Species identification and delimitation
The 7870 specimens from the Churchill collection were assigned to 1630 MOTUs at the 2% cutoff range. On average, we identified a different species for every 4.8 individuals sampled (SD = 11.3). Most MOTU were sparsely represented, with a mode of 1 (n = 734) and a median of 2. Only 194 MOTUs included 10 or more individuals, and only three of these included 100 or more (Figure 6). For the Churchill 2010 subcollection, there were 722 MOTUs, and the mean number of individuals per MOTU was 2.9 (SD = 4.5, mode = median = 1, maximum = 55, but pre-sorting of this subcollection limited the number of replicates per MOTU). Singleton MOTUs made up a similar proportion of the 2010 subcollection as they did of the parent collection (362 MOTUs = 50% of the 2010 subcollection; 734 MOTUs = 45% of the parent collection).
Based on number of MOTUs, the most diverse families in the overall Churchill collection were the Ichneumonidae (915 MOTUs), Braconidae (313 MOTUs), Diapriidae (83 MOTUs) and Tenthredinidae (72 MOTUs). All of these families except the Diapriidae included some specimens that had been previously identified below the family level. We also resolved 132 MOTUs among the Chalcidoidea, which were not determined to family. Each of the other taxonomic groups in this study, including the entire superfamily Cynipoidea, yielded fewer than 30 MOTUs.
As determined by traditional morphological taxonomy, the 4787 specimens of Ichneumonidae in the overall collection encompassed at least 13 subfamilies, 34 genera, and 49 species. However, 1861 individuals (38.9%) carried no identification below the family level. At the 2% cutoff level, 915 MOTUs were resolved for this family; if this cutoff level provides a good approximation to interspecific divergence in Ichneumonidae, then morphological examination previously resolved only 5.4% of the diversity present. Similarly, for 1367 specimens of Braconidae, 8.6% had no identification below the family level and the number of resolved species was only 40.9% of the MOTU total. For 479 Tenthredinidae specimens, 23.2% had no identification below the family level, and the resolved species count was 25.0% of the MOTU total.
These values should be considered approximations, for several reasons. First, no single MOTU cutoff level can definitively delimit all species; the 2% cutoff level for MOTU definition is known to be insufficiently stringent for some hymenopteran subtaxa and too stringent for others [1, 2, 32, 33]. Second, even the better-identified families in this collection have not to date received the same amount of scrutiny by morphological taxonomists, and the most completely determined taxa as of May 2012 (bees, vespid wasps, and microgastrine braconids) made up only a small proportion of the collection.
The second issue – unequal attention given to different taxa – is not a trivial one, because full morphological species identification requires considerable effort and expertise, and identifying a diverse collection requires a diverse assemblage of specialists. The low rate of morphological specimen identification says little about the ability of taxonomists to identify a specimen in hand, but much about the difficulty of placing every specimen from a large survey into the hands of the appropriate specialist [23, 34]. Specimens from this study may receive more attention from specialists in the future, and the availability of DNA barcodes will make it easier for taxonomists to delimit and perhaps even describe species in this collection via integrative methods [25, 35].
Rarefaction and richness estimates
Rarefaction/accumulation curves show no clear approach to asymptotes for either the complete Churchill collection (abundance-based) or the 2010 subcollection (incidence-based over 12 sites) The Chao 1 richness estimate at 2% cutoff is 2624 MOTUs, with a 95% CI of 2446 through 2840. By this estimate, the 1630 MOTUs defined from our collection cover approximately 62% ± 5% of total richness. Since this estimate was based on MOTUs rather than on morphological determinations, it is independent of the amount of attention each subtaxon received from a morphological taxonomist. It should still be considered exploratory, because we did not sequence every one of the many thousands of individual specimens originally collected, and because the overall collection was not controlled for variation in sampling effort at different locations throughout the collection range.
Abundance-based measures may also be biased by over- or under-representation of some species independent of site selection or sampling effort. For example, our collection contained social insects (ants, bumble bees, and vespines) that can be oversampled when traps are set near nests or other sites of high colony activity. As individual specimens, social Hymenoptera were among the most abundant members of the collection, but also among the least diverse, and this was verified by both sequence data and morphological examination. Ants comprised 7 MOTUs, including Camponotus herculaneus (n = 27), two apparently distinct members of the Formica fusca complex (n1 = 45 and n2 = 282,) one species of Myrmica (Myrmica alaskensis, n = 9,) and up to three members of the genus Leptothorax (n1 = 1, n2 = 4, n3 = 5). Similarly, the collection included seven identified species of Bombus whose sample sizes ranged from a minimum of two (B. flavifrons) to a maximum of 44 (B. sylvicola), and which formed eight MOTUs (jMOTU split B. mixtus and B. sylvicola each into two MOTUs, and pooled B. frigidus and B. jonellus into one MOTU). Three species of Vespinae were also present, with sample sizes ranging from eight (Vespula intermedia) to 87 (Dolichovespula albida). Vespine wasps were assigned to three MOTUs that were congruent with morphological identifications.
Similarity among sites
For the 2010 subcollection, values of the Chao-Sørensen-Est abundance-based similarity index were highly variable among different site pairs, ranging from 0.020 to 0.867. However, these values were not significantly correlated with the linear distances between site pairs (Figure 2).
Almost all specimens in the Hymenoptera collection were of winged insects or their larvae, and we expect these species to disperse widely among sites. The only exceptions were worker ants and female Dryinidae; however, since ant reproductives and male dryinids are winged, these species can still disperse via flight. The wide range of between-site similarities could be related to microhabitat differences, variation among sampling methods, or sampling at different times throughout the season. For example, even in the short sub-Arctic summer, the Trichoptera of the Churchill region show some seasonal variation in species presence and abundance [4], and this may be true of Hymenoptera as well. The Chao-Sørensen-Est similarity model should mitigate some of these effects, since it includes an estimate of unseen species and also accounts for sample size difference (the latter indirectly, via species/MOTU counts at each pair of sites). Whether or not methodological or microhabitat differences affected incidence or abundance measures of some MOTUs, the 2010 subcollection showed no evidence for distance-dependent substructure across the study area.
Feeding ecology and the importance of parasitoids
The hymenopteran fauna of the Churchill region is dominated by parasitoids, and the parasitoid fauna of Churchill is dominated by the Ichneumonoidea (Ichneumonidae + Braconidae). We observed this pattern not only in the overall collection, but also in the 2010 subcollection, in which specimens were deliberately selected for inclusion by a method designed to maximize taxonomic breadth.
Worldwide, the Ichneumonidae and Braconidae are thought to be the first and second most diverse families of Hymenoptera [5, 36], so this result is not unexpected in any regional survey of the order. Only about 12% of identified parasitoid MOTUs were non-ichneumonoids, but these represented at least six different hymenopteran superfamilies, including members of the families Diapriidae (Diaprioidea), Dryinidae (Chrysidoidea), Megaspilidae (Ceraphronoidea), Platygastridae (Platygastroidea), Proctotrupidae (Proctotrupoidea), plus multiple families of Chalcidoidea.
Still, parasitoids cannot reproduce without hosts, so the diversity of parasitoids in this sub-Arctic location implies a high level of host diversity. Many members of the superfamily Ichneumonoidea are parasitoids of immature Lepidoptera, but they also parasitize hosts from other holometabolous orders, nymphal Hemimetabola, and arachnids [5, 37, 38]. At least four ichneumonid subfamilies (Campopleginae, Ctenopelmatinae, Tersilochinae, and Tryphoninae) target sawflies, and others (e.g. Mesochorinae) include hyperparasitoids. Non-ichneumonoid parasitoids can also exploit a wide range of hosts, including Diptera (by Diapriidae), Hemiptera (by Dryinidae), and the variety of hosts that are susceptible to attack by Chalcidoidea [5, 37]. A more detailed list of Churchill parasitoid subtaxa and their presumed hosts is included in Additional file 1.
Parallel studies of the Churchill fauna reveal no shortage of potential hosts. Our Hymenoptera collection includes more than 70 presumed species of tenthredinoid sawflies. The comprehensive Churchill terrestrial arthropod collection includes approximately 1800 species of Diptera, 315 of Lepidoptera, 300 of Coleoptera, 90 of Hemiptera, and 200 of Araneae [S.J. Adamowicz et al. pers. comm.]. These numbers are probably conservative, since they are based on observed MOTU or species richness rather than on extrapolated richness. If true species richness of these other orders is similar to that of Hymenoptera (e.g. approximately one-third of the true richness is not accounted for in the collection), then we would expect that more than 4000 species from these groups are potential hosts for hymenopteran parasitoids in the Churchill region. Details of host-parasitoid interactions are far beyond the scope of this survey, so we cannot tell whether some parasitoids attack multiple hosts, or whether some hosts are attacked by multiple parasitoids. However, DNA barcode data can provide the earliest clues to the presence of cryptic host-parasitoid interactions that can be targeted for future taxonomic and ecological study [39, 40].
Non-parasitoids in this collection were dominated by obligate herbivores, including bees (Andrenidae + Apidae + Halictidae + Megachilidae, 20 MOTUs) and sawflies (Tenthredinidae, 73 MOTUs). Predators were much less diverse (Crabronidae, six MOTUs, Pompilidae, one MOTU, and Vespidae, seven MOTUs). Other aculeates included Chrysididae (four MOTUs) and Formicidae (seven MOTUs). The chrysidids – all members of the cleptoparasitic subfamily Chrysidinae – are likely to invade nests of host taxa that were represented in this collection [41]. One chrysidine, Omalus aeneus, is a known cleptoparasite in nests of Pemphredon spp. (Crabronidae), a genus represented by two individuals in the overall collection. The other chrysidines were not identified to species, but some Nearctic chrysidines are specialized cleptoparasites of the cavity nests of eumenine vespids [41]. The overall collection contained three determined species of cavity-nesting Eumeninae: Ancistrocerus albophaleratus, A. waldenii, and Euodynerus leucomelas[42].
Geography and climate
Some previous surveys of regional ichneumonid fauna showed an unusual pattern – a higher diversity of Ichneumonidae (but not necessarily other parasitoid taxa) in some temperate regions than in the tropics [8, 43]. However, this conclusion has been challenged [13, 44], because in practice, it is difficult to design and implement a sampling protocol free of methodological bias. As a result, spurious patterns of diversity can be traced to over- or under-sampling of some subtaxa or habitats that share characteristics other than latitude [12, 13, 45]. Some of these factors are additionally complicated by uncertain taxonomy. For example, molecular evidence may reveal that some “generalist” parasitoid species are actually made up of multiple cryptic species with more narrow host specializations than previously recognized [15, 39, 40].
These issues make it difficult to define a priori expectations of parasitoid diversity in a sub-Arctic environment, and to compare it with diversity at other latitudes. For example, the earliest descriptions of anomalous latitudinal gradients in parasitoid diversity pre-date the routine use of both large-scale trapping [14, 46] and molecular methods [47, 48] in biodiversity research. The MOTU approach is not always sufficient for species delimitation, and is not sufficient to describe new species, but it has the advantage of being quantitative, reproducible, and feasible even when morphological determinations are impractical [24]. As such, barcode-based diversity studies can reveal large-scale geographic or ecological patterns of cryptic diversity, and can also identify specific taxa in need of further study by specialist taxonomists or conservation biologists.