Establishing Silphids in the invertebrate DNA toolbox: a proof of concept.

Environmental DNA (eDNA) analyses are an increasingly popular tool for assessing biodiversity. eDNA sampling that uses invertebrates, or invertebrate DNA (iDNA), has become a more common method in mammal biodiversity studies where biodiversity is assessed via diet analysis of different coprophagous or hematophagous invertebrates. The carrion feeding family of beetles (Silphidae: Coleoptera, Latreille (1807)), have not yet been established as a viable iDNA source in primary scientific literature, yet could be useful indicators for tracking biodiversity in forested ecosystems. Silphids find carcasses of varying size for both food and reproduction, with some species having host preference for small mammals; therefore, iDNA Silphid studies could potentially target small mammal communities. To establish the first valid use of iDNA methods to detect Silphid diets, we conducted a study with the objective of testing the validity of iDNA methods applied to Silphids using both Sanger sequencing and high throughput Illumina sequencing. Beetles were collected using inexpensive pitfall traps in Alberta, Michigan in 2019 and 2022. We successfully sequenced diet DNA and environmental DNA from externally swabbed Silphid samples and diet DNA from gut dissections, confirming their potential as an iDNA tool in mammalian studies. Our results demonstrate the usefulness of Silphids for iDNA research where we detected species from the genera Anaxyrus, Blarina, Procyon, Condylura, Peromyscus, Canis, and Bos. Our results highlight the potential for Silphid iDNA to be used in future wildlife surveys.

Large carrion and burying beetles evolved from Staphylinidae (Coleoptera, Staphylinidae, Silphinae): a review of the evidence.

Large carrion beetles (Silphidae) are the focus of ongoing behavioral ecology, forensic, ecological, conservation, evolutionary, systematic, and other research, and were recently reclassified as a subfamily of Staphylinidae. Twenty-three analyses in 21 publications spanning the years 1927-2023 that are relevant to the question of the evolutionary origin and taxonomic classification of Silphidae are reviewed. Most of these analyses (20) found Silphidae nested inside Staphylinidae (an average of 4.38 branches deep), two found Silphidae in an ambiguous position, and one found Silphidae outside Staphylinidae, as sister to Hydrophilidae. There is strong evidence supporting the hypothesis that large carrion beetles evolved from within Staphylinidae and good justification for their classification as the subfamily Silphinae of the megadiverse, and apparently now monophyletic, Staphylinidae. Considerable uncertainty remains regarding the interrelationships and monophyly of many staphylinid subfamilies. Nonetheless, the subfamily Tachyporinae was found to be the sister of Silphinae in more analyses (7) than any other subfamily.

Drastic changes in ground-dwelling beetle communities following high-intensity deer culling: insights from an island ecosystem.

The overabundance of large herbivores can have detrimental effects on the local environment due to overgrazing. Culling is a common management practice implemented globally that can effectively control herbivore populations and allow vegetation communities to recover. However, the broader indirect effects of culling large herbivores remain relatively unknown, particularly on insect species such as ground-dwelling beetles that perform key ecosystem processes such as decomposition. Here we undertook a preliminary investigation to determine how culling sika deer on an island in North Japan impacted ground-beetle community dynamics. We conducted pitfall trapping in July and September in 2012 (before culling) and again in 2019 (after culling). We compared beetle abundance and community composition within 4 beetle families (Carabidae, Scarabaeidae, Geotrupidae, and Silphidae), across seasons and culling treatments. We found each family responded differently to deer culling. Scarabaeidae displayed the greatest decline in abundance after culling. Silphidae also had reduced abundance but to a lesser extent compared to Scarabaeidae. Carabidae had both higher and lower abundance after culling, depending on the season. We found beetle community composition differed between culling and season, but seasonal variability was reduced after culling. Overall, the culling of large herbivores resulted in a reduction of ground-dwelling beetle populations, particularly necrophagous species dependent on dung and carrion for survival. Our preliminary research highlights the need for long-term and large-scale experiments to understand the indirect ecological implications of culling programs on ecosystem processes.

First record of Diamesus osculans (Vigors, 1825) (Coleoptera: Silphidae) colonization on a human corpse.

A partially skeletonized human corpse was found in bushes in Selangor, Malaysia in June 2020. Entomological evidence was collected during the autopsy and sent to the Department of Medical Microbiology and Parasitology, Faculty of Medicine, Universiti Teknologi MARA (UiTM) for minimum postmortem interval (PMImin) analysis. Standard protocols were applied when processing preserved and live insect specimens of both larval and pupal stages. Entomological evidence revealed that the corpse was colonized by Chrysomya nigripes Aubertin, 1932 (Diptera: Calliphoridae) and Diamesus osculans (Vigors, 1825) (Coleoptera: Silphidae). Chrysomya nigripes was chosen as the PMImin indicator as this fly species is an earlier colonizer compared to D. osculans beetle larvae which their presence is the indicative of late stage of decomposition. For the present case, the pupae of C. nigripes were the oldest insect evidence collected and based on the available developmental data, the estimated minimum PMI was established between 9 and 12 days. It is noteworthy to highlight that this is the first record of D. osculans colonization on a human corpse.

Establishing Age-Based Color Changes for the American Burying Beetle, Nicrophorus americanus Olivier, with Implications for Conservation Efforts.

The American burying beetle, Nicrophorus americanus Olivier, is a federally protected insect that once occupied most of eastern North America. Adult beetles feature distinct, recognizable markings on the pronotum and elytra, and color changes with age have been observed. Among the challenges faced by research scientists and conservation practitioners is the ability to determine beetle age in the field between and including teneral (young) and senescent (old) adult stages. Using 20 (10 male and 10 female) captive-bred beetles, we characterized the change in greyscale and red, green, and blue (RGB) color channels over the lifespan of each beetle for field-aging applications. Individual beetles were photographed at set intervals from eclosion to death, and color data were extracted using open-source ImageJ Version 1.54f software. A series of linear mixed-effects models determined that red color showed the steepest decrease among all color channels in the pronotum and elytral markings, with a more significant decrease in the pronotum. The change in greyscale between the pronotum and elytral markings was visibly different, with more rapid darkening in the pronotum. The resulting pronotum color chart was tested under field conditions in Oklahoma, aging 299 adult N. americanus, and six age categories (day range) were discernable by eye: teneral (0-15), late teneral (15-31), early mature (31-45), mature (45-59), early senescent (59-76), and senescent (76-90). The ability to more precisely estimate age will improve population structure estimates, laboratory breeding programs, and potential reintroduction efforts.

Behavioral dominance interactions between two species of burying beetles (Nicrophorus orbicollis and Nicrophorus pustulatus).

Closely related species with ecological similarity often aggressively compete for a common, limited resource. This competition is usually asymmetric and results in one species being behaviorally dominant over the other. Trade-offs between traits for behavioral dominance and alternative strategies can result in different methods of resource acquisition between the dominant and subordinate species, with important consequences for resource partitioning and community structure. Body size is a key trait thought to commonly determine behavioral dominance. Priority effects (i.e., which species arrives at the resource first), however, can also determine the outcome of interactions, as can species-specific traits besides size that give an advantage in aggressive contests (e.g., weapons). Here, we test among these three alternative hypotheses of body size, priority effects, and species identity for what determines the outcome of competitive interactions among two species of burying beetles, Nicrophorus orbicollis and N. pustulatus. Both overlap in habitat and seasonality and exhibit aggressive competition over a shared breeding resource of small vertebrate carrion. In trials, we simulated what would happen upon the beetles' discovery of a carcass in nature by placing a carcass and one beetle of each species in a container and observing interactions over 13 h trials (n = 17 trials). We recorded and categorized interactions between beetles and the duration each individual spent in contact with the carcass (the key resource) to determine which hypothesis predicted trial outcomes. Body size was our only significant predictor; the largest species won most aggressive interactions and spent more time in contact with the carcass. Our results offer insight into the ecology and patterns of resource partitioning of N. orbicollis and N. pustulatus, the latter of which is unique among local Nicrophorus for being a canopy specialist. N. pustulatus is also unique among all Nicrophorus in using snake eggs, in addition to other carrion, as a breeding resource. Our results highlight the importance of body size and related trade-offs in ecology and suggest parallels with other coexisting species and communities.

Culture-independent analyses of carrion beetle (Coleoptera: Silphidae) secretion bacterial communities.

IMPORTANCE: The manuscript explores the secretion bacterial community of carrion and burying beetles of the central plains of North America. A core secretion microbiome of 11 genera is identified. The host subfamily, secretion type, and collection locality significantly affects the secretion microbiome. Future culture-dependent studies from silphid secretions may identify novel antimicrobials and nontoxic compounds that can act as meat preservatives or sources for antimicrobials.

When the red-lined carrion beetle disrupts successional dynamics on large vertebrate carcasses.

In a field study of insect colonization on domestic pig carcasses, the typical colonization sequence took a different turn midway through decomposition when Necrodes surinamensis (Fabricius) (Coleoptera: Silphidae) dramatically increased in abundance on some carcasses. This provided an opportunity to test the prediction that N. surinamensis would aggregate at clumped carcasses but not as much at dispersed carcasses, thus leaving enough resources in less heavily colonized habitat patches to support weaker competitors, as predicted by the aggregation model of coexistence. Larger aggregations of this silphine beetle resulted in the collapse of blow fly (Diptera: Calliphoridae) maggot populations due to intraguild predation, and a decline or exclusion of late-successional insect species. This phenomenon has been reported in a European Necrodes species but it is the first time it is documented with N. surinamensis, a widespread species in North America. Substantial increases in adult and larval populations of N. surinamensis were first documented on clumped carcasses and were not as pronounced on dispersed carcasses. The specifics of N. surinamensis aggregations are discussed with reference to ecological and forensic implications, including the fact that feeding by N. surinamensis larvae makes the remains appear to have been altered by scavengers and decomposing for longer than they have.

Necrodes littoralis (Coleoptera: Silphidae) visiting and breeding on a carcass in Italy

@#The community that progressively colonizes a decaying corpse can be considered a small ecosystem mostly composed of sarcosaprophagous arthropods belonging to the orders Diptera and Coleoptera. Studies on these species are often performed through animal models to obtain data on their succession, behaviour and life cycle, together with information on habitat, corpse conditions, season and association with other species. These data may be relevant for forensic investigations, especially concerning the estimation of Post Mortem Interval (PMI). An investigation on the sarcosaprophagous insect community in a rural area was set in Calabria (Southern Italy), using a pig, Sus scrofa Linnaeus, 1758 (Artiodactyla: Suidae) as experimental model. Analyses of the community of Diptera and Coleoptera revealed the massive presence of Necrodes littoralis (Linnaeus, 1758) (Coleoptera: Silphidae). Adults of this species reached the carcass during the bloated stage and a large amount of larvae was detected from the decay stage onwards, simultaneous to the sharp decrease in dipteran larvae and pupae. The occurrence and the activity of N. littoralis should be considered to avoid misinterpretation and errors in estimating PMI in forensic investigation.

First report of the presence of Necrodes littoralis (L.) (Coleoptera: Silphidae) on a human corpse in Italy.

The colonization of a human body by Necrodes littoralis (L.) (Coleoptera: Silphidae) is reported for the first time in Italy. This species is both necrophagous and predator of necrophagous fauna. The body colonized by the coleopteran was found indoors, in an advanced decomposition stage, in a suburban area of Cosenza (Calabria, Southern Italy) in November. Insects (adults, puparia and larvae) were collected on and around the body. Puparia and larvae were raised in the laboratory until the adult stage for morphological identification, which was carried out through taxonomical keys. Besides N. littoralis, also the presence of Calliphora vicina Robineau-Desvoidy, Chrysomya albiceps (Wiedemann) (Diptera: Calliphoridae), Hydrotaea dentipes (Fabricius) (Diptera: Muscidae), and Creophilus maxillosus (L.) (Coleoptera: Staphylinidae) was detected. Necrodes littoralis is a species of forensic interest because it may colonize human and vertebrate corpses and has been reported elsewhere in Europe.

The complete mitochondrial genome of a burying beetle, Nicrophorus nepalensis Hope, 1831 (Coleoptera: Silphidae).

The complete mitochondrial genome of burying beetle Nicrophorus nepalensis Hope, 1831 was reported in this study. It was 17,299 bps in length and included 13 protein-coding genes (PCGs), 22 tRNA genes, 2 rRNA genes, and a 2693 bp A + T-rich control region. Phylogenetic analysis using 13 PCGs from 38 Staphyliniform beetle species revealed that N. nepalensis was clustered in Silphidae, which together with Staphylinidae formed one monophyletic clade within Staphylinoidea.

emOiceoptoma/em emtangi/em (Coleoptera: Silphidae: Silphinae), a new species of carrion beetle from eastern China.

A new species, Oiceoptoma tangi Sommer, Ruzicka, Nishikawa Schneider, new species, from Zhejiang Province, China, is described and illustrated. The new species is closely related to O. subrufum (Lewis, 1888), distributed in Central and North-eastern China, the Korean Peninsula, Far East of Russia and Japan, and to O. nigropunctatum (Lewis, 1888), endemic to Japan. All three species are keyed, and distribution maps of all three species are presented.

Heads or Tails? Dung Beetle (Coleoptera: Scarabaeidae: Scarabaeinae and Aphodiinae) Attraction to Carrion.

Necrophilous insects occupy an ecologically interesting niche because carrion is a highly desirable but ephemeral food source. Dung beetles (Coleoptera: Scarabaeidae: Scarabaeinae and Aphodiinae) within temperate regions are frequently found at carrion, but little is known about their attraction to this resource. Are dung beetles attracted to the carrion itself or are they indirectly attracted due to the exposed gastrointestinal contents? We investigated the association between dung beetles and carrion by examining the distribution of dung beetles on the cranial and caudal end of rat carcasses, delimiting a resource more attractive to necrophagous insects (cranial end) from a resource more attractive to coprophagous insects (caudal end). Dung beetle distribution on rat carcasses was compared with the distribution of carrion beetles (Coleoptera: Silphidae), which serve as a null model of distribution patterns for a taxon known to directly target carrion. Results demonstrated that dung beetles show higher attraction to the cranial end of rat carrion. A similar distribution pattern was found in carrion beetles, suggesting that similar resources were targeted. When dung beetles were grouped by behavioral guilds, rollers and tunnelers also shared this pattern of greater abundance at the cranial end, but dwellers showed no discernible difference.

Behavioral dominance interactions between Nicrophorus orbicollis and N. tomentosus burying beetles (Coleoptera: Silphidae).

Asymmetric interference competition, where one species is behaviorally dominant over another, appears widespread in nature with the potential to structure ecological communities through trade-offs between competitive dominance and environmental tolerance. The details of how species interact and the factors that contribute to behavioral dominance, however, are poorly known for most species, yet such details are important for understanding when and why trade-offs occur. Here, we examine behavioral interactions between two species of burying beetles (Coleoptera: Silphidae) that compete for limited breeding resources (i.e., small vertebrate carcasses) in nature, to identify behaviors involved in interference competition and to test if large body size, species identity, or time of arrival best predict behavioral dominance among species. To test these ideas, we placed same-sex individuals of Nicrophorus orbicollis (early to mid-summer breeder) and N. tomentosus (late summer to fall breeder) into an enclosure together with a 25-30 g mouse carcass (Mus musculus). We then video-recorded all behaviors, including neutral and aggressive interactions, for 13 h per trial (N = 14 trials). For each interaction, we assigned a winner based on which beetle retained its position instead of fleeing or retained possession of the carcass; the overall behavioral dominant was determined as the individual that won the most interactions over the length of the trial. We found that large body size was the best predictor of behavioral dominance. In most interactions, N. orbicollis was larger and dominant over N. tomentosus; however, when N. tomentosus was larger they outcompeted smaller N. orbicollis, illustrating the importance of body size in aggressive contests. The order of arrival to the carcass (priority effects) did not predict behavioral dominance. The larger size and abundance of N. orbicollis in nature suggest a competitive asymmetry between the species, supporting the idea that N. orbicollis constrains the ability of N. tomentosus to breed earlier in the summer.

Standardized Laboratory Methodology for the Evaluation of Foraging Strategies in Necrophilous Beetles: A Case Study of Necrophila (Calosilpha) brunnicollis (Coleoptera: Silphidae).

Precise data regarding feeding habits of necrobiont species are a key element of food web and evolutionary ecology. They can also be used to assess the utility and value of those species for forensic entomology, where obligatory necrophagous species in particular are considered good bioindicators of postmortem or preappearance interval. However, the feeding habits of many species are known only from anecdotal field observations, often reduced to vaguely defined categories-predatory, necrophagous, or omnivorous. To address this issue, we designed a simple, in vitro behavioral experiment allowing the quantification of food preferences. Next, we applied it on Necrophila (Calosilpha) brunnicollis (Kraatz, 1877), which is a common carrion beetle of East Asia with unresolved food preferences. The results suggest that this species is preferentially necrophagous, thus valuable for forensic research. Importantly, however, our experimental design allowed us to reveal that it also readily feeds on larvae of Diptera, although they compose a minor proportion of its diet. This methodology can be applied to other species, and it could provide evidence for future decision making in forensic research.

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.

Larval morphology of Nicrophorus (Nicrophorus) nepalensis Hope (Coleoptera: Silphidae: Nicrophorinae).

Larval morphology of all three instars of Nicrophorus (Nicrophorus) nepalensis Hope, 1831 (Coleoptera: Silphidae: Nicrophorinae) is described and illustrated, based on reared larvae. The eastern Palaearctic and Oriental N. (N.) nepalensis is similar to the western Palaearctic N. (N.) humator (Gleditsch, 1767) in a number of larval characters. This is congruent with recent classification of the genus Nicrophorus Fabricius, 1775 by Sikes, who suggested a close phylogenetic affinity of the N. nepalensis species group (with 16 species) with the N. humator species group (2 species). The generic description of larvae of Nicrophorus Fabricius, 1775 is expanded, based on detailed SEM observation.

Ecological divergence of burying beetles into the forest canopy.

Closely related species with overlapping geographic ranges encounter a significant challenge: they share many ecological traits and preferences but must partition resources to coexist. In Ontario, potentially eleven species of carrion beetles (Coleoptera: Silphidae) live together and require vertebrate carrion for reproduction. Their reliance on an ephemeral and uncommon resource that is unpredictable in space and time is thought to create intense intra- and interspecific competition. Evidence suggests that burying beetle species reduce competition by partitioning carrion for breeding across different habitats, temperatures, and seasons. Here, we test predictions of an alternative axis for partitioning carrion: vertical partitioning between the ground and forest canopy. We conducted a survey of carrion beetles from May to July 2016 at the Queen's University Biological Station across 50 randomly generated points using baited lethal traps at zero and six metres. Ground traps yielded more species and individuals compared to those in the canopy, and the number of individuals and species caught increased through the season in both trap types. Ground and canopy traps were accurately distinguished by the presence or absence of three species: ground traps contained more Nicrophorus orbicollis and Necrophila americana, while canopy traps contained more Nicrophorus pustulatus. We trapped 253 N. pustulatus in the canopy, but only 60 on the ground. N. pustulatus is thought to be rare across its geographic range, but our results suggest it is uniquely common in canopy habitats, demonstrating a vertical partitioning of habitat and resources. Our results are consistent with N. pustulatus having diverged into canopy habitats as a strategy to coexist with closely related sympatric species when competing for similar resources. We still, however, do not know the traits that allow N. pustulatus to flourish in the canopy, exactly how N. pustulatus uses canopy resources for breeding, or the factors that restrict the expansion of other burying beetles into this habitat.

Larval identification key to necrophagous Coleoptera of medico-legal importance in the western Palaearctic.

Several necrophagous Coleoptera species are frequently collected on cadavers, may occasionally act as intermediate or paratenic hosts of parasites, as vectors of pathogens or as allergens, and can also represent major pests of preserved animal products. However, despite their medical, veterinary and economic importance, there is a lack of reliable species identification tools for the larval stages (usually the only entomological evidence associated with medicolegal investigations), thus severely limiting their potential application as forensic indicators. Here, we provide an identification key to the larvae of the necrophagous Coleoptera species which have been recorded on carrion in the western Palaearctic region, based on easily observable morphological characters. In total, we provide diagnostic characters for the reliable identification of 23 necrophagous Coleoptera species within four different families (Cleridae, Dermestidae, Nitidulidae and Silphidae). In addition to the aforementioned families, we provide diagnostic characters for the identification of the larvae of families Histeridae, Staphylinidae and Trogidae, which can also be collected on cadavers. It is expected that the present key will facilitate the identification of larval material of necrophagous Coleoptera collected either in carrion succession studies or during medicolegal investigations, in order to further advance in the potential use of this insects as forensic tools.

Nesting strategies and disease risk in necrophagous beetles.

While the effects of carcass decomposition on microorganisms have been demonstrated in recent years, little is known of how this impacts necrophagous insects. A common assumption is that insects that exploit carcasses are exposed to a high density of potentially harmful microorganisms, but no field data have so far validated this. Necrophagous beetles such as the Scarabaeinae have complex nesting behaviors with elaborate parental care. So here, we begin to explore whether this conjunction of life history and nesting behavior represents an adaptive response to the threat posed by microbes in these environments, mainly by entomopathogens. We evaluated the density and distribution of fungi and bacteria from soil near the carcasses, and their ability to infect and kill insects that are in contact with this soil during the decomposition process. Our data showed an increase in the density and activity of opportunistic or facultative pathogens during the apex of decomposition, when there is a predominance of necrophagous insects. Meanwhile, the survivorship of bait insects decreased when in contact with soil from this period of decomposition, indicating a potential risk of infection. However, the density and activity of these microorganisms decreased with distance from the carcass, mainly with depth, which would benefit tunneller beetles in particular. We have thus provided the first field data to show that necrophagous insects are indeed exposed to high densities of potentially harmful microorganisms. Furthermore, we propose that some parental care strategies may have arisen not only as a response to competition, but also as adaptations that reduce the risks of disease. Although we have focused on carrion feeders, we suggest that the same occurs with coprophagous beetles, as both carrion and dung are nutrient-rich resources.