Eye-background contrast as a quantitative marker for pupal age in a forensically important carrion beetle Necrodes littoralis L. (Silphidae).

Insect pupae sampled at a death scene may be used to estimate the post-mortem interval. The pupal age is however difficult to estimate, as there are no good quantitative markers for the age of a pupa. We present a novel method for pupal age estimation based on the quantification of contrast in intensity between the eyes of a pupa and the middle grey photography card as a standard background. The intensity is measured on a standardized scale from 0 (perfect black) to 255 (perfect white) using computer graphical software and pictures of the eye and the background taken with a stereomicroscope. Eye-background contrast is calculated by subtracting the average intensity of the eye from the average intensity of the background. The method was developed and validated using pupae of Necrodes littoralis (Linnaeus, 1758) (Coleoptera: Silphidae), one of the most abundant beetle species on human cadavers in Central Europe. To develop the model, pupae were reared in 17, 20 and 23 °C, with a total of 120 specimens. The method was validated by three raters, using in total 182 pupae reared in 15, 17, 20, 23 and 25 °C. We found a gradual increase in eye-background contrast with pupal age. Changes followed generalized logistic function, with almost perfect fit of the model. Using our method pupal age was estimated with the average error of 8.1 accumulated degree-days (ADD). The largest error was 27.8 ADD and 95% of age estimates had errors smaller than 20 ADD. While using the method, different raters attained similar accuracy. In conclusion, we have demonstrated that eye-background contrast is a good quantitative marker for the age of N. littoralis pupae. Contrast measurements gave accurate estimates for pupal age. Our method is thus proven to be a candidate for a reliable approach to age insect pupae in forensic entomology.

The quality of developmental reference data in forensic entomology: Detrimental effects of multiple, in vivo measurements in Creophilus maxillosus L. (Coleoptera: Staphylinidae).

Although in vivo measurements of larval insects are often performed during developmental studies of necrophilous beetles, their impact on development has not been studied. During measurements insects are taken out of the incubator for a few minutes at room temperature, which may affect the development and eventually the quality of the developmental reference data. Additionally, while being measured larvae are under stress which may have an effect on their development. We conducted an experiment using predatory beetle species Creophilus maxillosus L. (Coleoptera: Staphylinidae) which often occurs and breeds on large vertebrate carcasses. We tested the hypothesis that multiple, in vivo measurements affect the development of C. maxillosus by increasing its duration and changing adult insect size at emergence. As a consequence, we predicted that the multiple insect measurement protocol will affect the accuracy of age estimates using the resultant reference developmental data. Development of C. maxillosus was studied at 7 constant temperatures. All individuals were inspected for developmental landmarks; half of them were also repeatedly measured and weighed. Measured beetles developed longer than non-measured beetles (e.g. 1.59 days longer at 22.5 °C) and at emergence were distinctly smaller (e.g. 1.5 mm shorter and 22 mg lighter at 22.5 °C). The accuracy of age estimates was greater while using the model for non-measured beetles. These results support the claim that multiple in vivo measurements of insects reduce the quality of resultant developmental data. The measurements were particularly detrimental for the adult insect size. Consequently, particular attention should be paid to isomegalen diagrams which are based on insect length. Our findings indicate that these diagrams, when based on multiple, in vivo measurements of larval beetles, will systematically overestimate their age.

Estimation of postmortem interval (PMI) based on empty puparia of Phormia regina (Meigen) (Diptera: Calliphoridae) and third larval stage of Necrodes littoralis (L.) (Coleoptera: Silphidae) - Advantages of using different PMI indicators.

On 16 July 2015, a body of a 64-year-old man in advanced decomposition was found in an open area of the suburb of Srem (western Poland). Postmortem interval (PMI) was estimated by forensic pathologist for 3-6 weeks. Insects were sampled from the cadaver and the soil from below the cadaver. Empty puparia of Phormia regina were the most developmentally advanced specimens of blowflies. Moreover, third instar larva of Necrodes littoralis was collected directly from the cadaver. For the estimation of minimum PMI from puparia of P. regina, thermal summation method was used to estimate the total immature development interval of this species. In the case of larval N. littoralis, the pre-appearance interval (PAI) was estimated using temperature method and the development interval (DI) using thermal summation method. Average daily temperatures from the nearby weather station were used, as well as the weather station temperatures corrected by 1 °C and 2 °C. The estimates were as follows: 36-38 days using empty puparia of P. regina and 37-40 days using larva of N. littoralis (for the uncorrected temperatures), 31-34 days using both P. regina and N. littoralis (temperatures corrected by +1 °C), 24-27 days using P. regina and 28-29 days using N. littoralis (temperatures corrected by +2 °C). It was concluded that death occurred 24-40 days before the body was found and most probably 24-34 days before the body was found. This is the first report when PMI was approximated by the age estimates combined with the PAI estimates. Moreover, the case demonstrates the advantages of using different entomological indicators and an urgent need for the more robust developmental model for N. littoralis, as it proved to be highly useful for the estimation of PMI.

On the importance of skewed offspring distributions and background selection in virus population genetics.

Many features of virus populations make them excellent candidates for population genetic study, including a very high rate of mutation, high levels of nucleotide diversity, exceptionally large census population sizes, and frequent positive selection. However, these attributes also mean that special care must be taken in population genetic inference. For example, highly skewed offspring distributions, frequent and severe population bottleneck events associated with infection and compartmentalization, and strong purifying selection all affect the distribution of genetic variation but are often not taken into account. Here, we draw particular attention to multiple-merger coalescent events and background selection, discuss potential misinference associated with these processes, and highlight potential avenues for better incorporating them into future population genetic analyses.

Long-term study of pig carrion entomofauna.

Long-term changes of carrion entomofauna are poorly understood. No single carrion study lasted longer than one year. We studied entomofauna of large pig carcasses in the second and the third year postmortem. Ten carcasses were exposed in xerothermic grasslands of Western Poland in spring, early and late summer of 2012. Entomofauna was monitored until September of 2014. 72 species were found in the second year, and six in the third year. In the second or the third year carcasses from the late summer block revealed larger number of taxa compared to carcasses from the spring or the early summer block. Taxa differed in pattern of multiple larval colonizations. Only necrophagous taxa breeding in long-lasting carrion parts as well as predators feeding on arthropods present in these parts were found to recolonize. Recolonizations were always recorded in the second or the third year after death. Patterns of multiple colonization depended on the time of carcass exposure in the first year. Residencies of larvae were unexpectedly long in the second year. Several forensically important flies overwintered on carcasses in the larval stage. Current results support the view that insect fauna of cadavers with long post-mortem interval (PMI) may be used for PMI estimation.