This study is the first research using a large dataset of real cases for systematic study with a focus on the spatio-temporal and altitudinal distribution of blowflies in Switzerland. All 10 species of the family Calliphoridae, which were present on the bodies, belong among common necrophagous European fauna. The most common species of our dataset was by far C. vicina (69 %), while in some other ecological studies from Switzerland or Germany first place often belongs to L. sericata [3, 12, 34]. In our case, L. sericata was present in 30% of cases (while still being the second-most dominant species of our dataset). The occurrence of L. sericata and some other focal species is surprisingly different in comparison with a similar dataset of Bernhardt et al. , who analyzed the Diptera assemblages of 51 human remains in the city of Frankfurt. The biggest discrepancies between the two datasets were among the following species: L. sericata (30 % vs. 86 %), P. regina (5 % vs. 43 %), and L. ampullacea (1 % vs. 45 %) (Table 1). Some of the reasons for such substantial differences can be the different years of sampling (1993–2007 in our case vs. 2014–2016 in Bernhardt’s dataset) or the proportion of indoor vs. outdoor cases. There were only 9 out of 51 cases (i.e., 18 %) found outdoor in Frankfurt , while we localized 58 out of 145 cases (i.e., 39 %) outdoor in and around Lausanne. The geographical distribution of P. regina and L. ampullacea in Europe during the years 1993–2007 could also be different from recent years due to climate change [27, 35, 36]. However, we think that the most probable reason might be the character of the local habitat composition and the topography of the compared areas. While Lausanne is situated 495 m above sea level surrounded by high mountains, Frankfurt is a low altitude city (112 m above sea level). If we take the altitude as a surrogate of the climate, we can assume that higher altitudes will be more suitable for cold-tolerating species like C. vicina, while lower altitudes fit better the thermophilous species such as L. sericata.
Due to the local topography, the cases described herein are ranging from 287 m above sea level up to 2620 m above sea level (Fig. 5). Such a large altitudinal gradient allowed us to evaluate the ability of blowflies to breed and inhabit high mountain habitats. The knowledge of the blowfly distribution along the altitudinal gradient can be an important information as it can contribute to the understanding of the geographical distribution of many species as well as their local diversity . There were only 3 species able to colonize human remains above 1500 m: C. vomitoria, C. vicina, and L. caesar. These results are in agreement with the results of , who studied the distribution of blowflies along an altitudinal gradient in central Spain. Baz et al.  determined C. vomitoria and C. vicina as species of high altitudes (most abundant in between 1600–1800 m), while L. caesar as a species of intermediate elevations. Unfortunately, there were only 7 bodies localized outside in high altitudes (above 1200 m). Calliphora vomitoria was present in all of the cases, C. vicina was present in 3 cases, and L. caesar in 1 case. Even with such a small amount of appearances in the highest altitudes, the affinity to altitude was significant for L. caesar and C. vomitoria as the trend was set in lower altitudes. Calliphora vomitoria is a thermophobic species [12, 39], which could explain its altitudinal preferences if we consider elevational gradient as a surrogate of climate . Indeed, the temperature profile in high elevations will always be one of the limiting factors, which means that thermophilous species will visit higher altitudes only occasionally during the warmest days of the year. The occurrence of the thermophilous species in high elevations is thus also season dependent as the upper distributional limits of species may fluctuate on a seasonal or annual basis [40, 41]. All other species in our dataset preferred low elevations. The affinity to low altitudes is not surprising for thermophilous species such as L. sericata and C. albiceps (in fact, no species of Lucilia was able to pass 1200 m except for L. caesar). However, unlike L. sericata, C. albiceps is occasionally capable of reaching high elevations . The highest locality where C. albiceps was present in our dataset was at 1230 m, where 2 living pupae were found in the forest. Phormia regina is the only species, which did not pass the altitude of 750 m. Our data thus suggest a strong negative correlation to altitude. However, since P. regina was also the least observed species of our analyses, the results of its altitudinal gradient might be incomplete. Indeed, the presence of P. regina was already reported as high as 2835 m on remains of a human stillborn infant  and up to 3566 m on rabbit carcasses  (both cases from Colorado). The local altitudinal distribution of this species in Switzerland should be further monitored to complete the missing data and confirm its presence in higher elevations.
The habitat preferences of all 8 analyzed species are shown in Fig. 2 as well as in Fig. 6. The results generally confirm the ecology of detected species known from previous experiments conducted in Europe. Calliphora vicina and L. sericata preferred apartments (i.e., indoor cases), which is supported by the results of many other authors [3, 44,45,46]. Calliphora vicina was present in 42 % of indoor cases, which makes it the most important species from the point of forensic investigations of indoor cases in and around Lausanne. Lucilia sericata was often found also in the fields or parks near human settlements. The distribution of L. sericata in our study is therefore mostly synanthropic, which is similar to the findings of [4, 47]. On the contrary, C. vomitoria is considered an indicator of natural environments [4, 47, 48] and rural sites . The occurrence of C. vomitoria in our area was distributed among all observed habitats, with a preference for meadows, forests, and alpine habitats. Such broadscale of distribution excludes this species as a biogeographic indicator . The distribution of C. vomitoria was partially copied by the occurrence of L. caesar. Lucilia caesar also prefers shaded locations in rural environments [12, 21, 47] and it was positively correlated with forest habitats. However, even L. caesar was found 13 times (15 %) in the apartments. The explanation probably is the overall majority of the indoor cases in our dataset (Table 2). As mentioned above, 61 % of all the cases were found in the apartments. All analyzed species of blowflies were found indoor (Fig. 7). Lucilia illustris was significantly related to parks and fields, i.e., in more synanthropic habitats than those preferred by L. caesar. The habitat preference and synanthropic level for L. illustris are somehow in between L. sericata and L. caesar. It is a heliophilic species less abundant in the shady forest localities (in comparison to L. caesar) as well as in the indoor cases (in comparison with L. sericata) [4, 12]. It has to be noted that the identification of L. caesar and L. illustris can be tricky even in adult stages, which can lead to a certain bias in results or even omittance of the species differentiation for certain analyses . The non-indigenous blowfly C. albiceps is often described as tropical and subtropical species; however, its occurrence in Europe is already well established and it serves as a forensically important indicator for European criminal investigations for years [21, 49]. Its occurrence on human remains in and around Lausanne was rather occasional with some tendencies to prefer more natural habitats as forests and meadows. Recent data however show a bigger percentual proportion of this species on human cadavers, which can point at its increasing importance as a species of forensic importance in Switzerland (data of real cases from 2019 to 2020 in Lausanne - Hodecek unpublished). The rest of the observed species did not show any significant dependence on habitat type. This includes P. regina with only 7 observations and P. terraenovae, which was present in all habitat types except fields and alpine environment.
The seasonal occurrence of blowflies in Europe has already been studied in several experiments (Portugal ; Spain [6, 48]; England ; Germany ; Switzerland ; Italy ). However, the activity of geographically distant populations within the same species can differ due to their local adaptations [51,52,53]. It is thus very important to monitor the activity of the forensically important blowflies locally. In our dataset, there was only one species with occurrence in each month—C. vicina. It is a species well adapted for cold weather and during the winter season, it is often the only species found on the bodies . In southern parts of Europe, it can be missing during the warm part of the year due to its upper developmental threshold of 30 °C [47, 50]. Calliphora vomitoria was active during the whole year except February. Both species are thermophobic with supercooling points (SCP) of − 8 °C (C. vicina) and – 11 °C (C. vomitoria) for overwintering adult flies . The SCP is higher for C. vicina; however, due to its preference for urban habitats, it can stay active even in the coldest winters with many opportunities for warm refuges . The occurrence of all other species was focused to warm summer and autumn months. Such results are supported also by [4, 12, 34, 48]. Thermophilous species of genus Lucilia were mostly present in August with the highest activity from May to October. Only L. sericata occurred also during the cold months of February and November; however, all of these cases were located indoors. Interestingly, live pupae were found in the case from February 1997 and the PMImin was calculated to be 63 days, which would mean an oviposition at the end of December. Even though L. sericata is considered to be a “summer” species, it can occasionally be active even during the winter months when it seeks the warmth of human settlements. Similar to C. vicina, this species is also related to urban areas. All species except C. vicina and C. vomitoria were the most active during August; however, this is likely supported by the fact that August was the busiest month for the police investigators (Fig. 3). Chrysomya albiceps and P. terraenovae were irregularly distributed during the year with a peak of their activity in summer.
There was, however, a case with living pupae of P. terraenovae found in December 1997 in the forest. The PMImin for this case was estimated to be 105 days, which would mean oviposition at the beginning of September. Some living pupae of C. albiceps were also found during cold months of February and April 1996 in forested areas. The PMImin was calculated to be more than 6 months for both cases, which means that pupae of C. albiceps could be able to overwinter and survive very low temperatures. While P. terraenovae is considered a cold-tolerating species [20, 54], C. albiceps is a strictly thermophilous species [41, 49, 55]; therefore, such findings are an interesting observation. Phormia regina was found only in May, August and September, but the seasonal activity cannot be determined just from 7 occurrences.
The study limitations
Similar to other studies evaluating insect data from real cases, it is important to note that they do not represent the results of a planned ecological study and while such a dataset has certain advantages, we have to be also aware of its disadvantages . One of the most important differences between data collected during carefully designed experiments and real case scenarios is that we will never be able to retrieve the same amount of replications out of the latter. For example, in the presented dataset 61 % of cases were found indoor, while only 39% were outdoor. This is a typical scenario as most cases of legal investigations are conducted on bodies found inside . In this article, we also decided to analyze the habitat preferences of the focal species. Due to the majority of cases being located inside, the most numerous habitat types in our dataset was “apartment” with 88 cases, while only 5 bodies were found in parks (Table 2). The same applies for the altitudinal distribution as most of the cases (77) were between 0 and 500 m, while there was only 1 case between 2000 and 2750 m (Table 3). Due to the discrepancies between these variables, we could not compare them equally. Other not-tested variables as interspecific competition or potential presence and influence of xenobiotics were not taken into consideration.