Chrysomya Nigripes Research Articles

Geometry of posterior larval spiracles to identify medically and forensically important calliphorids in Thailand

Fly identification is the primary step of analysis in forensic entomology. Although morphology and molecular techniques are considered satisfactory methods, some constraints may arise from a financial or even human point of view. Over the past decade, the geometric morphometric approach has been increasingly advocated for the classification and identification of arthropods. This study explored the method for species identification of 800 third-instar larvae of eight blow fly species of medical and forensic importance: Chrysomya chani Kurahashi, Chrysomya megacephala (Fabricius), Chrysomya (Ceylonomyia) nigripes Aubertin, Chrysomya pinguis (Walker), Chrysomya (Achoetandrus) rufifacies (Macquart), Hemipyrellia ligurriens (Wiedemann), Lucilia cuprina (Wiedemann), and Lucilia porphyrina (Walker). Based on the posterior spiracles geometry, the cross-validation revealed a relatively high percentage of correct classification in most species, ranking from 86% to 100%. The results of this study confirmed that the geometric morphometric (GM) analysis of posterior spiracles might be utilized as a larva identification tool. Therefore, this GM method represents one way of overcoming difficulties with the identification of blow fly larvae and can support further studies of these flies.

Developmental Indicators of Chrysomya nigripes Aubertin under Different Constant Temperature Conditions and an Application Case for Estimating the PMImin.

Chrysomya nigripes Aubertin, 1932, is a Calliphoridae species that colonize the carcass after the bloat phase and remains for long periods. Some early sarcosaprophagous insects complete one generation of development and are no longer associated with the corpse and surrounding environment, while C. nigripes larvae and pupae remain, providing a basis for the estimation of the minimum postmortem interval (PMImin) for highly decomposed or skeletonized carcasses. However, data on the growth and development of this species are not yet complete. As a result, we studied the developmental patterns of C. nigripes at eight constant temperatures ranging from 16-37 °C and constructed various developmental models, including the isomorphen diagram, isomegalen diagram, linear thermal summation model, nonlinear thermodynamic Optim SSI model, and logistic regression model. Chrysomya nigripes could not complete the entire developmental process at 16 °C, although it could be completed at other temperatures. The mean developmental times (±SD) of C. nigripes from egg to adult at 19 °C, 22 °C, 25 °C, 28 °C, 31 °C, 34 °C, and 37 °C were 644.9 ± 36.8 h, 422.9 ± 20.1 h, 323.1 ± 13.9 h, 246.6 ± 11.2 h, 202.5 ± 1.8 h, 191.5 ± 3.8 h, and 191.8 ± 2.0 h, respectively. The thermal summation constant (K) and lower critical thermal threshold (TL) derived from the linear thermal summation models were 4083.00 ± 293.39 degree hours and 12.52 ± 0.83 °C, respectively. In addition, TL, intrinsic optimum temperature (TΦ), and upper critical thermal threshold (TH) estimated by the optimized nonlinear thermodynamic Optim SSI model were 15.76 °C, 24.88 °C, and 38.15 °C, respectively. This study provides more comprehensive developmental data of C. nigripes for PMImin estimation.

Identification of medically and forensically relevant flies using a decision treelearning method.

Blow flies, flesh flies, and house flies can provide excellent evidence for forensic entomologists and are also essential to the fields of public health, medicine, and animal health. In all questions, the correct identification of fly species is an important initial step. The usual methods based on morphology or even molecular approaches can reach their limits here, especially when dealing with larger numbers of specimens. Since machine learning already plays a major role in many areas of daily life, such as education, business, industry, science, and medicine, applications for the classification of insects have been reported. Here, we applied the decision tree method with wing morphometric data to construct a model for discriminating flies of three families [Calliphoridae, Sarcophagidae, Muscidae] and seven species [Chrysomya megacephala (Fabricius), Chrysomya rufifacies (Macquart), Chrysomya (Ceylonomyia) nigripes Aubertin, Lucilia cuprina (Wiedemann), Hemipyrellia ligurriens (Wiedemann), Musca domestica Linneaus, and Parasarcophaga (Liosarcophaga) dux Thomson]. One hundred percent overall accuracy was obtained at a family level, followed by 83.33% at a species level. The results of this study suggest that non-experts might utilize this identification tool. However, more species and also samples per specimens should be studied to create a model that can be applied to the different fly species in Thailand.

Application of Sarcosaprophagous Insects to Estimate the Postmortem Interval in 11 Cases.

Objective To test the feasibility and accuracy of with sarcosaprophagous insects postmortem interval （PMI） estimation with sarcosaprophagous insects and provide references for estimation practice. Methods Eleven cases confirmed by the detection results, with complete entomological evidence were selected. The insect species, estimation results and true results involved in the cases were statistically analyzed and compared. Results Thirteen species of insects were found at the criminal scene, including Chrysomya megacephala （Fabricius）, Chrysomya rufifacies （Macquart）, Chrysomya nigripes （Aubertin）, Lucilia sericata （Meigen）, Hydrotaea spinigera Stein, Muscina stabulans （Fallén）, Sarcophagid （species were not identified）, Megaselia scalaris （Loew）, Hermetia illucens （Linnaeus）, Saprinus splendens （Paykull）, Creophilus maxillosus （Linnaeus）, Dermestes maculatus （De Geer） and Necrobia ruficollis （Fabricius）. The PMI of all eleven cases was within the range of estimated PMI. The estimated results of 72.73% cases were on the same day of the true results. Conclusion Sarcosaprophagous insects can estimate the PMI simply and conveniently. In cases where the PMI is within the time range of one generation of flies or beetles, the estimation results are relatively accurate. However, the estimation is less accurate when the PMI is beyond the time range.

The complete mitochondrial genome of Chrysomya nigripes (Diptera: Calliphoridae)

Chrysomya nigripes (Diptera: Calliphoridae) is a blow fly species of forensic importance. Here we demonstrated the complete mitochondrial genome of this species for the first time. Phylogenetic analyses indicated that entire mitochondrial genome sequences can provide more useful information for distinguishing C. nigripes from the other species.

Molecular Analysis of Forensically Important Blow Flies in Thailand.

Blow flies are the first insect group to colonize on a dead body and thus correct species identification is a crucial step in forensic investigations for estimating the minimum postmortem interval, as developmental times are species-specific. Due to the difficulty of traditional morphology-based identification such as the morphological similarity of closely related species and uncovered taxonomic keys for all developmental stages, DNA-based identification has been increasing in interest, especially in high biodiversity areas such as Thailand. In this study, the effectiveness of long mitochondrial cytochrome c oxidase subunit I and II (COI and COII) sequences (1247 and 635 bp, respectively) in identifying 16 species of forensically relevant blow flies in Thailand (Chrysomya bezziana, Chrysomya chani, Chrysomya megacephala, Chrysomya nigripes, Chrysomya pinguis, Chrysomya rufifacies, Chrysomya thanomthini, Chrysomya villeneuvi, Lucilia cuprina, Lucilia papuensis, Lucilia porphyrina, Lucilia sinensis, Hemipyrellia ligurriens, Hemipyrellia pulchra, Hypopygiopsis infumata, and Hypopygiopsis tumrasvini) was assessed using distance-based (Kimura two-parameter distances based on Best Match, Best Close Match, and All Species Barcodes criteria) and tree-based (grouping taxa by sequence similarity in the neighbor-joining tree) methods. Analyses of the obtained sequence data demonstrated that COI and COII genes were effective markers for accurate species identification of the Thai blow flies. This study has not only demonstrated the genetic diversity of Thai blow flies, but also provided a reliable DNA reference database for further use in forensic entomology within the country and other regions where these species exist.

Ultrastructure of male genitalia of blow flies (Diptera: Calliphoridae) of forensic importance

Male genitalia of blow flies (Diptera: Calliphoridae) are distinctive in their morphological features and are often used for species identification. The aim of this work was to investigate the male genitalia of blow flies of medical and forensic importance from Thailand at the ultrastructural level, using scanning electron microscopy (SEM). Flies in two subfamilies were examined: Chrysomyinae [Chrysomya bezziana Villeneuve, Chrysomya chani Kurahashi, Chrysomya nigripes Aubertin, Chrysomya pinguis (Walker), Chrysomya rufifacies (Macquart), Chrysomya thanomthini Kurahashi & Tumrasvin, and Chrysomya villeneuvi Patton] and Luciliinae [Hemipyrellia ligurriens (Wiedemann), Hypopygiopsis infumata (Bigot), Hypopygiopsis tumrasvini Kurahashi, Lucilia cuprina (Wiedemann), Lucilia papuensis Macquart, Lucilia porphyrina (Walker), and Lucilia sinensis Aubertin]. Particular attention was paid to the main distinguishing features such as the shapes of the cercus and the surstylus, and the complex structure of the distiphallus. The differentiation of the male genitalia of these species at the SEM level is discussed and compared to the conditions in closely related species such as Chrysomya megacephala (Fabricius). A key for the identification of 14 blow fly species based on male genitalia is provided.

Temperature-dependent development and the significance for estimating postmortem interval of Chrysomya nigripes Aubertin, a new forensically important species in China.

In forensic entomology, the developmental duration and larval body length of sarcosaprophagous flies are presently the two major approaches to estimate minimal postmortem interval (PMImin). A full characterization of the developmental stages of Calliphoridae, a predominating species on corpses, is especially important for PMI estimation. Chrysomya nigripes Aubertin, which appears on the corpse mainly during the decay and the post-decay stages, is of significant value for PMImin estimation. However, there are limited detailed reports on the development of such species. Hence, this study was conducted to address the development pattern of C. nigripes, in order to provide the basic data related to PMI estimation. In this study, the larvae of C. nigripes were collected from pig carcasses that were placed in the field. The colonies were cultured at constant temperatures of 16, 20, 24, 28, and 32 (±1) °C. The biological features of C. nigripes, the developmental pattern, and the body length with time at different temperatures were studied. The results showed that the average developmental duration of the C. nigripes larvae at 20, 24, 28, and 32 (±1) °C were 608.0 ± 68.0h, 327.0 ± 53.8h, 254.0 ± 36.5h, and 217.0 ± 28.0h, respectively. There were two phases of body length increment with developmental time: growing phase and plateau phase. The maximal body length was 11-12mm, and the relationship between body length and developmental time can be simulated using the following equation: L = a + bT + cT(2) + dT(3). The data for developmental duration and larval body length at 16°C were not obtained. In summary, this study comprehensively studied the developmental biology of C. nigripes, which has a significant value for estimating PMI from highly decayed carcasses.

The Importance of Habitat in the Ecology of Decomposition on Rabbit Carcasses in Malaysia: Implications in Forensic Entomology.

The stages of decomposition and the faunal succession on rabbit carcasses in three different habitats, namely jungle, rural, and highland areas, were studied. Three New Zealand White rabbit (Oryctolagus cuniculus) carcasses weighing ∼2 kg were sampled daily until the decomposition process was completed. Representative specimens of adult flies, larvae, pupa, and mites were collected from the carcasses and processed in the laboratory. There were differences in decomposition rate and faunal succession between the carcasses. The fastest rate of decomposition was recorded in rural area, and the slowest rate of decomposition was recorded in highland area. The carcasses exhibited the same pattern of colonization by adult flies, but the dominant species of larvae and adult flies on each carcass in specific habitats were different. The primary species of flies recorded in jungle were Chrysomya megacephala F., Chrysomya rufifacies (Macquart), Chrysomya chani Kurahashi, Chrysomya villenuevi Patton, Chrysomya nigripes Aubertin, Chrysomya pinguis (Walker), Hemipyrellia ligurriens (Wiedemann), Hemipyrellia tagaliana (Bigot), Hypopyiopsis fumipennis (Walker), Hypopygiopsis violacea (Macquart), and Hydrotaea spinigera Stein represented by both adults and larvae. Musca domestica L., Atherigona sp., Lioproctia pattoni (Senior-White), Lioproctia saprianovae Pape & Bänziger, and Seniorwhitea princeps (Wiedemann) were represented by adults only. The biodiversity of flies in the rural area were C. megacephala, C. rufifacies, H. ligurriens, Fannia canicularis L., Hydrotaea chalcogaster (Wiedemann), and Hyd. spinigera represented by both adults and larvae, meanwhile M. domestica, Atherigona sp., Boettcherisca peregrina (Robineau-Desvoidy), Parasarcophaga taenionota Wiedemann, Parasarcophaga scopariiformis Senior-White, and S. princeps were represented by adults only. The species of flies collected in the highland area were Lucilia porphyrina (Walker), C. megacephala, C. rufifacies, C. villenuevi, C. pinguis, H. ligurriens, Hyd. spinigera, Hyd. chalcogaster, F. canicularis, and Boettcherisca highlandica Kurahashi & Tan represented by both adults and larvae, whereas C. nigripes, Chrysomya thanomthini Kurahashi & Tumrasvin, M. domestica, Atherigona sp., Parasarcophaga albiceps Meigen, P. taenionota, Sepsidae, Phoridae, and Millichidae were represented by adults only. Faunal succession followed the sequence of dominant flies, i.e., Calliphoridae, Sarcophagidae, Muscidae, Sepsidae, and lastly Stratiomyidae for jungle, or Sepsidae for rural and highland studies. Mites, from suborders Mesostigmata, Prostigmata, Astigmatina, and Oribatida, were also recovered throughout decomposition, which could be used for future implementation in forensic investigations. The data obtained from this study could provide more accurate indicators for local forensic scientists in solving criminal cases especially on the determination of time and primary location of death.

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.

Ommatidia of blow fly, house fly, and flesh fly: implication of their vision efficiency

This work aims to elucidate the number of ommatidia or facets (the outwardly visible units of each ommatidium) for compound eyes in blow flies [Chrysomya megacephala (F.), Chrysomya rufifacies (Macquart), Chrysomya nigripes (Aubertin), Lucilia cuprina (Wiedemann)], house flies (Musca domestica L.), and flesh flies (Liosarcophaga dux Thomson) by manual counts of the corneal spreads. The head of the fly in each species was soaked in 20% potassium hydroxide solution at room temperature for 7 days, and the clear compound eye was dissected into six small parts, each of which was placed onto a slide and flattened using a coverslip. Images of each part were obtained using a microscope connected to a computer. The printed images of each part were magnified, and the total number of ommatidia per eye was manually counted. For males, the mean number of ommatidia was statistically different among all flies examined: L. dux (6,032) > C. rufifacies (5,356) > C. nigripes (4,798) > C. megacephala (4,376) > L. cuprina (3,665) > M. domestica (3,484). Likewise, the mean number of facets in females was statistically different: L. dux (6,086) > C. megacephala (5,641) > C. rufifacies (5,208) > C. nigripes (4,774) > L. cuprina (3,608) > M. domestica (3433). Scanning electron microscopy analysis of adult flies revealed the sexual dimorphism in the compound eye. Male C. megacephala had large ommatidia in the upper two thirds part and small ommatidia in the lower one third part, whereas only small ommatidia were detected in females. Dense postulate appearance was detected in the external surface of the corneal lens of the ommatidia of C. megacephala, C. rufifacies, and C. nigripes, while a mix of dense postulate appearance and variable groove array length was detected in L. cuprina and M. domestica. The probable functions of ommatidia are discussed with reference to other literature.

Identifying fly puparia by clearing technique: application to forensic entomology

In forensic investigations, immature stages of the fly (egg, larva, or puparia) can be used as entomological evidence at death scenes, not only to estimate the postmortem interval (PMI), analyze toxic substances, and to determine the manner of death but also to indicate the movement of a corpse in homicide cases. Of these immature stages, puparia represent the longest developmental time, which makes them of useful. However, in order for forensic entomologists to use puparia effectively, it is crucial that they are able to accurately identify the species of fly found in a corpse. Typically, these puparia are similar in general appearance, being coarctate and light brown to dark brown in color, which makes identification difficult. In this study, we report on the clearing technique used to pale the integument of fly puparia, thereby allowing observation of the anterior end (second to fourth segments) and the profile of the posterior spiracle, which are important clues for identification. We used puparia of the blowfly, Chrysomya megacephala (F.), as the model species in this experiment. With placement in a 20% potassium hydroxide solution daily and mounting on a clearing medium (Permount(R), New Jersey), the profile of the posterior spiracle could be clearly examined under a light microscope beginning on the fifth day after pupation, and the number of papillae in the anterior spiracle could be counted easily starting from the ninth day. Comparison of morphological features of C. megacephala puparia with those of other blowflies (Chrysomya nigripes [Aubertin], Chrysomya rufifacies [Macquart], Chrysomya villeneuvi [Patton], Lucilia cuprina [Wiedemann], and Hemipyrellia ligurriens [Wiedemann]) and a housefly (Musca domestica L.) revealed that the anterior ends and the profiles of the posterior spiracles had markedly distinguishing characteristics. Morphometric analysis of the length and width of puparia, along with the length of the gaps between the posterior spiracles of seven fly species, displayed differences among them. This provides a key to identifying the puparia of these seven fly species.

Forensic entomology cases in Thailand: a review of cases from 2000 to 2006

This paper presents and discusses 30 cases of cadavers that had been transferred for forensic entomology investigations to the Department of Forensic Medicine, Faculty of Medicine, Chiang Mai University, northern Thailand, from 2000 to 2006. Variable death scenes were determined, including forested area and suburban and urban outdoor and indoor environments. The fly specimens found in the corpses obtained were the most commonly of the blow fly of family Calliphoridae, and consisted of Chrysomya megacephala (F.), Chrysomya rufifacies (Macquart) Chrysomya villeneuvi Patton, Chrysomya nigripes Aubertin, Chrysomya bezziana Villeneuve, Chrysomya chani Kurahashi, Lucilia cuprina (Wiedemann), Hemipyrellia ligurriens (Wiedemann), and two unknown species. Flies of the family Muscidae [Hydrotaea spinigera Stein, Synthesiomyia nudiseta (Wulp)], Piophilidae [Piophila casei (L.)], Phoridae [Megaselia scalaris (Loew)], Sarcophagidae [Parasarcophaga ruficornis (F.) and three unknown species], and Stratiomyiidae (Sargus sp.) were also collected from these human remains. Larvae and adults of the beetle, Dermestes maculatus DeGeer (Coleoptera: Dermestidae), were also found in some cases. Chrysomya megacephala and C. rufifacies were the most common species found in the ecologically varied death scene habitats associated with both urban and forested areas, while C. nigripes was commonly discovered in forested places. S. nudiseta was collected only from corpses found in an indoor death scene.

Fine structure of Chrysomya nigripes (Diptera: Calliphoridae), a fly species of medical importance

The fine structure of Chrysomya nigripes Aubertin, a blow fly species of medical importance, is presented using scanning electron microscopy (SEM) to contribute information on the morphology of the adult of this fly species. The surface of the dome-shaped ommatidia exhibits a microscopic granulose appearance. The palpus is equipped with small sensilla basiconica and sensilla chaetica, which provide sensory reception for detecting environmental information. At the apex of the mouthparts, the labellum is endowed with large numbers of sensilla trichodea and basiconic-like sensilla of variable length. The anterior (mesothoracic) spiracle is elliptical in shape and covered with extensively ramified setae except for a small dorsal aperture. The posterior (metathoracic) spiracle is shaped like a rounded isosceles triangle and covered by two valves of unequal size. The larger valve covers the upper approximately 2/3 of the spiracular opening, whereas the smaller valve covers the lower approximately 1/3 of the opening. Extensively ramified setae line and cover the valves over the entire spiracle. SEM analyses of the haltere knob and the prosternal organs, located adjacent to the cervical sclerites, revealed a striking resemblance of the morphological features of their sensilla. Each sensillum emanates from a cuticular ring, is approximately 12-15 mum in length, has a smooth surface, and terminates in a sharp tip. Various types of sensilla were associated with the ovipositor including sensilla trichodea, sensilla basiconica, sensilla placodea and probably sensilla styloconica. The possible function of sensilla distributed in particular regions of the fly integument is discussed.

Observations of the third instar larva and puparium of Chrysomya bezziana (Diptera: Calliphoridae)

Observations on the ultrastructure of the third instar larva and puparium of the Old World screw-worm fly, Chrysomya bezziana, are presented utilizing both light microscopy (LM) and scanning electron microscopy (SEM). Results of this study indicate that the shape of the intersegmental spines between the pro- and mesothorax markedly differ from other blow fly species (Chrysomya megacephala, Chrysomya rufifacies, Chrysomya nigripes, Lucilia cuprina) in being broad-based with sharp recurved tips. Other characters such as the posterior spiracles, number of papillae on the anterior spiracles, oral grooves, and posterior spiracular hairs also differ. The strong and robust mouthhooks may explain the ability of larvae to penetrate deeply into human tissues. Perforated sieve plates covered with antler-like projections were observed within the anterior spiracles of the puparium of C. bezziana. The posterior spiracular discs each bear three spiracular slits with approximately 2-microm wide openings that were viewed either open or closed by a membrane underneath. This study expands our knowledge of the fine details of the external morphology of both the third instar larva and puparium of C. bezziana, which is an obligatory myiasis-producing species in many regions. A key to differentiate the third instar of C. bezziana from other blow flies in Thailand is given.

Ultrastructure of adhesive device in fly in families calliphoridae, muscidae and sarcophagidae, and their implication as mechanical carriers of pathogens

The ultrastructure of adhesive device or the pulvilli, pad-like structure between the tarsal claws of the legs, is presented in the blowfly (Calliphoridae), housefly and relatives (Muscidae), and flesh fly (Sarcophagidae) through scanning electron microscopy. The blowfly species were Chrysomya chani, Chrysomya nigripes, Chrysomya pinguis, and Chrysomya villeneuvi; while those of the housefly and relatives were Musca domestica and Hydrotaea chalcogaster, respectively. Fresh fly species included Boettcherisca peregrina and Liosarcophaga dux. Numerous tenent setae were observed on the ventral side of the pulvilli. Two features of the tip of the tenent setae were found as a spatula-like (in C. chani, C. pinguis, C. nigripes, M. domestica, H. chalcogaster, B. peregrina, and L. dux) and spoon-like tip (in C. villeneuvi). Transmission electron microscopy of the tenent setae in M. domestica revealed the electron-lucent centrally located, suggesting an adhesive substance. These results provided anatomical information that allow us to not only understand the successful attachment of flies to smooth surfaces but also clarify their role as a mechanical carrier of microorganisms.

Morphological observation of puparia of Chrysomya nigripes (Diptera: Calliphoridae) from human corpse

This article presents Chrysomya nigripes Aubertin as a blow fly species of forensic importance in Thailand, and morphological observation of fly puparia using scanning electron microscopy (SEM). Morphologically, we focused on the characteristics of puparia used to accurately identify fly species. Numerous puparia of C. nigripes were found aggregated, adhering side by side, on the tibia of a skeletonized corpse, which was recovered from a forested area of Chiang Mai, northern Thailand. In the triangular shape of the anterior end of the puparia, three thoracic segments and broad hairy patches beginning dorsolaterally at the sixth segments were distinguishing characteristics. This study, showing pupariation of the flies along the bone of a corpse as well as morphological features, provides important guidance in identifying C. nigripes puparia. A key to differentiate puparia of C. nigripes from the other flies of forensic importance in Thailand is given.

Larval Morphology of <I>Chrysomya nigripes</I> (Diptera: Calliphoridae), a Fly Species of Forensic Importance

The morphology of all instars of Chrysomya nigripes Aubertin, a blow fly species of forensic importance, is presented with the aid of both light microscopy and scanning electron microscopy (SEM). Morphological features of the cephalopharyngeal skeleton, anterior spiracle, posterior spiracle, and dorsal spines between the prothorax and mesothorax are highlighted. No consistent features were found, even using SEM, for distinguishing the first instar of C. nigripes from that of Chrysomya megacephala (F.) or Chrysomya rufifacies (Macquart), two other commonly associated blow fly species in corpses in Thailand. Several features observed in second and third instars proved to be valuable characteristics for separating these species.

Larval Morphology of Chrysomya nigripes (Diptera: Calliphoridae), a Fly Species of Forensic Importance

The morphology of all instars of Chrysomya nigripes Aubertin, a blow fly species of forensic importance, is presented with the aid of both light microscopy and scanning electron microscopy (SEM). Morphological features of the cephalopharyngeal skeleton, anterior spiracle, posterior spiracle, and dorsal spines between the prothorax and mesothorax are highlighted. No consistent features were found, even using SEM, for distinguishing the first instar of C. nigripes from that of Chrysomya megacephala (F.) or Chrysomya rufifacies (Macquart), two other commonly associated blow fly species in corpses in Thailand. Several features observed in second and third instars proved to be valuable characteristics for separating these species.

Differentiation of the Third Instar of Forensically Important Fly Species in Thailand

Differentiation of the third instar of forensically important fly species in Thailand was performed using light microscopy, based on their morphological criteria for fly identification. Four species of the family Calliphoridae [Chrysomya rufifacies (Macquart), Chrysomya megacephala (F.), Chrysomya nigripes Aubertin, and Lucilia cuprina (Wiedemann)] and two species of the family Muscidae [Musca domestica L. and Hydrotaea (=Ophyra) spinigera Stein] were examined in this study, with the features of the anterior spiracles, dorsal spines between the prothorax and mesothorax, and posterior spiracles being emphasized. The comparisons, presented herein, should be helpful for forensic practitioners to readily distinguish the third instars of fly species found associated with human cadavers, before their use for further forensic investigations.