoridae) and use of developmental data in determining time intervals in forensic entomology

Precise developmental data for forensic indicator blow fly species are essential for accuracy in the estimate of the post-mortem interval (PMI). Why, then, does the determination of the PMI result in conflicting time frames when published conspecific developmental data are used? To answer this question, we conducted constant temperature trials between the developmental minimum temperature and upper threshold temperatures (8-32 degrees C) on the forensically important blow fly species Phormia regina (Meigen) (Diptera: Calliphoridae). Flies were reared using two designs to quantify sources of variation. We measured rearing container temperatures and internal growth chamber temperatures by using thermocouples to accurately record temperatures experienced by larvae and to construct a degree-day model. Differences in experimental design, as seen across temperature studies for this fly species, did not significantly impact larval development. We also found that using set chamber temperatures rather than rearing container temperatures altered the final degree-day model. Using any minimum threshold (including an empirically determined true minimum) other than that from linear interpolation (x-intercept) violated degree-day assumptions and invalidated estimates of the PMI. We observed the minimum developmental temperature to be higher (14 degrees C) than that generated under the x-intercept method (5.46 degrees C) by using data from oviposition to adult emergence. This difference along with the noted difference in accumulated degree-days (using different base temperatures) suggests a need for additional experimentation on other forensically important fly species at low temperature thresholds to help with development of curvilinear models. Former and current estimates of the PMI may be inaccurate if the process to determine the time frame ignored degree-day model assumptions or was based upon questionable data sets.     

Surface ultrastructure of the puparia of the blow fly, Lucilia cuprina (Diptera: Calliphoridae), and flesh fly, Liosarcophaga dux (Diptera: Sarcophagidae)

Surface ultrastructure of the puparia of the blow fly, Lucilia cuprina, and flesh fly, Liosarcophaga dux, are presented utilizing scanning electron microscopy (SEM). Emphasis was focused on characteristic features of the puparia that could be used for differentiation from other forensically important fly species. The puparium of L. cuprina typically measures 6.2±0.2 mm in length and 2.3±0.1 mm in width. Each anterior spiracle contains a single row of 5–7 papillae. The intersegmental spines between the prothorax and mesothorax are triangular with constricted tips. A clustered bubble membrane, comprising about 30 mammillate structures, is positioned dorsolaterally on each side of the first abdominal segment in young puparia but is replaced by short, tubular respiratory horns in aged puparia. The posterior end of the puparium is broadly truncate and bears a pair of medially positioned posterior spiracles that each contains three straight spiracular slits. The puparium of L. dux is larger in comparison to L. cuprina and typically measures 9.9±0.3 mm in length and 3.8±0.2 mm in width. An anterior spiracle of this species contains 14–17 papillae. The intersegmental spines between the prothorax and mesothorax are broad and triangular. A convoluted structure of unknown function was observed at the dorsolateral edge of segments 5–11. In L. dux, the caudal segment of the puparium is slightly tapered and abruptly truncated and contains a pair of posterior spiracles that are located within a deep concavity in the terminal end. Each posterior spiracular disc appears D-shaped, with a pronounced medial projection and three vertically oriented long, narrow spiracular slits. The anatomical features presented herein may be useful for identification of fly puparia of these two species in future forensic entomological investigations.     

Wound myiasis in cattle infested by Lucilia caesar (Diptera: Calliphoridae)

A case report on facultative wound myiasis after sectio caesarea in a cow by larvae of Lucilia caesar in May 1986.     

Observations on morphology of immature Lucilia porphyrina (Diptera: Calliphoridae), a fly species of forensic importance

Lucilia porphyrina (Walker) is a blow fly of forensic importance, and shares its geographical distribution with a related forensically important species, Lucilia cuprina (Wiedemann). The immature stages of both species are similar in general appearance; therefore, correct identification should be given special consideration. This study highlighted the main features of L. porphyrina larvae, as observed under light microscopy and scanning electron microscopy. Particular attention is given to the anterior and posterior spiracles, cephalopharyngeal skeleton, and characteristics of the dorsal spines between the prothorax and mesothorax. In the third instar specifically, morphological information on L. porphyrina showed several features that are shared by L. cuprina, and therefore need certain identification to distinguish between them. Such key features are (1) greater posterior spiracle, (2) apparent inner projection between the middle and lower slits of the posterior spiracle, and (3) strongly sclerotized peritreme. The number of papillae on the anterior spiracle may be a supplement, five to nine and three to six in L. porphyrina and L. cuprina, respectively. The key for identifying third instar of forensically important flies in Thailand has been updated to include L. porphyrina.     

Components of developmental plasticity in a Michigan population of Lucilia sericata (Diptera: Calliphoridae)

Forensic entomologists rely on laboratory growth data to estimate the time of blow fly colonization on human remains. Several data sets exist for the development of the common blow fly Lucilia sericata (Meigen) (Diptera: Calliphoridae), and although they generally describe similar rates of preadult development, all vary. Such differences could be explained by genetic variation, environmental (rearing) variation, or both. In the study presented here, flies from a single population were reared under variable conditions of food moisture, substrate type, substrate freshness, and sampling, to determine the effect each had on developmental time. Cohorts were tested in a single incubator at a single temperature and humidity, to eliminate effects of undesired environmental variation. Fly developmental times were significantly influenced by multiple laboratory rearing treatments; food moisture, transferring postfeeding larvae to fresh substrate, and destructive sampling affected different stages of development. Developmental times ranged from 329 to 505.5 h, covering the spectrum of variation observed in published data sets. Growth was then compared with larval development on rat carcasses under the same environmental conditions, establishing a link between laboratory-controlled growth and development on carrion. Cohorts raised on rats matured to adulthood between 333 and 337 h, which was best mimicked by the fastest growth treatment observed under laboratory conditions. The large environmental influence on development observed in this study could affect forensic entomology casework and accentuates the need for a standardized means of rearing flies in a laboratory setting that is relevant to decomposition on a corpse.     

First report of human myiasis caused by Chrysomya megacephala and Chrysomya rufifacies (Diptera: Calliphoridae) in Thailand, and its implication in forensic entomology

We report a forensic entomology case associated with human myiasis in Chiang Mai Province, northern Thailand. The remains of a 53-yr-old-male were concurrently infested with third instars of the two blow fly species, Chrysomya megacephala (F.) and Chrysomya rufifacies (Macquart), near a severe tumor lesion presented on the lower right leg. The presence of third instars, approximately 5 d old, on the day following postmortem indicated that myiasis occurred before death. This is the first report of both fly species acting as a myiasis-producing agent in Thailand. Unsynchronized data between the age of fly larvae due to myiasis premortem and verified age/ condition of the corpse suggest a potential complication and error in the estimation of postmortem interval if other predisposition fly infestations are not considered.     

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.     

Population and temperature effects on Lucilia sericata (Diptera: Calliphoridae) body size and minimum development time

Understanding how ecological conditions influence physiological responses is fundamental to forensic entomology. When determining the minimum postmortem interval with blow fly evidence in forensic investigations, using a reliable and accurate model of development is integral. Many published studies vary in results, source populations, and experimental designs. Accordingly, disentangling genetic causes of developmental variation from environmental causes is difficult. This study determined the minimum time of development and pupal sizes of three populations of Lucilia sericata Meigen (Diptera: Calliphoridae; from California, Michigan, and West Virginia) at two temperatures (20 degrees C and 33.5 degrees C). Development times differed significantly between strain and temperature. In addition, California pupae were the largest and fastest developing at 20 degrees C, but at 33.5 degrees C, though they still maintained their rank in size among the three populations, they were the slowest to develop. These results indicate a need to account for genetic differences in development, and genetic variation in environmental responses, when estimating a postmortem interval with entomological data.     

Destruction of bacteria in the digestive tract of the maggot of Lucilia sericata (Diptera: Calliphoridae)

Green fluorescent protein-producing Escherichia coli were used to investigate the fate of bacteria in the alimentary tract of sterile grown maggots, Lucilia sericata (Meigen), using a laser scanning confocal microscope. A computer program was used to analyze the intensity of the fluorescence and to quantify the number of bacteria. The crop and the anterior midgut were the most heavily infected areas of the intestine. A significant decrease in the amount of bacteria was observed in the posterior midgut. The number of bacteria decreased even more significantly in the anterior hindgut and practically no bacteria were seen in the posterior end, near the anus. The viability of bacteria in the different gut sections was examined. It was shown that 66.7% of the crops, 52.8% of the midguts, 55.6% of the anterior hindguts, and 17.8% of posterior hindguts harbored living bacteria. In conclusion, during their passage through the digestive tract the majority of E. coli was destroyed in the midgut. Most of the remaining bacteria were killed in the hindgut, indicating that the feces were either sterile or contained only small numbers of bacteria.     

Interdisciplinary workshop yields new entomological data for forensic sciences: Chrysomya rufifacies (Diptera: Calliphoridae) established in North Carolina

ABSTRACT Workshops are used for educating law enforcement personnel on the application of entomological, anthropological, and botanical techniques to gather forensically important information from a body recovery site. From 8 to 11 June 2004, such a workshop was conducted in the metropolitan area of Charlotte, NC, by the American Academy of Applied Forensics at Central Piedmont Community College. For this workshop, three pig carcasses weighing individually between 40 and 60 kg were placed in the field 4 June, whereas three pigs similar in size were placed in the field 7 June. During the afternoon session on 11 June, workshop participants collected three Chrysomya rufifacies (Macquart) (Diptera: Calliphoridae) third instar larvae from a pig carcass placed in the field on 7 June. These data represent the first records of this forensically important insect in North Carolina; furthermore, these data provide evidence of this species expansion into new geographic regions of the United States. This finding is an example of the benefits derived from an experiential and interdisciplinary approach to educating death scene investigators. Learners acquired new knowledge, put it into practice through the "body" recovery exercise, and ultimately contributed to science by way of the discovery and first documentation of a forensically important insect previously not known to inhabit North Carolina.