Forensic entomological examination without a corpse

Source: Rom J Leg Med (2025) 33: 210–215; DOI: 10.4323/rjlm.2025.210

By Kristina Baumjohann, Mark Benecke

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Abstract: In an apartment, the previous tenant had died. His decomposition fluid had seeped into the wooden floorboards of the old building over a long period of time. The next tenant sent us samples of the wood and various insects that had come from the apartments’ floor. There was a noticeable decomposition odor in the apartment. The landlord wanted to immediately sublet the apartment and denied both the presence of the decomposition smell and a connection between this and the presence of the insects. We had to clarify whether the discoloration in the wooden floor was caused by seepage of corpse fluids and whether the appearance of the insects was related to the deceased previous tenant. The species composition of the insects and an ammonia test was indicative of the presence of at least decomposition fluid which may have come from a corpse.

Keywords: forensic entomology, forensic biology, stains, insects, decomposition.

CASE DESCRIPTION

The client had rented an apartment in which the previous tenant had died and been lying dead on the wooden floor in the apartment for a long time. The wooden floorboards there were discolored. There was a noticeable corpse odor in the apartment. Our client repeatedly found various insects in the now empty apartment. The landlord did not want to commission any extensive renovation work, as is usual in the city in question, owing to the housing shortage – he wanted to rent the apartment out again immediately. He stated that: - There was no corpse odor; only a smell of sanded wood. - The visible stains could have been caused by any liquid, e.g. coffee. - The flies found in the apartment had come from outside by chance, had not developed in the apartment and were not attracted by the decomposition smell of the deceased. Insect and wood samples were obtained to clarify whether the discoloration in the wooden floor was caused by corpse fluids seeping into it and whether the occurrence of the insects was related to the deceased previous tenant. We were able to establish a connection between the appearance of the insects and the corpse fluid that leaked and seeped into the wooden floor (and far below).

MATERIAL AND METHODS

Insects

Insects were examined with a stereomicroscope binocular (Leica MZ12.5, max. 100x) and identification keys [1-5].

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Floorboards/odor

Due to the time pressure and suffering of those affected, it was not possible for us to inspect and take samples on site. However, we had photos and a video of the apartment in very good quality. The samples of the wooden floorboards were each packed airtight in three-layer Ziploc bags, with another small airtight bag with wooden material inside.

Each sample of those samples was sent to us in a glass jar (Fig. 1). Some of the wood material had a distinctly dark, charred-looking discoloration (Fig. 2). The wood samples consisted of smaller and slightly larger pieces of wood (Fig. 3).

Ammonia test

A rapid ammonia test (Bosike/Like Sun Company, Essen, Germany) was conducted to exclude coffee and as an indication of putrification liquid [6, 7]. Therefore, a small piece of a darkly discolored area of the wooden floor was swirled in 3 ml of sterile water (disposable vial, AddiPak Company), and put in a sterile, DNA-free centrifuge tube (Greiner Company) for a few minutes (Fig. 4). The ammonia test strip was dipped into it.

RESULTS

Insects

The samples contained the following insect groups and species: - Flesh flies of the genus Sarcophaga spec. (Fig. 5); - Blow fly of the species Calliphora vicina (Fig. 6); - Carpet beetles (skin beetles), probably of the genera Attagenus spec. and Anthrenus spec. (Dermestidae) (Fig. 7); - Larder beetle larvae (Dermestidae) (Fig. 8); - Chalcid wasps, probably of the family Torymidae (Chalcidoidea) (Fig. 9).

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Flesh flies (n = 56) were most prevalent in the samples, followed by carpet beetles (n = 41) and chalcid wasps (n = 9), while only one blow fly was present (Table 1). The skin beetle larvae were numerous and we did not count them. Apart from the chalcid wasps, all insects found in the samples are associated with decomposition processes of human and animal bodies [8-11].

While blow flies visit dead bodies in earlier stages of decomposition [12, 13], flesh flies colonize corpses during the entire decomposition process [14-22]. In large cities, this is particularly true for corpses found in apartments.

Carpet and/or skin beetles and their larvae occur in later stages of decay (desiccation) and prefer drying organic tissue (including mummies, stuffed animals, etc.) on which they feed [23-26]. Chalcid wasps are predators of other insects that colonize corpses, such as the blow flies and flesh flies found in the samples [27]. They only appear later during the course of corpse colonization, as they do not colonize the corpse as such, but animals living on the corpse.

Individual adult, dead flies showed holes and erosions, which may have been caused by skin beetles and chalcid wasps. This is a further indication of prolonged colonization and/or the prolonged presence of insects colonizing corpses and attracted by the odor of corpses (Fig. 10).

Odor

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As soon as the glass lids (i.e. the outermost packaging of the multi-packaged samples) were opened, the laboratory room was immediately filled with the typical smell of decomposition. All samples already gave off a distinct putrefactive smell through the closed bags. The smell of coffee or similar odors could not be detected in any of the two samples – even when the bags with the wooden samples were opened and the odor was tested directly with the nose.

No chemical analysis were performed as this had neither been asked by the client nor had this been necessary for our investigation.

Ammonia

The rapid ammonia test as an indicator putrefactive liquid gave a clear result after just a few seconds of immersion; according to the manufacturer, this normally takes longer (Fig. 11).

Regarding the landlord’s statements

Corpse odor and “stains”

According to the client, “all employees... [of the landlord] who were in the apartment as well as the... [the landlord’s] crime scene cleaner and pest controller [...] stated that there was no corpse odor, only the smell of sanded wood.” This is not correct for the discolored wood: It smelled very distinctly of a decomposing corpse. Irrespective of this, there may have been a strong odor of sanded wood or another odor in the apartment. However, the odor of the corpse must have been perceptible in the immediate vicinity of the wood.

Flies

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The clients state: “On his second visit to the apartment, the crime scene cleaner and pest controller took about 3 - 5 specimens of the flies from the large room to have them identified by a biologist friend. We were not informed of the results of the investigation.” The landlord is said to have stated: “An expert confirmed to us that these flies must have come from outside and could not have developed in the apartment.” This statement is partly correct: The insects may have flown into the apartment from outside because they were attracted by the corpse odor serving as an attractant for these insects. They do not necessarily have to have developed there. They can also “only” feed or lay eggs on putrefying tissue. The presence of the skin beetles and adult flies indicates that the colonization/ flying of insects into the apartment, which was either littered with animal meat or had a human corpse, had been taking place for some time: These are so-called late corpse colonizers. All of the insects we examined only occur at corpse sites with decomposing corpses or in environments with a high meat content in old waste (animal or human corpse parts).

DISCUSSION

The insects studied here are either corpse colonizers or are clearly associated with decomposition processes or animals attracted by them.

We are not aware of any odor that is similar in composition to the type of corpse odor present here. A similar odor can occasionally arise from mountains of garbage, which must then consist mainly of meat scraps. Only then does this odor develop, and then only after sufficiently long “maturing”. The time required for this depends on the temperature: The warmer, the faster. Since the apartment in the video sent to us does not appear to be full of garbage, but empty, there would have to have been a large quantity of garbage with a high meat content.

The ammonia test is triggered by the presence of decomposable substances, not by “normal” wood floors or pieces. This shows that decomposition fluid (and not coffee) must have penetrated the floorboards.

We have a video from the apartment which shows that there are cracks in the floor, including where the body is supposed to have been lying. Decomposition fluid and pieces of tissue from the corpse may have entered the area under the floorboards through these cracks (puddle of fluid). Cadaveric fluid can penetrate the actual wood of unsealed floors and then remain there or under the wooden floorboards owing to the adhesive properties of the semi-liquefied cadaveric material.

From time to time, these kinds of contaminated floors are sealed to “encapsulate” the odor or treated with chemicals to dissolve or mask the odor (for example with the raspberry odor of Maskomal or similar). However, it makes more sense to remove the source of the odor; this is comparable to used baby diapers, which are also thrown away or cleaned and not covered up in order to mask the odor.

The collective occurrence of the insects from the apartment samples certainly indicates the presence of at least decomposition fluid, the odor of which attracted the animals. The putrefactive liquid must have been in the wooden floor in sufficient quantity and presumably over a longer period of time, otherwise only blow flies would be found, which colonize corpses at an early stage. Skin beetles and chalcid wasps appear as later corpse colonizers [28]: Carpet beetles and their larvae use dried organic tissue and dead insects (such as flies) as a food source. However, they are less commonly found in apartments with corpses and piles of garbage, as the soiled areas are usually cleared and cleaned quickly. Chalcid wasps in particular are then naturally no longer attracted: They colonize the juvenile stages of other insects.

The absence of blow fly larvae and their pupae in the samples and on the photos indicate that there is now no fresh, new organic material available for the blow fly larvae to feed on. None was found anyway.

As blow fly larvae usually seek out dark and narrow “hiding places” to pupate, fly pupae and larvae could also be found under floorboards and behind carpet rails. This can occur from time to time and sometimes leads to strange occurrences of flies for the residents. For example, an apartment can be cleared and cleaned, but the pupae of the flies are still hidden in cracks and the flies hatch from the pupae later (see case 2 „Empty Apartment“ [29]).

The “mixture” of dead flies, beetles and chalcid wasps in the apartment concerned here means that the animals were attracted by the smell of corpses and died in the apartment or that there was sufficient food (including corpse fluid). It is irrelevant whether these flies developed in the apartment (from larvae) or whether adult insects have flown into the home. The main reason for their appearance is that they are attracted by the smell of decay.

Conflict of interest

The authors declare that they have no conflict of interest.

References

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11. Al-Khalifa MS, Mashaly AM, Al-Qahtni AH. Insect species colonized indoor and outdoor human corpses in Riyadh, Saudi Arabia. Journal of King Saud University. Science 2020; 32:1812-17.

12. Byrd JH, Castner JL. Forensic Entomology: The utility of arthropods in legal investigations. 1st edition. Boca Raton: CRC Press; 2001. p. 45-46.

13. Bugelli V, Forni D, Bassi LA, Di Paolo M, Marra D, Lenzi S, Toni C, Guisiani M, Domenici R, Gherardi M, Vanin S. Forensic entomology and the estimation of the minimum time since death in indoor cases. J Forensic Sci. 2015;60:525-531.

14. Byrd JH, Castner JL. Forensic Entomology: The utility of arthropods in legal investigations. 1st edition. Boca Raton: CRC Press; 2001. p. 50-51.

15. Cherix D, Wyss C, Pape T. Occurrences of flesh flies (Diptera: Sarcophagidae) on human cadavers in Switzerland, and their importance as forensic indicators. Forensic Sci Int. 2012; 220(1- 3):158-163.

16. Bonacci T, Greco S, Cavalcanti B, Brandmayr P, Vercillo V. The flesh fly Sarcophaga (Liopygia) crassipalpis Macquart 1839 as an invader of a corpse in Calabria (Southern Italy). J Forensic Sci Crimin. 2014;1:1-5.

17. Pinto Vairo K, da Cruz Canepro MF, Correa RC, Preti D, Moura MO. Can Sarcophagidae (Diptera) be the most important entomological evidence at a death scene? Microcerella halli as a forensic indicator. Rev Bras Entomol. 2017;61: 275-276.

18. Battan-Horenstein M, Mulieri PR, Gleiser RM. Evaluation of environmental and local factors on urban sarcosaprophagous Diptera: a case study with Sarcophagidae. Med Vet Entomol. 2020;35(2):158-168.

19. Gunn A. The colonisation of remains by the sarcophagid fly Sarcophaga argyrostoma (Robineau-Desvoidy) (Diptera: Sarcophagidae). Forensic Sci Int. 2020;315:110465.

20. Barbosa TM, Carmo RFR, Mello-Patiu CA, Leal IR, Dias Vasconcelos S. Effects of chronic anthropogenic disturbances on flesh fly (Diptera, Sarcophagidae) assemblages in areas of seasonally dry tropical forest. J Insect Conserv. 2021;25:485-498.

21. Afifi FM, Abdelfattah EA, El-Bossiony GM. Impact of using Sarcophaga (Liopygia) argyrostoma (Robineau-Desvoidy, 1830) as a toxicological sample in detecting clonazepam for forensic investigation. Egypt J Forensic Sci 2022;12:37.

22. Hediyeloo S, Akbarzadeh K, Rezaei M, Oshaghi MA. Colonization pattern and thermal needs of immature phases of Sarcophaga argyrostoma (Diptera: Sarcophagidae): significance for estimating postmortem interval. Heliyon 2024;10:e26576.

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24. Charabidze D, Colard T, Vincent B, Pasquerault T, Hedouin V. Involvement of larder beetles (Coleoptera: Dermestidae) on human cadavers: a review of 81 forensic cases. Int Journal Legal Med. 2013;128(6):1021-1030.

25. Gunn A. The exploitation of fresh remains by Dermestes maculatus De Geer (Coleoptera, Dermestidae) and their ability to cause a localized and prolonged increase in temperature above ambient. JFLM 2018;59:20-29.

26. Kadej M, Szleszkowski L, Thannhäuser A, Jurek T. A mummified human corpse and associated insects of forensic importance in indoor conditions. Int J Legal Med. 2020;134:1963-1971.

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29. Benecke M. Six forensic entomology cases: description and commentary. J Forensic Sci. 1998;43:797-805.

Estimation of post mortem intervals in humans

Source: Tagungsband der 12. Wissenschaftlichen Tagung der Gesellschaft für Entwicklungsbiologie e. V. vom 11. - 14. März 1997 in Köln (GER)

A macroscopic application of developmental biology called Forensic Entomology

From Mark Benecke

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Insect development is well-studied and thus can be used as a marker for estimation of time intervals. Blowflies and several beetles together with their larvae are used in forensic science to help diagnosing how long a corpse has been exposed to defined environmental conditions.

Most times the forensic entomologist is asked if the insect fauna of an corpse can tell the time and date of a person's dead.

For various reasons, pest control experts as weIl as entomologists and (forensic) medicinal practitioners have studied the correlations between temperature, humidity and speed of development of several insects.

Forensic scientists put together those facts and calculated curves which now allow a precise estimation of the living time of an insect found associated to a corpse. In most cases, length, weight, and macroscopic state (e.g., filling of intestines) of larvae are determined and compared to the standard data curve.

Because of several insects visiting corpses only at a defined state of decomposition, further information about the post mortem interval is given by determination of species. Sometimes it is quite hard to determine species just by their pupae or even worse by their larvae. That is why it can be necessary to breed larvae to adultship. In doing this, developmental biology once again is in focus of research and criminal investigations.

Only a lew laboratories in the world (Rosny sous Bois, Hawaii, Wien, Köln, Quantico among few others) perform(ed) studies concerning the forensic application of insect development. We present our aims to establish a database of developmental processes of insects which are specific for the western German fauna of human corpses, especially for the diptera Lucilia caesar, Calliphora vicina and Calliphora erythrocephala.

Forensic Entomological Examinations for Animal Welfare Offices under Suboptimal Preservation Conditions

Source: Forensic Sci. 4:387–395. https:// doi.org/10.3390/forensicsci4030023

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Kristina Baumjohann & Mark Benecke

Abstract

A female dog had allegedly been alive one day before its death. The veterinary office thought about accusing the dog’s owner for animal cruelty and commissioned forensic entomological expertise for the calculation of the time of insect colonization on the dog’s body. The statement of the dog’s owner was proven false by us on the grounds of (a) the advanced state of decomposition and (b) the minimal developmental time of fly larvae found on the dog’s body. The darkening and deformation of the fly maggots as well as insufficient temperature data made case work trickier than usual. We worked through the case by creating forensic entomological temperature scenarios. The court used the entomological evidence and issued a penalty order.

Keywords

forensic entomology; insect traces; minimum time since death; neglect; sampling

1. Introduction

Insects, mostly flies and beetles, can colonize human and animal wounds or bodies both during life and after death. If colonization takes place during life, it is referred to as myiasis. Myiasis may occur in cases of neglect; in such cases, it is sometimes possible to calculate the period of neglect using the development of insects [1–3].

The time of development of insects is temperature-dependent: low temperatures slow down insect development and high temperatures accelerate it. Different insect species develop at different rates at the same temperature [4].

The preference of adult flies to deposit eggs in wounds and body openings applies equally to humans and wild or domestic animals [5,6]. In cases of myiasis, eggs are also laid in the soiled diaper area [1–3]. This applies to soiled cushions in dog baskets and soiled blankets, too.

In neglect cases, the collection and subsequent preservation of fly maggots should be carried out separately according to the place of collection (diaper area, open wounds, or natural body openings) in order to record the possibly different developmental ages of the animals at the different colonization sites of the same body.

In cases of animal cruelty and neglect in wild and domestic animals, the entomological evidence collected from living or deceased animals may also provide investigating authorities with information on the circumstances of death [7–12].

The condition of the preserved insect specimens that reach us do not always allow for easy species determination. The case presented here shows that, despite poorly preserved insect material, the difficulty in calculating a possible development time of fly maggots can be narrowed down by creating various temperature scenarios.

Unlike in high-profile forensic cases, the veterinary office initially left the question open if time since death or time since the beginning of neglect including a possible maggot infestation had to be determined by us. We communicated that in this case, it would be best to operate with colonization time, irrespective of whether this was the colonization time of wounds of the living dog or the colonization of the dead dog.

2. Case Description

In connection with the death of an approximately two-year-old, female French bulldog, a German veterinary authority issued an order to calculate the minimum colonization period of the fly maggots collected from the dead dog.

The dog owner stated that she had left her dog in her uncle’s apartment several days before the dog’s death and had looked after him there every day during her work breaks. She had allegedly last provided the dog with water and food on 23 July 2022; she claimed that the dog was still alive at that time. When she went to pick up her dog from her uncle the next evening, 24 July 2022, the dog had died.

That same evening, the animal mortician collected the dead dog and froze it in the funeral home at −3 °C, according to his statement. The mortician had noticed “heaps of maggots” on the dead dog’s body when he handed it over for examination, i.e., the dog had already been severely decomposed at this point. The veterinary office received the corpse on 25 July 2022; the dog was frozen there at unknown temperatures. On 30 August 2022, the animal’s corpse was sent to an Institute for Veterinary Pathology and examined there on 2 September 2022. The fly maggots collected from the dog’s body were then frozen at −20 °C until shipment.

The owner’s statements were made to officers of the local veterinary authority. Police was not involved since the case was considered to be low-key. The veterinary office then contacted the prosecutors’ office. The dog owner did not have to give a sworn statement since it was clear that the case would be handled by the district court and the penalty would be very low because the dog owner had no criminal record and most of such cases are not prosecuted at all in Germany.

Figure 1. Condition of the emaciated dog’s body on delivery to the mortician; the dog had allegedly been alive and well the day before its death. (Note: the insufficient image quality of Figures 1 and 2 is due to the fact that the veterinary pathologist did not allow the use of original images, so copies from the report of the veterinary office had to be used.).

3. Veterinary Pathological Examination of the Dog

According to the veterinary pathologist, the dog was already in a high state of “autolysis to putrefaction”. There were numerous fly maggots on the dog’s body, there were nits (louse eggs) in the fur, and the beginning of skeletonization at the right upper jaw was noted.

The dog had no subcutaneous fat and no structural fat deposits: kidney capsule fat, coronary fat, and intestinal mesentery fat were missing. The stomach was empty; the animal was in a highly reduced nutritional state (Figures 1,2).

4. Forensic Entomological Examinations

4.1. Material and Methods

Figure 2. Eyes and brain of the dog missing due to the feeding activity of fly maggots and advanced decomposition. (Photo quality: see remark in Figure 1.)

The sample from the Institute of Veterinary Pathology included 146 individual fly maggots and eight clusters of several maggots that were connected (as if glued) together.

The previously frozen fly maggots were sent to us by the veterinarian in 96% ethanol and reached us on 16 January 2023. The thawed maggots were predominantly brown to black in color (Figure 3), the tissue was rubbery, and the animals were predominantly deformed. The discoloration and deformations made it difficult to determine the fly species, as certain body characteristics must be visible to do so. The length of a stretched maggot is used to determine age and cannot be measured correctly if the animals are bent and contorted. The proper and immediate storage of the fly maggots at the mortician’s office could have prevented the discoloration and tissue deformation [13].

Species determination was performed based on morphological features using stereomicroscopes (Leica Mz 12.5, Leica S9E, Wetzlar, Germany) and a light microscope (Leica DM LM, Wetzlar, Germany) with identification keys from Szpila [14,15].

One third of the maggots were therefore placed in an 8% potassium hydroxide solution (KOH) at room temperature. The tissue of the maggots was so firm and tough that it only became soft enough to micro-dissect after about a week in the softening solution. The bleaching effect of the potassium hydroxide made body features relevant to determination largely visible again (Figure 4).

The length of 20 blow fly maggots was measured, the stage of development determined, and their species identified (see Section 4.2). After examination, the maggots—with some body parts removed during the examination (anal plate, head capsule, and mouth parts)—were each transferred to a reaction tube (1.5 mL).

The remaining maggots left in the potassium hydroxide solution were placed in a container of methylated spirits for further storage. The sample also contained a maggot of a flesh fly species: this maggot was also placed in 8% KOH solution for one week at room temperature and then examined microscopically.

4.2. Results of the Species Identification

All 20 blow fly maggots examined belonged to the species Lucilia sericata (Meigen, 1826); the average length of the animals was 1.3 cm. All maggots had reached the third and thus last larval stage of development. It was not possible to determine whether the animals had already emptied the intestinal contents in preparation for the subsequent pupation phase (so-called postfeeders) or whether they still possessed them at the time of preservation due to the dark tissue discoloration.

Figure 3. Discoloration and deformation of the fly maggots to be examined

The single, 2 cm long flesh fly larva belonged to the species Sarcophaga argyrostoma (Robineau-Desvoidy, 1830) in the third (and last) larval stage of development.

4.3. Results of the Calculations of a Possible Egg-Laying Time

4.3.1. Temperature Data

Fly maggots develop depending on the surrounding temperature. The temperatures at which the animals developed until the body was found are therefore required.

Figure 4. Dark discoloration of the flesh fly maggot (top) and clear tissue after treatment with KOH (bottom)

Normally, the following steps are necessary to recalculate the temperatures in the best case [16]:

• Comparison of hourly temperatures over a certain period of time (e.g., three days or longer) between the location and a nearby weather station;

• Calculation of temperature deviations between these locations;

• Calculation of a correction factor;

• Recalculation of the temperatures for the location where a corpse was found for the time before it was found = time in which the insects developed on the body.

The temperature data from a weather station for this period and a previously calculated correction factor are used for this purpose. In our case, the German Weather Service (DWD) provided daily average, maximum, and minimum temperature values for the period from the beginning of June to the end of July, but no hourly temperature readings.

However, a privately operated weather station, which was located 6.8 km away from the dog owner’s uncle’s home, transmitted hourly air temperature values for the period from the beginning of June 2022 to the end of July 2022.

4.3.2. Calculating the Development Time of Fly Maggots

The steps listed above for calculating the development times of the fly species could not be carried out in this case due to the lack of data and information. The development time of the fly maggots could still be approximated by creating various “temperature scenarios” based on the hourly temperature data from the private weather station and developmental data from the literature.

4.3.3. Development Data for Lucilia sericata and Sarcophaga argyrostoma

The calculation of the time of oviposition of Lucilia sericata was carried out using the developmental data of Wang et al. 2020 [17]. Table 1 shows the results of the calculation of the development time of the maggots of Lucilia sericata under the influence of different temperatures.

Due to the dark coloration of the fly maggots, it could not be determined whether intestinal contents were present at the time of collection. Fly maggots empty their intestinal contents before pupation (postfeeding larvae; so-called postfeeders): maggots without intestinal contents are therefore older than those with intestinal contents. A possible influence of the KOH solution on any remaining intestinal contents (which were no longer visible after the KOH treatment because the tissue was brightened too much) could not be ruled out due to the long soaking time.

Data from Wang et al. [17] for the developmental transition from the second to the third developmental stage did not fully fit because the maggots were clearly in the third larval stage; any transitional stage would have been recognizable by morphological characteristics, e.g., half-shed skin and breathing spiracles. The maggots examined corresponded best with the data for transition from the third to the postfeeder developmental stage. As postfeeders, the maggots migrate from the dead body to pupate, so later developmental stages are not expected on a corpse.

Females of flesh flies, to which Sarcophaga argyrostoma belongs, lay live young larvae on decaying tissue in the first stage of development [18]. Since flesh fly larvae of this species do not randomly feed on a living organism but are attracted to decomposed tissue, we calculated the development time as a possible colonization time for this maggot based on the temperature and development data of Grassberger and Reiter (2002) [19] (Table 2).

4.4. Answering the Client’s Questions

Our calculations were based on both the fluctuating daily temperatures of the weather station 6.8 km away from the uncle’s home and constant temperatures, e.g., 15 °C for a possible colonization in the basement room, 24 °C for an indoor room in summer (during the day), and 30 °C daytime temperature for the outdoor colonization in that summer.

Further influences on egg laying and larval development such as rain [20–23], night, and dark conditions [23–29] were disregarded since we were told that the case took place inside or close to the apartment.

Since the maggots of both fly species had reached the third and last stage of development as larvae (before pupation), the larvae could not have developed within one day (e.g., from 23 to 24 July 2022) from oviposition.

We do not know whether the dog was still alive at the time the eggs were laid. It is possible that the dog was neglected and that its wounds were colonized by fly maggots during its lifetime. Lucilia sericata and Sarcophaga argyrostoma are fly species that may colonize living yet neglected bodies [9].

Since we also did not know whether the fly maggots sent in for examination were the oldest maggots that had developed on the dog, the calculated time periods were the minimum development time of the larvae.

The extensive maggot infestation of the oral cavity, the loss of substance on the muzzle due to autolysis, and the strong odor of decomposition before the body was frozen also spoke against the statement that the dog had been healthy and alive on 23 July 2022.

5. Discussion

5.1. Temperatures

Temperature data from the colonization site of the dog were missing. Therefore, the temperatures at which the insects colonized the dog before 24 July 2022 could not be mathematically reconstructed.

If the owner claimed to have visited the dog at about the same time on both days, the maximum PMI hypothesis to test would be approximately 24 h. Assuming the most rapid development rates in the reference papers [17,19], both a 13 mm third larval instar of L. sericata and a 20 mm third larval instar of S. argyrostoma would be too old for the owner to have told the truth, irrespective of the temperatures of the dog carcass.

We decided to use local weather data as well as the most recent developmental data including ADH information to build and check our temperature scenarios (Table 1). Since only one Sarcophaga larva was sent to us and since no recent developmental ADH data were available, we decided to use an older data set for this species that did cover the temperatures we used in our scenarios. This was sufficient because the statement of the owner of the dog was found to be false in all our calculations.

We decided to use the most current data for Lucilia sericata that also include ADH values. Since we observe a massive impact of climate change in Europe, we considered the most modern data to be the most reliable in this particular case. For Sarcophaga argyrostoma, we had to rely on the older data set because the most recent data sets did not cover the temperature ranges that we needed to include in our “check the scenarios” tests.

5.2. Colonization Site of the Dog

It remained unclear whether the dog was colonized inside the house or outside and if windows were closed or not. “Closed” doors in Germany often provide access points for flies, as the adults can squeeze through old keyholes or gaps between the door and the floor.

5.3. Fly Maggots

It is unknown to which colonization wave the fly maggots collected from the dead dog belonged, and especially, if there were older developmental stages of the flies or other insects in the vicinity of the dog. In a strict court room setting, one could also question if the maggots had been alive on the dog. The color changes and conservation state of the maggots did not allow a reliable length measurement. Our measured lengths were minimum lengths.

Concerning a possible lack of information in the scientific literature relating to the variability of the postfeeding stage, we restricted ourselves to the information contained in the sources that we used [17,19]. In the Lucilia larvae as well as the single Sarcophaga larva, we saw three slits in the abdominal (posterior) spiracles. Therefore, we decided that any developmental interval beginning from the transition from larval stage 2 to larval stage 3 until the possible postfeeding stage should be considered. Our minimum developmental estimate already excluded the dog owner’s statement, so in this particular case the question was answered without further examination of a possible postfeeding stage.

We did not aim for the inclusion of larval length data because on the one hand, we wanted to support the veterinary office even though hardly any budget was available and we thought that a discussion about larval lengths might lead to further unpaid work. On the other hand, our approach using scenarios sufficiently covered the questions that we were asked to answer. Since exact environmental information (the dog’s exact place of death, etc.) were unknown, we decided to work on the simplest and safest level so that a possible defense could not use a confusion strategy over numbers. Finally, in our lab, we are hesitant to work in an overprecise manner when larvae arrive in a hardened state. In higher profile cases, we would naturally determine the minimal developmental time from shrunken, hardened maggots, but this case had to be handled under minimalistic conditions, yet with simple and safe conclusions due to the circumstances described above.

5.4. The Dog Owner’s Statement

Strictly speaking, it is unknown whether the dog may have died elsewhere and was then transported to the uncle’s apartment.

5.5. Conclusions

Despite all limitations, our measurements show that the dog could not have been healthily alive on the evening of 23 July 2022. This entomological exclusion matches the observation that the dog’s brain was severely decomposed and largely missing.

The dog was colonized by cadaver flies on the morning of 21 July 2022 at the latest; if the dead or living animal had been in a colder environment than 30 ◦C outside temperature, then colonization could even have taken place much earlier.

In the trial, the court warned the dog owner and ordered her to pay 1200 Euro to a charitable organization. She was banned from keeping animals for one year. After that, the owner may legally own animals again.

Even though the larvae were in poor condition and not all data were available, the question of the animal welfare office could be answered in a useful, legal way. This allowed the office to go on trial.

We believe that this case might remind veterinarians and veterinary pathologists to preserve and document entomological traces in the best possible way. In more difficult cases, a better preservation of the maggots would have been necessary. Here, the relevant question could be answered sufficiently

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