Effects of population variations and temperature on Chrysomya megacephala (Diptera: Calliphoridae) development: implications for estimating the postmortem interval.

Forensic entomology requires knowledge of the developmental rates of the species that colonize a body after death to estimate the postmortem interval (PMI). These developmental rates may vary depending not only on the species but also on the geographic location due to population differences. Therefore, the objectives of this work were to determine the developmental duration of the forensically important fly Chrysomya megacephala under constant controlled and field condition temperatures and to compare these results, through a meta-analysis, with data reported by other authors on populations from different localities. For this, C. megacephala colonies were established in the laboratory, and the duration of the life cycle was studied at two controlled temperatures (25 °C and 27 °C) and field conditions (27.5 ± 3.2 °C). Analysis of variance was performed to determine differences in developmental time and larval length between constant laboratory temperatures and field conditions. A generalized linear model was performed with predictor variables extracted from the literature (diet, relative humidity, latitude, longitude) to evaluate the effect of population variation on developmental times. The results showed significant differences in developmental times between 25 and 27 °C. As expected, the complete life cycle of C. megacephala was shorter at 27 °C. Finally, the meta-analysis suggested differences between the developmental times of different populations, based on temperature and geographic location. The results of this study provide fundamental developmental data to use C. megacephala in PMI estimations. Finally, we suggest that, when making expert reports, information from local populations should be used to determine a more accurate and reliable PMI.

Forensic significance of Roundup Full® II effect on the development of Dermestes maculatus (Coleoptera: Dermestidae) and Lucilia sericata (Diptera: Calliphoridae).

The most widely used herbicides in the world are glyphosate formulations. These have been used in self-poisoning and accidental poisoning. Dermestes maculatus (Coleoptera: Dermestidae) and Lucilia sericata (Diptera: Calliphoridae) are associated with decomposing human and animal remains. The aim of the study was to evaluate Roundup Full® II effects on saprophagous insects´ development. For this, green bottle flies and hide beetles were exposed to different doses of the herbicide. The duration of life cycles was recorded by observations made every 8 and 24 h. To register possible variations in size, external morphology, and female proportion, specimens of all developmental stages were collected. The duration of developmental stages in L. sericata did not change due to the herbicide, but this reduced all size parameters of puparium with pupae inside, the weight of L1, and of adults. The larval stage and total duration in D. maculatus were decreased by the highest dose, while size parameters of hide beetles remained unchanged at all development stages. In both flies and beetles, there was no evidence of abnormal external morphology and the female proportion did not change in response to the herbicide. Finally, this study exhibited an impact on L. sericata and D. maculatus, so the possibility that the herbicide may affect other forensically important species should be considered, but more work is needed. Taxonomic determination could be done as usual; however, caution should be exercised when estimating the postmortem interval. This study represents the first to analyze a glyphosate-based herbicide associated with Forensic Entomology.

Postmortem Interval Estimation and Validation Through a Comparative Study of South American Flies Reared in the Field Versus Laboratory Conditions.

Studies under constant temperatures are the most common to estimate the Postmortem Interval (PMI). It is imperative that forensic sciences have data from studies carried out in the field. Therefore, this work aims to: (1) evaluate the parameters (weight, length, development time) associated with the life cycles of Lucilia ochricornis (Wiedemann) (Diptera: Calliphoridae) and Lucilia purpurascens (Walker) under experimental conditions in the field considering fluctuating temperatures, and (2) compare these results with those known and published by the same authors for cultures realized in the laboratory under constant temperatures; which will permit us to contrast the most widely used existing methodologies for forensic application in estimating the minimum postmortem interval (PMImin). For each season of the year, cultures of both species were made in the field, collecting information on temperature, humidity, and photoperiod to perform laboratory cultures, later comparing: development time, length, weight, and Accumulated Degree-Hours (ADH) in both types of cultures. Methods for estimating the PMI were obtained and validated with the information of the cultures grown in the field. The two types of cultures showed differences between each other for both species. The forensic use methods to estimate PMI were enhanced and their precision increased when maximum larval length data were used, and it was also concluded that feeding larval stages are the most accurate to be used in making estimates because the larva is growing. The estimation of the PMI through the use of necrophagous flies development remains reliable for obtaining the PMImin.

Estimation of the Postmortem Interval Through the Use of Development Time of Two South American Species of Forensic Importance of the Genus Lucilia (Diptera: Calliphoridae).

Obtaining the specific development time of each species of forensic interest is crucial for the estimation of an accurate and reliable Minimum Postmortem Interval (PMImin). In Argentina, Lucilia ochricornis (Wiedemann) and Lucilia purpurascens (Walker) (Diptera: Calliphoridae) were masked under the name Lucilia cluvia (Walker) for a long time still in forensic expertise. For this reason, the objective of this work is to deepen the study of the development time of these species and utilize this relevant information in the generation of different associated methods that can be used in forensics to estimate the PMI. Immature stages of L. ochricornis and L. purpurascens were reared in a brood chamber according to the following temperature treatments: 13.4, 15.1, 22.3, and 23.6°C. The development time of each stage/state of these flies was recorded as well as the resulting accumulated degree-hours (ADH), to build isomorphen diagrams and thermal summation models for each species. The development time and ADH were different between both species and their development stages. On the other hand, the methods provided for estimating PMImin provide the forensic entomologist more tools to reach accurate and reliable estimates.

Dermestes maculatus (Coleoptera: Dermestidae) development under fluoxetine effect using two drug administration models.

Fluoxetine is a selective serotonin reuptake inhibitor, commonly used for the treatment of a variety of psychopathological conditions. As such, fluoxetine may be expected to appear in clinical and forensic cases. Dermestes maculatus De Geer (Coleoptera: Dermestidae) has been recognized as a relevant component of the insect fauna associated with decomposing human and animal remains. Experiments were conducted to study the effect of fluoxetine on developing D. maculatus using two-drug administration models: a non-living animal model (pork muscle) and a living one (Sus scrofa L. pigs). We assessed the duration of immature stages and total life cycle, as well as morphological parameters (body length, cephalic width, and weight). The effect of fluoxetine was studied at an overdose concentration: In the non-living animal model the drug was mixed with macerated pork muscle (2000 mg/kg) and in the living animal model, pigs were given the drug orally (833 mg/kg). A control was used for each model. Daily observations were performed from the beginning to the end of the experiments. GC-MS was used for drug detection and quantification. There were no statistically significant differences in the duration of immature stages, life cycle, larval mortality, morphological parameters, or sex ratio, between treatment and control, regardless of the drug administration model. Given that fluoxetine had no detectable effect on the development of D. maculatus, detection of this drug in forensic situations would not compromise the accuracy of PMI estimations.

Contributions to the Estimation of the Postmortem Interval Through the Length and Body Weight of Two Indigenous Species of South America: Lucilia ochricornis (Diptera: Calliphoridae) and Lucilia purpurascens.

The early arrival and colonization of species belonging to the family Calliphoridae (Insecta: Diptera) on a corpse represent one of the most reliable means of estimating minimum postmortem interval (PMImin). However, information on the development and life cycles of some Argentine species in this family is not complete. The objective of this work was to contribute new information regarding the larval body size of neotropical species that allow, through the construction of forensic methods, the estimation of a more precise and specific PMImin. This work was conducted on laboratory cultures of larvae of Lucilia ochricornis (Wiedemann) and Lucilia purpurascens (Walker) using as average temperatures: 13.4, 15.1, 22.6, and 23.3°C, which represent the four seasons of the year for the province of Salta. With this information, we constructed isomegalen diagrams and growth models for the obtained variables of larval length and body weight. The mean values of length and body weight differ between both species, indicating that L. purpurascens exceeded L. ochricornis in both variables. In contrast, within each species the mean length and weight remained unchanged between culture temperatures for the three larval instars. Isomegalen diagrams can be used for the entire range of temperatures worked in the laboratory, but the body size entered is approximate. The growth models allow the use of point data but are specific for each culture temperature used.

An initial study of insect succession on pig carcasses in open pastures in the northwest of Uruguay.

The succession of carrion fauna and the decomposition stages were studied in summer for two years in open pastures of the Department of Paysandú, Uruguay (32°21'5″S; 58°3'37″). Each year, three pig carcasses (Sus scrofa) were placed in wire mesh cages, 100 m apart from each other. Each carcass was surrounded by pitfall traps, and a modified Malaise trap was placed above. Daily samplings were carried out to collect the insects present in the carcasses and the traps, and body and environmental temperatures were measured. The composition of carrion entomofauna and the decomposition patterns were markedly similar in both sampling years. The decomposition process was rapid, that is, the time to reach the remains stage was 7 days in all treatments, both years. The mosaic nature of decomposition was observed. Both Chrysomya albiceps (Wiedemann) and Cochliomyia macellaria (Fabricius) behaved as primary invaders of the carcasses, although the former species and its larvae were dominant. The first adult blow flies emerged after 6 days. The remains stage was characterized mainly by both adults and larvae of Dermestes maculatus. This preliminary study represents the first contribution to the knowledge of cadaveric succession in Uruguay. Its seasonal replication supports the succession pattern obtained.

Depressions of Dermestes maculatus (Coleoptera: Dermestidae) on Bones Could be Pupation Chambers.

Dermestes maculatus DeGeer beetles feed and can leave marks on bones. This species is of medical, veterinary, and forensic importance. The aim of this work was to determine if the depressions produced on bones by these beetles could be product of pupation. We studied under controlled conditions the behavior of D. maculatus larvae maintained at 24°C ± 3°C, 52.7% average relative humidity, and a 12:12-hour light/dark cycle and fed on pig carcasses. Observations were made, and photographs were taken. The results indicated that depressions produced by D. maculatus were consequence of feeding or pupation. This is the first time that pupation chambers on bones are confirmed.

Determination of fluoxetine in Dermestes maculatus (Coleoptera: Dermestidae) by a spectrophotometric method.

The aims of this study were to detect and quantify fluoxetine, an antidepressant, from entomological samples. Larvae, pupae and adults of Dermestes maculatus (Coleoptera, Dermestidae) were reared on pig muscle previously treated with fluoxetine. The concentration selected, 2000mg/kg, emulates a fluoxetine overdose lethal to humans and laboratory animals. Thirty larvae on the fourth and fifth stages, 50 adults and several exuviae were analyzed for fluoxetine content. Detection of fluoxetine was performed by UV spectrophotometry at 270 and 277nm. All developmental stages of D. maculatus and exuviae were positive for fluoxetine. We also quantified the drug and no significant differences were found either between the days or the stages in the general model, but at 277nm a tendency of the concentration to decrease with time was observed. Concentrations of fluoxetine at 277nm were almost equal or greater than those at 270nm. This is the first study to detect and quantify fluoxetine from entomological samples and, in particular, from D. maculatus beetles.

Primer registro de Talaromyces udagawae relacionado con cuerpos humanos en descomposición, en suelo de la Argentina / First record of Talaromyces udagawae in soil related to decomposing human remains in Argentina

The morphologic features of Talaromyces udagawae Stolk and Samson are here described and illustrated. This teleomorphic Ascomycota fungus was isolated from soil obtained in Buenos Aires province (Argentina) from beneath a human cadaver in an advanced state of decomposition. After washing and serial dilution of the soil along with moist-chamber techniques for fungal cultivation, T. udagawae formed very restricted colonies of bright yellow color on different growth media with 8-ascospored asci. The ascospores were ellipsoidal and ornamented. The anamorphic state was not observed. Molecular-genetic techniques identified the species. The present record is the first of the species in Argentina, pointing it as a tool to identify soils where cadaver decomposition occurs.

Se describen e ilustran las características morfológicas de Talaromyces udagawae Stolk y Samson. Se aisló el estado teleomórfico de este hongo Ascomycota de suelo obtenido en la provincia de Buenos Aires (Argentina), por debajo de un cadáver humano en avanzado estado de descomposición. Las muestras de suelo fueron analizadas mediante lavado, dilución seriada y cámaras húmedas, técnicas ampliamente usadas para el estudio de hongos de suelo. T. udagawae formó colonias muy restringidas de color amarillo brillante en diferentes medios de cultivo, con ascos con 8 ascosporas. Las ascosporas eran elipsoidales y ornamentadas. No fue hallado el estado anamórfico. La especie también fue identificada mediante técnicas moleculares. El presente registro es el primero de la especie en la Argentina y el único que la postula como herramienta para identificar suelos donde ocurre una descomposición cadavérica.

First record of Talaromyces udagawae in soil related to decomposing human remains in Argentina.

The morphologic features of Talaromyces udagawae Stolk and Samson are here described and illustrated. This teleomorphic Ascomycota fungus was isolated from soil obtained in Buenos Aires province (Argentina) from beneath a human cadaver in an advanced state of decomposition. After washing and serial dilution of the soil along with moist-chamber techniques for fungal cultivation, T. udagawae formed very restricted colonies of bright yellow color on different growth media with 8-ascospored asci. The ascospores were ellipsoidal and ornamented. The anamorphic state was not observed. Molecular-genetic techniques identified the species. The present record is the first of the species in Argentina, pointing it as a tool to identify soils where cadaver decomposition occurs.

The Effect of Temperature and Laboratory Rearing Conditions on the Development of Dermestes maculatus (Coleoptera: Dermestidae).

Experiments were conducted to study the life cycle of Dermestes maculatus and to establish the total developmental time and the developmental time of immature stages, in relation with six different temperatures. We also analyzed the variations in size, morphology, and other indicators of temporal variation during life cycle of D. maculatus, in relation with temperature. One hundred larvae were selected per experiment, reared individually. The remaining larvae were reared to evaluate and establish temporal variations among the instars (length, cephalic width, and dry weight). In all trials, survivorship was greater than 50% and seven larval instars were found. Data of the average developmental time of immature stages and of the total cycle, at different temperatures, are provided. This is of relevance when estimating particularly, a minimum PMI. No relation between morphometric parameters and temperature was found, suggesting that other random factors may have been involved. Thus, this indicates that the method of isomegalen diagrams could not be used for calculating PMI.

Modification of postmortem wounds by Dermestes maculatus (Coleoptera: Dermestidae) activity: A preliminary study.

The purpose of this analysis was to evaluate the effects of insect activity on hacking trauma which was inflicted with a small cleaver and a razor blade under controlled conditions. Three pig hooves were each subjected to a blow with a small cleaver and a cut with a razor blade prior to insect exposure. We used Dermestes maculatus DeGeer 1774 species. These beetles made principally depressions and destruction on both wounds, and bites were observed on the edges of the wounds. As time passed and insects fed and refuge, chop marks were deformed and disappeared, taking this less than a month. Thus, D. maculatus could mask postmortem wounds and probably premortem wounds, and so the cause of death.

Associational Patterns of Scavenger Beetles to Decomposition Stages.

Beetles associated with carrion play an important role in recycling organic matter in an ecosystem. Four experiments on decomposition, one per season, were conducted in a semirural area in Bahía Blanca, Argentina. Melyridae are reported for the first time of forensic interest. Apart from adults and larvae of Scarabaeidae, thirteen species and two genera of other coleopteran families are new forensic records in Argentina. Diversity, abundance, and species composition of beetles showed differences between stages and seasons. Our results differed from other studies conducted in temperate regions. Four guilds and succession patterns were established in relation to decomposition stages and seasons. Dermestidae (necrophages) predominated in winter during the decomposition process; Staphylinidae (necrophiles) in Fresh and Bloat stages during spring, summer, and autumn; and Histeridae (necrophiles) and Cleridae (omnivores) in the following stages during those seasons. Finally, coleopteran activity, diversity and abundance, and decomposition rate change with biogeoclimatic characteristics, which is of significance in forensics.

Marks caused by the scavenging activity of Necrobia rufipes (Coleoptera: Cleridae) under laboratory conditions.

Insects are an important group involved in carrion consumption and are thus of forensic interest. In the laboratory we studied the taphonomic marks that Necrobia rufipes (Cleridae) can produce. Pig trotters were exposed to adult beetles at 21 ± 3 °C and 12:12 h day/night cycle. We made observations and took photographs every 4-5 days for 12 months. Marks were noted after a month. We found scratches, pits, holes, and tunnels in several kinds of tissue such as integumental, connective and muscular. This work contributes preliminary data of significant application in biology, ecology, anthropology and forensics. Until now, no study has provided taphonomic information with N. rufipes.

Trophic roles of scavenger beetles in relation to decomposition stages and seasons

Carcasses represent a trophic and reproductive resource or shelter for arthropods, which are a representative component of the decomposition process. Four experiments, one per season, were conducted in a semi-rural area of Bahía Blanca, Argentina, to study the trophic roles of cadaveric beetles, evaluating the abundance, composition and dominance during all decomposition stages and seasons. Species of necrophagous, necrophilous and omnivorous habits were found. Abundance, composition and dominance of beetles in relation to their trophic roles changed according to seasons and decomposition stages. Guilds and patterns of succession were established in relation to those periods. Trophic roles could be an indicator of beetle associations with decomposition stages and seasons.

An illustrated key to and diagnoses of the species of Staphylinidae (Coleoptera) associated with decaying carcasses in Argentina.

A key to 24 Staphylinidae species associated with decaying carcasses in Argentina is presented, including diagnoses, illustrations, distributional and bionomical data for these species. This article provides a table of all species associated with carcasses, detailing the substrate from which they were collected and geographical distribution by province. All 24 Staphylinidae species recorded are grouped into three subfamilies: Aleocharinae (three species of Aleochara Gravenhorst and one species of Atheta Thomson), Oxytelinae (one species of Anotylus Thomson) and Staphylininae (18 species, two belonging to the tribe Xantholinini and 16 species belonging to the tribe Staphylinini). A discussion is presented on the potential forensic importance of some species collected on human and pig carcasses.

Ultramorphological characteristics of mature larvae of Nitidula carnaria (Schaller 1783) (Coleoptera: Nitidulidae), a beetle species of forensic importance.

Beetles of the genus Nitidula Fabricius are forensically important, and their adults and larvae have been found associated with human corpses and animal carcasses in many places of the world. The external morphology of the larvae of Nitidula carnaria (Schaller 1783) was examined by scanning electron microscopy (SEM) to provide a description enabling identification of this forensically important species. The ultrastructure of the head was examined, antennae, mandibles, epipharynx, maxillary and labial palpi, spiracles, thorax, legs, and abdominal segments (especially segments 9 and 10); the tegument was also emphasised in this examination. Several types of sensilla were observed on the maxillary and labial palpi, including sensilla basiconica, sensilla styloconica, and perhaps a different type of sensilla digitiformia. In abdominal segment 10, a sensilla campaniformia was observed. Two types of plates were noticed in the abdominal tegument. The characteristics described here can be used to identify this species. No other study of the ultrastructure of Nitidulidae larvae is available for comparison. This is the first report of N. carnaria in carcasses in Chile.

First report of myiasis caused by Cochliomyia hominivorax (Diptera: Calliphoridae) in a diabetic foot ulcer patient in Argentina.

Myiasis is usually caused by flies of the Calliphoridae family, and Cochliomyia hominivorax is the etiological agent most frequently found in myiasis. The first case of myiasis in a diabetic foot of a 54-year-old male patient in Argentina is reported. The patient attended the hospital of the capital city of Tucumán Province for a consultation concerning an ulcer in his right foot, where the larval specimens were found. The identification of the immature larvae was based on their morphological characters, such as the cylindrical, segmented, white yellow-coloured body and tracheas with strong pigmentation. The larvae were removed, and the patient was treated with antibiotics. The larvae were reared until the adults were obtained. The adults were identified by the setose basal vein in the upper surface of the wing, denuded lower surface of the wing, short and reduced palps, and parafrontalia with black hairs outside the front row of setae. The main factor that favoured the development of myiasis is due to diabetes, which caused a loss of sensibility in the limb that resulted in late consultation. Moreover, the poor personal hygiene attracted the flies, and the foul-smelling discharge from the wound favoured the female's oviposition. There is a need to implement a program for prevention of myiasis, in which the population is made aware not only of the importance of good personal hygiene and home sanitation but also of the degree of implication of flies in the occurrence and development of this disease.

Taphonomic marks on pig tissue due to cadaveric Coleoptera activity under controlled conditions.

The aim of this work was to study taphonomic marks that cadaveric coleopteran can produce under controlled conditions. To evaluate this, pig trotters were initially exposed to adults of Dermestes maculatus De Geer at 21 ± 5°C and a 12:12-h day/night cycle. Observations were made and photographs taken every 4-5 days for 9 months. When feeding and reproducing, D. maculatus produced, in both adult and larvae stages, different types of marks such as holes, striations, scratches, and pits in several kinds of tissue such as integumental, connective, and muscular, in both their fresh and dried stages. Bite marks were also evident. The results in this study provide not only taphonomic but also biological and forensic information. This is the first time that this kind of experiment has been performed.