A Preliminary Study of Insect Succession in Al‐Ahsaa Oasis,in the Eastern Region of the Kingdom of Saudi Arabia

Rabbit carcass decomposition was examined in four seasons in Al‐Ahsaa Oasis. Decomposition rate was significantly faster in summer compared with other seasons. Fourteen insect species from four orders and seven families were recorded: Calliphoridae, Muscidae, Sarcophagidae, Formicidae, Histeridae, Dermestidae, and Tenebrionidae. Most of the fly maggots collected were Calliphoridae with only one species from both Muscidae and Sarcophagidae. Maggots of Calliphora vicina (R.‐D.) and Chrysomya albiceps (Wiedemann) colonized the carcasses in all seasons while both Ch. bezziana (Villeneuve) and Lucilia sericata (Meigen) were found in all seasons except for winter. Maggots of Chrysomya megacephala (F.) and Phormia regina (Meigen) were only collected in the summer and autumn, respectively. Four coleopteran species were collected: Hymenorus sp., Saprinus chacites (Illiger), Dermestes maculatus DeGeer and Blaps sp. One formicidian species, Pheidole megacephala (F.), was observed in all seasons. Insect richness was greater, and decomposition rate was faster in summer compared with other seasons.     

Comparison of Decomposition Rates and Faunal Colonization of Carrion in Indoor and Outdoor Environments

Abstract: Decomposition and insect colonization of pig (Sus scrofa L.) carcasses were observed over a 42‐day period inside and outside a house in a suburban region of Edmonton, Alberta. Three freshly killed pig carcasses were placed outdoors on grass and three carcasses were placed in separate rooms inside a house in a suburban residential area. The carcasses were examined and sampled regularly. Outside carcasses were colonized rapidly by Calliphora vicina (R‐D), Lucilia sericata (Meigen), Lucilia illustris (Meigen), Eucalliphora latifrons (Hough), Cynomya cadaverina (R‐D), Phormia regina (Meigen), and Protophormia terraenovae (R‐D). There was a delay of 5 days before inside carcasses were first colonized but all species except E. latifrons and L. illustris readily colonized at this time. Many more insects colonized the outside carcasses, and these were rapidly skeletonized. Inside, much fewer insects were present and decomposition was slowed and colonization extended. Dispersal patterns of postfeeding larvae inside the house on hard substrate were greatly expanded compared with those from carcasses on grass.     

Forensic Entomology and the Estimation of the Minimum Time Since Death in Indoor Cases

Eight cases that occurred indoors in which the insects played an important role in the mPMI estimation are presented. The bodies of socially isolated people and old people living alone were discovered in central Italy between June and November. mPMI ranged from a few days to several weeks. Insects were collected during the body recovery and the postmortem. Climatic data were obtained from the closest meteorological stations and from measurements performed on the site. Sarcophagidae and Calliphoridae species were present in 75% of the cases with Lucilia sericata and Chrysomya albiceps collected in 50% of the cases. Chrysomya albiceps was always found in association with Lucilia species. Scuttle flies (Phoridae) were found in 37.5% of the cases, confirming the ability of these species in indoor body colonization. We show that if sealed environment may delay, the insect arrival dirty houses may create the environment where sarcosaprophagous insects are already present.     

An Analysis of Forensically Important Insects Associated with Human Remains in Indiana,U.S.A.*

Research documenting insect colonization of human remains is limited in North America, and currently nonexistent for the American Midwest. Such research is essential for forensic entomologists to identify species of research interest in a region. In this study, we collected insects from human remains in 24 cases across Indiana from June 2016 through September 2018. We analyzed species composition across scene type and season. Eight species of blow flies were collected as larvae from human remains, with Phormia regina and Lucilia sericata as the two predominant colonizers. Phormia regina was the most numerous species collected from outdoor scenes (73.6% of total collections) while L. sericata was the most numerous from the indoor scenes (60.4% of total collections). With scene types pooled, Calliphora vicina and Cochliomya macellaria were the predominant species in the fall (55.6% and 42.2%, respectively); P. regina was the dominant colonizer in the spring (68.6%); and P. regina and L. sericata were the predominant colonizers (46.5% and 44.4%, respectively) in the summer. In addition to these findings, we confirmed the first record of Lucilia cuprina colonizing human remains in Indiana having collected this species from three cases. A single adult Chrysomya megacephala was collected from an indoor scene in southern Indiana, which represents its second collection in the state. Beetles belonging to the families Staphylinidae, Silphidae, Histeridae, Cleridae, Trogidae, Dermestdae, and Nitidulidae were also collected from two outdoor scenes. This study provides important baseline data for forensic entomologists in Indiana, as well as surrounding states with similar environments.     

Correlation of Molecular Expression with Diel Rhythm of Oviposition in Calliphora vicina (Robineau-Desvoidy) (Diptera: Calliphoridae) and Implications for Forensic Entomology

This study explored the molecular mechanisms potentially underlying blow fly nocturnal oviposition. A behavioral study revealed that Calliphora vicina (Robineau-Desvoidy) (Diptera: Calliphoridae) possesses a diel rhythm of oviposition in light under 12:12 light/dark conditions. Reversal to 12:12 dark/light resulted in oviposition behavior changing to align with the adjusted regime in most females, but four of 59 experimental females lacked a diel rhythm of oviposition (were arrhythmic). Real-time PCR was used to monitor the molecular expression levels of known circadian genes per and tim in C. vicina to determine whether gene expression and behavior correlated. As with behavior, reversing light/dark conditions changed rhythmic gene expression to align with an adjusted light regime. This suggests that although it is unlikely that C. vicina will colonize dead bodies at night, arrhythmic females and oviposition in the dark was demonstrated.     

Decomposition and Arthropod Succession in Whitehorse,Yukon Territory,Canada

Forensic arthropod succession patterns are known to vary between regions. However, the northern habitats of the globe have been largely left unstudied. Three pig carcasses were studied outdoors in Whitehorse, Yukon Territory. Adult and immature insects were collected for identification and comparison. The dominant Diptera and Coleoptera species at all carcasses were Protophormia terraneovae (R‐D) (Fam: Calliphoridae) and Thanatophilus lapponicus (Herbst) (Fam: Silphidae), respectively. Rate of decomposition, patterns of Diptera and Coleoptera succession, and species dominance were shown to differ from previous studies in temperate regions, particularly as P. terraenovae showed complete dominance among blowfly species. Rate of decomposition through the first four stages was generally slow, and the last stage of decomposition was not observed at any carcass due to time constraints. It is concluded that biogeoclimatic range has a significant effect on insect presence and rate of decomposition, making it an important factor to consider when calculating a postmortem interval.     

Alteration of expirated bloodstain patterns by Calliphora vicina and Lucilia sericata (Diptera: Calliphoridae) through ingestion and deposition of artifacts

Abstract: Bloodstain pattern analysis can provide insight into a sequence of events associated with a violent crime. However, bloodstain pattern analysis can be confounded by the feeding activity of blow flies. We conducted two laboratory experiments to investigate the relationships between Lucilia sericata (green bottle fly) and Calliphora vicina (blue bottle fly), expirated bloodstains, and pooled bloodstains on a range of surfaces (linoleum, wallpaper, textured paint). C. vicina and L. sericata changed bloodstain pattern morphology through feeding and defecation. They also deposited artifacts in rooms where blood was not present originally. Chemical presumptive tests (Hemastix^®, phenolphthalein, leucocrystal violet, fluorescein) were not able to differentiate between insect artifacts and bloodstains. Thus, C. vicina and L. sericata can confound bloodstain pattern analysis, crime scene investigation, and reconstruction. Crime scene investigators should be aware of these fundamental behaviors, and the effects that blow flies can have on expirated and pooled bloodstain patterns.     

A Checklist of Calliphoridae Blowflies (Insecta,Diptera) Associated with a Pig Carrion in Central Brazil*

Abstract: Although the Cerrado is the second major Brazilian biome, few studies have been undertaken about its entomofauna. Blowflies have an important role in forensic entomology, helping in the determination of postmortem intervals. The main goal of this exploratory study was to identify and to catalog the blowfly species associated with a pig carcass. The study was conducted in a pasture in Brasília, Distrito Federal. A pig (Sus scrofa) was killed with a .22 caliber shot in the frontal region of the head. Adult blowflies were surveyed daily from June 1 through 30, 2004. A total of 14,910 adult calliphorids were collected, representing eight species: Chrysomya albiceps, C. megacephala, Cochliomyia macellaria, Chloroprocta idioidea, Hemilucilia semidiaphana, H. segmentaria, Lucilia cuprina, and L. eximia. C. albiceps was the most frequent species, amounting to 94.76% of the catch. Five decomposition stages were observed, and for calliphorids, the most attractive stage was the bloated one.     

Internal Morphological Analysis for Age Estimation of Blow Fly Pupae (Diptera: Calliphoridae) in Postmortem Interval Estimation*

Abstract: Larvae and pupae of blow fly species are frequently used in postmortem interval estimation, their age indicating minimum time since death. Most studies have considered age estimation of larvae, neglecting study of pupae. Relative development of external pupal features is useful, but there are also internal changes during metamorphosis that may be indicators of age, utilizing histological techniques. This study aimed to optimize preservation and histological analysis of blow fly pupae, specifically Calliphora vicina Robineau‐Desvoidy and Lucilia sericata (Meigen), and to examine internal features with potential for age estimation. Effect of hot‐water‐killing and different preservatives were examined. It was determined that blow fly pupae should be pierced through the three tagma, hot‐water‐killed, and preserved in 80% ethanol as the optimal preservation for subsequent analyses. Hematoxylin and eosin stained pupal sections revealed differences in brain and thoracic muscle development throughout the pupal stage with potential for age estimation.     

The Effect of Clothing on the Rate of Decomposition and Diptera Colonization on Sus scrofa Carcasses

Twenty Sus scrofa carcasses were used to study the effect the presence of clothing had on decomposition rate and colonization locations of Diptera species; 10 unclothed control carcasses were compared to 10 clothed experimental carcasses over 58 days. Data collection occurred at regular accumulated degree day intervals; the level of decomposition as Total Body Score (TBS_surf), pattern of decomposition, and Diptera present was documented. Results indicated a statistically significant difference in the rate of decomposition, (t₄₂₇ = 2.59, p = 0.010), with unclothed carcasses decomposing faster than clothed carcasses. However, the overall decomposition rates from each carcass group are too similar to separate when applying a 95% CI, which means that, although statistically significant, from a practical forensic point of view they are not sufficiently dissimilar as to warrant the application of different formulae to estimate the postmortem interval. Further results demonstrated clothing provided blow flies with additional colonization locations.     

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.     

Forensic Use of A Subtropical Blowfly: The First Case Indicating Minimum Postmortem Interval (mPMI) in Southern Brazil and First Record of Sarconesia Chlorogaster from a Human Corpse

Southern Brazil is unique due to its subtropical climate. Here, we report on the first forensic entomology case and the first record of Sarconesia chlorogaster (Wiedemann) in a human corpse in this region. Flies' samples were collected from a body indoors at 20°C. Four species were found, but only Chrysomya albiceps (Wiedemann) and S. chlorogaster were used to estimate the minimum postmortem interval (mPMI). The mPMI was calculated using accumulated degree hour (ADH) and developmental time. The S. chlorogaster puparium collected was light in color, so we used an experiment to establish a more accurate estimate for time since initiation of pupation where we found full tanning after 3 h. Development of C. albiceps at 20°C to the end of the third instar is 7.4 days. The mPMI based on S. chlorogaster (developmental time until the third instar with no more than 3 h of pupae development) was 7.6 days.     

A Preliminary Examination of Differential Decomposition Patterns in Mass Graves

Five pairs of mass graves, each containing carcasses of 21 rabbits, were used to examine differential decomposition at four locations within the burial: surface, deep, mid‐outer, and core. Every 100 accumulated degree days (ADD), a pair of graves was exhumed, and total body score (TBS) and internal carcass temperature of each rabbit were recorded. Decomposition did not differ for core‐ and deep‐positioned carcasses (p = 0.13); differences were significant (p < 0.001) for all other location comparisons. Decomposition occurred fastest in shallow carcasses, followed by mid‐outer carcasses; deep and core carcasses decomposed slowest and at rates not significantly different from one another. Adipocere formation was minimal and confined to deep carcasses. Carcass location within the mass grave significantly influenced internal carcass temperature (p < 0.001); a mean internal temperature difference of ca. 1°C existed between deep and shallow carcasses (30 cm apart). Effects of mass compactness and oxygenation require further investigation.     

An Experiment to Characterize the Decomposer Community Associated with Carcasses (Sus scrofa domesticus) on Oahu,Hawaii

Pig (Sus scrofa domesticus) carcasses were decomposed on the soil surface of a terrestrial habitat on the island of Oahu, Hawaii to begin characterizing the decomposer community. Results showed that carcasses can decompose rapidly on Oahu, primarily due to the activity of fly larvae, with ~80% of mass lost by 8 days (~220 ADD) postmortem. Scavenging was conducted exclusively by the small Indian mongoose (Herpestes javanicus), first feeding on larvae then feeding on the remains. Carcasses were habitats of warm temperature, little to no oxygen, slightly acidic/neutral pH, and high sodium concentration. Larval masses selected for a microbial community comprised of multiple bacterial taxa from phyla Firmicutes and Proteobacteria, particularly genera Clostridium, Proteus, and Providencia. These larval masses were well established from 3 to 8 days (~90 to ~220 ADD) postmortem. These data provide helpful, novel insight into the structure and activity of carcass decomposer communities on Oahu.     

Preliminary Observations of Arthropods Associated with Buried Carrion on Oahu

Several studies in Hawaii have focused on arthropod succession and decomposition patterns of surface remains, but the current research presents the first study to focus on shallow burials in this context. Three domestic pig carcasses (Sus scrofa L.) were buried at the depths of 20–40 cm in silty clay loam soil on an exposed ridge on the leeward side of the volcanically formed Koolau Mountain Range. One carcass was exhumed after 3 weeks, another after 6 weeks, and the last carcass was exhumed after 9 weeks. An inventory of arthropod taxa present on the carrion and in the surrounding soil and observations pertaining to decomposition were recorded at each exhumation. The longer the carrion was buried, the greater the diversity of arthropod species that were recovered from the remains. Biomass loss was calculated to be 49% at the 3‐week interval, 56% at the 6‐week interval, and 59% at the 9‐week interval.     

Effect of Temperature on Six Different Developmental Landmarks within the Pupal Stage of the Forensically Important Blowfly Calliphora vicina (Robineau‐Desvoidy) (Diptera: Calliphoridae)

This study investigates the pupal development times of the blow fly Calliphora vicina, which were studied in the laboratory at six different constant temperatures (15, 20, 23, 25, 28, and 30°C each ± 1°C). Lower thresholds (t_L) for development were estimated from the linear regression of the developmental rates on each temperature. These data have made it possible to calculate the accumulated degree days (ADD) necessary for C. vicina to complete the larval stage and to achieve adult emergence. The minimal duration of development from oviposition to adult emergence was found to be inversely related to temperature. Additionally, six landmarks in pupal development are showed, and for each of the landmarks, the ADD value was calculated for every rearing temperature involved. These data assist in calculating the duration of the pupal stage based on morphological characteristics and would be of great value for future forensic entomological casework.     

Volatile Emission of Decomposing Pig Carcasses (Sus scrofa domesticus L.) as an Indicator for the Postmortem Interval

This study aimed at correlating selected carcass borne volatile organic compounds (VOCs) with the postmortem interval (PMI). Selected volatiles should 1st be reliably emitted during vertebrate decay, 2nd be emitted at high concentrations, and 3rd show a reproducible quantitative dynamic during the decaying process. Four pigs (Sus scrofa domesticus L.) were placed in a deciduous forest in different seasons and volatiles emitted during the decaying process were sampled. Seventeen compounds were identified and quantified by GC-MS. Electrophysiological experiments on the antenna of female Calliphora vicina and additional data of Dermestes maculans were used as an evolutionary tuned information filter to evaluate the 1st criterion. The relative quantitative emission of hexanal, nonanal, dimethyl disulfide, dimethyl trisulfide, 1-butanol, and phenol were correlated with the PMI, and the observed stages of decay and the limitations of this model were discussed.     

An Evaluation of Differentially Spliced Genes as Markers of Sex for Forensic Entomology,,

Blow flies (Calliphoridae) are important medically and economically and are commonly used in forensics as temporal markers in death investigations. While phenotypic traits in adult flies can be sexually dimorphic, sex identification in immatures is difficult. Consequently, little is known about how sex may result in developmental disparities among sexes even though there are indications that they may be important in some instances. Since genetic mechanisms for sex are well studied in model flies and species of agricultural and medical importance, we exploit the sex-specifically spliced genes transformer (tra) and doublesex (dsx) in the sex determination pathway to optimize a sex identification assay for immatures. Using known primer sets for tra and with a novel one for dsx, we develop PCR assays for identifying sex in four forensically relevant Calliphoridae species: Lucilia sericata (Meigen), Lucilia cuprina (Wiedemann), Cochliomyia macellaria (Fabricius), and Chrysomya rufifacies (Macquart) and evaluated their performance. Band detection rates were found to range from 71 to 100%, call rates ranged from 90 to 100%, and no error was found when bands could be called. Such information is informative for purposes of testimony and in preparation for development studies. The developed assays will assist in further differentiating sexually dimorphic differences in development of the Calliphoridae and aid in more accurately estimating insect age when age predictive markers (size, development time, molecular expression) are sexually dimorphic.     

Spread and habitat selection of Chrysomya albiceps (Wiedemann) (Diptera Calliphoridae) in Northern Italy: forensic implications

Abstract: Habitat selection exploited by Chrysomya albiceps during its initial spread in Northern Italy was analyzed in relation to landscape structure. The results of two short studies and a case report are here discussed. C. albiceps was not found on experimental pig carcasses in the urban area of Pavia. It was missing in the woody mountains surrounding Lecco, but it was found in the same area, at a lower altitude, within the typical “urban sprawl” landscape. It was then recorded in a natural reserve, among a rich carrion‐fly population. Indications coming from habitat selection suggest that C. albiceps has not yet saturated its potential ecological niche in newly colonized areas of Northern Italy. Factors like temperature, altitude, and interspecific competition can act as limiting factors, affecting habitat selection and distribution in newly colonized areas; the influence of those factors has to be taken into account for forensic purposes.     

A Preliminary Forensic Entomological Study of Beetles (Coleoptera) in Interior Alaska,USA*

Forensic entomology uses knowledge of arthropod ecology to help solve crimes. There has been no published forensic entomological research in Alaska. We used one piglet carcass split in half to create two carcass plots in Fairbanks (~64.8°N, subarctic) that were sampled over a period of 59 days in 2019. Four pitfall traps were placed around each carcass, and four similarly arranged pitfall traps were placed 40 m distant as controls. Traps were emptied approximately weekly covering the first four stages of decomposition. We focused on adults of the larger-bodied (>9 mm) families and subfamilies of Coleoptera: Staphylinidae (subfamily Staphylininae), Carabidae, and Silphidae. A total of 621 specimens were collected and processed: 29 staphylinines, 210 carabids, and 382 silphids. A one-way ANOVA showed no significant difference between the mean numbers of staphylinines or carabids caught in carcass versus control traps. Silphids showed significantly greater mean number of beetles caught in carcass traps relative to control traps. Four species of Silphidae were documented, but contrary to similar studies, the vast majority of specimens belonged to two species of Nicrophorus (N. vespilloides Herbst and N. investigator Zetterstedt). Each of the three target taxa showed a peak in the number of specimens collected during the bloat stage of decomposition despite the carabid peak being driven by a phytophagous species.